The Preparation Technology And Preliminary Pharmacodynamics Study Of Toad Skin Gels

Object Toad skin preparation possesses dual functions including antitumor and enhance immune function for cancer patients,and it has been widely confirmed in clinic.There are many reports about external application of toad skin in the folk and the effect is satisfying.In this project,the toad skin were transformed to gel preparation,the systematic study including preformulation study,formulation screening and preparation craft?quality standard?preliminary pharmacodynamics study.Method preformulation study:skin peeled off from toad first,and then took water content as index,applied freeze drying and routine dry technology to manufacture dry toad skin and fresh toad skin.Ultramicro grinding powder technology was employed to break the skin cell,so as to quicken the dissolution of active constituent.The differences between dry toad skin and fresh toad skin in effective component content and antitumor activity were conducted.The component content and pharmacologic action were compared systematically.Formulation screening and preparation craft:took appearance?viscosity?coat malleable?degree of uniformity as indicator to preliminary screen the dosage of gel forming polymer materials such as carbopol?hydroxypropyl methyl cellulose?polyving akohol and sodium carboxymethylcellulose.In the same time,took in vitro skin through quantity as indicator and used single factor and orthogonal test to screen the type and dosage of best transdermal enhancer and ensure the last recipe.Screened the adding orders of excipients in recipe and confirmed the best preparation craft.Quality standard:three batches of fresh toad skin samples of gels were prepared and TLC qualitative identification of cinobufagin and bufogenins were conducted.In content determination test of preparation,selected high performance liquid chromatography(HPLC)to detect the cinobufagin and bufogenins content of toad skin gel.In vitro skin through quantity test investigated the accumulated through transit dose of three batches samples.Preliminary stability study:the influence factors for three batches of fresh samples of toad skin gels were investigated.Accelerated stability were observed as well.Preliminary pharmacodynamics study:took anti-tumor rate as indicator and as the same time investigated spleen index and thymus index systematically at the same time.Result the result of preformulation study demonstrated that toad skin dried continually for 18h in freezer dryer(T:-50±5?,P:20±5Pa)and the water content could meet requirement.After dried,the fresh toad skin and dry toad skin were ultramicro grinded continually for 5min and cell broken rate is up to 98.43%and 95.22%.The result of content determination showed that the total content of cinobufagin and bufogenins in fresh toad skin is remarkably higher than dry toad skin.In the pharmacodynamics test,the tumor control rate for S180 mice model of fresh toad skin was higher than dry toad skin.So in this project,the fresh toad skin was used as main drug.The result of formulation screening and preparation craft showed that the last mass ratio in recipe was as below:main drug10%,molding matrix HPMC K4M 2%,HPMC K100M 0.6%,transdermal enhancer menthol crystal 1%,azone 3%,propylene glycol 1%,humectant glycerol 5%,preservative methylparaben 0.1%.Though this study,the toad skin gel possessed good appearance?coat malleable and transmittance.The result of quality standard manifested that the samples was semi-solid preparations and the specificity in TLC qualitative identification was strong.High performance liquid chromatography(HPLC)was used to detect the cinobufagin and bufogenins content of toad skingel.Chromatographic condition:chromatographic column:kromasilC18(4.6mm×250mm,5?m)(Sweden);flowing phase:ACN(A)-water(B)(48:52);flow rate:1 mL·min-1;column temperature 30?;UV detection wavelength:296nm.Under this condition of research,the total cinobufagin and bufogenins content was not lower than 380?g/g of fresh toad skin gels.The determination method was simple and easy,precision and repeatability was good.The result of preliminary stability study showed that high temperature factor would affect the preparation quality and it should be stored in low-temperature environment and sealed.The preparation was placed in cold condition for 3 months,and there were no distinct changes of detection index compared with 0 month.The result of preliminary pharmacodynamics study:effects of fresh toad skin gel on S180 ascites solid tumor mice model were studied.By compare anti-tumor rate after drug administration of each group,the result showed that anti-tumor rate presented positive correlations with dosage of drug delivery;and compared with model group,the high dose and middle dose group possessed significant difference.Conclusion This paper finished the entire contents of fresh toad skin gel preparation craft study?quality standard study?preliminary stability study and preliminary pharmacodynamics study,and supplied reliable quality control methods.It established firm theoretical basis for the development of action mechanism and industrialization study for the future.