基于LOX-HPL途径的武夷肉桂加工中香气物质的形成与调控
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Formation and Regulation of Aroma-Related Volatiles During the Manufacturing Process of Wuyi Rougui Tea via LOX-HPL Pathway
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    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,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)和果酯类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检测发现: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,qRT-PCR)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,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,是武夷肉桂馥郁香气形成的重要基础。

    Abstract:

    Wuyi Rougui tea is a kind of oolong tea made from the fresh leaves of Cinnamomum cassia(Camellia sinensis cv. Rougui), which is a provincial improved tea tree variety in Fujian Province and deeply favored by consumers because of the characterization of 'rock in the throat, flower in the mouth'. Besides fresh tea leaves, the aroma quality of Wuyi Rougui tea depend greatly on the manufacturing process. However, studies on the metabolic mechanism of aliphatic aroma components represented by green leaf volatiles (GLVs) and fruit esters during process have not been reported. Fresh tea leaves, solar-withering leaves and turn-over leaves of Wuyi Rougui tea were used as test samples. The detection of aliphatic aroma components in samples during manufacture was done by headspace solid phase microextraction (HS-SPEM) combined with gas chromatography time-of-flight mass spectrometry (GC-TOF-MS) . The aliphatic aldehydes were significantly reduced during the turn-over process, while a large amount of aliphatic alcohols and esters were formed and accumulated. Among them, leaf alcohol esters, hexenols and hexenyl esters could responded to water loss and mechanical damage, respectively. The key enzyme activities in the LOX-HPL pathway during process was detected by UV/visible spectrophotometry. The results indicated that the activities of lipoxygenase(LOX), alcohol dehydrogenase(ADH) and alcohol acyltransferases (AAT) reached their peak values in the third time(3T), the fourth time(4T) and the fifth time(5T) of turn-over process, respectively. The results of quantitative reverse transcription polymerase chain reaction(qRT-PCR) demonstrated that CsLOX2 and CsLOX5 genes had a higher relative expression levels in the initial stage of turn over (2T, 3T), and the up-regulation of relative expression of CsADH2-1 and CsAAT genes was observed during the late stage of turn over (4T, 5T) when the process advanced. The correlation between aliphatic aroma, enzyme activity and gene expression during manufacturing pocess of Wuyi Rougui tea was evaluated by Pearson coefficient. 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 significant(P<0.05) or extremely significant(P<0.01) positive correlation with the CsADH2-1 gene. Meanwhile, CsAAT gene had similar expression pattern with CsLOX2 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 manufacture, laying an important basis for the formation of the aroma-related volatiles of Wuyi Rougui tea.

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周子维,刘宝顺,武清扬,毕婉君,倪子鑫,赖钟雄,孙云.基于LOX-HPL途径的武夷肉桂加工中香气物质的形成与调控[J].食品与生物技术学报,2021,40(1):100-111.

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  • 在线发布日期: 2021-04-07

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