TWI721262B - Use of Lactobacillus paracasei strain for preparing composition for improving intestinal microflora - Google Patents

Abstract

The present invention relates to the use of a Lactobacillus paracasei strain for preparing a composition for improving the intestinal microflora, wherein the Lactobacillus paracasei strain is Lactobacillus paracasei Yb (Lactobacillus paracasei Yb) with a deposit number of BCRC 910415. The composition prepared by the Lactobacillus paracasei strain of the present invention has the effect of improving the intestinal microflora.

Description

Use of Lactobacillus paracasei strain for preparing composition for improving intestinal microflora

The present invention relates to the use of a Lactobacillus paracasei strain, in particular to the use of a Lactobacillus paracasei Yb (Lactobacillus paracasei Yb) with a deposit number of BCRC 910415 for preparing a composition for improving intestinal microflora.

The intestine is the most important digestive organ and immune organ of the human body. There are about 100 trillion intestinal bacteria in the intestine, including probiotics, bad bacteria and intermediate bacteria. Probiotics can help in the intestines. The synthesis of multiple vitamins, regulate the intestinal immune system and help smooth defecation and avoid the accumulation of toxins in the body; these intestinal bacteria compete with each other in the intestine to form a dynamic balance. Such intestinal bacteria symbiotically live in the intestinal ecology. It is called "intestinal flora". Because the intestinal flora changes dynamically through diet, if the ratio of probiotics to bad bacteria in the intestine is changed, the intestinal flora will be out of balance, which can lead to diseases such as irritable bowel, allergies, odorous dermatitis or malignant tumors. .

Lactobacillus is a kind of probiotics. The most frequently mentioned lactic acid bacteria are mainly: (1) Lactobacillus sp., such as Lactobacillus acidophilus , Lactobacillus casei and Lactobacillus casei ( Lactobacillus paracasei ); and (2) Bifidobacterium sp., such as Bifidobacterium longum and Bifidobacterium bifidum . Many of these studies have pointed out that Lactobacillus paracasei has the ability to lower blood cholesterol, regulate immune response or treat allergy-related diseases. The Republic of China Patent No. I403327 Announcement A Lactobacillus paracasei with the deposit number BCRC 910482 and its use in the preparation of a food composition or pharmaceutical composition for lowering blood cholesterol; The Republic of China Patent No. I418356 Announcement can be used as an immunomodulator Lactobacillus paracasei LT12; Republic of China Patent No. I398259 announced a composition of Lactobacillus paracasei GMNL-133 for improving atopic dermatitis or other allergic diseases and its use. However, it is not seen in the prior art that Lactobacillus paracasei strain Yb has the effect of improving the intestinal microflora.

Although lactic acid bacteria have been researched and developed for many years, there are still many strains that are still under identification and classification; even if they are the same strains, their biochemical characteristics and achievable effects will be different because of the different gene sequences of different strains. Not the same.

In response to the public's demand for maintaining intestinal health, the purpose of the present invention is to provide a Lactobacillus paracasei strain for preparing a composition for improving the intestinal microflora, wherein the composition has the effect of improving the intestinal microflora.

In order to achieve the above-mentioned object of the invention, the technical means adopted by the present invention is to provide a Lactobacillus casei strain for preparing a composition for improving the intestinal microflora, wherein the Lactobacillus casei strain is Lactobacillus paracasei strain Yb ( Lactobacillus paracasei Yb), the registration number is BCRC 910415.

Preferably, the composition contains 10 ⁷ CFU/mL to 10 ⁸ CFU/mL of the Lactobacillus casei strain Yb.

Preferably, the Lactobacillus paracasei strain is a live strain.

According to the present invention, the improvement of the intestinal flora refers to the tendency of increasing the bacterial count of the intestinal lactic acid bacteria and reducing the bad bacterial count in the intestinal tract. As exemplified in the present invention, the improvement of the intestinal flora can be obtained by administering a composition containing Lactobacillus paracasei in a specific range and measuring the changes in the intestinal flora within a specific time range.

Preferably, the said composition is a drink, a snack or a health food. More preferably, the beverage is fermented milk.

Preferably, the said composition comprises 15% to 17% (weight to volume ratio) of skimmed milk powder, 0.05% to 0.2% (weight to volume ratio) of yeast powder and 8% to 10% (volume ratio) of The high fructose liquid is dissolved in water and mixed uniformly to obtain a milk. The milk ^{is homogenized at 180 kg/cm 2} and then sterilized at 120ºC for 4 seconds; at 90ºC to 100ºC (preferably 93ºC to 95ºC) Carry out the browning reaction on the milk for 1.5 hours to 3 hours (preferably 2 hours) to obtain a browned milk; then ^{add the lactobacillus paracasei strain Yb with an inoculation amount of 10 6} CFU/mL to 10 ⁷ CFU/mL The browned milk is fermented at 34°C to 36°C for 48 hours to obtain a composition.

More preferably, the aforementioned composition is fermented for 48 hours under a fermentation condition of 35±1ºC, and the acidity of the composition at the end of the fermentation is 2.4±0.2%, and then it is stirred and cooled to below 10ºC to obtain the composition.

According to the present invention, the browning reaction refers to a non-enzymatic browning reaction. During the heating process of the milk, the sugars in the milk will undergo Maillard reaction and caramelization with amino acids. caramelization), so that the browned milk has a burnt taste and a strong taste.

The Lactobacillus paracasei strain Yb of the present invention has the effect of improving the intestinal microflora, and the present invention proves that the composition containing the Lactobacillus paracasei strain Yb can increase the flora of Lactobacillus in the intestine to improve the intestinal microflora.

The following describes the technical means adopted by the present invention to achieve the objective in conjunction with the drawings and the preferred embodiments of the present invention, but these embodiments are not limited to the content disclosed in the foregoing of the present invention. Those who learn the skills of the present invention, Some improvements and modifications can be made without departing from the scope of the present invention.

Preparation example 1 To Lactobacillus paracasei Yb Preparation of composition for improving intestinal microflora

Dissolve 15% to 17% skimmed milk powder, 0.05% to 0.2% yeast powder, and 8% to 10% high fructose liquid in water and mix well to obtain a milk; brown the milk at 90ºC to 100ºC After 2 hours to 3 hours, a browned milk is obtained; then, the lactobacillus paracasei strain Yb is inoculated with an amount of 10 ⁶ CFU/mL to 10 ⁷ CFU/mL into the browned milk at 34ºC to 36ºC. Fermentation is carried out for 48 hours to obtain a composition containing Lactobacillus paracasei strain Yb.

Example 1 To Lactobacillus paracasei Yb The effect of prepared fermented milk on improving intestinal flora changes

In this example, 30 subjects with moderate to severe constipation symptoms are invited to perform a 40-day experiment: Before the experiment starts, it is necessary to pass a stable period of 10 to 15 days. During the stable period, the subjects It is necessary to cooperate with dietary instructions, obey the contents of the "Daily Dietary Guidelines" and "National Dietary Guidelines" of the subjects' brochures, and avoid overeating or other special food intakes, and do not take intestinal and stomachic medicines and supplements , Antibiotics, histamine-2 antagonists, antioxidants, functional foods that can regulate the intestines and stomach, or Chinese herbal medicines and other unknown drugs. If the subject is unable to continue the experiment due to severe constipation, cold or other diseases, the gastroenterologist participating in this study must evaluate the diagnosis and determine whether to withdraw from the study. This experiment is a single-blind experiment, that is, after the subjects have passed the screening, they need to cooperate with dietary guidance during the experiment. The experimental schedule is planned for the first, middle and last stages. That is, the 10 days before the experiment is the stable period, that is, the experiment from day -10 to day 0. It is to adjust the dietary habits of the subjects and not violate the dietary guidelines of the briefing session, and start to fill in the defecation questionnaire to observe the cooperation of the subjects Those with poor coordination will be strictly selected and excluded during this period; followed by the placebo drinking period. Days 1 to 10 are the placebo period (10 days in total). The subjects will be treated daily during the placebo period. Drink 300 mL of fermented milk without Lactobacillus casei strain Yb; then enter the experimental period from day 11 to day 30 (20 days in total), and the subject will drink 300 mL of Lactobacillus casei strain daily during the experiment The fermented milk of Yb (the bacterial count of Lactobacillus paracasei strain Yb is 10 ⁷ -10 ⁸ CFU/mL); finally enters the emptying period from the 31st to the 40th day (10 days in total), and the subjects emptied Stop drinking fermented milk containing Lactobacillus paracheis strain Yb within 10 days from the start of the period. In the subject during the experiment for collecting stool, the stool specimen detecting a change in the number belonging to the probiotics Lactobacillus bacteria (Lactobacillus sp.); Bad bacteria belonging to the Clostridium perfringens (Clostridium perfringens) and Escherichia coli (Escherichia coli ) changes in the number of bacteria, and perform statistical analysis.

The change in the number of Lactobacillus sp. is shown in Figure 1. It can be seen that compared with the placebo period, the number of Lactobacillus bacteria in the stool samples of the subjects entering the experimental period increased significantly. The fermented milk of Lactobacillus casei strain Yb has the effect of increasing the number of lactic acid bacteria in the human intestinal tract, that is, it has the effect of increasing the probiotics in the intestinal tract to improve the intestinal microflora. In addition, the number of lactobacilli in the stool sample during the emptying period was also significantly increased compared with the placebo group, and the results were similar to the number of lactobacilli in the experimental group, which can be inferred from the Lactobacillus paracasei strain Yb It can grow steadily in the human intestine during the period, and further balance the intestinal flora. The changes in the bacterial counts of Clostridium perfringens and Escherichia coli are shown in Figure 2 and Figure 3, respectively. It can be seen that the two bad bacteria gradually decreased after the subjects drank the fermented milk containing Lactobacillus paracasei strain Yb. Trend, it can be inferred that besides increasing the number of probiotics, Lactobacillus paracasei strain Yb also has the effect of reducing bad bacteria.

Example 1 Lactobacillus paracasei Yb Test of acid resistance and bile salt resistance of prepared fermented milk

In this embodiment, the fermented milk containing the Lactobacillus accessory casei strain Yb, the fermented milk containing the Daitian bacteria (commercially available Yakult) and the fermented milk containing the probiotic LP33 were further tested for acid resistance and bile resistance.

The acid resistance test procedure is a well-known technique in the field, so the following briefly describes the experimental procedure: pick a single colony and inoculate it in MRS broth, incubate overnight at 37ºC, then replace 1 ml of the test solution, centrifuge at 6000 rpm for 5 minutes, and Add different pH values (adjusted with 0.1 N hydrochloric acid) to 9 ml PBS, mix well, and incubate at 37ºC for 1 hour. After the culture is completed, take 1 ml of the mixed solution and add 9 ml of sterile water to dilute, and then take 100 µl of the diluted bacteria solution and inoculate it on the MRS culture plate, incubate at 37ºC for 48 hours, and finally calculate the number of surviving colonies.

The bile salt resistance test procedure is a well-known technique in the field, so the following brief description of the experimental procedure: pick a single colony and inoculate it in MRS broth, incubate overnight at 37ºC, then replace 1 ml of the test solution, centrifuge at 6000 rpm for 5 minutes , And add them to MRS broth containing 0.3% bile salts. After culturing for 0, 4, 8 and 24 hours, take 1 ml of the mixed solution and dilute with 9 ml of sterile water, and then inoculate 100 µl of the diluted bacterial solution on the MRS culture plate. Incubate at 37ºC for 48 hours, and finally count the number of surviving colonies, measure the number of remaining bacteria, test the survival of lactic acid strains under the culture conditions containing bile salts, and evaluate whether their growth is inhibited in the medium containing bile salts.

The results are shown in Table 1 and Table 2 below. Regardless of the results of the acid tolerance test or the bile salt tolerance test, the Lactobacillus paracasei strain Yb has better acid tolerance and bile salt tolerance than the other two strains.

Table 1 Acid resistance test results

Table 2 Results of bile salt tolerance test

Figure 1 is a histogram of the changes in the number of Lactobacillus bacteria in the feces of subjects during different experimental periods; the placebo period is the period of drinking fermented milk without Lactobacillus paracasei Yb (total 10 days), and the experimental period is drinking The period of fermented milk containing Lactobacillus paracasei Yb (total 20 days), the emptying period is the period of drinking the fermented milk containing Lactobacillus paracasei Yb for 20 consecutive days (total 10 days); the letters in the figure indicate different There is a significant difference between the two at p <0.05. Figure 2 is a histogram of Clostridium perfringens in the feces of subjects during different experimental periods; the same as the experimental description in Figure 1. Figure 3 is a histogram of E. coli in the feces of subjects during different experimental periods; the same as the experimental description in Figure 1.

The Lactobacillus casei strain Yb was deposited in the Bioresource Conservation Center of the Food Industry Development Institute on December 17, 2008. The deposit code of the Lactobacillus casei strain Yb is BCRC 910415.

Claims (4)

The use of a Lactobacillus paracasei strain for preparing a composition for improving intestinal microflora, wherein the Lactobacillus paracasei strain is Lactobacillus paracasei Yb (Lactobacillus paracasei Yb), and the deposit number is BCRC 910415, wherein the composition is The Lactobacillus casei strain Yb containing 10 ⁷ CFU/mL to 10 ⁸ CFU/mL, and the improvement of the intestinal flora includes increasing the number of fecal Lactobacillus bacteria. The use according to claim 1, wherein the Lactobacillus casei strain is a live strain. The use according to claim 1, wherein the composition is a drink, a snack or a health food. The use according to claim 3, wherein the beverage is fermented milk.

Images