Gölzer P, Janzowski C, Pool-Zobel B L, Eisenbrand G
Department of Chemistry, University of Kaiserslautern, Germany.
Chem Res Toxicol. 1996 Oct-Nov;9(7):1207-13. doi: 10.1021/tx9600107.
(E)-2-Hexenal (hexenal), a natural flavor compound, acts as directly genotoxic agent and forms cyclic 1,N2-propano adducts with deoxyguanosine. Formation of this adduct in isolated DNA and in cells was studied with a modified 32P-postlabeling procedure including HPLC separation, nuclease P1 enrichment, two-dimensional TLC of adducted nucleotide bisphosphates on PEI-cellulose, and quantification of adduct spots by liquid scintillation counting. Adduct formation with the more reactive crotonaldehyde was included for comparison. Synthesized adducted dG-3'-phosphates served as external standards for identification and quantification. In calf thymus DNA, hexenal (0.2 mM) shows a time dependent formation of adducts, yielding 1.55 pmol/mumol of DNA at 5 h incubation. With crotonaldehyde (0.2 mM) the adduct rate was about 10-fold higher. Hexenal also generated 1,N2-propano-dG adducts in the human lymphoblastoid Namalva cell line (0.2 mM, 1 h, 86 fmol/mumol of DNA) and in primary rat colon mucosa cells (0.4 mM, 30 min, 50 fmol/mumol of DNA). In primary colon mucosa cells from rats and humans, hexenal and crotonaldehyde (0.4 mM, 30 min) induced DNA damage, detected by single cell microgel electrophoresis (comet assay). In primary rat gastric mucosa cells, hexenal was only weakly active, inducing detectable DNA damage in 20% of cells at 0.8 mM concentration. In contrast, primary mucosa cells from rat esophagus were as sensitive as colon cells. After single oral application of hexenal to rats (up to 320 mg/kg body wt) DNA damage was not detectable in gastrointestinal mucosa. Analysis of hexenal in selected flavored foods revealed concentrations up to 14 ppm (0.14 mM) that are comparable to its natural occurrence in some fruits and vegetables (up to 30 ppm). Thus, the concentration range selected for the toxicological studies described here clearly is relevant: Hexenal, at concentrations found in food, exerts genotoxic effects in cells from rat and human gastrointestinal tract.
（E）-2-己烯醛（己烯醛）是一种天然风味化合物，作为直接的基因毒性剂，可与脱氧鸟苷形成环状1,N2-丙烷加合物。采用改良的32P后标记程序研究了该加合物在分离的DNA和细胞中的形成，该程序包括高效液相色谱分离、核酸酶P1富集、在聚乙烯亚胺纤维素上对加合的核苷酸二磷酸进行二维薄层层析，以及通过液体闪烁计数对加合物斑点进行定量。为作比较，还研究了与反应性更强的巴豆醛形成加合物的情况。合成的加合dG-3'-磷酸用作鉴定和定量的外标。在小牛胸腺DNA中，己烯醛（0.2 mM）显示加合物的形成呈时间依赖性，孵育5小时时产生的加合物为1.55 pmol/μmol DNA。对于巴豆醛（0.2 mM），加合速率约高10倍。己烯醛还在人淋巴母细胞Namalva细胞系（0.2 mM，1小时，86 fmol/μmol DNA）和原代大鼠结肠黏膜细胞（0.4 mM，30分钟，50 fmol/μmol DNA）中产生1,N2-丙烷-dG加合物。在大鼠和人类的原代结肠黏膜细胞中，己烯醛和巴豆醛（0.4 mM，30分钟）诱导DNA损伤，通过单细胞微凝胶电泳（彗星试验）检测到。在原代大鼠胃黏膜细胞中，己烯醛活性较弱，在0.8 mM浓度下仅在20%的细胞中诱导可检测到的DNA损伤。相比之下，大鼠食管的原代黏膜细胞与结肠细胞一样敏感。给大鼠单次口服己烯醛（最高320 mg/kg体重）后，在胃肠道黏膜中未检测到DNA损伤。对选定调味食品中己烯醛的分析显示，其浓度高达14 ppm（0.14 mM），与某些水果和蔬菜中的天然含量（高达30 ppm）相当。因此，此处所述毒理学研究选择的浓度范围显然具有相关性：在食品中发现的浓度下己烯醛对大鼠和人类胃肠道细胞具有基因毒性作用。
