切花茎基端酚类代谢与采后茎堵塞的关系及其机理研究

Cut flowers are very prone to stem blockage during their postharvest period, which directly disrupts their ornamental quanlity and commodity value. In our preliminary study, it was found that the phenolic metabolism induced by cutting wound in the stem-ends of cut flowers is evidently involved in the physiological blockage in the tested cut flowers. But so far there are few research reports on the phenolic metabolism in the stem-ends of cut flowers, and its influencing mechnism on the water transportation in their stems need to be elucidated. Hence, according to this research thought and our related preliminary experimental results, the present project aims at exploiting and understanding the inherent relationships between phenolic metabolism and stem blockage in cut flowers and its mechanism, based on physical and chemical analyses of main phenolic compounds and metabolites, the investigations in histochemistry and microstructure (i.e. the spatial-temporal feature of phenolic compounds and their metabolites deposition in the cut ends and its effects on the morphology and structure of water-conducting tissue in cut flowers), biochemistry and molecular biology (i.e. the induction of the activities and gene expressions of PAL, PPO and POD, the key enzymes in the phenolic metabolism, and its relation to the formation of phenolic compounds and their metabolites in cut stem-ends), and postharvest physiology (i.e. the effects of various postharvest treatments and its control on phenolic metabolism and their effects on stem blockage and ornamental quality of cut flowers), using cut lily and rose flowers as the main experimental materials. The results from this project will help to reveal the features of phenolic metabolism in stem-ends of cut flowers and its regulation mechanism, and to address a new approach to inhibit phenolic metabolism and to alleviate the stem blockage, and thereby to improve the water transportation in cut flower stems.

切花采后极易发生茎堵塞，并直接影响其采后品质和商品价值。在本人前期研究中发现采切创伤引起的茎基端酚类代谢可能参与切花采后的生理性堵塞。但目前国际上缺乏对切花茎基端酚类代谢的研究报道，且对其影响切花采后水分输导功能的机理尚不清楚。为此，基于这一思路和前期研究基础，本项目以采后茎基端酚类代谢和茎堵塞均突出的百合、月季等主流切花为研究材料，采取理化分析、组织化学与显微结构观察、酶活性分析、基因克隆与表达分析等技术手段，并结合相关采后处理（外源酚、酚类代谢关键酶抑制剂、茎基端热水处理和不同贮运环境条件等）对切花茎基端酚类代谢的调控作用以及对切花采后茎堵塞和观赏品质的影响进行研究，系统探讨和揭示茎基端酚类代谢与切花采后茎堵塞的内在关系及其机理，并为形成和提出有效控制切花采后茎堵塞的新策略与处理技术提供科学依据。此外，研究成果对果蔬贮运与加工、植物组培等酚类代谢发生与调控研究也具有重要的借鉴意义。

采切创伤诱导的茎基端酚类代谢被视为切花采后生理性茎堵塞的重要原因。本项目着重研究部分切花采后茎基端酚类代谢特点以及它们酚类代谢和茎堵塞的内在关系及其调控机制。首先建立了切花酚类代谢关键酶PPO和POD的测定系统，并用之分析显示月季、香石竹等切花茎基端POD和PPO活性在模拟贮藏期间变化的趋势大体一致，即：0~12 h内各切花茎基端PPO和POD活性虽有一些波动但总体活性水平并不高，12 h后快速上升，至48 h或72 h后降低。利用HPLC-DAD、LC-ESI-MS/MS及HRMS分析表明，月季和百合切花茎基端酚类物质种类存在较大差异，月季切花茎基端酚类物质主要是酚酸类、单宁类和黄酮类，百合切花茎基端酚类物质主要是酚酸类和黄酮类。从百合切花共克隆得到LhPPO1、LhPPO2和LhPPO3共3个PPOs基因全长序列，在LhPPO1和LhPPO2基因上游启动子序列均存在植物启动子基本转录元件及创伤、病原菌、植物激素等响应元件。转录组学研究表明，共有7 805个差异表达Unigenes参与百合‘蒂伯’切花茎基端对创伤的响应，创伤早期（6 h内）有大量Unigenes上调表达，随后下调表达基因的数目则逐渐增多。其中富集程度高且可靠的前3个途径分别为植物激素信号转导、植物病原体互作和苯丙烷生物合成，意味着这些途径在百合切花响应切割创伤中起着极为重要的作用。另外，一系列创伤反应相关的转录因子如WRKY, NAC, ERF和MYB等在采切创伤后明显上调。在月季、百合等切花瓶插和模拟贮藏期间茎基端褐变进程中，可见到酚类代谢物在切口及邻近处集聚和侵占切花的水分输导组织。采用石蜡切片和扫描电镜可明显观察到香石竹和唐菖蒲等切花在瓶插期间其茎基端木质部导管被Pb(CH3COO)2染成黑色物质的醌类物质存在。研究证实适宜浓度的多种POD和PPO抑制剂采后处理可有效减轻月季切花和绢毛相思切叶瓶插期间的茎堵塞，明显改善其水分关系和观赏品质，并延长瓶插寿命。总之，本项目研究所取得的有关成果有助于阐释切花茎基端酚类代谢发生与调控机制，并可望为形成相应的切花采后保鲜与处理技术提供科学依据。