liquid chromatography–tandem mass spectrometry method for the quantitation of actarit in rabbit plasma: application to pharmacokinetics and metabolic stability

• Published in: Journal of mass spectrometry. Vol. 51; no. 1; pp. 69 - 78
• Main Authors: Ramakrishna, Rachumallu, Bhateria, Manisha, Puttrevu, Santosh kumar, Durga Prasad, Yarra, Singh, Rajbir, Bhatta, Rabi Sankar
• Format: Journal Article
• Language: English
• Published: England Wiley 2016; Blackwell Publishing Ltd; Wiley Subscription Services, Inc
• Subjects: acetonitrile; actarit; Administration, Oral; ammonium acetate; Animals; Antirheumatic Agents - administration & dosage; Antirheumatic Agents - blood; Antirheumatic Agents - metabolism; Chromatography; Chromatography, High Pressure Liquid - methods; Coumaric Acids - chemistry; Drugs; Food and Drug Administration; guidelines; ionization; LC-MS/MS; Limit of Detection; liquid chromatography; Liquid-liquid extraction; Liquid-Liquid Extraction - methods; Liquids; liver microsomes; Male; Mass spectrometry; metabolic stability; Metabolism; methanol; Microsomes, Liver - metabolism; monitoring; New Zealand White rabbit; oral administration; p-coumaric acid; pharmacokinetics; Pharmacology; Phenylacetates - administration & dosage; Phenylacetates - blood; Phenylacetates - metabolism; Propionates; rabbit plasma; Rabbits; Reproducibility of Results; rheumatoid arthritis; Scientific imaging; Stability; tandem mass spectrometry; Tandem Mass Spectrometry - methods; United States; pharmacokinetics; LC-MS/MS; actarit; rabbit plasma; rheumatoid arthritis; metabolic stability
• ISSN: 1076-5174; 1096-9888
• DOI: 10.1002/jms.3730

Actarit (ATR), 4‐acetylaminophenylacetic acid is an orally effective disease‐modifying anti‐rheumatic drug widely prescribed for the treatment of rheumatoid arthritis. The present study demonstrates the first report on a selective and sensitive liquid chromatography–tandem mass spectrometry method for the quantification of ATR in rabbit plasma using p‐coumaric acid as an internal standard (IS). Following liquid–liquid extraction, chromatographic separation of the reconstituted samples was achieved isocratically on a Syncronis‐C18 column with a mobile phase consisting of aqueous ammonium acetate (10 mM, pH 4)‐ methanol and acetonitrile mixture (8 : 92, v/v) at a flow rate of 0.6 ml/min. ATR and IS were detected using electrospray ionization operated in negative multiple reaction monitoring mode. The calibration curve was linear (r² ≥ 0.990) over the concentration range of 1–4000 ng/ml with a lower limit of quantitation of 1 ng/ml. The mean extraction recovery of ATR and IS from rabbit plasma was greater than 85%. The method complied well with US Food and Drug Administration guidelines for selectivity, sensitivity, accuracy, precision, matrix effect, dilution integrity, carry‐over effect and stability. The method was successfully applied to in vitro metabolic stability (using rabbit liver microsomes) and in vivo pharmacokinetic study after oral administration of ATR at a dose of 10 mg/kg in New Zealand rabbits. Copyright © 2015 John Wiley & Sons, Ltd.