title: Quantitative analysis of 23-hydroxybetulinic acid in mouse plasma using electrospray liquid chromatography/mass spectrometry.
abstract: 23-Hydroxybetulinic acid is a newly isolated derivative of betulinic acid. The agent exhibits potential anti-tumor activity and functions in this regard via apoptosis. In support of pharmacokinetic and toxicological evaluations, a new assay based on liquid chromatography/mass spectrometry (LC/MS) was developed for the quantitative analysis of 23-hydroxybetulinic acid. Sample preparation consisted of extraction of the plasma by the addition of methylene chloride followed by centrifugation. Aliquots of the supernatant were analyzed using an isocratic reversed-phase high-performance liquid chromatography (HPLC) system coupled to a negative ion electrospray mass spectrometer. Molecules of 23-hydroxybetulinic acid and the internal standard limonin were detected using selected ion monitoring at m/z 471 and 469, respectively. The limit of detection of 23-hydroxybetulinic acid was 0.05 pg (0.11 fmol) injected on-column (10 pg/mL, 5 microL injection volume), and the limit of quantitation was 10 pg (21.19 fmol, 2 ng/mL, 5 muL injection volume). 23-Hydroxybetulinic acid was stable in plasma samples at -20 degrees C for at least 3 weeks. The intra-day and inter-day coefficients of variation of the assay were 3.0 and 4.8%, respectively. The utility of the assay was demonstrated by measuring 23-hydroxybetulinicacid in mouse plasma following intragastric administration (IG) in vivo. Pharmacokinetic parameters were calculated using the 3P97 pharmacokinetic software package. A two-compartment, first-order model was selected for pharmacokinetic modeling. The result showed that after IG of 200 mg/kg 23-hydroxybetulinic acid, the plasma concentrations reached peaks at 2 h with C(max) of 3.1 microg/mL. The 200 mg/kg 23-hydroxybetulinic acid suspension IG doses were found to have long elimination half-lives of 25.6 h and low bioavailability of 2.3%. No interference was noted due to endogenous substances. These analytical methods should be of value in future studies related to the development and characterization of 23-hydroxybetulinic acid.
authors.affiliation: Research Center of Pharmacokinetics, China Pharmaceutical University, Nanjing 210009, P.R. China. ymzfk@163.com
authors.forename: Min
authors.initials: M
authors.lastname: Yang
authors.forename: Guang Ji
authors.initials: GJ
authors.lastname: Wang
authors.forename: Su Jun
authors.initials: SJ
authors.lastname: Wang
authors.forename: Xiao Tian
authors.initials: XT
authors.lastname: Li
authors.forename: Yu Ping
authors.initials: YP
authors.lastname: Xu
authors.forename: Song Pei
authors.initials: SP
authors.lastname: Wang
authors.forename: Jing De
authors.initials: JD
authors.lastname: Xiang
authors.forename: Shang Ren
authors.initials: SR
authors.lastname: Pan
authors.forename: Guo Xian
authors.initials: GX
authors.lastname: Cao
authors.forename: Wen Cai
authors.initials: WC
authors.lastname: Ye
mesh_terms: D000818:Animals; D000970:Antineoplastic Agents / Q000097:blood* / Q000493:pharmacokinetics; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004365:Drugs, Chinese Herbal; D051379:Mice; D008813:Mice, Inbred ICR; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D014315:Triterpenes / Q000097:blood* / Q000493:pharmacokinetics
chemical_list: C468651:23-hydroxybetulinic acid; D000970:Antineoplastic Agents; D004365:Drugs, Chinese Herbal; D014315:Triterpenes