CN111493325A - Application of plant selenium peptide in preparation of composition for improving obesity and metabolic syndrome - Google Patents
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Abstract
The invention discloses an application of a plant selenium peptide in preparing a composition for improving obesity and metabolic syndrome, which comprises food, health care products, medicines, nutritional supplements, food additives and beverages, wherein the plant selenium peptide is preferably an organic selenium source extracted from cruciferous plants, the content of organic selenium is higher than 90%, the content of peptide is higher than 10%, and the plant selenium peptide can improve oxidative stress, enhance the activities of antioxidase such as GPx, SOD, CAT and TAOC and reduce the MDA level, inhibit the expression levels of inflammatory factors I L-6, TNF- α, I L-1 β and MCP-1, inhibit the expression levels of genes FABP4, FAS, PPAR-gamma, SREBP1, HMX and ACAT, promote the expression of lipolysis genes PPAR- α 1- α, L P L, GPX and L CAT, maintain the integrity of intestinal mucosa, tightly regulate the expression levels of zon-1 and ccludin, regulate the composition of intestinal microorganisms, promote the beneficial bacteria Blautia and have wide application prospects in producing Blaulautus and L.
Description
Technical Field
The application relates to the field of selenium peptide application, in particular to application of plant selenium peptide in preparing a composition for improving obesity and metabolic syndrome.
Background
With the rapid development of economy and the change of life style of people, obesity has become the most important public health problem in the world. According to the report of 'lancets' in 2017, over 22 hundred million people are overweight globally, and 13 hundred million adults are obese. In addition, there is increasing evidence that obesity is a high risk factor for many chronic diseases including type 2 diabetes, non-alcoholic fatty liver, cardiovascular disease and many cancers, severely affecting quality of life and increasing the global medical burden. Therefore, it is of great practical significance to develop more effective methods of preventing obesity and its complications.
The clinical features of obesity include not only massive fat accumulation, but also dyslipidemia, insulin resistance, chronic inflammation and excessive oxidative stress, excessive fat exposure causes organ disorders, thereby activating the immune system, causing immune cell infiltration, and generating inflammatory factors TNF-a and I L-6, thereby increasing oxidative stress, and at the same time, obesity can promote the formation of ROS, aggravate oxidative stress, further promote the release of inflammatory factors, and form vicious circle.
In addition, numerous studies have shown that intestinal microorganisms are closely associated with obesity and metabolic syndrome. High fat diet-induced intestinal micro-ecological disorders can affect host energy balance, cause abnormal glucose metabolism, affect related hormone secretion and produce inflammation. The regulation of intestinal microorganisms and the promotion of the growth of intestinal beneficial bacteria become important means for improving obesity and related diseases.
Selenium is an essential trace element for human body, and has various beneficial biological functions, including antioxidation, cardiovascular protection and cancer prevention, however, the potential mechanism and molecular mechanism are not clear. Selenium exists in two forms of inorganic selenium and organic selenium in nature, the safe dose of the inorganic selenium is narrow, and selenium poisoning can be caused when the dose exceeds a certain dose. It is generally accepted that organic selenium is safer and more available for absorption than inorganic selenium.
Selenium plant strengthening is an effective means for supplementing selenium at present, and plants can convert inorganic selenium into organic selenium. The plant selenium peptide (CSP) is prepared by hydrolyzing and processing cardamine violifolia, a high-selenium plant of Cruciferae by compound protease, has high organic selenium content of more than 90%, exists in a specific organic selenium form, and has higher safe dose. In vitro studies show that the plant selenium peptide has strong capacity of eliminating oxidative free radicals. The literature "research progress of iron, zinc, copper, selenium, manganese and metabolic syndrome" zhuchui et al, journal of the aged science in china, discloses the correlation between selenium and "metabolic syndrome", "insulin" and "obesity". Reviewing the relationship of selenium to diabetes, the possible mechanism is by increasing glutathione antioxidant enzyme activity, which is a common consensus for selenium as it is the active site of the enzyme. In addition, selenium is reported to possibly enhance the antioxidant effect or promote insulin secretion, but the action mechanism is not clear, and in addition, inorganic selenium exceeding a certain dosage can have certain side effects on diabetes and even cause the diabetes. The patent system explains the action mechanism of the plant organic selenium and selenium peptide for improving obesity and metabolic syndrome, and is realized by synergistically regulating intestinal microorganisms and related metabolic genes through antioxidation and anti-inflammation, so that the change of blood sugar is included, and the blood lipid disorder is also improved. Meanwhile, the cardamine violifolia plant selenium peptide is applied for the first time in the field, and has a better effect under a high-dose effect. At present, no clear report shows that the plant selenium peptide has protection effect on obesity and metabolic syndrome and can improve oxidative stress.
Disclosure of Invention
The invention aims to find a new application of plant selenium peptide derived from cruciferous high-selenium plant cardamine violifolia in preparing food or special medical food for improving obesity and metabolic syndrome.
The invention is realized by the following technical scheme:
the invention provides an application of plant selenium peptide in preparing a composition for improving obesity and metabolic syndrome; the plant selenium-titanium has an organic selenium content (mass) preferably higher than 90%, and a peptide content (mass) preferably higher than 10%.
In the application, the preparation method of the plant selenium peptide is a conventional method in the field, such as Chinese patent ' a preparation method of cardamine violifolia selenium polypeptide with high organic selenium content ' (publication number: CN 108157579A) or Chinese patent ' an antioxidant selenium polypeptide derived from cardamine violifolia and a preparation method thereof ' (publication number: CN 110483619A) ' to prepare the obtained plant selenium peptide; the plant selenium peptide is preferably organic selenium source extracted from plants of Brassicaceae (such as Cardamine cordifolia).
Furthermore, in the composition for improving obesity and metabolic syndrome, the dosage of the added plant selenium peptide is equivalent to 200-800 ug/day of the dosage taken by a human body.
Further, the composition for improving obesity and metabolic syndrome includes, but is not limited to, one or more of food, health product, medicine, nutritional supplement, food additive, and beverage; the food, health product, medicine, nutritional supplement and food additive all have the effects of improving obesity and metabolic syndrome, especially have the effects of improving chronic inflammation caused by high fat diet, improving lipid accumulation, improving intestinal metabolic disorder, improving non-alcoholic fatty liver, improving insulin resistance, preventing and improving central obesity and improving one or more of type II diabetes.
In the present application, the food includes a special medical food, a functional food (health food), a functional food ingredient, and the like; the term "special medical food" generally refers to a formula food specially processed and prepared for meeting the special needs of people with limited food intake, digestive absorption disorder, metabolic disorder or specific disease states for nutrients or diet, such as the food types specified in the general rules of GB 29922 and 2013 food safety national standard special medical application formula food; the pharmaceutical product generally comprises one or more pharmaceutically acceptable carriers, diluents or excipients, such as a powder, granules, tablets, etc.
The embodiment of the invention discloses application of plant selenium peptide (the content of organic selenium is more than 90 percent and the content of peptide is more than 10 percent) prepared from cardamine violifolia of cruciferae in preparing a composition for preventing and improving obesity and metabolic syndrome, and experimental results show that the plant selenium peptide can obviously reduce weight gain, reduce fat accumulation, obviously improve obesity, blood lipid metabolic disorder and insulin resistance and restore the metabolic stability of blood sugar within a certain dosage range; and under the action of very high selenium content, the selenium-enriched tea is still safe and effective, and has no obvious adverse reaction.
The other examples show that the plant selenium titanium can effectively maintain the barrier integrity of intestinal mucosa, increase the expression of tight junction protein, regulate the composition of intestinal microorganisms, increase the expression of beneficial bacteria, increase the abundance of beneficial lactobacillus in the intestinal tract, increase the abundance of bacteria Blautia generated by short-chain fatty acid, particularly increase the abundance of L actinobacillus and better improve lipid accumulation and insulin resistance.
Drawings
FIG. 1 is a graph showing the results of measurement of weight loss and fat accumulation in mice fed a high fat diet with plant selenium peptide (CSP);
wherein, A is a schematic diagram of Weight and Weight gain (Body Weight); b is a schematic diagram of the Weight of the white fat and the brown fat of the liver and the epididymis (Weight); c is a Food intake (Food intake) diagram; d is a schematic diagram of energy intake (Energeinak); e is a weight gain specific value (Body weight gain).
The assay data are shown as mean ± sem (n ═ 10); the statistical method is one way ANOVA testwithTukey; p <0.05, P <0.01, P <0.001 vs NCD; # P <0.05, # P <0.01,
###p<0.001 vs HFD。
FIG. 2 is a schematic diagram showing the results of detecting the fat and liver tissue hypertrophy of mice fed with high fat by cardamine hirsute plant selenium peptide (CSP);
wherein, A is the picture of the result of HE staining (200x) of white adipose tissues of epididymis; b is the result photograph of oil red staining (200X) of liver tissue.
FIG. 3 is a schematic diagram of plant selenium peptide (CSP) for improving dyslipidemia and insulin resistance in hyperlipidemic-fed mice;
wherein A is a schematic diagram of a detection result of blood fat level (TG, TC, HD L-C and L D L-C), B is a schematic diagram of an oral glucose tolerance (OGTT) detection result, C is an area under an OGTT curve (OGTT-Auc), D is a schematic diagram of fasted Fasting blood glucose (fasted Fasting glucose), E is a schematic diagram of fasted insulin (fasted insulin), and F is a schematic diagram of insulin resistance index (HOMA-IR);
the data above are shown as mean ± sem (n ═ 10), statistical method one way ANOVA test with tukey; # P <0.05, # P <0.01, # P <0.001 vs NCD, # P <0.05, # P <0.01, # P <0.001 vs hfd; # P & P <0.001 vs CSP L.
FIG. 4 is a schematic diagram showing the results of the measurement of the level of oxidative stress and inflammation in mice fed with high fat by plant selenium peptide (CSP);
wherein, A is a schematic diagram of the detection result of MDA level, B is a schematic diagram of the detection result of GSH-Px activity, C is a schematic diagram of the detection result of SOD activity, D is a schematic diagram of the detection result of CAT activity, E is a schematic diagram of the detection result of TAOC activity, and F is a schematic diagram of the detection result of serum Inflammation level (Inflammation L evel, I L-6, I L-1 β - α, and MCP-1);
data are shown as mean ± sem (n ═ 6), statistical method one way ANOVA test with tukey; p <0.05, P <0.01, P <0.001 vs NCD; # P <0.05, # P <0.01,
###p<0.001 vs HFD。
FIG. 5 is a schematic diagram showing the results of the expression of genes involved in the regulation of hepatic fat and cholesterol metabolism by plant selenopeptide (CSP);
wherein, A is a schematic diagram of the expression result (Relative mRNA expression) of GPX and FABP4 related mRNA, B is a schematic diagram of the expression result (Relative mRNA expression) of adipogenic genes FAS, PPAR-gamma and SREBP1C related mRNA, C) is a schematic diagram of the expression result (Relative mRNAexpression) of lipolytics PPAR- α 1- α and L P L related mRNA, and D) is a schematic diagram of the expression result (Relative mRNA expression) of cholesterol metabolism genes (L CAT, HMGR and ACAT related mRNA;
data are shown as mean ± sem (n ═ 6); the statistical method is one way ANOVA test with Tukey; p <0.05, P <0.01, P <0.001 vs NCD; # P <0.05, # P <0.01,
###p<0.001 vs HFD。
FIG. 6 is a schematic diagram showing the results of plant selenium peptide (CSP) improving the integrity of intestinal tract and regulating the gene expression of tight junction proteins, cholesterol ZO-1 and occludin;
wherein A is a schematic diagram of the ileum HE staining result of 4 groups of mice; b is a schematic diagram of the expression results (Relative mRNA expression) of ZO-1 and occludin related mRNAs;
data are shown as mean ± sem (n ═ 6), statistical method one way ANOVA test with tukey; p <0.05, P <0.01, P <0.001 vs NCD; # P <0.05, # P <0.01, # P <0.001 vs HFD.
FIG. 7 is a graph showing the results of measurements of the composition of gut microbes in mice fed a high fat diet by plant selenium peptide (CSP);
wherein, A is a wain graph containing unique OTU among different groups, B is a main component analysis result graph of OTU level, C is a relative abundance detection result graph of mouse intestinal microorganisms at a portal level, D is a heat map analysis result graph of bacteria with higher abundance at a genus level, E is a Cladogram analysis result graph, and F is an L EfSe analysis result graph (L EfSeanalysis, &lTtT transition = L "&gTt L &/T &gTt DA > 4).
FIG. 8 is a schematic representation of species of gut microbes with significant differences between different groups (. SP <0.05,. SP <0.01) analyzed by genus-level comparison using the Wilcoxon rank-sum test statistical method;
wherein, A is a comparison diagram of CK group and HFD group microorganisms, B is a comparison diagram of HFD group and CSP L group microorganisms, and C is a comparison diagram of HFD group and CSPH group.
FIG. 9 is a thermal schematic of the correlation between gut microbiome composition and obesity correlations;
the correlation of intestinal microorganisms with body weight (Bw), liver weight (L w), epididymal fat weight (Efw), TG, TC, HD L D L-1 (MCP1), I L-1 β (I L1), TNF- α (TNF), I L-6 (I L6), leptin and adiponectin is analyzed by using Spearman's method, the blue color is negative correlation, the red color is positive correlation, and the statistical analysis shows that the two-tail Student' st-test has p <0.05, p <0.01 and p < 0.001.
Detailed Description
Example 1 preparation of organic selenium plant selenium peptide (CSP)
Cardamine violifolia powder (protein 14.42%, total selenium 1421ppm) was immersed in NaOH solution (pH8.0) at a substrate concentration of 1:20, followed by addition of alkaline protease (Stanny-Jones) at an enzyme-to-protein material ratio of 2:100(g/g), enzymatic hydrolysis at 50 deg.C for 3h, followed by addition of neutral protease (Stanny-Jones) at an enzyme-to-protein material ratio of 1:100(g/g), pH adjustment to 7.0, and hydrolysis at 50 deg.C for 3 h. Inactivating enzyme in boiling water bath for 10min, centrifuging at 4000r/min for 10min, recovering supernatant, further ultrafiltering, concentrating, and freeze drying to obtain organic selenium plant selenium peptide (CSP).
Through detection, the total selenium content of the plant selenium peptide obtained in the embodiment is 1177ppm, the organic selenium content exceeds 90%, and the peptide content exceeds 10%.
In the embodiment, cardamine violifolia powder is derived from Enshi Deyuan health science and technology development Limited, cardamine violifolia is subjected to root fertilization by using sodium selenite, harvested after three months of growth, dried and crushed into powder, and the cardamine violifolia powder is disclosed by Chinese patent CN 201810015528.4; in addition, the preparation methods for preparing the CSP in the present example are all conventional methods in the art, such as the method disclosed in Chinese patent CN201810015528.4, and generally, the object of the present invention can be achieved as long as the organic selenium content in the CSP is ensured to exceed 90% (by mass) and the peptide content is ensured to exceed 10% (by mass).
Example 2 model establishment of obesity plant selenium peptide (CSP) to alleviate high fat diet-induced obesity in mice
40 SPF grade C57B L/6J mice (male, 6 weeks old, weight 19-21 g) were purchased from Beijing Wintonlifa laboratory animal technology, Inc. the animal experiments involved in the study procedure were approved by the animal welfare and ethics committee of the Beijing laboratory animal center.
The mouse model establishment method comprises the following steps:
experimental animals were acclimatized for one week (mice were given NC diet (basal diet, purchased from southerton teflon feed science and technology limited, TP23300) with a standard light-dark cycle of 12h, temperature of 23 ± 2 ℃, relative humidity of 55 ± 5%), after acclimatization 40C 57B L/6J mice were randomly divided into four groups of 10 animals each:
(1) NCD group (CK control group), continued to be fed with NC feed and sterile water for 6 weeks;
(2) HFD group (high fat group), HFD diet (high fat diet, available from south ton tamiflu feed science ltd, TP23302) and sterile water;
(3) HFD + CSP L group (CSP L group for short) HFD feed and sterile water were administered together with CSP obtained in example 1 (22mg/kg. bw, containing 26ug/kg. bw of selenium, equivalent to a human dose of 200 ug/day);
(4) HFD + CSPH group (CSPH group for short): HFD feed and sterile water were administered and CSP (88mg/kg. bw, 104ug/kg. bw of selenium, equivalent to a human dose of 200 ug/day) obtained in example 1 was administered.
Fecal samples were collected weekly and stored at-80 ℃ until week 9, wherein Oral Glucose Tolerance Test (OGTT) was performed on 4 groups of mice, respectively, after fasting for 12h, gavage of glucose solution (2.0g/kg,100g/m L, D-glucose) per mouse, after sampling blood from the tail vein at time points 30 min, 60 min, 90 min, 120 min, 150 min, 180 min before and after gavage (example 4), after week 10, animals were fasted for 12h, blood was collected from the aorta, and the animals were subsequently euthanized.
The livers, small intestines, brown fat, and epididymal white adipose tissues of 4 groups of mice were taken, weighed immediately, and then stored by quick freezing with liquid nitrogen at-80 ℃.
As shown in FIG. 1A, after ten weeks of high fat diet feeding, the high fat group (HFD) significantly increased body weight compared to the normal diet (NCD) control group, with an average body weight increase of 11.99g, while the CSP L and CSPH supplemented groups significantly decreased body weight increase compared to the high fat group, with final extra body weight gains of 9.61g and 9.11g, respectively, and the 4 groups of mice increased body weight specific values (Bodyweight gain), i.e., specific values of final body weight minus initial body weight (FIG. 1E), while the CSP supplemented group (CSP L, CSPH) significantly decreased high fat diet-induced liver weight (L iver) and epididymal white tissue (eWAT) weight gain, increasing brown fat weight (BAT) (FIG. 1B).
Furthermore, there was no significant difference in food intake versus energy intake between the high fat diet group and the CSP supplemented group (FIG. 1C, FIG. 1D), indicating that CSP L and CSPH reduced weight gain and fat accumulation not due to food changes.
Example 3 plant selenium peptide (CSP) reduction of high fat diet-induced liver and adipose tissue hypertrophy
Liver tissues and white adipose tissues of epididymis, which were freshly isolated from four groups of mice of example 2, respectively, were fixed in a 10% neutral formalin solution, then embedded with paraffin, cut into 5 μm tissue slices, and then HE-stained. Another part of liver tissue is cut into tissue slices with the size of 6 mu m and oil red staining is carried out. Tissue sections were analyzed under a microscope (200 x).
Microscopic examination results as shown in fig. 2A, HE staining revealed that normal group (NCD) white adipose tissue was of normal size, while high fat diet group (HFD) developed severe fat hypertrophy, and a large number of individual fat vesicles, however, CSP supplementation (CSP L, CSPH) restored high fat diet-induced adipose tissue abnormalities, particularly CSPH supplementation group (CSPH).
Similar results were also seen in the oil red staining of liver tissue (fig. 2B), with a significant increase in the area of oil red staining in the high lipid group CSP L supplementation (a mild relief of liver fat accumulation, while CSPH supplementation significantly improved liver fat accumulation, indicating that CSP supplementation (CSP L, CSPH) may protect the body from high lipid diet-induced steatohepatitis.
Example 4 plant selenium peptide (CSP) improves high fat diet-induced dyslipidemia and glucose homeostasis
This example performed an Oral Glucose Tolerance Test (OGTT) on 4 groups of mice from example 2 (9 weeks of feeding). after fasting for 12h, each mouse was gavaged with a glucose solution (2.0g/kg,100g/m L, D-glucose). blood was collected from the tail vein at time points 30 minutes, 60 minutes, 90 minutes, 120 minutes, 150 minutes, 180 minutes before and after gavage.
Serum lipid profile TG (total triglyceride), TC (total cholesterol), HD L-C (high density lipoprotein cholesterol), L D L-C (low density lipoprotein cholesterol) was tested using a commercially available kit from Nanjing Kangji corporation measuring mouse blood glucose levels using a sanno glucometer, calculating the area under the blood glucose curve (AUC) using the software GraphPadPrism 5.0 to quantify the cumulative change in blood glucose response fasting blood glucose levels were determined using a glucose oxidase kit, fasting serum insulin levels were determined using a Wuxi Chengzhi Bio E L ISA kit, and HOMA-IR index was calculated using the themaths formula to assess insulin resistance, specifically the formula HOMA-IR fasting glucose levels (mmol/L) × fasting insulin levels (μ M/M L)/22.5.
As shown in FIG. 3A, CSP L and CSPH were able to significantly reduce HFD-induced dyslipidemia, lower TG, TC, &lTtT transfer = L "&gTt L &lTt/T &gTt D L-C, and significantly increase HD L-C levels without significant dose dependence.
The oral glucose tolerance test showed that both CSP L and CSPH were able to significantly improve HFD-induced impaired glucose tolerance with some dose dependence (fig. 3B, fig. 3C).
Furthermore, CSP supplemented groups were able to significantly inhibit and reverse fasting glucose and insulin elevation (fig. 3D, fig. 3E), presenting a lower HOMA-IR index (fig. 3F) compared to HFD group. These results indicate that CSP supplementation can effectively ameliorate HFD-induced dyslipidemia and glucose metabolism disorders, and inhibit insulin resistance.
Example 5 plant selenium peptide (CSP) improves high fat diet-induced oxidative stress and chronic inflammation
The 4 groups of mice in example 2 were tested for serum MDA (malondialdehyde), TAOC (total antioxidant capacity), GPx (glutathione peroxidase), SOD and CAT (catalase) activities using an antioxidant assessment series kit from tokyo founding company.
The 4 groups of mice in example 2 were subjected to quantitative detection of serum MCP-1, TNF- α, I L-6, and I L-1 β levels using Nanjing's completed E L ISA kit.
And (3) detection results: as shown in fig. 4: compared with a normal control group, the MDA of the high-fat feeding group is remarkably increased, and the activity of TAOC, GPx, SOD and CAT is remarkably reduced. However, after CSP supplementation, these antioxidant enzyme activities were significantly increased and MDA was significantly decreased. No significant CSP dose-dependence was exhibited (fig. 4A-E).
At the same time, high fat feeding significantly increased the levels of inflammatory factors MCP-1 (monocyte chemotactic protein 1), TNF- α (tumor necrosis factor), I L-6 (interleukin 6), and I L-1 β (interleukin 1 β) over the normal control group-in contrast, CSP supplementation reversed this trend, significantly reducing the levels of these inflammatory factors (FIG. 4F).
Example 6 plant selenium peptide (CSP) modulation of expression of related genes involved in lipid and cholesterol metabolism
The genes related to the liver participating in lipid and cholesterol metabolism of the 4 groups of mice in example 2 were analyzed by real-time fluorescent quantitative PCR (qRT-PCR), and total RNA frozen in liver tissue at-80 ℃ was extracted using TRIzol, a total RNA extraction kit from Invitrogen, USA. Reverse transcription for cDNA synthesis and qRT-PCR analysis were performed using the promega reverse transcription kit. The forward and reverse primers for the genes tested are listed in table 1. Use 2-ΔΔCtThe method performs relative quantitative analysis of mRNA.
TABLE 1 primers for real-time fluorescent quantitative PCR analysis
As shown in FIGS. 5A to 5D, the HFD group significantly increased the expression of lipid transporter gene FABP4, lipid production genes FAS, PPAR- γ, and SREBP1C, as well as cholesterol synthesis-related genes HMGR and ACAT, reduced antioxidant selenoprotease GPx, lipid catabolic genes PPAR- α 1- α, and L P L, and cholesterol esterification gene L CAT, as compared to the normal control group, the CSP supplementation significantly increased the expression of GPX, PPAR- α 1- α P L, and L CAT, inhibited FAB4, FAS, PPAR- γ, SREBP1C, HMGR, and ACGR consistent with the results of reduced fat accumulation.
Example 7 phytoselenopeptide (CSP) restoration of high-fat diet-induced intestinal mucosal barrier leakage and increased claudin Gene expression
Ileum of 4 groups of mice in example 2 were each fixed in 10% neutral formalin solution, embedded in paraffin, sliced into 5 μm tissue slices, and then HE-stained and analyzed by microscopy. The claudin ZO-1 and occludin were analyzed by qRT-PCR.
As a result, the villi height of the small intestine was significantly shortened and the crypt depth was deepened with high-fat diet compared to the normal control group, whereas CSP L and CSPH significantly improved HFD-induced intestinal injury (FIG. 6A).
In addition, the expression level of the relative genes of the marker molecules of the intestinal integrity, namely the zon-1 and occludin, is consistent with the intestinal tissue morphology result, the high-fat diet remarkably reduces the expression of the zon-1 and occludin, and the CSP L and CSPH supplement groups obviously up-regulate the expression of the zon-1 and occludin compared with the HFD group (figure 6B), wherein CSPH promotes the expression of the ZO-1 better.
Example 8 plant selenium peptide (CSP) regulates high fat diet-induced intestinal microbial composition
Bacterial DNA in feces was collected from each of the 4 groups of mice in example 2 at week 10 using the Qiagen QIAamp DNA pool Mini Kit in Germany, and then subjected to quality control by agarose gel electrophoresis. Subsequently, the V3-V4 region of bacterial 16SrRNA was PCR amplified using primers 341F (5 '-CCTACGGGRSGCAGCAG-3', SEQ ID NO.29) and 806R (5 '-GGACTACVSGGGTATCTAAT-3' SEQ ID NO.30), followed by DNA library construction using the TruSeqDNA PCR-free sample preparation kit from Illumina, USA. Then, the DNA fragment was transferred to the American Gilg biomedical technology company in Shanghai for 16SrDNA sequencing and analysis on the Illumina MiSeq platform.
Results 352 OTUs were shared among all OTUs tested, 82 OTUs were unique among NCD, HFD, CSP L, and CSPH, 18 OTUs were unique among NCD, HFD, CSP L (FIG. 7A). PCA and PCoA analysis based on Bray-Curtis distance showed significant differences in intergroup microbial composition, and β -diversity was different between CSP supplemented and HFD groups (FIG. 7B).
Gut microbiota did not differ significantly at phylum level for each group, with a significant increase in firmicutes for all high lipid groups (fig. 7C), but at genus level, with heat map analysis showing the abundance of flora at genus level between groups (fig. 7D), L EfSe analysis showing key differences between groups (fig. 7E).
Furthermore, as shown in fig. 8A to 8C, the HFD group was significantly increased compared to the normal control group
unclassified _ f __ L achnolospiraceae and Allobaculum, reduction of Faecalibacilum, Blauia, norank _ f __ Erysipelotrichaceae, and Bifidobacterium. CSP L partially reversed these changes increasing Blauia and unclassified _ f __ L achnolospiraceae, whereas CSPH supplementation not only increased Blauia, a short chain fatty acid producing bacterium, but also significantly increased Blauia
L actinobacillus, reduced Staphylococcus and Dubosiella.
Example 9 Spearman correlation analysis to investigate the correlation of gut microbial changes with obesity-related metabolic index
The 4 groups of mice in example 2 were analyzed by heat map for their metabolic-related indices, body weight, liver weight, epididymal fat weight, HOMA-IR index, leptin and adiponectin levels, serum TG, TC, HD L D L-1, I L-1 β - α, and I L-6 relationship to gut microbes.
Analysis results as shown in fig. 9, high lipid group-enriched unclassified _ f __ L achnospiraceae was significantly and positively correlated with body weight, epididymal fat weight, HOMA-IR index, serum MCP-1, I L-1L, TNF-L, and I L-6, and the other enriched Allobaculum was nearly positively correlated with all indicators except liver weight, L D L, and MCP-1, in addition, these two bacteria were significantly and negatively correlated with adiponectin levels, however, Blautia enriched in CSP L and CSPH groups was significantly and negatively correlated with liver weight, serum TG, serum ttc, <tttransition &l "&tttl/ttt 6D L-C, I8-1L, TNF-L and I β -6 and blaustt β L &/ttt 5966D 2-3D 5967-1, and also showed significant and positive correlation with leptin receptor metabolism in weight, hct-9-11, hct-9-6, and hct 11-9-7-1-6, and in addition to the positive and negative correlation with weight of fat, hct 11, hct-7-6 and cswt-11 and cswt.
The present example shows that both CSP L and CSPH supplementation can effectively improve high fat diet-induced weight gain, excessive fat accumulation, lipid metabolism disorder, insulin resistance and hepatic steatosis, the underlying mechanism may be associated with improving oxidative stress, inhibiting inflammation, and thus regulating expression of related genes involved in fat and cholesterol metabolism, at the same time, CSP supplementation can regulate intestinal microbial composition changes, increase abundance of short-chain fatty acid producing bacteria Blautia.
In general, the studies of the above examples show that the plant selenium peptide can improve oxidative stress, enhance the activities of antioxidase such as GPx, SOD, CAT and TAOC, reduce MDA level, inhibit the expression levels of inflammatory factors I L-6, TNF- α, I L-1 β and MCP-1, inhibit the expression levels of lipid transport gene FABP4, lipid production gene FAS, PPAR-gamma and SREBP1C, and cholesterol synthesis genes HMGR and ACAT, promote the expression levels of lipid decomposition genes PPAR- α 1- α and L P L, antioxidative selenoprotein gene GPX and cholesterol esterification gene L CAT, maintain the integrity of intestinal mucosa, up-regulate the expression levels of claudin ZO-1 and ccludin, regulate the composition of intestinal microorganisms, promote the production of beneficial bacteria Blauia at low dosage, promote the production of beneficial bacteria Blauia and L actactin at high dosage, and can be used for preparing health-care products (such as obesity and obesity syndrome).
Sequence listing
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Claims (4)
1. Application of plant selenium peptide in preparing composition for improving obesity and metabolic syndrome is provided.
2. The use of claim 1, wherein the composition comprises one or more of a food product, a nutraceutical product, a pharmaceutical product, a nutritional supplement, a food additive, a beverage.
3. The use of claim 1, wherein the amount of the plant selenopeptide added to the composition is 200-800 ug/day.
4. The use of claim 1, wherein the plant selenopeptide is comprised of selenium in an amount greater than 90%.
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