,1,2, 吴萌,1, 李桂龙1, 江春玉1, 刘明1,2, 陈瑞蕊1,2, 李忠佩,1,2Slow-Release Synergistic Effect of Humic Acid on Low Concentration Chlorothalonil
WEI ShiPing,1,2, WU Meng,1, LI GuiLong1, JIANG ChunYu1, LIU Ming1,2, CHEN RuiRui1,2, LI ZhongPei,1,2通讯作者:
第一联系人:
收稿日期:2018-07-10接受日期:2018-08-17网络出版日期:2019-01-01
| 基金资助: |
Received:2018-07-10Accepted:2018-08-17Online:2019-01-01
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魏世平, 吴萌, 李桂龙, 江春玉, 刘明, 陈瑞蕊, 李忠佩. 腐殖酸对低浓度百菌清的缓释增效作用[J]. 中国农业科学, 2019, 52(1): 65-72 doi:10.3864/j.issn.0578-1752.2019.01.007
WEI ShiPing, WU Meng, LI GuiLong, JIANG ChunYu, LIU Ming, CHEN RuiRui, LI ZhongPei.
0 引言
【研究意义】尖镰孢（Fusarium oxysporum）是引起多种农作物根腐病、枯萎病的主要真菌致病菌,严重危害农作物的生长和粮食安全[1,2]。针对该致病菌的防治技术包括农业防治、物理防治、化学防治和生物防治,其中化学防治仍然是目前主要的防治手段,百菌清（chlorothalonil）、多菌灵（carbendazim）、代森锰锌（mancozeb）等杀菌剂是农业上常用的化学防治药剂[3]。作为一种广谱高效的保护性杀菌剂,百菌清被广泛应用于农业生产中,但随着使用年限的增加,防治效果逐年下降;且百菌清在水体、土壤中的累积残留量给生态环境和人类带来了新的威胁[3,4,5]。农业生产中如何提高农药药效,进而减少农药的使用量,成为一种研究趋势[6]。农药增效剂是解决这一生态环境问题的重要途径[7]。【前人研究进展】越来越多的绿色农药增效剂应用到农业生产中,如植物油、生物碱、萜烯、茶皂素、腐殖酸等[8,9,10,11]。研究证明腐殖酸能够通过理化作用与农药形成复合物,降低农药毒性,减少环境污染,是一种较好的农药缓释增效剂。腐殖酸类物质在结构单元组成上包含大量的活性基团,如羧基、羰基、醇羟基、醌基等,这些活性基团使得腐殖酸-农药复配体系在杀菌、杀虫等过程中发挥了重要的作用[12]。研究发现,腐殖酸类物质可作为甲基托布津、多菌灵等农药的增效剂[13]。王任明等[14]通过室外盆栽接虫和田间小区试验,得出黄腐酸对杀虫脒存在较好的增效作用;金平等[15]用1% NaOH提取出泥炭的腐殖酸,将其与克露复配喷施黄瓜叶片,从生理机制上证明了腐殖酸对克露起到增效作用。【本研究切入点】目前还未见腐殖酸对百菌清缓释增效作用的报道。另外,田间接种和生理指标测定的试验方法存在工作量大、耗时长等弊端,因此,探索一种新的试验方法实现对农药增效剂的有效筛选意义重大。等温微量热技术在医学上已经普遍用于检测药物活性及其定量构效关系的研究（quantitative structure-activity relationship,QSAR）[16,17],但利用等温微量热技术来筛选农药增效剂的研究未见报道。【拟解决的关键问题】将腐殖酸与农药百菌清同时添加至纯菌培养基中,通过测定病原菌菌丝体的净生长量以及孢子体数量,探索腐殖酸-百菌清复配体系对尖镰孢体外生长的直接影响,同时利用等温微量热仪监测复配体系及单独添加处理生长代谢过程中的热排放,实时监测微生物代谢活性的变化,为农药增效剂的选择提供新的筛选方法以及衡量指标,为扩大腐殖酸在农业领域的应用提供理论依据,为我国煤矿、泥炭等腐殖酸材料来源的充分利用提供新的途径。1 材料与方法
试验于2017年7月至2018年6月在中国科学院南京土壤研究所土壤与农业可持续发展国家重点实验室完成。1.1 腐殖酸材料来源及提取、纯化
供试腐殖酸材料来源于褐煤,购自内蒙古自治区乌海市同升商贸有限责任公司,其腐殖酸的提取、纯化过程参考文献[18,19]。经元素分析仪测定该腐殖酸含碳565.5 g·kg-1,氮15.1 g·kg-1,硫5.1 g·kg-1,氧339.4 g·kg-1。13C固态核磁光谱显示,其化学组成中各碳组分的相对比例为烷基碳11.6%,甲氧基/酰胺碳0.6%,烷氧碳0.7%,异头碳3.4%,芳香碳64.8%,芳香族碳氧8.5%,羰基碳10.5%。1.2 病原菌来源
供试菌种尖镰孢购自中国农业科学院农业资源与农业区划研究所中国农业微生物菌种保藏管理中心,为花生根腐病致病菌株。1.3 菌丝体体外平板培养试验
试验设置4个处理,分别为PSA（potato sugar agar）培养基对照、单独添加低浓度百菌清处理、单独添加腐殖酸处理以及同时添加腐殖酸和低浓度百菌清的处理。PSA固体培养基配方：200 g马铃薯加蒸馏水煮沸,小火煮20 min,加20 g蔗糖和18 g琼脂粉,定容至1 L,115℃灭菌,30 min。PSA液体培养基的配制不添加琼脂粉。
将腐殖酸和百菌清用二甲基亚砜（dimethyl sulfoxide,DMSO）溶解,并用含有200 μg·mL-1乳化剂的无菌蒸馏水稀释至相应浓度,腐殖酸添加浓度为50 mg·L-1,百菌清添加浓度为5 mg·L-1。吸取1 mL检测液至无菌培养皿中（直径9 cm）,加入9 mL的PSA固体培养基。菌丝接种环的直径约5 mm,将致病真菌接种至各处理培养皿的中间位置,25℃培养72 h,测量菌丝体的生长直径,每处理 3—4个重复。
1.4 孢子体计数
试验处理同1.3,但固体培养基改为不添加琼脂的PSA液体培养基。将致病菌从平板上接种至PSA液体培养基中,30℃摇菌培养48 h（摇菌时间根据孢子浓度适当调整）,吸取一定体积的菌悬液至含有新鲜培养基的不同处理中,接种后孢子初始浓度控制在2.5×105个/mL[17,20],将各处理菌悬液置于25℃摇菌,分别于摇菌培养的第3、7、14、33天采样进行孢子数计数。使用血球计数板进行计数,血球计数板规格为25个中方格×16个小方格,选择左上、左下、右上、右下以及中央的中方格进行孢子计数,对于临界的孢子体需遵循数上不数下,数左不数右的原则,最后折算为各摇菌瓶中的孢子总数。1.5 热动力学试验
试验处理同1.4,4 mL的安培瓶内最终上机液体体积为3 mL,其中包含各处理相应量的菌悬液、DMSO、腐殖酸、百菌清及PSA液体培养基。菌悬液配制同1.4,最终上机真菌孢子数同样为2.5×105个孢子/mL。将安培瓶置于TAM III（TA Instruments,Utah,USA）检测通道中,设定油浴温度为25℃恒温,代谢反应过程中的功率-时间曲线被连续监测72 h（根据试验结果可适当调整）。用TAM assistant软件对热排放曲线进行处理,汇总各热排放曲线的相关参数,包括最大热功率（Pmax）、最大热功率时间（Tmax）、整体发热量（Q）和生长速率常数k[21,22,23]。1.6 数据处理与统计分析
根据真菌菌丝体净生长量计算抑菌率（inhibition rate,IR）,计算公式：IR（%）=100×（对照菌丝体净生长量-处理菌丝体净生长量）/对照菌丝体净生长量。腐殖酸-百菌清复配处理的相对增效（%）=100×（复配处理抑菌率-百菌清处理抑菌率）/百菌清处理抑菌率。
热功率-时间曲线中生长速率常数k是由OriginPro8.0中Exponential进行指数模型拟合得到。各处理抑菌率、孢子数以及热排放曲线相关参数的单因素方差分析和Tukey B、Duncan多重比较由SPSS 25.0完成。
2 结果
2.1 腐殖酸对百菌清抑制病原菌菌丝体生长的作用
将对照抑菌率（IR）设为0,得到各处理对尖镰孢的IR（表1）。表1中百菌清、腐殖酸以及腐殖酸-百菌清复配处理对尖镰孢的IR均大于0,即各处理均存在抑制作用。同时,腐殖酸-百菌清复配处理的IR较百菌清处理有显著的提高,相对增效为25.33%,表明腐殖酸对低浓度百菌清抑制尖镰孢菌丝体生长存在显著的增效作用。Table 1
表1
表1不同处理对尖镰孢的抑菌率
Table 1
| 处理 Treatment | 抑菌率IR (%) |
|---|---|
| 对照 Control | 0d |
| 百菌清Chlorothalonil | 40.62b |
| 腐殖酸Humic acid | 6.60c |
| 腐殖酸+百菌清Humic acid+chlorothalonil | 50.91a |
新窗口打开|下载CSV
2.2 腐殖酸对百菌清抑制病原菌孢子体数量增加的作用
利用血球计数板对各处理不同培养时间采样的真菌孢子体进行计数,结果见图1。摇菌培养的第3天,对照与腐殖酸处理的孢子数极显著大于百菌清处理与腐殖酸-百菌清复配处理,而百菌清处理与腐殖酸-百菌清复配处理的孢子数之间无显著差异;摇菌培养第7天,对照的孢子数极显著大于腐殖酸处理及腐殖酸-百菌清复配处理,而百菌清处理的孢子数极显著小于其他处理;摇菌培养第14天,腐殖酸-百菌清复配处理的孢子数显著小于其他处理,而百菌清处理与对照之间无显著差异。以上结果表明腐殖酸对低浓度百菌清抑制尖镰孢孢子数增加的效应存在显著的延长作用。图1
新窗口打开|下载原图ZIP|生成PPT图1不同培养时间下各处理尖镰孢孢子体数量
图中同一培养时间下处理间不同字母为Duncan多重比较结果。*：差异显著（P<0.05）;**：差异极显著（P<0.01）
Fig. 1F. oxysporum spore number of treatments in different liquid shake cultured times
Different letters of the same cultured time in figure are the results of multiple comparisons analysis with Duncan.*: Significant difference (P<0.05); **: Extremely significant difference (P<0.01)
2.3 腐殖酸对百菌清抑制病原菌生长代谢的作用
各处理尖镰孢热功率-时间曲线见图2。热功率-时间曲线图显示腐殖酸处理与对照的热排放曲线接近,表明两个处理中病原菌的生长代谢过程相近,即腐殖酸对病原菌在生长代谢过程中的热量排放未检测到抑制作用。而百菌清处理与对照、腐殖酸处理差异较大,在监测初期热量排放曲线接近直线,百菌清对病原菌的生长起到了很好的抑制作用,这一过程主要来自百菌清杀灭病原菌的作用,随着监测时间的延长,热排放峰又重新出现,表明百菌清对病原菌的药效减弱,病原菌重新开始生长产生代谢热。腐殖酸-百菌清复配处理从监测初期开始热排放一直呈现直线状态,表明代谢热的排放始终受到抑制。从图3也可以看出,4个处理经微量热仪72 h、25℃恒温监测培养后,只有腐殖酸-百菌清复配处理的安培瓶中未见明显的菌丝,这与热功率-排放曲线的结果一致。图2
新窗口打开|下载原图ZIP|生成PPT图2不同处理尖镰孢的热功率-时间曲线
Fig. 2Heat flow-time curve of F. oxysporum of different treatments
图3
新窗口打开|下载原图ZIP|生成PPT图3不同处理尖镰孢恒温监测培养后的菌悬液
1：对照Control;2：百菌清处理Chlorothalonil treatment;3：腐殖酸处理Humic acid treatment;4：腐殖酸和百菌清复配处理Humic acid-chlorothalonil combined treatment
Fig. 3Incubated suspension of F. oxysporum of different treatments monitored by constant temperature
热功率-时间曲线的Pmax、Tmax、Q和k是用于评估微生物代谢活性的相关参数,其中Pmax、Q和k与代谢活性成正相关,而Tmax与代谢活性成负相关。各处理热功率-时间曲线的相关参数（图4）经进一步的统计分析得到,百菌清处理的Pmax显著低于对照和腐殖酸处理,而Tmax显著高于对照和腐殖酸处理,表明病原菌生长代谢活性受到百菌清的显著抑制。腐殖酸-百菌清复配处理的Pmax、Q、k均显著低于其他处理,表明该处理病原菌的生长代谢活性显著低于其他处理,即病原菌生长代谢受到抑制的程度最大。腐殖酸处理与对照各参数之间无显著差异,表明病原菌的生长代谢活性没有受到抑制,同时说明腐殖酸-百菌清复配处理中生长代谢受到的抑制作用与腐殖酸本身无关。由于腐殖酸-百菌清复配处理的热排放曲线接近直线,故Tmax在初始时最大。综上,腐殖酸对百菌清杀灭尖镰孢存在显著的缓释增效作用。
图4
新窗口打开|下载原图ZIP|生成PPT图4不同处理尖镰孢热功率-时间曲线相关参数的分布
柱上不同字母表示Tukey B多重比较差异显著（P<0.05）
Fig. 4Distribution of related parameters of F. oxysporum heat flow-time curve under different treatments
Different letters on the bar indicate significantly different at P<0.05 level by Tukey B multiple comparisons
3 讨论
通过尖镰孢体外培养和热动力学技术,本研究证实了腐殖酸对低浓度百菌清的缓释增效作用,增效作用提高的抑菌率与前人总结的10%—20%的增效范围相符[12]。目前在农业生产上,腐殖酸的主要应用包括作为土壤调节剂来改善土壤属性,以及作为植物刺激素来促进植物生长和增加作物产量[24,25,26]。而本研究的结果为拓展腐殖酸的应用范围提供了理论依据,即将腐殖酸作为农药增效剂来增加和延长百菌清的药效。早期的研究中,李善祥[12]就指出腐殖酸作为一种高效、无公害、低成本、多功能的农药增效剂有着广阔的发展前景,并且将腐殖酸的增效机理概括为：（1）腐殖酸与农药之间的化学作用,使得农药的化学结构发生变化从而导致其活性结构的改善;（2）腐殖酸没有明显影响农药的化学结构,但增加了农药的表面活性、膜透性、生理生化作用以及吸附作用等。金平等[15]研究证实了腐殖酸对克露的增效作用,其生理机制主要是因为腐殖酸-克露复配处理降低了黄瓜叶片脯氨酸、丙二醛的含量以及质膜透性、黄瓜霜霉病的病情指数,从而促进了黄瓜的生长,提高了黄瓜的产量及谷胱甘肽的含量;于宏伟等[9]认为黄腐酸是一种胶体物质,同时具有表面活性剂的作用,对有机农药具有强烈的吸附作用,并且使难溶于水相的农药更好地分散,以此来增加杀菌剂的杀菌效果;ABDEL- MONAIM 等[27]利用可溶性腐殖酸钾颗粒与苯并噻二唑（benzothiadiazole,BTH）浸泡大豆种子,在大田种植条件下,腐殖酸钾-苯并噻二唑复配体系浸泡后的处理能够有效地抵抗尖镰孢对大豆植株的危害;复配处理植株的过氧化物酶（PO）、多酚氧化酶（PPO）、苯丙氨酸解氨酶（PAL）3种酶活性和酚含量最高。腐殖酸钾增效的机理为增加的PO和PPO活性可以产生氧化形式的醌,这可能有助于增加防御能力,因为氧化形式的醌可以灭活病原菌产生的果胶酶。PAL是一种苯丙类化合物的关键酶,它会导致各种与防御有关的植物次生代谢物的产生,如水杨酸、植物抗毒素和类木质素聚合物等。本研究中,百菌清处理的热功率-时间曲线从生长代谢方面较好地体现了百菌清的重要杀菌机理,即通过对病原菌糖酵解过程产生影响而起到杀灭致病菌的作用。糖酵解过程中,3-磷酸甘油醛在磷酸甘油醛脱氢酶（GAPDH）的催化下,转变为1,3-二磷酸甘油酸,同时将氧化过程产生的能量贮存到三磷酸腺苷（ATP）分子中。而百菌清分子中的活性氯能够与磷酸甘油醛脱氢酶上的-SH结合,生成一种复合物,从而使酶的活性丧失,中断糖酵解过程中3-磷酸甘油醛的脱氢氧化,影响ATP的产生[28,29]。等温培养初期百菌清抑制了病原菌的糖酵解过程,没有热量排放,故热排放曲线接近直线,后期随着百菌清药效的降低,病原菌糖酵解过程恢复,重新出现热排放峰。腐殖酸-百菌清复配处理,其热功率-时间曲线始终为近似直线排放,笔者推断在腐殖酸存在的条件下,百菌清对病原菌糖酵解过程的影响增大,且对能量产生的抑制作用更持久,从而无法出现热排放峰。这与张国生等[7]总结的农药增效剂主要作用机理相一致,即农药增效剂是通过抑制或弱化靶标（害虫、杂草、病菌等）对农药活性的解毒作用,延缓药剂在防治对象内的代谢速度,从而增加防治效果。后续还需对腐殖酸的结构以及对腐殖酸-百菌清复配体系中的相关酶活性、谷胱甘肽含量等进行研究测定,从而对作用机理有更深入的了解。同时,为了更好地诠释腐殖酸缓释增效作用的机理,下一步还需要涉及更多不同来源、结构组成的腐殖酸。
本研究中等温微量热技术与病原菌离体培养试验结果的一致性表明等温微量热技术应用于农药增效剂筛选具有可行性。在应用生物学和医学领域,等温微热技术的非标记和被动属性可以对复杂的三维结构进行测量（如微观组织）,以此来代替破坏性的常规分析（如DNA或蛋白质的量化）[30]。在土壤学领域,微量热技术与高效气液相色谱、高通量测序等技术的结合,有望在土壤微生物活性与结构的地理属性、气候变化、土壤质量的空间大尺度比较和外源污染物的生物毒性评价等方面发挥其重要作用[31]。TAFIN等[17]在纯培养条件下,利用等温微量热技术估算两性霉素B、卡泊芬净等组合对两株曲霉（Aspergillus fumigatus和Aspergillus terreus）的抑制活性,研究证实微量热技术具有实时评估药物组合抑真菌活性的潜力。等温微量热技术同时还具备操作方便、检测灵敏、材料消耗少且重现性好等优势,具有很好的应用前景。
4 结论
腐殖酸对低浓度百菌清存在显著的缓释增效作用。在低浓度百菌清体系中添加腐殖酸,可以显著增加低浓度百菌清对尖镰孢的抑菌率,抑制尖镰孢孢子体数量的增加和生长代谢过程。将腐殖酸作为百菌清的农药增效剂具有生产和商业应用潜力。等温微量热技术作为筛选农药增效剂新的方法及衡量指标具有可行性。参考文献 原文顺序
文献年度倒序
文中引用次数倒序
被引期刊影响因子
URLMagsci [本文引用: 1]
<p>为明确尖孢镰刀菌种内各菌株间的遗传差异与亲缘关系，采用ISSR分子标记技术对33株地理来源不同的尖孢镰刀菌进行了遗传多样性分析。结果表明：利用筛选出的11条引物扩增出105条条带，其中多态性条带91条，多态性位点比例为86.7%；遗传相似性与聚类分析结果〖JP2〗供试菌株间的遗传相似系数为0.606～0.962，平均0.756，当遗传相似系数为0.962时，供试的33株菌可被全部区分开。表明，尖孢镰刀菌基因组在SSR区域具有丰富的多态性。寄主来源相同的供试菌株间的遗传相似性与其地理来源有一定的相关性。</p>
URLMagsci [本文引用: 1]
<p>为明确尖孢镰刀菌种内各菌株间的遗传差异与亲缘关系，采用ISSR分子标记技术对33株地理来源不同的尖孢镰刀菌进行了遗传多样性分析。结果表明：利用筛选出的11条引物扩增出105条条带，其中多态性条带91条，多态性位点比例为86.7%；遗传相似性与聚类分析结果〖JP2〗供试菌株间的遗传相似系数为0.606～0.962，平均0.756，当遗传相似系数为0.962时，供试的33株菌可被全部区分开。表明，尖孢镰刀菌基因组在SSR区域具有丰富的多态性。寄主来源相同的供试菌株间的遗传相似性与其地理来源有一定的相关性。</p>
[本文引用: 1]
DOI:10.7606/j.issn.1004-1389.2016.04.020URL [本文引用: 2]
草莓枯萎病是目前危害北京草莓产业的主要病害,为筛选出安全有效的杀菌剂,采用组织分离法从北京昌平地区采集的草莓枯萎病株中分离到引起草莓枯萎病的病原菌。经形态学和分子生物学手段鉴定该病原菌为尖孢镰刀菌草莓专化型(Fusarium oxysporum Schlecht f.sp.fragariae Winks et Williams)。采用菌丝生长速率法对14种杀菌剂进行筛选,其中,206.7g/L噁酮·氟硅唑、寡雄腐霉、φ=25%腈菌唑、w=60%嘧菌酯、w=70%代森锰锌、w=75%百菌清、w=50%异菌脲7种杀菌剂对草莓枯萎病具有显著的抑菌效果,其EC50值分别为0.003、0.63、0.73、5.91、8.90、78.54、229.09mg/L;另外,206.7g/L噁酮·氟硅唑、寡雄腐霉、w=70%代森锰锌和w=75%百菌清4种杀菌剂的药效可持续96h以上,其他3种药剂处理48h后对草莓枯萎病的抑制效果显著下降。因此,206.7g/L噁酮·氟硅唑、寡雄腐霉、w=70%代森锰锌和w=75%百菌清是草莓枯萎病防治的优先选择。
DOI:10.7606/j.issn.1004-1389.2016.04.020URL [本文引用: 2]
草莓枯萎病是目前危害北京草莓产业的主要病害,为筛选出安全有效的杀菌剂,采用组织分离法从北京昌平地区采集的草莓枯萎病株中分离到引起草莓枯萎病的病原菌。经形态学和分子生物学手段鉴定该病原菌为尖孢镰刀菌草莓专化型(Fusarium oxysporum Schlecht f.sp.fragariae Winks et Williams)。采用菌丝生长速率法对14种杀菌剂进行筛选,其中,206.7g/L噁酮·氟硅唑、寡雄腐霉、φ=25%腈菌唑、w=60%嘧菌酯、w=70%代森锰锌、w=75%百菌清、w=50%异菌脲7种杀菌剂对草莓枯萎病具有显著的抑菌效果,其EC50值分别为0.003、0.63、0.73、5.91、8.90、78.54、229.09mg/L;另外,206.7g/L噁酮·氟硅唑、寡雄腐霉、w=70%代森锰锌和w=75%百菌清4种杀菌剂的药效可持续96h以上,其他3种药剂处理48h后对草莓枯萎病的抑制效果显著下降。因此,206.7g/L噁酮·氟硅唑、寡雄腐霉、w=70%代森锰锌和w=75%百菌清是草莓枯萎病防治的优先选择。
DOI:10.1016/j.saa.2010.04.020URLPMID:20537938 [本文引用: 1]
NIR FT-Raman and IR spectra of the biologically active molecule, chlorothalonil have been recorded and analyzed. The molecular geometry and vibrational wavenumbers of the title compound in the ground state have been calculated by density functional theory (DFT) with 6-31G(d) basis set. In order to obtain the information about the influence of intramolecular interaction on the molecule, the calculated geometries of chlorothalonil molecule was compared with the experimental data. The results of the optimized molecular structure gave clear evidence for the intramolecular charge transfer (ICT). Time-dependent density functional theory (TD-DFT) calculation of the electronic spectra has been performed and compared with the experimental UV–visible spectrum. Mulliken's net charges have been calculated and compared with the atomic natural charges. The effects of chlorine and cyanide group substituent in benzene ring in the vibrational wavenumbers have been analyzed. NBO analysis is useful to understand the intramolecular hyperconjugative interaction between lone pair Cl and σ*(C–C) bond orbital.
DOI:10.1016/j.yrtph.2016.05.035URLPMID:27240750 [本文引用: 1]
61An non-parameter probabilistic model was developed to assess the risk of dietary exposure to chlorothalonil.61The health risk of chlorothalonil residues to 2 consumer groups via vegetable exposure was evaluated.61The Young children was more sensitive to chlorothalonil residues.61The results would provide useful information for re-evaluating and revising the pesticides.
DOI:10.1021/acs.jafc.6b05034URL [本文引用: 1]
A semi-automated qualitative method for target screening of 448 pesticide residues in fruits and vegetables was developed and validated using ultra-high performance liquid chromatography coupled with electrospray ionization quadrupole Orbitrap high resolution mass spectrometry (UHPLC/ESI Q-Orbitrap). The Q-Orbitrap Full MS/dd-MS2 (data dependent acquisition) was used to acquire product-ion spectra of individual pesticides to build a compound database or an MS library, while its Full MS/DIA (data independent acquisition) was utilized for sample data acquisition from fruit and vegetable matrices fortified with pesticides at 10 and 100 碌g/kg for target screening purpose. Accurate mass, retention time and response threshold were three key parameters in a compound database that were used to detect incurred pesticide residues in samples. The concepts and practical aspects of in-spectrum mass correction or solvent background lock-mass correction, retention time alignment and response threshold adjustment are dis...
DOI:10.3969/j.issn.1006-4184.2000.04.010URL [本文引用: 2]
介绍了农药增效剂的作用机理、现状及应用前景，以及不同增效剂在相应农药中的应用情况。
DOI:10.3969/j.issn.1006-4184.2000.04.010URL [本文引用: 2]
介绍了农药增效剂的作用机理、现状及应用前景，以及不同增效剂在相应农药中的应用情况。
DOI:10.3321/j.issn:0578-1752.1998.02.006URLMagsci [本文引用: 1]
经室内毒力测定及田间药效试验研究结果表明：茶皂素对20%灭多威乳油杀灭菜青虫增效6.33倍,对2.5%三氟氯氰菊酯乳油杀灭菜缢管蚜增效14.83倍,对5%尼索朗乳油杀灭桔全爪螨卵增效61.8倍,对20%哒螨灵(WP)杀灭桔全爪螨成螨增效18.67倍。茶皂素与上述化学农药原药复方乳油的田间杀虫效果均稳定保持在90%以上，与常规对照农药效果经DMRT法显著性分析，在5%水平上无显著差异；而且可以降低化学农药原药有效成分用量50%～75%。对于减少化学农药污染环境，开发我国及世界极其丰富的茶皂素资源，具有极显著的社会、生态和经济效益。
DOI:10.3321/j.issn:0578-1752.1998.02.006URLMagsci [本文引用: 1]
经室内毒力测定及田间药效试验研究结果表明：茶皂素对20%灭多威乳油杀灭菜青虫增效6.33倍,对2.5%三氟氯氰菊酯乳油杀灭菜缢管蚜增效14.83倍,对5%尼索朗乳油杀灭桔全爪螨卵增效61.8倍,对20%哒螨灵(WP)杀灭桔全爪螨成螨增效18.67倍。茶皂素与上述化学农药原药复方乳油的田间杀虫效果均稳定保持在90%以上，与常规对照农药效果经DMRT法显著性分析，在5%水平上无显著差异；而且可以降低化学农药原药有效成分用量50%～75%。对于减少化学农药污染环境，开发我国及世界极其丰富的茶皂素资源，具有极显著的社会、生态和经济效益。
DOI:10.3969/j.issn.1002-1302.2010.02.060URL [本文引用: 2]
综述报道了绿色农药增效剂在杀虫剂、杀菌剂、除草剂等方面的应用进展。并简述了植物油类、生物碱类、萜烯类等绿色农药增效剂在杀虫剂方面的应用,茶皂素和黄腐酸在病毒防治方面的应用,烷基多糖苷和黄腐酸在除草剂方面的应用。
DOI:10.3969/j.issn.1002-1302.2010.02.060URL [本文引用: 2]
综述报道了绿色农药增效剂在杀虫剂、杀菌剂、除草剂等方面的应用进展。并简述了植物油类、生物碱类、萜烯类等绿色农药增效剂在杀虫剂方面的应用,茶皂素和黄腐酸在病毒防治方面的应用,烷基多糖苷和黄腐酸在除草剂方面的应用。
[本文引用: 1]
(in Chinese)
[本文引用: 1]
DOI:10.1016/j.pmpp.2017.05.004URL [本文引用: 1]
Evaluation of monocrotophos and Azadirachta indica oil, herein referred to as ‘Vijayneem’, were combined in ratios of 10% (10: 90), 20% (20: 80), 30% (30: 70), 40% (40: 60) and 50% (50: 50) [Monocrotophos: Vijayneem] under laboratory conditions to determine efficacy for properties of pesticidal, repellency, attraction, stimulant of consumption, development and reproduction. Asa biopesticide, Vijayneem oil (LC 50 =10.619%), caused greater mortality in dermal toxicity bioassays (LC 50 =18.045%) over ingestion. The greatest repellent activity was observed in third (Access Proportion Index=610.25), fourth (API=610.52) and fifth instar larvae (API=610.67) in sprays of cotton leaves with monocrotophos. The detoxification enzymes like esterase (0.3, 11.1μl/min/mg) and lactate dehydrogenase (6.8, 13.8μl/min/mg) levels increased postingestion of treatments. Insects which ingested treated leaves showed an increase in carbohydrate (27.0, 18.0%), protein (4.7, 7.7%) and lipid (4.0, 10.4%) content. The greatest deformation of larvae, pupae and adults occurred in monocrotophos alone treatments, while the least deformities were recorded for monocrotophos+Vijayneem treatments. The male and female ratio and percentage of adult emergence was high in control than in A.indica oil alone treated S.litura . The greatest fecundity was observed in control (1620/4 female). In conclusion, the synergistic effects of A.indica oil+monocrotophos can be used for use the management of S.litura .
URL [本文引用: 3]
本文分析了腐植酸的组成结构和胶体、吸附、络合、化学反应以及表面活性性能,研究了腐植酸类物质与农药的相互作用,探讨其作用机制及效果.
URL [本文引用: 3]
本文分析了腐植酸的组成结构和胶体、吸附、络合、化学反应以及表面活性性能,研究了腐植酸类物质与农药的相互作用,探讨其作用机制及效果.
URL [本文引用: 1]
介绍了黄腐酸的抗菌抗病功能，与某些农药复配后的增效作用，以及增效机理与对农药的降毒作用。
URL [本文引用: 1]
介绍了黄腐酸的抗菌抗病功能，与某些农药复配后的增效作用，以及增效机理与对农药的降毒作用。
DOI:10.3969/j.issn.1008-3103.2001.01.012URL [本文引用: 1]
黄腐酸可刺激作物生长,作者提出了腐植酸和杀虫剂的混合方法,作为一种新剂型,它比单用农药效果好.根据水稻试验结果,作者确信此混合剂和空白对照比较可增产17%.
DOI:10.3969/j.issn.1008-3103.2001.01.012URL [本文引用: 1]
黄腐酸可刺激作物生长,作者提出了腐植酸和杀虫剂的混合方法,作为一种新剂型,它比单用农药效果好.根据水稻试验结果,作者确信此混合剂和空白对照比较可增产17%.
DOI:10.3969/j.issn.1005-9369.2003.01.002URL [本文引用: 2]
为探讨腐殖酸增效防病的生理机制 ,测定了脯氨酸、丙二醛、谷胱甘肽含量、膜透性等生理指标。结果表明 ,腐殖酸复配克露降低了叶片脯氨酸、丙二醛含量、质膜透性及病情指数 ,而且促进了黄瓜的生长 ,提高了产量及谷胱甘肽含量。证明腐殖酸作为农药增效剂确实提高了黄瓜的抗病能力 ,起到对农药的增效作用
DOI:10.3969/j.issn.1005-9369.2003.01.002URL [本文引用: 2]
为探讨腐殖酸增效防病的生理机制 ,测定了脯氨酸、丙二醛、谷胱甘肽含量、膜透性等生理指标。结果表明 ,腐殖酸复配克露降低了叶片脯氨酸、丙二醛含量、质膜透性及病情指数 ,而且促进了黄瓜的生长 ,提高了产量及谷胱甘肽含量。证明腐殖酸作为农药增效剂确实提高了黄瓜的抗病能力 ,起到对农药的增效作用
[本文引用: 1]
DOI:10.1016/j.diagmicrobio.2013.06.004URLPMID:23871413 [本文引用: 3]
We evaluated the activity of antifungals alone or in combination against Aspergillus fumigatus and Aspergillus terreus by real-time measurement of fungal growth-related heat production. Amphotericin B, voriconazole, caspofungin, and anidulafungin were tested alone or in combination. Heat production was measured in Sabouraud dextrose broth containing 10(5) Aspergillus conidia/mL for 48 h at 37 degrees C. Antifungal activity was evaluated by measuring the heat detection time relative to the growth control. Against A. fumigatus, the voriconazole-echinocandin combination demonstrated longer heat detection time than each antifungal alone. Against A. terreus, the combination amphotericin B-echinocandin prolonged the heat detection time, compared to each antifungal alone. In contrast, the echinocandin-voriconazole combination did not increase the heat detection time, compared to voriconazole alone. None of the antifungal combinations decreased the heat detection time compared to the antifungals alone (e.g. antagonism was not observed). Microcalorimetry has the potential for real-time evaluation of antifungal combinations against Aspergillus spp. (c) 2013 Elsevier Inc. All rights reserved.
DOI:10.1038/srep32858URLPMID:5011764 [本文引用: 1]
In the background of rapid expansion of plastic greenhouse vegetable production in China, many environmental risks have emerged in recent years. In this study, the soils with a chronosequence in greenhouse vegetable fields were collected and the soil humic acids (HAs) and fluvic acids (FAs) were extracted and purified. The soil HAs and FAs were found to show inhibition activities against phytopathogenic fungi for the first time. Fourier transform infrared spectroscopy was performed to investigate the chemical structures of HAs and FAs. The variation of relative peak areas indicated the chemical structure of HAs become more complex and stable under continuous cultivation. The PCA analysis showed HAs and FAs could be distinctly separated from each other and cultivation years mainly determined the variation. Mantel test and RDA analysis indicated the active components (aliphatic peaks for HAs and COOH, OH peaks for FAs) had positive correlation with the inhibition rates of HAs and FAs against phytopathogenic fungi. According to our research, the active fungicidal components in soil HAs and FAs decreased along with the extension of cultivation years, which made the soil suffer more risk to phytopathogenic fugi. So we believe continuous cultivation too many years in PGVP systems is inadvisable.
DOI:10.1021/acs.jafc.8b01931URLPMID:29987927 [本文引用: 1]
DOI:10.3321/j.issn:1000-4343.2007.01.018URL [本文引用: 1]
应用微量热法研究了配合物RE（CTH5O3）2（C9H6NO）（RE代表La，Sm和 Nd）对真菌的抗菌作用。在TAM Air热导式等温微量量热仪上，分别测定了桔青霉菌和黑曲霉菌在不同浓度不同稀土配合物及空白条件下生长代谢热谱曲线，并计算得到了真菌在不同条件下的生 长代谢速率常数k和传代时间G等热动力学参数。实验表明：3种稀土水杨酸8-羟基喹啉三元配合物对桔青霉菌和黑曲霉菌均有抑制作用，其抑制效果依次 为：Sm（Hsal）2（hq）〉La（Hsal）2（hq）〉Nd（Hsal）2（hq）。
DOI:10.3321/j.issn:1000-4343.2007.01.018URL [本文引用: 1]
应用微量热法研究了配合物RE（CTH5O3）2（C9H6NO）（RE代表La，Sm和 Nd）对真菌的抗菌作用。在TAM Air热导式等温微量量热仪上，分别测定了桔青霉菌和黑曲霉菌在不同浓度不同稀土配合物及空白条件下生长代谢热谱曲线，并计算得到了真菌在不同条件下的生 长代谢速率常数k和传代时间G等热动力学参数。实验表明：3种稀土水杨酸8-羟基喹啉三元配合物对桔青霉菌和黑曲霉菌均有抑制作用，其抑制效果依次 为：Sm（Hsal）2（hq）〉La（Hsal）2（hq）〉Nd（Hsal）2（hq）。
DOI:10.1016/0040-6031(95)90686-XURL [本文引用: 1]
Microcalorimetry has been used to study the effect of moisture on soil microbial activity. Different moisture regimes produce changes on heat flow rate-time curves recorded for soil samples, affecting also certain parameters, such as the total heat evolution Q tot , the microbial growth rate constant and growth yield Y , which are calculated by the microcalorimetric method. A positive correlation was found between the percentage humidity, the total heat evolution and the microbial growth rate constant. The values of these parameters decrease with the dryness of the soil. Field capacity moisture appears to be the optimum moisture level for soil microbial activity.
DOI:10.3969/j.issn.0253-9829.2012.04.002URL [本文引用: 1]
一切生命过程都伴随着热效应。微量热法可对这些热效应进行精确的跟踪，并表征其过程。微量热法已广泛应用于医药、生化等研究领域，但在土壤微生物研究中却鲜有应用。本文简要介绍微量热法的原理和实验方法，综述其在土壤微生物研究中的应用现状井展望其应用前景，以期为土壤微生物研究提供新的思路和技术。
DOI:10.3969/j.issn.0253-9829.2012.04.002URL [本文引用: 1]
一切生命过程都伴随着热效应。微量热法可对这些热效应进行精确的跟踪，并表征其过程。微量热法已广泛应用于医药、生化等研究领域，但在土壤微生物研究中却鲜有应用。本文简要介绍微量热法的原理和实验方法，综述其在土壤微生物研究中的应用现状井展望其应用前景，以期为土壤微生物研究提供新的思路和技术。
DOI:10.1016/j.soilbio.2016.11.014URL [本文引用: 1]
Despite our current understanding of soil C and N interactions, less is known about the response and feedback of C mineralization to soil P availability driven by microorganisms. To better understand these interactions, soils with long-term P-sufficient NPK fertilization (available P: 13.4mgkg 1 ) and P-deficient NK fertilization (available P: 0.96mgkg 1 ) were incubated with and without glucose. CO 2 emissions were monitored to characterize C mineralization during the incubation. The soil bacterial community structure, quantity and metabolic activity were evaluated using high-throughput sequencing, qPCR and microcalorimetric dynamics. Compared with P-sufficient soils, P-deficient soils had significantly lower basal respiration, but a significantly higher net mineralization of added glucose, probably due to higher energy cost of soil microorganisms. Glucose addition promoted microbial biomass and activity, particularly in P-deficient soils, and this improvement was maintained for at least 70 days. Shifts in bacterial community composition were induced by a predominance of several specific taxonomic groups, all of which were capable of solubilizing P in soils. P-deficiency decreases the retention of exogenous labile C into soil. Adding labile C to P-deficient soils may shift P from relatively unavailable soil-bound pools into microbial biomass pools through pool cycling. Our results indicated negative effects of P-deficiency on soil C retention, as well as positive effects of labile C on soil P availability in arable soils.
DOI:10.1016/j.fuel.2014.06.011URL [本文引用: 1]
The H2O2 degradation of humic acids and lignite from M r mine (South Moravian region, Czech Republic) resulted in oxidation of aromatic structures and cleavage of aromatic units. The hydrophilic fraction largely dominated the oxidation digest in both parental materials. The presence of polycyclic biomarkers -Phyllocladane, De-A-lupane confirms the high plant contribution. Hopanoid structures and branched C15 and C17 (iso and anteiso) fatty acids show slight bacterial inputs. Short chain n-fatty acids (<C20) are ubiquitous in living kingdom, whereas long chain homologs are probably derived from higher plants. Phosphoric acid production through the H2O2 oxidation probably results from hydrolyses of pyrophosphate. From hydrophilic fractions of lignite and humic acids, malonic acid and succinic acid are dominant compounds and could indicate that methylene and ethylene chains are main bridge bonds connecting between the aromatic rings in the structure. The H2O2 oxidation of lignite was very selective for producing malonic acid and succinic acid in high yields and the oxidation degradation method appears to be a suitable new way for utilizing South Moravian lignite. For production of malonic acid and succinic acid, the results indicated that the appropriate condition especially was at 50 C for 4h and 40 C for 4h, respectively. But oxidation followed by reoxidation of lignite at 30 C for 1h could be also a reasonable way to produce that kind of molecules (particularly malonic acid). Lipophilic compounds represented the minor GC-amenable fraction of the total products obtained by oxidation of lignite.
DOI:10.1016/j.scienta.2014.05.010URL [本文引用: 1]
Humic acid (HA) is natural biological organic fertilizer derived from organic waste materials, which shows a highly promoting effect on plant growth and development. However, the mechanisms responsible for the promoting effect of humic acid on the growth and development are poorly understood. In this study, the physiological mechanisms of foliar humic acid (FHA) fertilizer on chrysanthemum growth and development from the viewpoint of photosynthesis and chloroplast ultrastructure were investigated. Seedlings of chrysanthemum were sprayed with the same volume of distilled H2O (control), inorganic NPK fertilizer and organic foliar humic acid fertilizer every 15 days (15, 30, 45, 60 days after transplanting). The results showed that the morphological indices (stem diameter, fresh weights of shoots and roots, the root to shoot ratios, dry weights of shoots and roots, leaf area, flower diameter), the net photosynthesis rate, the chlorophyll fluorescence, the content of chlorophyll and the chloroplast ultrastructure of chrysanthemum improved obviously after foliar application of humic acid compared with those of the control and the NPK fertilizer. The promoting effect of the humic acid fertilizer showed an obvious dose-effect. The results from correlation analysis indicated that the responses of the foliar humic acid fertilizer on growth and development of chrysanthemum could be related with a constitutive increased net photosynthetic rate due to the high content of chlorophyll and the improved chloroplast ultrastructure.
DOI:10.1021/acssuschemeng.7b02094URL [本文引用: 1]
DOI:10.5941/MYCO.2011.39.4.290URLPMID:3385127 [本文引用: 1]
The ability of benzothiadiazole (BTH), humic acid (HA) and their combination when used as seed soaking to induce systemic resistance against a pathogenic strain of Fusarium oxysporum was examined in four soybean cultivars under greenhouse conditions. Both inducers and their combination were able to protect soybean plants against damping-off and wilt diseases compared with check treatment. These results were confirmed under field conditions in two different locations i. e Minia and New Valley governorates. The tested treatments significantly reduced damping-off and wilt diseases and increased growth parameters, except number of branches plant-1, and seed yield. Application of BTH (0.25) + HA (4 g/l) was the most potent in this respect treatment. Soybean seed soaking in BTH+ HA recorded the highest activities of the testes of oxidative enzymes followed by BTH in the four soybean cultivars. Whereas, HA treatment was recorded the lowest increased of these oxidative enzymes. Also, similar results were obtained in case of total phenol but HA increased the total phenol more than BTH in all tested cultivars.
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[本文引用: 1]
DOI:10.3724/SP.J.1260.2013.20175URLMagsci [本文引用: 1]
甘油醛-3-磷酸脱氢酶(glyceraldehyde-3-phosphate dehydrogenase,GAPDH)是细胞内的一个组成性表达的蛋白质,主要参与细胞的糖酵解途径。但近年来越来越多的研究表明,GAPDH不是一个纯粹简单的糖酵解酶。它是一种多功能蛋白,参与许多亚细胞水平活动,包括催化微管聚合、参与蛋白磷酸化修饰、参与膜融合和膜转运、促进细胞凋亡、调节蛋白质的表达、参与DNA损伤修复、作为转铁蛋白受体等。该文主要就GAPDH的结构特点及其功能的研究进展做一简要综述。
DOI:10.3724/SP.J.1260.2013.20175URLMagsci [本文引用: 1]
甘油醛-3-磷酸脱氢酶(glyceraldehyde-3-phosphate dehydrogenase,GAPDH)是细胞内的一个组成性表达的蛋白质,主要参与细胞的糖酵解途径。但近年来越来越多的研究表明,GAPDH不是一个纯粹简单的糖酵解酶。它是一种多功能蛋白,参与许多亚细胞水平活动,包括催化微管聚合、参与蛋白磷酸化修饰、参与膜融合和膜转运、促进细胞凋亡、调节蛋白质的表达、参与DNA损伤修复、作为转铁蛋白受体等。该文主要就GAPDH的结构特点及其功能的研究进展做一简要综述。
DOI:10.1002/biot.201400494URLPMID:4406140 [本文引用: 1]
Abstract Isothermal microcalorimetry is a label-free assay that allows monitoring of enzymatic and metabolic activities. The technique has strengths, but most instruments have a low throughput, which has limited their use for bioassays. Here, an isothermal microcalorimeter, equipped with a vessel holder similar to a 48-well plate, was used. The increased throughput of this microcalorimeter makes it valuable for biomedical and pharmaceutical applications. Our results show that the sensitivity of the instrument allows the detection of 3 × 104 bacteria per vial. Growth of P. mirabilis in Luria Broth medium was detected between 2 and 9 h with decreasing inoculum. The culture released 2.1J with a maximum thermal power of 76 μW. The growth rate calculated using calorimetric and spectrophotometric data were 0.60 and 0.57 h–1, respectively. Additional insight on protease activities of P. mirabilis matching the last peak in heat production could be gathered as well. Growth of tumor microtissues releasing a maximum thermal power of 2.1 μW was also monitored and corresponds to a diameter increase of the microtissues from ca. 100 to 428 μm. This opens new research avenues in cancer research, diagnostics, and development of new antitumor drugs. For parasitic worms, the technique allows assessment of parasite survival using motor and metabolic activities even with a single worm.
URL [本文引用: 1]
土壤微生物代谢活性与土壤功能直接相关,是评价土壤质量的灵敏的、普适的综合指标.物质的理化或生物变化过程都伴随着热效应,等温微量热技术可精确测定和跟踪此过程,其研究领域广范并带动土壤微生物的代谢热力学研究蓬勃发展.本文首先简要介绍微量热技术、普遍采用的底物诱导法揭示的微生物代谢活性和国内外研究团队.然后重点综述微量热技术在土壤微生物学的主要研究成果,包括代谢热力学可量化微生物对土壤有机碳的同化效率;土壤微生物在短时间内达到最大热释放值,伴随高的生长速率,表明其代谢活性高;代谢热熵（单位微生物热释放值）可指示微生物对土壤有机碳的转化和利用效率,是指示土壤质量的良好指标;外源污染物总体上抑制土壤微生物代谢活性;检测过程中应重视微量热测定的影响因素与条件优化.讨论测定过程中微生物生长条件的标准化和底物问题.最后展望潜在的研究方向,微量热技术与各种技术的有机结合,有望在土壤微生物结构与活性的地理属性、气候变化、土壤质量的空间大尺度比较和外源污染物的生物毒性评价等方面发挥其重要作用.
URL [本文引用: 1]
土壤微生物代谢活性与土壤功能直接相关,是评价土壤质量的灵敏的、普适的综合指标.物质的理化或生物变化过程都伴随着热效应,等温微量热技术可精确测定和跟踪此过程,其研究领域广范并带动土壤微生物的代谢热力学研究蓬勃发展.本文首先简要介绍微量热技术、普遍采用的底物诱导法揭示的微生物代谢活性和国内外研究团队.然后重点综述微量热技术在土壤微生物学的主要研究成果,包括代谢热力学可量化微生物对土壤有机碳的同化效率;土壤微生物在短时间内达到最大热释放值,伴随高的生长速率,表明其代谢活性高;代谢热熵（单位微生物热释放值）可指示微生物对土壤有机碳的转化和利用效率,是指示土壤质量的良好指标;外源污染物总体上抑制土壤微生物代谢活性;检测过程中应重视微量热测定的影响因素与条件优化.讨论测定过程中微生物生长条件的标准化和底物问题.最后展望潜在的研究方向,微量热技术与各种技术的有机结合,有望在土壤微生物结构与活性的地理属性、气候变化、土壤质量的空间大尺度比较和外源污染物的生物毒性评价等方面发挥其重要作用.
