We investigated the degradation and racemization of zopiclone (ZOP) enantiomers in plasma and partially aqueous solutions (ethano1:phosphate buffer). Degradation and racemization increased with increasing pH and temperature. Degradation products were identified by means of mass spectrometry, which revealed hydrolysis of the carbamate function and opening of the pyrrolidone ring. In plasma, neither degradation nor racemization occurred after 6 months of storage at -20°C and subsequent extraction. KEY WORDS: chirality, cyclopyrrolone, isomers, racemization, stability, zopiclone Zopiclone (ZOP) is a c h a l cyclopyrrolone ( Fig. 1) with hypnotic properties. It is administered therapeutically as a racemic mixture. ' The racemate has been extensively studied, and several analytical methods based on liquid chromatography or gas chromatography have been proposed for its determination in plasma. 2-7 Two chiral methods have been reported for the study of ZOP enantiomers in plasma and urine, using liquid chromatography with spectrofluorimetric detection. ' ,' It has recently been demonstrated that ZOP pharmacokinetics are stereoselective: higher plasma concentrations of the (+) enantiomer were observed following oral administration of the racemic mixture to humans.Enantiomers of certain chiral compounds, such as oxazepam, 11*12 thalidomide, l3 hyoscyamine, l 4 scopolamine, l4 and thymidine glycol, l5 undergo racemization in vitro in aqueous or partially aqueous solutions. It has been shown that compounds such as thalidomide and nonsteroidal antiinflammatory agents can also racemize in vivo. 13*16 In vitro or in vivo racemization may complicate stereoselectivity studies. When studying the stereoselective kinetics of a chiral compound, it is essential to prove that racemization does not occur during storage or treatment of biological samples.Given the presence of a carbamate function on the asymmetric carbon, we investigated the stability and racemization of ZOP enantiomers and the racemic mixture in conditions usually employed for the study of ZOP pharmacokinetic stereoselectivity . EXPERIMENTAL Materials and MethodsSemipreparative chromatography. The HPLC system for semipreparative enantiomer separation consisted of an LC-6A pump (Shimadzu, Touzart et Matignon, Vitry, France), a Rheodyne 7125 injection valve equipped with a 1 ml loop, a Lambda-Max Model 480 UV detector (Waters, Mdford, USA), and a C-R6A integrator (Shimadzu). The column was a semipreparative Chiralcel OD (10 x 250 mm) (Daicel, J.T. Baker France, Paris, France). A 300 ~1 aliquot of freshly prepared ruc-ZOP solution in ethanol (2 mg/ml) was injected into the semipreparative column and eluted with ethanokhexane (6040, v/v) at a flow rate of 2 mumin.Mobile phase fractions containing (-)-ZOP or (+)-ZOP were separately evaporated to dryness under nitrogen. After reconstitution with ethanol, the purity of the separated enan-