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

作为环境和生物标志物的致癌杂环胺蛋白加合物的分析

• 批准号: 12672171
• 负责人: KATAOKA Hiroyuki
• 金额: $ 1.47万
• 依托单位: OKAYAMA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2000
• 资助国家: 日本
• 起止时间: 2000 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12672171/
• 关键词: heterocyclic amine; protein adduct; biomarker; medaka; carcinogenicity; NPD-gas chromatogranhy; river water; epidemiological analysis

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

迄今为止，人们仅从食物中分离出具有强致突变性和致癌性的杂环胺。随后，在大气、河水等一般环境中也发现了这些胺。因此，从卫生化学的角度把握这些胺的分布、动态相和生物效应是非常重要的。然而，人类和野生动物在多大程度上暴露于杂环胺，以及这些胺的水平是否安全，都是没有解决的问题。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来源的河水进行测定，以掌握杂环胺在野生动物和人体中的污染情况。根据这些结果，利用流行病学技术计算了日本人杂环胺的估计摄入量。此外，杂环胺-蛋白加合物作为暴露评估的生物标志物进行了研究。通过Blue-rayon萃取、衍生化、GC-NPD分析、IQ、Trp-P-1、MeIQx、PhIP等方法对河水、熟鱼、肉进行了鉴定。特别是在烤鱼中检测到高浓度的PhIP，且PhIP在鱼皮部分的含量高于肉的含量。此外，根据对肉类和鱼类摄入的自我报告食物频率问卷调查，对来自四个地理区域的日本人的杂环胺的每日个人摄入量进行了估计。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有效指标。其次，我们注意到杂环胺-蛋白质加合物，以直接估计暴露的杂环胺的量。建立了测定蛋白质加合物含量的方法，在选定的时间内，在杂环胺水溶液中培养水母菌，通过提取蛋白质部分、酸水解和释放胺的GC-NPD进行加合物量的测定。当用该方法检测杂己胺暴露与蛋白质加合物形成的相关性时，加合物形成与暴露浓度和暴露时间成正比。结果表明，杂环胺在持续摄入过程中积累，蛋白质加合物是有效的暴露指标。另一方面，证明单次给药大鼠血液中可以形成蛋白质加合物，但这些加合物在短时间内消失。此外，在低剂量重复给药中没有观察到蛋白质加合物的积累。从脏器分布检查来看，杂环胺蛋白加合物主要在肝脏中检出，而在血液中未检出。从这些结果可以看出，如果杂环胺一直存在于水中，虽然蛋白质加合物在持续暴露的鱼体内积累，但在主要通过膳食摄取杂环胺的动物包括人体内很难积累。为了将血液中的蛋白质加合物作为人体杂环胺暴露的生物标志物，未来有必要发展除亚胺经酸水解成为原始胺的亚胺外的加合物的测定方法、更灵敏的检测方法和样品浓度法等。少

项目成果

Minatsu Kobayashi: "Estimation of dietary HCA intakes in a large-scale population-based prospective study in Japan"Mutation Research. (in press). (2002)

Minatsu Kobayashi：“日本大规模人群前瞻性研究中膳食 HCA 摄入量的估算”突变研究。

Hiroyuki Kataoka, et al.: "Selective detectors with gas chromatography for environmental amine analysis"Encyclopedia of Analytical Chemistay : Instrumentation and Applications. 3. 2587-2618 (2000)

Hiroyuki Kataoka 等人：“用于环境胺分析的气相色谱选择性检测器”分析化学百科全书：仪器和应用。

Yukari Totsuka: "In vitro and in vivo formation of aminophenylnorharman from norharman and aniline"Mutation Research. (in press). (2002)

Yukari Totsuka：“在体外和体内从去甲哈尔曼和苯胺形成氨基苯基去哈尔曼”突变研究。

Yukari Totsuka, et al.: "In vitro and in vivo formation of aminophenylnorharman from norharman and aniline"Mutation Research. (in press). (2002)

Yukari Totsuka等人：“在体外和体内从去甲哈尔曼和苯胺形成氨基苯基去哈尔曼”突变研究。

Hiroyuki Kataoka, et al.: "Analysis of mutagenic heterocyclic amines in cooked food samples by gas chromatography with nitrogen-phosphorous detector"Bulletin of Environmental Contamination and Toxicology. 64 (in press). (2002)

Hiroyuki Kataoka 等人：“用氮磷检测器气相色谱法分析熟食样品中的致突变杂环胺”环境污染与毒理学通报。