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文章摘要
基于LOX-HPL途径的武夷肉桂加工中香气物质的形成与调控
The Formation and Regulation of Aroma-related Volatiles During the Manufacturing Process of Wuyi Rougui tea from LOX-HPL Pathway
投稿时间:2019-10-05  修订日期:2019-10-18
DOI:
中文关键词: 脂肪氧合酶途径  武夷岩茶  加工过程  香气  酶活力  基因
英文关键词: LOX pathway, Wuyi RockTea, Manufacturing process, Aroma, Enzyme activity, Gene
基金项目:国家现代农业(茶叶)产业技术体系建设专项(CARS-19);福建农林大学科技创新(CXCX2017178)
作者单位E-mail
周子维 福建农林大学园艺学院 zwchow92@126.com 
刘宝顺 武夷山市幔亭岩茶研究所  
武清扬 福建农林大学园艺学院  
毕婉君 福建农林大学园艺学院  
倪子鑫 福建农林大学园艺学院  
赖钟雄 福建农林大学园艺植物生物工程研究所  
孙云 福建农林大学园艺学院 sunyun1125@126.com 
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中文摘要:
      武夷肉桂是以福建省级茶树良种肉桂(Camellia sinensis cv. Rougui)鲜叶为原料制成的乌龙茶,具有“岩骨花香”的品质特征,而深受消费者青睐。除茶树鲜叶外,武夷肉桂的品质形成很大程度上取决于加工工艺,但生产加工过程中以绿叶挥发物(Green Leaf Volatiles, GLVs)和弥散果酯类(Fruit esters)为代表的脂肪族类香气成分代谢机理的研究,尚未见报道。以武夷肉桂的鲜叶、晒青叶及做青过程叶为试材,采用顶空固相微萃取法(Headspace Solid-Phase Microextraction, HS-SPEM)结合气相色谱与高通量飞行时间质谱联用技术(Gas Chromatography-Time-of-Flight Mass Spectrometry, GC-TOF-MS)对加工过程样中的脂肪族类香气成分进行检测,结果表明,脂肪族醛类在做青过程中显著降低,而大量的脂肪族醇类和酯类显著积累,其中叶醇酯类和己烯醇、酯类分别对静置失水和机械损伤存在响应;利用紫外/可见分光光度法检测加工过程样中LOX-HPL途径中关键酶活力,结果表明,脂肪氧合酶(Lipoxygenase, LOX)、乙醇脱氢酶(Alcohol dehydrogenase, ADH)及乙醇酰基转移酶(Alcohol acyltransferases, AAT)的活力分别在第三次摇青叶(3T)、第四次摇青后叶(4T)及第五次摇青后叶(5T)中达到最大值;实时荧光定量聚合酶链式反应(Quantitative Real-Time Polymerase Chain Reaction, RT-qPCR)结果表明,CsLOX2、CsLOX5基因主要在做青初始阶段(2T、3T)相对表达量较高,而随着工艺过程的推进,CsADH2-1、CsAAT基因在做青后期(4T、5T)开始上调表达;通过Pearson系数评估武夷肉桂加工过程中的脂肪族类香气、酶活力和基因表达量间的相关性,结果发现,正己醇、戊酸叶醇酯、己酸己酯、丁酸甲基叶醇酯、柳酸叶醇酯等大量具有天然花果香的脂肪族醇类和酯类与CsADH2-1基因具有显著(P<0.05)或极显著(P<0.01)的正相关,CsAAT基因与CsLOX2、CsLOX5基因在加工过程中具有相似的表达模式(r=0.762,r=0.732)。LOX-HPL途径关键基因的共表达,驱动了脂肪族类香气在加工过程中的协同积累,这可能是武夷肉桂馥郁香气形成的重要基础。
英文摘要:
      Abstact: Wuyi Rougui is a kind of oolong tea made from the fresh leaves of Camellia sinensis cv. Rougui which is an improved tea tree variety of Fujian Province,?and characterized with ‘rock in the throat, flower in the mouth', deeply favored by consumers. Besides fresh tea leaves, the aroma quality of Wuyi Rock tea depend greatly on manufacturing process. However, studies on the metabolic mechanism of aliphatic aroma components represented by green leaf volatiles (GLVs) and fruit esters during manufacturing process have not been reported. Fresh tea leaves, solar-withering leaves and turn-over leaves of Wuyi Rougui were used as test materials. We use headspace solid phase microextraction (HS-SPEM) combined with gas chromatography time-of-flight mass spectrometry (GC-TOF-MS) to detect aliphatic aroma components in manufacturging process samples. The result showed that the aliphatic aldehydes were significantly reduced during the turn-over process, and a large amount of aliphatic alcohols and esters were formed and accumulated. Among them, leaf alcohol esters and hexenyl esters and alcohols might have responses to water loss and mechanical damage respectively during the manufacturing proecess. The results of UV/visible spectrophotometry for the detection of key enzyme activities in the LOX-HPL pathway in manufacturing process samples indicated that The activities of lipoxygenase(LOX), alcohol dehydrogenase(ADH) and alcohol acyltransferases (AAT) reach their peak values in the third time , the fourth time and the fifth time turn-over leaves respectively. The results of quantitative reverse transcription polymerase chain reaction(qRT-PCR) demonstrated that CsLOX3-2 and CsLOX5 genes had a higher relative expression levels in the initial stage of turn over (2T, 3T), and with the advancement of the manufacturing process, relative expression of CsADH2-1、CsAAT genes were up-regulated during the late stage of turn over (4T, 5T). Evaluating the correlation between aliphatic aroma, enzyme activity and gene expression during manufacturing pocess of Wuyi Rougui by Pearson coefficient, we found that a large number of aliphatic alcohols with natural floral and fruity flavors such as 1-Hexanol, cis-3-Hexenyl isovalerate, Hexanoic acid, hexyl ester, cis-3-Hexenyl-methylbutyrate and cis-3-Hexenyl salicylate, had significantly(P<0.05) or extremely significantly (P<0.01) positive correlation with the CsADH2-1 gene; meanwhile, CsAAT gene had similar expression pattern with CsLOX3-2 and CsLOX5 genes during manufacturing process(r=0.762,r=0.732). Co-expression of key genes in the LOX-HPL pathway drove the synthesis and accumulation of aliphatic aromas during manufacturing process, laying an important basis for the formation of the aroma-related volatiles of Wuyi Rougui.
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