内源抑制物对加拿大紫荆种子萌发的影响

Effects of endogenous inhibitors on seed germination of Cercis canadensis

【目的】探究内源抑制物质对加拿大紫荆(Cercis canadensis)种子萌发的影响,为揭示加拿大紫荆种子的休眠原因及其解除机制提供依据。【方法】以加拿大紫荆种子为试验材料,先采用80℃热水预处理,再经低温层积处理解除种子的生理休眠,对不同层积阶段的种子进行发芽率和生物活性测定。结合气质联用(GC-MS)、高效液相色谱(HPLC)技术与标准品有效半抑制浓度(IC_(50))方法,分析加拿大紫荆种子内源抑制物种类及休眠解除过程中内源抑制物含量的变化。【结果】内源抑制物是引起加拿大紫荆种子休眠的主要原因,随着层积时间延长,休眠逐渐解除。加拿大紫荆种子种皮和胚乳各分离相提取液对白菜籽发芽抑制作用由大到小依次为乙醚相>甲醇相>水相≈乙酸乙酯相>石油醚相,表明种皮和胚乳中的内源抑制物主要存在于乙醚相、甲醇相、乙酸乙酯相提取液中。采用GC-MS技术测得加拿大紫荆种皮和胚乳分离相提取液中主要有油酸、亚油酸、棕榈酸、硬脂酸、2,6-二叔丁基对甲酚(BHT)、邻苯三酚6种可能对加拿大紫荆种子具有抑制作用的物质。对6种物质标准品进行有效半抑制浓度(IC_(50))测定发现:油酸对白菜籽发芽无抑制作用,较高浓度的亚油酸、棕榈酸、硬脂酸才能抑制白菜籽发芽,而较低浓度的BHT和邻苯三酚对白菜籽发芽就有很强的抑制作用。种子休眠解除过程中,亚油酸含量呈先下降后上升再下降的趋势,而棕榈酸、硬脂酸、BHT、邻苯三酚的含量逐渐下降,层积后的4种物质含量下降幅度分别为42.43%、52.00%、5.77%、96.14%。【结论】结合IC_(50)及层积过程中内源抑制物含量的变化,认为邻苯三酚可能是引起加拿大紫荆种子休眠的主要抑制物质。

【Objective】 The relationship between endogenous inhibitors and seed dormancy of Cercis canadensis was explored to provide evidence for revealing its seed dormancy and the mechanisms for breaking dormancy.【Method】 Stratification treatments at cold temperatures were applied to break the seed dormancy of C.canadensis after 80 ℃ water pretreatment.The germination percentage and the bioactivity of seeds at different stratification stages were determined.Combined with GC-MS/HPLC and effective semi-inhibitory concentration(IC_(50)),the endogenous inhibitor types in the seed of C.canadensis and the content of these endogenous inhibitors in the dormancy breaking process were analyzed.【Result】 The endogenous inhibitors in the seed of C.canadensis resulted in seed dormancy.The dormancy was gradually released along with the time of stratification.The inhibition of the C.canadensis seed coat and the endosperm separation on the germination of cabbage seeds was ether phase > methanol phase > water phase ≈ ethyl acetate phase > petroleum ether phase.This indicated that endogenous inhibitors in the seed coat and the endosperm mainly existed in the ether phase,the methanol phase,and the ethyl acetate phase of the seed coat-endosperm extraction.Six inhibitor types in the seed coat-endosperm separation of C.canadensis were identified using GC-MS,including oleic acid,linoleic acid,palmitic acid,stearic acid,2,6-di-tert-butyl-p-cresol(BHT) and pyrogallol.The IC_(50) was performed to determine these six standards and results showed that oleic acid had no inhibitory effect on the seed germination of cabbage.A higher concentration of linoleic acid,palmitto acid,and stearic acid inhibited cabbage germination,while lower concentrations of BHT and pyrogallol strongly retarded cabbage germination.In the dormancy release process,linoleic acid content resulted in a “decrease-increase-decrease” trend.Meanwhile,the content of palmitic acid,stearic acid,BHT and pyrogallol presented a decreasing trend,decreasing by 42.43%,52.00%,5.77% and 96.14%,respectively.【Conclusion】 By combining IC_(50) and the changes of inhibitor content in the stratification process,it was concluded that pyrocatechol may be the main inhibitor of C.canadensis seed dormancy.