Analysis of carcinogenic heterocyofic amine-protein adduct as environmental and biological marker

迄今为止，仅从食物中分离出强致突变和致癌的杂环胺。随后，这些胺也被发现在一般环境中，如大气和河水等，因此，它是非常重要的，掌握这些胺的分布和动态相和生物效应的卫生化学观点。然而，人类和野生动物暴露于杂环胺的程度以及这些胺的水平是否安全尚未解决。本研究通过对生活环境中杂环胺含量的测定，特别是对作为饮用水和熟食供应源的河水的测定，了解杂环胺对野生动物和人类的污染情况。根据这些结果，根据流行病学技术计算了日本人的杂环胺估计摄入量。此外，还研究了杂环胺-蛋白质加合物作为暴露评价生物标志物的可能性。 ...更多信息 经气相色谱-氮磷检测器（GC-NPD）分析，在河水、熟鱼和熟肉中检出IQ、Trp-P-1、MeIQx和PhIP等。特别是在烤鱼中检测到高浓度的PhIP，并且其在皮中的含量高于肉。此外，基于肉类和鱼类摄入量调查的自我报告食物频率问卷，估计了来自四个地理区域的日本人的杂环胺每日个体摄入量水平。日本人主要服用PhIP，大陆受试者的摄入水平高于冲绳受试者。这些结果被认为是评价饮食习惯和致癌风险相关性的一个有效指标。其次，我们注意到杂环胺-蛋白质加合物可以直接估计暴露的杂环胺的量。青鳉鱼在杂环胺水溶液中繁殖一定时间后，通过提取蛋白质组分、酸水解、GC-NPD法测定蛋白质加合物的含量，建立了青鳉鱼蛋白质加合物的测定方法。当杂环胺暴露与蛋白质加合物形成之间的相关性使用这种方法进行检查时，加合物形成与暴露浓度和暴露时间成比例地增加。结果表明，杂环胺在持续摄入过程中积累，蛋白质加合物是一个有效的暴露指标。另一方面，已证实，在单次给予高剂量杂环胺时，可证实大鼠血液中形成蛋白加合物，但这些加合物在短时间内消失。此外，在低剂量重复给药中未观察到蛋白加合物蓄积。从器官分布检查来看，杂环胺-蛋白加合物主要在肝脏中检出，而在血液中未检出。根据这些结果，如果杂环胺总是存在于水中，则尽管蛋白质加合物在持续暴露的鱼中积累，但在主要通过膳食摄入杂环胺的动物（包括人）中难以积累。为了将血液中的蛋白质加合物作为人类杂环胺暴露的生物标志物，今后有必要开发除亚磺酰胺外的加合物的检测方法、更灵敏的检测方法和样品浓缩方法等。少

Powerful mutagenic and carcinogenic heterocyclic amines had been isolated from only food until now. Subsequently, these amines were also found in the general environment such as atmosphere and river water, etc. Therefore, it is very important to grasp the distribution and dynamic phase and biological effects of these amines for the hygiene chemistry viewpoint. However, it is unsolved how degree human and wildlife are exposed to the heterocydic amines, and whether the levels of these amines are safe. In this study, we measured heterocyclic amine contents in living environment, especially river water as supply source of drinking water and cooked foods in order to grasp heterocyclic amine pollution in the wildlife and human. From these results, an estimated intake of heterocyclic amine in Japanese was calculated based on the epidemiological technique. Furthermore, the heterocyclic amine-protein adduct as a biomarker for exposure evaluation was examined.By Blue-rayon extraction, derivatiza … More tion and GC-NPD analysis, IQ, Trp-P-1, MeIQx and PhIP, etc, were identified in river water, and cooked fish and meat. Especially, PhIP was detected in the grilled fish at high concentration, and it was included in the part of skin than that of flesh at high content. In addition, daily individual intake level of heterocyclic amine in Japanese was estimated for Japanese from four geographic areas based on a self-reported food frequency questionnaire for meat and fish intake survey. Japanese mainly took PhIP, and the intake levels for the subjects in the mainland were higher than those for the subjects in Okinawa. It is considered that these results become an effective index for evaluating the relevance to eating habit and carcinogenic risk.Next, we noticed the heterocyclic amine-protein adducts to directly estimate amount of the exposed heterocyclic amine. The method for measuring protein-adduct was established by extraction of protein fraction, acid hydrolysis and amount of adduct by the subsequent GC-NPD of the liberated amine, after it breeds the medaka in heterocyclic amine aqueous solution in selected time period. When the correlation between heterocychc amine exposure and the protein adduct formation was examined using this method, adduct formation increase in proportion to the exposure concentration and exposure period. The result suggest that heterocyclic amines are accumulated in the continually intake, and protein adduct is an effective index for exposure. On the other hand, it was proven that it could confirm protein-adduct formation in rat blood in the single administration of the high dose heterocyclic amine, but these adducts disappear short period. Furthermore, the accumulation of protein-adduct was not observed in low dose repetitive administration. From examination of organ distribution, the heterocyclic amine-protein adducts were mainly detected in liver, but not blood. From these results, it is difficult to be accumulated in the animal including the human who mainly takes the heterocyclic amine through the meal, though the protein adducts are accumulated in the fish continually exposed, if the heterocyclic amine always exists in the water. In the future, the development of measuring method of adducts except for sulfinamide which becomes original amine by acid hydrolysis, more sensitive detection method and samples concentration method, etc. seem to be necessary in order to use protein adducts in blood as biomarker for the heterocyclic amine exposure in human. Less