No abstract

A sensitive and rapid liquid chromatography-tandem mass spectrometry (LC–MS/MS) method has been developed for the simultaneous determination of lisinopril (LIS) and hydrochlorothiazide (HCTZ) in human plasma using their labeled internal standards (ISs). Sample pre-treatment involved solid phase extraction on Waters Oasis HLB cartridges using 100 µL of plasma, followed by liquid chromatography on Hypersil Gold C18 (50 mm×3.0 mm, 5 µm) column. The analytes were eluted within 2.0 min using acetonitrile-5.0 mM ammonium formate, pH 4.5 (85:15, v/v) as the mobile phase. The analytes and ISs were analyzed in the negative ionization mode and quantified using multiple reaction monitoring. The method showed excellent linearity over the concentration range of 0.50–250.0 ng/mL for both the analytes. The intra-batch and inter-batch precision (% CV) was ≤5.26% and their extraction recoveries were in the range of 96.6%–103.1%. Matrix effect evaluated in terms of IS-normalized matrix factors ranged from 0.97 to 1.03 for both the analytes. The validated method was successfully applied to determine the plasma concentration of the drugs using 10 mg lisinopril and 12.5 mg hydrochlorothiazide fixed dose formulation in 18 healthy Indian volunteers.

A sensitive and selective method has been proposed for the simultaneous determination of amlodipine (AML), valsartan (VAL) and hydrochlorothiazide (HCTZ) in human plasma by liquid chromatography–tandem mass spectrometry (LC–MS/MS). The analytes and their deuterated analogs were quantitatively extracted from 100 µL human plasma by solid phase extraction on Oasis HLB cartridges. The chromatographic separation of the analytes was achieved on a Chromolith RP18e (100 mm × 4.6 mm) analytical column within 2.5 min. The resolution factor between AML and VAL, AML and HCTZ, and VAL and HCTZ was 2.9, 1.5 and 1.4, respectively, under isocratic conditions. The method was validated over a dynamic concentration range of 0.02–20.0 ng/mL for AML, 5.00–10,000 ng/mL for VAL and 0.20–200 ng/mL for HCTZ. Ion-suppression/enhancement effects were investigated by post-column infusion technique. The mean IS-normalized matrix factors for AML, VAL and HCTZ were 0.992, 0.994 and 0.998, respectively. The intra-batch and inter-batch precision (% CV) across quality control levels was ≤ 5.56% and the recovery was in the range of 93.4%–99.6% for all the analytes. The method was successfully applied to a bioequivalence study of 5 mg AML + 160 mg VAL + 12.5 mg HCTZ tablet formulation (test and reference) in 18 healthy Indian males under fasting. The mean log-transformed ratios of Cmax, AUC0–120h and AUC0-inf and their 90% CIs were within 90.2%–102.1%. The assay reproducibility was demonstrated by reanalysis of 90 incurred samples.

Abstract:A selective and sensitive ultra performance liquid chromatography-tandem mass spectrometry method was developed for the simultaneous determination of losartan (LOS), EXP-3174, which is an active metabolite LOS carboxylic acid, and hydrochlorothiazide (HCTZ) in human plasma. Solid-phase extraction was carried out on Oasis HLB cartridges with 100 µ L of plasma to give an extraction recovery in the range of 88.5%-102.5% for the three analytes.Chromatography on a BEH C18 column afforded baseline separation of all the analytes within 2.4 min using 1.0% formic acid in water and acetonitrile (15:85, v/v) as the mobile phase. Quantitation was performed with multiple reaction monitoring in the negative ionization mode. The response of the method was linear over a dynamic range of 0.5-500, 1.0-750, and 0.25-150 ng/mL for LOS, EXP-3174, and HCTZ, respectively. Extent of signal suppression/enhancement was examined through postcolumn infusion. The effect of matrix components was evaluated by postextraction spiking and calculation of the slope of calibration lines. The method was successfully applied to a bioequivalence study of 50 mg losartan and 12.5 mg hydrochlorothiazide tablet formulation in 65 healthy human subjects. Reproducibility of the method was shown by reanalysis of 213 incurred samples.

A single LC–MS/MS assay has been developed and validated for the simultaneous determination of metformin and dapagliflozin in human plasma using ion‐pair solid‐phase extraction. Chromatographic separation of the analytes and their internal standards was carried out on a reversed‐phase ACE 5CN (150 × 4.6 mm, 5 μm) column using acetonitrile–15 mm ammonium acetate, pH 4.5 (70:30, v/v) as the mobile phase. To achieve higher sensitivity and selectivity for the analytes, mass spectrometric analysis was performed using a polarity switching approach. Ion transitions studied using multiple reaction monitoring mode were m/z 130.1 [M + H]+/60.1 for metformin and m/z 467.1 [M + CH3COO]−/329.1 for dapagliflozin in the positive and negative modes, respectively. The linear calibration range of the assay was established from 1.00 to 2000 ng/mL for metformin and from 0.10 to 200 ng/mL for dapagliflozin to achieve a better assessment of the pharmacokinetics of the drugs. The limit of detection and limit of quantitation for the analytes were 0.39 and 1.0 ng/mL for metformin and 0.03 and 0.1 ng/mL for dapagliflozin, respectively. There was no interference of plasma matrix obtained from different sources, including hemolyzed and lipemic plasma. The method was successfully applied to study the effect of food on the pharmacokinetics of metformin and dapagliflozin in healthy subjects.