A new, validated HPLC-MS/MS method for the simultaneous determination of the anti-cancer agent capecitabine and its metabolites: 5'-deoxy-5-fluorocytidine, 5'-deoxy-5-fluorouridine, 5-fluorouracil and 5-fluorodihydrouracil, in human plasma.
A rapid and selective liquid chromatography/tandem mass spectrometric method was developed for the simultaneous determination of capecitabine and its metabolites 5'-deoxy-5-fluorocytidine (5'-DFCR), 5'-deoxy-5-fluorouracil (5'-DFUR), 5-fluorouracil (5-FU) and dihydro-5-fluorouracil (FUH(2)) in human plasma. A 200 microL human plasma aliquot was spiked with a mixture of internal standards fludarabine and 5-chlorouracil. A single-step protein precipitation method was employed using 10% (v/v) trichloroacetic acid in water to separate analytes from bio-matrices. Volumes of 20 microL of the supernatant were directly injected onto the HPLC system. Separation was achieved on a 30 x 2.1 mm Hypercarb (porous graphitic carbon) column using a gradient by mixing 10 mm ammonium acetate and acetonitrile-2-propanol-tetrahydrofuran (1 : 3 : 2.25, v/v/v). The detection was performed using a Finnigan TSQ Quantum Ultra equipped with the electrospray ion source operated in positive and negative mode. The assay quantifies a range from 10 to 1000 ng/mL for capecitabine, from 10 to 5000 ng/mL for 5'-DFCR and 5'-DFUR, and from 50 to 5000 ng/mL for 5-FU and FUH(2) using a plasma sample of 200 microL. Correlation coefficients (r(2)) of the calibration curves in human plasma were better than 0.99 for all compounds. At all concentration levels, deviations of measured concentrations from nominal concentration were between -4.41 and 3.65% with CV values less than 12.0% for capecitabine, between -7.00 and 6.59% with CV values less than 13.0 for 5'-DFUR, between -3.25 and 4.11% with CV values less than 9.34% for 5'-DFCR, between -5.54 and 5.91% with CV values less than 9.69% for 5-FU and between -4.26 and 6.86% with CV values less than 14.9% for FUH(2). The described method was successfully applied for the evaluation of the pharmacokinetic profile of capecitabine and its metabolites in plasma of treated cancer patients.
Vainchtein
Liia D
LD
Astellas Pharma Europe B.V., Exploratory Development Department, Elisabethhof 1, Leiderdorp, The Netherlands. liia.vainchtein@eu.astellas.com
Rosing
Hilde
H
Schellens
Jan H M
JH
Beijnen
Jos H
JH
D000433:1-Propanol / Q000737:chemistry; D000097:Acetonitriles / Q000737:chemistry; D000964:Antimetabolites, Antineoplastic / Q000097:blood* / Q000737:chemistry; D000069287:Capecitabine; D011232:Chemical Precipitation; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000097:blood / Q000737:chemistry; D004355:Drug Stability; D005467:Floxuridine / Q000097:blood / Q000737:chemistry; D005472:Fluorouracil / Q000031:analogs & derivatives* / Q000097:blood / Q000737:chemistry; D005663:Furans / Q000737:chemistry; D006801:Humans; D016014:Linear Models; D011743:Pyrimidines / Q000097:blood* / Q000737:chemistry; D015203:Reproducibility of Results; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014238:Trichloroacetic Acid / Q000737:chemistry; D014498:Uracil / Q000031:analogs & derivatives / Q000737:chemistry; D014740:Vidarabine / Q000031:analogs & derivatives / Q000737:chemistry
D000097:Acetonitriles; D000964:Antimetabolites, Antineoplastic; D005663:Furans; D011743:Pyrimidines; D005467:Floxuridine; D003841:Deoxycytidine; C018674:tetrahydrofuran; D014498:Uracil; D014238:Trichloroacetic Acid; D000069287:Capecitabine; C019389:5-fluorodihydrouracil; C010889:5-chlorouracil; C418208:5'-deoxy-5-fluorocytidine; D000433:1-Propanol; D014740:Vidarabine; C024352:fludarabine; D005472:Fluorouracil; C025034:doxifluridine; C032159:acetonitrile