CN115353988A - Lactobacillus paracasei LC-37 with digestion promoting effect and application thereof - Google Patents
Lactobacillus paracasei LC-37 with digestion promoting effect and application thereof Download PDFInfo
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- CN115353988A CN115353988A CN202210574483.0A CN202210574483A CN115353988A CN 115353988 A CN115353988 A CN 115353988A CN 202210574483 A CN202210574483 A CN 202210574483A CN 115353988 A CN115353988 A CN 115353988A
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- lactobacillus paracasei
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- lactobacillus
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- acid
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- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L2/00—Non-alcoholic beverages; Dry compositions or concentrates therefor; Their preparation
- A23L2/02—Non-alcoholic beverages; Dry compositions or concentrates therefor; Their preparation containing fruit or vegetable juices
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- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
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Abstract
The invention belongs to the field of microbial fermentation, and particularly relates to Lactobacillus paracasei (Lactobacillus paracasei) LC-37 with a digestion promoting effect and application thereof, wherein the provided Lactobacillus paracasei (Lactobacillus paracasei) LC-37 is preserved in the China general microbiological culture Collection center (CGMCC), and the preservation number is CGMCC NO.14055. The lactobacillus paracasei LC-37 is separated from intestinal contents of healthy and long-life old people in Guangxi Bama, and the lactobacillus paracasei LC-37 has the function of promoting digestion, particularly improves dyspepsia symptoms such as abdominal pain, eructation, pantothenic acid, abdominal distension, inappetence, diarrhea or constipation and the like of dyspepsia people, and can increase the content of short-chain fatty acids in intestinal tracts.
Description
Technical Field
The invention belongs to the field of microbial fermentation, and particularly relates to Lactobacillus paracasei (Lactobacillus paracasei) LC-37 with a digestion promoting effect and application thereof.
Background
Lactobacillus paracasei (Lactobacillus paracasei) is one of the groups in Lactobacillus, is an important probiotic and is often applied to the production of dairy products, health food, beverages, ice cream and the like. Numerous studies have shown that lactobacillus paracasei has the functions of regulating intestinal flora, enhancing immunity and the like. For example, in chinese patent document CN110892989A, a lactobacillus paracasei ET-22 with constipation relieving function is disclosed, wherein a lactobacillus paracasei ET-22 with constipation relieving function is introduced, and the main functions of the strain are to improve the intestinal tract propulsion rate and shorten the intestinal tract running time, thereby playing a role in relieving constipation. Further, as disclosed in chinese patent document CN110882280A, a new application of lactobacillus paracasei K56 is introduced, wherein the efficacy of lactobacillus paracasei CGMCC15139 in significantly alleviating and/or preventing constipation is introduced, which can improve the intestinal tract propulsion rate and shorten the defecation time.
With the development of society, the rhythm of life is accelerated continuously, the working pressure is increased, and the dyspepsia problem caused by the problems that the eating on time, the fast-food eating and the like cannot be guaranteed is gradually highlighted. Eating a food with a digestive effect is an important and convenient means to improve the problem of dyspepsia. However, the digestion promoting effect of lactobacillus paracasei is currently studied less, and effective evaluation of digestion promoting-related functions is lacking.
Disclosure of Invention
Therefore, the technical problem to be solved by the invention is to provide Lactobacillus paracasei (Lactobacillus paracasei) LC-37 with the function of promoting digestion and application thereof.
The invention provides a Lactobacillus paracasei (Lactobacillus paracasei) strain LC-37, which is preserved in China general microbiological culture Collection center (CGMCC for short), and the preservation address is as follows: west road No.1, north west of the township, beijing, ministry of sciences, china, institute of microbiology, zip code: 100101, its preservation number is CGMCC NO.14055.
The invention provides a microbial preparation, which takes the lactobacillus paracasei LC-37, the inactivated substance of the lactobacillus paracasei LC-37 or the processed substance of the lactobacillus paracasei LC-37 as an active component;
the processed product of the lactobacillus paracasei LC-37 is at least one of a culture, a disrupted product and an extract of the culture of the lactobacillus paracasei LC-37.
The microbial preparation further comprises at least one excipient suitable for microbial preparation; the microbial preparation is powder, pills, capsules, granules, tablets, liquid preparations or gels.
The invention provides a preparation method of lactobacillus paracasei LC-37 powder, which comprises the steps of centrifuging fermentation liquor of lactobacillus paracasei LC-37, collecting thalli, adding a freeze-drying protective agent into the obtained thalli, and carrying out vacuum freeze-drying to obtain freeze-dried powder.
In the preparation method, the fermentation steps of the lactobacillus paracasei LC-37 are as follows: inoculating the seed liquid of the lactobacillus paracasei LC-37 into a fermentation culture medium in an inoculation amount of 2-5% (v/v), culturing at 33-40 ℃, keeping the pH of the culture medium at 5.0-6.5 with a stirring rotation speed of 100-200 rpm, and fermenting for 14-20 hours.
In the preparation method, the centrifugation conditions are as follows: rotating at 8000-10000 rpm, centrifuging for 1-2 hours.
In the preparation method, the freeze-drying protective agent comprises the following raw materials in parts by weight: 1 to 3 portions of trehalose, 1 to 3 portions of skim milk powder, 2 to 4 portions of maltodextrin and 0.1 to 0.5 portion of sodium ascorbate.
The mass ratio of the addition amount of the freeze-drying protective agent to the thalli obtained by centrifugation is 0.2-0.9, the vacuum freeze-drying condition is that the temperature is increased from minus 50 ℃ to 30 ℃ within 40-60 hours, and the vacuum degree is 3-50 Pa.
The preparation method also comprises the following steps of: inoculating the frozen strain of the lactobacillus paracasei LC-37 into 10-50 mL of culture medium, and culturing for 12-24 hours at 33-40 ℃.
In the preparation method, the auxiliary materials are added into the freeze-dried powder, and the auxiliary materials can be maltodextrin, whey powder, fructo-oligosaccharide or resistant starch and the like.
The invention provides the lactobacillus paracasei LC-37 bacterial powder prepared by the preparation method of the lactobacillus paracasei LC-37 bacterial powder.
The invention provides the application of the lactobacillus paracasei LC-37, the microbial preparation, the bacterial powder prepared by the method or the bacterial powder in preparing medicines or foods with digestion promoting effect.
In the present invention, the drug can be prepared into different forms according to different administration routes, for example, powder, tablet, granule, capsule, solution, emulsion, suspension, etc.; the medicament may also include a pharmaceutically acceptable adjuvant. The food product includes any type of food product such as solid beverages, soy products, juices, dairy products, ice cream, candies, cookies, etc.; the food may also contain conventional additives, nutrition enhancer, and adjuvants, such as essence, flavor, stabilizer, thickener, antiseptic, minerals, vitamins, maltodextrin, etc.
The invention provides a preparation method of a lactobacillus beverage containing lactobacillus paracasei LC-37, which comprises the steps of using the lactobacillus paracasei LC-37, a microbial preparation, and bacterial powder prepared by the method or the bacterial powder.
In the preparation method, lactobacillus paracasei LC-37 powder is inoculated in the fermentation step of the beverage base material, and the inoculation amount is 10 4 -10 7 CFU/mL, fermentation temperature of 33-40 deg.C, fermentation timeIs 72-100 hours.
In the preparation method, the viable count range of the lactobacillus paracasei LC-37 in the prepared lactobacillus beverage is more than or equal to 5 multiplied by 10 7 CFU/mL。
In the preparation method, the beverage base material comprises the following raw materials in parts by weight: 110-140 parts of skim milk powder, 15-25 parts of glucose and 35-50 parts of white granulated sugar.
In the preparation method, the beverage base material is subjected to sterilization and browning before fermentation, wherein the sterilization and browning conditions are 90-97 ℃ and 2-5 hours.
In the above preparation method, the auxiliary material is added to the fermented milk obtained in the fermentation step. The auxiliary materials can be sugar liquor and/or essence and the like.
The invention provides a lactobacillus beverage containing lactobacillus paracasei LC-37 prepared by the method.
Compared with the prior art, the invention has the following advantages:
1. the Lactobacillus paracasei (Lactobacillus paracasei) LC-37 provided by the invention has been preserved in the China general microbiological culture Collection center of China Committee for culture Collection of microorganisms, and the preservation number is CGMCC NO.14055. The lactobacillus paracasei LC-37 is separated from intestinal contents of healthy and long-life old people in Guangxi Bama, and the lactobacillus paracasei LC-37 has the effect of improving dyspepsia, particularly improving dyspepsia symptoms such as abdominal pain, eructation, pantothenic acid, abdominal distension, inappetence, diarrhea or constipation and the like of dyspepsia people, and simultaneously increasing the content of short-chain fatty acids in intestinal tracts.
2. The lactobacillus beverage containing the lactobacillus paracasei LC-37 provided by the invention utilizes the lactobacillus paracasei strain LC-37 as a fermentation strain, and the prepared lactobacillus beverage has the function of promoting digestion, can improve the indigestion symptoms such as abdominal pain, eructation, pantothenic acid, abdominal distension, inappetence, diarrhea or constipation and the like, and can increase the content of short-chain fatty acid in intestinal tracts.
3. The lactobacillus paracasei LC-37 powder provided by the invention has the function of promoting digestion, can improve dyspepsia symptoms such as abdominal pain, eructation, pantothenic acid, abdominal distension, inappetence, diarrhea or constipation and the like, and can increase the content of short-chain fatty acid in intestinal tracts.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the embodiments or the prior art descriptions will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.
FIG. 1 shows the score change of abdominal pain symptoms in subjects of the experimental examples of the present invention;
FIG. 2 is a graph showing the score change of belching symptoms of a subject in an experimental example of the present invention;
FIG. 3 is a change in the score for pantothenic acid symptoms in subjects in the experimental examples of the present invention;
FIG. 4 shows the change in the score of the abdominal distension symptom of the subject in the experimental example of the present invention;
FIG. 5 shows the change in the scores of appetite symptoms in subjects in the experimental examples of the present invention;
FIG. 6 shows the change of scores of diarrhea and diarrhea symptoms in subjects in the experimental examples of the present invention;
FIG. 7 is a graph of the change in the total clinical symptom score of dyspepsia for a subject in an experimental example of the present invention;
FIG. 8 is a graph showing the change in acetic acid content before and after drinking by the subjects in the experimental examples of the present invention;
FIG. 9 is a graph showing the change in propionic acid content before and after drinking by the subjects in the experimental examples of the present invention;
FIG. 10 is a graph showing the change in butyric acid content before and after drinking by the subjects in the experimental examples of the present invention;
FIG. 11 is a graph showing the change in total acid content before and after drinking by the test subjects in the experimental examples of the present invention;
FIG. 12 is a graph of a standard curve in an experimental example of the present invention.
Detailed Description
The following examples are provided to further understand the present invention, not to limit the scope of the present invention, but to provide the best mode, not to limit the content and the protection scope of the present invention, and any product similar or similar to the present invention, which is obtained by combining the present invention with other prior art features, falls within the protection scope of the present invention.
The examples do not show the specific experimental steps or conditions, and can be performed according to the conventional experimental steps described in the literature in the field. The reagents or instruments used are not indicated by manufacturers, and are all conventional reagent products which can be obtained commercially.
The embodiments of the present invention are illustrated below by specific examples, and unless otherwise indicated, the experimental methods disclosed in the present invention are performed by using conventional techniques in the art, and reagents and raw materials used in the examples are commercially available.
EXAMPLE 1 acquisition of Lactobacillus paracasei strain LC-37
1. Isolation of Lactobacillus paracasei strain LC-37
Collecting intestinal contents, namely feces, of the Guangxi Bama healthy elderly with long life, performing gradient dilution on intestinal feces samples of the Guangxi Bama healthy elderly with long life by using a diluent, separating lactobacillus by using an MRS (lactic acid bacteria culture medium), picking bacterial colonies with different forms, selecting gram-positive bacillus by microscopic examination, and continuously purifying strains by using a pour plate method until pure cultures of the strains are obtained by separation. And (3) obtaining the lactobacillus strain through the separation process.
The preparation method of the diluent used in the dilution step comprises the following steps: 4.5g of sodium dihydrogen phosphate, 6.0g of disodium hydrogen phosphate, 0.5g of L-cysteine, 0.5g of agar, 0.5mL of Tween 80 and 1000 mL of distilled water are heated and dissolved, the pH value is adjusted to 7.4-7.6, and the mixture is sterilized at 121 ℃ for 15min for later use.
The pouring flat plate method comprises the following steps: selecting bacterial colonies with different forms, inoculating the bacterial colonies into a liquid MRS culture medium, culturing for 12-16h, continuously carrying out 7 times of 10-time gradient dilution on 1mL of bacterial liquid by using a diluent, adding 1mL of diluted bacterial liquid into a sterile plate, pouring a proper amount of MRS solid culture medium, inverting the culture medium after the culture medium is solidified, culturing for 48h in an incubator at 37 ℃, and selecting a single bacterial colony to obtain a pure culture of the bacterial strain.
2. Identification and preservation
The lactobacillus obtained by separation is identified as lactobacillus paracasei by 16S rRNA, named as lactobacillus paracasei LC-37, and is preserved in the China general microbiological culture Collection center, with the preservation address: west road No.1, north west of the township, beijing, ministry of sciences, china, institute of microbiology, zip code: 100101, preservation number is CGMCC No.14055.
EXAMPLE 2 Lactobacillus paracasei LC-37 powder
The embodiment provides a preparation method of lactobacillus paracasei LC-37 powder, which specifically comprises the following steps:
1. expanded culture of strains
Inoculating the frozen strain of lactobacillus paracasei LC-37 into 10mL of MRS culture medium, and culturing for 12 hours at 37 ℃ to obtain a seed culture solution suitable for inoculation, wherein the form of the strain examined by a microscope is consistent with the unique form of the strain, and no visible mixed bacteria exist.
2. Fermentation of
The seed culture obtained above was inoculated into a fermentation medium (MRS medium in this example) at 33 ℃ in an amount of 2% to 5% (v/v) (2% v/v was selected in this example), and the mixture was fermented for 20 hours at a stirring speed of 200rpm with the pH of the medium being maintained at pH 5.0. And (3) after the fermentation is finished, centrifuging the culture solution by using a disc-type centrifuge at the rotating speed of 8000rpm for 2 hours, and collecting the thalli.
3. Freeze-drying
Adding a freeze-drying protective agent into the collected thalli, wherein the freeze-drying protective agent comprises the following raw materials in parts by weight: 1kg of trehalose, 3kg of skim milk powder, 2kg of maltodextrin and 0.5kg of sodium ascorbate. And (3) adding the freeze-drying protective agent, wherein the mass ratio of the added freeze-drying protective agent to the thalli obtained by centrifugation is 0.9, placing the thalli into a freeze-drying tray, and carrying out vacuum freeze-drying under the conditions that the temperature is increased from-50 ℃ to 30 ℃ within 40 hours and the vacuum degree is 3Pa so that the water activity of the final bacterial powder is lower than 0.25, and finishing the vacuum freeze-drying. Followed by crushing and packaging to form a lyophilized powder of Lactobacillus paracasei LC-37, based on Lactobacillus paracaseiThe viable count of the LC-37 freeze-dried powder is mixed with maltodextrin in proportion, so that the viable count of the finally obtained lactobacillus paracasei LC-37 powder is 5 multiplied by 10 9 CFU/g。
EXAMPLE 3 Lactobacillus paracasei LC-37 powder
The embodiment provides a preparation method of lactobacillus paracasei LC-37 powder, which specifically comprises the following steps:
1. expanded culture of strains
Inoculating the frozen strain of lactobacillus paracasei LC-37 into 10mL of MRS culture medium, and culturing for 12 hours at 37 ℃ to obtain a seed culture solution suitable for inoculation, wherein the form of the strain examined by a microscope is consistent with the unique form of the strain, and no visible mixed bacteria exist.
2. Fermentation of
The seed culture obtained above was inoculated into a fermentation medium, MRS medium in this example, in an amount of 2 to 5% (v/v) (selected to 5% v/v in this example), cultured at 40 ℃ with a stirring rotation speed of 100rpm and a medium pH maintained at pH6.0, and fermented for 14 hours. And (3) after the fermentation is finished, centrifuging the culture solution by using a disc-type centrifuge, wherein the rotating speed of the centrifuge is 10000rpm, centrifuging for 1 hour, and collecting thalli.
3. Freeze-drying
Adding a cryoprotectant into the collected thallus, wherein the cryoprotectant comprises the following raw materials in parts by weight in the embodiment: 3kg of trehalose, 1kg of skim milk powder, 4kg of maltodextrin and 0.1kg of sodium ascorbate, wherein the ratio of the addition amount of the freeze-drying protective agent to the thalli obtained by centrifugation is 0.5, the mixture is placed into a freeze-drying tray, the temperature of the vacuum freeze-dried strip is increased from-50 ℃ to 30 ℃ within 60 hours, the vacuum degree is 50Pa, the water activity of the final bacterial powder is lower than 0.25, and the vacuum freeze-drying is finished. Then crushing and packaging to form lactobacillus paracasei LC-37 freeze-dried powder, and mixing maltodextrin in proportion according to the viable count of the lactobacillus paracasei LC-37 freeze-dried powder to ensure that the viable count of the finally obtained lactobacillus paracasei LC-37 powder is 5 multiplied by 10 10 CFU/g。
EXAMPLE 4 Lactobacillus paracasei LC-37 powder
The embodiment provides a preparation method of lactobacillus paracasei LC-37 powder, which specifically comprises the following steps:
1. expanded culture of strains
Inoculating the frozen strain of lactobacillus paracasei LC-37 into 10mL of MRS culture medium, and culturing for 12 hours at 37 ℃ to obtain a seed culture solution suitable for inoculation, wherein the form of the strain examined by a microscope is consistent with the unique form of the strain, and no visible mixed bacteria exist.
2. Fermentation of
The seed culture obtained above was inoculated into a fermentation medium (MRS medium in this example) at an inoculum size of 2% to 5% (v/v) (3% v/v was selected in this example), and the mixture was cultured at 37 ℃ with a stirring rotation speed of 150rpm and a pH of the medium maintained at pH 5.8, and fermented for 17 hours. After the fermentation is finished, the culture solution is centrifuged by using a disk centrifuge, the rotating speed of the centrifuge is 9000rpm, the culture solution is centrifuged for 1.5 hours, and the thalli are collected.
3. Freeze-drying
Adding a cryoprotectant into the collected thallus, wherein the cryoprotectant comprises the following raw materials in parts by weight in the embodiment: 2kg of trehalose, 2kg of skim milk powder, 3kg of maltodextrin and 0.3kg of sodium ascorbate. The ratio of the addition amount of the freeze-drying protective agent to the thalli obtained by centrifugation is 0.2, the thalli are placed in a freeze-drying tray, the vacuum freeze-drying condition is that the temperature is increased from-50 ℃ to 30 ℃ within 50 hours, the vacuum degree is 25Pa, the water activity of the final bacterial powder is lower than 0.25, and the vacuum freeze-drying is finished. Then crushing and packaging to form lactobacillus paracasei LC-37 freeze-dried powder, and mixing maltodextrin in proportion according to the viable count of the lactobacillus paracasei LC-37 freeze-dried powder to ensure that the viable count in the finally obtained lactobacillus paracasei LC-37 powder is 5 multiplied by 10 11 CFU/g。
Example 5 Lactobacillus paracasei LC-37-containing lactic acid bacteria drink
The embodiment provides a preparation method of a lactobacillus beverage containing lactobacillus paracasei LC-37, which comprises the following steps:
taking skim milk powder (110 kg) as a raw material, adding glucose (25 kg) and white granulated sugar (35 kg), sterilizing and browning, namely sterilizing and browning at 90 ℃ for 5 hours, then cooling to 33 ℃, and inoculating the lactobacillus paracasei prepared in the example 2LC-37 strain powder with the inoculation amount of 10 7 CFU/mL (even if the final concentration of Lactobacillus paracasei LC-37 in the feed solution is 10 7 CFU/mL), the fermentation temperature is 33 ℃, the fermentation is finished after 72 hours of long-time fermentation, and then the obtained fermented milk is taken as the main raw material, and a proper amount of auxiliary materials such as sugar solution, essence and the like are added, so that the content of viable bacteria in the lactobacillus beverage containing lactobacillus paracasei LC-37 is finally prepared: 5X 10 7 CFU/ml。
Example 6 Lactobacillus paracasei LC-37-containing lactic acid bacteria drink
The embodiment provides a preparation method of a lactobacillus beverage containing lactobacillus paracasei LC-37, which comprises the following steps:
taking skim milk powder (140 kg) as a raw material, adding glucose (15 kg) and white granulated sugar (50 kg), sterilizing and browning, namely sterilizing and browning at 97 ℃ for 2 hours, then cooling to 40 ℃, and inoculating the lactobacillus paracasei LC-37 strain powder prepared in the example 3, wherein the inoculation amount is 10 4 CFU/mL (even if the final concentration of Lactobacillus paracasei LC-37 in the feed solution is 10 4 CFU/mL), the fermentation temperature is 40 ℃, the fermentation is finished after long-time fermentation for 100 hours, and then the obtained fermented milk is taken as a main raw material, and a proper amount of auxiliary materials such as sugar solution, essence and the like are added, so that the content of viable bacteria in the lactobacillus beverage containing lactobacillus paracasei LC-37 is finally prepared: 1X 10 8 CFU/ml。
Example 7 Lactobacillus paracasei LC-37-containing lactic acid bacteria beverage
The embodiment provides a preparation method of a lactobacillus beverage containing lactobacillus paracasei LC-37, which comprises the following steps:
adding glucose (20 kg) and white granulated sugar (42 kg) into skim milk powder (125 kg) as a raw material, sterilizing and browning at 90-97 deg.C for 2-5 hr, cooling to 33-40 deg.C, inoculating the Lactobacillus paracasei LC-37 powder prepared in example 4 with an inoculation amount of 10 5 CFU/mL (to give a final concentration of 10 for Lactobacillus paracasei LC-37 in the feed solution) 5 CFU/mL), the fermentation temperature is 37 ℃, the fermentation is finished after the long-term fermentation for 86 hours, and then the obtained fermented milk is used as the main raw material, and a proper amount of auxiliary materials such as sugar solution, essence and the like are added, so that the fermented milk containing the side product is finally preparedThe viable count content of the lactobacillus beverage of the lactobacillus casei LC-37 is as follows: 5X 10 8 CFU/ml。
Examples of the experiments
1. Test subject
1.1 Inclusion criteria
Selecting voluntary subjects with functional dyspepsia, long-term gastrointestinal discomfort, complaints of inappetence, early satiety, excessive qi, gastrointestinal fullness, vomiting, chronic diarrhea or constipation of unknown reason and the like.
1.2 exclusionary criteria
(1) Acute diarrhea.
(2) Dyspepsia due to severe organic lesions.
(3) The weak constitution can not receive the testee.
(4) Patients with serious systemic diseases such as cardiovascular diseases, liver diseases, kidney diseases, hemopoietic diseases, etc.
(5) If the test sample is not taken as required, the result of drinking can not be judged.
(6) Lactose intolerance or milk allergy.
1.3 design of the experiment
The total 120 people of the group-entering population are divided into 4 groups of 30 people after screening of the experiment. Wherein 55 men and 65 women; age 20-60 years, mean age 47.8 years, mean BMI index 24.4.BMI index: body Mass Index (Body Mass Index, BMI for short).
1.4 dosage and time
Storing the sample in refrigerator at 2-10 deg.C, and taking after lunch or dinner every day, wherein group A and group C take one bag of Lactobacillus paracasei LC-37 powder every day, and group B and group D take one bottle of lactobacillus beverage containing Lactobacillus paracasei LC-37 every day. The administration is continued for 28 days. The original diet habit and normal diet were not changed during the test period.
Group A: lactobacillus paracasei LC-37 powder 1 (viable count content prepared in example 2: 5X 10) 9 CFU/g, daily intake: 2g) (ii) a
Group B: lactobacillus paracasei LC-37-containing lactic acid bacterium beverage 1 (viable cell count content: 5X 10 prepared in example 5) 7 CFU/ml, daily intakeQuantity: 200 ml)
Group C: lactobacillus paracasei LC-37 powder 2 (viable count content: 5X 10 prepared in example 3) 10 CFU/g, daily intake: 2g) (ii) a
Group D: lactobacillus paracasei LC-37-containing lactic acid bacterium beverage 2 (viable cell count content: 5X 10 prepared in example 7) 8 CFU/ml, daily intake: 200 ml)
1.5 Experimental procedure
The emptying period lasted for 7 days. During the period, the volunteers could not drink any fermented product (including yogurt, cheese, active lactic acid bacteria beverage, etc.), but could drink milk. Day 7 is the time of first stool sample and blood collection, and clinical symptom scoring and filling out an intestinal health survey. The drinking period lasts 28 days, and feces are collected for the second time on day 14, scored for clinical symptoms, and filled in a health questionnaire. Stool samples and blood were collected on day 28, scored for clinical symptoms, and filled in an intestinal health questionnaire.
1.6 Observation index
1.6.1 safety index
And (3) detecting blood routine and blood biochemical indexes of blood samples collected before and after drinking. The conventional blood biochemical indexes such as white blood cell count, hemoglobin, mean red blood cell volume, mean red blood cell hemoglobin concentration, platelet count, thrombocyte deposition, lymphocyte absolute value, neutrophil absolute value, basophil absolute value, monocyte percentage, eosinophil percentage, erythrocyte count, hematocrit, mean red blood cell hemoglobin amount, red blood cell volume distribution width, monocyte absolute value, eosinophil absolute value, lymphocyte percentage, neutrophil percentage, basophil percentage, urea, uric acid, glutamic-oxaloacetic transaminase, total protein, white-to-globular ratio, triglyceride, creatinine, glutamic-pyruvic transaminase, albumin, globulin, total cholesterol, blood sugar and the like in each intervention group blood sample are all within normal value ranges, and the lactobacillus paracasei LC-37 and the lactobacillus beverage containing the lactobacillus paracasei LC-37 have no adverse effect on human bodies. Specifically, see tables 1-4 below:
TABLE 1 group A conventional and biochemical blood treatment of each group before drinking
TABLE 2B groups of conventional and biochemical blood treatment
TABLE 3C groups blood routine and blood biochemistry of each group before drinking
TABLE 4D groups of conventional and biochemical blood treatment
1.6.2 Observation of efficacy
Clinical symptoms of dyspepsia before and after the test of the subject were accurately recorded, quantitative scores were given according to table 5, and changes in scores of symptoms of dyspepsia before and after the test were compared.
TABLE 5 clinical symptom score
And adding the scores of the symptoms of abdominal pain, eructation, pantothenic acid, abdominal distension, appetite, diarrhea or constipation to obtain the dyspepsia clinical symptom score. Statistical analysis was performed on the clinical symptom scores of the population, and the results are shown in fig. 1-7. Fig. 1-6 show the score change of each dyspepsia symptom, and fig. 7 shows the change of the total clinical symptom score of dyspepsia. As can be seen from fig. 1 to fig. 7, at day 0, the scores of the dyspepsia symptoms such as abdominal pain, belching and acid regurgitation are similar to the total score of the dyspepsia symptoms, and no significant difference exists, which indicates that the groups have comparability. At 14 days of intervention, a significant reduction in dyspepsia score was observed for each group. In addition, the effect of the high dose of lactobacillus paracasei is better than that of the low dose of lactobacillus paracasei, and the lactobacillus beverage containing the same dose of lactobacillus paracasei is better than that of the lactobacillus paracasei powder. At day 28 of the intervention period, the dyspepsia symptoms score for each group was further reduced, with the loss of appetite symptoms for all groups and the loss of abdominal pain, belching, and acid regurgitation symptoms for group D. The total symptom score of dyspepsia of each group is reduced from the initial 4.18-4.71 to 0.17-0.71, and extremely remarkable improvement appears.
1.6.3 detection of short-chain fatty acid content in feces
Establishing a standard curve of the short-chain fatty acid: a solution of acetic acid (5 mmol/L), propionic acid (5 mmol/L), butyric acid (5 mmol/L) and heptanoic acid (5 mmol/L) in ether was prepared in a volumetric flask, and diluted in two-fold to obtain standard solutions containing acetic acid, propionic acid, butyric acid and heptanoic acid (2.5, 1.25,0.625,0.312, 0.156mmol/L), respectively. And respectively transferring 1 mu L of standard solution to perform gas chromatography analysis, repeatedly measuring each concentration for 3 times to obtain peak areas and corresponding concentrations of each concentration, thereby preparing a standard curve and solving a regression equation and a correlation coefficient.
Extracting short-chain fatty acids in the excrement: an appropriate amount of glass beads (2 to 3 mm) was put into a 1.5mL screw-port collection tube, and 1mL of an ether solution containing heptanoic acid (concentration: 1 mmol/L) and 50. Mu.L of a hydrochloric acid solution (concentration: 1 mmol/L) were added and weighed. Subjects fecal samples (about 0.1-0.2 g) were carefully added to the collection tube with forceps, weighed again, and the weight of the feces added was recorded. Homogenizing for 1min with homogenizer, and centrifuging for 2min at 10000 r/min. Transfer supernatant into a vial with an internal cannula. Because of the strong volatility of the ether, the above operations are all carried out rapidly in an ice bath or at low temperature.
Chromatographic conditions for detecting the concentration of short-chain fatty acid in the fecal sample are as follows: the detector is an FID hydrogen flame ion detector; a chromatography column DB-FFAP chromatography column; temperature rising procedure: maintaining at 50 deg.C for 1min, heating to 140 deg.C at 10 deg.C/min, maintaining for 1min, heating to 240 deg.C at 30 deg.C/min, and maintaining for 2min; carrier gas (N) 2 ) The pressure is 260kPa; the nitrogen flow rate is 40mL/min, the hydrogen flow rate is 40mL/min, and the air flow rate is 400mL/min; the sample volume is 2 mu L; the detection temperature was 230 ℃. The N2000 chromatography workstation collects the signals and detects them.
Detecting according to the above gas chromatography conditions, calculating peak area of corresponding short chain fatty acid, making standard curve by external standard method (as shown in FIG. 12), correcting with internal standard heptanoic acid concentration, calculating concentration of acetic acid, propionic acid and butyric acid in the detected sample, and converting to content in fecal sample.
The results of the tests are shown in FIGS. 8-11, and the contents of acetic acid, propionic acid, butyric acid and total short chain fatty acids in the feces of the tested human bodies were not significantly changed after 14 days of eating the powder of Lactobacillus paracasei LC-37 and the lactic acid bacteria beverage containing Lactobacillus paracasei LC-37. After eating for 28 days, the acetic acid, propionic acid, butyric acid and total short chain fatty acid in each group are increased to different degrees. Shows that the edible lactobacillus paracasei LC-37 powder and the lactobacillus beverage containing the lactobacillus paracasei LC-37 can increase the content of short-chain fatty acid in the feces.
It should be understood that the above examples are only for clarity of illustration and are not intended to limit the embodiments. Other variations and modifications will be apparent to persons skilled in the art in light of the above description. And are neither required nor exhaustive of all embodiments. And obvious variations or modifications therefrom are within the scope of the invention.
Claims (11)
1. The application of Lactobacillus paracasei (Lactobacillus paracasei) LC-37 in preparing products for improving abdominal pain, eructation, pantothenic acid, abdominal distension, inappetence, diarrhea or constipation is disclosed, wherein the Lactobacillus paracasei LC-37 is preserved in the China general microbiological culture Collection center (CGMCC), and the preservation number is CGMCC NO.14055.
2. The application of Lactobacillus paracasei (Lactobacillus paracasei) LC-37 in a product for increasing the content of short-chain fatty acids in intestinal tracts is disclosed, wherein the Lactobacillus paracasei LC-37 is preserved in the China general microbiological culture Collection center (CGMCC) with the preservation number of CGMCC NO.14055.
3. Use according to claim 2, wherein the short chain fatty acids are total short chain fatty acids; or, the short chain fatty acid is at least one of acetic acid, propionic acid and butyric acid.
4. Use according to any one of claims 1 to 3, wherein the product is a food or a medicament.
5. Use according to claim 4, wherein the food products include, but are not limited to, solid beverages, soy products, fruit juices, dairy products, ice cream, candies and biscuits.
6. The use according to claim 4, wherein the food product is a food product comprising Lactobacillus paracasei LC-37 powder.
7. The use according to claim 6, wherein the food product is a lactic acid bacteria drink; the viable count of the lactobacillus paracasei LC-37 in the lactobacillus beverage is more than or equal to 5 multiplied by 10 7 CFU/mL。
8. The use according to any one of claims 4 to 7, wherein the food further comprises raw materials and auxiliary materials, and the auxiliary materials in the raw materials and auxiliary materials include but are not limited to additives and/or nutrition enhancers.
9. Use according to any one of claims 4 to 7, wherein the auxiliary materials in the food product include, but are not limited to, at least one of sugar solutions, flavors and fragrances, stabilizers, thickeners, preservatives, minerals, vitamins, maltodextrins, whey powder, fructooligosaccharides and resistant starch.
10. The use according to claim 4, wherein the medicament is in a dosage form including, but not limited to, powders, tablets, granules, capsules, solutions, emulsions or suspensions.
11. The use according to claim 10, wherein the medicament further comprises a pharmaceutically acceptable adjuvant.
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