title: 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.
abstract: 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.
authors.affiliation: Key Laboratory of Drug Metabolism & Pharmacokinetics, China Pharmaceutical University, 24# Tong-Jia Lane, Nanjing 210009, China. gooey@vip.sina.com
authors.forename: Yi
authors.initials: Y
authors.lastname: Gu
authors.forename: Guang-Ji
authors.initials: GJ
authors.lastname: Wang
authors.forename: Jian-Guo
authors.initials: JG
authors.lastname: Sun
authors.forename: Xiao-Yan
authors.initials: XY
authors.lastname: Zhang
authors.forename: Yang
authors.initials: Y
authors.lastname: Sai
mesh_terms: 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
chemical_list: D003692:Delayed-Action Preparations; D004337:Drug Carriers; D013607:Tablets; D019357:Potassium Citrate