Fortification of staple foods with zinc for improving zinc status and other health outcomes in the general population

Abstract

Background

Zinc deficiency is a global nutritional problem, particularly in children and women residing in settings where diets are cereal based and monotonous. It has several negative health consequences. Fortification of staple foods with zinc may be an effective strategy for preventing zinc deficiency and improving zinc‐related health outcomes.

Objectives

To evaluate the beneficial and adverse effects of fortification of staple foods with zinc on health‐related outcomes and biomarkers of zinc status in the general population.

Search methods

We searched the following databases in April 2015: Cochrane Central Register of Controlled Trials (CENTRAL, Issue 3 of 12, 2015, the Cochrane Library), MEDLINE & MEDLINE In Process (OVID) (1950 to 8 April 2015), EMBASE (OVID) (1974 to 8 April 2015), CINAHL (1982 to April 2015), Web of Science (1900 to 9 April 2015), BIOSIS (1969 to 9 April 2015), POPLINE (1970 to April 2015), AGRICOLA, OpenGrey, BiblioMap, and Trials Register of Promoting Health Interventions (TRoPHI), besides regional databases (April 2015) and theses. We also searched clinical trial registries (17 March 2015) and contacted relevant organisations (May 2014) in order to identify ongoing and unpublished studies.

Selection criteria

We included randomised controlled trials, randomised either at the level of the individual or cluster. We also included non‐randomised trials at the level of the individual if there was a concurrent comparison group. We included non‐randomised cluster trials and controlled before‐after studies only if there were at least two intervention sites and two control sites. Interventions included fortification (central/industrial) of staple foods (cereal flours, edible fats, sugar, condiments, seasonings, milk and beverages) with zinc for a minimum period of two weeks. Participants were members of the general population who were over two years of age (including pregnant and lactating women) from any country.

Data collection and analysis

Two review authors independently assessed the eligibility of studies for inclusion, extracted data from included studies, and assessed the risk of bias of the included studies.

Main results

We included eight trials (709 participants); seven were from middle‐income countries of Asia, Africa, Europe, and Latin America where zinc deficiency is likely to be a public health problem. Four trials compared the effect of zinc‐fortified staple foods with unfortified foods (comparison 1), and four compared zinc‐fortified staple foods in combination with other nutrients/factors with the same foods containing other nutrients or factors without zinc (comparison 2). The interventions lasted between one and nine months. We categorised most trials as having unclear or high risk of bias for randomisation, but low risk of bias for blinding and attrition. None of the studies in comparison 1 reported data on zinc deficiency.

Foods fortified with zinc increased the serum or plasma zinc levels in comparison to foods without added zinc (mean difference (MD) 2.12 µmol/L, 95% confidence interval (CI) 1.25 to 3.00 µmol/L; 3 studies; 158 participants; low‐quality evidence). Participants consuming foods fortified with zinc versus participants consuming the same food without zinc had similar risk of underweight (average risk ratio 3.10, 95% CI 0.52 to 18.38; 2 studies; 397 participants; low‐quality evidence) and stunting (risk ratio (RR) 0.88, 95% CI 0.36 to 2.13; 2 studies; 397 participants; low‐quality evidence). A single trial of addition of zinc to iron in wheat flour did not find a reduction in proportion of zinc deficiency (RR 0.17, 95% CI 0.01 to 3.94; very low‐quality evidence). We did not find a difference in serum or plasma zinc levels in participants consuming foods fortified with zinc plus other micronutrients when compared with participants consuming the same foods with micronutrients but no added zinc (MD 0.03 µmol/L, 95% CI ‐0.67 to 0.72 µmol/L; 4 studies; 250 participants; low‐quality evidence). No trial in comparison 2 provided information about underweight or stunting.

There was no reported adverse effect of fortification of foods with zinc on indicators of iron or copper status.

Authors' conclusions

Fortification of foods with zinc may improve the serum zinc status of populations if zinc is the only micronutrient used for fortification. If zinc is added to food in combination with other micronutrients, it may make little or no difference to the serum zinc status. Effects of fortification of foods with zinc on other outcomes including zinc deficiency, children’s growth, cognition, work capacity of adults, or on haematological indicators are unknown. Given the small number of trials and participants in each trial, further investigation of these outcomes is required.

Plain language summary

The effects of fortification of common foods with zinc on health outcomes

Background

It is believed that zinc deficiency is widespread globally, particularly in children and women living in low‐ and middle‐income countries. Cereal‐based foods are rich in dietary fibre and phytates, which reduce absorption of zinc from the intestine. As people in low‐income households eat a lot of cereal‐based foods, they are more likely to develop zinc deficiency. Fortification of common staple foods with zinc alone or in combination with other vitamins and minerals has been proposed as an intervention to increase intake of zinc in populations who consume these foods.

Review question

We evaluated the effects of fortification of staple foods with zinc on blood zinc levels and health‐related outcomes in the general population above two years of age.

Study characteristics

We included eight trials (709 participants); seven were from middle‐income countries of Asia, Africa, Europe, and Latin America where zinc deficiency is likely to be a public health problem.

Key results and quality of the evidence

The effect of fortification of foods with zinc on incidence of zinc deficiency is uncertain. Fortification of foods with zinc may slightly improve the blood zinc levels of populations if zinc is the only micronutrient used for fortification. Zinc added to food in combination with other micronutrients may make little or no difference to blood zinc levels. The fortification of foods with zinc may make little or no difference on incidence of underweight or stunting. The effects of fortification of foods with zinc on other outcomes, including wasting and weight/height, are unknown. Fortification of foods with zinc does not seem to have any adverse effect on indicators of iron or copper status. Most studies included in this review involved a small number of participants and were judged to be at low or unclear risk of bias. There were also some inconsistencies in the results across different studies.

Background

Description of the condition

Nutritional deficiencies continue to affect many people in low‐ and middle‐income countries. Insufficient consumption of protein, energy, and micronutrients, particularly iodine, iron, vitamin A, and zinc, puts these people at an increased risk of disease and death (WHO 2009a).

Zinc is a vital constituent of many metallo‐enzymes important for the normal functioning of the human reproductive, neurologic, immune, dermatologic, and gastrointestinal systems. This nutrient is widely present in foods; the highest concentrations are found in meat, fish and shellfish, nuts, seeds, legumes, and whole‐grain cereals, and lower concentrations are found in tubers. Animal products contain substantial amounts of zinc in a readily absorbable form. Conversely, zinc from whole‐grain cereals and legumes is absorbed less efficiently because uptake by the intestine is inhibited by other dietary components, such as fibre and phytate (Gibson 2012). As most diets in low‐income households are rich in these foods, this population is more likely to develop zinc deficiency. Groups that are especially vulnerable to zinc deficiency include infants and young children who are receiving unfortified complementary foods, children recovering from malnutrition, adolescents, pregnant and lactating women, and the elderly.

It is believed that zinc deficiency is widespread globally, particularly in children and women residing in low‐ and middle‐income countries. Data from these countries indicate that zinc deficiency occurs at a sufficient scale to be classified as a significant public health problem (Kapil 2011; Shah 2011;Wessells 2012). Estimates based on dietary zinc and phytate intakes suggest that more than 25% of the population in Latin America and the Caribbean, South and Southeast Asia, and sub‐Saharan Africa is at risk of inadequate zinc intake (IZiNCG 2004). Zinc deficiency in combination with other micronutrient deficiencies such as iron or vitamin A, childhood undernutrition, and suboptimal breastfeeding causes 7% of deaths and 10% of the total disease burden (WHO 2009a). The combined burden from these nutritional risks is considered to be equivalent to the entire disease and injury burden of high‐income countries (WHO 2009a).

The manifestation and severity of zinc deficiency varies based on age. Zinc is ubiquitous, and because it is involved in so many core areas of metabolism, the features of mild zinc deficiency are frequently nonspecific. Zinc deficiency in childhood may lead to diarrhoea (Lazzerini 2012), impairment of cognitive function and behavioural problems (Gogia 2012), hair loss, inflammation of the eyelids and conjunctiva, growth retardation (Imdad 2011; Levenson 2011), and recurrent infections (Yakoob 2011). Fertility, reproductive performance, and work capacity are also affected in adolescents and adults (Atig 2012; Bernhardt 2012; Kawade 2012; Lukaski 2000; Shah 2006), while infections are recurrent among the elderly (Pae 2012).

Most dietary zinc is absorbed by the small intestine, particularly in the jejunum, and the efficiency of intestinal zinc absorption is increased when there is zinc depletion. Once absorbed, it is bound to albumin, and transported from the intestine into the body. Zinc is found in all body tissues, with approximately 85% of the whole‐body zinc residing in muscle and bone and 11% in the skin and liver. The remaining 2% to 3% of the whole‐body zinc is found in the other tissues, including only 0.1% in the plasma (King 2000). Homeostatic regulation of zinc metabolism is achieved by adjusting zinc absorption and endogenous intestinal excretion from pancreatic and intestinal cell secretions (King 2000).

Biochemical and functional markers of zinc status are needed to assess the impact of programmes aimed at improving the zinc nutrition of populations. At present there are no simple, quantitative markers of zinc status that can be used to identify zinc deficiency in individuals. However, the International Zinc Nutrition Consultative Group recommends serum or plasma zinc as the best available biomarker of the zinc status in populations (IZiNCG 2004), because of its correlation with dietary zinc intake and a consistent response to zinc supplementation (de Benoist 2007; Hess 2009a). Serum zinc concentrations range between 80 µg/dL and 100 µg/dL (12 µmol/L to 15 µmol/L) in healthy adults, although concentrations differ by sex, age, infection, and fasting status. Currently there are reference data for most age and sex groups, and it has been proposed that if more than 20% of the population (or population subgroup) falls below the corresponding cutoff, the whole population (or subgroup) should be considered to be at risk of zinc deficiency (IZiNCG 2004).

Other biomarkers have also been suggested for assessing zinc status, including erythrocyte (red blood cell) zinc, hair zinc, or alkaline phosphatase activity, but many issues are still pending regarding their sensitivity, specificity, reliability, and feasibility of use in the field. However, stunting (low height‐ or length‐for‐age) remains as the preferred functional marker of zinc status, as it is often responsive to supplemental zinc, and is widely used in most health‐ and nutrition‐monitoring activities (de Benoist 2007).

Description of the intervention

Fortification is defined as the process of adding nutrients to commonly eaten foods, beverages, or condiments during food processing at the industrial (central) level, with the goal of improving the quality of the diet. Fortification programmes represent promising long‐term strategies to combat various micronutrient deficiencies among recipients. Fortification has played a major role in increasing the dietary intake of some vitamins and minerals and has contributed to the elimination of vitamin and mineral malnutrition in certain settings, and is postulated to have the advantages of better compliance, long‐term sustainability, and potential to reach the intended population (WHO/FAO 2006). Iron and folic acid are two nutrients commonly added to staple foods and condiments. Fortification of foods with iron has been proved to improve iron status and haemoglobin and result in a significant reduction of anaemia in the general population (Gera 2012). Fortification of cereal products, including wheat flour, with folic acid has also contributed to a reduction of neural tube defect‐affected pregnancies (for example spina bifida) in some countries (Castillo‐Lancellotti 2012). It is thus postulated that fortification of staple foods with zinc could result in an increased daily zinc intake, which may prevent deficiency and improve zinc‐related health outcomes.

The World Health Organization recommends the addition of zinc to wheat and maize (also known as corn) flours (WHO 2009b). However, the addition of this micronutrient to foods has generally been confined to infant formula milks (in the form of zinc sulphate), complementary foods, and ready‐to‐eat breakfast cereals. In some countries, such as Indonesia, it is mandatory to add zinc to wheat noodles, while other countries like Mexico have voluntary fortification programmes where zinc and other micronutrients are added to wheat and corn flours used for preparing bread and tortillas, milk, and food supplements provided in social programmes. More recently, several Latin American countries have expressed some interest in fortifying cereal flours with zinc (Brown 2010). According to the Food Fortification Initiative, in 2012 at least 20 countries had mandatory zinc fortification for wheat flour, and three countries had it for maize flour, although the level of implementation varied among countries (Food Fortification Initiative 2012). From a public health perspective, mandatory fortification of staple foods with zinc has the potential to reach everyone in a population, particularly vulnerable groups. Effective fortification of staple foods with zinc can help ensure access and equity to adequate zinc in the diets of all children and women, especially those who are less well off.

How the intervention might work

Staple foods are those eaten regularly in such quantities that they constitute the dominant part of the diet and supply a major proportion of energy and nutrient needs (FAO 2012). Staple foods are acceptable vehicles for a fortification programme as they are consumed by a large portion of the target population in relatively constant amounts. Potentially suitable staple food vehicles for zinc fortification in public health include cereal grains or their flours (wheat, rice, maize); condiments and seasonings; and powdered or liquid milk (WHO/FAO 2006). Appropriate fortification of a staple food has the potential to ensure a predictable and fairly stable level of intake of the added nutrient. Moreover, it does not require changes in the dietary habits of the population, thereby simplifying the implementation process.

Zinc is a suitable micronutrient for fortification of staple foods, as total daily zinc absorption has been shown to increase following an increase in dietary zinc (Hess 2009b). Although phytates in food reduce zinc absorption, the total amount of zinc absorbed from fortified foods is greater than the amount absorbed from unfortified foods. Thus, fortification of staple foods with zinc has the potential to improve the zinc nutrition of populations, resulting in better health outcomes, such as improved childhood growth and reduction in infectious morbidity. However, the impact of zinc fortification could be dependent upon the baseline zinc status of populations, choice of food vehicle, dose and duration of the intervention, and selection of the fortificant.

The choice of a particular chemical form of zinc to be used in food fortification should be based on its solubility in water, intragastric (within the stomach) solubility, taste, cost, side effects, and safety (IZiNCG 2004). In general, water‐soluble compounds, such as zinc‐EDTA, zinc acetate, zinc gluconate, and zinc sulphate, are considered to be more readily absorbable than compounds with limited solubility at neutral pH. Zinc sulphate and zinc oxide are thought to be the least expensive and most commonly used by the food industry (Brown 2007; IZiNCG 2007).

In addition to efficacy, the effectiveness of fortification of foods with zinc in public health depends on several factors related to policies and legislation including: production and supply of the fortified food; development of delivery systems; development and implementation of external and internal food quality control systems; and the development and implementation of strategies for information, education, and communication for behavioural change among consumers. We have presented a generic logic model for micronutrient interventions in public health that depicts these processes in Figure 1 (De‐Regil 2013; WHO/CDC 2011).

1.

WHO/CDC logic model for micronutrient interventions in public health (with permission from WHO)

The potential concerns of adding zinc to food include its possible interactions with iron and copper. Anaemia may occur in zinc‐supplemented populations due to interference with iron absorption, and inhibited iron transport via decreased copper absorption (Dekker 2010). Common metal transporters are involved in active transport of iron, copper, and zinc (Espinoza 2012). In‐vitro models have demonstrated the reduction in bioavailability of each of these minerals by the presence of other minerals in the diet (Arredondo 2006). Negative effects of zinc supplementation on indices of iron and copper status have been reported (Sandstrom 2001). However, a systematic review reported that zinc supplementation, at doses typically used in randomised trials, does not decrease the haemoglobin concentrations (Dekker 2010). On the other hand, a few randomised controlled trials have documented a beneficial effect of adding zinc to iron treatment on haemoglobin response and iron indices in anaemic women, in Kolsteren 1999, and in children, in Alarcon 2004. The beneficial effects of zinc on haemoglobin concentration can be explained by its role in DNA synthesis (Ishido 1999), modulation of erythropoiesis (red blood cell production), and vitamin A metabolism (Alarcon 2004).

Why it is important to do this review

There are very few national surveys reporting on zinc status. Calculations based on diet and stunting suggest that zinc deficiency is widespread in low‐ and middle‐income countries, especially among women and children. For example, in 2004 zinc deficiency was estimated to be responsible for 260,502 deaths in Africa; 182,546 in Asia; and 10,159 in Latin America, accounting for 4.4% of all childhood deaths between six months and five years of age (Fischer Walker 2009). Recent mathematical models based on national food balance data that the Food and Agriculture Organization of the United Nations obtained suggest that the estimated global prevalence of inadequate zinc intake varied between 12% to 66%, depending on which methodological assumptions were applied (Wessells 2012). Studies from low‐ and middle‐income countries report zinc deficiency of sufficient magnitude to be a significant public health problem (Kapil 2011; Shah 2011). Interventions to improve population zinc status are thus recommended.

Fortification of food with zinc appears to be a promising strategy because of its relatively low cost and long‐term sustainability. For example, zinc concentrations in rural Chinese women improved after 24 months of intervention (Hess 2009b; Huo 2011; Huo 2012). Zinc intake and absorption increase when some zinc‐fortified foods are consumed, but the impact as a public health intervention remains unknown (WHO/FAO 2006). There is a paucity of systematic assessments of the benefits and safety of fortification of foods with zinc to inform public health policy. This systematic review attempted to address the gap in the information on this subject.

Objectives

To evaluate the beneficial and adverse effects of fortification of staple foods with zinc on health‐related outcomes and biomarkers of zinc status in the general population.

Methods

Criteria for considering studies for this review

Types of studies

We included randomised controlled trials that were randomised at the level of the individual or cluster in this review. In anticipation of non‐availability of sufficient trials of this nature, we planned to include data from the following additional study designs.

• Non‐randomised trials at the level of the individual or cluster. To be eligible, such trials should have had a concurrent comparison group (as defined later), preferably with adjustment for baseline characteristics and confounders.

• Controlled before‐after (CBA) studies where allocation to the different comparison groups is not made by the investigators. Outcomes of interest were measured in both intervention and control groups before zinc fortification intervention was introduced, and again after introduction of the intervention.

We included cluster‐randomised trials, non‐randomised cluster trials, and CBA studies with at least two intervention sites and two control sites.

We included eligible studies irrespective of the date of publication, language of publication, or publication status.

Types of participants

Members of the general population who were over two years of age (including pregnant and lactating women), from any country. We chose this age group because the types of interventions being evaluated (see below) would be predominantly applicable to this population. We excluded trials on: (i) participants with a critical illness or severe comorbidities; (ii) participants who were being tube‐fed or are nil‐by‐mouth; and (iii) older adults in long‐term care facilities who would be receiving regular nutrition intervention and monitoring for clinical status. Although the risk of zinc deficiency is greater in low‐ and middle‐income countries, it can also occur in relatively wealthy countries. We therefore included trials conducted in any country irrespective of the income status, and performed a subgroup analysis (low‐ and middle‐income countries versus others) to probe any differences with this categorisation.

Types of interventions

We included trials in which common staple foods (for example wheat flour, maize flour or cornmeal, oils, milk, pulses, bread, sauces, and beverages) or condiments or seasonings had been industrially (centrally) fortified with zinc, irrespective of the fortification technology (or compound) used. The intervention must have been in operation for a minimum period of two weeks. We considered studies with co‐interventions (for example nutrition education, deworming, additional food supplementation, diarrhoea control, and so forth) for inclusion if the only difference between the two comparison arms was micronutrient fortification (zinc alone or in combination with additional micronutrients).

We included the following staple foods and condiments in this review.

• Edible oils for household use (Codex 1981a; Codex 1981b; Codex 1999a).

• Edible fats for household use (Codex 1981a).

• White or brown sugar for household use (Codex 1999b).

• Wheat flour or its subproducts (Codex 1985a).

• Rice, including paddy rice, milled rice, parboiled rice, husked rice, glutinous rice (Codex 1995a).

• Maize flour or its subproducts (Codex 1985b; Codex 1985c).

• Condiments and seasonings for household use (Codex 1995b).

• Milk and dairy products for household use (Codex 1999c).

• Fruit juices and nectars (Codex 2005).

We made the following comparisons.

• Food fortified with zinc versus same food without added zinc (comparison 1).

• Food fortified with zinc plus other micronutrients versus food fortified with other micronutrients without zinc (comparison 2).

• Food fortified with zinc versus no intervention (comparison 3).

We evaluated the comparisons mentioned above separately. We did not include comparisons of food fortification with zinc versus other forms of micronutrient interventions (that is supplementation or dietary diversification). We did not include studies examining point‐of‐use fortification of foods with micronutrient powders, biofortification, or provision of zinc supplements. We also excluded trials involving fortification of drinking water with zinc, but considered beverages fortified with zinc alone or with other vitamins and minerals (fruit juices, milk). We did not include studies evaluating multiple micronutrient fortification if zinc was not the only difference between intervention and control groups.

Types of outcome measures

The primary outcomes across all populations in this review were zinc deficiency, serum plasma zinc concentrations, and undernutrition.

Primary outcomes

Children (24 to 59 months of age)

• Zinc deficiency (as defined by authors, depending on the age and sex).

• Serum or plasma zinc (in µmol/L).

• Stunting (author defined).

• Underweight (author defined)

Children (5 to 11.9 years of age)

• Zinc deficiency (as defined by authors, depending on the age and sex).

• Serum or plasma zinc (in µmol/L).

Adolescent girls and boys (12 to 18.9 years of age)

• Zinc deficiency (as defined by authors, depending on the age and sex).

• Serum or plasma zinc (in µmol/L).

Pregnant and lactating women (any age)

• Zinc deficiency (as defined by authors, depending on the age).

• Serum or plasma zinc (in µmol/L).

Adult males and females (19 years of age or older)

• Zinc deficiency (as defined by authors, depending on the age and sex).

• Serum or plasma zinc (in µmol/L).

Secondary outcomes

Secondary outcomes of interest may differ according to participant group; we have listed these accordingly.

All participants

• Diarrhoea (as defined by authors).

• Pneumonia (as defined by authors).

• All‐cause morbidity.

• Haemoglobin (in g/dL).

• Anaemia (as defined by authors, depending on age and gender).

• Adverse effect (iron status measured as serum ferritin in µg/L).

• Adverse effect (iron status measured as serum transferrin receptor in mg/L).

• Adverse effect (copper status as measured by serum or plasma copper level in µg/dL).

• Vomiting (as defined by authors).

• Any adverse effect reported (as defined by authors).

Children 24 to 59 months of age

• Weight (in kg).

• Height or length (in cm).

• Mid‐upper arm circumference (in cm).

• Cognitive and motor skill development (as assessed by trialists, including use of indexes such as the Bayley Mental Development Index (MDI), Bayley Psychomotor Development Index (PDI), Stanford‐Binet Test, and DENVER II Developmental Screening Test).

• All‐cause death.

Male and female adults

• Cognitive and work performance (as defined by authors).

Search methods for identification of studies

Electronic searches

We searched the following international and regional sources:

International databases

• Cochrane Central Register of Controlled Trials (CENTRAL) Issue 3 of 12 (2015): Date of search 9 April 2015

• MEDLINE & MEDLINE In Process (OVID) 1950 to 8 April 2015: Date of search 9 April 2015

• EMBASE (OVID) 1974 to 8 April 2015: Date of search 9 April 2015

• Web of Science (ISI) 1900 to 9 April 2015: Date of search 10 April 2015

• CINAHL (EBSCO) 1982 to May 2013: Date of search 10 April 2015

• POPLINE 1970 to April 2015: Date of search 12 April 2015

• AGRICOLA (http://agricola.nal.usda.gov/) to April 2015: Date of search 12 April 2015

• BIOSIS (ISI) 1969 to 9 April 2015: Date of search 10 April 2015

• OpenGrey (grey literature resource) to April 2015: Date of search 12 April 2015

• BiblioMap and TRoPHI (Trials Register of Promoting Health Interventions) to April 2015: Date of search 12 April 2015

Regional databases

• IBECS (http://ibecs.isciii.es/) to April 2015: Date of search 13 April 2015

• SciELO (http://www.scielo.br/) to April 2015: Date of search 13 April 2015

• Global Index Medicus ‐ AFRO (includes African Index Medicus); EMRO (includes Index Medicus for the Eastern Mediterranean Region) to April 2015: Date of search 17 April 2015

• LILACS to April 2015: Date of search 19 April 2015

• PAHO (Pan American Health Organization) Library to April 2015: Date of search 17 April 2015

• WHOLIS (WHO Library) to April 2015: Date of search 17 April 2015

• WPRO (includes Western Pacific Region Index Medicus) to April 2015: Date of search 17 April 2015

• IMSEAR (Index Medicus for the South‐East Asian Region) to April 2015: Date of search 17 April 2015

• IndMED (http://indmed.nic.in/) to April 2015: Date of search 17 April 2015

• Native Health Database (https://hscssl.unm.edu/nhd/) to April 2015: Date of search 18 April 2015

For theses we searched WorldCat, NDLTD (Networked Digital Library of Theses and Dissertations), DART‐Europe E‐theses Portal, ADT (Australasian Digital Theses) Program, Theses Canada Portal, and ProQuest Dissertations & Theses.

The search employed keyword and controlled vocabulary (when available), using the search terms set out. We have presented the search strategy for databases in Appendix 1. We did not apply language or date restrictions for any database.

We handsearched all the issues published in last 12 months of the five journals with the highest number of studies eligible for inclusion in this review after search of all above databases, to capture any article that may not have been indexed in the databases at the time of the search. We contacted authors of included studies and checked reference lists of included papers to identify additional records.

We searched the World Health Organization International Clinical Trials Registry Platform (ICTRP), which includes CTRI (Clinical Trials Registry ‐ India), for any ongoing or planned trials, and contacted authors of such studies to obtain further information or eligible data if available.

We commissioned translations of any articles written in a language other than English. If this was not possible, we sought advice from the Cochrane Public Health Group. We planned to store such articles in the 'Awaiting assessment' section of the review until a translation was available.

Searching other resources

For assistance in identifying ongoing or unpublished studies, we contacted the Department of Nutrition for Health and Development and the regional offices of the World Health Organization (WHO), as well as the nutrition section of the Centers for Disease Control and Prevention (CDC), the United Nations Children's Fund (UNICEF), the World Food Programme (WFP), the Micronutrient Initiative (MI), Global Alliance for Improved Nutrition (GAIN), Helen Keller International (HKI), World Vision, Sight and Life, PATH, premix producers DSM and BASF, Food Fortification Initiative (FFI), and the International Zinc Nutrition Consultative Group (IZiNCG).

We also reviewed the reference lists of identified articles and handsearched reviews and abstracts of international micronutrient conferences of the past three years. We contacted experts in the field to identify any additional or ongoing trials.

Data collection and analysis

Selection of studies

Two review authors (DS and TG) independently screened all citations and abstracts retrieved by the search strategy to identify potentially eligible trials. We obtained the full text of potentially eligible trials and two review authors (DS and TG) assessed the full text for inclusion in the review. We resolved any differences in opinion by simultaneous review, or if necessary by discussion with a third review author (LMD). We excluded studies that did not meet the eligibility criteria and documented the reasons for their exclusion in the Characteristics of excluded studies section.

Data extraction and management

We removed duplicates from the retrieved records, and if multiple reports of the same study were available, we consolidated these so that the unit of interest was the study. Two review authors (DS and TG) independently extracted all data using a tailored and pretested data extraction form. We extracted data on study design, context, participant characteristics, interventions (including costs and process data where available), outcomes, source of funding, and PROGRESS framework characteristics (Campbell and Cochrane Equity Methods Group Equity Checklist items). PROGRESS is an acronym for Place of Residence, Race/Ethnicity, Occupation, Gender, Religion, Education, Socioeconomic Status, and Social Capital, and Plus represents additional categories such as Age, Disability, and Sexual Orientation (Campbell and Cochrane Equity Methods Group Equity Checklist items).

We considered the last recorded time point for the evaluated outcome measurements while the intervention was still operative or had just been terminated. If similar outcomes (for example zinc deficiency) had been reported using multiple measures, we averaged them. One review author entered data into Review Manager software (RevMan 2014), and a second review author carried out checks for accuracy. We resolved any discrepancies between the extracted data by discussion and, if required, referral to a third review author (LMD). If data were unclear or not presented in the study papers, we attempted to contact the study authors for further details.

Assessment of risk of bias in included studies

We assessed the quality and risk of bias in each included study using the criteria outlined in the Cochrane Handbook for Systematic Reviews of Interventions (Higgins 2011). For randomised controlled trials, non‐randomised controlled trials, and controlled before‐after studies these criteria included: (i) sequence generation (selection bias), (ii) allocation sequence concealment (selection bias), (iii) blinding of participants and personnel (performance bias), (iv) blinding of outcome assessment (detection bias), (v) incomplete outcome data (attrition bias), (vii) selective outcome reporting (reporting bias), (viii) comparability of baseline outcome and characteristics, (ix) protection from contamination, and (x) other potential sources of bias.

For cluster‐randomised trials we considered the following biases: (i) recruitment bias, (ii) baseline imbalance, (iii) loss of clusters, (iv) incorrect analysis, and (v) comparability with individually randomised trials (Higgins 2011). For each entry we assessed the risk of bias as ‘low risk’, ‘high risk', or ‘unclear risk’, with the last category indicating either lack of information or uncertainty over the potential for bias. We created plots of ‘Risk of bias’ assessments in Review Manager (RevMan 2014). Two review authors (DS and TG) independently assessed risk of bias, resolving any disagreements by mutual or group discussion, if required. The risk of bias assessment provided essential input for data synthesis in the 'Summary of findings' table.

Measures of treatment effect

For two‐group comparisons, we gave preference to pooling risk ratios (RR) or comparable measures including hazard ratios and odds ratios. For computing the summary risk ratio, we used individual study risk measures and 95% confidence intervals (CI) or standard error (SE).

We gave preference to the risk measure stated by authors, in a hierarchical fashion, with a recheck of the calculations from the stated numbers, if possible.  If RR was not stated, we computed with the following preference order for the denominator: (i) numbers with definite outcome known (complete follow‐up) until completion of intervention period, or (ii) number randomised.

For continuous data, we used weighted mean difference (WMD) when outcomes were measured in the same way between trials. We used the standardised mean difference (SMD) to combine trials that measured the same outcome with different methods.

Unit of analysis issues

For cluster‐randomised trials, we used the stated cluster‐adjusted RR and 95% CI, irrespective of the method employed in their calculation. If the analyses had not been adjusted for cluster‐design effect, we used the intracluster correlation coefficient (ICC) derived from the trial (if available), or from another source (for example using the ICCs derived from other, similar trials) and then calculated the design effect as recommended (Higgins 2011). We then recalculated the risk measures using a design effect inflation of SE from these external sources (Higgins 2011).

In factorial trials and in multi‐arm designs yielding two or more intervention groups (different dosage or administration regimens) and a single control group, we pooled and compared the data in the intervention groups, including the variation in the intervention characteristic, against the single control group to prevent unit‐of‐analysis error.

Dealing with missing data

We contacted the authors of relevant trials when information about any outcome was unclear or had not been fully reported.

We preferred to use intention‐to‐treat analyses (author reported) when a per‐protocol analysis had also been reported. If the author had not reported an intention‐to‐treat analysis, we used the reported analysis, and judged the study to be at risk of bias due to these criteria.

When assessing adverse events, adhering to the principle of 'intention‐to‐treat' may be misleading, therefore we attempted to relate the results to the treatment received ('per protocol' or 'as observed'). This means that for side effects, we attempted to base the analyses on the participants who actually received the intervention and the number of adverse events that were reported in the studies.

Assessment of heterogeneity

We intended to assess contextual heterogeneity on the basis of information collected on the context in which the intervention was implemented. We assessed for variability in the participants, interventions, and outcomes studied to identify clinical heterogeneity, and for variability in study design to describe methodological diversity. We checked and measured for statistical heterogeneity using I² and Chi² as recommended by the Cochrane Handbook for Systematic Reviews of Interventions (Section 9.5.2) (Higgins 2011). A rough guide used for interpretation was:

• 0% to 40%: might not be important;

• 30% to 60%: may represent moderate heterogeneity;

• 50% to 90%: may represent substantial heterogeneity;

• 75% to 100%: considerable heterogeneity.

In anticipation of few included trials, we used a P value of 0.10 from the Chi² test to determine statistical significance with regard to heterogeneity.

If we identified moderate, substantial, or considerable heterogeneity, we explored it by prespecified subgroup effect analyses.

Assessment of reporting biases

If sufficient numbers of trials were available, we planned to evaluate the presence of reporting bias in the extracted data visually using the funnel plot. If 10 or more trials were available per outcome, we also planned to use formal statistical tests for funnel plot asymmetry, namely Begg’s and Egger’s with the user‐written 'metabias' command in Stata® (version 9) software (Steichen 1998; Sterne 2001).

Data synthesis

We carried out meta‐analysis with Review Manager software to provide an overall estimate of treatment effect when more than one study examined the same intervention (RevMan 2014), provided that studies used similar methods and measured the same outcome in similar ways in similar populations.

In anticipation of significant contextual heterogeneity between studies, we used the random‐effects model for pooling data. We calculated pooled estimates of RR with 95% CIs using the generic inverse variance method. For continuous variables, we used the inverse variance method, and for dichotomous variables we used the method proposed by Mantel‐Haenszel (Higgins 2011). We used the SMD to combine trials that measured the same outcome with different methods. If we found too few studies, or the studies could not be pooled, we summarised the results related to outcomes in a narrative form.

We have presented the main results as a 'Summary of findings' table for important outcomes (listed primary outcomes and adverse effects) and incorporated the GRADE assessment according to recommendations (Balshem 2011; GRADE 2013). The factors included in the GRADE assessment that can decrease the quality of evidence are limitation in study design or execution (risk of bias), inconsistency of results, indirectness, imprecision, and publication bias. The factors that can increase the quality of evidence (for observational studies only) are large magnitude of effect, dose‐response gradient, and effect of plausible residual confounding (GRADE 2013).

The clinical relevance and importance of statistically significant results is determined from the 'Summary of findings' table, giving due consideration to the quality of evidence and functional outcomes in addition to biochemical outcomes.

Subgroup analysis and investigation of heterogeneity

When a sufficient number of trials were available, we explored significant heterogeneity (I² greater than 30%) and performed subgroup analyses only for the primary outcomes zinc deficiency and serum zinc, as a hypothesis‐generating exercise. The prespecified subgroup analyses included analysis by:

• sex: males versus females, versus mixed;

• duration of intervention: less than six months; six months to one year; more than one year;

• type of food vehicle: oils and fats versus sugar versus wheat flour or wheat flour subproducts versus maize flour and cornmeals or maize flour subproducts versus rice or rice subproducts, versus condiments/seasonings, versus milk/dairy products, versus fruit juices/nectars, versus others;

• type of zinc compound: zinc sulphate versus zinc oxide, versus other forms of zinc, versus unknown/unreported;

• dose of zinc added per 100 g of food;

• development status of country: low‐ and middle‐income countries versus others (World Bank 2012).

We limited subgroup analysis to primary outcomes for which three or more trials contributed data.

We examined differences between subgroups by visual inspection of the subgroups’ CIs; non‐overlapping CIs suggest a statistically significant difference in treatment effect between the subgroups. When there were two or more subgroups, we conducted a formal statistical analysis, as per the method in Borenstein 2008 incorporated in Review Manager (RevMan 2014).

If a sufficient number of trials (equal to or greater than 10) was available, we planned to explore the contribution of these variables to heterogeneity by meta‐regression using the 'metareg' command in Stata® (version 9.0) software with the restricted maximum likelihood option (Sharp 1998).

We also used narrative synthesis, guided by the data extraction form, in terms of the ways in which studies were grouped and summarised in this review to explore intervention implementation (using information about resource use and findings from process evaluations) and to describe the impact of interventions by socio‐demographic characteristics known to be important from an equity perspective based on the PROGRESS framework, where this information was available.

Sensitivity analysis

We conducted a sensitivity analysis taking into account the trial quality components (allocation concealment, blinding, and attrition), with each of these domains being considered separately (low risk versus others). We conducted sensitivity analyses for the primary outcomes, provided three or more trials contributed data to the outcome.

Results

Description of studies

Results of the search

We identified 6582 references (after removing duplicates) for possible inclusion, 6337 through international databases and the remaining through regional databases and other resources. We have described the study selection process in Figure 2 in the form of a PRISMA flowchart. We included nine reports from eight trials (Aaron 2011; Badii 2012; Haibin 2001; Hambidge 1979; Hettiarachchi 2004; Kilic 1998; Lopez de Romaña 2005; Sanchez 2014), and excluded 41 references (39 studies) after reading the full text. One included trial was translated into English from the Chinese language (Haibin 2001), and one was translated into English from the Spanish language (Sanchez 2014). No trials are awaiting assessment, and one study is still ongoing or unpublished (Moretti 2014).

2.

Study flow diagram.

Included studies

We have presented the details of included studies, including participants, intervention, outcomes, source of funding, and results of contact with the authors, in the Characteristics of included studies section. We have also provided a summary of the general characteristics of the included studies in Table 3.

1. Summary of characteristics of included studies.

Study and Year	Participants	Type of food vehicle	Zinc salt and dose of elemental zinc	Duration of intervention	Development status of country
Comparison 1 (foods fortified with zinc alone versus same foods without zinc)
Badii 2012	80 zinc‐deficient women (age 19 to 49 y) with low serum zinc (≤ 70 µg/dL)	Taftoon bread prepared from wheat flour, salt, and yeast.	Zinc sulphate Group 1: unfortified bread Group 2: bread containing fortified flour 50 mg zinc/100 g of flour, providing mean zinc content of 5.72 mg and 10.1 mg elemental zinc per 100 g of bread	1 month	Upper middle income (Iran)
Hambidge 1979	96 healthy young children (age 33 to 90 mo; mean age 58 mo; 56 boys and 40 girls) from a private preschool and kindergarten	Ready‐to‐eat breakfast cereals	Zinc oxide Group 1: 3.75 mg of zinc per ounce (13.23 mg/100 g) serving of cereals Group 2: non‐zinc‐fortified cereals	9 months	High income (United States of America)
Kilic 1998	24 schoolchildren (age 7 to 11 y; 10 boys and 14 girls) with serum zinc concentrations below 65 µg/dL	Bread prepared from wheat flour, salt, and yeast	Zinc acetate providing 400 mg of elemental zinc per loaf of bread Group 1: zinc‐fortified bread providing 2 mg/kg/day elemental zinc Group 2: unfortified bread	3 months	Upper middle income (Turkey)
Sanchez 2014	301 healthy children (age 2 to 5 y) from child day‐care centres who received more than 80% of their daily food intake from the childcare, and liked milk	Milk	Zinc sulphate and zinc amino chelate Group 1: milk fortified with elemental zinc (as zinc sulphate) Group 2: milk fortified with elemental zinc (as zinc amino chelate) Group 3: unfortified milk The dose of elemental zinc varied according to the age of the participants. Children 2 to 3 y received 7 mg zinc daily, and children 4 to 5 y received 9.45 mg elemental zinc	4 months (16 weeks)	Upper middle income (Colombia)
Comparison 2 (foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc)
Aaron 2011	130 healthy men (age ≥ 18 y; mean age 24.8 y)	Bread prepared from wheat flour, salt, yeast, and baking enzyme additive	Zinc oxide Group 1: bread fortified with iron and folic acid but not zinc, and 10 mL liquid multivitamin supplement between meals *Group 2: bread fortified with iron and folic acid but not zinc, and the same 10 mL multivitamin supplement with 15 mg zinc added Group 3: bread fortified with iron, folic acid, and zinc (3.75 mg elemental zinc/100 g), and the same 10 mL multivitamin supplement without zinc Group 4: bread fortified with iron, folic acid, and zinc (7.5 mg elemental zinc/100 g), and the same 10 mL multivitamin supplement without zinc	1 month (4 weeks)	Lower middle income (Senegal)
Haibin 2001	313 healthy women with their first pregnancy (mean age 24.7 y)	Biscuits	*Group 1: 3 biscuits per day; 8 g each containing 400 IU vitamin D Group 2: 3 biscuits per day; 8 g each containing 400 IU vitamin D and 400 mg carbonate calcium Group 3: 3 biscuits per day; 8 g each containing 400 IU vitamin D, 400 mg carbonate calcium, and 10 mg lactate zinc Group 4: 3 biscuits per day; 8 g each containing 400 IU vitamin D, 400 mg carbonate calcium, 10 mg ferrous lactate, and 50 mg vitamin C Group 5: 3 biscuits per day; 8 g each containing 400 IU vitamin D, 400 mg carbonate calcium, 10 mg ferrous lactate, 50 mg vitamin C, and 10 mg lactate zinc	6 months (24 weeks)	Upper middle income (China)
Hettiarachchi 2004	53 schoolchildren (age 6 to 10 y; 25 boys and 28 girls)	Local dish called ‘halapa’, prepared from rice flour, grated coconut, sugar, and salt	Zinc oxide Group 1: rice flour fortified with 60 mg elemental iron/kg (as ferrous sulphate) and folic acid (2 mg/kg) Group 2: rice flour fortified with 60 mg elemental iron/kg (as ferrous sulphate), 385.08 mg/kg of Na2EDTA in dry powder form, and folic acid (2 mg/kg) Group 3: rice flour fortified with 60 mg elemental iron/kg (as ferrous sulphate), folic acid (2 mg/kg), and 60 mg elemental zinc/kg (as zinc oxide powder) Group 4: rice flour fortified with 60 mg elemental iron/kg (as ferrous sulphate), 385.08 mg/kg of Na2EDTA in dry powder form, folic acid (2 mg/kg), and 60 mg elemental zinc/kg (as zinc oxide powder)	1 month (4 weeks)	Lower middle income (Sri Lanka)
Lopez de Romaña 2005	58 stunted, moderately anaemic children (age 3 to 4 y) residing in a poor community	Wheat products (biscuits, noodles)	Zinc sulphate Group 1: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and no zinc Group 2: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and 3 mg elemental zinc per 100 g flour Group 3: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and 9 mg elemental zinc per 100 g flour	2 months (70 days)	Upper middle income (Peru)

*These groups not eligible for inclusion in current review.

Study designs

Four of the eight included studies compared zinc‐fortified foods versus same food without zinc (comparison 1) (Badii 2012; Hambidge 1979; Kilic 1998; Sanchez 2014). Of these, three were controlled trials at the level of the individual (with variable description of randomisation and blinding), whereas one was a cluster‐randomised trial (Sanchez 2014). One of these trials had three arms (one unfortified food and two fortified food with different zinc content) (Badii 2012), while the other studies compared two arms (zinc‐fortified versus unfortified food).

Four of the eight included studies provided data comparing zinc fortification in combination with other micronutrients versus the same food with other micronutrient(s) but no added zinc (comparison 2) (Aaron 2011; Haibin 2001; Hettiarachchi 2004; Lopez de Romaña 2005). All of these studies were controlled trials at the level of the individual, and had multiple arms. The trial by Lopez de Romaña 2005 had three groups; each received daily, at breakfast and lunch, 100 g wheat products fortified with 3 mg iron (as ferrous sulphate) and 0, 3, or 9 mg zinc (as zinc sulphate) per 100 g flour.

The trial by Aaron 2011 had four groups: (i) 200 g per day of wheat bread fortified with iron and folic acid but no zinc, and a liquid multivitamin supplement without zinc between meals; (ii) the same bread but 15 mg zinc added to multivitamin supplement; (iii) bread fortified with 7.5 mg zinc in addition to iron and folic acid, and same multivitamin supplement without zinc; or (iv) bread fortified with 15 mg zinc in addition to iron and folic acid, and the same multivitamin supplement without zinc. Three of these four groups provided data for this review. A trial among Sri Lankan schoolchildren also had four groups that consumed a local dish prepared with 25 g of fortified rice flour labelled with one of the following: (i) ferrous sulphate; (ii) ferrous sulphate plus Na₂EDTA; (iii) ferrous sulphate plus zinc oxide; or (iv) ferrous sulphate plus Na₂EDTA plus zinc oxide (Hettiarachchi 2004).

The trial among pregnant women in China had five groups: biscuits fortified with (i) vitamin D; (ii) vitamin D and calcium; (iii) vitamin D, calcium, and zinc; (iv) vitamin D, calcium, iron, and vitamin C; and (v) vitamin D, calcium, iron, zinc, and vitamin C (Haibin 2001). The last four arms of this study provided data for this review.

No study compared zinc‐fortified foods versus no intervention (comparison 3).

We did not find any observational study satisfying the inclusion criteria, for any of the comparisons.

Participants

Three studies were done in children (n = 455) between the ages of 24 months and 59 months (Hambidge 1979; Lopez de Romaña 2005; Sanchez 2014); two of these enrolled apparently healthy children, whereas one included stunted and moderately anaemic children (Lopez de Romaña 2005). Two studies were conducted in older children (5 to 11.9 years of age; n = 77) (Hettiarachchi 2004; Kilic 1998); one of these selected participants after diagnosing them as zinc‐deficient (Kilic 1998). One study was conducted in apparently healthy pregnant women (n = 313) (Haibin 2001). Two studies enrolled adult participants (n = 224) (Aaron 2011; Badii 2012); one of these selected participants after diagnosing them as zinc‐deficient (Badii 2012).

Intervention

Seven included trials used cereal‐based foods (bread in three trials: Aaron 2011, Badii 2012, and Kilic 1998; biscuits in one trial: Haibin 2001; breakfast cereals in one trial: Hambidge 1979; rice flour in one trial: Hettiarachchi 2004; and wheat products in one trial: Lopez de Romaña 2005) as vehicles for zinc‐fortification; one trial used milk fortified with zinc (Sanchez 2014). Zinc salts used for fortification were zinc oxide in three studies (Aaron 2011; Hambidge 1979; Hettiarachchi 2004), zinc sulphate in three studies (Badii 2012; Lopez de Romaña 2005, Sanchez 2014), zinc acetate in one study (Kilic 1998), zinc lactate in one study (Haibin 2001), and zinc amino chelate in one study (Sanchez 2014). The dose of elemental zinc ranged from 3 mg to 40 mg per 100 g of food. See other details in the Characteristics of included studies table and Table 3 (Summary of characteristics of included studies).

Settings

Table 4 compares the PROGRESS‐Plus parameters of equity in the included trials. Seven of the included studies were carried out in low‐ and middle‐income countries, where zinc deficiency is a public health problem, whereas one study was conducted in the USA. Of the seven studies from low‐ and middle‐income countries, two were carried out in lower‐middle‐income countries (Senegal and Sri Lanka), and five were carried out in upper‐middle‐income countries (China, Colombia, Iran, Peru, and Turkey).

2. PROGRESS‐Plus equity checklist of included studies.

Study	Place	Race/Ethnicity	Occupation	Gender	Religion/ culture/education	Socio‐economic status	Social status	Others/ disability/ age/ sexual orientation	Overall PROGRESS‐Plus
Aaron 2011	Dakar, Senegal (lower middle income)	Senegalese (no more details provided)	Not available	Male	Not available	Low income Quote: “in a community clinic based in a low‐income urban neighbourhood”	Not available	Apparently healthy Age > 18 y No details on sexual orientation	The study enrolled apparently healthy adult males reporting to a feeding centre in low‐income urban area of a lower‐middle‐income country in Africa
Badii 2012	Isfahan (a large city located 340 km south of Tehran), Iran (upper middle income)	No details	Staff and students of university	Female	No details for religion and culture / university staff or students	Not available	Not available	Healthy non‐pregnant, non‐lactating women during follicular phase of their menstrual cycle. No details about sexual orientation	The study enrolled healthy non‐pregnant, non‐lactating women teaching/studying in a technical university of a major city of an upper‐middle‐income country of Middle East (Asia)
Haibin 2001	Xinxiang city, Henan Province, China (upper middle income)	No details	No details	Female	No details	No details	No details	Pregnant women seeking antenatal care at the hospital	The study enrolled pregnant women seeking care at the antenatal clinic of a hospital in a major city of an upper‐middle‐income country of Asia
Hambidge 1979	Denver, USA (high income)	No details	Preschool and kindergarten children	Males and females	Not available	Middle‐income families Quote: "Though details of individual family income were unknown, the population was essentially derived from middle‐income families"	Not available	Healthy children enrolled in private preschools	The study was conducted amongst preschool children from middle‐income families from a city in a high‐income country (USA)
Hettiarachchi 2004	Galle (a major city situated 119 km from Colombo), Sri Lanka (lower middle income)	Sinhalese (no more details provided)	Children (7 to 10 y)	Males and females	Not available	Not available	Not available	Apparently healthy children (7 to 10 y) recruited through public advertising	The study enrolled apparently healthy children (7 to 10 y) recruited through public advertising in an urban area of a lower‐middle‐income country in South Asia
Kilic 1998	An elementary school in Ankara, the capital city of Turkey (upper middle income)	Turkish (no more details provided)	Children (7 to 11 y)	Males and females	No details for religion and culture / schoolchildren	Average Quote: “their socio‐economic levels were average”	Not available	24  zinc‐deficient (S. zinc < 65 µg/dL) children were selected from 101 children	The study selected  zinc‐deficient schoolchildren from an elementary school in an urban area of a upper‐middle‐income country in Europe
Lopez de Romaña 2005	Periphery of Lima, Peru (upper middle Income), considered to be at high risk of zinc deficiency	Peruvian (no more details provided)	Children (3 to 4 y)	Males and females	Not available	Residing in poor community	Not available	Stunted and moderately anaemic (Hb 9 to 11 g/L) children selected after screening 765 children (3 to 4 y)	The study selected an urban poor population from an upper‐middle‐income country of South America. Children at high risk of zinc deficiency were selected from within this population for inclusion in the study
Sanchez 2014	Medellin, Colombia	No details	Children (2 to 5 y), beneficiaries of child care	Males and females	No details for religion and culture / preschool children	51.5% in lower socio‐economic strata	61.8% is subsidised	Children 2 to 5 years of age attending healthcare centres affiliated with Fundación de Atención a la Niñez (FAN)	The study selected children between 2 and 5 years of age from 6 child day‐care centres in Medellin, Colombia, who attended 8 hours per day and received more than 80% of their meals in the centre, and who liked milk

Hb: haemoglobin

Four of the studies described the included population as zinc‐deficient or having high risk of zinc deficiency. Most studies did not provide any information on ethnicity, religion, education, social status, or occupation of participants or their parents (in case of children). Two studies identified participants as being from low‐income or poor communities, whereas another two studies reported that their participants belonged to middle‐ or average‐income families. Most studies enrolled apparently healthy participants.

Outcomes

Of the eight included studies, seven provided data on serum/plasma zinc concentrations after a variable period of intervention. The only cluster‐randomised trial included in this review did not contribute data to the outcome (Sanchez 2014).

In comparison 1 (foods fortified with zinc alone versus same foods without zinc), all three studies contributing data to the outcome of serum/plasma zinc evaluated this as the main outcome (Badii 2012; Hambidge 1979; Kilic 1998). No trial in this comparison provided information about frequency of zinc deficiency in the participants after intervention. Two trials (Hambidge 1979; Sanchez 2014), both in comparison 1, provided information about stunting and underweight in children. One trial provided data on haemoglobin and serum ferritin (Kilic 1998), and no study in this comparison provided information about the prevalence of anaemia in study participants after they received fortified foods. Two trials provided serum/plasma copper levels after intervention (Hambidge 1979; Kilic 1998). Only one trial provided information related to individual morbidities such as pneumonia and diarrhoea (Sanchez 2014), whereas one trial provided information on all‐cause infectious morbidity (Kilic 1998). This was reported in another publication (Saldamli 1996) from the results of this study (Kilic 1998).

All four studies in comparison 2 (foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc) provided data for the outcome serum/plasma zinc. Of these four studies, only one study evaluated plasma zinc as the main outcome (Aaron 2011). The other three studies were mainly designed to evaluate fractional absorption of iron/zinc, in Hettiarachchi 2004 and Lopez de Romaña 2005, or prevalence of anaemia, in Haibin 2001.

Only one study provided data about frequency of zinc deficiency after intervention (Lopez de Romaña 2005). No trial in this comparison provided information about stunting and underweight in children, whereas one trial provided the weight and height of participants without data relating to stunting or overweight (Lopez de Romaña 2005). Three studies provided data on haemoglobin and serum ferritin (Haibin 2001; Hettiarachchi 2004; Lopez de Romaña 2005), but only two of these provided information about prevalence of anaemia in study participants after receiving fortified foods (Haibin 2001; Lopez de Romaña 2005). No study in this comparison provided information related to morbidities such as pneumonia and diarrhoea, or any adverse effect of intervention.

Funding

Most included studies were funded by government sector, academic bodies, or independent non‐government organisations. In three studies, fortified food or fortificants were provided/donated by the industrial sector. The trial by Aaron 2011 was funded by Global Alliance for Improved Nutrition (Geneva, Switzerland) and the Michael and Susan Dell Foundation (Austin, USA), but the fortificants were donated by DSM Nutritional Products and breads were prepared by Les Grands Moulins de Dakar. In a trial from the USA (Hambidge 1979), Kellogg Company provided the ready‐to‐eat cereals, although the study was sponsored by the National Institute of Arthritis, Metabolic and Digestive Diseases and General Clinical Research Centers Program of the Division of Research Resources, National Institutes of Health, USA. In the study by Hettiarachchi 2004, the food‐grade minerals were supplied free of charge through Percy Ranasinghe of the Greenfields International, Sri Lanka. The provider of ferrous sulphate and zinc oxide was Paul Lohmann, Germany. The iron‐EDTA was provided by AkzoNobel, Netherlands and folic acid by Glaxo Wellcome. In a study from Ankara, Turkey (Kilic 1998), the breads were made by the Food Engineering Department of Hacettepe University, Ankara, Turkey. Three members of this department also contributed to authorship. The funding details were not clear in one trial (Haibin 2001).

Excluded studies

We excluded 41 full‐text articles providing information about 39 studies. We excluded 21 studies that used other micronutrients along with zinc for fortification, and zinc was not the only difference between intervention and control groups. We excluded four studies that provided information about fortification of milk formula or complementary feeds for use in children younger than two years. We excluded four studies that reported only outcomes related to absorption or acceptability, or both. We excluded five studies in which zinc was used as a medicine that was given along with food or mixed in food at point‐of‐use. We excluded one study that was of a non‐randomised pretest, post‐test cluster design with two intervention and one control clusters (Ohiokpehai 2009).

One excluded study, Haibin 2001b, that provided outcomes related to placental weight, cord blood nutrients, and birthweight in relation to antenatal fortification with zinc in combination with other nutrients was actually another report from the included trial Haibin 2001. We excluded one study that was reported as a cross‐over design, but there was no randomisation, and comparison for morbidity was done with a group who refused to participate in the trial. We excluded two studies (both unpublished) in which water was used as vehicle for delivery of zinc. We have provided detailed description of the studies and the reasons for exclusion in the Characteristics of excluded studies table. We have provided details of an ongoing study in the Characteristics of ongoing studies table.

Risk of bias in included studies

The Characteristics of included studies section along with Figure 3 and Figure 4 present risk of bias for each included trial as well as an overall summary of the risk of bias.

3.

Risk of bias graph: review authors' judgements about each risk of bias item presented as percentages across all included studies.

4.

Risk of bias summary: review authors' judgements about each risk of bias item for each included study.

Allocation

Three studies adequately described sequence generation (Aaron 2011; Lopez de Romaña 2005; Sanchez 2014), and only two studies adequately described allocation concealment (Lopez de Romaña 2005; Sanchez 2014). Three and five studies were at unclear risk of bias for random sequence generation and allocation concealment, respectively. Two studies were at high risk of bias for sequence generation (Haibin 2001; Hambidge 1979), whereas one study was at high risk of bias for allocation concealment, as allotment was done on the basis of order of visit to hospital (Haibin 2001).

Blinding

Five trials reported blinding of participants, personnel, and outcome assessors. Two trials did not provide any details about blinding and were therefore classified as unclear risk for this domain (Badii 2012; Hettiarachchi 2004); one of these mentioned "double‐blind" in the title of the study with no subsequent details in the full text (Badii 2012). We classified one trial that allocated participants based on their order of visit to hospital as at high risk of bias for blinding of participants and investigators (Haibin 2001).

Incomplete outcome data

We assessed three trials as having a high attrition (more than one‐third of participants, without providing reasons) for the primary outcome (Haibin 2001; Hambidge 1979; Lopez de Romaña 2005). The other five studies had a low risk of attrition bias.

Selective reporting

There was no indication of selective reporting by any of the studies from published reports, however we did not have the access to the study protocols. Only one trial provided a trial registration number for the protocol (Sanchez 2014).

Other potential sources of bias

We could not identify any other potential source of bias in the included studies. Two studies were industry sponsored (manufacturer of breakfast cereals and manufacturer of micronutrient mixes, respectively) (Hambidge 1979; Sanchez 2014). The cluster randomised trial included in this review was at low risk of bias for recruitment of clusters, baseline imbalance, or loss of clusters (Sanchez 2014). However, we rated the risk of bias for incorrect analysis as unclear as the authors provided no information about adjustment for clusters in the analyses. Similarly, we rated the risk of bias as unclear for comparability with individual randomised controlled trials as no other trial provided information on outcomes reported in this study (Sanchez 2014).

Effects of interventions

See: Table 1; Table 2

Summary of findings for the main comparison. Foods fortified with zinc alone compared to same foods without adding zinc for improving zinc status and other health outcomes in the general population.

Foods fortified with zinc alone compared to same foods without adding zinc for improving zinc status and other health outcomes in the general population
Patient or population: General population (over 2 years of age, including pregnant and lactating women) Settings: Denver, USA; Isfahan, Iran; Ankara, Turkey; Medellin, Colombia Intervention: Foods fortified with zinc alone Comparison: Same foods without adding zinc
Outcomes	Illustrative comparative risks* (95% CI)		Relative effect (95% CI)	No of Participants (studies)	Quality of the evidence (GRADE)	Comments
	Assumed risk	Corresponding risk
	Same foods without adding zinc	Foods fortified with zinc alone
Serum or plasma zinc (in µmol/L)	The mean serum zinc level ranged across control groups from 9.6 to 10.9	The mean serum or plasma zinc (in µmol/L) in the intervention groups was 2.12 higher (1.25 to 3 higher)	‐	158 (3 studies)	⊕⊕⊝⊝ low1,2	Fortification of foods with zinc may improve the serum zinc levels of populations (low‐quality evidence)
Stunting (author defined)	Study population		RR 0.88 (0.36 to 2.13)	397 (2 studies)	⊕⊕⊝⊝ low3,4	Fortification of foods with zinc may make little or no difference to the incidence stunting in children (low‐quality evidence)
	76 per 1000	67 per 1000 (28 to 163)
	Moderate
	78 per 1000	69 per 1000 (28 to 166)
Underweight (author defined)	Study population		RR 3.1 (0.52 to 18.38)	397 (2 studies)	⊕⊕⊝⊝ low3,4	Fortification of foods with zinc may make little or no difference to the incidence of underweight in children (low‐quality evidence)
	7 per 1000	22 per 1000 (4 to 128)
	Moderate
	10 per 1000	31 per 1000 (5 to 184)
*The basis for the assumed risk (e.g. the median control group risk across studies) is provided in footnotes. The corresponding risk (and its 95% confidence interval) is based on the assumed risk in the comparison group and the relative effect of the intervention (and its 95% CI). CI: confidence interval; RR: risk ratio
GRADE Working Group grades of evidence High quality: Further research is very unlikely to change our confidence in the estimate of effect. Moderate quality: Further research is likely to have an important impact on our confidence in the estimate of effect and may change the estimate. Low quality: Further research is very likely to have an important impact on our confidence in the estimate of effect and is likely to change the estimate. Very low quality: We are very uncertain about the estimate.

¹Downgraded by one for serious risk of bias: Of the three included trials, two were at low to unclear risk of bias. One trial, Haibin 2001, had a high risk of selection, attrition, performance and detection bias.
 ²Downgraded by one for imprecision: The total population size is 158, which is less than the threshold rule‐of‐thumb value of 400 of optimal information size for downgrading for imprecision for continuous outcomes.
 ³Downgraded by one for serious risk of bias: The included trial, Hambidge 1979, was at unclear risk for selection bias and high risk for attrition bias (96 participants were enrolled, 93 completed the study; results available for only 59 at the end of study).
 ⁴Downgraded by one for imprecision for the wide 95% confidence interval around the estimate of effect that includes both no effect and appreciable benefit or harm.

Summary of findings 2. Foods fortified with zinc plus other micronutrients compared to foods fortified with other micronutrients without zinc for improving zinc status and other health outcomes in the general population.

Foods fortified with zinc plus other micronutrients compared to foods fortified with other micronutrients without zinc for improving zinc status and other health outcomes in the general population
Patient or population: General population (over 2 years of age, including pregnant and lactating women) Settings: Lima, Peru; Dakar, Senegal; Xinxiang city, China; Galle, Sri Lanka Intervention: Foods fortified with zinc plus other micronutrients Comparison: Foods fortified with other micronutrients without zinc
Outcomes	Illustrative comparative risks* (95% CI)		Relative effect (95% CI)	No of Participants (studies)	Quality of the evidence (GRADE)	Comments
	Assumed risk	Corresponding risk
	Foods fortified with other micronutrients without zinc	Foods fortified with zinc plus other micronutrients
Zinc deficiency (as defined by authors, depending on age and gender)	Study population		RR 0.17 (0.01 to 3.94)	30 (1 study)	⊕⊝⊝⊝ very low1,2,3	It is uncertain whether fortification of foods with zinc in addition to other micronutrients leads to any change in incidence of zinc deficiency
	100 per 1000	17 per 1000 (1 to 394)
	Moderate
	100 per 1000	17 per 1000 (1 to 394)
Serum or plasma zinc (in µmol/L)	The mean serum zinc level ranged across control groups from 10.7 to 12.6	The mean serum or plasma zinc (in µmol/L) in the intervention groups was 0.03 higher (0.67 lower to 0.72 higher)	‐	250 (4 studies)	⊕⊕⊝⊝ low4,5	It may make little or no difference to serum zinc levels when foods are cofortified with zinc in addition to other micronutrients (low‐quality evidence)
*The basis for the assumed risk (e.g. the median control group risk across studies) is provided in footnotes. The corresponding risk (and its 95% confidence interval) is based on the assumed risk in the comparison group and the relative effect of the intervention (and its 95% CI). CI: confidence interval; RR: risk ratio
GRADE Working Group grades of evidence High quality: Further research is very unlikely to change our confidence in the estimate of effect. Moderate quality: Further research is likely to have an important impact on our confidence in the estimate of effect and may change the estimate. Low quality: Further research is very likely to have an important impact on our confidence in the estimate of effect and is likely to change the estimate. Very low quality: We are very uncertain about the estimate.

¹Downgraded by one for serious risk of bias: The included trial was at unclear risk of selection, performance and detection bias and high risk of attrition bias (final results available for only 34 out of 58 enrolled).
 ²Downgraded by one for indirectness: The study population is stunted, moderately anaemic children 3 to 4 years of age from Peru. These findings are not easily generalised to the entire general population in the same or other settings.
 ³Downgraded by one for imprecision: There is only one included trial, with the total sample for this outcome being 30, which is less than the optimal information size; and the wide 95% confidence interval around the estimate of effect that includes both no effect and appreciable benefit.
 ⁴Downgraded by one for serious risk of bias: Of the four included trials, two were at low to unclear risk of bias. One trial, Haibin 2001, was at high risk of selection, attrition, performance and detection bias, while one trial, Lopez de Romaña 2005, was at high risk for attrition bias.
 ⁵Downgraded by one for imprecision: The total population size is 250, which is less than the threshold rule‐of‐thumb value of 400 of optimal information size for downgrading for imprecision for continuous outcomes.

See: Table 1 and Table 2

We included eight trials in this review; four compared effect of zinc‐fortified foods with unfortified food (comparison 1), and four compared zinc‐fortified food in combination with other nutrients/factors with the same food containing other nutrients/factors without zinc (comparison 2). We found no study that compared effect of zinc‐fortified food with no intervention (comparison 3). We have organised the summary results by comparison, and by primary and secondary outcomes. As we did not have sufficient number of trials in each age group, we have not presented age‐wise categorisation of effects, as listed in Types of outcome measures. Many prespecified outcomes in this review were not assessed by any of the included trials. As all results showed significant heterogeneity that could not be explained by standard sensitivity analyses, including quality assessment, we analysed the results using a random‐effects model.

See the Data and analyses section for detailed results on the primary and secondary outcomes.

Comparison 1 (zinc‐fortified foods versus unfortified foods)

Four studies provided data for this comparison (Badii 2012; Hambidge 1979; Kilic 1998; Sanchez 2014). Two studies were performed in apparently healthy children between the ages of 24 months and 59 months (Hambidge 1979; Sanchez 2014). One study was conducted in older children (5 to 11.9 years of age) who were known to be zinc‐deficient (Kilic 1998). One study enrolled adult participants who were diagnosed as zinc‐deficient (Badii 2012). See Table 3 (Summary of characteristics of included studies).

Primary outcomes

Zinc deficiency

None of the four studies in this comparison reported the number of participants found to be zinc‐deficient after the intervention.

Serum or plasma zinc

Three of the four included trials in this comparison evaluated this outcome (Badii 2012; Hambidge 1979; Kilic 1998). One cluster‐randomised trial did not report this outcome (Sanchez 2014). Participants consuming foods fortified with zinc as a single micronutrient had higher serum zinc levels (μmol/L) than those consuming unfortified foods (mean difference (MD) 2.12 μmol/L, 95% confidence interval (CI) 1.25 to 3.00 μmol/L; 3 studies; 158 participants) after a follow‐up period ranging from one to nine months (Analysis 1.1). The heterogeneity was high (Chi² = 8.34, I² = 76%; P = 0.02) mainly due to differences in effect size among the studies (Analysis 1.1), but the direction of benefit was the same in all three studies. We rated the evidence as low quality due to the small number of participants in each arm and high risk of attrition bias in one trial (Hambidge 1979). There was no more than one study in each prespecified age group within each comparison group; pooled analysis was therefore not possible for the prespecified age groups.

1.1. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 1 Serum or plasma zinc (in µmol/L).

In the subgroup analysis for comparison 1 (zinc‐fortified foods versus unfortified foods) (Analysis 1.2; Analysis 1.3; Analysis 1.4; Analysis 1.5; Analysis 1.6), the rise in serum zinc was not seen with a study, Hambidge 1979, conducted in a high‐income country (Analysis 1.6), using more than 6 months of supplementation (Analysis 1.2) of maize‐based cereals (Analysis 1.3) with zinc‐oxide for fortification (Analysis 1.4). The subgroup analysis according to dose of zinc (≤ 10 mg/100 g of food or > 10 mg/100 g of food) suggested no evidence of effect of dose of zinc in fortified foods on serum zinc status. However, the analysis was very underpowered to reach any conclusions for subgroup effects, with often only one study in each subgroup.

1.2. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 2 Serum or plasma zinc (in µmol/L) (by duration of intervention).

1.3. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 3 Serum or plasma zinc (in µmol/L) (by type of food vehicle).

1.4. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 4 Serum or plasma zinc (in µmol/L) (by type of zinc compound).

1.5. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 5 Serum or plasma zinc (in µmol/L) (by dose of zinc added to the food).

1.6. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 6 Serum or plasma zinc (in µmol/L) (by development status of country).

We could not perform sensitivity analysis for low risk of bias for allocation concealment as none of the three studies providing data for this outcome were at low risk of selection bias. For blinding, the inclusion of only two studies at low risk of bias, Hambidge 1979 and Kilic 1998, also showed a significant increase in serum zinc levels with zinc fortification (MD 1.94 μmol/L, 95% CI 0.10 to 3.77 μmol/L; 2 studies; 83 participants). Sensitivity analysis by excluding one study at high risk of attrition bias, Hambidge 1979, also showed a beneficial effect of zinc fortification on serum zinc levels (MD 2.46 μmol/L, 95% CI 2.05 to 2.87 μmol/L; 2 studies; 99 participants; I² = 8%; P = 0.30).

Stunting

Two studies in preschool children reported this outcome (Hambidge 1979; Sanchez 2014), and there was no evidence of effect of zinc fortification on stunting (risk ratio (RR) 0.88, 95% CI 0.36 to 2.13; 2 studies; 397 participants; I² = 26%; P = 0.24; low‐quality evidence) (Analysis 1.7). In a study evaluating the effect of zinc‐fortified milk on height and weight status of preschool children attending day‐care centres (Sanchez 2014), the frequency of stunting (height‐for‐age z‐score < ‐2) was comparable in children receiving zinc‐fortified or unfortified milk (RR 0.60, 95% CI 0.23 to 1.57) after 16 weeks of intervention. In another study (Hambidge 1979), after 9 months of consuming zinc‐fortified cereals, 7 of 13 children who were stunted (height for age below 10th percentile) at baseline had increased their height above that percentile. Two out of six children stunted at baseline in the control group were no more stunted after receiving unfortified cereals for nine months.

1.7. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 7 Stunting (author defined).

Underweight

Two studies in comparison 1 reported this outcome in preschool children (Hambidge 1979; Sanchez 2014). No effect of fortification of food with zinc could be documented in these studies (RR 3.10, 95% CI 0.52 to 18.38; 2 studies; 397 participants; I² = 0%; P = 0.96; low‐quality evidence) (Analysis 1.8). In the study on fortification of milk with zinc (Sanchez 2014), 3 out of 205 children receiving fortified milk and nil out of 96 children receiving unfortified milk were underweight (weight‐for‐age < ‐2 standard deviation (SD)) after 16 weeks of intervention. In the other study (Hambidge 1979), after 9 months of consuming zinc‐fortified cereals, 12 of 15 children who were underweight (weight for age below 10th percentile) at baseline had increased their weight above that percentile. Three out of four children who were underweight at baseline in the control group were no more so after receiving unfortified cereals for nine months.

1.8. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 8 Underweight (author defined).

Secondary outcomes

Pneumonia, diarrhoea, and all‐cause morbidity

Only two trials provided data related to the occurrence of morbidity in children receiving foods fortified with zinc (Kilic 1998; Sanchez 2014). A trial on the fortification of milk reported fewer occurrences of acute respiratory infections in children receiving milk fortified with zinc salts in comparison to those receiving unfortified milk (21 episodes in 205 children in the intervention group versus 20 episodes in 96 children in the control group; RR 0.49, 95% CI 0.28 to 0.86) (Sanchez 2014). The same trial did not document any benefit of zinc fortification on frequency of diarrhoea (7 episodes in 205 children in the intervention group versus 3 episodes in 96 children in the control group; RR 1.09, 95% CI 0.29 to 4.13) (Table 5).

3. Summary of outcomes reported in single studies only.

Outcome	Study reference (food vehicle)	Participants	Zinc salt and dose of elemental zinc	Duration of intervention	Main results
Comparison 1 (foods fortified with zinc alone versus same foods without zinc)
Acute respiratory infection (ARI) defined as ≥ 2 of the following symptoms: cough, runny nose, shortness of breath, and pharyngeal pain for ≥ 2 days	Sanchez 2014 (milk)	301 healthy children (age 2 to 5 y) from child day‐care centres who received more than 80% of their daily food intake from the childcare, and who liked milk	Zinc sulphate and zinc amino chelate Group 1: milk fortified with elemental zinc (as zinc sulphate) Group 2: milk fortified with elemental zinc (as zinc amino chelate) Group 3: unfortified milk The dose of elemental zinc varied according to the age of the participants. Children 2 to 3 y of age received 7 mg zinc daily, and children 4 to 5 y of age received 9.45 mg elemental zinc	4 months (16 weeks)	9/93 children in Group 1, 12/112 children in Group 2, and 20/96 children in Group 3 developed ARI. Incidence rates: Group 1: 1.42/1000 child‐years Group 2: 1.57/1000 child‐years Group 3: 3.30/1000 child‐years
Acute diarrhoeal disease defined as ≥ 3 liquid stools in 24 hours with > 2 days of evolution and < 14 days duration	Sanchez 2014 (milk)	301 healthy children (age 2 to 5 y) from child day‐care centres who received more than 80% of their daily food intake from the childcare, and who liked milk	Zinc sulphate and zinc amino chelate Group 1: milk fortified with elemental zinc (as zinc sulphate) Group 2: milk fortified with elemental zinc (as zinc amino chelate) Group 3: unfortified milk The dose of elemental zinc varied according to the age of the participants. Children 2 to 3 y of age received 7 mg zinc daily, and children 4 to 5 y of age received 9.45 mg elemental zinc	4 months (16 weeks)	1/93 children in Group 1, 6/112 children in Group 2, and 3/96 children in Group 3 developed diarrhoea. Incidence rates: Group 1: 0.15/1000 child‐years Group 2: 0.78/1000 child‐years Group 3: 0.49/1000 child‐years
Infectious morbidity (lower and upper respiratory infections, acute diarrhoea, and pyoderma)	Kilic 1998 (bread)	24 schoolchildren (age 7 to 11 y; 10 boys and 14 girls) with serum zinc concentrations below 65 µg/dL	Zinc acetate providing 400 mg of elemental zinc per loaf of bread Group 1: zinc‐fortified bread providing 2 mg/kg/day elemental zinc Group 2: unfortified bread	3 months	Number of episodes 1.0 ± 0.9 in Group 1 and 2.3 ± 1.6 in Group 2 (P < 0.05)
Haemoglobin	Kilic 1998 (bread)	24 schoolchildren (age 7 to 11 y; 10 boys and 14 girls) with serum zinc concentrations below 65 µg/dL	Zinc acetate providing 400 mg of elemental zinc per loaf of bread Group 1: zinc‐fortified bread providing 2 mg/kg/day elemental zinc Group 2: unfortified bread	3 months	Mean ± SD haemoglobin levels (mmol/L) in Group 1 (1.98 ± 0.17) not significantly (P > 0.05) different from Group 2 (1.92 ± 0.22)
Serum ferritin	Kilic 1998 (bread)	24 schoolchildren (age 7 to 11 y; 10 boys and 14 girls) with serum zinc concentrations below 65 µg/dL	Zinc acetate providing 400 mg of elemental zinc per loaf of bread Group 1: zinc‐fortified bread providing 2 mg/kg/day elemental zinc Group 2: unfortified bread	3 months	Mean ± SD ferritin levels (µg/L) in Group 1 (30.4 ± 9.7) not significantly (P > 0.05) different from Group 2 (24.1 ± 11.7)
Vomiting (defined as any episode within 20 minutes of ingesting the milk)	Sanchez 2014 (milk)	301 healthy children (age 2 to 5 y) from child day‐care centres who received more than 80% of their daily food intake from the childcare, and who liked milk	Zinc sulphate and zinc amino chelate Group 1: milk fortified with elemental zinc (as zinc sulphate) Group 2: milk fortified with elemental zinc (as zinc amino chelate) Group 3: unfortified milk The dose of elemental zinc varied according to the age of the participants. Children 2 to 3 y of age received 7 mg zinc daily, and children 4 to 5 y of age received 9.45 mg elemental zinc	4 months (16 weeks)	5/93 children in Group 1, 12/112 children in Group 2, and 10/96 children in Group 3 developed vomiting (P = 0.246)
Any adverse effect	Sanchez 2014 (milk)	301 healthy children (age 2 to 5 y) from child day‐care centres who received more than 80% of their daily food intake from the childcare, and who liked milk	Zinc sulphate and zinc amino chelate Group 1: milk fortified with elemental zinc (as zinc sulphate) Group 2: milk fortified with elemental zinc (as zinc amino chelate) Group 3: unfortified milk The dose of elemental zinc varied according to the age of the participants. Children 2 to 3 y of age received 7 mg zinc daily, and children 4 to 5 y of age received 9.45 mg elemental zinc	4 months (16 weeks)	6/93 children in Group 1, 16/112 children in Group 2, and 13/96 children in Group 3 developed any adverse effect (P = 0.170)
*Weight	Sanchez 2014 (milk)	301 healthy children (age 2 to 5 y) from child day‐care centres who received more than 80% of their daily food intake from the childcare, and who liked milk	Zinc sulphate and zinc amino chelate Group 1: milk fortified with elemental zinc (as zinc sulphate) Group 2: milk fortified with elemental zinc (as zinc amino chelate) Group 3: unfortified milk The dose of elemental zinc varied according to the age of the participants. Children 2 to 3 y of age received 7 mg zinc daily, and children 4 to 5 y of age received 9.45 mg elemental zinc	4 months (16 weeks)	Mean ± SD weight (kg) in fortified group (Group 1 + 2) 16.03 ± 2.61 higher than that 14.99 ± 2.24 in unfortified group (Group 3) (MD 1.04 kg, 95% CI 0.47 to 1.61 kg) and height (MD 3.15 cm, 95% CI 1.48 to 4.82 cm)
*Height	Sanchez 2014 (milk)	301 healthy children (age 2 to 5 y) from child day‐care centres who received more than 80% of their daily food intake from the childcare, and who liked milk	Zinc sulphate and zinc amino chelate Group 1: milk fortified with elemental zinc (as zinc sulphate) Group 2: milk fortified with elemental zinc (as zinc amino chelate) Group 3: unfortified milk The dose of elemental zinc varied according to the age of the participants. Children 2 to 3 y of age received 7 mg zinc daily, and children 4 to 5 y of age received 9.45 mg elemental zinc	4 months (16 weeks)	Mean ± SD height (cm) in fortified group (Group 1 + 2) 99.43 ± 6.32 higher than that 96.28 ± 7.17 in unfortified group (Group 3) (MD 3.15 cm, 95% CI 1.48 to 4.82 cm)
Comparison 2 (foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc)
Zinc deficiency (defined as plasma zinc < 65 µg/dL)	Lopez de Romaña 2005 (wheat products; biscuits and noodles)	58 stunted, moderately anaemic children (age 3 to 4 y) residing in a poor community	Zinc sulphate Group 1: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and no zinc Group 2: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and 3 mg elemental zinc per 100 g flour Group 3: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and 9 mg elemental zinc per 100 g flour	2 months (70 days)	1/10 children in Group 1, and no children in Group 2 (n = 8) or Group 3 (n = 12) had zinc deficiency (P = 0.046)
Weight	Lopez de Romaña 2005 (wheat products; biscuits and noodles)	58 stunted, moderately anaemic children (age 3 to 4 y) residing in a poor community	Zinc sulphate Group 1: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and no zinc Group 2: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and 3 mg elemental zinc per 100 g flour Group 3: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and 9 mg elemental zinc per 100 g flour	2 months (70 days)	Mean ± SD weight (kg) in Group 1: 14.8 ± 1.4; Group 2: 15.2 ± 2.3; Group 3: 14.5 ± 1.4; P = 0.83
Height	Lopez de Romaña 2005 (wheat products; biscuits and noodles)	58 stunted, moderately anaemic children (age 3 to 4 y) residing in a poor community	Zinc sulphate Group 1: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and no zinc Group 2: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and 3 mg elemental zinc per 100 g flour Group 3: 2 meals/d of 100 g wheat products fortified with 3 mg elemental iron and 9 mg elemental zinc per 100 g flour	2 months (70 days)	Mean ± SD height (cm) in Group 1: 93.5 ± 3.7; Group 2: 94.7 ± 5.5; Group 3: 91.7 ± 4.6; P = 0.97

*Data supplied by authors on request.

CI: confidence interval
 MD: mean difference
 SD: standard deviation

The other trial reported all‐cause morbidity data related to zinc fortification of foods (Kilic 1998). The data were reported in another publication (Saldamli 1996) out of this study (Kilic 1998). In this study on 24 zinc‐deficient (serum zinc < 65 µg/dL) children aged 7 to 11 years, the mean (SD) number of infections (diarrhoea, upper respiratory infections, lower respiratory infections, and pyoderma) in the intervention group after 90 days of consuming zinc‐fortified bread was lower in comparison to the control group who consumed unfortified bread (1.0 (0.9) versus 2.3 (1.6); P < 0.05). However, the very small sample size of the study makes it difficult to draw any meaningful conclusions (Table 5).

Haemoglobin

Only one trial in this comparison provided information on haemoglobin of participants after 90 days of intervention (Kilic 1998). There was no effect of zinc fortification on haemoglobin levels of participants (standardised mean difference (SMD) 0.29, 95% CI ‐0.51 to 1.10) (Table 5).

Anaemia

None of the trials in this comparison provided number of anaemic participants after intervention.

Adverse effect (iron status measured as serum ferritin in µg/L)

One study evaluated the effect of zinc fortification on iron status (Kilic 1998). There was no significant impact of zinc fortification on serum ferritin levels in the study participants (Table 5).

Adverse effect (copper status as measured by serum or plasma copper level in µg/dL)

Two studies in this comparison measured serum copper after zinc fortification (Hambidge 1979; Kilic 1998). There was no significant effect of zinc fortification on serum copper levels (MD ‐8.73 µg/dL, 95% CI ‐18.03 to 0.58 mcg/dL; 2 studies; 82 participants; I² = 0%; P = 0.81) (Analysis 1.14).

1.14. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 14 Adverse effect (copper status as measured by serum or plasma copper level in μg/dL).

Adverse effect (vomiting)

Only one study reported the outcome vomiting (Sanchez 2014). The occurrence of vomiting in children receiving milk fortified with zinc salts (17 out of 205 children) did not differ from those receiving unfortified milk (10 out of 96 children). The same study also reported the occurrence of any adverse effect in study children and found no difference between children receiving zinc‐fortified milk (22 out of 205) and unfortified milk (13 out of 96) (Table 5).

Weight and height in children 24 months to 59 months of age

Only one study in this comparison reported weight and height of children (between the ages of 2 and 5 years) at the end of the follow‐up period with fortification (Sanchez 2014). This study on zinc‐fortified milk reported significantly higher weight (MD 1.04 kg, 95% CI 0.47 to 1.61 kg) and height (MD 3.15 cm, 95% CI 1.48 to 4.82 cm) in the intervention group after 16 weeks of receiving zinc‐fortified milk in comparison to the control group who received unfortified milk for the same duration (Table 5). Another study conducted in this age group reported growth velocities of children during the period of fortification (Hambidge 1979), and found no significant difference in the mean increments for height and weight between children receiving zinc‐fortified cereals in comparison to those receiving unfortified cereals.

Other outcomes

No study provided data on the following secondary outcomes: mid‐upper arm circumference (for children 24 to 59 months of age), cognitive and motor skill development (for children 24 to 59 months of age), adverse effect (iron status measured as serum transferrin receptor in mg/L), all‐cause death (for children 24 to 59 months of age), and cognitive and work performance (for adults). Hambidge 1979 reported that there were no significant differences between test and control children for mean increments in arm muscle circumference during the study period; no data were provided.

Comparison 2 (zinc‐fortified food in combination with other nutrients/factors versus same food containing other nutrients/factors without zinc)

Four out of eight included studies provided data for this comparison (Aaron 2011; Haibin 2001; Hettiarachchi 2004; Lopez de Romaña 2005). One of these studies evaluated stunted and moderately anaemic children aged less than 59 months (Lopez de Romaña 2005). One study was conducted in apparently healthy older children (5 to 11.9 years of age) (Hettiarachchi 2004), and one study was conducted in apparently healthy pregnant women (Haibin 2001). One study enrolled apparently healthy adult participants (Aaron 2011). See Table 3 (Summary of characteristics of included studies).

Primary outcomes

Zinc deficiency

Only one study contributed data to this outcome (Lopez de Romaña 2005). In this study with factorial design (wheat products fortified with iron only or iron and 1 of 2 amounts of zinc), the number of participants (moderately anaemic, stunted children aged 3 to 4 years) with zinc deficiency (plasma zinc < 65 μg/L) in the intervention group was reduced from 5 out of 26 (19.2%) at baseline to nil out of 20 (0%) after 70 days of consuming food products made up of zinc‐fortified (in addition to iron) wheat flour. The proportion of children with zinc deficiency in the control group (iron‐only fortification) was reduced from 3 out of 13 (23.1%) at baseline to 1 out of 10 (10%). Overall, a benefit of adding zinc to iron in wheat flour in terms of reducing the incidence of zinc deficiency could not be documented (RR 0.17, 95% CI 0.01 to 3.94), however we rated the evidence as very low quality due to the small number of children in the only trial reporting this outcome (Table 5).

Serum or plasma zinc

All four studies in this comparison evaluated serum or plasma zinc of participants after a variable period of intervention. Two studies with factorial design provided two comparison groups each (with a different set of controls for each group) for this outcome (Haibin 2001; Hettiarachchi 2004). The evidence was of low quality due to the small number of participants in each arm, and high risk of attrition bias in one trial (Haibin 2001). There was no significant increase in serum zinc levels (μmol/L) in participants consuming foods fortified with zinc in combination with other factors/nutrients in comparison to those consuming food fortified with other factors/nutrients (MD 0.03 μmol/L, 95% CI ‐0.67 to 0.72 μmol/L; 4 studies; 250 participants; low‐quality evidence) with no significant heterogeneity (I² = 20%; P = 0.28) (Analysis 2.2). There was no more than one study in each prespecified age group within each comparison group; pooled analysis was therefore not possible for the prespecified age groups.

2.2. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 2 Serum or plasma zinc (in µmol/L).

In subgroup analysis (Analysis 2.3; Analysis 2.4; Analysis 2.5; Analysis 2.6; Analysis 2.7), visual examination of the forest plots suggested that there were no clear differences between groups for duration of intervention (< 6 months versus ≥ 6 months), type of zinc compound used for fortification, type of food vehicle, or development status of the country. We did not perform subgroup analysis for gender in view of obvious differences in the age groups and physiological status of participants (for example pregnancy in Haibin 2001).

2.3. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 3 Serum or plasma zinc (in µmol/L) (by duration of intervention).

2.4. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 4 Serum or plasma zinc (in µmol/L) (by type of food vehicle).

2.5. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 5 Serum or plasma zinc (in µmol/L) (by type of zinc compound).

2.6. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 6 Serum or plasma zinc (in µmol/L) (by dose of zinc added to the food).

2.7. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 7 Serum or plasma zinc (in µmol/L) (by development status of country).

We could not perform sensitivity analysis for low risk of bias for allocation concealment as we assessed only one of the four studies included in this comparison to be at low risk of selection bias. For blinding, inclusion of only two studies at low risk of bias, Aaron 2011 and Lopez de Romaña 2005, did not show a significant increase in serum zinc levels with zinc fortification (MD ‐0.28 μmol/L, 95% CI ‐0.94 to 0.37 μmol/L; 2 studies; 134 participants). Sensitivity analysis by excluding two studies at high risk of attrition bias, Haibin 2001 and Lopez de Romaña 2005, also did not show a beneficial effect of zinc fortification on serum zinc levels (MD ‐0.31 μmol/L, 95% CI ‐0.86 to 0.24 μmol/L; 2 studies; 149 participants; I² = 4%; P = 0.35).

Stunting

No study evaluating effect of zinc fortification in combination with other nutrients (comparison 2) reported this outcome.

Underweight

No study evaluating effect of zinc fortification in combination with other nutrients (comparison 2) reported this outcome.

Secondary outcomes

Pneumonia, diarrhoea, and all‐cause morbidity

No study evaluating effect of zinc fortification in combination with other nutrients (comparison 2) reported outcomes related to morbidity.

Anaemia

Only two trials provided data on anaemia among study participants (Haibin 2001; Lopez de Romaña 2005). Lopez de Romaña 2005 used iron‐fortified wheat products with or without zinc fortification, whereas Haibin 2001 used a factorial design to evaluate the effect of fortification of wheat flour with calcium, vitamin D, zinc, vitamin C, and iron in various combinations. Lopez de Romaña 2005 enrolled 41 stunted, moderately anaemic (haemoglobin 9 to 11 g/dL) children in their study. The proportion of children having anaemia (haemoglobin 11 g/dL) decreased from 100% (27 out of 27) to 13.6% (3 out of 22) in the zinc‐added group in comparison to a reduction from 100% (14 out of 14) to 0% (0 out of 12) in the iron‐fortified group, after 70 days of supplementation. Haibin 2001 enrolled pregnant women in their study; the baseline anaemia rate was 35.2%. Prevalence of anaemia in the groups of Ca + Fe + Zn + VD and Ca + Fe + VD was 35.3% and 40.7%, respectively, before fortification and was reduced to zero and 4.0%, respectively, after participants consumed fortified biscuits for 24 weeks. Pooled results from these two trials did not suggest any positive/negative effect of fortification of zinc on anaemia (RR 0.89, 95% CI 0.35 to 2.28; I² = 0%; P = 0.55) (Analysis 2.8).

2.8. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 8 Anaemia.

Haemoglobin

Three trials in this comparison group provided information on haemoglobin after the end of the fortification period (Haibin 2001; Hettiarachchi 2004; Lopez de Romaña 2005). Two studies provided two arms each (with a separate control group for each arm) for this outcome, owing to their factorial design (Haibin 2001; Hettiarachchi 2004). There was no effect of zinc fortification on haemoglobin levels of participants (MD 0.13 g/dL, 95% CI ‐0.33 to 0.59 g/dL; 3 studies; 186 participants; I² = 38%; P = 0.17) (Analysis 2.9).

2.9. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 9 Haemoglobin (g/dL).

Adverse effect (iron status measured as serum ferritin in µg/L)

Two studies in this comparison group evaluated serum ferritin after zinc fortification as an outcome (Hettiarachchi 2004; Lopez de Romaña 2005). There was no significant impact of zinc fortification on serum ferritin levels in the study participants (I² = 0%; P = 0.47) (Analysis 2.10).

2.10. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 10 Adverse effect (iron status measured as serum ferritin in µg/L) (Ln transformed).

Adverse effect (copper status as measured by serum or plasma copper level in µg/dL)

No study in this comparison provided data for this outcome.

Adverse effect (vomiting)

No study in this comparison reported this outcome.

Weight and height in children 24 months to 59 months of age

One study with a factorial design (iron‐fortified wheat flour with no zinc, low zinc (3 mg/100 g), or high zinc (9 mg/100 g)) reported no difference in the magnitude of changes in weight‐for‐age, weight‐for‐height, or height‐for‐age z‐scores (Lopez de Romaña 2005). Mean (SD) weights of children in these three groups at the end of the 70‐day follow‐up period were 14.8 (1.4) kg, 15.2 (2.3) kg, and 14.5 (1.4) kg, respectively (P = 0.83). The pooling of results for the low‐ and high‐zinc group did not document any evidence (low level of evidence) of benefit of zinc fortification on weight (MD 0.02 kg, 95% CI ‐1.09 to 1.13 kg), in comparison to the group with no zinc. Mean (SD) heights of the children in these three groups (no zinc, low zinc and high zinc) at the end of 70‐day follow‐up period were 93.5 (3.7) cm, 94.7 (5.5) cm, and 91.7 (4.6) cm, respectively (P = 0.97). The pooling of results for the low‐ and high‐zinc group did not document any evidence of benefit of zinc fortification on height (MD ‐0.44 cm, 95% CI ‐3.40 to 2.52 cm), in comparison to the group with no zinc (Table 5).

Other outcomes

No study provided data on the following secondary outcomes: mid‐upper arm circumference (for children 24 to 59 months of age), cognitive and motor skill development (for children 24 to 59 months of age), and cognitive and work performance (for adults).

Comparison 3 (foods fortified with zinc versus no intervention)

There was no study in this comparison group.

Discussion

Summary of main results

We included eight trials in this review, of which four compared zinc‐fortified foods with unfortified foods and four compared foods fortified with zinc in combination with other nutrients and foods fortified with other nutrients minus zinc. Only one of the eight trials reported the primary outcome of zinc deficiency, and there was no significant difference in incidence of zinc deficiency after fortification of foods with zinc in combination with other micronutrients (very low‐quality evidence). We found that zinc‐fortified foods may improve the serum zinc levels of populations (low‐quality evidence). However, it may make little or no difference to the serum zinc levels when foods are cofortified with other micronutrients (low‐quality evidence).

The fortification of foods with zinc may make little or no difference to the incidence of underweight or stunting in children (low‐quality evidence). We found limited evidence of reduced occurrence of acute respiratory infection and all‐cause morbidity during the period of supplementation with zinc‐fortified foods (one study). Fortification of foods with zinc may not have a significant effect on haematological indicators or serum copper levels. We found no studies documenting the effects of zinc‐fortified foods on other parameters such as cognition and development of children, or work capacity of adults.

Overall completeness and applicability of evidence

Fortification of foods with zinc is considered to be a potentially useful strategy for improving the zinc status of deficient populations, and thus control its deficiency. Most of the studies in this review were from lower‐ or upper‐middle‐income countries with a population described as zinc‐deficient or having high risk of zinc deficiency. The chemical forms of zinc evaluated in five of the included studies were either zinc oxide or zinc sulphate, which are considered to be the two cheapest zinc formulations and are recognised as safe for human consumption (Brown 2010). The vehicles selected for fortification were the typical high‐phytate cereal‐based foods consumed by these populations. However, most studies included a small number of participants, and two of the included studies were primarily designed to determine the absorption of zinc or iron, or both, from fortified diets using zinc isotopic tracers (Hettiarachchi 2004; Lopez de Romaña 2005). We found other studies evaluating the bioavailability/acceptability of zinc from fortified diets (Herman 2002; Islam 2013; Khoshgoftarmanesh 2010; Olivares 2013), but we did not include these studies in the present review as they provided no information about zinc or health status of the participants. These studies have shown in general that the additional zinc provided by fortification decreases fractional absorption of zinc from phytate‐rich foods, but increases total zinc absorption from these foods (Brown 2010; Hess 2009b).

Four of the studies included in this review used additional fortificants, but zinc was the only nutrient that differed between the two groups. This review suggests (low‐quality evidence) that there is evidence of no significant increase in serum zinc levels in populations where additional fortificants were added to the food. It is plausible that cofortification with other micronutrients (for example iron) interfered with absorption of zinc from fortified food. Although there are no direct data about cofortificants interfering with zinc absorption from foods, similar findings have been reported from trials of multimicronutrient powders. In a community‐based study on a home‐based fortification program in 6‐ to 18‐month‐old, moderately anaemic Ghanaian children (Zlotkin 2003), significantly fewer children in the zinc‐plus‐iron group had zinc deficiency (serum zinc less than 70 µg/dL) than in the iron group, but there were no significant differences in final mean serum zinc concentrations. On the contrary, fortification of food with zinc does not seem to affect iron or copper status, as is evident from the findings of this review (low‐quality evidence) and reviews of absorption studies (Brown 2010; Hess 2009b).

There are many studies on the fortification of foods with multiple micronutrients, but we excluded these from the review as zinc was not the only nutrient that differed between the groups (see Characteristics of excluded studies). It is not possible to attribute a health benefit to zinc if it is only one of the nutrients used to fortify foods. Some of these trials evaluated serum zinc as one of the outcomes, and the results were inconsistent (Hess 2009b).

Several other studies have evaluated the effect of zinc fortification of milk, infant feeding formulas, and complementary foods in children younger than two years of age. We did not include these in the review as children aged under two years of age are not the targets of mass fortification programs, for which the selected food vehicle should be consumed regularly in fairly constant amounts by a large segment of the general population (Hess 2009b). The results of this review are thus applicable only to fortification of foods fit for consumption by older children and adults. There was also no study available from adolescent age group, who might have a different level of nutrient intake from fortified foods related to their consumption habits.

It must be noted that most of the information obtained for this review was from small‐scale studies using targeted fortification. No information is available from large‐scale programmatic effectiveness trials. As most countries using mandatory zinc fortification are using multiple micronutrients for fortification, this information might be difficult to obtain in the near future. Evaluations of such programs in Chinese women have reported higher serum zinc levels in those consuming products made up of multiple micronutrient‐fortified flour (Huo 2011).

Quality of the evidence

We obtained most of the information in this review from trials categorised as being at unclear bias for key elements of randomisation, but low bias for blinding and attrition. The reporting of randomisation details in most trials was inadequate; we attempted to contact authors to obtain more data, and used the information provided by those who responded (details in Characteristics of included studies) for final rating of quality of evidence. We classifed one study as being at high risk of bias for randomisation (allocation and concealment) and blinding. Three studies had high attrition bias for key outcomes. Studies had significant clinical and statistical heterogeneity. According to the GRADE assessment the quality of the evidence was low for most outcomes due to the small number of participants in each arm and high attrition bias of some studies. Further research is very likely to have an important impact on our confidence in the estimate of effect and is likely to change the estimate.

Potential biases in the review process

Two review authors independently assessed eligibility for inclusion, carried out data extraction using similar pro forma, and assessed risk of bias. As many trials did not provide details about the process of randomisation or blinding, we contacted study authors for further information. As the process of assessing risk of bias includes many personal judgements, especially in the absence of precise details in published papers, we settled the issue by mutual discussion. The risk of publication bias was minimised by searching for grey literature, trial registries, and by contacting agencies involved in this intervention. Language bias was minimised by not limiting the searches to any language and by obtaining translations of the abstracts/full texts to English where required.

Agreements and disagreements with other studies or reviews

A systematic review of randomised controlled trials and quasi‐randomised trials evaluated the health outcomes of women and children who received zinc‐fortified food (Das 2013). This review included 11 studies, of which 6 pertained to use of formula feeds in neonates and 1 evaluated zinc fortification of complementary food for use in infants. This review only included trials where zinc was the sole micronutrient used for fortification. Trials evaluating zinc fortification along with other micronutrients were not included, even if zinc was the only nutrient that differed in the comparison groups. The authors reported that zinc fortification was associated with significant improvements in plasma zinc concentrations (SMD 1.28, 95% CI 0.56 to 2.01). An improvement in height velocity assumed borderline significance after removing one trial as outlier (SMD 0.52, 95% CI 0.01 to 1.04); most of the impact related to height velocity was seen in a subgroup of very low birthweight infants. There was no evidence of impact of zinc fortification on serum copper, haemoglobin, or weight gain. The authors concluded that zinc fortification is associated with an increased serum concentration of the micronutrient, but overall evidence of the effectiveness of this approach is limited.

Our review focused on a fortification strategy intended for an age group of over two years, as infants are usually not the target of mass fortification strategies meant for the general population. We were able to identify more studies in this age group due to the inclusion of studies using other cofortificants, where the only difference between the comparison groups was zinc. It is important to include these studies, as they provide important data related to the effect of zinc that may be independent of other micronutrients included in fortification. Even if there is an interference in the effect of zinc because of cofortification, it is important to document the same by including such trials in the analysis, and analysing them separately.

Authors of a comprehensive desk review on zinc fortification concluded that zinc fortification can increase dietary zinc intake and total daily zinc absorption without adversely affecting the absorption of other minerals such as iron (Hess 2009b). The authors also reported that despite the positive effect of zinc fortification on total zinc absorption, the data regarding positive impact of zinc fortification on serum zinc concentrations or functional indicators of zinc status were inconsistent. The studies evaluating other functional outcomes in this review were too few for any valid interpretation.

Authors' conclusions

Implications for practice.

Fortification of staple foods is proposed as an effective strategy to combat micronutrient deficiencies in populations. This review suggests that there may be a marginal increase (low‐quality evidence) in serum zinc levels (an indicator of zinc status) with zinc‐fortified cereal food staples. It is uncertain whether this intervention can reduce zinc deficiency or improve functional outcomes such as height, weight, infectious morbidity, or prevalence of malnutrition (wasting and stunting).

There appears to be a reduction in infectious morbidity (very low‐quality evidence; one trial) with use of zinc as a fortificant. The effect of zinc fortification in raising serum zinc levels may be negated if the food is simultaneously fortified with other nutrients. The practice of many countries of implementing mandatory fortification of cereal flours with zinc in combination with other micronutrients may therefore not be the best strategy to combat zinc deficiency in these populations.

This review suggests that fortification of foods with zinc may make little or no difference to the iron status (anaemia, haemoglobin, serum ferritin), copper status, or occurrence of vomiting after intervention. We are unable to recommend the dose of zinc to be used for fortification due to the small number of trials, and even amongst these trials, there was no difference in effect on serum zinc with dose.

We are unable to provide any evidence related to fortification of non‐cereal‐based foods (for example beverages, sugar, edible oils, condiments) due to the absence of any study assessing these interventions.

Implications for research.

As there was limited and low‐quality evidence for most outcomes related to zinc fortification of foods, we have identified many research gaps. The following areas merit further research.

1. Most information about zinc fortification of foods is available from small‐scale studies using targeted fortification. Large‐scale programmatic effectiveness trials and controlled evaluation of ongoing fortification programs need to be conducted to provide precise data related to zinc fortification of foods in large populations.

2. Future zinc fortification studies must include functional outcomes such as growth, stunting and wasting of under‐five children (based on current World Health Organization definitions), cognitive development, and work capacity (for adults). The target population should also include adolescents, pregnant/lactating women, and the elderly.

3. Future studies need to compare fortification of foods with zinc in isolation versus in combination with other micronutrients, with special emphasis on micronutrient interactions.

4. The feasibility of using other food vehicles, such as edible oils, sugars, and beverages, for zinc fortification should be explored.

5. The safe and effective dose of zinc in fortified foods needs to be determined by using trials with factorial design.

6. Monitoring and evaluation of ongoing food fortification programmes to identify the acceptability, adequacy of consumption (especially in children and adolescents), safety, and cost‐effectiveness.

Appendices

Appendix 1. Appendix 1

Search Strategy for Databases

CENTRAL

#1           MeSH descriptor: [Zinc] explode all trees

#2           MeSH descriptor: [Zinc Compounds] explode all trees

#3           (zn or zinc):ti,ab,kw  (Word variations have been searched)

#4           MeSH descriptor: [Food, Fortified] explode all trees

#5           ((fortif* or enrich* or enhanc* or boost*) near/2 (food* or rice or flour or corn or wheat or semolina or pulse* or bread* or sauce* or beverage* or maize or oat* or sugar or salt or oil* or fat* or condiment* or seasoning* or spice* or milk or dairy or juice* or nectar*)):ti,ab,kw  (Word variations have been searched)

#6           #1 or #2 or #3

#7           #4 or #5

#8           #6 and #7

 

MEDLINE (OVID)

1 exp Zinc/

2 exp Zinc Compounds/

3 (zn or zinc).tw.

4 exp Food, Fortified/

5 ((fortif* or enrich* or enhanc* or boost*) adj2 (food* or rice or flour or corn or wheat or semolina or pulse* or bread* or sauce* or beverage* or maize or oat* or sugar or salt or oil* or fat* or condiment* or seasoning* or spice* or milk or dairy or juice* or nectar*)).tw.

6 or/1‐3

7 4 or 5

8 6 and 7

9 exp animals/ not humans.sh.

10 8 not 9

EMBASE (OVID)

1 exp Zinc/

2 exp Zinc Compounds/

3 (zn or zinc).tw.

4 exp Food, Fortified/

5 ((fortif* or enrich* or enhanc* or boost*) adj2 (food* or rice or flour or corn or wheat or semolina or pulse* or bread* or sauce* or beverage* or maize or oat* or sugar or salt or oil* or fat* or condiment* or seasoning* or spice* or milk or dairy or juice* or nectar*)).tw.

6 or/1‐3

7 4 or 5

8 6 and 7

9 exp animal/ not human.sh.

10 8 not 9

CINAHL (EBSCO)

S8 S4 AND S7

S7 S5 OR S6

S6 ((fortif* or enrich* or enhanc* or boost*) N2 (food* or rice or flour or corn or wheat or semolina or pulse* or bread* or sauce* or beverage* or maize or oat* or sugar or salt or oil* or fat* or condiment* or seasoning* or spice* or milk or dairy or juice* or nectar*))

S5 (MH "Food, Fortified")

S4 S1 OR S2 OR S3

S3 (zn or zinc)

S2 (MH "Zinc Compounds+")

S1 (MH "Zinc")

Web of Science (SCI, SSCI & CPCI) & BIOSIS (ISI)

# 1 Topic=(((fortif* or enrich* or enhanc* or boost*) near/2 (food* or rice or flour or corn or wheat or semolina or pulse* or bread* or sauce* or beverage* or maize or oat* or sugar or salt or oil* or fat* or condiment* or seasoning* or spice* or milk or dairy or juice* or nectar*)))

#2 Topic=((zn or zinc))

#3 #1 and #2

Popline

(zinc or zn) and fortif*

(zinc or zn) and enhanc*

(zinc or zn) and enrich*

(zinc or zn) and boost*

AGRICOLA

Search = (zinc zn)[ in Title ]OR(zinc zn)[ in Abstract ]AND( fortif? enrich? enhanc? boost?)[ in Abstract ]OR( fortif? enrich? enhanc? boost?)[ in Title ]

OpenGrey

Zinc or ZN

Bibliomap & TROPHI

1.Freetext: zinc

2.Freetext: zn

3. 1 or 2

4. Freetext: fortif*

5. Freetext: enrich*

6. Freetext: enhanc*

7. Freetext: boost*

8. 4 or 5 or 6 or 7

9. 3 and 8

IBECS, WHOLIS, PAHO and LILACS

(zinc or zn) [Words] and (fortif$ or enrich$ or enhanc$ or boost$) [Words] and (food$ or rice or flour or corn or wheat or semolina or pulse$ or bread$ or sauce$ or beverage$ or maize or oat$ or sugar or salt or oil$ or fat$ or condiment$ or seasoning$ or spice$ or milk or dairy or juice$ or nectar$)

SCIELO

(zinc or zn) and (fortif$ or enrich$ or enhanc$ or boost$) and (food$ or rice or flour or corn or wheat or semolina or pulse$ or bread$ or sauce$ or beverage$ or maize or oat$ or sugar or salt or oil$ or fat$ or condiment$ or seasoning$ or spice$ or milk or dairy or juice$ or nectar$) [All indexes]

WPRO

zinc AND fortified

zinc AND fortify

zinc AND enhanced

zinc AND enhance

zinc AND enrich

zinc AND enriched

zinc AND boost

zinc AND boosts

zinc AND boosted

IMSEAR

zinc AND (fortify or fortified)

INMED

zinc or zn AND fortify or fortified or enhance or enhanced or enrich or enriched or boost or boosted

NATIVE HEALTH RESEARCH DATABASE

KEYWORDS for: (zinc OR zn) AND (fortified or fortify or enhanced or enhance or enrich or enriched or boosted)

AFRO and EMRO

(zinc OR zn) AND (fortified or fortify or enhanced or enhance or enrich or enriched or boosted)

 

Appendix 2. Relevant assumptions, calculations, and imputations for extracting specific data in some individual trials

1. Aaron 2011

(a)    This trial with factorial design provided changes in plasma zinc following intervention and baseline serum zinc levels in each group separately. Post‐intervention mean plasma zinc levels were obtained by adding the mean change in each group to its baseline mean. SD for post‐intervention serum zinc levels in each intervention group was assumed to be same as baseline SD.

(b)   The post‐intervention plasma means in two intervention groups (FZn7.5 and FZn15) were pooled by using the individual means and numbers in each group. The pooled SD of two groups was calculated using the following SPSS syntax:

*Pooled SD haste pre‐David method.

compute psd=sqrt(((n1‐1)*s1*s1 + (n2‐1)*s2*s2)/((n1‐1) + (n2‐1))).

variable label psd 'Pooled SD'.

exe.

 

Badii 2012

This trial with factorial design provided post‐intervention serum zinc levels in two groups (low‐zinc and high‐zinc) separately. The post‐intervention plasma means in two intervention groups (FZn7.5 and FZn15) were pooled using the individual means and number of participants in each group. The pooled SD of two groups was calculated using the following SPSS syntax:

*Pooled SD haste pre‐David method.

compute psd=sqrt(((n1‐1)*s1*s1 + (n2‐1)*s2*s2)/((n1‐1) + (n2‐1))).

variable label psd 'Pooled SD'.

exe.

 

Haibin 2001

(a)    This study was translated from Chinese to English by WHO translations department.

(b)   Number of anaemic participants at the end of trial were calculated from the proportions provided in Table 4 using the number of subjects in which haemoglobin were done at the end of the trial (Column 2 of Table 4)

 

Hambidge 1979

This trial provided changes in plasma zinc and copper following intervention and baseline serum zinc and serum copper levels for both groups (intervention and control) combined.

(a)    Baseline mean and SD serum zinc and serum copper levels in each group (intervention and control) were assumed to be same as that for the combined group.

(b)   Post‐intervention mean plasma zinc levels were obtained by adding the mean change in each group to its baseline mean. SD for post‐intervention serum zinc levels in each intervention group was assumed to be same as baseline SD.

 

Kilic 1998

(a)    Levels on day‐90 of study considered for this systematic review as per protocol.

(b)   Haemoglobin values were provided in mmol/L, and standardized mean difference used for analysis.

(c)    Serum copper values (µmol/L) were converted into µg/dL by multiplying with 6.3532.

 

Lopez de Romaña 2005

This trial with factorial design compared two doses of zinc‐fortified (in addition to iron) foods versus iron‐fortified foods.

(a)    The post‐intervention plasma means in two intervention groups (Zn‐3 and Zn‐9) were pooled by using the individual means and numbers in each group. The pooled SD of two groups was calculated using the following SPSS syntax:

*Pooled SD haste pre‐David method.

compute psd=sqrt(((n1‐1)*s1*s1 + (n2‐1)*s2*s2)/((n1‐1) + (n2‐1))).

variable label psd 'Pooled SD'.

exe.

(b)   The study reported Geometric mean and SD for plasma ferritin values. However, the SD values were very high for these to be geometric. Clarification was sought from the authors, and the authors provided the actual means and standard deviations of the logarithm of ferritin values.

SPSS Syntax used for converting arithmetic means and SD to geometric means and SDs for serum ferritin

* Raw or arithmetic mean and SD to geometric mean and SD.

 

if (meandatacode=0) rawmeancas= tmremncas.

if (meandatacode=0) rawsdcas= tmresdcas.

if (meandatacode=0) rawmeancon= tmremncon.

if (meandatacode=0) rawsdcon= tmresdcon.

compute sigmacas=sqrt(ln(1+(rawsdcas/rawmeancas)**2)).

compute sigmacon=sqrt(ln(1+(rawsdcon/rawmeancon)**2)).

compute geomeancas=rawmeancas/exp(sigmacas*sigmacas/2).

compute geosdcas=exp(sigmacas).

compute geomeancon=rawmeancon/exp(sigmacon*sigmacon/2).

compute geosdcon=exp(sigmacon).

 

variable labels

 geomeancas 'geometric mean for cases'

/ geomeancon 'geometric mean for controls'

/ geosdcas 'geometric sd for cases'

/ geosdcon 'geometric sd for controls'.

 

Data and analyses

Comparison 1. Foods fortified with zinc alone versus same foods without added zinc.

Outcome or subgroup title	No. of studies	No. of participants	Statistical method	Effect size
1 Serum or plasma zinc (in µmol/L)	3	158	Mean Difference (IV, Random, 95% CI)	2.12 [1.25, 3.00]
2 Serum or plasma zinc (in µmol/L) (by duration of intervention)	3	158	Mean Difference (IV, Random, 95% CI)	2.12 [1.25, 3.00]
2.1 less than 6 months	2	99	Mean Difference (IV, Random, 95% CI)	2.46 [2.05, 2.87]
2.2 six months to one year	1	59	Mean Difference (IV, Random, 95% CI)	0.98 [‐0.02, 1.98]
3 Serum or plasma zinc (in µmol/L) (by type of food vehicle)	3	158	Mean Difference (IV, Random, 95% CI)	2.12 [1.25, 3.00]
3.1 Wheat flour or derivatives	2	99	Mean Difference (IV, Random, 95% CI)	2.46 [2.05, 2.87]
3.2 maize flour and corn meals or derivatives	1	59	Mean Difference (IV, Random, 95% CI)	0.98 [‐0.02, 1.98]
4 Serum or plasma zinc (in µmol/L) (by type of zinc compound)	3	158	Mean Difference (IV, Random, 95% CI)	2.12 [1.25, 3.00]
4.1 Zinc oxide	1	59	Mean Difference (IV, Random, 95% CI)	0.98 [‐0.02, 1.98]
4.2 Zinc sulphate	1	75	Mean Difference (IV, Random, 95% CI)	2.35 [1.93, 2.77]
4.3 Others	1	24	Mean Difference (IV, Random, 95% CI)	2.85 [2.01, 3.69]
5 Serum or plasma zinc (in µmol/L) (by dose of zinc added to the food)	3	158	Mean Difference (IV, Random, 95% CI)	2.12 [1.25, 3.00]
5.1 up to 10 mg/100 g	1	75	Mean Difference (IV, Random, 95% CI)	2.35 [1.93, 2.77]
5.2 more than 10 mg/100g	2	83	Mean Difference (IV, Random, 95% CI)	1.94 [0.10, 3.77]
6 Serum or plasma zinc (in µmol/L) (by development status of country)	3	158	Mean Difference (IV, Random, 95% CI)	2.12 [1.25, 3.00]
6.1 Low and middle income countries	2	99	Mean Difference (IV, Random, 95% CI)	2.46 [2.05, 2.87]
6.2 Others	1	59	Mean Difference (IV, Random, 95% CI)	0.98 [‐0.02, 1.98]
7 Stunting (author defined)	2	397	Risk Ratio (M‐H, Random, 95% CI)	0.88 [0.36, 2.13]
8 Underweight (author defined)	2	397	Risk Ratio (M‐H, Random, 95% CI)	3.10 [0.52, 18.38]
9 Pneumonia	1		Risk Ratio (M‐H, Random, 95% CI)	Totals not selected
10 Diarrhoea (as defined by authors)	1		Risk Ratio (M‐H, Random, 95% CI)	Totals not selected
11 All‐cause morbidity	1		Mean Difference (IV, Random, 95% CI)	Totals not selected
12 Haemoglobin	1		Std. Mean Difference (IV, Random, 95% CI)	Totals not selected
13 Adverse effect (iron status measured as serum ferritin in μg/L) (Ln transformed)	1		Mean Difference (IV, Random, 95% CI)	Totals not selected
14 Adverse effect (copper status as measured by serum or plasma copper level in μg/dL)	2	82	Mean Difference (IV, Random, 95% CI)	‐8.73 [‐18.03, 0.58]
15 Adverse effect (vomiting)	1		Risk Ratio (M‐H, Random, 95% CI)	Totals not selected
16 Weight (kg)	1		Mean Difference (IV, Random, 95% CI)	Totals not selected
17 Height or length (cm)	1		Mean Difference (IV, Random, 95% CI)	Totals not selected
18 Any adverse effect	1		Risk Ratio (M‐H, Random, 95% CI)	Totals not selected

1.9. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 9 Pneumonia.

1.10. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 10 Diarrhoea (as defined by authors).

1.11. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 11 All‐cause morbidity.

1.12. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 12 Haemoglobin.

1.13. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 13 Adverse effect (iron status measured as serum ferritin in μg/L) (Ln transformed).

1.15. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 15 Adverse effect (vomiting).

1.16. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 16 Weight (kg).

1.17. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 17 Height or length (cm).

1.18. Analysis.

Comparison 1 Foods fortified with zinc alone versus same foods without added zinc, Outcome 18 Any adverse effect.

Comparison 2. Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc.

Outcome or subgroup title	No. of studies	No. of participants	Statistical method	Effect size
1 Zinc deficiency (as defined by authors, depending on the age and gender)	1		Risk Ratio (M‐H, Random, 95% CI)	Totals not selected
2 Serum or plasma zinc (in µmol/L)	4	250	Mean Difference (IV, Random, 95% CI)	0.03 [‐0.67, 0.72]
3 Serum or plasma zinc (in µmol/L) (by duration of intervention)	4	250	Mean Difference (IV, Random, 95% CI)	0.03 [‐0.67, 0.72]
3.1 less than 6 months	4	250	Mean Difference (IV, Random, 95% CI)	0.03 [‐0.67, 0.72]
4 Serum or plasma zinc (in µmol/L) (by type of food vehicle)	5	309	Mean Difference (IV, Random, 95% CI)	0.36 [‐0.40, 1.12]
4.1 Wheat flour or derivatives	3	201	Mean Difference (IV, Random, 95% CI)	‐0.01 [‐1.01, 0.98]
4.2 maize flour and corn meals or derivatives	1	59	Mean Difference (IV, Random, 95% CI)	0.98 [‐0.02, 1.98]
4.3 Rice or derivatives	1	49	Mean Difference (IV, Random, 95% CI)	0.64 [‐0.78, 2.07]
5 Serum or plasma zinc (in µmol/L) (by type of zinc compound)	4	250	Mean Difference (IV, Random, 95% CI)	0.03 [‐0.67, 0.72]
5.1 Zinc oxide	2	149	Mean Difference (IV, Random, 95% CI)	‐0.31 [‐0.86, 0.24]
5.2 Zinc sulphate	1	34	Mean Difference (IV, Random, 95% CI)	0.31 [‐0.90, 1.52]
5.3 Others	1	67	Mean Difference (IV, Random, 95% CI)	3.99 [‐0.87, 8.85]
6 Serum or plasma zinc (in µmol/L) (by dose of zinc added to the food)	4	250	Mean Difference (IV, Random, 95% CI)	0.03 [‐0.67, 0.72]
6.1 up to 10 mg/100 g	3	183	Mean Difference (IV, Random, 95% CI)	‐0.24 [‐0.70, 0.22]
6.2 more than 10 mg/100g	1	67	Mean Difference (IV, Random, 95% CI)	3.99 [‐0.87, 8.85]
7 Serum or plasma zinc (in µmol/L) (by development status of country)	5	309	Mean Difference (IV, Random, 95% CI)	0.36 [‐0.40, 1.12]
7.1 Low and middle income countries	4	250	Mean Difference (IV, Random, 95% CI)	0.03 [‐0.67, 0.72]
7.2 Others	1	59	Mean Difference (IV, Random, 95% CI)	0.98 [‐0.02, 1.98]
8 Anaemia	2	137	Risk Ratio (M‐H, Random, 95% CI)	0.89 [0.35, 2.28]
9 Haemoglobin (g/dL)	3	186	Mean Difference (IV, Random, 95% CI)	0.13 [‐0.33, 0.59]
10 Adverse effect (iron status measured as serum ferritin in µg/L) (Ln transformed)	2	79	Mean Difference (IV, Random, 95% CI)	‐0.08 [‐0.35, 0.19]
11 Weight (24 to 59 mo)	1	34	Mean Difference (IV, Random, 95% CI)	0.02 [‐1.09, 1.13]
12 Height or length (24 to 59 mo)	1	34	Mean Difference (IV, Random, 95% CI)	‐0.44 [‐3.40, 2.52]

2.1. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 1 Zinc deficiency (as defined by authors, depending on the age and gender).

2.11. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 11 Weight (24 to 59 mo).

2.12. Analysis.

Comparison 2 Foods fortified with zinc plus other micronutrients versus foods fortified with other micronutrients without zinc, Outcome 12 Height or length (24 to 59 mo).

Characteristics of studies

Characteristics of included studies [ordered by study ID]

Aaron 2011.

Methods	Double‐blind randomised trial with factorial design (2 different doses of fortified zinc, 1 medicinal zinc and 1 control) (computer‐generated block randomisation) carried out from August 2009 to December 2009
Participants	144 apparently healthy men 18 years or older (71.5% zinc‐deficient) in a community clinic based in a low‐income urban neighbourhood in Dakar, Senegal with haemoglobin concentrations higher than 100 g/L, no history of chronic illnesses, no acute illnesses or medication use for 2 wk preceding the intervention, no use of vitamin or mineral supplements, and no consumption of commercially available zinc‐fortified foods. All men attending the screening sessions received a single 400 mg dose of the anthelmintic drug albendazole
Interventions	Participants were randomly assigned to 1 of 4 groups during 4 weeks: Group 1: (n = 39) received 200 g/day of wheat bread fortified with iron and folic acid (15 ppm elemental iron as ferrous fumarate and 1.5 ppm folic acid), but not zinc, and 10 mL liquid multivitamin supplement (containing 0.4 mg vitamin B6; 0.8 mg vitamin B12; 9.9 mg biotin; 29.7 mg vitamin C; 5.3 mg niacin; 0.4 mg riboflavin; 1.7 mg pantothenic acid; and 0.4 mg thiamin) between meals; Group 2 (n = 34) received 200 g/day of wheat bread fortified with iron and folic acid (15 ppm elemental iron as ferrous fumarate, 1.5 ppm folic acid, 0 ppm added zinc) and the same 10 mL multivitamin supplement with 15 mg zinc as zinc sulphate monohydrate added to be consumed between meals (containing 0.4 mg vitamin B6; 0.8 mg vitamin B12; 9.9 mg biotin; 29.7 mg vitamin C; 5.3 mg niacin; 0.4 mg riboflavin; 1.7 mg pantothenic acid; 0.4 mg thiamin; and 15 mg zinc) ; Group 3 (n = 36) received 200 g/day of wheat bread fortified with iron and folic acid and zinc as ZnO (15 ppm elemental iron as ferrous fumarate, 1.5 ppm folic acid, 63 ppm of zinc as zinc oxide) and the same 10 mL multivitamin supplement without zinc (containing 0.4 mg vitamin B6; 0.8 mg vitamin B12; 9.9 mg biotin; 29.7 mg vitamin C; 5.3 mg niacin; 0.4 mg riboflavin; 1.7 mg pantothenic acid; and 0.4 mg thiamin) ; or Group 4 (n = 35) received 200 g/day of wheat bread fortified with iron and folic acid and zinc (15 ppm elemental iron as ferrous fumarate, 1.5 ppm folic acid, 126 ppm of zinc as zinc oxide) and the same 10 mL multivitamin supplement without zinc (containing 0.4 mg vitamin B6; 0.8 mg vitamin B12; 9.9 mg biotin; 29.7 mg vitamin C; 5.3 mg niacin; 0.4 mg riboflavin; 1.7 mg pantothenic acid; and 0.4 mg thiamin). Breads were served with fruit jam or butter, depending on individual preference, and a non‐zinc–containing juice prepared from purified water and fruit juice concentrate. The liquid multivitamin (or multivitamin + zinc) supplement was given to participants after a 90‐min fast following the meal. We considered only groups 1, 3, and 4 for purposes of the comparisons in this review. Participants consumed under supervision the respective bread products during a morning meal
Outcomes	Change in plasma zinc at d 15 and 29 of the intervention, plasma concentrations of the acute phase proteins a1‐acid‐glycoprotein (AGP) and C‐reactive protein (CRP), self reported morbidity, cigarette smoking, anaemia
Notes	sex: males duration of intervention: less than 6 months type of food vehicle: wheat flour or derivatives type of zinc compound: zinc oxide dose of zinc added per 100 g of food: 3.75 or 7.5 mg zinc 100 g serving of bread (groups 3 and 4, respectively) development status of country: lower middle income Source of funding: Global Alliance for Improved Nutrition (Geneva, Switzerland) and the Michael and Susan Dell Foundation (Austin, USA). DSM Nutritional Products donated the fortificants, and Les Grands Moulins de Dakar provided the flour and prepared the breads for the study. Contact details: Corresponding author contacted for some clarifications. No response
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Low risk	Computer‐generated block randomisation with block size of 4
Allocation concealment (selection bias)	Unclear risk	No details reported
Blinding of participants and personnel (performance bias) All outcomes	Low risk	Double‐blind randomised clinical trial. Group assignments remained masked until all biochemical and statistical analyses were completed
Blinding of outcome assessment (detection bias) All outcomes	Low risk	Double‐blind randomised clinical trial. Group assignments remained masked until all biochemical and statistical analyses were completed
Incomplete outcome data (attrition bias) All outcomes	Low risk	10% attrition; 129 out of 144 completed the study. 100% compliance to product(s)
Selective reporting (reporting bias)	Low risk	No additional outcome recorded; authors could have reported proportion with zinc deficiency at end of intervention too
Other bias	Low risk	No other apparent bias. This trial was registered at ClinicalTrials.gov as NCT00944723

Badii 2012.

Methods	Double‐blind randomised clinical trial with factorial design (2 different doses of zinc fortification and 1 control)
Participants	80 zinc‐deficient (serum zinc ≤ 70 μg/dl) non‐pregnant, non‐lactating women aged 19 to 49 years from the staff and students of the Isfahan University of Technology, Iran. Exclusion criteria: Women had a known chronic disease, were participating in any similar programme, or were currently taking any vitamin or mineral supplements
Interventions	Participants were randomly assigned to 1 of 3 groups: Group 1 (n = 25) a non‐fortified taftoon bread daily; Group 2 (n = 25) received a low‐zinc (50 ppm of zinc sulphate‐fortified flour) taftoon bread; and Group 3 (n = 25) received a high‐zinc (100 ppm of zinc sulphate‐fortified flour) taftoon bread. After fermentation of the dough for 1.5 to 2 hours, the total mass was divided into equal portions, each about 200 g. Portions of this dough were fermented for 10 to 15 minutes, rolled into balls, flattened, and baked at 300°C for 90 seconds, following which the breads were put in individual labelled plastic containers until they were distributed among the participants. It was assumed that there was 130 g of flour in a loaf of taftoon bread, therefore each participant received 1 bread every day for 1 month and was controlled to ensure that the whole bread was consumed. The bread is prepared with 90% extraction wheat flour, salt, and active dried yeast
Outcomes	Serum zinc and iron were measured by atomic absorption before and after the study. Zinc and phytate content of prepared breads were also analysed
Notes	sex: females duration of intervention: less than 6 months type of food vehicle: wheat flour or derivatives type of zinc compound: zinc sulphate dose of zinc added per 100 g of prepared food: group 2 (low zinc): 5.72 mg per 100 g taftoon bread; group 3 (high zinc): 10.1 mg per 100 g taftoon bread development status of country: upper middle income Source of funding: Isfahan University of Technology provided financial support for this study. Contact details: Corresponding author contacted for some clarifications. No response
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Unclear risk	Participants were randomly divided into 3 groups, but method of sequence generation not mentioned
Allocation concealment (selection bias)	Unclear risk	Unclear allocation concealment
Blinding of participants and personnel (performance bias) All outcomes	Unclear risk	No details of blinding. Who was blinded remains unclear
Blinding of outcome assessment (detection bias) All outcomes	Unclear risk	Reported as "double‐blind" in title; no details thereafter
Incomplete outcome data (attrition bias) All outcomes	Low risk	Outcome recorded in 75 out of 80 women enrolled
Selective reporting (reporting bias)	Low risk	No apparent selective reporting
Other bias	Low risk	No other apparent bias

Haibin 2001.

Methods	Non‐randomised (allocation based on order of visit to hospital) clinical trial with factorial design
Participants	313 healthy women with their first pregnancy (mean (SD) age 24.7 (2.4) yrs) from Xinxiang City, Henan Province, China with no digestive system disease or pregnancy complications, and no history of taking calcium, iron, zinc, or other nutritionally fortified food
Interventions	Participants were allocated to 1 of 5 groups: Group 1 (n = 70): women received 3 biscuits per day; 8 g each, containing 400 IU vitamin D; Group 2 (n = 62): women received 3 biscuits per day; 8 g each, containing 400 IU vitamin D and 400 mg carbonate calcium; Group 3 (n = 64): women received 3 biscuits per day; 8 g each, containing 400 IU vitamin D, 400 mg carbonate calcium, and 10 mg lactate zinc; Group 4 (n = 57): women received 3 biscuits per day; 8 g each, containing 400 IU vitamin D, 400 mg carbonate calcium, 10 mg ferrous lactate, and 50 mg vitamin C; and Group 5 (n = 40): women received 3 biscuits per day; 8 g each, containing 400 IU vitamin D, 400 mg carbonate calcium, 10 mg ferrous lactate, 50 mg vitamin C, and 10 mg lactate zinc. The biscuits (3 biscuits per day; 8 g each) fortified with different nutritional elements were provided from the 5th month of gestation until delivery (24 weeks in total). Another group (n = 20) served as control. We only considered 2 comparisons for purposes of this review: group 3 versus group 2, and group 5 versus group 4. In both comparisons the difference is the addition of zinc
Outcomes	Women (before delivery): haemoglobin; anaemia rate; plasma calcium, iron and, zinc; RBC calcium, iron and zinc; bone mineral density Neonates: birthweight, birth height, z‐scores for weight‐for‐age, height‐for‐age, and weight‐for‐height; placenta weight, placental content of calcium, iron, and zinc; umbilical cord haemoglobin; umbilical cord plasma alkaline phosphatase, calcium, iron, and zinc; umbilical cord RBC calcium, iron, and zinc
Notes	sex: females duration of intervention: less than 6 months type of food vehicle: wheat flour or derivatives type of zinc compound: zinc sulphate dose of zinc added per 100 g of food: not reported but calculated to be approximately 40 mg development status of country: upper middle income Source of funding: No information available in translated version of the paper. Contact details: Authors could not be contacted because of lack of electronic contact details
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	High risk	Quote: "They were divided into 5 groups based on the order in which they visited hospital for antenatal care" Comment: No sequence generation
Allocation concealment (selection bias)	High risk	Quote: "They were divided into 5 groups based on the order in which they visited hospital for antenatal care" Comment: No allocation concealment
Blinding of participants and personnel (performance bias) All outcomes	High risk	Not mentioned Comment: Blinding unlikely in absence of allocation concealment
Blinding of outcome assessment (detection bias) All outcomes	High risk	Not mentioned Comment: Blinding unlikely in absence of allocation concealment
Incomplete outcome data (attrition bias) All outcomes	High risk	High (> 50%) attrition rates for most outcomes
Selective reporting (reporting bias)	Low risk	All outcomes reported
Other bias	Low risk	No other significant bias

Hambidge 1979.

Methods	Double‐blind clinical trial
Participants	96 healthy young children 33 to 90 months of age (mean age 58 mo) from a private preschool and kindergarten in Denver, United States of America
Interventions	Participants were assigned to 1 of 2 groups: Group 1: received either zinc‐fortified ready‐to‐eat cereal (3.75 mg of zinc (as zinc oxide) per 1 ounce serving), providing 25% United States Recommended Dietary Allowance per 1 ounce serving (test children); Group 2: received non‐zinc‐fortified ready‐to‐eat cereal (controls) for a 9‐month period. The test children were calculated on average to receive an additional 2.57 mg of zinc per day from this programme. The ready‐to‐eat cereals were provided to the families for consumption by the children in their homes. The mean number of servings consumed by the participants was 5.8 servings per week during the school year
Outcomes	Plasma zinc, hair zinc, urine zinc, saliva zinc, zinc‐dependent enzymes (alkaline phosphatase and erythrocyte carbonic anhydrase) serum vitamin A, serum copper, serum proteins
Notes	sex: mixed duration of intervention: 6 months to 1 year type of food vehicle: wheat flour or derivatives type of zinc compound: zinc oxide dose of zinc added per 100 g of food: 13.23 mg development status of country: high income Source of funding: National Institute of Arthritis, Metabolic and Digestive Diseases grant 2RO 1 AM12432‐10 and by Grant RR‐69 from General Clinical Research Centers Program of the Division of Research Resources, National Institutes of Health, USA. Kellogg Company provided the ready‐to‐eat cereals for this trial. Contact details: Authors could not be contacted because of lack of electronic contact details
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	High risk	There is no mention that this study is randomised. Quote: "Of the children (test group) 50% received zinc‐fortified cereals, the remainder (control group) received non zinc‐fortified cereals." Comment: Procedure of sequence generation not described
Allocation concealment (selection bias)	Unclear risk	No details Comment: Unclear allocation concealment
Blinding of participants and personnel (performance bias) All outcomes	Low risk	Quote: The study was designed and implemented as a double‐blind controlled investigation. Comment: Probably done
Blinding of outcome assessment (detection bias) All outcomes	Low risk	Quote: The study was designed and implemented as a double‐blind controlled investigation. Comment: Probably done
Incomplete outcome data (attrition bias) All outcomes	High risk	96 enrolled, 93 completed the study; plasma zinc results available only in 59 at the end of study
Selective reporting (reporting bias)	Low risk	All outcomes reported
Other bias	Low risk	Ready‐to‐eat cereals were supplied by Kellogg, but the study was funded by a government agency

Hettiarachchi 2004.

Methods	Randomised clinical trial with 4 arms
Participants	53 Sri Lankan schoolchildren aged 7 to 10 yrs recruited through public advertising. Children who had a chronic medical condition or were taking micronutrient supplements were excluded
Interventions	Participants received 75 g fortified rice flour/d (in form of a local dish 'halapa' made of 25 g of fortified rice) to be consumed for a period of 2 wk. Participants were randomly assigned to 1 of 4 groups: Group 1: rice flour fortified with 60 mg elemental iron/kg (as ferrous sulphate) and folic acid (2 mg/kg); Group 2: rice flour fortified with 60 mg elemental iron/kg (as ferrous sulphate), 385.08 mg/kg of Na2EDTA in dry powder form, and folic acid (2 mg/kg); Group 3; rice flour fortified with 60 mg elemental iron/kg (as ferrous sulphate), folic acid (2 mg/kg), and 60 mg of elemental zinc/kg (as zinc oxide powder); Group 4: rice flour fortified with 60 mg elemental iron/kg (as ferrous sulphate), 385.08 mg/kg of Na2EDTA in dry powder form kg, folic acid (2 mg/kg), and 60 mg of elemental zinc/kg (as zinc oxide powder). All study participants received oral mebendazole (100 mg twice daily for 3 consecutive days). For purposes of this review, we only considered 2 comparisons in which we could isolate the effect of zinc fortification: group 3 versus group 1, and group 4 versus group 2
Outcomes	Primary: iron and zinc absorption after 2 weeks of intervention Secondary: blood chemistry (Hb, serum ferritin, serum zinc, and serum folate) after 4 weeks of intervention
Notes	sex: mixed duration of intervention: 1 month type of food vehicle: rice type of zinc compound: zinc oxide dose of zinc added per 100 g of food: 6 mg development status of country: lower middle income Source of funding: International Atomic Energy Agency (IAEA contract SRL‐11958). The food grade minerals were supplied free of charge through Percy Ranasinghe of the Greenfields International, Sri Lanka. The provider of ferrous sulphate and zinc oxide was Paul Lohmann, Germany. The iron‐EDTA was provided by AkzoNobel, Netherlands and folic acid by Glaxo Wellcome. Contact details: Corresponding author was contacted for some clarifications. No response
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Unclear risk	Quote: "The subjects were randomly divided into 4 groups based on type of fortification and stratified by gender" Comment: Unclear sequence generation
Allocation concealment (selection bias)	Unclear risk	Quote: "The subjects were randomly divided into 4 groups based on type of fortification and stratified by gender" Comment: No details of allocation concealment
Blinding of participants and personnel (performance bias) All outcomes	Unclear risk	No details Comment: Unclear
Blinding of outcome assessment (detection bias) All outcomes	Unclear risk	No details Comment: Unclear
Incomplete outcome data (attrition bias) All outcomes	Low risk	Outcome presented for 49 out of 53 enrolled participants
Selective reporting (reporting bias)	Low risk	All outcomes presented
Other bias	Low risk	No other apparent bias

Kilic 1998.

Methods	Randomised trial with 2 arms
Participants	24 healthy 7‐ to 11‐year‐old children with asymptomatic zinc deficiency (defined as serum zinc < 65 µg/dL) and no evidence of acute or chronic illnesses in an elementary school in Ankara, Turkey
Interventions	Participants were randomly assigned to 1 of 2 groups: Group 1 (n = 12): received zinc‐fortified bread providing 2 mg/kg/day elemental zinc acetate for 90 days; Group 2 (n = 12): received the same quantity unfortified bread (no zinc added). The breads were made by the Food Engineering Department of Hacettepe University. The flour sample used in this trial contained 10.9% protein, 58% ash, 30.7% wet gluten, 10.2% dry gluten, and intrinsic zinc content of 6.7 ppm
Outcomes	Serum zinc, leukocyte zinc, haemoglobin, serum ferritin, serum copper, serum alkaline phosphatase, calcium, phosphorus, protein and albumin, cellular immune function
Notes	sex: mixed duration of intervention: less than 6 months type of food vehicle: wheat flour or derivatives type of zinc compound: zinc acetate dose of zinc added per 100 g of food: unable to estimate development status of country: upper middle income Source of funding: The breads were made by the Food Engineering Department of Hacettepe University, Ankara, Turkey. Three members of this department contributed to authorship. No other source of funding disclosed. Contact details: Corresponding author was contacted for some clarifications. No response
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Unclear risk	No details Comment: Unclear sequence generation
Allocation concealment (selection bias)	Unclear risk	No details Comment: Unclear allocation concealment
Blinding of participants and personnel (performance bias) All outcomes	Low risk	Quote: "The study was blind to investigators, parents, and teachers, who assured that the children ate all of the bread given to them at meals in school." "The two kinds of breads were indistinguishable in appearance and taste." "The students were unable to observe any difference in appearance between the two types of bread throughout the experiment." Comment: Probably done
Blinding of outcome assessment (detection bias) All outcomes	Low risk	Quote: "The study was blind to investigators, parents, and teachers, who assured that the children ate all of the bread given to them at meals in school." Comment: Probably done
Incomplete outcome data (attrition bias) All outcomes	Low risk	Information available for all 24 participants
Selective reporting (reporting bias)	Low risk	No apparent selective reporting
Other bias	Low risk	No other apparent bias

Lopez de Romaña 2005.

Methods	Clinical trial with factorial design
Participants	58 stunted, moderately anaemic children 3 to 4 yr of age, residing in a poor community on the periphery of Lima, Peru, and who were considered as at high risk for zinc deficiency
Interventions	Participants were randomly (block randomisation with block size of 3) assigned to 1 of 3 groups: Group 1 (n = 19) received in 2 meals/d (at breakfast and lunch), a total of 100 g wheat products fortified with 3 mg elemental iron (as ferrous sulphate) and no zinc; Group 2 (n = 19): received in 2 meals/d (at breakfast and lunch), a total of 100 g wheat products fortified with 3 mg elemental iron (as ferrous sulphate) and 3 mg elemental zinc (as zinc sulphate) per 100 g flour; Group 3 (n = 20): received in 2 meals/d (at breakfast and lunch), a total of 100 g wheat products fortified with 3 mg elemental iron (as ferrous sulphate) and 9 mg elemental zinc (as zinc sulphate) per 100 g flour. The intervention lasted 70 days
Outcomes	Fractional absorption of zinc (FAZ) was measured on days 2–3 and 51–52; meal‐specific absorption of zinc was calculated as the product of FAZ and zinc intake Haemoglobin, serum ferritin, plasma zinc on day 70
Notes	sex: mixed duration of intervention: less than 6 months type of food vehicle: wheat flour or derivatives type of zinc compound: zinc sulphate dose of zinc added per 100 g of food: 3 and 9 mg development status of country: upper middle income Source of funding: The Bill & Melinda Gates Foundation, the Fogarty International Center, and the National Institute of Child Health and Human Development of the US National Institutes of Health (research grant D43 TW01267). Contact details: Senior author (Kenneth H Brown) of the paper was contacted to provide clarification about units of central tendency and units of variability for serum ferritin values. Authors provided us with corrected values, which we included in this systematic review. Authors also provided clarifications related to randomisation and blinding
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Low risk	Quote: "were randomly assigned to receive..." Clarification from author: The children were stratified by sex and then randomly assigned to 1 of 3 groups using a block randomisation technique with fixed block length of 3. Comment: Adequate
Allocation concealment (selection bias)	Low risk	Quote: "were randomly assigned to receive..." Clarification from author: The randomisation list was received by an investigator in Peru who was not directly involved in the study. Comment: Adequate
Blinding of participants and personnel (performance bias) All outcomes	Low risk	No details in published paper Clarification from author: The study diets were masked to the participants and investigators, as well as the laboratory analysts. Comment: Adequate blinding
Blinding of outcome assessment (detection bias) All outcomes	Low risk	No details in published paper Clarification from author: The study diets were masked to the participants and investigators, as well as the laboratory analysts. Comment: Adequate blinding
Incomplete outcome data (attrition bias) All outcomes	High risk	Final results available for only 34 out of 58 enrolled participants. No details about missing cases
Selective reporting (reporting bias)	Low risk	No apparent selective reporting
Other bias	Low risk	No other apparent bias

Sanchez 2014.

Methods	Cluster‐randomised trial with 3 arms Randomisation unit: child day‐care centres
Participants	301 children between 2 and 5 years of age from 6 child day‐care centres in Medellin, Colombia managed by a foundation (Fundacion FAN) who attend 8 hours daily, received more than 80% of their daily food intake from the childcare, and liked milk. Exclusion criteria: children with a medical diagnosis of recurrent pneumonia, cystic fibrosis, digestive malformations, any‐cause persistent diarrhoea, inflammatory bowel disease, lactose intolerance, and children who at the time of initiation of the intervention had respiratory infection or acute diarrhoeal illness diagnosed by physician as well as children who had received zinc supplements within 2 months before enrolment.
Interventions	Participants were assigned to 1 of 3 groups depending on their care centre. 2 centres were considered a conglomerate and allocated to: Group 1: received milk fortified with elemental zinc (as zinc sulphate); Group 2 received milk fortified with elemental zinc (as zinc amino chelate); Group 3: received unfortified milk. The dose of elemental zinc varied by age of the participants. Children 2 to 3 received 7 mg zinc daily, and children 4 to 5 received 9.45 mg elemental zinc. The milk was prepared by Nutreo (Rionegro, Colombia) and did not differ in colour, odour, or taste. The daily dose was delivered in 2 doses, 1 at breakfast and another snack, Monday to Friday for 16 weeks. The laboratory Nutreo produced the fortified and unfortified milks each month and delivered them to the appropriate care of children's centre also monthly, masking allocation. The baseline prevalence of zinc deficiency was not measured in this study
Outcomes	Diarrhoea episodes, acute respiratory infection, adverse effects, vomiting, abdominal pain, weight, height, weight‐for‐age z‐scores, weight‐for‐height z‐score, height‐for‐age z‐score, and body mass index for age
Notes	sex: mixed duration of intervention: less than 6 months type of food vehicle: milk/dairy products type of zinc compound: zinc sulphate and zinc amino chelate dose of zinc added per 100 g of food: unable to estimate development status of country: upper middle income Source of funding: Universidad CES (academia) and Nutreo, S.A.S., Rionegro, Antioquia (private sector). Contact details: We contacted the first author of the paper (Juliana Sánchez) to provide additional details related to some of the outcomes of this review, and we received the details. However, regarding another query related to further clarification of adjustment for cluster, we received no response
Risk of bias
Bias	Authors' judgement	Support for judgement
Random sequence generation (selection bias)	Low risk	Randomly assigned through ballot, 6 clusters so that 2 children's centres were in each of the 3 study groups. One group received zinc sulphate; another group received zinc amino acid chelate; and the third group received placebo
Allocation concealment (selection bias)	Low risk	The laboratory Nutreo produced the fortified and unfortified milks each month and delivered them to the appropriate care of children's centre also monthly, masking allocation. The 3 milks were similar in colour, smell, and taste.
Blinding of participants and personnel (performance bias) All outcomes	Low risk	The study was triple blind throughout the procedure and analysis of information
Blinding of outcome assessment (detection bias) All outcomes	Low risk	The study was triple blind throughout the procedure and analysis of information
Incomplete outcome data (attrition bias) All outcomes	Low risk	17 (4.76%) children were lost to follow‐up, and 39 were excluded from the final analysis after ingesting milk for less than 75% of the observation time defined by the protocol. The analysis was thus done with 301 children: 93 of them took milk fortified with zinc amino acid chelate, 112 milk fortified with zinc sulphate, and 96 received placebo. This represents about 16% attrition rate
Selective reporting (reporting bias)	Low risk	Stratum variables, type of family and housing tenure were controlled during analysis and not found to act as potential confounding variables when comparing the crude risk ratio and adjusted risk ratio. ClinicalTrials.gov identifier: NCT01791608
Other bias	Low risk	No other apparent bias

Hb: haemoglobin

IU: international units
 RBC: red blood cell
 SD: standard deviation

Characteristics of excluded studies [ordered by study ID]

Study	Reason for exclusion
Aaron 2011b	This double‐blind, placebo‐controlled trial determined the effects of a multimicronutrient beverage on biochemical and anthropometric indicators of nutritional status among schoolchildren participating in a pilot school feeding program in Nasarawa State, Nigeria. Children received 1 of 2 interventions 5 d/wk during school hours: 1) 250 mL/d of a multimicronutrient beverage that included vitamin A, iron, and zinc (micronutrient); or 2) an isoenergetic control beverage (control). At baseline, 566 children 5 to 13 yrs old were randomised to groups (micronutrient: n = 288; control: n = 278). Height, weight, haemoglobin, and serum concentrations of C‐reactive protein, ferritin, retinol, and zinc were measured at baseline and at the end of the study. A total of 270 children in the micronutrient group and 264 children in the control group completed the study. Self reports of vomiting increased in both groups at 6 mo, however the prevalence tended to be greater in the micronutrient group (21%) compared to the control group (14%) (P = 0.06). Biochemical changes were greater in the micronutrient group compared to control for serum retinol (0.1060.02 mmol/L vs 0.0260.02 mmol/L; P = 0.016) and zinc (1.060.2 mmol/L vs 0.660.2 mmol/L; P = 0.031). The intervention did not significantly affect haemoglobin or serum ferritin concentrations. The study was excluded as the beverage was fortified with iron and vitamin A in addition to zinc
Abrams 2003	This study evaluated the efficacy of a fortified (12 micronutrients) beverage in improving the nutritional status of children in Botswana. 311 lower‐income urban schoolchildren, age 6 to 11 yrs in 2 primary schools were screened. Children were given seven 240 mL servings weekly of either an experimental beverage fortified with 12 micronutrients or an isoenergetic placebo drink for 8 weeks. Weight, mid‐upper arm circumference, haemoglobin, retinol, ferritin, vitamin B12, folate, and riboflavin status were measured at baseline and at the end of the study. Plasma zinc and serum transferrin receptors were also measured at study end. A total of 145 children in the experimental group and 118 in the control group completed the trial. The changes in mid‐upper arm circumference, weight for age, and total weight were significantly better in the experimental group than in the control group (P < 0.01). Ferritin, riboflavin, and folate status were significantly better in the experimental group at study end (P < 0.01), but serum vitamin B12 was not. Zinc was significantly higher, and transferrin receptors were significantly lower at the conclusion of the study in the experimental group (P < 0.001). Mean plasma retinol concentrations, which were low (0.7 mol/L) in both groups, did not change. The study was excluded as the beverage was fortified with many other micronutrients in addition to zinc
Angeles‐Agedeppa 2011	This study determined the effects of a non‐carbonated fortified juice drink on the iron, zinc, and nutritional status of schoolchildren in San Juan, Manila, Philippines. 100 randomly selected anaemic children were randomly allocated into 2 groups in a doubly masked, placebo‐controlled manner: Group 1 received the fortified juice and Group 2 received the non‐fortified juice for 100 days, 5 days a week under strict supervision. The juice drink was fortified with vitamin A, zinc, iron, vitamin C, and lysine. The non‐fortified juice was fortified only with vitamin C. All children were dewormed prior to the intervention. The basal prevalence of anaemia was significantly reduced in both the fortified group (100% to 13%) and the non‐fortified group (100% to 40%) at end line. The mean plasma ferritin levels were similar in both groups at baseline and end line. At end line, mean plasma zinc in the fortified group significantly increased by 20 μg/dL from a baseline value of 83.9 μg/dL to 103.9 μg/dL, while the non‐fortified group remained at similar levels with baseline. Basal weight and height significantly increased among all children at end line. The study was excluded as juice was fortified with other micronutrients in addition to zinc
Arsenault 2007	This study, from low‐income peri‐urban communities of Peru, compared energy intake, reported appetite, and body composition of 6‐ to 8‐mo‐old Peruvian children with stunting (length‐for‐age z‐score (LAZ) < ‐0.5 SD) who were randomly assigned to receive daily for 6 mo: 1) 3 mg/d zinc in a liquid supplement; 2) 3 mg/d zinc in a fortified porridge (150 mg zinc/kg dry weight); or 3) no extra zinc in either the supplement or porridge. There were no group‐wise differences in changes in dietary energy intakes or body composition or in the prevalence of reported poor appetite. However, among children with an initial LAZ less than the median (‐1.1 SD), those who received zinc as a liquid supplement had a 0.41 kg greater increase in fat‐free mass than those who did not receive zinc (P < 0.05). The authors concluded that daily provision of 3 mg of supplemental zinc did not affect energy intake or reported appetite. Among children with initial mild‐to‐moderate stunting, those who received the zinc supplement had a greater increase in fat‐free mass than those who did not receive additional zinc, and zinc supplements may be more efficacious than the same dose of zinc provided in fortified food. The study was excluded as the intervention was performed in children below two years of age
Ash 2003	In this randomised, double‐blind, placebo‐controlled efficacy trial, children were assigned to receive a fortified beverage (10 micronutrients) or an unfortified beverage at school for 6 mo. There were non‐significant differences at baseline between children in the fortified and non‐fortified groups in iron status, serum retinol, and anthropometry. At the 6‐mo follow‐up, among children with anaemia (haemoglobin < 110 g/L), there was a significantly larger increase in haemoglobin concentration in the fortified group than in the non‐fortified group (9.2 and 0.2 g/L, respectively). Of those children who were anaemic at baseline, 69.4% in the non‐fortified group and 55.1% in the fortified group remained anaemic at follow‐up (Risk ratio 0.79), a cure rate of 21%. The prevalence of children with low serum retinol concentrations (< 200 g/L) dropped significantly from 21.4% to 11.3% in the fortified group compared with a non‐significant change (20.6% to 19.7%) in the non‐fortified group. At follow‐up, mean incremental changes in weight (1.79 compared with 1.24 kg), height (3.2 compared with 2.6 cm), and BMI (0.88 compared with 0.53) were significantly higher in the fortified group than in the non‐fortified group. This study was excluded as the beverage was fortified with multiple micronutrients
Bardosono 2009	This study evaluated the effect of milk supplementation enriched with iron and zinc on indicators of growth, physical capacity, and cognitive performance in underweight schoolchildren from Jakarta and Surakarta, Indonesia. A total of 245 underweight schoolchildren (7 to 9 yrs) were randomly allocated to receive milk fortified with iron and zinc (n = 121) or milk not fortified with iron and zinc (n = 124) daily for 6 months. There was no difference in haemoglobin and serum ferritin increase between groups. Decrease in serum zinc was also found in both groups. There was a significantly higher increase in body weight and higher improvements in the proportions of underweight children in the fortified milk group as compared to the non‐fortified milk group. There was a significant improvement in cognitive performance, i.e. coding test‐score, but no significant difference in physical work capacity between groups. The study was excluded as the milk was fortified with iron in addition to zinc
Bates 1993	In this study, 110 rural Gambian children aged between 0.57 and 2.30 years were divided into 2 matched groups: the placebo group received a fruit‐flavoured drink shown to contain negligible amounts of Zn, and the supplemented group received the same drink with 70 mg added Zn, initially as zinc acetate, but subsequently as zinc gluconate (since compliance was better). The drinks were given twice weekly under supervision for 15 months. Growth and mid‐upper arm circumference were measured at weekly intervals throughout the study, and illnesses were monitored. Capillary blood and urine samples were collected at 0, 2, and 8 weeks. Body weights and arm circumferences showed a linear increase, plus a seasonal effect (rainy season faltering). There was no significant overall effect of the supplement for body weight. A very small (2%) but significant (P < 0.01) difference favoured the supplemented group for arm circumference. Zinc supplementation also did not affect the immunological markers. The trial was excluded as age of included children was 0.57 to 2.3 years, and majority were below 2 years of age. Moreover, the fortification/supplementation was at point‐of‐use
Chen 2008	This study evaluated different combinations of nutrient‐fortified diets to improve blood levels of micronutrients in preschool children (2 to 6 yrs) in Chongqing, China. 226 children from nurseries were randomly assigned to 3 groups: Group 1: vitamin A; Group 2: vitamin A plus iron; and Group 3: vitamin A plus iron, thiamine, riboflavin, nicotinamide, folic acid, zinc, and calcium. The fortified diets were given for 6 months; blood levels of micronutrients were measured at the beginning and end of the study period. The study was excluded as the foods were fortified with other micronutrients in addition to zinc
Costarelli 2014	This study evaluated the efficacy of consumption of zinc‐fortified skimmed milk for a period of 2 months in comparison to standard non‐fortified milk on some biochemical parameters, zinc status, inflammatory/immune response, and on a key parameter of the T cell‐mediated immunity (thymulin hormone) in healthy very old (age > 82 y) people. 21 participants from a nursing home in central Italy were enrolled. Elderly with degenerative age‐related diseases (cancer, severe infections, diabetes, severe cardiopathies, and neurodegenerative diseases) were excluded from the study. In the reported cross‐over design, participants received 250 mL of zinc‐fortified (12 mg/L of zinc gluconate) milk or unfortified milk, each for a period of 60 days. A control group, for comparison of clinical events during the follow‐up period (1 year), comprised 23 people who did not participate in the cross‐over study. No significant differences existed in plasma zinc (Zn) and copper (Cu) before and after treatment with zinc‐fortified milk. The study was excluded as there was no randomisation for deciding intervention at baseline; all participants in the cross‐over design received unfortified milk simultaneously, followed by zinc‐fortified milk. Zinc levels were not collected from the participants before and after each intervention arm to be considered for controlled before‐after study. For clinical outcomes, the control group was decided on the basis of not participating in the main study design
de Romaña 2009	This was a conference abstract related to review of zinc fortification strategies
Ebrahimi 2006	In this randomised controlled trial from Yasuj city of Iran, 804 schoolchildren (age 8 to 11 yrs) were randomised to receive daily supplements of zinc or placebo, 6 days/wk for 7 months. Anthropometry was performed at enrolment, and at monthly intervals thereafter. Zinc supplementation resulted in a significantly better increase in weight and height in comparison to placebo. The study was excluded as zinc was given as a medicine (syrup) and not as a fortificant
Evans 1992	This is a conference abstract of the published study Bates 1993, which we excluded as the majority of included children were below 2 years of age, and fortification/supplementation was at point‐of‐use
Feng 1993	382 healthy children aged between 2 and 6 yrs were chosen (185 males and 195 females) at Longfeng Chang Xi Kindergarten, Daqing City, China with a low zinc status (hair zinc < 100 ug/g) were randomly divided into 3 groups: Group 1 (n = 91) were put in a low‐zinc control group; Group 2 (n = 97) received a zinc supplement; and Group 3 (n = 88) received a zinc and calcium supplement. A group 4 (n = 106) were children with a high‐zinc status (hair zinc ≥ 100 mg/g) chosen for a high‐zinc control group. The children took half of their meals at home and the other half in kindergarten. As food was purchased collectively, the food consumed in each family was similar. This study on supplementation through tablets is outside the scope of this review
Golden 1981	In this study from Jamaica, 16 children were evaluated 4 to 12 wk after admission for severe malnutrition. During recovery, 10 children were given a high‐energy soy protein‐based diet containing 52 micromil/L of zinc and 1.33 mmol phytic acid per litre, and 6 children were given a cow’s milk protein‐based formula diet containing 69 micromol/L zinc. When the children's rate of weight gain decreased steadily as they approached the expected weight for a normal child of the same height, the diets were supplemented with medicinal zinc. Zinc acetate solution (15 mM) was added to the feed 4 times daily, to supply an additional 25 to 150 micromol zinc/kg body weight per day. The study was excluded as it was a non‐randomised comparison, zinc supplements were given to both groups, and the feeds were supplemented with medicinal zinc at point‐of‐use
Haibin 2001b	This publication reported information on placental weight, birthweight, birth length, and haemoglobin of infants born to mothers who received foods fortified with zinc (along with other micronutrients) during pregnancy. The primary study reporting outcomes related to women's health, Haibin 2001, has been included in this review
Herman 2002	This study aimed to examine the effect of zinc fortification of flour on iron bioavailability or the optimum form of zinc supplementation. Supplementation was in the form of wheat flour dumplings containing 25 g flour fortified with 60 mg Fe/kg, either alone or with 60 mg Zn/kg as zinc oxide or zinc sulphate. 90 children aged 4 to 8 yrs were recruited from a rural outreach clinic in Situ Udik (a small village around 70 km from Jakarta, Indonesia) and assigned randomly to the 3 groups; 86 completed the study. Iron and zinc absorption were measured with established stable isotope methods. Iron absorption from the flour fortified with iron only was good (15.9 ± 6.8%), but when corrections were made for haemoglobin concentrations, it was significantly lower from the flour cofortified with zinc sulphate (11.5 ± 4.9%; P < 0.05) but not from the flour cofortified with zinc oxide (14.0 ± 8.9%). Zinc absorption was not significantly different between the zinc oxide and zinc sulphate cofortified flours (24.1 ± 8.2% compared with 23.7 ± 11.2%; P = 0.87). The study was excluded because outcomes were only related to iron and zinc absorption. The outcomes of interest were not measured after intervention
Huo 2011	This study was designed to evaluate the effectiveness of fortified flour (vitamins A, B1, B2, niacin, folic acid, iron, and zinc in mg/kg at 2, 3.5, 3.5, 35, 1, 20, and 25, respectively) on micronutrient status in poor rural adult women from Weichang, a county in Hebei province, located in the northwest of China. A total of 4700 farmers as the intervention group were supplied with multinutrient‐fortified wheat flour for 3 years (2004 to 2007), while 2750 farmers as the control group were supplied with unfortified wheat flour. Blood samples were taken at baseline and annually from about 300 volunteer adult females aged 20 to 60 years in each group. Haemoglobin, serum retinol, serum iron, free erythrocyte protoporphyrin, and serum zinc were measured annually and a dietary survey conducted every 6 months. Intervention groups showed a significant increase in terms of Hb levels from 24 mo to 36 mo, and anaemia rate decreased from 15.1% at baseline to 10.8% at 36 mo. Serum iron levels of the intervention group significantly increased from 12 mo to 36 mo, and erythrocyte protoporphyrin decreased from 24 mo to 36 mo. Serum retinol and serum zinc of intervention group improved significantly from 12 mo to 36 mo compared with baseline and the control group. The results showed that the fortified flour could improve micronutrient status of adult females in a poor rural region. The study was excluded as wheat flour was fortified with multiple micronutrients in addition to zinc
Hyder 2007	This study evaluated the effect of a multiple‐micronutrient‐fortified beverage on haemoglobin (Hb) concentrations, micronutrient status, and growth among adolescent girls in rural Bangladesh. A total of 1125 girls enrolled in a randomised, double‐blind, placebo‐controlled trial and were allocated to either a fortified or non‐fortified beverage of similar taste and appearance. The beverage was provided at schools 6 d/wk for 12 mo. The fortified beverage increased the Hb and serum ferritin and retinol concentrations at 6 mo (P = 0.01). Adolescent girls in the non‐fortified beverage group were more likely to suffer from anaemia, iron deficiency, and low retinol concentrations. The fortified beverage group had greater increases in weight, mid‐upper arm circumference, and BMI over 6 mo (P = 0.01). Consuming the beverage for an additional 6 mo did not further improve the Hb concentration, but the serum ferritin level continued to increase. The study was excluded as the beverage was fortified with multiple micronutrients
Islam 2013	The study participants were non–breastfed children 36 to 59 mo of age who were recruited from a low‐income, peri‐urban community near Dhaka, Bangladesh. The study was designed to permit within‐child comparisons of zinc absorption from mixed diets containing either high‐zinc rice cultivar or conventional rice, with or without added zinc, by using the dual stable isotope tracer ratio technique. The study was excluded because the intervention lasted only for 4 days, and the outcomes were only related to zinc absorption
Khoshgoftarmanesh 2010	This study from Iran investigated the effects of fortification of 2 different flours with 60 ppm Fe, 60 ppm Fe plus 2 ppm folic acid, 60 ppm Fe plus 30 ppm Zn, and 30 ppm Zn. Formulations of iron and Zn were ferrous sulphate and zinc sulphate, respectively. Fortification treatments had significant effects on Zn and Fe concentrations in the flour, dough, and bread. Addition of Zn to the flour significantly (P < 0.05) decreased the molar ratio of phytic acid to Zn in bread. Fortification of flour with ferrous sulphate impaired the colour, taste, and overall acceptability of Khabbazi bread. The study was excluded as evaluated outcomes were sensory characteristics and acceptability; the outcomes of interest were not evaluated
Mahloudji 1975	This study investigated the effects of supplementation of the diets of 6‐ to 12‐year‐old children in a village near Shiraz in Iran with zinc plus iron or iron alone, together with the indispensable amino acids in the form of egg white protein, vitamins, minerals, and corn oil. 75 children attending the school in Kherak, a village of about 900 inhabitants located 55 km west of Shiraz, were divided into 3 groups of 25 each. Group 1 received, in addition to their regular diet, 20 mg of elemental zinc as carbonate together with 20 mg of iron as ferrous fumarate, vitamin, mineral, egg white and corn oil supplements. Group 2 received 20 mg of iron as ferrous fumarate together with the same supplement as Group 1, except for zinc. Group 3 received placebo capsules containing lactose and simulated supplement. The liquid supplement had corn oil, egg protein, starch, and multiple micronutrients besides traces of zinc that were also present in the control supplement. The study was excluded as zinc supplements (intervention) were provided in medicinal form along with liquid dietary supplement containing vitamins, minerals, proteins, fats, and traces of zinc
Makola 2003	This study evaluated the effect of a micronutrient‐fortified beverage containing 11 micronutrients (iron, iodine, zinc, vitamin A, vitamin C, niacin, riboflavin, folate, vitamin B12, vitamin B6, and vitamin E) on the haemoglobin, iron, and vitamin A status of pregnant women in Tanzania. A group of 259 pregnant women with gestational ages of 8 to 34 wk were enrolled in a randomised double‐blind controlled trial in which study they received 8 wk of supplementation. The supplement resulted in a 4.16 g/L increase in haemoglobin concentration and a 3 g/L increase in ferritin and reduced the risk of anaemia and iron deficiency anaemia by 51% and 56%, respectively. The risk of iron deficiency was reduced by 70% among women who had iron deficiency at baseline and by 92% among those who had adequate stores. The study was excluded as the beverage was fortified with other micronutrients in addition to zinc
Manger 2008	This randomised controlled trial of 569 children (5.5 to 13.4 yrs), from 10 schools in poorest subdistricts of northeast Thailand, compared the efficacy of a seasoning powder fortified with or without 5 mg Fe, 5 mg Zn, 50 g I, and 270 g vitamin A per serving consumed with a school lunch 5 d/wk. The intervention had no statistically significant effect on anthropometric measures over 31 wk, but reduced the incidence of respiratory‐related illnesses (rate ratio (RR) 0.83, 95% confidence interval (CI) 0.73 to 0.94), symptoms of runny nose (RR 0.80, 95% CI 0.70 to 0.92), cough (RR 0.80, 95% CI 0.66 to 0.96), and diarrhoea (RR 0.38, 95% CI 0.16 to 0.90). For the visual recall test, children in the fortified group recalled 0.5 more items (95% CI 0.1 to 0.9) than the controls. There were no statistically significant differences between groups in the results of the digits forward and backward tests or in school grades at the conclusion of the 2 semesters. The study was excluded as seasoning powder was fortified with other micronutrients in addition to zinc
Mendez 2012	This controlled trial included 108 schoolgirls (12 to 18 years old) from northwest Mexico, randomly assigned to receive 500 mL/day of micronutrient‐fortified milk (n = 53) in addition to regular diet or only regular diet (n = 55). Age, weight, and height were measured at the beginning of the study. Food intake by the 24‐hour recall method and plasma zinc levels assessed by absorption spectrophotometry were determined before and after 27 days of fortified milk intake. Plasma zinc improved in the girls receiving fortified milk (116.6 ± 26.9 μg/dL; P < 0.01) compared with the control group (98.5 ± 26.6 μg/dL). The study was excluded as the fortified milk had other micronutrients (iron, folic acid, vitamins, etc.) in addition to zinc
NCT01481181	An efficacy trial of a gravity‐fed household water treatment device as a delivery system for zinc in zinc‐deficient children and women from low‐income settings Methods: Randomised controlled trial Participants: 270 zinc‐deficient Kenyan children aged 2 to 5 years. The study population living in an area with low‐zinc status and sharing the same unimproved water supply will be included in the trial and randomly assigned to 1 of the treatment groups for 12 months. Interventions: All households will receive hygiene practice recommendations. In addition, participants will be assigned to 1 of 3 groups: Group 1 will receive the point‐of‐use water filtration system called LifeStraw® Family device with the zinc delivery system; Group 2 will receive hygiene practice recommendations and a point‐of‐use water filtration system called LifeStraw® Family without zinc delivery system, and Group 3 will receive hygiene practice recommendations. Outcomes: At baseline, midpoint, and endpoint, anthropometrics (weight, height, mid‐upper arm circumference) and 7 mL whole blood will be collected from preschool children for determination of serum zinc, C‐reactive protein, haemoglobin, and serum ferritin. Children and adult participants remaining zinc deficient at completion of the trial will receive zinc supplements. Rates of diarrhoea and growth in preschool children will be compared in three groups. Starting date: August 2011 Contact information: Michael Zimmermann, ETH Zurich, Wageningen University Sponsors and collaborators: Wageningen University, Maseno University, School of Public Health, Kenya. The study was excluded as zinc was provided through the water treatment device
NCT01790321	Water‐based zinc efficacy trial in Beninese schoolchildren. Double‐blind, randomised controlled trial with 2 arms. 278 school‐age (5 to 10 yrs) children enrolled in a primary school equipped with a water pump, in the rural area of the commune of Natitingou, Benin. Children with severe anaemia (Hb < 70 g/L), who consume a supplement or dietary supplement containing zinc, use drugs that affect the metabolism of zinc, or are suffering from a chronic disease affecting the metabolism of zinc were excluded. Participants were randomly assigned to 1 of 2 groups: Group 1 consumed a defined quantity of pump‐water outside of meals that was previously filtered and zinc‐fortified (zinc + filter, delivering 2.8 mg Zn/day over the entire study period); Group 2: children received water that was filtered only (filter, 0 mg Zn/day). Zinc fortification and filtration of water was achieved through a gravity‐fed point‐of‐use filter by Vestergaard S.A. (LifeStraw® Family). Primary outcomes: zinc status, zinc concentration in plasma, and prevalence of zinc deficiency Secondary outcomes: occurrence of diarrhoea episodes; measurement of height and weight for the assessment of height‐for‐age, weight‐for‐age, and BMI‐for‐age z‐scores. Results: Plasma zinc in the zinc + filter group over the study period was significantly higher than in the filter group (P = 0.006). Prevalence of Zn deficiency in the zinc + filter group over the study period was significantly lower than in the filter group (P = 0.046). There was no significant time by treatment effect on diarrhoeal morbidity or growth indices. The study was excluded as water (and not food) was used as the vehicle for zinc fortification
Nga 2009	This randomised, double‐blind, placebo‐controlled trial conducted among 510 primary schoolchildren (6 to 8 yrs) evaluated the efficacy of school‐based intervention using multimicronutrient–fortified biscuits with or without deworming on anaemia and micronutrient status in Vietnamese schoolchildren. Non‐fortified or multimicronutrient–fortified biscuits including iron (6 mg), zinc (5.6 mg), iodine (35 mg), and vitamin A (300 mg retinol equivalents) were given 5 d/wk for 4 mo. Multimicronutrient fortification significantly improved concentrations of haemoglobin (11.87 g/L, 95% confidence interval (CI) 0.78 to 2.96), plasma ferritin (17.5 mg/L, 95% CI 2.8 to 12.6), body iron (10.56 mg/kg body weight, 95% CI 0.29 to 0.84), plasma zinc (10.61 mmol/L, 95% CI 0.26 to 0.95), plasma retinol (10.041 mmol/L, 95% CI 0.001 to 0.08), and urinary iodine (122.49 mmol/L, 95% CI 7.68 to 37.31). Fortification reduced the risk of anaemia and deficiencies of zinc and iodine by 40%. The study was excluded as biscuits were fortified with other micronutrients in addition to zinc
Ohiokpehai 2009	This experimental study examined the effect of corn‐soy‐fortified complementary food on serum zinc levels among primary schoolchildren (6 to 9 years) in 3 schools of Suba district, Kenya. Children in 2 schools (Mbita and Sindo) were fed with corn‐soy blend for 3 months. The fortified complementary food provided 5.0 mg of zinc per 100 g. Children in a third school (Ong’ayo primary) were the control and were not fed with the corn‐soy blend. Assessments of serum zinc were performed before (n = 156) and after (n = 138) 3 months of feeding. At baseline, nearly all (95.7%) children were found to be deficient, with low serum zinc (< 10.7 μm/L). There was a significant reduction (P = 0.0421) in the number of zinc‐deficient cases to 70.2% after feeding for 3 months on corn‐soy blend, with the mean serum zinc having improved from 8.4 to 10.2 μm/L (P = 0.002). The study was excluded as it was a non‐randomised controlled pretest, post‐test design with 2 intervention but only 1 control cluster
Olivares 2013	This study assessed the effect of Zn on Fe absorption from bread prepared with wheat flour fortified with Fe and graded levels of Zn fortificant. 12 non‐pregnant women aged 30 to 43 years from Santiago, Chile received on 4 different days, after an overnight fast, 100 g of bread made with wheat flour (70% extraction) fortified with 30 mg Fe/kg as ferrous sulphate (A) or prepared with the same Fe‐fortified flour but with graded levels of Zn, as zinc sulphate: 30 mg/kg (B), 60 mg/kg (C), or 90 mg/kg (D). Fe radioisotopes of high specific activity were used as tracers, and Fe absorption iron was measured by the incorporation of radioactive Fe into erythrocytes. The geometric mean and range of ±1 SD of Fe absorption were: A = 19.8% (10.5% to 37.2%), B = 18.5% (10.2% to 33.4%), C = 17.7% (7.7% to 38.7%), and D = 11.2% (6.2% to 20.3%). The authors concluded that Fe is well absorbed from low‐extraction flour fortified with 30 mg/kg of Fe, as ferrous sulphate, and up to 60 mg/kg of Zn, as Zn sulphate. A statistically significant reduction of Fe absorption was observed at a Zn fortification level of 90 mg Zn/kg. The study was excluded as evaluated outcomes were only related to absorption; outcomes of interest not evaluated
Osei 2009	This study assessed the effectiveness of micronutrient fortification of meals cooked and fortified at school on anaemia and micronutrient status of schoolchildren in Himalayan villages of India. In this placebo‐controlled, cluster‐randomised study, 499 schoolchildren (6 to 10 yrs) received either multiple micronutrients (treatment group) or placebo (control group) as part of school meals (6 d/wk) for 8 mo. Both groups were dewormed at the beginning of the study. The micronutrient premix provided 10 mg iron, 375 mg vitamin A, 4.2 mg zinc, 225 mg folic acid, and 1.35 mg vitamin B12 for each child per day. Blood samples drawn before and after the intervention were analysed for haemoglobin, ferritin, retinol, zinc, folate, and vitamin B12. Postintervention, fewer children in the treatment group had lower serum retinol (odds ratio (OR) 0.57, 95% confidence interval (CI) 0.33 to 0.97) and folate (OR 0.47, 95% CI 0.26 to 0.84) than the control group. The serum vitamin B12 concentration decreased in both groups, but the magnitude of change was less in the treatment than in the control group (P < 0.05). Total body iron increased in both groups, however the change was greater in the treatment than in the control group (P < 0.05). The study was excluded as the intervention consisted of multiple micronutrients in addition to zinc. Moreover, fortification was at point‐of‐use in the form of a premix powder
Osendarp 2007	These were 2 randomised double‐blind trials using 2‐by‐2 factorial design conducted in Adelaide, South Australia and at 6 primary schools in Jakarta, Indonesia. A total of 396 children (aged 6 to 10 yrs) in Australia and 384 children in Indonesia were randomly allocated to receive a drink with a micronutrient mix (iron, zinc, folate, and vitamins A, B6, B12, and C), with docosahexaenoic acid (DHA, 88 mg/d) and eicosapentaenoic acid (EPA, 22 mg/d), or with both or placebo 6 d/wk for 12 mo. Biochemical indicators were determined at baseline and 12 mo. Cognitive performance was measured at baseline, 6 mo, and 12 mo. The micronutrient treatment significantly improved plasma micronutrient concentrations in Australian and Indonesian children. DHA + EPA treatment increased plasma DHA and total plasma n‐3 fatty acids in both countries. The micronutrient treatment resulted in significant increases in scores on tests representing verbal learning and memory in Australia (estimated effect size 0.23, 95% confidence interval (CI) 0.01 to 0.46). A similar effect was observed among Indonesian girls (estimated effect size 0.32, 95% CI ‐0.01 to 0.64), but not in boys (estimated effect size ‐0.04, 95% CI: ‐0.38 to 0.29). No effects were found on tests measuring general intelligence or attention. The study was excluded as the intervention included other nutrients in addition to zinc. Moreover, the fortification was done at point‐of‐use using powders
Rameshwar Sarma 2006	This double‐blind, placebo‐controlled, matched‐pair, cluster‐randomised study in semi‐urban middle‐income residential schoolchildren (6 to 16 yrs) evaluated the effect of a micronutrient‐fortified beverage on growth and morbidity in apparently healthy schoolchildren. After 14 mo of supplementation, there was a significant increase in mean increments of height and weight z‐scores of 0.04 and 0.02, respectively, in the supplemented group compared with 0.14 and 0.09 in the placebo group. Velocity of weight (3.56 versus 3.00) was significantly (P < 0.01) higher with supplementation. Although there were no differences in the incidence of common childhood diseases such as fever, cough and cold, diarrhoea, and ear infections, the mean duration of illness (calculated per person per year) was significantly shorter in the supplemented group (5.0 d) than in the placebo group (7.4 d). The study was excluded as the beverage was fortified with multiple micronutrients
Sazawal 2007	This community‐based, double‐blind randomised controlled trial, from a peri‐urban settlement in North India, evaluated the efficacy of milk fortified with specific multiple micronutrients (providing additional 7.8 mg zinc, 9.6 mg iron, 4.2 g selenium, 0.27 mg copper, 156 g vitamin A, 40.2 mg vitamin C, 7.5 mg vitamin E per day) on morbidity in children compared with the same milk without fortification. 633 children (1 to 3 yrs) were randomly allocated to receive fortified milk (n = 316) or control milk (n = 317) for 1 year. Mean number of episodes of diarrhoea per child was 4.46 (SD 3.8) in the intervention (fortified milk) group and 5.36 (SD 4.1) in the control group. Mean number of episodes of acute lower respiratory illness was 0.62 (SD 1.1) and 0.83 (SD 1.4), respectively. The fortified milk reduced the odds for days with severe illnesses by 15% (95% confidence interval (CI) 5% to 24%), the incidence of diarrhoea by 18% (95% CI 7% to 27%), and the incidence of acute lower respiratory illness by 26% (95% CI 3% to 43%). The study was excluded as milk was fortified with other micronutrients in addition to zinc
Sazawal 2013	This double‐masked randomised controlled trial was conducted in 4 primary schools of Northern Bangladesh. 1010 children from classes 1 to 4 (age 6 to 9 years) were randomly allocated to receive either micronutrient‐fortified yoghurt (FY, n = 501) or non‐fortified yoghurt (NFY, n = 509). For 1 year, children were fed with 60 g yoghurt every day providing 30% recommended dietary allowance (RDA) for iron, zinc, iodine, and vitamin A. Anthropometric measurements and blood/urine samples were collected at base‐, mid‐, and end‐line. Children in the FY group showed improvement in Hb (mean difference (MD) 1.5, 95% confidence interval (CI) 0.4 to 2.5; P = 0.006) as compared to the NFY group. Retinol‐binding protein (MD 0.05, 95% CI 0.002 to 0.09; P = 0.04) and iodine levels (MD 39.87, 95% CI 20.39 to 59.35; P < 0.001) decreased between base‐ and end‐line, but the decrease was significantly less in the FY group. Compared to the NFY group, the FY group had better height gain velocity (MD 0.32, 95% CI 0.05 to 0.60; P = 0.02) and height‐for‐age z‐scores (MD 0.18, 95% CI 0.02 to 0.33; P = 0.03). There was no difference in weight gain velocity, weight‐for‐age z‐scores, or BMI z‐scores. The study was excluded as yoghurt was fortified with other micronutrients in addition to zinc
Schlesinger 1992	This study from nutrition recovery centres of Santiago, Chile, evaluated the effect of zinc supplementation in marasmic infants during nutritional rehabilitation using a controlled double‐blind design in which 19 infants fed a zinc‐fortified formula (15 mg/L) were compared with 20 infants fed the same non‐supplemented formula. Evaluation of immunocompetence, growth, and zinc, copper, and iron status was performed on admission and at 30, 60, and 105 d of nutritional rehabilitation. Zinc‐supplemented infants had significantly higher linear growth gain, and their immune function improved as demonstrated by conversion of their delayed hypersensitivity skin reactions, enhanced lymphoproliferative response to phytohaemagglutinin, and increased salivary immunoglobulin A concentrations. The study was excluded as the intervention consisted of fortified milk formula involving children less than 2 years of age
Thankachan 2012	This randomised, double‐blind, controlled trial determined the efficacy of extruded rice grains fortified with multiple micronutrients on the prevalence of anaemia, micronutrient status, and physical and cognitive performance in 6‐ to 12‐year‐old low‐income schoolchildren in Bangalore, India. 258 children were assigned to 1 of 3 intervention groups to receive rice‐based lunch meals fortified with multiple micronutrients with either low‐iron (6.25 mg) or high‐iron (12.5 mg) concentrations or identical meals with unfortified rice. The meals were provided 6 d/wk for 6 mo. After 6 mo, plasma vitamin B12 and homocysteine concentrations (both P < 0.001) as well as physical performance (P < 0.05) significantly improved in the intervention arms. No between‐group differences were observed in haemoglobin concentration, anaemia, and deficiencies of other micronutrients or cognitive function after 6 mo, but paired analyses revealed a small reduction in anaemia prevalence in children in the low‐iron group. This study was excluded as rice was fortified with multiple micronutrients
Villalpando 2006	This study assessed the efficacy of whole cow's milk fortified with ferrous gluconate and zinc oxide, along with ascorbic acid, in reducing the prevalence of anaemia and improving iron status of low‐income children 10 to 30 mo of age. Healthy children were randomly assigned to drink 400 mL/d of whole cow's milk, either fortified milk (FM) with 5.8 mg/400 mL of iron as ferrous gluconate, 5.28 mg/400 mL of zinc as zinc oxide, and 48 mg/400 mL of ascorbic acid, or non‐fortified milk (NFM) with 0.2 mg iron/400 mL, 1.9 mg zinc/400 mL, and 6.8 mg ascorbic acid/400 mL. Haemoglobin, serum ferritin, soluble transferrin receptors (TfR), and C‐reactive protein concentrations were measured at baseline and 6 mo after intervention. The prevalence of anaemia declined from 41.4 to 12.1% (P < 0.001), or 29 percentage points, in the FM group; there was no change in the NFM group. Haemoglobin (coefficient = 0.22, P < 0.01) was positively and TfR (coefficient = ‐0.29, P < 0.001) negatively associated with treatment, controlling for children's respective baseline values, age, and gender. Treatment with FM was negatively associated with the likelihood of being anaemic after 6 mo of intervention.  This study was excluded as the milk was fortified with iron and vitamin C in addition to zinc. Moreover, the fortification was at point‐of‐use using premix powder
Walravens 1976	This double‐blind controlled study determined the effects of supplementing a milk formula with iron and zinc (4 mg/L). By 6 months of age, mean growth increments for the supplemented male infants were 2.1 cm greater in length (P < 0.025) and 535 g greater in weight (P < 0.05) than for male controls. Growth increments for female test and control infants did not differ significantly. Plasma zinc levels at 3 months of age were significantly higher for both male and female supplemented infants. By 6 months, only the male supplemented infants maintained significantly higher plasma zinc levels (P < 0.025). The addition of zinc was associated with a lower incidence of disturbed gastrointestinal function (P < 0.005) and not accompanied by any signs of toxicity. The study was excluded as the intervention involved fortification of milk formula used in neonates
Walravens 1983	This double blind, pair‐matched controlled study was conducted in 40 children with low growth percentiles in order to evaluate the effect of zinc on growth velocity. Participants were low‐income Spanish‐American children, 2 to 6 yrs of age with heights below the 10th percentile and nutritional or biochemical evidence of zinc deficiency. Test children were assigned to receive 5 mg zinc (as ZnSO4) twice daily in 5 mL of cherry‐flavoured syrup. The control children received the placebo syrup alone. After 1 yr, the mean height velocity of the zinc‐supplemented children was slightly, but significantly (P < 0.005), greater than that of the control children. This effect was primarily due to a greater height achievement of the zinc‐supplemented boys. Increases in height‐for‐age z‐scores were also significant for the supplemented males (P < 0.001) and for the combined sexes (P < 0.05). The study was excluded as zinc was provided in medicinal form (syrup)
Winichagoon 2006	This study evaluated the efficacy of a micronutrient‐fortified seasoning powder served with a school lunch on reducing anaemia and improving the micronutrient status of children in rural northeastern Thailand. Children (n = 569) aged 5.5 to 13.4 yrs from 10 schools were randomly assigned to receive a seasoning powder either unfortified or fortified with zinc (5 mg), iron (5 mg), vitamin A (270 mg), and iodine (50 mg) (per serving) and incorporated into a school lunch prepared centrally and delivered 5 d/wk for 31 wk. For the primary outcome, anaemia (based on haemoglobin), no intervention effect was apparent (odds ratio (OR) 1.02, 95% confidence interval (CI) 0.69 to 1.51) after adjustment for design strata. The odds of zinc (based on serum zinc) and urinary iodine deficiency in the fortified group were 0.63 (95% CI 0.42 to 0.94) and 0.52 (95% CI 0.38 to 0.71) times those in the unfortified group, respectively. Fortification had no effect on deficiencies of serum retinol (OR 0.61, 95% CI 0.25 to 1.51), ferritin (OR 1.12, 95% CI 0.43 to 2.96), or mean red cell volume (OR 1.16, 95% CI 0.82 to 1.64). The study was excluded as the seasoning powder was fortified with other micronutrients in addition to zinc
Xiuwen 1993	Randomised trial in 382 children (age 2 to 6 yrs) from Longfeng Chang Xi Kindergarten, Daqing City, China. Children were divided into low‐zinc or high‐zinc group based on their hair zinc content. Low‐zinc group were randomly divided into 3 groups: 91 in a low‐zinc control group, 97 in a zinc supplement group, and 88 in a zinc and calcium supplement group. 106 children with high‐zinc status (hair zinc ≥ 100 mg/g) were chosen for a high‐zinc control group. For the zinc supplement group, a 2 mg/kg/d zinc sulphate tablet was used. The trial was excluded as it involved medicinal zinc supplementation in the form of a tablet

Hb: haemoglobin
 BMI: body mass index
 SD: standard deviation

Characteristics of ongoing studies [ordered by study ID]

Moretti 2014.

Trial name or title	The effect of zinc on iron bioavailability from extruded rice fortified with ferric pyrophosphate
Methods	Randomised trial
Participants	20 women 18 to 45 years of age with body mass index in the range of 18.5 to 25, serum ferritin level < 15 µg/L, and body weight < 65 kg
Interventions	Women will be randomly assigned to 1 of 3 groups: Group 1: rice fortified with ferric pyrophosphate and zinc oxide (ZnO); Group 2: rice fortified with ferrous sulphate (ZnSO4); Group 3: rice fortified with ferric pyrophosphate and zinc sulphate. Iron‐fortified rice will be administered to all women, and women will act as their own controls
Outcomes	Change in isotopic ratio of iron in blood at 2, 4, and 6 weeks Haemoglobin, plasma ferritin, and C‐reactive protein at 16th, 30th, and 44th day
Starting date	December 2014
Contact information	Laura S Hackl +41 44 632 87 40 Swiss Federal Institute of Technology laura.hackl@hest.ethz.ch Dr Diego Moretti +41 44 632 84 36 Swiss Federal Institute of Technology diego.moretti@hest.ethz.ch
Notes	Sponsors and collaborators: Swiss Federal Institute of Technology and PATH

Differences between protocol and review

We were unable to access Food Science and Technology Abstracts (FSTA), and therefore did not search this database.

We dropped the comparison 'Food fortified with zinc plus other micronutrients versus no intervention', as it would not have resulted in estimation of effects due to zinc fortification. This comparison was mentioned in the protocol but was not included in the review. Moreover, we did not find any study providing data for this comparison.

For definitions of underweight and stunting, we used 'author defined' rather than less than ‐2 SD.

For units of haemoglobin, we used g/dL rather than g/L.

Contributions of authors

Dheeraj Shah drafted an initial protocol with technical input from Harshpal S Sachdev, Tarun Gera, Luz Maria De‐Regil, and Juan Pablo Peña‐Rosas. Luz Maria De‐Regil, Harshpal S Sachdev, and Juan Pablo Peña‐Rosas developed the methods of the protocol. Dheeraj Shah and Tarun Gera independently extracted all data using a tailored and pretested data extraction form. All review authors provided input into and contributed to drafting the final version of the protocol.

Disclaimer: Juan‐Pablo Peña‐Rosas is full‐time staff member of the World Health Organization. The review authors alone are responsible for the views expressed in this publication, and they do not necessarily represent the decisions, policy, or views of the World Health Organization.

Sources of support

Internal sources

• Evidence and Programme Guidance, Department of Nutrition for Health and Development, World Health Organization, Switzerland.

• Micronutrient Initiative, Canada.

External sources

• Evidence and Programme Guidance, Department of Nutrition for Health and Development, World Health Organization, Switzerland.

• Bill & Melinda Gates Foundation, USA.

  WHO acknowledges the financial support from the Bill & Melinda Gates Foundation for the development of systematic reviews of the evidence on the effects of nutrition‐specific and nutrition‐sensitive interventions

• Micronutrient Initiative, Canada.

  WHO acknowledges the financial support from the Micronutrient Initiative to the Evidence and Programme Guidance, Department of Nutrition for Health and Development for the development of systematic reviews of the evidence on the effects of nutrition‐specific and nutrition‐sensitive interventions

Declarations of interest

Dheeraj Shah ‐ none
 Harshpal S Sachdev ‐ none
 Tarun Gera ‐ none
 Luz Maria De‐Regil ‐ none
 Juan Pablo Peña‐Rosas ‐ none

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