Fractionation of [14C]Metolachlor Metabolites by Centrifugal Partition Chromatography

Metolachlor (2-chloro-N-(2′-ethyl-6′-methylphenyl)-N-(2-methoxy-l-methylethyl)acetamide) is one of the most widely used and important herbicides throughout the world. In order to understand the biodegradation pathways for this compound it is essential to develop large scale separation and purification procedures for its metabolites. However, analyzing crude plant extracts for these compounds poses several problems. In particular, the complex matrix and low levels of many metabolites requires a preparative separation and detection procedure that is both sensitive and robust. The use of [14C] labeled compounds and scintillation counting satisfies the sensitivity requirement but creates an additional problem when using preparative LC. Irreversible adsorption to the stationary phase frequently occurs when analyzing “real-world” biological samples or extracts thereof. This is particularly undesirable if the matrix contains radioactive components. This problem can be avoided by using a technique with a liquid stationary phase such as centrifugal partition chromatography (CPC). CPC is used for the preparative fractionation of [14C]metolachlor metabolites contained in crude corn plant extract. Also, a rapid analytical HPLC method is developed for the separation of standard non-radiolabeled metolachlor metabolites on a dimethylphenyl-derivatized β-cyclodextrin stationary phase.