| Camellia sinensis (L.) is a kind of plant that can concentrate fluoride in nature; it can absorb a large amount of fluorine from the soil, air and water, higher than other plants of 2 to 3 orders-of-magnitude. However, research of fluoride on the physiological and biochemical of tea plant are less. In this experiment Chinese cultivars Fu ding da bai was used as the material to made a study of fluoride effect on the tea quality components, cell ultrastructure, physiological indicators of tea, analysed the the role and mechanism of antioxidant system and ascorbic acid-glutathione cycle in fluoride resistance of tea, analysis, investigated the fluoride distribution in sub-cellular and the effects on the interaction among fluoride, aluminum and calcium in tea. The results of this study are as follows:1. Effect of F on the chemical composition in tea leaves.Seedlings of the tea plant, Camellia sinensis (L.), were grown hydroponically for 30 days to study the effect of fluoride (F) on the chemical composition in the leaves. Polyphenols, protein, total catechins and monomeric catechins decreased significantly with increasing F concentration. On the other hand, the content of amino acids and soluble-sugars increased significantly, but the differences in caffeine and water-soluble extracts were not statistically significant, in the same time, most of the amino acid composition are first increased and then decreased. Metallic elements content of Ca, K, Cu, Zn decreased significantly, except for content of Mn increased significantly yet Fe first increased and then decreased and Mg appeared in contradiction to Fe, However, differences among treatments were not significant. The changing trend of the relative content of aroma components was different; however, the relative content of total aroma components was decreased. These results suggestted that quality of tea was related to fluoride level and the main chemical constituents of tea leaves decreased with F increasing.2. Effect of F on tea leaves photosynthesis feature and ultrastructureSeedlings of Camellia sinensis were grown hydroponically for 30 d in order to study the effect of fluoride (F) on photosynthesis and leaf ultrastructure. Chl content and PN increased slightly at low F concentration (2 mg/L) but not significantly. However, these two parameters decreased significantly (P<0.05) with increasing F. concentration. The cell ultrastructure had little change under 0-4 mg/L F, e.g. plastoglobulus and thylakoid expanded slightly, however, an irreversible destruction under the concentration of F higher than 6 mg/L was observed, such as membrane rupture, thylakoid expantion and even disintegration was found. These results were interpreted that the destruction of F to tea leaves ultrastructure leaded the decreased of Chl content and photosynthesis.3. Effect of F on metabolism of active oxygen in tea plant.3.1 Effect on metabolism of active oxygen in tea plant under different fluoride concentrationsSeedlings of Camellia sinensis were grown hydroponically for 30 d in order to study the effect of fluorine (F) on antioxidant defence and ascorbate-glutathione cycle system. MDA and H2O2 content did not have a significant difference compared to control at low F concentration (2 mg/L), however, these two parameters increased significantly with increasing F concentration. SOD activity was significantly decreased, which was detrimental to remove ROS, POD and CAT activity that was first increased then decreased, and the maximum at 6 and 4 mg/L, proline increased significantly, indicating that at low F concentration the antioxidant system had made a positive action to fluoride stress. In the ASA-GSH cycle system, APX, GR, MDHAR activities increased first and decreased afterwards, reached maximun under 4,2,2 mg/L, respectively, DHAR activity was significantly decreased; ASA increased first and then decreased, which showed that the antioxidant enzymes had made a positive action to fluoride stress in a degree and protected cells from ROS. However, the balance between formation and detoxification of ROS was lost with increasing of F concentrations. GSH content, ASA/DHA and GSH/GSSG were significantly decreased, and also showed an excessive accumulation of ROS resulting in the destruction of tea seedlings. Therefore, our results suggestted that at low F concentrations, the leaf antioxidant defence system and ascorbate-glutathione cycle system can scavenge reactive oxygen species and sufficiently protect cells from free radical injury. However, antioxidant defence system and ascorbate-glutathione cycle system couldn't scavenge excessive reactive oxygen species to protect the tissue from free radical injury under the stress of higher F. 3.2 Dynamic response of activity oxygen metabolism in tea plant under fluoride stressThrough nutrient fluid culturing, annual cuttings were exposed to 16 mg/L fluoride concentrations and cultivited for 0,6,12,24,48,72 h (without fluoride as control), the results indicated that with the time increasing, MDA and H2O2 had no significant difference, the activities of SOD decreased significantly, activities of POD, CAT, APX, GR increased first and then decreased, reached the highest at 48 h, the activities of antioxidases above MDHAR,DHAR and antioxidant ASA increased significantly, GSH had no obvious difference, ASA/DHA and GSH/GSSG increased, all these suggested that to protect tea from injury, tea could eliminate redundant ROS in the short term through antioxidases and AsA-GSH cycle, this is one of the mechanisms of fluoride-resistant of tea.4. Cause of Ca can alleviate the toxicity of Al-F to tea seedlings and fluoride-resistant mechanisms of teaUltrastructure of tea leaves and roots were severely damaged no matter under the separated treating of Al and For or the interaction of them, yet after adding Ca, it can significantly reduced the damage degree of the ultrastructure of leaves and roots, which explained that Ca could effectively reduce the damage of Al-F to tea cell. Under the processing of Al or interaction treatment of F-Al, content of pectin in root cell wall were both higher than the control. After adding Ca, when tea under the treatment of Ca-Al, the content of pectin in root cell wall decreased significantly than Al treatment alone, whereas under the treatment of F-Al-Ca, pectin content was significantly decreased than under interaction treatment of Al-F. This showed that Ca could reduce the damage of F-Al to tea may be due to Ca could change the composition of tea cell wall.Through the treating of F-Al of different proportions, the results showed that during the range, Al did not reduce the damage of F to tea seedlings, actually the damage was worsened. Therefore, the argument of F-Al compounds that could detoxify the damage needed further verification. In this study also found that, no matter cultivars Fu an da bai and Wu niu zao cultivated in field, or Fu ding da bai cultured in nutrient solution, Fluoride distribution in the sub-cells were Cell wall (F1)>Nuclear and chloroplast (F2) > Soluble fraction containing ribosomal (F4)>Mitochondria (F3). Most of the fluoride (60%) was fixed in the cell wall, so that cell wall could prevent F get into the cell which was an important mechanism of fluoride-resistant of tea. Combining with the phenomenon that field-cultivated tea bush containing amount of F but did not appear F-poisoning, while solution-cultured tea was poisoned by F, we inferred that the mechanism of fluoride-resistant in tea besides the protection of cell wall of tea leaves, also had relationship with some other components existing in soil, so that F-poisoning occurred in solution-cultured tea just because of lacking this composition in the solution. Therefore, we believed that fluoride-some components in the soil-the cell wall may be the fluoride-resistant mode of tea.
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