USE OF NARROW-BORE HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY DIODE-ARRAYDETECTION FOR THE ANALYSIS OF INTERMEDIATES OF THE BIOLOGICAL DEGRADATION OF 2,4,6-TRINITROTOLUENE

A single method was developed for the separation and quantitation of h exahydro-1,3,5-trinitro-1,3,5-triazine, 2,4,6-trinitrotoluene (TNT), a nd most of the known and suspected biodegradation intermediates of TNT by RP-HPLC and diode array detection. The known biodegradation interm ediates of TNT analyzed were 2-amino-4,6-dinitrotoluene, 4-amino-2,6-d initrotoluene, 2,6-diamino-4-nitrotoluene, 2,4-diamino-6-nitrotoluene, 2,4,6-triaminotoluene, 2,2',6,6'-tetranitro-4,4'-azoxytoluene, and 4,4 ',6,6'-tetranitro-2,2'-azoxytoluene. The suspected biodegradation inte rmediates of TNT included 1,2,3-benzenetriol (pyrogallol), 1,3,5-benze netriol (phloroglucinol), 2-methyl-1,3 ,5-benzenetriol (methyl phlorog lucinol) and 4 methylphenol (p-cresol). Mobile phases consisting of aq ueous buffers adjusted to three different pH values in a gradient with acetonitrile were examined for their efficiency in separating the int ermediate compounds and for the minimization of speciation of the ioni zable intermediates (e.g. 2,4,6-triaminotoluene). A final aqueous buff er pH of 3.2 was selected to minimize the interference to the separati on caused by 2,4,6-triaminotoluene speciation. Solvent consumption was minimized by the use of a narrow-bore column. All of the known reduct ion products as well as p-cresol and methylphloroglucinol were identif ied in culture supernatants from TNT-degrading cultures while pyrogall ol and phloroglucinol were not.