Study on Endocrine Disruption of Polycyclic Aromatic Hydrocarbons and Their Metabolites and Its Assay

近年来，国内外对柴油机尾气颗粒物（DEPE）及其成分多环芳烃（PAH）的雌激素和抗雌激素作用进行了研究。今年，我们对DEPE和PAH的雄激素和抗雄激素活性进行了检测。将DEPE应用于人PC3/AR培养细胞荧光素酶报告基因检测，DEPE无雄激素活性，但有抗雌激素活性。DEPE中含有的4环或5环多环芳烃具有较强的抗雄激素活性。DEPE和PAHs均为4环或5环，也是人烯丙基烃受体（AhR）的激动剂。这些结果表明，DEPE具有抗雄激素活性，其部分活性来源于DEPE中含有的多环芳烃。此外，几种硝基多环芳烃在酵母双杂交实验中显示出异体生殖活性。其次，为了增加单羟基多环芳烃（OHPAH）作为分析物的数量，对HPLC/荧光检测方法的条件进行了修改。为了分析尿液中OHPAHs及其偶联物，建立了水解偶联物的预处理方法。结果表明，吸烟者尿液中2-羟基芴的浓度高于非吸烟者，证明2-羟基芴是比1-羟基芴更有效的吸烟生物标志物。泰国受试者尿OHPAH浓度高于日本受试者。作为一个可能的原因，泰国房屋室内使用的木柴燃料产生的烟雾被认为是原因之一。

Last year, both estrogenic and antiestrogenic actions of diesel exhaust particulate(DEPE) and polycyclic aromatic hydrocarbon(PAH), which was one of the constituent of DEPE, were clarified. This year, both androgenic and antiandrogenic activities of DEPE and PAH were examined. By applying DEPE to the luciferase reporter gene assay in human PC3/AR culture cells, DEPE did not show androgenic activity but antiestrogenic activity. Several PAHs having 4 or 5 rings, which are included in DEPE, showed strong antiandrogenic activity. Both DEPE and PAHs having 4 or 5 rings, were also agonists of human allyl hydrocarbon receptor(AhR). These results suggested that DEPE showed antiandrogenic activity and that a part of this activity was derived from PAHs which were included in DEPE. In addition, several nitropolycyclic aromatic hydrocarbons showed aniandorgenic activity in the yeast two-hybrid assay. Next, to increase the number of monohydroxy polycyelic aromatic hydrocarbons(OHPAH) as analytes, conditions of our HPLC/fluorimetric detection method were modified. And, to anayze both urinary OHPAHs and their conjugates, the pretreatment method including hydrolysis of the conjugates was developed. As the result, the urinary concentration of 2-hydroxyfluorene in smokers' urine samples was higher than that in nonsmokers', and it was proven that 2-hydroxyfluorene was a more effective biomarker for smoking than 1-hydroxypyrene. Urinary OHPAH concentrations of Thai subjects were higher than those of Japanese. As a possible reason for this, the smoke of the firewood fuel used in the indoor of Thai house was considered.