Caribik Yarrowia mayer yeast and application thereof in vinasse treatment

Abstract

The invention belongs to the technical field of biology, and particularly relates to Meyerozyma caribba XX2120, and the preservation number of the Meyerozyma caribba XX2120 is GDMCC No: 67777. The invention further relates to a preparation method of the Meyerozyma caribba XX2120. The strain is obtained by taking saccharomycetes X2120 as a starting strain, carrying out ARTP mutagenesis treatment and screening, has relatively strong acid resistance, relatively high salt stress resistance and certain alcohol resistance and aldehyde resistance, can effectively utilize monosaccharide, disaccharide and trisaccharide, has a wide utilization range on carbohydrate and has a relatively good utilization rate on inorganic nitrogen. Moreover, the strain can be used for producing mycoprotein through liquid fermentation by taking Luzhou-flavor distillers' grains as a raw material, and also can be used for performing solid fermentation on the Luzhou-flavor distillers' grains, so that the true protein content of the distillers' grains is increased, the nutritional value of the distillers' grains is improved, and the recycling of distillers' grain resources is effectively promoted.

Description

Technical Field

[0001] This invention belongs to the field of biotechnology, specifically relating to a type of *Calibic Mayer's yeast* and its application in the treatment of distiller's grains. Background Technology

[0002] With the improvement of people's living standards, the rapid growth in residents' demand for meat, eggs, and dairy products has driven the continuous expansion of the livestock industry, leading to an ever-increasing demand for feed protein. Therefore, the development of new feed protein alternatives is urgently needed. Yeast protein is a novel protein with high essential amino acid content, ideal distribution, and rich branched-chain amino acids. It is also rich in fats, carbohydrates, minerals, and vitamins, possessing nutritional value no less than conventional protein sources. Furthermore, it features high efficiency, low resource utilization, short production cycles, and sustainable production, gradually becoming an economical and eco-friendly alternative to traditional animal and plant proteins. However, current traditional yeast protein production mainly relies on sugar raw materials such as corn-derived glucose and sugarcane molasses. These raw materials compete with human food resources. To ensure the sustainability of yeast protein supply, it is necessary to develop unconventional substrates.

[0003] Rice husks are added as an auxiliary material during the fermentation process of baijiu (Chinese liquor), resulting in a high content of cellulose, hemicellulose, and lignin in the lees. This severely affects the palatability and digestibility of the lees, hindering its application in livestock and poultry farming. However, after certain pretreatment, these lignocellulose biomass can be converted into a mixed sugar solution consisting mainly of glucose, xylose, hexoses, and pentoses, providing a cheap carbon source for yeast growth.

[0004] In the process of producing microbial protein from lignocellulosic biomass (such as corn stalks, rice straw, and distiller's grains), the biomass must first undergo physicochemical pretreatment. During this pretreatment, several fermentation inhibitors inevitably form, such as furfural, phenols, and weak acid compounds. The accumulation of these toxic byproducts significantly inhibits microbial growth and fermentation activity, thereby reducing the yield and process stability of the target product. Secondly, the carbon sources produced from lignocellulosic biomass are complex sugars. Most yeast strains exhibit selective utilization of complex carbon sources, exhibiting a catabolite repression (CCR) effect of the phosphoenolpyruvate transferase system. This leads to the inactivation of non-glucose transport and catabolism systems in the presence of glucose, limiting the microorganisms' utilization of the complex carbon sources from lignocellulosic biomass. Therefore, breeding yeast strains that can utilize complex carbon sources, possess a certain degree of tolerance, and have high protein content and high biomass is crucial for producing protein from distiller's grains biomass resources. Summary of the Invention

[0005] The purpose of this invention is to screen out a type of Calibick Mayer's yeast, in order to obtain yeast protein from the lees of strong-aroma baijiu (Chinese liquor) and promote the application of strong-aroma baijiu lees in the livestock and poultry farming industry.

[0006] To achieve the above objectives, the present invention provides: a Calibick Mayer's yeast ( Mycoplasma Caribbean )XX2120, its accession number is GDMCC No:67777.

[0007] The present invention also provides a microbial agent comprising the above-mentioned *Calibic Mayer's yeast* ( Mycoplasma Caribbean )XX2120 or its culture. Among them, *Calibic Mayer's yeast* ( Meyerozyma caribbica )XX2120 culture is a culture of Calibick Mayer's yeast ( Meyerozyma caribbica The substance obtained by culturing XX2120 in a culture medium includes Calibick Mayer's yeast ( Meyerozyma caribbica XX2120 (the cell itself) and Calibick Mayer's yeast ( Meyerozyma caribbica Metabolites of XX2120. The term "culture" refers to any liquid or solid culture medium that has grown a microbial community after artificial inoculation and cultivation. It is the product obtained by growing and / or amplifying microorganisms; it can be a biologically pure culture of microorganisms, or it can contain a certain amount of culture medium, metabolites, or other components produced during the cultivation process. The term "culture" also includes passaged cultures obtained by subculturing microorganisms; these can be cultures of a single generation or a mixture of several generations.

[0008] This invention also provides the above-mentioned Calibick Mayer's yeast ( Meyerozyma caribbica The preparation method of XX2120 includes the following steps: S1. Yeast strain X2120 was used as the starting strain. The starting strain was subjected to ARTP mutagenesis. The ARTP mutagenesis conditions were: high-purity helium as the working gas, power of 100 W, working gas flow rate of 10 LSM, irradiation distance of 2 mm, and mutagenesis time of 90-150 s. S2. The strains treated with ARTP mutagenesis were cultured in droplets, and droplets with OD600 > 10.5 were screened. The droplets were then expanded and cultured to obtain strains with OD600 > 5. The expansion culture was continued, and strains with relatively high cell mass and protein yield were screened using cell mass and protein yield as indicators. S3. The strains screened in step S2 are expanded into larger-scale cultures. Using cell volume, protein content, and protein yield as indicators, *Calibic Mayer's yeast* is selected. Meyerozyma caribbica )XX2120.

[0009] This invention also provides the above-mentioned Calibick Mayer's yeast ( Meyerozyma caribbica Application of XX2120 or the above-mentioned microbial agents in any of the following: (1) Production of yeast protein from baijiu lees through liquid fermentation; (2) Solid-state fermentation of baijiu lees.

[0010] This invention also provides a method for producing yeast protein through liquid fermentation of baijiu lees, comprising the following steps: 1) Pretreatment: Add hydrogen peroxide to the strong-aroma baijiu lees. The mass concentration of hydrogen peroxide added to the strong-aroma baijiu lees is 0.55-0.61%. Treat at 36-37℃ for 1.8-2 hours to obtain pretreated baijiu lees. 2) Enzymatic hydrolysis: Adjust the pH of the pretreated baijiu lees to 5 using ammonia water, add cellulase, and enzymatically hydrolyze at 120-150 r / min for 72-80 h, then centrifuge at 8000 r / min for 10 min to obtain the supernatant. 3) Fermentation: Inoculate the supernatant with the above-mentioned *Calibic Myersia* ( Meyerozyma caribbica ) XX2120 or the above-mentioned inoculum was fermented at 30℃ and 180 r / min for 48 h, and then centrifuged at 7500 r / min for 10 min to obtain a precipitate. The precipitate was dried or freeze-dried to obtain yeast protein.

[0011] Optionally, in step 2), the amount of cellulase added is 1.5 to 2% of the weight of the pretreated baijiu lees.

[0012] This invention also provides a method for solid-state fermentation of baijiu lees, comprising the following steps: a) Pretreatment: Add hydrogen peroxide to the strong-aroma baijiu lees. The mass concentration of hydrogen peroxide added to the strong-aroma baijiu lees is 0.55-0.61%. Treat at 36-37℃ for 1.8-2 h to obtain pretreated baijiu lees. b) Fermentation: Adjust the pH of the pretreated baijiu mash to 5 using ammonia water, add ammonium sulfate, soybean meal, wheat bran, and corn cob, mix well, then add cellulase and inoculate with the above-mentioned Calibick Mayer's yeast ( Meyerozyma caribbica Mix XX2120 or the above-mentioned inoculum, and ferment at 30°C for 72–96 h to obtain the fermented product.

[0013] Optionally, in step b), the amount of ammonium sulfate added is 0.5-1% of the weight of the pretreated distillers' grains, the amount of soybean meal added is 2-5% of the weight of the pretreated distillers' grains, the amount of wheat bran added is 7.5% of the weight of the pretreated distillers' grains, the amount of corn cob added is 10% of the weight of the pretreated distillers' grains, and the amount of cellulase added is 0.5-1% of the weight of the pretreated distillers' grains.

[0014] The beneficial effects of this invention are as follows: This method screened out *Calibic Mayer's yeast* ( Meyerozyma caribbica XX2120, after crystal violet staining, exhibits the following morphological characteristics: single oval cells, some showing budding; single colonies on YPD plates are milky white, glossy, smooth, and have regular edges. The *Calibicula cambogia* (Calibicula cambogia) used in this protocol... Mycoplasma Caribbean In YPD medium, XX2120 has a cell protein content of up to 49.82%, a cell weight of 13.26 g / L, and a protein yield of 6.6 g / L. Its growth is unaffected under pH ≥ 2 conditions, and it has a long adaptation period under pH = 1.5 conditions. It has strong acid resistance, high salt resistance, and certain resistance to alcohol and aldehydes. It can effectively utilize monosaccharides, disaccharides, and trisaccharides, has a wide range of carbohydrate utilization, and has a good conversion ability for inorganic nitrogen.

[0015] The *Calibic Mayer's yeast* in this protocol ( Meyerozyma caribbica XX2120 can be inoculated into the supernatant obtained after enzymatic hydrolysis and centrifugation of baijiu lees, utilizing the reducing sugars in the supernatant to produce yeast protein, with a yield of up to 3.63 g / L. Furthermore, it can be directly inoculated into pretreated baijiu lees, and through solid-state fermentation, increase the true protein content of the baijiu lees, improve its nutritional value, effectively promote the recycling of baijiu lees resources, and alleviate the shortage of protein feed in livestock and poultry farming to some extent. Attached Figure Description

[0016] Figure 1 The image shows the detection results of the 57 strains screened in Example 1 of this invention; Figure 2 This is a morphological image of strain 2-C1 in Example 2 of the present invention after being stained with crystal violet; Figure 3 This is a distribution diagram of strain 2-C1 on a YPD plate in Example 2 of the present invention; Figure 4 This is a magnified image of a single colony of strain 2-C1 in Example 2 of the present invention; Figure 5 This is a phylogenetic tree of the 26S rRNA of strain 2-C1 in Example 2 of the present invention; Figure 6 This is a graph showing the acid resistance test results of strain 2-C1 in Example 3 of the present invention; Figure 7 This is a graph showing the salt tolerance test results of strain 2-C1 in Example 3 of the present invention; Figure 8 This is a graph showing the alcohol resistance test results of strain 2-C1 in Example 3 of the present invention; Figure 9 This is a graph showing the furfural resistance test results of strain 2-C1 in Example 3 of the present invention; Figure 10 The images are scanning electron microscope (SEM) images of the liquor lees before and after pretreatment in Embodiment 6 of the present invention. Detailed Implementation

[0017] The present invention will now be described in further detail with reference to the accompanying drawings, so that those skilled in the art can implement it based on the description. The following description is intended to disclose the present invention and enable those skilled in the art to implement it. The preferred embodiments described below are merely examples, and those skilled in the art can conceive of other obvious modifications. The basic principles of the invention defined in the following description can be applied to other embodiments, modifications, improvements, equivalents, and other technical solutions that do not depart from the spirit and scope of the invention. In this invention, unless otherwise specified, the raw materials and equipment used are commercially available or commonly used in the art. The methods in the following embodiments, unless otherwise specified, are conventional methods in the art.

[0018] Example 1: Obtaining Meyerozyma caribbica XX2120 Yeast strain X2120 (isolated and screened from the surface of fruit peel in the laboratory) and other inclusions, preserved in glycerol tubes at -80℃, were aseptically transferred to a 250 mL shake flask containing 50 mL of YPD liquid medium. The flask was incubated at 200 r / min and 30℃ for 24 h. Subsequently, 2–3 loops of the culture were aseptically streaked onto YPD plates and incubated at 30℃ for 48 h. Single colonies of the activated strain were picked from the YPD plates and incubated in 3 mL of YPD liquid medium with shaking at 200 r / min and 30℃ for 16–18 h until the logarithmic growth phase. The YPD liquid medium formulation was: 5.0 g yeast extract, 10.0 g peptone, 20.0 g glucose, and 1 L distilled water, sterilized at 121℃ for 15 min (unless otherwise specified, all YPD liquid medium formulations are as described).

[0019] Centrifuge an appropriate amount of the cultured bacterial solution at 6000 r / min for 10 min, discard the supernatant, resuspend in physiological saline, wash three times, add 1 mL of glycerol and resuspend again. Dilute the resuspended bacterial solution 10 times and set aside for later use. Place 10 μL of the diluted bacterial solution on a mutagenesis slide and perform mutagenesis using an SRTP mutagenesis breeding instrument (ARTP-C1-13, Tianmu Biotechnology). The mutagenesis conditions are: high-purity helium working gas, power 100 W, working gas flow rate 10 LSM, and irradiation distance 2 mm.

[0020] Four time points with a mutagenic lethality rate above 80% were selected: 90 s, 100 s, 110 s, and 150 s. After these times, the culture was eluted in YPD liquid medium. After elution, 100 μL was collected for counting, and the remainder was mixed together and cultured in a high-throughput micro-scale droplet culture omics system (MISS-cell, Tianmu Biotechnology). After 42 h of incubation at 30℃, droplets with OD600 > 10.5 were collected to construct a mutagenic library. A total of 783 droplets were screened, and 564 of them were selected. These droplets were inoculated into 96-well deep-well plates containing 1 mL of YPD liquid medium for scale-up culture at 30℃, 220 r / min, for 48 h. After culture, the OD600 of the bacterial culture was measured using a multi-functional microplate reader (Synergy H1, Biotek), and strains with OD600 > 5 were screened, resulting in 57 strains. 50 μL of each of the 57 bacterial strains was placed in 50 mL of YPD liquid medium and cultured at 30℃ for 48 h. The OD600 of the strains was then measured, followed by centrifugation at 7500 r / min for 10 min to determine cell weight, cell protein content, and protein yield. The results are as follows: Figure 1 As shown. Figure 1 Among the strains, 18 had a cell protein content >60%, but these strains generally had low cell weights, with most having cell weights below 6 g / L and protein yields below 4.5 g / L. While these strains accumulated high amounts of protein, their growth was limited, resulting in low protein production efficiency. Conversely, 23 strains had a cell weight >14.5 g / L. These strains generally had cell protein content between 40% and 46%, which was relatively low, but they had high cell biomass and protein yields all >6.5 g / L, indicating higher protein production efficiency. Therefore, 13 strains with high biomass and high protein yield were selected for another round of screening.

[0021] Thirteen selected strains were cultured in 50 mL YPD liquid medium, with each strain inoculated at 50 μL in triplicate. After culturing at 30℃ for 48 h, the OD600 of the strains was measured. Subsequently, the cells were centrifuged at 7500 r / min for 10 min to determine cell weight, cell protein content, and protein yield. The results are shown in Table 1. Yeast strain X2120 (the starting strain) was used as a control group for screening. The only difference between the starting strain X2120 and the other 13 strains was that it had not undergone ARTP mutagenesis treatment; all other procedures were the same. Strains with superior protein content, cell weight, and protein yield were selected.

[0022] Table 1 Results of the second round of screening experiments for the mutagenic strains

[0023] Table 1 shows that the protein yield of all the mutagenized strains was increased. Strain 2-C1 had the highest protein yield, reaching 6.6 g / L, an increase of 12.6% compared to the original strain; its protein content was 49.82%, an increase of 3.1% compared to the original strain; and its cell mass was 13.26 g / L, an increase of 9.3% compared to the original strain. Therefore, the glycerol tubes of strain 2-C1 were stored at -80℃.

[0024] Example 2: Strain Identification After activation, strain 2-C1 was cultured in shake flasks. Under aseptic conditions, it was centrifuged at 5000 r / min for 5 min to obtain bacterial cells. Morphological characteristics of the cells after crystal violet staining were: single oval cells, some with budding linear structures, such as... Figure 2 As shown; single colonies on YPD plates are milky white, glossy, smooth, and have neat edges, as indicated. Figure 3 and Figure 4 As shown.

[0025] Genomic DNA of strain (2-C1) was extracted using the HyperMB Extraction-Free Fungal Genome PCR Direct Amplification Kit (Shanghai Sangon Biotech Co., Ltd., catalog number: B690021-0100). Amplification was then performed using NL1:5'-GCATATCAATAAGCGGAGGAAAAG-3' and NL4:5'-GGTCCGTGTTTCAAGACGG-3' as upstream and downstream primers. After the amplification products passed the tests, 26S rRNA sequencing was performed. The sequencing results were as follows:

[0026] Homology comparison analysis was performed in the NCBI-Genbank database. Strains with high homology were selected, and phylogenetic trees were constructed using MEGA 7.0 software. The 26S rRNA phylogenetic tree of strain (2-C1) is shown below. Figure 5 As shown, the results indicate that strain (2-C1) is *Calibic Mayer's yeast*. Meyerozyma caribbica ), named Calibick Mayer's yeast ( Meyerozyma caribbica)XX2120, this strain was deposited on February 2, 2026 at the Guangdong Provincial Center for Microbial Culture Collection, located at No. 100 Xianlie Middle Road, Yuexiu District, Guangzhou City, Guangdong Province, with accession number GDMCC No:67777.

[0027] Example 3: Study on the stress resistance of the strain The strain (2-C1) obtained in Example 1 (i.e., *Calibic Mayer's yeast*) was used. Mycoplasma Caribbean (XX2120), take a single colony and put it into 50 mL of YPD liquid medium. Culture for 16-18 h until the logarithmic phase to obtain the primary seed culture. Inoculate it into the medium of different treatments at an inoculation rate of 1%.

[0028] (I) Acid resistance study: The pH of YPD liquid culture medium was adjusted to 1, 1.5, 2, 3, and 4 using 5 mol / L hydrochloric acid solution, and the OD600 was measured at 6 h, 12 h, 18 h, 24 h, and 48 h of incubation. The results are as follows: Figure 6 As shown.

[0029] (II) Salt resistance study: 10 g / L, 30 g / L, 50 g / L, and 70 g / L NaCl were added to YPD liquid culture medium, and OD600 was measured at 6 h, 12 h, 18 h, 24 h, and 48 h of incubation. The results are as follows: Figure 7 As shown.

[0030] (III) Ethanol Tolerance Study: Anhydrous ethanol with volume fractions of 0%, 4%, 8%, 12%, 16%, and 20% was added to YPD liquid culture medium, and OD600 was measured at 6 h, 12 h, 18 h, 24 h, and 48 h of incubation. The results are as follows: Figure 8 As shown.

[0031] (IV) Furfural tolerance study: Furfural was added to YPD liquid culture medium at concentrations of 0.5 g / L, 1 g / L, 1.5 g / L, 2 g / L, and 2.5 g / L, and OD600 was measured at 6 h, 12 h, 18 h, 24 h, and 48 h, respectively. The results are as follows: Figure 9 As shown.

[0032] pass Figure 6 It can be seen that *Calibic Mayer's yeast* ( Meyerozyma caribbica The XX2120 strain showed no growth susceptibility at pH ≥ 2, but exhibited a longer adaptation period at pH = 1.5, with slow growth occurring after 24 hours, demonstrating strong acid resistance. Figure 7 It can be seen that *Calibic Mayer's yeast* ( Meyerozyma caribbicaThe growth of strain XX2120 was largely unaffected under ≤50 g / L NaCl conditions. While growth was slower under 70 g / L NaCl conditions compared to salt-free conditions (YPD liquid medium), it still grew rapidly, demonstrating high salt tolerance. Figure 8 It can be seen that *Calibic Mayer's yeast* ( Meyerozyma caribbica The XX2120 strain can tolerate 4% (v / v) ethanol and can also grow slowly under 8% (v / v) ethanol conditions, demonstrating a certain degree of alcohol tolerance. (Through...) Figure 9 It can be seen that *Calibic Mayer's yeast* ( Meyerozyma caribbica The growth of strain XX2120 was not affected under stress of 1 g / L furfural. Under stress of 1.5 g / L furfural, the lag phase of the strain was prolonged, but after adaptation, the bioaccumulation was not affected, indicating a certain degree of tolerance to furfural.

[0033] Example 4: Utilization of common carbohydrates by the strain The procedure was performed according to the API ID32C kit requirements (bioMérieux (Shanghai) Biopharmaceuticals Co., Ltd.) to study *Callibik Mayer's yeast* (…). Meyerozyma caribbica The utilization of common compounds by strain XX2120 is shown in Table 2. Table 2 shows that *Calibic Myersella calibickensis* (…) Meyerozyma caribbica The XX2120 strain can effectively utilize monosaccharides, disaccharides, and trisaccharides, and has a wide range of carbohydrate utilization capabilities.

[0034] Table 2. Utilization of common carbohydrates by strains

[0035] Example 5: The ability of strains to generate proteins using inorganic nitrogen The YPD medium formula is as follows: 5.0g yeast extract, 10.0g peptone, 20.0g glucose, add 1 L of distilled water, and sterilize at 121℃ for 15 min.

[0036] YPD medium was used as the starting medium, with a nitrogen content of 0.39%. The nitrogen was successively replaced by ammonium sulfate (1.84%), ammonium chloride (1.49%), urea (0.84%), ammonium carbonate (1.34%), ammonium bicarbonate (2.2%), and ammonia water (added fresh each time, 1.9%, analytical grade ammonia water with an ammonia content of 25-28%), while the carbon source remained unchanged. Simultaneously, 1.0 g of K₂HPO₄, 0.5 g of MgSO₄, 0.01 g of FeSO₄, and 0.5 g of NaCl were added to 1 L of medium. The pH of the medium was adjusted to 7 using 1 mol / L hydrochloric acid solution, and the medium was sterilized at 121°C for 15 min.

[0037] The Calibick Mayer's yeast prepared in Example 3 ( Meyerozyma caribbica XX2120 primary seed culture was inoculated at a 1% inoculum into each of the above-mentioned inorganic nitrogen source media, with three replicates for each inorganic nitrogen source medium. The cultures were incubated at 30℃ and 200 r / min for 48 h. After incubation, the cultures were centrifuged at 7500 r / min for 10 min, and the cell weight, cell protein content, and protein yield were measured. The results are shown in Table 3. Table 3 shows that *Calibic Mayer's yeast* (… Meyerozyma caribbica XX2120 exhibits good conversion efficiency for inorganic nitrogen, capable of converting common inorganic nitrogen sources into bacterial protein. It is even better than the strain isolated by Cao Xinong et al. (Isolation, identification and stress resistance study of high ammonia nitrogen conversion yeast, Journal of Animal Nutrition, 2025, 37(4): 2763-2771) which achieved a total protein yield of 1.76 g / L under ammonium sulfate as the sole nitrogen source.

[0038] Table 3. Ability of strains to convert proteins using inorganic nitrogen

[0039] Example 6: Preparation of yeast protein by liquid fermentation of strains 1) Pretreatment: Hydrogen peroxide was added to the strong-aroma baijiu mash (dry basis) at a concentration of 0.608% by mass. The mash was treated at 36.46℃ for 1.81 h to obtain pretreated baijiu mash. The oxidizing properties of hydrogen peroxide were utilized to open the lignin structure in the baijiu mash, providing a space for subsequent cellulase hydrolysis or bacterial fermentation. SEM analysis was performed on the structures of the strong-aroma baijiu mash before and after pretreatment. The results are shown below. Figure 10 As shown, after pretreatment, the morphology and structure of the baijiu lees changed significantly. The originally regular, smooth, long fibers showed obvious cracks, and some areas peeled off in flakes, which is beneficial for subsequent enzymatic hydrolysis and fermentation.

[0040] 2) Enzymatic hydrolysis: The pH of the pretreated baijiu lees was adjusted to 5 using ammonia water (conventional industrial ammonia water, generally an aqueous solution containing 25-28% ammonia). Cellulase Cellic CTec3 HS (purchased from Novozymes (Denmark)) was added, and the amount of cellulase added was 2% of the mass of the pretreated baijiu lees. After enzymatic hydrolysis at 120 r / min for 80 h, the mixture was centrifuged at 8000 r / min for 10 min to obtain the supernatant. The reducing sugar content in the supernatant was measured, and the results are shown in Table 4.

[0041] 3) Fermentation: The primary seed culture of *Meyerozyma caribbica* strain XX2120 obtained in Example 3 was inoculated into YPD liquid medium at a 1% inoculation rate and cultured until the end of the logarithmic phase to obtain the secondary seed culture of the strain; 50 mL of the supernatant from step 2) was taken and inoculated into *Meyerozyma caribbica* strain XX2120 at a 1% inoculation rate. Meyerozyma caribbica Secondary seed culture of strain XX2120 was fermented at 30℃ and 180 r / min for 48 h, then centrifuged at 7500 r / min for 10 min. The reducing sugar content of the supernatant was determined, and the precipitate was dried to obtain yeast protein. The protein content was weighed and determined, and the results are shown in Table 4. Table 4 shows that *Calibic Myersella* (… Mycoplasma Caribbean XX2120 can basically utilize the reducing sugar obtained from enzymatic hydrolysis of baijiu lees to generate 9.17 g / L of bacterial cells, with a bacterial protein content of 39.62% and a protein yield of 3.63 g / L.

[0042] Table 4. Changes in fermentation indicators of strong-aroma baijiu lees hydrolysate

[0043] Example 7 Solid-state fermentation of strong-aroma baijiu mash a) Pretreatment: Hydrogen peroxide was added to the strong-aroma baijiu lees. The mass concentration of hydrogen peroxide added to the strong-aroma baijiu lees (dry basis) was 0.608%. The mixture was treated at 36.46℃ for 1.81 h to obtain pretreated baijiu lees.

[0044] b) Fermentation: Adjust the pH of the pretreated baijiu mash to 5 using ammonia water (conventional industrial ammonia water, generally an aqueous solution containing 25-28% ammonia). Based on the mass of the pretreated baijiu mash, add 0.5% ammonium sulfate, 2% soybean meal, 7.5% wheat bran, and 10% corn cob. Mix well, then add 1% cellulase Cellic CTec3 HS (purchased from Novozymes (Denmark)) and inoculate with 5% of the Calibick Mayer's yeast prepared in Example 6. Meyerozyma caribbica The secondary seed culture of XX2120 was mixed and fermented at 30℃ for 72 hours to obtain the fermented product. The nutrient composition before and after fermentation was determined, as shown in Table 5. Table 5 shows that *Calibic Myersella* (…) Meyerozyma caribbica XX2120 can effectively increase the crude protein and true protein content of baijiu lees, improve the nutritional value of baijiu lees, and effectively promote the recycling and utilization of baijiu lees resources.

[0045] Table 5. Changes in nutritional components of strong-aroma baijiu mash before and after fermentation (oven-dry basis)

[0046] The above are merely embodiments of the present invention. The invention is not limited to the fields covered by these embodiments. Commonly known structures and characteristics are not described in detail here. Those skilled in the art are aware of all common technical knowledge in the field prior to the application date or priority date, are aware of all existing technologies in that field, and have the ability to apply conventional experimental methods prior to that date. Those skilled in the art can improve and implement this solution based on the guidance provided in this application and their own capabilities. Some typical known structures or methods should not be obstacles for those skilled in the art to implement this application. It should be noted that those skilled in the art can make several modifications and improvements without departing from the structure of the present invention. These should also be considered within the scope of protection of the present invention, and will not affect the effectiveness or practicality of the invention. The scope of protection claimed in this application should be determined by the content of its claims. The specific embodiments described in the specification can be used to interpret the content of the claims.

Claims

1. A type of Calibick Mayer's yeast ( Meyerozyma caribbica XX2120, characterized in that: Its accession number is GDMCC No:67777.

2. A microbial agent, characterized in that: Contains the *Calibic Mayer's yeast* as described in claim 1. Meyerozyma caribbica )XX2120 or its culture.

3. The Calibick Mayer's yeast as described in claim 1 ( Meyerozyma caribbica The preparation method of XX2120 is characterized by: Includes the following steps: S1. Yeast strain X2120 was used as the starting strain. The starting strain was subjected to ARTP mutagenesis. The ARTP mutagenesis conditions were: high-purity helium as the working gas, power of 100 W, working gas flow rate of 10 LSM, irradiation distance of 2 mm, and mutagenesis time of 90-150 s. S2. The strains treated with ARTP mutagenesis were cultured in droplets, and droplets with OD600 > 10.5 were screened. The droplets were then expanded and cultured to obtain strains with OD600 > 5. The expansion culture was continued, and strains with relatively high cell mass and protein yield were screened using cell mass and protein yield as indicators. S3. The strains screened in step S2 are expanded into larger-scale cultures. Using cell volume, protein content, and protein yield as indicators, *Calibic Mayer's yeast* is selected. Meyerozyma caribbica )XX2120.

4. The Calibick Mayer's yeast as described in claim 1 ( Meyerozyma caribbica The use of XX2120 or the microbial agent as described in claim 2 in any of the following: (1) Production of yeast protein from baijiu lees through liquid fermentation; (2) Solid-state fermentation of baijiu lees.

5. A method for producing yeast protein through liquid fermentation of baijiu lees, characterized in that: Includes the following steps: 1) Pretreatment: Add hydrogen peroxide to the strong-aroma baijiu lees. The mass concentration of hydrogen peroxide added to the strong-aroma baijiu lees is 0.55-0.61%. Treat at 36-37℃ for 1.8-2 hours to obtain pretreated baijiu lees. 2) Enzymatic hydrolysis: Adjust the pH of the pretreated baijiu lees to 5 using ammonia water, add cellulase, and enzymatically hydrolyze at 120-150 r / min for 72-80 h, then centrifuge at 8000 r / min for 10 min to obtain the supernatant. 3) Fermentation: Inoculate the supernatant with the *Calibic Myers sinensis* as described in claim 1. Meyerozyma caribbica )XX2120 or the inoculum as described in claim 2, fermented at 30°C and 180 r / min for 48 h, then centrifuged at 7500 r / min for 10 min to obtain a precipitate, and the precipitate was dried to obtain yeast protein.

6. The method for producing yeast protein by liquid fermentation of baijiu lees according to claim 5, characterized in that: In step 2), the amount of cellulase added is 1.5 to 2% of the weight of the pretreated baijiu lees.

7. A method for solid-state fermentation of baijiu lees, characterized in that: Includes the following steps: a) Pretreatment: Add hydrogen peroxide to the strong-aroma baijiu lees. The mass concentration of hydrogen peroxide added to the strong-aroma baijiu lees is 0.55-0.61%. Treat at 36-37℃ for 1.8-2 h to obtain pretreated baijiu lees. b) Fermentation: Adjust the pH of the pretreated baijiu mash to 5 using ammonia water, add ammonium sulfate, soybean meal, wheat bran, and corn cob, mix well, then add cellulase and inoculate with the Calibick Mayer's yeast as described in claim 1. Meyerozyma caribbica )XX2120 or the bacterial agent as described in claim 2, after being mixed, is fermented at 30°C for 72-96 h to obtain the fermented product.

8. The solid-state fermentation method for baijiu lees according to claim 7, characterized in that: In step b), the amount of ammonium sulfate added is 0.5-1% of the weight of the pretreated distillers' grains, the amount of soybean meal added is 2-5% of the weight of the pretreated distillers' grains, the amount of wheat bran added is 7.5% of the weight of the pretreated distillers' grains, the amount of corn cob added is 10% of the weight of the pretreated distillers' grains, and the amount of cellulase added is 0.5-1% of the weight of the pretreated distillers' grains.

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