Mucosal mucilaginous lactobacillus ccfm1490 transformed with cassia glycosides and enhancing its efficacy in relieving liver lipid accumulation and application thereof
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- JIANGNAN UNIV
- Filing Date
- 2026-03-03
- Publication Date
- 2026-06-09
Smart Images
Figure CN122168464A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to a strain of *Lactobacillus mucosae* CCFM1490 that is transformed with cassiaside and enhanced to alleviate lipid accumulation in the liver, and its application, belonging to the fields of microbial technology and pharmaceutical technology. Background Technology
[0002] Nonalcoholic fatty liver disease (NAFLD) has become one of the most common chronic liver diseases worldwide, characterized by excessive accumulation of lipids in the liver (hepatic steatosis), which can progress to hepatitis, liver fibrosis, and even cirrhosis in severe cases. Its occurrence is closely related to obesity, insulin resistance, and lipid metabolism disorders. Currently, there are no specific therapeutic drugs, and management is mainly achieved through lifestyle interventions. Therefore, developing safe and effective adjunctive strategies to alleviate hepatic lipid accumulation has significant clinical and social implications.
[0003] Cassia seed, a traditional Chinese medicine, has been proven to possess pharmacological activities such as lipid-lowering and liver-protecting effects. Cassia glycosides are one of its main active ingredients; however, their glycoside content leads to low bioavailability, requiring metabolism by intestinal flora to convert them into cassia aglycones before they can be better absorbed and exert their effects. However, individual differences in intestinal flora structure result in significant variations in the metabolic capacity of cassia glycosides and the resulting health benefits among different populations, greatly limiting the stability of the efficacy of cassia seed products.
[0004] Utilizing specific probiotics for in vitro fermentation of cassia seeds to pre-convert them into cassia aglycones is an effective strategy to enhance their efficacy. Currently, some microorganisms capable of converting glycosides have been reported, but none have been reported to convert cassia aglycones. Furthermore, existing research largely focuses on the conversion efficiency itself or only on its general lipid-lowering effects. The liver is the core organ for lipid metabolism in the body, but probiotic strains that can specifically enhance the liver's lipid accumulation-relieving function of cassia seeds are currently lacking.
[0005] Mucosal lactobacilli ( Limosilactobacillus mucosae Lactobacillus mucosa is a common commensal bacterium in the gut, attracting attention due to its excellent intestinal colonization ability and immunomodulatory function. However, to date, no Lactobacillus mucosa has been reported to be able to enhance the regulatory effect of hepatic lipid metabolism by converting cassiaside.
[0006] Therefore, there is an urgent need in this field for a dedicated strain capable of efficiently converting cassiaside and significantly enhancing its targeting of the liver, alleviating hepatic steatosis, and protecting hepatocyte function. Such a strain would have significant clinical application value for developing next-generation probiotics or functional products targeting NAFLD and related liver diseases. Summary of the Invention
[0007] To address the shortcomings and deficiencies of the existing technology, this invention provides *Lactobacillus mucosae* (…). Limosilactobacillus mucosae The application of CCFM1490 in products that enhance the efficacy of cassiaside in alleviating hepatic lipid accumulation aims to address the current lack of a dedicated strain capable of efficiently converting cassiaside and significantly enhancing its targeting of the liver, alleviating hepatic steatosis, and protecting hepatocyte function.
[0008] The first technical solution provided by this invention is a strain of *Lactobacillus mucosae* (…). Limosilactobacillus mucosae CCFM1490 was deposited at the Guangdong Provincial Center for Microbial Culture Collection on April 11, 2025, with accession number GDMCC NO: 66137.
[0009] In one embodiment of the present invention, after culturing the *Lactobacillus mucosa* CCFM1490 on MRS medium for 48 h, the colonies are round, convex, or lenticular, slightly white, opaque, and have a smooth to mucous-like soft surface. Fermentation of cassiaside using this bacterium can significantly increase the content of cassiaside aglycone, thereby enhancing the effect of cassiaside in alleviating liver lipid accumulation.
[0010] The second technical solution provided by the present invention is a microbial preparation containing the *Lactobacillus mucosae* CCFM1490 described in the first technical solution.
[0011] In one embodiment of the present invention, the concentration of *Lactobacillus mucosa* CCFM1490 in the microbial preparation is at least 1 × 10⁻⁶. 6 CFU / mL or 1×10 6 CFU / g.
[0012] In one embodiment of the present invention, the concentration of *Lactobacillus mucosa* CCFM1490 in the microbial preparation is at least 1 × 10⁻⁶. 9 CFU / mL or 1×10 9 CFU / g.
[0013] The third technical solution provided by the present invention is a synbiotic preparation containing live bacteria of Lactobacillus mucosa CCFM1490 and cassia seed glycosides as described in the first technical solution.
[0014] The fourth technical solution provided by the present invention is an epigenetic agent, wherein the epigenetic agent is the fermentation broth of the *Lactobacillus mucosa* CCFM1490 described in the first technical solution or the microbial preparation described in the second technical solution, which is fermented in a fermentation system containing cassiaside and then subjected to heat treatment and high-pressure homogenization.
[0015] The present invention also provides a method for preparing the postgenetic agent, comprising the following steps: (1) Activate the mucosal lactobacillus CCFM1490 to obtain a bacterial solution; (2) Inoculate the bacterial solution of (1) into the fermentation substrate containing cassia glycosides and carry out fermentation; (3) After fermentation is completed, the fermentation liquid is heat-treated and then homogenized under high pressure to obtain the final product.
[0016] In one embodiment of the present invention, step (1) involves streaking *Lactobacillus mucosae* CCFM1490 onto MRS solid medium, inverting the plate for culture (preferably inverting at 37°C for 48 hours), and then picking a single colony and inoculating it into MRS liquid medium (preferably 5 mL) for culture (preferably incubating at 37°C for 48 hours) to obtain a bacterial solution.
[0017] In one embodiment of the present invention, the concentration of cassiaside in the fermentation substrate containing cassiaside in step (2) is 500-1000 mg / L (preferably 500 mg / L).
[0018] In one embodiment of the present invention, the inoculation amount of bacterial solution in step (2) is 2-8% (v / v).
[0019] In one embodiment of the present invention, the fermentation in step (2) is a constant temperature fermentation at 30-40℃ for 36-72 hours (preferably a constant temperature fermentation at 37℃ for 48 hours).
[0020] In one embodiment of the present invention, the heat treatment conditions in step (3) are 60-70°C for 25-35 min (preferably 65°C for 30 min).
[0021] In one embodiment of the present invention, the high-pressure homogenization conditions in step (3) are 300 to 1500 Bar, and the bacterial strain is circulated 4 to 8 times (preferably 1200 Bar, 6 times).
[0022] The fifth technical solution provided by the present invention is the application of the *Lactobacillus mucosae* CCFM1490 described in the first technical solution or the microbial preparation described in the second technical solution in the preparation of products for converting cassiaside.
[0023] In one embodiment of the present invention, the product is a pharmaceutical product.
[0024] The sixth technical solution provided by the present invention is a method for converting cassiaside, which uses the *Lactobacillus mucosae* CCFM1490 described in the first technical solution or the microbial preparation described in the second technical solution to ferment a substrate containing cassiaside.
[0025] In one embodiment of the present invention, the fermentation is carried out at 30-40°C for 36-72 hours; the concentration of cassiaside is 0.5-1 mg / mL; and the inoculum size is 2-8% (v / v).
[0026] The seventh technical solution provided by this invention is a method for enhancing the medicinal efficacy of cassiaside, wherein cassiaside is treated with any of the following methods: (1) Mix cassia glycoside with the *Lactobacillus mucosae* CCFM1490 described in the first technical solution; (2) Add cassiaside to the fermentation medium of Lactobacillus mucosa CCFM1490 as described in the first technical solution for culturing.
[0027] The eighth technical solution provided by the present invention is the application of the *Lactobacillus mucosa* CCFM1490 described in the first technical solution, the microbial preparation described in the second technical solution, the synbiotic preparation described in the third technical solution, or the postbiotic described in the fourth technical solution in the preparation of a drug for lowering blood lipids and / or alleviating non-alcoholic fatty liver disease.
[0028] In one embodiment of the present invention, the product includes at least one of the following functions: (a) Reduce serum triglyceride levels; (b) Reduce serum total cholesterol levels; (c) Reduce serum alanine aminotransferase and / or aspartate aminotransferase levels; (d) Improves fatty degeneration of liver tissue.
[0029] In one embodiment of the present invention, the product contains *Lactobacillus mucosa* CCFM1490, a carrier, and / or excipients.
[0030] The ninth technical solution provided by this invention is the application of the *Lactobacillus mucosa* CCFM1490 described in the first technical solution, the microbial preparation described in the second technical solution, the synbiotic preparation described in the third technical solution, or the postbiotic described in the fourth technical solution in the preparation of a drug for alleviating liver damage caused by a high-fat diet.
[0031] Compared with the prior art, the beneficial effects of the present invention are as follows: 1. The *Lactobacillus mucosae* CCFM1490 of the present invention can grow in a fermentation substrate rich in cassiaside, converting cassiaside into cassia aglycone.
[0032] 2. The synergistic preparation of *Lactobacillus mucosae* CCFM1490 and cassiaside, or the post-biotic prepared by fermentation in the presence of cassiaside, can promote the efficacy of cassiaside, thereby achieving the effects of lowering blood lipids, alleviating liver damage, and alleviating liver lipid accumulation.
[0033] Products that enhance the efficacy of alleviating liver lipid accumulation can be produced using *Lactobacillus mucosa* CCFM1490 and cassiaside. The production process uses cassiaside as a raw material to promote its efficacy. This process has the advantages of being safe, efficient, low-cost, and having a mild reaction, making it suitable for large-scale industrial production.
[0034] Preservation of biological materials Mucosal lactobacilli ( Limosilactobacillus mucosae (CCFM1490, taxonomically named) Limosilactobacillus mucosae It was deposited on April 11, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC No: 66137, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou. Attached Figure Description
[0035] Figure 1 High-performance liquid chromatography (HPLC) chromatograms of *Lactobacillus mucosae* CCFM1490 before and after fermentation.
[0036] Figure 2 The effects of CCFM1490 combined with cassiaside on obesity characteristics in mice; (A) pre-dissection body weight; (B) weight gain during intervention; (C) liver weight; (D) epididymal fat weight.
[0037] Figure 3 The effects of CCFM1490 combined with cassia seed glycoside on blood lipids in mice were as follows: (A) serum triglyceride (TG) content; (B) serum total cholesterol (TC) content; (C) serum low-density lipoprotein cholesterol (LDL-C) content; (D) serum high-density lipoprotein cholesterol (HDL-C) content; and (E) the ratio of LDL-C to HDL-C in serum.
[0038] Figure 4 Effects of CCFM1490 combined with cassiaside on liver injury indicators and liver pathological morphology in mice; serum (A) serum alanine aminotransferase (ALT) content; (B) serum aspartate aminotransferase (AST) content; (C) liver oil red section; (D) H&E section; scale bar: 100 μm.
[0039] Figure 5 This is a high-performance liquid chromatogram of cassia glycosides fermented by the comparative strain.
[0040] "*" indicates a significant difference from the Model group (p<0.05), "**" indicates p<0.01 from the Model group, and "***" indicates p<0.001 from the Model group. Detailed Implementation
[0041] The preferred embodiments of the present invention are described below. It should be understood that the embodiments are for better explanation of the present invention and are not intended to limit the present invention.
[0042] Test method: HPLC analysis of the content of cassiaside and cassia aglycone in the fermentation system: (1) Sample pretreatment: Cassia glycoside detection: After fermentation, the supernatant of the fermentation broth was collected by centrifugation. The supernatant was extracted with dichloromethane and ethyl acetate, respectively. The dichloromethane phase and ethyl acetate phase were evaporated by vacuum centrifugation to remove organic solvents. Then, chromatographic grade methanol was added for redissolution. Finally, the sample was filtered through a 0.22 μm organic filter membrane and then tested.
[0043] (2) Preparation of standard products Accurately weigh 10.0 mg of each cassiaside standard and dissolve it in methanol, then dilute to volume. Prepare a 1.0 mg / mL standard stock solution using methanol, and dilute it to concentrations of 20, 40, 80, 160, and 320 g / mL, respectively. Filter the solutions through a 0.22 μm filter membrane, and take 10 μL samples for HPLC analysis. Plot three standard curves with peak area (mAU) on the ordinate and cassiaside concentration (g / mL) on the abscissa.
[0044] (3) Sample testing Take 10 mL of the pretreated fermentation product (sample to be tested), filter it through a 0.45 μm membrane, and then dilute it to 100 mL with analytical grade 50% methanol solution. Inject the reference solution and the sample solution, and determine the content of cassiaside in the fermentation product under the above chromatographic conditions. Calculate the content using the external standard method.
[0045] The chromatographic conditions were as follows: Column: X Bridge RC18 (250×4.6mm, 5μm); Mobile phase: Water (pH 2.85 + 0.6% (v / v) acetic acid): Methanol = 19:81 (v / v); Detector: Ultraviolet detector (UV) 254nm; Injection volume: 10μL; Column temperature: 35℃; Elution conditions: Flow rate 1.0mL / min.
[0046] Raw materials used in the examples: The C57BL / 6J mice used in the following examples were purchased from Beijing Vital River Company.
[0047] The culture media involved in the following examples are as follows: MRS liquid culture medium: yeast extract 5.0 g / L, beef extract 10.0 g / L, peptone 10.0 g / L, glucose 20.0 g / L, anhydrous sodium acetate 2.0 g / L, diammonium citrate 2.0 g / L, dipotassium hydrogen phosphate 2.6 g / L, manganese sulfate monohydrate 0.25 g / L, magnesium sulfate heptahydrate 0.5 g / L, and Tween-80 1 mL / L, pH 6.2–6.4.
[0048] MRS solid medium: yeast extract 5.0 g / L, beef extract 10.0 g / L, peptone 10.0 g / L, glucose 20.0 g / L, anhydrous sodium acetate 2.0 g / L, diammonium citrate 2.0 g / L, dipotassium hydrogen phosphate 2.6 g / L, manganese sulfate monohydrate 0.25 g / L, magnesium sulfate heptahydrate 0.5 g / L, Tween-80 1 mL / L, and agar 20.0 g / L, pH 6.2–6.4.
[0049] Fermentation substrate containing cassiaside (g / L): cassiaside 1 g / L, peptone 10 g / L, yeast extract 5 g / L, beef extract 10 g / L, glucose 10 g / L, anhydrous sodium acetate 2 g / L, diamine hydrogen citrate 2 g / L, K2HPO4·3H2O 2.6 g / L, MgSO4·7H2O 0.5 g / L, MnSO4·7H2O 0.25 g / L, Tween-80 1 g / L, cysteine 1 g / L.
[0050] The culture medium raw materials used in the examples were all purchased from Sinopharm Chemical Reagent Co., Ltd.
[0051] Example 1: Isolation, screening, identification and preservation of *Lactobacillus mucosa* CCFM1490 The specific steps are as follows: 1. Screening The samples were obtained from feces of healthy adults in Shanghai. After pretreatment, the samples were stored in 20% glycerol at -80°C. After thawing, the samples were mixed and 0.5 mL of the sample was added to 4.5 mL of physiological saline. The samples were serially diluted with physiological saline and then spread on MRS solid medium. The samples were incubated at 37°C for 48 h. Typical colonies of *Lactobacillus mucosa* were picked and streaked onto MRS solid medium for purification. Single colonies were picked and transferred to MRS liquid medium for enrichment. The samples were preserved in 30% glycerol to obtain the strain, which was named CCFM1490. The typical colonies of *Lactobacillus mucosa* were round, white, and smooth.
[0052] 2. Identification The genome of strain CCFM1490 was extracted, and the 16S rDNA of strain CCFM1490 was amplified and sequenced (performed by Biotechnology Co., Ltd., and the nucleotide sequence of the amplified 16S rDNA of CCFM1490 is shown in SEQ ID NO.1). The sequence was compared with the nucleic acid sequence in NCBI, and the results showed that the strain was Lactobacillus mucosa, and it was named Lactobacillus mucosa CCFM1490.
[0053] 3. Preservation of microbial strains Inoculate 5 mL of MRS liquid medium with *Lactobacillus mucosa* CCFM1490 and incubate at 37°C for 24 h. Take 1 mL of bacterial culture into a sterile centrifuge tube, centrifuge at 8000 r / min for 3 min, discard the upper culture medium, and resuspend the bacterial sludge in 30% glycerol solution and store at -80°C.
[0054] 4. Preservation of bacterial strains Lactobacillus mucosa CCFM1490 was deposited on April 11, 2025 at the Guangdong Provincial Center for Microbial Culture Collection, with accession number GDMCC NO: 66137, located at 5th Floor, Building 59, No. 100 Xianlie Middle Road, Guangzhou.
[0055] Example 2: Transformation of cassiaside by *Lactobacillus mucosa* CCFM1490 The specific steps are as follows: (1) The *Lactobacillus mucosae* CCFM1490 from Example 1 was streaked on MRS solid medium and the plate was incubated upside down at 37°C for 48 h. A single colony was picked and inoculated into 5 mL of MRS liquid medium and incubated at 37°C for 48 h to prepare seed culture.
[0056] (2) Preparation of fermentation substrate containing cassiaside: Cassia glycoside 1 mg / mL, peptone 10 g / L, yeast extract 5 g / L, beef extract 10 g / L, glucose 20 g / L, anhydrous sodium acetate 2 g / L, diamine hydrogen citrate 2 g / L, K₂HPO₄·3H₂O 2.6 g / L, MgSO₄·7H₂O 0.58 g / L, MnSO₄·7H₂O 0.25 g / L, Tween-80 1 g / L, distilled water, cysteine hydrochloride 0.5 g / L.
[0057] Add 5% (v / v) of the bacterial culture of *Lactobacillus mucosa* CCFM1490 to the fermentation substrate containing cassiaside, and ferment at a constant temperature of 37°C for 48 hours.
[0058] (3) After fermentation, centrifuge at 8000 r / min for 15 min, take the fermentation supernatant to obtain the fermentation product of fermented cassia glycosides, and store it at 4℃.
[0059] (4) Take the fermentation product (sample to be tested) obtained in step (3) and filter it through a 0.45 μm membrane. Inject the reference solution and the sample test solution, and determine the chromatographic conditions as described above. The results are as follows: Figure 1 As shown: The results showed that the content of cassiaside was significantly reduced after fermentation with *Lactobacillus mucosa* CCFM1490, and a significant chromatographic peak was found in the elution time of cassiaside. After fermentation of cassiaside with *Lactobacillus mucosa* CCFM1490 of the present invention, the content of cassiaside was significantly increased, and the amount of cassiaside generated was found to be 24.28 mg / L by standard curve analysis.
[0060] Example 3: Effects of a synergistic preparation of *Lactobacillus mucosa* CCFM1490 and cassiaside on mice on a high-fat diet and its post-biotic effects 1. The method for preparing post-generics of cassia seed glycosides from *Lactobacillus mucosa* fermentation is as follows: (1) Streaking of *Lactobacillus mucosa* CCFM1490 on MRS solid medium, incubating at 37°C upside down for 48 h, then picking a single colony and inoculating it into 5 mL of MRS liquid medium and incubating at 37°C for 48 h to obtain bacterial culture; (2) Inoculate the bacterial solution from (1) into the fermentation substrate containing cassiaside, and ferment at a constant temperature of 37°C for 48 hours until the viable count reaches 1×10⁻⁶. 9 CFU / mL; (3) After fermentation, the fermentation broth was heat-treated at 65°C for 30 min, followed by high-pressure homogenization at 1200 Bar for 6 times to obtain the final product. The obtained solution was freeze-dried and resuspended in physiological saline before gavage according to the required dose (2.25 mg cassiaside / kg body weight, calculated based on the cassiaside concentration before fermentation).
[0061] The preparation method of the *Lactobacillus mucosae*-cassiaside fermentation synergistic preparation is as follows: (1) Activate the mucosal lactobacillus CCFM1490 to obtain a bacterial solution; (2) In MRS medium, inoculate the bacterial suspension from (1) with a viable count of 5 × 10⁶. 7 CFU / mL, cultured in an anaerobic environment until the strain reaches the logarithmic growth phase to obtain a bacterial culture of *Lactobacillus mucosae*. (3) Collect bacterial cells from liquid culture medium by centrifugation, mix with freeze-drying protectant (12% skim milk powder), and freeze-dry to obtain freeze-dried bacterial powder; (4) Adjust the dosage according to the mouse's body weight (in kg), and mix to a final concentration of 5 × 10⁻⁶. 9 The product is obtained by adding CFU (carbohydrate lactobacillus mucosa) CCFM1490 + 2.25 mg cassiaside / kg body weight.
[0062] 2. Animal Experiment Design Eighty-eight healthy male C57BL / 6J mice aged 6 weeks were randomly divided into 11 groups of 8 mice each, as shown in Table 3. The 11 groups were: a blank control group, a model group, a positive control group, and a group of *Lactobacillus mucosa* (1 × 10⁻⁶ bacteria per mouse per day via gavage). 9 CFU (Colombia mucosa) and cassia seed glycoside group (2.25 mg cassia seed glycoside / kg body weight), and high-dose CFU-cassia seed glycoside synergistic preparation group (1×10 mg / kg body weight per mouse by gavage daily) were administered to mice. 9 The following groups were prepared by gavage: * *Lactobacillus mucosa* CCFM1490 + 2.25 mg cassiaside / kg body weight (the lyophilized strain powder was mixed with the cassiaside solution and diluted to the rated volume before gavage); *Lactobacillus mucosa*-cassiaside synergistic preparation group (dose was half of the high dose); *Lactobacillus mucosa*-cassiaside synergistic preparation group (dose was one-quarter of the high dose); *Lactobacillus mucosa*-fermented cassiaside post-biotic group (gavage samples were prepared according to the above-mentioned fermented cassiaside post-biotic preparation method, and the dose was calculated based on the cassiaside concentration before fermentation, which was 2.25 mg cassiaside / kg body weight); *Lactobacillus mucosa*-fermented cassiaside post-biotic group (dose was half of the high dose); and *Lactobacillus mucosa*-fermented cassiaside post-biotic group (dose was one-quarter of the high dose).
[0063] The experiment lasted 13 weeks: After one week of acclimatization, mice were fed a low-fat, low-sugar diet starting from the second week, while the other groups were fed a high-fat, low-sugar diet. The modeling period was 8 weeks. From the 9th week, the positive control group was administered 5.2 mg / kg simvastatin at a dose of 0.2 mL / mouse via gavage. Each experimental group was administered a mixture of lyophilized bacterial strain powder and cassia seed glycosides or fermented lyophilized powder (dissolved in physiological saline at the appropriate dose) at a dose of 0.2 mL / mouse daily via gavage. The blank control group and the model group were administered an equal volume of physiological saline as controls until the end of the experiment. All groups had free access to water and food.
[0064] Table 1 Animal Experiment Design
[0065] 3. Indicator Testing After intervention, the mice were observed daily for their mental state and weighed weekly. During dissection, intact liver tissue and epididymal adipose tissue were sampled, washed with phosphate-buffered saline (PBS), blotted dry on filter paper, and then their wet weight was measured.
[0066] The levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), total cholesterol (TG), and triglycerides (TC) in serum were measured using a fully automated bioanalyzer.
[0067] The preparation of histopathological sections was handled by Nanjing Saiwei. H&E staining included, in sequence: tissue fixation, trimming, dehydration, clearing, paraffin infiltration, embedding, sectioning, mounting, dewaxing, rehydration, staining, dehydration and mounting, and microscopic examination. Oil Red O staining included, in sequence: embedding, sectioning, isopropanol treatment, staining, fixation, and microscopic examination.
[0068] 4. Results The weight gain, liver weight, and epididymal fat content of the mice after intervention were as follows: Figure 2 As shown; serum ALT, AST, TG, and TC levels are as follows Figure 3 As shown; liver H&E sections and Oil Red O stained sections are as follows. Figure 4 As shown.
[0069] (1) Effects on obesity phenotype Depend on Figure 2 The results showed that the weight gain during the intervention period was 5.04 g in the model group, while the weight gain in the high-dose CCFM1490-cassia glycoside synergistic preparation group was only 2.13 g, a decrease of 58% compared to the model group (p<0.001); the weight gain in the high-dose post-biotic group was 1.44 g, a decrease of 71% (p<0.001). The weight gain in the cassia glycoside alone gavage group was 2.97 g, a decrease of only about 41%, which was comparable to the effect of the low-dose synergistic preparation (3.19 g) and post-biotic (3.01 g), indicating that CCFM1490 can significantly improve the weight control effect of cassia glycoside.
[0070] Regarding liver weight, the model group had a weight of 1.67g, while the high-dose synbiotic and post-biotic groups showed reductions of 1.21g and 1.15g, respectively, representing decreases of approximately 27% and 31% (p<0.01), which was superior to the cassia seed glycoside intervention group alone (1.29g, a decrease of approximately 23%). Epididymal fat weight was 2.56g in the model group, while the high-dose synbiotic and post-biotic groups showed reductions of 1.93g and 1.87g, respectively, representing decreases of approximately 25% and 27% (p<0.01), whereas cassia seed glycoside intervention alone had no significant effect (2.12g, P>0.05), further demonstrating the synergistic fat-reducing effect of CCFM1490 combined with cassia seed glycoside.
[0071] (2) Effects on blood lipid levels Depend on Figure 3 The results showed that serum triglyceride (TG) in the model group was 1.16 mmol / L, while in the high-dose synbiotic and post-biotic groups it decreased to 0.86 mmol / L and 0.88 mmol / L, respectively, representing reductions of approximately 26% and 24% (p<0.001), showing a dose-dependent effect. In the cassia seed glycoside intervention group, TG was 0.91 mmol / L, a reduction of approximately 21%.
[0072] The serum total cholesterol (TC) level in the model group was 6.94 mmol / L. In the high-dose synbiotic preparation and post-biotic preparation groups, the levels decreased to 6.03 mmol / L and 5.84 mmol / L, respectively, representing reductions of approximately 13% and 16% (p<0.05), which were superior to intervention with cassia seed glycoside alone (6.32 mmol / L, a reduction of approximately 9%). Oral administration of CCFM1490 live bacteria alone had no significant effect on lipid indicators (P>0.05), indicating that its lipid-lowering effect depends on synergy with cassia seed glycoside.
[0073] (3) Effects on liver damage and lipid accumulation Depend on Figure 4 The serum ALT and AST levels in the model group were 77.8 U / L and 219.2 U / L, respectively. In the high-dose synbiotic group, ALT and AST levels decreased to 56.1 U / L and 159.7 U / L, respectively, representing reductions of approximately 28% and 27% (p<0.001). In the high-dose post-synbiotic group, ALT and AST levels were 55.1 U / L and 164.3 U / L, respectively, with similar reductions. In the cassia glycoside-only intervention group, ALT and AST levels were 62.6 U / L and 169.2 U / L, respectively, representing reductions of approximately 19% and 22%, indicating that CCFM1490 can significantly enhance the hepatoprotective effect of cassia glycoside.
[0074] Oil Red O sections of the liver showed extensive fusion of lipid droplets in hepatocytes in the model group, with a significantly elevated steatosis score. The high-dose combined treatment and post-suspension group showed a reduction of approximately 70%-75% in the number and area of lipid droplets (p<0.001), and hepatocyte morphology essentially returned to normal, demonstrating significantly better efficacy than the cassia seed glycoside-only intervention group (lipid droplet reduction of approximately 50%). Further H&E sections showed that in the high-dose intervention group, hepatocyte vacuolation and reticular structure almost disappeared, with only a few intercellular gaps observed, and hepatocyte morphology returned to near normal.
[0075] In conclusion, at the same intervention dose, compared with cassiaside alone, combining strain CCFM1490 with cassiaside can more effectively alleviate obesity, dyslipidemia, liver damage, and lipid accumulation induced by a high-fat diet in mice. Both the post-biotic and synergistic formulations showed superior effects compared to the same dose of cassiaside, exhibiting a clear dose-dependent effect, confirming the synergistic effect of CCFM1490 in enhancing the hepatoprotective efficacy of cassiaside.
[0076] Comparative Example 1: Effect of Microbial Strains on Cassia glycoside Conversion The specific implementation method is the same as in Example 2, except that Lactobacillus mucosa CCFM1490 is replaced with Lactobacillus mucosa ( Limosilactobacillus mucosae Fynlj51M3; Lactobacillus plantarum ( Lactiplantibacillus plantarumCCFM1274 (accession number GDMCC No. 62798, described in the patent application text with application (patent) number CN202311184003.0); *Lactobacillus plantarum* CCFM1366 (accession number GDMCC No. 63998, described in the patent application text with publication number CN118374396A); *Pediococcus lactis* ( Pediococcus acidilactici CCFM1364 (accession number GDMCC No: 63996, described in the patent text with publication number CN118360201A); Bifidobacterium breve ( Bifidobacterium breve FHNX48M6, Bifidobacterium animalis ( Bifidobacterium animalis subsp. lactis SCYA1M1, Bifidobacterium longum ( Bifidobacterium longum subsp. longum FGSYC28M5, Lactobacillus gasseri ( Lactobacillus gasseri FGSZY12L1 was obtained from the Food Microbiology Culture Collection Center of Jiangnan University. The changes in cassiasides in each component before and after fermentation, and the presence or absence of cassia aglycone formation, were detected.
[0077] The results are as follows Figure 5 As shown: the chromatograms of the comparative strains showed no significant chromatographic peaks at 32 minutes, indicating that these strains did not have the ability to convert cassiaside into cassia aglycone. This proves that the ability to convert cassiaside into cassia aglycone is strain-specific, and not all mucosal lactobacilli or lactobacilli have the ability to convert cassiaside into cassia aglycone.
[0078] Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Anyone skilled in the art can make various modifications and alterations without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be determined by the claims.
Claims
1. A strain of *Lactobacillus mucosa* ( Limosilactobacillus mucosae CCFM1490, characterized in that, It was deposited at the Guangdong Provincial Center for Microbial Culture Collection on April 11, 2025, with accession number GDMCC NO: 66137.
2. A microbial preparation, characterized in that, Contains the *Lactobacillus mucosae* CCFM1490 as described in claim 1.
3. The microbial preparation according to claim 2, characterized in that, The concentration of *Lactobacillus mucosa* CCFM1490 in the microbial preparation is at least 1 × 10⁻⁶. 6 CFU / mL or 1×10 6 CFU / g.
4. A synthetic preparation, characterized in that, It contains live bacteria of Lactobacillus mucosae CCFM1490 as described in claim 1 and cassiaside.
5. An epigenetic agent, characterized in that, The post-biotic is the fermentation broth of *Lactobacillus mucosae* CCFM1490 as described in claim 1 or the microbial preparation as described in any one of claims 2 to 3, after fermentation in a fermentation system containing cassiaside and subsequent heat treatment and high-pressure homogenization.
6. The use of the *Lactobacillus mucosae* CCFM1490 of claim 1 or the microbial preparation of any one of claims 2-3 in the preparation of a product for converting cassiaside.
7. A method for converting cassiaside, characterized in that, The substrate containing cassiaside is fermented using the *Lactobacillus mucosae* CCFM1490 as described in claim 1 or the microbial preparation as described in any one of claims 2 to 3.
8. The use of the *Lactobacillus mucosae* CCFM1490 of claim 1, the microbial preparation of any one of claims 2-3, the synbiotic preparation of claim 4, or the metabiotic of claim 5 in the preparation of a medicament for lowering blood lipids and / or alleviating non-alcoholic fatty liver disease.
9. The application according to claim 8, characterized in that, The product includes at least one of the following functions: (a) Reduce serum triglyceride levels; (b) Reduce serum total cholesterol levels; (c) Reduce serum alanine aminotransferase and / or aspartate aminotransferase levels; (d) Improves fatty degeneration of liver tissue.
10. The use of the *Lactobacillus mucosae* CCFM1490 of claim 1, the microbial preparation of any one of claims 2-3, the synbiotic preparation of claim 4, or the metabiotic of claim 5 in the preparation of a medicament for alleviating liver damage caused by a high-fat diet.