The Regulation Crucial Enzymes In Esters Biosynthesis For Aroma Foding Of Refrigerated ’Nanguo’ Pears

’Nanguo?pears are native to Liaoning province.The fruit,which is normally picked in early September when it has a crisp flesh with little juice and no aroma.After ripening,it matures to produce a ripe fruit with juicy and has soft and smooth flesh,a sour-sweet taste.However,the storage time of’Nanguo’ pears is short,they can just store for half a month to up optimal taste period,and quickly senesce.Nowadays,cold storage is the primary postharvest technology to prolong the storage period.However,the fruit exhibits reduced aroma after optimal taste period,resulting in reduced commodity value.Therefore,it is significant to study on the mechanism of reduced aroma quality of ’Nanguo’ pears.This study systematically analyzed the changes of aroma components,contents of the esters,activities of key enzymes and protein expressions during postharvest natural ripening and shelf life after cold storage of’Nanguo’ pears.Low temperature conditioning(LTC)treatments were applied to regulation of the reduced aroma of’Nanguo’ pears after cold storage.The main results are as follows:(1)Application of Western blot to analyze the activities and protein expressions in the aroma metabolism pathway of Nanguo pear,optimization of Western blot for the detection of protein expression in ’Nanguo’ pears.The optimal conditions of Western blot are as follows:Trizol was used to extract the total protein.The total protein was separated by polyacrylamide gel electrophoresis(SDS-PAGE)and the optimal sample size was 50μg.The best temperature and time for antibody incubation is first antibody at room temperature,shaking under the conditions of 2 hours,the second antibody incubate for 1 hour at room temperature under shaker conditions.ECL was used to analyze result,the optimal dilution ratio of Actin protein was determined to be 1:400,the optimum dilution ratio of LOX protein was 1:200,and the optimal dilution ratio of AAT protein was 1:600.(2)The variety and contents of esters significantly decreased of fruit in shelf life after cold stored for 90 d and 180 d.The longer the period of cold storage of ’Nanguo’ pears,the severer the degree of reduced aroma.The maximum of activity of LOX and AAT,and protein expression of LOX appeared in 3 days advance in the fruit cold stored for 180 d than 90 d.Compared to cold stored for 90 d,markedly lower activity of AAT and level of protein expression of LOX and AAT were observed in the pears cold stored for 180 d.The expression of LOX protein,the activity of AAT and the expression of corresponding protein were inhibited during optimal taste period after cold storage,and the inhibitory effect was more obvious with the prolongation of cold storage.These results indicated that long-term refrigeration inhibit the activity and protein expression of key enzymes to a certain extent,thus influence the generation of aroma-related esters.which reduce the aroma quality of cold stored ’Nanguo’ pears.(3)Low temperature condition treatment promoted the aroma-related esters of refrigerated fruit during refrigeration and shelf life after refrigeration,the main aroma components were significantly higher than that after cold storage.LTC increased the activity and protein expression of LOX and AAT,which may promote the aroma formation of’Nanguo’ pears to certain extent.