Methylation of 3,4-methylenedioxymethamphetamine in formalin-fixed human liver tissue.

3,4-Methylenedioxymethamphetamine (MDMA or ecstasy) is a commonly consumed recreational drug. As is the case with most secondary amines, MDMA reacts with formaldehyde under acidic conditions to form tertiary amines. This reaction is likely to occur in formalin-fixed tissue. In formalin solutions, MDMA is methylated producing 3,4-methylenedioxy-N,N-dimethylamphetamine (MDDA). MDDA standard was synthesized by treating methylenedioxyamphetamine HCl in formaldehyde solution. Structure confirmation was by electrospray ionization-mass spectrometry (MS) and MS-MS. Randomly chosen human liver pieces (100-200 mg) were injected with 2 microg of MDMA HCl. The liver pieces in centrifuge tubes were covered with 200 microL of formalin solution (20% v/v), held at room temperature for 24 h, and then homogenized. The resulting suspension was sonicated for 5 min and then centrifuged. Controls consisted of substitution of 200 microL of water in place of formalin solution. Supernatant aliquots (10 mciroL) were added to 500 microL of 0.1% formic acid in acetonitrile for MS analysis. Positive ion electrospray spectra recorded in MS, MS2, and MS3 modes were used to confirm the presence of methylated MDMA. Liver tissue containing added MDMA HCl but not treated with formalin did not show a detectable level of methylated MDMA.