PT  - JOURNAL ARTICLE
AU  - Sun, Zejin
AU  - Murry, Daryl J.
AU  - Sanghani, Sonal P.
AU  - Davis, Wilhelmina I.
AU  - Kedishvili, Natalia Y.
AU  - Zou, Qin
AU  - Hurley, Thomas D.
AU  - Bosron, William F.
TI  - Methylphenidate Is Stereoselectively Hydrolyzed by Human Carboxylesterase CES1A1
AID  - 10.1124/jpet.104.067116
DP  - 2004 Aug 01
TA  - Journal of Pharmacology and Experimental Therapeutics
PG  - 469--476
VI  - 310
IP  - 2
4099  - http://jpet.aspetjournals.org/content/310/2/469.short
4100  - http://jpet.aspetjournals.org/content/310/2/469.full
SO  - J Pharmacol Exp Ther2004 Aug 01; 310
AB  - Methylphenidate is an important stimulant prescribed to treat attention-deficit hyperactivity disorder. It has two chiral centers, but most current commercial formulations consist of the racemic mixture of the threo pair of methylphenidate isomers (d-, l-threo-methylphenidate). The d-isomer is the pharmacologically active component. Numerous studies reported that oral administration of the methylphenidate racemate undergoes first-pass, stereoselective clearance in humans with l-methylphenidate being eliminated faster than d-methylphenidate. Accordingly, the kinetics of hydrolysis of individual enantiomers by purified native and recombinant human liver carboxylesterases CES1A1 and CES2 and a colon isoenzyme CES3 were examined with a liquid chromatography/mass spectrometry assay. The expression of CES1A1, CES2, and CES3 in Sf9 cells and the methods for purification of the three isoenzymes are reported. CES1A1 has a high catalytic efficiency for both d- and l-enantiomers of methylphenidate. No catalytic activity was detected with CES2 and CES3 for either enantiomer. The catalytic efficiency of CES1A1 for l-methylphenidate (kcat/Km = 7.7 mM–1 min–1) is greater than that of d-methylphenidate (kcat/Km = 1.3–2.1 mM–1 min–1). Hence, the catalytic efficiency of CES1A1 for methylphenidate enantiomers agrees with stereoselective clearance of methylphenidate reported in human subjects. Both enantiomers of methylphenidate can be fit into the three-dimensional model of CES1A1 to form productive complexes in the active site. We conclude that CES1A1 is the major enzyme responsible for the first-pass, stereoselective metabolism of methylphenidate. The American Society for Pharmacology and Experimental Therapeutics