A new method for plasma citrate determination by reversed-phase high-performance liquid chromatography after ultrafiltration extraction: an example for bioequivalence evaluation of a medicinal endogenous substance.
Bioequivalence studies of pharmaceutical preparations of medicinal endogenous substances are generally lacking, as the endogenous background is the main obstacle for both experimental design and drug analysis. We conducted a single-dose, self-control, 3-period crossover study to evaluate the bioequivalence of a sustained-release versus an immediate-release preparation of potassium citrate for the treatment of urinary tract stones. This study included a placebo period to monitor the dynamics of endogenous plasma citrate, which could therefore be subtracted during the data processing. Notably, a new convenient method for plasma citrate determination utilizing ultrafiltration extraction and direct reversed-phase high-performance liquid chromatography (HPLC) was successfully applied in the study after validation. To our knowledge, this is the first report using simple reversed-phase HPLC to analyze plasma citrate, and ultrafiltration also significantly simplified the plasma extraction procedure. HPLC was performed using an ODS column, with acetonitrile and 0.02 mol/l sulfuric acid (3:97 v/v) as the mobile phase and ultraviolet detection at 210 nm. Results showed linearity from 10 to 200 microg/ml, an extraction recovery of more than 90% and analysis variability within 15% for additive citrate. The concentration of dog endogenous plasma citrate was determined to be about 30-40 microg/ml. The sustained-release preparation was bioequivalent to the immediate-release preparation in terms of the extent of absorption, but it could not achieve full effects due to a tmax issue.
Gu
Yi
Y
Key Laboratory of Drug Metabolism & Pharmacokinetics, China Pharmaceutical University, 24# Tong-Jia Lane, Nanjing 210009, China. gooey@vip.sina.com
Wang
Guang-Ji
GJ
Sun
Jian-Guo
JG
Zhang
Xiao-Yan
XY
Sai
Yang
Y
D000284:Administration, Oral; D000818:Animals; D019540:Area Under Curve; D001682:Biological Availability; D002681:China; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D018592:Cross-Over Studies; D003692:Delayed-Action Preparations / Q000008:administration & dosage / Q000737:chemistry / Q000493:pharmacokinetics; D004285:Dogs; D004337:Drug Carriers / Q000737:chemistry; D005260:Female; D008297:Male; D010949:Plasma / Q000737:chemistry*; D019357:Potassium Citrate / Q000097:blood* / Q000737:chemistry / Q000493:pharmacokinetics*; D011897:Random Allocation; D015203:Reproducibility of Results; D052616:Solid Phase Extraction / Q000379:methods; D013607:Tablets; D013678:Technology, Pharmaceutical / Q000379:methods; D013810:Therapeutic Equivalency; D014462:Ultrafiltration / Q000379:methods
D003692:Delayed-Action Preparations; D004337:Drug Carriers; D013607:Tablets; D019357:Potassium Citrate