Study On The Browning Mechanism And Regulation Of Upper Tobacco Leaves Based On Lipid Metabolism

Cuticular wax is a secondary metabolic product formed during the long-term resistance physiology of the plant,and is an important cause of ash hanging on the upper leaves and reduced tobacco availability.In order to safeguard the level of raw material and economic efficiency of tobacco,solving the problem of upper leaf ash hanging has become an inevitable need.It is important to understand the physiological mechanisms of lipid metabolism on tobacco browning to control browning.In this study,we systematically analysed the pattern of yellowing and browning and the changes of lipid metabolism-related indexes during tobacco drying,and used the upper leaves of Yunnan Tobacco 87 and Zhongchuan 208 varieties as test materials.In order to further clarify the effect of curing process on the browning and lipid metabolism of tobacco leaves,the high temperature and medium humidity curing process was used to reveal the effect of lipid oxidation on the relationship between lipid oxidation and browning of tobacco leaves under different curing processes at the molecular level and to explore the technology of tobacco leaf browning regulation.The main findings are as follows:1.The pattern of yellowing and browning of upper tobacco leaves during curing and the changes of lipid metabolism-related indicators was clarified.The drying rate of leaves and main veins decreased,chlorophyll in the leaves was degraded,colour value of a* increased and the leaves started to brown;the fatty acid content of tobacco leaves was effectively reduced during curing,and the proportion of medium to top grade tobacco and light browning increased.2.The relationship between upper leaf lipid metabolism and tobacco browning was analysed.Further transcriptome GO analysis and KEGG enrichment analysis showed that the upper leaves of Zhongchuan 208 were more enriched in lipid metabolism(linolenic acid and linolenic acid metabolism).The results of the GO analysis and KEGG enrichment analysis showed that the upper leaves of Zhongchuan 208 were more enriched in lipid metabolism(linolenic acid and linolenic acid metabolism)than the upper leaves of Yunnan Tobacco 87,indicating that lipid oxidation caused by fatty acid metabolism was an important factor in the browning of the upper leaves.3.The effects of different picking and curing methods on lipid Browning in upper leaves of flue-cured tobacco were studied.The browning rate of tobacco leaves in different harvesting treatments were ranked as follows: 6 leaves with stem curing treatment < 4 leaves with stem treatment < 2 leaves with stem curing treatment <normal harvesting treatment,and the browning reaction was inhibited in the stem curing treatment.The ultrastructure observations showed that the cuticle thickness decreased with increasing number of leaves during curing in the stemmed treatment.The expression of genes related to fatty acid metabolism was significantly lower in the six-leaf stemmed treatment compared to the CK treatment(P<0.05),and the relative expression of key genes for lipid oxidation was down-regulated in the six-leaf stemmed treatment.Correlation analysis showed that cuticle thickness and LOX activity were highly significantly and positively correlated with oleic and linolenic acids,the result showed that fatty acid metabolism is one of the important causes of tobacco browning.4.The effects of different curing processes on the regulation of lipid browning in the upper leaves of cured tobacco were compared.The proportion of upper and middle grade tobacco increased under the high-temperature medium-humidity process,the proportion of tobacco leaf browning decreased and the browning response was suppressed.Further study by transcriptome sequencing revealed that the differential genes were significantly enriched in lipid metabolism pathways in addition to phenylalanine,flavonoids and sugar metabolism pathways.A significant reduction in multiple fatty acid and lipoxygenase(LOX)activities was observed in the high-temperature medium-humidity curing process compared to the control treatment.Compared with the control treatment,the high-temperature medium-humidity curing process was more effective in maintaining the leaf color,significantly inhibiting browning reactions,reducing LOX activity and improving the grade of tobacco leaves.The results provide a further theoretical basis for research on the control of quality in tobacco curing and the relationship between browning and lipid metabolism.