CN110559319A - Application of lactobacillus paracasei N1115 to improvement of diabetes, corresponding milk powder and application - Google Patents
Application of lactobacillus paracasei N1115 to improvement of diabetes, corresponding milk powder and application Download PDFInfo
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- CN110559319A CN110559319A CN201910901638.5A CN201910901638A CN110559319A CN 110559319 A CN110559319 A CN 110559319A CN 201910901638 A CN201910901638 A CN 201910901638A CN 110559319 A CN110559319 A CN 110559319A
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- A—HUMAN NECESSITIES
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Abstract
The invention provides an application of lactobacillus paracasei N1115 for improving diabetes, corresponding milk powder and an application, wherein the lactobacillus paracasei N1115 is used for preventing or relieving the diabetes; the milk powder prepared by using the lactobacillus paracasei N1115 can effectively prevent or relieve diabetes, and is reasonable in formula and rich in nutrition.
Description
Technical Field
The invention belongs to the field of bioengineering, and relates to an application of a strain for improving diabetes, in particular to an application of lactobacillus paracasei N1115 for improving diabetes; the invention also provides milk powder prepared by using the lactobacillus paracasei N1115 for improving diabetes and application thereof.
Background
Diabetes is a chronic metabolic disease caused by genetic factors, environmental factors and living habits. The main reasons are a syndrome of elevated blood glucose levels as a result of the inability of the human pancreas to produce insulin normally (insulin deficiency) or the body to utilize insulin normally (insulin resistance). Hyperglycemia is caused by a defect in insulin secretion or an impaired biological action, or both. Diabetes is a long-standing hyperglycemic condition that can lead to chronic damage to, and dysfunction of, various tissues, particularly the eye, kidneys, heart, blood vessels, nerves. If the disease condition is not well controlled, complications such as heart disease, coronary heart disease, cerebrovascular disease, retinal vascular disease, renal arteriosclerosis, limb arteriosclerosis and the like can be caused in the future. According to the latest data released by the national institutes of health on day 11/12 in 2016, the following are shown: the total number of Chinese diabetics reaches 1 hundred million and will continue to increase, and China now becomes the world with the largest number of diabetics. How to prevent diabetes in advance becomes a current problem.
Research shows that probiotics has preventive, relieving and therapeutic effects on diabetes, and potential intervention mechanisms of the probiotics comprise:
1 Activity for inhibiting alpha-glucosidase
Alpha-glucosidase is a key enzyme in the intestinal lumen and brush border membrane that promotes carbohydrate hydrolysis, which degrades starch and polysaccharides into absorbable monosaccharides. Inhibiting the activity of alpha-glucosidase, reducing the absorption of glucose and lowering the postprandial blood glucose level.
2 regulating intestinal flora
Reports indicate that the intestinal flora of diabetics is unbalanced, but probiotics can colonize the intestinal tract, regulate the intestinal flora and increase the expression of adhesion proteins, and form a natural biological barrier of the intestinal tract together with intestinal mucosa. The barrier can reduce the number of harmful bacteria in the intestinal tract, increase the number of beneficial bacteria, and maintain the balance of intestinal flora, thereby reducing the content of lipopolysaccharide in endotoxin and the permeability of epithelial cells in the intestinal tract, reducing the release of inflammatory factors and oxidative stress reaction, and further reducing the damage to beta cells of pancreatic islets.
3 antioxidation
Diabetes is often accompanied by oxidative stress, i.e.: excessive production of free radicals and destruction of the antioxidant defense system. This will lead to cellular and enzymatic damage, increased lipid peroxidation, and development of insulin resistance. It also destroys islet beta cells leading to diabetes. The antioxidant action of the strain has been proved by research, and the antioxidant mechanism can be divided into active oxygen scavenging, metal ion chelating, activity of inhibiting pro-oxidase, reduction activity and oxidation of ascorbic acid inhibiting.
4 regulating immunity and reducing inflammation
Diabetes is often associated with long-term low-grade inflammation that can lead to diabetes by disrupting endothelial cell structure and function, causing a barrier to insulin transport in tissue cells, and leading to islet beta cell structure and dysfunction. The probiotics can regulate the immune function of the organism by adhering epithelial cells, and reduce inflammation by regulating the secretion of anti-inflammatory factors, thereby preventing and controlling the occurrence of diabetes.
However, no product for preventing or relieving diabetes by using probiotics exists in the market at present, so that the market is blank at present.
Disclosure of Invention
The invention aims to solve the technical problem of providing an application of lactobacillus paracasei N1115 in improving diabetes, and the lactobacillus paracasei N1115 is used for preventing or relieving diabetes. The lactobacillus paracasei N1115 is separated and screened from traditional fermented milk products in inner Mongolia of China, is disclosed in patent CN201110357058.8, has been preserved in the strain collection center of the institute of microbiology of Chinese academy of sciences in 2011 for 3 months, and has the preservation number of CGMCC 4691.
The invention also provides milk powder for improving diabetes, which is prepared by adding lactobacillus paracasei N1115 into the raw materials, and the milk powder can effectively prevent or relieve diabetes.
The invention also provides an application of the milk powder for improving diabetes, which is used for preventing or relieving diabetes.
In order to solve the technical problems, the technical scheme adopted by the invention is as follows: the application of lactobacillus paracasei N1115 in improving diabetes, wherein the lactobacillus paracasei N1115 is used for preventing or relieving diabetes.
The invention also provides milk powder for improving diabetes, which is prepared from the following raw materials in parts by weight: 40-45 parts of raw milk powder, 28-33 parts of desalted whey powder, 5-8 parts of white granulated sugar, 4-4.5 parts of anhydrous cream, 3-4 parts of soybean oil, 3-4 parts of lactose, 2-4 parts of concentrated whey protein powder, 1.5-2 parts of galacto-oligosaccharide, 1-1.5 parts of fructo-oligosaccharide, 0.5-1 part of phospholipid, 0.1-0.2 part of DHA algal oil powder, 0.1-0.3 part of arachidonic acid powder, 0.05-0.2 part of choline chloride, 0.1-0.8 part of calcium carbonate, 0.02-0.06 part of nucleotide, 0.1-0.5 part of compound vitamin, 0.1-0.3 part of compound mineral substance and 0.1-1 part of probiotic powder capable of improving diabetes; wherein the content of Lactobacillus paracasei N1115 in the probiotic powder capable of improving diabetes is 1 × 108CFU/g-9×1010CFU/g。
The invention also provides an application of the milk powder for improving diabetes, which is used for preventing or relieving diabetes.
Compared with the prior art, the invention has the following advantages:
1. The lactobacillus paracasei N1115 can prevent or relieve diabetes.
2. The lactobacillus paracasei N1115 can regulate immunity, reduce the damage of oxidation to insulin secreting cells, increase intestinal barrier function and reduce the occurrence of long-term low-grade inflammation.
3. The lactobacillus paracasei N1115 can improve the symptoms of thirst and polydipsia by improving the diabetic milk powder.
Description of the drawings:
FIG. 1 is a graph showing the variation of the N1115 TEER value in the evaluation of intestinal barrier function in example 1 of the present invention.
FIG. 2 is a graph showing the change in LGG TEER values in the evaluation of intestinal barrier function in example 1 of the present invention.
FIG. 3 shows the hydroxyl radical scavenging ability of probiotic bacteria in the evaluation of the anti-oxidative ability according to example 1 of the present invention.
FIG. 4 is a graph showing the ability of probiotic bacteria to scavenge superoxide anions in the evaluation of the anti-oxidative ability in example 1 of the present invention.
Fig. 5 is a graph of the DPPH scavenging ability of probiotic bacteria in the evaluation of the anti-oxidative ability according to example 1 of the present invention.
FIG. 6 shows the immunomodulatory capacity of active N1115 in the assessment of immunomodulatory capacity in example 1 of the present invention.
FIG. 7 is a graph showing the immunoregulatory ability of heat-inactivated N1115 in the evaluation of the immunoregulatory ability in example 1 of the present invention.
Detailed Description
The present invention is described in further detail below with reference to specific examples, which are intended to be illustrative only and not limiting.
Example 1: application of lactobacillus paracasei N1115 to improvement of diabetes
1. Evaluation of intestinal barrier function by Lactobacillus paracasei N1115
Based on the aim that some probiotics in the intestinal tract can maintain the activity of intestinal tract cells by regulating the permeability of the intestinal tract, the aim of maintaining the tight connection structure between the intestinal tract cells is fulfilled, and the intestinal epithelial cell research model established in vitro adopts a method of forming a single-layer membrane barrier in a Transwell cell chamber by Caco-2 cells, so that the intestinal epithelial cell research model is a mature in vitro intestinal barrier model.
Selecting a 6-hole plate with a 0.4 mu m Transwell filter membrane, and culturing the cultured cells wellCaco-2 was placed in the upper chamber to give a cell volume of about 5X 105One/well, at 37 ℃ and 5% CO2Culturing in an incubator, gradually contacting and differentiating Caco-2 cells with each other along with the extension of the culture time, finally forming a tight connection structure between the cells, simultaneously increasing the measured transmembrane resistance value, completing model establishment, replacing culture solution in a Transwell cell chamber, and adding probiotic suspension (the concentration is about 1 × 10) into an upper chamber8CFU/well), 5% CO at 37 deg.C2And co-incubating for 6 h in the incubator, measuring the transmembrane resistance values in 2 h, 4 h and 6 h respectively, and calculating the relative transmembrane resistance values according to a formula.
The intercellular tight junction structure is the basis of the intestinal mechanical barrier, and in the Transwell Caco-2 cell model, if the tight junction structure is broken, the resistance to ion flow is reduced, and the transmembrane resistance value is reduced.
The experimental results are shown in fig. 1-2, which indicate that the probiotics have a certain maintaining effect on intercellular tight junction structures, wherein lactobacillus paracasei N1115 has a better effect of maintaining intestinal barrier functions by maintaining and protecting the tight junction structures, thereby alleviating damage to islet beta cells and improving the conditions of the diabetic patients.
Evaluation of the anti-oxidizing ability of Lactobacillus paracasei N1115
Diabetes is often accompanied by oxidative stress, i.e.: excessive production of free radicals and destruction of the antioxidant defense system. This will lead to cellular and enzymatic damage, increased lipid peroxidation, and development of insulin resistance. It also destroys islet beta cells leading to diabetes. The oxidation resistance of probiotics has been proven by various in vivo and in vitro experiments, and for the current research, the possible oxidation resistance mechanisms of probiotics mainly include four mechanisms, namely resistance to oxygen stress, elimination of a free radical system, regulation of a signal path, inhibition of lipid peroxidation and influence of metal ions. Free radicals are mainly produced by mitochondria and released into cytoplasm, and excessive free radicals can generate chains with organismsThe formula (II) causes cell damage. The free radicals in the organism mainly comprise the following components: hydroxyl radical (OH)-) Superoxide anion radical (O)2-) DPPH radical, hydrogen peroxide (H)2O2) Singlet oxygen (O)2) And the like. The existing data show that part of probiotics have stronger free radical scavenging capacity through the research of an in-vitro simulated oxidation system.
Activating the strain, inoculating in MRS liquid culture medium, culturing at 37 deg.C for 24 hr, collecting bacterial liquid, centrifuging at 4000 rpm and 4 deg.C for 10 min, collecting thallus, and washing with normal saline for 1 time. Resuspending the cells, adjusting the solubility to 1X 107CFU/mL. The hydroxyl radical scavenging capacity (Fenton method), superoxide anion scavenging capacity (o-phenol autoxidation method) and DPPH scavenging capacity of the probiotics are respectively detected. The specific method is shown in table 1:
TABLE 1 detection method for oxidation resistance of probiotics
The experimental results are shown in fig. 3-5, and the results show that N1115 has better scavenging capacity for DPPH, hydroxyl radicals and superoxide anions, which indicates that N1115 has good antioxidant capacity and can relieve the oxidative stress effect accompanying diabetes.
Evaluation of immunomodulatory Capacity by Lactobacillus paracasei N1115
Diabetes is also associated with an immunoinflammatory response. The probiotics are proved to have the functions of promoting the growth and development of immune cells, tissues and organs, stimulating the organism to generate immune regulation factors such as antibodies and the like, and enhancing the activity of phagocyte, thereby improving the immune function of a host. Interleukin-1 b (IL-1 b), interleukin-6 (IL-6), interleukin-10 (IL-10) and interleukin-12 a (IL-12 a) play an important role in the immune response process of a human body. Through a real-time fluorescent quantitative PCR technology, the influence of the N1115 incubated macrophage J774A.1 on the expression levels of the immune regulatory factors IL-1b, IL-6, IL-10 and IL-12a mRNA in cells can be evaluated. This test was conducted separatelyThe activity and heat inactivated N1115 strain are evaluated, and the bacterial suspension concentration is adjusted to 106CFU/mL, and after incubating with J774A.1 cells for 3 h, detecting the change of the mRNA expression levels of IL-1b, IL-6, IL-10 and IL-12a in macrophage J774A.1.
The internal reference gene GAPDH and 4 kinds of cell factors are subjected to fluorescence quantitative detection, and the relative expression change of the 4 kinds of cell factor mRNA in macrophage J774A.1 is compared, and the results are shown in FIGS. 6 and 7. Whether the strain N1115 has activity has a different effect on the cytokine of macrophage j774a.1. The control group was a blank cell group without addition of bacteria. The relative expression quantity of mRNA of each cytokine of J774A.1 under the action of activity and heat inactivation N1115 is increased or reduced in different degrees. From this, it is known that lactobacillus paracasei N1115 has a regulatory effect on the mRNA expression levels of interleukin-1 b (IL-1 b), interleukin-6 (IL-6), interleukin-10 (IL-10) and interleukin-12 a (IL-12 a), thereby enhancing the immunity of the body, reducing the occurrence of inflammation, and preventing and controlling the occurrence of diabetes.
Examples 2-5 milk powder for improving diabetes prepared using Lactobacillus paracasei N1115
Examples 2 to 5 are milk powders for improving diabetes prepared using lactobacillus paracasei N1115, respectively. The raw materials have the same composition, but the different dosage of the raw materials is different, and the specific mixture ratio of the raw materials is shown in the following table 2:
TABLE 2 raw material ratios of examples 2-5
The milk powder for improving the diabetes of the corresponding embodiment is obtained by uniformly mixing the raw material components.
Example 6 use of the milk powder provided in examples 2-5 to improve diabetes
120 subjects were recruited in the study, all suffering from type II diabetes, 60 subjects in the control group, and 60 subjects in the drink test group. By the end of the test, 55 persons were counted in the control group and 53 persons were counted in the trial group. The average age of the subjects was 56.9 + -6.7 years, and the average body weight was 71.5 + -12.5 kg.
The trial drinking group drinks the milk powder for improving the diabetes mellitus prepared in the embodiment 2 of the invention every day, the dosage of the milk powder is 50g, warm water is added into the milk powder, the milk powder is uniformly mixed to obtain 500mL of milk, and the milk is continuously drunk for 60 days. The control group was blank.
The statistical method comprises the following steps: data entry was performed using EpiData 3.1 and statistical analysis was performed using SPSS13.0 software. The self-contrast data is analyzed by single-factor repeated measurement variance, two-by-two comparison is performed by LSD-t test, two-group mean comparison is performed by t test in groups, and 2-group rate comparison is performed by x test2And (6) checking. The metrology data is expressed as mean ± standard deviation or median (25 decimals per cent, 75 decimals per cent). P<0.05 indicates that the difference is significant.
The improvement rate of thirst and polydipsia symptoms of the test drinking group is higher than that of the control group (P = 0.032) when the test drinking is carried out for 2 months, and the improvement rates of polyphagia, bulimia, diuresis, lassitude, hypodynamia, soreness and weakness of waist and knees and insomnia of the test drinking group have no statistical significance with the difference of the control group (P > 0.05).
TABLE 3 comparison of the improvement rate of diabetic symptoms in the control group after 2 months of trial drinking and the trial drinking group after 2 months of trial drinking
*: the difference is statistically significant.
Claims (3)
1. The application of lactobacillus paracasei N1115 for improving diabetes is characterized in that: the lactobacillus paracasei N1115 is used for preventing or relieving diabetes.
2. The milk powder for improving the diabetes is characterized by comprising the following raw materials in parts by weight: 40-45 parts of raw milk powder, 28-33 parts of desalted whey powder, 5-8 parts of white granulated sugar, 4-4.5 parts of anhydrous cream, 3-4 parts of soybean oil, 3-4 parts of lactose, 2-4 parts of concentrated whey protein powder, 1.5-2 parts of galacto-oligosaccharide, 1-1.5 parts of fructo-oligosaccharide, 0.5-1 part of phospholipid, 0.1-0.2 part of DHA algal oil powder, 0.1-0.3 part of arachidonic acid powder, 0.05-0.2 part of choline chloride, 0.1-0.8 part of calcium carbonate, 0.02-0.06 part of nucleotide and 0.1 part of compound vitamin0.5 part, 0.1-0.3 part of composite mineral substance and 0.1-1 part of probiotic powder capable of improving diabetes; wherein the content of Lactobacillus paracasei N1115 in the probiotic powder capable of improving diabetes is 1 × 108CFU/g-9×1010CFU/g。
3. The use of milk powder for improving diabetes as claimed in claim 2, wherein: the milk powder for improving diabetes is used for preventing or relieving diabetes.
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Application publication date: 20191213 |