The metabolism of membrane lipid participates in the occurrence of chilling injury in cold-stored banana fruit.

Banana fruit is highly vulnerable to chilling injury (CI) during cold storage, which results in quality deterioration and commodity reduction. The purpose of this study was to investigate the membrane lipid metabolism mechanism underlying low temperature-induced CI in banana fruit. Chilling temperature significantly induced CI symptoms in banana fruit, compared to control temperature (22 °C). Using physiological experiments and transcriptomic analyses, we found that chilling temperature (7 °C) increased CI index, malondialdehyde content, and cell membrane permeability. Additionally, chilling temperature upregulated the genes encoding membrane lipid-degrading enzymes, such as lipoxygenase (LOX), phospholipase D (PLD), phospholipase C (PLC), phospholipase A (PLA), and lipase, but downregulated the genes encoding fatty acid desaturase (FAD). Moreover, chilling temperature raised the activities of LOX, PLD, PLC, PLA, and lipase, inhibited FAD activity, lowered contents of unsaturated fatty acids (USFAs) (γ-linolenic acid and linoleic acid), phosphatidylcholine, and phosphatidylinositol, but retained higher contents of saturated fatty acids (SFAs) (stearic acid and palmitic acid), free fatty acids, phosphatidic acid, lysophosphatidic acid, diacylglycerol, a lower USFAs index, and a lower ratio of USFAs to SFAs. Together, these results revealed that chilling temperature-induced chilling injury of bananas were caused by membrane integrity damage and were associated with the enzymatic and genetic manipulation of membrane lipid metabolism. These activities promoted the degradation of membrane phospholipids and USFAs in fresh bananas during cold storage.

The Changes in Metabolisms of Membrane Lipids and Phenolics Induced by Phomopsis longanae Chi Infection in Association with Pericarp Browning and Disease Occurrence of Postharvest Longan Fruit.

This study investigated the changes in metabolisms of membrane lipids and phenolics caused by Phomopsis longanae Chi infection in association with pericarp browning and fruit disease occurrence of postharvest longans. Compared with the uninoculated-longans, the longans inoculated by P. longanae exhibited higher cellular membrane permeability; higher PLD, lipase, and LOX activities; and higher levels of saturated fatty acids (SFAs) and phosphatidic acid but lower levels of phosphatidylinositol, phosphatidylcholine, and unsaturated fatty acids (USFAs). Additionally, the longans inoculated by P. longanae showed higher activities of POD and PPO but a lower amount of total phenolics. These findings suggested that infection of P. longanae enhanced activities of PLD-, lipase-, and LOX- stimulated degradations of membrane lipids and USFAs, which destroyed the integrity of the cell membrane structure, resulting in enzymatic browning by contact of phenolics with POD and PPO, and resulting in reduction of resistance to pathogen infection and accordingly accelerated disease occurrence of postharvest longan fruit.

Expression and Characterization of a Novel Thermostable and pH-Stable ß-Agarase from Deep-Sea Bacterium Flammeovirga Sp. OC4.

A novel gene (aga4436), encoding a potential agarase of 456 amino acids, was identified in the genome of deep-sea bacterium Flammeovirga sp. OC4. Aga4436 belongs to the glycoside hydrolase 16 ß-agarase family. Aga4436 was expressed in Escherichia coli as a fusion protein and purified. Recombinant Aga4436 showed an optimum agarase activity at 50-55 °C and pH 6.5, with a wide active range of temperatures (30-80 °C) and pHs (5.0-10.0). Notably, Aga4436 retained more than 90%, 80%, and 35% of its maximum activity after incubation at 30 °C, 40 °C, and 50 °C for 144 h, respectively, which exhibited an excellent thermostability in medium-high temperatures. Besides, Aga4436 displayed a remarkable tolerance to acid and alkaline environments, as it retained more than 70% of its maximum activity at a wide range of pHs from 3.0 to 10.0 after incubation in tested pHs for 60 min. These desirable properties of Aga4436 could make Aga4436 attractive in the food and nutraceutical industries.

Hydrogen Peroxide Induced Changes in Energy Status and Respiration Metabolism of Harvested Longan Fruit in Relation to Pericarp Browning.

Energy status and respiration metabolism of "Fuyan" longan fruit treated by hydrogen peroxide (H2O2) and their relationship to pericarp browning were studied. The results displayed that H2O2 significantly increased the respiration rate, increased activities of respiratory terminal oxidases like cytochrome C oxidase (CCO) and ascorbic acid oxidase (AAO), decreased NAD kinase activity, maintained lower contents of NADP and NADPH as well as higher amounts of NAD and NADH, and accelerated the decrease of energy charge. These results gave convincing evidence that the treatment of H2O2 for accelerating longan pericarp browning was due to an increase of energy deficiency, an increase of respiratory metabolic pathways of Embden-Meyerhof pathway (EMP) and tricarboxylic acid (TCA) cycle, a decrease of pentose phosphate pathway (PPP) of respiratory pathway, and an increase of activities of respiratory terminal oxidases like CCO and AAO.

Effect of Monascus aged vinegar on isoflavone conversion in soy germ by soaking treatment.

Soy germ rich in isoflavones has attracted much attention for health-promoting characteristics. An effective approach via Monascus aged vinegar soaking was adopted to enhance the aglycone amount. The profiles and interconversion of soy germ isoflavones via Monascus aged vinegar soaking were investigated, and the distribution in vinegars were also explored. The aglycones were dramatically increased by 40.76 times. Concomitantly, ß-glycosides and malonylglycosides were significantly decreased. The proportion of aglycones presented a sharp increase with the endogenous ß-glucosidase activity at the initial 4h incubation. There appeared to be correlations between ß-glucosidase activity and the hydrolysis of conjugated isoflavones. The results demonstrated that the reactions of decarboxylation, de-esterification and de-glycosylation were involved in the Monascus aged vinegar soaking, supporting synergistic effects of enzymolysis by endogenous ß-glucosidase from soy germ and acid hydrolysis of vinegars. Soaking by vinegar is a promising pathway for preparing aglycone-rich soy germ.

Simultaneous determination of caffeine and some selected polyphenols in Wuyi Rock tea by high-performance liquid chromatography.

The primary taste and healthy benefits of tea are mainly attributed to tea polyphenols and caffeine. Due to very many kinds of flavonoid glycosides in tea and the lack of commercial standards of flavonoid glycosides, it is critical to develop a rapid and cheap method for determining flavonoid glycosides of tea. Contents of myricetin glycosides and quercetin glycosides in Wuyi Rock tea were determined by detecting contents of corresponding myricetin and quercetin. Optimizing hydrolysis conditions for hydrolyzing flavonoid glycosides to their corresponding flavonols including quercetin and myricetin in Wuyi Rock tea was a key technology for detecting contents of corresponding myricetin and quercetin. The results showed that hydrolysis at 2 mol/L HCl solution and at 90 °C for 1 h was an optimizing condition for hydrolyzing flavonoid glycosides to myricetin and quercetin in Wuyi Rock tea. Caffeine and seven kinds of polyphenols (GA, EGC, C, EGCG, EC, ECG, and CGA) in 20 samples of Wuyi Rock tea were simultaneously determined using a simple and fast reverse-phase high-performance liquid chromatography procedure coupled with photodiode array detector (RP-HPLC-PDAD). The results indicated that there were significant (P < 0.05) differences of ECG, CGA, ECG, and myricetin glycosides in 'Wuyi Rougui' and 'Wuyi Shuixian', which were credited with causing the difference in taste between these two cultivar of Wuyi Rock tea. The study may be useful for clarifying the cause of "cultivated varieties flavor" of Wuyi Rock tea.

Inhibitory effects of propyl gallate on tyrosinase and its application in controlling pericarp browning of harvested longan fruits.

Tyrosinase (EC 1.14.18.1), also known as polyphenol oxidase (PPO), is a key enzyme in pigment biosynthesis of organisms. The inhibitory effects of propyl gallate on the activity of mushroom tyrosinase and effects of propyl gallate on pericarp browning of harvested longan fruits in relation to phenolic metabolism were investigated. The results showed that propyl gallate could potently inhibit diphenolase activity of tyrosinase. The inhibitor concentration leading to 50% activity lost (IC50) was determined to be 0.685 mM. Kinetic analyses showed that propyl gallate was a reversible and mixed type inhibitor on this enzyme. The inhibition constants (K(IS) and K(I)) were determined to be 2.135 and 0.661 mM, respectively. Furthermore, the results also showed that propyl gallate treatment inhibited activities of PPO and POD in pericarp of harvested longan fruits, and maintained higher contents of total phenol and flavonoid of longan pericarp. Moreover, propyl gallate treatment also delayed the increases of browning index and browning degree in pericarp of harvested longan fruits. Therefore, application of propyl gallate may be a promising method for inhibiting tyrosinase activity, controlling pericarp browning, and extending shelf life of harvested longan fruits.

Inhibitory effects of naphthols on the activity of mushroom tyrosinase.

Tyrosinase (EC 1.14.18.1), a copper-containing multifunctional oxidase, was known to be a key enzyme for biosynthesis in fungi, plants and animals. In this work, the inhibition properties α-naphthol and ß-naphthol toward the activity of tyrosinase have been evaluated, and the effects of α-naphthol and ß-naphthol on monophenolase and diphenolase activity of tyrosinase have been investigated. The results showed that both α-naphthol and ß-naphthol could potently inhibit both monophenolase activity and diphenolase activity of mushroom tyrosinase, and that ß-naphthol exhibited stronger inhibitory effect against tyrosinase than α-naphthol. For monophenolase activity, ß-naphthol could not only lengthen the lag time but also decrease the steady-state activity, while α-naphthol just only decreased the steady-state activity. For diphenolase activity, both α-naphthol and ß-naphthol displayed revisible inhibition. Kinetic analyses showed that both α-naphthol and ß-naphthol were competetive inhibitors.

Two GH3 genes from longan are differentially regulated during fruit growth and development.

In the present work, two full length cDNAs of GH3 genes, named DlGH3.1 and DlGH3.2 were cloned from pericarp and aril tissues of the longan fruit, respectively. Three conserved motifs, SSGTSAGERK, YASSE and YRVGD, as a characteristic of the acyladenylate/thioester forming enzyme superfamily were observed in DlGH3.1 and DlGH3.2 proteins. DlGH3.1 mainly expressed in pericarp tissues while DlGH3.2 accumulated in both the pericarp and aril tissues during fruit growth and development. In addition, NAA treatment induced the expression of DlGH3.1 and DlGH3.2 in the pericarp tissues at 21 and 77days after anthesis (DAA), while only DlGH3.2 in the aril tissues could be induced by NAA at 77DAA. More importantly, ABA and ethrel treatments suppressed the accumulations of DlGH3.1 and DlGH3.2 in the pericarp tissues of longan fruit at 21DAA (a rapid growth stage of pericarp), but enhanced DlGH3.2 expression in the aril tissues at 77DAA (a fruit ripening stage). Furthermore, the expression patterns of DlGH3.1 and DlGH3.2 showed different tissue specificity. Thus, our results suggest that DlGH3.1 gene expression might be associated with pericarp growth, while DlGH3.2 accumulation is likely to be related to both pericarp growth and fruit ripening, and the responses of DlGH3s to plant growth hormones are different and dependent on fruit development stage and fruit tissue.

[Changes in cell wall components and cell wall-degrading enzyme activities of postharvest longan fruit during aril breakdown].

Longan (Dimocarpus longan Lour.) fruits are very susceptible to pericarp browning and aril breakdown, and postharvest aril breakdown is one of the most important factors degrading the quality and shorting storage life of longan fruit. Changes in aril breakdown index, cell wall components and cell wall-degrading enzyme activities in aril of longan cv. Fuyan fruits using sealed polyethylene film bags (0.015 mm thick) at (10+/-1) degrees C were investigated. The main results were as follows. Development of aril breakdown was higher with storage time (from day 0 to day 36). Aril breakdown index was positively and significantly correlated with storage time (P<0.01). During development of aril breakdown, the total dry weight of the cell wall materials, protopectin, cellulose, semicellulose and cell wall protein contents of aril decreased progressively. The total dry weight of the cell wall materials, contents of protopectin, cellulose, semicellulose and cell wall protein of aril were all negatively correlated with aril breakdown index. There were low beta-galactosidase activity, and high activities of pectinesterase (PE), polygalacturonase (PG) and cellulase in aril of harvested fruit. PE activity in aril gradually decreased during development of aril breakdown. The activities of PG and cellulase in aril increased significantly during storage from day 6 to day 12 and from day 0 to day 12, respectively. The peaks enzyme activities of both PG and cellulase appeared on the 12th day after harvest, then the enzyme activity decreased; whereas, the activities of PE, PG and cellulase changed little from day 0 to day 24, and then rapidly decreased. The beta-galactosidase activity in aril decreased slightly during storage from day 0 to day 24. However, the beta-galactosidase activity increased significantly after day 24. Especially, the beta-galactosidase activity increased rapidly after 30 d of storage, in the meantime, the activities of PE, PG and cellulase almost disappeared. From the results it can be seen that the development of aril breakdown was due to the degradation of cell wall components such as protopectin, cellulose, semicellulose and cell wall protein. The early and middle phases of development of aril breakdown were mainly brought about by the action of PE, PG and cellulase, whereas, beta-galactosidase played the key role at the late phase of aril breakdown.

[The relationship between the desiccation-induced browning and the metabolism of active oxygen and phenolics in pericarp of postharvest longan fruit].

Longan (Dimocarpus longan Lour.) fruits are very susceptible to water loss and pericarp browning, and postharvest pericarp browning is the most important factors degrading the quality of longan fruit and shorting storage life. Pericarp browning has been attributed to desiccation, chilling, heat stress, senescence and pest or pathogen attack. Desiccation is the most main factor of induced-pericarp browning in longan. The relationship between water loss from pericarp and pericarp browning in longan cv. Fuyan fruits using open plastic punnets and sealed polyethylene bags at 10 degrees C +/-1 degrees C and 50% relative humidity, and the effect of pericarp water loss of the fruit on active oxygen metabolism and phenolics metabolism were investigated. Water loss resulted in rapid pericarp browning. Development of pericarp browning was higher with higher rate of water loss from pericarp and storage time (from 0 to 6 days). Water loss from pericarp was positively correlated with pericarp browning index significantly (P<0.01). Water loss from pericarp resulted in reduced activities of reactive-oxygen-scavenging enzymes (SOD, CAT, APX and GR), decreased amounts of endogenous antioxidant substances (AsA and GSH), and increased rates of O(-.)(2) production, MDA content and relative leakage rate, which showed that membrane structure was broken. Water loss from pericarp resulted in an increase in activity of PPO, and obvious reductions in total phenolic and flavonoid contents, whereas there was not obvious change in anthocyanin content. These results show that phenolics and flavonoids are the main substrates for PPO during desiccation-induced browning. Water loss from pericarp caused a significant increase in activity of POD, which also plays an important role in desiccation-induced browning in pericarp of longan fruit. Water loss from pericarp caused an increase in pH value, which resulted in changes in anthocyanin structure and color, the degradation of anthocyanin became easier. The results suggest that desiccation-induced browning of longan pericarp may be due to a decrease in activities of reactive-oxygen-scavenging enzymes and amounts of endogenous antioxidant substances, an accumulation of active oxygen, an increase in membrane lipid peroxidation, an injury of the integrity of cellular membrane structure, which, in turn, may cause cellular decompartmentation, resulted in PPO and POD, located in plastid and other organelle, to come into contact with phenolic and flavonoid substrates, located in vacuole, to form brown polymers.