Study On The Aroma Evaluation Of Citron Resources In Yunnan And Tibet Region Of China And The Causes Of Lemon Fruit Appearance Defects

Yunnan and Tibet region of China possesses abundant citrus resources,including common cultivated varieties and some special wild and semi-wild citrus germplasm resources.In recent years,citron（Citrus medica L.）and lemon（Citrus limon L.）,as featured citrus germplasms in this region,have received plenty of attention in the targeted poverty alleviation of China.One distinguishing feature of lemon and citron is that their peel and pulp are both edible,and the flavor and appearance of the peel are the most important sensory quality traits that directly affect the preference of the consumers.However,due to the remote location and the limitations of local research conditions,little research has been conducted on these citron and lemon resources and the key factors that restrict the industrial development in this region.On the one hand,the abundant citron resources remain largely unexplored,and the lack of investigations on the unique aroma substances in citron fruit seriously restricts the industrialization and integration of these resources.On the other hand,the defects in the appearance of fruit surface caused by uneven degreening and oleocellosis have become a bottleneck that restricts the development of green lemon industry in Yunnan.In this study,to better utilize the characteristic citrus resources in this region and promote the development of the local citrus industry,the volatile components of these characteristic citric citrus resources were systemically analyzed by GC-MS,and the causes of the uneven degreening of citron and oleocellosis of green lemon were also studied.The research results are as follows:1 Differences in peel volatile components between citron and lemon from different regions（1）The composition of volatile substances in the fruit peel of 179 citron and 30lemon germplasms showed certain similarities,with high accumulations of monoterpenes,particularly d-limonene.However,the total content of volatile substances in the peel of most lemon germplasms,such as monoterpene,monoterpene aldehyde,monoterpene alcohol and sesquiterpene,was higher than that of citron germplasms.Least-squares analysis（PLS-DA）identified nine characteristic substances with VIP>1.4,which could significantly distinguish lemon and citron.Among them,the four substances with VIP>1.50 were d-limonene,β-myrcene,eremophilene,and perillaldehyde.Among all the nine characteristic substances,eremophilene,perillaldehyde,and sabinene hydrate were only present in lemon peel;while 3-carene,α-phellandrene and isoterpinolene were exclusively detected in citron peel.（2）Environment showed significant effects on the composition and content of volatile substances in lemon and citron germplasms.The results of principal component analysis（PCA）and PLS-DA analysis on volatile substances in the peel can significantly distinguish the lemon germplasms from Yunnan and Tibet,as well as those from dry&hot areas and humid&hot areas in Yunnan.3-Carene,α-phellandrene and isoterpinolene were not detected in 129 samples from Yunnan,while 3-carene was detected in a high content from the Tibetan germplasm,which can be used as a characteristic volatile substance to distinguish citron from Yunnan and Tibet.The total content of volatile substances in the peel of lemon germplasms in the humid&hot regions was significantly higher than that in the dry&hot regions of Yunnan,but eremophilene,citronellol acetate and geraniol only showed significant accumulations in the lemon germplasms from the dry&hot regions.2 Causes and regulatory mechanism of uneven degreening of postharvest green lemon（1）The color index（CI）and the chlorophyll a and chlorophyll b content of the degreening part were significantly lower than those of the green part in uneven degreening green lemon.The degreening part showed an increase in surface wax and a significant reduction of stomata density.Intact chloroplasts were observed in the green part.However,in the degreening part,the chloroplasts gradually changed to chromoplasts,and there were significant accumulations of peel phytofluene and phytoene and increases in ABA content,while the IAA,BR and GA₃contents showed significant decreases.CEPA treatment accelerated the degreening of lemon peel,and at the same time increased the ABA and Me-JA content in the degreening part of the peel.The above results suggest that the uneven degreening of green lemon may be closely related to the uneven distribution of peel hormones.（2）The best exogenous hormone treatment for lemon was 50 mg/L GA₃and1000 mg/L CEPA treatment.CEPA treatment reduced the content of chlorophyll and its derivatives as well as resulted in the accumulation of Phytofluene and Phytoene,and at the same time induced the synthesis of wax,the accumulation of endogenous hormone ABA and reduction of IAA content.Moreover,CEPA treatment significantly up-regulated the expression of PPH,NYC1（the chlorophyll degradation pathway）,PSY,ZED（the carotenoid synthesis pathway）,ACO1,ACO5,ACS8,ACS1 and the ethylene receptor ETR2（the ethylene synthesis and signal pathway）,and LOX,AOC4,JMT（the jasmonate synthesis pathway）.In addition,CEPA treatment significantly up-regulated the expression levels of different genes related to ABA synthesis in peel,namely NCED1 and ZEP,and significantly down-regulated the expression levels of ABA catabolism-related genes CYP707A2 and CYP707A4.The GA₃treatment showed exactly opposite effects of the CEPA treatment.The above results indicate that exogenous GA₃and CEPA treatment regulate endogenous hormone and pigment content by regulating the expression of endogenous genes related to hormone synthesis,signal transduction and pigment metabolism,thereby affecting the peel color.3 Cause of lemon oleocellosis（1）There were significant decreases in primary metabolites（particularly glucose）and increases in secondary metabolites（particularly lignin）in oleocellosis-damaged flavedo.The biosynthesis of phenylpropanoid was significantly activated,and there were 79 DEGs related to lignin accumulation,among which 65 genes（82.3%of the total DEGs）were significantly up-regulated,especially the PAL,C4H,CCR,F5H POD and UGT72E genes.Oleocellosis-damaged flavedo showed higher lignin accumulation and enhanced defense against pathogenic fungi.（2）Oleocellosis disorder induced the accumulation of C16 and C18 free saturated and unsaturated fatty acids in lemon flavedo.The total content of epicuticular wax showed no significant differences between healthy and oleocellosis-damaged flavedo,but oleocellosis significantly increased the total alkane content and decreased the aldehyde content in the flavedo.The expression of Fab Z,FAD2 and SAD6 involved in the accumulation C16 and C18 fatty acids and CER1 gene involved in the accumulation of VLC fatty alkanes was up-regulated.In summary,this study established the volatile substance spectrum in the fruit flavedo of 179 citron and 30 lemon germplasms in Yunnan and Tibet,and obtained the distribution and accumulation characteristics of main aroma components.It was found that the uneven degreening of green lemon may be closely related to the uneven distribution of peel hormones.Exogenous GA₃and CEPA treatment regulate endogenous hormone and pigment content by regulating the expression of endogenous genes related to hormone synthesis,signal transduction and pigment metabolism,thereby affecting the peel color.The effect of oleocellosis on metabolite accumulation in pericarp of green lemon was analyzed.The results provided a theoretical basis for the development and utilization of the characteristic citrus resources and the research and development of the prevention and treatment technology of the uneven degreening and oleocellosis of green lemon after harvest.