Method for preparing rhizoma polygonati wine through combined fermentation of composite strains

Abstract

The invention discloses a method for preparing rhizoma polygonati wine through combined fermentation of composite strains. The composite strain leavening agent is firstly provided, the leavening agent is composed of lactobacillus plantarum, lactic acid-producing bacillus and saccharomyces cerevisiae according to the mass ratio of (2-4): (1-2): (50-100), the composite strain leavening agent can effectively balance and improve the original bitter taste of rhizoma polygonati medicinal materials through the synergistic effect of composite strains, and the comprehensive quality of rhizoma polygonati wine is remarkably improved; the fermentation process is accelerated, the fermentation period is remarkably shortened, the production efficiency is improved, and the production cost is reduced. The invention further provides a method for preparing polygonatum kingianum wine based on the composite strain leavening agent, and the method has remarkable effects in the aspects of improving the taste, improving the fermentation efficiency, enhancing the product stability, reducing adverse metabolites, improving the nutrition and health-care value and the like, and can meet the pursuit of consumers for high-quality polygonatum kingianum health-preserving drinks.

Description

Technical Field

[0001] The invention belongs to the technical field of fermented wine, and in particular relates to a method for preparing polygonatum wine by combined fermentation of composite strains. Background Art

[0002] Polygonatum sibiricum is the dried rhizome of Polygonatum kingianum Coll.et Hemsl., Polygonatum sibiricum Red. or Polygonatum cyrtonema Hua, a plant of the Liliaceae family. It is a Chinese medicinal material that can be used as both medicine and food. Polygonatum sibiricum contains many ingredients that are beneficial to the human body, such as polysaccharides, saponins, flavonoids, alkaloids, lignin, anthraquinone, and many other essential amino acids and trace elements for the human body. Polygonatum sibiricum has many functions such as nourishing the kidney and essence, nourishing yin and moistening dryness, delaying aging, resisting fatigue, resisting oxidation, enhancing immune function, lowering blood sugar and blood lipids, and has a significant therapeutic effect on diseases such as diabetes, coronary heart disease, and hypertension.

[0003] Based on the health-care and health-preserving effects of Polygonatum sibiricum, it is often used to make various health-care wines, namely "Polygonatum sibiricum wine". The production methods of Polygonatum sibiricum wine mainly include soaking method or extraction and preparation method of Polygonatum sibiricum extract. However, the Polygonatum sibiricum wine prepared by these two methods is often difficult to remove the bitter taste of the medicinal materials, resulting in relatively poor taste and flavor. In particular, when Polygonatum sibiricum is soaked or prepared with other medicinal materials to prepare composite Polygonatum sibiricum wine, the medicinal taste is stronger, most of them have poor taste, and the color is darker, and the visual pleasure is poor. In addition, the stability of Polygonatum sibiricum wine prepared by soaking or preparation method is also poor. Because the macromolecular components in Polygonatum sibiricum (such as polysaccharides, proteins, polypeptides, etc.) have low solubility in wine and poor stability, precipitation is easy to occur after being placed for a period of time, which greatly affects the sensory quality of Polygonatum sibiricum wine.

[0004] The traditional fermentation method can also be used to prepare Huangjing wine. This method can improve the taste and color of the wine to a certain extent, but it also has some shortcomings. Specifically: (1) Long fermentation cycle: Traditional fermentation requires a certain amount of time, and the temperature, humidity and other conditions during this period must be strictly controlled, otherwise it is easy to cause fermentation failure or produce odor. (2) Not rich enough taste: If only traditional fermentation is used for fermentation, the chemical components in the fermentation product may not be rich enough, thus affecting the layering of flavor. (3) Poor quality stability: The number and types of microorganisms in traditional fermentation are different, and it is impossible to ensure that the number and types of microorganisms in each batch of fermentation are exactly the same. At the same time, the metabolic activities of microorganisms during the fermentation process are complex and changeable, making it difficult to accurately control the fermentation process of traditional fermentation, which in turn affects the quality stability of the final Huangjing wine. (4) Possible generation of undesirable metabolites: Some undesirable metabolites, such as methanol, may be produced during the fermentation of traditional fermentation. This is because traditional fermentation is composed of a mixture of various microorganisms, including yeast, mold and other bacteria. Among them, molds, such as Aspergillus niger, can decompose pectin in the raw materials to produce methanol. Higher concentrations of adverse metabolites such as methanol may have negative effects on human health.

[0005] There are also methods on the market that use a single strain of bacteria, such as brewer's yeast, to ferment and prepare Huangjing wine. However, this method often produces Huangjing wine with a single taste, low fermentation efficiency, and the quality of the final product is easily affected by environmental changes.

[0006] In summary, the current Huangjing wine products and their preparation and brewing processes still have some defects, which seriously affect their sensory evaluation and overall quality. Therefore, improving the taste of Huangjing wine and improving its stability and safety are issues that still need to be solved in Huangjing wine fermentation. Summary of the invention

[0007] The purpose of the present invention is to overcome the above defects and shortcomings in the prior art and provide a composite bacterial fermentation agent.

[0008] The second object of the present invention is to provide a method for preparing polygonatum wine by combined fermentation of composite bacteria.

[0009] The above-mentioned object of the present invention is achieved through the following technical solutions:

[0010] The invention firstly provides a composite bacterial fermentation agent, which is composed of plant lactobacillus, lactic acid-producing bacillus and brewer's yeast in a mass ratio of 2-4:1-2:50-100.

[0011] The present invention innovatively uses three strains of plant lactobacillus, lactic acid-producing bacillus and brewer's yeast as composite strain fermentation agents, and prepares Huangjing wine by co-fermentation in a specific ratio (2-4:1-2:50-100). Through the synergistic effect of the composite strains, the multi-strain co-fermentation technology can effectively balance and improve the original bitter taste of Huangjing medicinal materials, and significantly improve the comprehensive quality of Huangjing wine. The Huangjing wine prepared by the composite strain fermentation agent has a refreshing taste, moderate sweetness and sourness, and a fragrant mellow aroma. At the same time, it preserves the original fragrance of Huangjing medicinal materials, has rich taste layers, and is mellow, sweet and sour, and has a long aftertaste. The wine is clear and transparent, and the color is light yellow. Its taste, aroma, and color have reached a high level. In contrast, under the same amount of fermentation agent, the Huangjing wine prepared by fermentation using traditional wine koji, one or two of the above strains is insufficient in taste and flavor. Furthermore, the synergistic effect of multiple strains can accelerate the fermentation process, significantly shorten the fermentation cycle, improve production efficiency, and thus reduce production costs. In particular, lactic acid-producing Bacillus has strong resistance to stress and can resist adverse external factors, further promoting the rapid progress of the fermentation process. Composite bacterial fermentation technology can optimize the microbial metabolic pathway, reduce the generation of adverse metabolites (such as methanol), improve the safety of Huangjing wine, and reduce potential risks to human health.

[0012] Furthermore, the mass ratio of the plant lactobacillus, the lactic acid-producing bacillus and the brewer's yeast is 2-3:1-1.5:50-75.

[0013] Furthermore, the viable count of the plant lactobacillus is 10 10 ~10 12 CFU / g, the viable count of lactic acid-producing Bacillus and Saccharomyces cerevisiae is 10 9 ~10 11 CFU / g.

[0014] Preferably, the viable count of the plant lactobacillus is 10 11 CFU / g, the number of viable lactic acid-producing Bacillus is 10 10 CFU / g, the number of viable bacteria of Saccharomyces cerevisiae is 10 10 CFU / g.

[0015] Furthermore, the plant lactobacillus is plant lactobacillus LP45, the lactic acid-producing bacillus is lactic acid-producing bacillus DU-106, and the brewer's yeast is Angel wine yeast RV171.

[0016] The present invention provides the use of any of the above-mentioned composite bacterial strain fermentation agents in the preparation of polygonatum odoratum wine.

[0017] The present invention provides a method for preparing polygonatum wine by combined fermentation of composite bacteria, comprising the following steps:

[0018] S1. Activating any of the above-mentioned composite bacterial fermentation agents to obtain a composite bacterial solution;

[0019] S2. saccharify the Polygonatum sibiricum tablets, mix them with distilled water and the composite bacterial solution, and ferment them at 25 to 35° C. for 5 to 10 days to obtain fermented wine;

[0020] S3. Remove impurities from the fermented wine, filter and sterilize it, and remove macromolecular compounds with a molecular weight greater than 10,000 to obtain Polygonatum odoratum wine.

[0021] The present invention realizes precise control of the fermentation process through scientific and reasonable strain ratio and fermentation conditions (fermentation time, temperature and filtration sterilization process), reduces the complexity and variability of microbial metabolic activities during the fermentation process, and significantly improves the quality stability of Huangjing wine, so that the product can maintain stable quality for a long time (120 days). At the same time, multiple filtration processes are used to remove impurities such as macromolecular polysaccharides and proteins in the wine body, ensure the clarity and stability of the wine body, and avoid secondary precipitation after long-term storage.

[0022] Furthermore, the activation in step S1 is to add the composite bacterial fermentation agent into a mixed solution of physiological saline and sugar water.

[0023] Preferably, the volume ratio of physiological saline to sugar water in the mixed solution is 1:1.

[0024] Preferably, the sugar water is 4-6 wt% sugar water.

[0025] Preferably, the sugar water is 5% wt % sugar water.

[0026] Furthermore, the activation condition in step S1 is 25-35° C. for 25-35 minutes.

[0027] Preferably, the activation condition in step S1 is activation at 30° C. for 30 minutes.

[0028] Furthermore, the polygonatum sibiricum slices in step S2 are used after being soaked, cleaned and boiled.

[0029] Furthermore, in step S2, the mass ratio of the polygonatum sibiricum tablets, sugar, distilled water and composite bacterial liquid is 1:10-20:35-70:1.

[0030] Preferably, the mass ratio of the polygonatum sibiricum tablets, sugar, distilled water and composite bacterial liquid in step S2 is 1:10-15:35-52.5:1.

[0031] Furthermore, the fermentation condition in step S2 is 25-35° C. and the fermentation time is 5-10 days.

[0032] Furthermore, the impurity removal method in step S3 is to use a coarse fiber filter bag to perform preliminary coarse filtration to remove large particle impurities; secondly, further refine the filtration effect through a plate and frame filter; and finally use a diatomaceous earth filter to deeply remove tiny suspended matter.

[0033] Furthermore, the filtration sterilization in step S3 is performed by using 0.4-0.5 μm and 0.1-0.2 μm microporous filter membranes in sequence.

[0034] Preferably, the filtration sterilization is performed by sequentially filtering using 0.45 μm and 0.15 μm microporous filter membranes.

[0035] Furthermore, the removal of compounds with a molecular weight greater than 10,000 in step S3 is performed by ultrafiltration using an ultrafiltration membrane with a molecular weight cutoff of 10,000.

[0036] Compared with the prior art, the present invention has the following beneficial effects:

[0037] (1) The composite bacterial fermentation agent of the present invention is composed of plant lactobacillus, lactic acid-producing bacillus and saccharomyces cerevisiae in a specific ratio. The composite bacterial fermentation agent can efficiently transform and fully utilize the complex metabolic components in polygonatum, and produce a series of small molecular flavor substances in the metabolic process through the interaction between bacterial species. These substances can greatly enrich the taste level and flavor characteristics of polygonatum wine. It can effectively balance and improve the original bitter taste of polygonatum medicinal materials, and significantly improve the comprehensive quality of polygonatum wine. The polygonatum wine prepared by the composite bacterial fermentation agent has a refreshing taste, moderate sweetness and sourness, and a fragrant mellow aroma. At the same time, it preserves the original fragrance of polygonatum medicinal materials, has rich taste levels, mellow sweetness and sourness, and a long aftertaste. The wine is clear and transparent, and the color is light yellow. Its taste, aroma, and color have reached a high level. Furthermore, the composite bacterial fermentation agent can also significantly shorten the fermentation cycle, effectively improve the stability of the product, optimize the microbial metabolic pathway, and reduce the generation of potential adverse metabolites such as methanol, thereby improving the safety of polygonatum wine and reducing the potential risks to human health.

[0038] (2) The method for preparing Huangjing wine by combined fermentation of composite strains of the present invention realizes precise control of the fermentation process through scientific and reasonable strain ratio and fermentation condition control, reduces the complexity and variability of microbial metabolic activities during the fermentation process, and significantly improves the quality stability of Huangjing wine, so that the product can maintain stable quality for a long time (120 days). At the same time, the plate and frame filter, diatomaceous earth filter, microfiltration, ultrafiltration and other equipment and technologies are used to remove impurities such as macromolecular polysaccharides and proteins in the wine, ensure the clarity, transparency and stability of the wine, and avoid secondary precipitation after long-term storage. BRIEF DESCRIPTION OF THE DRAWINGS

[0039] Figure 1These are the methanol detection results of Huangjing wine of Examples 1 to 3 and Comparative Example 1; wherein A is the sample of Example 1, B is the sample of Example 2, C is the sample of Example 3, and D is the sample of Comparative Example 1.

[0040] Figure 2 Sensory evaluation of Huangjing wine during storage ( n=10).

[0041] Figure 3 These are the results of measuring the alcohol content of Polygonatum odoratum wine during storage.

[0042] Figure 4 This is the determination result of total acid during the storage process of Huangjing wine.

[0043] Figure 5 This is the measurement result of the color difference value (ΔE) during the storage of Huangjing wine.

[0044] Figure 6 These are the results of measuring the transmittance of Polygonatum odoratum wine during storage. DETAILED DESCRIPTION

[0045] The present invention is further described below in conjunction with the accompanying drawings and specific examples, but the examples do not limit the present invention in any form. Unless otherwise specified, the reagents, methods and equipment used in the present invention are conventional reagents, methods and equipment in the art.

[0046] Unless otherwise specified, the reagents and materials used in the following examples are commercially available.

[0047] The traditional koji was purchased from “Liangshi-plant sweet koji” produced by Hubei Quzixiang Food Co., Ltd.

[0048] Lactobacillus plantarum LP45 (100 billion CFU / g) was purchased from Hebei Yiran Biotechnology Co., Ltd.; Saccharomyces cerevisiae was Angel wine yeast RV171 (10 billion CFU / g) purchased from Angel Yeast Co., Ltd.; Lactobacillus DU-106 (10 10 CFU / g) was provided by the School of Food Science and Engineering, South China Agricultural University.

[0049] Example 1

[0050] (1) Raw material processing:

[0051] Weigh 200g of Polygonatum sibiricum slices, soak and wash them in distilled water for several minutes to thoroughly remove impurities on the surface of Polygonatum sibiricum. Then, add distilled water again to cover the Polygonatum sibiricum, boil for 3 to 5 minutes, remove the Polygonatum sibiricum slices, and put them into a fermentation tank for later use.

[0052] (2) Saccharification:

[0053] Take 1000mL of distilled water and boil it, then add 2000g of white sugar and stir until the sugar is completely dissolved. Then, transfer the dissolved sugar water into the fermentation tank prepared earlier.

[0054] (3) Selection and activation of bacterial strains:

[0055] The strains are selected from plant lactobacillus, lactic acid-producing bacillus DU-106 and saccharomyces cerevisiae. Appropriate amounts of physiological saline and 5% sugar water of the same volume are prepared respectively, mixed, and sterilized at 121°C in an autoclave for 15 minutes. Then, 0.1g of plant lactobacillus, 0.05g of lactic acid-producing bacillus DU-106 and 2.5g of saccharomyces cerevisiae are added to each 100mL solution. Finally, activation treatment is carried out at a temperature of 30°C for 30 minutes to obtain an activated composite strain.

[0056] (4) Inoculation and fermentation:

[0057] Add 6000 mL of distilled water to the fermentation tank and wait for the solution to cool to room temperature. Then, add 200 g of the composite bacterial solution activated in step (3) (i.e., the amount of the composite bacterial solution added is equal to the amount of the Polygonatum sibiricum medicinal material added) to the fermentation tank and stir evenly. After that, seal the fermentation tank and keep the temperature at 25°C for 5 days of fermentation.

[0058] (5) Filtration and sterilization:

[0059] After fermentation, the wine is coarsely filtered through a coarse fiber filter bag to remove the polygonatum medicinal materials and larger impurities. Next, a plate and frame filter is used to remove larger suspended matter and impurities to reduce the burden for subsequent filtration. Then, a diatomaceous earth filter is used to further remove tiny particles and suspended matter in the wine, including some plant fibers, colloids, etc. After that, 0.45μm and 0.15μm microporous filter membranes are used for filtration in turn to achieve a sterile effect. Finally, the macromolecular compounds are removed by intercepting an ultrafiltration membrane with a molecular weight of 10,000 to prevent the precipitation of macromolecular compounds such as polygonatum polysaccharides and proteins and the appearance of flocculent precipitates, thereby ensuring the clarity and stability of the wine.

[0060] (6) Filling:

[0061] The filtered Polygonatum fermented wine is transferred into bottles for filling.

[0062] Example 2

[0063] (1) Raw material processing:

[0064] Weigh 200g of Polygonatum sibiricum slices, soak and wash them in distilled water for several minutes to thoroughly remove impurities on the surface of Polygonatum sibiricum. Then, add distilled water again to cover the Polygonatum sibiricum, boil for 3 to 5 minutes, remove the Polygonatum sibiricum slices, and put them into a fermentation tank for later use.

[0065] (2) Saccharification:

[0066] Take 1500mL of distilled water and boil it, then add 3000g of white sugar and stir until the sugar is completely dissolved. Then, transfer the dissolved sugar water into the fermentation tank prepared earlier.

[0067] (3) Selection and activation of bacterial strains:

[0068] The strains are selected from plant lactobacillus, lactic acid-producing bacillus DU-106 and saccharomyces cerevisiae. Appropriate amounts of physiological saline and 5% sugar water of the same volume are prepared respectively, and sterilized at 121°C in an autoclave for 15 minutes. Then, 0.15g of plant lactobacillus, 0.075g of lactic acid-producing bacillus DU-106 and 3.75g of saccharomyces cerevisiae are added to each 100mL solution. Finally, activation treatment is carried out at a temperature of 30°C for 30 minutes to obtain an activated composite strain.

[0069] (4) Inoculation and fermentation:

[0070] Add 9000 mL of distilled water to the fermentation tank and wait for the solution to cool to room temperature. Then, add 200 g of the composite bacterial solution activated in step (3) (i.e., the amount of the composite bacterial solution added is equal to the amount of the Polygonatum sibiricum medicinal material added) to the fermentation tank and stir evenly. After that, seal the fermentation tank and keep the temperature at 30°C for 7 days of fermentation.

[0071] (5) Filtration and sterilization:

[0072] After fermentation, the wine is coarsely filtered through a coarse fiber filter bag to remove the polygonatum medicinal materials and larger impurities. Next, a plate and frame filter is used to remove larger suspended matter and impurities to reduce the burden for subsequent filtration. Then, a diatomaceous earth filter is used to further remove tiny particles and suspended matter in the wine, including some plant fibers, colloids, etc. After that, 0.45μm and 0.15μm microporous filter membranes are used for filtration in turn to achieve a sterile effect. Finally, the macromolecular compounds are removed by intercepting an ultrafiltration membrane with a molecular weight of 10,000 to prevent the precipitation of macromolecular compounds such as polygonatum polysaccharides and proteins and the appearance of flocculent precipitates, thereby ensuring the clarity and stability of the wine.

[0073] (6) Filling:

[0074] The filtered Polygonatum fermented wine is transferred into bottles for filling.

[0075] Example 3

[0076] (1) Raw material processing:

[0077] Weigh 200g of Polygonatum sibiricum slices, soak and wash them in distilled water for several minutes to thoroughly remove impurities on the surface of Polygonatum sibiricum. Then, add distilled water again to cover the Polygonatum sibiricum, boil for 3 to 5 minutes, remove the Polygonatum sibiricum slices, and put them into a fermentation tank for later use.

[0078] (2) Saccharification:

[0079] Take 2000mL of distilled water and boil it, then add 4000g of white sugar and stir until the sugar is completely dissolved. Then, transfer the dissolved sugar water into the fermentation tank prepared earlier.

[0080] (3) Selection and activation of bacterial strains:

[0081] The strains used are plant lactobacillus, lactic acid-producing bacillus DU-106 and saccharomyces cerevisiae. Appropriate amounts of physiological saline and 5% sugar water of the same volume are prepared respectively, and sterilized at 121°C in an autoclave for 15 minutes. Then, 0.2g of plant lactobacillus, 0.1g of lactic acid-producing bacillus DU-106 and 5.0g of saccharomyces cerevisiae are added to each 100mL solution. Finally, activation treatment is carried out at a temperature of 30°C for 30 minutes to obtain an activated composite strain.

[0082] (4) Inoculation and fermentation:

[0083] Add 12000mL of distilled water to the fermentation tank and wait for the solution to cool to room temperature. Then, add 200g of the composite bacterial solution activated in step (3) (i.e., the amount of the composite bacterial solution added is equal to the amount of the Polygonatum sibiricum medicinal material added) to the fermentation tank and stir evenly. After that, seal the fermentation tank and keep the temperature at 35°C for 10 days of fermentation.

[0084] (5) Filtration and sterilization:

[0085] After fermentation, the wine is coarsely filtered through a coarse fiber filter bag to remove the polygonatum medicinal materials and larger impurities. Next, a plate and frame filter is used to remove larger suspended matter and impurities to reduce the burden for subsequent filtration. Then, a diatomaceous earth filter is used to further remove tiny particles and suspended matter in the wine, including some plant fibers, colloids, etc. After that, 0.45μm and 0.15μm microporous filter membranes are used for filtration in turn to achieve a sterile effect. Finally, the macromolecular compounds are removed by intercepting an ultrafiltration membrane with a molecular weight of 10,000 to prevent the precipitation of macromolecular compounds such as polygonatum polysaccharides and proteins and the appearance of flocculent precipitates, thereby ensuring the clarity and stability of the wine.

[0086] (6) Filling:

[0087] The filtered Polygonatum fermented wine is transferred into bottles for filling.

[0088] Comparative Example 1

[0089] This comparative example does not use a specific fermentation strain, but uses traditional distiller's yeast for natural fermentation. Specifically, in the selection and activation of strains, the plant lactobacillus, lactic acid-producing Bacillus DU-106 and saccharomyces cerevisiae used in Example 1 are replaced with traditional distiller's yeast of equal quality for fermentation. Except for the above strain replacement, all other fermentation steps and production conditions are the same as those in Example 1.

[0090] Comparative Example 2

[0091] This comparative example adopts single strain fermentation, that is, only plant lactobacillus is used for fermentation. In the selection and activation of strains, the three strains mixed in proportion in Example 1 are replaced with plant yeast of equal mass for fermentation. In addition, all fermentation steps and production conditions are the same as those in Example 1.

[0092] Comparative Example 3

[0093] This comparative example adopts a single strain fermentation, that is, only lactic acid-producing Bacillus DU-106 is used for fermentation. In the selection and activation of strains, the three strains mixed in proportion in Example 1 are replaced with lactic acid-producing Bacillus DU-106 of equal mass for fermentation. In addition, all fermentation steps and production conditions are the same as those in Example 1.

[0094] Comparative Example 4

[0095] This comparative example adopts a single strain fermentation, that is, only Saccharomyces cerevisiae is used for fermentation. In the selection and activation of strains, the three strains mixed in proportion in Example 1 are replaced with Saccharomyces cerevisiae of equal mass for fermentation. In addition, all fermentation steps and production conditions are the same as those in Example 1.

[0096] Comparative Example 5

[0097] This comparative example adopts double strain fermentation, and the specific strains selected are Lactobacillus plantarum and lactic acid-producing Bacillus DU-106. In the selection and activation stage of strains, the three strains mixed in proportion in Example 1 are replaced with equal masses of Lactobacillus plantarum and lactic acid-producing Bacillus DU-106, and the ratio is kept the same as in Example 1. Except for the strain adjustment, all fermentation steps and production conditions are the same as in Example 1.

[0098] Comparative Example 6

[0099] This comparative example adopts dual-strain fermentation, and the specific strains selected are lactic acid-producing Bacillus DU-106 and saccharomyces cerevisiae. In the strain selection and activation stage, the three strains mixed in proportion in Example 1 are replaced with lactic acid-producing Bacillus DU-106 and saccharomyces cerevisiae of equal mass, and the proportion is kept the same as in Example 1. Except for the strain adjustment, all fermentation steps and production conditions are the same as in Example 1.

[0100] Comparative Example 7

[0101] This comparative example adopts double strain fermentation, and the strains specifically selected are plant lactobacillus and saccharomyces cerevisiae. In the selection and activation stage of strains, the three strains mixed in proportion in Example 1 are replaced with plant lactobacillus and saccharomyces cerevisiae of equal mass, and the ratio is kept the same as in Example 1. Except strain adjustment, all fermentation steps and production conditions are the same as in Example 1.

[0102] Comparative Example 8

[0103] This comparative example uses three strains for fermentation, but unlike Example 1, the mixing ratio of these three strains (Lactobacillus plantarum, lactic acid-producing Bacillus DU-106 and Saccharomyces cerevisiae) is 1:1:1, and the total mass remains the same as that of Example 1. In the selection and activation stage of strains, the strains are mixed according to a ratio of 1:1:1. In addition, all fermentation steps and production conditions are the same as those of Example 1.

[0104] Comparative Example 9

[0105] This comparative example uses three strains for fermentation, but unlike Example 1, the mixing ratio of these three strains (Lactobacillus plantarum, lactic acid-producing Bacillus DU-106 and Saccharomyces cerevisiae) is 1:10:50, and the total mass remains the same as Example 1. In the selection and activation stage of strains, the strains are mixed according to the ratio of 1:10:50. In addition, all fermentation steps and production conditions are the same as Example 1.

[0106] Comparative Example 10

[0107] This comparative example uses three strains for fermentation, but unlike Example 1, the mixing ratio of these three strains (Lactobacillus plantarum, lactic acid-producing Bacillus DU-106 and Saccharomyces cerevisiae) is 50:10:1, and the total mass remains the same as Example 1. In the selection and activation stage of strains, the strains are mixed according to a ratio of 50:10:1. In addition, all fermentation steps and production conditions are the same as Example 1.

[0108] Comparative Example 11

[0109] The filtration sterilization method of this comparative example is different from that of Example 1. In the filtration step, the wine after fermentation is coarsely filtered through a coarse fiber filter bag, and then a plate and frame filter is used to remove larger suspended matter and impurities, and then a diatomaceous earth filter is used to further remove tiny particles and suspended matter in the wine, including some plant fibers, colloids, etc. However, in this comparative example, microporous filter membranes and ultrafiltration membranes are no longer used for deep filtration. In addition, the remaining steps and production conditions are the same as those of Example 1.

[0110] Comparative Example 12

[0111] The filtration sterilization method of this comparative example is different from that of Example 1. In the filtration step, the wine after fermentation is coarsely filtered through a coarse fiber filter bag, and then a plate and frame filter is used to remove larger suspended matter and impurities, and then a diatomaceous earth filter is used to further remove tiny particles and suspended matter in the wine, including some plant fibers, colloids, etc. Then, it is filtered with 0.45μm and 0.15μm microporous membranes in turn. However, in this comparative example, an ultrafiltration membrane is no longer used for deep filtration. In addition, the remaining steps and production conditions are the same as those of Example 1.

[0112] Test Example 1

[0113] Sensory Evaluation of Polygonatum odoratum Wine

[0114] The Huangjing wines of Examples 1 to 3 and Comparative Examples 1 to 12 were subjected to sensory evaluation from the four aspects of aroma, flavor, color, and clarity. The specific sensory evaluation standards are shown in Table 1. The evaluation scores are shown in Table 2.

[0115] 1. Purpose of the experiment

[0116] Through systematic sensory evaluation, the Huangjing wines of Examples 1 to 3 and Comparative Examples 1 to 12 were systematically evaluated from four aspects: aroma, taste, color, and clarity, so as to objectively compare the qualities of different Huangjing wines.

[0117] 2. Experimental materials and personnel

[0118] Experimental materials: the Huangjing wine samples of Examples 1 to 3 and Comparative Examples 1 to 12, a clean and odorless tasting cup, a light source for clarity observation and a transparent glass cup.

[0119] Personnel: 10 trained sensory evaluators with keen sense of smell, taste and visual identification, familiar with the process and standards of wine sensory evaluation.

[0120] 3. Experimental Procedure

[0121] 3.1 Preparation stage:

[0122] Ensure that the evaluation environment is clean and tidy, free of odor interference, and that the temperature and humidity are controlled within an appropriate range (room temperature 20-25°C, humidity 50%-60%).

[0123] All samples of Polygonatum odoratum wine were numbered and randomly placed to ensure that the evaluators were unaware of the source of the samples.

[0124] Prepare a tasting cup, a light source and a clear glass.

[0125] The evaluators first tasted a glass of pure water to clear any residual taste from their mouths.

[0126] Pour the Huangjing wine samples into tasting cups and transparent glass cups respectively to ensure that the amount of sample in each cup is consistent.

[0127] 3.2 Sensory evaluation process:

[0128] Aroma evaluation: The evaluator sniffs the sample and records its aroma characteristics (such as intensity, purity, and whether there is any peculiar smell). Then, the evaluator sniffs the sample again after a certain interval (30 seconds) to evaluate the aroma persistence and gives a score according to the standards in Table 1.

[0129] Taste evaluation: The evaluator takes a small sip of the sample, paying attention to the balance and coordination of the sweetness, sourness, bitterness, astringency and other flavors, as well as whether the aftertaste is long, and scores according to the standards in Table 1.

[0130] Color evaluation: Under a uniform light source, observe the color of each Huangjing wine sample, record its depth, transparency and uniformity, and score according to the standards in Table 1.

[0131] Clarity evaluation: Place the sample in a transparent glass cup, observe its clarity, the presence of suspended matter or sediment, and score according to the standards in Table 1.

[0132] 3.3 Notes:

[0133] Before the evaluation, the evaluator should avoid eating irritating foods and keep the oral cavity clean.

[0134] During the evaluation process, you should rinse your mouth with clean water after tasting each sample to avoid residual taste affecting subsequent evaluation.

[0135] Evaluators should evaluate independently to avoid mutual interference.

[0136] For each evaluation indicator, each evaluator needs to repeat it three times to improve the accuracy of the results.

[0137] 4. Data Recording and Analysis

[0138] Data recording: Collect scores from 10 evaluators on each sample in terms of aroma, taste, color and clarity.

[0139] Result analysis: Calculate the sum of the scores of each sample in each evaluation index as the final comprehensive score of the sample. The higher the comprehensive score, the better the overall quality of the Huangjing wine.

[0140] Table 1 Evaluation criteria for sensory indexes of Huangjing wine

[0141]

[0142]

[0143] Table 2 Sensory evaluation results of Huangjing wine of Examples 1 to 3 and Comparative Examples 1 to 12 ( n=10)

[0144]

[0145]

[0146] Comparative Examples 1 to 12 were compared with Example 1 for sensory evaluation. The sensory evaluation results are as follows:

[0147] Compared with Example 1, the quality of the Huangjing wine prepared in Comparative Example 1 is significantly different from that of the Huangjing wine prepared in Example 1. First, the Huangjing wine prepared in Comparative Example 1 has a distinct bitter taste of the Huangjing medicinal material, and the fermentation cycle needs to be significantly extended. Specifically, the alcohol content of the Huangjing wine prepared in Example 1 and Comparative Example 1 was measured, and the alcohol content of the Huangjing wine obtained in Comparative Example 1 was still relatively low after fermentation for 5 days, and it needed to be extended to about 15 days to reach an alcohol content level similar to that of Example 1 (about 8.7°), indicating that the synergistic effect of multiple strains of the present invention can accelerate the fermentation process and significantly shorten the fermentation cycle. In addition, due to the complexity and uncertainty of the natural fermentation environment, competition and interaction between multiple microorganisms may lead to the generation of adverse metabolites.

[0148] Compared with Example 1, the "Huangjing wine" prepared in Comparative Example 2 has no wine taste, only sour taste. This is because the main metabolite of Lactobacillus plantarum during the fermentation process is lactic acid rather than alcohol. It shows that using Lactobacillus plantarum alone is not suitable for fermenting and preparing Huangjing wine with a typical wine taste, and it is more likely to obtain a sour product similar to "Huangjing vinegar".

[0149] Compared with Example 1, the "Huangjing wine" prepared in Comparative Example 3 has no wine taste, only sour taste. This is because the main metabolite of lactic acid-producing Bacillus DU-106 is similar to Lactobacillus plantarum, and lactic acid rather than alcohol is produced during the fermentation process. This shows that the use of lactic acid-producing Bacillus DU-106 alone is not suitable for fermenting and preparing Huangjing wine with a typical wine taste, and it is more likely to obtain a sour product similar to "Huangjing vinegar".

[0150] Compared with Example 1, the Huangjing wine prepared in Comparative Example 4 is deficient in both taste and aroma. Specifically, due to the use of a single strain of bacteria for fermentation, the Huangjing wine is relatively monotonous in taste, lacking the sour, sweet and smooth balance in Example 1, and the bitter taste of the Huangjing is more prominent, affecting the overall drinking experience. This shows that in the fermentation process of Huangjing wine, the synergistic effect of multiple strains is crucial to improving the taste and aroma of the product.

[0151] Compared with Example 1, the "Huangjing Wine" prepared in Comparative Example 5 shows significant differences in flavor. Specifically, the product lacks the aroma of traditional Huangjing wine, and is replaced by a strong sour taste, and the sour taste is more prominent. This phenomenon can be attributed to the fact that the main metabolites of Lactobacillus plantarum and lactic acid-producing Bacillus DU-106 during the fermentation process are lactic acid, not alcohol. Therefore, it is difficult to obtain Huangjing wine with a typical wine flavor by fermenting only these two strains, and it is more likely to obtain a drink with a stronger sour taste.

[0152] Compared with Example 1, the Huangjing wine prepared in Comparative Example 6 showed certain differences in flavor. Specifically, the Huangjing wine had a lighter sour taste and a higher sweetness. At the same time, its flavor complexity and layering were insufficient, and the rich and coordinated taste of the Huangjing wine in Example 1 could not be fully demonstrated.

[0153] Compared with Example 1, the Huangjing wine prepared in Comparative Example 7 has some changes in flavor. Specifically, the Huangjing wine has a lighter taste, a lighter sour taste, and a higher sweetness. However, this flavor adjustment also leads to a lack of flavor complexity and layering of the Huangjing wine, and fails to fully show the rich and coordinated taste of the Huangjing wine in Example 1.

[0154] Compared with Example 1, the "Huangjing wine" prepared in Comparative Example 8 has a significant change in flavor. The product exhibits sharp acidity, high sweetness, but extremely low alcohol content, resulting in a taste that is more sour and refreshing rather than the mellow aroma of traditional Huangjing wine. Overall, its flavor characteristics are closer to a fermented vinegar drink with a distinct sour and refreshing taste rather than the traditional "Huangjing wine".

[0155] Compared with Example 1, the Huangjing wine prepared in Comparative Example 9 tastes sour and not sweet enough, which reflects the possible incoordination problem in the adjustment of the strain ratio, resulting in the failure to achieve an ideal balance between acidity and sweetness.

[0156] Compared with Example 1, the "Huangjing wine" prepared in Comparative Example 10 has a significant imbalance in flavor. The product has a distinct sour taste, which almost covers the mellow flavor of the wine, making the overall taste tend to be extreme, and failing to achieve the flavor balance that Huangjing wine should have.

[0157] Compared with Example 1, the taste, aroma and color of the "Huangjing Wine" prepared in Comparative Example 11 are highly consistent with those of Example 1. However, in terms of the clarity of the wine, the "Huangjing Wine" of Comparative Example 11 is not transparent enough and is slightly turbid. After being placed at room temperature for a few days, the wine will have obvious flocculent precipitation. In contrast, the Huangjing Wine prepared in Example 1 has no obvious precipitation after being placed at room temperature for three months. This shows that the filtration sterilization method, especially the microfiltration and ultrafiltration steps, will affect the clarity and stability of Huangjing Wine.

[0158] Compared with Example 1, the taste, aroma and color of the "Huangjing Wine" prepared in Comparative Example 12 are highly consistent with those in Example 1. When it is just prepared, the clarity of the wine is not significantly different from that in Example 1. However, after being placed at room temperature for 3 months, a small amount of flocculent precipitation will appear in the Huangjing Wine of Comparative Example 12, while in contrast, the Huangjing Wine prepared in Example 1 remains clear without obvious precipitation. This phenomenon shows that the filtration and sterilization method, especially the ultrafiltration step, has an important influence on the long-term clarity and stability of Huangjing Wine.

[0159] Through systematic sensory evaluation analysis, it can be clearly observed from Table 2 that the Huangjing wine of Examples 1 to 3 exhibits excellent performance in many aspects such as aroma, taste, color and clarity, and its average comprehensive score is significantly higher than that of Comparative Examples 1 to 12, and this difference is statistically significant, which fully proves that the Huangjing wine of Examples 1 to 3 has better quality and better production process.

[0160] This significant advantage can be attributed to the innovative use of three strains of plant lactobacillus, lactic acid-producing Bacillus DU-106 and saccharomyces cerevisiae in the production process of Huangjing wine in Examples 1 to 3, which were co-fermented in a specific ratio (2-4:1-2:50-100). In contrast, the Huangjing wines in Comparative Examples 1 to 10 have obvious differences in the use of strains or strain ratios, some of which do not use the above strain combination (such as Comparative Example 1), some only use some strains (such as Comparative Examples 2 to 7), and some use three strains but in an improper ratio (such as Comparative Examples 8 to 10).

[0161] The flavor and taste of the Huangjing wine of Examples 1 to 3 are richer and more coordinated than that of the Huangjing wine prepared in the comparative example, and the flavor and taste are the embodiment of the overall chemical composition of the wine, which also illustrates that the chemical composition of the Huangjing wine prepared in Examples 1 to 3 is richer and more coordinated, and the synergistic effect of the three strains of Lactobacillus plantarum, lactic acid-producing Bacillus DU-106 and Saccharomyces cerevisiae promotes the efficient transformation and utilization of complex metabolites in Huangjing. Therefore, it is clear that the precise ratio and synergistic fermentation mechanism of Lactobacillus plantarum, lactic acid-producing Bacillus DU-106 and Saccharomyces cerevisiae play a vital role in improving the overall quality of Huangjing wine. The absence or imbalance of any strain component will have an adverse effect on the fermentation process, thereby weakening the quality of the final product.

[0162] In addition, Examples 1 to 3 are also carefully designed in the filtration and sterilization process, using a multi-level filtration method of "coarse fiber filter bag coarse filtration - plate and frame filter filtration - diatomaceous earth filter filtration - microporous membrane filtration - ultrafiltration membrane filtration with a molecular weight cutoff of 10,000" to ensure the clarity of the wine. Even after being left at room temperature for 3 months, no precipitation was observed. Comparative Examples 11 and 12 are simplified in the filtration method. Comparative Example 11 omits the microporous membrane filtration and ultrafiltration membrane filtration steps, while Comparative Example 12 omits the ultrafiltration membrane filtration step, which results in their sensory evaluation scores for clarity being significantly lower than those of Examples 1 to 3. This comparison fully illustrates the key role of microporous membrane filtration and ultrafiltration membrane filtration with a molecular weight cutoff of 10,000 in improving the clarity of Huangjing wine.

[0163] In summary, Examples 1 to 3 significantly improve the quality of Polygonatum odoratum wine through precise strain ratio, synergistic fermentation mechanism and multi-level filtration and sterilization process.

[0164] Test Example 2

[0165] Methanol determination of Examples 1 to 3 and Comparative Example 1.

[0166] The main component of wine is ethanol. However, due to the simplification of brewing methods, the low quality of raw materials, the lack of brewing technology or the illegal addition of industrial methanol and other improper behaviors, the methanol content in some wines may be abnormally high. Methanol will be converted into formaldehyde and formic acid during metabolism in the body. The toxicity of these two substances is far greater than that of methanol itself. Excessive intake may quickly cause retinal damage, and in severe cases even cause blindness or death.

[0167] The formation of methanol is affected by many factors, including the brewing process, the types of raw materials used, and fermentation temperature. Specifically, the selection of inferior alcohol and raw materials, as well as failure to follow standardized production processes, are important reasons for the increase in methanol content. If the alcohol fermentation process is insufficient or the brewing technology is improperly used, the methanol content may exceed the safety standard. Controlling the conditions in the brewing process, such as temperature and fermentation conditions, and using high-quality raw materials are the key to reducing methanol production.

[0168] 1. Experimental purpose:

[0169] Evaluate whether the composite strain fermentation technology adopted in the present invention can optimize the metabolic pathway of microorganisms, effectively reduce the production of adverse metabolites (methanol), and thus improve the safety of Polygonatum odoratum wine.

[0170] 2. Detection method:

[0171] A methanol detection kit was used. The kit was produced by Shenyang Kaicheng Technology Co., Ltd.

[0172] 3. Detection principle:

[0173] During the test, the methanol in the wine sample is oxidized to formaldehyde under the action of a specific oxidant and a decolorizing agent. Subsequently, the formaldehyde reacts chemically with the developer in the kit to generate a purple quinone-type pigment. The color depth of this reaction product is proportional to the methanol content in the wine sample, that is, the darker the color, the higher the methanol content. By comparing the color depth with the standard color scale, the methanol content in the wine sample can be roughly determined, and based on this, it can be judged whether it exceeds the national safety standard value.

[0174] 4. Detection steps:

[0175] (1) Take 100 mL of the Huangjing wine sample to be tested and place it in the distillation bottle of the rotary evaporator. Set the water bath temperature to 80°C and the cooling circulation pump temperature to -20°C. Do not start the vacuum pump during this process and keep the air pressure in the rotary evaporator at normal pressure. Continue distillation until no more liquid flows into the receiving bottle, and collect the resulting distillate. Afterwards, add an appropriate amount of distilled water to the collected distillate to make it 100 mL as the subsequent sample to be tested. This step is intended to distill out the alcohol substances in the Huangjing wine and remove as much as possible the pigments, sugars and other substances that may interfere.

[0176] (2) Use a pipette to take 0.1 ml of the sample to be tested and inject it into the test tube. Then, add distilled water to the test tube until the liquid level reaches the 1.0 ml mark (i.e., the 1.0 mark on the test tube), and immediately cover the lid and shake gently to mix thoroughly.

[0177] (3) Add 2 drops of test solution A (oxidant) to the mixed test tube. Then, shake the test tube for about 1 minute and let it stand for 10 minutes to allow for a complete reaction.

[0178] (4) After 10 minutes, add 1 drop of test solution B (decolorizer) to the test tube. After closing the lid again, shake it up and down several times to ensure that the solution is fully mixed. Then, wait for the solution to fade on its own. If the solution does not completely fade within 10 minutes, add another drop of decolorizer.

[0179] (5) After the solution has completely faded to light yellow or colorless, add 3 drops of test solution C (color developer) to the test tube. Cover the lid again and shake gently to mix. Then, let it stand for 30 minutes (this time point is critical, and the results must be observed at 30 minutes). Finally, observe the color change of the solution. If the solution is purple, it means that the sample contains a large amount of methanol; and the darker the color, the higher the methanol concentration. By comparing with the colorimetric card, the methanol content in the wine sample can be roughly determined.

[0180] 5. Evaluation criteria:

[0181] According to the relevant provisions of GB 2757-2012 "National Food Safety Standard Distilled Spirits and Blended Spirits", the methanol content of spirits brewed with non-grain raw materials shall be ≤2.0g / L.

[0182] 6. Experimental Results

[0183] The methanol detection results of Examples 1 to 3 and Comparative Example 1 are as follows: Figure 1 shown. Figure 1 The color of the final test sample solution of Huangjing wine in Examples 1 to 3 is intuitively presented, which is significantly lighter than the color of the final test sample solution in Comparative Example 1. Through accurate comparison with the colorimetric card supporting the methanol detection tube, it can be confirmed that the methanol content of Huangjing wine in Examples 1 to 3 is lower than 0.1g / L, while the methanol content of Huangjing wine in Comparative Example 1 falls within the range of 0.1-0.2g / L. Although it meets the limit standard of methanol content in fermented wine, its methanol content is significantly higher than that of the Huangjing wine prepared in Example 1. This result shows that the composite strain fermentation technology of the present invention can inhibit the generation of undesirable metabolite methanol by finely regulating the metabolic pathways of microorganisms.

[0184] It is worth noting that the methanol content of Huangjing wine in Examples 1 to 3 is far lower than the limit specified in the national food safety standard GB2757-2012 (i.e., no more than 2.0 g / L). This result proves that Huangjing wine brewed using the technology of the present invention not only meets national safety standards in terms of methanol control, but also shows extremely high safety, providing consumers with a healthier choice.

[0185] In summary, the composite strain fermentation technology involved in the present invention has significant effects in reducing the methanol content in Polygonatum odoratum wine, and provides a solid technical foundation and guarantee for improving the overall quality and safety of Polygonatum odoratum wine.

[0186] Test Example 3

[0187] Stability evaluation of Huangjing wine during storage:

[0188] The Huangjing wine of Examples 1 to 3 was stored at room temperature for 0 days, 10 days, 30 days, 60 days, 90 days and 120 days, respectively, and a comprehensive sensory evaluation as well as alcohol content, total acidity, color and clarity were performed to evaluate the stability of the Huangjing wine during storage.

[0189] 1. Sensory evaluation of Huangjing wine during storage

[0190] The method, steps and evaluation criteria for the sensory evaluation of Huangjing wine all followed the method and steps in Test Example 1. The evaluation results are shown in Figure 2 The sensory evaluation score can comprehensively and intuitively reflect the changes of Huangjing wine during the storage process. Figure 2 The results clearly show the changes in the sensory scores of Huangjing wine in Examples 1 to 3 at different storage days. The results show that as the storage days gradually increase, the sensory scores of Huangjing wine do not fluctuate significantly, which indicates that the flavor and quality of Huangjing wine remain relatively stable during storage for up to 120 days at room temperature.

[0191] 2. Determination of Alcohol Content of Huangjing Wine during Storage

[0192] According to the alcohol content determination method in "GB5009.225-2023 National Food Safety Standard Determination of Ethanol Concentration in Wine and Edible Alcohol", the alcohol meter method was used to determine the alcohol content of the Huangjing wine of Examples 1 to 3.

[0193] principle:

[0194] The non-volatile substances in the sample are removed by distillation, and the alcohol volume fraction is measured with an alcohol meter. The temperature correction is performed by consulting the conversion table between the alcohol meter temperature and the ethanol concentration (alcohol content) at 20°C to obtain the ethanol concentration (alcohol content) of the sample at 20°C.

[0195] Instruments and equipment:

[0196] (1) Precision alcohol meter: the graduation value is 0.1% vol.

[0197] (2) All-glass distiller: 500mL, 1000mL.

[0198] (3) Measuring cylinder: 100mL, 200mL, 500mL, 1000mL.

[0199] (4) Thermometer: Graduation value is 0.1℃.

[0200] Analysis steps:

[0201] (1) Sample processing

[0202] After the clean, dry 100mL volumetric flask and sample are kept at a constant temperature of 20℃, accurately measure 100mL of sample into a 500mL distillation flask, rinse the volumetric flask with 50mL of water three times, add the washing liquid into the 500mL distillation flask, add a few zeolites, connect the serpentine condenser, use the original volumetric flask used for sampling as a receiver (with an ice bath), turn on the cooling water (cooling water temperature is lower than 15℃), slowly heat and distill, and collect the distillate. When it is close to the scale, remove the volumetric flask, cover the stopper, keep it warm in a 20℃ water bath for 30 minutes, then add water (20℃) to the scale, mix well, and set aside.

[0203] (2) Determination of sample solution

[0204] Pour the sample solution into a clean, dry measuring cylinder and let it stand for several minutes. After the bubbles disappear during the test, put in a clean, dry alcohol meter and press it gently. It should not touch the wall of the measuring cylinder. At the same time, insert the thermometer and balance for about 5 minutes. Observe horizontally, read the scale indication at the point where it is tangent to the meniscus, and record the temperature.

[0205] (3) Analysis results presentation

[0206] According to the measured alcohol meter indication and temperature, check Appendix B of "GB5009.225-2023 National Food Safety Standard Determination of Ethanol Concentration in Wine and Edible Alcohol" and convert it to the alcohol content of the sample at 20°C, expressed as volume fraction "%vol". Expressed as the arithmetic mean of two independent measurement results obtained under repeatability conditions, the result is retained to 1 decimal place. The absolute difference between two independent measurement results obtained under repeatability conditions shall not exceed 0.5%vol. Results are shown in Figure 3 .

[0207] Alcohol content is one of the core indicators for evaluating wine quality, and its stability is particularly important during the storage process. If the alcohol content can remain constant during storage, it is not only an important sign of the wine's maturity and quality, but also a key manifestation of the wine's excellent storage potential.

[0208] Figure 3The figure shows the change of alcohol content of Huangjing wine in Examples 1 to 3 after storage for different days at room temperature. It can be clearly seen from the figure that the alcohol content of Huangjing wine does not fluctuate significantly with the extension of storage time, which fully demonstrates that the alcohol content of Huangjing wine in Examples 1 to 3 remains highly stable and remains at a relatively constant level during the 120 days of storage at room temperature.

[0209] 3. Determination of total acid during storage of Huangjing wine

[0210] The test is carried out according to the acid-base indicator titration method in "GB / T1GB12456-2021 National Food Safety Standard Determination of Total Acid in Food".

[0211] principle:

[0212] According to the acid-base neutralization principle, the acid in the test solution is titrated with alkali solution, and phenolphthalein is used as an indicator to determine the titration endpoint. The total acid content in the food is calculated based on the consumption of alkali solution.

[0213] Reagents and preparation:

[0214] Phenolphthalein indicator solution (10 g / L): Weigh 1 g of phenolphthalein, dissolve it in ethanol (95%), and dilute it to 100 mL with ethanol (95%).

[0215] Sodium hydroxide standard titration solution (0.1 mol / L): prepared and calibrated in accordance with the requirements of GB / T 5009.1.

[0216] Analysis steps:

[0217] Use a pipette to draw 25.0 mL of Huangjing wine sample into a 250 mL volumetric flask, dilute to the mark with carbon dioxide-free water, and shake well. Filter with fast filter paper and collect the filtrate as the test solution for determination.

[0218] Use a pipette to draw 25mL of test solution, place it in a 250mL conical flask, add 2 to 4 drops (10g / L) of phenolphthalein indicator solution, and titrate with 0.1mol / L sodium hydroxide standard titration solution until it turns slightly red and does not fade for 30s. Record the volume value of 0.1mol / L sodium hydroxide standard titration solution consumed. At the same time, use the same volume of carbon dioxide-free water instead of the test solution to perform a blank test, and record the volume value of the sodium hydroxide standard titration solution consumed.

[0219] Calculation results:

[0220] The total acid content in the sample is calculated according to formula (1).

[0221]

[0222] Where:

[0223] X——the total acid content in the sample, in grams per liter (g / L);

[0224] c——the concentration of the standard sodium hydroxide titration solution, in moles per liter (mol / L);

[0225] V1——The volume of sodium hydroxide standard titration solution consumed when titrating the test solution, in milliliters (mL);

[0226] V2——The volume of sodium hydroxide standard titration solution consumed in the blank test, in milliliters (mL);

[0227] K——conversion coefficient of acid. Huangjing wine mainly contains lactic acid, and the conversion coefficient of lactic acid is 0.090;

[0228] F——the dilution factor of the test solution, which is 10 in this formula;

[0229] m——the volume of the sample sucked, in milliliters (mL);

[0230] 1000——Conversion factor.

[0231] The calculation results are expressed as the arithmetic mean of two independent determination results obtained under repeatability conditions. Figure 4 .

[0232] During the storage process of Huangjing wine, the stability of total acid content plays a key role in maintaining its unique variety characteristics and excellent quality. The flavor and taste of Huangjing wine, as a direct reflection of its core quality, are deeply affected by total acid, an important component of the wine. Total acid not only enriches the flavor level of the wine, but also profoundly shapes its unique taste experience. Therefore, during storage, the total acid content can be maintained stable, and the flavor and taste of Huangjing wine can be effectively preserved. In addition, an appropriate amount of total acid content can effectively inhibit the potential harmful microbial activities in Huangjing wine, thereby extending its shelf life and ensuring the safety and quality of the wine. On the contrary, if the total acid content is insufficient, it may provide a breeding ground for microorganisms, thereby posing a threat to the flavor, taste and even safety of Huangjing wine. Therefore, the stability of total acid is of great significance for the long-term preservation of Huangjing wine.

[0233] from Figure 4 It can be seen that during the storage of the Huangjing wine of Examples 1 to 3 for up to 120 days at room temperature, the total acid content did not fluctuate significantly, but was stably maintained at a relatively constant level.

[0234] 4. Color evaluation of Huangjing wine during storage

[0235] Based on the fact that the colorimeter can simulate the process of human eyes observing colors, the color stability of the Huangjing wine of Examples 1 to 3 during storage was analyzed by accurately measuring and quantifying the subtle differences in colors.

[0236] By regularly measuring the color difference values ​​of Huangjing wine at different storage times, we can observe how the color of Huangjing wine changes with the extension of storage time.

[0237] The colorimeter measures the L, a, and b values ​​of the standard sample (newly brewed Huangjing wine, i.e. Huangjing wine stored for 0 days) and the sample to be tested (Huangjing wine stored for different times) to calculate the color difference (ΔE) between the two. The formula is shown in Formula 2. In the formula, L represents brightness, and the larger the value, the brighter it is; the a value represents red and green, and the larger the positive value of a, the darker the red, and the larger the negative value, the darker the green; the b value represents yellow and blue, and the larger the positive value of b, the darker the yellow, and the larger the negative value, the darker the blue. If the ΔE value is small, it means that the color does not change much and the stability is good; on the contrary, if the ΔE value is large, it means that the color changes significantly and the stability is poor.

[0238]

[0239] in:

[0240] △L——The difference between the L values ​​measured twice before and after

[0241] △a——The difference between the two measured a values

[0242] △b——The difference between the b values ​​measured twice before and after

[0243] The color remains stable during the storage of Huangjing wine, which is of immeasurable importance to its quality. The constancy of color directly reflects the stability of Huangjing wine quality. If the color can remain the same or only slightly change during the storage process, it means that most of the components in the wine have maintained good stability, avoiding significant oxidation and degradation processes, thereby ensuring the overall quality of Huangjing wine is good and stable.

[0244] There is a close connection between color and flavor. Stable color often indicates that the loss of flavor components in the wine is minimal, and the unique medicinal and wine aroma of Polygonatum odoratum can be preserved intact. On the contrary, if the color changes significantly, such as deepening or fading, this is often a sign of damage to the flavor substances, which may be accompanied by the generation of unpleasant flavors, thus affecting the overall flavor of Polygonatum odoratum wine.

[0245] Figure 5 The color difference of Huangjing wine in Examples 1 to 3 after being stored for different days at room temperature is specifically shown. The size of the △E value in the figure directly reflects the degree of color difference. The larger the △E value, the more significant the color difference. Figure 5It can be seen that with the passage of storage time, the color difference value of Huangjing wine did not show significant fluctuations, which fully proves that the color of Huangjing wine remained highly stable within 120 days of storage at room temperature.

[0246] 5. Evaluation of the clarity of Huangjing wine during storage

[0247] Based on transmittance, a physical indicator that measures the ability of light to penetrate a medium, the dynamic changes in liquid transparency can be directly observed, thereby effectively evaluating whether the Huangjing wine of Examples 1 to 3 maintains a clear and stable state during storage.

[0248] Transmittance (T) is the ability of light to pass through a medium. It is the percentage of the luminous flux passing through a transparent or translucent body to the incident luminous flux. When parallel monochromatic light passes through a uniform, non-scattering medium, part of the light is absorbed, part of it passes through the medium, and part of it is reflected by the surface of the medium. The transmittance T is equal to the ratio of the intensity of the transmitted light It to the intensity of the incident light I0, that is, T = It / I0. The greater the transmittance, the stronger the medium's ability to transmit light and the higher the transparency.

[0249] The clarity of Huangjing wine is closely related to its transparency. During storage, if Huangjing wine becomes turbid or precipitates, its transparency will decrease, and the light transmittance will also decrease. In view of this, the light transmittance of Huangjing wine can be tested regularly using a spectrophotometer to intuitively grasp the trend of its transparency changes and further evaluate the stability of its clarity.

[0250] Combined with the test range of the spectrophotometer and related research, 680nm was selected as the wavelength to measure the clarity of Huangjing wine. Take an appropriate amount of Huangjing wine sample and measure its transmittance at a wavelength of 680nm. By comparing the transmittance data of Huangjing wine at different storage times, the change in its clarity can be intuitively evaluated. Generally speaking, the higher the transmittance value, the better the clarity of Huangjing wine; conversely, the lower the transmittance, the lower its clarity. See the results. Figure 6 .

[0251] Figure 6 The light transmittance of the Huangjing wine of Examples 1 to 3 at room temperature changes with the number of days of storage. It can be clearly observed from the figure that with the extension of storage time, the light transmittance of the Huangjing wine does not fluctuate significantly, which proves that the clarity of the Huangjing wine of Examples 1 to 3 remains highly stable within 120 days of storage at room temperature.

[0252] In summary, the Huangjing wine of Examples 1 to 3 was stored at room temperature for up to 120 days, and after comprehensive stability evaluation, including sensory evaluation, alcohol content, total acid content, color and clarity, it showed excellent stability. Its flavor and quality changed little during storage, the alcohol content and total acid content remained constant, the color was stable without significant changes, and the clarity was also maintained well. These results show that the Huangjing wine of Examples 1 to 3 has excellent storage characteristics under room temperature storage conditions, and the quality remains stable for 120 days.

Claims

1. A composite bacterial fermentation agent, characterized in that: It is composed of plant lactobacillus, lactic acid-producing bacillus and brewer's yeast in a mass ratio of 2-4:1-2:50-100.

2. The composite bacterial fermentation agent according to claim 1, characterized in that: The mass ratio of the plant lactobacillus, the lactic acid-producing bacillus and the brewing yeast is 2-3:1-1.5:50-75.

3. A method for preparing Huangjing wine by combined fermentation of composite bacteria, characterized in that: The following steps are involved: S1. Activating the composite bacterial fermentation agent according to claim 1 or 2 to obtain a composite bacterial solution; S2. saccharify the Polygonatum sibiricum tablets, mix them with distilled water and the composite bacterial solution, and ferment them at 25 to 35° C. for 5 to 10 days to obtain fermented wine; S3. Remove impurities from the fermented wine, filter and sterilize it, and remove macromolecular compounds with a molecular weight greater than 10,000 to obtain Polygonatum odoratum wine.

4. The preparation method according to claim 3, characterized in that: The activation in step S1 is performed by using a mixed solution of physiological saline and sugar water.

5. The preparation method according to claim 4, characterized in that: The sugar water is 4-6wt% sugar water.

6. The preparation method according to claim 3, characterized in that: The activation condition in step S1 is 25-35° C. for 25-35 minutes.

7. The preparation method according to claim 3, characterized in that: The polygonatum sibiricum slices described in step S2 are used after being soaked, cleaned and boiled.

8. The preparation method according to claim 3, characterized in that: The mass ratio of the polygonatum slices, sugar, distilled water and composite bacterial liquid in step S2 is 1:10-20:35-70:

1.

9. The preparation method according to claim 8, characterized in that: The mass ratio of the polygonatum slices, sugar, distilled water and composite bacterial liquid in step S2 is 1:10-15:35-52.5:

1.

10. The preparation method according to claim 3, characterized in that: The filtration sterilization in step S3 is performed by using 0.4-0.5 μm and 0.1-0.2 μm microporous filter membranes in sequence.

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