Lactobacillus fermentum Lee working fermenting agent product and application thereof to prevent constipation and keep healthy

Abstract

The invention discloses a lactobacillus fermentum Lee working fermenting agent product and application thereof to prevent constipation and keep healthy. Lactobacillus fermentum Lee is preserved in China Center for Type Culture Collection with the preservation number of CCTCCM2013512. Lactobacillus fermentum Lee is capable of normally growing in human intestinal tracts. Mouse in-vivo tests show that lactobacillus fermentum Lee is capable of alleviating the mouse body weight mitigating trend caused by constipation, accelerating melaena discharging time, improving intestinal propulsive rate, increasing levels of MTL, Gas, ET, AchE, SP and VIP in serum to different degrees, reducing SS factor level, increasing food intake and water intake, and reducing the decrease trend of defecation amount, defecation particle quantity and water content in faeces, gives play to effects similar to that of constipation-treating medicines, and shows certain constipation prevention effect. The optimum usage dosage is 1.0*10<9> CFU for per kg body weight.

Description

technical field

[0001] The present invention relates to the technical field of microorganisms, more specifically, the present invention relates to a kind of Lactobacillus fermentum that can regulate intestinal movement and prevent constipation and use the bacterial strain as a starter Or added fermented milk, health food and animal health products. Background technique

[0002] Lactobacillus is closely related to human life and is one of the beneficial microorganisms widely used in food fermentation, industrial lactic acid fermentation and medical care. Many of the now recognized Lactobacillus and Bifidobacterium genera have specific favorable properties and genetic traits, showing extremely strong probiotic properties. Lactobacillus can not only improve the nutritional value of food, improve the flavor of food, prolong the storage time, but also improve the functional properties of food. It can improve the digestibility and biological value of food and promote digestion a...

Examples

Embodiment 1

[0047] Example 1: Isolation, purification and preliminary identification of Lactobacillus fermentum Lee

[0048] This embodiment is carried out according to the following steps:

[0049] (1) Isolation and purification of Lactobacillus fermentum Lee

[0050] Aseptically draw 500 μL of the sample and add it to 5 mL of sterilized saline to make a uniform dilution of 1:10, and continue to make a certain proportion of dilution. Select the appropriate gradient diluent, draw 100 μL each into the MRS solid plate medium with a sterile pipette tip, and incubate at 30°C for 48-72h, observe and record the colony morphology ( figure 1 ). Use an inoculation loop (or a sterilized toothpick) to pick different colonies from the surface and inside of the plate and inoculate them in MRS liquid medium, culture them on a 300r shaker at 30°C for 24-48 hours; Ran's staining microscope ( figure 2 ), determined to be the continued activation of G+ bacteria until pure colonies were obtained (micro...

Embodiment 2

[0055] Embodiment 2: In vitro screening of Lactobacillus fermentum Lee

[0056] (1) Screening of probiotics resistant to pH 3.0 artificial gastric juice

[0057] Preparation of artificial gastric juice: 0.2% NaCl, 0.35% pepsin, adjust the pH value to 3.0 with 1M HCl, filter and sterilize with a vacuum pump in an aseptic operating table for later use.

[0058] Determination of the tolerance of probiotics to artificial gastric juice: take 5mL of the activated strain culture solution, pour it into a sterilized 10mL centrifuge tube in a sterile operating table, collect the bacteria by centrifugation at 3000r / min for 10min, add 5mL of sterilized Mix normal saline to make bacterial suspension, mix 1mL bacterial suspension with 9mL pH3.0 artificial gastric juice, shake well, place in a constant temperature shaker (37°C, 300r), and take samples at 0h and 3h respectively, Incubate at 37°C for 48h with MRS agar medium. Measure the number of viable bacteria with the plate counting meth...

Embodiment 3

[0070] Example 3: Activated carbon induced constipation experiment

[0071]Feeding healthy Kunming mice, 6 weeks old, weighing 23±2g, 60 females. After feeding the basic diet for 5 days, the mice were randomly divided into 5 groups according to body weight, namely: normal group, constipation control group, 5% RS3 group, 10% RS3 group and 15% RS3 group, with 12 mice in each group. Raise them in stainless steel cages, keep room temperature at 24±2°C, relative humidity at 50±10%, and light-dark rotation for 12 hours (lighting at 8:00-20:00). During the whole experimental period, the normal group and the constipation control group were fed with the basal diet, and the mice in the experimental group were fed with 1.0×10 9 CFU / kg gavage probiotics. Two weeks later, at 9:00 in the morning, the mice in the other five groups except the normal group were gavaged with 10% activated carbon ice water at 2°C, 0.2 mL / mouse, once a day for 3 consecutive days. Until the last day of the expe...

technical field

[0001] The present invention relates to the technical field of microorganisms, more specifically, the present invention relates to a kind of Lactobacillus fermentum that can regulate intestinal movement and prevent constipation and use the bacterial strain as a starter Or added fermented milk, health food and animal health products. Background technique

[0002] Lactobacillus is closely related to human life and is one of the beneficial microorganisms widely used in food fermentation, industrial lactic acid fermentation and medical care. Many of the now recognized Lactobacillus and Bifidobacterium genera have specific favorable properties and genetic traits, showing extremely strong probiotic properties. Lactobacillus can not only improve the nutritional value of food, improve the flavor of food, prolong the storage time, but also improve the functional properties of food. It can improve the digestibility and biological value of food and promote digestion a...

Examples

Embodiment 1

[0047] Example 1: Isolation, purification and preliminary identification of Lactobacillus fermentum Lee

[0048] This embodiment is carried out according to the following steps:

[0049] (1) Isolation and purification of Lactobacillus fermentum Lee

[0050] Aseptically draw 500 μL of the sample and add it to 5 mL of sterilized saline to make a uniform dilution of 1:10, and continue to make a certain proportion of dilution. Select the appropriate gradient diluent, draw 100 μL each into the MRS solid plate medium with a sterile pipette tip, and incubate at 30°C for 48-72h, observe and record the colony morphology ( figure 1 ). Use an inoculation loop (or a sterilized toothpick) to pick different colonies from the surface and inside of the plate and inoculate them in MRS liquid medium, culture them on a 300r shaker at 30°C for 24-48 hours; Ran's staining microscope ( figure 2 ), determined to be the continued activation of G+ bacteria until pure colonies were obtained (micro...

Embodiment 2

[0055] Embodiment 2: In vitro screening of Lactobacillus fermentum Lee

[0056] (1) Screening of probiotics resistant to pH 3.0 artificial gastric juice

[0057] Preparation of artificial gastric juice: 0.2% NaCl, 0.35% pepsin, adjust the pH value to 3.0 with 1M HCl, filter and sterilize with a vacuum pump in an aseptic operating table for later use.

[0058] Determination of the tolerance of probiotics to artificial gastric juice: take 5mL of the activated strain culture solution, pour it into a sterilized 10mL centrifuge tube in a sterile operating table, collect the bacteria by centrifugation at 3000r / min for 10min, add 5mL of sterilized Mix normal saline to make bacterial suspension, mix 1mL bacterial suspension with 9mL pH3.0 artificial gastric juice, shake well, place in a constant temperature shaker (37°C, 300r), and take samples at 0h and 3h respectively, Incubate at 37°C for 48h with MRS agar medium. Measure the number of viable bacteria with the plate counting meth...

Embodiment 3

[0070] Example 3: Activated carbon induced constipation experiment

[0071]Feeding healthy Kunming mice, 6 weeks old, weighing 23±2g, 60 females. After feeding the basic diet for 5 days, the mice were randomly divided into 5 groups according to body weight, namely: normal group, constipation control group, 5% RS3 group, 10% RS3 group and 15% RS3 group, with 12 mice in each group. Raise them in stainless steel cages, keep room temperature at 24±2°C, relative humidity at 50±10%, and light-dark rotation for 12 hours (lighting at 8:00-20:00). During the whole experimental period, the normal group and the constipation control group were fed with the basal diet, and the mice in the experimental group were fed with 1.0×10 9 CFU / kg gavage probiotics. Two weeks later, at 9:00 in the morning, the mice in the other five groups except the normal group were gavaged with 10% activated carbon ice water at 2°C, 0.2 mL / mouse, once a day for 3 consecutive days. Until the last day of the expe...

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