TWI719691B - Use of lactobacillus reuteri gkr1 for preparing composition of reducing uric acid - Google Patents

Abstract

The present invention relates to a use of Lactobacillus reuteri GKR1 for preparing a composition of reducing uric acid. The composition including Lactobacillus reuteri GKR1 is administered to reduce the uric acid in the serum, alleviate and/or prevent hyperuricemia of a subject.

Description

Lactobacillus reuteri strain Use of GKR1 for the preparation of a composition for reducing uric acid

The present invention relates to the use of a single Lactobacillus strain for the preparation of a composition for reducing uric acid, in particular to the use of a Lactobacillus reuteri strain GKR1 for the preparation of a composition for reducing uric acid.

Purines in the human body are mainly excreted by the kidneys after uric acid is synthesized in the liver. In recent years, due to changes in dietary patterns, the proportion of patients with hyperuricemia has also been on the rise. When exposed to high-purine diets (such as seafood, animal offal, broth, beer, etc.), high-fat diets (such as fried foods), stress, overwork, sugary drinks and/or intense exercise in life, it is possible Increase serum uric acid concentration. When the serum uric acid concentration exceeds the normal value, it is called hyperuricemia, which is a high risk factor leading to gouty arthritis, nephropathy, and cardiovascular disease. It is generally believed that a female serum uric acid concentration greater than 6 mg/dL or a male serum uric acid concentration greater than 7 mg/dL can be preliminarily determined as hyperuricemia.

The drugs currently used to reduce serum uric acid concentration can be divided into two categories. One is drugs that inhibit the synthesis of uric acid (such as Allopurinol tablets and Febuxostat film-coated tablets), and the other is to promote Drugs that excrete uric acid (for example, Probenecid tablets and Benzbromarone tablets under the trade names). However, long-term use of these drugs is prone to the following side effects. For example, long-term use of Anloprinol tablets may cause Stevens-Johnson syndrome or toxic epidermolysis, and the fatality rate of toxic epidermolysis is as high as 20% to 25%. Long-term use of Fubitong film-coated tablets can easily cause cardiovascular-related symptoms, such as heart disease or stroke. Long-term use of Piloxis tablets and Youlikang tablets are prone to urinary calculi, which is not suitable for patients with poor renal function.

Past studies on probiotics have shown that the use of composite strains (such as Lactobacillus acidophilus and Lactobacillus rhamnosus ) can indeed reduce hyperuricemia. However, the interaction or antagonism between the compound strains is very complicated. For compound strains composed of different strains, it is impossible to predict which combination of compound strains is effective for specific indications without experimental confirmation. As for the treatment of hyperuricemia with a single strain of probiotics, there are few relevant studies so far.

In view of this, it is urgent to provide a single probiotic strain for reducing serum uric acid to solve the above problems.

Therefore, one aspect of the present invention is to provide a use of Lactobacillus reuteri strain GKR1 for preparing a composition for lowering uric acid.

According to the above aspect of the present invention, a use of Lactobacillus reuteri strain GKR1 for preparing a composition for lowering uric acid is proposed. The composition contains an effective dose of Lactobacillus reuteri strain GKR1, and the Lactobacillus reuteri strain GKR1 was deposited on February 12, 2018 at the Biological Resource Center (BCRC, Food Industry Development Institute, No. 331, Food Road, Hsinchu, Taiwan). ), the registration number is BCRC 910827.

According to the above-mentioned embodiment of the present invention, the above-mentioned composition includes a culture containing Lactobacillus reuteri strain GKR1, a liquid containing the culture, a liquid concentrate and/or a dried substance.

According to the above-mentioned embodiment of the present invention, the above-mentioned composition is an oral composition.

According to the above-mentioned embodiment of the present invention, the above-mentioned composition is a food composition or a medical composition.

According to the above-mentioned embodiments of the present invention, the above-mentioned composition further comprises food or pharmaceutically acceptable carriers, excipients, diluents, adjuvants and/or additives.

According to the above-mentioned embodiment of the present invention, the dosage form of the above-mentioned composition is aqueous solution, suspension, dispersion, emulsion, hydrogel, gel, solid lipid nanoparticle, lozenge, granule, powder and/or capsule.

According to the above-mentioned embodiment of the present invention, when the composition is administered to mice, the effective dose of Lactobacillus reuteri strain GKR1 in the composition is at least 0.2 g per kilogram of body weight per day.

According to the above-mentioned embodiment of the present invention, when the composition is administered to an adult, the effective dose of Lactobacillus reuteri strain GKR1 in the composition is at least 0.016 g per kilogram of body weight per day.

After the composition of the present invention containing Lactobacillus reuteri strain GKR1 is administered to a subject, serum uric acid can be reduced, thereby effectively slowing down and/or preventing hyperuricemia.

101/103/105/107/109/111/201/203/205/207/209/211‧‧‧Straight

In order to make the above and other objectives, features, advantages and embodiments of the present invention more obvious and understandable, the detailed description of the attached drawings is as follows:

[Fig. 1] is a bar graph showing the survival rate of strains in a culture solution of different pH according to an embodiment of the present invention.

[Figure 2] is a bar graph showing the survival rate of strains in a 0.3 wt% bile salt culture solution according to an embodiment of the present invention.

[Figure 3] is a bar graph showing the serum uric acid concentration according to an embodiment of the present invention.

Based on the foregoing, the present invention provides a use of Lactobacillus reuteri strain GKR1 for preparing a composition for reducing uric acid, wherein the composition contains an effective dose of Lactobacillus reuteri strain GKR1, and Roy The Lactobacillus strain GKR1 was deposited at the Biological Resource Center (BCRC) of the Food Industry Development Research Institute, No. 331 Food Road, Hsinchu, Taiwan on February 12, 2018, with the deposit number BCRC 910827.

In one embodiment, the above-mentioned composition may be a culture containing Lactobacillus reuteri strain GKR1, a liquid containing the culture, a liquid concentrate and/or a dried substance. The aforementioned culture refers to the product obtained after the Lactobacillus reuteri strain GKR1 undergoes a culture step. The cultivation method is not particularly limited, and may be conventional solid-state cultivation or liquid cultivation. The aforementioned liquid may be a liquid obtained by adding nutrient solution or similar substances to the culture after the culture step. The aforementioned concentrate refers to the product obtained after the culture and/or liquid is subjected to a conventional concentration treatment method (such as an evaporation concentration method or a membrane concentration method). The aforementioned dried product refers to the product obtained by the culture and/or liquid after the conventional water removal treatment method (such as freeze-drying method, vacuum drying method or spray drying method).

The aforementioned composition may include an oral composition, and the type of the oral composition is not particularly limited, as long as it contains the Lactobacillus reuteri strain GKR1.

In one embodiment, the composition may be, for example, a food composition or a medical composition. In one embodiment, the composition may optionally include food or pharmaceutically acceptable carriers, excipients, diluents, adjuvants and/or additives, such as solvents, emulsifiers, suspending agents, disintegrating agents, Binders, stabilizers, chelating agents, diluents, gelling agents, preservatives, lubricants and/or absorption delaying agents, etc.

The dosage form of the composition of the present invention is not particularly limited. In one embodiment, the dosage form of the composition is an aqueous solution, suspension, dispersion, emulsion (single Phase or multiphase dispersion system, single or multilamellar liposomes), hydrogels, gels, solid lipid nanoparticles, lozenges, granules, powders and/or capsules, etc.

The aforementioned food composition can be, for example, but not limited to, cereal products, fruit products, vegetable products, meat products, fish products, egg products, dairy products, beverages, health foods, health foods, functional foods, and nutritional supplements. Food or special nutritious food.

When the above composition is used, the route of administration is not particularly limited, and it can be administered, for example, orally, and adjusted according to actual needs. The dosage and number of dosages of the above-mentioned composition can also be adjusted flexibly according to demand. In one embodiment, when the above composition is administered to mice, the effective dose of Lactobacillus reuteri strain GKR1 in the composition is at least 0.2 g per kilogram of body weight per day. When the composition is administered to an adult, the effective dose of Lactobacillus reuteri strain GKR1 in the composition is at least 0.016 g per kilogram of body weight per day. [Estimating the maximum safe starting dose in initial clinical trials for therapeutics in adult healthy volunteers announced by the U.S. Food and Drug Administration in 2005, based on the recommended intake per kilogram of body weight per day (/kg bw/ The 12.3 times of d) is converted to 1 times the dose of mice]. The number of doses can be administered once a day or divided into several doses.

Experiments have confirmed that the composition of the present invention containing Lactobacillus reuteri strain GKR1 can reduce serum uric acid after being administered to a subject, thereby effectively slowing and/or preventing hyperuricemia.

Several embodiments are used below to illustrate the application of the present invention, but they are not used to limit the present invention. Those with ordinary knowledge in the technical field of the present invention can make various modifications and changes without departing from the spirit and scope of the present invention. Retouch.

1. The nature of Lactobacillus reuteri strain

(1) Preparation of Lactobacillus reuteri strain GKR1

The Lactobacillus reuteri strain GKR1 was deposited in the Bioresource Center (BCRC) of the Food Industry Development Research Institute, No. 331 Food Road, Hsinchu, Taiwan on February 12, 2018, with the deposit number BCRC 910827.

(2) Evaluation of the acid tolerance of Lactobacillus reuteri strain GKR1

The Lactobacillus reuteri strain GKR1 and the commercially available Lactobacillus reuteri strains (BCRC 80379, BCRC 80063, BCRC 17477, BCRC 16091 and BCRC 14625) were tested for acid resistance. First, add or not add HCl to MRS broth (De Man, Rogosa & Sharpe broth, MRS broth), so that the pH of the broth is pH 6.5 (without HCl group), pH 3.2, pH 2.4 and pH 2.0. Then, after the aforementioned strains were activated, they were added to culture solutions with different pH values, and after culturing at 37°C for 3 hours, the number of colonies was counted. Taking the survival rate of the strain at pH 6.5 (without adding HCl) as 100%, the survival rate of the strain in the culture solution of different pH value is calculated.

Please refer to Figure 1, which shows a bar graph of the survival rate of strains in the culture medium with different pH according to an embodiment of the present invention, where the X axis from left to right is the pH of the culture medium pH 6.5, pH 3.2, pH 2.4 and pH 2.0, The Y-axis is the cell survival rate (%), the bar 101 represents the survival rate of Lactobacillus reuteri strain GKR1, the bar 103 represents the survival rate of Lactobacillus reuteri strain BCRC 80379, and the bar 105 represents the survival rate of Lactobacillus reuteri The survival rate of strain BCRC 80063, bar 107 represents the survival rate of Lactobacillus reuteri strain BCRC 17477, bar 109 represents the survival rate of Lactobacillus reuteri strain BCRC 16091 and bar 111 represents the survival rate of Lactobacillus reuteri strain BCRC Survival rate of 14625.

The results in Figure 1 show that when the pH of the culture solution is pH 3.2, pH 2.4, and pH 2.0, the cell survival rate of the Lactobacillus reuteri strain GKR1 (straight 101) of the present invention is higher than 90%, 67% And 8%, its survival rate is higher than that of commercially available Lactobacillus reuteri.

(3) Evaluation of the bile salt tolerance of Lactobacillus reuteri strain GKR1

The Lactobacillus reuteri strain GKR1 and commercially available Lactobacillus reuteri strains (BCRC 80379, BCRC 80063, BCRC 17477, BCRC 16091 and BCRC 14625) were tested for bile resistance. First, add or not add 0.3% by weight of bile salt to MRS (De Man, Rogosa & Sharpe broth, MRS) culture solution, and after soaking at 37°C for half an hour, count the number of colonies. The survival rate of the strain in the culture solution without bile salt is 100%, and the survival rate of the strain in the culture solution containing 0.3% by weight of bile salt is calculated.

Please refer to FIG. 2, which shows a bar graph of the survival rate of strains in a 0.3% by weight bile salt culture solution according to an embodiment of the present invention, where the X-axis from left to right is without bile salt and bile salt added. Salt culture solution, the Y axis is the cell survival rate (%), the bar 201 represents the survival rate of Lactobacillus reuteri strain GKR1, and the bar 203 represents the survival rate of Lactobacillus reuteri strain BCRC 80379. Viability, bar 205 represents the survival rate of Lactobacillus reuteri strain BCRC 80063, bar 207 represents the survival rate of Lactobacillus reuteri strain BCRC 17477, and bar 209 represents the survival rate of Lactobacillus reuteri strain BCRC 16091 And the bar 211 represents the survival rate of Lactobacillus reuteri strain BCRC 14625.

The results in Figure 2 show that the cell survival rate of the Lactobacillus reuteri strain GKR1 of the present invention in the culture medium supplemented with 0.3% by weight of bile salt (as shown in the bar 201) is significantly higher than that of the commercially available Lactobacillus reuteri .

2. Establish the animal model of hyperuricemia mice

(1) Induction of hyperuricemia mice

The mouse was purchased from the Philadelphia Institute of Cancer Research (Institute of Cancer Research) ICR strain of BioLASCO (BioLASCO Taiwan Co., Ltd., Taiwan). The mouse was 5 weeks old and weighed 27.5. g to 32.5g. The mice were kept at a temperature of 22±3°C, a humidity of 40% to 70%, 12 hours of light and 12 hours of dark cycle light, and provided sufficient feed and sterile reverse osmosis water for the mice to eat freely. After 1 week of domestication and adaptation, animal experiments were carried out. The experiment process was carried out in accordance with the guidelines for the care and use of laboratory animals of the Agriculture Committee and the relevant regulations of the Taiwan Animal Protection Law.

Before the animal experiment, the mice were randomly divided into 4 groups, 8 in each group, one cage for each 4, and 2 cages for each group. The control group was given normal feed intake. In the negative control group, 2.5% by weight of potassium oxonate and 1% by weight of RNA were given daily to mice mixed with feed to induce hyperuricemia, but no drugs were given. The positive control group uses the same method as the negative control group to induce hyperuricemia, and is fed 5 tubes a day mg/kg b.w. of Anloprinol tablets. The Lactobacillus reuteri (GKR1) group used the same method as the negative control group to induce hyperuricemia, and was fed 0.205g/kg b.w. Lactobacillus reuteri (GKR1) daily.

The experiment was conducted for 7 days. After the experiment, 0.5 mL to 0.8 mL of blood was collected from the orbital socket and the serum uric acid concentration was analyzed.

(2) Assess the basic physiological parameters of mice

During the experiment, the weight change and food intake of the mice were recorded every day, and the changes of various basic physiological parameters of the mice were observed. Table 1 shows the weight change of the mice during the experiment period, in grams (g). The data statistics use the Duncan test method. Different lowercase English letters indicate significant differences between groups (p<0.05), and the same lowercase English letters indicate that there is no significant difference between the groups (p>0.05). Table 1 shows that on the 0th day and the 7th day, there was no significant difference in the body weight of the mice between each group.

Table 1

Table 2 shows the weight changes of the mice on the 7th day of the experiment, and the food intake of the mice in each group has no significant difference.

Table 2

The above physiological parameters showed that the ingestion of Lactobacillus reuteri strain GKR1 did not affect the growth status of mice.

3. The effect of Lactobacillus reuteri strain GKR1 on reducing serum uric acid

The serum uric acid concentration was measured with a commercially available uric acid quantitative kit (QuantiChrom Uric Acid Assay Kit, BioAssay Systems, USA) to evaluate the mouse serum uric acid (UA) concentration.

Please refer to FIG. 3, which shows a bar graph of serum uric acid concentration according to an embodiment of the present invention. The X axis from left to right is the control group, negative control group, positive control group and Lactobacillus reuteri (GKR1) group. The Y axis is the serum uric acid concentration in mg/dL.

The results in Figure 3 show that compared with the negative control group, the serum uric acid concentration of the Lactobacillus reuteri (GKR1) group was significantly reduced.

It can be seen from the above-mentioned embodiments that the use of the Lactobacillus reuteri strain GKR1 of the present invention for the preparation of a composition for reducing uric acid has the advantage that it contains a single strain of Lactobacillus reuteri strain GKR1 After being administered to a subject, the composition can reduce serum uric acid, thereby effectively slowing down and/or preventing hyperuricemia.

It should be understood that although the present invention uses male mice to establish an animal model of hyperuricemia, it should not be used to limit the sex of the subject to which the Lactobacillus reuteri strain GKR1 is administered. Anyone with ordinary knowledge in the technical field to which the present invention pertains should understand that the effect of reducing serum uric acid by Lactobacillus reuteri strain GKR1 has been confirmed in male mice, and that Lactobacillus reuteri strain GKR1 has the same or similar effect to female mice. Effect.

It should be added that although the present invention takes a specific manufacturing process and/or a specific analytical method as an example to illustrate the use of the Lactobacillus reuteri strain GKR1 of the present invention for the preparation of a composition for reducing uric acid, the present invention Anyone with ordinary knowledge in the technical field to which the present invention belongs will know that the present invention is not limited to this. Without departing from the spirit and scope of the present invention, the composition of the present invention containing Lactobacillus reuteri strain GKR1 can also be used Other processes, other analytical methods or other instruments.

Although the present invention has been disclosed in several embodiments as above, it is not intended to limit the present invention. Anyone with ordinary knowledge in the technical field to which the present invention belongs can make various modifications without departing from the spirit and scope of the present invention. Modifications and modifications, therefore, the scope of protection of the present invention shall be subject to the scope of the attached patent application.

【Biological Material Deposit】

The Lactobacillus reuteri strain GKR1 was deposited at the Biological Resource Center (BCRC), Food Industry Development Research Institute, No. 331, Food Road, Hsinchu, Taiwan on February 12, 2018, and the deposit number is BCRC 910827.

Claims (8)

A use of Lactobacillus reuteri strain GKR1 for preparing a composition for reducing uric acid, wherein the composition comprises an effective dose of Lactobacillus reuteri strain GKR1, and the Lactobacillus reuteri strain GKR1 It was deposited at the Biological Resource Center (BCRC), Food Industry Development Research Institute, No. 331, Food Road, Hsinchu, Taiwan on February 12, 2018, and the deposit number is BCRC 910827. The use of the Lactobacillus reuteri strain GKR1 described in item 1 of the scope of patent application for preparing a composition for lowering uric acid, wherein the composition includes a culture containing the Lactobacillus reuteri strain GKR1, containing the culture A liquid of the substance, a concentrate of the liquid and/or a dry substance. The use of the Lactobacillus reuteri strain GKR1 described in item 1 of the scope of patent application for preparing a composition for lowering uric acid, wherein the composition is an oral composition. The use of the Lactobacillus reuteri strain GKR1 described in item 1 of the scope of patent application for preparing a composition for reducing uric acid, wherein the composition is a food composition or a pharmaceutical composition. The use of the Lactobacillus reuteri strain GKR1 described in item 4 of the scope of patent application for the preparation of a composition for lowering uric acid, wherein the composition further comprises a food or a pharmaceutically acceptable carrier and excipient , A diluent, an adjuvant and/or an additive. The use of the Lactobacillus reuteri strain GKR1 described in item 1 of the scope of patent application for the preparation of a composition for reducing uric acid, wherein one of the dosage forms of the composition is an aqueous solution, a suspension, a dispersion, an emulsion, One hydrogel, one gel, one solid lipid nanoparticle, one lozenge, one granule, one powder And/or a capsule. The use of the Lactobacillus reuteri strain GKR1 described in item 1 of the scope of patent application for preparing a composition for lowering uric acid, wherein when the composition is administered to a mouse, the Lactobacillus reuteri strain GKR1 is in the The effective dose of the composition is at least 0.2 g per kilogram of body weight per day. The use of the Lactobacillus reuteri strain GKR1 described in item 1 of the scope of patent application for preparing a composition for lowering uric acid, wherein when the composition is administered to an adult, the Lactobacillus reuteri strain GKR1 is in the composition The effective dose of the substance is at least 0.016 g per kilogram of body weight per day.

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