H_2O_2对白木香细胞生长和挥发性次生产物的影响
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摘要
【目的】本文研究了外源H_2O_2对白木香[Aquilaria sinensis(Lour.)Gilg]细胞生长和细胞挥发性次生产物组分及相对含量的影响,为白木香结香机制提供依据。【方法】采用不同浓度H_2O_2与白木香细胞共培养的方法,检测H_2O_2对白木香细胞的鲜重、活力、过氧化酶(POD)活性、细胞沉降体积(SCV)、丙二醛(MDA)含量、DNA琼脂糖凝胶电泳和挥发性次生产物的影响。【结果】H_2O_2对白木香细胞的生长、细胞沉淀体积、活力均较对照呈降低的影响;丙二醛含量随H_2O_2浓度的增加而升高;低浓度处理的白木香细胞POD活性在生长前期显著升高,在生长后期急剧降低,高浓度处理细胞的POD活性在培养期内均较低、中浓度处理的低,生长后期POD活性基本丧失;白木香细胞的DNA均出现降解,低浓度组降解条带较亮,高浓度组降解条带较暗;低浓度处理组(1 mmol/L)的芳香族化合物和烷烃类化合物含量均高于其他组,10 mmol/LH_2O_2处理组检测出了新产生的长叶烯(3.27%)和角鲨烯(3.05%)2种萜类,高浓度处理组(30和50 mmol/L)有机酸和芳香族类化合物的组分和含量显著下降。【结论】在不同浓度的H_2O_2影响下,白木香细胞生长、活力和挥发性次生产物的组分与相对含量都会发生变化。
【Objective】This paper aimed to study the effect of exogenous H_2O_2 on the growth, vitality, MDA content, POD activity and the components and relative content of volatile secondary products in Aquilaria sinensis(Lour.) Gilg cells, which provided a basis for the mechanism of the Aquilaria sinensis as materials.【Method】The effects of H_2O_2 on fresh weight, vitality, peroxidase(POD) activity, cell sedimentation volume(SCV), malondialdehyde(MDA) content, DNA agarose gel electrophoresis and volatile secondary production of Aquilaria sinensis cells were detected by co-culture of its different concentrations. 【Result】Compared with the control group, H_2O_2 had a decreased effect on the growth of Aquilaria sinensis cells, its SCV and viability of suspension cells. MDA content increased with the increase of H_2O_2 concentration. The activity of POD in Aquilaria sinensis cells treated by low concentration of H_2O_2 showed a significant increase change in the early growth stage and decreased rapidly at the late growth stage, while the POD activities of the cells treated by high concentration H_2O_2 were more than that treated by low concentration and middle concentration H_2O_2 during the culture period. The DNA of Aquilaria sinensis cells was degraded, and the degradation bands were lighter in the low concentration group and darker in the high concentration group. Aromatic compounds and alkanes in low concentration treatment group(1 mmol/L) were higher than other groups, two new compounds terpenes(3.27 %) and squalene(3.05 %) were found under the condition of 10 mmol/L H_2O_(2 )treated cells, and the species constituents and content of organic acids and aromatic compounds in the high concentration treatment group(30 and 50 mmol/L) decreased significantly. 【Conclusion】With the treatment of different concentrations of H_2O_2, the cell growth and volatile secondary production components and relative content of Aquilaria sinensis would change.
【Objective】This paper aimed to study the effect of exogenous H_2O_2 on the growth, vitality, MDA content, POD activity and the components and relative content of volatile secondary products in Aquilaria sinensis(Lour.) Gilg cells, which provided a basis for the mechanism of the Aquilaria sinensis as materials.【Method】The effects of H_2O_2 on fresh weight, vitality, peroxidase(POD) activity, cell sedimentation volume(SCV), malondialdehyde(MDA) content, DNA agarose gel electrophoresis and volatile secondary production of Aquilaria sinensis cells were detected by co-culture of its different concentrations. 【Result】Compared with the control group, H_2O_2 had a decreased effect on the growth of Aquilaria sinensis cells, its SCV and viability of suspension cells. MDA content increased with the increase of H_2O_2 concentration. The activity of POD in Aquilaria sinensis cells treated by low concentration of H_2O_2 showed a significant increase change in the early growth stage and decreased rapidly at the late growth stage, while the POD activities of the cells treated by high concentration H_2O_2 were more than that treated by low concentration and middle concentration H_2O_2 during the culture period. The DNA of Aquilaria sinensis cells was degraded, and the degradation bands were lighter in the low concentration group and darker in the high concentration group. Aromatic compounds and alkanes in low concentration treatment group(1 mmol/L) were higher than other groups, two new compounds terpenes(3.27 %) and squalene(3.05 %) were found under the condition of 10 mmol/L H_2O_(2 )treated cells, and the species constituents and content of organic acids and aromatic compounds in the high concentration treatment group(30 and 50 mmol/L) decreased significantly. 【Conclusion】With the treatment of different concentrations of H_2O_2, the cell growth and volatile secondary production components and relative content of Aquilaria sinensis would change.
引文
[1]黄玮婷,孔凡芹,王海燕,等. 沉香属植物繁殖研究进展[J]. 世界林业研究,2017,30(1):44-48.
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[4]何梦玲,何芳,孟京兰,等. 3种诱导子对白木香根悬浮培养细胞中2-(2-苯乙基)色酮化合物形成的影响[J]. 中成药,2013,35(7):1367-1371.
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[7]董闪,李明,唐堃,等. 白木香愈伤组织的诱导及培养[J]. 湖北农业科学,2014,53(15):3673-3677.
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[9]王跃华,刘益丽,马良良,等. TTC—脱氢酶还原法测定滇重楼细胞活力优化条件筛选[J]. 成都大学学报(自然科学版),2011(1):1-3.
[10]史昆,杨模华,李志辉,等. 马尾松胚性细胞悬浮增殖培养体系的建立[J]. 中南林业科技大学学报, 2014(1):64-68.
[11]李玲,李娘辉,江素梅,等. 植物生理学模块实验指导[M]. 北京:科学出版社,2009.
[12]张娟芳,戚树源,何梦玲,等. 几种生理因子对印楝细胞悬浮培养生长的影响[J]. 广西植物, 2003(6):549-552.
[13]Miku A A, Niedzielski M, Rybczyski J J. The use of TTC reduction assay for assessment of Gentiana spp. cell suspension viability after cryopreservation[J]. Acta Physiologiae Plantarum, 2006, 28(4):315-324.
[14]刘进元,丛靖莉,潘明祥. 植物科学进展[M]. 北京:高等教育出版社,1998.
[15]Jones M. Programmed cell death in development and defense[J]. Plant Physiol, 2002,125: 94-97.
[16]梅文莉,曾艳波,刘俊,等. 五批国产沉香挥发性成分的GC-MS分析[J]. 中药材,2007(5): 551-555.
[17]Houot V, Etienne P, Petitot A S,et al. Hydrogen peroxide induces programmed cell death features in cultured tobacco BY-2 cells, in a dose-dependent manner[J]. J. Exp. Bot., 2001,52(361): 1721-1730.
[18]张园园,吕秀军,张辉,等. 外源H2O2处理对唐古特白刺愈伤组织脯氨酸代谢的影响[J]. 植物研究,2011,31(2):213-217+226.
[19]Solomon M, Belenghi B, Delledonne M, et al. The involvement of cysteine proteases and protease inhibitor gene in the regulation of programmed cell death in plants[J]. Plant Cell, 1999, 11: 431-443.
[20]董金皋,韩建民. 植物与病原物互作中的活性氧代谢及其作用[J]. 沈阳农业大学学报,2000,31(5):427-431.
[21]蔡凤香,陈豆豆,杨飞,等. 云H2O2对水稻幼苗生长和生理的调节[J]. 江苏农业科学,2016,44(3):74-77.
[22]刘建新,王金成,王瑞娟,等. 外源过氧化氢提高燕麦耐盐性的生理机制[J]. 草业学报,2016,25(2):216-222
[23]陈贵华,石岭,王萍,等. 外源过氧化氢对苦菜耐盐生理作用的影响[J]. 内蒙古农业大学学报(自然科学版),2017,38(6):8-11.
[2]李明月,沈华杰,何海珊,等. 白木香解剖构造及抽提物化学成分分析[J]. 西南林业大学学报(自然科学),2017,37(5):208-213.
[3]张争,杨云,魏建和,等. 白木香茎中内源茉莉酸类和倍半萜类物质对机械伤害的响应[J]. 园艺学报,2013,40(1):163-168.
[4]何梦玲,何芳,孟京兰,等. 3种诱导子对白木香根悬浮培养细胞中2-(2-苯乙基)色酮化合物形成的影响[J]. 中成药,2013,35(7):1367-1371.
[5]NEILL J, DESIKAN R, CLARKE A, et al. Hydrogen peroxide and nitric oxide as signaling molecules in plants[J]. EXP BIOL MED, 2002, 53: 1237-1247.
[6]Hancock J T, Desikan R, Clarke A, et al. Cell signalling following plant/pathogen interactions involves the generation of reactive oxygen and reactive nitrogen species[M].Elsevier Masson SAS, 2002: 611-617.
[7]董闪,李明,唐堃,等. 白木香愈伤组织的诱导及培养[J]. 湖北农业科学,2014,53(15):3673-3677.
[8]Miku A A, Niedzielski M, Rybczyski J J. The use of TTC reduction assay for assessment of Gentiana spp.cell suspension viability after cryopreservation[J]. Acta Physiologiae Plantarum, 2006, 28(4): 315-324.
[9]王跃华,刘益丽,马良良,等. TTC—脱氢酶还原法测定滇重楼细胞活力优化条件筛选[J]. 成都大学学报(自然科学版),2011(1):1-3.
[10]史昆,杨模华,李志辉,等. 马尾松胚性细胞悬浮增殖培养体系的建立[J]. 中南林业科技大学学报, 2014(1):64-68.
[11]李玲,李娘辉,江素梅,等. 植物生理学模块实验指导[M]. 北京:科学出版社,2009.
[12]张娟芳,戚树源,何梦玲,等. 几种生理因子对印楝细胞悬浮培养生长的影响[J]. 广西植物, 2003(6):549-552.
[13]Miku A A, Niedzielski M, Rybczyski J J. The use of TTC reduction assay for assessment of Gentiana spp. cell suspension viability after cryopreservation[J]. Acta Physiologiae Plantarum, 2006, 28(4):315-324.
[14]刘进元,丛靖莉,潘明祥. 植物科学进展[M]. 北京:高等教育出版社,1998.
[15]Jones M. Programmed cell death in development and defense[J]. Plant Physiol, 2002,125: 94-97.
[16]梅文莉,曾艳波,刘俊,等. 五批国产沉香挥发性成分的GC-MS分析[J]. 中药材,2007(5): 551-555.
[17]Houot V, Etienne P, Petitot A S,et al. Hydrogen peroxide induces programmed cell death features in cultured tobacco BY-2 cells, in a dose-dependent manner[J]. J. Exp. Bot., 2001,52(361): 1721-1730.
[18]张园园,吕秀军,张辉,等. 外源H2O2处理对唐古特白刺愈伤组织脯氨酸代谢的影响[J]. 植物研究,2011,31(2):213-217+226.
[19]Solomon M, Belenghi B, Delledonne M, et al. The involvement of cysteine proteases and protease inhibitor gene in the regulation of programmed cell death in plants[J]. Plant Cell, 1999, 11: 431-443.
[20]董金皋,韩建民. 植物与病原物互作中的活性氧代谢及其作用[J]. 沈阳农业大学学报,2000,31(5):427-431.
[21]蔡凤香,陈豆豆,杨飞,等. 云H2O2对水稻幼苗生长和生理的调节[J]. 江苏农业科学,2016,44(3):74-77.
[22]刘建新,王金成,王瑞娟,等. 外源过氧化氢提高燕麦耐盐性的生理机制[J]. 草业学报,2016,25(2):216-222
[23]陈贵华,石岭,王萍,等. 外源过氧化氢对苦菜耐盐生理作用的影响[J]. 内蒙古农业大学学报(自然科学版),2017,38(6):8-11.