Physicochemical, microbiological, and sensory quality of industrialized and fresh coconut water commercialized in Petrolina, Pernambuco

doi:10.21203/rs.3.rs-1523554/v1

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Physicochemical, microbiological, and sensory quality of industrialized and fresh coconut water commercialized in Petrolina, Pernambuco

https://doi.org/10.21203/rs.3.rs-1523554/v1

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Coconut water is widely consumed in Brazil. However, due to its nutritional qualities, this product is susceptible to microbiological contamination when good hygiene practices are not adopted. From this perspective, this study aimed to evaluate the physicochemical and microbiological quality of industrialized and fresh coconut water samples purchased in the municipality of Petrolina-PE, and perform the sensory analysis of the studied samples. The microbiological properties were analyzed by evaluating the concentration of total and thermotolerant coliforms, Escherichia coli, Salmonella ssp., molds, and yeasts, whereas the physicochemical properties were analyzed by evaluating the pH, total titratable acidity, ash content, and turbidity. The sensory analysis was performed by an acceptance test using a 9-point hedonic scale. The results of the evaluations demonstrated that all samples were within the standards of the current regulations. However, the presence of total coliforms and molds suggests that the products were contaminated by fecal material and/or had problems in the production process, which could result in health damage to consumers. Sensory evaluation revealed that fresh coconut water was preferred among tasters. In this scenario, it is essential to adopt good practices among traders and bottling companies and greater participation of the public sector to ensure the quality of food products commercialized in the region, thus reducing the risks associated with the consumption of contaminated foods.

Coconut Water

Food Quality

Coliforms

Escherichia coli

Consumer Behaviour

Favored by climatic conditions, coconut cultivation is widely encouraged in Brazil, especially in the North and Northeast regions, which concentrate approximately 75% of the national coconut production (Santos 2018). Since the 1990s, the greater awareness about the benefits of natural products has favored the cultivation of green dwarf coconuts for water consumption, a natural product with outstanding nutritional qualities (Martins and Jesus-Junior 2014; Gonçalves and Souza 2019).

Coconut water is usually commercialized within the fruit. However, this form of consumption might not be viable in locations far from producing regions due to transportation, storage, and shelf-life problems. Therefore, in order to allow its consumption in distant areas, bottling techniques for coconut water have become a viable alternative to decrease the transported volume and weight and increase its shelf life (Silva et al. 2017).

Although sterile when inside the fruit, coconut water favors microbial development due to its chemical composition and abundance of easily assimilated nutrients, becoming a food product with a high risk of disease transmission when not properly handled (Lima and Silva 2019). With regard to the bottling system, although microbiological control methods exist in the production sector, small industries usually have no access to qualified professionals with specific knowledge about how to improve food quality standards, favoring public health problems (FAO/WHO 2007; Lima 2013).

In this scenario, the city of Petrolina, Pernambuco, on the banks of the São Francisco River, stands out in the irrigated agriculture sector, with more than 2.2 thousand hectares of coconut farming areas and a total production of approximately six million fruits per month (Lima Júnior and Souza 2015; Globo Rural 2019). In that municipality, it is possible to observe both street vendors, who sell coconut water in the streets, and small processing industries, which bottle coconut water and commercialize it in the municipality and other locations. Although an acceptable quality standard exists for both forms of sale, it is often neglected, negatively affecting consumer health (Silva et al. 2017).

From this perspective, this study aimed to evaluate the physicochemical and microbiological quality of industrialized and fresh (in natura) coconut water samples in the municipality of Petrolina-PE and evaluate the sensory perception and predilection of the local population with regard to the coconut water available for consumption.

2.1. Sample collection

The experiments were conducted using industrialized and fresh coconut water samples commercialized in Petrolina-PE and collected from July to December 2019. The fresh samples were purchased from street vendors, stored in sterile amber glass vials, placed in coolers containing ice to maintain the temperature, and immediately sent to the Laboratory of Microbiology of the Federal University of Vale do São Francisco (Univasf), Central Campus. The industrialized products were purchased from commercial establishments and sent to the laboratory in their original containers.

2.2. Colimetric analyses

Microbiological analysis was performed using the multiple-tube method with a series of three tubes (Hitchins et al., 1998). For the presumptive test, the samples were sequentially diluted in test tubes containing 9 mL of Lactose Broth to obtain the dilutions of 10^− 1, 10^− 2, and 10^− 3, in triplicate, with incubation at 36ºC for 48h. The tubes considered positive were then inoculated in Bright Green Bile Lactose Broth for confirmatory analysis of total coliforms, in Escherichia coli broth (EC) for confirmation of thermotolerant coliforms, and incubated at 36ºC and 44.5ºC, respectively, for another 48h. The Most Probable Number (MPN/mL) of total and thermotolerant coliforms was determined using the three-tube series table (Garthright 2001).

2.3. Identification of Escherichia coli

For the isolation of E. coli, an aliquot taken from the tubes containing EC broth and showing a positive result was streaked on Petri dishes containing Eosin Methylene Blue agar medium. The colonies with characteristic growth were then identified through the following biochemical tests: Indole, motility, H₂S production, carbohydrate fermentation, gas production, glucose fermentation, and Simmons citrate (Koneman 2001).

2.4. Identification of Salmonella ssp.

For the isolation and identification of the strains of Salmonella sp., 25 mL coconut water aliquots were incubated at 37 ºC for 24 h in 225 mL of Buffered Peptone Water as a pre-enrichment phase. Next, for the selective enrichment, 0.1 mL of Buffered Peptone Water was transferred to test tubes containing 10 mL of Rappaport-Vassiliadis soya broth and then incubated at 41.5 ºC for another 24 h. After this period, the cultures were then continuously streaked on Petri dishes containing Salmonella-Shigella agar medium, followed by incubation at 37 ºC for 24 h. The typical Salmonella colonies were subjected to biochemical assays for genus confirmation (Koneman 2001).

2.5. Count of molds and yeasts

Molds and yeasts were analyzed based on the methodology described in Brasil (2003). For that purpose, 1 mL of each sample was diluted in 9 mL of sterile saline (0.85%). Subsequently, 100 µL aliquots from the dilutions were transferred to sterilized and previously identified Petri dishes. Next, each plate received 20 mL of previously fused Potato Dextrose Agar and cooled at 45°C. The palates were then slightly homogenized in 8-shaped movements. After solidification at ambient temperature, the dishes were incubated in an oven at 35 ± 1ºC for 48 hours. All assays were performed in triplicate.

2.6. Potential of Hydrogen (pH)

The potential of hydrogen (pH) of the samples was measured using an MS Tecnopon® pH meter, model mPA-210. The pH value was measured in 200 mL of sample following the method developed by the Adolfo Lutz Institute (2008).

2.7. Total Titratable Acidity

The acidity was determined by titration with sodium hydroxide and expressed as NaOH per 100 mL of coconut water according to the analytical guidelines of the Adolfo Lutz Institute Lutz (2008).

2.8. Waste by incineration

The fixed mineral residue (ash) was determined by the gravimetric incineration method in a QUIMIS muffle furnace (Q-318S24) at 500°C according to the methodology of the Adolfo Lutz Institute Lutz (2008).

2.9. Turbidity

The turbidity was measured with an MS TECNOPON benchtop turbidimeter, model TB-1000. The results were expressed as NTU (Nephelometric Turbidity Units).

2.10. Sensory analysis

The industrialized and fresh coconut water samples were sensorily analyzed at time zero in a double-blind study by applying an acceptance test using a 9-point hedonic scale (9 = like extremely and 1 = dislike extremely), in which the subject expresses its feelings about consuming a certain product (IAL 2008).

The sensory evaluation was performed at the Central Campus of UNIVASF in a classroom reserved for the experiment to ensure the safety and privacy of the participants and the secrecy and confidentiality of the information obtained. Any person interested in participating could take part in the study after signing the Free Consent Form and attesting to having no intolerance or allergy to any of the evaluated foods. The data obtained in the acceptance test were statistically analyzed by ANOVA, and data significance was verified by the Tukey test at 5% of significance.

The evaluation was arranged so that each participant could taste the different samples sequentially (differentiated order among the participants), among industrialized and fresh samples offered in disposable plastic cups (50mL of coconut water per sample) without identifying their origin. The samples were presented in each position an equal number of times.

In order to reduce the risk of food poisoning, the industrialized samples were opened and put in the disposable cups for up to five minutes before the test, thus maintaining the characteristics of the product and avoiding contamination. The fresh samples were obtained by purchasing green coconuts, which were opened by the researchers and had their water stored for up to one hour in a vacuum bottle. Then, the fresh water samples were put in disposable cups for up to five minutes before experimentation. The samples were sealed and stored in a refrigerator at 5ºC until the moment of the assays. Finally, in order to prevent allergic reactions during the interviews, all ingredients contained in the products were informed to the interviewees before consumption.

This study was analyzed by the Research Ethics Committee of the Federal University do Vale do São Francisco (CEP/UNIVASF) under the CAAE protocol No. 97252818.4.0000.5196.

Ten fresh and twenty industrialized coconut water samples of four different brands were analyzed during the study, with five samples of the same batch per brand. The results of the colimetric analyses (Table 1) signaled total coliforms in all fresh samples and in five samples of brand D. Although total coliforms are not addressed by RDC 12/2001 of ANVISA as a microbiological quality parameter, the results revealed the presence of exogenous contaminants in these samples since coconut water remains sterile while inside the fruit (Dias et al. 2015).

Table 1

Result of the microbiological assays for industrialized (samples A – D) and fresh (samples 01–10) coconut water samples.
Sample	Total coliforms (NMP/mL)	Thermotolerant coliforms (NMP/mL)	Escherichia coli (NMP/mL)	Salmonella ssp. (+/-)	Molds and yeasts (UFC/mL)
A	0*	0*	0*	-	> 300
B	0*	0*	0*	-	> 300
C	0*	0*	0*	-	> 300
D	5*	0*	0*	-	> 300
01	745	11	0	-	> 300
02	1100	0	0	-	> 300
03	475	0	0	-	> 300
04	1980	20	8	-	> 300
05	475	9	9	-	> 300
06	168	0	0	-	> 300
07	216	0	0	-	> 300
08	334	0	0	-	> 300
09	1040	11	0	-	> 300
10	324	0	0	-	> 300
*Results expressed as number of products from the same batch with MPN > 200/mL.

Thermotolerant coliforms and Escherichia coli were found in four of the ten fresh samples, although within the 200 NMP/mL limit established by the regulations. No samples were contaminated by Salmonella sp. Although the values observed were within the range considered acceptable for consumption, the presence of total coliforms suggests that these samples were directly or indirectly contaminated by human or animal feces, characterizing a potential health risk (Araujo 2017).

Moreover, although the samples purchased from street vendors were within the safety limits established for consumption, good hygiene practices are crucial to ensure social well-being and prevent disease outbreaks. From this perspective, the scientific literature contains several studies with similar results, whose levels of thermotolerant coliforms are within the established limits (Serejo, Neves and Brito 2010; Souza and Souza 2019). However, various other studies have also found levels above the permitted (Michelin et al. 2014; Silva et al. 2017), showing that this issue deserves attention by both the population and public authorities.

Molds and yeasts were detected in all analyzed samples, with microorganism counts above the test limit of 300 Colony-Forming Units (UFC) per milliliter. For foods subjected to microbial reduction processes that require refrigeration, the maintenance of this condition is essential to prevent the proliferation of microorganisms, with refrigeration failure acting as the main factor responsible for increasing the microbial load. Furthermore, the commercialization of fresh coconut water raises concerns since street vendors often open the fruits at home and store the water in jars or vacuum bottles, where it remains for several hours during commercialization.

Amid the economic and social problems observed in Brazil, the informal economic sector has grown steadily in recent years, with food trade by street vendors standing out in that regard (Viviani et al. 2020). However, most people in this segment have little or no knowledge about good hygiene and food handling practices, which is directly responsible for the findings observed in the present study (Nascimento et al. 2017). According to Boaventura (2017), the presence of contaminants in foods is often associated with precarious personal hygiene, equipment, and utensils.

With regard to physicochemical analyses, the products belonging to brands A, B, and C agreed with Normative Instruction No. 9 of January 30, 2020, with regard to the analyzed parameters (Table 2). On the other hand, the total titratable acidity of brand D was above the recommended, which should range between 0.06% and 0.18%. Furthermore, although the new regulation established 4 as the minimum level for pH analysis, the previous regulation (Normative Instruction No. 39 of July 22, 2009) established 4.3 as the minimum pH for coconut water, which would disagree with the observed parameter.

These analyses are essential in the production of bottled coconut water since they directly interfere with the final quality of the product (Lima et al. 2015). According to Vasconcelos et al. (2015), attention with regard to the pH of coconut water is essential to ensure its desirable sweet taste, which is improved when the pH approaches 5.5. On the other hand, coconut water samples with low pH suggest the use of premature fruits, affecting the final quality.

Table 2

Physicochemical parameters of industrialized products
Sample
	pH	Turbidity	Ash (g/100mL)	Titratable acidity (g/100mL)
A	4.9	14	0.36	0.07
B	5	102	0.45	0.06
C	5	86	0.43	0.07
D	4.2	13	0.56	0.19

Similar to pH, acidity also influences the characteristic taste and aroma of the food product. Titratable acidity refers to the content of malic acid present in coconut water: when the value is high, it indicates that immature fruits were used since the acid content decreases throughout maturity, negatively affecting the organoleptic characteristics (Vasconcelos et al. 2015).

In the studied samples, the turbidity ranged from 13 to 102, with brands B and C showing the highest values of 102 and 86, respectively. The water samples of these brands were visually whiter than the remainder, which was responsible for their higher turbidity values. These results may be related to both the maturity stage of the fruit used, since turbidity tends to increase throughout maturation, and the shelf-life of the product, since turbidity increases during storage (Kwiatkowski, Oliveira and Clemente, 2012; Lima 2015).

With regard to the ash content, the Nutritional Table of Food Composition (TBCA 2020) determined a mean value of 0.46% for every 100g of food, a value very close to those obtained for brands B and C. On the other hand, lower values such as those observed for the brand A samples could indicate the use of immature fruits during the production processes, whereas high values could imply the addition of inorganic matter to the product (IAL 2008; Vasconcelos et al. 2015). However, the presence of exogenous materials in the analyzed samples was not evaluated in the present study.

After evaluating the physicochemical parameters, although all four brands were fit for consumption, brands B and C showed the best evaluations with regard to consumption conditions. Brands A and D, on the other hand, showed results that suggest the use of immature fruits, which can compromise the final characteristics of the products, reducing their acceptance by consumers and causing financial damage to the company.

For the sensory evaluations, 64 individuals were interviewed to assess their predilection for the coconut water samples of brands A, B, C, and fresh coconut water. Due to the presence of coliforms in the products of brand D, these samples were not included in the sensory analysis.

As a result of the evaluations, the tasters preferred the fresh coconut water samples, which received 100% approval (grades between 6–9). The analysis of variance of the acceptance values revealed a significant difference (p < 0.05) between industrialized and fresh coconut water samples. Table 3 shows the mean acceptance of the four forms of presentation of coconut water.

Table 3

Mean acceptance of coconut water samples.
Sample	Mean	% Approval	% Rejection
A	5.1a	55	33
B	5.6a	67	33
C	3.8b	28	69
In natura	8.7c	100	0
Note: Means with the same letter do not differ statistically (p < 0.05).

Among the brands analyzed, A and B did not differ significantly with regard to consumer acceptance and were classified in the hedonic scale close to the category "neither like/nor dislike", obtaining an average acceptance index. Sample C showed the lowest acceptance among tasters and was classified close to the category “dislike moderately”. On the other hand, the fresh coconut water samples obtained the highest acceptance rates, with 100% approval, and were classified close to the category “like very much”.

Since it is a minimally processed food, fresh coconut water keeps its original organoleptic characteristics, thus justifying its high acceptability. The samples that underwent processing before bottling and commercialization were less accepted with regard to palatability than the natural product, constituting a risk to the population since all analyzed samples had some contamination.

From this perspective, coconut water should be obtained directly from the fruit opened at the moment of consumption since the food will be less subject to microbial contamination and will maintain all its organoleptic characteristics. If this form of consumption is not possible, the industrialized product is more advantageous from the food safety perspective due to its sterilization prior to bottling. However, consumers should be aware of shelf life and packaging characteristics, avoiding crumpled and swollen products or with any violation.

Furthermore, public authorities should regulate and monitor street vendors who sell this product to minimize the risks associated with contaminated foods and continuously or periodically offer courses on food safety and handling to allow the safe trade of foods in the municipality.

According to the current regulations, all coconut water samples analyzed were fit for consumption from the microbiological and physicochemical perspectives. However, coliforms in the fresh samples suggest their contamination by fecal material. The contamination observed in the samples belonging to brand D suggests issues in the aseptic chain during production, highlighting the need for greater attention with regard to products before their acquisition.

Sensory analysis revealed that fresh coconut water was preferred by tasters over bottled samples, highlighting the need for good hygiene and handling practices by traders in order to ensure the quality of the product and reduce risks associated with contamination.

CRediT authorship contribution statement

Carlos Henrique Araujo Dias: Conceptualization, Methodology, Investigation, Writing - Original Draft, Supervision. Dairan Santos França: Conceptualization, Formal analysis, Investigation, Writing - Original Draft. Monica Cristina Rezende Zuffo Borges: Formal analysis, Investigation, Writing - Review & Editing. Rita de Cássia Rodrigues de Souza: Methodology, Resources, Writing - Review & Editing. Marlos Gomes Martins: Methodology, Resources, Writing - Review & Editing.

Funding Source Declaration

The authors declare that no funding was received for this work.

Declaration of competing interest

The authors declare that there is no conflict of interest.

Data Availability Statement

No datasets were generated or analyzed during the current study.

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No competing interests reported.

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Physicochemical, microbiological, and sensory quality of industrialized and fresh coconut water commercialized in Petrolina, Pernambuco

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Abstract

1. Introduction

2. Material And Methods

2.1. Sample collection

2.2. Colimetric analyses

2.3. Identification of Escherichia coli

2.4. Identification of Salmonella ssp.

2.5. Count of molds and yeasts

2.6. Potential of Hydrogen (pH)

2.7. Total Titratable Acidity

2.8. Waste by incineration

2.9. Turbidity

2.10. Sensory analysis

3. Results And Discussion

4. Conclusions

Declarations

References

Additional Declarations

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