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摘要
[目的]通过对水稻苗前期盐的处理后，测定盐胁迫条件下生理指标的变化，确定盐害对水稻的影响。[方法] 测定了一种盐敏感水稻品种（丽江新团黑谷）经过低浓度盐胁迫预处理后在盐害条件下叶片和根中活性氧H2O2的产生和CAT活力。[结果] 50 mmol/L盐胁迫能导致其叶片和根中H2O2含量显著上升，而CAT活力则显著下降。经过低浓度盐（20 mmol/L）预先处理的水稻在盐胁迫下H2O2含量上升幅度则较小，而CAT活力较高。此外，盐胁迫预处理的水稻在盐害条件下生长情况更好，其叶片中叶绿素含量更高。[结论] 活性氧H2O2及其清除酶CAT参与水稻对盐害的应答反应，并且这种预处理能增强水稻抗盐害胁迫能力。
关键词水稻；盐害；过氧化氢；过氧化氢酶；预处理
中图分类号S511文献标识码A文章编号0517-6611（2014）36-12814-02
Effects of NaCl Pretreatment on H2O2 Content and Catalase Activity of Rice under Salt Stress
GAO Yu1，2，XU Chunying1，WANG Dan1， JIN Xuehui1* et al
（1.Agricultural College， Heilongjiang Bayi Agricultural University， Daqing ，Heilongjiang 163319； 2.Heilongjiang Daqing Academy of Science， Daqing， Heilongjiang 163712）
Abstract[Objective]In order to study the effects of salt injury to rice， based on early rice seedlings after pretreatment of salt， the changes of physiological indexes under salt stress were determine. [Method]The salt sensitive rice variety of LTH was pretreated by low concentration of salt stress. And H2O2 content and catalase activity in the leaves and the roots were studied. [Result] 50 mmol/L salt stress could significantly increase H2O2 content in the leaves and the roots， but significantly decrease CAT activity. The rise of H2O2 content of rice pretreated by low concentration of salt stress （20 mmol/L） was low， but that of CAT activity was high. Otherwise， the growth of the rice pretreated by salt stress under the condition of salt stress was good， and chlorophyll content in the leaves was higher. [Conclusion] H2O2 and CAT were engaged in the response reaction of rice to salt stress. And the pretreatment could improve the salt stress resistance of rice.
Key wordsRice； Salt stress； Hydrogen peroxide； Catalase； Pretreatment
在植物生长过程中，其细胞在生理活动中不可避免地产生活性氧分子，如单线态氧、超氧阴离子、过氧化氢以及羟自由基等。这些活性氧会对植物的生物大分子造成伤害，但植物在亿万年进化过程中会产生活性氧自由基防卫系统如抗氧化酶（如清除H2O2的过氧化氢酶）和大分子抗氧化物质（如抗坏血酸等），从而将活性氧自由基的危害降低到最低程度。各种环境胁迫因子如干旱、盐害、强光等都能导致植物细胞中活性氧的过度产生[1]。我国有大量盐碱地，对农作物生长有明显的危害作用，如抑制植物生长，作物产量下降。水稻是一种盐敏感植物，也是我国主要的农作物之一。前人发现，离体水稻叶片在盐害作用下H2O2没有显著上升。此外，对于水稻植物在盐害条件下H2O2是否会上升有不同的结论，如盐敏感水稻H2O2会显著上升，而盐抗性水稻H2O2上升并不明显。然而，在自然界中很少有抗盐水稻品种。这里，笔者对一种盐敏感水稻品种（丽江新团黑谷）用低浓度NaCl预处理后，试图导致其抗盐能力增强，同时研究了后天盐害预处理的水稻在盐害条件下，其活性氧如H2O2产量及其主要清除酶过氧化氢酶（CAT）活性。该研究工作会促进人们对水稻在盐害条件下活性氧代谢产生新的认识。
1材料与方法
1.1植物材料培养及处理
将水稻品种丽江新团黑谷先用浓度0.5%次氯酸钠消毒液处理10 min，然后置于清水中24 h让其发芽。接着，转入营养土中使其生长，光照条件为12 h/12 h，75%湿度下生长。待长到2片叶后，将一部分水稻幼苗用20 mmol/L NaCl预处理24 h，对照为清水处理。然后，将水稻苗重新转入营养土中培养生长48 h。挑选健康和大小一致的2种处理后水稻苗（盐预处理和对照处理的）进行盐害（50 mmol/L NaCl）处理24 h，对照为清水处理。最后，将处理后的水稻苗挑选出来，用自来水轻轻冲洗干净，用于H2O2含量和CAT活力测定。 [3] GAY C，COLLINS J，GEBICKI J.Hydroperoxide assay with the ferricxylenol orange complex[J].Anal Biochem，1999，273：149-155.
[4] THORDALCHRISTENSEN H，ZHANG Z，WEI Y，et al.Subcellular localization of H2O2 in plants.H2O2 accumulation in papillae and hypersensitive response during the barleypowdery mildew interaction[J].Plant J，1997，11：1187-1194.
[5] ZHANG J，KIRKHAM M.Droughtstressinduced changes in activities of superoxide dismutase，catalase，and peroxidase in wheat species[J].Plant Cell Physiol， 1994，35：785-791.
[6] ZAKA R，VANDECASTEELE C，MISSET M.Effects of low chronic doses of ionizing radiation on antioxidant enzymes and G6PDH activities in Stipa capillata（Poaceae）[J].J Exp Bot， 2002，53：1979-1987.
[7] LIU C L， DU W C，CAI H S，et al.Trivalent chromium pretreatment alleviates the toxicity of oxidative damage in wheat plants exposed to hexavalent chromium[J].Acta Physiol Plant， 2014，36：787-794.
[8] DENG B L，DONG H S.Ectopic expression of riboflavinbinding protein gene TsRfBP paradoxically enhances both plant growth and drought tolerance in transgenic Arabidopsis[J].J Plant Growth Regul， 2013，32：170-181.
[9] DENG B L.Antioxidative response of golden agave leaves with different degrees of variegation under high light exposure[J].Acta physiol Plant， 2012，34：1925-1933.
[10] MITTLER R.Oxidative stress，antioxidants and stress tolerance[J].Trends Plant Sci， 2002，7：405-410.
[11] GILL S，TUTEJA N.Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants[J].Plant Physiol Biochem， 2010，48：909-930.
[12] AMIRJANI M.Effect of NaCl on some physiological parameters of rice[J].Eur J Biol Sci， 2010，3：6-16.
[13] LEE D，KIM Y，LEE C.The inductive responses of the antioxidant enzymes by salt stress in the rice（Oryza sativa L.）[J].J Plant Physiol，2001，158：737-745.
[14] LIN C，KAO C.Effect of NaCl stress on H2O2 metabolism in rice leaves[J].Plant Growth Regul， 2000，30：151-155.
[15] VAIDYANATHAN H，SIVAKUMAR P，CHAKRABARTY R，et al.Scavenging of reactive oxygen species in NaClstressed rice（Oryza sativa L.）differential responses in salttolerant and sensitive varieties[J].Plant Sci， 2003，165：1411-1418.
[16] WILLEKENS H，CHAMNONGPOL S，DAVEY M，et al. Catalase is a sink for H2O2 and is indispensable for stress defence in C3 plants[J].EMBO J， 1997，16：4806-4816.
[17] SHIM I，MOMOSE Y，YAMAMOTO A，et al. Inhibition of catalase activity by oxidative stress and its relationship to salicylic acid accumulation in plants[J].Plant Growth Regul， 2003，39：285-292.
[18] QUAN L，ZHANG B，SHI W，et al. Hydrogen peroxide in plants：a versatile molecule of reactive oxygen species network[J].J Integr Plant Biol， 2008，50：218.
[目的]通过对水稻苗前期盐的处理后，测定盐胁迫条件下生理指标的变化，确定盐害对水稻的影响。[方法] 测定了一种盐敏感水稻品种（丽江新团黑谷）经过低浓度盐胁迫预处理后在盐害条件下叶片和根中活性氧H2O2的产生和CAT活力。[结果] 50 mmol/L盐胁迫能导致其叶片和根中H2O2含量显著上升，而CAT活力则显著下降。经过低浓度盐（20 mmol/L）预先处理的水稻在盐胁迫下H2O2含量上升幅度则较小，而CAT活力较高。此外，盐胁迫预处理的水稻在盐害条件下生长情况更好，其叶片中叶绿素含量更高。[结论] 活性氧H2O2及其清除酶CAT参与水稻对盐害的应答反应，并且这种预处理能增强水稻抗盐害胁迫能力。
关键词水稻；盐害；过氧化氢；过氧化氢酶；预处理
中图分类号S511文献标识码A文章编号0517-6611（2014）36-12814-02
Effects of NaCl Pretreatment on H2O2 Content and Catalase Activity of Rice under Salt Stress
GAO Yu1，2，XU Chunying1，WANG Dan1， JIN Xuehui1* et al
（1.Agricultural College， Heilongjiang Bayi Agricultural University， Daqing ，Heilongjiang 163319； 2.Heilongjiang Daqing Academy of Science， Daqing， Heilongjiang 163712）
Abstract[Objective]In order to study the effects of salt injury to rice， based on early rice seedlings after pretreatment of salt， the changes of physiological indexes under salt stress were determine. [Method]The salt sensitive rice variety of LTH was pretreated by low concentration of salt stress. And H2O2 content and catalase activity in the leaves and the roots were studied. [Result] 50 mmol/L salt stress could significantly increase H2O2 content in the leaves and the roots， but significantly decrease CAT activity. The rise of H2O2 content of rice pretreated by low concentration of salt stress （20 mmol/L） was low， but that of CAT activity was high. Otherwise， the growth of the rice pretreated by salt stress under the condition of salt stress was good， and chlorophyll content in the leaves was higher. [Conclusion] H2O2 and CAT were engaged in the response reaction of rice to salt stress. And the pretreatment could improve the salt stress resistance of rice.
Key wordsRice； Salt stress； Hydrogen peroxide； Catalase； Pretreatment
在植物生长过程中，其细胞在生理活动中不可避免地产生活性氧分子，如单线态氧、超氧阴离子、过氧化氢以及羟自由基等。这些活性氧会对植物的生物大分子造成伤害，但植物在亿万年进化过程中会产生活性氧自由基防卫系统如抗氧化酶（如清除H2O2的过氧化氢酶）和大分子抗氧化物质（如抗坏血酸等），从而将活性氧自由基的危害降低到最低程度。各种环境胁迫因子如干旱、盐害、强光等都能导致植物细胞中活性氧的过度产生[1]。我国有大量盐碱地，对农作物生长有明显的危害作用，如抑制植物生长，作物产量下降。水稻是一种盐敏感植物，也是我国主要的农作物之一。前人发现，离体水稻叶片在盐害作用下H2O2没有显著上升。此外，对于水稻植物在盐害条件下H2O2是否会上升有不同的结论，如盐敏感水稻H2O2会显著上升，而盐抗性水稻H2O2上升并不明显。然而，在自然界中很少有抗盐水稻品种。这里，笔者对一种盐敏感水稻品种（丽江新团黑谷）用低浓度NaCl预处理后，试图导致其抗盐能力增强，同时研究了后天盐害预处理的水稻在盐害条件下，其活性氧如H2O2产量及其主要清除酶过氧化氢酶（CAT）活性。该研究工作会促进人们对水稻在盐害条件下活性氧代谢产生新的认识。
1材料与方法
1.1植物材料培养及处理
将水稻品种丽江新团黑谷先用浓度0.5%次氯酸钠消毒液处理10 min，然后置于清水中24 h让其发芽。接着，转入营养土中使其生长，光照条件为12 h/12 h，75%湿度下生长。待长到2片叶后，将一部分水稻幼苗用20 mmol/L NaCl预处理24 h，对照为清水处理。然后，将水稻苗重新转入营养土中培养生长48 h。挑选健康和大小一致的2种处理后水稻苗（盐预处理和对照处理的）进行盐害（50 mmol/L NaCl）处理24 h，对照为清水处理。最后，将处理后的水稻苗挑选出来，用自来水轻轻冲洗干净，用于H2O2含量和CAT活力测定。 [3] GAY C，COLLINS J，GEBICKI J.Hydroperoxide assay with the ferricxylenol orange complex[J].Anal Biochem，1999，273：149-155.
[4] THORDALCHRISTENSEN H，ZHANG Z，WEI Y，et al.Subcellular localization of H2O2 in plants.H2O2 accumulation in papillae and hypersensitive response during the barleypowdery mildew interaction[J].Plant J，1997，11：1187-1194.
[5] ZHANG J，KIRKHAM M.Droughtstressinduced changes in activities of superoxide dismutase，catalase，and peroxidase in wheat species[J].Plant Cell Physiol， 1994，35：785-791.
[6] ZAKA R，VANDECASTEELE C，MISSET M.Effects of low chronic doses of ionizing radiation on antioxidant enzymes and G6PDH activities in Stipa capillata（Poaceae）[J].J Exp Bot， 2002，53：1979-1987.
[7] LIU C L， DU W C，CAI H S，et al.Trivalent chromium pretreatment alleviates the toxicity of oxidative damage in wheat plants exposed to hexavalent chromium[J].Acta Physiol Plant， 2014，36：787-794.
[8] DENG B L，DONG H S.Ectopic expression of riboflavinbinding protein gene TsRfBP paradoxically enhances both plant growth and drought tolerance in transgenic Arabidopsis[J].J Plant Growth Regul， 2013，32：170-181.
[9] DENG B L.Antioxidative response of golden agave leaves with different degrees of variegation under high light exposure[J].Acta physiol Plant， 2012，34：1925-1933.
[10] MITTLER R.Oxidative stress，antioxidants and stress tolerance[J].Trends Plant Sci， 2002，7：405-410.
[11] GILL S，TUTEJA N.Reactive oxygen species and antioxidant machinery in abiotic stress tolerance in crop plants[J].Plant Physiol Biochem， 2010，48：909-930.
[12] AMIRJANI M.Effect of NaCl on some physiological parameters of rice[J].Eur J Biol Sci， 2010，3：6-16.
[13] LEE D，KIM Y，LEE C.The inductive responses of the antioxidant enzymes by salt stress in the rice（Oryza sativa L.）[J].J Plant Physiol，2001，158：737-745.
[14] LIN C，KAO C.Effect of NaCl stress on H2O2 metabolism in rice leaves[J].Plant Growth Regul， 2000，30：151-155.
[15] VAIDYANATHAN H，SIVAKUMAR P，CHAKRABARTY R，et al.Scavenging of reactive oxygen species in NaClstressed rice（Oryza sativa L.）differential responses in salttolerant and sensitive varieties[J].Plant Sci， 2003，165：1411-1418.
[16] WILLEKENS H，CHAMNONGPOL S，DAVEY M，et al. Catalase is a sink for H2O2 and is indispensable for stress defence in C3 plants[J].EMBO J， 1997，16：4806-4816.
[17] SHIM I，MOMOSE Y，YAMAMOTO A，et al. Inhibition of catalase activity by oxidative stress and its relationship to salicylic acid accumulation in plants[J].Plant Growth Regul， 2003，39：285-292.
[18] QUAN L，ZHANG B，SHI W，et al. Hydrogen peroxide in plants：a versatile molecule of reactive oxygen species network[J].J Integr Plant Biol， 2008，50：218.