CN106668635B - New application of children's seven-star tea - Google Patents
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Abstract
The invention relates to a new application of children's Qixing tea, the invention proves the application of children's Qixing tea in preparing anti-inflammatory and antibacterial medicines for protecting intestinal mucosa immune barrier and mechanical barrier, improving low intestinal mucosa immune function caused by high fat diet, regulating intestinal flora and improving intestinal mucosa cell morphology change and distribution disorder caused by high fat diet; the compound has the effects of resisting inflammation and bacteria, improving the change and distribution disorder of intestinal mucosa cell morphology, regulating intestinal flora and protecting intestinal mucosa barrier.
Description
Technical Field
The invention relates to the field of medicine application, in particular to new application of children's Qixing tea.
Background
The children Qixing tea is prepared from coix seeds, rice sprouts, hawthorn, lophatherum gracile, uncaria, cicada slough and liquorice, mainly aims at children food retention, has the main functions of appetizing, eliminating stagnation, clearing heat and arresting convulsion, and is used for children food retention, relieving heat, dyspepsia, poor appetite, dysphoria, easy convulsion, insomnia, unsmooth defecation and scanty and brownish urine.
The children's seven-star tea is mainly used in pediatrics and has the effects of promoting appetite, removing food stagnation, clearing heat and arresting convulsion, the hawthorn and the rice sprout are used as monarch drugs for treating main symptoms, the coix seed strengthens the function of the monarch drugs for treating the main symptoms, and the liquorice is used as a messenger drug for harmonizing the effects of the other drugs and can resist various' fire sources 'in children' bodies more safely and comprehensively. Wherein, the coix seed promotes diuresis and eliminates dampness, mainly attacking spleen fire and stomach fire; the hawthorn has the effects of invigorating stomach and helping digestion and mainly attacks stomach fire; the rice sprouts clear heat and relieve restlessness, and the lophatherum gracile promotes urination, mainly attacking heart fire; cicada slough is hydrophobic and erupts, mainly attacking heart fire and spleen fire; ramulus Uncariae cum uncis has effects of calming liver and calming endogenous wind; licorice root, radix Glycyrrhizae tonifies liver and qi, clears away heat and toxic material and mainly attacks liver fire. It is understood that whether the prescription is intended to address the anti-inflammatory and antibacterial effects is not considered when the prescription is first prepared.
In addition, under normal conditions, intestinal epithelial cells constitute a mechanical barrier and the intestinal flora forms a micro-ecological environment. On one hand, the intestinal symbiotic flora competes with pathogenic bacteria for nutrient substances to inhibit the proliferation of the pathogenic bacteria; on the other hand, the catabolism of saccharides produces short-chain fatty acids mainly comprising acetic acid, thereby inhibiting the translocation of toxins in the intestinal tract.
The intestinal flora permanent symbiotic bacteria take firmicutes (main members are clostridiales) and bacteroidetes (main members are bacteroides) as main dominant bacteria, and comprise bacteroides, previa, porphyromonas, clostridium, tenella, eubacterium, ruminococcus, lactobacillus and the like. Other sub-dominant bacteria such as Actinomycetes, mollicutes, etc. And the pathogenic bacteria comprise proteobacteria, verrucomicrobia, Elusimicrobi and the like. Proteobacteria are present in small amounts in the normal intestinal tract, often in proportions below 1%. Most bacteria of proteobacteria are facultative anaerobes, are difficult to adapt to strict anaerobic environment in intestinal tracts, and are mostly pathogenic bacteria. Common pathogenic enteromorpha bacteria are enterobacteriaceae, vibriaceae and pseudomonadaceae of the gamma-proteobacteria class. Sulphate-reducing bacteria of the class Proteobacteria, such as Desulfovir, are also the more common pathogenic bacteria in the intestine.
Mucosal immunity is an important component of the human body's entire immune system, performs mainly local specific immune functions, and is widely distributed in the gastrointestinal tract, respiratory tract, urogenital tract mucosa, and lymphoid tissues at the exocrine glands. Mucosal immunity is the first line of defense against infection because mucosal surfaces are in direct contact with external pathogens and pathogenic microorganisms are inactivated by sIgA and the like before invading body tissues.
When the intestinal mucosal immune barrier and the mechanical barrier are damaged by infection, disease, and other causes, the digestive tract is damaged. Recent studies have indicated that a high fat diet can lead to an impaired intestinal epithelial barrier and cause an inflammatory response in the gut.
Due to the complexity of disease treatment mechanism, the application of the children's Qixing tea in preparing medicaments for diminishing inflammation, resisting bacteria, protecting intestinal mucosa immune barrier and mechanical barrier, regulating intestinal flora, improving intestinal mucosa cell morphology change and distribution disorder and the like is not disclosed for a long time.
Disclosure of Invention
Therefore, in order to solve the problems, a new application of the children Qixing tea is needed.
The specific technical scheme is as follows:
an application of a Chinese medicinal composition in preparing anti-inflammatory and antibacterial medicines is prepared from Coicis semen, fructus oryzae Germinatus, fructus crataegi, folium Bambusae, ramulus Uncariae cum uncis, periostracum Cicadae, and Glycyrrhrizae radix.
In some embodiments, the traditional Chinese medicine composition is prepared from the following raw materials in parts by weight: 870-905 parts of coix seeds, 870-905 parts of rice buds, 435-460 parts of hawthorn, 655-685 parts of lophatherum gracile, 322-347 parts of uncaria, 100-124 parts of cicada slough and 100-124 parts of liquorice.
The invention also discloses application of the traditional Chinese medicine composition in preparing a medicine for protecting an intestinal mucosa immune barrier and a mechanical barrier.
The invention also discloses application of the traditional Chinese medicine composition in preparing a medicine for improving intestinal mucosa immunologic hypofunction caused by high fat diet.
The invention also discloses application of the traditional Chinese medicine composition in preparing a medicine for improving intestinal mucosa cell morphology change and distribution disorder caused by high fat diet.
The invention also discloses application of the traditional Chinese medicine composition in preparing a medicine for regulating intestinal flora.
The invention also discloses application of the traditional Chinese medicine composition in preparing a medicine for improving intestinal dysbacteriosis caused by high-fat diet.
Compared with the prior art, the invention has the following beneficial effects and advantages:
the invention proves the application of the children's Qixing tea in preparing the medicines for diminishing inflammation and resisting bacteria, protecting the intestinal mucosa immune barrier and the mechanical barrier, improving the low intestinal mucosa immune function caused by high-fat diet, regulating the intestinal flora and improving the intestinal mucosa cell shape change and distribution disorder caused by high-fat diet; the compound has the effects of resisting inflammation and bacteria, improving the change and distribution disorder of intestinal mucosa cell morphology, regulating intestinal flora and protecting intestinal mucosa barrier.
Drawings
Fig. 1 is a schematic diagram of the effect of pediatric seven-star tea (EXQ) on pathological tissue changes of a high-fat diet model mouse.
Detailed Description
The present invention is further illustrated by the following examples, which should not be construed as limiting the scope of the invention, which is defined by the claims.
The raw materials and equipment used in the following examples are all commercially available common raw materials and equipment.
Example 1
The traditional Chinese medicine composition in the following examples is a children seven-star tea, and the raw material formula and the preparation method thereof are as follows:
the formula is as follows: 893 parts of coix seeds, 893 parts of rice sprouts, 446 parts of hawthorn, 670 parts of lophatherum gracile, 335 parts of uncaria, 112 parts of cicada slough and 112 parts of liquorice.
(2) The extraction process comprises the following steps: decocting Coicis semen and fructus oryzae Germinatus in water for 2 times, each for 2 hr, filtering, mixing filtrates, concentrating to relative density of 1.08-1.12(55 deg.C), adding ethanol to make ethanol content reach 45%, standing, filtering, recovering ethanol from filtrate, concentrating into soft extract, decocting the rest 5 Chinese medicinal materials including fructus crataegi, folium Bambusae, ramulus Uncariae cum uncis, periostracum Cicadae, and Glycyrrhrizae radix in water for 2 times, each for 2 hr, filtering, mixing filtrates, concentrating to appropriate amount, and mixing with the above soft extract.
Example 2
Anti-inflammatory effect of children's Qixing tea
Effect of P-xylene on mouse ear swelling
1. Experimental Material
1.1 experimental animal SPF level Kunming mouse 40, each half of male and female, the weight 18 ~ 22g, provided by Guangzhou university of traditional Chinese medicine experimental animal center, the certification number: 4405900448.
1.2 experimental medicines of distilled water, aspirin (90mg/kg, Jiangsu Enhua pharmaceutical products Co., Ltd., lot number: 20090902), xylene (Shantou Guanghua chemical plant, lot number: 20040530), low dose of pediatric seven-star tea (3.84g/kg), medium dose of pediatric seven-star tea (7.67g/kg) and high dose of pediatric seven-star tea (15.34 g/kg).
1.3 Experimental apparatus JJ600 electronic weighing scale (Gao-Shi Shuangjie apparatus factory), Sartorius CP225D analytical balance (Germany, Sartorius Co.)
2. Grouping animals
Taking 40 Kunming mice with the weight of 18-22 g, and dividing the Kunming mice into 5 groups, wherein each group comprises 8 mice and each half of the mice is male and female; the group 1 is a model control group, the group 2 is an aspirin positive control group, the group 3 is a pediatric seven-star tea high-dose group, the group 4 is a pediatric seven-star tea medium-dose group, and the group 5 is a pediatric seven-star tea low-dose group.
3. Test method
The administration groups are respectively administrated by intragastric administration according to corresponding dose, and the model control group is administrated by intragastric administration of distilled water and aspirin positive control group which have the same volume as the intragastric administration of the aspirin suspension once a day for 7 days. On day 7 of administration, fasting was not prohibited for 12 h. After the last administration, xylene (0.04 ml/mouse) was uniformly applied to the right and the left of the right ear of each mouse 40min after the last administration to cause inflammation, the left ear was not used as a control, the cervical vertebrae were removed 1h after the inflammation to sacrifice the mice, both ears were cut off along the base line of the auricle, and the left and right ears were taken out at the same position by a punch with a diameter of 8 mm. Weighing on an analytical balance, and calculating swelling degree and inhibition rate.
Swelling degree-right ear mass-left ear mass;
the inhibition ratio (%) (average degree of swelling in model group-average degree of swelling in administration group)/average degree of swelling in model group × 100%
4. The statistical method adopts the sps software to perform T-test analysis on experimental data, and differences among groups are compared.
5. Experimental results the experimental results are shown in table 1.
As can be seen from Table 1, compared with the model control group, the positive aspirin control group can significantly inhibit the swelling degree of mouse auricle caused by xylene (P < 0.05); the low, medium and high dose groups of the children's seven-star tea have a tendency of inhibiting auricle swelling caused by P-xylene with the increase of the dose, and the high dose group obviously inhibits the mouse auricle swelling degree caused by xylene (P is less than 0.05). The experimental result shows that the children's seven-star tea has an inhibiting effect on the swelling degree of mouse auricle caused by xylene.
TABLE 1 influence of pediatric Qixing tea on mouse auricle swelling caused by xylene
Note: p <0.05 compared to model control group
(II) Effect on carrageenan-induced toe swelling in rats
1. Experimental Material
1.1 Experimental animals
40 SD rats, half male and half female, with a weight of 180-220 g, provided by Guangzhou university of traditional Chinese medicine laboratory animal center, and with a certification number: 4400590000031.
1.2 Experimental drugs
Distilled water, carrageenan (1%, Sigma, lot: 127H1227), indomethacin (5mg/kg, Guangdong Enhua pharmaceutical Co., Ltd., lot: 20090902), pediatric seven-star tea low dose (3.84g/kg), pediatric seven-star tea medium dose (7.67g/kg), and pediatric seven-star tea high dose (15.34 g/kg).
1.3 Experimental apparatus JJ600 electronic weighing scale (Gao-Shi Shuangjie apparatus factory), self-made toe volume measuring instrument
2. Grouping animals
Taking 40 SD rats with the weight of 180-220 g, dividing the SD rats into 5 groups with 8 rats in each group and half of male and female rats. The group 1 is a model group, the group 2 is an indomethacin positive control group, the group 3 is a pediatric seven-star tea high-dose group, the group 4 is a pediatric seven-star tea medium-dose group, and the group 5 is a pediatric seven-star tea low-dose group.
3. Test method
The administration groups are respectively administrated by intragastric administration according to corresponding dose, the model group is intragastric infused with distilled water with the same volume, and the positive group (indometacin) is intragastric infused with indometacin suspension, and the intragastric administration is carried out once a day for 7 days continuously. On day 7 of administration, fasting was not prohibited for 12 h. Before administration, the volume of the right hind paw of each group of rats was measured by toe volume measuring instrument to obtain the volume of the ante-inflammatory paw. Rats in each group are respectively administrated by intragastric administration, a model group is intragastric administered with distilled water with the same volume, and a positive group is intragastric administered with indomethacin suspension. After 0.5h of administration, each group of rats was inflamed by injecting 0.l ml of carrageenan under the thenar membrane of the right hind paw, and the volume of the paw was measured at 1h after the model was made, so that the difference between the volume of the paw before and after inflammation was taken as the degree of swelling.
4. The statistical method adopts the sps software to carry out paired T-test analysis on experimental data, and the differences among groups are compared.
5. Experimental results the experimental results are shown in table 2.
As shown in Table 2, compared with the model group, after the carrageenan causes the toe swelling, the positive drug indometacin and the children seven-star tea can obviously inhibit the foot swelling of the rats within 1h of inflammation (P is less than 0.05) at low, medium and high doses. The experimental result shows that the children's Qixing tea has the effect of transiently inhibiting the swelling of feet.
TABLE 2 influence of pediatric Qixing tea on carrageenan-induced swelling of toes in rats
Note: p <0.05, P <0.01, compared to model group
The anti-inflammatory experimental results show that: the children's Qixing tea can inhibit rat foot swelling caused by carrageenan, and can inhibit mouse auricle swelling caused by xylene at high dose, thus suggesting that the children's Qixing tea has anti-inflammatory effect.
Antibacterial effect of Qixing tea for children
(I) antibacterial test
1. Experimental Material
1.1 Experimental drugs
Distilled water, infantile Qixing tea, Staphylococcus aureus, beta hemolytic streptococcus, alpha hemolytic streptococcus, Pseudomonas aeruginosa, Staphylococcus epidermidis, Escherichia coli, and Salmonella typhi (all from the biological product research institute of Beijing Ministry of health).
1.2 test methods
In vitro antibacterial assay (liquid test tube method)
The infantile seven-star tea is prepared from nutrient broth, contains 9 concentrations of crude drugs of 400, 200, 100, 50, 25, 12.5, 6.25, 3.123 and 1.5625mg per mL, and is sterilized by steam in a tube of 1 mL. For streptococcus experiments, 1% of glucose needs to be added into a sterilized solution. The strain control is culture medium without drug and test bacteria, and the drug control is liquid medicine without test bacteria. 0.1mL of experimental bacterial liquid (8h culture) in a ratio of 1:2000 was added to each concentration tube and strain control tube of each liquid, and the mixture was cultured at 37 ℃. After 18 hours, the result is observed, and the sterile growth of each tube is visually observed by taking turbidity as an index, so that the Minimum Inhibitory Concentration (MIC) is determined. The results are shown in Table 3.
4. Results of the experiment
As can be seen from Table 3, the Qixing tea for children has different degrees of bacteriostasis on tested strains (mainly common pathogenic bacteria and conditional pathogenic bacteria causing respiratory tract infection and intestinal tract infection), has MIC (minimal inhibitory concentration) of 12.5-50 mg/mL for each strain, and has certain bacteriostasis on most of tested strains.
TABLE 3 infantile minimal inhibitory concentration of Qixing tea
The antibacterial experiment result shows that the children's Qixing tea has different degrees of bacteriostasis on tested strains (mainly common pathogenic bacteria and conditional pathogenic bacteria causing respiratory tract infection and intestinal tract infection); the conclusion of the antibacterial experiment indicates that the children's Qixing tea has the function of in vitro bacteriostasis.
Example 3
Immune function regulating effect of children's Qixing tea on high fat diet model mouse and mechanism research thereof
(I) regulating effect on immune function of high-fat diet model mouse and mechanism research thereof
1. Experimental Material
1.1 Experimental animals
SPF-level Kunming young mouse, male, 8 ~12 g, provided by Guangzhou university of traditional Chinese medicine laboratory animal center, license number: SCXK (Yue) -2013-0020.
1.2 animal feed
The common feed is a complete pellet feed for common mice, which is provided by the laboratory animal center of Guangzhou Chinese medicinal university;
self-made feed is high in calorie and high in protein: the raw materials are milk powder (Nestle whole milk powder, specification 400 g/bag), bean powder (southern brand bean powder, specification 600 g/bag), flour (XueJian brand flour, specification 1 kg/bag), and fish floss (Weiyi brand flag fish floss, specification 200 g/can).
1.3 laboratory instruments and apparatus
The instrument No. 6 mouse is a stomach irrigator, an injector (1ml), a dissecting forceps, an elbow forceps, a dissecting scissors, a centrifuge tube, a homogenate tube and a micropipettor.
The JJ600 electronic weighing scale (double Jie instruments factory in ever-maturing city), the electrothermal constant temperature blast drying box (model DG20-002), the centrifuge (model KA-IOOO), the homogenizer (model IKAT 18).
1.4 reagents and drugs
Reagent: picric acid, 75% alcohol, 4% paraformaldehyde, normal saline, PBS (PH7.4), xylene (100%), paraffin, concentrated hydrochloric acid, hematoxylin (china guangzhou chemical reagent factory), eosin (china guangzhou chemical reagent factory), aluminum potassium sulfate (china guangzhou chemical reagent factory), sodium iodate (china guangzhou chemical reagent factory), glacial acetic acid (china guangzhou chemical reagent factory), glycerol (china guangzhou chemical reagent factory).
Medicine preparation: low dose of infantile seven-star tea, medium dose of infantile seven-star tea, high dose of infantile seven-star tea, and 52% milk solution.
The kit comprises: mouse secretory immunoglobulin A (sIgA) enzyme-linked immunoassay kit (Beijing Chenglin Biotechnology, Inc.), mouse TNF-alpha kit (eBioscience), and mouse IL-10 kit (eBioscience).
2. Test method
2.1 animals 40 male Kunming mice are selected in groups, the weight is 8-12 g, the animals are divided into 5 groups, and each group comprises 8 animals. The group 1 is a normal group, the group 2 is a high fat diet model group, the group 3 is a pediatric seven-star tea low dose group (0.75g extract/kg), the group 4 is a pediatric seven-star tea medium dose group (1.5g extract/kg), and the group 5 is a pediatric seven-star tea high dose group (3g extract/kg).
2.2 feed preparation
High protein and high calorie feed: grinding the dried fish floss into powder, mixing the dried fish floss, flour, milk powder and bean powder according to the weight ratio of 1:1:1:2, adding a proper amount of water, uniformly stirring, cutting the feed into small pieces with the same size as that of a common feed, putting the small pieces into an electric heating constant-temperature blast drying oven, heating and baking at the temperature of 40-45 ℃, drying, and storing in a refrigerator at the temperature of 4 ℃ for later use after the dried fish floss is manufactured.
High protein high calorie solution: the milk powder and the water are mixed according to the weight ratio of 52:48, and are stirred uniformly to prepare milk solution with the concentration of 52 percent, and the milk solution is prepared before each intragastric administration.
2.3 preparation of the medicine, 3.75g, 7.5g and 15g of the extract of the children's Qixing tea are respectively weighed and dissolved to 100ml by distilled water to prepare low, medium and high dosage solutions of the children's Qixing tea.
2.4 preparation of reagents
Mayer's hematoxylin stain: the corresponding reagents were prepared as in table 4.
TABLE 4 Mayer's hematoxylin stain formula
Dissolving hematoxylin in absolute ethanol (solution A), dissolving potassium aluminum sulfate in distilled water, heating to promote dissolution (solution B), adding solution A into solution B, mixing, adding sodium iodate, adding glacial acetic acid after hematoxylin is oxidized into mauve, adding glycerol, filtering, and oxidizing.
0.8% eosin dye solution: 8g of eosin is dissolved in 1000mL of distilled water, mixed uniformly, and then 10 drops of concentrated glacial acetic acid are added dropwise, and bottled for later use.
0.5% hydrochloric alcohol: 800mL of 95% alcohol is used for complementing distilled water to 1000mL, namely 75% alcohol, 5mL of concentrated hydrochloric acid is added dropwise, and the mixture is uniformly mixed for later use.
2.5, molding: mice in the high-fat diet model group and the children seven-star tea administration groups are fed with special high-protein high-calorie feed and freely eat and drink water. On the 2 nd to 8 th day of the experiment, the patient is fed with 52% milk solution (0.2mL/10g, gavage) 2 times a day.
After high-fat diet, the mice have lassitude, reduced activity, crouched and piled, and yellow and lusterless fur; the food consumption is reduced; the abdomen is swollen and full, the urine is yellow, the feces are sticky, greasy and soft, and the color is brown yellow; the dissection can be seen: most of them are the enlargement of the stomach and intestine cavity and the flatulence. Basically accords with the clinical manifestations of inflammation caused by high fat diet of children, and indicates that the model building is successful.
2.6 administration: on the 2 th to 8 th days of the experiment, the low, medium and high doses of the children's Qixing tea (equivalent to 0.75g, 1.5g, 3.0g of extract/kg, 0.2ml/10g) are respectively given to the children's Qixing tea component, 1 time per day and 7 days are continuously given. The normal group and the high fat diet model group were perfused with equal amounts of distilled water.
2.7 anatomical material selection: on the 9 th day of the experiment, weighing, picking up eyeballs and blood, killing the mice by dislocation of cervical vertebrae after blood flowing out, opening the abdominal cavity, observing the change of gastrointestinal tracts, picking up thymus and spleen, weighing and recording, and calculating the organ index according to the following formula.
Organ index (weight of organ/body weight) × 1000 (unit: mg/g)
Finding the cecum, taking the cecum excrement, cutting the colon from the ileocecum to the rectal part, removing residual excrement in the colon by using dissecting forceps, and washing the colon by using normal saline for later use.
2.8 observation indexes and detection:
relative abundance of intestinal flora: total DNA was extracted from 100mg of the sample according to the operation manual of the fecal genome extraction kit omega D40-15. After the DNA concentration was detected by a nucleic acid concentration detector, the sample was stored at-80 ℃. Amplifying the 16SrRNA V1-V3 regions on an Illuminaseq platform, and performing double-End sequencing by constructing a small fragment library by using a double-End sequencing (paired-End) method.
The specific process is as follows: 1) PCR amplification of target fragments, quality inspection after mixing samples, and re-amplification of unqualified samples. 2) And (3) operating the qualified sample on a computer to carry out Illumina high-throughput sequencing, wherein the sequencing read length is 2 multiplied by 250 bp. 3) The data is sorted by barcode and the PE reads are spliced into Tag. A total of 880700 Tags were obtained for all samples, with an average of 29356 for each sample and an SD of 190; the average length of Tag is 252bp, and the SD value is 0 bp. The relative abundance of each species was analyzed using 16S. And (3) sequencing 16SrDNA to detect the intestinal flora, comparing the intestinal flora level with the integral intestinal flora level of an individual, and comparing the differences of phyla and genus levels of the flora.
Intestinal tissue and serum sIgA: keeping the colon tissue specimen at the temperature of 2-8 ℃, cutting the colon tissue specimen into fine blocks with the size of about 1 cubic millimeter, respectively adding a certain amount of PBS (PH7.4), and fully homogenizing the fine blocks by a homogenizer; centrifuging at 3000 rpm for 20min, and collecting supernatant; standing the blood sample at room temperature after collection, and naturally coagulating for 20 minutes; centrifuging at 3000 r/min for 20min, collecting supernatant, and determining intestinal tissue and serum sIgA content according to kit instructions.
Intestinal tissue TNF- α, IL-10: colon homogenate was prepared as above and assayed according to kit instructions.
Pathological observation of colon tissues: after the colon tissue is taken, immediately soaking the colon tissue in 4 percent paraformaldehyde, pouring out the fixing solution, trimming the colon tissue into 4cm flat blocks, and washing and soaking the colon tissue with running water; pouring out water, sucking the residual liquid by using filter paper, and dehydrating by using absolute ethyl alcohol; carrying out transparent treatment by using a dimethylbenzene solution, and then immersing the transparent treated product into paraffin for embedding; fixing the wax block on a slicing machine for slicing, wherein the slicing thickness is controlled to be 5-8 mu m; sections were stained with hematoxylin and eosin after deparaffinization (H & E staining) as per table 5; sealing the sheet with gum; and (4) observing by using an optical microscope.
TABLE 5H & E staining procedure
2.9 statistical methods One-Way ANOVA One-Way analysis of variance was performed on the experimental data using SPSS software, and LSD or Dunnett test compared the differences between groups.
3. Experimental results the experimental results are shown in tables 6 to 10 and figure 1.
As can be seen from Table 6, according to phyla level analysis, compared with the normal group, the relative abundance values of firmicutes, mollicutes and spirochetes in the high-fat diet model group are significantly reduced, while the relative abundance values of bacteroidetes, proteobacteria, verrucomicrobia and traceable bacteria are significantly increased, which indicates that the high-fat diet causes the flocculation of intestinal flora. Compared with a high-fat diet model group, the medium and low dose groups of the children Qixing tea can obviously improve the relative abundance values of the phylum mollicutes and the phylum spirochaetes and reduce the relative abundance values of the phylum bacteroidetes, the phylum verrucomicrobia and the phylum traceback. The high-dose group of the children Qixing tea can obviously reduce two pathogenic bacteria of verrucomicrobia and traceable bacteria, but has no regulating effect on dominant bacteria. The medium and low dose of the children Qixing tea has the function of obviously regulating intestinal flora, and the high dose of the children Qixing tea can inhibit part of pathogenic bacteria.
As can be seen from Table 7, according to the analysis of the genus levels, the relative abundance values of the genera Vibrio desulphuricus, Bacteroides, Prevotella and Sarcodon in the high-fat diet model group are obviously increased, while the relative abundance values of the genera Treponema twigs, Ruminococcus and Treponema are obviously decreased, which indicates that the high-fat diet causes the flocculation of the intestinal flora. Compared with a high-fat diet model group, the relative abundance values of vibrio desulfovis and prevotella can be reduced in each dose group of the children Qixing tea, and the relative abundance values of spirochete fibrillation bacteria, ruminococcus and bacteroides are improved. In addition, the middle and low dose groups of the pediatric Qixing tea can obviously increase the treponema and reduce the relative abundance value of the sarterium, while the high dose group has no regulating effect on the two bacteria. The medium and low dose of the children's Qixing tea has the function of obviously regulating intestinal flora, and the high dose of the children's Qixing tea can regulate most intestinal bacteria.
As can be seen from table 8, there was no significant difference in both spleen index and thymus index in the high fat diet model group compared to the normal group, but there was an upward trend. Compared with a high-fat diet model group, spleen indexes and thymus indexes of each dose group of the children's Qixing tea have no significant difference, but have a descending trend which is close to that of a normal group.
As can be seen from Table 9, compared with the normal group, the colonic tissue sIgA content of the high-fat diet model group is significantly reduced (P <0.05), which indicates that the colonic mucosal immune function of the high-fat diet model mouse is significantly reduced; compared with a high-fat diet model group, the sIgA content of each dose group of the pediatric Qixing tea is increased in different degrees and is in a dose-dependent relationship, wherein the sIgA content of colon tissues of high-fat diet mice can be obviously increased (P is less than 0.05) by the medium dose group and the high dose group, and is close to that of a normal group, and the experimental result shows that the pediatric Qixing tea can improve the colon mucosal immune function of the high-fat diet model mice.
As can be seen from Table 10, colon TNF-alpha and IL-10 were significantly increased in the high fat diet model group compared to the normal group (P < 0.05); compared with a high-fat diet model group, the children seven-star tea can reduce the levels of the two cytokines in each dose group, wherein the medium and high doses have statistical significance (P < 0.05).
Pathological tissue section observation results: the results of the pathological section are shown in FIG. 1.
As can be seen from FIG. 1, the colon tissue structure of Normal group (Normal) mice is clear and intact; the columnar epithelial cells and goblet cells in a single layer are arranged in order; epithelial cells are not denatured, necrotic or exfoliated; the intestinal mucosa has no pathological changes such as congestion, edema and inflammatory cell infiltration. Colon tissue structure is complete in a high fat diet Model group (Model); disorganization or loss of monolayer columnar epithelial cell alignment; goblet cell enlargement and disorganization; inflammatory cell infiltration and inflammatory tissue are observed among tissue cells. The colon tissue structure of each dosage group (EXQ 0.75.75 g/kg, EXQ 1.5.5 g/kg and EXQ 3g/kg) of the children's Qixing tea is complete, the number of monolayer columnar epithelial cells and goblet cells of high-fat diet groups is reduced, and the arrangement is tidy; the intestinal mucosa is substantially free of inflammatory cell infiltration.
TABLE 6 Regulation of intestinal flora levels in high fat diet model mice by pediatric Qixing tea
Note: in comparison with the normal group,#P<0.05; p compared to the high-fat diet model group<0.05
TABLE 7 Regulation of intestinal flora levels in high fat diet model mice by pediatric Qixing tea
Note: in comparison with the normal group,#P<0.05; p compared to the high-fat diet model group<0.05
TABLE 8 influence of pediatric Qixing tea on immune organ index of high fat diet model mouse
TABLE 9 Effect of pediatric Qixing tea on intestinal tissue sIgA of high fat diet model mouse
Note: in comparison with the normal group,#P<0.05; p compared to the high-fat diet model group<0.05
TABLE 10 influence of pediatric Qixing tea on TNF-alpha and IL-10 in intestinal tissues of high fat diet model mice
Note: compared to the normal group, # P < 0.05; p <0.05 compared to the high fat diet model group.
And (3) knotting: high fat diets can cause a disorganization of intestinal flora, a reduction in intestinal mucosal immune function, a disorganization of intestinal mucosal epithelial columnar cells and an increase in goblet cells, and cause inflammatory reactions.
The children's Qixing tea can effectively regulate intestinal flora and improve the intestinal mucosal immunity of high-fat diet mice; and can relieve pathological changes and inflammation of intestinal mucosa single-layer columnar epithelial cells and goblet cells caused by high fat diet.
The intestinal flora can regulate intestinal movement and secretion, decompose macromolecular compound polysaccharide in food, participate in digestion and absorption of nutrient substances, maintain the integrity of intestinal epithelial barrier, promote and maintain normal development and activity of immune system, and the like. The experimental result shows that the relative abundance value of part of dominant bacteria in the high-fat diet model group is obviously reduced, and the relative abundance value of pathogenic bacteria is obviously increased, which indicates that the high-fat diet can cause the flocculation of intestinal flora. The Qixing tea group for children can obviously improve the relative abundance value of the dominant bacteria and reduce the relative abundance value of part of pathogenic bacteria. The children's Qixing tea has obvious function of regulating intestinal flora.
Mucosal immunity is an important component of the human body's entire immune system, performs mainly local specific immune functions, and is widely distributed in the gastrointestinal tract, respiratory tract, urogenital tract mucosa, and lymphoid tissues at the exocrine glands. Mucosal immunity is the first line of defense against infection because mucosal surfaces are in direct contact with external pathogens and pathogenic microorganisms are inactivated by sIgA and the like before invading body tissues. SIgA is a marker for the development of a mucosal immune response, is secreted by plasma cells, inactivates pathogenic microorganisms by agglutination, and stimulates the intestinal tract to secrete mucus, thereby excluding bacteria and endotoxins. After local mucosa is stimulated, sIgA can be generated by itself without the participation of the central immune system to carry out immune response. Experimental results show that the content of SIgA in intestinal tissues of a high-fat diet model mouse is remarkably reduced, which indicates that the intestinal tissue mucosa immune function is low due to high-fat diet, the body resistance is reduced, the infantile Qixing tea can enhance the mucosa immune function of the intestinal tissues, and the intestinal tract is prevented from being damaged by high-fat diet.
TNF-alpha is mainly secreted by monocyte-macrophage, normal physiological level of TNF-alpha of human body can promote tissue repair, play an important role in killing and eliminating tumor cells and the like, and a large amount of released TNF-alpha can induce inflammatory reaction, even cause tissue damage due to over-immune response. IL-10 is mainly produced by immune cells such as T lymphocytes, B lymphocytes, monocyte-macrophages and the like, can inhibit macrophages from secreting proinflammatory factors such as IL-6, IL-12, TNF-alpha and the like, and TNF-alpha in intestinal tissues of a high fat diet model mouse is remarkably increased, while the content of TNF-alpha can be remarkably reduced by the children's Qixing tea, which shows that the children's Qixing tea has a regulating effect on intestinal mucosa immunity. The change trend of IL-10 in the model of high fat diet is consistent with the change trend of TNF-alpha and the function of inhibiting macrophage, and the immunoregulation function of the pediatric Qixing tea is also exactly explained.
Thymus and spleen are important immune organs, and the visceral index of the thymus and spleen depends on the proliferation degree of lymphocytes in the thymus and spleen, and can reflect the strength of immune functions of the body to a certain extent. The main functions of thymus are to produce T lymphocytes and to secrete thymosin, mainly involved in cellular immunity; spleen is rich in lymphocytes and macrophages, but the proportion of B lymphocytes is slightly larger, so that the spleen is a place where T cells and B cells are colonized and also a place for immune response, and is closely related to the immune function of the body. The thymus and spleen of the mouse of the high-fat diet model are slightly increased, which indicates that the high-fat diet may cause slight inflammatory reaction, and immune cells proliferate, thereby causing the index of immune organs to be increased.
However, after the children's Qixing tea is administrated, the indexes of two organs are in a descending trend, and are probably related to the reduction of inflammatory reaction of the children's Qixing tea, and the results of intestinal tract pathological histology and inflammatory factor detection also prove that the indexes are the same.
In conclusion, the research preliminarily proves that the pediatric seven-star tea has the function of protecting the intestinal mucosa immune barrier by measuring the intestinal sIgA of a high-fat diet model mouse.
Through observation of pathological tissues, the children's Qixing tea can improve the morphological change and distribution disorder of intestinal mucosa cells caused by high fat diet and improve inflammatory conditions, thereby protecting the intestinal mucosa barrier.
The technical features of the embodiments described above may be arbitrarily combined, and for the sake of brevity, all possible combinations of the technical features in the embodiments described above are not described, but should be considered as being within the scope of the present specification as long as there is no contradiction between the combinations of the technical features.
The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the present invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. Therefore, the protection scope of the present patent shall be subject to the appended claims.
Claims (3)
1. Application of a Chinese medicinal composition in preparing medicine for improving intestinal dysbacteriosis caused by high fat diet;
the traditional Chinese medicine composition is children's seven-star tea;
the dosage of the children's Qixing tea is that 0.75g of children's Qixing tea extract is added into each 1kg of water or 1.5g of children's Qixing tea extract is added into each 1kg of water;
the children Qixing tea extract is prepared from the following raw materials in parts by weight: 870-905 parts of coix seeds, 870-905 parts of rice buds, 435-460 parts of hawthorn, 655-685 parts of lophatherum gracile, 322-347 parts of uncaria, 100-124 parts of cicada slough and 100-124 parts of liquorice.
2. The application of claim 1, wherein the pediatric Qixing tea extract is prepared from the following raw materials in parts by weight: 893 parts of coix seeds, 893 parts of rice sprouts, 446 parts of hawthorn, 670 parts of lophatherum gracile, 335 parts of uncaria, 112 parts of cicada slough and 112 parts of liquorice.
3. The application of claim 2, wherein the extraction method of the pediatric Qixing tea extract comprises the following steps:
decocting Coicis semen and fructus oryzae Germinatus in water for 2 times, each for 2 hr, filtering decoction, mixing filtrates, concentrating to relative density of 1.08-1.12 at 55 deg.C, adding ethanol to make ethanol content reach 45%, standing, filtering, recovering ethanol from filtrate, concentrating into soft extract, decocting 5 Chinese medicinal materials including fructus crataegi, folium Bambusae, ramulus Uncariae cum uncis, periostracum Cicadae, and Glycyrrhrizae radix in water for 2 times, each for 2 hr, filtering decoction, mixing filtrates, concentrating the filtrate to appropriate amount, and mixing with the soft extract.
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