Effects Of Aluminum On Uptake, Distribution And Accumulation Of Fluorine In Tea Plants And Its Mechanism

Tea(Camellia sinensis), as a fluorine (F) accumulator, accumulates F in mature tea leaves, which was100-1000times higher than other plant species. F is an important element to human health and a moderate intake of F is confirmed to be beneficial to alleviation of dental caries. However, excessive accumulation of F in human body can cause diseases such as fluorosis. The drinking-tea born fluorosis was reported to be caused by long-term consumption of a large quantities of tea containing extremely high level of F. Many researchers attempted to exploit measures to control F concentration in tea and tea liquor, including breeding cultivars with low-F level, improving cultivation methods, optimizing processing procedures and developing defluoride agent. This works studied F accumulation characteristics by investigating root morphology parameters, root vitality, cell membrane penetration, and antioxidase activities and malondialdehyde (MDA) content in tea leaves under F stress; The relationship between F accumulation and organic acid secretion was studied using high performance liquid chromatography (HPLC). F distribution differences between cultivars, organs and subcellular fractions under various treatment combination of F-and Al3+were investigated by isolating of cell fraction using differential centrifugation and transmission electron microscopy-energy dispersive X-ray (TEM-EDX). Various chemical species of aluminum and their subcellular distribution in tea leaves were also studied by using differential centrifugation technique and chemical extraction method; Kinetic and extraction model of F leaching from tea were studied using the second-order Fickian equation based on the kinetic behaviors and extraction model and a two-step infusion method was developed to control F level in tea liquor.1. Low level of F-promoted roots tips prolongation of tea plant. However, when tea plant was exposed to a high concentration of F-such as24mg·L-1, the root tip prolongation was inhibited, and root cell membrane was damage, which resulted in the cell membrane penetration increased. Level of free radical scarvenge activity decreased under F-stress. In this case, thylakoid membrane of the chloroplast was attacked and chloroplast membrane was injured, resulting in devrease in chlorophyll content.2. The major organic acids in tea root exudates were malic and oxalic acid, but malic, oxalic acid, pyruvic acid, ascorbic acid and a-ketoglutaric acid in tea leaf. Organic acids of tea plant under solution cultivation increased with increase in F concentration. These characteristics of tea root exudates might affect rhizosphere F chemical species and bioavailability. Al3+promoted the secretion of root exudate organic acids when tea plant was co-treated with F-and Al3+. The changes of organic acids might be a physiological response of tea plant to the stress of F-and Al3+.3. When tea plants were cultivated under high level F solution conditions, the F uptake rate by tea plant reached the highest at early24th h, and then declined. F might be passively absorbed in the form of F-when the tea plant was exposed to high concentration of F-upto24mg·L-1. F translocation rate in tea plant (from roots to stems and leaves) increased with increase in solution F-concentration. However, the F bioenrichment coefficients decreased with the increase in F concentration in culture solution. It is assumed that F accumulation in tea leaves was increasingly processed and it was inhibited by low concentration of Al. However, F accumulation in shoots and tender leaves (3rd,4th and5th) was enhanced by high concentration of Al3+in culture solution.4. Studies on subcellular distribution of F in tea plant leaves by isolation of cell fraction using differential centrifugation and TEM-EDX showed the following sequence:cell wall faction> protoplast fraction> chloroplast fraction in both tender shoots and mature leaves. Capacity of F accumulation in vacuole was quite limited. F distributed in subcellular fractions increased with the increase in F concentration of culture solution. The increase in F accumulation in cell wall was higher than in protoplast and chloroplast. In F" and A13+combination treatments, the accumulation of F in cell wall of mature leaves was the highest when tea plant was exposed to high F and low Al level. The absorbed F in tea leaves might be aggregated with Al in the cell wall where there are amylase and protein which are abundant in metal ion coordination groups such as hydroxide, carboxyl, aldehyde, amidogen and phosphate. These inhibited F-and Al3+cross-membrane transport, and decreased the F concentration in the protoplast. These might be a mechanism of tea plant tolerance to F. Under F starving condition, F in cell wall can be dissociated into free species which will be transported to young growing shoots and leaves of tea plants.5. Chemical species of aluminum and their subcellular distribution in tea leaves was studied by the differential centrifugation technique and chemical extraction method. It showed that organic aluminum amounted86.88%-88.11%of total aluminum in tea leaves, among which36.87%-56.82%was stable form and31.28%-51.08%unstable one. Inorganic aluminum was11.89%-13.12%of total aluminum in tea leaves, including species Al3-, Al(OH)2+and Al(OH)2+. There was significant difference in total aluminum level but no significant difference in ratio of organic aluminum to inorganic aluminum between various tea cultivars. The absorbed aluminum was mainly distributed in cell wall fraction which accounted for64.4%and83.24%of the total aluminum in the tender shoots and the matured leaves, respectively. The aluminum species of tea leaves is majorly in the organic forms and aluminum concentration in the cell wall fraction increased with the maturity of leaves. These might be contributed to lowering toxicity of aluminum and the aluminum tolerance of tea plant.6. Kinetics of F leaching from tea were studied. The leaching process of F could be divided into initial fast stage and later slow stage. The duration of initial fast stage depended on leaf particle size and temperature. The higher the temperature and the smaller the leaf particle size, the shorter the duration of the initial fast stage. Based on the test results, a two-step infusion method was developed to control F level in tea liquor. When the tea was infused in50℃water for20s (first infusion) and then the residue leaf was re-infused in boiling water for5min (second infusion), more than26percent of total F was removed during first infusion, but more than80percent of catechins and caffeine of the teas were remained in the second infusions. The F level in the second infusion was0.70mg·L-1or less. It is considered that the two-step infusion method is an alternative to control excessive F exposure for tea drinkers.