| Neonatal abstinence syndrome (NAS) is defined as neonatal withdrawal symptoms due to antepartum maternal substance abuse. Substances that are most likely to cause NAS are opioids, alcohol and central nervous system (CNS) stimulants or depressants, such as cocaine and benzodiazepines, respectively. Opioid addiction in adults as well as neonates is treated with methadone or buprenorphine. In severe maternal addiction and neonatal withdrawal cases, morphine is administered as an additive to methadone or buprenorphine. Mothers who are on methadone or buprenorphine treatment are often found to have plasma traces of alprazolam, a common benzodiazepine prescribed for the treatment of anxiety. However, the in vitro metabolism of these drugs, either alone or when taken together, has never been well established in fetuses or neonates. Here, we analyze the activity of methadone, buprenorphine, morphine, and alprazolam in fetal, neonate and adult female livers. Since the cytochrome P450 3A enzymes (CYP3A4 and CYPA5) are common and major routes of metabolism for these drugs, the resulting metabolite of each can be measured to assess the activity. This was done by in vitro incubations with human liver microsomes (HLM) from adult females (20-75 years old) and neonates (male and female, 13 days -- 11 months old). The metabolites were identified by high-performance liquid chromatography (HPLC). We found there was no significant difference in alprazolam, buprenorphine or methadone metabolite formation rate between adults and neonates. Morphine showed inhibition of alprazolam metabolite formation at high concentrations in both female adult and neonatal liver microsomes. We also saw high variability between the individual microsomes and performed a multiple linear regression analysis to determine if age, CYP content, and alcohol or cigarette use could be responsible for this variability. From these analyses, CYP content was a significant variable for predicting metabolite formation in all groups except neonates in 4-hydroxyalprazolam (4-OHALP) and 2-ethylidene-1,5-dimethyl-3,3-dipehnylpyrrolidine (EDDP) production. Additionally, norbuprenorphine (NBUP) and EDDP formation rate was partially related to alcohol use in adult females. From these results, we can conclude: 1) the metabolite formation rate differences of 4-OHALP, BUP, and EDDP between adult females and neonates are not predictors of neonatal abstinence syndrome onset, but in vitro studies with fetal microsomes still need to be done and 2) the interaction between morphine and alprazolam occurs at a concentration too high to be of pharmacokinetic importance, but there may be an underlying pharmacodynamic mechanism leading to additive effects. |
