Simultaneous determination of pioglitazone and its two active metabolites in human plasma by LC-MS/MS.
A liquid chromatography/tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous determination of pioglitazone (PIO) and its two metabolites: M-III (keto-derivative) and M-IV (hydroxy-derivative) in human plasma. Human plasma samples of 0.2 ml were extracted by a single step liquid-liquid extraction procedure and analyzed using a high performance liquid chromatography (HPLC) electrospray tandem mass spectrometer system. The compounds were eluted isocratically on a C-18 column, ionized using a positive ion atmospheric pressure electrospray ionization source and analyzed using multiple reaction monitoring mode. The ion transitions monitored were m/z 357-->134 for PIO, m/z 371-->148 for M-III, m/z 373-->150 for M-IV and m/z 413-->178 for the internal standard. The chromatographic run time was 2.5 min per injection, with retention times of 1.45, 1.02 and 0.95 min for PIO, M-III and M-IV, respectively. The calibration curves of pioglitazone, M-III and M-IV were well fit over the range of 0.5-2000 ng/ml (r(2)>0.998759) by using a weighted (1/x(2)) quadratic regression. The inter-day precisions of the quality control samples (QCs) were </=10.5% (N=15), coefficient of variation (CV) and the inter-day accuracy (%Nominal) ranged from 84.6 to 103.5% for PIO, 94.4 to 104.0% for M-III, and 96.8 to 101.0% for M-IV. All three analytes demonstrated acceptable short-term, long-term, and freeze/thaw stability. The method is simple, rapid and rugged, and has been applied successfully to sample analysis for clinical studies.
Lin
Zhongping John
ZJ
Avantix Laboratories, Inc., 57 Read's Way, New Castle, DE 19720, USA. linj@avantixlabs.com
Ji
Weihua
W
Desai-Krieger
Daksha
D
Shum
Linyee
L
D001711:Biotransformation; D002138:Calibration; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D007004:Hypoglycemic Agents / Q000097:blood*; D007202:Indicators and Reagents; D013058:Mass Spectrometry; D000077205:Pioglitazone; D011786:Quality Control; D015203:Reproducibility of Results; D013056:Spectrophotometry, Ultraviolet; D045162:Thiazolidinediones / Q000097:blood*
D007004:Hypoglycemic Agents; D007202:Indicators and Reagents; D045162:Thiazolidinediones; D000077205:Pioglitazone