| 1. Development of Fexofenadine Hydroehloride tablets analysis methodA RP-HPLC method has been developed to determine Fexofenadine Hydrochloride and its related substances. The chromatographic system consisted of a Hitachi model L-7110 pump and six port switching injector equipped with a 20μL sample loop. The chromatographic separation was performed on the colume of Diamonsil C18 (200mm×4.6mm, 5μm) with UV detection at 210nm, and the mobile phase was water (consisted of 0.4％phosphate, 0.7％triethylamine)-acetonitrile-methanol in the ratio 50:40:10(v/v/v, adjusted pH=3.0 with phosphate), flow rate at 1.0 mL·min-1. The content of Fexofenadine Hydrochloride was calculated by internal standard method. There is a good linear relationship for Fexofenadine Hydrochloride within the range of 1.0～20.0mg·L-1(r=0.9998). The average recovery was 100.5％and intra-run precision of the method was within 2％. The LOD of Fexofenadine Hydrochloride was 5.0μg·L-1. The labeled quantity percentage content of 3 batches of Fexofenadine Hydrochloride tablets were 93.0％, 92.9％and 93.8％, respectively. The method of main component self-compare with no calibration factor has been used to determine the related substances of Fexofenadine Hydrochloride tablets. The result indicated that the maximal impurity was not more than 0.15％, the sum of impurities were not more than 0.43％. The method is simple, rapid, accurate and is suitable for the determination of Fexofenadine Hydrochloride and its related substances and can be used to quality control in Fexofenadine Hydrochloride tablets.2. The pharmacokinetics of Fexofenadine Hydrochloride in Wistar ratA HPLC-fluorescence method has been established for determination of Fexofenadine Hydrochioride concentration in Wistar rat' plasma and tissues. A Diamonsil C18(200×4.6mm, 5μm)column with a mobile phase composed of water (consisted of 0.4％phosphate, 0.7％triethylamine)-acetonitrile-methanol in the ratio 55:35:10(v/v/v, adjusted pH=3.0 with phosphate)was used in separation. The detection wavelength wasλex=230nm andλem=280nm and the flow rate was 1.0mL·min-1. The biological samples were analysed after deprotaining by acetonitrile, and the concentration of Fexofenadine Hydrochloride was calculated by calibration curve accompanied. The calibration curve of Fexofenadine Hydrochloride in plasma have a good linear between 0.05～4.0 mg·L-1(r=0.9993). The intra-day precision were13％,8.5％and 4.8％, respectively. The inter-day precision were 15％,14％,9.2％,and the accuracy were 99.0％,96.2％and 98.7％, respectively. The recovery of extraction was more than 60％. The calibration curve of Fexofenadine Hydrochloride in liver was linear between 0.1～10.0μg·g-1(r=0.9951). The intra-day precision of QC samples was 9.2％, 11％and 7.3％, respectively. The inter-day precision were 14％,15％and 11％, and the accurate 97.3％,101.8％and 108.3％, respectively. The recovery of extraction was more than 50％.Dose of 15,30,60 mg·kg-1were administrated to 3-dose group male rats by i.g. There had a maximal plasma concentration between 1.67～2.33h, and the ratio between Cmax,AUC0-1 and dose(P＜0.05) had a linear correlation. The ratio of AUC0-t and dose had insignificant difference after analysis of variance (P＞0.05), which showed that the process of Fexofenadine Hydrochloride in Wistar rats conformed with linear pharmacokinetics.Fexofenadine Hydrochloride distributed rapidly in tissues after absorbing into plasma. The concentration in stomach and intestine were highest, next liver, kidney, spleen, lung, heart and the concentration in brain was the lowest, which was proportioned with dose, and that in stomach and intestine was more 30 times than that in plasma. The t1/2 in stomach, intestine, lung, heart and spleen were not longer than that in plasma, which showed that Fexofenadine Hydrochloride was not accumulated in tissues, but the t1/2 in liver was a little longer than that in plasma. The ratio of AUC0-t in tissues and AUC0-t in plasma indicated that distribution in stomach and intestine were the widest and in brain least. Fexofenadine Hydrochloride was uneasy to pass the blood-brain barrier. Dose of 30 mg·kg-1were administrated to female rats by i.g. Tmax was 2.33±0.58h, Cmax was0.95±0.13mg·L-1, AUC0-t wasl 1.92±2.62mg·h·L-1 and t1/2 was 14.82±2.96h. Vd/F and Cl/F in female rats were 9.05±2.05L and 0.46±0.12L·h-1, respectively, which were 30％lower than that in plasma. In relation to Cmax and AUC0-t in male and female plasma, there was no significant different observed by analysis of variance. Fexofenadine Hydrochloride widly distribute in female rats after administrating. Cmax and AUC0-t in male and female tissues had insignificant different except Cmax in kidney by analysis of variance.Plasma protein binding of Fexofenadine Hydrochloride in Wisatar rat was measured after laying 72h when a equilibrium has achieved between in and out dialysis membrane laying at 4℃. At the rat plasma concentrations of 0.5,1,2μg·mL-1, plasma protein binding of Fexofenadine Hydrochloride were more than 50％.3. The pharmacokinetics and bioavailability of Fexofenadine Hydrochloride in humanA HPLC-fluorescence method has been established for determination Fexofenadine Hydrochloride concentration in human plasma. Diamonsil C18(200mm×4.6mm, 5μm)cohunn with mobile phase composed of water (consisted of 0.4％phospbate, 0.7％triethylamine)acetonitrile-methanol in the ratio 55:35:10(v/v/v, adjusted pH=3.0 with phosphate) was used in separation. Fluorescence detection with the excitation wavelength at 230nm and emission wavelength at 280nm was used to determine. The flow rate was 1.0mL·min-1 and column temperature was 35℃. Liquid-liquid extraction was used to pretreat plasma sample. Internal standard tehnisartan and plasma sample were added into the turbe. The mixture was vortexed, then ethylether-ethyl acetate(1:4,v/v) of 1.6mL was add into the mixture, vortexed for 5 rain and centrifuged at 10000 r·min-1 for 5 min. Supematanty transferred into another turbe was evaporated to dryness at 40℃water bath under stream of nitrogen. The residue was reconsitituted with 50μL methanol. After vortexing 1 min supernatant of 20μL was injected. The typical chromatogram of Fexofenadine Hydrochloride showed that no endogenous interference was observed at the peak of the analytes. This demonstrated that the method was specific to determine Fexofenadine Hydrochloride in human plasma. The concentration of Fexofenadine Hydrochloride was calculated by calibration curve accompanied. The calibration curve in plasma was linear between 5.0～500.0μg·L-1(r=0.9996). Accuracy and precision of method was evaluated by assaying 6 QC samples and on 3 successive days, the results were as follows: The intra-day precision of QC samples were 4.4, 4.6％and 4.0％, respectively; the inter-day precision were 9.2％, 8.1％and 7.7％, respectively; and the accurate was 101.6％, 103.7％and 104.5％, respectively, The recovery of extraction of Fexofenadine Hydrochloride and internal standard telmisartan was more than 80％.A single oral dose of 120mg Fexofenadine Hydrochloride test and reference preparations were administrated to 20 healthy volunteers in a randomized cross-over design to study pharmacokinetics and relative bioavailability of Fexofenadine Hydrochioride capsule in healthy male volunteers. Plasma concentration of Fexofenadine Hydrochloride was determined by HPLC-fluorescence method, and pharmacokinetics and relative bioavailability were evaluated by DAS 1.0. Main pharmacokinetic parameters for single dose were as follows: Tmax were 2.55±0.72 and 2.60±0.84h ; Cmax were370.8±84.7 and 354.5±88.3μg·L-1; t1/2 were 5.34±1.15 and 5.62±1.23h; Cl/F were 51.0±8.1 and 53.8±9.4L h-1; Vd/F were 390.6±96.8 and 438.4±122.4L; MRT0-t were 6.61±0.82 and 6.56±0.87h;AUC0-t were 2290.1±368.1 and 2159.5±372.8μg·h·L-1 for the test and reference drugs, respectively. The relative bioavailability of the test drug is 107.6±17.3％.The main pharmacokinetic parameters were analyzed by ANOVA after log transformation. It showed that AUC0-t,AUC0-∞ and Cmax for the test and reference drugs of Fexofenadine Hydrochloride has no significant difference in preparations and periods, and has significant difference between individual. Adopted two one-side test and (1-2α) confidential interval test, AUC0-t,AUC0-∞ and Cmax of test preparation both rejection the assumption of inequivalence. AUC0-t of test preparation was 99.7～113.3％of reference preparation by 90％confidence interval test. So the test and reference preparations were bioequivalence.The result of concentration-time curve was fitting by DAS 1.0. The pharmacokinetic behavior of Fexofenadine Hydrochloride in human conformed with two-compartment model... |
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