ABSTRACT
Objective To develop and validate a LC-MS/MS method to quantify oxybutynin in rabbit plasma and evaluate the bioequivalence of self-prepared oxybutynin chloride gel and Gelnique. Methods The plasma sample was submitted to liquid-liquid extraction using methyl t-butyl ether after alkalified by 0.5 mol/L NaOH, with di-phenhydramine as the internal standard. Chromatographic separation was performed on a Kinetex C18column with the mobile phase consisting of 10 mmol/L ammonium acetate(1‰formic acid)-acetonitrile(50 : 50,v/v). Oxy-butynin and diphenhydramine were ionized with an ESI source operated in positive ion mode,and the detected ions were m/z 358→142 (oxybutynin),m/z 256→167(diphenhydramine). The validated method was then applied to the drug determination in rabbit plasma following single dermal topical administration of oxybutynin gel. Results Calibration curve was liner over the concentration range of 1~200 μg/L in rabbit plasma.For quality control samples, the intra-and inter-day precision was in the range of 1.67%~9.79%, and accuracy was within 92.9% to 103%. Self-prepared oxybutynin chloride gel and Gelnique were proved to be bioequivalent. Conclusions It was validated that the LC-MS/MS method is simple,strong specificity and high sensitivity,which could be successfully applied to pharmacokinetic study and bioequivalence evaluation of transdermal oxybutynin formulations in rabbit.
ABSTRACT
<p><b>OBJECTIVE</b>To modify the isolation and culture method of Sertoli cells and investigate its' effects on xeno-lymphocytes apoptosis.</p><p><b>METHODS</b>Sertoli cells which was isolated from 2 - 4 week-old Sprague Dawley (SD) rats, were successfully prepared by collagenase type V, trypsin and DNase I and then identified by electron microscope. Viability and apoptosis of cultured cells were measured by flow cytometry. The apoptosis rates of Balb/c mouse lymphocytes were examined which were co-cultured with Sertoli cells of SD rats by flow cytometry, too. The expression of FasL, TGF-beta(1) and clusterin on Sertoli cells were detected by immunocytochemistry.</p><p><b>RESULTS</b>In the co-cultured system, Sertoli cells accounted for more than 90%. The viability of Sertoli cells was above 95% and the apoptosis rate was 10.87% +/- 3.87% in this study. The lymphocytes apoptosis ratio was 15.52% +/- 0.17% (P < 0.01). Streptavidin-biotin-peroxidase-complex immunochemistry staining showed that the Sertoli cells could express FasL, TGF-beta(1) and clusterin, respectively.</p><p><b>CONCLUSIONS</b>It indicates that the expression of FasL, TGF-beta(1) on the Sertoli cells might relate to the immune privilege, and it supposed to be benefit for xenotransplantation.</p>