Different electrolytic treatments for food sanitation and conservation simulating a wash process at the packinghouse

Abstract

Microorganisms are predominantly responsible for food deterioration, necessitating the sanitization and removal of these entities from food surfaces. The packinghouse employs free chlorine in the sanitization process; however, free chlorine's propensity to react with organic matter, forming potentially toxic compounds, has led to its restriction or outright prohibition in several European countries. Therefore, this study aims to assess various washing methods, emulating packinghouse conditions, utilizing diverse forms of electrolyzed water to impede microbial proliferation and significantly enhance the food's shelf life. The subject of investigation was cherry tomatoes. The findings revealed that electrolyzed water containing NaCl exhibited superior efficacy compared to electrolysis with Na₂SO₄. Both forms of electrolyzed water demonstrated noteworthy effectiveness in inhibiting microorganisms, resulting in a reduction of 2.0 Log CFU mL⁻¹ for bacteria and 1.5 Log CFU mL⁻¹ for fungi. The electrolyzed water also exhibited a comparable capability to free chlorine in removing fecal coliforms from the tomato surfaces. Notably, both electrolyzed water treatments extended the shelf life of cherry tomatoes by at least three days, accompanied by minimal or negligible residues of free chlorine. Consequently, the electrolyzed water formulations proposed in this study present themselves as promising alternatives to traditional packinghouse sanitizers.

Supplementary Information

The online version contains supplementary material available at 10.1007/s13197-023-05882-1.

Introduction

The demand from consumers for fresh, organic, minimally processed, and chemical-free foods is on the rise, impacting their availability and supply in grocery stores and markets. In North America and Europe, the search for and sale of fresh, organic, and chemical-free foods increased by 90% (Arbenz et al. 2015; Rahman et al. 2021). Furthermore, regulations governing the use of various agricultural plant protection products have become more stringent, aiming to ensure the safety of these fresh, minimally processed foods.

Fresh, minimally processed, and organic farming foods pose greater conservation challenges due to the risk of contamination by pathogenic microorganisms, which can compromise product quality and, notably, shelf life (Kim and Song 2017). Particularly, fruits may lose essential properties and decay before reaching consumers, rendering even specific fruits unsuitable for sale (Dilarri et al. 2016). Moreover, bacteria classified as fecal coliforms, such as Escherichia coli, can lead to severe foodborne infections in humans, posing a significant food contamination concern (Martin et al. 2016).

To ensure consumer safety, the post-harvest process involves the packinghouse, where sanitization and controlled packaging procedures take place. The primary sanitizing agent used in most countries is chlorinated compounds, primarily sodium hypochlorite (NaOCl), often at a concentration of 200 ppm or higher (Weng et al. 2016; Dilarri et al. 2022). However, the continuous application of chlorine and chlorine-associated compounds raises concerns among public health organizations, leading to restrictions on its use or oversight of chlorine residue on food surfaces (EFSA 2015; Meireles et al. 2016).

The European Union has introduced a new category, 'Endocrine Disrupters' (EDs), for sanitizing products used in foods intended for human consumption. This category imposes limits on the maximum chlorate compounds allowed on fresh fruits (European Commission 2018; Dilarri et al. 2022). The focus on chlorine residue stems from the formation of byproducts (trihalomethanes and chloroform) that are known carcinogens and toxic compounds (São José and Vanetti 2012; Wang et al. 2013; EFSA 2015).

Electrolyzed water results from the electrolysis of a salt solution, with variations in time and electric current, yielding a water product with diverse properties and compounds. A broad spectrum of electrolyzed water exists, including neutral electrolyzed water, acid electrolyzed water, and alkaline electrolyzed water. Additionally, factors such as the type of salt utilized (NaCl, Na₂SO₄, ZnSO₄, and CaCl₂), in conjunction with the electrolysis equipment, electric current, and electrolysis time, can produce various types of electrolytic water characterized by distinct properties and by-products (Rahman et al. 2016; Ampiaw et al. 2021; Zhao et al. 2021).

The application of electrolyzed water has proliferated across diverse areas, finding utility in the clinical field of human health for the elimination of pathogenic microorganisms (Yan et al. 2021), as well as in the preservation and sanitation of foods (Rahman et al. 2016; Zhao et al. 2021). Electrolysis processes involving water are also deployed in environmental applications, such as the removal of dyes from the textile industry (Setodeh et al. 2021) and the degradation of organic matter present in sewage (Azizi et al. 2015).

The most widely used electrolyzed water involves NaCl, constituting the most extensively studied electrolysis process, given its low cost and high efficiency in microbial elimination. Conversely, electrolysis with Na₂SO₄ has not undergone extensive testing in the sanitization and preservation of food. To date, no study has explored the use of Na₂SO₄ in food sanitation simulating a packinghouse scenario against fecal coliforms, such as Escherichia coli.

It is essential to highlight that, unlike electrolysis with NaCl, which generates free chlorine, electrolysis with Na₂SO₄ does not produce free chlorine. The antimicrobial components in electrolyzed water with Na₂SO₄ consist of reactive oxygen species (ROS), including peroxides, superoxide, hydroxyl radicals, singlet oxygen, and alpha-oxygen. Consequently, aside from its effectiveness in eliminating microorganisms, it leaves no residue in the food.

The objectives of this study were to evaluate electrolyzed water with both NaCl and Na₂SO₄ in the sanitization and preservation of food. Both electrolyzed waters were compared with the traditional NaOCl at 200 ppm used in conventional packinghouses. The electrolysis generated with both salts underwent characterization and was employed in immersion wash assays of cherry tomatoes, simulating a commercial packinghouse. The microorganisms on the food surface were analyzed both before and after the wash treatment. Additionally, E. coli was utilized as the fecal coliform contaminant evaluated in this study. Observations were made on the shelf life of cherry tomatoes after the sanitization treatment with the two electrolysis methods, and all data were subjected to statistical analysis to ascertain the efficacy and differences between the two proposed electrolyzed water solutions.

Materials and methods

Electrolysis

The electrolysis was conducted using a deionized water solution of Na₂SO₄ at 0.08 M or NaCl at 0.08 M. The process took place in a cell containing titanium (Ti) electrodes coated with positively and negatively charged TiO₂ and RuO₂, each with a geometric area of 40.6 cm² for the anode and cathode. The energy system was connected to an electric power source generated by the Dawer model FCC-3005D. A magnetic stirrer was attached to the electrolytic system for agitation and homogenization of the electrolyzed water (Dilarri et al. 2016). The setup scheme of the electrolytic system is illustrated in Fig. S1 of the Supplementary Material. All electrolysis procedures were carried out for 3 min at 1 A and 8.7 V.

Conductivity, temperature, pH, and free chlorine levels of the produced electrolyzed water were monitored before and after electrolysis. Conductivity, temperature, and pH were measured using the Benchtop M1000 pH/mV/Conductivity/ISE/DO—Meters by LabForce (Muttenz, Switzerland). Free chlorine was quantified using the DPD colorimetric method with the Test Kit Model DL-DPDT by Del Lab (Araraquara, Brazil), and the absorbance of the color solution was measured using the UV–Vis spectrophotometer Shimadzu—Model 2401-PC (Kyoto, Japan). The electrolysis processes were executed ten times, with all parameters analyzed and compared during their generation.

Fecal coliform evaluated

The fecal coliform employed in this study was the Gram-negative bacterium E. coli (ATCC 8739). Escherichia coli cells were cultured in a nutrient broth medium containing 5 g L⁻¹ of peptone and 3 g L⁻¹ of beef extract, supplemented with 15 g L⁻¹ of bacterial agar for solid medium. The cultivation was conducted at 35 °C for 12 h in a shaker operating at 200 rpm. All reagents utilized were sourced from Himedia Laboratories Ltd. (Mumbai, India).

Food selection and sanitization assays

The vegetable selected for this investigation was the cherry tomato (Solanum lycopersicum var. cerasiforme). All specimens were directly procured from the producer, bypassing any sanitization processes in a packinghouse. The cherry tomatoes were meticulously divided into three distinct groups, ensuring uniformity in size, weight, and the absence of punctures or deformities. Rigorous control measures were implemented to maintain the utmost consistency among the evaluated groups.

Sanitization assays were conducted with three independent replicates, each comprising three groups of 10 vegetables subjected to different treatments. This approach aimed to establish a robust dataset for subsequent statistical analyses. To mitigate the risk of immediate spoilage, the sanitization assays were performed promptly after the collection of tomatoes.

Prior to conducting the sanitization assays, wherein the contamination of tomatoes by E. coli cells was simulated to mimic fecal coliform contamination, a series of steps were executed. Initially, E. coli cells were cultured in a nutrient broth medium until reaching a concentration of 10⁸ CFU mL⁻¹. Subsequently, the medium underwent centrifugation at 6000 × g for 7 min, resulting in the discarding of the supernatant, and the cells were then resuspended in a saline solution (NaCl 0.96%). Following this, tomatoes were spray-inoculated with the E. coli suspension at a concentration of 10⁸ CFU mL⁻¹ until the run-off point, after which they were allowed to air-dry at room temperature for 4 h.

The sanitization tests encompassed the evaluation of four distinct treatments: Negative Control (NC) utilizing autoclaved tap water; Positive Control (PC) employing a NaClO solution (Chemical Abstract Service number 7681-52-9, purchased from Sigma-Aldrich, Taufkirchen, Germany) with a concentration of 200 ppm of free chlorine; Treatment 1 (T1) employing electrolyzed water with NaCl; and Treatment 2 (T2) utilizing electrolyzed water with Na₂SO₄. Sanitization assays involved immersion washing in 500 mL of the respective treatment for duration of 2 min. Subsequently, the tomatoes were rapidly dried using a non-heated airflow from a hair dryer for 30 s and then placed into sterile Becker flasks covered by a porous membrane.

The Becker flasks were maintained at room temperature (1 ± 26 °C), and individual analyses of vegetable treatments were conducted every 24 h. The assessments included checking for the state of deterioration based on parameters such as degree of deterioration, fungus coverage, color, and consistency. Values ranging from 0 to 4 were assigned, with 0 indicating no change, 1 assigned to changes covering up to 1/4 of the food, 2 to changes covering up to 1/2 of the food, 3 to changes covering up to 3/4 of the food, and 4 to changes affecting the entire food item. Additionally, the appearance of each type of vegetable was scrutinized, considering external color categorized as alive (A) for foods maintaining their appearance and dimmed (D) for those that lost their color.

Microbiological analysis

Microbiology analyses were conducted after the food sanitization assays. Sterile swabs were employed to sample the treated foods, and these samples were inoculated onto various culture media specified in the present study. Nutrient Agar (NA) was employed for bacterial growth, while Sabouraud agar, supplemented with 0.15% v/v chloramphenicol, was utilized for fungal growth. Subsequently, the plates were incubated for 24 h at 30 ± 1 °C for NA media and 72 h at 26 ± 1 °C for Sabouraud media. Following the incubation period, colony-forming units (CFU) of fungi and bacteria were enumerated for each treatment. Microbiology assays were conducted immediately after the sanitization process (time zero) and after twelve days.

A total of 9 plates with NA and 9 with Sabouraud were made and counted for each treatment evaluated (NC, PC, T1, and T2). Each treatment in the sanitization assays consisted of three equal and independent groups, resulting in the utilization of 3 plates of medium for each independent group and a total of 9 medium plates. Given the presence of three independent triplicates in the sanitization assays, a cumulative total of 27 medium plates of NA and Sabouraud medium were assessed for time zero and the final time, respectively.

To confirm and identify the presence of the fecal coliform used as contamination (E. coli cells), bacterial colonies exhibiting growth similar to that of E. coli on agar medium were subjected to diagnosis by PCR, following the protocol outlined by Tsen et al. (2002). This approach allows for the verification of the efficacy of treatments PC, T1, and T2 in eliminating all E. coli cells. The same was done for the NC, seeking to validate the results with the rescue of E. coli cells from tomatoes surface.

Statistical analysis

All experimental assays were conducted with three independent groups, each comprising 10 vegetables per group. All assays were executed in triplicate and subjected to standard deviation analysis (SD) (Eq. 1) to assess experimental errors. All datasets underwent non-parametric statistical analysis using the Kruskal–Wallis (Dunn) test, with three degrees of freedom, performed using BioEstat 5.30 software.

$$SD=\sqrt{\frac{1}{N-1}\sum _{i=1}^N{\left(\frac{Qie-Qic}{\mathit{Qie}}\right)}^2}$$ 1

where Qie and Qic (µg mg⁻¹) are experimental and calculated data, and N is the number of measurements made.

Results and discussion

The results of temperature, conductivity, and pH revealed clear distinctions between the two types of electrolyzed water produced in this study and tap water (Table 1).

Table 1.

Physicochemical values of the two kinds of electrolyzed water produced in the present study and the tap water

	Free chlorine (ppm)	Conductivity (mS cm−1)	pH	Temperature (°C)
Tap water	1.4 ± 0.3	0.8106 ± 0.18	6.02 ± 0.06	23 ± 1.00
Electrolyzed water (NaCl)	67.15 ± 2.1	73.04 ± 0.48	8.02 ± 0.90	33 ± 0.80
Electrolyzed water (Na2SO4)	0	69.88 ± 1.59	4.80 ± 0.66	31 ± 0.78

Tap water exhibited consistent properties in terms of conductivity, pH, and temperature, despite the presence of free chlorine, which remained at very low levels. The Brazilian water treatment includes a chlorination stage, yet the concentrations are regulated to ensure that the level of free chlorine in the water does not exceed 2 ppm for consumers. In contrast, electrolyzed water with NaCl demonstrated significantly higher levels of free chlorine, potentially exerting antimicrobial effects at this concentration. Additionally, it displayed an alkaline pH and greater conductivity compared to tap water. These characteristics of electrolyzed water with NaCl arise from the generation of superoxide radicals and other antimicrobial agents such as Cl₂, ClO⁻, NaClO, HO₂, and HOCl (Rahman et al. 2016). Although the levels of free chlorine generated by NaCl electrolysis raise health concerns (EFSA 2015; Meireles et al. 2016), registering 67.15 ppm, this concentration is comparatively low when juxtaposed with traditional food sanitization in packinghouses using free chlorine at 200 ppm (Weng et al. 2016; Dilarri et al. 2022). Therefore, the electrolyzed water generated and applied in this study is unlikely to result in high concentrations of chlorine residue on the food surface post-application.

NaOCl is known to be corrosive, posing risks to individuals involved in packinghouse or sanitization processes, warranting caution in its use (São José and Vanetti 2012; Dilarri et al. 2022). However, at the levels of NaOCl produced in this work, it was deemed safe for use with low corrosive potential. The NaCl electrolysis proves highly effective in sanitizing food and is authorized and employed in various countries worldwide (Gil et al. 2015; Rahman et al. 2016). Notably, due to the NaCl concentration and electrolysis time, the generated electrolyzed water with NaCl predominantly contains a free chlorine amount below 100 ppm and maintains a pH value close to neutrality. This aligns with recommendations from the US Environmental Protection Agency and the United States Department of Agriculture (Hricova et al. 2008). Therefore, it can be affirmed that the NaCl electrolyzed water produced in this study is less aggressive compared to other studied and applied food sanitizers (Gil et al. 2015; Rahman et al. 2016; Ampiaw et al. 2021; Dilarri et al. 2021, 2022).

Electrolysis using Na₂SO₄ avoids the generation of free chlorine, and additionally, it imparts an acidic pH to the solution owing to the formation of free H₂ and H⁺ in the electrolyzed water. Na₂SO₄ electrolysis produces reactive oxygen species (ROS), including peroxides, superoxide, hydroxyl radical, singlet oxygen, and alpha-oxygen, all serving as potent antimicrobial agents that induce damage to proteins, nucleic acids, and cytoplasmic membranes (Li et al. 2010; Dharmaraja 2017; Yan et al. 2021). Crucially, the ROS compounds do not foster resistance or mutations when applied against microorganisms, presenting an additional advantage of Na₂SO₄ electrolyzed water (Li et al. 2010; Dharmaraja 2017).

While the conductivity of both types of electrolyzed water did not exhibit significant differences, they consistently maintained higher levels than tap water due to the ions formed. This elevated conductivity may also contribute to an inhibitory effect against certain microorganisms.

The cherry tomatoes exhibited signs of decay after 8 days, particularly following exposure to tap water washing (Table 2). Since these vegetables had not undergone any packinghouse processing or sanitization, their accelerated decay is expected. Despite rotting within a week at room temperature post-tap water washing, it is likely that the Negative Control (NC) treatment (2 min of immersion in tap water) merely removed surface dirt and debris without imparting any sanitizing activity. Notably, during the initial five days, the NC showed no significant difference compared to PC, T1, and T2, with distinctions becoming apparent only on the sixth day (Table 2). Remarkably, by the conclusion of the 12-day assay, the characteristics of the NC once again aligned with those of T1 and T2 (Table 2). This outcome underscores the pivotal role of washing processes in packinghouses, significantly extending the shelf life of foods (Kaewklin et al. 2018; Belgacem et al. 2021).

Table 2.

Classification of the vegetables after the sanitization assay

Cherry tomatoes	Degree of deterioration				Fungus coverage				Consistency				Color				Kruskal–Wallis (Dunn)
Days	NC	PC	T1	T2	NC	PC	T1	T2	NC	PC	T1	T2	NC	PC	T1	T2	NC	PC	T1	T2
1	1	1	1	1	1	1	1	1	1	1	1	1	A	A	A	A	n.s	n.s	n.s	n.s
2	1	1	1	1	1	1	1	1	1	1	1	1	A	A	A	A	n.s	n.s	n.s	n.s
3	1	1	1	1	1	1	1	1	1	1	1	1	A	A	A	A	n.s	n.s	n.s	n.s
4	2	1	1	1	1	1	1	1	2	1	1	1	A	A	A	A	n.s	n.s	n.s	n.s
5	2	1	1	1	1	1	1	1	2	1	1	1	A	A	A	A	n.s	n.s	n.s	n.s
6	3	2	1	2	1	1	1	1	3	1	1	2	D	A	A	A	n.s	p > .05	p > .05	p > .05
7	3	2	1	2	2	1	1	1	3	1	1	2	D	A	A	A	n.s	p > .05	p > .05	p > .05
8	4	3	2	3	4	2	2	2	4	2	2	3	D	A	A	A	n.s	p > .05	p > .05	p > .05
9	4	3	3	3	4	3	3	3	4	3	3	3	D	A	A	A	n.s	p > .05	p > .05	p > 0.5
10	4	3	2	3	4	3	3	3	4	3	3	3	D	A	A	D	n.s	p > .05	p > .05	n.s
11	4	4	3	4	4	4	3	4	4	4	3	4	D	D	D	D	n.s	n.s	n.s	n.s
12	4	4	4	4	4	4	4	4	4	4	4	4	D	D	D	D	n.s	n.s	n.s	n.s

n.s. Not significant

After six days of exposure to T1 and T2, cherry tomatoes exhibited favorable characteristics when compared with the Negative Control (NC) and Positive Control (PC), being statistically different from the NC. However, a distinction emerged between T1 and T2, with T2 manifesting signs of deterioration, loss of color, and a decrease in food consistency from day eight, indicative of a shorter shelf life compared to T1 and PC. By the tenth day, cherry tomatoes from T2 had already decayed, displaying all characteristics identical to those of the NC on the eighth day. While T2 extended the shelf life of cherry tomatoes by three days (a commendable outcome) it remained inferior to T1 and PC. Conversely, T1 increased the shelf life of cherry tomatoes by four days, proving more effective than T2 and statistically equivalent to PC. The color and food consistency of cherry tomatoes, following sanitization with T1 and PC, began to decline after the ninth day, attesting to their efficiency in sanitization and shelf life extension when compared with the NC.

The T1 contained antimicrobial compounds such as ClO⁻, HO₂, Cl₂, HOCl, NaClO, and superoxides, all proven to be efficient in inhibiting microorganisms and eliminating them without inducing resistance or mutation (Xuan and Ling 2019; Kim et al. 2019; Ampiaw et al. 2021). Notably, ClO⁻, HO₂, HOCl, and free chlorine act by damaging cytoplasmic membrane structures, ultimately leading to cell death (Dilarri et al. 2021). Consequently, for an organism to develop resistance to these compounds, mutations in multiple genes related to the structure of the cytoplasmic membrane would be necessary, rendering the emergence of mutants resistant to halogen compounds highly improbable. While the specific quantity of electrolysis byproducts was not identified in this study, the quantification of free chlorine, along with other parameters such as temperature, pH, and conductivity, affirms the generation of antimicrobial derivatives during the electrolysis of T1 and T2.

T2 demonstrated lower efficacy when compared to T1. Reactive Oxygen Species (ROS) are the primary agents in electrolyzed water with Na₂SO₄ against deteriorating microorganisms (Li et al. 2010). These agents are volatile, losing their effectiveness rapidly after electrolysis. Although the washing experiment was conducted immediately after electrolysis to minimize the loss of antimicrobial agents formed in the water, some may have still volatilized during the immersion washing process, diminishing the impact of T2 compared to T1. It is essential to recognize the capacity of certain microorganisms to resist ROS (Dharmaraja 2017; Ren et al. 2021), thus reducing the sanitizing effect of T2 and potentially resulting in a higher number of microorganisms on the food surface even after treatment. However, T2 demonstrated a significant extension in the shelf life of cherry tomatoes compared to the NC, establishing itself as a viable alternative for the sanitization process in packinghouses to enhance the shelf life of vegetables. Another advantage of T2 lies in its non-generation of chlorine residues or derivatives on the food surface.

Microbiological assays revealed a significant difference between the electrolytic treatments and the NC, confirming that washing with the two types of electrolyzed water led to a substantial reduction in the number of bacterial colonies on the food surface. Both T1 and T2 were statistically equivalent to the PC, representing a reduction of 2 Log CFU mL⁻¹ in bacterial numbers in both electrolytic treatments when compared to the NC (Fig. 1). However, by the final time point (12 days), the number of bacterial colonies in each treatment had returned to the levels observed in the NC (Fig. 1). Surprisingly, even the traditional sanitization with free chlorine at 200 ppm exhibited statistical equivalence to the NC after twelve days, implying that it provides a similar level of protection and microorganism elimination as T1 and T2.

Fig. 1.

Microbiological assays depicting bacteria colony numbers following washing treatments: a Bacteria isolation after the wash (time zero), and b Bacteria isolation at the final assay time (12 days). NC represents washing with tap water, PC denotes washing with a free chlorine solution at 200 ppm, T1 corresponds to washing with electrolyzed water with NaCl, and T2 signifies treatment with electrolyzed water with Na₂SO₄. Boxes illustrate the data distribution per treatment, with lines above and below representing the minimum and maximum values in the datasets; plus signs indicate the averages calculated for each distribution. Asterisks denote statistical differences among treatments, as determined by Kruskal–Wallis (Dunn) analysis

The fungi isolation results paralleled those of bacterial isolation, demonstrating a reduction of 1.5 Log CFU mL⁻¹. Although this reduction was less pronounced compared to bacteria, it still exerted a significant effect on removing fungi from the surface of cherry tomatoes (Fig. 2). Both electrolytic treatments were statistically equivalent at both time zero and after twelve days (Fig. 2). Additionally, there were consistently more yeast colonies observed than molds or mycelium fungi on all treatment plates, which constitutes an intriguing result.

Fig. 2.

Microbiological assays displaying fungi colony numbers following washing treatments: a Fungi isolation after the wash (time zero), and b Fungi isolation at the final assay time (12 days). NC stands for washing with tap water, PC refers to washing with a free chlorine solution at 200 ppm, T1 corresponds to washing with electrolyzed water with NaCl, and T2 signifies treatment with electrolyzed water with Na₂SO₄. Boxes illustrate the data distribution per treatment, with lines above and below representing the minimum and maximum values in the datasets; plus signs indicate the averages calculated for each distribution. Asterisks denote statistical differences among treatments, as determined by Kruskal–Wallis (Dunn) analysis

The results of microorganism isolation from the sanitization assays during the final storage period indicated a persistent presence of the microbial population even after the treatments. This suggests that both the electrolytic treatments and the PC (which are commonly recommended sanitation agents in packinghouses) predominantly inhibited microorganisms rather than eliminating them entirely from the surface of cherry tomatoes. Notably, electrolytic water and NaOCl at 200 ppm exhibit limited effectiveness in eliminating bacterial biofilms (Dilarri et al. 2016; Zhao et al. 2021; Yan et al. 2021), potentially contributing to the resurgence of microbial growth towards the end of the food storage period. Bacterial and fungal spores may also persist on the food surface, leading to renewed microbial growth once the antimicrobial compounds have completely volatilized. It is well-known that electrolyzed water is not effective in eliminating microbial spores (Pintaric et al. 2015).

Moreover, even at time zero (immediately after sanitizing), isolated microorganisms were detected on the surface of tomatoes washed with T1, T2, and PC (Fig. S2 of the Supplementary Material). Since the tomatoes were not kept in a sterile environment and were exposed to external air, other microorganisms or microbial spores might have come into contact with them, simulating real-world market storage conditions. It's noteworthy that the byproducts generated by the electrolysis of Na₂SO₄ exhibit a fast volatilization effect, resulting in a less sustained residual effect compared to electrolysis with NaCl or NaOCl at 200 ppm. Electrolysis with NaCl produces volatile compounds, but free chlorine persists for a longer duration on the food surface compared to the products from Na₂SO₄ electrolysis (SO₄⁻ and H⁺) (Li et al. 2010; Pintaric et al. 2015; Rahman et al. 2016). This is supported by the observed increase in the shelf life of cherry tomatoes by one additional day in T1 and PC compared to T2. These findings underscore that electrolytic treatments and NaOCl do not exhibit prolonged residual effects on microbial inhibition, and microorganisms reemerge once their biocidal agents have volatilized from the food surface.

The elimination of fecal coliform cells, represented by E. coli in the present study, was confirmed by PC, T1, and T2, with a reduction of more than 2 logs of E. coli cells rescued (Fig. 3). This result is particularly noteworthy given that the contamination with E. coli involved a saline solution with 10⁸ CFU mL⁻¹, an excessively high number for real-life contamination scenarios. Nonetheless, T1, T2, and PC significantly reduced the bacterial load compared to NC (3.34 log of CFU mL⁻¹ of E. coli cells), which only involved a wash with sterile tap water (Fig. 3).

Fig. 3.

Escherichia coli cells rescued from the tomato surface after the sanitization wash of each treatment at time zero. Bars represent averages of isolated cells. Data showing the same capital letters are not significantly different from each other based on Kruskal–Wallis Dunn analysis

Escherichia coli, as a key fecal coliform and a widely used bacterial species for analyzing food contamination (Martin et al. 2016; Kim and Song 2017; Kim et al. 2019), underscores the food safety aspect of using electrolytic water of NaCl and Na₂SO₄ for food sanitization, completely eliminating this fecal coliform from the food surface. Although this study did not analyze other clinical pathogens such as Salmonella enterica, Salmonella typhi, Staphylococcus aureus, and Enterococcus faecalis, Kim et al. (2019) demonstrated success in eliminating a range of pathogens with electrolyzed water. Similarly, Gil et al. (2015) investigated the application of electrolytic water in the fresh-cut industry and concluded that it is as effective as NaOCl in food disinfection, offering an eco-friendly and non-hazardous technology.

The present work revealed that many microorganisms associated with food rot regrew, indicating the persistence of microbiota in the food even after the sanitization treatments. This phenomenon was also observed in the PC during the final storage time, emphasizing that the shelf life and fecal coliform removal evaluated by electrolytic water were comparable to NaOCl. Consequently, based on the findings of this study and other investigations (Li et al. 2010; Rahman et al. 2016; Gil et al. 2015; Kim et al. 2019), it is reasonable to consider that foods sanitized with electrolytic water are as safe for consumption as those sanitized by the traditional NaOCl used in the food industry.

In summary, the electrolysis with Na₂SO₄ and NaCl has demonstrated efficacy as a viable sanitizer, effectively reducing a significant quantity of microorganisms and fecal coliform on the food surface. Moreover, it extends the shelf life by an additional three days without leaving any residual free chlorine, a notable advantage considering the increasing demand for food sanitizers devoid of chlorine residue (EFSA 2015).

It is crucial to recognize that while NaOCl at 200 ppm remains the most widely employed sanitizer in industrial settings due to its lower operational costs compared to electrolytic water, the constraints imposed by EU regulations on endocrine disruptors (EDs) will likely curtail its application. This limitation justifies the exploration of alternatives to NaOCl in packinghouses. Notably, water electrolysis stands out as a technology easily transferrable to large-scale industries (Chatenet et al. 2022). Many industries are already adapting their sanitization processes to comply with stringent regulations regarding chlorine residues. Consequently, the study's findings emphasize the comparative aspects of applying electrolytic water in contrast to NaOCl, revealing minimal differences that could position electrolytic water as a viable alternative in packinghouses.

Conclusions

The effectiveness of both electrolytic waters in inhibiting microorganisms on the food surface, leading to a four-day extension in the shelf life of cherry tomatoes, has been established. The inhibitory effects of the evaluated electrolyzed waters were transient, with a limited residual impact on the food surface post-application. It is noteworthy that a residual microbiota persisted on the food surface even after the sanitization process, indicating that the effect of the electrolytic water did not eliminate all microorganisms, ultimately resulting in the rotting of cherry tomatoes after 12 days. Interestingly, similar results were observed in the case of the free chlorine solution at 200 ppm.

Both electrolyzed waters also demonstrated significant reductions in the tested fecal coliforms, with results statistically equivalent to those achieved with the free chlorine solution at 200 ppm. Consequently, electrolytic water derived from NaCl and Na₂SO₄ exhibited comparable efficacy to traditional free chlorine sanitation, suggesting it as a viable alternative to NaClO at 200 ppm for disinfecting fresh produce in packinghouses.

Supplementary Information

Below is the link to the electronic supplementary material.

Author contributions

CRM and GD: carried out the experiments, conceptualization, methodology, validation, formal analysis, writing original draft; RNM: conceptualization, methodology, resources; EDB: conceptualization, visualization, writing/review and editing, supervision, project administration, funding acquisition.

Funding

C. R. Mendes received a Ph.D. scholarship from Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)—Brazil. This work was funded by Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)—process 150092/2022–9.

Data availability

The data that support the findings of this study are available from the corresponding authors upon request.

Declarations

Conflict of interest

The authors declare that they have no conflict of interest.