Selective antibodies to methadone enantiomers: Synthesis of(R)- and (R,S)-methadone conjugates and determination by an immunoenzymatic method in human serum

Selective antibodies to (R)-methadone (Mtd) and to its racemate were produc ed in rabbits by immunization with conjugates of (R)- or (R,S)-hemisuccinyl methadol-bovine serum albumin, respectively. A hapten was first prepared by reduction of (R)- or (R,S)-Mtd with sodium borohydride, followed by esteri fication with succinic anhydride. The conjugation of hapten with albumin wa s achieved by the mixed anhydride method. After immunization of rabbits, th e titers and specificity of each antibody were determined by ELISA. The ant ibodies obtained were tested with (R)-, (S)-, (R,S)-Mtd, its major metaboli te (EDDP), and some drugs of abuse (morphine, codeine, cocaine). The sensit ivities of antibodies to (R)- and (R,S)-Mtd were about 1 and 2 ng/ml, respe ctively. Selective (R)-antibodies recognized (R)-Mtd about 40 times more av idly than the (S)-isomer, while an antiserum against (RS)-Mtd recognized (R )- and (S)-isomers to about the same degree. Both selective antibodies show ed little interference (about 0.5%) with EDDP metabolite and no crossreacti vity with morphine, codeine, and cocaine. These two selective antibodies we re used to develop an immunoenzymatic method (ELISA) for the determination of (R)- and (R,S)-Mtd in serum samples of patients under maintenance treatm ent for narcotic addiction. Chirality 13:187-192, 2001. (C) 2001 Wiley-Liss , Inc.