KR20240102128A - Composition for promoting absorption of water soluble vitamins by synbiotic - Google Patents

Abstract

The present invention relates to a new strain of Lactobacillus Plantarum Beansflora 1712, a new domestic soybean variety of soybean, developed as a result of research on new breed breeding and cultivation, and specifically to a new strain of Lactobacillus Plantarum Beansflora 1712 isolated from soybean feed. It relates to the lactic acid bacterium Lactobacillus Plantarum Vinceflora 1712 strain (microorganism accession number KCTC19014P).
The strain of the present invention is a new lactic acid bacterium with the characteristics of fermentation ability of various carbohydrates, production ability of various enzymes, antioxidant activity, and enhanced absorption of water-soluble vitamin nutrients, and can be included or fermented in probiotic probiotic preparations, food, cosmetics, and feed. It is very useful as a food fermentation bacteria. It relates to a composition for promoting the proliferation and growth of hair papilla cells containing a fermented soybean extract fermented using. The neulpum soybeans of the present invention have a higher amino acid content than existing soybeans and are highly safe and stable for use as a food raw material. It is easy to use as food, health functional food, and cosmetics.

Description

Method for producing a synbiotic composition for enhancing absorption of water-soluble vitamins {Composition for promoting absorption of water soluble vitamins by synbiotic}

The present invention relates to a new strain of lactic acid bacteria Lactobacillus Plantarum Vinceflora 1712, and specifically to a new strain of Lactobacillus Plantarum Beansflora 1712 isolated from soybean feed (microorganism accession number KCTC19014P).

Recently, as interest in health has increased, intestinal health has begun to receive attention. There are more than 1 trillion bacteria in the human intestine, and the number of bacteria in the intestine is about 10 times more than the number of cells in the human body. The intestinal bacterial community continues to change quantitatively and qualitatively depending on the internal environment such as pH, bile acids, digestive enzymes, and mucins, the external environment such as nutrients and medicines, and bacterial factors. Generally, beneficial bacteria account for 30% and harmful bacteria account for 5 to 10%. Among these, beneficial bacteria are called probiotics.

In a healthy state, most bacteria act as prebiotics that have beneficial effects on the human body. It has been reported that prebiotics not only help intestinal function when consumed in sufficient amounts, but are also effective in activating the immune system, suppressing cancer development, suppressing hypersensitive immune responses, and regulating the microbial balance in the digestive tract.

Probiotics can be defined as microorganisms that are beneficial to health, as the word means ‘for life’ in Greek. So, just as Hippocrates wrote 2,400 years ago that all diseases originate from the intestines, interest in probiotics is increasing even today as many studies have shown that they are closely related to specific diseases.

Today, researchers are not only uncovering the important physiological functions of probiotics, but are also continuously attempting to discover, mass produce, and commercialize new uses of probiotics. Probiotic microorganisms that can be produced using standardized methods can not only be used commercially on their own, but also have a wide range of applications, including use as fermenting bacteria for fermented foods. In addition to discovering new animal-based lactic acid bacteria from dairy products as before, research to secure vegetable-based lactic acid bacteria, which are reported to have strong acid resistance and bile resistance and thus have a high rate of reaching the intestines and colonization, is also receiving great attention.

Accordingly, the present inventors screened new microorganisms for the excellent properties of probiotics from the newly discovered soybean variety, Neulpumkong, and as a result, the new vegetable lactic acid bacterium Lactobacillus Plantarum Beansflora 1712 strain has the fermentation ability of various carbohydrates and the production of various enzymes. The present invention was completed after confirming that it has excellent probiotic properties such as production capacity, antioxidant activity, and enhanced absorption of water-soluble vitamin nutrients.

Accordingly, while researching a new substance that can replace existing hair loss-related preparations, the present inventors discovered a product containing fermented soybean extract fermented using Lactobacillus Plantarum Beansflora 1712 (accession number) lactic acid bacteria as an active ingredient. The present invention was completed by confirming that cell growth was promoted when the composition was administered to human dermal papilla cells.

The purpose of the present invention is to provide a composition for promoting the proliferation and growth of hair papilla cells containing a fermented extract of Neulpum soybean fermented using Lactobacillus Plantarum Beansflora 1712.

Therefore, the main purpose of the present invention is to provide a new strain with excellent properties as a probiotic. Another object of the present invention is to make probiotics using the above strain, and it is very useful for inclusion in food, cosmetics, feed, or as a fermenting bacteria for fermented foods.

Lactobacillus plantarum Vinceflora 1712 strain, a new lactic acid bacterium of the present invention, was isolated from Neulpoom soybean and deposited in the Microorganism Bank (KACC: Korean Agricultural Culture Collection) under the microorganism accession number KCTC19014P.

A plate on which sufficient colonies of the new lactic acid bacteria strain of the present invention had grown was submitted to a 16s rRNA base sequence analysis agency, and as a result, it was confirmed to be a new strain belonging to Lactobacillus plantarum. The strain of the present invention can ferment carbohydrates such as xylose, glucose, and mannose. Therefore, materials containing these carbon sources can be used for fermentation.

The strain of the present invention can produce enzymes such as a-galactosidase and b-glucosidase. Therefore, they can be used in fields that require the activity of these enzymes.

The strain of the present invention can survive in a gastric fluid environment of pH 2-3 and can survive in conditions where bile salts are present at high concentrations, so it has the possibility of surviving while passing through digestive organs such as the stomach and small intestine and reaching the intestines alive. This is high.

The strain of the present invention has excellent antioxidant activity and can be usefully used as a probiotic with antioxidant properties.

The strain of the present invention has properties that enhance the absorption of water-soluble vitamins in the body, and can be usefully used as a nutrient absorption enhancer.

As described above, the strain of the present invention has excellent characteristics as a probiotic and can be used as a probiotic preparation. At this time, the probiotic may be composed of the strain itself or a culture of the strain of the present invention, and may be in the form of excipients, etc. added for ease of formulation.

Additionally, the strain of the present invention or its culture can be used as an ingredient or additive in foods, health functional foods, cosmetics, and feed. At this time, the culture can be used in a form that does not contain bacteria, but in order to demonstrate the effect of probiotics, it is preferable to apply it in a form that contains bacteria.

Additionally, the strain of the present invention can also be used as a fermented strain for fermented foods, fermented cosmetics, or fermented feed. At this time, it is preferable to use raw materials containing ingredients that can ferment the strain of the present invention for fermented food or fermented feed.

Hereinafter, the present invention will be described in more detail through examples. Since these examples are merely for illustrative purposes of the present invention, the scope of the present invention is not to be construed as limited by these examples.

[Example]

1. Method

1-1. Isolation of strains

Samples of neulpoom bean provided by Seoul National University in Seoul, Korea were collected. The area for pre-processing the feed and all consumables to be used were disinfected with 70% alcohol to prevent contamination. The samples were fermented using soaked and unsoaked soybeans at 37 degrees for the growth of lactic acid bacteria for about 24 hours.

Each culture medium was incubated at a concentration of 10³~10? using a sterilized 0.85% NaCl solution. After diluting to , each diluted sample was plated and inoculated on MRS agar plate medium (BPB medium). The pure separation process was conducted so that only one strain could be isolated by culturing for 48 hours under anaerobic conditions at 37 degrees.

The plage with sufficient colonies of the pure cultured strain was submitted to a 16s rRNA sequence analysis agency and sequence analysis was completed. Specifically, the genomic DNA of the strain was extracted and purified using the QIAquick kit (Qiagen Ltd., Crawley, UK), and universal primers 27F (5'-AGA GTT TGA TCG TGG CTC AG-3') and 1492R (3'-GCT) The sequence of the amplification product was analyzed using TAC CTT GTT ACG ACT T-5'). The 16s rRNA sequence was aligned and analyzed using BLAST in the NCBI gene bank database.

Next, only the strains that could be used were selected, inoculated on MRS agar medium, cultured, and when enough colonies had grown, a stock was prepared and completed.

1-2. Resistance analysis to pH environment

Resistance to low pH of Lactobacillus plantarum Vinspolora 1712 strain was investigated according to the method of Ilavenil et al. (J Sci Food Agric. 96:593-601). The cultured strains were inoculated into MRS cultures of various pH and cultured at 37°C for 48 hours. After culturing, 100㎕ of each culture was inoculated into MRS medium using the pour plate method, and the colonies were counted by culturing at 37°C for 48 hours.

1-3. Tolerance analysis for above pH

Resistance to gastric juice of Lactobacillus plantarum Vincellora 1712 strain was investigated according to the method of Charteris et al. (J Appl Microbiol. 84:759-768). Gastric fluid was prepared using pepsin (3 mg/ml), 0.5% (w/v) NaCl, and water, and the pH was adjusted to about 2 and 3 using 1 mol/L HCl. 30 ml of strain culture was centrifuged at 6000xg for 20 minutes at 4°C, and the pellet was obtained and washed three times with K2HPO4 (50 mmol/L, 6.5 pH). After resuspending the pellet, 1 ml of suspension was mixed with 9 ml of gastric fluid and incubated at 37°C for 3 hours. Total viable bacteria before and after culture were counted.

1-4. Analysis of tolerance to concentrated bile salt environment

The culture of Lactobacillus Plantarum Vinspolora 1712 strain was inoculated into MRS liquid medium containing bile salts such as 0.5% sodium deoxycholate and 0.3% Oxgal (DCA Sigma, St Louis, MO, USA) and incubated at 37°C. It was cultured in . Culture liquid was aliquoted at regular time intervals (after 24 and 48 hours) and absorbance was measured at 600 nm. MRS medium without bile salts was used as a control.

1-5. Free radical scavenging power analysis by DPPH method

The DPPH (2,2-Diphenyl-1-picrylhydrazyl) radical scavenging activity of Lactobacillus plantarum Vincellora 1712 strain was measured by applying a slight change to the method of Li et al. (Food Chemistry, 135:1914-1919). .

Various concentrations (10, 20, 30, 40, and 50 μg) of the strain culture (10 9 CFU/ml) were mixed with 1 ml of DPPH solution (0.05mM) and incubated in the dark at room temperature for 30 minutes. DPPH solution was used as a control. The mixture of methanol and strain cells was used as a blank. Absorbance was measured at 517 nm. DPPH radical scavenging activity was calculated using the following formula:

DPPH scavenging activity (%) = [(Sample - Blank) / Control] x 100

1-6. Analysis of water-soluble vitamin absorption enhancement ability

The following experiment was conducted to confirm whether the Lactobacillus plantarum Vinspolora 1712 strain of the present invention has the ability to enhance water-soluble vitamin absorption. Six-week-old C57Bl/6 male rats were acclimated to regular solid feed for 3 days, then divided into the following two groups (control group, experimental group 1 and 2) and fed the experimental diet for 4 days. Afterwards, blood was collected and the biotin content in the blood was measured through HPLC analysis.

Control group: regular solid feed

Experimental group 1: General solid feed + biotin 10mg/kg

Experimental group 2: General solid feed + biotin 10mg/kg + Lactobacillus Plantarum Vinspolora 1712 strain 1x10? concentration powder

2. Results

2-1. Separation and screening

Lactobacillus Plantarum Vincellora 1712 strain was isolated based on distinct morphological characteristics. As a result of analyzing the molecular characteristics of Lactobacillus Plantarum Vincellora 1712 through 16S rRNA sequence analysis, it was found to belong to Lactobacillus plantarum. This 16S rRNA sequence was deposited under the microorganism accession number KCTC19014P.

2-2. Acidic conditions and bile salt tolerance

As a result of measuring whether the Lactobacillus plantarum Vinspolora 1712 strain of the present invention is resistant to various pH environments, it showed excellent resistance even in gastric fluid environments of pH 2 and pH 3. It also showed excellent tolerance even in a bile salt environment with high concentrations of 0.3% oxgall and 0.5% sodium deoxycholate (see Figure 1).

2-3. Antioxidative effect

As a result of measuring the DPPH radical scavenging ability of the Lactobacillus plantarum Vinspolora 1712 strain of the present invention, it was found to have superior free radical scavenging activity compared to vitamin C, a representative antioxidant nutrient (see Figure 2).

2-4. Strengthening effect of water-soluble vitamin absorption

It can be seen that the biotin content in the blood of experimental groups 1 to 2 that consumed biotin increased compared to the control group that did not consume biotin. In particular, it can be seen that the biotin concentration in the blood of experimental group 2 administered with the Lactobacillus plantarum Vinspolora 1712 strain of the present invention increased by about 33% compared to experimental group 1. Through these results, it can be confirmed that the Lactobacillus plantarum Vinspolora 1712 strain of the present invention is effective in increasing the bioavailability of biotin (see Figure 3)

In order to achieve the above object, a fermented extract of Neulpoom soybean fermented using Lactobacillus Plantarum Beansflora 1712 is provided. Hereinafter, the composition according to the present invention will be described in detail.

Since the present invention can be modified in various ways and can take on various forms, embodiments will be described in detail in the text. However, this is not intended to limit the present invention to a specific disclosed form, and should be understood to include all changes, equivalents, and substitutes included in the spirit and technical scope of the present invention.

The composition for enhancing the absorption of water-soluble vitamins according to the present invention contains Lactobacillus Plantarum Vinspolora 1712 strain and can promote absorption of water-soluble vitamins in vivo and increase bioavailability.

In addition, the food composition containing water-soluble vitamins according to the present invention can be implemented in various dosage forms (tablets, capsules, powders, liquids, granules, pills, flakes, pastes, syrups, gels, jellies, caramels, gummies, bars, etc.) Therefore, the convenience of ingestion increases, satisfying consumer preferences, and since there are no restrictions on the ingredient mixing ratio, it can be manufactured as a high-content vitamin preparation or health functional food with multiple functions.

Figure 1 shows the results for acidic conditions and bile salt tolerance.
Figure 2 shows the results of measuring antioxidant efficacy.
Figure 3 shows the results of promoting absorption of biotin, a water-soluble vitamin.

Accordingly, while researching a new substance that can replace existing hair loss-related preparations, the present inventors discovered a product containing fermented soybean extract fermented using Lactobacillus Plantarum Beansflora 1712 (accession number) lactic acid bacteria as an active ingredient. The present invention was completed by confirming that cell growth was promoted when the composition was administered to human dermal papilla cells.

The purpose of the present invention is to provide a composition for promoting the proliferation and growth of hair papilla cells containing a fermented extract of Neulpum soybean fermented using Lactobacillus Plantarum Beansflora 1712.

Therefore, the main purpose of the present invention is to provide a new strain with excellent properties as a probiotic. Another object of the present invention is to make probiotics using the above strain, and it is very useful for inclusion in food, cosmetics, feed, or as a fermenting bacteria for fermented foods.

Claims (5)

Composition for enhancing water-soluble vitamin absorption comprising Lactobacillus Plantarum Vinspolora 1712 strain or a culture thereof The food composition according to claim 1, wherein the food composition is in the form of tablets, capsules, powders, liquids, granules, pills, flakes, pastes, syrups, gels, jellies, caramels, gummies, and bars. The health functional food composition according to claim 1, wherein the food composition is in the form of tablets, capsules, powders, liquids, granules, pills, flakes, pastes, syrups, gels, jellies, caramels, gummies, and bars. The cosmetic composition according to claim 1, wherein the food composition is used for an application selected from cleansing cream, massage cream, nutritional cream, essence, lotion, or pack. The composition according to claim 1, wherein the food composition is used as a fermentation control additive and pellets for animal breeding.