Osamu Endo

Collaborative study using the preincubation Salmonella typhimurium mutation assay for airborne particulate matter in Japan. A trial to minimize interlaboratory variation

A collaborative study has been performed over a period of 3 years to develop a suitable method for monitoring the mutagenicity of airborne particulate matter. The study was organized with 8 laboratories and performed in the following steps: (1) selection of a suitable technique for each process involved in the mutagenicity monitoring, (2) developing a tentative protocol by combining systematically the selected techniques, (3) evaluation of the protocol by intra- and inter-laboratory studies, (4) modification of the protocol according to the evaluation, and (5) evaluation of the modified protocol by conducting an interlaboratory study. We found a suitable method for mutagenicity monitoring of particles in the atmosphere. Airborne particles were sampled with a high-volume sampler, the samples were stored at -80 degrees C, extracted by sonication using dichloromethane, solvent-exchanged, and assayed by the preincubation method using Salmonella typhimurium TA98 and TA100. The observed mutagenic activity was normalized with that of an internal standard. Round robin tests revealed that the method resulted in excellent reproducibility. The coefficient of variation for mutagenic activities of airborne particulate samples collected in various districts of Japan were in the range of 14.7 +/- 6.6% to 19.6 +/- 4.0% for strains TA98 and TA100 with and without metabolic activation. We also found that the plate incorporation method was equivalent to the preincubation method for airborne particulate extracts.

Estimation of Indoor Air PAH Concentration Increases by Cigarette, Incense-Stick, and Mosquito-Repellent-Incense Smoke

Abstract Smoke from cigarette smoking and burning of incense sticks and mosquito-repellent incense, which is particularly used in summer, are thought to be primary factors of indoor air pollution by polycyclic aromatic hydrocarbons (PAHs) in Japanese houses. In this study, these respective smokes as indoor air pollutants were evaluated by quantifying their particulate matter (PM) and 7 PAHs. PM and PAHs in smoke from 5 popular brands of Japanese cigarettes were collected on the glass fiber filter by the international smoking mode. PM was quantified by weighing the filter before and after trapping them. PAHs were analyzed by HPLC after extracting the filter with ethanol/benzene (1:3, by vol.). Similarly, evaluation of smoke respectively generated by natural burning of 4 brands of incense sticks and 3 brands of mosquito-repellent incenses was also conducted. As the results of these experiments, the average concentrations of PM and benzo[a]pyrene in a 35 m3 closed room were estimated to increase 1630 μg/m3 and 8.2 ng/m3, respectively, by smoking of 3 cigarettes; 1320 μ/m3 and 4.1 ng/m3, respectively, by burning of 2 incense sticks; and 2510 μg/m3 and 17.3 ng/m3, respectively, by burning of 8 cm of mosquito-repellent incense.

Mutagenicity of alkylhydrazine oxalates in Salmonella typhimurium TA100 and TA102 demonstrated by modifying the growth conditions of the bacteria.

Alkylhydrazines are important carcinogens. However, they show generally only weak mutagenicity and the activities reported from different laboratories are contradictory. We have developed a sensitive method to detect the mutagenicity of alkylhydrazines. The method is based on a modified preculturing procedure in the Ames test, the emphasis in the modification being a change in the growth period of tester strains. The optimal growth periods were found to be 11 h in Salmonella typhimurium TA100 and 5 h in Salmonella typhimurium TA102. We tested the mutagenic activity of 12 alkylhydrazines; 1,2-dimethylhydrazine, 1,2-diethylhydrazine, 1,2-dipropylhydrazine, 1,2-dibutylhydrazine, 1,1-dimethylhydrazine, 1,1-diethylhydrazine, 1,1-dipropylhydrazine, 1,1-dibutylhydrazine, methylhydrazine, ethylhydrazine, propylhydrazine and butylhydrazine. All 12 alkylhydrazines were clearly mutagenic in Salmonella typhimurium TA102, and 10 hydrazines were mutagenic in Salmonella typhimurium TA100, both in the absence of S9 mix. The mutagenicity was inhibited by the addition of S9 mix or bovine serum albumin. This suggests deactivation of the mutagens by proteins.

Human Exposure to Airborne Mutagens Indoors and Outdoors Using Mutagenesis and Chemical Analysis Methods

The lung cancer mortality rate has been steadily increasing in industrialized countries of the world. There are many risk factors for the lung cancer induction in our environments. Airborne carcinogens and mutagens are considered to be one of the major risk factors. However, quantitative contribution of airborne carcinogens and mutagens to the lung cancer induction has not yet been accurately evaluated, because of the lack of data on long-term human exposure to these chemicals in many areas which are in different pollution levels. For this purpose, there is an urgent need to develop methodologies suitable for monitoring long-term exposure to these carcinogens and mutagens.

Broad range observation of particle deposition on greased and non-greased impaction surfaces using a line-sensing optical microscope

Abstract A computer-automated optical microscope combined with a line-sensing camera was used to capture the entire range of a particle deposit downstream of an individual acceleration nozzle in a hi-volume Andersen sampler. To investigate the particle bounce and reentrainment, particles collected on collocated greased and non-greased Teflon plates on the inlet stage (d a >7 μm ) were observed by an automated particle counting, locating and sizing method. The result confirmed reproducible collection characteristics among nozzles even though application of the grease increased collection efficiency and altered the size distribution of collected particles to the larger side. In these experiments, assuming spherical particles with uniform density, approximately 65% of particle mass and 50% by number were lost from non-greased plates at 54% RH, while 45% by mass and 25% by number were underestimated at 84% RH. The spatial investigation showed that particles were densely deposited around the center of deposition on greased plates while on non-greased plates they were dispersedly distributed. Particle dispersions on the smooth impaction plate were due to bounce and/or reentrainment of small particles especially with d PA μm .

TEMPORAL VARIATION OF RADIOCARBON CONCENTRATION IN AIRBORNE PARTICULATE MATTER IN TOKYO

The temporal radiocarbon variation (in terms of percent Modern Carbon: pMC) of size-fractionated airborne particulate matter (APM) collected in Tokyo between April 2002 and February 2003 was analyzed in order to get an insight into the sources of carbonaceous particles. Results indicated significant biogenic origins (approximately 40 pMC on average). In general, the seasonal and particle size variations in pMC were relatively small, with 2 exceptions: elevated pMC in coarse particles in April and October 2002, and relatively low pMC in the finest particle size fraction collected in August 2002. The former finding could be tentatively attributed to the abundance of coarse particles of biological origins, such as pollen; the latter might be due to an increased fraction of anthropogenic secondary particles.

Results of a comparative study on the Salmonella pre-incubation and plate incorporation assays using test samples from the IPCS collaborative study.

Three complex mixtures (air particles, diesel particles and a coal tar fraction) and two pure compounds (benzo[a]pyrene and 1-nitropyrene) were tested in both the pre-incubation and the plate incorporation assay employing Salmonella typhimurium TA98 and TA100. Each experiment was conducted independently 2 or 4 times in duplicate in the presence and absence of metabolic activation. The mutagenic activities were calculated by least squares linear regression from the slope of the linear portion of each dose-response curve. Although slightly higher mutagenic activity was observed in the pre-incubation assay for the two pure compounds and with the plate incorporation assay for the diesel particulate sample, the overall data from both assays gave similar values and good correlations in TA100 and TA98. The results indicate that the pre-incubation assay could be used for these samples instead of the plate incorporation assay.

Mutagenic activities of a chlorination by-product of butamifos, its structural isomer, and their related compounds.

The mutagenic activities of 5-methyl-2-nitrophenol (5M2NP), a chlorination by-product of butamifos, its structural isomer 2-methyl-5-nitrophenol (2M5NP), and related compounds were evaluated by the Ames assay. The mutagenic activities of 5M2NP and 2M5NP were negative or not particularly high. However, those of their chlorinated derivatives were increased in Salmonella typhimurium strain TA100 and the overproducer strains YG1026, and YG1029 in the absence and/or presence of a rat liver metabolic activation system (S9 mix), particularly for YG1029. The mutagenic activities of 6-chloro-2-methyl-5-nitrophenol (6C2M5NP) in YG1029 in the absence and presence of S9 mix were 70000 and 110000 revertants mg(-1), respectively. When nitro functions of 6C2M5NP and 4-chloro-5-methyl-2-nitrophenol (4C5M2NP) were reduced to amino functions, their mutagenic activities were markedly decreased. The mutagenic activities of 5M2NP and 4C5M2NP were lower than those of 2M5NP and 6C2M5NP, respectively. Thus, it was shown that substituent position is a key factor for the mutagenic activities of methylnitrophenols (MNPs) and related compounds. The mutagenic activities of the extracts of 2M5NP in chlorination increased early during the reaction time and then decreased. The main chlorination by-product contributing to the mutagenic activities of the extracts of 2M5NP in chlorination was 6C2M5NP. The results of chlorination of 2M5NP suggested that MNPs were present as their dichlorinated derivatives or further chlorination by-products in drinking water.

POLYCYCLIC AROMATIC HYDROCARBON CONCENTRATIONS OF AIRBORNE PARTICLES IN URBAN AIR OVER THE PAST TWENTY YEARS

To estimate human exposure to hazardous chemicals, it is important to measure the annual trends regarding the concentrations of these compounds in the environment. Airborne particulate samples were collected over the past 20 years (1980–2001) from ambient air using a quartz fiber filter with the help of a high-volume air sampler placed on the top of the National Institute of the Public Health building, Minato-ku, Tokyo, Japan. In the present study, portions of these filter samples were analyzed to investigate the trends regarding the concentration of PAHs in urban air over the sampling period. The concentrations of various PAHs per unit air volume were determined. The concentrations of seven PAHs decreased within the sampling periods, especially in the late 1980s, and slowly decreased from the middle 1990s. The concentrations of seven PAHs were higher in autumn and winter than in the spring and summer.

GAS/PARTICLE AND SIZE DISTRIBUTIONS OF POLYCYCLIC AROMATIC HYDROCARBONS IN AMBIENT AIR IN TOKYO 2001–2002

By using an Andersen sampler equipped with a low-pressure impactor, samples of 12 size-classified (>0.13 μm to <12 μm) airborne particles and samples of gaseous components were taken from the air in Tokyo for continuous periods of 19 weeks in the summer of 2001 and 17 weeks in the winter of 2001–2. The sampling filters were changed weekly. The concentrations of eight polycyclic aromatic hydrocarbons (PAHs) in the particulate and gas-phase samples were measured by reverse-phase high performance liquid chromatography (HPLC) with fluorescence detection. Pyrene was detected in the gas phase in both summer and winter: 59% of the total pyrene detected was present in the gas phase in summer, but this fraction decreased to 40% in winter. In the particle fractions, the summer levels of benzo[k]fluoranthene (BkF), dibenz[a,h]anthracene (dBahA), and benzo[a]anthracene (BaA) peaked in particles of diameter 1.25 μm, and benzo[ghi]perylene (BghiP), benzo[a]pyrene, benzo[b]chrysene (BbC), and dibenzo[a,e]pyrene (dBaeP) peaked in particles of diameter 0.76 μm. In winter, BkF, BghiP, BaA, BbC, and dBaeP levels peaked in particles of diameter 0.52 μm, whereas dBahA peaked in particles of diameter 0.76 μm.