LC-MS/MS method for the determination of rosiglitazone on rat dried blood spots and rat urine: Application to pharmacokinetics.

A bioanalytical method for the quantification of rosiglitazone on rat dried blood spots (DBS) and rat urine using liquid chromatography, electrospray ionization coupled with tandem mass spectrometry (LC-ESI-MS/MS) was developed and validated. The chromatographic separation was achieved on a Nova-Pak C18 Column (150 mm × 4.6 mm i.d., 4 µm), using 30 mM ammonium acetate (pH 4.0 adjusted with acetic acid) and acetonitrile (75:25, v/v) as a mobile phase at ambient temperature. LC-MS detection was performed with selected ion monitoring using target ions at m/z 358 and m/z 356 for rosiglitazone and pioglitazone respectively. The calibration curve showed a good linearity in the concentration range of 0.05-100 ng/mL. The effect of hematocrit, blood volume and punch location for DBS samples was studied. The mean recoveries of rosiglitazone from DBS and urine were 93.30% and 92.49% respectively. The intra and inter-day precisions of RSD were less than 4.82% in DBS as well as urine. The limit of detections and quantifications were 0.015 and 0.052 ng/mL in DBS and 0.023 and 0.075 ng/mL in urine samples respectively. The method was validated as per FDA guidelines and successfully applied to a pharmacokinetic study of rosiglitazone in rats.

Development of LC-MS/MS method for the determination of dapiprazole on dried blood spots and urine: application to pharmacokinetics.

A rapid and highly sensitive liquid chromatography­tandem mass spectrometric (LC-MS/MS) method for determination of dapiprazole on rat dried blood spots and urine was developed and validated. The chromatographic separation was achieved on a reverse-phase C18 column (250 × 4.6 mm i.d., 5 µm), using 20 mm ammonium acetate (pH adjusted to 4.0 with acetic acid) and acetonitrile (80:20, v/v) as a mobile phase at 25 °C. LC-MS detection was performed with selective ion monitoring using target ions at m/z 326 and m/z 306 for dapiprazole and mepiprazole used as internal standard, respectively. The calibration curve showed a good linearity in the concentration range of 1­3000 ng/mL. The effect of hematocrit on extraction of dapiprazole from DBS was evaluated. The mean recoveries of dapiprazole from DBS and urine were 93.88 and 90.29% respectively. The intra- and inter-day precisions were <4.19% in DBS as well as urine. The limits of detection and quantification were 0.30 and 1.10 ng/mL in DBS and 0.45 and 1.50 ng/mL in urine samples, respectively. The method was validated as per US Food and Drug Administration guidelines and successfully applied to a pharmacokinetic study of dapiprazole in rats.

LC-MS/MS method for the characterization of the forced degradation products of Entecavir.

A rapid, specific, and reliable isocratic LC-MS/MS method has been developed and validated for the identification and characterization of the stressed degradation products of Entecavir (ETV). ETV, an antiviral drug, was subjected to hydrolysis (acidic, alkaline, and neutral), oxidation, photolysis and thermal stress, as per the international conference on harmonization specified conditions. The drug showed extensive degradation under oxidative and acid hydrolysis stress conditions. However, it was stable to thermal, acidic, neutral, and photolysis stress conditions. A total of five degradation products were observed and the chromatographic separation of the drug and its degradation products were achieved on a Waters Symmetry C18 (250 mm × 4.6 mm, id, 5 µm) column using 20 mM ammonium acetate (pH 3)/acetonitrile (50:50, v/v) as a mobile phase. The degradation products were characterized by LC-MS/MS and its fragmentation pathways were proposed. The LC-MS method was validated with respect to specificity, linearity, accuracy, and precision. No previous reports were found in the literature regarding the degradation behavior of ETV.

LC-MS/MS characterization of forced degradation products of zofenopril.

A rapid, specific and reliable isocratic LC-MS/MS method has been developed and validated for the identification and characterization of stressed degradation products of Zofenopril. Zofenopril, an anti-hypertensive drug, was subjected to hydrolysis (acidic, alkaline and neutral), oxidation, photolysis and thermal stress, as per ICH-specified conditions. The drug showed extensive degradation under oxidative and base hydrolysis stress conditions. However, it was stable to thermal, acid, neutral and photolysis stress conditions. A total of 6 degradation products were observed and the chromatographic separation of the drug and its degradation products were achieved on Phenomenex (Luna) C18 (250mm×4.6mm, i.d., 5µm) column using 20mM ammonium acetate: acetonitrile (50:50, v/v) as a mobile phase. The degradation products were characterized by LC-MS/MS and its fragmentation pathways were proposed. The LC-MS method was validated with respect to specificity, linearity, accuracy and precision. No previous reports were found in the literature regarding the degradation behavior of zofenopril.

Development and validation of a stability-indicating RP-HPLC assay method and stress degradation studies on dapiprazole.

Dapiprazole (DPZ) was subjected to different stress conditions prescribed by the International Conference on Harmonization. A stability-indicating high-performance liquid chromatography method was developed for the analysis of the drug in the presence of its degradation products. The degradation was found to occur in hydrolytic, and to some extent, photolytic conditions, however, the drug was stable to oxidative and thermal stress. The drug was particularly labile under neutral and alkaline hydrolytic conditions. The assay was involved an isocratic elution of DPZ in a Kromasil 100C18 column using a mobile phase composition of water (pH 6.5, 0.05%, w/v, 1-heptane sulfonic acid) and acetonitrile (40:60, v/v). The flow rate was 0.8 mL/min and the detection was conducted at 246 nm. The assay method was found to be linear from 5 to 30 µg/mL. The method was validated for linearity, range, precision, accuracy, specificity, selectivity, limit of detection and limit of quantitation.