Analysis of ecgonine and other cocaine biotransformation products in postmortem whole blood by protein precipitation-extractive alkylation and GC-MS

Ecgonine is formed by the hydrolysis of both ester linkages in cocaine and is of interest as both a potential metabolite and a postmortem and postcollection artifact in biological specimens. Its extreme polarity and water solubility make its isolation very difficult, particularly from complex matrices such as postmortem whole blood. This procedure describes an initial protein precipitation followed by two derivatization steps, the first of which is to propylate organic acids and primary and secondary amines and the second of which is to form the p-nitrobenzoyl ester of organic alcohols. The resultant derivatized products are then subjected to cleanup using a standard extraction with n-butyl chloride as the solvent. Analysis of the extracts is performed by gas chromatography-mass spectrometry using deuterated internal standards, although the adducts would also be suitable for analysis by high-performance liquid chromatography. Significant quantities of ecgonine have been identified in postmortem whole blood samples using this method.