GC-MS and GC-MS/MS measurement of ibuprofen in 10-μL, aliquots of human plasma and mice serum using α-methylo-2H3]ibuprofen after ethyl acetate extraction and pentafluorobenzyl bromide derivatization: Discovery of a collision energy-dependent H/D isotope effect and pharmacokinetic application to inhaled ibuprofen-arginine in mice

GC-MS and GC-MS/MS methods were developed and validated for the quantitative determination of ibuprofen (do-ibuprofen), a non-steroidal anti-inflammatory drug (NSAID), in human plasma using alpha-methyl-H-2(3)-4-(isobutyl)phenylacetic acid (d(3)-ibuprofen) as internal standard. Plasma (10 mu L) was diluted with acetate buffer (80 mu L, 1 M, pH 4.9) and d(0)- and d(3)-ibuprofen were extracted with ethyl acetate (2 x 500 mu L). After solvent evaporation and d(0)- d(3)-ibuprofen were derivatized in anhydrous acetonitrile by using pentafluorobenzyl (PFB) bromide and N,N-diisopropylethylamine as the base catalyst. Under electron-capture negative-ion chemical ionization (ECNICI), the PFB esters of d(0)- and d(3)-ibuprofen readily ionize to form their carboxylate anions [M-PFB](-) at m/z 205 and m/z 208, respectively. Collision-induced dissociation (CID) of m/z 205 and m/z 208 resulted in the formation of the anions at m/z 161 and m/z 164, respectively, due to neutral loss of CO2 (44 Da). A collision energy-dependent H/D isotope effect was observed, which involves abstraction/elimination of H- from d(0)-ibuprofen and D- from d(3)-ibuprofen and is minimum at a CE value of 5 eV. Quantitative GC-MS determination was performed by selected-ion monitoring of m/z 205 and m/z 208. Quantitative GC-MS/MS determination was performed by selected-reaction monitoring of the mass transitions m/z 205 to m/z 161 for d(0)-ibuprofen and m/z 208 to m/z 164 for d(3)-ibuprofen. In a therapeutically relevant concentration range (0-1000 mu M) d(0)-ibuprofen added to human plasma was determined with accuracy (recovery, %) and imprecision (relative standard deviation, %) ranging between 93.7 and 110%, and between 0.8 and 4.9%, respectively. GC-MS (y) and GC-MS/MS (x) yielded almost identical results (y = 4.00 + 0.988x, r(2) = 0.9991). In incubation mixtures of arachidonic acid (10 mu M), d(3)-ibuprofen (10 mu M) or d(0)-ibuprofen (10 mu M) with ovine cyclooxygenase (COX) isoforms 1 and 2, the concentration of d(3)-ibuprofen and d(0)-ibuprofen did not change upon incubation at 37 degrees C up to 60 min. The trough pharmacokinetics of an inhaled arginine-containing ibuprofen preparation in mice was studied after once-daily treatment (0.0, 0.07, 0.4 and 2.5 mg/kg body weight) for three days. A linear relationship between ibuprofen concentration in serum (10 mu L) and administered dose 24h after the last drug administration was observed. (C) 2016 Elsevier B.V. All rights reserved.