Effect of Different Quantities of Miracle Fruit on Sour and Bitter Beverages

Amanda Cristina Andrade, Marina Böhme Martins, Jéssica Ferreira Rodrigues,

Sandra Bragança Coelho , Ana Carla Marques Pinheiro , Sabrina Carvalho Bastos

Abstract

Currently, there is a great demand for natural sweeteners. Miracle fruit has become a good alternative for a sweetener. However, it is important to optimize its quantities and check its application viability as a sweetener for bitter drinks. This study aimed to assess the effect of different miracle fruit quantities on the temporal sensory profile and overall taste of lemonade and green tea, to evaluate its effect in bitter beverages and to determine the optimal quantity to be used in these products. The lemonade results showed that the quantities of 150 mg, 300 mg and 600 mg of miracle fruit were effective in reducing acidity and promoting sweet perception. Furthermore, through the acceptance test, 300 mg miracle fruit showed to be an efficient substitute for sucrose and sucralose in lemonade. Miracle fruit was also effective for bitter beverages, but its effect was less pronounced. Regarding the sensory acceptance, similar results were obtained among the samples, indicating a potential use of miracle fruit as a natural sweetener for bitter beverages as well.

Introduction

A high sugar intake may cause harmful effects to health. These effects are associated with obesity and the onset of chronic diseases. Due to their high occurrence in recent years, they have become a public health problem, being necessary to find ways to reduce the population's sugar consumption. In this way, sweeteners have been used by industries to promote sweetness in foods, reducing their sugar and caloric value. To apply a sweetener effectively, it is necessary to be healthy, safe, to have a pleasant sensory characteristics, a similar sweetness of sucrose and an absence of off-flavors. Thus, it constitutes in a great challenge for the industries. However, studies have demonstrated that some artificial sweeteners could promote collateral effects to health, like the development of glucose intolerance (through changes in the intestinal macrobiotic), bladder cancer and cardiac insufficiency. Therefore, the use of natural substances has gained a great interest like miracle berries.

Knowing that many sweeteners are also applied in bitter beverages, such as teas, coffee, etc., it would be advisable to investigate its effect on these products. Mainly on teas that have several nutritional properties of interest, but also have a pronounced bitter taste (often not appreciated by consumers).

Different miracle fruit quantities on the temporal sensory profile and overall liking of lemonade and green tea, evaluating its effect on bitter beverages and determining the optimal quantity to be used as a sweetener in these products.

Material and methods

2.1. Experiment overview

The research was performed on two different products (lemonade and green tea) in different sessions, following the same logistics to evaluate the effect of different quantities of miracle fruit on sour (lemonade) and bitter (green tea) beverages. Sachets of green tea (Lipton– 2g), lemons, sucrose and miracle fruit were used to prepare the samples of this study. Six treatments of each product were established to evaluate the different miracle fruit quantities effect on the temporal sensory profile (TDS and TI) of lemonade and green tea . The miracle fruit quantities (600 mg) and based on pretests, aiming to find the lowest possible quantity of miracle fruit to promote positive effects as a sugar substitute in lemonade and green tea. During the analysis of the products sweetened with the miracle fruit, it was served 5 min before the ingestion of unsweetened lemonade/ green tea. The panelist was informed to place the miracle fruit on their tongue and roll it around very slowly until it completely dissolved.

2.2. Lemonade preparation

The lemonades were prepared according to in a proportion of 3:1 (water: lemon) (w/w). The lemons were squeezed using a juicer. Thereafter, water was added, and the lemonade was then stirred with a glass rod for 5 min.

2.3. Green tea preparation

The teas were prepared according to the product's specifications. Boiling filtered drinking water (100 ml) was used for each sachet. It was infused during 2 min and it was filled with ice water (100 ml). The

contents were stirred with a previously sterilized glass rod, 1 min for pure tea and 5 min when sucrose and sucralose were added.

2.4. Sensory analysis

Sensory analyses were performed at the sensory analysis laboratory of the Federal University of Lavras. The temporal tests (Time Intensity and Temporal Dominance of Sensations) were performed with the same panel in different sessions (one session for each product - lemonade/ green tea), but at the same conditions. Acceptance tests were performed by consumers of each product (lemonade and green tea) in two different sessions (one for each product).

2.4.1. Time-intensity (TI)

The time-intensity analysis was performed by eleven panelists (eight women and three men, between 20 and 35 years old) selected based on their good sensory abilities, following the procedures used by. Thereafter, they performed the basic taste identification tests and triangular test, being selected trough sequential Wald Analysis.

After the training, the panelists performed the Time-Intensity tests in triplicate, with four sections for each attribute (sourness and sweetness) for the four treatments. The samples were served in plastic

cups labeled with randomly selected 3-digit numbers and they were presented in a monadic way, using a balanced complete block design . In the case of L1/T1 (unsweetened lemonade/green tea), L5/T5 (lemonade/green tea prepared with sucrose) and L6/T6 (lemonade/green tea prepared with sucralose) the individuals were instructed to drink the product (30 ml) all at once and immediately start the evaluation for 50s. For L2/T2, L3/T3 and L4/T4 (ingestion of different miracle fruit quantities followed by the ingestion of lemonade/green tea), the panelists were instructed to place the miracle fruit on their tongue and roll it around very slowly until it's completely dissolved and then, drink the green tea at once and immediately start the evaluation. Then, they recorded the intensity of the attribute (sourness or sweetness) using a graphic interface in the form of a 10-point scale, with 0 meaning no perception and 10 an extreme perception of the attribute. After performing the lemonade tests, the same logistic was used to perform the green tea tests.

2.5. Temporal Dominance of sensations (TDS)

The same selected panelists that participated in the TI tests performed the TDS tests. Thus, eleven panelists performed the TDS tests of each treatment in triplicate, totaling 33 evaluations per session, being one session for each treatment. The attributes analyzed in the TDS sensory test were sweet, sour, bitter, unpleasant, and no taste as established. The samples were served in disposable white plastic cups identified with three-digit numbers in a monadic, balanced order and the evaluation was performed during 50 s. After performing the lemonade tests, the same logistic was used to perform the green tea tests.

2.6. Overall liking

As L4/T4 (600 mg miracle) and L3/T3 (300 mg miracle) had similar temporal profiles (see the results section) for both products (lemonade and green tea), and considering the cost of miracle fruit, only L2/T2 (150 mg miracle) and L3/T3 (300 mg miracle fruit) were submitted to acceptance tests. L5/T5 and L6/T6 (lemonade/green tea sweetened with sucrose and sucralose at optimal quantities) were also evaluated. The sucralose sample was added in order to compare the acceptance of the miracle fruit samples with the most commonly used and recognized sugar substitute. Thereafter, the tests were performed in different steps divided by product category: one test was performed with eighty lemonade consumers (37 male and 43 female, aged from 20 to 40 years old), with a minimum consumption frequency of lemonade of once a week. Another test was performed with eighty green tea consumers (19 male and 61 female, aged from 20 to 40 years old), with a minimum consumption frequency of green tea of once a week. They evaluated each sample in a separate session in relation to its overall liking, using a nine-point hedonic scale, ranging from

2.7. Statistical analysis

The TI results were analyzed by ANOVA (sources of variation: treatments, panelists and treatments*panelist interactions) and Tukey's test using the Sensomaker software. The TI curves were plotted using Microsoft Excel 2012. In the graphs, the horizontal axis denoted the time, while the vertical axis displayed the intensity values. The TDS results were assessed through the TDS curves using the Sensomaker software. Three TDS parameters (DRmax– maximum dominance rate, TDRmax– time at which maximum dominance occurs, and Plateau – duration of the attribute, i.e., the time range over which the dominance rate is equal to or higher than 90% of the maximum dominance rate) were also computed trough the curves . Overall liking results were assessed by ANOVA and Internal Preference Mapping  followed by Tukey's test using the Sensomaker software.

Results

3.1. Lemonade

3.1.1. Time-intensity (TI)

The time-intensity curves for sourness and sweetness intensities over the time (50 s) for the unsweetened lemonade (L1) and the ingestion of different quantities of miracle fruit (150 mg – L2, 300 mg – L3 and 600 mg – L4) followed by the ingestion of lemonade and for lemonades sweetened with sucrose (L5) and sucralose (L6) different quantities of miracle fruit (L2, L3 and L4) behaved in a similar way, i.e., they had a similar curve profile, showing to be effective on lemonade, reducing the perception of acidity.

According to the TI parameters, the pure lemonade (L1) obtained significantly higher acidity (p ≤ 0.005), while the lemonade samples with quantities of 300 mg and 600 mg of miracle fruit (L3 and L4) did not differ significantly from each other. However, they were significantly different (p ≤ 0.005) compared to the sample with 150 mg of miracle fruit (L2), sucrose (L5) and sucralose (L6) in relation to the TI parameters (Imax, and area under the curve). Moreover, these quantities (300 mg and 600 mg of miracle fruit) were more effective in reducing the lemonade's acidity. In the sweetness results evaluation, ANOVA showed that there was no significant difference in the maximum intensity of the lemonade samples prepared with 300 mg (L3) (Imax = 6.94) and 600 mg (L4) (Imax = 7.3) miracle fruit, with them being 6.12 and 5.75 times, respectively, sweeter than pure lemonade (L1). Lemonade followed by the previous ingestions of 150 mg (L2) (Imax 5.77) of miracle fruit varied significantly (p < 0.005) in relation to L3 and L4, but did not differ from the lemonade samples with sucrose (L5), being 4.58 times sweeter than pure lemonade. In addition, the lemonade samples with sucralose (L6) were significantly less sweet. Regarding the area under the curve, it was found that L4 obtained a higher value (278.28), being significantly different (p ≤ 0.005) in relation to the other samples, followed by L2 and L3 (217.94 and 236.63, respectively). Therefore, it seems that 300 mg and 600 mg of miracle fruit behaved in a similar way regarding the time-intensity profile and were more

effective than sucrose and sucralose in the lemonade taste modifying.

3.1.2. Temporal Dominance of sensations (TDS)

The TDS profiles obtained for unsweetened lemonade (L1), different quantities of miracle fruit (150 mg – L2, 300 mg – L3 and 600 mg – L4) and lemonade sweetened with sucrose (L5) and sucralose (L6). Each curve represents the variation of the dominance profile for each attribute assessed over time. The bottom line is the chance line – it means that the values were randomly marked; the top line corresponds to the significance line – the results above this line indicate that they were significantly perceived as dominant. The lemonade had no taste after 35 s. In the lemonade samples with previous ingestion of different quantities of miracle fruit, the sour sensation was not significantly perceived. Considering the curves below the chance level, L2 had higher dominance rate (0.33) and duration (12.5 s) of sour taste compared to the samples with L3 and L4. They obtained a maximum dominance rate of 0.21 and 0.26 and duration of 3.50 s and 6.30 s respectively, having similar sensorial profiles in the reduction of sourness perceptionery lemonade sample with previous ingestion of miracle fruit, while sour taste was detected as dominant until the 8s mark in the lemonade samples with sucralose and it predominated in the sample with sucrose. The sweetness taste was detected over approximately 40s.

3.1.3. Overall liking

The acceptance test was performed only with the 150 mg and 300 mg quantities in order to verify the consumers’ preference. The first principal component explained 38.28% of the data variability and the second component explained 26.65% of it, totaling 64.93%. Consumers showed a greater acceptance for the samples with sucralose, sucrose and 300 mg of miracle fruit (L3). On the other hand, the lemonade followed by the previous ingestion of 150 mg of miracle fruit (L2) and the pure lemonade (L1) were the less accepted samples.

3.2. Green tea

3.2.1. Time-intensity (TI)

All the others samples with miracle fruit ingestion were able to reduce the green tea bitterness. 150 mg, 300 mg, 600 mg of miracle fruit (T2, T3 and T4) and sucrose (T5) did not differ in relation to the maximum bitter intensity but the sucralose was the most effective taste modifier. Regarding the

area under the curve, the miracle fruit samples had a similar behavior, being the sucralose (T6) and sucrose (T5) more effective in reducing the bitterness. T5 provided the highest (p ≤ 0.05) sweet intensity (4.76), followed by the other samples regarding the area under the curve, T3, T4 and T6 did not differ significantly from each other, but differed from T2; which was the less effective in promoting sweetness in green tea. Moreover, this sample took a longer time to reach the maximum sweetness intensity compared to T2 and T3, achieving a higher sweetness intensity after 20 s, while those with 300 mg and 600 mg of miracle fruit reached maximum intensity 10 s after the test.

3.2.2. Temporal Dominance of sensations (TDS)

It was noted that as the miracle fruit quantity was increased, the bitterness maximum dominance rate (0.72; 0.67; 0.5 respectively and the duration decreased. It was also noted that T4 had a shorter maximum dominance rate for bitter sensation in relation to sugar and sucralose (0.61). However, there was the dominance of sweet taste only in the sugar sample.

3.2.3. Overall liking

Despite the miracle fruit samples acceptance having not been similar to sugars, their sensory acceptance was similar to sucralose's 9, one of the most used sweeteners.

Discussion

4.1. Lemonade

It was observed that all tested quantities of miracle fruit (150 mg, 300 mg and 600 mg) were efficient in reducing the perception of acidity and providing sweetness to lemonade (TI results). The sour into sweet taste conversion occurred due to the miraculin present in the miracle fruit. Miraculin binds itself to the sweet receptors of taste cells in the presence of protons (H+), changing its structural conformation, causing the perception of sweet taste. Consequently a lower quantity of miraculin) had a smaller effect in this modulation, reaching lower sweetness intensities.

However, L3 and L4 obtained similar behavior in relation to these attributes. In the sucrose substitution, the sweet taste intensity and persistence, as well as the presence or absence of residual flavors, are important factors that influence the product acceptance. The absence of unpleasant flavors was observed, even at lower quantities, in the TDS profiles of the different quantities (150 mg, 300 mg and 600 mg) of miracle fruit evaluated. In addition, the sweet sensation was the only sensation detected as significantly dominant. This is an indicative of the potential use of miracle fruit as a sweetener for sour beverages even at lower quantities. However, it seems that lower quantities promote a shorter duration of sweetness perception. Ingestion of 300 mg of miracle fruit before the lemonade ingestion was able to reduce the acidity and promote the sweet taste perception. In addition, this quantity provided a sweetness profile similar to sucralose's – one of the sweeteners most used by the food industry, due to its sweetness-promoting properties, besides the bitter taste absence. However, the beverage's acceptance was not evaluated. Therefore, in this study, the lemonades acceptance with different miracle fruit quantities was assessed in relation to the lemonade prepared with sucrose and sucralose. Although 150 mg of miracle fruit was effective in promoting the sweet taste. The lemonade sample with previous intake of 300 mg of miracle fruit obtained a similar acceptance to the lemonade samples prepared with sucrose and sucralose. This probably occurred due to the greater intensity and duration of sweetness noted in the TI results. Currently, miracle fruit is marketed in capsules with 600 mg. 400 mg of miracle fruit improves sweetness in a similar way to sucrose in an acid dessert. In this study, a smaller quantity (300 mg) was adequate to be used as a natural sweetener for lemonade. However, the mechanisms in which the miracle fruit is involved when inducing sweetness differ from the other common sweeteners. Miracle fruit cannot be added directly to the food. It is necessary that miraculin binds itself to the receptor of sweet taste to promote the perception of sweetness and, therefore, it must be consumed before the food's ingestion. Another important point to extending the miracle fruit application to other sour beverages is considering their acid composition. Therefore, complementary research should be conducted in order to evaluate its effectiveness in other products.

4.2. Green tea

Miracle fruit effects on sourness had already been studied by several authors. In this work, it was noted that the miracle fruit have an effect on sour beverages, providing the sweet taste, but at a lower intensity than the sour taste effect. Miracle fruit was found to be twice as sweet when compared to sugar when eaten, miraculin binds to the taste buds receptors on the tongue and tricks sour,bitter and savory receptors to detect sweetness. The sweet receptors are then heightened. At neutral pH, miraculin binds and blocks the receptors. Nevertheless, at low pH levels, which are categorized as acidic and are caused in the mouth when eating sour or bitter foods, miraculin activates the sweet receptors. Thus, its effect is more pronounced as lowest as the pH, which justifies the results founded in this study for lemonade and green tea. They noted the bitterness rating of broccoli and saltiness rating of the Goldfish crackers did not change after the berry was consumed, while the others presented alterations. However, the perceived sweetness for each acidic food item (lemon, grapefruit, lime, sour candy and cider vinegar) significantly increased after the berry was eaten.The miracle fruit ingestion could be a good alternative to reduce the metallic taste, promoting a more pleasant taste in with cancer patients treated with chemotherapy.

Green tea is a widely consumed beverage in the world. Its flavor generally comprises in bitterness, astringent, umami and sweet, but it is primarily bitter and astringent due to the catechins presence. Tea consumption is usually accompanied by sugar or milk, or both. In addition, consumers usually add sweeteners to teas, due to the rejection of their bitter taste. Considering these points, the use of sugar or sweeteners during the tea consumption is very common. Thus, miracle fruit would be a natural alternative to be employed. Sucralose is an artificial sweetener and artificial sweeteners are associated with some detrimental health effects, such as glucose intolerance due to changes in the intestinal microbiota, tumor development, heart failure, among others. Thus, the use of natural sweeteners, such as the miracle fruit, is of great interest to promote sweet taste without providing caloric value. Despite the lower effects of miracle fruit on green tea's bitterness, its use in quantities of 300 and 600 mg provided an acceptance similar to sucralose, making it a natural sweetener alternative to be applied in diet green tea and other bitter beverages. However, it is important to highlight that more studies investigating miracle fruit effects on bitter taste modulation are necessary to better elucidate its application as a sweetener in bitter beverages.

Conclusion

Temporal results showed that 150 mg, 300 mg and 600 mg of miracle fruit were effective in reducing acidity and promoting sweet perception in lemonade. Furthermore, 300 mg and 600 mg obtained similar profiles. Through the acceptance test, 300 mg of miracle fruit was shown to be an efficient substitute for sucrose and sucralose in lemonade, as they had similar hedonic values between 6 –“I liked it lightly” and 7 –“I liked it moderately”. Therefore, 300 mg of miracle fruit is adequate to be used as a natural sweetener of lemonade. Miracle fruit was also effective in bitter beverages, but its effect was less pronounced. TI and TDS results showed that higher miracle fruit quantities (600 mg) are necessary to achieve similar profiles of sucralose's in green tea. Regarding the sensory acceptance, similar results were obtained among the samples, with scores between 5 – “I do not know” and 6 – “I liked it slightly”, indicating a potential use of miracle fruit as a natural sweetener for bitter beverages as well. It is important to highlight that it is an initial study and other researches investigating the optimization of the process of keeping the lyophilized fruit, the purity and contents of miraculin must be realized.

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