The Quality, Fingerprint And Pharmacokinetics Research Of Traditional Chinese Medicine Toad Skin

Objective:To set up the standard, identify the active ingredients, clarify thepharmacokinetics as well as provide the basic data for further application oftraditional Chinese medicine toad skin.Methods:（1）The experiments of physicochemical property and thin-layerchromatography were used to identify the medicinal materials of toad skin, themoisture content, ash content and the amount of cinobufagin, resibufogeninwere investigated to established the standards;（2）In the fingerprint research,the technique of gradient elution was adopted in high performance liquidchromatography（HPLC）, in order to determine the commen peak,10samplesform different regions were analysed comparing with9given bufadienolides, the results of relative retention time（RRT）and the relative peak area（RPA）were evaluated with the medicine chromatographic fingerprint similarityevaluation software;（3）By gastric perfusion, the cinobufagin, resibufogenin,bufalin and extraction of toad skin were dosed to SD rats, gathered the plasmasamples and detected the concentration by HPLC, to process the results withpharmacokinetic software, the concentration-time curve and relevantparameters were acquired.Results:（1） With reference to the Chinese pharmacopoeia standards andrequirements, moisture content should not over12.0%, total ash shoud notmore than14.5%, acidinsoluble ash shoud not more than5.0%and the contentof cinobufagin should not less than0.02%, resibufogenin should not less than0.01%respectively;（2） The fingerprint of toad skin was established,29commen peaks contained9known bufadienolides including gamabufotalin（GBT）, arenobufagin （ABG）, telocinobufagin （TCB）,desacetylcinobufotalin（DCB）, bufotalin（BFT）, cinobufotalin（CBG）, bufalin（BFL）, cinobufagin（CBG）, resibufogenin（RBG） were screened out, itrevealed that the major ingredients of toad skin from different regions weresimilar to each other, but there were great differences in the contents, forexample, the total content of9ingredients in toad skin from Xi’an was themost（19.0μg/g）, whereas the total content in toad skin from Ji’nan was thelowest（5.2μg/g）, among10samples, the content of CBG is the highest（2.13μg/g）, the others were BFL（1.45μg/g）, RBG（1.18μg/g）, BFT（0.96μg/g）, GBT（0.82μg/g）, CBG（0.82μg/g）, DCB（0.62μg/g）,ABG（0.43μg/g）and TCB（0.26μg/g）;（3）After cinobufagin and resibufogenin oral administration, the prototype were not detected, they all exist in forms ofmetabolic products. The pharmacokinetic parameters of BFL after singlecomponent oral administration were: AUC(0-t)=975.89mg/L·min,AUC(0-∞)=1067.69mg/L·min, MRT(0-t)=234.21min, MRT(0-∞)=608.08min,t1/2z=375.76min, Tmax=22.50min, CLz/F=0.008L/min/kg, Vz/F=3.51L/kg,Cmax=4.66mg/L; and the pharmacokinetic parameters of BFL aftermulticomponent oral administration were: AUC(0-t)=476.87mg/L·min,AUC(0-∞)=518.43mg/L·min, MRT(0-t)=160.69min, MRT(0-∞)=221.46min,t1/2z=146.76min, Tmax=26.25min, CLz/F=0.023L/min/kg, Vz/F=4.56L/kg,Cmax=3.00mg/L. Compared the two groups, we found that the P-value ofAUC(0-t), AUC(0-∞), MRT(0-t), MRT(0-∞), t1/2z, Tmax, CLz/F, Vz/Fand Cmaxwere0.0279,0.0021,0.0045,0.0201,0.0246,0.5536,0.0029,0.3251and0.1894respectively, which indicated that there were no difference in the Cmax, Tmaxand Vz/F, however, the AUC(0-t), AUC(0-∞), MRT(0-t), MRT(0-∞), t1/2zand CLz/Fall displayed significant difference.Conclusion:There were significantly regional characteristics of toad skin, greatdifferences existed in the bufadienolides contents of toad skin from differentregions; Compound medicine of toad skin could speed up the metabolic rate,shorten the half-time period and MRT, reduce the AUC, affect the absorption,distribution, metabolism and excretion of BFL.