JPH02249482A - Extract of lactic acid bacterium multiplication-accelerating plant - Google Patents

Abstract

PURPOSE:To provide the subject extract accelerating the multiplication of valuable enterobacteria such as Bifidobacterium selectively inhibiting the multiplication of enterobacteria such as Clostridium bacteria and having remarkably high safety, comprising the solvent extract of an Araliaceae plant. CONSTITUTION:An Araliaceae plant (e.g. Panax ginseng) is extracted with a solvent (e.g. a lower alcohol such as methyl alcohol) to provide th objective extract. The Araliaceae plant is concretely extracted with the solvent at room temperature and the extracted solution is filtered, followed by distilling off the solvent to provide the objective extract.

Description

DETAILED DESCRIPTION OF THE INVENTION [Industrial Application Field] This invention relates to a lactic acid bacteria growth-promoting plant extract comprising a solvent extract of a plant of the Araliaceae family.

[Conventional technology]

Many types of bacteria live in the human intestine, forming the local flora, which is an extremely complex ecosystem.The composition of this intestinal flora reflects intestinal metabolism, which in turn has various effects. In other words, the dietary components that are ingested every day and the biological components secreted and excreted in the intestines are converted into various substances, and as a result, they have an extremely large impact on the host's nutrition, physiological function, aging, carcinogenesis, immunity, etc. The intramural flora can have both beneficial and disadvantageous effects on the host's health, increasing the proportion of beneficial intestinal bacteria such as monobifidobacteria in the composition of the intestinal flora, and conversely decreasing the proportion of harmful intestinal bacteria. It is becoming clear that suppression is important for human health and longevity.

Human health is closely related to the bacteria that make up the intestinal flora, and a steady intestinal flora suppresses pathogenic bacteria, but when pathogenic bacteria become dominant, it can lead to a diseased state. Among the intestinal bacteria, beneficial bacteria such as Bifidobacterium are known to have beneficial effects on host infection prevention, nutrition, and intestinal cleansing, while harmful bacteria such as Rostridia spp. are directly or indirectly related to the onset of cancer, liver disease, arteriosclerosis, hypertension, etc. Based on such knowledge, various bifidobacteria-containing preparations and dairy products have been developed in recent years to increase intestinal bifidobacteria.

However, even if Bifidobacterium is taken orally, -
Bifidobacterium does not colonize during boat fishing, and intestinal flora often returns to its original state when intake is discontinued. There is a growing interest in substances that are effective in promoting the growth of bifidobacteria.

Lactulose and other indigestible oligosaccharides are known to have the effect of promoting the growth of bifidobacteria, which are intestinal bacteria closely related to human health. It is commercialized as a functional food.

The growth-promoting effect of saccharides is related to their assimilation by bifidobacteria, but it has been found that for substances other than sugars, pantethine-related compounds in edible carrots have a growth-promoting effect of bifidobacteria.

[Problem to be solved by the invention]

Highly safe substances that promote the growth of beneficial bacteria in the intestine while suppressing the growth of harmful bacteria in the intestine and are highly safe are useful as ingredients for health-oriented functional foods, but such substances are still lacking. No substance has been developed.

[Means to solve the problem]

The present inventors have worked diligently to search for substances that are extremely safe, promote the growth of beneficial bacteria in the intestines, and inhibit the growth of harmful bacteria in the intestines. We have discovered that a plant solvent extract has the effect of promoting the growth of Bifidobacteria, which is a beneficial bacteria in the intestine, and at the same time has the effect of suppressing the growth of Clostridium bacteria, which is a harmful bacteria in the intestine.

Previous reports have not found any evidence that plant components of the Araliaceae family have the effect of regulating bacterial growth, and this is the first discovery made by the present inventors. It's arrived.

In the present invention, the Araliaceae plants include, for example, Korean ginseng (Panax ginseng), Bamboo-knot ginseng (Panax japonicum), Panax ginseng (Panax nodoginseng), Gokapi (Acanthopanax spp.), and Talano tree (Aralia).・
It refers to living alone (alaria cordata), etc., and the details are shown in Table 1.

Table 1 Six species of Araliaceae plants shown in Table 1 are each extracted with alcohol at room temperature. After extraction, the alcoholic extract is separated by filtration according to a conventional method. As the alcohol used for extraction, lower alcohols such as methyl alcohol, ethyl alcohol, and n-propyl alcohol are preferred from the viewpoint of operability and extraction efficiency.

For the residue remaining after alcohol extraction, add a sufficient amount of distilled water and heat extraction at 80° C. for 1 hour.

After extraction, the extract is separated by filtration to obtain an extract. After extraction, the extract was concentrated under reduced pressure and freeze-dried to obtain an aqueous extract. [Example] (Example 1) 500 g of ginseng root was added with 4N methyl alcohol and extracted by immersion at room temperature for 2 days. The extract was heated in an oven, the methyl alcohol was distilled off, and the extract was dried to obtain 908 g of an extract (alcoholic extract of Korean ginseng).

(Example 2) To the entire amount of the residue after the methyl alcohol extraction of the ginseng root in Example 1, distilled water of No. 51 was added and heated to 80°C.

Extraction was carried out by heating for 1 hour. After filtering the extract, concentrate under reduced pressure,
Freeze-drying yielded 185.5 g of extract (aqueous ginseng extract).

(Example 3) 25g of bamboo knot ginseng root and 500ml of methyl alcohol! was added and extracted by immersion at room temperature for 2 days. The extract was filtered, the methyl alcohol was distilled off, and the extract was dried to obtain 5.3 g of an extract (alcoholic extract of Bamboo ginseng).

(Example 4) 1! Add distilled water and heat to 80℃.

Extraction was carried out by heating for 1 hour. After filtering the extract, the vacuum wave line,
The extract was freeze-dried to obtain 8.4 g of extract (aqueous extract of Bamboo ginseng).

(Example 5) 500 m2 of methyl alcohol was added to 25 g of Panax ginseng root, and the mixture was extracted by immersion at room temperature for 2 days. The extract was filtered, and the methyl alcohol was distilled off and dried to obtain 4.3 g of extract (alcoholic extract of Panax ginseng).

(Example 6) 1j! Distilled water was added thereto and extracted by heating at 80"C for 91 hours. After heating the extract, it was concentrated under reduced pressure and freeze-dried to obtain 12.1 g of extract (water extract of Panax ginseng).

(Example 7) Methyl alcohol 1N was added to 100 g of the trunk bark and root bark of Gokapi, and the mixture was immersed and extracted at room temperature for 2 days. The extract was heated in an oven, the methyl alcohol was distilled off, and the extract was dried to obtain 14.3 g of an extract (alcoholic extract of Gokapi).

(Example 8) 1j! Add distilled water to 80"C, 1
Extracted by heating for hours. After treating the extract with P, it was concentrated under reduced pressure and lyophilized to obtain 8.2 g of extract (water extract of Gokapi).
.

(Example 9) 50g of bark and root bark of tarano tree and 500% of methyl alcohol
mj! The extract was extracted by immersion at room temperature for 2 days.
The methyl alcohol was distilled off and dried to obtain extract 6.2.
(alcoholic extract of tarano tree).

(Example io) 1N distilled water was added to the entire amount of the residue after the methyl alcohol extraction of the bark and root bark of the tarano tree in Example 9, and the mixture was heated at 80°C for 1 hour.
Extracted by heating for hours. After treating the extract with P, it was concentrated under reduced pressure and lyophilized to obtain 3.1 g of extract (aqueous extract of Talano tree).

(Example 11) Add 1 l of methyl alcohol to 50 g of independent roots, and add 1 l of methyl alcohol to
Extracted by immersion for 1 day. The extract was heated in an oven, the methyl alcohol was distilled off, and the extract was dried to obtain 5.8 g of an extract (independent alcohol extract).

(Example 12) 1! Distilled water was added thereto, and extraction was carried out by heating at 80°C for 1 hour. After treating the extract with P, it was concentrated under reduced pressure and lyophilized to obtain 5.6 g of extract (independent water extract).

Test Example 1 The aqueous extract and methyl alcohol extract of the Araliaceae plant of the present invention obtained in Example 1 and Example 12 were tested for their growth promoting effects on Bifidobacterium. The following two types of basal media were used.

[Medium 1] K, HPOa 2.5g lactose 35. OgCH, CO
ONa・3Ht O25, OgBacto Casa
min.

Ac 1ds (Vi tami M-free, Di
rco) 5.0g alanine, L-cysteine.

Tryptophan 200mg each Asparagine 100mg Adenine, Guanine.

Uracil, xanthine 10mg each thiamine
HC1200gg Riboflavin 200gg Pyridoxine-MCI 1200μg Nicotinic acid
600 μg p-aminobenzoic acid 1 folic acid.

Biotin 12.5 μg each Mg 30
4 ・7H* 0 200mgFe5Oa
・7H, 010mg NaC 110mg Mn5Oa 6.74mg Add distilled water to make 1N (adjust to pH 6,8 II), 11s°
Moist heat sterilization for Cl2O minutes.

[Medium 2. Glycolysis test medium (PYF medium) Cotrypticase (BBL) 10g yeast extract (
Di f'co) 10g Digested Blood
40mj2L-Sistine-
HCl, H! 0 0.5g salt solution'
40ml purified water
920mA! Humid heat sterilization at pH 7.6 at 115°C for 20 minutes.

×The composition of the salt solution is CaC1* 0.2g, Mg804
0.2g, Km HPO41g, KHz PO21g
, NaHCOs 10g, NaC 12g and purified water 1
00mj! It is.

For medium 1, to search for non-carbon growth factors,
In the case of medium 2, it was used to search for growth factors derived from hypodermis.

Bifidobacterium longum (Bi
fidobacterium longum E1
94b) and Bifidobacterium breve(9,1)y
Bacteria grown in Briges River Broth or EGF Pross (37"C, 48 hours) were collected by centrifugation (3,000 rpm, 10 minutes), and then added to 10 ml of sterile physiological saline (0, 85% NaCl
, 0.1% L-cysteine-HCl, 0.1% thioglycolic acid) and suspended in 5 mj2 of the above saline solution. Add 2 drops of suspension to both media. A sterile test sample solution and a moist heat sterilized (115°C, 20 minutes) ascorbic acid solution (34 g of ascorbic acid, 10.7 g of JQ acid soda, L-cysteine-MCI Ig dissolved in distilled water xoomj!) were added. to reduce the final liquid volume to 1
0m1. Anaerobically (100% CO
, or 80% Nm + 15% CO* + 5% H,) After culturing, the pH of the culture solution is measured and used as an index of bacterial growth.

The degree of proliferation is divided into four stages: ++ (strong, pH 4, 5 to 5).
．． 0), + (medium, pH 5, 1-5.5), (+) (weak,
pH 5, 6 to 6.0), - (no growth).

Samples (extracts prepared by the methods of Examples 1 to 12) 1. Alcohol extract of Korean ginseng root 2, Water extract of Korean ginseng root 3, Alcohol extract of bamboo knot ginseng root 4, Water extract of bamboo knot ginseng root Product 5, Alcohol extract of 37 ginseng root 6, Water extract of 37 ginseng root 7, Alcohol extract of 37 ginseng root bark and root bark 8, Water extract of 37 ginseng root bark and root bark 9 , alcoholic extract of the bark and root bark of the tarano tree 10, water extract of the bark and root bark of the tarano tree 11, alcoholic extract of the dokkatsu board 12, water extract of the dokkatsu board. The results of the growth response to longum and Bifidobacterium plebe are shown in Table 2.

Bifidobacterium breve was selected because it is the predominant bacterial species in the intestines of infants, and the aqueous extract of the Araliaceae plant is resistant to both bacterial species. It showed a moderate growth promoting effect. However, with regard to methyl alcohol extracts, the growth promoting effect was not observed or only weak in the case of Korean ginseng, Bamboo ginseng, and Ginseng ginseng, but the effect was only weak in promoting growth, whereas Talano ginseng showed a strong effect.

Test Example 2 The test results shown in Test Example 1 show that ginseng has a strong and stable growth-promoting effect on bifidobacteria, and its pharmacological action is well known. ) and the aqueous extract of Example 2 (Sample 2), the relationship between dose and growth response was tested for five representative Bifidobacterium species. The five species are Bifidobacterium longum (Note 1: Test bacterial species Note 1: Basal medium Bifidobacterium longum is Phytobacterium infantis (B, i) in adults, and the results are It is shown in Table 3.

Table 3 Note 1: Basal medium Note 1: Amount of sample added (%) When the water extract of Korean ginseng (test sample 2) is used as the basic medium, 0.1% or more of Bifidobacteria A moderate or higher growth promoting effect was observed on the four test bacterial species except U. bifidum, but when medium 2 was used as the basal medium, no effect was observed at an addition amount of 0.1%.

The methyl alcohol extract of Korean ginseng (test sample 1) only had a weak growth-promoting effect on some bacteria.

Test Example 3 The growth promoting effect of the ginseng water extract of Example 2 (Test Sample 2) with the majority strain number ζ belonging to various bacterial species forming the intestinal flora including Bifidobacterium was tested.

[Test bacterial species] 1. Bifidobacterium adolescentis (B, a
dolescentis) E-298b eE-
319a, M-101-4゜M-601, M-602, S-601゜S-602, U
-601 2, Bifidobacterium breve (B, brave) l-53-8w, S 4
63. Bifidobacterium longum (Dan,!!!!!J!!!!ri Kd-5-6wM-10
1-2. M-601. S-3゜-aoi 4, Bifidobacterium infantes (B, 1
nfantis) I-10-55, Bifidobacterium bifidum (B, bifidum) A
-234-4.

E-319, M-601, S-28a.

S-601, S-602 6, Lactobacillus acidophilus
s) AT CC-4356, Omf'1 7, Lactobacillus livarius (L, 5alivarius) ATCC-117
41.

ATCC-11742 8. Lactobacillus casei (L, cas@i) ATCC-7469.

IFO-3425 9, Lactobacillus gazelli (L, :) F-164, M-601゜10, Bacteroides fragilis 3
876.

11. Bacteroides setaiutamicron (B, t
hetaiotamicron) AS-126
12. Bacteroides pulgaras (Dan, 恕小憇') F-92, S-601゜S-6
02, S-603, S-604゜5i-605, S-6
06 13, Bacteroides distasonis (B, distasonis) B-26, M
-602.

M-603, S-601, U-604 14, /< Cutiloides uniformis (B, uni
B. formis) M-60115, B. melaninogeni
cus) N CT C-933716, Clostridium putilicum 14823, S-601 17, Clostridium coccoides (C, coccoides) B-218, Clostridium difficile (C, difficile) AT CC-9689
19, Clostridium innocuum (C, inn
ocum) M-60120, Clostridium parabutrificum (9, boxed four types and soldiers) B-3-
4゜B-78, VPI-6372 21, Clostridium verfringens (9, snakes) ATC・C-13124゜B-165-16
, C-01 22, Clostridium ramosum (C, ramosum) ATCC-25582.

23. Niebacterium ofaciens (Eu
bacterium aerofaciens)
M-608.

M-609, M-610, 8-601゜S-602, S
-603, 3-604.

S-605, S-606 24, Niebacterium lentum (E, lentum) M-60125, Niebacterium limosum (E, limosum) ATCC-8486.

E-1, VPI-1939 26, Nibacterium nitritogenes (E, ni
tritogenes) ATCC-2554727
, Nibacterium tortuosum (E, tort
uosum) ATCC-2554828, Escherichia coli.

(Escherichia coli) B r −
608.

E-605, M-602, 0-601 29, Mitsuokella multiacidus
NCT C-10934, NCTC-10935
゜P-208-58 30, Mitsuochera murgasidas scold, 憇■煕平4) Fl-376 31, Pebtostrebtococcus anaerobiu (Pe
tostreptococcus anaerobi
us) X-3632, Bebutostrebutococcus
Productus (P, productus) ATC
C-2734033, Pebtostrbutococcus bar plus axis, 1 concave) 1612 34, Pebtostrbutococcus brebochyi (P,
prevotii) AT CC-932135,
Pebtostrebtococcoliticus φ Asaccharolyticus (P, asaccharolyticus) V P
I-5045A36, Streptococcus faecalis
IFO-37, Propionibacterium ae
nes) ATCC-6919, ATCC-11
829 38, Fusobacterium varium P
103-11239, Fusibacterium necrophorum (F, necrophorum) W-12,
201340, Fusibacterium biacutus (F, biacutus) PAS-447
641, Megasphaera aisdenii F
1-37542, Rikenella m1crofusus N
CT C - The number of strains that showed each degree of growth under each condition is shown in Table 4.

Table 4 Note 1: The results on the basal medium and above show that, with the exception of 6 strains of Bifidobacterium bifidum in which no growth was observed, many strains of Bifidobacterium were found in the aqueous extract of ginseng (sample). 2)
It has been shown that proliferation is promoted by Regarding other bacteria, it showed a growth inhibiting effect on Lactobacillus, but no growth or only poor growth was observed against Escherichia coli and Clostridium strains, which are harmful bacteria in the intestines, in the two media.

Test Example 4 Next, the methyl alcohol extract (Sample 1) and water extract (Sample 2) of Korean ginseng obtained in Examples 1 and 2 were tested for their growth inhibiting effects on various bacteria. That is, one platinum loop of test bacteria was suspended in 1 mff1 of physiological saline. 01 ml of this suspension was mixed with 5% horse fluid (Wh
Brucella Agar (
Dif'c.

) applied to the surface of a flat plate. 10mg of sample 1 and sample 2
methyl alcohol (100 μl and water (100 aff)
The solution dissolved in i) was added to a paper disk (Toyo Paper, diameter 8 mm) using a capillary tube. After evaporating the solvent, the disks were placed on the agar surface. The flat plate was heated at 37℃ under a mixed gas of 80% nitrogen gas, 15% carbon dioxide gas, and 5% hydrogen.
, and kept at constant temperature for 2 days. When a growth inhibiting area of 10 mm or more was observed, the inhibitory effect was determined to be positive +, and when it was 10 mm or less, the inhibitory effect was determined to be negative.

[Test strain 1, Bifidobacterium adolescentis (B, a
dolescentis) E-194a2, Bifidobacterium pleve (B, breve) S 1 3, Bifidobacterium longum (Dan, eclipse) E194b 4, Bifidobacterium infantis (B, 1)
nfantis) S 125, Lactobacillus acidophilus, 5 flights 9 hikibuki) ATCC-43566, B. fragilis (B, fragilis) 3676, M-
6017, Bacteroides setaiutamicron (B,
thetaiotamicron) AS-1
268, Bacteroides vulgatus (B, vulgatus) F-92, S-6
019, Bacteroides distasonis (B, distasonis) M-602, S
-60110, Clostridium bifermentans (C, bifermentans) B 1
11. Clostridium butyricum (C, butyricum) ATCC-14823
.

12. Clostridium fucoi°des (C, cocc)
oides) B-213, Clostridium difficile (C, difficile) AT CC-9689
14. Clostridium innocuum (C, inn
ocum) M-60115, Clostridium parabutrificum (C, araputrificum)
m) B-78, VP I-16, Clostridium barfringiens deaf, 4φ force chamber) ATCC
-13124°B-165-16, C-0f 17, Clostridium ramosum (C, ramosum) ATCC-25582.

C-0O 18, Niebacterium aureofaciens (E,
aerofaciens) S-601, S-
603, 8-605 19, Escherichia coli (E, coli) Bf'-606, E-605°F
-604, M-602, 0-601.

V-603° Table 5 shows the number of strains belonging to each classification under each condition.

The results shown in Table 5 indicate that ginseng methyl alcohol extract (sample 1) specifically inhibits the growth of many Clostridium strains, but does not inhibit the growth of Bifidobacteria and many other bacteria. It is shown that.

〔Effect of the invention〕

Claims (4)

[Claims] (1) Lactic acid bacteria growth-promoting plant extract consisting of a solvent extract of Araliaceae plants (2) The lactic acid bacteria growth-promoting plant extract according to claim 1, wherein the solvent is alcohol. (3) The lactic acid bacteria growth-promoting plant extract according to claim 1, which is a residual aqueous extract after alcohol extraction. (4) The lactic acid bacteria growth-promoting plant extract according to claim 1, wherein the lactic acid bacteria are bacteria of the genus Bifidus and Lactobacillus.