title: Determination of atrasentan by high performance liquid chromatography with fluorescence detection in human plasma.
abstract: Atrasentan is an endothelin antagonist selective for the ET(A) receptor in development at Abbott Laboratories for the treatment of cardiovascular disease and cell proliferation disorders. A simple and sensitive chromatographic method for the determination of atrasentan in human plasma has been developed and validated. The analytical method involves acidification of the plasma samples with 0.3 N HCl prior to extraction with 1:1 (v:v) hexane/tert-butylmethylether. The organic extract was evaporated to dryness, reconstituted with 20:80 (v:v) acetonitrile/0.05 M K(2)HPO(4) and washed with 75:25 (v:v) hexane/tert-butylmethylether. The organic layer was discarded and the aqueous layer was injected into the HPLC. Atrasentan and internal standard (ABT-790) were separated from interference using a 250 x 4.6 mm, 5 microm, 120 A Phenomenex Spherisorb C(8) analytical column with a 50 x 4.6 mm, Alltech Absorbosphere 5 microm CN guard cartridge using a mobile phase consisting of 25:15:5:55 (v:v:v:v) acetonitrile/isopropanol/methanol/0.05 M K(2)HPO(4), pH 7.0, at a flow rate of 1.0 mL/min. Fluorescence detection was achieved using lambda(ex) 278 nm and lambda(em) 322 nm. For a 1.0 mL plasma sample volume, the limit of quantitation was approximately 200 pg/mL. The method was linear from 0.2 to 1300 ng/mL (r(2) = 0.9986). Inter- and intra-day assay RSD (n = 6) were less than 10%. Mean accuracy determinations showed the quality control samples to range between 94 and 99% of the theoretical concentration.
authors.affiliation: Drug Analysis Department, Abbott Laboratories, 100 Abbott Park Road, Abbott Park, IL 60064-6118, USA.
authors.forename: P D
authors.initials: PD
authors.lastname: Bryan
authors.forename: L B
authors.initials: LB
authors.lastname: Sapochak
authors.forename: M M
authors.initials: MM
authors.lastname: Tames
authors.forename: R J
authors.initials: RJ
authors.lastname: Padley
authors.forename: T A
authors.initials: TA
authors.lastname: El-Shourbagy
mesh_terms: D000970:Antineoplastic Agents / Q000032:analysis* / Q000097:blood / Q000493:pharmacokinetics; D000077868:Atrasentan; D002138:Calibration; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004355:Drug Stability; D019332:Endothelin-1 / Q000037:antagonists & inhibitors*; D005453:Fluorescence; D006801:Humans; D015394:Molecular Structure; D011759:Pyrrolidines; D011786:Quality Control; D012015:Reference Standards; D012680:Sensitivity and Specificity; D013696:Temperature
chemical_list: D000970:Antineoplastic Agents; D019332:Endothelin-1; D011759:Pyrrolidines; D000077868:Atrasentan