Cocaine (COC) is rapidly hydrolyzed to benzoylecgonine (BE), a major vasoactive metabolite measured in screening infants for prenatal exposure to COC. Recently, detection of meta-hydroxy benzoylecgonine (m-OH BE) has been suggested to improve identification of these infants. However the pathway of m-OH BE formation is unknown. This study was designed to determine if m-OH BE is a late metabolite derived directly from COC or via BE. Pregnant Dunkin-Hartley guinea pigs at 63 d gestation were injected i.p. with 10 mg/kg COC or BE. Amniotic fluid (AF) and meconium (MEC) from each fetus were collected upon sacrifice at 24 and 48 h post-cocaine; and 24, 48, and 72 h post BE injection. COC metabolites were quantitated by gas chromatography-mass spectroscopy. After maternal COC injection, recovery of COC in AF (75%) and MEC (88%) decreased from 24 (n=12) to 48 h (n=9). While BE levels post-COC injection declined in both AF (58%) and MEC (69%) from 24 (n=11) to 48 h(n=7), m-OH BE recovery in MEC increased (1600%). Similarly, after maternal BE injection, mean recovery of BE decreased from 24 (n=11) to 48 h (n=7) in both MEC (69%) and AF (64%), while m-OH BE recovery increased (633%) in MEC. By 72 h (n=11), BE recovery decreased another 85% in both AF and MEC, however m-OH BE was still present in MEC and unchanged from 48 h. There was no significant recovery of m-OH BE in AF after COC or BE injection. Colon from BE exposed fetuses were independently analyzed to determine if m-OH BE is formed in MEC or released into MEC. BE was present at 24 h and levels were undetectable by 72 h, however no m-OH was found. These results indicate that 1) m-OH BE is a metabolite of BE rather than of COC; and 2) levels of m-OH BE may be detectable longer in MEC than BE. Finally, failure to test for m-OH BE in MEC may result in underreporting of cocaine exposure. Funded by: Kapiolani MC& Children's Miracle Network, US Army HSC.No financial support received from US Drug Testing Labs.