| Objective:To establish the rapid HPLC methods for the determination of the components of MFXD on different compatibility,to provide support for transdermal research of the effective chemical composition;by investigating the basic physical and chemical characters of compatibility of different groups and extractive,to provide basis for further preparation development of transdermal drug delivery;the use of modified Franz diffusion cell methods in vitro,to investigate the effects of compatibility of different groups on percutaneous penetration,explore the reasonability of compatibility on traditional Chinese medicine for external use.Methods:(1)Using the method of HPLC,The analysis methods of the alkaloids(ephedrine,pseudoephedrine),aconite alkaloids(benzoylmesaconine,benzoylhypaconine and benzoylaconine),asarum volatile oil(methyl eugenol,asarinin)were established.The reliability of the analytical method is determined by examining the precision,stability,repeatability and sample recovery.(2)The equilibrium dissolution of ephedra aconite extract,aconite extract and its components in water was determined by precipitation method and index method.Determining the apparent oil/water partition coefficients in the n-octanol/water system by shaking flask method.(3)Franz diffusion cells method was used to determine the permeation rate and 12h accumulation of ephedrine alkaloids,benzoylmesaconine,benzoylhypaconine and benzoylaconine,methyl eugenol and asarinin in the different compatibility of MFXD in vitro.The results were analyzed by SPSS 17.0 software,and the results showed that the compatibility of dierent groups in MFXD had an affect on transdermal permeation properties.Result:(1)To establish HPLC chromatographic conditions of ephedrine and pseudoephedrine were:the mobile phase consisted of acetonitrile-0.1%phosphoric acid solution(2:98,100ml each was added with 0.04ml of tri-citrate);the chromatographic column was Diamonsil C18(5?m,250 mm×4.6 mm);detection wavelength of 190nm;flow rate of 1ml/min;column temperature of 30?;injection volume of 10?l.To establish HPLC chromatographic conditions of the alkaloids of aconite were:the mobile phase consisted of B acetonitrile-tetrahydrofuran(25:15)-A 0.1 mol/L ammonium acetate solution(0.5 ml of glacial acetic acid was added to each 1000ml).The chromatographic column was Diamonsil C18(5?m,250 mm x 4.6 mm),the optimal gradient elution mode was as follows:0-48min,(15%-24%B);detection wavelength of 245nm,flow rate of 1ml/min,column temperature:30?;injection volume of 10?l.To establish HPLC chromatographic conditions of methyl eugenol and asarinin were:the mobile phase was acetonitrile-water(60:40).The column was Diamonsil C18(5?m,250mm×4.6mm);The column temperature was 40?,and the detection wavelength was 287nm.The injection volume was 10?l.(2)The equilibrium solubility of ephedrine in water at 37? was 27-5.19?g/ml,the equilibrium solubility of pseudoephedrine in water was 132.31?g/ml,ephedrine and pseudoephedrine had optimum solubility in buffered solution of 7.4,which were 344.27?g/ml and 162.22?g/ml.The n-octanol/water partition coefficients of ephedrine and pseudoephedrine in the Mahuang sample were 53.85(logP=1.73)and 19.38(logP=-0.99),respectively.The n-octanol/water partition coefficients were 0.101(logP=-0.99),0.076(logP=-1.12)in the Mafu sample,respectively.The n-octanol/water partition coefficients were 2.91(logP=0.46)and 3.43(logP=0.54)in the Maxi sample,respectively,and the n-octanol/water partition coefficients in the whole formula sample were 5.59(logP=0.75)and 5.98(logP=0.78),respectively.The equilibrium solubility of benzoylmesaconine,benzoylhypaconine and benzoylaconine in water were 11.0?g/ml,2.07?.g/ml and 4.36?g/ml.The solubility of benzoylmesaconine and benzoylhypaconine at 37? was higher in buffered solution of 7.4.The n-octanol/water partition coefficients of benzoyl aconitine,benzoyl aconitine and benzoyl aconitine in the Fuzi sample were 0.022(logP?-1.65),0.091(logP=-1.04),0.094(logP=-1.03),respectively.The n-octanol/water partition coefficients were 0.021(logP=-1.68),0.09(logP=-1.05),0.094(logP=-1.03)in the Mafu sample,respectively.The n-octanol/water partition coefficients were 0.037(logP=-1.43),0.095(logP=-1.02),0.097(logP=-1.01)in the Fuxi sample,respectively.The n-octanol/water partition coefficients were 0.024(logP=-1.62),0.09(logP=-1.05),0.094(logP=-1.03)in the whole formula sample,respectively.The n-octanol/water partition coefficients of methyl eugenol and asarum were 0.94(logP=-0.027),1.02(logP=0.009)in the Xixin sample,respectively.The n-octanol/water partition coefficients were 0.027(logP=-1.57)and 0.172(logP=-0.77)in the Maxi sample,respectively.The n-octanol/water partition coefficients were 0.116(logP=-0.94),0.338(logP=-0.41)in the Fuxi sample,respectively.The n-octanol/water partition coefficients in the whole formula sample were 0.118(logP=-0.93)and 0.383(logP=-0.42),respectively.(3)The effects of different compatibility in ephedrine alkaloids of MFXD on transdermal permeation were as follows:transdermal permeation rate(68.864?g-cm-2·h-1?28.088?g.cm-2·h-1)of ephedrine alkaloids in the whole group and 12h accumulation(820.8?g-cm-2,335.4?g·cm-2)were greater than other compatibility group,the difference was extremely significant.The transdermal permeation rate and the 12h accumulation of the compatibility group were in the order of whole fumula>Maxi>Mahuang>Mafu.The effects of benzoylmesaconine,benzoylhypaconine and benzoylaconine of different compatibility groups on transdermal permeation were as follows:the permeation rate of benzoylmesaconin and the cumulative infiltration of 12 h(7.122?g·cm-2·h-1,88.8?g-cm-2)in Fuxi group were higher than Fuzi group,Mafu group,whole fumula group.The transdermal rate of benzoylhypaconine was higher than Fuzi group,Mafu group,whole group,the 12h cumulative infiltration of benzoylhypaconin in Fuxi group was higher than Mafu group,whole fumula group.The transdermal rate of benzoylaconine in Fuxi group was higher than the single group,Mafu group,whole fumula group,The transdermal rate of whole fumula group,Maxi group and the cumulative permeation rate of 12h were lower than Fuxi group,aconite single group.Mafu group compared with whole fumula group,there was no significant difference in transdermal rate and 12 h cumulative permeation of benzoylmesaconine,benzoylhypaconine and benzoylaconine.The effects of methyl eugenol and asarum of different compatibility groups on transdermal permeation were as follows:the transdermal rate of methyl eugenol and the cumulative permeation of 12h in asarum single group were 76.391?g-cm-2·h-1 and 788.6?g·cm-2,which were significantly higher than the other groups.There was no significant difference in transdermal rate and 12 h cumulative permeation of methyl eugenol between whole fumula group and Fuxi group.There was no significant difference in transdermal rate and 12 h cumulative permeation rate between whole fumula group and the Maxi group,Fuxi group and asarum single group.Conclusion:In this paper,the analysis methods of various components in MFXD of different compatibility were established.The method is simple,accurate and reliable.The water solubility of ephedrine,pseudoephedrine and aconite alkaloids are better.The results of oil and water partition coefficient determination show that,ephedrine alkaloids,aconite alkaloids,methyl eugenol and asarinin above the ingredients have a high biofilm permeability.The effects of compatibility on the percutaneous permeation of various active ingredients in MFXD were different.Oil and water partition coefficient values close to or greater than zero,the drug composition showed fat-soluble,percutaneous absorption increased.The physical and chemical properties of different compatibility groups showed a certain correlation with percutaneous absorption,changes of oil-water partition coefficient in different compatible groups,which can promote the transdermal absorption of some components and reduce the transdermal rate and cumulative permeation rate of some toxic components.It has certain attenuation and synergistic effect,which proves the scientific compatibility of external medicine. |
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