A study of the distribution of 7-ethoxycoumarin O-de-ethylase activity in human placental subcellular fractions.

Human placental homogenates from maternal smokers and non-smokers were fractionated using differential centrifugation techniques. Yields of the various subfractions were determined and their homogeneity assessed using electron microscopic procedures. The distribution and response of 7-ethoxycoumarin O-de-ethylase activity towards inhibition by dimethylsulphoxide, alpha-naphthoflavone and 9-hydroxyellipticine inhibitors in the placental subfractions were investigated. The low yield of microsomal protein obtained following differential centrifugation of placental homogenates (2.5 +/- 0.2 mg protein per g placenta) highlights the extremely refractory nature of human placental tissue towards homogenization. Enzymic studies showed that the majority (75 per cent) of the original O-de-ethylase activity in homogenates from smokers and non-smokers was to be found in the crude nuclear fraction. The 7-ethoxycoumarin O-de-ethylase activity present in both homogenate and crude nuclear preparations from a maternal smoker was found to be inhibited by both alpha-naphthoflavone and 9-hydroxyellipticine to a lesser extent than the O-de-ethylase activity which was present in both mitochondrial and microsomal fractions. While this observation suggests the existence of more than one induced O-de-ethylase activity in the human placenta, the possibility that such differences in inhibitory response may be due to other factors (e.g. inhibitor solubility effects) cannot be excluded. Studies using the above inhibitors also confirmed the results of earlier work by demonstrating that the O-de-ethylase activity in placental homogenates and subfractions from non-smokers is qualitatively different from the O-de-ethylase activities induced as a result of maternal smoking.