Composition for increasing selenium content in liquiritin, glycyrrhiza polysaccharide and liquorice and application of composition

Abstract

The invention discloses a composition for increasing the selenium content in liquiritin, glycyrrhiza polysaccharide and liquorice and application of the composition. The technical problem to be solved by the invention is how to increase the content of selenium and active substances in liquorice. To this end, the present invention provides a composition containing a rhizobium and a selenite, the rhizobium being a mesorhizium temperulum 01578 (i.e., a mesorhizium temperulum CCBAU 01578), and a method for producing the same, and the present invention provides a composition containing the rhizobium, i.e., a mesorhizium temperulum CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU CCBAU. Experiments prove that when the rhizobium and exogenous selenium are jointly applied, the content of liquiritin, glycyrrhiza polysaccharide, selenium, fat, protein, flavone, saponin and / or glycyrrhizic acid in liquorice can be effectively increased, and a reference basis is provided for increasing the medicinal value of liquorice and the like.

Description

Technical Field

[0001] The invention belongs to the field of biotechnology, and particularly relates to a composition for increasing the selenium content in liquiritin, glycyrrhizic polysaccharide and licorice, and application thereof. Background Art

[0002] Licorice root Glycyrrhiza uralensis Fisch., Glycyrrhiza inflata Glycyrrhiza inflata Bat. or Glycyrrhiza glabra Glycyrrhiza glabra The dried roots and rhizomes of Licorice (Licorice) are a perennial herbaceous plant in the genus Glycyrrhizae, Leguminosae, used for both medicinal and edible purposes. It primarily grows in arid desert environments. Its roots and rhizomes are used medicinally to tonify the spleen and replenish qi, relieve phlegm and relieve coughs, and harmonize various other medicines. Licorice's active ingredients, including flavonoids, saponins, and polysaccharides, contribute to its widespread value. Its medicinal value includes components such as glycyrrhizic polysaccharides, which can enhance immune function and help prevent and treat a variety of diseases. Glycyrrhizic acid and glycyrrhetinic acid exhibit anti-inflammatory properties. Licorice also acts on the respiratory mucosa to relieve cough symptoms and is an important ingredient in many cough and expectorant medications. According to the Chinese Pharmacopoeia, glycyrrhizin is a key component of licorice, and its content is crucial for its quality. Furthermore, as a nationally designated plant or herb for both medicinal and edible purposes, glycyrrhizic polysaccharides are widely used as natural sweeteners.

[0003] Selenium is a trace element that plays a crucial role in plant growth and physiological functions. It participates in physiological processes such as photosynthesis, respiration, and antioxidant defense, making it an essential trace element for the growth and development of both animals and plants. Selenium's antioxidant properties can enhance plant resistance to stress, reduce the incidence of pests and diseases, and increase agricultural yields. Within plants, selenium exists in various forms, such as selenocysteine and selenomethionine. These selenium compounds have significant impacts on plant metabolism and growth and development. Plants can effectively utilize these non-metallic elements to produce selenium-rich products to meet their growth and development needs. Selenium deficiency in plants may lead to growth retardation and decreased stress resistance. Selenium also plays an important role in human immunity, anti-tumor activities, and antioxidant defenses. Compared to inorganic selenium, organic selenium is safer for the human body, has higher absorption rates, and offers greater nutritional value. Research by Li Fangzheng et al. has shown that organic selenium, which is primarily formed through biotransformation and combines with organic compounds within the body, has cardiovascular, anti-tumor, anti-pathogen, immune, and nervous system protective effects. Rhizobia can live in the soil or in nodules formed on the roots of legumes. In the nodules, they form a symbiotic relationship with the legumes, where the plants provide energy to the rhizobia through photosynthesis, and the rhizobia fix atmospheric nitrogen in the nodules, converting it into a form that the plants can use and improving the soil microbial environment. Summary of the Invention

[0004] The technical problem to be solved by the present invention may be how to increase the contents of liquiritin, glycyrrhiza polysaccharide, selenium, fat, protein, flavonoid, saponin and / or glycyrrhizic acid in licorice. The technical problem to be solved by the present invention may also be to enhance the improvement effect of rhizobia on the contents of the above components in licorice.

[0005] To solve the above problems, the present invention provides a composition, which contains rhizobia and selenite, and the rhizobia may be Mesorhizobium temperatum ( Mesorhizobium temperatum ).

[0006] The Mesorhizobium temperatum ( Mesorhizobium temperatum ) may be Mesorhizobium temperatum ( Mesorhizobium temperatum ) 01578 (i.e., Mesorhizobium temperatum ( Mesorhizobium temperatum ) CCBAU 01578).

[0007] The composition may be a composition for increasing the contents of liquiritin, glycyrrhiza polysaccharide, selenium, fat, protein, flavonoid, saponin and / or glycyrrhizic acid in licorice.

[0008] The ratio of rhizobia to selenite in the composition may be 4.88×10 6 CFU rhizobia: 578 μmol selenite.

[0009] In the above composition, the selenite may be a water-soluble selenite salt. The water-soluble selenite salt may be sodium selenite.

[0010] The composition may have at least one of the following properties: A1) Increasing the content of liquiritin in licorice, A2) Increasing the content of polysaccharide in licorice, A3) Increasing the content of selenium in licorice, A4) Increasing the content of fat in licorice, A5) Increasing the content of protein in licorice, A6) Increasing the content of flavonoid in licorice, A7) Increasing the content of saponin in licorice, A8) Increasing the content of glycyrrhizic acid in licorice.

[0011] In the composition, the rhizobia and the selenite may be independently packaged.

[0012] The composition may be a microbial agent or a fertilizer.

[0013] The above microbial agent refers to a live microbial preparation prepared by expanding the target microorganisms and using a carrier as an adsorbent to adsorb the fermentation broth or solid fermentation product of the microbial cells.

[0014] Among the above-mentioned microbial agents, the dosage form of the microbial agent can be various dosage forms, including but not limited to liquid agents, emulsions, suspensions, powders, granules, wettable powders, water-dispersible granules, etc.

[0015] As needed, the microbial agent may further include a carrier. The carrier can be a solid carrier or a liquid carrier.

[0016] The above-mentioned fertilizer can be a microbial nitrogen fertilizer, which can convert nitrogen in the air into ammonia that can be utilized by plants.

[0017] Specifically, the above-mentioned carrier can be at least one of calcium phosphate, rice husk powder, bran, rice bran, skimmed milk powder, maltodextrin, sucrose, glycerol, and starch.

[0018] The fertilizer can be used for soil application.

[0019] The present invention also provides the application of the above-mentioned composition in increasing the content of active ingredients in licorice, and the active ingredients are at least one of the following: B1) liquiritin, B2) polysaccharide, B3) selenium, B4) fat, B5) protein, B6) flavonoid, B7) saponin, B8) glycyrrhizic acid.

[0020] The licorice polysaccharide is a polysaccharide derived from licorice. The glycyrrhizic acid belongs to the saponin category. The liquiritin belongs to the flavonoid category.

[0021] The present invention also provides the application of the above-mentioned composition in the preparation of a product for increasing the content of active ingredients in licorice, and the active ingredients are at least one of the following: C1) liquiritin, C2) polysaccharide, C3) selenium, C4) fat, C5) protein, C6) flavonoid, C7) saponin, C8) glycyrrhizic acid.

[0022] The present invention also provides a method for increasing the content of active ingredients in licorice. The method includes the step of treating licorice with the above-mentioned composition to increase the content of active ingredients in licorice, and the active ingredients can be at least one of the following: D1) liquiritin, D2) polysaccharide, D3) selenium, D4) fat, D5) protein, D6) flavonoid, D7) saponin, D8) glycyrrhizic acid.

[0023] The present invention also provides a method for preparing a product for increasing the content of active ingredients in licorice. The method includes the step of using the above-mentioned composition as a component of the product, and the active ingredients can be at least one of the following: E1) liquiritin, E2) polysaccharide, E3) selenium, E4) fat, E5) protein, E6) flavonoid, E7) saponin, E8) glycyrrhizic acid.

[0024] The present invention also provides a method for preparing a composition, the composition is the above-mentioned composition, and the method includes the step of using the rhizobia and selenite as components of the composition.

[0025] In the above-described composition, the above application, and the above method, the improvement is a comparison carried out under comparable conditions. The comparable conditions may be that all other operations are the same except that the licorice is treated with the rhizobia instead of the composition. The comparable conditions may also be that all other conditions are the same except that the licorice is treated with water instead of the composition.

[0026] This paper studied the effect of exogenous selenium combined with rhizobia on the active ingredients of licorice. Using two-year-old licorice as the experimental material, a CK blank group, a Nano treatment group, a Nano+G treatment group, a SeIV treatment group, a SeIV+G treatment group (the composition treatment group of the present invention), etc. were set up, where G78 is Mesorhizobium temperatum Mesorhizobium temperatum 01578. Modern analytical techniques such as the Kjeldahl method, Soxhlet extraction, and high-performance liquid chromatography were used to determine the active ingredients such as crude protein, crude fat, glycyrrhizic acid, liquiritin, polysaccharide, and saponin in licorice.

[0027] The results showed that: in terms of selenium content, the selenium contents of the blank group and the rhizobium group were 0.094 mg / kg and 0.092 mg / kg respectively. The selenium contents of the SeIV group and the SeIV+G group treated with sodium selenite were approximately 30.43 times (2.86 mg / kg) and 32.06 times (3.01 mg / kg) that of the CK group, with extremely significant differences (P<0.01). The selenium contents of the Nano group and the Nano+G group treated with nano-selenium were approximately 1.65 times (0.155 mg / kg) and 2.3 times (0.217 mg / kg) that of the CK group. Among them, the Nano group was not significant, while the Nano+G group was extremely significant. In terms of fat content, the fat contents of the CK group, G78 group, SeIV group, SeIV+G group, Nano group and Nano+G group were 1.18%, 1.55%, 1.91%, 2.35%, 2.55% and 1.85% respectively. The G78 group was approximately 30.81% higher than the CK group, which was extremely significant. The SeIV group and the Nano group with single application of exogenous selenium were approximately 61.07% and 115.01% higher than the CK group respectively, both of which were extremely significant. In terms of protein content, the protein content of the G78 group was 5.04%, and that of the CK group was 3.77%. The G78 group was 33.73% higher than the CK group, which was extremely significant. The four groups with added exogenous selenium were also significantly higher than the CK group. In terms of flavonoid content, the flavonoid contents of the CK group and the G78 group were 27.89 mg / g and 36.94 mg / g respectively. The G78 group was approximately 32.44% higher than the CK group, which was extremely significant. The SeIV group and the Nano group of the exogenous selenium treatment group were approximately 12.01% and 14.37% higher than the CK group respectively, both of which were extremely significant. In terms of polysaccharide content, the polysaccharide contents of the CK group and the G78 group were 106.99 mg / g and 148.77 mg / g respectively. The G78 group was approximately 39.04% higher than the CK group, which was extremely significant. The SeIV group and the Nano group with single application of exogenous selenium were approximately 44.06% and 7.03% higher than the CK group respectively. In terms of saponin content, the saponin contents of the CK group and the G78 group were 0.96 mg / g and 69.17 mg / g respectively. The G78 group was approximately 22.6% higher than the CK group, which was extremely significant. In terms of glycyrrhizic acid content, the glycyrrhizic acid contents of the CK group and the G78 group were 2.03% and 2.39% respectively. The G78 group was approximately 17.56% higher than the CK group, which was extremely significant. The nano-selenium showed an obvious increase when single application of exogenous selenium. In terms of liquiritin content, the liquiritin contents of the CK group and the G78 group were 0.58% and 0.73% respectively. The G78 group was approximately 24.46% higher than the CK group, which was extremely significant. Sodium selenite showed an obvious increase when single application of exogenous selenium.

[0028] In the present invention, each treatment group meets the requirements of the Chinese Pharmacopoeia, providing a reference basis for improving the edible and medicinal values of licorice. The combination of the optimized rhizobium strain and exogenous selenium is more adaptable to the local soil environment in Xinjiang, effectively promoting the growth and quality improvement of licorice, and playing the best effect on the production of high-quality selenium-rich licorice in Xinjiang. Description of the Drawings

[0029] Figure 1 Comparison of selenium content in different treatment groups such as CK group, G78 group, SeIV treatment group, SeIV+G treatment group, Nano treatment group, Nano+G treatment group, etc.

[0030] Figure 2 Comparison of fat content in different treatment groups such as CK group, G78 group, SeIV treatment group, SeIV+G treatment group, Nano treatment group, Nano+G treatment group, etc.

[0031] Figure 3 Comparison of protein content in different treatment groups such as CK group, G78 group, SeIV treatment group, SeIV+G treatment group, Nano treatment group, Nano+G treatment group, etc.

[0032] Figure 4 Comparison of flavonoid content in different treatment groups such as CK group, G78 group, SeIV treatment group, SeIV+G treatment group, Nano treatment group, Nano+G treatment group, etc.

[0033] Figure 5 Comparison of polysaccharide content in different treatment groups such as CK group, G78 group, SeIV treatment group, SeIV+G treatment group, Nano treatment group, Nano+G treatment group, etc.

[0034] Figure 6 Comparison of saponin content in different treatment groups such as CK group, G78 group, SeIV treatment group, SeIV+G treatment group, Nano treatment group, Nano+G treatment group, etc.

[0035] Figure 7 Chromatogram of reference substance

[0036] Figure 8 Chromatogram of sample

[0037] Figure 9 Comparison of glycyrrhizic acid content in different treatment groups such as CK group, G78 group, SeIV treatment group, SeIV+G treatment group, Nano treatment group, Nano+G treatment group, etc.

[0038] Figure 10 Comparison of liquiritin content in different treatment groups such as CK group, G78 group, SeIV treatment group, SeIV+G treatment group, Nano treatment group, Nano+G treatment group, etc.

[0039] Figure 11 Correlation analysis of main components in licorice Specific implementation method

[0040] The present invention will be further described in detail below in conjunction with specific embodiments. The provided embodiments are only for clarifying the present invention and not for limiting the scope of the present invention. The following provided embodiments can be used as a guide for those of ordinary skill in the art to make further improvements and do not constitute any limitation to the present invention in any way.

[0041] In the following embodiments, the experimental methods are all conventional methods unless otherwise specified, and are carried out according to the techniques or conditions described in the literature in this field or according to the product instructions. The materials, reagents, etc. used in the following embodiments can be obtained from commercial channels unless otherwise specified.

[0042] Rhizobium sp. strain 78 in the following embodiments is Mesorhizobium temperatum Mesorhizobium temperatum ) 01578 (i.e., Mesorhizobium temperatum Mesorhizobium temperatum ) CCBAU 01578), which was kindly donated by Associate Professor Chen Wenfeng of China Agricultural University and is recorded in Table 1 of the following literature: Li M, Li Y, Chen WF, Sui XH, Li Y Jr, Li Y, Wang ET, Chen WX. Genetic diversity, community structure and distribution of rhizobia in the root nodules of Caragana spp. from arid and semi-arid alkaline deserts, in the north of China. Syst Appl Microbiol. 2012 Jun;35(4):239-45. doi: 10.1016 / j.syapm.2012.02.004. The public can obtain this biological material from Xinjiang Agricultural University. This biological material is only used for repeating the experiments of the present invention and cannot be used for other purposes.

[0043] In the following embodiments, SPSS 11.5 statistical software was used to process the data. The experimental results are expressed as mean ± standard deviation, and One-way ANOVA test was used. P < 0.05 (*) indicates significant difference, and P < 0.01 (**) indicates extremely significant difference.

[0044] 1. Materials and Instruments 1.1 Materials The experimental materials are two-year-old Glycyrrhiza uralensis Fisch. collected from the planting base of Lianqiang Group, the fifth group of Dongtan Village, Ershidian Town, Hutubi County, Changji Hui Autonomous Prefecture, Xinjiang Uygur Autonomous Region (44°17' N, 87°6' E, altitude 464.1 m). It was identified as Glycyrrhiza uralensis Fisch. by Associate Professor Ma Shengjun of Xinjiang Agricultural UniversityGlycyrrhiza uralensis Fisch.).

[0045] 1.2 Main reagents

[0046] 1.3 Test instruments

[0047] Example 1. Preparation of Rhizobium agent No. 78 1. Slant culture of Rhizobium No. 78 Pick Rhizobium No. 78 and inoculate it on a solid medium for slant culture. Culture it at 28 °C for 56 hours to obtain slant-cultured Rhizobium No. 78. The preparation method of the solid medium is as follows: 5 mL of glycerol, 5 g of mannitol, 1 g of yeast extract powder, 0.5 g of K2HPO4, 0.2 g of anhydrous MgSO4, 0.2 g of CaSO4·2H2O, 0.1 g of NaCl, 1% (NH4)6Mo7O 24 ·4H2O 1 mL, 1% H3BO3 1 mL, 20 g of agar, make up the volume to 1000 mL with water, adjust the pH value to 6.8 - 7.0, and sterilize at 121 °C for 30 min.

[0048] 2. Preparation of Rhizobium agent No. 78 Inoculate Rhizobium No. 78 into LB liquid medium at an inoculation amount of 1%, and place it in a shaker at 200 rpm and 30 °C for 12 h to obtain a fermentation broth. Centrifuge the fermentation broth to collect the cells, and then dilute the cells with sterile water to 1.95×10 5 CFU / ml to obtain a Rhizobium agent No. 78 with a Rhizobium content of 1.95×10 5 CFU / ml. Among them, the LB liquid medium is prepared as follows: 10 g of peptone, 5 g of yeast extract, 10 g of sodium chloride, adjust the pH to 7.0 with 5 mol / L NaOH, and make up the volume to 1 L with deionized water. Autoclave with distilled water at 121 °C for 20 min.

[0049] Example 2. Effects of different treatments on the contents of selenium and active substances in Glycyrrhiza uralensis 1. Preparation of nano-selenium Prepare nano-selenium according to the method of Kang et al. (Kang L, Wu Y, Jia Y, et al. Nano-selenium enhances melon resistance to Podosphaera xanthi by enhancing the antioxidant capacity and promoting alterations in the polyamine, phenylpropanoid and hormone signaling pathways [J]. Journal of Nanobiotechnology, 2023, 21(01): 377.). Add 10 mL of 20 mmol / L selenious acid solution and 4 mL of 1% ascorbic acid solution to 20 mL of 1% chitosan solution. Stir the mixture at 25 °C and 500 rpm for 3 h until the liquid turns transparent red, obtaining nano-selenium colloid.

[0050] 2. Specific methods for different treatments Take two-year-old licorice planted in the large field of the planting base of Lianqiang Group, the fifth group of Dongtan Village, Ershidian Town, Hutubi County, Changji Hui Autonomous Prefecture, Xinjiang Uygur Autonomous Region, China as the research object to study the effects of different treatments on licorice. The experiment adopts a randomized complete block design, with three plots. Each plot is set with a CK treatment group, a G78 treatment group, a SeIV treatment group, a SeIV+G treatment group, a Nano treatment group, and a Nano+G treatment group, with 100 licorice plants in each treatment group. The medicament applied to the CK treatment group is clear water (blank group), the medicament applied to the G78 treatment group is Rhizobium strain No. 78, the medicament applied to the SeIV treatment group is 4 mg / ml sodium selenite solution, the medicament applied to the SeIV+G treatment group is 4 mg / mL sodium selenite solution and Rhizobium strain No. 78, the medicament applied to the Nano treatment group is 4 mg / mL nano-selenium solution, and the medicament applied to the Nano+G treatment group is 4 mg / mL nano-selenium solution and Rhizobium strain No. 78. Except for the different medicaments applied in the above treatment groups, other operations are the same.

[0051] Start the following pesticide application treatment from May 1, 2024: CK treatment group: Irrigate 50 mL of clear water for each licorice plant, and then irrigate 50 mL of clear water for each licorice plant every 30 days, with a total of 4 irrigations to complete the pesticide application treatment.

[0052] G78 treatment group: Irrigate 50 mL of the Rhizobium strain No. 78 obtained in Example 1 with a content of 1.95×10 5The rhizobium agent of Rhizobium sp. No. 78 at 9.75×10 6 CFU / mL was applied to each Glycyrrhiza uralensis Fisch. plant, with the application amount of the rhizobium agent treatment reaching 9.75×10

[0053] SeIV treatment group: Each Glycyrrhiza uralensis Fisch. plant was irrigated with 50 mL of sodium selenite solution at a concentration of 4 mg / mL (the solvent was water), so that the application amount of sodium selenite treatment for each Glycyrrhiza uralensis Fisch. plant reached 200 mg (1156 µmol). Then, this irrigation was repeated once every 30 days for a total of 4 times to complete the pesticide application treatment.

[0054] SeIV+G treatment group: Each Glycyrrhiza uralensis Fisch. plant was irrigated with 25 mL of the rhizobium agent of Rhizobium sp. No. 78 with a content of 1.95×10 5 CFU / mL and 25 mL of sodium selenite solution with a concentration of 4 mg / mL, so that the application amount of Rhizobium sp. No. 78 for each Glycyrrhiza uralensis Fisch. plant reached 4.88×10 6 CFU, and the application amount of sodium selenite reached 100 mg (578 µmol). Then, this irrigation was repeated once every 30 days for a total of 4 times to complete the pesticide application treatment.

[0055] Nano treatment group: Each Glycyrrhiza uralensis Fisch. plant was irrigated with 50 mL of nano-selenium solution at a concentration of 4 mg / mL, so that the application amount of nano-selenium treatment for each Glycyrrhiza uralensis Fisch. plant reached 200 mg (2532 µmol) of nano-selenium. Then, this irrigation was repeated once every 30 days for a total of 4 times to complete the pesticide application treatment.

[0056] Nano+G treatment group: Each Glycyrrhiza uralensis Fisch. plant was irrigated with 25 mL of the rhizobium agent of Rhizobium sp. No. 78 with a content of 1.95×10 5 CFU / mL and 25 mL of nano-selenium solution with a concentration of 4 mg / mL, so that the application amount of Rhizobium sp. No. 78 for each Glycyrrhiza uralensis Fisch. plant reached 4.88×10 6 CFU, and the application amount of nano-selenium reached 100 mg (1266 µmol). Then, this irrigation was repeated once every 30 days for a total of 4 times to complete the pesticide application treatment.

[0057] On October 1, 2024, the following index detections were carried out.

[0058] 3 Effects of different treatments on selenium content in Glycyrrhiza uralensis Fisch. The selenium content in Glycyrrhiza uralensis Fisch. was determined according to the "National Food Safety Standard Determination of Selenium in Foods" (GB 5009.93—2017). The effects of different treatments on the selenium content in Glycyrrhiza uralensis Fisch. were as follows Figure 1As shown in the figure. The selenium contents of the CK group and the G78 group were 0.094 mg / kg and 0.092 mg / kg respectively. Since no exogenous selenium was applied, licorice only enriched the selenium in the soil, resulting in a low selenium content. For the SeIV group and the SeIV+G group of licorice treated with the commonly used exogenous selenium sodium selenite in selenium-rich agricultural products, the selenium contents were 2.86 mg / kg and 3.01 mg / kg respectively, which were approximately 30.43 times and 32.06 times that of the CK group respectively, and the difference was extremely significant (P<0.01). As a non-toxic exogenous selenium, nano-selenium was not as effective as sodium selenite in enriching selenium in licorice. The content of the Nano group was 0.155 mg / kg, which was approximately 1.65 times that of the CK group, but there was no significant difference ( P> 0.05). The content of the Nano+G group was 0.217 mg / kg, which was higher than that of the CK group and the difference was extremely significant (P<0.01), and it was approximately 2.3 times that of the CK group. For the treatment groups with co-application of exogenous selenium and rhizobia, the selenium contents of the corresponding single-application selenium groups were all increased. The selenium content of the SeIV+G group was increased by approximately 5.36% compared with the SeIV group, and the difference was extremely significant (P<0.01). The selenium content of the Nano+G group was increased by approximately 39.81% compared with the Nano group, but there was no significant difference ( P> 0.05). It can be seen from this that sodium selenite has a higher selenium enrichment efficiency as an exogenous selenium than nano-selenium. The reason may be that the sodium selenite solution is more easily infiltrated into the soil and absorbed and transformed by licorice. As a single selenium with a small particle size, although nano-selenium has low toxicity, its transformation efficiency and absorption efficiency are both lower than those of sodium selenite, resulting in a smaller selenium enrichment effect than sodium selenite. Rhizobia played a promoting role in the absorption and transformation of both sodium selenite and nano-selenium.

[0059] 4. Effects of different treatments on the crude fat content in licorice Determined by the Soxhlet extraction method (Chen Xiao, Yi Xueping, Ni Sui. Optimization study on the process of extracting peony seed oil by Soxhlet extraction method [J]. Chinese Wild Plant Resources, 2020, 39(05):29-33, 37.). Weigh 2 g of the uniformly mixed sample, accurate to 0.001 g, and put it into a filter paper tube. Then put the filter paper tube into the extraction tube of the fat analyzer. Its lower end is connected to a receiving bottle dried to a constant weight. Add petroleum ether to two-thirds of the volume above the condenser of the extractor, and carry out water bath heating to make the petroleum ether continuously reflux and extract (6 times / h - 8 times / h). After 7 hours of extraction, it ends. Use a glass rod to pick up a drop of the extract. If there is no oil spot, it indicates that the extraction is complete. Then take down the receiving bottle. When the solvent in it remains 2 ml, place it on a water bath to evaporate to dryness. After drying for 1 h, place it in a desiccator to cool for 30 min, and weigh it.

[0060] The fat in licorice plays a role in storing energy substances and protection. It is also a good solvent and storage depot for active substances such as flavonoids, regulating the anabolic metabolism of other medicinal components. Therefore, the fat content in licorice can indirectly evaluate the quality of licorice. The effects of different treatments on the fat content in licorice are as Figure 2 shown. The fat contents of the CK group, G78 group, SeIV group, SeIV+G group, Nano group, and Nano+G group are 1.18%, 1.55%, 1.91%, 2.35%, 2.55%, and 1.85% respectively. From the data, it can be seen that G78 increased by about 30.81% compared with the CK group, and the difference was extremely significant ( P <0.01). The fat contents of the exogenous selenium single application groups, the SeIV group and the Nano group, increased by about 61.07% and 115.01% respectively compared with the CK group, and both differences were extremely significant ( P <0.01). The Nano group had a higher promotion effect on the fat content in licorice than the SeIV group. It is speculated that the non-toxic mildness of nano-selenium as exogenous selenium ensured the transformation stability of the fat synthesis pathway in licorice. The SeIV+G group in the co-application group increased by about 23.28% compared with its corresponding single application group, the SeIV group, and the difference was extremely significant ( P <0.01), indicating that the co-application of Rhizobium sp. No. 78 and sodium selenite had a positive effect on the fat content in licorice. The fat content of the Nano+G group in the co-application group decreased by about 27.29% compared with its corresponding single application group, the Nano group, which was opposite to the obvious promotion effect of the SeIV+G group. The specific reason is unknown. It is speculated that the microscopic transformation path of rhizobium and nano-selenium coincides with the fat transformation, resulting in an inhibitory effect.

[0061] 5. Effects of different treatments on the protein content in licorice Determined by the Kjeldahl method (Sun Yujie, Chen Lu, Ye Shanrong, et al. Research on the method for determining the total nitrogen content in plant samples [J]. Shandong Chemical Industry, 2024, 53(15): 199-201, 215.). Weigh 1 g of the solid sample to be measured, accurate to 0.001 g, transfer it into a dry 250 ml Kjeldahl flask, add 0.4 g of copper sulfate, 6 g of potassium sulfate, and 20 mL of concentrated sulfuric acid, shake gently, then place a small funnel on the bottle mouth, and tilt the bottle body at 45° and place it on a closed small electric furnace with an asbestos net. Heat it in a fume hood until it boils continuously for nitrification for 1.5 h, showing a blue-green color. Take it out and let it cool (if it shows a blue-green color after cooling, the nitrification is complete; if it returns to a black-brown color after cooling, the nitrification is not complete, then put it back into the closed electric furnace and continue heating for nitrification until it is fully nitrified). Then add 20 mL of distilled water, transfer it to a 100 mL volumetric flask after cooling, and make up the volume to the scale line with distilled water, and shake well. At the same time, do a reagent blank.

[0062] Set up the nitrogen distillation device. Add water to the steam generator up to 2 / 3 of its volume, add 5 glass beads, add 2 mL of concentrated sulfuric acid and a few drops of methylated ethanol solution until it turns purplish-red to keep the aqueous solution acidic, and carefully heat the water in the steam generator until it boils continuously. Then add 10 mL of boric acid to the receiving flask, add 1 - 2 drops of Indicator A (10 mL of methyl red and 5 mL of methylene blue), insert the lower end of the condenser below the liquid level in the receiving flask, and clamp it with a clip. Inject 5 mL of the test sample into the reaction chamber, rinse it with 10 mL of distilled water and then inject it into the reaction chamber as well. Then add 10 mL of NaOH solution, stopper the upper end with a glass rod-shaped stopper, seal it with water, and clamp it with a clip to carry out distillation. After distilling for 10 minutes, remove the conical flask and titrate it with 0.05 mol / L H2SO4 until the color turns grayish-blue as the end-point color, record the volume of H2SO4 consumed, and calculate the protein content based on this.

[0063] Proteins can regulate immune functions and physiological activities such as photosynthesis during the growth process of licorice. Moreover, after exogenous selenium is absorbed and transformed by licorice, it is stored in licorice in the form of organic selenium proteins. Therefore, the protein content in licorice is one of the indicators to measure the quality of licorice and the effect of selenium enrichment. The effects of different treatments on the protein content in licorice are as Figure 3 shown. The protein content in the G78 group is 5.04%, and the protein content in the CK group is 3.77%. The protein content in the G78 group increased by 33.73% compared with the CK group, and the difference was extremely significant (P < 0.01). The protein contents of the four groups of licorice with added exogenous selenium also increased to varying degrees compared with the CK group, and all had extremely significant differences compared with the CK group ( P <0.01). The protein contents in the two groups of licorice with co-application of rhizobia and exogenous selenium were also greater than those in the corresponding groups with only selenium application. The protein contents in the SeIV+g group and the SeIV group were 4.67% and 4.09% respectively. The protein content in the group with rhizobia application increased by 14.25%, and the difference between the two groups was extremely significant (P < 0.01). The protein contents in the Nano+G group and the Nano group were 4.73% and 4.47% respectively. The protein content in the group with rhizobia application increased by 5.64%, and the difference was extremely significant (P < 0.01). The protein content in the Nano group increased by 9.36% compared with the SeIV group, with an extremely significant difference ( P <0.01). The protein content in the corresponding group with co-application of rhizobia increased by 1.11%, but there was no significant difference ( P>0.05). The data indicate that rhizobia can effectively promote the absorption and transformation of proteins by Glycyrrhiza uralensis Fisch. The nodulation and nitrogen fixation of rhizobia can convert atmospheric nitrogen into ammonia useful for plants, which is then converted by symbiotic plants into nutrients such as amino acids, and further into proteins, increasing the protein content, meeting the growth requirements of plants and improving the quality. Exogenous selenium promotes the conversion of inorganic selenium by Glycyrrhiza uralensis Fisch. into organic non-toxic selenoprotein and stores it in the plant body, thereby increasing the total protein content of Glycyrrhiza uralensis Fisch. In terms of enhancing the protein content, sodium selenite is stronger than nano-selenium. It is speculated that this may be because sodium selenite, as exogenous selenium, has a better effect on enriching selenium in Glycyrrhiza uralensis Fisch., resulting in a high content of selenoprotein.

[0064] 6. Effects of Different Treatments on the Flavonoid Content in Glycyrrhiza uralensis Fisch. Determined by ultraviolet visible-spectrophotometry (Zhu Yichao, Shang Qingwen. Determination of the Total Flavonoid Content in Chrysanthemum morifolium Ramat. cv. Taoju by Ultraviolet Spectrophotometry [J]. Special Economic Animal and Plant, 2024, 27(11): 11-13.). Weigh 0.3 g of the sample powder, accurate to 0.001 g. Add 50 mL of 60% ethanol solution, then perform ultrasonic treatment for 90 min, filter by suction, shake the test solution well, pour the filtrate into a 50 mL round-bottom centrifuge tube, and make up the volume to 50 mL with 60% ethanol solution, and shake well. Pipette 1 mL of the filtrate into a 25 mL volumetric flask, add 1 mL of 5% sodium nitrite solution, shake well, let it stand for 6 min, then add 1 mL of 10% aluminum nitrate solution, shake well, let it stand for 6 min, then add 10 mL of 1 mol / L NaOH solution, shake well, make up the volume to the mark with 30% ethanol solution, shake well, let it stand for 15 min, and then obtain the crude extract of flavonoids. Measure the absorbance X at a wavelength of 510 nm and record it. Calculate the flavonoid content in 6 kinds of Glycyrrhiza uralensis Fisch. powder according to the absorbance and the measured standard curve.

[0065] Flavonoid is one of the important active ingredients in Glycyrrhiza uralensis Fisch. Flavonoid is closely related to the anti-inflammatory and antibacterial effects of Glycyrrhiza uralensis Fisch. etc. Through the standard curve of the flavonoid standard product (Y = 13.5x + 0.021, R 2 = 0.9992), the flavonoid content of each treatment group can be obtained. The effects of different treatments on the flavonoid content in Glycyrrhiza uralensis Fisch. are as Figure 4As shown. The flavonoid contents of the CK and G78 groups were 27.89 mg / g and 36.94 mg / g, respectively. The flavonoid content of the G78 group increased by approximately 32.44% compared with the CK group, and the difference was extremely significant (P < 0.01). This shows that rhizobium treatment can increase the flavonoid content of licorice. The flavonoid contents of the SeIV and Nano groups treated with exogenous selenium were 31.24 mg / g and 31.89 mg / g, respectively, which were approximately 12.01% and 14.37% higher than those of the CK group, respectively. Both showed extremely significant differences (P < 0.01). This shows that the application of exogenous selenium can increase the flavonoid content of licorice. The flavonoid content of the corresponding co-application groups was also increased compared with the single application groups. The SeIV+G group increased by about 17.43% compared with the SeIV group, and the difference was extremely significant (P<0.01). The Nano+G group increased by about 4.13% compared with Nano, but there was no significant difference (P>0.05). When rhizobia and exogenous selenium were co-applied, rhizobia could enrich licorice with selenium and increase the flavonoid content at the same time. It may be that the microbial environment of the soil changed during co-application, which adjusted the soil environment for licorice growth and increased the flavonoid content. It may also be that the symbiotic nitrogen fixation of rhizobia and the enrichment of selenium by licorice adjusted the secondary metabolism and enzyme content of licorice, thereby improving the synthesis efficiency of flavonoids.

[0066] 7. Effects of different treatments on the polysaccharide content in licorice The polysaccharide was measured by the phenol-concentrated sulfuric acid method (Gao Xin, Guo Yimei, Zhou Jun, et al. Study on the determination of polysaccharide content in red ginseng by the phenol-concentrated sulfuric acid method [J]. Laboratory Science, 2018, 21(01): 29-30.). 1 g of sample powder was weighed to the nearest 0.001 g and placed in a 50 mL conical flask. 30 mL of distilled water was added and the mixture was placed in a 90°C water bath for 90 min. After shaking, the mixture was poured into a 15 mL centrifuge tube and centrifuged at 5000 r for 5 min. 0.01 mL of the supernatant was taken and placed in a test tube. 1 mL of 5% phenol solution was added and then 6 mL of concentrated sulfuric acid was slowly added while shaking. The mixture was thoroughly mixed and cooled. After 30 min, the absorbance was measured at a wavelength of 490 nm. The polysaccharide content in the treated licorice powder was calculated based on the polysaccharide standard curve and the absorbance.

[0067] The polysaccharide in licorice regulates its pharmacological effects such as immune regulation and antioxidant, and is one of the important active ingredients in licorice. The standard curve of glucose standard (Y=69.829x+0.0313, R 2, =0.9994) to obtain the polysaccharide content of each treatment group. The effects of different treatments on the polysaccharide content in licorice are as follows: Figure 5As shown in the figure. Comparing the polysaccharide contents of the CK group and the G78 group, the polysaccharide contents of the CK group and the G78 group were 106.99 mg / g and 148.77 mg / g respectively. The polysaccharide content of the G78 group increased by about 39.04% compared with the CK group, and the difference was extremely significant (P<0.01). Rhizobium No. 78 could significantly increase the polysaccharide content in licorice. The polysaccharide contents of the single exogenous selenium treatment groups, the SeIV group and the Nano group, were 154.13 mg / g and 177.45 mg / g respectively, which were about 44.06% and 7.03% higher than that of the CK group respectively. The increase in the polysaccharide content of the SeIV group was significantly different (P<0.01), while there was no significant difference in the Nano group (P>0.05). Analysis showed that sodium selenite as an exogenous selenium treatment could significantly increase the polysaccharide content in licorice. It was speculated that inorganic selenium might be stored in licorice in the form of selenium polysaccharide after being transformed by licorice, resulting in a significant increase in the polysaccharide content. As an exogenous selenium, nano-selenium had a lower effect on selenium enrichment in licorice, so the polysaccharide content in licorice was lower than that in the sodium selenite treatment group. Comparing the co-application group and the single-application group, the polysaccharide content of the SeIV+G group was 177.45 mg / g, which was 15.13% higher than that of the SeIV group after the co-application of rhizobium, with a significant difference (P<0.05). The polysaccharide content of the Nano+G group was 177.45 mg / g, which was 37.19% higher than that of the Nano group, with an extremely significant difference (P<0.01). It can be seen from this that the synergistic effect of rhizobium in the co-application group could significantly increase the polysaccharide content in licorice. It was speculated that it was related to the fact that rhizobium could promote the absorption and transformation of exogenous selenium, and exogenous selenium was further transformed into organic selenium such as selenium polysaccharide and other substances stored in licorice, resulting in an increase in the licorice polysaccharide content.

[0068] 8. Effects of Different Treatments on the Saponin Content in Licorice It was determined by the vanillin-perchloric acid method (Liu Zhen, Wang Zhunian, Wang Maoyuan, et al. Determination of the Total Saponin Content and Antioxidant Activity in the Roots of Millettia speciosa Champ. with Different Growth Years [J]. Journal of Tropical Biology, 2025, 1(03): 1-11.). 0.1 g of the well-mixed sample was weighed accurately to 0.001 g. Then 20 mL of 80% ethanol solution was added, and after ultrasonic treatment for 70 min, it was filtered. The filtrate was poured into a 30 mL round-bottom centrifuge tube, and the 80% ethanol solution was added dropwise to 20 mL. 0.5 mL of the test solution was accurately pipetted into a 25 mL beaker, and the solvent was evaporated in a water bath at 60℃. Then 0.2 mL of 5% vanillin-glacial acetic acid solution was added to dissolve it. After that, 0.8 mL of perchloric acid solution was added, and it was heated in a water bath at 60℃ for 20 min. It was immediately taken out and placed in an ice-water bath for 5 min. Finally, 5 mL of glacial acetic acid solution was added, and the absorbance was measured at 545 nm. The saponin content in the licorice powder was calculated based on the standard curve of saponin and the absorbance.

[0069] Saponins are one of the active ingredients with relatively high content in licorice. Saponins are closely related to the anti-inflammatory and antiviral effects of licorice and are one of the important indicators for evaluating the quality of licorice. Through the standard curve of oleanolic acid reference substance (Y = 90.978x + 0.0265, R 2 = 0.9992), the saponin contents of each treatment group can be obtained. The effects of different treatments on the saponin content in licorice are as Figure 6 shown. Comparing the saponin contents of the CK group and the G78 group, the saponin contents of the CK group and the G78 group are 0.96 mg / g and 69.17 mg / g respectively. The saponin content of the G78 group is about 22.6% higher than that of the CK group, and the difference is extremely significant (P < 0.01). The data indicate that Rhizobium sp. No. 78 can significantly increase the saponin content in licorice. The saponin contents of the single exogenous selenium treatment groups, the SeIV group and the Nano group, are 56.9639 mg / g and 68.65 mg / g respectively, which are about 0.96% and 21.67% higher than that of the CK group respectively. The increase in the saponin content of the SeIV group is extremely small and there is no significant difference (P > 0.05). The increase in the saponin content of the nano-selenium group is obvious and the difference is extremely significant (P < 0.01). It shows that as an exogenous selenium, nano-selenium can significantly increase the saponin content in licorice. However, sodium selenite does not have a significant effect on increasing the saponin content. Comparing the co-application group and the single-application group, the saponin content of the SeIV + G group is 58.81 mg / g, and the content increases by about 3.25% after the co-application of rhizobium, and there is no significant difference (P > 0.05). The saponin content of the Nano + G group is 71.87 mg / g, and the content increases by about 4.69%, and there is a significant difference (P < 0.05). It can be seen that the co-application group of rhizobium and exogenous selenium can slightly increase the saponin content in licorice, but the effect is not obvious, and there is a significant difference in the promotion effect between the co-application of nano-selenium and rhizobium.

[0070] 9. Effects of different treatments on glycyrrhizic acid content and liquiritin content in licorice The contents of liquiritin and glycyrrhizic acid in licorice powder were determined by high performance liquid chromatography (Yin Xingzhang, Jiang Daiquan. Simultaneous determination of benzoic acid and glycyrrhizic acid in compound glycyrrhiza mixture by HPLC [J]. Yunnan Chemical Technology, 2024, 51(09): 96 - 99.). Acetonitrile was used as mobile phase A, and 0.05% phosphoric acid solution was used as mobile phase B. Gradient elution was carried out according to the regulations in Table 1. The detection wavelength was set at 237 nm, and the time was 60 min. The contents of different concentration standard solutions of glycyrrhizic acid and liquiritin, the contents of glycyrrhizic acid and liquiritin in 6 samples to be tested, and the contents of glycyrrhizic acid and liquiritin in the reference substances were compared and analyzed to determine the contents in the samples to be tested. Among them, the glycyrrhizic acid used as the reference substance is the compound with CAS: 1405 - 86 - 3, and the liquiritin is the compound with CAS: 551 - 15 - 5.

[0071] Table 1 Gradient elution settings

[0072] Preparation of the standard solution: Accurately weigh 0.002 g of the reference substance powders of glycyrrhizic acid and liquiritin respectively, place them in a 10 mL volumetric flask, add 70% ethanol solution, and make up to the scale line to prepare a solution containing 20 μg of liquiritin and 0.2 mg of glycyrrhizic acid per 1 mL, that is, the standard solution is obtained. The chromatogram of the reference substance is as Figure 7 shown.

[0073] Preparation of the test solution: Weigh accurately 0.2 g of the sample powder to be tested to 0.001 g, place it in a stoppered conical flask, accurately add 100 mL of 70% ethanol solution, then filter by suction, and take the subsequent filtrate for sample treatment. Then, add the sample, pipette 10 μL of the sample to be tested for injection, measure the absorbance of the sample to be tested, perform gradient elution using the gradient elution setting table in Table 1 above, the detection wavelength of the detector is 237 nm, the running time is 60 min, the flow rate is 1 mL / min, collect the baseline, compare the results with the standards of glycyrrhizic acid and liquiritin, and analyze the results. The chromatogram of the test sample (sample) is as Figure 8 shown.

[0074] Glycyrrhizic acid is a special active ingredient in licorice, which has various functions such as anti-tumor, anti-inflammatory, and immunomodulatory effects. It is a sweet substance in licorice and is also one of the indicators to measure whether licorice meets the medicinal material standards. Through the standard curve of the glycyrrhizic acid reference substance (y = 973478x + 133.82, R 2 = 0.9953), the glycyrrhizic acid content of each treatment group can be obtained. The effects of different treatments on the glycyrrhizic acid content in licorice are as Figure 9As shown in the figure. Comparing the glycyrrhizic acid content of the CK group and the G78 group, the glycyrrhizic acid contents of the CK group and the G78 group were 2.03% and 2.39% respectively. The glycyrrhizic acid content of the G78 group increased by about 17.56% compared with the CK group, showing a highly significant difference (P<0.01). The above data indicate that irrigating the roots with Rhizobium sp. No. 78 can significantly increase the glycyrrhizic acid content of licorice. The glycyrrhizic acid contents of the SeIV group and the Nano group in the single exogenous selenium treatment group were 2.09% and 2.27% respectively, which were about 2.91% and 11.76% higher than that of the CK group respectively. The increase in the glycyrrhizic acid content of the SeIV group was not obvious and the difference was not significant (P>0.05), while the increase in the nano-selenium group was obvious and showed a highly significant difference (P<0.01). It can be analyzed that nano-selenium as an exogenous selenium treatment can significantly increase the glycyrrhizic acid content in licorice. Comparing the co-application group and the single-application group, the glycyrrhizic acid content of the SeIV+G group was 2.02%, and the content decreased by about 3.38% after the co-application of rhizobia, without a significant difference (P>0.05). The glycyrrhizic acid content of the Nano+G group was 2.05%, and the content decreased by about 11.08%, showing a highly significant difference (P<0.01). It can be seen that the synergistic effect of rhizobia in the co-application group will slightly reduce the glycyrrhizic acid content in licorice. The Chinese Pharmacopoeia stipulates that the glycyrrhizic acid content of dried licorice should not be less than 2.0%. All of the above six groups meet the national regulations.

[0075] Liquiritin is a special active ingredient present in licorice, which has various functions such as antioxidant and anti-inflammatory effects, and is also one of the indicators to measure whether licorice meets the standards. Through the standard curve of the liquiritin standard (Y = 240600x + 91.54, R 2 = 0.9947), the liquiritin contents of each treatment group can be obtained. The effects of different treatments on the liquiritin content in licorice are as follows Figure 10As shown in the figure. Comparing the liquiritin contents of the CK group and the G78 group, the liquiritin contents of the CK group and the G78 group were 0.58% and 0.73% respectively. The liquiritin content of the G78 group increased by about 24.46% compared with the CK group, showing a highly significant difference (P<0.01). The above data indicate that irrigating the roots with Rhizobium No. 78 can significantly increase the liquiritin content of Glycyrrhiza uralensis. The liquiritin contents of the SeIV group and the Nano group in the single exogenous selenium treatment group were 0.67% and 0.56% respectively, which were about 14.92% and 3.39% higher than that of the CK group respectively. The increase in the liquiritin content of the SeIV group was obvious, with a highly significant difference (P<0.01); there was no significant difference in the Nano group (P>0.05). It can be analyzed that sodium selenite as an exogenous selenium treatment can significantly increase the liquiritin content in Glycyrrhiza uralensis. It is speculated that it may be the effect of the conversion of inorganic selenium by Glycyrrhiza uralensis that leads to a significant increase in the liquiritin content. While nano-selenium, as a stable exogenous selenium, has a low bioavailability and a low effect of enriching selenium in Glycyrrhiza uralensis, so the liquiritin content in Glycyrrhiza uralensis is lower than that of the sodium selenite treatment group. Comparing the co-application group and the single-application group, the liquiritin content of the SeIV+G group was 0.79%. After the co-application of rhizobia, the liquiritin content increased by about 17.63% compared with the SeIV group, showing a highly significant difference (P<0.01). The liquiritin content of the Nano+G group was 0.62%, which was about 10.71% higher than that of the Nano group, also showing a highly significant difference (P<0.01). It can be seen that the synergistic effect of rhizobia in the co-application group can significantly increase the liquiritin content in Glycyrrhiza uralensis. It is speculated that Rhizobium No. 78 can not only promote the absorption and conversion of exogenous selenium, but also significantly increase the liquiritin content in Glycyrrhiza uralensis. The Chinese Pharmacopoeia stipulates that the liquiritin content of the dried product of Glycyrrhiza uralensis shall not be less than 0.5%. All the above six groups meet the national regulations.

[0076] Further correlation analysis of the main components in Glycyrrhiza uralensis, the results are as Figure 11 shown. Selenium showed a highly significant strong positive correlation with fat and protein. As the selenium content increased, the fat and protein contents were likely to increase accordingly. This association may be because they have a synergistic effect in certain physiological processes or substance synthesis pathways. During the growth of Glycyrrhiza uralensis, selenium may participate in regulating the metabolic pathways related to the synthesis of fat and protein. There was a significant or highly significant positive correlation between protein and flavonoid and polysaccharide. It shows that the content changes of protein and these two components in the samples have a certain synchrony. It may be that during the growth of Glycyrrhiza uralensis, protein may show specific pharmacological activities in synergy with flavonoid and polysaccharide.

[0077] The present invention has been described in detail above. For those skilled in the art, without departing from the spirit and scope of the present invention and without the need for unnecessary experiments, the present invention can be implemented within a relatively wide range under equivalent parameters, concentrations, and conditions. Although specific embodiments of the present invention are given, it should be understood that the present invention can be further improved. In short, according to the principle of the present invention, this application intends to cover any modifications, uses, or improvements to the present invention, including changes made using conventional techniques known in the art that depart from the scope disclosed in this application.

Claims

1. A composition, characterized in that, The composition contains rhizobia and selenite, and the rhizobia are Mesorhizobium temperatum ( Mesorhizobium temperatum ).

2. The composition according to claim 1, wherein The ratio of rhizobia to selenite in the composition is 4.88×10 6 CFU rhizobia: 578 µmol selenite.

3. The composition according to claim 1 or 2, characterized in that, The composition has at least one of the following properties: A1) Increasing the liquiritin content in licorice; A2) Increasing the polysaccharide content in licorice; A3) Increasing the selenium content in licorice; A4) Increasing the fat content in licorice; A5) Increasing the protein content in licorice; A6) Increasing the flavonoid content in licorice; A7) Increasing the saponin content in licorice; A8) Increasing the glycyrrhizic acid content in licorice.

4. The composition according to any one of claims 1-3, characterized in that: The composition is a microbial inoculum or a fertilizer.

5. Use of the composition according to any one of claims 1-4 in increasing the content of active ingredients in licorice, wherein the active ingredient is at least one of the following: B1) Liquiritin; B2) Polysaccharide; B3) Selenium; B4) Fat; B5) Protein; B6) Flavonoid; B7) Saponin; B8) Glycyrrhizic acid.

6. Use of the composition according to any one of claims 1-4 in the preparation of a product for increasing the content of active ingredients in licorice, wherein the active ingredient is at least one of the following: C1) Liquiritin; C2) Polysaccharide; C3) Selenium; C4) Fat; C5) Protein; C6) Flavonoid; C7) Saponin; C8) Glycyrrhizic acid.

7. A method for increasing the content of active ingredients in licorice, characterized in that, The method includes the step of treating licorice with the composition according to any one of claims 1-4 to increase the content of active ingredients in licorice, wherein the active ingredient is at least one of the following: D1) Liquiritin; D2) Polysaccharide; D3) Selenium; D4) Fat; D5) Protein; D6) Flavonoid; D7) Saponin; D8) Glycyrrhizic acid.

8. A method for preparing a product with increased content of active ingredients of licorice, characterized in that, The method includes the step of using the composition according to any one of claims 1-4 as a product ingredient, wherein the active ingredient is at least one of the following: E1) Liquiritin; E2) Polysaccharide; E3) Selenium; E4) Fat; E5) Protein; E6) Flavonoid; E7) Saponin; E8) Glycyrrhizic acid.

9. A method for preparing a composition, characterized in that, The composition is the composition according to any one of claims 1-4, and the method includes the step of using the rhizobia and selenite described in claim 1 as composition ingredients.

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