Biosynthesis And Accumulation Of Phenolic Compounds In Fresh-Cut Carrot And Its Possible Regulatory Mechanisms

Fresh-cut fruits and vegetables are popular for consumers as a result of its convenient,fresh,nutritional and high edible values.As compared with un-cut produce,fresh-cut fruits and vegetables are very perishable due to mechanical damage.The changes in fresh-cut product include physiological metabolic disorders,accelerated quality deterioration and shortened shelf-life.When wounding stress occurs,fruits and vegetables usually synthesize and accumulate phenolic compounds to resist mechanical damage and to repair the damaged tissues.However,polyphenols are prone to oxidative polymerization under the catalysis of polyphenol oxidase(PPO)and peroxidase(POD),leading to browning of cutting surfaces.For a long time,domestic and foreign researchers pay more attention to control the adverse effects of tissue browning induced by phenolic compounds,but ignore its potential health-promoting effects,such as anti-oxidation and anti-tumor.Under the premise of ensuring the appearance and safety of fresh cut fruits and vegetables,the use of cutting as a physical stress factor to improve the contents of active ingredients in fresh-cut fruits and vegetables will be helpful for taking full advantages of antioxidant activity in fresh-cut fruits and vegetables.The present study was focus mainly on the phenomenon of the biosynthesis and accumulation of phenolic compounds in fresh-cut fruits and vegetables during storage.In this study,the effects of storage temperature and cut type on total phenolics content and antioxidant capacity of different fresh-cut vegetables during storage were investigated.Furthermore,carrot tissue was selected as the experimental material because of its remarkable accumulation of phenolic compounds and used to evaluate the interrelationship among respiratory metabolism,sugar metabolism,energy metabolism and phenolics biosynthesis,to explore the regulatory role of reactive oxygen species(ROS)and nitric oxide(NO)in modulating wound-induced biosynthesis and accumulation of phenolic compounds in carrot and to elucidate the molecular regulatory mechanisms of wound-induced biosynthesis and accumulation of phenolic compounds by transcriptome.The information generated in this study may supply a theoretical foundation for enhancing the biosynthesis and accumulation of phenolic compounds in fresh-cut fruits and vegetables.The results were as follows:1.In order to evaluate the effects of storage temperature and cut type on total phenolics content and antioxidant capacity of fresh-cut vegetables,8 different common vegetables were cut into slices,pies,and shreds and stored at 4 ? for 7 d and 20 ? for 2 d.Our results indicated that the total phenolics content and antioxidant capacity were influenced by vegetable species,storage temperature and cut type.In general,storage at 20? was more conductive for the accumulation of phenolic compounds compared with storage at 4 ?,and shredding was more conductive to promote the content of phenolic compounds compared with slicing and dicing.Among the 8 species of vegetables,an obvious increase of total phenolis content and antioxidant capacity was observed in fresh-cut onion,fresh-cut Chinese spring onion,fresh-cut zucchini,fresh-cut carrot and fresh-cut sweet potato,no evident changes of total phenolis content and antioxidant capacity were observed in fresh-cut cucumber and fresh-cut Pleurotus eryngii,while an obvious decrease of total phenolis content and antioxidant capacity was observed in fresh-cut green bell pepper.The largest increasement of total phenolics content and antioxidant capacity and the most evident phenomenon of biosynthesis and accumulation of phenolic compounds were both shown in fresh-cut carrot.2.The effect of cut type(whole,0 cm2 g-1;slices,4.5 cm2 g-1;pies,6.2 cm2 g-1;shreds,19.1 cm2 g-1)on respiratory metabonlisn,sugar metabolism,energy metabolism and phenolics biosynthesis of fresh-cut carrot stored at 20 ? for 2 d was investigated.We found that wounding induced the sharp increase of phenylalanine ammonium lyase(PAL)activity and caused the increase of total phenolics content and antioxidant capacity in carrot.Moreover,increased wounding intensity strengthened the increasing trend of these three parameters.The HPLC data indicated that wounding induced the increase of chlorogenic acid,caffeic acid,3,4-dicaffeoylquinic acid,3,5-dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid contents in carrot,among which chlorogenic acid showed the largest increasement after 2 days storage.Moreover,wounding induced the sharp increase of and respiration rate,the decrease of glucose,fructose and sucrose contents and the enhancement of ATP content and energy charge(EC)in carrot.Besides,increased wounding intensity strengthened the activation of respiratory metabolism,sugar metabolism and energy metabolism.The application of fructose,glucose and ATP treatments enhanced the accumulation of phenolic compounds and the increase of antioxidant capacity in fresh-cut carrot.These observations suggested that wound-induced consumption of carbohydrate and ATP's synthesis are related to the biosynthesis of phenolic compounds,and enhancing the supply of carbohydrate and energy wound be helpful for the biosynthesis and accumulation of phenolic compounds.3.The effect of cut type on ROS levels of fresh-cut carrot and the signaling role of ROS in regulating wound-induced biosynthesis and accumulation of phenolic compounds in carrot were investigated.The results indicated that increased wounding intensity induced the generation of O2·-and the accumulation of H2O2 in carrot.The application of NADPH oxidase inhibitor-diphenyliodonium iodide(DPI)reduced ROS generation,suppressed phenylpropanoid metabolism-related enzymes activities of PAL,cinnamate-4-hydroxylase(C4H)and 4-coumarate:coenzyme A ligase(4CL)and restrained the accumulation of phenolic compounds and the increase of antioxidant capacity in shredded carrots.In contrast,the use of H2O2 generator-glucose/glucose oxidase(G/GO)promoted the accumulation of H2O2,induced phenylpropanoid metabolism-related enzymes activities of PAL,C4H and 4CL and enhanced the accumulation of phenolic compounds and the increase of antioxidant capacity in shredded carrots.These observations suggested that ROS play a key role as a signaling molecule for the wound-induced biosynthesis and accumulation of phenolic compounds in carrot,reducing or enhancing ROS levels can successfully modulate the content of phenolic compounds in fresh-cut carrot.4.The effect of NO and the relationship between NO and ROS in regulating wound-induced biosynthesis and accumulation of phenolic compounds in carrot were investigated.The results indicated that increased wounding intensity induced the reduction of NO content and NOS activity in carrot.The application of NO inhibitor-Carboxy-PTIO(cPTIO)reduced NO content,and had no effect on H2O2 content,activities of PAL,C4H and 4CL and the accumulation of phenolic compounds in shredded carrots.The use of NO donor-sodium nitroprusside(SNP)promoted the content of NO in shredded carrots accompanied by enhancing the H2O2 content,activities of PAL,C4H and 4CL and the accumulation of phenolic compounds.However,the positive role of SNP was eliminated by adding DPI,and no significant difference was observed between SNP treatment and combined treatment of SNP and DPI.These observations suggested that ROS are crucial for the biosynthesis and accumulation of phenolic compounds in fresh-cut carrot,exogenous NO can induce the increase of total phenolics content in shredded carrots through enhancing their ROS levels.5.The changes of transcriptional levels of shredded carrots at four stages(0(whole)6^ 24 and 48 h)during storage at 20? were investigated by transcriptome.The results indicated that wounding resulted in the extensive up-regulation of genes related with respiratory metabolism,sugar metabolism,energy metabolism and phenolics biosynthesis,which was in consistent with the results of changes in respiration rate,soluble sugars contents,energy status and total phenolics content.Moreover,many genes involved in the conversion of sugars to phenolics were up-regulated,which elucidates the molecular mechanisms of the conversion of sugars to phenolics in wounded carrot.Besides,the qRT-PCR data basically matched with the RNA-Seq data,suggesting the reliability of results from RNA-Seq.