| Objective:By researching the effect of compatibility on the effective components and visceral hypersensitivity animal model of IBS with deficiency of liver and spleen,to explore the role of AMR-PRA in Tongxie Yaofang.It lays a foundation for the study of the mechanism of Tongxie Yaofang and the theory of compatibility of traditional Chinese medicine.Methods:(1)The Thermo Fisher C18 column was used to screen the best detection conditions and to establish a HPLC-PDA method for simultaneous determination of five effective components(atractylenolide Ⅰ,Ⅱ,Ⅲ,paeoniflorin and paeoniflorin)in AMR,PRA,AMRPRA herbal pairs and Tongxie Yaofang.(2)To reproduce the model of IBS visceral hypersensitivity by stimulating the rectal dilatation of Wister rats and taking the mother-infant transient separation method.The model rats were randomly divided into 7 groups:normal control group,model group,prescription group,AMR group,PRA group,AMR-PRA group,SR-CRP group.By inspecting the changes of general state,weight,urine D-xylose excretion rate,abdominal withdrawal reflex test,black and white box test of the model rats,to analyze the effects on general state,excretion function,visceral sensitivity and animal behavior.To investigate the effect of compatibility on IBS of liver and spleen deficiency.Result:(1)The HPLC-PDA method:the chromatographic column was Thermo Fisher C18 column and five active components was performed with acetonitrile(A)-water(B)as the mobile phase gradient elution at flow rate of 1 mL·min-1;The column temperature was 30℃;The detection wavelength was set at 222 nm(atractylenolide Ⅰ,Ⅲ),278 nm(atractylenolide Ⅱ)and 232 nm(albiflorin and paeoniflorin).Average sample recovery rates of atractylenolide Ⅰ,Ⅱ,Ⅲ,albiflorin and paeoniflorin respectively were 97.15%(RSD=2.1%),98.36%(RSD=2.4%),99.87%(RSD=1.9%),99.26%(RSD=1.1%),99.33%(RSD=0.8%);The results of precision test,repeatability test and stability test showed that the RSD of the peak area of the five active components was less than 2.0.The results showed that the content of atractylenolide Ⅰ,Ⅱ,Ⅲ were increased after compatibility(p<0.05),however,paeoniflorin and paeoniflorin were reduced(p<0.05).Compared AMRPRA group with prescription group,the contents of,except atractylenolide I,four compounds in prescription were higher than in AMR-PRA,but there was little difference between them;The contents of paeoniflorin and paeoniflorin were lower than those of in prescription(P<0.05).(2)Compared with the normal control group,the overall evaluation results of the model group showed that the rats were small in size,languid and motionless,lost color of hair,loose stool,decreased excretion rate of D-xylose,and caused lower threshold arch volume of abdominal elevation and back.Behavioral experiments showed a decrease in the length of stay in the light box and the number of shuttles between the two boxes.After 2 weeks of administration,both the prescription group and the AMR-PRA group could significantly activate the rats,increase their weight,increase the excretion rate of D-xylose and increase volume threshold of causing abdominal elevation and back arched.The behavioral experiment showed that the residence time in the light box and the number of shuttle between the two boxes increased significantly compared with the model group(p<0.05),the other indexes except the food quantity in the drug treatment group and the normal control group were significantly different(p<0.05);The AMR group,the PRA group and the SR-CRP group,compared with the normal control group,were significantly different(p<0.05),and compared with the model group,the results of AWR test and animal behavior experiment was significantly different(p<0.05).Conclusion:In this paper,a method for simultaneous determination of five active components in single drug,herbal pair and prescription was established,which was simple,accurate,reproducible and reliable.After compatibility,the atractylenolide components were significantly increased;Prescription group and the AMR-PRA group could regulate the general state,reduce the visceral hypersensitivity,enhance the function of absorption and excretion,and regulate mood to produce good therapeutic effect on IBS model.Compatibility can increase the content of active ingredients,thereby enhancing the efficacy of Tongxie Yaofang.AMR-PRA herbal pairs was the main support of its therapeutic effect. |
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