- 學位論文
以核磁共振為主的代謝體學探討順丁烯二酸重複暴露下對於大鼠之影響
1H NMR-based Metabolomics to Study Repeated Exposure to Maleic Acid in Rats
摘要
順丁烯二酸 (Maleic acid)，過去常用於樹脂原料、殺蟲劑與工業黏著劑的原料。2013年，台灣發生的「毒澱粉事件」就是在市售澱粉製品中檢測出違法的食品添加物順丁烯二酸，其製品包括黑輪、肉圓、米粉和粉圓等等。添加目的是為了提升澱粉的穩定性、口感和降低成本，而在食用的化製澱粉上，我國核准的有21種，但不包含順丁烯二酸。過去在動物實驗上，順丁烯二酸的暴露會對腎臟造成影響，出現范可尼氏症候群 (Fanconi syndrome)的症狀，影響近端小管的再吸收功能和腎臟的酸鹼平衡，造成有磷酸鹽尿 (Phosphaturia)和糖尿 (Glucosuria)的產生。然而，長期暴露下是否會對其他器官造成影響與在體內的影響機制能有待確認。 本次的研究對象為Sprague-Dawley (SD) 品系的大鼠，依據暴露劑量不同分為四個組:低 (6 mg/kg)、中 (20 mg/kg)、高 (60 mg/kg)和控制組。在生物監測的部份，採集特定時間點早晚的尿液(第0、7、14、21和28天)，並利用600 MHz 的核磁共振儀來獲得大鼠尿液中的代謝物profile。最後使用主成分分析、單變量無母數分析多樣本中位數差異檢定和Dunn式事後檢定來分析代謝物和暴露劑量之間的關係。 研究結果顯示，高劑量暴露下與控制組的代謝體差異有所變化，而分界點從14天之後有明顯不同，暴露組在第28天尿液中的代謝物如acetoacetate和 hippurate相較於控制組有顯著上升；而 alanine 和acetate 則是相較於控制組有顯著下降，這些代謝物的改變可能跟能量產生受到影響與器官損害是有相關的，這部分都可以和過去腎臟與肝臟的切片結果相互呼應，除此之外，代謝物影響的效果隨著暴露劑量越高、暴露時間越久，所造成的影響更加明顯。 藉由研究大鼠在暴露順丁烯二酸後尿液中代謝物的變化，可以更為了解順丁烯二酸所引起的毒理機制，在未來，若能進一步的在其他動物上或是血液介質裡做探索，相信可以提供更多生物資訊以推估人體暴露後所產生良健康效應。
並列摘要
More and more food safety issues are noticed by the public. In 2013, the Ministry of Health and Welfare (MOHW) in Taiwan declared that some starch-processed foods were illegally added food addictive, maleic acid/maleic anhydride. Modified starch can enhance favorable properties, such as viscosity, texture, and elasticity in food. Accidental consumption of maleic acid at low levels does not cause significant adverse health effects; however, long term exposure of high levels of maleic acid can induce kidney damage. The molecular effects of repeated maleic acid exposure are still largely unknown. In this study, we intend to understand metabolic effects of repeated exposure to maleic acid in rats using 1H NMR-based metabolomic approach. Rat urinary metabolome were examined to study time-course and dose-response of maleic acid. Adult male Sprague-Dawley (SD) rats were divided into control, low-dose (6 mg/kg), medium-dose (20 mg/kg), and high-dose (60 mg/kg) and treated with vehicle or maleic acid via oral gavage daily. Urine samples were collected twice a day (once during daytime and once at night) on day 0, 7, 14, 21, and 28 and then examined by high-resolution 1H nuclear magnetic resonance (NMR) followed by multivariate statistical analysis. The principle component analysis (PCA) score plots from the anlaysis of urinalry metabolome showed changes of metabolome patterns within different exposure groups. Clear metabolome seperation between high-dose and the control groups were observed from the night samples of day 14 and later. The increased levels of acetoacetate and hippurate, and decreased levels of alanine and acetate in the treatment groups were observed in the night samples of day 28. Changes of metabolites are related with environment stress and energy metabolism. Metabolic effects of maleic acid exposure are obvious on rats in high dose group and at last time points. By investigating the perturbation of urinary metabolome in SD rats can assist urinary biomarker discovery for maleic acid and find out possible toxic mechanisms induced by maleic acid.
參考文獻
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