Isolation Of Mitochondria From Pulp And Components Assay Of Essential Oil From Peel Of Citrus Fruit

Citrus is originated and widely cultivated in China. The main citrus cultivated in China are used for delicious food on table. With the development of economy in China, higher requirements on the internal and external quality of citrus fruits were put forward by people. The internal quality mainly includes fruit flavor, mineral nutrition, soluble sugar content, organic acid content and so on. The external quality mainly includes fruit shape, the ratio of transverse diameter to longitudinal diameter, the color and lustre and so on. After harvest, the nutrition supply was cut off and citrus fruits begin to senesce. During this stage, some senescence related physiological indexs change significantly which will result in fruits quality deterioration. As an important organelle in animal and plant cells, mitochondrial participate in cell senescence. Although the morphology and physiological state of mitochondrial have close relationship with the biological characteristics of citrus fruits after harvest, we still lack the direct experimental evidence. So, it is meaningful to establish the method for mitochondrial isolation. On the other hand, oleocellosis, a kind of physiological disorder, will significantly reduce the external quality of citrus fruits. However, the mechanism of oleocellosis is still not clear.In our study, Citrus unshiu “Guoqing No.1” and valencia orange(Citrus sinensis Osbeck) were choosed as materials for mitochondrial isolation task. At the same time, we choosed satsuma orange, “newhall” navel orange and “zaohong” navel orange as materials to analysis their essential oil components. The main experimental results are as follows:1 Different citrus cultivars need different buffer with different sucrose concentration. For valencia orange, the suitable buffer for mitochondria isolation should contain 0.4 M sucrose. For Citrus unshiu “Guoqing No.1”, however, the suitable buffer for mitochondria isolation should contain 0.5 M sucrose. By one time nycodenz purification, the western blot was performed to estimate the purity of mitochondrial. We found that most mitochondrial isolated by our method were intact and without chromoplast, peroxisome, cytoplasm contamination. However, in our results, we detected V-ATPase signal, which indicated that the mitochondrial isolatd by our method contaminated by vacuole.2 By the comparison of different development stages of “zaohong” navel orange, we found that γ-terpinene, terpinolene, β-linalool, citronellal, neral, perillal and α-caryophyllene were up-regulated accumulated in “zaohong” navel orange during the degreen stage. By the comparison of different varieties during the degreen stage, we found that sabinene, myrcene, β-linalool, neral, geranial, β-cubebene, β-farnesene,(Z,E)-α-farnesene, 1-octanol, octanal, nonanal, decanal and dodecanal were up-regulated accumulated in “zaohong” navel orange.3 The results of fruit surface treatment using essential oil extracted from “zaohong” navel orange during the degreen stage indicated that the satsuma oranges before degreen stage were more sensitive to essential oil than that after degreen stage. After the application of essential oil, the lesion area on the fruits before degreen stage will be larger than that on the fruits after degreen stage. The browning degree on the fruits before degreen stage will also be more serious than that on the fruits after degreen stage. By ultraviolet light irradiation, we observed yellow fluorescence on the oleocellosis area. The light microscopy images showed that the shape of parenchyma cells in oleocellosis area changed significantly.4 By the lesion diameter measurement of “zaohong” navel orange, we found that essential oils extracted from different kinds of citrus fruits during four stages did not show different effects on citrus fruit. However, the fruits in different development stages showed different oleocellosis sensitivities.