Simultaneous determination of 2-phenylbenzotriazole-type mutagens, PBTA-1 through -8, in river water by liquid chromatography-tandem mass spectrometry.

We describe a method for the simultaneous determination of eight kinds of phenylbenzotriazole-type mutagens (PBTA-1, -2, -3, -4, -5, -6, -7 and -8) in river water based on liquid chromatography-tandem mass spectrometry (LC/MS/MS). The application of dopant-assisted atmospheric pressure photoionization (APPI) for the detection of the PBTAs was studied. The APPI technique provided higher PBTA signal intensities than those obtained with an electrospray ionization (ESI) source, and the APPI method was used for the determination of the PBTAs. A solid-phase extraction procedure was used for the extractions of PBTA-1 through -8 from river water. The procedure was rapid and the relative standard deviations were below 15%. The detection limits of PBTA-1 through -8 in river water using the proposed method were found to range from 0.04 to 0.5 ng L(-1) and PBTAs were successfully detected in river water at sub-ng L(-1) levels.

Use of a high-throughput umu-microplate test system for rapid detection of genotoxicity produced by mutagenic carcinogens and airborne particulate matter.

In the present study, we developed a rapid umu-microplate test system that uses the nitroreductase- and O-acetyltransferase-overproducing Salmonella typhimurium strain NM3009 and the O-acetyltransferase-overproducing S. typhimurium strain NM2009 to detect genotoxic activity in small volume samples. The assay was used to test the genotoxicity of several standard mutagens and environmental samples. Exponentially growing cultures of NM3009, NM2009, and the parental strain TA1535/pSK1002 were incubated in 96-well microplates with test chemicals both in the presence and in the absence of rat liver S9. The relative beta-galactosidase activities were then determined colorimetrically using either chlorophenol red-beta-D-galactopyranoside (CPRG) or O-nitrophenyl-beta-D-galactopyranoside (ONPG) as a measure of umuC gene induction activity. The sensitivities of NM3009 without S9 mix and NM2009 with S9 mix to nitroarenes and aromatic amines were up to 24- to 75-fold higher than those of the parent strain. Induction of umuC gene expression was detected more readily with CPRG than ONPG. The umu-microplate assay also detected genotoxicity in organic extracts of particulate matter from air samples collected in Osaka City, Japan. The pattern of the responses suggested that the genotoxic activity of the particulate extract was due primarily to nitrated polycyclic aromatic hydrocarbons. Our results indicate that the umu-microplate assay may be a useful way of carrying out rapid screens for genotoxicity in small-volume environmental samples.

Determination of aromatic amine mutagens, PBTA-1 and PBTA-2, in river water by solid-phase extraction followed by liquid chromatography-tandem mass spectrometry.

We describe a novel method for the determination of two kinds of aromatic amine mutagens, 2-[2-(acetylamino)-4-[bis(2-methoxyethyl)-amino]-5-amino-7-bromo-4-chloro-2H-benzotriazole (PBTA-1) and 2-[2-(acetylamino)-4-[bis(2-cyanoethyl)-ethylamino]-5-amino-7-bromo-4-chloro-2H-benzotriazole (PBTA-2), in river water based on liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS-MS). A solid-phase extraction procedure was used for the extraction of PBTA-1 and PBTA-2 from river water. The procedure was rapid and the relative standard deviations were below 4%. The detection limits of PBTA-1 and PBTA-2 in river water using the proposed method were found to be 1 and 2 ng/l, respectively. The compounds were detected by this method in river water taken from two sites in the Yodo River system at the ng/l level.

Detection of genotoxicity of atmospheric particles using a high-throughput microplate umu-test system.

A previously developed and highly sensitive umu-microplate test system based on the nitroreductase- and O-acetyltransferase-overproducing strain Salmonella typhimurium NM3009 and the O-acetyltransferase-overproducing strain S. typhimurium NM2009 was applied to the detection of genotoxic activity in atmospheric particles in urban areas using a relatively small sample load. The results showed that the test system was able to detect slight increases in induced genotoxicity in atmospheric particles and that genotoxicity was detected mainly in the fine fraction but also partially in the coarse fraction. The present sensitive microplate test system has potential for application to the screening of various other environmental samples.