Yeast Strain for Rapid Fermentation of Sugar-Free Rehmannia Root and Its Use

Abstract

The present invention provides a yeast strain for rapid fermentation of sugar-free dough and its use. The budding yeast strain AMCC31248 (Saccharomyces cerevisiae AMCC31248) provided by the present invention has been deposited with the China Center for Type Culture Collection (CCTCC), and the deposit number is CCTCC NO: M 20211686. The budding yeast strain AMCC31248 provided by the present invention has good fermentation performance in sugar-free dough and can rapidly ferment sugar-free dough.

Description

Technical Field

[0001] The present invention relates to the field of microorganisms, and specifically to a yeast strain for rapid fermentation of sugar-free dough and its use.

Background Art

[0002] From the discovery of the first commercial baker's yeast to the formal establishment of the yeast manufacturing process and the development of the current active dry yeast, the types of yeast products have been increasing day by day, the quality of the products has been continuously improved, and it can be used to manufacture various wheat flour foods. Wheat flour fermented foods, as one of the traditional staple foods, occupy an important position in the diet structure of the common people. According to statistics, more than about half of the population in China mainly eats wheat flour foods, and most of them are sugar-free dough fermented wheat flour foods such as steamed buns and steamed stuffed buns. From the perspective of health, sugar-free or low-sugar wheat flour foods are more acceptable to consumers.

[0003] In the manufacture of wheat flour foods, the dough fermentation process always plays an important role. It not only affects the softness, texture, nutritional value, etc. of wheat flour food products, but also determines the speed of dough fermentation. The stronger the fermentation ability of yeast, the less time required until the start of dough fermentation. As a result, the fermentation period is shortened, the manufacturing process of wheat flour foods is accelerated, and the efficiency of industrial mass production is increased. Therefore, developing a budding yeast strain with good fermentation performance in sugar-free dough has great practical significance.

Summary of the Invention

Problems to be Solved by the Invention

[0004] In the manufacturing process of wheat flour foods, since the environmental temperature is very high, yeast fermentation often starts early during kneading, and the quality of the products often deteriorates. In order to lower the temperature of the dough, it is common to add cold water to cool it, but yeast cells will be in a shock state when entering a low-temperature environment, which affects the normal growth of yeast cells. Therefore, strong low-temperature osmotic shock resistance of yeast is strongly required.

[0005] Therefore, in view of the problems of the prior art that the efficiency of sugar-free dough fermentation by budding yeast is low and the low-temperature osmotic shock resistance is poor, the present invention provides a yeast strain for rapid fermentation of sugar-free dough with low-temperature osmotic shock resistance.

Means for Solving the Problems

[0006] In a first aspect, the present invention provides a budding yeast strain, wherein the budding yeast strain is the budding yeast strain AMCC31248 (Saccharomyces cerevisiae AMCC31248), which is deposited at the China Center for Type Culture Collection (CCTCC) with the deposit number CCTCC NO: M 20211686.

[0007] In a second aspect, the present invention provides a method for fermentatively producing a budding yeast agent, the method comprising culturing the budding yeast strain.

[0008] Preferably, the production method comprises (1) a step of expanding the culture of the budding yeast strain; and (2) a step of adding the product obtained in step (1) to a liquid medium and performing fermentation culture at 26 to 32 °C.

[0009] In a third aspect, the present invention provides an agent containing the budding yeast strain AMCC31248 (Saccharomyces cerevisiae AMCC31248).

[0010] Preferably, the agent is obtained by the fermentation production method.

[0011] In a fourth aspect, the present invention also provides the use of the budding yeast strain or the agent in fermentation.

[0012] In a fifth aspect, the present invention also provides the use of the budding yeast strain or the agent in dough.

[0013] In a sixth aspect, the present invention provides a dough, and the dough contains the budding yeast strain or the microbial agent.

[0014] Preferably, the dough contains wheat flour and the budding yeast strain in a mass ratio of 100:0.5 to 5.

[0015] In a seventh aspect, the present invention also provides a method for producing the dough, and the production method includes a step of kneading wheat flour with water at 0 to 35°C.

[0016] In the present invention, the water at 0°C may be ice at 0°C, or an ice-water mixture at 0°C, or liquid water at 0°C.

[0017] In the process for producing the dough according to the present invention, water, wheat flour, and budding yeast may be added in any order. For example, the wheat flour and yeast may be uniformly mixed, and then water at 0 to 35°C may be added and kneaded. Wheat flour may be added to water at 0 to 35°C, and then budding yeast may be added, or budding yeast may be added to water at 0 to 35°C, and then wheat flour may be added.

[0018] In an eighth aspect, the present invention also provides a wheat flour product obtained by the method for producing the dough.

[0019] Preferably, the wheat flour product is steamed bun, baozi, bread, biscuit, noodle, fried dumpling, etc.

Advantages of the Invention

[0020] The budding yeast strain AMCC31248 (Saccharomyces cerevisiae AMCC31248) provided by the present invention has good fermentation performance in sugar-free dough, can rapidly ferment sugar-free dough, and has good low-temperature osmotic shock resistance.

[0021] (Strain Deposit Information) The budding yeast strain AMCC31248 (Saccharomyces cerevisiae AMCC31248) provided by the present invention was deposited with the China Center for Type Culture Collection (CCTCC) on December 29, 2021, with the deposit number CCTCC NO: M 20211686. The depository institution is Wuhan University, Wuhan, China, with the postal code 430072 and the telephone number 027 - 68754052.

[0022] The budding yeast strain AMCC30010 (Saccharomyces cerevisiae AMCC30010) used in the present invention was deposited with the China Center for Type Culture Collection (CCTCC) on March 29, 2022, with the deposit number CCTCC NO: M 2022340. The depository institution is Wuhan University, Wuhan, China, with the postal code 430072 and the telephone number 027 - 68754052.

[0023] The budding yeast strain AMCC32101 (Saccharomyces cerevisiae AMCC32101) provided by the present invention was deposited with the China Center for Type Culture Collection (CCTCC) on March 29, 2022, with the deposit number CCTCC NO: M 2022341. The depository institution is Wuhan University, Wuhan, China, with the postal code 430072 and the telephone number 027 - 68754052.

Brief Description of the Drawings

[0024]

Figure 1

Figure 2

Figure 3

Embodiments for Carrying Out the Invention

[0025] In the present invention, using the budding yeast AMCC30010 strain and the budding yeast AMCC32101 strain as parent strains, the budding yeast AMCC31248 strain provided by the present invention is obtained by the method of microscopic hybridization.

[0026] The budding yeast AMCC30010 (Saccharomyces cerevisiae AMCC30010) strain is a budding yeast strain bred by Angel Yeast Co., Ltd., and the original strain was collected in Yichang City, Hubei Province. When observed under an optical microscope, the cells of this budding yeast strain are elliptical in shape, reproduce by budding, and the single colonies grown on the solid plate are spherical with a slightly raised center, milky white, smooth surface, and neat edges. Through morphological observation with a high-magnification microscope and molecular biological identification, it was determined to be a strain of the Saccharomyces cerevisiae species and is edible. It was deposited with the China Center for Type Culture Collection (CCTCC) on March 29, 2022, and the deposit number is CCTCC NO:M 2022340 (i.e., CCTCC M 2022340).

[0027] The budding yeast AMCC32101 (Saccharomyces cerevisiae AMCC32101) strain is a budding yeast strain bred by Angel Yeast Co., Ltd., and the original strain was collected in Ulanqab City, Inner Mongolia Autonomous Region. When observed under an optical microscope, the cells of this budding yeast strain are elliptical in shape, reproduce by budding, and the single colonies grown on the solid plate are spherical with a slightly raised center, milky white, smooth surface, and neat edges. Through morphological observation with a high-magnification microscope and molecular biological identification, it was determined to be a strain of the Saccharomyces cerevisiae species and is edible. It was deposited with the China Center for Type Culture Collection (CCTCC) on March 29, 2022, and the deposit number is CCTCC NO:M 2022341 (i.e., CCTCC M 2022341).

[0028] The budding yeast strain AMCC31248 provided by the present invention has good fermentation performance in sugar-free dough and can quickly ferment sugar-free dough.

[0029] The production of some sugar-free bread, soda crackers, steamed buns, etc. mainly involves fermenting sugar-free dough. Since most of the wheat flour is starch and is converted to maltose by the action of amylase in the wheat flour, the level of maltose utilization ability of yeast determines the time required until the fermentation of sugar-free dough starts. The maltose utilization enzyme system of yeast includes maltose hydrolase and maltose permease. Yeast with high maltose utilization ability is called rapid fermentation yeast.

[0030] Sugar-tolerant yeast refers to yeast that has high tolerance to sucrose in sugar-containing dough, that is, yeast whose growth and fermentation performance in sugar-containing bread are both higher than those of ordinary yeast.

[0031] Low-sugar tolerant yeast is used in a sucrose dough system of about 7%, and high-sugar tolerant yeast is used in a dough system with a higher sucrose concentration, and the content reaches even 25%. Generally, sucrose cannot be directly utilized by microorganisms. Among budding yeasts, sucrose hydrolase that can decompose sucrose is contained. It acts on β-1,2 glycosidic bonds to hydrolyze sucrose into D-glucose and D-fructose. Subsequently, glucose and fructose enter the glucose degradation pathway by the glycolysis system and are utilized by yeast. At the same time, the glucose and fructose generated by the rapid decomposition of sucrose increase the osmotic pressure around the yeast cells. The yeast cell membrane is a selectively semi-permeable biological membrane, and the level of external concentration affects the activity of yeast cells. When the cell is in an environment with high osmotic pressure, the water and protoplasm inside the cell exude from the cell membrane, dehydrating or killing the cell. Therefore, the hypertonic environment surrounding budding yeast in high-sugar dough affects both its growth and fermentation performance. Therefore, the gas generation ability of sugar-free yeast is determined by the activity of maltose utilization enzymes, and the gas generation ability of sugar-tolerant yeast is determined by the activity of sucrase.

[0032] The source information of the reagents and equipment used in the examples of the present invention is as shown in Table 1 and Table 2.

Table 1

Table 2

[0033] The composition of the sporulation medium used in the examples of the present invention is 1% potassium acetate, 0.1% powdered yeast extract, 0.05% glucose, and 2% agar.

[0034] In the examples of the present invention, each budding yeast strain was activated using YPD solid medium. The composition of the YPD solid medium is 1% powdered yeast extract, 2% peptone, 2% glucose, and 2% agar.

[0035] In the examples of the present invention, each budding yeast strain was cultured using YPD liquid medium. The composition of the YPD liquid medium is 1% powdered yeast extract, 2% peptone, and 2% glucose.

[0036] (Example 1: Construction and identification of strains) The parental budding yeast strains AMCC30010 and AMCC32101 were each activated, sporulation was induced, and after enzymatic degradation of the cells, single spores were picked using a yeast micromanipulator. Single spores from two different parental strains were brought into contact and then cultured at 30°C. The morphology of the spores was observed. When the hybridization of the two single spores was successful, they were continuously cultured at 30°C. This was the first-generation strain. The obtained first-generation strain was also hybridized with the budding yeast strain AMCC30010 to obtain a second-generation strain, and a sporulation test was performed, and hybrid strains were selected for subsequent screening.

[0037] Using the fully automatic growth curve analyzer Bioscreen C, the growth curves of the hybrid new strains obtained by hybridization were measured, and the top 20 hybrid strains with higher growth efficiency than the parental strains were selected.

[0038] Fermentation tests were carried out on 20 hybrid strains with higher growth efficiency obtained from the resulting parent strains using shaking flasks. Using the net dry weight of the strains and their fermentation activity in a 0% sugar dough system of fresh yeast as screening indicators, hybrid strains were selected in which the net dry weight of yeast milk could reach 95 - 105% of any parent strain and the fermentation activity of the 0% sugar dough reached 95 - 150% of any parent strain.

[0039] Subsequently, the hybrid strains obtained by screening in the above step were cultured in a fermenter in a 45L system, and active dry yeast was prepared respectively with the obtained yeast cells. The fermentation activity of the active dry yeast in a 0% sugar dough system was measured. New strains were selected in which there were no obvious abnormalities in the dry yeast production process and the fermentation activity of the active dry yeast in a 0% sugar dough reached 95 - 120% of any parent strain.

[0040] Finally, low - temperature osmotic shock resistance screening was carried out on the preferred hybrid strains selected in the above step. Kneading with crushed ice at 0 °C to make a 0% sugar dough containing hybrid strain active dry yeast, measuring the fermentation time of the dough, and taking the hybrid strain with the shortest fermentation time of the dough as the target strain. In this way, hybrid strains with low - temperature osmotic shock resistance were screened.

[0041] A hybrid strain numbered AMCC31248 was obtained through the above screening. This strain has high fermentation activity in a 0% sugar dough and good low - temperature osmotic shock resistance. This strain was identified, and the identification results were as follows.

[0042] Observed under an optical microscope, the strain cells were oval in shape, reproduced by budding, and the single colonies grown on the solid plate were spherical with a slightly raised center, milky white, with a loose texture, easy to pick with an inoculation loop, smooth and dry on the surface, and with a neat edge. Figure 1 shows the colonies of the hybrid strain AMCC31248.

[0043] When the spore formation of the hybrid strain AMCC31248 was examined under a microscope, as shown in Figure 2, there were many spores with complete morphology in the microscopic field of view, indicating that it had the ability of spore formation, that is, it was a hybrid strain.

[0044] The obtained hybrid strain AMCC31248 was named Saccharomyces cerevisiae AMCC31248 strain. The Saccharomyces cerevisiae AMCC31248 strain was deposited at the China Center for Type Culture Collection (CCTCC) on December 29, 2021, and the deposit number was CCTCC NO: M 20211686 (i.e., CCTCC M 20211686).

[0045] (Example 2: Measurement of growth efficiency) The Saccharomyces cerevisiae AMCC31248 strain obtained in Example 1, and the parental strains Saccharomyces cerevisiae AMCC30010 and Saccharomyces cerevisiae AMCC32101 were inoculated into wort medium (purchased from Qingdao Hi-tech Industrial Park Hope Bio-technology Co., Ltd.), cultured at 30 °C for 48 hours, and the OD 600 values of each strain at different time points were measured with a high-throughput by an automatic growth curve analyzer Bioscreen C. With time (h) on the horizontal axis and the corresponding OD 600 value on the vertical axis, the growth curve was plotted, and the test data was analyzed to calculate the growth efficiency of the strain by the following formula. Growth efficiency = (OD 2 - OD 1 ) / (t 2 - t 1 ) OD 1 : OD 1 value corresponding to the strain at time t 600 OD 2 : OD 2 value corresponding to the strain at time t 600 t 1 : Starting point of the logarithmic growth phase t 2 : Ending point of the logarithmic growth phase

[0046] The growth curve of budding yeast AMCC31248 is shown in Figure 3. It can be seen that this strain can grow rapidly in wort medium. The following Table 3 shows the growth efficiencies of the parent strain and the budding yeast AMCC31248 strain.

Table 3

[0047] As can be seen from Table 3, the growth efficiency of the obtained budding yeast AMCC31248 strain is significantly higher than that of the parent strains, budding yeast AMCC30010 and budding yeast AMCC32101.

[0048] (Example 3: Measurement of the fermentation activity of fresh yeast) The budding yeast AMCC31248 obtained in Example 1 was inoculated into an Erlenmeyer flask containing a fermentation medium and cultured at 30°C. The precipitate collected after centrifugation was yeast milk. The weight of the yeast milk was weighed and the moisture content of the yeast milk was measured. The net dry weight (g / L) of each strain in the Erlenmeyer flask was calculated by the following formula. Net dry weight (g / L) = weight of yeast milk × (1 - moisture content)

[0049] A dough was made with a 0% sugar dough system shown in Table 4, and the fermentation activity of the budding yeast AMCC31248 strain was measured. The mass of the yeast milk added to the budding yeast AMCC31248 strain and the parent strains, budding yeast AMCC30010 and budding yeast AMCC32101, was calculated and weighed. Wheat flour, salt, and water were weighed according to the dough formulation shown, and uniformly mixed in a kneader to make a dough. The total amount of carbon dioxide gas generated by yeast fermentation for 1 hour at 30°C of 280 g of the dough made from the system shown in Table 4 was directly measured using an SJA fermentation apparatus, and this was taken as the fermentation activity of the strain, and the results were shown in milliliters (mL).

Table 4

[0050] The relative percentage of the net dry weight in Table 5 was calculated by the following formula. Relative percentage of net dry weight (%) = (Net dry weight of the hybrid new strain / Net dry weight of the parent strain) × 100% The relative percentage of the dough fermentation activity of the budding yeast AMCC31248 strain against its parent strain in Table 6 was calculated by the following formula. Relative percentage of dough fermentation activity (%) = Dough fermentation activity of the hybrid new strain / Dough fermentation activity of the parent strain × 100%

[0051] As shown in Table 5, the net dry weights of the budding yeast AMCC31248 strain were 95.7% and 109.9% of those of the parent strains, budding yeast AMCC30010 and budding yeast AMCC32101, respectively. As shown in Table 6, the fermentation activities of the fresh yeast of the budding yeast AMCC31248 strain in the 0% sugar dough were all higher than those of the two parent strains, with a difference of about 10%.

Table 5

Table 6

[0052] (Example 4: Measurement of Fermentation Activity of Active Dry Yeast) After activating and culturing the budding yeast AMCC31248 strain, it was expanded in a fermentation tank in a 45 L system, and then further separated, washed, pressure-filtered, and dried to obtain active dry yeast. With the dough formulation shown in Table 7, wheat flour, salt, water, and the prepared active dry yeast were weighed respectively, and using the SJA method, the total amount of carbon dioxide gas generated by fermenting 280 g of dough made from the system shown in Table 7 with active dry yeast for 1 hour was measured, and this was taken as the dough fermentation activity of the active dry yeast.

Table 7

[0053] For the parent strain AMCC32101, it is very difficult to dry in the active dry yeast manufacturing process and there is no corresponding fermentation activity data. Therefore, only the dry yeast activity of the parent strain AMCC30010 was used as a control here. The budding yeast AMCC31248 strain showed no obvious abnormality in the active dry yeast manufacturing process. For the measurement data of the fermentation activity of its active dry yeast in the 0% sugar dough, please specifically refer to Table 8. Here, the calculation method of the relative percentage of fermentation activity in Table 8 was as follows. Relative percentage of fermentation activity = (fermentation activity of the hybrid new strain / fermentation activity of the parent strain AMCC 30010) × 100% [Table 8]

[0054] As is clear from the results, compared with the parent strain budding yeast AMCC30010 strain, the active dry yeast of the budding yeast AMCC31248 strain still has a 11.5% difference in the dough fermentation activity under the condition of 0% sugar. Therefore, it was shown that the budding yeast AMCC31248 strain has a certain drought tolerance.

[0055] (Example 5: Measurement of the low-temperature osmotic shock resistance of active dry yeast) According to the dough formulation shown in Table 9, crushed ice at 0 °C was put into a kneader, active dry yeast was added and mixed at a low speed. Next, wheat flour and salt were added, and mixing was continued while stirring until it became uniform to make the dough. After preparing 400 g of the dough, it was put into a fermenter and fermented. The temperature was limited to 38 ± 1 °C and the humidity was limited to 85 - 90%. The time required for the fermented dough to reach the same set height was recorded, and this was the fermentation time. [Table 9]

[0056] For the fermentation time of the dough corresponding to the budding yeast AMCC31248 strain and the parent strain budding yeast AMCC30010 strain, please refer to Table 10. Here, the calculation method of the relative percentage of fermentation time in Table 10 was as follows. Relative percentage of fermentation time = (Fermentation time of the hybrid new strain / Fermentation time of the parent strain AMCC30010) × 100%

Table 10

[0057] As is clear from the results, the fermentation time of the budding yeast AMCC31248 strain is only 70.0% of that of the parent budding yeast AMCC30010 strain, with a 30% difference. Therefore, under the same conditions, it was shown that the budding yeast AMCC31248 strain ferments the dough faster and has excellent low-temperature osmotic shock resistance.

[0058] What is described above is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any corrections, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention shall be included within the protection scope of the present invention.

[0059] (Appendix) (Appendix 1) A budding yeast strain, which is the budding yeast AMCC31248 strain (Saccharomyces cerevisiae AMCC31248), deposited with the China Center for Type Culture Collection (CCTCC), and has a deposit number of CCTCC NO: M 20211686.

[0060] (Appendix 2) A method for fermentatively producing a budding yeast agent, characterized by including the step of culturing the budding yeast strain described in Appendix 1.

[0061] (Appendix 3) (1) The step of subculturing the budding yeast strain described in Appendix 1; and (2) The step of adding the product obtained in step (1) to a liquid medium and performing fermentation culture at 26 to 32°C.

[0062] (Appendix 4) A microbial agent, characterized by containing the budding yeast strain AMCC31248 (Saccharomyces cerevisiae AMCC31248) described in Supplementary Note 1.

[0063] (Supplementary Note 5) The microbial agent described in Supplementary Note 4, characterized by being obtained by the fermentation production method described in Supplementary Note 2 or 3.

[0064] (Supplementary Note 6) Use of the budding yeast strain described in Supplementary Note 1 or the microbial agent described in Supplementary Note 4 or 5, characterized by being used in fermentation.

[0065] (Supplementary Note 7) Use of the budding yeast strain described in Supplementary Note 1 or the microbial agent described in Supplementary Note 4 or 5, characterized by being used in dough.

[0066] (Supplementary Note 8) Dough, characterized by containing the budding yeast strain described in Supplementary Note 1 or the microbial agent described in Supplementary Note 4 or 5.

[0067] (Supplementary Note 9) The dough described in Supplementary Note 8, characterized by containing flour and the budding yeast strain in a mass ratio of 100:0.5 - 5.

[0068] (Supplementary Note 10) A method for producing the dough described in Supplementary Note 8 or 9, characterized by including the step of kneading flour with water at 0 - 35°C.

[0069] (Supplementary Note 11) A wheat flour product obtained by the method for producing dough described in Supplementary Note 8 or 9.

[0070] (Supplementary Note 12) The wheat flour product described in Supplementary Note 11, which is steamed buns, baozi, bread, biscuits, noodles, or fried dumplings.

Claims

1. A budding yeast strain, which is the budding yeast strain AMCC31248 (Saccharomyces cerevisiae AMCC31248), deposited with the China Center for Type Culture Collection (CCTCC), with the deposit number CCTCC NO: M 20211686, characterized by being a budding yeast strain.

2. A method for fermentatively producing a budding yeast agent, characterized by including the step of culturing the budding yeast strain according to Claim 1.

3. (1) The step of subculturing the budding yeast strain according to Claim 1, and (2) The step of adding the product obtained in step (1) to a liquid medium and performing fermentation culture at 26 to 32 °C, characterized by including the above steps, the production method according to Claim 2.

4. An agent, characterized by containing the budding yeast strain AMCC31248 (Saccharomyces cerevisiae AMCC31248) according to Claim 1.

5. The agent according to Claim 4, characterized by being obtained by the fermentation production method according to Claim 2 or 3.

6. The use of the budding yeast strain according to Claim 1 or the agent according to Claim 4 or 5, characterized by being used in fermentation.

7. The use of the budding yeast strain according to Claim 1 or the agent according to Claim 4 or 5, characterized by being used in dough.

8. Dough, characterized by containing the budding yeast strain according to Claim 1 or the agent according to Claim 4 or 5.

9. The dough according to Claim 8, characterized by containing wheat flour and the budding yeast strain with a mass ratio of 100:0.5 to 5.

10. A method for producing the dough according to Claim 8 or 9, characterized by including the step of kneading wheat flour with water at 0 to 35 °C.

11. A wheat flour product obtained by the method for producing dough according to Claim 8 or 9.

12. The wheat flour product according to Claim 11, which is steamed buns, baozi, bread, biscuits, noodles, or pot stickers.

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