The major side effects of racemic oxybutynin (OXY), which is used in the treatment of urinary incontinence are dry mouth (xerostomia) and blurred vision (mydriasis). Highly purified enantiomers of OXY [(R)OXY, (S)OXY] were compared with the racemate both in vitro in functional studies and in vivo in guinea pigs to evaluate their pharmacological action relative to their adverse effects. The affinity of (R)OXY and (S)OXY for different muscarinic receptor subtypes was determined using field stimulated rabbit vas deferens (M1) and guinea pig atria (M2) or bladder (M3) strips. Stereoselective antimuscarinic effects [(R)OXY greater than or equal to (R/S) OXY much greater than (S)OXY] were evident at all three receptor subtypes; the isomeric ratio [(S)OXY/(R)OXY] ranged from 12 to 88. Both (R)OXY and (R/S)OXY were slightly more selective (2-4-fold, P less than .01) for M1 and M3 relative to M2 muscarinic receptors. Stereoselectivity was also evident in vivo for volume-induced urinary bladder contractions as measured by cystometrogram parameters [(S)OXY/(R)OXY approximately 21], mydriasis [(S)OXY/(R)OXY approximately 136] and salivary gland secretory responses [(S)OXY/(R)OXY approximately 30]. The absolute potencies of (R)OXY or (R/S)OXY for mydriasis and salivation were similar to those for inhibition of intravesical bladder pressure. Also, (R)OXY and (R/S)OXY equipotently antagonized cholinergic-mediated CNS effects in mice. Collectively, the data suggest that the activity of (R/S)OXY resides predominantly in the (R)-enantiomer. However, it appears that (R)OXY may offer no significant pharmacological advantage over (R/S)OXY in terms of its principal therapeutic and side effect profile.