NaCl胁迫下Ca~(2+)调控发芽大豆生理代谢和γ-氨基丁酸含量变化

为研究NaCl胁迫下Ca~(2+)对发芽大豆主要生理指标和γ-氦基丁酸(gamma-aminobutyric acid,GABA)富集的调控作用,利用CaCl_2和乙二醇二乙醚二胺四乙酸(ethylene glycol tetraacetic acid,EGTA)处理发芽大豆,研究NaCl胁迫下外源和内源Ca~(2+)对发芽大豆主要生理代谢和GABA含量的影响。结果显示,发芽大豆经NaCl联合CaCl_2处理,其芽长和呼吸速率显著增加,表明CaCl_2缓解了NaCl对发芽大豆生长的抑制,同时过氧化氢酶及过氧化物酶活力显著提高,说明CaCl_2可能是通过提高抗氧化酶活力来缓解NaCl胁迫下发芽大豆的抑制效应,而施用EGTA则呈相反的变化趋势;NaCl联合CaCl_2处理后发芽大豆中GABA含量与单独NaCl处理无显著差异,但显著高于对照组;在NaCl联合CaCl_2或EGTA基础上施用氦基胍,发芽大豆子叶中GABA含量分别下降17.1%和9.8%,胚中分别下降26.5%和8.5%,表明NaCl胁迫下施用CaCl_2在促进发芽大豆中GABA富集的同时还可保证生物产量,且CaCl_2和EGTA处理下多胺降解途径对GABA富集贡献降低。     

低氧联合NaCl胁迫下外源Ca2+对发芽苦荞γ-氨基丁酸富集的影响

采用在通气的培养液中添加外源Ca~(2+)培养的方式,研究了低氧联合NaCl胁迫下外源Ca~(2+)对发芽苦荞GABA富集的影响。低氧联合NaCl胁迫对苦荞的芽长有抑制作用,GAD活力和GABA的含量随着NaCl浓度的增大均呈先上升后下降的趋势,在10 mmol/L NaCl溶液处理时均达到最高值,分别是单纯低氧胁迫的1.65倍和1.10倍;在低氧联合NaCl胁迫的基础上添加低浓度的Ca~(2+)可缓解NaCl胁迫对苦荞芽长的抑制作用,添加3 mmol/L Ca~(2+)处理可使GAD活力和GABA的富集量达到最大,分别为91.44 U/g FW和492.31μg/g DW,分别是低氧联合NaCl胁迫的1.12倍和1.16倍;低氧联合NaCl胁迫下添加Ca~(2+)螯合剂和通道阻断剂后发芽苦荞芽长显著降低,GAD活力显著下降,GABA富集量减少。外源Ca~(2+)可降低低氧联合NaCl胁迫对苦荞芽长的抑制作用,并可通过提升GAD活力,增强发芽苦荞GABA的富集。     

钙离子通道抑制剂处理下发芽大豆主要生理生化和γ-氨基丁酸的代谢变化

为研究钙离子通道抑制剂对Na Cl胁迫下发芽大豆生理代谢和γ-氨基丁酸(GABA)富集的影响,通过Na Cl联合质膜钙离子通道抑制剂(La Cl3)和内膜钙离子通道抑制剂(Heparin)处理大豆籽粒,分析发芽期间主要生理生化和GABA代谢酶活力的变化。结果显示,发芽大豆经Na Cl联合La Cl3处理后,其生长发育较单独Na Cl处理进一步受到抑制,芽长显著降低,经Heparin处理芽长无显著性变化,而二者呼吸速率均显著增加。相较单独Na Cl处理,Na Cl联合La Cl3或Heparin处理后其子叶中GABA代谢酶活力均显著下降,La Cl3处理后子叶和胚中GABA含量分别为Na Cl处理的50%和79%,而Heparin处理则分别为70%和66%,表明抑制内源钙库的释放对Na Cl胁迫下GABA富集的影响小于抑制质膜钙离子通道。     

大豆发芽富集γ-氨基丁酸的培养液组分优化及盐胁迫下的富集机理

以东北大豆为原料,研究培养液组分对大豆发芽富集γ-氨基丁酸(γ-aminobutyric acid,GABA)的影响,利用响应面法优化了大豆发芽富集GABA的培养液组分,在此基础上对低盐胁迫下大豆发芽富集GABA的机理进行研究。结果表明:优化后有效的培养液组分为谷氨酸钠1.0 mg/mL、磷酸吡哆醛2.0 mmol/L、CaCl_2 2.0 mmol/L、NaCl 100 mmol/L,在此条件下,富集得到的发芽大豆中GABA含量较高,为(269.93±4.73)mg/100 g,比大豆发芽前提高了约10倍;盐胁迫下,发芽大豆谷氨酸脱羧酶(glutamate decarboxylase,GAD)活性和GABA含量随Na Cl浓度加大和胁迫时间延长而提高,同时大豆发芽期间GABA含量与其他指标之间相关性分析表明,盐胁迫下发芽大豆GABA含量与芽长、GAD活性、游离氨基酸和可溶性蛋白含量之间呈显著正相关,在低盐胁迫下,大豆发芽受到抑制,但促进了GAD活性的升高,游离氨基酸和可溶性蛋白质含量增加,富集产生了较多的GABA。     

NaCl胁迫联合Ca2+调控糙米发芽富集GABA的工艺优化

为优化NaCl胁迫联合Ca~(2+)调控下糙米发芽富集γ-氨基丁酸(γ-aminobutyric acid,GABA)的培养条件,通过单因素和Box-Behnken响应面试验考察NaCl浓度、Ca~(2+)浓度、发芽温度及发芽时间4个因素对GABA含量的影响,得出糙米发芽最佳工艺条件。结果表明,发芽糙米在NaCl胁迫联合Ca~(2+)处理下富集GABA的最佳培养条件为NaCl浓度7.50 mmol/L,Ca~(2+)浓度15.0 mmol/L,发芽温度29℃,发芽时间2.3 d,在此条件下发芽糙米中GABA含量为144.98mg/100g。研究结果为糙米健康食品的研究提供了一定的理论依据。     

两步酶法生产γ-氨基丁酸的研究

提出两步酶法(包括发芽和均相反应2个阶段)生产γ-氨基丁酸(GABA)的新方法。以大豆为原料,采用两步酶法生产GABA,讨论了Ca2+对大豆发芽和均相反应过程中蛋白酶和谷氨酸脱羧酶活力,以及谷氨酸游离和GABA生成的影响。实验结果表明,不同浓度的Ca2+对这两种酶分别表现出激活和抑制作用。在发芽过程中,蛋白酶和谷氨酸脱羧酶的活力,以及GABA和游离谷氨酸的含量随发芽时间呈先上升后下降的趋势。在浸泡液CaCl2浓度3.0mmol/L、发芽时间60h的条件下,发芽大豆中GABA和游离谷氨酸的含量分别为111.9mg/(100g干物大豆)、224.8mg/(100g干物大豆)。在均相反应过程中,GABA和游离谷氨酸的含量增加。在固体含量20g/100mL、反应时间2h的条件下,游离谷氨酸含量在料液CaCl2浓度为1.0mmol/L时达到最大值369.9mg/(100g干物大豆),GABA含量在CaCl2浓度为3.0mmol/L时达到最大值316.1mg/(100g干物大豆)。采用两步酶法可充分发挥大豆内源酶的活力,生产出游离谷氨酸和GABA含量都较高的食品材料。     

低氧联合NaCl胁迫下蚕豆发芽富集γ-氨基丁酸培养条件优化

以蚕豆(启豆2号)为原料,研究了低氧联合NaCl胁迫下培养条件对γ-氨基丁酸(GABA)富集的影响。结果显示:非胁迫培养时间、培养pH和胁迫培养时间显著影响发芽蚕豆GABA积累。蚕豆发芽富集GABA最佳培养条件是非胁迫培养1.5 d、培养液pH 3.5和低氧联合NaCl胁迫4 d,在此条件下其GABA含量可达1.06mg/g DW,为原料蚕豆的7.57倍。     

响应面法优化盐胁迫发芽苦荞富集γ-氨基丁酸的培养条件

为优化盐胁迫条件下发芽苦荞富集γ-氨基丁酸(γ-aminobutyric acid,GABA)的最优培养条件,在单因素试验的基础上,采用响应面法探讨NaCl浓度、发芽时间和发芽温度对发芽苦荞中GABA含量的影响。结果表明:发芽苦荞在盐胁迫条件下富集GABA的最佳培养条件为:NaCl浓度34 mmol/L、发芽时间5 d、发芽温度31℃,在此条件下发芽苦荞中GABA富集量为250.06μg/g(以干质量计)。方差分析及验证实验显示,模型具有极显著的可靠性和拟合度(R~2=0.9611),可准确预测盐胁迫条件下苦荞发芽过程中GABA的富集量。     

冷等离子体联合L-谷氨酸与盐胁迫对红小豆萌发富集γ-氨基丁酸的效果及工艺条件

为探究等离子体联合盐胁迫对红小豆萌发后γ-氨基丁酸(γ-Aminobutyric acid,GABA)含量的富集作用及效果。本实验以红小豆为原料,考察大气冷等离子电压、频率、时间处理种子对其发芽过程中GABA含量的影响,同时采用L-谷氨酸(L-Glu)联合盐胁迫的发芽方法,通过考察单因素(发芽时间、CaCl₂、L-Glu和NaCl浓度)对GABA富集量的影响及响应面优化试验确定该法富集GABA最佳工艺。结果表明,大气冷等离子体技术处理种子对其萌发富集γ-氨基丁酸有促进作用,电压90 kV、频率120 Hz、时间20 min条件下大气冷等离子体处理效果较好。在发芽时间为58 h、CaCl₂浓度为4.4 mmol/L、L-Glu浓度为3.2 mg/mL、NaCl浓度为66 mmol/L时,发芽红小豆GABA含量为160.23±2.91 mg/100 g,是未发芽红小豆的7.12倍。该方法高效可靠且成本低,为富含GABA食品的工厂化生产提供技术参考。     

低氧胁迫下大豆发芽富集γ-氨基丁酸品种筛选及培养条件优化

比较理想M-7、95-优1、苏青1号和YH-NJ大豆品种培养4 d后的生理生化变化情况,结果表明:YH-NJ的发芽率、芽长和呼吸强度显著高于其他品种,发芽4 d后YH-NJ的蛋白酶活力、游离氨基酸含量的增加量也显著高于其他品种,γ-氨基丁酸(γ-aminobutyric acid,GABA)产量及增加量均为最大。由此筛选出大豆YH-NJ是发芽富集GABA的良好品种。以YH-NJ为原料,在单因素试验结果的基础上,通过正交试验,得到发芽大豆富集GABA的最佳培养条件为:正常培养4 d、低氧胁迫培养48 h、培养液p H 5.0、发芽温度30℃。在此条件下,GABA产量达到1.97 mg/g,是随机组的1.56倍。     

GABA shunt and polyamine degradation pathway on γ-aminobutyric acid accumulation in germinating fava bean (Vicia faba L.) under hypoxia

GABA shunt and polyamine degradation pathway on γ-aminobutyric acid (GABA) accumulation in germinating fava bean under hypoxia was investigated. GABA content, GAD and DAO activity were significantly increased under hypoxia treatment. Glu and polyamine contents enhanced largely and thus supplied as sufficient substrates for GABA formation. In contrast, GABA content decreased, mainly in the embryo, after removing the hypoxia stress. DAO activity, Glu and polyamines contents decreased, while an increment of GAD activity was observed. This indicated that GAD activity can be not only regulated by hypoxia, but by the rapid growth of embryo after the recovery from hypoxia stress. When treated with AG, DAO activity was almost inhibited completely, and the GABA content decreased by 32.96% and 32.07% after treated for 3 and 5 days, respectively. Hence, it can be inferred that about 30% of GABA formed in germinating fava bean under hypoxia was supplied by polyamine degradation pathway.     

NaCl stress and supplemental CaCl2 regulating GABA metabolism pathways in germinating soybean

Effects of NaCl stress and supplemental CaCl₂ on soybean growth and gamma-aminobutyric acid (GABA) accumulation pathways in germinating soybean were investigated. Germinating under NaCl stress, GABA accumulation was caused not only by the activation of glutamate decarboxylase (GAD), diamine oxidase (DAO) and aminoaldehyde dehydrogenase (AMADH) existing in seed but also by their gene expression up-regulation during germination. However, after CaCl₂ (6 mM) application, the harmful effect of NaCl stress (50 mM) was removed. GAD activity significantly increased, while its gene expression levels had no significant changes with the control, indicating that the increment in GAD activity was caused by the stimulation of exogenous Ca²⁺. Meanwhile, GABA was still accumulated although DAO and AMADH activities and their gene expression levels had no significant difference compared with the control. It can be concluded that CaCl₂ addition decreased the contribution of polyamine degradation pathway to GABA accumulation compared with NaCl stress, and the increased GABA was mainly synthesized via GABA shunt.     

未熟粒糙米低氧胁迫发芽期间生理生化变化研究

以未熟粒糙米为试材,研究其低氧胁迫发芽期间主要生理生化变化。结果表明：经低氧胁迫发芽48 h,未熟粒糙米的发芽率达79.3%,芽长为1.92 mm,呼吸强度提高5.6 倍,容重达0.79 g/mL;发芽期间其含水量快速增加后趋于平缓,淀粉含量持续下降,还原糖和游离氨基酸含量则逐渐增加,均与完熟粒糙米变化趋势一致且无显著性差异;未熟粒糙米中γ-氨基丁酸（γ-aminobutyric acid,GABA）含量随发芽时间延长呈逐渐增加趋势,发芽48 h时GABA含量增加1.08 倍,且谷氨酸脱羧酶（glutamate decarboxylase,GAD）活力和谷氨酸（glutamic acid,Glu）含量与对照组无显著差异。低氧胁迫下未熟粒发芽糙米中GABA含量与芽长、呼吸强度、游离氨基酸和GAD活力均呈极显著正相关（P＜0.01）,与淀粉和谷氨酸含量呈极显著负相关（P＜0.01）。低氧胁迫发芽改善了未熟粒糙米营养品质,是富集GABA的良好原料。     

低氧与低温胁迫对发芽玉米籽粒中γ-氨基丁酸富集的影响

以玉米籽粒为实验材料,研究低氧胁迫下发芽时间、低温胁迫与回温解冻下的温度及时间对发芽玉米中γ-氨基丁酸(γ-aminobutyric acid,GABA)含量的影响,对其低氧和低温胁迫工艺进行了优化,同时对胁迫期间发芽玉米籽粒中GABA代谢酶活性的变化进行了研究。结果表明:玉米经低氧胁迫发芽72 h后,在-18℃冷冻6 h和25℃回温4 h条件下,发芽玉米中GABA含量增加29.9倍,达到1.52 mg/g(以干质量计);低氧胁迫下发芽玉米籽粒主要是通过GABA支路富集GABA的。玉米籽粒是富集GABA的良好原料,且低氧与低温胁迫是富集发芽玉米中GABA的有效方式。     

Accumulation of γ-aminobutyric acid in germinated soybean (<Emphasis Type="Italic">Glycine max</Emphasis> L.) in relation to glutamate decarboxylase and diamine oxidase activity induced by additives under hypoxia

High levels of γ-aminobutyric acid (GABA) accumulate in plant tissues under various stresses and exogenous additives. The purpose of this research is to provide an effective finding that can prove a rapid accumulation of GABA in germinated soybean (Glycine max L.) in response to different additives under hypoxia. Hypoxia-induced GABA accumulation in soybean embryo resulted in part from polyamine oxidation. Response to different concentration of glutamate (Glu), pyridoxal phosphate, arginine, CuCl₂, NaCl, and CaCl₂, a significant difference including GABA accumulation, changes of Glutamate decarboxylase (GAD), and Diamine oxidase activity (DAO) activity in germinated soybean under hypoxia occurred (p < 0.05) and the maximum accumulation of GABA were 4.07, 3.02, 3.50, 3.26, 4.00, and 3.30 g kg⁻¹ DW respectively, which were significantly higher than those germinated soybean under normal culture (CK) and hypoxia culture (CK₀) (p < 0.05). The GAD and DAO have different distributions in cotyledon and embryo of germinated soybean, and the enzyme activity mainly located in embryo of germinated soybean. Germinated soybean is a good resource of GABA-rich food. Different additives have significant effects on GABA production, among which Glu and NaCl are ideal material for GABA accumulation.     

低氧胁迫下发芽玉米淀粉特性及高γ-氨基丁酸玉米饮料开发

以糯玉米（京甜紫花糯）为试材,研究低氧胁迫下其籽粒发芽期间淀粉特性变化,筛选了高γ-氨基丁酸（γ-aminobutyric acid,GABA）含量的玉米保健饮料的稳定剂及产品配方。结果表明,玉米经低氧胁迫发芽72 h,GABA含量增加12.2 倍,达0.65 mg/g,淀粉含量显著下降,淀粉糊透光率降低27.8%,淀粉持水性、持油性和乳化性均增强,淀粉乳化稳定性降低7.4%;玉米匀浆最佳料液比1∶8,最优复合稳定剂为羧甲基纤维素钠0.03%、藻酸丙二醇酯0.09%、海藻酸钠0.04%和黄原胶0.06%,其饮料静置沉淀率最低,仅为3.7%;白砂糖添加量为4.8 g/100 mL和体系pH值为4.6 时,模糊综合评价表明该高GABA玉米饮料达到“容易接受”级别,符合消费者饮用要求。     

活性化糙米富集γ-氨基丁酸的雾化液组分优化

通过单因素试验和正交试验,优化了糙米湿润活性化富集γ-氨基丁酸(γ-aminobutyric acid,GABA)的雾化液p H值、VB6及Ca Cl2浓度,同时研究了最优条件下活性化糙米谷氨酸脱羧酶活力及主要物质含量变化。结果表明:最优培养条件为雾化液p H 3.5、VB6浓度2 mmol/L和Ca Cl2浓度10 mmol/L;在此条件下,活性化糙米中GABA含量可达7.67 mg/100 g,为原料糙米的2.74倍。随着活性化时间的延长,糙米中游离氨基酸、还原糖含量及谷氨酸脱羧酶活力呈现逐渐增加的趋势,可溶性蛋白及淀粉含量呈现逐渐下降趋势。相关性分析表明,GABA含量与谷氨酸、游离氨基酸、可溶性蛋白含量之间均显著相关。     

玉米籽粒低氧胁迫发芽期间主要生理生化和γ-氨基丁酸含量变化

以玉米籽粒为材料研究低氧胁迫下籽粒发芽过程中主要生理生化和γ-氨基丁酸(γ-aminobutyric acid,GABA)含量变化,筛选出最适富集GABA的玉米品种。结果表明:5个品种玉米籽粒在72 h低氧胁迫发芽期间,芽长增长15.5~26.3 mm,呼吸强度提高1.7~3.2倍,干物质损失35.6%~40.6%,淀粉消耗15.5%~28.9%,还原糖和游离氨基酸含量分别增加1.7~4.7倍和7.4~13.3倍,GABA含量提高9.3~13.2倍,不同品种玉米籽粒发芽能力和GABA富集量呈显著差异,以京甜紫花糯(ZHN)品种发芽率最高,达到85.5%,且GABA富集量达到0.65mg/g(以干质量计),ZHN是富集GABA的最适玉米品种。低氧胁迫下发芽玉米中GABA含量与芽长、呼吸强度、游离氨基酸呈极显著正相关(P0.01),玉米经低氧胁迫发芽能提升玉米营养品质。