CA3110913A1

CA3110913A1 - Functional soy whey beverage having refreshing flavor and manufacturing method thereof

Info

Publication number: CA3110913A1
Application number: CA3110913A
Authority: CA
Inventors: Li Li; Yongtao FEI; Suijun LI; Ke Huang
Original assignee: South China University of Technology SCUT
Current assignee: South China University of Technology SCUT
Priority date: 2017-08-31
Filing date: 2018-08-22
Publication date: 2019-03-07
Anticipated expiration: 2038-08-22
Also published as: WO2019042197A1; CN107751389A; AU2018325047A1; CA3110913C; AU2018325047B2; CN107751389B

Abstract

A functional soy whey beverage having a refreshing flavor and a manufacturing method thereof are provided. The method comprises: selecting fresh fermented soybean yellow serofluid as a raw material for manufacturing the functional soy whey beverage, and adding sucrose thereto; sterilizing after the sucrose is completely dissolved; adding freshly squeezed pineapple juice at a volume ratio of 20:1-2 in a sterile environment; and after cooling adding an activated fermentation starter at a ratio of 2-6%. The cultures used are Streptococcus thermophilus and probiotic Lactobacillus amylolyticus or Lactobacillus rhamnosus, and are cultured at 35-45°C after being inoculated into the soybean yellow serofluid and are then refrigerated so that a fermented soy whey beverage is obtained.

Description

DESCRIPTION
FUNCTIONAL SOY WHEY BEVERAGE HAVING REFRESHING FLAVOR
ANT) MANUFACTURING METHOD THEREOF
FIELD OF THE INVENTION
The present invention relates to the technical field of the soy whey beverage production process, in particular to a soy whey beverage with light yellow color, refreshing taste, pleasant flavor and significant antioxidation activity, and a manufacturing method thereof.
BACKGROUND OF THE INVENTION
Whey is a by-product of using animal milk to make cheese. It contains a large amount of acid-soluble nutrients and is usually further processed into a variety of high-value products. Similarly, in the process of using soybean milk to make bean curd in China, yellow water will also be produced, commonly known as soybean yellow serofluid. The differences between whey and soy whey are mainly as follows: CD Different sources: Whey comes from the supernatant separated out of casein in animal milk when the casein coagulates due to the acid produced by Lactobacillus fermentation, while soy whey comes from the supernatant separated out of glycinin in plant soybean milk when the glycinin coagulates due to calcium or magnesium salts or acids. 0 Different chemical compositions:
Whey contains a lot of nutrients such as whey protein and lactose, while soy whey contains a lot of soy whey protein, prebiotic such as raffinose and stachyose, and phytochemicals such as isoflavone, with the soy whey from the brine bean curd production also containing a large amount of calcium and magnesium ions. 0 Different flavors: There is still a small amount of milk fat in whey, which thus has a slight milky flavor, while there is still the oxidized products of soybean oil in soy whey, which thus has an obvious beany smell. It can be seen that soy whey contains a variety of functional ingredients that are beneficial to health and can be further used like whey.
However, because the whey of brine bean curd contains a large amount of metal ions and has a beany smell and other defects, it is mainly discharged directly as wastewater or treated to reduce BOD and COD
before being discharged.
The main reason for the above problems is that soy whey has a large amount but a low concentration, and the need to remove calcium and magnesium ions. Therefore, the cost of industrial application (concentration, refinement, and transportation) of soy whey is very high, and it is urgent to solve this technical problem.

Date Recue/Date Received 2021-02-26 CONTENTS OF THE INVENTION
In order to solve the above-mentioned problems in the prior art, a purpose of the present invention is to provide a method for producing a Lactobacillus-fermented soy whey beverage with simple process, unique flavor and significant antioxidation effect.
For reducing the cost and simplifying the process, the present invention selects the filtrate produced from the preparation of acidified serofluid-treated bean curd as a fermentation raw material.
Acidified serofluid-treated bean curd is a kind of bean curd made with a coagulant of acidified serofluid, which becomes acidic due to natural fermentation. Due to the unique flavor and large amount of probiotic Lactobacillus, the acidified serofluid-treated bean curd is popular among more and more modern consumers, which makes the demand for it increasing. When the protein in soybean milk coagulates after meeting with acid and is then pressed, the soluble soy whey protein, raffinose, stachyose, isoflavone, saponin and the like will drain out with water under an acidic condition.
Therefore, this soy whey is chemically an acidic extract of soybean not containing excessive calcium and magnesium ions, and can be used as an excellent beverage raw material without ion exchange.
In order to promote the fermentation of Lactobacillus and strengthen the nutrition and flavor of the beverage, the present invention also adds a certain proportion of pineapple juice to the soy whey.
Pineapple fruit is rich in fructose, glucose, sucrose, vitamin A, vitamin B, vitamin C, and calcium, phosphorus, iron, and other ingredients. These ingredients are not only trace nutrients, but also promoters of Lactobacillus growth. Besides, the combination of these ingredients also has a pleasant taste and aroma, which is generally favored by consumers.
In order to obtain a high-performance fermentation starter, this research group screened and optimized the adaptive cultures that had been separated and stored in the early stage, and obtained 2 groups of soy whey fermentation starter. Among them, Streptococcus thermophilus (S. thermophilus) was isolated, domesticated and screened by this laboratory from the fermented soybean milk using a directed Vat Set starter culture XPL-1 (a sample from Chr Hansen Company), and Lactobacillus amylolyticus (L. amylolyticus) and Lactobacillus rhamnosus (L. rhamnosus) were isolated by this laboratory from naturally acidified serofluid.
The key of the present invention was the selection of fermentation raw materials and the optimization of the fermentation starter.
First, yellow serofluid of an acidified serofluid-treated bean curd was selected as the fermentation raw material, and paired with pineapple juice from South China. With pineapple juice having a unique sweet taste and smell, addition of a proper proportion of pineapple juice to the yellow serofluid of an acidified serofluid-treated bean curd can partially mask the beany smell of the yellow serofluid and

2 Date Recue/Date Received 2021-02-26 make the flavor fresh and fragrant. In addition, with pineapple juice rich in vitamin A, vitamin B2, vitamin C, and mineral nutrients, addition of pineapple juice to yellow serofluid can supplement the micronutrients in the yellow serofluid and promote the growth of Lactobacillus.
Second, a soy whey-fermentation starter combination was obtained through screening and optimization. Having strong a-galactosidase activity and a very high utilization rate of raffinose and stachyose, S. thermophilus has good fermentation performance in soy whey, and can produce a large amount of extracellular polysaccharides. Having strong 13-glucosidase activity and very strong ability to convert isoflavones into aglycones, L. rhamnosus can improve the bioavailability of isoflavones in fermented soy whey. L. amylolyticus has strong a-galactosidase and 13-glucosidase activities at the same time. S. thermophilus was combined with L. amylolyticus and L. rhamnosus, respectively, to form a fermentation starter to apply to the preparation of whey beverages. By utilizing the characteristics of complementary metabolism of different cultures, a soy whey beverage with an increased aglycone isoflavone content and significant antioxidation activity could be obtained.
Moreover, other metabolites of the above cultures, such as lactic acid, extracellular polysaccharide, and diacetyl, would also make the soy whey beverage have intestinal health effect, enhanced immunity and good flavor.
Through the above selection and application of the yellow serofluid of an acidified serofluid-treated bean curd and pineapple juice, as well as the screening and optimization of the fermentation starter, the technical problems such as poor flavor and bad fermentation of soy whey beverages have been overcome, which provides a new and effective solution to avoid environmental pollution and waste of resources and improve production efficiency.
In view of the above findings, the present invention achieves its object by the following technical solution:
A functional soy whey beverage having a refreshing flavor and a manufacturing method thereof are provided. The method comprises the following steps:
1) With kilogram and liter respectively as the unit of mass and volume, adding sucrose at a mass-to-volume ratio of 0 to 6% to a fresh yellow serofluid of an acidified serofluid-treated bean curd, fully dissolving, and filtering the fresh yellow serofluid of an acidified serofluid-treated bean curd containing sucrose;
2) treating the fresh yellow serofluid of an acidified serofluid-treated bean curd containing sucrose at a temperature of 110 C to 121 C for 5-20 min, and cooling same;

3) sterilizing freshly squeezed pineapple juice, and cooling same;

4) mixing the sterilized yellow serofluid of an acidified serofluid-treated bean curd obtained in step 2) with the sterilized pineapple juice obtained in step 3) in a sterile environment at a volume ratio Date Recue/Date Received 2021-02-26 of 20:1 to 25:2; and

5) inoculating an activated fermentation starter into the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd mixture obtained in step 4) at an inoculation amount of 2% to 6%
of the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd mixure, then culturing at 35 C to 45 C for 12-36 h, centrifuging to obtain a supernatant, and refrigerating the supernatant to obtain a fermented pineapple juice-soy whey beverage.
The fermentation starter is a mixture of S. thermophilus and one of L.
amylolyticus and L.
rhamnosus at a volume ratio of 1:1-4.
The culture collection number of the S. thermophilus is CGMCC NO. 11943;
the culture collection number of the L. amylolyticus is CGMCC NO. 9090; and the culture collection number of the L. rhamnosus is GDMCC 60224.
To further achieve the purpose of the present invention, it is preferred that the freshly squeezed pineapple juice is sterilized at 110 C to 115 C for 1-15 min.
Preferably, the volume ratio of the yellow serofluid of an acidified serofluid-treated bean curd to the pineapple juice is 20:1-4.
Preferably, the method for preparing the pineapple juice is as follows:
squeezing pineapple pulp and filtering same through four layers of gauze to collect a filtrate.
Preferably, the method for activating S. thermophilus is as follows:
inoculating 3% to 8% of the glycerol-preserved S. thermophilus into a sterilized Elliker medium to culture at 37 C to 45 C for 10-24 h, repeatedly activating for 2-3 times, and putting aside for later use.
Preferably, the method for activating L. amylolyticus and L. rhamnosus is as follows: inoculating 3% to 8% of the glycerol-preserved L. amylolyticus or L. rhamnosus into a sterilized MRS liquid medium to culture at 32 C to 47 C for 10-24 h, repeatedly activating for 2-3 times, and putting aside for later use.
Preferably, the refrigeration is carried out at 4 C.
A functional soy whey beverage having a refreshing flavor is manufactured by the above method.
The present invention has the following advantages compared to the prior art:
1. The lactic acid bacteria fermented pineapple juice-soy whey beverage produced by the present invention not only integrates the nutritional value of pineapple juice and yellow serofluid, but also imparts the unique aroma and refreshing taste to the soy whey beverage, which provides the possibility for the market development of the yellow serofluid beverage.
2. Compared with fresh soybean yellow serofluid, the lactic acid bacteria fermented pineapple juice-soy whey beverage produced by the present invention has significantly higher antioxidation Date Recue/Date Received 2021-02-26 activity indexes such as DPPH free radical scavenging ability, and generates a large amount of active ingredients such as lactic acid and extracellular polysaccharide.
3. The fermentation starter used in the present invention is a mixture of S.
thermophilus and one of L. amylolyticus and L. rhamnosus, having extremely high adaptability and a synergistic fermentation effect in the yellow serofluid. The raw materials used in the production are pure plant sources, and the resulting products have probiotic effects such as regulating intestinal flora and favoring cardiovascular health.
BRIEF DESCRIPTION OF THE DRAWINGS
Fig. 1 shows the existing mode of isoflavones in Examples 2,4 and 5.
DETAILED DESCRIPTION OF THE EMBODIMENTS
In order to better understand the present invention, it will be further described below in conjunction with drawings and examples; however, the embodiments of the present invention are not limited thereto.
In the following examples, the acidity of the fermented yellow serofluid was measured according to the method described in GB 5413.34-2010. The specific operation is as follows: accurately weighing 5.0 g of the sample, putting it into a 50 ml beaker, adding 20 mL of CO2-free distilled water, and mixing well; then adding 0.5 mL of phenolphthalein indicator, and titrating with a 0.1 N NaOH
reference solution until the solution was reddish and did not fade within 0.5 min, which was taken as the end point of titration; the acidity value was calculated according to the following formula:
X = (CxVx100)/(mx0.1) wherein:
X ¨ acidity value of the sample, expressed in Thorner degrees ( T);
C ¨ concentration of the sodium hydroxide reference solution (mol/L);
V ¨ volume of the sodium hydroxide reference solution consumed (m1); and m ¨ mass of the sample (g).
In the following examples, the measurement of DPPH = free radical for the antioxidation activity of the fermented yellow serofluid was made according to the method described in the literature "Use of a free radical method to evaluate antioxidation activity", and modified appropriately. The specific method was as follows: Taking 2 mL of each group of samples and 2 mL of a DPPH
solution (dissolved in absolute ethanol at 0.2 mmol/L) and mixing thoroughly; a reaction was carried out in the dark at room temperature for 30 min, avoiding light; the absorbance of the sample was measured at 517 nm; 2 ml of DPPH and 2 ml of absolute ethanol were used as a control, and tocopherol (Trolox) was used as a reference material; the DPPH free radical scavenging ability of the fermented Date Recue/Date Received 2021-02-26 pineapple-yellow serofluid mixture was expressed as a p.M Trolox/L sample, in which a reference curve, y = -0.012x + 1.174, r2 = 0.9996 (y ¨ absorbance value, x ¨ Trolox concentration in prnol), was determined with tocopherol (Trolox) as the reference material.
In the following examples, the measurement of the ferric reducing ability of plasma (FRAP) for the antioxidation activity of the fermented yellow serofluid was made according to the literature "The ferric reducing ability of plasma (FRAP) as a measure of `antioxidation power': the FRAP assay', and modified appropriately. The specific method was as follows: Weighing 0.0781 g of TPTZ, and dissolving it with 0.04 mol/L HC1 to make up to 25 mL; mixing the TPTZ
solution, FeCl3 solution (20 mmol/L) and acetate buffer solution (300 mmol/L, pH 3.6) at a volume ratio of 1:1:10 to prepare a FRAP solution; adding 100 !IL of various samples to 3 mL of the FRAP solution, allowing to stand at 37 C for 10 min, and then measuring the absorbance at 593 nm; Trolox was used as the reference material; the ferric reducing ability of the fermented yellow serofluid was expressed as an mM
Trolox/mL sample, and the reference curve obtained from the preparation was y = 0.001x + 0.511, r2 =
0.9993 (y ¨ absorbance value, x ¨ Trolox concentration in mmol).
In the following examples, the content of isoflavones in the fermented yellow serofluid was determined by high-performance liquid chromatography specifically as follows:
Pretreatment of samples: Accurately taking 2 mL of each group of samples and placing them in a 10 mL volumetric flask, and making up to 10 mL with a 80% methanol solution; ultrasonically extracting at 60 C for lh, and centrifugating at 10,000 g for 10 min; taking about 1 mL of the supernatant, and filtering it through a 0.45 um filter membrane (organic phase); injecting the filtrate into a sample bottle, and storing at -20 C for later use. It was used for the analysis of isoflavone.
Chromatographic conditions:
Phenomenex Analytical Guard Cal _____________________________________________ tiidge System (KJO-4282), 8-core (AJO-4286); chromatographic column: Phenomenex Luna 8 (2) 100A column (inner diameter 4.6 mm, length 250 mm, filler particle size 5 pm); mobile phase: A represented a 0.1% acetic acid solution, and B
represented a 0.1%
acetonitrile acetate solution; flow rate: 1 mL/min; column temperature: 36 C;
Thermo-Ultimate 3000 liquid chromatograph; detector: VWD 3100 UV detector; detection wavelength:
254 nm; injection volume: 20 0_4 separation time: 26 min. Gradient elution conditions were in accordance with GB/T
26625-2011. Drawing of a standard curve: Weighing 1 mg of reference materials genistin, daidzein, genistein and glycitein, respectively, and making up to 10 mL with methanol to prepare a reference stock solution; taking a certain amount of each reference stock solution, diluting to a certain concentration with methanol, and performing on-line analysis; performing linear regression with the mass concentration (x) and peak area (y) of each component, and drawing the linear regression equations of 5 reference materials.

6 Date Recue/Date Received 2021-02-26 In the following examples, the fermentation starter seed liquid was prepared according to the following method specifically as follows: 1) S. thermophilus seed liquid:
Taking a 250 mL triangular flask and adding 100 mL of an Elliker medium thereto, sterilizing at 121 C for 15 min, inoculating S.
thermophilus in a ratio of 5% to culture at 37 C to 45 C for 12-24 h, and continuously activating 2-3 times to prepare a seed medium for later use. 2) L. amylolyticus and L.
rhamnosus seed liquids: Taking two 250 mL triangular flasks, respectively adding 100 mL of an MRS liquid medium, sterilizing at 121 C for 15 min, inoculating L. amylolyticus and L. rhamnosus respectively in a ratio of 5% to culture at 32 C to 47 C for 12-24 h, and continuously activating 2-3 times to prepare a seed medium for later use.
In the following examples, the sensory evaluation of the fermented yellow serofluid was made according to the method described in GB 19302-2010 and modified appropriately.
According to a 9-point system, 1 is the worst and 9 is the best. Eight reviewers were invited to rate the odor, color, taste, and overall acceptability, and describe and analyze the odor, color, and taste.
Table 1 shows the sensory score of the yellow serofluid beverage.
Item Feature Score 1 2 3 4 5 6 7 8 Odor Pineapple juice odor, and light beany smell of the 9-7 yellow serofluid Pineapple juice odor, but heavy beany smell of 7-5 the yellow serofluid No pineapple juice odor 5-3 Abnormal odor 3-1 Color Transparent, bright and uniform light yellow .. 9-7 Non-transparent, non-bright and uniform dark 7-5 yellow or too light color Non-transparent, non-bright and non-uniform 5-3 dark yellow or too light color Abnormal color 3-1 Taste Pineapple juice taste, sweet, sour, and strong 9-7 Pineapple juice taste, soft 7-5 Light pineapple juice taste, weak and having no 5-3 aftertaste Abnormal taste 3-1 Overall Very good 9-7 acceptability Good 7-5

7 Date Recue/Date Received 2021-02-26 Poor 5-3 Very poor 3-1 Example 1: Isolation and identification of L. rhamnosus and characteristics of fermented soy whey (1) L. rhamnosus was isolated from the fermented serofluid of Kaiping, Guangdong. Its isolation and purification method was as follows: Taking 1 ml of the supernatant of acidified serofluid, diluting and spreading on an MRS agar medium, inverting to culture at 37 C overnight, and picking out a single colony on the plate to obtain cultures.
(2) Using a DNA extraction kit to extract genomic DNA, performing agarose gel electrophoresis, and then designing 16S rDNA primers for PCR amplification; sequencing the purified and recovered 16S rDNA fragment, with the sequencing result shown in the sequence listing SEQ.ID.N01; then using the BLAST software to compare the obtained 16S rDNA with the reported 16S rDNA. The comparison results showed that the similarity between the DNA sequence and L.
rhamnosus reached 99%, and so it was identified as L. rhamnosus.
(3) L. rhamnosus cultures were deposited on Aug. 25, 2017 in Guangdong Microbial Culture Collection Center at Guangdong Institute of Microbiology, 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou, having the culture collection number of GDMCC 60224.
(4) Taking 300 mL of the fresh yellow serofluid of an acidified serofluid-treated bean curd, filtering to remove insoluble matter, treating with high-pressure steam at 121 C for 15 min, and cooling to room temperature; inoculating L. rhamnosus in a sterile environment at an inoculation amount of 5%, and culturing at 37 C for 36 h. After testing and analysis, it was found that:
CD the acidity values at 12 h, 24 h and 36 h of fermentation were 50.79 T, 65.15 T and 70.25 T, respectively; CD at 12 h of fermentation, the glycoside isoflavones (daidzein and genistin) had been completely converted, the content of aglycone isoflavone glycitein had increased from 0 to 197.18 mg/L, and the content of genistein had increased from 0 to 37.06 mg/L.
Example 2 (Comparative Example): Unfermented soy whey beverage (1) Taking 2 x 300 mL of the fresh yellow serofluid of an acidified serofluid-treated bean curd and adding 15 g of sucrose thereto, fully dissolving, filtering to remove insoluble matter, treating with high-pressure steam at 121 C for 15 min, cooling, and putting in a refrigerator at 4 C to stay overnight.
Taking 300 ml to subpackage, and performing physical and chemical analysis and sensory evaluation, with the findings as follows: CD the soy whey beverage got 16.49 T for the acidity value, and 5.63

8 Date Recue/Date Received 2021-02-26 points for the sensory evaluation overall acceptability score, of which the odor score was 5.25 points, the color score was 8.25 points, and the taste score was 5.75 points; and 0 the fermented yellow serofluid got 13.81 jtmol Trolox/L for the DPPH free radical scavenging ability, and 436 jtmol Trolox/L
for the ferric reducing ability.
(2) Squeezing fresh pineapple pulp, filtering to remove the filter residue, treating with high-pressure steam at 115 C for 15 min, and cooling for later use.
(3) Taking 15 mL of the pineapple juice prepared in (2) above to add to the yellow serofluid prepared in (1) above in a sterile environment, mixing well, and putting in a refrigerator at 4 C to stay overnight.
(4) After the physical and chemical analysis and sensory evaluation, it was found that: CD the product got 26.62 T for the acidity value, and 6.50 points for the sensory evaluation overall acceptability score, of which the odor score was 6.88 points, the color score was 7.13 points, and the taste score was 6.88 points; 0 the product got 19.78 jimol Trolox/L for the DPPH free radical scavenging ability, and 409 junol Trolox/L for the ferric reducing ability;
and 0 isoflavones existed mainly in the form of daidzein (122.29 mg/L) and genistin (109.63 mg/L) (Fig.
1).
Example 3: S. thermophilus combined with L. amylolyticus to ferment soy whey beverage (1) Taking 300 mL of the fresh yellow serofluid of an acidified serofluid-treated bean curd and adding 15 g of sucrose thereto, fully dissolving, filtering to remove insoluble matter, treating with high-pressure steam at 121 C for 15 min, and cooling for later use.
(2) Inoculating respectively 3 mL of the S. thermophilus seed culture and 3 mL
of the L.
amylolyticus seed culture into the yellow serofluid prepared in (1) above, shaking well, and subpackaging in a 50 mL beaker to ferment at 42 C for 36 h. S. thermophilus was deposited on Dec.28, 2015 at an address of No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, having the culture collection number of CGMCC NO. 11943.
(3) Centrifuging the above fermented soy whey beverage, and taking the supernatant to store in a refrigerator at 4 C overnight for later physical and chemical analysis and sensory evaluation. The results were as follows: CD the product got 79.72 T for the acidity value, and 7.00 points for the sensory evaluation overall acceptability score, of which the odor score was 6.50 points, the color score was 8.00 points, and the taste score was 6.37 points; and 0 the product got 26.92 jtmol Trolox/L for the DPPH free radical scavenging ability, and 637 junol Trolox/L for the ferric reducing ability.
Example 4: S. thermophilus combined with L. amylolyticus to ferment pineapple juice-soy whey beverage

9 Date Recue/Date Received 2021-02-26 (1) Taking 300 mL of the fresh yellow serofluid of an acidified serofluid-treated bean curd and adding 15 g of sucrose thereto, fully dissolving, filtering to remove insoluble matter, treating with high-pressure steam at 121 C for 15 min, and cooling for later use.
(2) Squeezing fresh pineapple pulp, filtering to remove the filter residue, treating with high-pressure steam at 115 C for 15 min, and cooling for later use.
(3) Taking 15 mL of the pineapple juice prepared in (2) above to add to the yellow serofluid prepared in (1) above in a sterile environment, and mixing well.
(4) Inoculating respectively 3 mL of the S. thermophilus seed culture and 3 mL
of the L.
amylolyticus seed culture into the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd prepared in (3) above, shaking well, and subpackaging in a 50 mL
beaker to ferment at 42 C
for 36 h. S. thermophilus was deposited on Dec.28, 2015 at an address of No.
3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Postcode 100101, having the culture collection number of CGMCC
NO. 11943. L. amylolyticus was deposited on Apr. 25, 2014 at an address of No.
3, Yard 1, Beichen West Road, Chaoyang District, Beijing, Postcode 100101, having the culture collection number of CGMCC NO. 9090.
(5) Centrifuging the above fermented soy whey beverage, and taking the supernatant to store in a refrigerator at 4 C overnight for later physical and chemical analysis and sensory evaluation. The results were as follows: CD The product got 91.11.72 T for the acidity value, and 7.50 points for the sensory evaluation overall acceptability score, of which the odor score was 7.88 points, the color score was 8.38 points, and the taste score was 7.75 points; 0 the product got 34.14 jtmol Trolox/L for the DPPH free radical scavenging ability, and 660 jtmol Trolox/L for the ferric reducing ability; and 0 isoflavones were converted, with daidzein 95.55 mg/L, genistin 32.92 mg/L, glycitein 62.10 mg/L, and genistein 24.02 mg/L (Fig. 1).
Example 5: S. thermophilus combined with L. rhamnosus to ferment pineapple juice-soy whey beverage (1) Taking 300 mL of the fresh yellow serofluid of an acidified serofluid-treated bean curd and adding 15 g of sucrose thereto, fully dissolving, filtering to remove insoluble matter, treating with high-pressure steam at 121 C for 15 min, and cooling for later use.
(2) Squeezing fresh pineapple pulp, filtering to remove the filter residue, treating with high-pressure steam at 115 C for 15 min, and cooling for later use. The culture collection number of L.
rhamnosus is GDMCC 60224.
(3) Taking 15 mL of the pineapple juice prepared in (2) above to add to the yellow serofluid Date Recue/Date Received 2021-02-26 prepared in (1) above in a sterile environment, and mixing well.
(4) Inoculating respectively 3 mL of the S. thermophilus seed culture and 3 mL
of the L.
rhamnosus seed culture into the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd prepared in (3) above, shaking well, and subpackaging in a 50 mL beaker to ferment at 42 C for 36h.
(5) Centrifuging the above fermented soy whey beverage, and taking the supernatant to store in a refrigerator at 4 C overnight for later physical and chemical analysis and sensory evaluation. The results were as follows: 0 The product got 92.22 T for the acidity value, and 7.75 points for the sensory evaluation overall acceptability score, of which the odor score was 8.00 points, the color score was 8.13 points, and the taste score was 8.25 points; 0 the product got 40.47 umol Trolox/L for the DPPH free radical scavenging ability, and 731 umol Trolox/L for the ferric reducing ability; and 0 isoflavones were converted, with daidzein 54.87mg/L, genistin 18.57 mg/L, glycitein 157.17 mg/L, and genistein 36.80 mg/L (Fig. 1).
Example 6: S. thermophilus combined with L. amylolyticus to ferment pineapple juice-soy whey beverage (1) Same as in Example 4.
(2) Same as in Example 4.
(3) Same as in Example 4.
(4) Inoculating respectively 3 mL of the S. thermophilus seed culture and 3 mL
of the L.
amylolyticus seed culture into the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd prepared in (3) above, shaking well, and subpackaging in a 50 mL
beaker to ferment at 42 C
for 24 h.
(5) Centrifuging the above fermented yellow serofluid, and taking the supernatant to store in a refrigerator at 4 C overnight for later physical and chemical analysis and sensory evaluation. The results were as follows: 0 The product got 79.33 T for the acidity value, and 7.88 points for the sensory evaluation overall acceptability score, of which the odor score was 7.63 points, the color score was 8.50 points, and the taste score was 7.88 points; 0 the product got 31.31 umol Trolox/L for the DPPH free radical scavenging ability, and 647 umol Trolox/L for the ferric reducing ability.
Example 7: S. thermophilus combined with L. rhamnosus to ferment soy whey beverage (1) Same as in Example 5.
(2) Same as in Example 5.
(3) Same as in Example 5.

Date Recue/Date Received 2021-02-26 (4) Inoculating respectively 3 mL of the S. thermophilus seed culture and 3 mL
of the L.
rhamnosus seed culture into the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd prepared in (3) above, shaking well, and subpackaging in a 50 mL beaker to ferment at 37 C for 48 h.
(5) Centrifuging the above fermented yellow serofluid, and taking the supernatant to store in a refrigerator at 4 C overnight for later physical and chemical analysis and sensory evaluation. The results were as follows: 0 The product got 105.85 T for the acidity value, and 7.00 points for the sensory evaluation overall acceptability score, of which the odor score was 7.25 points, the color score was 7.25 points, and the taste score was 6.75 points; and 0 the product got 41.92 umol Trolox/L for the DPPH free radical scavenging ability, and 752 umol Trolox/L for the ferric reducing ability.
It can be seen from the above examples that: 0 the flavor of the yellow serofluid of an acidified serofluid-treated bean curd was not good; if the yellow serofluid of an acidified serofluid-treated bean curd was eaten directly with sugar, its sensory score would be very low and it would be difficult to be accepted by consumers; however, after a small amount of raw pineapple juice was added, its overall acceptability was greatly improved, and its antioxidation activity was correspondingly enhanced (Example 2); nevertheless, the improvement of flavor and antioxidation activity by fermentation was more significant, with its DPPH free radical scavenging ability doubled compared with that before fermentation (Example 3); 0 when S. thermophilus and L. amylolyticus were combined to ferment the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd, due to the adaptability of the cultures and the promotion of pineapple juice to the culture growth, the overall acceptability score of the fermented product was further increased to 7.50, of which the odor score was 7.88 points, the DPPH free radical scavenging ability was increased to 34.14 urnol Trolox/L, and the ferric reducing ability was 660 urnol Trolox/L (Example 4); CD when S. thermophilus and L.
rhamnosus were combined to ferment the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd, the overall acceptability of the fermented product was higher, reaching 7.75 points, of which the taste score was 8.25 points, the DPPH free radical scavenging ability reached 40.47 urnol Trolox/L, and the ferric reducing ability was 731 urnol Trolox/L, almost doubled compared with the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd before fermentation (Example 5).
These indicated that pineapple juice and fermentation had a significant effect on improving the flavor, nutrition and function of the yellow serofluid of an acidified serofluid-treated bean curd.
The cultures had significant effects on the flavor and antioxidation activity of the fermented soy whey beverage. L. rhamnosus, having a very strong ability to convert isoflavones, could completely Date Recue/Date Received 2021-02-26 convert all isoflavones into aglycones when fermenting soy whey alone at 37 C
for 12 h (Example 1).
When L. rhamnosus was combined with S. thermophilus for fermentation, the content of glycitein reached 157.17 mg/L and the content of genistein reached 36.80 mg/L (Example 5), which were higher than those obtained when L. amylolyticus was combined with S. thermophilus for fermentation (Example 4), indicating the stronger isoflavone conversion ability of L.
rhamnosus. S. thermophilus has a very high utilization rate of stachyose, and can produce extracellular polysaccharides. The combined application of cocci and bacilli can well improve the flavor of the fermented soy whey beverage, and greatly enhance the acid production and antioxidation activity. In summary, the fermented pineapple juice-soy whey beverage prepared by this patented technology has the characteristics of good flavor and outstanding antioxidation, etc., having broad market development and application prospects.

Date Recue/Date Received 2021-02-26

Claims

CA 03110913 2021-02-26

1. A method for manufacturing a functional soy whey beverage having a refreshing flavor, characterized in that: the method comprises the following steps:
1) with kilogram and liter respectively as the unit of mass and volume, adding sucrose at a mass-to-volume ratio of 0 to 6% to a fresh yellow serofluid of an acidified serofluid-treated bean curd, fully dissolving, and filtering;
2) treating the fresh yellow serofluid of an acidified serofluid-treated bean curd containing sucrose at a temperature of 110 C to 121 C for 5-20 min, and cooling;
3) sterilizing freshly squeezed pineapple juice, and cooling;
4) mixing the sterilized yellow serofluid of an acidified serofluid-treated bean curd obtained in step 2) with the sterilized pineapple juice obtained in step 3) in a sterile environment at a volume ratio of 20:1 to 25:2; and 5) inoculating an activated fermentation starter into the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd mixture obtained in step 4) at an inoculation amount of 2% to 6%
of the pineapple juice-yellow serofluid of an acidified serofluid-treated bean curd mixture, then culturing at 35 C to 45 C for 12-36 h, centrifuging to obtain a supernatant, and refrigerating the supernatant to obtain a pineapple juice-soy whey beverage; wherein the fermentation starter is a mixture of Streptococcus thermophilus and one of Lactobacillus amylolyticus and Lactobacillus rhamnosus at a volume ratio of 1:1-4;
the culture collection number of the S. thermophilus is CGMCC NO. 11943;
the culture collection number of the L. amylolyticus is CGMCC NO. 9090; and the culture collection number of the L. rhamnosus is GDMCC 60224.

2. The method for manufacturing a functional soy whey beverage having a refreshing flavor according to claim 1, characterized in that: the freshly squeezed pineapple juice is sterilized at 110 C to 115 C for 1-15 min.

3. The method for manufacturing a functional soy whey beverage having a refreshing flavor according to claim 1, characterized in that: the yellow serofluid of an acidified serofluid-treated bean curd and the pineapple juice are mixed at a volume ratio of 20:1-4.

4. The method for manufacturing a functional soy whey beverage having a refreshing flavor according to claim 1, characterized in that the method for preparing the pineapple juice is as follows:

Date Recue/Date Received 2021-02-26 squeezing pineapple pulp and filtering same through four layers of gauze to collect a filtrate.

5. The method for manufacturing a functional soy whey beverage having a refreshing flavor according to claim 1, characterized in that the method for activating S.
thermophilus is as follows:
inoculating 3% to 8% of the glycerol-preserved S. thermophilus into a sterilized Elliker medium to culture at 37 C to 45 C for 10-24 h, repeatedly activating for 2-3 times, and putting aside for later use.

6. The method for manufacturing a functional soy whey beverage having a refreshing flavor according to claim 1, characterized in that the method for activating the L.
amylolyticus and the is as follows: inoculating 3% to 8% of the glycerol-preserved L. amylolyticus or into a sterilized MRS
liquid medium to culture at 32 C to 47 C for 10-24 h, repeatedly activating for 2-3 times, and putting aside for later use.

7. The method for manufacturing a functional soy whey beverage having a refreshing flavor according to claim 1, characterized in that: the refrigeration is carried out at 4 C.

8. A functional soy whey beverage having a refreshing flavor prepared by the method according to any of claims 1 to 7.
Date Recue/Date Received 2021-02-26