Evaluation of vitamin D medicines and dietary supplements and the physicochemical analysis of selected formulations

Abstract

Context

Vitamin D is purported to offer wide ranging and numerous health benefits leading to increased interest from manufacturers of medicines and dietary supplements. Elderly patients frequently require vitamin D supplementation due to reduced sun exposure and dietary intake. There are ever increasing numbers of vitamin D formulations in the global market. However, due to a lack of regulatory restrictions for some of these products the quality of these dosage forms can be of some concern.

Objectives

To study vitamin D formulations available in the global market and evaluate physic-chemical properties of selected formulations from the New Zealand market.

Method

The first component of this study consisted of a search for different vitamin D formulations available in selected countries. The second component of the study involved assaying selected vitamin D formulations available in New Zealand. Vitamin D was extracted from capsule, tablet and emulsion formulations and quantified using a validated High Performance Liquid Chromatography (HPLC) method.

Results

Of the 14 analysed formulations, only 60% were within 100+10 % of the label claim. The two registered, prescription formulations available exhibited vitamin D levels of 90+4% and 97±2% of the labeled amount, while non-registered, non-prescription dietary supplements had vitamin D levels ranging from 8±2% to 201±29% of the labeled amount.

Conclusions

Dietary supplements do not require strict regulation and showed a large variation in the percentage label claim of vitamin D. Prescription formulations which are more strictly regulated gave content values within standard acceptance ranges. Vitamin D has proven health benefits and also the potential to cause harm, therefore there is a need for tougher regulations of dietary supplements to ensure acceptable quality.

Introduction

Globally, interest in vitamin D has grown dramatically in recent years due to the promising health benefits it has displayed for a range of conditions (1). Manufacturers of medicines and dietary supplements have viewed this as an immense opportunity with many formulations available in the market.

Vitamin D is a fat-soluble vitamin that can be obtained through dietary means and is transformed in the skin to its active form following exposure to UV light. Vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol), the two most prevalent forms of the vitamin, once activated act as a regulator of mineral and skeletal homeostasis (2., 3., 4., 5.). Increased vitamin D intake has shown many benefits such as decreasing a patient’s risk of osteoporosis, rheumatoid arthritis, multiple sclerosis, cancer and diabetes (6). Specific health claims of vitamin D have recently been commented on by the European Food Safety Agency (EFSA). A cause and effect relationship can be established between vitamin D intake (800 IU or 20 µg (40 IU = 1 µg) daily from all sources) and a reduction in falls in men and women over 60 years (7). In addition a cause and effect relationship was established between vitamin D and calcium intake and a reduction in the loss of bone mineral density (BMD) in women 50 years and over (8). Both a reduction and falls and a reduction in BMD loss lower the risk of bone fracture.

Vitamin D plays an important role in preventing and treating a range of chronic diseases in the elderly (9). However, the elderly have a reduced capacity to activate vitamin D in the skin, frequently coupled with less UV light exposure due to lifestyle changes (9, 10). In addition, dietary intake of vitamin D is often reduced in the elderly (10). The UK National Diet and Nutrition Survey found increasing prevalence of vitamin D insufficiency with increasing age. As a result, vitamin D is one of the most common supplements administered to the elderly population (10).

Vitamin D2 (ergocalciferol) and vitamin D3 (cholecalciferol) are practically insoluble in water, freely soluble in alcohol and soluble in fatty oils (11). These are sensitive to air, heat and light allowing a reversible isomerisation to pre-vitamin D to take place in solution depending on temperature and time (12). As a result of these sensitivities, stability is an important issue to address with vitamin D formulations.

In New Zealand, vitamins are managed under the 1985 Dietary Supplements Regulations as part of the Food Act 1981 (13). Dietary supplements have fewer restrictions placed on them than registered medicines. These restrictions include labelling requirements, highest permissible daily dose and the inability to make healing claims. The maximum permitted daily dose of vitamin D is 1000 IU (25 µg) as a dietary supplement for adults (13). Since current food regulations do not provide assurance of product quality and efficacy, New Zealand consumers have to rely entirely on the manufacturer (14). Any product containing more than 1000 IU of vitamin D is classified as a medicine and must be registered with the New Zealand Medicines and Medical Devices Safety Authority, Medsafe.

In contrast, Australia regulates all vitamins as ‘low risk medicines’ under the Therapeutics Goods Act 1989 (15). This ensures the quality and safety of therapeutic goods to consumers. The Therapeutic Goods Administration regulates the licensing and auditing of manufacturers, pre-market assessment of products, reporting of adverse reactions and testing in the laboratory (15). Thus there is a more stringent pre-market approval of vitamins in Australia compared to New Zealand. In the United Kingdom, dietary supplements are not usually considered medicinal products, and thus, do not require a marketing authorisation for sale in the UK (16). Vitamins are usually only subject to the Food Safety Act 1990 and the Food Labelling Regulations 1996 (16). The Dietary Supplements Health and Education Act of 1994 controls dietary supplements in the United States (17). Dietary supplements do not require pre-approval from the Food and Drug Administration (FDA) before they enter the market because they are considered as a category of foods and not as drugs (18). However the FDA has implemented a current Good Manufacturing Practice (cGMP) policy to ensure dietary supplements are of good quality (19). In Europe, Directive 2002/46/EC guides the use of Vitamin D as a food supplement (20). Food supplements are regulated as food, however if claims are made for the treatment, cure or prevention of disease then the supplements would be regulated as medicines. The Scientific Committee of Food (SCF) and European Food Safety Agency (EFSA) were directed to provide scientific opinion on tolerable upper levels of vitamins to “be contained within foodstuffs and proposed a maximum safety level for Vitamin D of 1400 IU or 35 µg (21).

The objectives of this study were to identify vitamin D formulations available in the global market and to evaluate selected products in the New Zealand market for vitamin D content, and compare to the label claim.

Methods

Market research

The availability of vitamin D products in different countries was determined by performing an Internet search. This consisted of utilising web based search engines such as Google, using the keywords ‘vitamin D3’, ‘vitamin D2’, ‘ergocalciferol’, ‘cholecalciferol’ or ‘vitamin D’. Websites including regulatory agencies and online pharmacies were investigated. The markets in India, Japan, Canada, United States of America, United Kingdom, Malaysia, Australia and New Zealand were explored in-depth as products could be identified online. These countries were expected to contain representative formulations from the major pharmaceutical companies.

To narrow down the global market search, a criterion of formulations not exceeding three actives was implemented. In addition to searching online, the New Zealand market was investigated by visiting various pharmacies, health food shops and supermarkets.

Product selection

A comprehensive selection criteria was used to shortlist the products for the quantitative analysis from the New Zealand market. The product selection criteria included formulations containing vitamin D3, strength of ≥ 500 IU and containing no more than three active ingredients. For the procured products there was a difference in time between the date of manufacture and the date of testing. However, all products were within their expiry date and therefore expected to meet expiry specifications of 90±10 %, which is a commonly accepted range in the pharmaceutical industry.

Sample analysis

Analytical method development

Working Standards

Standard solutions of vitamin D3 (provided by API Pharmaceuticals) in the range of 1 ppm to 100 ppm were prepared for the analysis of samples. A stock standard solution of 100 ppm strength (100 mg/L) was prepared. Individual standards at a concentration of 1 mg/L (1 ppm) to 50 mg/L (50 ppm) were prepared using the 100 ppm stock standard solution. These standards as well as the 100 ppm standard solution were used to prepare the standard calibration curve.

100% methanol was used as diluent for the analysis of tablets, hard gelatine capsules and emulsion. A mixture of 5 mL n-hexane and 95 mL of 2-propanol was used for testing the soft gelatine capsules.

HPLC Method

All analytical work was carried out in AnQual, a Good Laboratory Practices (GLP) compliant analytical facility using validated protocols. Reverse phase HPLC was performed for the quantification of vitamin D3 in the sample solutions. A Shimadzu Prominence series HPLC (Shimadzu Scientific Instruments) comprising an autosampler maintained at 4°C (SIL-20A/C), a quaternary pump and UV detector at 265 nm was used. Data acquisition was by LC Solutions software (Shimadzu). A Waters 5 micron Symmetry column (C18, 150 mm x 3.9 mm) maintained at 25°C was operated under isocratic conditions at 67 bar and a flow rate of 0.5 ml per minute. The injection volume was 20 µL, the sample run time was 40 minutes with vitamin D3 eluting at 25 minutes. The mobile phase comprised 82 % acetonitrile, 13 % methanol and 5 % milliQ water.

Formulation Analysis

Multiple dosage forms for vitamin D were investigated i.e. tablets, hard and soft gelatine capsules, and emulsion, therefore different extraction methods were used. A generalised method is described here.

All formulations were analysed in triplicate under sodium light to avoid any light induced degradation. The mass of the sample that provides the approximate final theoretical sample solution concentration was calculated and taken for analysis. For each sample replicate, the sample mass/volume wastransferred to a volumetric flask. The sample solution was then sonicated for 30 minutes to achieve complete extraction of vitamin D3 followed by cooling to room temperature for fifteen minutes. Once the sample solution reached room temperature the volumetric flask was made up to volume and the aliquot of supernatant was transferred to the HPLC vial for subsequent analysis.

Since all vitamin D3 formulations were different and each had different label claims, individual calculation formula was used for the determination of percentage of label claim. It was decided that a percentage of label claim of 100 ± 10 % was deemed acceptable.

Results and Discussion

Market research

The majority of formulations found in the global market contain vitamin D3. Other formulations contain vitamin D2, calcitriol and alfacalcidol. A number of studies have been carried out comparing vitamin D3 and D2, with vitamin D3 shown to be more efficacious (22).

Globally there are more non-prescription formulations of vitamin D3 available compared to prescription only. In New Zealand, 29 formulations were found to be non-prescription, in comparison to only 2 prescription products.

The recommended daily intake for all age groups up to the age of 70 which is required to prevent deficiency is approximately 600 IU and 800-1000 IU for those without adequate sun exposure (23). This is in accordance with the majority of formulations found in New Zealand, Australia, UK, Canada, India and Malaysia. However, the recommended dietary intake increases when an adult has a comorbid medical condition such as hyperparathyroidism or osteomalacia (23). The dose of vitamin D needed to meet these requirements is greater than 1000 IU. The different strengths of vitamin D3 available are divided into categories and represented in Figure 1. In Japan and the US the majority of products contain more than 1000 IU. In New Zealand, only registered prescription medications contain 1000 IU or more due to the maximum permitted dose set by the Food Act 1981 (13).

Figure 1.

Strengths of vitamin D3 formulations available in different countries. Total number of formulations per country; New Zealand = 31, Australia = 33, USA = 48, UK = 29, Japan = 79, Canada = 58, India = 40 and Malaysia = 15. Note, 40 IU = 1 µg

Tablets are the main dosage form of vitamin D3 found in Australia, UK, Canada and India. Conversely, in New Zealand, USA and Japan the most common dosage forms are capsules, followed by tablets. Other dosage forms found include chewable tablets, powder, emulsion drops, sublingual formulations, oral sprays and effervescent tablets. Research has also shown that vitamin D is most stable when it is dissolved in fats or oils, giving reason to why capsules were one of the main dosage forms found (24).

Sample analysis

The analysis of vitamin D3 formulations involved selecting 14 products widely available in the New Zealand market and in addition, a reference vitamin D3 formulation and analysing their percentage of label claim. In this study, six tablet formulations, two hard gelatine capsules, six soft gelatine capsules and one emulsion were analysed. Of the 14 products selected from the New Zealand market, 2 were registered prescription medicines while 12 weren’t registered as medicines. No registered non-prescription medicines met the selection criteria. A percentage of label claim within 100 ± 10% was deemed acceptable.

The average percentage of label claim for all the 15 analysed formulations from the New Zealand market are graphically presented in Figure 2. The percentages of label claims of these formulations vary significantly. The two registered formulations; TAB1 and TAB2 produced 90 ± 4% and 97 ± 2% respectively. Overall the registered formulations were considered to have acceptable levels. Out of the remaining non-registered formulations 6 formulations were far from the acceptable range. These were TAB5, TAB6, HGC1, SGC1, SGC4 and EML1 at 29 ± 11%, 201 ± 29%, 8 ± 2%, 21 ± 8%, 156 ± 6%, and 133 ± 9% respectively. The rest of the non-registered formulations were within the acceptable range.

Figure 2.

Average percentage of label claim for various formulations; TAB = tablet, SGC = soft gellatin capsule, HGC1 = hard gellatin capsule and EML = emulsion (error bars represents standard deviation, n=3). Dark grey bars represent formulations within 100 ± 10% of the label claim, black bars formulations above 110% and light grey bars formulations under 90%

The acceptable results found for the two registered prescription products TAB1 and TAB2 were predicted as registered medications in New Zealand are tightly regulated under the Medicines act 1981. Due to stringent regulations, registered medications must constantly and consistently meet the requirements specified by the law. Before a medicine is approved by Medsafe, the pharmaceutical company must give clear reports of the tests that are put into place to ensure the strength, quality, purity and safety of the medicine. If a medication fails to meet these standards the regulators can suspend any consent given to the pharmaceutical company to sell or supply the medicine (25).

Conversely, there was large variation of percentage label claim for the non-registered products (8 ± 2% to 201 ± 29%). In new Zealand non-registered products are not required to undergo a strict pre-market approval (13). The lack of pre-market restrictions indicates there are no tests put into place to guarantee the strength and quality of the active ingredients. HGC1, SGC1 and TAB5 had a substantially lower percentage label claim of 8 ± 2%, 21 ± 8% and 29 ± 11% respectively. Conversely, EML1, SGC4 and TAB6 had a much higher percentage label claim of 133 ±9%, 156 ± 6% and 201 ± 29 respectively. These products have the potential to cause harm through toxic side effects, which may occur unwittingly as the label claim is well over 100%. According to Section 10 of the Food Law 1981, no person shall sell a product that has a false label claim (13). Any person who contravenes this law has committed an offence and is subsequently heavily penalized (13). Non-registered products that contain more than 1000 IU of vitamin D3 are also in violation of the new Zealand Food (Supplemented Food) Standard 2010 which states that the maximum concentration of vitamin D3 is 1 000 IU (26). Therefore, to ensure that dietary supplements are of acceptable quality, there needs to be a greater enforcement of regulations on these types of formulations.

Conclusion

The increasing interest in vitamin D due to its numerous health benefits has led to an increased interest from manufacturers of medicines and dietary supplements. Based on the global market research undertaken it was found that the majority of formulations contained the D3 form of the vitamin as the key active ingredient (22). In the US and Japan the majority of formulations contained over 1000 IU, while in new Zealand, australia, UK, Canada, india and Malaysia the majority of formulations contained under 1000 IU. Globally, there were more non-prescription formulations of vitamin D3 compared to prescription formulations. In most of the countries investigated non-prescription products are less tightly regulated than prescription products leading to questions over quality for the bulk of formulations.

not all formulations available in the new Zealand market contained the specified amount of vitamin D3. It was found that registered, prescription formulations were within the acceptable range of 100 ± 10%, whereas, unregistered formulations had a much wider variability in regards to percentage of label claim (8 ± 2% to 201 ± 29%). There is a greater need for tighter regulations when it comes to unregistered dietary supplements in new Zealand to ensure formulations comply with their label claims. Vitamin D has proven health benefits, however, also the potential to cause harm through toxicity or lack of efficacy, therefore there is a need for stricter regulations for all vitamin D formulations to ensure acceptable quality and safety.