DeMarini D M, Shelton M L, Bell D A
Environmental Carcinogenesis Division, U.S. Environmental Protection Agency, Research Triangle Park, NC 27711, USA.
Mutat Res. 1996 Jan 17;349(1):1-20. doi: 10.1016/0027-5107(95)00074-7.
Using an ion-exchange procedure coupled to a microsuspension Salmonella assay, we fractionated the dichloromethane-extractable particulate organics emitted by a municipal waste incinerator. Most (80-95%) of the mutagenic activity resided in the neutral/base fraction; however, the polar neutral fraction accounted for 12% of the direct-acting mutagenic activity. The mutagenic potencies of the whole extract and the various fractions were 4-15 times greater in the absence than in the presence of S9. Results with strains deficient in classical nitroreductase (TA98NR) and transacetylase (TA98/1,8-DNP6) indicated that a majority of the direct-acting mutagenicity was due to nitroarenes. This was confirmed by bioassay-directed subfractionation of the neutral/base faction by a cyanopropyl/HPLC method. The mutations in -3,000 revertants (approximately 400 each induced in TA98 by the whole extract, the neutral/base and polar neutral fractions from the ion-exchange column and 3 of the neural/base subfractions from the HPLC column; along with 200 revertants each induced by the model nitroarene 1-nitropyrene (1NP) in strains TA98, TA1538 and TA100) were analyzed by probe hybridization and PCR/DNA sequence analysis. The results indicated that nitroarenes such as 1NP that eluted in the neutral/base fraction accounted for at least 50% of the direct-acting mutagenicity and induced only a hotspot 2-base deletion in the sequence (CG)4 in TA98. In contrast, most of the complex frameshifts (a frameshift with a flanking base substitution) induced by the whole extract were induced by nitroarenes other than 1NP that were activated by transacetylation and that eluted in the polar neutral fraction. This study (1) identifies nitroarenes as an important contributor to the mutagenic activity of the emissions from municipal waste incinerators; (2) confirms our previous conclusion that the mutation spectrum of a complex mixture reflects the dominance of particular classes of chemical mutagens within the mixture; and (3) demonstrates the possibility of isolating certain chemical fractions of a complex mixture that induce certain classes of mutations produced by the whole, unfractionated mixture.
我们采用离子交换程序结合微悬浮沙门氏菌试验，对城市垃圾焚烧炉排放的二氯甲烷可萃取颗粒有机物进行了分级分离。大部分（80 - 95%）的诱变活性存在于中性/碱性组分中；然而，极性中性组分占直接作用诱变活性的12%。在无S9时，整个提取物和各个组分的诱变效力比有S9时大4 - 15倍。对缺乏经典硝基还原酶（TA98NR）和转乙酰酶（TA98/1,8 - DNP6）的菌株的研究结果表明，大部分直接作用诱变活性是由硝基芳烃引起的。这通过氰丙基/HPLC方法对中性/碱性组分进行生物测定导向的亚分级分离得到了证实。对约3000个回复突变体（整个提取物、离子交换柱的中性/碱性和极性中性组分以及HPLC柱的3个中性/碱性亚组分在TA98中各自诱导产生约400个；模型硝基芳烃1 - 硝基芘（1NP）在TA98、TA1538和TA100菌株中各自诱导产生200个）中的突变进行了探针杂交和PCR/DNA序列分析。结果表明，在中性/碱性组分中洗脱的硝基芳烃如1NP至少占直接作用诱变活性的50%，并且在TA98中仅在序列（CG）4中诱导一个热点2 - 碱基缺失。相比之下，整个提取物诱导的大多数复杂移码突变（伴有侧翼碱基取代的移码突变）是由1NP以外的硝基芳烃引起的，这些硝基芳烃通过转乙酰化被激活并在极性中性组分中洗脱。本研究（1）确定硝基芳烃是城市垃圾焚烧炉排放物诱变活性的重要贡献者；（2）证实了我们之前的结论，即复杂混合物的突变谱反映了混合物中特定化学诱变剂的主导地位；（3）证明了分离复杂混合物中某些化学组分的可能性，这些组分可诱导未分级的整个混合物产生的特定类型的突变。
