Use of lactobacillus plantarum tci378 strain and metabolites thereof for fat reduction

Abstract

The present application relates to the field of microorganism, especially the application of lactobacillus plantarum TCI378 strain and its metabolite for reducing fat. The present application discloses a metabolite of lactobacillus plantarum, which comprises a compound selected from the group consisting of tryptophyl pyroglutamic acid, 1-methyl-1,2,3,4-tetrahydro-beta-carboline-3-carboxylic acid, 3-phenyllactic acid (as shown in formula III), or any combination thereof. The present application also discloses the use of the lactobacillus plantarum for preparing a fat-reducing composition or a fat-reducing drug. The lactobacillus plantarum and its metabolite can inhibit the accumulation of fat in fat cells, effectively reduce the content of fat in cells, and achieve the goal of reducing fat and obesity.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of microorganisms, in particular, the application of Lactobacillus plantarum TCI378 strain and its metabolites for reducing fat. BACKGROUND

[0002] The World Health Organization (WHO) describes the rapid spread of obesity as an "infectious disease" and calls it "Globesity."

[0003] Therefore, obesity is one of the most serious diseases in modern society, and the main cause of obesity is excessive intake of fat. In addition to causing physical defects, forming psychological and social barriers, and affecting work ability, obesity can also cause many physiological symptoms, including edema, cardiac hypertrophy, fatty liver, biliary and urinary calculi, musculoskeletal pain, gynecological breast or uterine tumors, hyperuricemia (gout), hyperlipidemia, angina, diabetes, hypertension, stroke, etc. Among them, eight causes of death, including malignant tumors, heart disease, cerebrovascular disease, diabetes, chronic lower respiratory tract disease, hypertension, chronic liver disease and cirrhosis, and chronic kidney disease, are related to obesity. Therefore, maintaining a healthy weight and normal body fat content is a goal that modern people must strive for.

[0004] However, the most effective method for treating obesity at present is surgical treatment, and other legal drugs (currently only Roche), exercise, calorie control, and low-calorie meal replacement have also been proven to be effective ways. However, in addition to surgical treatment, most patients use other methods to lose weight, and most of them regain weight after the weight loss treatment ends, so the phenomenon of losing weight and gaining weight again (yo-yo effect) causes more harm to the body.

[0005] In summary, in response to the obesity and overall health problems caused by obesity faced by modern people due to changes in lifestyle and eating habits, and based on the improvement of modern people's living standards and the improvement of health concepts, it is necessary to develop a composition that can effectively reduce the body fat content of an effective ingredient. SUMMARY

[0006] Therefore, an object of the present application is to provide a metabolite of Lactobacillus plantarum, which comprises a compound selected from the group consisting of pyroglutamyl-tryptophan, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, 3-phenyllactic acid, or any combination thereof.

[0007] Another object of the present invention is to provide the use of the metabolites of Lactobacillus plantarum as described above for the preparation of a fat-reducing composition.

[0008] Another object of the present invention is to provide a pharmaceutical composition for use in the preparation of a weight-loss pharmaceutical product, wherein the pharmaceutical composition comprises a compound selected from the group consisting of: tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, 3-phenyllactic acid, or any combination thereof, and a pharmaceutically acceptable carrier.

[0009] In one embodiment of the present invention, the Lactobacillus plantarum has the registration number DSM32451; and the metabolite of the Lactobacillus plantarum is the secretion of the Lactobacillus plantarum, comprising the culture medium in which the Lactobacillus plantarum is cultured; the metabolite of the Lactobacillus plantarum comprises an extract obtained by extracting the metabolite of the Lactobacillus plantarum with methanol, and the metabolite of the Lactobacillus plantarum comprises a compound selected from the group consisting of: tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, 3-phenyllactic acid, or any combination thereof; wherein the concentration of the metabolite of the Lactobacillus plantarum is at least 1 ppm.

[0010] In another embodiment of the present invention, the tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, or 3-phenyllactic acid is obtained by isolating and purifying the metabolites of Lactobacillus plantarum as described above; and the concentration of the tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, or 3-phenyllactic acid is at least 10 μg / mL.

[0011] In another embodiment of the invention, the fat reduction is achieved by inhibiting the accumulation of fat in fat cells, thereby reducing the fat content in fat cells.

[0012] The *Lactobacillus plantarum* or its metabolites of the present invention effectively inhibit fat accumulation in adipocytes and reduce the fat content in cells. The methanol extract of the metabolites of *Lactobacillus plantarum* 8 contains tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid, which also effectively inhibit fat accumulation in adipocytes and reduce the fat content in cells. Therefore, *Lactobacillus plantarum*, its metabolites, and the tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid purified from its metabolites can be used to prepare a composition for fat reduction, which is a pharmaceutical product or a food product, and can be administered to an individual by oral administration or other means.

[0013] The following will further illustrate the embodiments of the present invention with reference to the accompanying drawings. The examples listed below are for illustrative purposes only and are not intended to limit the scope of the present invention. Anyone skilled in the art can make some modifications and refinements without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention shall be determined by the appended claims. Attached Figure Description

[0014] Figure 1 This is a hydrogen nuclear magnetic resonance spectrum of TCI378-1 purified from the metabolites of Lactobacillus plantarum TCI378 according to an embodiment of the present invention.

[0015] Figure 2 This is a mass spectrum of purified TCI378-1 from the metabolites of Lactobacillus plantarum TCI378 according to an embodiment of the present invention;

[0016] Figure 3 This is a hydrogen nuclear magnetic resonance spectrum of TCI378-2 purified from the metabolite of Lactobacillus plantarum TCI378 according to an embodiment of the present invention.

[0017] Figure 4 This is a mass spectrum of purified TCI378-2 from the metabolites of Lactobacillus plantarum TCI378 according to an embodiment of the present invention;

[0018] Figure 5 This is a hydrogen nuclear magnetic resonance spectrum of TCI378-3 purified from the metabolites of Lactobacillus plantarum TCI378 according to an embodiment of the present invention.

[0019] Figure 6 This is a mass spectrum of purified TCI378-3 from the metabolites of Lactobacillus plantarum TCI378 according to an embodiment of the present invention;

[0020] Figure 7 The histogram of purified TCI378-1, TCI378-2, and TCI378-3 metabolites from Lactobacillus plantarum TCI378 in an embodiment of the present invention on the inhibition of fat accumulation, *p value < 0.05. Detailed Implementation

[0021] The values ​​used in this paper are approximate, and all experimental data are expressed within a 20% range, a better range of 10%, and a best range of 5%.

[0022] Statistical analysis was performed using Excel software. Data are expressed as mean ± standard deviation (SD), and differences between groups were analyzed using Student's t-test.

[0023] definition

[0024] The Lactobacillus plantarum of this invention is a probiotic bacteria that can reduce fat. This invention relates to a novel *Lactobacillus plantarum*, named TCI378, which was deposited on March 13, 2017, at the German Collection of Microorganisms and Cell Cultures (DSMZ), located at 7B Inhofenstrasse, Braunschweig 38124, Germany, with accession number DSM32451. In vitro experiments have demonstrated that *Lactobacillus plantarum* TCI378, its metabolites, and the compounds purified from these metabolites—tryptophan, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid—can inhibit fat accumulation in adipocytes. This invention demonstrates the use of Lactobacillus plantarum TCI378 and its metabolites in the preparation of a composition for weight loss, wherein the composition is a pharmaceutical or a food product that can be administered to an individual by oral administration or other means.

[0025] Probiotics (or probiotic bacteria) are microorganisms whose cells, mixed strains, extracts, or metabolites have a positive effect on the host. They are usually live bacteria that are beneficial to gut health and are derived from the human body. They can also refer to certain microorganisms that are added from outside the body and may be beneficial to the body. The metabolites of a probiotic strain are the secretions of the probiotic strain and include the culture medium in which the bacteria are cultured.

[0026] According to the present invention, three compounds purified from the metabolites of *Lactobacillus plantarum* TCI378 of the present invention by column chromatography and thin layer chromatography (TLC) are tryptophan, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid, and will be named TCI378-1, TCI378-2, and TCI378-3, respectively.

[0027] According to the present invention, the operating procedures and parameter conditions for bacterial culture fall within the professional competence and routine technical scope of those skilled in this art.

[0028] As used in this article, the term "metabolite" refers to the substance secreted into the bacterial culture medium after being metabolized by the bacteria during bacterial culture, including the culture medium in which the bacteria are cultured.

[0029] According to the present invention, pharmaceutical products can be manufactured using techniques known to those skilled in the art into dosage forms suitable for parenterally or topically administration, including, but not limited to: injections [e.g., sterile aqueous solutions or dispersions], sterile powders, external preparations, and the like.

[0030] According to the present invention, the pharmaceutical product may further comprise a pharmaceutically acceptable carrier widely used in pharmaceutical manufacturing techniques. For example, the pharmaceutically acceptable carrier may comprise one or more reagents selected from the following: solvent, buffer, emulsifier, suspending agent, decomposer, disintegrating agent, dispersing agent, binding agent, excipient, stabilizing agent, chelating agent, diluent, gelling agent, preservative, wetting agent, lubricant, absorption delaying agent, liposome, and the like. The selection and quantity of these reagents fall within the scope of professional competence and routine practice of those skilled in the art.

[0031] According to the present invention, the pharmaceutically acceptable carrier comprises a solvent selected from the group consisting of: water, normal saline, phosphate buffered saline (PBS), an aqueous solution containing alcohol, and combinations thereof.

[0032] According to the present invention, the pharmaceutical product can be administered via a parenteral route selected from the group consisting of: subcutaneous injection, intraepidermal injection, intradermal injection, and intralesional injection.

[0033] According to the present invention, pharmaceutical products can be manufactured using techniques known to those skilled in the art as external preparations suitable for topical application to the skin, including, but not limited to: emulsions, gels, ointments, creams, patches, liniments, powders, aerosols, sprays, lotions, serums, pastes, foams, drops, suspensions, salves, and bandages.

[0034] According to the present invention, the external formulation is prepared by mixing the pharmaceutical product of the present invention with a base known to those skilled in the art.

[0035] According to the present invention, the substrate may contain one or more additives selected from the following: water, alcohols, glycols, hydrocarbons [such as petroleum jelly and white petrolatum], waxes [such as paraffin and yellow wax], preserving agents, antioxidants, surfactants, absorption enhancers, stabilizing agents, gelling agents [such as...] 974P ( The additives include 974P, microcrystalline cellulose, and carboxymethyl cellulose; active agents; humectants; odor absorbers; fragrances; pH adjusting agents; chelating agents; emulsifiers; occlusive agents; emollients; thickeners; solubilizing agents; penetration enhancers; anti-irritants; colorants; and propellants. The selection and quantity of these additives fall within the scope of professional expertise and routine techniques of those familiar with this technology.

[0036] According to the present invention, food products can be used as food additives, added during the preparation of raw materials or during the production of food by conventional methods, and formulated with any edible material to form food products for human and non-human animal consumption.

[0037] According to this invention, the types of food products include, but are not limited to: beverages, fermented foods, bakery products, health foods, and dietary supplements.

[0038] Chemical Analysis Materials

[0039] The compounds were separated using methanol and acetonitrile as solvents. Chemical structure analysis of the compounds was performed using deuterated methanol d4 (99.5% deuteration) as solvent. All materials were purchased from Merck Taiwan.

[0040] Chemical analysis instruments

[0041] The separation of compounds utilizes column chromatography and thin-layer chromatography (TLC). A medium-pressure liquid chromatography (MPLC) system is used. Rf + (Teledyne ISCO, Lincoln, NE); the columns were selected from Sephadex LH-20 (Pharmacia, Piscataway, NJ, USA), Diaion HP-20 (Mitsubishi Chemical Co., Japan), and Silica gel 0.040-0.023mm and... RP-18 (0.040-0.023mm) (Merck, EMDMillopore Co., Germany). The High Performance Liquid Chromatography (HPLC) system is an Agilent 1200 series; the degassing unit is an Agilent vacuum gas storage unit 1322A; the extraction solvent delivery system is an Agilent quaternary pump G1311A; the variable wavelength detector (MWD) is an Agilent G1314B; the diode array detector (DAD) is an Agilent 1260Infinity DADVL G1315D, with detection wavelengths of 210nm, 280nm, 320nm, and 365nm (Agilent Germany). The column is... 5μmC18(2) (250 x 10 mm, Phenomenex, USA). Thin-layer chromatography slides were Silica Gel 60F. 254 (0.25mm; Merck, EMD Millopore Co., Germany) or RP-18F 254 -S (0.25mm; Merck, EMD Millopore Co., Germany) aluminum sheet.

[0042] The chemical structure of the compounds was analyzed using mass spectrometry (MS) and nuclear magnetic resonance spectrometry (NMR). The mass spectrometer (MS) was a tandem mass spectrometer-two-dimensional ion trap tandem Fourier transform mass spectrometer and ESI-MS / MS: measurements were performed using a BrukeramaZon SL system, with units measured in m / z. The nuclear magnetic resonance spectrometer (NMR) used an Ascend 400MHz (Bruker Co., Germany) for both 1D and 2D spectra, with chemical shift expressed as δ, in ppm. The rotary vacuum evaporator, used for solvent removal, was Laborota 4000, Heidolph Instruments GmbH & Co. KG Germany.

[0043] According to the present invention, the operating procedures and parameter conditions for the chemical separation and chemical structure analysis of the mixture fall within the professional competence and routine technical scope of those skilled in this art.

[0044] This invention provides *Lactobacillus plantarum* TCI378, its metabolites or purified from its metabolites, namely, tryptophan, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, or 3-phenyllactic acid. The present invention relates to the use of tryptophan metabolites obtained from the culture medium of *Lactobacillus plantarum* TCI378, which contain the active ingredients tryptophan pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid. *Lactobacillus plantarum* TCI378, its metabolites, and tryptophan pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid purified from its metabolites can be used to inhibit fat accumulation in adipocytes.

[0045] Meanwhile, the composition of the present invention for weight loss may also contain an effective amount of Lactobacillus plantarum TCI378, its metabolites, tryptophan-based pyroglutamamide purified from its metabolites, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, or 3-phenyllactic acid, and a pharmaceutically acceptable carrier. The composition is a pharmaceutical product or a food product.

[0046] The following details the preparation method of the metabolites of *Lactobacillus plantarum* TCI378 of the present invention, the detailed method for isolating the active substances tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid from the metabolites of *Lactobacillus plantarum* TCI378 of the present invention, and the efficacy test of these active ingredients in inhibiting fat accumulation in adipocytes. This demonstrates that *Lactobacillus plantarum* TCI378 of the present invention, its metabolites, and tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid purified from its metabolites have the effect of inhibiting fat accumulation in adipocytes and can be used to prepare fat-reducing compositions.

[0047] Example 1: Preparation method of Lactobacillus plantarum TCI378 metabolites of the present invention

[0048] In this embodiment of the invention, the cryopreserved strain of Lactobacillus plantarum TCI378 was activated by a single culture and then cultured at a concentration of 1% in MRS (de Man, Rogosa and Sharpe, BD Difco)TM In Lactobacilli MRS Broth medium, preferably 0.1 mL of activated bacterial culture is added to 10 mL of MRS, and after culturing at 37°C for 18 hours, the culture solution is centrifuged at 5000 rpm for 10 minutes, and the supernatant is collected, which is the metabolite of Lactobacilli TCI378 of the present invention.

[0049] Example 2: Analysis of the active ingredients in the metabolites of *Lactobacillus plantarum* TCI378 of the present invention

[0050] One embodiment of the present invention involves the analysis of the active components in the metabolites of *Lactobacillus plantarum* TCI378. During the separation and purification of the active compound, the selection of layering, sub-layering, and sub-sub-layering was based on a bioassay-guided fractionation method (results not shown). First, 10 liters of the aforementioned *Lactobacillus plantarum* TCI378 metabolites were concentrated under reduced pressure to remove most of the water, yielding 2 liters of concentrate. This concentrate was then subjected to column chromatography (70 cm x 7 cm) using Diaion HP-20 macroporous resin, with an extraction gradient of 100% water, 20% methanol aqueous solution, 40% methanol aqueous solution, 60% methanol aqueous solution, and 100% methanol, resulting in five layers. Layer 3 was then subjected to RP-C18 rapid column chromatography with a linear gradient from 20% methanol to 100% methanol, followed by thin-layer chromatography analysis to obtain eight sub-layers. The 7th sub-layer was subjected to column chromatography using Sephadex LH-20 gel, followed by thin-layer chromatography analysis to obtain 8 sub-layers. Sub-layer 7 was purified by RP-HPLC (methanol / water = 2 / 3) to obtain compounds TCI-378-1 and TCI-378-2; and sub-layer 5 was purified by RP-HPLC (methanol / water = 2 / 3) to obtain compound TCI-378-3.

[0051] TCI378-1 was analyzed by hydrogen nuclear magnetic resonance (NMR) Figure 1 ), C13-NMR (results not shown), HSQC resonance (results not shown), HMBC resonance (results not shown), COSY resonance (results not shown), and electrospray ionization mass spectrometry (negative ion mode) ( Figure 2 After analyzing its chemical structure, it was confirmed to be pyroglutamyl-tryptophan, which is the first known compound obtained in nature.

[0052] TCI378-2 was analyzed by hydrogen nuclear magnetic resonance (NMR) Figure 3 ) and electrospray ionization mass spectrometry (negative ion mode) Figure 4After analyzing its chemical structure, it was confirmed to be 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, which is a known compound.

[0053] TCI378-3 underwent hydrogen nuclear magnetic resonance (NMR) Figure 5 ), C13-NMR (results not shown), HSQC resonance (results not shown), HMBC resonance (results not shown), COSY resonance (results not shown), and electrospray ionization mass spectrometry (negative ion mode) ( Figure 6 After analyzing its chemical structure, it was confirmed that it is 3-phenyllactic acid, a known compound.

[0054] The chemical structures of compounds TCI378-1, TCI378-2, and TCI378-3 were determined by mass spectrometry and nuclear magnetic resonance spectroscopy. Their names and structural formulas are shown in Table 1 below.

[0055] Table 1. Chemical structural formulas of compounds TCI378-1, TCI378-2, and TCI378-3

[0056]

[0057] Example 3: The efficacy of TCI378-1, TCI378-2, and TCI378-3, metabolites of Lactobacillus plantarum TCI378 of the present invention, in inhibiting fat accumulation.

[0058] In this embodiment, mouse bone marrow stromal cells (OP9 cells) were used to test the efficacy of TCI378-1, TCI378-2, and TCI378-3, metabolites of *Lactobacillus plantarum* TCI378, in inhibiting fat accumulation. The mouse bone marrow stromal cells were purchased from the American Center for Type Culture Collection (USA), catalog number CRL-2749. TMThe cells were cultured before differentiation in pre-adipocyte expansion medium, which contained 90% MEMAM (Minimum Essential Medium Alpha Medium, purchased from Gibco, USA, FBS:Cat#10437-028) cell culture medium, 20% fetal bovine serum (purchased from Gibco, USA), and 0.1% penicillin-streptomycin (purchased from Gibco, USA). Mouse bone marrow stromal cells were then differentiated using differentiation medium, which contained 90% MEMAM cell culture medium, 20% fetal bovine serum, and 0.1% penicillin-streptomycin. Lipids in cells were stained with Oil Red O staining reagent (purchased from Sigma, USA, catalog number O0625), in which a 3 mg / mL Oil Red O stock solution was prepared with 100% isopropanol, and the stock solution was prepared into a 60% reaction solution with ddH2O.

[0059] To demonstrate that the metabolites TCI378-1, TCI378-2, and TCI378-3 of *Lactobacillus plantarum* TCI378 of this invention have the effect of inhibiting fat accumulation, mouse bone marrow stromal cells were first differentiated into adipocytes, and 8 x 10⁸ adipocytes were then used. 4 Mouse bone marrow stromal cells were cultured in 24-well culture dishes containing 0.5 μL of the above-mentioned preadipocyte expansion culture medium and cultured at 37°C for 7 days, with the differentiation culture medium being replaced with fresh medium every 3 days. After 7 days, the formation of lipid droplets was observed under a microscope to ensure that the cells had fully differentiated. The cells were then divided into the following four groups: (1) control group containing only cell culture medium, (2) experimental group containing 10 μg / mL TCI378-1, (3) experimental group containing 10 μg / mL TCI378-2, and (4) experimental group containing 10 μg / mL TCI378-3. The cells were cultured at 37°C for 7-10 days, with the differentiation culture medium being replaced with fresh medium every 3 days.

[0060] Next, Oil Red O was used to stain the lipids in the cells to assess whether the active components in the metabolites of *Lactobacillus plantarum* TCI378 of the present invention could indeed reduce lipid accumulation. First, the culture medium was gently removed and the cells were washed twice with 1 mL of phosphate buffered saline (PBS). Then, 1 mL of 10% formaldehyde (purchased from Echo Chemical, Taiwan, Cat.TG1794-4-0000-72NI) was added and reacted at room temperature for 30 minutes to fix the cells. After removing the formaldehyde, the cells were gently washed twice with 1 mL of PBS. Then, 1 mL of 60% isopropanol (purchased from Echo Chemical, Taiwan, PH-3101) was added to each well of the cells and reacted for 1 minute. After removing the isopropanol, 1 mL of Oil Red O reaction solution was added and reacted at room temperature for 1 hour. Then, the Oil Red O solution was removed and the cells were quickly destained with 1 mL of 60% isopropanol for 5 seconds. Finally, the cells were photographed and quantified using a microscope. Next, 100% isopropanol was added to the stained cells, and the mixture was placed on a shaker for 10 minutes to dissolve the stain. Then, 100 μL was transferred to a 96-well culture dish, and the OD values ​​of each group were measured using an ELISA reader. 510 nm Read the values ​​to quantify Oil Red O. Then use Excel software to perform a student t-test to determine whether there is a statistically significant difference between the two sample groups (*p value < 0.05; **p value < 0.01; ***p value < 0.001).

[0061] The results of testing the inhibitory effects of purified TCI378-1, TCI378-2, and TCI378-3 metabolites from *Lactobacillus plantarum* TCI378 metabolites on fat accumulation are as follows: Figure 7 As shown, treatment with TCI378-1 of the present invention significantly reduced cellular lipid accumulation by approximately 11.5% compared to the control group; treatment with TCI378-2 of the present invention significantly reduced cellular lipid accumulation by approximately 12.4% compared to the control group; and treatment with TCI378-3 of the present invention significantly reduced cellular lipid accumulation by approximately 21.5% compared to the control group. These results indicate that the metabolites of *Lactobacillus plantarum* TCI378 of the present invention, namely TCI378-1, TCI378-2, and TCI378-3, can effectively inhibit lipid accumulation in adipocytes and reduce the lipid content in cells.

[0062] In summary, *Lactobacillus plantarum* TCI378 or its metabolites of the present invention effectively inhibit fat accumulation in adipocytes and reduce the fat content in cells. The methanol extract of the metabolites of *Lactobacillus plantarum* TCI378 contains tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid, which also effectively inhibit fat accumulation in adipocytes and reduce the fat content in cells. Therefore, *Lactobacillus plantarum* TCI378, its metabolites, and the tryptophan-based pyroglutamamide, 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid, and 3-phenyllactic acid purified from its metabolites can be used to prepare a composition for fat reduction, which is a pharmaceutical product or a food product, and can be administered to an individual via oral administration or other means.

Claims

1. Use of a tryptophan-based pyroglutamamide in the preparation of a weight-loss composition, wherein the chemical structural formula of the tryptophan-based pyroglutamamide is as follows (I): Equation (I).

2. The use according to claim 1, characterized in that, The tryptophan-based pyroglutamamide is derived from a metabolite of Lactobacillus plantarum, whose accession number is DSM32451.

3. The use according to claim 2, characterized in that, The metabolites of *Lactobacillus plantarum* are secretions of *Lactobacillus plantarum*, including the culture medium in which *Lactobacillus plantarum* is cultured.

4. The use according to claim 2, characterized in that, The metabolites of the plant lactobacillus include an extract obtained by extracting the metabolites of the plant lactobacillus with methanol.

5. The use according to claim 4, characterized in that, The metabolites of the plant lactobacillus further include 3-phenyllactic acid and 1-methyl-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid.

6. The use of a pharmaceutical ingredient in the preparation of a weight-loss pharmaceutical product, characterized in that, The pharmaceutical composition comprises tryptophan-based pyroglutamamide and a pharmaceutically acceptable carrier, the chemical structural formula of which is as follows (I): Equation (I).

7. The use according to claim 6, characterized in that, The concentration of the tryptophan-based pyroglutamamide is at least 10 μg / mL.

8. The use according to claim 1 or 6, characterized in that, The fat reduction mentioned above is to inhibit the accumulation of fat in fat cells, thereby reducing the fat content in fat cells.

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