Mechanism Of Postharvest Apple With Fresh Bruises Based On Metabolomics

Apple is one of the major types of fruit cultivated in China and it plays an important role in modern agricultural production and widely consumed.During the processes of picking,sorting,and transporting,apple fruits are subject to extrusion,oscillation,striking,and collision,which can cause mechanical damage to apple tissue.Fresh bruise usually occurs under apple skin,which are difficult to distinguish with the naked eye during an early stage.As the time elapses,the apple tissue with fresh bruise is susceptible to bacterial and fungal infections,eventually leading to apple rot,and even reducing the quality of the same batch of high-quality fruit.Because of this,it brings huge quality loss and economic loss.Therefore,it is necessary to research the early metabolic mechanism of fresh bruise apples after harvest.Based on the plant metabolomics strategy,this paper studied the primary metabolites,some small molecular metabolites and volatile metabolites of healthy and fresh bruise apples during the early storage process.This study has not only expounded the primary metabolism and volatile metabolism mechanism in fresh bruised apple,but also determined the optimal time of reducing loss of apple after fresh bruise.The main contents and results of this study are as follows:（1）We analyzed the primary metabolic response in apple fruits during 24 h storage after fresh bruise by using non-targeted metabonomics method based on GC-MS and multivariate statistical analysis.We selected seven time points to monitor the metabolic disorder and delineated disorder-induced physiological changes and their primary metabolic mechanisms.Based on the discrimination and similarity analysis by PLS-DA,we observed the maximum separation of healthy and bruised apple samples at 12 h;this indicates that the levels of metabolites in apple fruits changed significantly at 12 h after damage,and provides an exact time for reducing loss and retaining freshness of apple post-harvest.Through the metabonomics analysis,56 compounds were identified from healthy and bruised apple tissues.We found no significant effects on the types of major metabolites within 24 h after compared with before damage,yet the contents of these metabolites markedly differed across different storage time points.While sugars and alditols increased within 24 h,amino acids,fatty acids,fatty acid lipids all decreased.We observed less accumulation of Sur,Fru,Glu,Glyce,and Man in bruised tissue than healthy tissue,and this was related to an enhancement of respiration in apple tissue after fresh bruise.Mal,Sal,and Man accumulation in bruised tissue was lower than that in healthy tissue,which was attributable to the increase of ROS in apple tissue after fresh bruise.There was a larger decrease of Gly in bruised compared with healthy tissue,indicating the synthesis of glutathione was accelerated in apple tissue with fresh bruise,aimed at eliminating the adverse effects of ROS on the cells.Less accumulation of quinate in bruised tissue than healthy tissue prompts that secondary metabolism was activated after fresh bruise.The accumulation of inositol in bruised tissue was less than that in healthy tissue,which was related to the repair of cell walls after fresh bruise.The decrease of Ole in bruised tissue was greater than that in healthy tissue,suggesting cell senescence and apoptosis was accelerated after fresh bruise.（2）Raman spectroscopy was used to study the small molecule metabolites in apple fruits after fresh bruise.We used the savizky-golay smoothing method and wavelet transform algorithm to remove the noise.The results showed that the first-order three-time 15-point savizky-golay smoothing method had the best denoising effect.Small molecular metabolites,such as fructose,lycopene,chlorophyll a,carotene and phenolic compounds,were identified by analyzing the Raman spectra of healthy and fresh bruise apples.Compared with the Raman spectra of healthy apples,the Raman spectra of fresh bruise apples at 455,645,824,871,1270,1521 and 1631 cm^-1 showed a decrease in the intensity of characteristic peaks or even no characteristic peaks.The results indicated that the contents of carotene,lycopene,fructose,phenolic compounds,etc,in the apple flesh tended to decrease,providing an effective means for the development of a complete metabolic network in apple fruits after fresh bruise.（3）HS-SPME-GC-MS was used to analyze the content changes of volatile metabolites and the secondary metabolic mechanism of fresh bruise apples.There was a significant difference in the volatile matter spectra of bruise and healthy tissues,and a transitional change occurred at 12 h by PLS-DA.This is consistent with the results of primary metabolism studies,and further indicates that 12 h is determined to be the optimal time for loss reduction in fresh bruised apple.A total of 55 substances were identified through qualitative and semi-quantitative analysis of volatile substances in tissues by TCI diagrams.According to their different properties,they can be divided into aldehydes,ketones,esters,terpenes,alcohols and others.Within 0-24 h,the content of volatile metabolites of alcohols and esters（mainly hexyl esters）first increased and then decreased in the damaged tissues,which was related to the oxidation of unsaturated fatty acids,but limited by the content of unsaturated fatty acids.The increase of farnesene content in the injured tissue is mainly related to the activation of secondary metabolism after minor injury.The content of volatile metabolites of aldehydes and ketones increased in the damaged tissues,especially the decanal showed a monotonic increasing trend with time,so it can be used as an iconic metabolite.