Abstract
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/K m = 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.
| Original language | English |
|---|---|
| Pages (from-to) | 469-476 |
| Number of pages | 8 |
| Journal | Journal of Pharmacology and Experimental Therapeutics |
| Volume | 310 |
| Issue number | 2 |
| DOIs | |
| State | Published - Aug 2004 |
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ASJC Scopus subject areas
- Pharmacology
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Methylphenidate is stereoselectively hydrolyzed by human carboxylesterase CES1A1. / Sun, Zejin; Murry, Daryl J.; Sanghani, Sonal P.; Davis, Wilhelmina I.; Kedishvili, Natalia Y.; Zou, Qin; Hurley, Thomas; Bosron, William F.
In: Journal of Pharmacology and Experimental Therapeutics, Vol. 310, No. 2, 08.2004, p. 469-476.Research output: Contribution to journal › Article
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TY - JOUR
T1 - Methylphenidate is stereoselectively hydrolyzed by human carboxylesterase CES1A1
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
AU - Bosron, William F.
PY - 2004/8
Y1 - 2004/8
N2 - 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/K m = 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.
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/K m = 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.
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U2 - 10.1124/jpet.104.067116
DO - 10.1124/jpet.104.067116
M3 - Article
VL - 310
SP - 469
EP - 476
JO - Journal of Pharmacology and Experimental Therapeutics
JF - Journal of Pharmacology and Experimental Therapeutics
SN - 0022-3565
IS - 2
ER -