内源抑制物对乌桕种子萌发的影响

Effect of Endogenous Inhibitors on Seed Germination of Sapium sebiferum

2020年11月份,以采集于江苏省南京市情侣园的乌桕(Sapium sebiferum)种子为供试材料,将种子与湿润细沙按体积比V(种子)∶V(湿润细沙)为1∶3混匀,置于0~5℃低温层积处理(0(未层积)、40、80、120 d),在光照培养箱中(25℃)进行发芽试验,计算发芽率;用甲醇溶液(分析纯),在0~4℃对低温层积处理不同阶段种子胚乳密闭浸提,用系统分离法对提取液进行分离为5个组分(石油醚相、乙醚相、乙酸乙酯相、甲醇相、水相),并以市售白菜矮脚黄(Brassica rapa)种子为对照,在培养皿中加入提取液(以相同体积蒸馏水为对照)进行发芽试验,计算发芽指数,对各分离相的提取液进行乌桕种子浸提液生物活性测定;应用高效液相色谱仪(美国Agelient HP1100)对层积不同阶段种子胚乳中2,6-二叔丁基对甲酚和邻苯二酚的质量浓度进行动态检测、应用气相色谱质谱联用仪(美国Agelient 6890N/5973N)对层积不同阶段种子胚乳中棕榈酸和亚油酸的质量浓度进行动态检测;筛选种子休眠的内源抑制物,分析内源抑制物质对乌桕种子萌发的影响。结果表明:胚乳各分离相中,甲醇相对白菜籽发芽的抑制作用最强,其次是乙醚相。从这两相中筛选出邻苯二酚、棕榈酸、2,6-二叔丁基对甲酚、亚油酸4种抑制种子萌发的物质。在休眠解除过程中,这4种物质的质量浓度,下降幅度均大于50.0%。除亚油酸外,其他3种物质的质量浓度与层积时间、发芽率呈显著负相关。结合发芽抑制物质的有效抑制中质量浓度(即半抑制质量浓度)及层积过程中抑制物质质量浓度的变化,在试验范围内邻苯二酚、棕榈酸是抑制乌桕种子萌发的主要物质,2,6-二叔丁基对甲酚有较弱的抑制作用。

Sapium sebiferum seeds collected in Nanjing Lovers’Garden in November 2020 were used as test materials.The seeds were mixed with moist fine sand at a volume ratio of V(seed)∶V(moist fine sand)of 1∶3.The seeds were treated with low temperature stratification at 0-5℃(0(non stratification),40,80,120 d).The germination test was conducted in a light incubator(25℃)to calculate the germination rate.The methanol solution(analytical pure)was used to extract the endosperm of seeds at different stages of low-temperature stratification treatment in a closed manner at 0-4℃.Use the system separation method to separate the extract into five components(petroleum ether phase,ether phase,ethyl acetate phase,methanol phase,and water phase).With commercially available Brassica rapa seeds as the control,we added the extract(with distilled water of the same volume as the control)in the Petri dish for germination test,and calculated the germination index.The bioactivity of the extracts of S.sebiferum seeds was determined;The contents of BHT and catechol in seed endosperm at different stages of stratification were dynamically detected by high performance liquid chromatography(American Agelite HP1100),and the contents of palmitic acid and linoleic acid in seed endosperm at different stages of stratification were dynamically detected by gas chromatography mass spectrometry(American Agelite 6890N/5973N).The endogenous inhibitors of seed dormancy were screened,and the effects of endogenous inhibitors on seed germination of S.sebiferum were analyzed.The results showed that methanol had the strongest inhibitory effect on the germination of Brassica rapa,followed by ether.From these two phases,catechol,palmitic acid,BHT and linoleic acid were screened to inhibit seed germination.In the process of dormancy release,the contents of these four substances decreased by more than 50.0%.Except linoleic acid,the mass concentrations of the other three substances were negatively correlated with the stratification time and germination rate.Combined with the change of the mass concentration(i.e.semi inhibitory mass concentration)of the effective inhibition of the germination inhibitor and the mass concentration of the inhibitor during the stratification process,it is inferred that catechol and palmitic acid are the main substances that inhibit the germination of S.sebiferum seeds,and BHT has a weak inhibitory effect on cresol.