| In the present study, Hydrilla verticillata (Linn.f.) Royle, a submerged macrophyte widely distributed in China was used as experimental material. Cd, Cu and Zn were selected as the stress factors; the former is unessential heavy metal element, the latter two ions are essential elements for the normal development and growth of plants. H. verticillata was cultivated in water containing elevated concentrations of Cd, Cu and Zn (up to 10 mg/1) for 7 d under lab condition and response of H.a verticillata to Cd, Cu and Zn pollution in water were systemically studied from the following aspects: (1) the bioconcentration of Cd, Cu and Zn in H.a verticillata were determined by Atomic Absorption Spectrometry (AAS) and X-ray energy spectrum analysis; (2) Distribution of Cd, Cu and Zn in different subcellular fraction was measured by AAS; (3) the Cd. Cu and Zn chemical forms in leaf cell were analyzed using sequential extraction; (4) Determination of macronutrients (P, K) and micronutrients (Ca, Mg, Na, Mn, Fe, Cu, Zn) in H. verticillata by Inductively Coupled Plasma-Atomic Emission Spectrometry (ICP); (5) Generation of active oxygen (O2) and response of antioxidant enzymes (SOD, POD, CAT) in H. verticillata under Cd, Cu and Zn stress were investigated; (6) Changes of chlorophyll and soluble protein contents in H. verticillata were studied; (7) Changes of auto-fluorescence in H. verticillata leaf under Cd, Cu and Zn stress were analyzed by Laser Scanning Confocal Microscope (LSCM); (8) Damages of Cd, Cu and Zn to morphologic structure and ultrastructure were observed by scanning and transmission election microscope; (9) Subcellular distribution of Cd, Cu and Zn in leaf cells was observed by a modified sulfide-silver method; (10) Zn-binding protein in H. verticillata under Zn stress was isolated and purified by fractional precipitation with ammonium sulfate and gel filtration on Sephadex G-100 and G-75. The results showed that: (1) Aquatic plants are known to accumulate and bioconcentrate heavy metals. Contents of Cd, Cu and Zn in H. verticillata were nearly in linear increase with rise of metal concentration in solution, Statistical analysis indicated that they allreached significant positive levels It was found that H. verticillata fronds was able to absorb and accumulate Cd, Cu and Zn differently, the bioconcentration factor was ranged from 193 to 307, 571 to 1328 and 346 to 830, respectively. The absorption capability of H. verticillata to Cu was maximal among the three elements and low concentration of heavy metals was good for absorption. (2) Contents of Cd, Cu and Zn in subcellular fraction ascended with augment of content of the three heavy metals in H. verticillata. Both Cd and Zn occurred in different parts of leaf cell with the following sequence: Cd, Zn: cell wall > soluble fraction > organelles; Cu: cell wall > organelles > soluble fraction. Which revealed that cell wall played an important role in the H. verticillata' absorption of Cd, Cu and Zn. (3) Sequential extraction indicated that the ratio of different Cd, Cu and Zn chemical forms were different markedly, of which NaCl extractable Cd and Zn, Ethanol extractable Cu were predominated as compared with the other 5 forms; which could be seen in the following order: for Cd: for Zn: FfxJaC FHAC FHC! FEthanol (4) Cd, Cu and Zn affected the concentration of nutrient elements in H. verticillata differently. For macronutrient, such as P and K, the three heavy metals all inhibited its absorption and all reached significant negative correlation with the increase of concentration in the media. As for micronutrient, Cd accelerated the absorption of C Cu, content of Mg and Na increased first and decreased afterwards; This is the same with the effects of Cu and Zn on Ca and Mn, both Cu and Zn all promoted the H. verticillata' absorption of Fe significantly. Cu content in H. verticillata enhanced when treated with Zn, and Zn content in H. verticillata declined when treated with Cu. This suggested that the balance of ion in cell was broke. (5) Cd, Cu and Zn acceler...
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