Sensory evaluation of a novel sustainable probiotic beverage based on tofu whey

Abstract

The reuse of agro-industrial residues aligns with sustainability purposes and the circular economy; in this context, tofu whey - a nutrient-rich by-product of tofu production - is a promising substrate for functional beverages. This study aimed to assess the sensory attributes of a potential probiotic plant-based beverage from tofu whey fermented using Lactiplantibacillus plantarum and Kluyveromyces marxianus. The strains were cultivated separately and inoculated at 1×10⁶ CFU/mL, followed by 48 h of co-culture fermentation at 37 °C. Samples were analyzed for pH, microbial growth, and metabolites via HPLC. Formulations were prepared with concentrated apple juice or demerara sugar and compared to non-fermented tofu whey in a sensory evaluation with 98 panelists. Fermentation reduced the pH from 5.92 to 3.95, promoted acetic acid production, and consumed residual sugars, while inhibiting ethanol formation. Beverages with demerara sugar presented the highest sensory acceptance due to their balanced sweetness and smoothness, whereas apple juice formulations were moderately rated. The results suggested that enhancing sweetness and texture while minimizing acidity and bitterness can improve acceptance, demonstrating tofu whey’s potential as a sustainable and functional ingredient for innovative plant-based beverages.

Keywords:
Agro-industrial by-products; Sensory acceptance; Lactiplantibacillus plantarum; Kluyveromyces marxianus; Fermentation; Co-culture; Check-All-That-Apply (CATA)

Highlights

Co-culture fermentation reduced pH and enhanced probiotic viability

Tofu whey was successfully valorized into a sustainable probiotic beverage

Demerara sugar improved sensory acceptance and overall liking

1 Introduction

In today’s world, where healthier and more sustainable lifestyles are increasingly valued, there is a growing need to develop alternative natural products that can be applied on a large scale in the food industry and beyond. Agro-industrial residues from soy have emerged as promising raw materials for producing value-added bioproducts through biotechnological and fermentative processes (Chua & Liu, 2019).

Tofu is a traditional Asian food whose popularity has been increasing worldwide and is produced by coagulating soy milk similarly to cheese-making. Such a rising tofu consumption has led to an increase in the volume of tofu whey, a nutrient-rich by-product, thus raising environmental concerns due to the disposal of this wastewater (Corzo-Martínez et al., 2016).

The reuse of tofu whey aligns with sustainability purposes and the circular economy. Transforming agro-industrial residues into new food products not only adds value and creates alternative income streams but also reduces environmental impacts while promoting more efficient use of natural resources (Silva et al., 2023; Oliveira & Mendes, 2021). Rich in proteins, carbohydrates, and minerals, tofu whey is an attractive substrate for fermented beverages and other biotechnological applications (Penãs et al., 2006).

Microorganisms of interest include lactic acid bacteria (LAB) such as Lactiplantibacillus plantarum, known for metabolizing sugars, organic acids, proteins, and fats into key metabolites including lactic acid, alcohol, and acetic acid, which can enhance flavor, texture, and viscosity in fermented products (De Vuyst & Degeest, 1999; Ruas-Madiedo et al., 2002; Ferreira et al., 2022). Yeasts, such as Kluyveromyces marxianus, also exhibit significant biotechnological potential, capable of producing lactic acid, enzymes, flavor compounds, proteins, and alcohol, thereby representing versatile agents for sustainable fermentation processes (Tabanelli et al., 2016). The use of co-cultures, combining two or more microorganisms, allows synergistic interactions that optimize metabolic activity, product yield, and quality. For example, Herkenhoff et al. (2024) used co-cultures and probiotic strains in complex and acidic matrices and found that yeast–bacteria interactions significantly influenced metabolite production and the development of sensory-related compounds during the first stages of fermentation, as revealed by HS-SPME/GC–MS analysis. Furthermore, this approach is widely applied in biotechnology and the production of fermented foods and beverages (García-Ochoa et al., 2009; Mills & Fleet, 2020).

Plant-based fermented beverages have gained increasing attention as functional foods due to their potential health benefits and ability to serve as carriers of probiotic microorganisms. Studies have demonstrated that soy-based substrates are particularly suitable for fermentation processes, supporting microbial growth while enhancing nutritional and sensory properties. Agarbati et al. (2023) evaluated selected yeast consortia in soy milk and reported efficient fermentation performance, probiotic viability, and desirable sensory characteristics. Similarly, Ruiz de la Bastida et al. (2023) reported that soy-based beverages can act as effective carriers of probiotic Lactobacillus strains, improving microbial viability, increasing isoflavone bioavailability, enhancing antioxidant activity, and maintaining good consumer acceptance. Furthermore, Rana et al. (2024) developed fermented soy milk beverages co-cultured with probiotic strains of Lactiplantibacillus plantarum and Lactobacillus acidophilus, reporting improved sensory attributes, reduced beany flavor, and viable probiotic counts above 10⁶ CFU/mL under simulated gastrointestinal conditions. Together, these studies highlight the growing potential of fermented soy-based beverages as functional products with desirable technological, nutritional, and sensory properties.

Sensory analysis plays a crucial role in product development by evaluating taste, aroma, texture, and appearance, guiding the design of products with higher consumer acceptance (Cerqueira et al., 2022). In Brazil, the preference for sweet flavors is dominant, with 82.1% of individuals favoring sweet foods, while choices for savory foods are influenced by salt sensitivity and body weight (Santos et al., 2019). External factors, such as sleep patterns, can also influence sensory perception and flavor preferences, underscoring the complexity of consumer food choices (Camargo, 2020). Similar findings have been reported for probiotic fermented beverages supplemented with fruit juices and by-products, with the incorporation of these ingredients contributing to the development of distinct aroma and flavor profiles while maintaining satisfactory sensory acceptance (Praia et al., 2022). Apple juice has been used in food products to replace commercial sugar, enhancing micronutrients, polyphenols, and fiber content (Esperança et al., 2025). Additionally, according to Hutu & Amariei (2024), commercial sugar can be replaced with apple sauce in pastry products without compromising consumer acceptance, thus showing the potential of apple products as a sugar substitute; however, it has not been explored in fermented beverages.

This study aimed to assess the sensory attributes, sensory characteristics, or sensory descriptors of a plant-based fermented beverage from tofu whey using a co-culture of potentially probiotic microorganisms, Lactiplantibacillus plantarum and Kluyveromyces marxianus. We also aimed to evaluate the physicochemical properties and microbial viability to assess its potential as a functional and probiotic product.

2 Materials and methods

2.1 Tofu whey

Tofu whey was provided by a local company located in the city of Viamão, state of Rio Grande do Sul, Brazil. The whey resulting from the tofu production process, considered a liquid by-product, was collected in a volume of 10 L. The pH of the whey was not adjusted using a pH meter, as its natural value was already approximately 6.0, allowing the use of tofu whey without further adjustments. It was stored under refrigeration (4 °C) to preserve its quality until the time of fermentation. Before fermentation, tofu whey was sterilized in an autoclave (Phoenix Vertical Autoclave) at 121 °C under 1 atm of pressure for 15 min.

2.2 Microorganisms

This study used microorganisms in co-culture: Lactiplantibacillus plantarum BL011 and Kluyveromyces marxianus B0399. The lactic acid bacterium L. plantarum BL011 was obtained from the Microbiological Culture Collection of BiotecLab (UFRGS, Porto Alegre, Brazil), and the yeast K. marxianus was kindly provided by the company Turval (Udine, Italy). Microorganisms were stored in a glycerol solution at -20 °C. For immediate use, L. plantarum was cultivated in Man, Rogosa and Sharpe (MRS) medium, whereas K. marxianus was cultivated separately in Yeast Malt Agar/Yeast Malt Extract (YM) medium, according to Fabricio et al. (2025).

2.3 Co-culture fermentation

The starter culture used in this work was developed in a previous study (Fabricio et al., 2025). In co-culture experiments, equal concentrations of L. plantarum and K. marxianus (1 × 10⁶ CFU/mL) were inoculated and cultivated using a broth-to-flask ratio of 1:5 in an Erlenmeyer flask and incubated in a shaker incubator (Shaker Incubator SL – 222, Solab) at 37 °C with agitation at 180 rpm. Optical density was measured at 600 nm using a spectrophotometer (UV-6100 Double Beam Spectrophotometer, Pró-Análise). Subsequently, the L. plantarum and K. marxianus cultures were combined and centrifuged (Refrigerated Centrifuge SL – 703, Solab) at 3500 rpm for 15 min, discarding the supernatant. The cells were washed with 0.9% sterile saline solution, and the total microbial pellet was resuspended in tofu whey for fermentation. Fermentations were performed in 1 L Erlenmeyer flasks containing 500 mL of tofu whey inoculated with the co-culture and incubated statically at 37 °C for 48 h. Samples were collected for microbiological and physicochemical analysis at 0 h and 48 h.

2.4 Microbiological analysis

The viability of yeast and bacteria (CFU) was determined by plate count and the results were expressed in CFU/mL. For bacterial count, tofu whey was enumerated on MRS agar plates supplemented with fluconazole (0.128 g/L) to inhibit yeast growth. Yeast counts were performed on YM agar plates supplemented with the antibiotic chloramphenicol (0.034 g/L) to inhibit bacterial growth. The CFU was quantified following the SP-SDS (Single Plate-Serial Dilution Spotting) method (Thomas et al., 2015), with the plate divided into six quadrants (10¹ to 10⁶); in addition, a 20 μL aliquot of the corresponding dilution was plated in each quadrant. The plates were incubated at 30 °C for 48 h.

2.5 Physicochemical analysis

Analysis was performed at 0 h and 48 h. The pH was measured using a pH meter (DM-22, Digimed). Collected samples were centrifuged and the supernatant was filtered using a 0.22 μm membrane pore size for further analysis of glucose, acetic acid, and alcohol content by HPLC.

The HPLC system was equipped with a Refractive Index Detector (RID) and a Rezex ROA-Organic Acid H+ (8%) column from Phenomenex. The mobile phase consisted of ultrapure water with sulfuric acid (H₂SO₄) 0.005 N, flowing at a rate of 0.6 mL/min. The column was maintained at 65 °C, and the detector at 40 °C to optimize analysis conditions.

2.6 Sensory analysis

Five samples were prepared for the sensory analysis, consisting of three fermented tofu whey samples and two non-fermented tofu whey samples:

• Sample NFA: non-fermented tofu whey with 8% apple juice;

• Sample NFS: non-fermented tofu whey with 10% demerara sugar;

• Sample F: fermented tofu whey;

• Sample FS: fermented tofu whey with 10% demerara sugar;

  • Sample FA: fermented tofu whey with 8% apple juice.

Different juice concentrations were previously tested by the research group before determining the concentration of 8% (data not shown). For the preparation of fermented samples, apple juice or demerara sugar was added after a 48-hour fermentation period. The concentrated apple juice was kindly provided by Golden Sucos (Farroupilha – RS), with 70 °Brix, while the demerara sugar (União, São Paulo, Brazil) was purchased from a local supplier. For preparation, the fermented tofu whey was supplemented with 8% concentrated apple juice or 10% demerara sugar (20 g in 200 mL of tofu whey).

For the consumer test, 98 individuals attended a central location test at the UERGS campus in Porto Alegre, in groups of up to 10 people. The following recruitment criteria were applied: i) aged 18 years or older; ii) no food allergies or intolerances; iii) not pregnant or breastfeeding; iv) interest in participating in a fermented beverage sensory test. Prior to the beginning of the samples evaluation, participants were briefed on the study procedures and signed the consent to participate in line with ethics procedures. At individual tables, consumers evaluated samples of 20 mL at 7 °C, served into plastic cups, coded with three-digit numbers. The project was approved by the Research Ethics Committee (CEP), protocol CAAE 73088423.0.0000.5347.

Samples were presented to consumers monadically and randomly following a Williams Latin Square design. After tasting each sample, they indicated their hedonic acceptance using a structured nine-point scale anchored with the conditions from “I really disliked” to “I really liked”. Then, they were presented with a list of 17 attributes and asked to check all that applied (CATA) to each beverage. All attributes were selected from studies previously reported in the literature (Treviso et al., 2024) and clustered by appearance (yellow color, white color, or uniform color), aroma (acid aroma, sour aroma, sweet aroma, or cooked aroma), flavour (acid, salty, sweet, sour, or bitter), texture (smooth, sandy, or sticks to the roof of the mouth), and residual flavour (bitter and salty). CATA terms were presented randomly among the volunteers, but in the same order for each sample.

In the end, consumers were asked about their sociodemographic profile (gender, age and frequency of tofu consumption).

2.7 Statistical analysis

2.7.1 Physicochemical data

To evaluate the statistical differences among tofu whey, means of triplicates from two independent experiments (repetition of the analysis was performed three times from two different samples) were submitted to one-way ANOVA followed by Tukey’s Honestly Significant Difference (HSD) test.

2.7.2 Sensory data

To evaluate the statistical differences in volunteers’ acceptance rates, means were evaluated by two-way ANOVA followed by Tukey’s test. Samples were considered fixed effects and consumers, random effects. Data normality and homogeneity of the variances were evaluated by Shapiro-Wilk and Hartley’s maximum F tests, respectively.

Frequency of mention for each sensory term was determined by counting the number of consumers that check to describe each sample, and the non-parametric Cochran’s Q followed by the post hoc pairwise Sheskin’s Critical Difference, was applied to detect differences in consumers’ perception of the evaluated samples. The perceived sensory maps of the samples were obtained by correspondence analysis (CA) based on chi-squared distances and the test of independence between rows and columns used on the frequency table containing the number of consumers who checked each term from the CATA tasks to describe each beverage. Penalty-lift analysis was performed for each CATA term to determine the mean impact of the term on liking (Meyners et al., 2013). A threshold of 10% citation was used.

Cochran’s Q test, penalty analysis, and CA were performed on XLSAT software (Addinsoft, New York, USA, version 2023.2.0) (Lumivero, 2023), and ANOVA analyses were carried out on R software version 4.0.5 (2021-03- 31) (R Development Core Team, 2021).

3 Results and discussion

3.1 Co-culture fermentation and pH

Cell growth was quantified by colony forming units per milliliter (CFU/mL) at 0 h and 48 h for both L. plantarum and K. marxianus. Initially, cell concentrations aligned with the inoculum level of approximately 6 log CFU/mL. Following 48 h of fermentation, viable cell counts increased significantly, reaching approximately 9 log CFU/mL for L. plantarum and 7 log CFU/mL for K. marxianus. These results demonstrate that both microorganisms remained viable and proliferated effectively during fermentation.

The observed increase indicates that the microorganisms achieved cell densities considered physiologically relevant for probiotics. Literature indicates that concentrations above 10⁶–10⁷ CFU/mL are generally required to confer health benefits by modulating gut microbiota and enhancing host defenses (Shah, 2000; Kandylis et al., 2016). Thus, the levels attained suggest the co-culture system’s potential to meet criteria for probiotic products.

Furthermore, co-culturing L. plantarum and K. marxianus can foster synergistic interactions, where yeast metabolism mitigates acid stress by consuming lactic acid, thereby stabilizing pH and supporting bacterial growth, while bacterially derived metabolites sustain yeast viability. This mutualism may enhance metabolite production, improve sensory attributes, sensory characteristics, or sensory descriptors, and promote functional stability in fermented foods.

These findings are consistent with studies showing that co-fermentation of soybean substrates with Lactiplantibacillus plantarum and complementary bacterial cultures can significantly improve flavor profiles by reducing undesirable compounds and enhancing pleasant sensory attributes, thereby increasing the overall acceptability of fermented soybean foods (Luo et al., 2023). Along with cell growth, pH changes were observed. A marked reduction in pH was observed during fermentation, decreasing from 5.92 to 3.95 after 48 h. This reduction in pH is characteristic of fermentation processes, in which microorganisms produce organic acids, such as lactic acid, thereby increasing the acidity of the medium. In the fermented samples, the pH decreased from 5.92 to 3.95 after 48 h of fermentation, confirming the metabolic activity of the co-culture. The non-fermented controls also exhibited acidic pH values. Sample NFA showed a pH of 3.80, which can be attributed to the addition of apple juice, a naturally acidic ingredient that also contains fermentable sugars such as fructose and glucose (Di Cagno et al., 2013). Similarly, sample NFS presented a pH of 3.88, likely reflecting the intrinsic characteristics of the tofu whey formulation rather than microbial fermentation.

From a microbiological standpoint, pH values below 4.0 are effective in inhibiting pathogenic and spoilage microorganisms, representing an advantage in terms of food safety (Liu, 2011). However, very low pH values may compromise the viability of lactic acid bacteria during storage, thus requiring their stability under highly acidic conditions (Gomes, 2015).

3.2 Sugars, acids, and alcohol

The concentration of acetic acid was measured at two fermentation time points, 0 h and 48 h. At the beginning, the level was 0.013 g/L, reaching 0.015 g/L after 48 h. Although there is a numerical difference, this variation is minimal and does not represent a real increase, indicating that acetic acid levels essentially remained constant throughout the fermentation process. This stability is expected since the microorganism used is primarily a lactic acid–producing bacterium, meaning its metabolic activity is directed toward lactic acid production rather than acetic acid synthesis. The consistency among samples reinforces that the fermentation system operated under controlled conditions with low variability. Low levels of acetic acid are typical for L. plantarum metabolism, which mainly produces lactic acid but can generate small amounts of acetic acid depending on environmental factors and substrate composition (Kandler & Weiss, 1986). Similar metabolic behaviour has been reported in acidic fermented beverages produced with probiotic cultures, with microorganisms exhibiting high adaptability and functional performance despite the challenging fermentation matrix. Using omics-based approaches, Herkenhoff et al. (2024), demonstrated that probiotic and starter cultures can successfully modulate metabolite production and fermentation pathways in sour beer, highlighting their metabolic robustness and supporting the selection of probiotic strains and fermentation strategies aimed at producing functional beverages with desirable sensory and technological characteristics.

Previous studies have demonstrated that Lactiplantibacillus plantarum and Kluyveromyces marxianus are capable of partially or fully metabolizing sucrose in substrates containing this disaccharide, as both microorganisms possess specific metabolic pathways enabling its utilization (Yin et al., 2018; Lertwattanasakul et al., 2011). Acetic acid was detected in all samples, ranging from 0.011 g/L in NFA to 0.017 g/L in F and FS, reflecting active fermentation processes. The NFA sample (tofu whey without co-culture plus 8% apple juice) exhibited low residual glucose (0.011 g/L), whereas the FA sample (tofu whey with co-culture and 8% apple juice) presented a higher glucose concentration (0.067 g/L), suggesting that the presence of the co-culture influenced the efficiency of sugar metabolism.

Samples NFS and FS, supplemented with 10% demerara sugar, as well as sample F (tofu whey with co-culture) showed no residual glucose. No ethanol was detected in any of the samples, which can be explained by the metabolic interplay between L. plantarum and K. marxianus. Competition between these microorganisms favored lactic acid production over alcoholic fermentation; in addition, the acidic pH generated by L. plantarum (below pH 4.0) inhibits K. marxianus fermentative activity and ethanol production. Similarly, Praia et al. (2022) reported that controlling alcohol production is an important factor for probiotic survival in fermented beverages, reinforcing the feasibility of developing functional products with high microbial viability and potential health-promoting properties.

In summary, metabolism in the co-culture fermentation of tofu whey was characterized by slight acetic acid production, near-complete sugar consumption, and suppressed ethanol formation. These outcomes align with findings that L. plantarum and K. marxianus are suitable for fermenting tofu whey, effectively converting sugars into organic acids while limiting alcohol formation, thus producing stable, probiotic-rich plant-based beverages (Ferreira et al., 2022).

3.3 Sensory analysis

3.3.1 Consumer profile

Consumer profiles showed that individuals were predominantly female (63.6%), with a mean age of 32.1 ± 12.1 years (range: 18–89 years). Tofu consumption was reported to be very low among the participants, with 94.9% indicating that they consumed tofu less than once a month or rarely. This profile suggests that most evaluators had limited prior exposure to tofu products, which may have influenced their sensory perceptions and acceptance. This observation is consistent with previous studies reporting that plant-based fermented products such as tofu remain relatively unfamiliar to many consumers, thereby affecting sensory evaluation outcomes (Yamahata et al., 2020). Manzanillo & Isidro (2025) also reported that tofu consumption is generally low in many populations, mainly in Western countries, and sensory evaluations frequently reveal that consumer preferences are shaped by factors including taste, texture, aroma, and appearance of tofu-based products.

3.3.2 Acceptance

Acceptance results (Figure 1) showed significant differences among samples (p < 0.0001). Samples NFS and FS achieved the highest acceptance scores (5.9 ± 0.3 and 5.6 ± 0.3, respectively), with no significant difference between them (p > 0.05). In contrast, NFA, FA, and F received significantly lower acceptance scores than NFS and FS (all below 5.0), indicating low consumer acceptance. Among these lower-scoring samples, NFA and FA did not differ significantly from each other (p > 0.05), meanwhile F sample presented the lowest liking scores (p < 0.05). These results indicate that sugar plays a critical role in consumer acceptance, as the highest acceptance was observed for samples with higher sugar content, regardless of the presence of the co-culture, whereas the lowest acceptance was observed for samples without added sugar. Dartora et al. (2023) and Treviso et al. (2024) reported that an adequate level of sweetness is essential for consumer acceptance of fermented beverages such as kombucha, corroborating the findings of this study. Although the apple juice-based beverages showed lower acceptance than those sweetened with commercial sugar, they exhibited higher acceptance than the formulation without any sweetener (F), in agreement with previous studies (Hutu & Amariei, 2024; Esperança et al., 2025). Further studies should investigate combinations of apple juice and commercial sugar to reduce the amount of added sugar in the beverage while maintaining consumer acceptance and enhancing its nutritional value.

Figure 1
Means of liking scores for each sample. ^a,b,c Different letters indicate significant differences among samples based on Tukey’s HDS test. NFA: non-fermented tofu whey with 8% apple juice; NFS: non-fermented tofu whey with 10% demerara sugar; F: fermented tofu whey; FS: fermented tofu whey with 10% demerara sugar; FA: fermented tofu whey with 8% apple juice.

3.3.3 CATA and correspondence analysis

Table 1 shows the differences among samples for perceived attributes, whereas Figure 2 illustrates the perceived sensory map. Results from CA (Figure 2) show that the first dimension (F1) explains 89.10% of the data variance, while the second dimension (F2), explains 5.15%. Thus, CA separated samples containing sugar (FS and NFS) in the positive side of F1, primarily based on sweet taste and aroma, and smooth texture. Samples without sugar (F, NFA and FA) were placed on the negative side of F1 and were primarily based on white color. Sample F was characterized by a higher intensity of this descriptor, as well as bitter taste and acid taste (which were stronger in samples FA and NFA).

Table 1
Contingency table of attributes for non-fermented tofu whey with 8% apple juice (NFA), non-fermented tofu whey with 10% demerara sugar (NFS), fermented tofu whey (F), fermented tofu whey with 10% demerara sugar (FS), and fermented tofu whey with 8% apple juice (FA) evaluated by the CATA method.

Attribute	Dimension	FS	NFA	F	NFS	FA	p-value
Appearance	Yellow	81ab	86a	75b	82ab	87a	0.046*
	White	6ab	5ab	14a	3b	6ab	0.008*
	Uniform	25	20	20	28	17	0.061
Aroma	Acid	15bc	27ab	31a	11c	29ab	0.001*
	Sour	31bc	34b	50a	17c	41ab	<0.001*
	Sweet	45ab	23bc	10c	5c	25bc	<0.001*
	Cooked	19b	32a	22ab	31a	22ab	0.0371*
Flavor	Acid	17c	30b	37b	17c	46ª	<0.001*
	Salty	11b	21ª	18ab	6c	13b	0.010*
	Sweet	68ª	14b	3c	74ª	13b	0.000*
	Bitter	14bc	31ª	25ab	10c	34ª	<0.001*
	Sour	7c	45ab	47a	17c	36ab	<0.001*
Texture	Smooth	74ab	57b	54b	80a	52b	<0.001*
	Sandy	12b	17ab	22ab	10b	27a	0.0007*
	Sticks to the roof of the mouth	8bc	17a	17a	5c	13ab	0.004*
Residual Flavor	Bitter	49b	63ª	59ab	47b	63ª	0.015*
	Salty	36	32	37	40	32	0.581

• ^a,b,c
  Different letters indicate statistical differences among samples for each attribute by Sheskin’s test. *Indicates statistical significance at the 5% significance level (p < 0.05).

Figure 2
Perceived sensory map based on correspondence analysis.

Uniform color and salty residual taste did not differ among samples (Table 1). Co-fermentation of soybean-based substrates with L. plantarum and complementary bacterial cultures significantly reduces undesirable off-flavors while enhancing overall sensory quality (Luo et al., 2023); however, results from frequency of citation showed no pattern.

The sensory perception of the beverages was closely related to their physicochemical characteristics. Sample F, which exhibited the lowest pH value after fermentation, showed the high frequencies of sour and acid aroma, acid taste, and sour flavor descriptors, indicating that acidification strongly influenced sensory perception. In contrast, FS and NFS, characterized by higher sugar contents, were more frequently associated with sweet taste and achieved the highest acceptance scores. These findings suggest that sweetness contributed positively to consumer liking, whereas excessive acidity negatively affected acceptance. Although acetic acid was detected in all samples, its concentration varied only slightly among treatments, indicating that the perception of sourness was likely influenced by the combined effect of organic acids produced during fermentation rather than acetic acid alone.

3.3.4 Penalty-lift analysis

Figure 3 presents the penalty-lift analysis, identifying the sensory attributes that significantly influenced liking. Results show that sweet taste and sweet aroma, uniform color, and smooth texture had a positive impact on acceptance (2.6, 1.7, 0.8 and 0.8, respectively). Yellow color (-0.8), acid, salty, bitter tastes (-0.5, -0.6, -0.8, respectively), sour flavor (-1.3), bitter residual taste (-0.7), sandy, and sticks to the roof of the mouth (-07, -08, respectively) were drivers of disliking in fermented tofu beverages. Dartora et al. (2023) and Treviso et al. (2024) also observed that sweet taste is critical for acceptance of fermented beverages, meanwhile acid taste and sour flavor and sour aroma impact negatively on liking scores, which are linked to sour and fermented flavors in the beverage sensory profile. These findings are consistent with previous studies showing that sweet taste is a driver of liking, whereas acid taste, sour flavor, sour aroma, and bitterness negatively influence consumer acceptance and the perceived quality of the product (Yamahata et al., 2020; Silva et al., 2023).

Figure 3
Drivers of liking (blue) and disliking (red) of tofu beverages based on penalty lift analysis.

F sample presented high frequency of citation of acid and sour aromas (Table 1), which were drivers of disliking (Table 1), and low citation of sweet taste (Table 1), which was a driver of liking, explaining the low acceptance shown in Figure 1. The rejection of samples may also be related to the acidification resulting from fermentation. Although low pH contributes positively to microbiological safety, excessive acidity can negatively affect flavor perception and mask desirable aromatic compounds. Similarly, residual bitterness and gritty mouthfeel are frequently reported as limiting factors in the acceptance of soybean-derived products. Therefore, formulations associated with these attributes received lower sensory scores, whereas samples characterized by sweetness, sweet aroma, and a softer texture showed greater consumer acceptance. These findings indicate that improving sweetness balance, reducing excessive acidity, and minimizing undesirable textural attributes may be important strategies for increasing the acceptance of fermented tofu whey beverages.

Additionally, Herkenhoff et al. (2024) demonstrated that fermented products containing probiotic cultures and cashew by-products positively influenced gut microbiota composition, supporting the development of probiotic or symbiotic foods with enhanced functional properties. Together, these findings reinforce the health benefits and functional potential of fermented soy beverages.

4 Conclusions

The co-culture fermentation of the plant-based beverage revealed Lactiplantibacillus plantarum’s robust growth and Kluyveromyces marxianus’s moderate performance, leading to a significant pH drop from 5.92 to 3.95, efficient sugar consumption, and enhanced acetic acid production, while suppressing ethanol formation. Sensory evaluation showed that the addition of demerara sugar, regardless of fermentation, maximized consumer acceptance, with sweetness, smooth texture, and uniform appearance driving preference. These findings demonstrate that a simple ingredient can improve the sensory acceptance of tofu whey, a nutrient-rich but underutilized agro-industrial byproduct, enabling the development of a functional and potentially probiotic plant-based beverage. Although the formulations containing sugar achieved the highest acceptance scores, overall consumer acceptance remained moderate, with maximum scores corresponding to “liked slightly” Therefore, further studies should focus on optimizing the formulation to improve sensory acceptability while preserving the beverage’s nutritional, functional, and sustainable characteristics. This work highlights the potential of sustainable plant-based fermentation to create innovative beverages with health-promoting and environmental benefits, providing the growing functional beverage market with a promising alternative.

Acknowledgements

The authors would like to thank the Fundação de Amparo à Pesquisa do Rio Grande do Sul (FAPERGS, RS, Brazil), grant 22/2551-0000397-4 (RITEs), for the financial support, and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES, Brazil).

Data Availability Statement

All data generated or analyzed in this study are included in this published article.

• Cite as:
  Jesus, M. G., Fabricio, M. F., Sant’Anna, V., Ayub, M. A. Z., & Hickert, L. R. (2026). Sensory evaluation of a novel sustainable probiotic beverage based on tofu whey. Brazilian Journal of Food Technology, 29, e2026015. https://doi.org/10.1590/1981-6723.0152026
• Funding:
  Fundação de Amparo à Pesquisa do Rio Grande do Sul (FAPERGS, RS, Brazil), grant 22/2551-0000397-4 (RITEs), and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES, Brazil).

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Edited by

• Associate Editor:
  Deborah dos Santos Garruti.

Publication Dates

• Publication in this collection
  25 Sept 2026
• Date of issue
  2026

History

• Received
  06 Feb 2026
• Accepted
  27 July 2026