葡萄化感物质的分离、鉴定及其作用机制研究
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摘要
本试验以晚红(V. vinifera L. cv. Red Globe)葡萄组培苗、山河二号(V. amurensis×V. riparia No.2)葡萄组培苗及贝达(V. riparia×V. labrusca cv. Beta)葡萄扦插苗为试材,采用室内葡萄组培苗和室外盆栽苗相结合的生物检测方法,分析了葡萄根系分泌物及腐解物的自毒作用,并对其中的化感物质进行了分离鉴定；采用盆栽试验,研究外源酚酸对葡萄幼苗生长及生理指标的影响,初步探讨了化感物质的作用机制。主要试验结果如下：
1.根系分泌物和腐解物显著抑制了晚红葡萄组培苗的生长。具体表现为植株高度、生根数、平均根长及地上、地下鲜重降低,随着处理浓度提高,抑制作用增强,0.5 g·mL-1根系分泌物处理后组培苗干枯死亡；0.1 g·mL-1根茬腐解物处理后组培苗生根受到抑制。
2.根系分泌物对盆栽贝达葡萄扦插苗生长表现为低浓度促进、高浓度抑制的作用趋势。低浓度根系分泌物处理后植株生长势增强,叶片净光合速率和根系呼吸速率提高，叶片可溶性糖、淀粉及蛋白质含量增加,MDA含量降低,而高浓度处理下则表现相反的变化趋势。
3.添加根茬残体显著抑制了盆栽贝达葡萄扦插苗的生长,添加量越高,抑制作用越强。随着根茬残体添加量的增加,叶片的净光合速率、蒸腾速率和气孔导度下降,可溶性糖、淀粉和蛋白质含量减少,根呼吸速率降低；SOD活性降低,MDA含量增加。
4.根系分泌物对土壤多酚氧化酶与转化酶活性表现为低浓度促进、高浓度抑制的作用趋势,且高浓度根系分泌物对酶活性的抑制作用显著。添加根茬残体会抑制土壤多酚氧化酶和转化酶活性,其中对土壤转化酶的抑制程度大于对土壤多酚氧化酶活性的影响,说明土壤转化酶对添加根茬残体腐解处理的自毒作用敏感性高。
5.根系分泌物处理使葡萄根际土壤中微生物数量显著减少,随着根系分泌物浓度的提高,细菌数量逐渐减少,而真菌占微生物总数的比例逐渐升高。添加根茬残体处理使土壤中微生物数量显著增加,但降低了根际土壤的真菌种群多样性,其中有益的木霉菌消失,出现了不利于植株生长的腐霉属、根串珠霉属和束梗孢霉属。
6. LC-MS检测结果表明葡萄根系分泌物中含有2种酚酸类物质(水杨酸、对羟基苯甲酸)；根茬腐解物中含有3种酚酸类物质(苯甲酸、水杨酸和苯丙酸)。组织培养试验表明,四种酚酸类物质具有显著的化感作用。
7.化感物质苯甲酸、苯丙酸和水杨酸通过干扰植物细胞膜及相关生理过程影响了葡萄的生长。盆栽试验表明,随着酚酸浓度提高、处理时间延长,对盆栽葡萄苗生长的抑制程度增强。除苯甲酸在0.1 mmol·L-1浓度处理时地上、地下鲜重比对照略有所提高外,随着苯甲酸、水杨酸、苯丙酸处理浓度的提高,地上、地下鲜重呈逐渐降低的趋势。同时外源酚酸处理增加了葡萄植株细胞膜透性、降低了SOD活性及植株根系活力,影响了光合作用和叶片物质积累等生理生化过程,致使植株生长发育受到抑制。
Using tissue culture seedling of Red Globe (V.vinifera L. cv.), tissue culture seedling of Shanhe No.2 (V. amurensis×V. riparia) grape and pot seedling of Beta (V. riparia×V. labrusca cv.) as experimental material, tissue cultured seedling of Red Globe in laboratory and cutting seedlings of Beta by pot experiment were used to examine the allelopathy of grape exudates and root decomposing products, and major allelochemicals were determined; Using potted experiment, the effects of phenolic acids on grape seedling growth and physiological index were studied, and the mechanism of allelochemicals was discussed. The major results were as follow:
1. Grape root exudates and root decomposing products were significantly inhibited tissue culture seedling growth of Red Globe grape. Plant height, root number, average root length, fresh weight of shoot and fresh weight of root decreased. With concentration increasing of root exudates and root decomposing products, the inhibition increased. Grape tissue culture seedling died when 0.5 g·mL-1 root exudates treated; and roots growth of tissue culture seedling was inhibited when 0.1 g·mL-1 root decomposing products treated.
2. Grape root exudates of low concentration promoted pot cottage seedling growth of Beta grape and high concentration inhibited its growth. Root exudates of low concentration made the growth enhanced; net photosynthetic rate and root respiration rate increased; soluble sugar content, starch content and protein content increased; MDA content decreased, while root exudates of high concentration showed opposite trend.
3. Decomposition substance from grape root stubble were significantly inhibited the growth of Beta pot cottage seedling. With concentration increasing of root stubble, the stronger inhibition showed. Plant height, stem diameter, fresh weight of shoot and fresh weight of root decreased. Net photosynthetic rate, transpiration rate, stomatal conductance of leaves decreased. Soluble sugar content, starch content, protein content and root respiration rate decreased. SOD activity of leaves decreased, while MDA content increased.
4. Grape root exudates of low concentration promoted soil PPO activity and invertase activity of Beta grape and high concentration inhibited its activity. Root exudates of high concentration inhibited enzyme activity significantly. Decomposition substance from grape root stubble were significantly inhibited soil PPO activity and invertase activity; the inhibition of soil invertase activity, was greater than soil PPO activity. Description of soil invertase activity had high sensitivity to the allelopathy of decomposition substance from grape root stubble.
5. With concentration increasing of root exudates, the microbial number of rhizosphere soil was significant decreased, the bacteria number was reduced, and increased the fungus proportion of the total. With concentration increasing of root stubble, the microbial number of rhizosphere soil was significant increased, decreased fungal diversity, made Trichoderma disappeared, the harmful plant growth of Pythium, Thielaviopsis and Stilbellales appeared.
6. Two phenolic acids (p-hydroxybenzoic acid, salicylic acid) were isolation and identification in grape root exudates by LC-MS, and three phenolic acids (coumaric acid, phenylpropionic acid and benzoic acid) were isolation and identification in grape root decomposing products, and they were important allelochemicals by verification of tissue culture experiment.
7. It was clear that phenylpropionic acid and salicylic acid inhibited plant growth through disturbing membrane and related physiological process. Based on pot experiment, the results indicated that with phenolic acids concentration increasing and treatment time prolonging, the degree of inhibition increased. With the concentrations rise of benzoic acid, salicylic acid and phenylpropionic acid, fresh weight of shoot and fresh weight of root decreased except benzoic acid under 0.1 mmol·L-1 concentration treatment which fresh weight of shoot and fresh weight of root were higher than control. In addition, phenolic acids enhanced cell membrane permeability, decreased SOD activity and root activity, affected photosynthesis and dry matter accumulation of grape leaves, and resulted in growth vigor weak.
1.根系分泌物和腐解物显著抑制了晚红葡萄组培苗的生长。具体表现为植株高度、生根数、平均根长及地上、地下鲜重降低,随着处理浓度提高,抑制作用增强,0.5 g·mL-1根系分泌物处理后组培苗干枯死亡；0.1 g·mL-1根茬腐解物处理后组培苗生根受到抑制。
2.根系分泌物对盆栽贝达葡萄扦插苗生长表现为低浓度促进、高浓度抑制的作用趋势。低浓度根系分泌物处理后植株生长势增强,叶片净光合速率和根系呼吸速率提高，叶片可溶性糖、淀粉及蛋白质含量增加,MDA含量降低,而高浓度处理下则表现相反的变化趋势。
3.添加根茬残体显著抑制了盆栽贝达葡萄扦插苗的生长,添加量越高,抑制作用越强。随着根茬残体添加量的增加,叶片的净光合速率、蒸腾速率和气孔导度下降,可溶性糖、淀粉和蛋白质含量减少,根呼吸速率降低；SOD活性降低,MDA含量增加。
4.根系分泌物对土壤多酚氧化酶与转化酶活性表现为低浓度促进、高浓度抑制的作用趋势,且高浓度根系分泌物对酶活性的抑制作用显著。添加根茬残体会抑制土壤多酚氧化酶和转化酶活性,其中对土壤转化酶的抑制程度大于对土壤多酚氧化酶活性的影响,说明土壤转化酶对添加根茬残体腐解处理的自毒作用敏感性高。
5.根系分泌物处理使葡萄根际土壤中微生物数量显著减少,随着根系分泌物浓度的提高,细菌数量逐渐减少,而真菌占微生物总数的比例逐渐升高。添加根茬残体处理使土壤中微生物数量显著增加,但降低了根际土壤的真菌种群多样性,其中有益的木霉菌消失,出现了不利于植株生长的腐霉属、根串珠霉属和束梗孢霉属。
6. LC-MS检测结果表明葡萄根系分泌物中含有2种酚酸类物质(水杨酸、对羟基苯甲酸)；根茬腐解物中含有3种酚酸类物质(苯甲酸、水杨酸和苯丙酸)。组织培养试验表明,四种酚酸类物质具有显著的化感作用。
7.化感物质苯甲酸、苯丙酸和水杨酸通过干扰植物细胞膜及相关生理过程影响了葡萄的生长。盆栽试验表明,随着酚酸浓度提高、处理时间延长,对盆栽葡萄苗生长的抑制程度增强。除苯甲酸在0.1 mmol·L-1浓度处理时地上、地下鲜重比对照略有所提高外,随着苯甲酸、水杨酸、苯丙酸处理浓度的提高,地上、地下鲜重呈逐渐降低的趋势。同时外源酚酸处理增加了葡萄植株细胞膜透性、降低了SOD活性及植株根系活力,影响了光合作用和叶片物质积累等生理生化过程,致使植株生长发育受到抑制。
Using tissue culture seedling of Red Globe (V.vinifera L. cv.), tissue culture seedling of Shanhe No.2 (V. amurensis×V. riparia) grape and pot seedling of Beta (V. riparia×V. labrusca cv.) as experimental material, tissue cultured seedling of Red Globe in laboratory and cutting seedlings of Beta by pot experiment were used to examine the allelopathy of grape exudates and root decomposing products, and major allelochemicals were determined; Using potted experiment, the effects of phenolic acids on grape seedling growth and physiological index were studied, and the mechanism of allelochemicals was discussed. The major results were as follow:
1. Grape root exudates and root decomposing products were significantly inhibited tissue culture seedling growth of Red Globe grape. Plant height, root number, average root length, fresh weight of shoot and fresh weight of root decreased. With concentration increasing of root exudates and root decomposing products, the inhibition increased. Grape tissue culture seedling died when 0.5 g·mL-1 root exudates treated; and roots growth of tissue culture seedling was inhibited when 0.1 g·mL-1 root decomposing products treated.
2. Grape root exudates of low concentration promoted pot cottage seedling growth of Beta grape and high concentration inhibited its growth. Root exudates of low concentration made the growth enhanced; net photosynthetic rate and root respiration rate increased; soluble sugar content, starch content and protein content increased; MDA content decreased, while root exudates of high concentration showed opposite trend.
3. Decomposition substance from grape root stubble were significantly inhibited the growth of Beta pot cottage seedling. With concentration increasing of root stubble, the stronger inhibition showed. Plant height, stem diameter, fresh weight of shoot and fresh weight of root decreased. Net photosynthetic rate, transpiration rate, stomatal conductance of leaves decreased. Soluble sugar content, starch content, protein content and root respiration rate decreased. SOD activity of leaves decreased, while MDA content increased.
4. Grape root exudates of low concentration promoted soil PPO activity and invertase activity of Beta grape and high concentration inhibited its activity. Root exudates of high concentration inhibited enzyme activity significantly. Decomposition substance from grape root stubble were significantly inhibited soil PPO activity and invertase activity; the inhibition of soil invertase activity, was greater than soil PPO activity. Description of soil invertase activity had high sensitivity to the allelopathy of decomposition substance from grape root stubble.
5. With concentration increasing of root exudates, the microbial number of rhizosphere soil was significant decreased, the bacteria number was reduced, and increased the fungus proportion of the total. With concentration increasing of root stubble, the microbial number of rhizosphere soil was significant increased, decreased fungal diversity, made Trichoderma disappeared, the harmful plant growth of Pythium, Thielaviopsis and Stilbellales appeared.
6. Two phenolic acids (p-hydroxybenzoic acid, salicylic acid) were isolation and identification in grape root exudates by LC-MS, and three phenolic acids (coumaric acid, phenylpropionic acid and benzoic acid) were isolation and identification in grape root decomposing products, and they were important allelochemicals by verification of tissue culture experiment.
7. It was clear that phenylpropionic acid and salicylic acid inhibited plant growth through disturbing membrane and related physiological process. Based on pot experiment, the results indicated that with phenolic acids concentration increasing and treatment time prolonging, the degree of inhibition increased. With the concentrations rise of benzoic acid, salicylic acid and phenylpropionic acid, fresh weight of shoot and fresh weight of root decreased except benzoic acid under 0.1 mmol·L-1 concentration treatment which fresh weight of shoot and fresh weight of root were higher than control. In addition, phenolic acids enhanced cell membrane permeability, decreased SOD activity and root activity, affected photosynthesis and dry matter accumulation of grape leaves, and resulted in growth vigor weak.
引文
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