Highland barley fermented beverage and preparation method thereof

Abstract

According to the highland barley fermented beverage and the preparation method thereof, highland barley is used as a raw material, leuconostoc lactis XP1, bacillus velezensis and pichia pastoris are mixed for highland barley beverage fermentation, specific raw material ratio and enzymolysis reaction conditions are adopted, and strain inoculation and fermentation process conditions are optimized, so that the highland barley fermented beverage which is fine and smooth in mouth feel, rich in nutrition and rich in nutrition is obtained. The highland barley fermented beverage is moderate in sourness and sweetness and has grain fragrance. The obtained fermented beverage has good content indexes of total dietary fiber, resistant starch and protein, and no escherichia coli is detected. The technical scheme provided by the invention can provide theoretical basis and technical support for industrial production of the fermented highland barley beverage.

Description

Technical Field

[0001] The present invention belongs to the technical field of food processing, specifically to the technical field of fermented foods, and more specifically to the technical field of a hulless barley fermented beverage and its preparation method. Background Art

[0002] Cereals are important calorie and nutrient sources in food and are the grains of Gramineae plants. The main products made from cereals include white liquor, sour porridge, cereal vinegar, fermented beverages, etc. Hulless barley, also known as naked barley, mainly grows in the northwest and southwest of China, especially in plateau areas such as Tibet, Qinghai, and Xinjiang. It is rich in protein, dietary fiber, and vitamins, while having very low fat and sugar contents. The starch content of hulless barley accounts for about 70% of the total mass. Among them, the contents of rapidly digestible starch (RDS, digested within 20 minutes), slowly digestible starch (SDS, digested within 20 - 120 minutes), and resistant starch (RS, not digested within 120 minutes) are approximately 96.19%, 1.54%, and 2.27% of the total starch content respectively. Resistant starch refers to starch that is not digested in the mouth, stomach, and small intestine but can be fermented by intestinal microbiota in the large intestine, and has many benefits such as reducing blood sugar and blood lipids, promoting intestinal health, and improving intestinal function.

[0003] Fermentation has a rich history in different food processing, and it changes food flavors, nutritional components, functional effects, etc. through the action of microorganisms. Existing research has shown that cereal beverages have great potential as carriers for probiotic microorganisms and can enhance the biological activity of strains. In the beverage Pito made by Ajiboye et al. with sorghum / millet as raw materials through fermentation, it has high antioxidant activity; Liu Ting et al. used black rice and purple rice as the main raw materials and through lactic acid bacteria fermentation, improved its antioxidant activity and α - glucosidase inhibitory activity; Gao Xiaomei et al. used black rice, red rice, and brown rice as the main raw materials, and through lactic acid bacteria fermentation, the types and contents of flavor substances in rice milk were significantly increased.

[0004] Due to its unique functional characteristics, lactic acid bacteria play an important role in the field of food fermentation, contributing to the improvement of food flavor, nutritional value, structural texture, etc. Leuconostoc lactis, as a type of lactic acid bacteria, has a relatively wide range of applications in the food industry and is mainly present in fermented dairy products, food raw materials, and functional foods. Pichia cactophila often appears in fermented foods. For example, Pichia cactophila exists in the dominant yeast genus in Inner Mongolia's koumiss; in the analysis of the microbial flora in traditional fermented sour soup in Shaanxi by Zhang Yong et al., Pichia cactophila was also found. Bacillus velezensis plays an important role in fermented soy products. It can not only decompose macromolecular substances in soybeans to produce flavor substances but also inhibit the growth of harmful bacteria, improving the safety of soy products. Moreover, Bacillus velezensis can also change the microbial community structure of Daqu, promote saccharification and ethanol fermentation, and produce flavor compounds, etc.

[0005] However, there are still some technical difficulties in existing cereal fermentations. For example, in terms of strain selection and how to exert the synergistic effect of multiple strains, there are still technical difficulties in actual production. Leuconostoc lactis, as a facultative anaerobic Gram-positive coccus or coccobacillus, is negative for catalase and belongs to the members of lactic acid bacteria. It can produce lactic acid during its growth and is a potential probiotic. Bacillus velezensis produces butyric acid during fermentation, and butyric acid has an unpleasant smell. There is also a certain technical difficulty in effectively avoiding the influence of bad smells and obtaining good sensory properties during cereal fermentation. The current application of Pichia cactophila mainly focuses on the fermentation of dairy products, mainly in the fermentation of koumiss and milk residue. There is currently no report on whether it exhibits fermentation characteristics in the fermentation of cereal products.

[0006] Therefore, how to select Leuconostoc lactis and apply it in compound fermentation with Bacillus velezensis and Pichia cactophila, and exert excellent fermentation characteristics in highland barley fermented beverages is of great significance for further enhancing the nutritional value of highland barley. Developing a highland barley fermented beverage with a suitable sweet and sour taste can meet consumers' needs for healthy beverages, bring new choices to the market, increase the added value of highland barley, and further extend the product chain. Summary of the Invention

[0007] In view of the technical problem that there is currently no research report on the preparation of highland barley fermented beverages by the compounding of Leuconostoc lactis XP1, Bacillus velezensis, and Pichia cactophila in the prior art, a highland barley fermented beverage and its preparation method are provided. This method uses highland barley as the raw material, undergoes mixed fermentation with 3 strains of bacteria, adopts specific raw material ratios and enzymatic hydrolysis reaction conditions, and optimizes the strain inoculation and fermentation process conditions to obtain a highland barley fermented beverage with a delicate and smooth taste, moderate sourness and sweetness, and a cereal aroma. The obtained fermented beverage has good total dietary fiber, resistant starch, and protein content indicators, and Escherichia coli is not detected. The technical solution provided by this application can provide a theoretical basis and technical support for the industrial production of fermented highland barley beverages.

[0008] To achieve this technical purpose, the present invention adopts the following technical solutions:

[0009] On the one hand, this application provides a highland barley fermented beverage, and the highland barley fermented beverage includes: highland barley, purified water, a mixed bacterial agent, a compound enzyme, and a saccharifying enzyme raw material; the mixed bacteria adopt Leuconostoc lactis XP1, Pichia cactophila, and Bacillus velezensis; the compound enzyme adopts α-amylase and saccharifying enzyme; the total dietary fiber in the highland barley fermented beverage is 1.72%; the resistant starch content is 0.80%; the protein content is 0.97%.

[0010] Further, among the mixed bacteria, the viable cell count of all 3 strains of bacteria is 10 8 CFU / mL, and the inoculation ratio of each strain of Bacillus velezensis:Pichia cactophila:Leuconostoc lactis XP1 in the mixed bacteria is 10:10:1.

[0011] Further, the material-to-water ratio of the highland barley to the purified water is set to: 1:(8 - 16).

[0012] Preferably, the material-to-water ratio of the highland barley to the purified water is set to: 1:8.

[0013] This application also provides a preparation method for the highland barley fermented beverage, and the highland barley fermented beverage is obtained by the following steps:

[0014] (1) Wash the selected plump and round highland barley grains 3 times with clean water and then drain the water. Stir-fry over low heat for 35 minutes until the highland barley emits a strong cereal aroma;

[0015] (2) Cool the highland barley stir-fried in step (1), crush it with a pulverizer, add water and pass it through a colloid mill for 15 minutes, and then boil it for 10 minutes at a certain material-to-water ratio;

[0016] (3) Add the cooked highland barley obtained in step (2) to the complex enzyme and enzymolyze for a certain period of time at the optimal enzyme reaction temperature. After cooling to room temperature, inoculate with the complex bacteria, ferment at the appropriate temperature and time, and then sterilize to obtain the mixed bacteria fermented highland barley beverage.

[0017] Preferably, the inoculation order of the complex bacteria is: inoculate Bacillus velezensis, Leuconostoc lactis XP1 and Pichia cactophila at the beginning of fermentation.

[0018] Preferably, the inoculation amount of the complex bacteria is 1%-9%, the fermentation temperature is 28°C-37°C, and the fermentation time is 24h-72h.

[0019] More preferably, the inoculation amount of the complex bacteria is 7%, the fermentation temperature is 35°C, and the fermentation time is 50h.

[0020] Preferably, the sterilization procedure is to sterilize at 121°C and 0.1 MPa for 3 min.

[0021] Preferably, in the enzymolysis step, the complex enzyme used is α-amylase and glucoamylase, and the addition ratio is 1:2; the addition amount of the complex enzyme is 0.8%, and under the condition that the enzyme reaction temperature is 60°C, the enzyme reaction time is 0.5 h.

[0022] On the other hand, the present application also provides the application of the highland barley fermented beverage or the highland barley fermented beverage prepared by the preparation method of the highland barley fermented beverage in the preparation of functional foods, and the function is to control blood sugar through resistant starch components.

[0023] Regarding the issue of using existing well-known common knowledge as the basis of the prior art for new inventions and creations. In fact, except for pioneering inventions, any invention and creation is inseparable from the prior art means and technical elements as the basis, and is the result of further innovation on the basis of the prior art means. Technologies such as cleaning, frying, grinding, colloid mill, adding bacteria for fermentation, and sterilization are all the basis of the prior art. However, how to fully retain the nutrients in hulless barley, how to coordinate the ratio between raw and auxiliary materials, and how to make the hulless barley fermented beverage obtain the characteristics of good taste, good flavor, and rich nutrition through a series of processes all need to be repeatedly verified through a series of unforeseeable scientific experiments. Due to these reasons, although this application uses conventional technical means as the basis and provides a hulless barley fermented beverage and its preparation method through scientific experiments on this basis, the various technical steps in the provided preparation method of the hulless barley fermented beverage form an integral whole, with overall indivisibility, and the technical steps cannot be arbitrarily disassembled. Only in this way can the prepared fermented beverage achieve the effect of having good total dietary fiber, resistant starch, and protein content indicators, not only overcoming the problems with adverse effects during the fermentation process of a single strain, but also improving the use value and nutritional function of the product. It meets the requirements of the development of the modern food industry and has broad practical value.

[0024] By implementing the technical solution of the present invention, the following beneficial effects can be achieved:

[0025] This application provides a preparation method of a hulless barley fermented beverage. The optimal process conditions for the pretreatment of the fermented hulless barley beverage are mixed fermentation with 3 strains of bacteria, a material-to-water ratio of 1:8, and an enzyme reaction time of 0.5 h; the optimal process conditions for the fermented hulless barley beverage are synchronous fermentation with 3 strains of bacteria, an inoculation ratio of 10:10:1, an inoculation amount of 7%, a fermentation temperature of 35 °C, and a fermentation time of 50 h. Under these process conditions, the taste is delicate, smooth, moderately sour and sweet, and there is a cereal aroma. Among them, the total dietary fiber is 1.72%; the resistant starch content is 0.80%; the protein content is 0.97%, and Escherichia coli is not detected. The technical solution provided by this application can provide a theoretical basis and technical support for the industrial production of the fermented hulless barley beverage. Description of the Drawings

[0026] Figure 1 Shown is the glucose standard curve diagram of the 1 mL system.

[0027] Figure 2 Shown is the diagram of the influence of different strain combinations on the resistant starch, non-resistant starch, and sensory score of the hulless barley fermented beverage.

[0028] In the figure, strain 1 represents Bacillus velezensis, strain 2 represents Leuconostoc lactis XP1, and strain 3 represents Pichia cactophila. The same expressions will be used hereinafter.

[0029] Figure 3 The figure shows the influence of different ratios of material to water on the contents of resistant starch and non-resistant starch and the sensory score of fermented highland barley beverage.

[0030] Figure 4 The figure shows the influence of different enzyme reaction times on the reducing sugar and starch contents of mixed-strain fermented highland barley beverage.

[0031] Figure 5 The figure shows the color change of the reaction between starch and iodine at different enzyme reaction times.

[0032] Figure 6 The figure shows the influence of inoculation sequence on the quality of highland barley fermented beverage.

[0033] Figure 7 The figure shows the influence of inoculation ratio on the quality of highland barley fermented beverage.

[0034] Figure 8 The figure shows the influence of inoculation amount on the quality of highland barley fermented beverage.

[0035] Figure 9 The figure shows the influence of fermentation temperature on the quality of highland barley fermented beverage.

[0036] Figure 10 The figure shows the influence of fermentation time on mixed-strain fermented highland barley beverage.

[0037] Figure 11 The figure shows the response surface plot and contour plot of the inoculation sequence and fermentation temperature on the comprehensive response value.

[0038] Among them, Figure A shows the response surface plot, and Figure B shows the contour plot.

[0039] Figure 12 The figure shows the response surface plot and contour plot of the inoculation sequence and fermentation time on the comprehensive response value.

[0040] Among them, Figure A shows the response surface plot, and Figure B shows the contour plot.

[0041] Figure 13 The figure shows the response surface plot and contour plot of the fermentation temperature and fermentation time on the comprehensive response value.

[0042] Among them, Figure A shows the response surface plot, and Figure B shows the contour plot. Detailed implementation method

[0043] The technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts belong to the scope of protection of the present invention.

[0044] In this embodiment, the reagents selected are: MRS broth medium, MRS agar medium, nutrient broth (NB) medium, yeast extract peptone dextrose agar medium (YPD), potato dextrose agar medium (PDA): Qingdao Haibo Biotechnology Co., Ltd.; α-amylase (biochemical reagent), Xingtai Wanda Bioengineering Co., Ltd.; glucoamylase (biochemical reagent), Shanghai Yuanye Bio-Technology Co., Ltd.; iodine (analytical pure), Tianjin Beilian Fine Chemicals Development Co., Ltd.; resistant starch and non-resistant starch content (enzymatic method) kit (product number: G0571W): Suzhou Grees Biotech Co., Ltd.

[0045] The instruments selected in this application are: SSW-420-2S electrothermal constant temperature water bath, Shanghai Boxun Industry Co., Ltd.; A1204 electronic analytical balance, Mettler-Toledo Group, Switzerland; Phs-25 pH meter, Shanghai Leici Instrument Factory; SW-CJ-2D double-person purification workbench, Zhejiang Fuxia Medical Technology Co., Ltd.; DHP-9162 constant temperature incubator, Shanghai Yiheng Scientific Instrument Co., Ltd.; BL-75A vertical pressure steam sterilizer, Shanghai Boxun Industry Co., Ltd.; HF-200 type pulverizer, Chenheshengfeng Industry and Trade Co., Ltd.

[0046] The Lactococcus lactis subsp. cremoris XP1 selected in this application is a strain self-screened by this laboratory in the early stage, which has been deposited in the China General Microbiological Culture Collection Center, with the deposit number of CGMCC No. 31789 and the deposit date of August 30, 2024. The general public can obtain it by contacting the applicant's research group or purchasing it through public channels such as the China General Microbiological Culture Collection Center (CGMCC).

[0047] The Bacillus velezensis and Pichia cactophila selected in this application are both conventional strains, and the general public can purchase them through public channels such as the China General Microbiological Culture Collection Center (CGMCC).

[0048] The highland barley was purchased from Taxkorgan Tajik Autonomous County, Xinjiang, located on the Pamir Plateau.

[0049] Unless otherwise specified in the following implementation cases, the technical means used are conventional means well known to those skilled in the art.

[0050] Example 1: A highland barley fermented beverage

[0051] The present application provides a hulless barley fermented beverage, which includes: hulless barley, purified water, a mixed bacterial agent, a compound enzyme, and a saccharifying enzyme raw material; the mixed bacteria are Leuconostoc lactis XP1, Pichia cactophila, and Bacillus velezensis; the compound enzyme is α-amylase and saccharifying enzyme; the total dietary fiber in the hulless barley fermented beverage is 1.72%; the resistant starch content is 0.80%; the protein content is 0.97%.

[0052] Example 2: A preparation method of a hulless barley fermented beverage

[0053] The selected plump and round hulless barley grains are rinsed 3 times with clear water and then drained, stir-fried over low heat for 35 minutes until the hulless barley emits a strong grain aroma; after the hulless barley cools, it is crushed with a pulverizer and then passed through a colloid mill with water for 15 minutes, and then boiled for 10 minutes at a certain solid-to-liquid ratio; after boiling, the compound enzyme is added to the hulless barley at the optimal enzyme reaction temperature for enzymatic hydrolysis for a certain period of time, cooled to room temperature, inoculated with the compound bacteria, fermented at a suitable temperature and time, and then sterilized to obtain a mixed bacteria fermented hulless barley beverage.

[0054] The inoculation sequence of the compound bacteria is: inoculate Bacillus velezensis, Leuconostoc lactis XP1, and Pichia cactophila at the beginning of fermentation.

[0055] The inoculation amount of the compound bacteria is 1% - 9%, the fermentation temperature is 28°C - 37°C, and the fermentation time is 24h - 72h.

[0056] The inoculation amount of the compound bacteria is 7%, the fermentation temperature is 35°C, and the fermentation time is 50h.

[0057] The sterilization procedure is to sterilize at 121°C and 0.1 MPa for 3 minutes.

[0058] In the enzymatic hydrolysis step, the compound enzyme is α-amylase and saccharifying enzyme, and the addition ratio is 1:2; the addition amount of the compound enzyme is 0.8%, and under the condition that the enzyme reaction temperature is 60°C, the enzyme reaction time is 0.5h.

[0059] Example 3: A preparation method of a hulless barley fermented beverage

[0060] The selected plump and round highland barley grains are rinsed 3 times with clear water and then drained. They are stir-fried over low heat for 35 minutes until the highland barley emits a strong grain aroma. After the highland barley cools, it is ground with a pulverizer and then passed through a colloid mill with water added for 15 minutes. Then, it is boiled for 10 minutes at a material-to-water ratio of 1:8. After boiling, a complex enzyme composed of α-amylase and glucoamylase in a ratio of 1:2 is added to the highland barley at 60 °C. The addition amount of the complex enzyme is 0.8%, and the reaction time is 0.5 h. After cooling to room temperature, a complex bacterium is inoculated. When starting fermentation, Bacillus velezensis, Leuconostoc lactis XP1, and Pichia cactophila are inoculated. The inoculation ratio of Bacillus velezensis:Pichia cactophila:Leuconostoc lactis XP1 is 10:10:1, and the inoculation amount is 7%. The fermentation temperature is 35 °C, and the fermentation time is 50 h. After fermentation, sterilization is carried out under the conditions of 121 °C and 0.1 MPa for 3 minutes to obtain the mixed-bacterium fermented highland barley beverage.

[0061] Example 4: A preparation method of a highland barley fermented beverage

[0062] The selected plump and round highland barley grains are rinsed 3 times with clear water and then drained. They are stir-fried over low heat for 35 minutes until the highland barley emits a strong grain aroma. After the highland barley cools, it is ground with a pulverizer and then passed through a colloid mill with water added for 15 minutes. Then, it is boiled for 10 minutes at a material-to-water ratio of 1:8. After boiling, a complex enzyme composed of α-amylase and glucoamylase in a ratio of 1:2 is added to the highland barley at 60 °C. The addition amount of the complex enzyme is 0.8%, and the reaction time is 1 h. After cooling to room temperature, a complex bacterium is inoculated. When starting fermentation, Bacillus velezensis, Leuconostoc lactis XP1, and Pichia cactophila are inoculated. The inoculation ratio of Bacillus velezensis:Pichia cactophila:Leuconostoc lactis XP1 is 1:10:1, and the inoculation amount is 1%. The fermentation temperature is 28 °C, and the fermentation time is 24 h. After fermentation, sterilization is carried out under the condition of sterilizing at 72 °C for 15 s - 30 min to obtain the mixed-bacterium fermented highland barley beverage.

[0063] Example 5: A preparation method of a highland barley fermented beverage

[0064] The selected plump and round highland barley grains are rinsed 3 times with clear water and then drained. They are stir-fried over low heat for 35 minutes until the highland barley emits a strong grain aroma. After the highland barley cools, it is ground with a pulverizer and then passed through a colloid mill with water for 15 minutes. Then, it is boiled for 10 minutes at a material-to-water ratio of 1:16. After boiling, a compound enzyme composed of α-amylase and glucoamylase at an addition ratio of 1:2 is added to the highland barley at 60°C. The addition amount of the compound enzyme is 0.8%, and the reaction time is 0.5 h. After cooling to room temperature, a compound bacterium is inoculated, and when starting fermentation, Bacillus velezensis, Leuconostoc lactis XP1, and Pichia cactophila are inoculated. The inoculation ratio of Bacillus velezensis:Pichia cactophila:Leuconostoc lactis XP1 is 10:1:10, the inoculation amount is 9%, the fermentation temperature is 37°C, and the fermentation time is 72 h. After fermentation, sterilization is carried out, and the sterilization conditions are sterilization at 100°C for 5 minutes to obtain the mixed-bacteria fermented highland barley beverage.

[0065] Example 6: Optimization of the conditions for fermenting highland barley beverage

[0066] Based on the descriptions in the above Examples 1 to 5, the conditions for fermenting highland barley beverage are optimized.

[0067] I. Test method

[0068] 1. Process flow of highland barley beverage

[0069] The selected plump and round highland barley grains are rinsed 3 times with clear water and then drained. They are stir-fried over low heat for 35 minutes until the highland barley emits a strong grain aroma, cooled, ground with a pulverizer and then passed through a colloid mill with water for 15 minutes. Then, it is boiled for 10 minutes at a certain material-to-water ratio, and liquefying enzyme and glucoamylase are added at the optimal enzyme reaction temperature for enzymatic hydrolysis for a certain time. After cooling to room temperature, a compound strain is inoculated, and fermentation is carried out at a suitable temperature and time to obtain the mixed-bacteria fermented highland barley beverage.

[0070] 2. Optimization of the pretreatment conditions for fermenting highland barley beverage

[0071] (1) Determination of fermentation strains

[0072] The material-to-water ratio is 1:20, the inoculation amount is 3%, the inoculation ratio is 1, 1:1, 1:1:1, the fermentation temperature is 35°C, and the fermentation time is 48 h. Different strain combinations are selected for synchronous inoculation, and their viable bacteria count, resistant starch and non-resistant starch contents, and sensory scores are evaluated. The control group is the beverage without adding strains. The strains are selected as shown in Table 1 below:

[0073] Table 1: Selection of strains

[0074] Bacterium No. 1 Bacterium No. 2 Bacterium No. 3 + - - + + - + - + + + +

[0075] Note: "+" indicates that the strain is selected for fermentation, and "-" indicates that the strain is not selected for fermentation.

[0076] In the table, Bacillus velezensis is represented by strain 1, Leuconostoc lactis XP1 is represented by strain 2, and Pichia cactophila is represented by strain 3. The same expressions are used hereinafter.

[0077] (2) Determination of the material-water ratio

[0078] Before inoculation, adjust the viable count of the 3 strains to 10 8 CFU / mL. Set the material-water ratios as: 1:8, 1:10, 1:12, 1:14, 1:16, the inoculation amount as 3%, the inoculation ratio as 1:1:1, the fermentation temperature as 35 °C, and the fermentation time as 48 h. Take the sensory score, resistant starch and non-resistant starch contents as evaluation indexes.

[0079] (3) Determination of the enzyme reaction time

[0080] Based on the previous research in the laboratory, under the conditions that the addition ratio of α-amylase to glucoamylase is 1:2, the addition amount of the complex enzyme is 0.8%, and the enzyme reaction temperature is 60 °C, optimize the enzyme reaction time. Measure the reducing sugar and starch contents at 0, 0.5, 1, 1.5, 2, 2.5 h of the enzyme reaction.

[0081] 3. Process optimization of fermented highland barley beverage

[0082] (1) Single-factor experiment

[0083] ① Determination of the strain inoculation order

[0084] On the basis of the pretreatment conditions of the fermented highland barley beverage, take the inoculation amount of 3%, the inoculation ratio of 1:1:1, the fermentation temperature of 35 °C, and the fermentation time of 48 h as the fermentation conditions, and determine the strain inoculation order with the resistant starch content and sensory score as the optimization values. The strain inoculation order is shown in Table 2 below:

[0085] Table 2: Strain inoculation order

[0086]

[0087] Note: "-" indicates that the strain is not selected.

[0088] ② Determination of the strain compounding ratio

[0089] The strain compounding ratios are shown in Table 3 below:

[0090] Table 3: Strain compounding ratio

[0091] Group Bacterium No. 1 Bacterium No. 3 Bacterium No. 2 1 1 10 1 2 1 1 10 3 10 1 1 4 10 10 1 5 1 10 10 6 10 1 10 7 1 1 1

[0092] ③ Determination of the inoculation amount

[0093] Set the inoculation amounts as: 1%, 3%, 5%, 7%, 9% respectively.

[0094] ④Determination of fermentation temperature

[0095] Set the fermentation temperatures to: 28°C, 30°C, 32°C, 35°C, 37°C respectively.

[0096] ⑤Determination of fermentation time

[0097] Set the fermentation times to: 24h, 36h, 48h, 60h, 72h respectively.

[0098] 4. Response surface experiment design

[0099] According to the results of single-factor experiments, select 3 factors (A: inoculation order, B: fermentation temperature, C: fermentation time) that have a greater impact on the sensory score and sugar-acid ratio of the mixed-strain fermented hulless barley beverage. According to the Box-Benhnken central composite experimental design principle of Design-Expert 13 software, use the response surface analysis method with 3 factors and 3 levels for design. To facilitate the implementation of the response surface experiment, record the 4th, 5th, and 6th groups of the inoculation order of strain in Table 2 in the above experiment as levels 1, 3, and 2 respectively, the fermentation temperature levels are 33°C, 35°C, 37°C, and the fermentation time levels are 36h, 48h, 60h. The factor and level design of the response surface analysis is shown in Table 4. The comprehensive response value Y is composed of two parts, where the sensory score Y1 accounts for 70%, and the sugar-acid ratio Y2 accounts for 30%. Taking the highest actual sensory score value as 70 points, the higher the actual sensory score, the higher the value of Y1. Taking the lowest actual sugar-acid ratio value as 30 points, the lower the actual sugar-acid ratio, the higher the value of Y2.

[0100] Table 4: Factors and levels of response surface analysis

[0101]

[0102] 5. Sensory evaluation of hulless barley fermented beverage

[0103] The sensory scores are shown in Table 5.

[0104] Table 5: Sensory score table

[0105]

[0106]

[0107] 6. Determination indexes and methods for process optimization

[0108] (1) Determination of resistant starch and non-resistant starch contents

[0109] Use the kits for determining the contents of resistant starch and non-resistant starch (enzymatic method) for determination.

[0110] (2) Determination of reducing sugar content

[0111] ① Drawing of glucose standard curve

[0112] The experimental design table for the glucose standard curve is shown in Table 6.

[0113] Table 6: Experimental design table for drawing glucose standard curve

[0114]

[0115]

[0116] Take appropriate amounts of the test solutions with different glucose contents into a 96-well plate, and measure their absorbance values under the condition of 540 nm. Zero the instrument with the test solution with a glucose content of 0 mg / mL. Draw a glucose standard curve with the glucose concentration as the abscissa and the absorbance value as the ordinate.

[0117] ② Determination of reducing sugar content

[0118] The specific method is as follows: Take 100 μL of the mixed bacteria-fermented highland barley beverage diluted by an appropriate multiple, add 100 μL of distilled water and 400 μL of DNS, mix well, immediately take it out after boiling water bath for 5 minutes and place it in a cold water bath for cooling, and then add 400 μL of distilled water after cooling. Take an appropriate amount of the test solution and add it to a 96-well plate, and measure its absorbance value at 540 nm.

[0119] ③ Glucose standard curve

[0120] The glucose standard curve is shown in the attached figure:

[0121] ④ Determination of other indicators

[0122] Determination of total acid: Potentiometric titration method with a pH meter, referring to GB 12456-2021;

[0123] Determination of total number of colonies and viable bacteria is carried out referring to GB 4789.2-2022.

[0124] 7. Determination of sterilization conditions

[0125] To avoid the influence of excessive microbial fermentation on the sensory score of the beverage, the beverage was sterilized in the experiment. Specifically: Sterilize the beverage at 72 °C for 15 s, 30 s, 1 min, 5 min, 10 min, 20 min, 30 min respectively, at 100 °C for 5 min, and at 121 °C, 0.1 MPa for 3 min, 5 min. Start timing after the temperature rises to the corresponding sterilization temperature, and measure the number of viable bacteria in the beverage after sterilization and cooling to determine the sterilization conditions of the beverage.

[0126] 8. Quality analysis of fermented highland barley beverage

[0127] Dietary fiber content: Determined by the enzymatic gravimetric method, referring to GB5009.88-2023; Resistant starch content: Determined by the enzymatic hydrolysis method, NY / T4358-2023; Protein content: Determined by the Kjeldahl method, GB5009.5-2016; Fat content: Determined by the Soxhlet extraction method, GB5009.6-2016.

[0128] 9. Data Analysis

[0129] In this example, all tests and analyses were repeated 3 times. Data processing and significance analysis (P < 0.05) were performed using SPSS 27, and the results were expressed as "mean ± standard deviation". Graphs were plotted using Origin 2022.

[0130] II. Experimental Results

[0131] 1. Optimization of Pretreatment Conditions for Fermented Hulless Barley Beverage

[0132] (1) Influence of Strain Selection on Fermented Hulless Barley Beverage

[0133] See Appendix Figure 2 It can be seen that after adding the strains, the resistant starch content increased significantly (P < 0.05). The resistant starch content was the highest when fermented with Bacillus velezensis strain 1 alone. As the proportion of Bacillus velezensis strain 1 decreased, the resistant starch content also decreased; when Leuconostoc lactis XP1 strain 2 was present in the combination, the resistant starch content decreased compared to other combinations. This may be because Leuconostoc lactis XP1 strain 2 decomposed the resistant starch.

[0134] After adding strains, the content of resistant starch decreased extremely significantly (P<0.01). Among them, the reduction of resistant starch in three combinations, namely Bacillus velezensis of strain 1, Bacillus velezensis of strain 1 + Pichia cactophila of strain 3, and Bacillus velezensis of strain 1 + Pichia cactophila of strain 3 + Leuconostoc lactis XP1 of strain 2, was relatively high. When fermented with Bacillus velezensis of strain 1 alone, the reduction of resistant starch was the highest, but there was no significant difference among the three combination methods; the content of resistant starch in the combination of Bacillus velezensis of strain 1 + Leuconostoc lactis XP1 of strain 2 was higher than that of other combinations. When fermented with Bacillus velezensis of strain 1 alone, the sensory score was the lowest. The main reason was that Bacillus velezensis would produce butyric acid during fermentation, and butyric acid had an unpleasant smell; when Leuconostoc lactis XP1 of strain 2 was present, the sour taste of the fermented highland barley beverage was too strong and could not cover the smell produced by the fermentation of Bacillus velezensis of strain 1, resulting in too low sensory score; the combination of Bacillus velezensis of strain 1 + Pichia cactophila of strain 3 + Leuconostoc lactis XP1 of strain 2 had the highest sensory score, mainly because Pichia cactophila of strain 3 belongs to the genus Pichia, and the genus Pichia has been proven to be an important contributor to the flavor of some fermented foods, so it could neutralize the smell of spores to a certain extent. The sensory scores of the highland barley beverages fermented by the four combinations did not exceed 50 points. One of the reasons was that the material-to-water ratio of 1:20 was too low, resulting in no cereal fragrance in the four groups of fermented highland barley beverages and the overall taste of the beverage not being thick enough. Therefore, in the subsequent optimization process, to further improve the sensory score, Bacillus velezensis of strain 1 needed to improve the decomposition, utilization and conversion rate of starch, lactic acid bacteria provided sour taste, and yeast enhanced the aroma of the fermented highland barley beverage. In summary, this application selected the mixed fermentation of three strains (Leuconostoc lactis XP1, Pichia cactophila and Bacillus velezensis) for highland barley grain beverage.

[0135] According to Table 7, the total colony counts of the fermented highland barley beverages with different strain combinations were all higher than 7 log CFU / mL. Among them, the combination of Bacillus velezensis of strain 1 + Pichia cactophila of strain 3 had the highest total colony count, which was 7.77±0.02 log CFU / mL; followed by the single fermentation of Bacillus velezensis of strain 1, and the total colony count was 7.63±0.08 log CFU / mL. When Leuconostoc lactis XP1 of strain 2 was present in the combination, the total colony count decreased, and the colony counts of lactic acid bacteria were all higher than those of other strains. This might be because Leuconostoc lactis XP1 of strain 2 produced acid during fermentation, resulting in a decrease in pH value and having an inhibitory effect on most bacteria. Therefore, it had a certain inhibitory effect on the growth of Bacillus velezensis of strain 1 and Pichia cactophila of strain 3. This corresponded to the results of the resistant starch content shown in the figure. Therefore, in the subsequent experiment, the inoculation order of lactic acid bacteria was placed at the end.

[0136] Table 7: Viable cell count of fermented highland barley beverage with different probiotic combinations (log CFU / mL)

[0137] Group Number of spore colonies Number of lactic acid bacteria colonies Number of yeast colonies Total number of colonies Bacterium No. 1 7.63±0.08 - - 7.63±0.08 Bacterium No. 1 + Bacterium No. 2 7.00±0.26 7.21±0.28 - 7.42±0.27 Bacterium No. 1 + Bacterium No. 3 7.74±0.02 - 6.64±0.19 7.77±0.02 Bacterium No. 1 + Bacterium No. 2 + Bacterium No. 3 6.78±0.34 7.22±0.12 6.56±0.07 7.49±0.16

[0138] (2) Effect of solid-to-liquid ratio on fermented highland barley beverage

[0139] See Appendix Figure 3 It can be seen that increasing the solid-to-liquid ratio can significantly reduce the aroma of the fermented highland barley beverage. Compared with the solid-to-liquid ratio of 1:20 in the experiment, the fragrance of the fermented highland barley beverage is extremely significantly improved, and the sensory score is the highest when the solid-to-liquid ratio is 1:10. The contents of resistant starch and non-resistant starch decrease with the increase of the solid-to-liquid ratio, and are the highest when the solid-to-liquid ratio is 1:8. This is because the solid-to-liquid ratio affects the substrate concentration of the enzymatic reaction, thereby affecting the enzymatic hydrolysis of proteins and starches in highland barley, and thus affecting the contents of resistant starch and non-resistant starch. Therefore, a solid-to-liquid ratio of 1:8 is selected for subsequent experiments.

[0140] (3) Effect of enzyme reaction time on fermented highland barley beverage

[0141] See Appendix Figure 4 It can be seen that with the extension of the enzyme reaction time, the reducing sugar content of the highland barley rice paste increases significantly, and the starch content decreases significantly. After 1 h of reaction, the starch content drops below 4 mg / mL and there is no significant difference. The iodine color reaction is carried out on the solutions at each enzyme reaction time. See Appendix Figure 5 It can be seen that when the enzyme reaction time is 1.5 h, the color reaction between starch and iodine begins to show reddish brown, indicating that the starch is fully liquefied at this time. Since the mixed bacteria also have a certain liquefaction ability for starch, and blood glucose needs to be controlled in this study, the enzyme reaction time is selected as 0.5 h. At this time, the starch content is 30.06 mg / mL and the reducing sugar content is 5.68 mg / mL.

[0142] 2. Process optimization of fermented highland barley beverage

[0143] (1) Effect of inoculation order on fermented highland barley beverage

[0144] See Appendix Figure 6It can be seen that when Pichia opuntiae strain No. 3 was inoculated first at the 0th hour, the sugar-acid ratio was relatively high, indicating that Pichia opuntiae strain No. 3 has relatively weak ability to produce acid by utilizing reducing sugar. When the three strains were inoculated simultaneously, the sugar-acid ratio was the lowest, indicating that simultaneous inoculation has a stronger ability to produce acid by utilizing reducing sugar. In terms of sensory evaluation scores, there were off-flavors in the first 4 groups, and there was a certain bitter and astringent taste in the 5th group. This shows that the flavor of segmented inoculation fermentation is not good, probably because the inoculum amount was too small at the beginning and the fermentation time was too short. The sensory evaluation score of the 6th group was the highest, and it was synchronous fermentation. The inoculation process was more convenient and fast, and at the same time, the risk of contamination could be greatly reduced. Therefore, synchronous fermentation was selected for subsequent experiments, that is, Lactococcus lactis subsp. cremoris XP1, Pichia opuntiae, and Bacillus velezensis were fermented synchronously for subsequent experiments.

[0145] (2) Influence of inoculation ratio on fermented highland barley beverage

[0146] It can be seen from Figure 7 that in the first group, when the single-strain ratio of Pichia opuntiae increased, the fruity aroma produced by the strain was too strong, resulting in a relatively low sensory evaluation score; in the third group, when the single-strain ratio of Bacillus velezensis increased, the sensory evaluation score was relatively low due to the poor smell. The sugar-acid ratio of the second group was the lowest, indicating that when the single-strain addition ratio of Lactococcus lactis subsp. cremoris XP1 increased, the ability to produce acid by utilizing reducing sugar was relatively strong, and the sensory evaluation score of this group was the best. In the second group, due to the low addition ratios of Bacillus velezensis and Pichia opuntiae, the overall smell of the beverage was light, the cereal aroma was prominent, and the taste was appropriate, resulting in a relatively high score. However, there was no significant difference in the sensory evaluation score and sugar-acid ratio between this group and the fourth group. Considering the reason of improving enzyme activity, the fourth group was selected for subsequent experiments, that is, the inoculation ratio of Bacillus velezensis, Pichia opuntiae, and Lactococcus lactis subsp. cremoris XP1 was 10:10:1.

[0147] (3) Influence of inoculum amount on fermented highland barley beverage

[0148] See appendix Figure 8 The results showed that the inoculum amount had no significant effect on the sugar-acid ratio. In the actual measurement results, as the inoculum amount increased, the reducing sugar content continued to decrease, and the total acid content also continued to decrease, but there was no significant difference. As the inoculum amount increased, the sensory evaluation score showed a trend of first increasing and then decreasing. The sensory evaluation score was the highest when the inoculum amount was 7%, which was 73.96±0.12. When the inoculum amount was 9%, the sensory evaluation score had no significant difference from that at 7%. Considering the cost, 7% was selected as the optimal inoculum amount for subsequent experiments.

[0149] (4) Influence of fermentation temperature on fermented highland barley beverage

[0150] See appendix Figure 9As shown, with the increase of fermentation temperature, the sugar-acid ratio of the highland barley fermented beverage shows an upward trend. At 28°C, the beverage has a strong aroma but a sour taste; at 32°C and 37°C, the sensory scores are low, which may indicate that the temperature is not suitable for the co-fermentation of the three strains; at 35°C, the sensory score is the highest. At this time, the beverage has a sweet smell and a moderate sweet and sour taste. In this application, 35°C with the highest sensory score is selected as the optimal temperature for fermenting highland barley beverage.

[0151] (5) Influence of fermentation time on fermented highland barley beverage

[0152] See the appendix Figure 10 As shown, with the extension of fermentation time, the sugar-acid ratio of the highland barley fermented beverage shows a trend of first decreasing and then increasing, reaching the lowest at the 48th hour. At 24 hours of fermentation, the highland barley beverage has a slightly sour smell without sweetness, and a sour and astringent taste with a bitter aftertaste; by the 48th hour, the fermented beverage has a sweet smell, a grain aroma, and a suitable sweet and sour taste. As the fermentation time continues to increase, the highland barley beverage has an alcohol smell, a sour taste with a slight bitterness. According to the test results, 48 hours is selected as the optimal fermentation time for the mixed-strain fermentation of highland barley beverage.

[0153] 3. Response surface experiment for fermentation process optimization

[0154] (1) Results and analysis of response surface experiment

[0155] The response surface method is a modeling method that integrates mathematical and statistical principles. This method analyzes the relationship between process parameters and response variables through a limited experimental design to optimize the extraction efficiency or the content of target compounds, and has the characteristics of high experimental efficiency, high precision, and short cycle. Based on the single-factor experiment, according to the Box-Benhnken central composite experimental design principle of Design-Expert 13 software, a 3-factor 3-level response surface analysis method is used for design. Three factors (A: inoculation order, B: fermentation temperature, C: fermentation time) that have a greater impact on the sensory score and sugar-acid ratio of the mixed-strain fermentation of highland barley beverage are selected as independent variables. The comprehensive response value Y is composed of two parts, where the sensory score Y1 accounts for 70% and the sugar-acid ratio Y2 accounts for 30%. The experimental design and results are shown in Table 8.

[0156] Table 8: Experimental design and results for response surface optimization

[0157]

[0158]

[0159] Performing multiple regression fitting on the experimental data, the obtained regression equation model is:

[0160] Y = -2771.79 + 109.65A + 150.11B + 5.52C - 0.20AB - 0.05AC

[0161] -0.03BC - 26.26A 2 -2.16B 2 -0.04C 2

[0162] As can be seen from the analysis of variance table 9, the regression model is highly significant (P < 0.0001), and the main effects of A, B, C and the quadratic terms of A 2 and B 2 and C 2 all reach the significant level, while the interaction terms AB, AC, and BC are not significant, indicating that the interaction among factors is weak within the current factor range. As can be seen from Table 10, the R 2 of the model is 0.9959, Adj R 2 is 0.9907, and Pred R 2 is 0.9752. The values of the three are close and all relatively high, indicating that the model has an excellent fitting degree and good prediction ability; the standard deviation of 1.49, the coefficient of variation of 1.91%, and AdeqPrecision of 34.9565 all show a large signal-to-noise ratio and high model robustness. In summary, this model can effectively explain the change of the response value, can be used for process optimization and parameter prediction, and the result of no significant interaction effect indicates that the influence of each factor on the response is more inclined to independent or quadratic effects.

[0163] Table 9: Results of the analysis of variance of regression simulation

[0164]

[0165]

[0166] Table 10: Feasibility analysis of the model

[0167]

[0168] (2) Response surface analysis of the interaction among factors

[0169] Three-dimensional and two-dimensional response surface plots can intuitively reveal the degree of interaction among factors and the changing trends of various factors. As can be seen from the three-dimensional surface plot and contour plot in See Appendix Figure 11 , the influence of the inoculation order (A) on the comprehensive response value is greater than that of the fermentation temperature (B). The inoculation order (A) and the fermentation temperature (B) show an obvious single-peak distribution for the comprehensive response value, and a relatively high comprehensive response value can be obtained in a certain intermediate interval of A and B. The contour lines near the highest point of the surface are elliptical, indicating that the two factors mainly have main effects within this range. When the inoculation order or fermentation temperature deviates from this optimal interval, the comprehensive response value decreases significantly, indicating that a reasonable inoculation strategy combined with an appropriate fermentation temperature can maximize the comprehensive response value.

[0170] See the appendix Figure 12 As can be seen from the three-dimensional surface and contour map Figure 12 , the inoculation order (A) and fermentation time (C) can synergistically increase the response value within a certain region, and the contour lines near the highest point are elliptical. This indicates that when both A and C are within a medium range, the comprehensive response value reaches its peak; if either factor deviates from this interval, the response value will decrease significantly, suggesting that the combination of the two factors within this region is most conducive to the improvement of the comprehensive response value.

[0171] See the appendix Figure 13 , the fermentation temperature (B) and fermentation time (C) have a significant impact on the comprehensive response value. Under medium temperature conditions, moderately extending the fermentation time can cause the response value to gradually increase and reach a peak at a certain time point, and then decrease as the time is further extended. The contour lines in the figure are elliptically distributed, indicating that the temperature and time mainly show separate main effects, and the interaction is not prominent, but there is a certain synergistic effect between them. Appropriate temperature is beneficial to improving the microbial activity, while appropriate fermentation time helps the accumulation of metabolites, and the two together determine the fermentation effect. When either factor deviates from the optimal range, the response value will decrease significantly.

[0172] (3) Verification of response surface optimization

[0173] According to the regression model, the predicted optimal experimental conditions are: inoculation order 1.90, fermentation temperature 35.01 °C, fermentation time 49.99 hours, and the predicted comprehensive response value under this condition is 97.81. In actual operation, the inoculation order was adjusted to 2, the temperature was set at 35 °C, and the time was set at 50 hours, and a verification test was carried out. After 3 repeated measurements, the obtained comprehensive response value was 96.37 ± 1.56, and the error from the predicted value was less than 5%, which proved the accuracy of the model. Therefore, the optimal processing technology of the mixed-strain fermentation beverage determined by response surface optimization is reasonable and feasible.

[0174] 4. Determination of sterilization conditions

[0175] Table 11 shows the sterilization conditions of the mixed-strain fermentation highland barley beverage. At 72 °C, within 30 min of sterilization time, the Bacillus in the beverage still survived and could not be killed, which is consistent with the results of previous studies, indicating that conventional pasteurization cannot effectively sterilize the mixed-strain fermentation beverage. When the sterilization conditions reached 121 °C, 0.1 MPa for 3 min, all the live bacteria in the beverage could be killed, and the results were the same with the extension of the sterilization time, but other active ingredients in the beverage would be lost with the extension of the sterilization time. To retain as many effective and nutritional components as possible, the final sterilization conditions were selected as: 121 °C, 0.1 MPa for 3 min.

[0176] Table 11: Sterilization results of the mixed-strain fermentation highland barley beverage

[0177]

[0178] 5. Quality Analysis of Fermented Hulless Barley Beverage

[0179] Using hulless barley as raw material, through mixed bacteria fermentation, the processing technology of fermented hulless barley beverage was optimized by single factor and response surface experiments. The final optimal process conditions were as follows: the ratio of material to water was 1:8, the enzyme reaction time was 0.5 h, the inoculation ratio of Bacillus velezensis:Pichia cactophila:Leuconostoc lactis XP1 was 10:10:1, the inoculation amount was 7%, the fermentation temperature was 35 °C, the fermentation time was 50 h, and the sterilization conditions were 121 °C, 0.1 MPa for 3 min. Under these conditions, the fermented hulless barley beverage had a delicate and smooth taste, moderate sour and sweet taste, and had a cereal aroma, with a sensory score of 81.76; Escherichia coli was not detected; among them, the total dietary fiber was 1.72 ± 0.04%; the resistant starch content was 0.80 ± 0.01%; the protein content was 0.97 ± 0.03%.

[0180] In summary, this application optimized the process conditions of the fermented hulless barley beverage. The optimal process conditions for the pretreatment of the fermented hulless barley beverage were mixed fermentation of 3 strains of bacteria, the ratio of material to water was 1:8, and the enzyme reaction time was 0.5 h; the optimal process conditions for the fermented hulless barley beverage were synchronous fermentation of 3 strains of bacteria, the inoculation ratio of Bacillus velezensis:Pichia cactophila:Leuconostoc lactis XP1 was 10:10:1, the inoculation amount was 7%, the fermentation temperature was 35 °C, and the fermentation time was 50 h. Under these process conditions, the taste was delicate and smooth, the sour and sweet taste was moderate, and there was a cereal aroma. Among them, the total dietary fiber was 1.72%; the resistant starch content was 0.80%; the protein content was 0.97%, and Escherichia coli was not detected. Therefore, the technical solution provided by this application can provide a theoretical basis and technical support for the industrial production of fermented hulless barley beverage.

[0181] The above embodiments are only to illustrate the technical concept and characteristics of the present invention in a specific scenario. The purpose is to enable those who need this technology to understand the content of the present invention and implement it, and does not limit the protection scope of the present invention. Any equivalent changes or modifications made according to the spirit and essence of the present invention should be covered within the protection scope of the present invention.

Claims

1. A hulless barley fermented beverage, characterized in that, The described highland barley fermented beverage includes: highland barley, purified water, mixed bacterial agents, composite enzymes, and saccharifying enzyme raw materials; the mixed bacteria used are Leuconostoc lactis XP1, Pichia cactophila, and Bacillus velezensis; the composite enzymes used are α-amylase and saccharifying enzyme; the total dietary fiber in the highland barley fermented beverage is 1.72%; the resistant starch content is 0.80%; the protein content is 0.97%.

2. The hulless barley fermented beverage according to claim 1, characterized in that Among the mixed bacteria, the viable cell counts of all three strains are 10 8 CFU / mL, and the inoculation ratio of Bacillus velezensis:Pichia opuntiae:Leuconostoc lactis XP1 in the mixed bacteria is 10:10:

1.

3. The hulless barley fermented beverage according to claim 1, wherein The material-water ratio of the highland barley to the purified water is set to: 1:(8 - 16).

4. The preparation method of the hulless barley fermented beverage according to any one of claims 1 to 3, characterized in that, The described highland barley fermented beverage is prepared by the following steps: (1) The selected plump and round highland barley grains are rinsed 3 times with clear water and then drained, and stir-fried over low heat for 35 minutes until the highland barley emits a strong grain aroma; (2) The highland barley stir-fried in step (1) is cooled, crushed by a pulverizer, and then passed through a colloid mill with water for 15 minutes, and then boiled for 10 minutes at a certain material-water ratio; (3) The boiled highland barley obtained in step (2) is added with composite enzymes and enzymolyzed for a certain time at the optimal enzyme reaction temperature, cooled to room temperature, inoculated with composite bacteria, fermented at a suitable temperature and time, and then sterilized to obtain the mixed bacteria fermented highland barley beverage.

5. The preparation method of the hulless barley fermented beverage according to claim 4, wherein, The inoculation sequence of the composite bacteria is: inoculate Bacillus velezensis, Leuconostoc lactis XP1, and Pichia cactophila at the beginning of fermentation.

6. The preparation method of the hulless barley fermented beverage according to claim 4, wherein, The inoculation amount of the composite bacteria is 1% - 9%, the fermentation temperature is 28°C - 37°C, and the fermentation time is 24h - 72h.

7. The preparation method of the hulless barley fermented beverage according to claim 6, characterized in that, The inoculation amount of the composite bacteria is 7%, the fermentation temperature is 35°C, and the fermentation time is 50h.

8. The preparation method of the hulless barley fermented beverage according to claim 4, characterized in that, The sterilization procedure selects sterilization at 121°C and 0.1MPa for 3 minutes.

9. The preparation method of the hulless barley fermented beverage according to claim 4, wherein, In the enzymolysis step, the composite enzymes used are α-amylase and saccharifying enzyme, and the addition ratio is 1:2; the addition amount of the composite enzymes is 0.8%, and under the condition that the enzyme reaction temperature is 60°C, the enzyme reaction time is 0.5h.

10. Use of the hulless barley fermented beverage according to any one of claims 1-9 or the hulless barley fermented beverage obtained by the preparation method of the hulless barley fermented beverage in the preparation of functional foods, characterized in that, The described function is to control blood sugar through the resistant starch component.

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