Verification Of Structure And Study On Quality Control And Pharmacokinetics And Bioequiavailability In Rats Of Lamivudine

It is widely believed that virosis is one of the serious diseases to harm mankind's healthy at present,especially HBV and HIV.The study of antiviral drugs becomes more and more important,and it is one of the hot spots in the modern medical research.Lamivudine(3TC) is a high performance,high selective.secure and low-adverse reaction antiviral drug.It has been used to treat HBV and HIV in clinic,and gotten satisfactory curative effect. Antiviral mechanism of action of lamivudine is restraining RNA of pregenome to negative strand DNA,blocking the chain of HBV-DNA or HIV-DNA,activeIy suppressing replication of HBV or HIV,finally killing virus.In this study, the structure of lamivudine was Conformed by NMR,MS,IR,UV and so on along with its physical-chemical property. At the same time, its purity was verified to the standard of raw drug by thermal analysis , element analysis and MS ,etc.Since the quality standard of lamivudine has been recorded into USP26, but the quality standard of lamivudine and its tablet has not been recorded into Chp yet, quality standard of lamivudine was studied on its chemical identification, determination of general and related substance and its physical-chemical property based on its structure property. The UV-Vis absorptive spectrum characteristic was studied, and the results were followed: the maximum absorption wavelength are 270nm. Absorptivities (E_1cm^1%) are 421.5 (270nm) respectively. Critical relative humidity is 84%. Nonaqueous titration was developed for the assay of reference substance and RP-HPLC was performed for the determination the content and related substance of raw drugs and tablet. Lamivudine and its related substance were separated on the Hypersil C₁₈ column (250×4.6mm, l0μm) with phosphate buffer (pH6.8, 20mmol/L)-methanol(91:9) as the mobile phase and detected at 270nm. The calibration curves were linear (r = 0.9999) within the range of 5.00 400.0 μg/ml for lamivudine, the detection limit was Ing (S/N=3 ) , the analysis precision RSD was less than 1.0 % (n=5), the repeatability precision RSD was 0.19% (n=6), and the average recovery was 99.9%. The limit of related substance is less than l%,then chiral HPLC is used to control the content of dextroisomer of lamivudine.This method was simple ,accurate and suitable for the quality control for lamivudine.Since the quality of experiment preparation comparing with the quality of reference preparation, bioequiavailability and pharmacokinetics of Lamivudine in rats in vivo were investigatedin this study. Protein precipitation method was chosen to pretreat plasma samples before RP-HPLC analysis. Lamivudine were separated on Hypersil Cigcolumn (250x4.6mm, 10um ) with the water and methanol as the mobile phase by gradient RP-HPLC and detected at 270nm. The calibration curves were linear (r > 0.9998) within the range of 0.10²0.00 mg/L for lamivudine. All the detection limits were Ing (S/N>3).. The extraction recovery and analytical recovery were all more than 95%. Intra-day and inter-day precision of three were all less than 6.0% (n=5). The reliability of this method is high enough to perform quantitative analysis of lamivudine in plasma samples and the study of bioequiavailability.The pharmacokinetics and bioequiavailability study of lamivudine was performed in 12 healthy rats after intragastric administration with single dosage,using experiment preparation in 6 rats and using reference preparation in the other 6 rats, by the established analytical method. The experimental data showed that the concentration-time curve of lamivudine in rat plasma could be fitted to two-compartment model, and the main pharmacokinetic parameters of experiment preparation were as follows: the area under the plasma concentration-time curve (AUCo-k) was 9.4872+0.7352 ( mg/L ) -h, (AUCo-*.) wasl4.4669±1.6987 ( mg/L ) h ,the peak plasma concentration (Cmax) was 2.0320±0.3701mg/L, the peak time (Tmax) was 0.9889±0.0629h, the absorption half life (txa. ka) was 0.2532+0.1867 h , the distribution half life (t1/2. a) was 1.391310.1912 h, the elimination half life (t1/2. p) was 11.839912.7824 h, the clearance (CL) was 1.2761+0.5069 mg/kg/h ( mg/L ) , and the volume of apparent distribution (V/F) was 20.7360+5.2227( mg/kg )/(mg/L). The main pharmacokinetic parameters of reference preparation were as follows: the area under the plasma concentration-time curve (AUCo->T) was 10.235511.0954 (mg/L) -h, (AUC0^) was 16.133612.5803 (mg/L) -h ,the peak plasma concentration (Cmax) was 2.765010.2931 mg/L, the peak time (Tmax) was 1.0056l0.0574h, the absorption half life (t,/2. ka) wasO.2004+0.1144 h , the distribution half life (tm. a) was 1.391310.1912 h, the elimination half life (t,/2. p ) wasl3.6527+2.3821 h, the clearance (CL) was 1.360010.3373 mg/kg/h ( mg/L ) , and the volume of apparent distribution (V/F) was 26.621417.2545 (mg/kg) /(mg/L) It was also proved that lamivudine was absorpted into blood and eliminated quickly.lt is bioequiavailability between experiment preparation and reference preparation in rats in vivo.These reseach could provide reference for clinical rational administration,quality evaluation of experiment preparation.and moreover offer medical data in human body for studying the pharmacokinetics and bioequiavailability of lamivudine.