| OBJECTIVE:under the guidance of the theory of external treatment of Mongolian medicine,a new compound preparation of pediatric antipyretic Mongolian medicine was developed.In this study,the dosage form of Mongolian Medicine Sanchen pill was improved,and its pharmacodynamics,mechanism of action,safety,quality standard and stability were systematically evaluated.METHODS:1 The extraction technology of Sanchen Xiaoer antipyretic paste was studied:The content of Hydroxysafflor pigment a and the amount of extract were used as indexes,and the extraction technology of safflower active ingredient was optimized by orthogonal test.2 study on the preparation technology of Sanchen antipyretic patch for children:the Matrix prescription of Sanchen antipyretic patch for children was selected by single factor experiment and orthogonal test with comprehensive sensory evaluation and adhesive force as evaluation indexes.(3)In vitro study on the parity of Sanchen children antipyretic paste,the release rate experiment:The cumulative transmittance of Hydroxysafflor a(Qn%)was used as the index to evaluate the drug release of the preparation;In Vitro transdermal test:the optimal proportion of Transdermal enhancers was selected by in vitro transdermal test with 24h total transdermal quantity(Qn)of hydroxy safflor a per unit area.4 preliminary experiment:SD Rats were randomly divided into 4 groups:Normal Group,Model Group,Sanchen Pill Group,Sanchen children antipyretic paste group,etc.,all the rats except the normal group were injected with dry yeast subcutaneously to induce fever.The rats were given the corresponding drugs at 4H and 7H after the MODEL was established.The Rats'body temperature was measured once every 1H for 10 times.The maximum elevation of body temperature(?T)and temperature response index(TRI6)were calculated as indexes to compare the therapeutic effects of oral and transdermal administration of Sanchen pill.Antipyretic effect of Sanchen Xiaoertuiretie on dry yeast-induced fever rats:SD rats were randomly divided into normal group,model group,sanchen children antipyretic paste high,medium and low dose(pharmacognosy 1:6,0.8,0.4 g/kg)group,Aspirin(100 mg/kg)group,blank group(Matrix Paste)Group 7 groups.The model of Dry yeast-induced fever was established,and the corresponding drugs were given to each group at 4H and 7H.Take your temperature every hour,10 times.Plot the average temperature rise curve,calculate ?T and TRI6,and evaluate the dose-effect relationship.5 The antipyretic mechanism of Sanchen Xiaoer antipyretic tie on dry yeast-induced fever rats,the effect of Sanchen Xiaoer antipyretic tie tie on the fever-related cytokines of dry yeast-induced fever rats:the serum levels of TNF-?,IL-6 and CRP of rats in each group were detected by Elisa,to evaluate the effect of Sanchen Xiaoer antipyretic paste on the expression of fever-related genes in rats induced by dry yeast:the Mrna expression levels of TNF-? and COX2 in hypothalamus of rats were detected by QRT-PCR,to evaluate the effect of Sanchen Xiaoer tuiretie on endogenous metabolites in rats with fever induced by dry yeast:the data of each group were compared and analyzed by using the method of UPLC-QE-MS and Multivariate statistics,to search for the potential biomarkers related to fever in plasma,explore the regulatory effects of Sanchen Xiaoer antipyretic patch on these endogenous components,and infer its possible metabolic pathways.6 Safety Evaluation Study of Sanchen antipyretic adhesive for children:to evaluate the irritation to intact skin and damaged skin of rats by the index of Erythema and Edema,and to evaluate the Anaphylaxis by the index of Erythema and Edema and systemic anaphylaxis.7 evaluation on quality standard of Sanchen Xiaoer antipyretic patch:the artificial bezoar and Tenjiku bezoar in the preparation were identified by Thin layer chromatography;According to the stability method of patch in Chinese Pharmacopoeia,the content,weight difference and adhesive force of the paste were examined,and the content of Hsya,cholic acid and deoxycholic acid in the patch were determined by HPLC,and the quality standard was established.8 stability of Sanchen Xiaoer Tuifetie:The accelerated stability and long-term stability of Sanchen Xiaoer Tuifetie were investigated according to the method of Chinese PHARMACOPOEIA.Results:1 the best extraction technology of Carthamus Tinctorius L.was 15 times 60%ethanol as solvent,extracting 3 times,soaking 1 hour each time.2 N46:GLYCOLATE:TARTARIC ACID:glycerol-7:0.15:0.15:30 was the most suitable matrix for Sanchen antipyretic patch for children.3 the release rate of HSYA was 50%in Q12%and 90%in Q24%.The Q24 of HSYA was 65.37 g/cm2 when the dosage of azone was 2%.4 PRE-EXPERIMENT OF PHARMACODYNAMICS EVALUATION:At the 4th hour,compared with the normal group,the body temperature of rats in each group increased,and there was a significant difference(p<0.01),which indicated that the Model Group was successful;There was a significant difference(p<0.01)between Sanchen Xiaoer tuiretie group and Sanchen pill group(p<0.01),the antipyretic effect of Sanchen Xiaoer Tuiretie group appeared 2 hours earlier than that of Sanchen pill.The antipyretic effect of Sanchen Xiaoertuiretie on dry yeast-induced fever rats:the model of Dry yeast-induced fever was completed after 4 hours,compared with the model group,the AT and TRI6 of the rats in each group were decreased,and there was a significant difference(p<0.01).Compared with aspirin group,the level of ?T in Sanchen group was lower than that in aspirin group,and TRI6 in Sanchen group was higher than that in Aspirin Group(p<0.01).There was no difference in ?Tbetween Aspirin Group and Sanchen group,but TRI6 OF ASPIRIN group and Sanchen group was lower than that of Sanchen group(p<0.01).5 The effects of Sanchen Xiaoertuiretie on the levels of cytokines related to fever in rats induced by dry yeast:compared with the normal group,the levels of IL-6,TNF-? and CRP in serum of rats in the model group were increased,and there were significant differences(p<0.01),compared with the model group,the levels of Serum IL-6,TNF-? and CRP in the other groups were significantly lower than those in the control group(p<0.01),there was no difference in IL-6 between the middle-dose group and the other groups(p<0.01;p<0.05),but the levels of L-6,TNF-? and CRP in the middle-dose group were higher than those in the Aspirin Group(p<0.01;p<0.05),the levels of L-6,TNF-?and CRP in the other groups were lower than those in the blank Matrix Patch Group(p<0.01;p<0.05).6 The effect of Sanchen Xiaoertuiretie on the expression of fever-related genes in rats with fever induced by dry yeast:compared with normal group,the expression of TNF-? and COX-2 genes in hypothalamus of model group were up-regulated,and there was significant difference(p<0.01)Compared with the model group,the expression of TNF-? and COX-2 genes in hypothalamus of the Sanchen children with antipyretic plaster group were down-regulated,and there was significant difference(p<0.01).Effects of Sanchen Xiaoer tuiretie on endogenous metabolites in rats with fever caused by dry yeast:7 potential plasma biomarkers have been identified in the plasma of rats in model group,the contents of Ketoleucine,SM(D17:1/24:1(15Z),SM(D18:1/24:1(15Z)),SM(D18:1/18:1(9Z)),PC(16:0/14:0),PC(18:0/15:0),PC(16:0/16:0)were all increased(p<0.01 orp<0.05)The content of Taurine and isocitrate in plasma was also increased(p<0.01 orp<0.05).Metabolic pathway analysis revealed Taurine and subtaurine metabolism,glyoxylic acid and dibasic acid metabolism,tricarboxylic acid cycle,valine,leucine,L-Isoleucine metabolism,primary bile acid biosynthesis,linolenic acid metabolism,niacin and nicotinamide metabolism,sphingolipids metabolism and other metabolic pathways are related to the antipyretic effect of Sanchen Xiaoer antipyretic paste in treating fever rats induced by dry yeast.6 Sanchen antipyretic adhesive for children has no irritation to intact skin and damaged skin,and has no hypersensitivity to skin.7 artificial bezoar and Tenjiku yellow showed the same color spots in the chromatogram corresponding to the Chromatogram of the control material and the control substance.The results showed that the average paste content of the preparation was 10.38 g,the initial viscosity was above 10 steel ball,the holding viscosity was above 10 s,and the weight difference and the formability were all in accordance with the regulations.The contents of Hsya,cholic acid and deoxycholic acid were 0.03%,0.26%and 0.27%.8 respectively.The results of stability test showed that the quality of Sanchen antipyretic paste was stable within 6 months.Conclusion:The extraction method of Carthamus Tinctorius is stable,repeatable,accurate,simple and feasible,and can be used for the extraction of Sanchen Xiaoer antipyretic paste.The sanchen infantile antipyretic paste prepared with the selected formula has good properties,moderate viscoelasticity and good release of active components.Sanchen Xiaoertuiretie has a stable antipyretic effect on rats with fever caused by dry yeast,and its antipyretic effect is better than Sanchen pill.The mechanism may be through reducing inflammation,reducing the production or accelerated degradation of serum cytokines related to fever,and down-regulating the expression of fever-related genes in the central nervous system.From the enzyme inhibition,fat metabolism,Amino Acids and energy metabolism and other multi-level coordination,multi-target cross-related.Sanchen Xiaoer antipyretic paste has no irritability and hypersensitivity to the skin,and can be used for many times and for a long time.According to the result of quality evaluation,a draft quality standard is drawn up.The content limit of paste is tentatively set at 9.8 g for each piece,the lower limit of initial viscosity is tentatively set at 9 steel balls,and the lower limit of holding viscosity is tentatively set at 9s,the content of HSYA is not less than 0.25 mg per 1g paste,and the content of cholic acid and deoxycholic acid is not less than 2.0 mg per 1g paste.The weight difference and endotype are in accordance with the relevant regulations of Chinese Pharmacopoeia.The method of identification and content determination of Sanchen antipyretic paste for children is simple,reliable,specific and without negative interference.To sum up,the sanchen child antipyretic paste developed in this paper is safe,effective,easy to use,stable in quality and worth popularizing. |
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