Sensitive and selective liquid chromatography-electrospray ionization tandem mass spectrometry analysis of hydrochlorothiazide in rat plasma.
A sensitive and selective method for the determination of hydrochlorothiazide (HCTZ) concentrations in rat plasma was developed using high performance liquid chromatography-electrospray ionization tandem mass spectrometry (LC-MS/MS). An aliquot of plasma (50 microl) was mixed with the solution of internal standard, hydrofluorothiazide (HFTZ), and extracted with tert-butyl methyl ether. The reconstituted extract was applied to the LC-MS/MS system with a reversed phase C8 column and eluted with distilled water/acetonitrile (85/15, v/v). To enhance negative ionization of HCTZ and HFTZ in the multiple reaction monitor (MRM), the solution consisting of acetonitlile/1% (v/v) ammonia solution (95/5, v/v) was delivered after column separation. This additional technique, so-called the post-column addition, increased sensitivity of HCTZ and HFTZ about 500- and 200-fold, respectively. The calibration curve showed good linearity (r = 0.999) over the range of 4-1000 ng/ml. Acceptable accuracy (100.8-113.1%) and precision (0.28-16.4%) were confirmed in the intra- and the inter-day analyses. It is indicated that this LC-MS/MS method is useful for pharmacokinetic studies of HCTZ in small animals, because it enabled the serial determination of plasma level of HCTZ in rats.
Takubo
Takatoshi
T
Drug Metabolism, Tsukuba Research Institute, Banyu Pharmaceutical Co Ltd, 3, Okubo, Tsukuba, Ibaraki 300-2611, Japan. takatoshi_tokubo@merck.com
Okada
Hiromasa
H
Ishii
Mikio
M
Hara
Ken-ichi
K
Ishii
Yasuyuki
Y
D000818:Animals; D019540:Area Under Curve; D002853:Chromatography, Liquid / Q000379:methods*; D004232:Diuretics; D006852:Hydrochlorothiazide / Q000097:blood* / Q000493:pharmacokinetics; D051381:Rats; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D049993:Sodium Chloride Symporter Inhibitors / Q000097:blood* / Q000493:pharmacokinetics; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D004232:Diuretics; D049993:Sodium Chloride Symporter Inhibitors; D006852:Hydrochlorothiazide