FR3135394A3 - FERMENTED TURMERIC WITH LACTICASEIBACILLUS PARACASEI FOR THE REGULATION OF METABOLIC DISEASES - Google Patents

Abstract

The invention relates to turmeric fermented with Lacticaseibacillus paracasei to regulate metabolic diseases. More specifically, turmeric fermented with Lacticaseibacillus paracasei obtained by a manufacturing process disclosed in the invention contains water-soluble curcuminoids and other compositions, and is capable of effectively inhibiting fat accumulation and lipogenesis in the body of an individual; and, therefore, by administering an effective amount of the Lacticaseibacillus paracasei fermented turmeric disclosed in the invention to an individual, an effectiveness in preventing or treating high-fat diet-induced metabolic diseases can be effectively achieved. .

Description

Turmeric Fermented With Lacticaseibacillus Paracasei For The Regulation Of Metabolic Diseases

BACKGROUND OF THE INVENTION

Field of the invention

The invention relates to a use of a fermented product and, more particularly, a turmeric fermented with Lacticaseibacillus paracasei for the regulation of metabolic diseases.

Related prior art

The number of obese people is increasing year by year, mainly due to poor diet and exercise habits. In addition to causing metabolic abnormalities in the body, obesity can also lead to other metabolic diseases such as diabetes and non-alcoholic fatty liver disease. At present, the World Health Organization, the United States, Canada, etc. have declared obesity as a chronic disease.

The main active ingredients of the turmeric rhizome (also called curcuma longa) are curcuminoids (also called curcumin, curcuma longa extract), which are natural polyphenols. Curcuminoids include curcumin, demethoxycurcumin and bis~demethoxycurcumin. Turmeric is used as a traditional medicinal plant in many Asian countries, and studies have shown that it has anti-oxidant, anti-inflammatory, and anti-cancer properties. Although many studies have pointed out that curcumin has strong biological activity, due to the structural characteristics of curcumin, it has problems such as poor water solubility, poor absorption, rapid metabolism and low bioavailability. Although coating technology or nanotechnology is currently used to overcome the problems of poor absorption and low water solubility of curcumin to improve the disadvantage of low bioavailability in vivo, the above-mentioned technologies still cannot to curcumin treated to exert its effectiveness in the body reliably.

The invention aims to provide a turmeric fermented with Lacticaseibacillus paracasei for regulating metabolic diseases, which is obtained by a special fermentation manufacturing process disclosed in the invention, has active ingredients such as water-soluble curcuminoids to be capable of effectively improving the bioavailability and absorption speed of curcumin, thereby being capable of achieving effectiveness in preventing or ameliorating metabolic diseases by regulating fat metabolism and blood glucose metabolism in an organism.

To this end, the invention relates to a use of a fermented turmeric for the regulation of metabolic diseases, the fermented turmeric being prepared by a fermentation reaction of a turmeric with a Lacticaseibacillus paracasei , which means that the administration of an effective amount of the fermented turmeric disclosed in the invention or a composition containing the effective amount of the fermented turmeric to an individual is capable of effectively ameliorating or preventing metabolic diseases or symptoms caused by a high-fat diet.

The Lacticaseibacillus paracasei may be the one called Lacticaseibacillus paracasei MC1-40, deposited with the Leibnitz-Institut DSMZ (Deutsche 20 Sammlung von Mikroagencen and Zellkulturen GmbH), Inhoffenstrasse 7B, 38124 Braunschweig, Germany, on April 28, 2022 (deposit number DSM 34254) .

In one embodiment of the invention, fermented turmeric contains water-soluble curcuminoids.

More specifically, one embodiment of the invention provides for the use of fermented turmeric to prepare a weight loss composition.

Another embodiment of the invention provides for the use of a fermented turmeric to prepare a composition intended to treat and/or prevent diseases linked to a metabolic imbalance.

Metabolic imbalance diseases can be caused by a high-fat diet, such as non-alcoholic fatty liver disease, hyperglycemia and the like.

shows the analysis results of oil droplet content in HepG2 cells after being cultured under different conditions.

shows the body weight changes of each group of mice in Example 2 during the test period.

shows the results of hematoxylin and eosin (H&E) staining of liver tissues from each group of mice in Example 2.

shows the results of analysis of the triglyceride content in the liver tissues of each group of mice in Example 2.

shows the H&E staining results of gonadal adipose tissues from each group of mice in Example 2.

shows the results of analysis of the expression levels of SirT1 in the liver tissues of each group of mice in Example 2.

shows the results of analysis of the expression levels of PGC ~ 1α in the liver tissues of each group of mice in example 2.

shows the results of blood glucose analysis measured by an oral glucose tolerance test in each group of mice in Example 2.

shows the results of analysis of the area under the glucose curve of the measured blood of each group of mice in the .

shows the results of analysis of GluT2 expression levels in the liver tissues of each group of mice in Example 2.

shows the results of analysis of the expression levels of pAkt/Akt in the liver tissues of each group of mice in Example 2.

DETAILED DESCRIPTION OF THE INVENTION

Disclosed is a turmeric fermented with Lacticaseibacillus paracasei for the regulation of metabolic diseases, the turmeric fermented with Lacticaseibacillus paracasei disclosed in the invention containing water-soluble curcuminoids to be capable of inhibiting fat accumulation and lipogenesis; therefore, administering an effective amount of the Lacticaseibacillus paracasei -fermented turmeric disclosed in the invention or a composition containing the Lacticaseibacillus paracasei -fermented turmeric disclosed in the invention to an individual is capable of effectively preventing or treating metabolic diseases induced by a high-fat diet, such as non-alcoholic fatty liver disease, obesity, hyperglycemia and diabetes.

Furthermore, the turmeric fermented with Lacticaseibacillus paracasei disclosed in the invention is obtained by a fermentation reaction of a turmeric with a Lacticaseibacillus paracasei , said Lacticaseibacillus paracasei referring to the species Lacticaseibacillus paracasei from a taxonomic point of view or to a strain classified in the species Lacticaseibacillus paracasei , such as Lacticaseibacillus paracasei MC1-40 (deposited with Leibnitz-Institut DSMZ on April 28, 2022); Generally speaking, the fermentation reaction conditions include a pH value of 4.0~6.0, fermentation temperature of 35~40°C and fermentation time of 21~36 hours, culture medium used is a culture medium which can be used for the growth of culture of lactic acid bacteria or Lacticaseibacillus paracasei which is well known to those skilled in the art to which the invention relates.

What is called "composition" in the invention refers to a composition containing from 0.1% to 100% of the turmeric fermented with Lacticaseibacillus paracasei disclosed in the invention, which can be a food, a medicine, a supplement nutritional, and can be prepared in different forms as needed, including but not limited to powder, liquid, lozenges, pills and capsules.

Below, in order to illustrate the technical characteristics and efficiencies of the invention, several examples will be provided and described together with the drawings as follows.

The Lacticaseibacillus paracasei MC1-40 used in the following examples has been deposited with the Leibnitz-Institut DSMZ (Deutsche 20 Sammlung von Mikroagencen and Zellkulturen GmbH). Inhoffenstrasse 7B, 38124 Braunschweig, Germany, April 28, 2022 (deposit number DSM 34254).

The cells used in the following examples, such as HepG2, are biological materials which can be easily obtained by one skilled in the art to which the invention relates, and therefore do not need to be deposited.

Example 1 : Preparation of fermented turmeric

Lacticaseibacillus paracasei MC1-40 is cultured in a medium containing 1~2% glucose, 1~2% peptone, 0.01~0.08% magnesium sulfate, when the lactic acid bacteria in the medium reach a phase of logarithmic growth, it is inoculated into main fermentation medium to achieve main fermentation, the main fermentation medium contains 2~4% glucose, 2~6% peptone, 0.01~0.08% magnesium sulfate and 3% turmeric powder, fermentation pH is controlled at 4.0~6.0, fermentation temperature is 35~40°C, fermentation time is 21~36 hours; after the end of fermentation, it is dried to obtain fermented turmeric powder.

Turmeric powder without microbial fermentation and fermented turmeric liquid are extracted with water and methanol as extraction solvents, respectively, to obtain methanolic turmeric extract, aqueous turmeric extract, methanolic turmeric extract fermented and an aqueous extract of fermented turmeric. The methanolic extract and the aqueous extract are analyzed by HPLC, the HPLC analysis conditions being as follows: the mobile phase consists of a 2% acetic acid solution and acetonitrile (6:4 (v/v) ) ; the analytical column is C18; the detection wavelength is 420 nm; the column temperature is 35 °C; the flow rate is 2 mL/min; The sample injection volume is 10 μL.

HPLC analysis results show that a total curcuminoid content in turmeric methanol extract is 0.51%, including 0.13% BDMC (bisdemethoxycurcumin), 0.18% DMC (demethoxycurcumin) and 0. 20% curcumin; in the methanolic extract of fermented turmeric, the curcuminoid content is 0.39%, including 0.11% BDMC, 0.16% DMC, 0.12% curcumin; and curcuminoids cannot be detected in turmeric aqueous extract, but a content of curcuminoids in fermented turmeric aqueous extract is 0.12%.

The above results show that curcumin in turmeric powder not processed by fermentation cannot be dissolved in water due to the hydrophobic nature of curcumin, and its content cannot be detected in aqueous solution; while fermented turmeric obtained by fermenting turmeric with lactic acid bacteria, such as L. paracasei , is analyzed to contain 0.12% curcuminoids in water. In other words, the fermented turmeric and its fermented composition disclosed in the invention are capable of effectively improving a disadvantage of low water solubility.

Example 2: cellular test

HepG2 cells (6×10 ⁴ cells/well) are cultured in a 96-well plate for 24 hours, the medium being DMEM medium containing 10% fetal bovine serum; the cultivation conditions are 37°C and 5% carbon dioxide; cultured until the cells grow to about 70~80% of full growth, add control medium, DMSO, 16mM FFA (free fatty acids, hereafter referred to as oleic acid), 16mM FFA and a standard product of curcumin (25 μM, dissolved in DMSO), 16mM FFA and aqueous solution of unfermented turmeric (25 μM), 16mM FFA and aqueous solution of fermented turmeric (25 μM), and aqueous solution of fermented turmeric (25 μM ), and cultured for 24 hours, the aqueous solution of fermented turmeric being prepared with reference to the method of preparation of using fermented turmeric and water as disclosed in Example 1. Coloring with Oil Red O is carried out on the cells in each well after the end of the culture to analyze a staining of intracellular oil droplets, and the results are presented in the .

The results of the show that HepG2 cells produced obvious oil droplets after being treated with oleic acid. After being treated with the standard curcumin product and unfermented turmeric aqueous solution, although an amount of fat accumulation in liver cells can be reduced, the effect is not as good as the effect of the aqueous solution of fermented turmeric disclosed in the invention to inhibit the generation of oil droplets in liver cells. The fermented turmeric aqueous solution is significantly better than the unfermented turmeric aqueous solution or the standard curcumin product. The above results show that the fermented turmeric produced by the fermentation manufacturing method disclosed in the invention is indeed capable of effectively increasing an effectiveness of turmeric and active ingredients thereof for inhibiting the generation of oil and fat, which means that the fermented turmeric disclosed in the invention is capable of regulating lipogenesis in hepatic cells, thereby being capable of achieving an effectiveness of inhibiting or slowing down the appearance of non-fatty hepatic steatosis. alcoholic or diseases linked to fat metabolism.

Example 3: animal test

Six-week-old male C57BL/6 black mice are divided into four groups and raised for 17 weeks under the following conditions:

Group 1: normal diet (food 5001M) and normal drinking water.

Group 2: 50% high fat foods and normal drinking water.

Group 3: 50% high-fat foods, normal drinking water, unfermented turmeric powder and inactivated bacteria ( Lacticaseibacillus paracasei MC1-40) up to 5% of the weight of the food.

Group 4: 50% high-fat foods, normal drinking water, fermented turmeric powder and inactivated bacteria ( Lacticaseibacillus paracasei MC1-40) at 5% of the weight of the foods, the fermented turmeric powder being prepared by reference to the manufacturing process disclosed in Example 1.

During the experiment, the changes in body weight of each group of mice are recorded, and the results are shown in the .

The results of the show that the weights of mice in group 2 are significantly higher than those of mice in group 1, indicating that the administration of high-fat food will effectively increase the weights of mice; the weights of mice in groups 3 and 4 are respectively lower than those of mice in group 2, a degree of increase in the weights of mice in group 4 is significantly slower than that of group 3, and the weights of mice in group 4 being almost the same as those of group 1.

The results of the show that the fermented turmeric disclosed in the invention is capable of effectively slowing down obesity caused by a high-fat diet, and that a weight gain inhibition effect is significantly better than that of turmeric not treated by fermentation; in other words, the fermented turmeric disclosed in the invention is capable of reliably inhibiting or slowing down weight gain induced by a high-fat diet in vivo, thereby being capable of effectively achieving weight loss effectiveness and prevent associated metabolic diseases.

Example 4: fabric cuts

Once each group of mice from Example 3 has been sacrificed, their liver tissues are removed, after obtaining paraffin-embedded sections, H&E staining is carried out, and a triglyceride content in the livers of each group of mice is analyzed, the results are shown in FIGS. 3A and 3B; Furthermore, gonadal adipose tissues from each group of mice are collected for obtaining tissue sections and H&E staining, the results are presented in the .

The results of the show that the cells in the liver tissues of group 1 mice are neatly arranged and there are no pathological changes; liver tissues of mice in group 2 show obvious white fat vacuoles (indicated by the arrows in the ) with an accumulation of lipids; white fat vacuoles in the liver tissues of mice in groups 3 and 4 are significantly fewer than those of mice in group 2. The results of the show that, compared with group 1 mice, the triglyceride content in the livers of group 2 mice is significantly increased, while triglyceride content in the livers of group 4 mice is significantly lower than that of group 4 mice. of group 2. The results of the show that a gonadal adipocyte size of mice in group 2 is significantly larger than that of mice in group 1, while a gonadal adipocyte size of mice in group 4 is significantly smaller than that of mice in groups 2 and 3.

The results of FIGS. 3 and 4 show that a high-fat diet can actually cause fat accumulation in the liver and other tissues, leading to the onset of individual obesity and diseases linked to abnormal lipid metabolism, such as non-fatty liver disease. alcoholic. On the other hand, the administration of the fermented turmeric disclosed in the invention to an individual is able to effectively decrease the accumulation of fats and the content of triglycerides in tissues induced by a high-fat diet, which means that the fermented turmeric disclosed in the invention is capable of preventing or treating diseases related to abnormal lipid metabolism caused by a high-fat diet, such as non-alcoholic fatty liver disease and obesity.

Example 5: protein expression

Once each group of mice from Example 3 was sacrificed, their liver tissues were removed, the expressions of SirT1 and PGC~1α in the liver tissues of each group of mice were analyzed by Western blotting and a commercially available ELISA kit, and the results are shown in FIGS. 5A and 5B.

The results of FIGS. 5A and 5B show that expression levels of Sirt1 and PGC~1α in the liver tissues of mice from group 4 are increased compared to those of mice from groups 2 and 3. Given that Sirt1 is a major regulator of mitochondrial biosynthesis capable of promoting expression of PGC~1α involved in energy metabolism, the results of FIGS. 5A and 5B show that the fermented turmeric disclosed in the invention is capable of promoting the oxidation of fatty acids by increasing the expressions of Sirt1 and PGC~1α, which means that the fermented turmeric disclosed in the invention is capable of effectively reducing a probability of the appearance of obesity or diseases linked to metabolism induced by a diet rich in fats.

Example 6: analysis of the ability to regulate and control blood sugar

In Example 3, each group of mice is subjected to an oral glucose tolerance test (OGTT) at the 16th week of the experiment, firstly, the fasting blood glucose levels of each group of mice are measured, then each group of mice receives 2 g/kg body weight of glucose solution by tube feeding, and blood glucose levels are measured at 30, 60, 90, 120 and 180 minutes after feeding, the results are presented in the , areas under the OGTT curves are analyzed, and the results are presented in the .

In addition, after the experiment of Example 2, the liver tissues of each group of mice are respectively collected, and the expression levels of proteins such as GluT2 and pAkt/Akt in the liver tissues of each group of mice are analyzed by Western blotting and a commercially available ELISA kit, and the results are shown in FIGS. 7A and 7B.

From the results of the , we can know that at 0 minutes, fasting blood sugar levels of mice in group 2 are higher than those of mice in group 1 and reach statistical significance. Blood glucose concentrations of group 2 mice begin to increase 30 minutes after provision of the glucose solution by probe, blood glucose concentrations of group 2 mice gradually decrease at 60, 90 and 120 minutes but are still higher than those of group 1 mice, and reached statistical significance at 90 minutes; blood glucose concentrations of group 4 mice, measured 30 minutes after provision of the glucose solution by probe, were lower than those of group 2 mice, while 60, 90 and 120 minutes after provision of the solution of glucose by probe, there is no significant difference between the blood glucose concentrations of mice in groups 3 and 4 respectively and those of mice in group 2. The results of the show that group 2 mice have a higher area under the curve (AUC), while the AUC of group 4 mice is significantly lower than that of group 2 mice.

Furthermore, the results of FIGS. 7A and 7B show that the fermented turmeric disclosed in the invention is capable of increasing a pAkt/Akt value and significantly increasing an expression level of the glucose transporter Glut2.

The results of FIGS. 6 and 7 show that a high-fat diet can slow down a blood sugar decline rate of the oral glucose tolerance test, but that the administration of the fermented turmeric disclosed in the invention is able to effectively improve a situation of rapid increase in blood sugar within 30 minutes of dieting, that is, the fermented turmeric disclosed in the invention has a better ability to metabolize glucose and regulate blood sugar, thereby stabilizing diet-induced blood sugar rich in fats, and effectively preventing the onset of diseases linked to poor blood sugar metabolism.

Claims (9)

- Use of a fermented turmeric to prepare a weight loss composition, in which the fermented turmeric is prepared by a fermentation reaction of a turmeric with a Lacticaseibacillus paracasei . - Use according to claim 1, in which the fermented turmeric contains water-soluble curcuminoids. - Use according to claim 1, in which Lacticaseibacillus paracasei is designated as Lacticaseibacillus paracasei MC1-40 deposited with the Leibnitz-Institut DSMZ (Deutsche 20 Sammlung von Mikroagencen and Zellkulturen GmbH), Inhoffenstrasse 7B, 38124 Braunschweig, Germany, on April 28, 2022 (DSM deposit number 34254). - Use of a fermented turmeric to prepare a composition for treating and/or preventing a disease linked to a metabolic imbalance, in which the fermented turmeric is prepared by a fermentation reaction of a turmeric with a Lacticaseibacillus paracasei . - Use according to claim 4, in which Lacticaseibacillus paracasei is designated as Lacticaseibacillus paracasei MC1-40 deposited with the Leibnitz-Institut DSMZ (Deutsche 20 Sammlung von Mikroagencen and Zellkulturen GmbH), Inhoffenstrasse 7B, 38124 Braunschweig, Germany, on April 28, 2022 (DSM deposit number 34254). - Use according to one of claims 4 or 5, in which the disease linked to a metabolic imbalance is induced by a diet high in fat. - Use according to one of claims 4 or 5, in which the disease linked to an imbalance in fat metabolism is non-alcoholic fatty liver disease. - Use according to one of claims 4 or 5, in which the disease linked to an imbalance in fat metabolism is hyperglycemia. - Use according to one of claims 4 or 5, in which the fermented turmeric contains water-soluble curcuminoids.