百日草對鎘毒害的反應與其鎘結合勝?的鑑定

Abstract

本實驗是以發芽後二十天或四十天的百日草（Zinnia elegans）為材料。以1 ppm，5 ppm及10 ppm等不同鎘濃度培養液處理植株十天。另以2 ppm鎘濃度分別進行間斷和連續給鎘培養十八天。鎘害現像是以檢視外表形態及利用超薄切片觀察超微構造的變化，同時以電泳分析法檢定解毒同功酵素的類型受鎘害的差異。鎘在植株內的分佈和累積則以原子吸收光譜儀測定。利用離子交換柱形色層和膠體過濾柱形色層法分離鎘結合勝?，並測定其氨基酸組成的比例。並配合酸性不穩定硫的測定，以區別鎘結合勝?的存在類型。 鎘害主要造成植株矮化、葉脈與附近葉片的黃化以及根部生長的抑制。培養液的鎘含量超過1 ppm時即呈鎘害。鎘濃度愈高受害程度愈嚴重，表現鎘害所需的時間愈短。 超微構造的觀察顯示受害植物以葉綠體受害最突出。油滴（Plastoglobuli）數目增加、葉綠餅間層（Intergranal thylakoid）膨脹鬆散、葉綠餅數目和疊膜層數減少及其膜腔膨大鬆散是常見之現象，隨鎘濃度的增高而明顯。根部及維管束組織細胞的細胞壁、原生質膜以及液胞則有極多的黑色小點聚集。 酸性磷酸酵素（Acid phosphatase）和過氧化氫酵素（Peroxidase）的同功酵素類型，其活性的表現隨鎘處理濃度的不同而異。超氧歧化酵素（Superoxide dismutase）同功酵素的表現則與鎘濃度無關。 百日草消耗培養液中的鎘量，以處理後六天達最大消耗量。消耗量的下降則隨處理鎘濃度的增高而加快。連續給鎘處理與間斷給鎘在實驗初期的耗鎘量相當，但六天後則呈不同消耗曲線。鎘進入植株後主要累積在根部，其累積量往往是地上部的4到10倍。鎘濃度愈高植物體內累績的鎘量愈多。另外，較成熟植株的根部比幼嫩者具有較高的聚集能力。 經柱形色層法純化得到兩群鎘結合蛋白質，檢定酸性不穩定硫含量及比較氨基酸組成的比例，推測小分子量蛋白質是鎘結合勝?（Phytochelatins）。

Young plants of Zinnia elegans in 20-day-old or 40-day-old were cultured in nutrient solutions containing 0, 1, 5, or 10 ppm cadmium for 10 days. Cakmium toxicity were reflected on morphological changes and isozyme patterns studied with electron microscopy and electrophoreses, respectively. The accumulation of cadmium in different parts of plant was detected by a Hitachi Z-6100 atomic absorption spectroscope. Phytochelatins were purified with both ion exchange and gel filtration column-chromatographic techniques and their amino acid composition was determined by aamino acid analyzer. The major morphological changes of cadmium-treated plants are the retardation of plant growth, chlorosis of leaves and shrivelling of plants. The durataion for toxicity expression is reversedly to cadmium concentration. Cadmium cause the swelling and looseness of thylakoid membrane, the increment of plastoglobuli quantity and the reduction of stack members of chroloplast grana. Besides, electron dense aggregates are often observed within cell wall, plasmalemma and vacuole of cells in vascular bundles of leaves and roots. The isozyme patterns of acid phosphatase and peroxidase are altered depending on the cadmium concentration, but those of superoxide dismutase remain uniformity. The quantity of cadmium accumulation on roots was 4-10 fold higher than that of shoots. The older plants occupy with the higher ability of cadmium accumulation than the younger ones. The depletion of cadmium in nutrient solution maximaly reveals at the sixth day after treatment. The depletion of cadmium in successive supplement is similiar to that of discontinuous treatment. Two groups of cadmium-binding peptides purified with column chromatography were identified by acid-labile sulfur test and amino acid composition. Only the smaller molecular weight fraction of cadmium-binding peptide is phytochelatin.