Use of Pecan Shell Extract and Green Tea in a Kombucha-Vinegar-Based Beverage with Enhanced Antioxidant Properties

Abstract

We aimed to develop kombucha-vinegar beverages inspired by switchel (a beverage that combines apple cider vinegar and ginger extract), using pecan shell aqueous extract (PSE) and green tea infusion (GTI) in the preparation of kombucha vinegar, and to assess its effects on physicochemical characteristics, antioxidant activity, and sensory acceptance. Combinations of PSE and GTI (100:0, 75:25, 50:50, 25:75, and 0:100) were tested as substrates to produce kombucha vinegar with an initial sugar concentration of 80 g/L. After, the initial sucrose concentration was tested (80 to 60 g/L) using two of the previous formulations (50% of PSE and 50% of GTI; 25% of PSE and 75% of GTI), that showed better results in antioxidant capacity and sensory characteristics, particularly bitterness, which was attributed to the addition of higher amounts of pecan nutshell extract (100 and 75%). The formulation with 60 g/L of sucrose and higher pecan shell extract (50%) was chosen, allowing a beverage with less sugar at the end of kombucha fermentation. An increase in antioxidant potential was observed during the fermentations, with this being a highlight of this study. Kombucha vinegar beverages inspired by switchel were developed (50% PSE and 50% GTI, 60 g/L of sucrose), with the use of ginger extract or juice fruits (apple, pineapple, or white grape), in order to make the beverage palatable to consumers. The samples without ginger showed the highest antioxidant capacity values. In the sensory evaluation using acceptability and the check all that apply method (CATA), the beverages without ginger showed acceptability ranging from 74.4% (addition of white grape juice) to 84.0% (addition of pineapple juice), being described as refreshing, healthy, and energizing.

1. Introduction

The functional beverage market is one of the fastest-growing segments within the functional food industry [1]. Consumers are increasingly interested in beverages that offer health benefits, with vinegar-based drinks being recognized for their potential antioxidant effects, anti-inflammatory properties, and support for digestive health [2]. Brands have diversified flavors to make these beverages more appealing, and consumer interest is driving the exploration of unconventional raw materials with the potential to enhance the functional properties of final products [3].

Switchel is a traditional fermented or acidified beverage made with water, vinegar (usually apple cider vinegar), a natural sweetener (such as molasses, honey, or maple syrup), and often ginger. It is historically known as “reaper’s punch” because it was consumed by farm workers for hydration. This beverage is now well accepted by health-conscious consumers [4]. Adding fruit juices to this drink is also an interesting approach to introducing new flavors into the beverage industry, which constantly demands innovation. This practice not only provides unique taste experiences but also highlights the diversity of local agricultural production.

Kombucha vinegar, a variation of traditional vinegar, is produced through prolonged fermentation using a kombucha culture [5]. Kombucha, a functional beverage that may offer numerous health benefits due to its active metabolites [6,7], is traditionally obtained by fermenting sweetened green or black tea with the help of a microbial colony known as SCOBY (Symbiotic Culture of Bacteria and Yeasts) [8,9]. The popularity of kombucha as a functional food is mainly driven by its antioxidant potential, cholesterol- and blood pressure-lowering effects, and its ability to improve immune function and gastrointestinal health [10,11]. Similar to kombucha, the functional and sensory characteristics of kombucha vinegar depend on the types of teas used and the flavors added during the fermentation process [12]. Therefore, fermentation in alternative substrates, when properly conducted, can result in distinct and appealing products for consumers seeking diverse functional and sensory experiences [8]. These trends reflect the ongoing evolution of consumer preferences toward options that are not only healthier but also more flavorful in the beverage market.

Alternative substrates may include different types of teas, such as herbal infusions [13], fruit infusions [14], or even infusions of specific plants [15], such as hibiscus [16] or yerba mate [17]. The aqueous extract of pecan shells has been explored in various applications in food and beverage production [18,19,20]. By using pecan shell aqueous extract, researchers aim not only to explore its health benefits but also to find sustainable ways to valorize agricultural residues, contributing to more eco-friendly and efficient practices within the production chain. Pecan shells contain a variety of compounds with potential health benefits, such as antioxidant and antimicrobial properties [21]. The relevance of these findings has the potential to create new revenue streams from shell by-products, thereby increasing the economic value of pecan cultivation [22].

The aim of our study was to develop kombucha-vinegar beverages inspired by switchel, using pecan shell aqueous extract and green tea in the preparation of kombucha vinegar (an alternative acidic base instead of apple vinegar present in switchel), and to assess its effects on physicochemical characteristics, antioxidant activity, and sensory acceptance. During the beverage development process, we studied the kombucha fermentation by gradually replacing green tea with pecan shells extract, adjusting the initial sugar concentration to obtain a final kombucha vinegar with reduced sugar content, and subsequently, we studied the addition of fruit juices or ginger to the formulations in order to make the beverage palatable to the consumer.

2. Materials and Methods

The study was conducted in four sequential steps. First, an aqueous extract from pecan shells was obtained and chemically characterized, with emphasis on physicochemical properties and antioxidant potential. In the second step, kombucha vinegar was produced using pecan shells extract (PSE) and green tea infusion (GTI) in various proportions, and subjected to physicochemical characterization to evaluate its suitability as an acidic base for beverage formulation. The third step involved the development of beverages inspired by switchel formulations using kombucha vinegar, ginger extract, or fruit juices, followed by physicochemical and antioxidant characterization of the resulting formulations. Finally, in step 4, sensory analysis was performed to assess consumer acceptance and overall perception of the developed beverages.

2.1. Obtaining Pecan Shell Extract (PSE)—Step 1

Pecan shells were obtained directly from the industrial processing stage in order to facilitate the use of the residue without the need for degreasing. The shells, supplied by the company “Agroindústria Pitol” (city of Anta Gorda—RS, Brazil), were separated by particle size (20 mesh). The aqueous extracts from pecan shells were prepared using 10% (w/v) pecan shells in water at 75 °C for 10 min. After preparation, the extracts were filtered through paper filters and cooled to room temperature. During this procedure, the extracts were protected from light and stored in amber bottles, as described by Trevizol et al. [23]. This extract was used in the preparation of the substrate for the production of the kombucha vinegar.

2.2. Kombucha-Vinegar Production—Step 2

The kombucha-vinegar production was studied in two experiments, as shown in

Table 1. Sequential experiments to study the kombucha-vinegar production.

Experiment 1 for Kombucha-Vinegar Production
Substrates	F1	F2	F3	F4	F5
PSE (%, v/v)	100	75	50	25	0
GTI (%, v/v)	0	25	50	75	100
Sucrose (g/L)	80 (in all formulations of Experiment 1)
Experiment 2 for Kombucha-Vinegar Production
Substrates		F6	F7	F8	F9
PSE (%, v/v)		50	50	25	25
GTI (%, v/v)		50	50	75	75
Sucrose (g/L)		80	60	80	60

Legend: PSE: pecan shell extract prepared according to item 2.1; GTI: green tea infusion prepared using 5 g of green tea leaves in 1 L of water at 80 °C for 10 min.

. The first experiment consisted of five formulations combining different levels of pecan shell aqueous extract (PSE) prepared according to item 2.1, and green tea infusion (GTI) (5 g of green tea leaves in 1 L of water at 80 °C for 10 min) as fermentation substrates to evaluate the physicochemical parameters of fermentation and antioxidant activity of the formulations. After mixing the substrates in 2.5 L fermentation flasks, the liquid of the flasks was supplemented with 15% fermentation starter liquid, 80 g/L sucrose, and 25 g/L SCOBY (Symbiotic Culture of Bacteria and Yeasts). In the second experiment, the sucrose concentration was reduced to 60 g/L in two of the formulations from experiment 1, with the justification for the selection of the two formulations to be presented in Section 3.

All experiments were performed in duplicate. Even though only two fermentations were performed for each operating condition, it was possible to obtain consistent responses regarding the behavior of the fermentation process. The increase in the number of fermentation process replicates was not carried out in this study because, in addition to investigating the fermentation process of kombucha supplemented with pecan nut extract, the authors also aimed to develop a final product from the obtained fermented material.

The formulations were characterized through analyses of total titratable acidity, pH, total soluble solids, total reducing sugars, total phenolic compounds, and antioxidant activity using the ABTS [(2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)] radical scavenging assay. Samples were collected on 0, 3, 5, 7, 14, 21, 28, and 35 days. All analyses were accomplished in replicates.

2.3. Development Kombucha-Vinegar Beverages Inspired by Switchel—Step 3

The preparation of the kombucha-vinegar beverages inspired by switchel was based on formulation 7 from the previous step, with additions of ginger extract and apple, pineapple, or white grape juices, as shown in

Table 2. Formulations of kombucha-vinegar beverages inspired by switchel.

Formulation	KV (% v/v)	GE (% v/v)	PAJ (% v/v)	AJ (% v/v)	WGJ (% v/v)
G1	20	20	60	0	0
G2	20	20	0	60	0
G3	20	20	0	0	60
V1	20	0	80	0	0
V2	20	0	0	80	0
V3	20	0	0	0	80

KV: kombucha vinegar prepared based on Formulation 7 of step 2; GE: ginger extract; PAJ: pineapple juice; AJ: apple juice; WGJ: white grape juice.

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The ginger extract was prepared using a ratio of 200 g of ginger to 300 mL of water and blended in an industrial blender for 5 min. Subsequently, the extract was filtered, pasteurized (80 °C for 20 min), and stored in an amber bottle under refrigeration for later use. The 100% pure juices, with no added sugars, were purchased from local stores.

The raw materials (ginger extract), juices, and the formulations of kombucha-vinegar beverages inspired by switchel were characterized by measurements of total titratable acidity, pH, total soluble solids, total reducing sugars, total phenolic compounds, and antioxidant activity using the ABTS method. All characterization assays were performed in triplicate.

2.4. Analytical Determinations

2.4.1. Total Titratable Acidity, pH, and Total Soluble Solids

Total titratable acidity was determined by volumetric analysis using a standard 0.1 mol/L sodium hydroxide solution and phenolphthalein as indicator [24]. The pH was measured using a calibrated bench pH meter, and total soluble solids (ºBrix) were determined by refractometry according to the methods of the Association of Official Analytical Collaboration (A.O.A.C.) [25].

2.4.2. Total Reducing Sugars

The concentration of total reducing sugars was determined by the Miller method [26], with modifications, using dinitrosalicylic acid (DNS) solution as the oxidizing agent. Two mL of the sample and 2 mL of 2 mol/L HCl were added to a test tube. The mixture was heated in a water bath at 100 °C for 30 min and then cooled to room temperature. Subsequently, 2 mL of 2 mol/L NaOH was added, and the tubes were centrifuged at 5000 rpm for 20 min. After centrifugation, 1 mL of the supernatant was diluted in a 25 mL volumetric flask. From this solution, 1 mL was added to a test tube along with 1 mL of DNS and 1 mL of 1 mol/L NaOH. The mixture was heated again at 100 °C for 5 min and then cooled to room temperature. Finally, the volume was completed to 7 mL with distilled water, and the sample transmittance was measured at 560 nm using a spectrophotometer. Results were calculated using the calibration curve equation and expressed as grams of glucose per liter of sample (g/L).

2.4.3. Total Phenolic Compounds

The total phenolic compounds content was determined by the Folin–Ciocalteu method, adapted from Singleton et al. [27]. The reaction mixture was prepared by mixing 1 mL of the sample, 1 mL of 96% ethanol, 5 mL of distilled water, 0.5 mL of Folin–Ciocalteu reagent, and 1 mL of 5% (w/w) sodium carbonate solution in a test tube. After incubation at room temperature for 1 h, absorbance was measured at 765 nm in a spectrophotometer. Gallic acid was used as a calibration standard, and results were expressed as gallic acid equivalents per liter of sample.

2.4.4. Antioxidant Activity

The antioxidant activity, evaluated by the ABTS radical scavenging method [2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid)], was determined using a methodology adapted from Re et al. [28]. Initially, the ABTS cation radical was prepared by reacting ABTS stock solution (7 µmol/L) with potassium persulfate solution (2.45 µmol/L) in a 1:1 v/v ratio. This solution was left to stand in the dark at room temperature for 12 to 16 h before use. Prior to analysis, the ABTS solution was diluted with 96% ethanol until an absorbance of approximately 0.700 ± 0.05 at 734 nm was obtained. For the assay, 20 µL of sample was added to 980 µL of the diluted ABTS radical solution, mixed thoroughly, and incubated for 6 min protected from light. Absorbance was measured at 734 nm against an appropriate blank. Radical scavenging activity was calculated as a percentage inhibition using Equation (1) [29].

$$\% \mathrm{Inhibition}={\displaystyle \frac{{\mathrm{A}}_{\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l}}-{\mathrm{A}}_{\mathrm{s}\mathrm{a}\mathrm{m}\mathrm{p}\mathrm{l}\mathrm{e}}}{{\mathrm{A}}_{\mathrm{c}\mathrm{o}\mathrm{n}\mathrm{t}\mathrm{r}\mathrm{o}\mathrm{l}}}}\times 100$$

(1)

where A_control is the absorbance of the ABTS solution without the sample, and A_sample is the absorbance in the presence of the sample. Antioxidant capacity was expressed as Trolox equivalent antioxidant capacity (µmol TEAC/L) using a Trolox calibration curve constructed under the same experimental conditions.

2.5. Sensory Evaluation—Step 4

Acceptance of the kombucha-vinegar beverages inspired by switchel was evaluated using a verbal hedonic scale composed of nine categories, ranging from “1” (disliked extremely) to “9” (liked extremely). The attributes assessed were appearance, color, aroma, flavor, and overall impression.

To better understand consumers’ sensory perceptions regarding the functional switchel-type beverages, the CATA (Check All That Apply) methodology was used. A questionnaire containing descriptive and affective terms was developed based on literature data about descriptive characterization of kombuchas [30]. The terms included in the CATA lists covered sensory, emotional, and functional aspects. Panelists were instructed to mark all terms they deemed relevant to characterize the analyzed samples.

This research was approved by the Research Ethics Committee of the University of Passo Fundo, Brazil, under opinion number 5.864.827. All participants signed a Free and Informed Consent Form prior to the sensory test. A total of 100 untrained panelists were selected. The number of tasters was determined following the methodology described by Dalla Nora [31]. The panelists were recruited from the University of Passo Fundo, being students, employees, and professors of the institution, of both sexes, over 18 years of age, and who declared themselves consumers of beverages with a health appeal.

2.6. Statistical Analysis

Data were analyzed according to the experimental strategy used for beverage development. The independent variables were the proportion of pecan shell aqueous extract replacing green tea infusion during kombucha fermentation, the initial sugar concentration adjusted to obtain kombucha vinegar with reduced residual sugars, and the beverage formulations prepared with fruit juices or ginger.

The dependent variables included physicochemical parameters (pH, total titratable acidity, alcohol content, reducing sugars, and ash), total phenolic compounds, antioxidant activity, and sensory acceptance. Data normality was verified prior to analysis. Differences among treatments were evaluated by analysis of variance (ANOVA) followed by Tukey’s test (p ≤ 0.05) using Statistica 7.0 software (StatSoft, Tulsa, OK, USA).

Sensory acceptance data were analyzed by ANOVA and Tukey’s test (p < 0.05). For the Check-All-That-Apply (CATA) descriptive analysis, term citation frequencies were compared using Cochran’s Q test, followed by Correspondence Analysis performed in SPSS software version 28 (IBM, Armonk, NY, USA).

3. Results and Discussion

3.1. Characterization of Pecan Shell Extract—Step 1

The pecan shell extracts produced under the described conditions in this study (10% of pecan shell, 75 °C, 10 min) exhibited a total phenolic content of 10.4 ± 0.2 mg GAE/g (gallic acid equivalents per gram of fresh weight sample) and antioxidant capacity of 74.6 ± 3.0 µmol TEAC/g measured by the ABTS radical scavenging assay. The extraction method influences the recovery of compounds with antioxidant properties. In this study, previously, we investigated the extraction of phenolic compounds and antioxidant activity by varying pecan shell concentrations from 10 to 30%, temperatures from 75 to 85 °C, and infusion times from 10 to 30 min. The selected condition was the one that yielded the best results.

Water extraction was chosen because it allows the extract to be used directly in beverage formulation; however, the solvent is reported in the literature as an important factor in the extraction of phenolic compounds, including tannins, which are the main constituents of pecan shells [32,33]. Extraction processes may be influenced by solvent type [34,35], temperature, and extraction time. Bottari et al. [32] extracted several phenolic compounds from Carya illinoinensis using water or ethanol and concluded that, in general, the ethanolic extract tended to show higher concentrations of phenolic compounds and flavonoids, except for catechin and epicatechin. For example, values of 192.4 mg GAE/g (total phenolics) and 2218.8 ± 0.8 µmol TEAC/g (antioxidant activity) have been reported by Müller et al. [36] and Reckziegel et al. [37] for this type of extract, obtained under infusion conditions using a particle size of 60 mesh, shell concentrations of 10–25 g/L, at 98 °C for 20 min.

The extraction of high amounts of tannins, the main phenolic compound of pecan shell, may result in increased bitterness and astringency, which can limit their application in foods and beverages [38]. These compounds may exhibit strong antimicrobial activity [39,40]. Therefore, the lower extraction yield resulting from the process conducted at 75 °C, a temperature below boiling conditions that may promote higher compound extraction, can be considered advantageous, since the extract was used in the fermentative production of kombucha vinegar. Therefore, considering the aim of this study to obtain beverages with acceptable palatability, and the fact that green tea also contributes to phenolic compound content and antioxidant activity, the extraction conditions applied in this work were considered sufficient.

3.2. Production of Kombucha Vinegar—Step 2

3.2.1. Results of Experiment 1—Step 2

Figure 1 shows the values of total titratable acidity, pH, total reducing sugars, and total soluble solids of the formulations at the initial and final fermentation times for Experiment 1 (Table 1). pH and total titratable acidity (TTA) are complementary and important parameters for kombucha quality control. pH is the main indicator of product safety and microbial selectivity, whereas TTA quantifies the amount of acids formed and helps define the end of fermentation as well as the sensory profile. Monitoring both parameters allows better control of quality, safety, and reproducibility of the fermentation process [41]. In kombucha production, a pH range between 2.5 and 4.2 is generally recommended; values below 2.5 may be harmful to consumers. However, in the present study, since kombucha vinegar was used as an ingredient in beverage formulations, acidification beyond this range would not have been considered a problem. TTA tends to increase continuously over time due to the formation of organic acids [42], and this parameter is used by several authors to determine the end of fermentation [43].

The results indicated a significant difference (p < 0.05) in total titratable acidity values among the formulations on the 35th day of fermentation. Acidic kombucha produced by Kaewkod et al. [44] showed organic acid concentrations of 57.26, 63.95, and 91.82 g/L using green, oolong, and black teas, respectively, after 10–15 days of fermentation. These values correspond to percentages ranging from 5.7 to 9.18%. In our study, percentages around 12% were obtained (Figure 1a) after 35 days of fermentation. The substrates used in kombucha-vinegar production influence acid formation [5,30,45]. Therefore, the use of pecan shell extract as a substrate, rich in compounds such as condensed tannins [46], due to its antimicrobial activity, may have reduced SCOBY bacterial activity, resulting in lower acid production.

The pH of the formulations decreased more rapidly during the first five days, and subsequently, the changes became slower. Acetic acid, responsible for the acidic aroma and vinegar-like flavors, is normally present in solutions fermented by kombucha culture and may have contributed to the pH reduction [15,47]. Since kombucha vinegar is intended to replace apple cider vinegar in our proposed beverage, these are desirable characteristics. After inoculation, the formulations had a pH ranging from 4.20 to 4.62, and these values decreased to between 2.84 and 2.70 at the end of fermentation. Previous studies have reported similar pH values in fermentations with kombucha culture to identify the end of the fermentation process [5,30,48].

Over 35 days, a decrease in total reducing sugars was observed in all formulations, with reductions ranging from 23.53% (F1) to 55.58% (F5) after fermentation (Figure 2). This decrease was expected, as the added sucrose was utilized by yeasts and acetic acid bacteria, which metabolize these sugars and convert them into ethanol and organic acids, mainly acetic acid [49]. In F1, which presented the highest percentage of pecan shell extract, the presence of tannins with antimicrobial properties may have influenced the fermentation process [35].

Total soluble solids, measured in ºBrix, consist not only of sugars but also of organic acids, and showed a reduction during the process (Figure 1). Previous authors have supported the idea that the bacteria and yeasts present in the SCOBY utilize sugars and organic acids as nutrients to promote their growth and metabolism; therefore, biomass yield may be directly related to the decrease in sugar content during fermentation [9,50].

Table 3. Total phenolic compounds (TPC) and antioxidant capacity determined by ABTS radical scavenging assay for the formulations from Stage 1 at initial time and after 35 days of fermentation.

Form.	F1	F2	F3	F4	F5
	TFC (mg EAG/L)
0 days	293.59 ± 6.58 c	408.70 ± 3.61 a	440.63 ± 4.48 a	654.28 ± 3.13 b	681.09 ± 5.96 b
35 days	222.25 ± 5.81 c	386.08 ± 44.08 a	462.67 ± 13.67 a	575.76 ± 8.20 b	667.09 ± 36.91 b
	ABTS (µmol TEAC/L)
0 days	1125.56 ± 142.40 c	2020.00 ± 179.00 a	3342.22 ± 259.27 a	4975.56 ± 218.54 b	6528.89 ± 117.85 b
35 days	3253.33 ± 36.14 a	5064.44 ± 13.35 ab	5747.78 ± 75.42 b	7920.00 ± 62.85 c	8320.00 ± 29.07 c
Var (%)	189.04 (+)	150.72 (+)	71.97 (+)	59.18 (+)	32.93 (+)

Different letters within rows indicate statistically significant differences between samples according to Tukey’s test (p < 0.05). Results are expressed as mean ± standard deviation (n = 3). Form: Formulation; Var: Variation; F1(100% PSE; 0% GTI); F2(75% PSE; 25% GTI); F3(50% PSE; 50% GTI); F4(25% PSE; 75% GTI); F5(0% PSE; 100% GTI). PSE: pecan shell extract; GTI: green tea infusion; TPC: total phenolic compounds; ABTS: antioxidant activity by the ABTS radical scavenging assay.

presents the concentrations of total phenolic compounds during the production of kombucha vinegars from Experiment 1, on 0 and 35 days; however, they showed differences among themselves (p < 0.05, Table 3). At both times, the highest contents were observed in formulations F4 and F5, which were prepared with 75% and 100% green tea, respectively, with no significant difference between these samples. Some studies report an increase in polyphenol content during kombucha fermentation [47,51,52], due to the biotransformation of molecules by enzymes produced by microorganisms present in the SCOBY and starter liquid. In our study, this was not observed, and the phenolic content was maintained without significant differences during the fermentation time. Little variation was observed, which may be associated with the oxidation of compounds, influenced by oxygen levels and the low pH value [52]. Furthermore, the total amount of polyphenols is related to the type of substrate used in each formulation [53]. When higher percentages of green tea were used, higher quantities of phenolics were observed, similar to the results reported by Jakubczyk et al. [54], where fermentation in green tea substrate showed higher polyphenol levels than fermentation in black tea substrate.

The antioxidant activity of the formulations measured by the ABTS radical scavenging assay was correlated with the phenolic content of the samples (Figure 2). Similar to total phenolic compounds, the highest antioxidant activities were also observed in formulations F4 and F5. Antioxidant activity of the extracts increased over the fermentation time (Table 3; p < 0.05), with variations varying from 32.9 to 189% (Table 3).

The content and composition of bioactive compounds in kombucha-fermented products depend on the raw materials used as well as the level and type of sugar added to the fermentation substrate [55]. In this study, the results indicate that the higher the level of green tea extract in the substrate, the greater the antioxidant activities of the formulations. Green tea is composed of flavonoids, such as catechins [56,57], while pecan nutshells mainly contain proanthocyanidins and ellagic acid [58]. Therefore, the differences in phenolic profiles reported in the literature for these two substrates (pecan shell and green tea) may explain the results obtained in this research.

These findings support previous studies indicating that catechins are degraded by bacterial and yeast activity, transforming into simpler molecules that enhance antioxidant effects [13,54]. In contrast to green tea, the aqueous extract of pecan shell is not a source of catechins, which may explain the lower antioxidant activity observed by ABTS radical scavenging assay in formulations containing this raw material as the major substrate. However, according to other authors, the aqueous extract of pecan shell contains other molecules, including gallic acid, ellagic acid, and condensed tannins, all with antioxidant and functional properties [22,33,58,59]. Additionally, it was observed that the positive variation in values between days 0 and 35, by the ABTS radical scavenging assay, was greater in samples containing higher amounts of aqueous extract of pecan shell. This highlights that the fermentation process using kombucha culture has potential effectiveness in enhancing the antioxidant activity of alternative substrates [13,50,60,61].

3.2.2. Results of Experiment 2—Step 2

Beverages with health claims are developed by adding beneficial ingredients, such as vitamins, and/or by reducing or removing negative ingredients like sugar. Sugar reduction in beverages is vital to promote healthier food choices, prevent diet-related diseases, and meet the growing consumer demand for products that contribute to a healthier lifestyle [62]. The residual total reducing sugar values of the kombucha vinegars from Experiment 1 were considered high. According to the World Health Organization (WHO), no more than 10% of daily calories should come from sugar consumption. Considering a 2000-calorie diet, this percentage equals 50 g of sugar per day (about 10 teaspoons).

In order to prevent the kombucha vinegar from contributing large amounts of sugars when used in switchel formulations, experiments were conducted to reduce the initial sucrose concentration. The concentration of sucrose in Experiment 1—Step 2 was 80 g/L (considered an ideal concentration for the beginning of fermentation) [63]. It was reduced to 60 g/L, based on the formulations from Experiment 1 (F3 and F4). The pecan extract was used at levels ranging from 25 (F4) to 50% (F3), while green tea remained at 50% (F3) to 75% (F4). These formulations presented a lower proportion of pecan extract in comparison with formulations F1 and F2, and this choice was justified by the bitterness it imparts to the beverage. Moreover, the use of 100% green tea in the formulation (F5) did not significantly increase antioxidant activity or phenolic compound content in comparison with the formulation F4 (p > 0.05), which further supports the selection of formulations F3 and F4 for the next stage.

Although a higher proportion of green tea was used, the incorporation of pecan allows for the valorization of a waste material in place of a more costly raw ingredient. Moreover, the fermentation process was shown to enhance the availability of antioxidant potential derived from the compounds present in the raw material blend. Thus, beyond developing a beverage with antioxidant potential, the study also aimed to promote the use of a waste-derived raw material, contributing to the advancement of the circular bioeconomy.

The physicochemical and antioxidant properties of the kombucha vinegars from Experiment 2—Step 2 (Table 1) are presented in

Table 4. Physicochemical and antioxidant properties of kombucha vinegars of the Experiment 2—Step 2 at the end of fermentation (35 days).

Parameter Evaluated	Formulation
	F6	F7	F8	F9
TTA (expressed as % acetic acid)	1.86 ± 0.11 a	1.80 ± 0.11 a	1.98 ± 0.12 a	1.84 ± 0.12 a
TRS (g/L)	10.65 ± 0.55 a	16.96 ± 0.08 b	8.10 ± 1.27 a	16.50 ± 0.68 b
TSS (ºBrix)	3.65 ± 0.07 c	5.25 ± 0.07 a	3.15 ± 0.21 b	4.95 ± 0.07 a
pH	2.68 ± 0.05 a	2.62 ± 0.03 a	2.76 ± 0.04 a	2.72 ± 0.02 a
TPC (mg EAG/L)	538.39 ± 15.05 a	522.03 ± 1.39 a	645.41 ± 3.62 b	640.88 ± 14.49 b
ABTS (µmol TEAC/L)	5361.64 ± 26.32 a	5081.11 ± 133.56 a	7078.30 ± 113.92 b	7031.11 ± 47.14 b

Different letters within rows indicate statistically significant differences among samples by Tukey’s test (p < 0.05). Results are expressed as mean ± standard deviation (n = 3). TTA: total titratable acidity; TRS: total reducing sugars; TPC: total phenolic compounds; ABTS: antioxidant capacity by ABTS radical scavenging assay; Formulations: F6 (50% PSE; 50% GTI; 80 g/L of sucrose); F7 (50% PSE; 50% GTI; 60 g/L of sucrose); F8 (25% PSE; 75% GTI; 80 g/L of sucrose); F9 (25% PSE; 75% GTI; 60 g/L of sucrose). PSE: pecan shell extract; GTI: green tea infusion.

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The results of the assays related to TTA (total titratable acidity) (%) and pH showed no significant difference between the samples (p > 0.05) after 35 days of fermentation (Table 4). The total reducing sugars (TTS) and total soluble solids (TSS) of the kombucha vinegars from Experiment 2—step 2 showed statistically significant differences among themselves (p < 0.05). The results were linear with respect to the sucrose content added at the start of fermentation, with the formulation containing the lowest sugar concentration showing the lowest residual sugar content, while the formulation with the highest sugar concentration showed the highest residual sugar content. The total soluble solids behaved in the same way. Several factors affect glucose consumption: the type of substrate, the microorganisms present in the SCOBY, and fermentation conditions (temperature and time). Therefore, comparison among kombucha fermentations produced from different raw materials is a difficult task due to variation in fermentation conditions and SCOBY, resulting in products with diverse characteristics. However, in this work, our results clarified that the levels of association between pecan shell aqueous extract and green tea did not influence the sugar consumption process (p > 0.05). Thus, reducing the amount of sucrose added to the fermentation substrate is beneficial for lowering total reducing sugars in kombucha vinegars, allowing the use of substrate associations of 50:50 (v/v) or 25:75 (v/v) (PSE:GTI) without affecting pH and acidity parameters.

Supporting the results of Experiment 1, our findings indicate that the total phenolic compound content and antioxidant capacity of the kombucha vinegars are strongly influenced by the type of substrate [64]. Total phenolic compounds and antioxidant capacities showed significantly different results (p < 0.05) between samples with different substrate association levels (Table 4). Contrary to other studies [9,55], the level of added sucrose was not significant (p > 0.05).

3.3. Results of Development Kombucha-Vinegar Beverages Inspired by Switchel—Step 3

Table 5. Characterization of ginger extract and fruit juices used as ingredients of kombucha-vinegar beverages inspired by switchel.

Ingredient	Physicochemical and Antioxidant Parameters
	TTA	TRS	TSS	pH	TPC mg EAG/L	ABTS
	% Acetic Acid	g/L	ºBrix			(µmol TEAC/L)
GE	0.15 ± 0.04 a	4.79 ± 0.02 b	1.45 ± 0.21 c	6.44 ± 0.01 d	393.76 ± 1.37 c	4521.11 ± 217.52 a
PAJ	0.51 ± 0.04 b	54.04 ± 0.92 a	11.90 ± 0.01 b	3.85 ± 0.01 b	310.78 ± 12.69 b	980.33 ± 4.41 c
AJ	0.27 ± 0.04 a	55.26 ± 0.50 a	12.70 ± 0.02 b	3.89 ± 0.02 c	489.11 ± 4.48 d	4018.33 ± 267.84 a
WGJ	0.51 ± 0.04 b	74.49 ± 0.65 c	15.20 ± 0.02 a	3.24 ± 0.02 a	234.90 ± 1.90 a	1978.33 ± 129.64 b

Different letters in the columns indicate statistically significant differences between samples by Tukey’s test (p < 0.05). TTA: total titratable acidity (%); TRS: total reducing sugars g/L; TSS: total soluble solids % (w/w); pH: potential of hydrogen; TPC: total phenolic compounds mg EAG/L; ABTS: antioxidant activity by the ABTS radical scavenging assay. GE: ginger extract; PAJ: pineapple juice; AJ: apple juice; WGJ: white grape juice.

presents the results of the characterization of the ginger extract and fruit juices used in the development of the beverages. All samples showed significantly different values among themselves (p < 0.05).

The pineapple juice and white grape juice samples showed the highest total titratable acidity. The white grape juice also recorded the highest values for both total reducing sugars and total soluble solids, while the lowest values were observed in the ginger extract. Regarding pH, the white grape juice showed the lowest value. Concerning functional properties, the apple juice presented the highest content of total phenolic compounds, while the white grape juice exhibited the lowest value. In terms of antioxidant activity evaluated by the ABTS radical scavenging assay, the ginger extract and apple juice demonstrated the highest values, whereas pineapple juice recorded the lowest values.

Table 6. Physicochemical and functional characterization of kombucha-vinegar beverages inspired by switchel.

Formulation	Physicochemical and Antioxidant Parameters
	TAA (Expressed as % Acetic Acid)	TRS (g/L)	TSS (ºBrix)	pH	TPC (mg EAG/L)	ABTS (µmol TEAC/L)
G1	0.57 ± 0.40 abc	30.81 ± 0.71 a	7.25 ± 0.07 a	3.55 ± 0.05 a	374.64 ± 10.51 c	2104.32 ± 293.09 a
G2	0.45 ± 0.40 a	41.26 ± 0.32 b	8.60 ± 0.14 a	3.44 ± 0.17 e	297.38 ± 7.85 b	2329.17 ± 514.98 a
G3	0.63 ± 0.40 bcd	32.22 ± 0.92 a	10.15 ± 0.21 a	3.11 ± 0.20 c	213.81 ± 6.26 a	2204.44 ± 114.97 a
V1	0.78 ± 0.40 d	44.16 ± 1.30 c	9.85 ± 0.07 a	3.52 ± 1.55 a	485.80 ± 7.82 e	2798.59 ± 528.65 b
V2	0.51 ± 0.40 ab	56.58 ± 1.38 d	10.40 ± 0.00 a	2.39 ± 1.43 d	404.99 ± 20.66 d	3348.89 ± 226.7 b
V3	0.72 ± 0.40 cd	42.14 ± 0.98 bc	13.10 ± 0.00 a	3.01 ± 1.71 b	272.54 ± 4.92 b	2204.44 ± 103.53 a

TTA: total titratable acidity; TRS: total reducing sugars; TSS: total soluble solids; pH: hydrogen potential; TPC: total phenolic compounds; ABTS: antioxidant capacity by the ABTS radical scavenging assay; KV: kombucha vinegar; GE: ginger extract; PAJ: pineapple juice; AJ: apple juice; WGJ: white grape juice. G1 (20% KV, 20% GE, and 60% PAJ). G2 (20% KV, 20% GE, and 60% AJ). G3 (20% KV, 20% GE, and 60% WGJ). V1 (20% KV, 80% PAJ). V2 (20% KV, 80% AJ). V3 (20% KV, 80% WGJ). Different letters in the columns indicate statistically significant differences between samples by Tukey’s test (p < 0.05). Results are expressed as means of three determinations ± standard deviation.

presents the results of the physicochemical characterization of the kombucha-vinegar beverages inspired by switchel. Formulation 7 from the previous stage was selected for beverage development because it used a higher amount of pecan shell extract (50% PSE and 50% GTI) and a lower amount of sugar at the beginning of fermentation (60 g/L), and it showed favorable physicochemical parameters and antioxidant characterization. The titratable acidity of the samples ranged from 0.45 to 0.78%, with significant differences (p < 0.05) among the formulations. The samples showed differences related to the addition of ginger extract in the formulation, with those containing this ingredient (G1, G2, and G3) presenting lower values. For the pH parameter, results varied from 2.39 to 3.55, with significant differences (p < 0.05) among the formulations. These values fall within the range considered safe for human consumption, which varies from 2.5 to 4.2. Values below pH 2.5 indicate a high concentration of acetic acid, posing a health risk to consumers. Similarly, pH values above 4.2 may compromise the microbiological safety of the beverage [65].

Regarding total soluble solids, expressed in ºBrix, no significant differences were observed among the samples (p > 0.05), although the beverages G3 and V3, flavored with grape juice, showed the highest values. The amount of total reducing sugars (g/L) showed significant differences (p < 0.05) and varied according to the type of juice used for flavoring the beverages, with beverages flavored with pineapple juice registering the lowest values (G1 and V1), while those with grape juice presented the highest values (G3 and V3).

Consumers show a strong preference for foods and beverages that offer health benefits. The analysis of phenolic compound content in kombucha-vinegar beverages inspired by switchel showed significant variation, ranging from 213.81 ± 6.26 mg EAG/L (G3) to 485.80 ± 7.82 mg EAG/L (V1) (Table 6). The results were influenced by the fruit juice added, and the beverages prepared without ginger extract (V1 and V2) presented the highest antioxidant capacity values. Beverages containing white grape juice (G3 and V3) presented the lowest values in the total phenolic compounds assay, with values of 213.81 ± 6.26 mg EAG/L and 272.54 ± 4.92 mg EAG/L, respectively.

3.4. Sensory Evaluation—Step 4

In addition to instrumental analysis, the evaluation of perceived attributes and consumer acceptance is important data for a deeper understanding of how processes affect food characteristics. Kombucha-vinegar beverages inspired by switchel were evaluated through an acceptance test using a 9-point hedonic scale and the CATA (Check-All-That-Apply) test. The results of the analysis of variance (ANOVA) for acceptance showed that the panelists found a significant difference (p < 0.05) only for the aroma attribute.

Table 7. Mean acceptance scores for Kombucha-vinegar beverages inspired by switchel regarding the attributes.

Formulation	Attributes					Acceptability Index (%)
	Appearance	Color	Aroma	Flavor	Overall Impression
G1	6.58 ± 1.80 a	6.58 ± 1.72 a	7.12 ± 2.13 ab	6.70 ± 2.13 a	6.94 ± 1.93 a	77.11
G2	6.68 ± 1.57 a	6.70 ± 1.54 a	6.76 ± 1.82 ab	6.50 ± 1.82 a	6.78 ± 1.84 a	75.33
G3	6.38 ± 1.76 a	6.48 ± 1.63 a	6.54 ± 2.04 ab	6.52 ± 2.04 a	6.70 ± 1.96 a	74.44
V1	6.72 ± 1.92 a	6.44 ± 2.05 a	7.46 ± 1.64 b	7.54 ± 1.64 a	7.56 ± 1.61 a	84.00
V2	7.30 ± 1.72 a	7.30 ± 1.63 a	6.46 ± 1.86 ab	7.48 ± 1.86 a	7.48 ± 1.69 a	83.11
V3	7.18 ± 1.79 a	7.12 ± 1.79 a	6.34 ± 1.80 a	6.92 ± 1.80 a	7.08 ± 2.08 a	78.67

KV: kombucha vinegar; GE: ginger extract; PAJ: pineapple juice; AJ: apple juice; WGJ: white grape juice). G1 (20% KV, 20% GE, and 60% PAJ). G2 (20% KV, 20% GE, and 60% AJ). G3 (20% KV, 20% GE, and 60% WGJ). V1 (20% KV, 80% PAJ). V2 (20% KV, 80% AJ). V3 (20% KV, 80% WGJ). Different letters in the columns indicate statistically significant differences between samples according to Tukey’s test (p < 0.05). Results are expressed as mean ± standard deviation.

presents the mean acceptance values for the attributes of appearance, color, aroma, flavor, and overall impression, as well as the acceptability index of the samples, standard deviations obtained, and significant differences identified by Tukey’s test.

Although they did not differ significantly from the other formulations, the appearance, color, and flavor of the beverages formulated without ginger extract (V1, V2, and V3) received higher ratings. For the aroma attribute, the beverages that stood out in quality were those containing pineapple juice in the formulation (G1 and V1), with no significant difference between the sample containing ginger extract and the one containing only kombucha vinegar. Scores for overall impression and, consequently, the acceptability index were higher for the samples without ginger extract in their composition. Regarding the developed formulations, it is relevant to highlight that most of the means obtained were within the range of the concept “liked slightly” (above 6) for all evaluated attributes. These results indicate good potential acceptance by consumers. On the other hand, the formulations containing ginger (G1, G2, and G3) showed means below 7 for all attributes. In contrast, samples prepared only with fruit juice and kombucha vinegar (V1, V2, and V3), with means above 7, exhibited the best sensory characteristics and were well accepted by consumers. Concerning the frequencies of the hedonic values assigned, there is a predominance of scores ranging from “liked slightly” to “liked very much”, common to all formulations.

In the consumer-based methodology employed in this work, sensory descriptor terms are divided by attributes. The results of Cochran’s Q test and multiple comparisons for the CATA questionnaire are presented in

Table 8. Frequency of descriptive terms and Cochran’s Q test results for the CATA questionnaire.

Attributes	Descriptive Terms	Formulations						Total
		G1	G2	G3	V1	V2	V3
Flavor	Acid	35 ab	30 ab	37 a	32 ab	22 b	33 ab	189
	Sweet	16 bc	19 b	14 bc	31 a	37 a	33 a	151
	Alcoholic	2 ab	1 ab	0 b	1 ab	7 a	7 a	18
	Bitter	9 ab	4 ab	9 ab	1 b	11 a	1 b	35
	Tasty	19 b	25 b	20 b	40 a	40 a	28 ab	172
	Vinegar	5 b	10 b	11 ab	11 ab	24 a	18 ab	79
	Strong	35 a	28 ab	23 ab	14 bc	10 c	20 b	130
	Sour	7 a	7 a	13 a	7 a	4 a	8 a	46
	Fermented	12 ab	12 ab	10 b	17 ab	15 ab	23 a	89
	Exotic	11 a	13 a	8 a	14 a	9 a	20 a	75
	Astringent	3 ab	3 ab	5 ab	10 a	2 b	3 ab	26
	Spicy	25 a	27 a	31 a	6 b	2 b	5 b	96
Aroma	Strong	18 a	16 a	16 a	22 a	12 a	16 a	100
	Alcoholic	1 b	0 b	2 b	9 a	14 a	9 a	35
	Tasty	30 ab	28 ab	28 ab	40 a	31 ab	24 b	181
	Vinegar	7 b	15 b	13 b	13 b	24 a	22 a	94
	Fermented	12 a	10 a	10 a	17 a	16 a	17 a	82
	Exotic	13 a	14 a	11 a	17 a	13 a	13 a	81
Appearance	Clear	17 b	18 b	16 b	16 b	38 a	35 a	141
	Translucent	1 b	0 b	1 b	8 b	24 a	26 a	60
	Dark	10 a	2 ab	4 ab	7 ab	1 b	0 b	26
	Cloudy	38 a	39 a	31 a	41 a	5 b	5 b	159
Impressions	Energizing	19 a	27 a	21 a	26 a	28 a	25 a	146
	For the whole family	10 b	4 b	6 b	24 a	23 a	13 ab	80
	Good for breakfast	3 b	4 ab	2 b	13 a	10 ab	7 ab	39
	A healthy option	25 ab	25 ab	28 a	14 b	31 a	31 a	174
	Good for physical activities	11 ab	6 b	11 ab	14 ab	21 a	9 ab	72
	Perfect for dieting	11 a	12 a	14 a	8 a	14 a	12 a	75
	Refreshing	33 a	33 a	35 a	44 a	43 a	40 a	228

KV: kombucha vinegar; GE: ginger extract; PAJ: pineapple juice; AJ: apple juice; WGJ: white grape juice). G1 (20% KV, 20% GE, and 60% PAJ). G2 (20% KV, 20% GE, and 60% AJ). G3 (20% KV, 20% GE, and 60% WGJ). V1 (20% KV, 80% PAJ). V2 (20% KV, 80% AJ). V3 (20% KV, 80% WGJ). Note: Different letters within rows indicate significant difference (p < 0.05) between formulations by Cochran’s Q test adjusted by the Bonferroni method.

. The Bonferroni adjustment was used to correct the results of the multiple comparison tests. This method aims to control the false positive rates by reducing the individual significance level for each test performed. This reduction is achieved by dividing the significance level by the total number of tests performed, thus generating a new adjusted significance level for each test.

In a general analysis, the terms with the highest frequency of citation for the six beverage formulations were, for the flavor attribute: acidic, tasty, sweet, and strong; for the aroma attribute: tasty, strong, and vinegar; for the appearance attribute: clear and cloudy; and for impressions: “refreshing”, “a healthy option”, and “energizing”.

The terms with the lowest frequency of citation for the six beverage formulations were, for the flavor attribute: alcoholic, astringent, bitter, and sour; for the aroma attribute: alcoholic; for the appearance attribute: dark; and for impressions: “good for breakfast”.

For the flavor attribute, no significant differences were observed among the samples for the terms “sour” and “exotic”. However, the samples containing ginger extract in the formulation (G1, G2, and G3) showed a significantly higher citation frequency for the descriptors “acidic”, “bitter”, “strong”, and “spicy”, while the terms “sweet”, “vinegar”, and “fermented” were more associated with the samples formulated only with kombucha vinegar and fruit juice (V1, V2, and V3). The results of the present study corroborate those found by Dartora et al. [5,30], where the frequency of the attributes “vinegar aroma”, “flavor”, and “alcoholic” increased in the sample subjected to prolonged fermentation of green tea by kombucha culture. The results for the terms “alcoholic” and “adstringent” were also significantly different among the samples.

For the aroma attribute, the terms “strong”, “fermented”, and “exotic” did not show significant differences in the results. This may be attributed to the raw materials used, since both beverages containing ginger and those containing vinegar have previously been described by these terms [66,67]. However, the terms “alcoholic” and “vinegar” showed significant differences between samples, with those not containing ginger extract in the formulation (V1, V2, and V3) having the highest citation frequency for these terms. These samples also had higher acid concentrations (Table 6), and these terms have been associated with volatile compounds frequently identified in beverages fermented by kombucha culture, such as acids, higher alcohols, aldehydes, ketones, and esters [30,68].

For the term “tasty”, the samples that stood out were those containing pineapple juice in the formulation (G1 and V1), regardless of whether they contained ginger extract or not. These results highlight the panelists’ preference for pineapple juice compared to apple and grape juices. This preference was also evidenced in other studies on the acceptance of functional beverages made from pineapple juice [69,70].

For the appearance attribute, all descriptive terms showed significant differences. The terms “clear” and “translucent” were more frequently cited for the samples that did not contain ginger extract in their formulation (V1, V2, and V3), whereas the terms “dark” and “cloudy” were more associated with the samples that did contain it (G1, G2, and G3). The inclusion of ginger extract in beverages may alter the color of the mixture, resulting in yellowish or brownish hues due to the gingerols naturally present in ginger. Additionally, the presence of fine particles or solids in the extract can affect the clarity of the beverage, potentially causing turbidity [67]. However, it is important to note that sample V1, which contained only kombucha vinegar and pineapple juice in its formulation, also received a high frequency of citations for these terms. Pineapple juice is known for its vibrant color, characterized by intense yellow tones. Nevertheless, variations in the pineapple juice extraction process can impact the turbidity of the final beverage [71].

Regarding citation frequencies for impressions, the terms “energizing” and “perfect for dieting” did not show significant differences. However, in samples containing only vinegar and juice in their formulation (V1, V2, and V3), the frequency of citations for the term “energizing” exceeded 50%. These results are consistent with the assessors’ association of vinegar-based beverages with a healthy lifestyle. The properties of vinegar have been widely studied and publicized in recent years, and its flavor creates a sensory experience associated with stimulation and alertness [72].

Among these studies, Bang et al. [73] reported that the acetic acid present in vinegar can activate enzymes related to metabolism, promoting efficient calorie burning and a sensation of increased energy [2]. Furthermore, vinegar can enhance satiety, making it appealing for those seeking low-calorie options during dieting [74]. Studies also indicate that vinegar consumption may help regulate blood glucose levels, providing more stable energy by avoiding peaks and drops [75,76].

The same samples (V1, V2, and V3) also showed higher citation frequencies and significant differences among samples for the impressions “for the whole family”, “good for breakfast”, “good for physical activities”, and “refreshing”. These results align with the insights from the Global Food and Drink Trends Guide [77], which highlights that consumers are concerned not only with the healthfulness of food but also with its sensory attributes. These trends promote the dissemination of innovative recipes, sparking growing interest in food and beverage pairings, as well as the exploration of new ingredients and flavors.

Correspondence analysis is widely used to visualize contingency tables of citation frequencies for attributes obtained through the CATA test, and the results showed that variability among samples could be explained by 83.73% for the flavor attribute, 95.36% for aroma, 97.78% for appearance, and 88.36% for impressions, using two main factors in the first and second dimensions, as illustrated in Figure 3.

For the flavor attribute, the formulations that included ginger extract (G1, G2, and G3) were described as acidic, strong, spicy, and sour, while sample V2, formulated with kombucha vinegar and apple juice, was classified as the sweetest and noted for its vinegar aftertaste. Samples V1 and V3 were associated with the terms “exotic” and “fermented”, while the term “tasty” was more frequently related to the samples that did not contain ginger extract (V1, V2, and V3).

For the aroma attribute, samples G1 and V1, both containing pineapple juice in their composition, were associated with the terms “strong” and “pleasant”. Samples V2 and V3 were related to the terms “fermented” and “vinegar”, respectively. Samples G2 and G3 were associated with the term “exotic”.

Regarding appearance, the terms “cloudy” and “dark” described samples G1 and V1, both containing pineapple juice. Samples G2 and G3, which included ginger extract in their formulation, were also classified as “cloudy”. In contrast, samples V2 and V3 were described by the attributes “translucent” and “clear”.

As for impressions, sample V1 was considered “ideal for breakfast”, while samples G2 and V3 were described as “energizing, healthy options, and refreshing”. The term “perfect for dieting” was associated with samples G1 and G3, and sample V2 was related to the terms “good to accompany meals”, “for the whole family”, and “good for physical activities”.

4. Limitations

This study presents some limitations that should be considered. First, the fermentation experiments were conducted in duplicate, which may limit the robustness and reproducibility of the results, especially considering the inherent variability of kombucha fermentation systems. However, it is important to highlight that the main objective of this work was the development of a new product using kombucha vinegar in a beverage inspired by switchel. In addition, the experiments were carried out under controlled laboratory conditions, which may not fully represent industrial-scale processes, where factors such as process variability, contamination risks, and operational constraints can influence product characteristics.

The sensory evaluation was conducted with untrained panelists; however, the method employed is appropriate for this type of panelist. Nevertheless, increasing the number of panelists would be necessary to better assess acceptance across broader populations with different cultural backgrounds and consumption habits. Additionally, an important limitation of the study is the lack of detailed characterization of antioxidant compounds, which would allow a better understanding of how the fermentation process acts on the raw materials, promoting an increase in antioxidant activity, especially in the case of the residual raw material used, the pecan shell extract.

As this is an exploratory study focused on the development of a new beverage, several opportunities for further research can be considered. The microbiological characterization of the SCOBY and the evaluation of fermentation dynamics could provide deeper insights into the metabolic pathways responsible for the observed changes. Furthermore, aspects related to shelf-life stability, safety over time, and storage conditions of the developed beverages were not evaluated, although they are essential for future industrial applications and product commercialization.

5. Conclusions

The results demonstrated that substrate composition significantly influenced the physicochemical characteristics, antioxidant activity, and sensory properties of the inspired switchel beverages developed with kombucha vinegars made with pecan shell aqueous extract and green tea.

Fermentation promoted changes in pH, acidity, and sugar content, confirming its role in defining the final physicochemical profile of kombucha vinegar. Antioxidant activity was influenced by both substrate composition and fermentation, with higher proportions of green tea leading to greater phenolic content and antioxidant capacity, while pecan shell extract contributed to antioxidant enhancement throughout fermentation.

The reduction of sucrose from 80 to 60 g/L decreased residual sugar content without significantly affecting acidity, pH, total phenolics, or antioxidant activity, indicating that lower-sugar formulations can be obtained without compromising functional properties. Formulations containing intermediate proportions of pecan shell extract and green tea (50:50 and 25:75, v/v) showed a balance between physicochemical stability and antioxidant activities.

Sensory evaluation indicated that the developed beverages were generally well accepted, with acceptability indices above 70%, and that formulation components, such as the presence of ginger or fruit juices, influenced consumer perception.

The findings demonstrate that the use of pecan shell extract and green tea is a viable approach for producing kombucha vinegar and switchel-inspired beverages.