環境流布濃度之鄰苯二甲酸二(2-乙基己基)酯對青鱂魚生殖毒性探討

Environmental Relevant Concentrations of Di(2-ethylhexyl)phthalate Cause Reproductive Toxicity in Medaka Fish (Oryzias Latipes)

林明慧(Ming-Huei Lin)

指導教授 ： 陳佩貞

國立臺灣大學/生物資源暨農學院/農業化學研究所/碩士(2014年)

https://doi.org/10.6342/NTU.2014.02633

摘要

鄰苯二甲酸酯類為全世界工業製造常用之塑化劑，添加至塑膠製品中可增加塑膠聚合物的柔軟度和延展性。其中，又以鄰苯二甲酸二(2-乙基己基)酯 [Di(2-ethylhexyl)phthalate, DEHP] 的使用量最大且最普遍。在世界各地包括臺灣水體環境中都曾偵測到DEHP的流布。近年來有文獻指出DEHP對小鼠或水生生物具有內分泌干擾效應，可能影響生殖器官的發育及功能，但前人研究多以高於環境流布濃度之劑量暴露模式生物，無法反映DEHP對環境生物之影響，此外DEHP之確切作用機制尚未釐清，不同研究之間解釋方向仍有歧異。本研究目的為探討環境流布濃度之DEHP暴露對青鱂魚生殖能力的影響及可能的毒性作用機制。實驗以FLFII (Female leucophore free II) 品系之青鱂魚 (Oryzias latipes) 作為模式生物，將成魚暴露於環境相關濃度 (20-200 μg/L) 之DEHP達21天後，觀察其生殖能力變化，並量測內分泌及氧化壓力相關指標，包含性荷爾蒙含量、抗氧化酵素活性和基因表現。結果顯示，暴露DEHP達21天會顯著降低青鱂魚產卵量，並抑制肝臟中cyp1a、cyp3a40等代謝外源化合物的一期 (Phase I) 酵素基因表現，以及鰓中CAT、GST等抗氧化酵素活性，其中GST也負責外源汙染物的二期 (Phase II) 代謝，顯示DEHP會對青鱂魚造成生殖毒性外，也會影響青鱂魚代謝DEHP此類親脂汙染物的能力。除此之外，青鱂魚肝臟中抗氧化酵素的基因表現也受到DEHP干擾，雌魚的cat表現被誘導，gpx則受到抑制，顯示雌魚肝臟中可能產生過氧化氫，且主要由過氧化氫酶 (Catalase) 負責清除；雄魚則是sod和gpx基因表現皆受到抑制，可能影響雄魚清除活性氧物種的能力。實驗也測量了性腺中抗氧化酵素的基因表現，結果顯示只有雌魚卵巢的基因表現受到干擾，其sod被誘導，cat、gpx則受到抑制，顯示卵巢組織可能產生氧化壓力，影響雌魚生理功能，進而造成產卵量下降。綜上所述，環境流布濃度之DEHP會造成青鱂魚產卵量下降，且影響抗氧化酵素活性及相關基因表現，並可能干預青鱂魚排除外源汙染物的能力並造成氧化壓力，損害正常生理功能。由於DEHP在環境中廣泛存在，因此對環境生物和生態的潛在衝擊不容忽視。

關鍵字

鄰苯二甲酸二(2-乙基己基)酯 ； 環境相關濃度 ； 生殖毒性 ； 氧化壓力 ； 青鱂魚

並列摘要

Phthalates are synthetic plasticizers commonly used in industry to enhance the flexibility and softness of plastic products. Among these phthalates, di(2-ethylhexyl)phthalate (DEHP) is the most common plasticizer in the world. It is easily leached into the environment and ubiquitously detected in surface water in many countries including Taiwan. Recent studies have indicated that DEHP is an endocrine disrupting chemical (EDC) in experimental animals. It is considered that DEHP exposure will retard development of reproductive organs and impair physiological function. However, studies directly linking to environmental relevant concentrations are still limited. The impact and associated toxic mechanism of DEHP on environmental aquatic life is unclear at present. The objective of the study is to investigate the toxic effect of DEHP on fish reproduction using medaka (Orzyias latipes) fish as the model organism. We have treated pairs of sexually matured adults of female leucophore free II (FLFII) medaka with DEHP at environmentally relevant concentrations 20, 100 and 200 μg/L for a 21-day aqueous exposure and assess their reproduction performance. Fish were used for analyses of plasma concentrations of sex hormone at the end of exposure, activities of antioxidant enzymes and gene expression in different organs. Our results show that DEHP at 20-200 μg/L decreased fecundity, and also inhibited the expression of phase I enzymes (cyp1a and cyp3a40) in liver. In gill, antioxidant enzymes such as catalase (CAT) and glutathione S-transferase (GST), the phase II enzyme, activities were decreased with 100-200 μg/L DEHP. DEHP may interfere xenobiotics metabolism of medaka through inhibiting expression of both phase I and phase II enzymes. Furthermore, gene expression of sod, cat and gpx were interfered in liver and gonads with DEHP treatments (20-200 μg/L). In female liver, cat was induced, but gpx was inhibited. On the other hand, both sod and gpx were inhibited in male liver, suggesting the defense against oxidative stress was altered. In addition, sod induction and cat and gpx inhibition were observed in ovaries. The DEHP-induced oxidative stress in ovaries may obstruct female reproductive function, and then decrease fecundity. In conclusion, environmentally relevant concentrations of DEHP caused reproductive toxicity in medaka. The activities and gene expression of antioxidant enzymes were altered by DEHP exposure in liver, gill and gonad, indicating oxidative stress and interference of xenobiotics metabolism. DEHP is an ubiquitous environment contaminant of emerging concern, so it is important to evaluate the risk to ecosystem.

並列關鍵字

Di(2-ethylhexyl)phthalate (DEHP) ； Environmental relevant concentration ； Reproduction toxicity ； Oxidative stress ； Medaka (Oryzias latipes)

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