Phaeolepiota aurea s-lwz20190428-3 and application thereof in preparation of rice fungus grain

Abstract

The present application belongs to the technical field of multifunctional fungus grain preparation. The present application provides a strain of Phylloporia pulla S-LWZ20190428-3CGMCC No. 40573. The strain has been preserved in the China General Microbiological Culture Collection Center (CGMCC) on April 12, 2023. The strain preservation number is CGMCC No. 40573. The address of the center is No. 1, Xibaheyi, Beijing, China. The present application provides a strain that can increase the total phenolic content in the product after rice fermentation by 77.94%, and the soluble protein content by about 15 times.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of multifunctional fungus grain preparation. BACKGROUND

[0002] Rice is one of the most commonly consumed grains in daily life, which can provide humans with nutrients such as protein, fatty acids, vitamins, and minerals. Due to the pursuit of taste, rice is over-processed, and as it is rolled more and more finely, it leads to a large loss of nutrients. Rice is rich in polyphenols, mainly in two forms of free and bound states, and the bound polyphenols are much higher than the free polyphenols. Most of the phenolic substances are not easily digested and absorbed by the human body because they are combined with polysaccharides, proteins and lipids in the cell wall in the form of bound state. In addition, the protein in rice often lacks lysine, methionine, threonine and tryptophan, at the same time, other amino acids cannot be fully utilized, so that the digestibility of protein and the nutritional value of grain are reduced, and rice nutrition strengthening has become a problem to be solved in the rice processing industry in China.

[0003] Phylloporia pulla belongs to Basidiomycota, Agaricomycetes, Hymenochaetales, Hymenochaetaceae and Phylloporia. The species of Phylloporia have a long history of medicinal use in China, especially in Shandong Province. They are mainly taken in the form of dried fruiting bodies and tea, and are believed to have the effects of treating pharyngitis and tonsillitis and enhancing immunity. Studies on the medicinal functions of Phylloporia fungi used by the public show that Phylloporia fungi have antitumor, antioxidant, immunomodulatory, antidiabetic and neurotrophic activities. Secondary metabolites such as polysaccharides, triterpenes and sterols play an important role in the process of the above functions of Phylloporia species. SUMMARY

[0004] Therefore, the application provides a Phylloporia pulla S-LWZ20190428-3CGMCC No.40573. The strain has been preserved in the China General Microbiological Culture Collection Center (CGMCC) on April 12, 2023, and the preservation number of the strain is CGMCC No.40573. The address of the center is No.1, Beichen West Road, Haidian District, Beijing.

[0005] The application also provides a composition comprising Phylloporia pulla S-LWZ20190428-3CGMCC No.40573.

[0006] The third aspect of the present application provides the application of Phylloporia pulla S-LWZ20190428-3 CGMCC No.40573 in preparing rice fungus food.

[0007] The present application at least includes the following beneficial effects:

[0008] Firstly, the flavonoid content of the rice fermented product inoculated with Phylloporia pulla in the present application is significantly increased, such as hesperidin, luteolin, which has various biological activities, such as antioxidant, anti-ototoxicity, anti-inflammatory, anti-atherosclerosis, and anti-hypertension.

[0009] Secondly, the lysine, threonine, tryptophan, and methionine contents of the rice fermented product inoculated with Phylloporia pulla in the present application are increased to different degrees, so that the essential amino acids lacking in rice are supplemented, and the nutritional value of protein is improved. DETAILED DESCRIPTION

[0010] Example 1

[0011] 1. Strain acquisition, isolation, and identification

[0012] The strain was collected on April 28, 2019, from Yongquan Village, Zhihe Town, Zichuan District, Zibo City, Shandong Province, obtained from fresh fruiting bodies growing on branches of living trees of Prunus sp. using tissue isolation method, named strain S-LWZ20190428-3, and after purification, cultured at 26-28℃ for 10-15 days.

[0013] The DNA of S-LWZ20190428-3 was extracted using lysis method and fungal genome rapid extraction kit, and its nLSU sequence was obtained by PCR amplification, the sequence information is shown as SEQ ID NO. 1 in the sequence table, through comparison with NCBI database BLAST and construction of phylogenetic tree, combined with morphological analysis, it was determined that the strain was identified as Phylloporia pulla.

[0014] Phylloporia pulla S-LWZ20190428-3 has been preserved in the China General Microbiological Culture Collection Center (CGMCC) on April 12, 2023, the center is abbreviated as CGMCC, the strain preservation number is CGMCC No.40573, and the address of the center is No. 1, Beichen West Road, Beijing City, Chaoyang District.

[0015] 2. Activated strain:

[0016] The preparation formula and method of mother culture medium (PDA) are as follows: 300 g / L of boiled potato liquid, 20 g / L of glucose, 5 g / L of peptone, 20 g / L of agar, 1 g / L of potassium dihydrogen phosphate, 0.5 g / L of magnesium sulfate heptahydrate, pH natural, pour into a conical flask after stirring, sterilize at 121℃ for 20 min, after the temperature decreases to room temperature, add 0.1% of streptomycin sulfate and ampicillin solution.

[0017] The low-temperature preserved Phylloporia pulla S-LWZ20190428-3 obtained in step (1) is transplanted into the mother culture medium (PDA) medium described above for subculture, and is cultured at 2℃ for 7-10 days in an incubator, to obtain the activated Phylloporia pulla mother culture.

[0018] 3, Preparation of rice fungus food:

[0019] Preparation of solid culture medium: wash the rice clean, soak in distilled water for 24 h, drain the surface water, divide into 15*30*5C polypropylene fungus bags, add 40% of the weight of the rice in distilled water, cover with 3.8 cm sponge and 0.22 μm microporous filter membrane, sterilize at 121℃ for 2.5 h, then cool to room temperature, to obtain the rice solid culture medium.

[0020] Inoculation culture: inoculate the activated strain S-LWZ20190428-3 in step (2) into the above rice solid culture medium, inoculate a 6 cm diameter PDA culture medium inoculum block, and place it in a 26℃ constant temperature incubator for culture for 45 days.

[0021] Preparation of fungus food: after the mycelium grows throughout the culture medium, dry it at 40-45℃ to constant weight, grind the rice with a grinder and pass it through a 90 mesh sieve, to obtain the rice functional fungus food, which is stored at -80℃ for standby use.

[0022] Comparative Example 1

[0023] Preparation of control group rice raw food: wash the rice clean, soak in distilled water for 24 h until the kernels become soft, drain the surface water, divide into 15*30*5C polypropylene fungus bags, add 40% of the weight of the rice in distilled water, cover with 3.8 cm sponge and 0.22 μm microporous filter membrane, sterilize at 121℃ for 2.5 h, then cool to room temperature, to obtain the rice solid culture medium. Dry the obtained solid culture medium at 40-45℃ to constant weight, grind the rice with a grinder and pass it through a 90 mesh sieve, to obtain the control group raw food, which is stored at -80℃ for standby use.

[0024] Experimental Example 1

[0025] Experiment I, extraction and content determination of active substances in Phylloporia pulla inoculated rice solid fermentation product and raw food:

[0026] Determination was carried out as follows:

[0027] 1. Determination of total phenol content in Phaeolepiota aurea inoculated rice solid fermentation product:

[0028] The determination of total phenol content in the sample was carried out by Folin phenol colorimetry (Wang WK, Lu N, Yan J, Fu LZ, Song JL, Yuan WD, Zhou ZF. Effects of growth duration on the nutrition, active ingredients and antioxidant activity of segment wood cultivated Baum's Sanghuang fruiting bodies. Mycosystema, 2021, 40(3): 668-680)

[0029] 2. Determination of soluble protein content in Phaeolepiota aurea inoculated rice solid fermentation product: determined by BCA protein content test kit

[0030] The total phenol and soluble protein contents in the Phaeolepiota aurea inoculated rice fermentation product were detected in the experimental example, and the specific data are shown in Table 1:

[0031] Table 1 Active substance content in examples and comparative examples

[0032] Comparative Example 1-1 Comparative Example 1-2 Comparative Example 1-3 Example 1-1 Example 1-2 Example 1-3 Total phenolics (g / mg) 1.09 0.99 2 2.43 2.7 2.13 Soluble protein (g / mg) 5.69 4.7 9 104.7 97.3 105.9

[0033] As can be seen from the above table, the total phenol and soluble protein contents of the three replicates of the examples are significantly higher than those of the comparative examples, indicating that inoculation of Phaeolepiota aurea has a significant effect on the increase of total phenol and soluble protein contents in the raw grain.

[0034] Experiment two, non-targeted detection of differential metabolites in Phaeolepiota aurea inoculated rice fermentation product and raw grain:

[0035] 1. Determination of differential metabolites in Phaeolepiota aurea inoculated rice fermentation product. Non-targeted metabolomics determination method (An MJ, Hong DS, Chang DD, Zhang CY, Fan H, Wang KY. Polymer amendment regulates cadmium migration in cadmium contaminated cotton field: Insights from genetic adaptation and phenotypic plasticity. Science of the Total Environment, 2022, 808: 151075) was used.

[0036] The content of active substances in the fermentation product of non-targeted detection of Phylloporia fasciata inoculated rice showed that, compared with the comparative example, the contents of hesperetin and luteolin in flavonoids were significantly increased, among which hesperetin had various biological activities, such as antioxidant, anti-ototoxicity, anti-inflammatory, anti-atherosclerosis, anti-hypertension and the like; luteolin had various pharmacological effects such as antioxidant, anti-inflammatory and anti-tumor and the like.

[0037] Compared with the comparative example, the content of β-caryophyllene, a sesquiterpene compound in the fermentation product inoculated with Phylloporia fasciata, was significantly increased, and the compound belongs to bicyclic sesquiterpene, and has pharmacological activities such as anti-inflammatory, antioxidant and anti-anxiety; in addition, nootkatone and capsidiol were also significantly increased.

[0038] Compared with the comparative example, the content of β-sitosterol in the fermentation product inoculated with Phylloporia fasciata was significantly increased, and β-sitosterol has the effect of lowering cholesterol, and further enhances the functional value of the fungus-grain.

[0039] Compared with the comparative example, the contents of lysine and tryptophan in the fermentation product inoculated with Phylloporia fasciata were significantly increased, effectively supplementing the lack of lysine, methionine, threonine and tryptophan in rice, so that other amino acids can be further utilized, improving the protein digestibility and nutritional value of the original grain.

[0040] The lysine includes: β-alanyl-L-lysine, N6-(L-1,3-dicarboxypropyl)-L-lysine, L-methionine and L-threonine.

[0041] The tryptophan includes: N-acetyl-D-tryptophan and tryptophan.

[0042] The specific data is shown in Table 2, and the percentage increase of the example compared with the comparative example is shown in Table 3:

[0043] Table 2 Quantification of differential metabolites in examples and comparative examples

[0044]

[0045]

[0046] Note: Fold change of metabolites in different groups Table III Increase in content of different active substances Examples over Comparative Examples

[0047]

[0048] Note: Increase in percentage values: Examples over Comparative Examples

Claims

1. Physisporinus vitreus (Fr.) Bond. et Sing. Phylloporia pulla ) S-LWZ20190428-3, with the accession number of CGMCC No. 40573.

2. Composition, characterized in that, The polypeptide of claim 1, wherein the polypeptide is a polypeptide having an amino acid sequence of SEQ ID NO: 1 Phylloporia pulla ) S-LWZ20190428-3.

3. The dark leaf pore fungus (Physisporinus sp.) of claim 1. Phylloporia pulla ) S-LWZ20190428-3 in the preparation of rice fungus grain.

4. The dark leaf pore fungus (Physisporinus vitreus) of claim 1, Phylloporia pulla ) Application of S-LWZ20190428-3 in improving the content of rice fermentation products, characterized in that, the rice fermentation product is one or more of flavonoids, soluble proteins, sesquiterpenes, and phytosterols; the flavonoids include one or more of hesperitin and luteolin; the soluble proteins include one or more of beta-alanyl-L-leucine, N6-(L-1,3-dicarboxypropyl)-L-lysine, methionine, threonine, N-acetyl-D-tryptophan, and tryptophan; the sesquiterpenes include one or more of caryophyllen, melilotate, and capsicodiol.

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