Simultaneous determination of irbesartan and hydrochlorothiazide in human plasma by ultra high performance liquid chromatography tandem mass spectrometry and its application to a bioequivalence study

• A UPLC–MS/MS method for the simultaneous determination of irbesartan and hydrochlorothiazide in human plasma was developed and validated.
• The advantages of this assay include simple sample preparation procedures, short analysis time (1.5 min per sample), and high sensitivity.
• This assay was used to support a clinical study.
• A relatively larger number of samples can be analyzed in a short time.

An ultra high performance liquid chromatography tandem mass spectrometry (U-HPLC–MS/MS) method was developed and validated to determine irbesartan (IRB) and hydrochlorothiazide (HCTZ) in human plasma simultaneously. Plasma samples were prepared using protein precipitation with acetonitrile, the two analytes and the internal standard losartan were separated on an Acquity U-HPLC BEH C18 column and mass spectrometric analysis was performed using a QTrap5500 mass spectrometer coupled with an electro-spray ionization (ESI) source in the negative ion mode. The MRM transitions of m/z 427.2 → 206.9 and m/z 296.1 → 204.9 were used to quantify for IRB and HCTZ, respectively. The linearity of this method was found to be within the concentration range of 5–3000 ng/mL for IRB, and 0.5–300 ng/mL for HCTZ in human plasma, respectively. The lower limit of quantification (LLOQ) was 5 ng/mL and 0.5 ng/mL for IRB and HCTZ in human plasma, respectively. The relative standard deviations (RSD) of intra and inter precision were less than 12% for both IRB and HCTZ. The analysis time of per sample was 2.5 min. The developed and validated method was successfully applied to a bioequivalence study of IRB (300 mg) with HCTZ (12.5 mg) tablet in Chinese healthy volunteers (N = 20).