title: Comparison of extraction procedures for assessment of matrix effect for selective and reliable determination of atazanavir in human plasma by LC-ESI-MS/MS.
abstract: A comparative study with three conventional extraction techniques namely protein precipitation (PP), liquid-liquid extraction (LLE) and solid phase extraction (SPE) has been demonstrated to assess the magnitude of matrix interference by post-column analyte infusion and post extraction analyte spiking for the determination of atazanavir from human plasma. Severe ion suppression observed in PP and to a lesser extent in LLE was circumvented by SPE on LiChrosep Sequence extraction cartridge. Based on these observations a selective, rugged and high throughput SPE-LC-MS/MS method has been developed for reliable determination of atazanavir in human plasma. The chromatographic separation was achieved on a Hypersil Gold C18 (50mm×4.6mm, 5μm) analytical column using 5mM ammonium formate in water:methanol (10:90, v/v) as the mobile phase under isocratic conditions. The method was validated over a wide dynamic concentration range of 10-6000ng/mL. The mean relative recovery and absolute matrix effect across quality controls were 84.9 and 93.2%, respectively. The precision value for relative matrix effect between eight different lots of plasma, expressed as %CV of the slopes of the calibration lines was 2.41. The stability of atazanavir under different storage conditions varied from -8.4 to 5.4%. The method was successfully applied to a bioequivalence study of 300mg atazanavir capsule formulation in 24 healthy Indian males under fasting condition.
authors.affiliation: Bioanalytical Research, CRO, Cadila Pharmaceuticals Ltd., Ahmedabad 387810, India; Chemistry Department, School of Sciences, KSKV Kachchh University, Bhuj 370001, India.
authors.forename: Manish
authors.initials: M
authors.lastname: Yadav
authors.forename: Vikas
authors.initials: V
authors.lastname: Trivedi
authors.forename: Vivek
authors.initials: V
authors.lastname: Upadhyay
authors.forename: Gaurang
authors.initials: G
authors.lastname: Shah
authors.forename: Girin A
authors.initials: GA
authors.lastname: Baxi
authors.forename: Sailendra
authors.initials: S
authors.lastname: Goswami
authors.forename: Pranav S
authors.initials: PS
authors.lastname: Shrivastav
mesh_terms: D000097:Acetonitriles; D000069446:Atazanavir Sulfate; D002853:Chromatography, Liquid / Q000379:methods*; D018592:Cross-Over Studies; D004355:Drug Stability; D006801:Humans; D019469:Indinavir / Q000097:blood; D008297:Male; D000432:Methanol; D009842:Oligopeptides / Q000097:blood* / Q000493:pharmacokinetics; D011725:Pyridines / Q000097:blood* / Q000493:pharmacokinetics; D015203:Reproducibility of Results; D052616:Solid Phase Extraction; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D013810:Therapeutic Equivalency
chemical_list: D000097:Acetonitriles; D009842:Oligopeptides; D011725:Pyridines; D000069446:Atazanavir Sulfate; D019469:Indinavir; D000432:Methanol; C032159:acetonitrile