嚼食檳榔者在世界各地約有6 億人口，它是繼吸煙、酒精及咖啡因之後全球第四大的
生活習性。2004 年IARC 已將單獨嚼食檳榔的行為(Betel quid without tobacco)及檳榔子
(Areca nut)皆列為人類致癌物Group 1 。儘管全球檳榔嚼塊的組成變化很大，檳榔子(Areca
nut)依然它們主要的共通點，其主成分中檳榔鹼(Areca nut alkaloids)及多酚類化合物
(Polyphenolic compounds)被視為重要致癌因子。儘管檳榔鹼的代謝對於研究檳榔暴露及其
潛在的致癌性十分重要；在過去的三十年來，人類對於檳榔鹼的代謝所知不多。直到今年
Giri et al. (2006)方才運用小鼠建立較完整的代謝地圖並證實N-methylnipecotic acid 是
arecoline 及arecaidine 兩者共同且最主要的代謝產物。檳榔鹼在體內可經亞硝化反應
(nitrosation) 形成檳榔特有亞硝胺(Areca-specific N-nitrosamines) ， 其中以
3-(N-nitrosomethylamino)propionitrile (NMPN)的致癌性最強，它經代謝後可造成DNA 甲基
化 (methylation)。而檳榔特有亞硝胺的主要代謝物係為N-nitrosonipecotic acid。此外，多酚
類成份在添加石灰的鹼性環境中(pH＞9.5)，很容易產生反應性含氧物質(Reactive oxygen
species, ROS)；這會在體內造成基因氧化傷害，而8-OxodGuo 為主要的基因氧化性傷害產
物。儘管上述生物偵測指標對於檳榔子成分的暴露或致癌機制研究深具意義但生物偵測方
法缺乏，尤其是針對檳榔鹼及檳榔特有亞硝胺的相關研究。在台灣大部分的嚼食檳榔者都
有抽煙，吸煙行為同樣也會造成DNA 的甲基化及氧化傷害。因此要釐清DNA 損傷與不同
暴露源的關連性，必須也要同時建立吸煙的暴露生物偵測指標，亦即尿液中nicotine 及其代
謝物cotinine 的測定。本計畫的目的在於運用連線固相萃取(On-line solid phase extraction)
搭配液相層析串聯質譜儀(LC-MS/MS)開發快速、靈敏、精準的檳榔暴露及效應生物偵測方
法，並運用於嚼食檳榔者之尿液分析，期以分子的角度探究檳榔對人體的潛在危害。
It has been estimated that, world wide, ~600000000 people chew areca nut. IARC (2004) has
evaluated betel quid without tobacco as causing oral cancer and has stated that there is sufficient
evidence in experimental animals for the carcinogenicity of areca nut. Arecoline and arecaidine
are reported to be the most abundant alkaloids in the areca nut. However, little is known about the
metabolism and disposition of arecoline and arecaidine in humans or in animals, until very
recently Giri et al. (2006) have reported the metabolic map of arecoline and arecaidine in the
mouse. The major metabolite of both arecoline and arecaidine appear to be N-methylnipecotic
acid. In the meantime, chemical nitrosation of arecoline leads to the formation of areca-specific
N-nitrosamines (ASNAs) such as 3-(N-nitrosomethylamino)propionitrile, N-nitrosoguvacoline
and N-nitrosoguvacine. The α-hydroxylation of these N-nitrosamines is an important activation
pathway mediated mainly by P450s. This pathway yields alkyldiazohydroxide ions which
alkylate DNA bases. It has been also reported that a major urinary metabolite of
N-nitrosoguvacoline and N-nitrosoguvacine was identified as N-nitrosonipecotic acid, which
could be used as a marker of ASNAs exposure. In addition to areca nut alkaloids, areca nut
polyphenols under alkaline conditions can release reactive oxygen species (ROS), which are
capable of inducing nucleotides modification and the formation of
8-oxo-7,8-dihydro-2’-deoxyguanosin (8-oxodGuo).
Taken together, although these biomarkers play important roles for assessing exposures and
risk from areca nut chewing, little effort has been made to develop analytical methods.
Therefore, the aims of this project are to firstly develop a serial “on-line SPE LC-MS/MS
methods” for sensitive and specific analysis of (i) arecoline, arecaidine and N-methylnipecotic
acid (biomarkers for areca alkaloids exposure); (ii) nicotine and cotinine (biomarkers for
cigarette smoke exposure) in body fluids.
