基于不同基质测定土壤碱性磷酸酶活性的比较

土壤磷酸酶对有机磷的矿化及植物的磷素营养有重要影响。目前测定土壤磷酸酶活性采用的基质主要有磷酸苯二钠(CAS编号:3279-54-7)和对硝基苯磷酸二钠(CAS编号:4264-83-9)两种。国际上对土壤磷酸酶活性的测定普遍采用对硝基苯磷酸二钠,而国内大部分学者主要采用磷酸苯二钠。由于不同类型的基质对磷酸酶活性的测定结果有很大影响,所以选择适宜的基质种类对磷酸酶活性进行测定有很大的不确定性。为探讨碱性磷酸酶活性的测定方法,以磷酸苯二钠和对硝基苯磷酸二钠为基质,选用酸性、中性和碱性土壤各10个土样,比较了依据《土壤酶及其研究法》(DPP1),《Method in Soil Biology》(DPP2)和《Methods of Soil Enzymology》(PNPP)书中所介绍的测定方法所获结果之间的差异。结果表明:三种方法所测的三种类型土壤磷酸酶活性的变化趋势基本一致,但不同土壤之间差异性的变化幅度有明显不同。在酸性土壤和碱性土壤中,DPP1和DPP2所测十个土样碱性磷酸酶活性之间的变异系数属于中度变异,而PNPP则属于高度变异;在中性土壤中,DPP1和DPP2属于低度变异,PNPP属于中度变异。总的来看,PNPP比DPP1和DPP2敏感。此外,由于PNPP具有高精确度、培养时间短和实验操作简便等优点,我们建议采用PNPP进行碱性磷酸酶活性的测定。     

缓冲液种类对土壤酸性磷酸酶活性的影响

土壤酸性磷酸酶影响土壤有机磷矿化。选择不同的缓冲液种类和不同p H值测定土壤酸性磷酸酶活性,可影响数据的可比性。为明确缓冲液及p H值对不同土壤磷酸酶活性的影响程度,选取p H值4～8的土壤样本,测定了在醋酸盐缓冲液、柠檬酸盐缓冲液和MUB缓冲液3种不同培养条件下酸性磷酸酶活性变化。结果表明,采用MUB缓冲液条件可获得较稳定的土样区分序列,且与缓冲液的p H值的变化无关。在醋酸盐缓冲液和柠檬酸盐缓冲液条件下,不同p H值范围的不同土样之间土壤酸性磷酸酶活性变异系数变化剧烈,而采用MUB缓冲液获得的变异系数较其他缓冲液大,且趋势平稳。综合考虑,进行土壤酸性磷酸酶活性测定,可优先选用MUB缓冲溶液。     

中药材龙胆GAP基地土壤酶活性的研究

分别采用硫代硫酸钠滴定法、高锰酸钾滴定法、TTC-脱氢酶活性测定法和磷酸苯二钠比色法测定龙胆GAP基地土壤中转化酶、过氧化氢酶、脱氢酶和磷酸酶的活性。对不同月份、种植不同年生龙胆根际土及非根际土中酶活性的测定结果进行分析表明:土壤过氧化氢酶活性在6月份出现一较高值,脱氢酶活性较高时期在7月份,土壤转化酶活性较大值出现在7、8月份,磷酸酶活性则逐月增高至9月份达到最大值,土壤总体酶活指标极大值出现在8月份;随着龙胆种植年限的增加,土壤过氧化氢酶和脱氢酶活性有所提高,而磷酸酶和转化酶活性没有明显变化,种植不同年生龙胆土壤中酶的活性随季节波动不尽相同。比较GAP基地龙胆根际土与非根际土中酶活性,土壤转化酶和过氧化氢酶的活性均表现为根际土非根际土,而土壤磷酸酶和脱氢酶的活性则为非根际土根际土;相关分析结果显示,过氧化氢酶活性与酸性及中性磷酸酶活性显著负相关;酸性、中性及碱性三种磷酸酶活性之间存在极显著正相关;总体酶活指标与转化酶、酸性磷酸酶、中性磷酸酶活性呈显著正相关。本研究完善了中药材龙胆GAP种植基地土壤酶的信息,为综合评价基地土壤肥力提供了科学数据,为药用植物龙胆的合理施肥、品质的提高提供了科学依据。     

丛枝菌根对三叶草根际磷酸酶活性的影响

以三叶草为材料,利用三室隔网培养方法,探讨了取自肥料长期定位试验中多年施用与不施用有机肥的田间小区土壤上,接种菌根菌（G.mosseae）对根际土壤酸性和碱性磷酸酶活性的影响。植物生长9周后,收获测定菌丝生长室土壤酸性磷酸酶和碱性磷酸酶活性,并对磷酸酶产生位点进行细胞化学定位。结果表明,接种丛技菌根菌对根际土壤酸性和碱性磷酸酶活性均有增强作用,但作用程度在有机服小区土壤上要大于无机肥小区土壤。根际土壤酸性磷酸酶原位化学定位结果表明,菌丝周围有明显的酸性磷酸酶的反应产物,说明报外丛枝菌根菌丝能直接向外分泌磷酸酶。     

丛枝菌根真菌群落对白三叶草植株生物量磷吸收和土壤磷酸单酯酶活性的影响

丛枝菌根真菌（AMF）在土壤与植物的磷素循环中发挥着关键的作用。采用盆栽实验研究了丛枝菌根真菌群落对白三叶草植株生物量、磷吸收和土壤磷酸单酯酶活性的影响。结果表明,接种不同AMF群落均能显著地促进白三叶草植株的生长及其对磷素的吸收,提高根际土壤磷酸单酯酶的活性。Mnp处理中,白三叶草生物量最大,白三叶草总生物量、茎叶生物量和根系生物量分别比对照处理（-M）提高64.48%、61.48%和84.91%。不同菌根处理中,Mck处理显著地提高白三叶草磷吸收和土壤磷酸单酯酶活性,白三叶草磷吸收总量和茎叶磷吸收量分别比对照（-M）提高107.18%和91.91%,土壤碱性磷酸单酯酶和酸性磷酸单酯酶活性相对对照（-M）分别提高54.33%和138.43%。碱性磷酸单酯酶活性与AMF群落中的Acaullospora属孢子数呈显著的正相关关系,而酸性磷酸单酯酶活性则主要受Paraglomus属孢子数的影响。说明接种AMF群落可显著地影响土壤的磷酸单酯酶活性,从而影响白三叶草的生长及其对磷素的吸收。     

长期施肥对黄壤稻田和旱地土壤磷酸酶活性的影响

  目的  明确施肥对黄壤稻田和旱地磷酸酶活性的影响及其主要影响因子。  方法  依托进行了22年的黄壤长期定位试验,研究长期施用化肥和有机肥对稻田和旱地土壤养分及酸性、中性和碱性磷酸酶活性的影响。  结果  长期不同施肥处理显著改变了稻田和旱地土壤养分含量,且土壤pH、有机质和有效磷含量变化因稻田、旱地不同所受影响差异较大。稻田和旱地土壤酸性磷酸酶（ACP）活性和中性磷酸酶（NEP）活性均高于碱性磷酸酶（ALP）活性。与不施肥处理相比,稻田上除个别处理外,施用化肥对磷酸酶活性无显著影响,施用有机肥处理NEP活性显著提高,增幅达15.8% ~ 27.1%;旱地上,长期施用氮肥的处理（NK、NP、NPK）ACP活性显著提高了14.2% ~ 29.0%,ALP活性显著降低了20.1% ~ 50.7%,施用有机肥处理ACP和ALP活性分别提高了11.7% ~ 17.7%和9.4% ~ 56.9%,NEP活性降低了10.5% ~ 32.3%。与平衡施肥处理相比,稻田上缺素施肥对磷酸酶活性影响不显著,施用有机肥处理ACP、NEP、ALP活性分别提高了9.1% ~ 18.5%、6.9% ~ 17.3% 和8.3% ~ 15.0%;旱地上, NK处理ACP和NEP分别显著提高了12.9%和12.9%,PK处理则分别显著降低了16.7%和18.9%,施用有机肥处理NEP活性降低了5.9% ~ 28.9%,ALP活性显著提高了50.1% ~ 115.3%。相关分析表明,NEP活性与各土壤养分含量显著相关,但在稻田和旱地上相关性相反;ALP活性在旱地和稻田上与有机质和微生物量磷均呈显著正相关。通径分析表明,除旱地上ALP活性受pH直接影响作用较大外,其他磷酸酶活性受土壤磷素的直接影响作用均较大。  结论  黄壤稻田和旱地土壤磷酸酶活性与土壤磷素有效性密切相关,稻田上施用有机肥是提高土壤磷酸酶活性的有效途径,旱地上长期施用化学氮肥尤其是氮钾处理可提高ACP活性,增加土壤磷素供应,施用有机肥可通过提高土壤pH增加ALP活性。     

根际pH对玉米利用磷酸单酯和双酯盐的影响

【目的】土壤有机磷在土壤全磷中占有很大比重,是植物潜在的有效磷源,但必须通过磷酸酶的水解作用释放出无机磷才能被植物利用。土壤中有机磷的主要形式为磷酸单酯和磷酸双酯。本研究中,我们探讨了无菌条件下不同形态的氮源引起的根际pH变化如何影响植物对这两种有机磷的活化利用。【方法】采用琼脂无菌培养体系种植玉米,向玉米植株供应两种形态的氮源和磷源, 氮源为硝态氮和铵态氮,磷源为植酸钙和卵磷脂,植酸钙属于磷酸单酯盐,卵磷脂属于磷酸双酯盐。不同的供氮形态会导致根际pH变化,进而研究根际pH变化对磷酸单酯盐和磷酸双酯盐的活化利用所产生的影响。【结果】当给玉米供应铵态氮时,根际pH从5.5降至4.0; 供应硝态氮时,根际pH升至6.6。测定玉米根际的琼脂中根系分泌的磷酸单酯酶和磷酸双酯酶活性发现,磷酸单酯酶活性在pH 6.0~7.0之间最高,磷酸双酯酶活性在pH 7.0~8.0之间达到最高。无论以植酸钙还是卵磷脂为有机磷源,相对于铵态氮处理,硝态氮处理能够使根际保持较高的磷酸单酯酶或磷酸双酯酶活性。有机磷的水解过程由磷酸酶活性和底物有效性两个因素控制,而植酸钙的水解受根际pH影响很大,在一定pH范围内,植酸钙的溶解度随根际pH值降低而升高,有效态磷浓度的增加,使得磷酸酶的底物有效性提高。在供应铵态氮时,根际pH值降低,玉米对植酸钙的利用效率高于硝态氮处理,尽管供硝态氮时磷酸单酯酶活性更高。同时,在供应铵态氮条件下,植株对植酸钙的利用率要显著高于卵磷脂,原因在于虽然磷酸双酯酶和磷酸单酯酶活性较低,由于植酸钙的溶解度较大,它的底物有效性更高。因此,植酸钙处理中植株的磷含量更高。相反,在供应硝态氮时,植酸钙溶解度减小而两种磷酸酶活性较高,卵磷脂处理中植株的磷含量更高。【结论】土壤中有机磷的水解过程由磷酸酶活性和有机磷底物有效性两个因素控制,酶活性与根际pH密切相关。本研究说明土壤有机磷的活化必须首先转化为溶解于水溶液中的状态,才能作为磷酸酶的底物被催化水解。我国长期施用化肥导致北方土壤大范围酸化,这种酸化无疑对土壤固有或随有机物料进入农田的有机磷的活化利用是具有重要贡献的,应该在北方土壤养分管理中应加以考虑。     

外源植酸酶对土壤磷酸酶活性和有效磷含量的影响

采用室内培养的方法,研究了不同用量外源植酸酶在不同培养时期对土壤磷酸酶活性和有效磷含量的影响.结果表明,添加外源植酸酶各处理的土壤酸性、中性磷酸酶活性和有效磷含量均高于对照,各处理间呈高量＞中量＞低量＞对照,其中土壤酸性、中性磷酸酶活性与对照相比增加的幅度分别为9.3%～70.2%、4.3%～73.1%,差异达显著水平;从培养时间来看,土壤磷酸酶活性和有效磷含量均随培养时间的延长呈先升高后略有回落的趋势,其中酸性磷酸酶活性至培养第10 d达最高值,此时各处理与对照相比提高了33.9%～70.2%,差异达极显著水平,而中性磷酸酶活性和有效磷含量至培养的第20 d达到最高值,此时与对照相比分别提高了49.9%～73.1%和26.6%～122.5%.     

用通径分析法研究土壤水解酶活性与土壤性质的关系

为正确评价土壤性质对酶活性的影响程度 ,运用通径分析方法研究了浙江 5种土壤 1 2个样品中的土壤脲酶、转化酶、酸性磷酸酶活性与土壤性质的关系。结果表明 :土壤有机质、全氮、全磷是制约脲酶、转化酶和酸性磷酸酶活性的主要因素 ,而阳离子交换量对三种酶活性的影响均很微弱 ;就通径分析的直接效应而言 ,pH对脲酶活性的影响最为显著 ,对转化酶和酸性磷酸酶活性的影响程度大致相当。然而pH对脲酶、转化酶、酸性磷酸酶的这种直接效应在很大程度上被通过其它因素对三种酶的间接效应所抵消 ;土壤粉粒对转化酶活性的作用尤为显著。此外 ,粘粒对酸性磷酸酶、砂粒对脲酶有一定影响且均主要体现为直接效应。     

八达岭山地次生林土壤养分与磷酸酶活性研究

 为探讨北京八达岭山地土壤肥力状况,以黑桦、糠椴、核桃楸3种次生林为对象,对其土壤养分和磷酸酶活性进行研究。结果表明:土壤养分有明显的表聚效应;土壤全磷含量仅为0.39～0.66g.kg-1之间,有可能出现缺磷现象;土壤磷酸酶的活性以酸性磷酸酶为主,呈垂直递减分布规律;各种土壤养分和磷酸酶活性均有明显的根际效应,根系/本土比值在1.02～5.25之间。用SPSS软件进行分析表明,土壤磷酸酶活性与土壤pH、有机质、全氮、有效氮、有效磷和速效钾显著正相关,可以作为土壤肥力评价的一个指标。     

Phosphatases in soils

Most studies on phosphatase activity in soils have been concerned with acid phosphatase. This study was conducted to determine the activity of phosphomonoesterases (acid and alkaline phosphatases), phosphodiesterase, and “phosphotriesterase”. The results indicate that acid phosphatase is predominant in acid soils and that alkaline phosphatase is predominant in alkaline soils. With universal buffer, the pH optima of phosphodiesterase and phosphotriesterase were at pH 10. The activities of these phosphatases in soils were much lower than those of the acid and alkaline phosphatases. Studies on the effects of various soil treatments on the activity of phosphatases in soils indicated that air-drying increased the activity of acid phosphatase and phosphotriesterase, decreased the activity of alkaline phosphatase, but did not affect the activity of phosphodiesterase. Steam sterilization of soils at 121 C for 1 h inactivated alkaline phosphatase, phosphodiesterase, and phosphotriesterase, but did not completely inactivate acid phosphatase. Addition of toluene to the incubation mixture did not markedly affect the activity of acid phosphatase, alkaline phosphatase, phosphodiesterase, but increased the activity of phosphotriesterase in soils.Studies of the kinetic parameters of phosphatases in the soils studied showed that the K_m values ranged from 1.11 to 3.40 mm for acid phosphatase. from 0.44 to 4.94 mm for alkaline phosphatase, and from 0.25 to 1.25 mm for phosphodiesterase. Expressed as μg p-nitrophenol released·h^?1·g^?1 soil, the V_max values ranged from 200 to 625 for acid phosphatase, from 124 to 588 for alkaline phosphatase, and from 46 to 127 for phosphodiesterase. The substrate of phosphotriesterase (tris-p-nitrophenyl phosphate) is insoluble in water; hence, the K_m and V_max values of this enzyme in soils could not be determined.     

不同AM菌根菌分泌的磷酸酶对根际土壤有机磷的影响

以三叶草为材料,利用3室隔网培养方法,研究了4种AM菌根菌侵染三叶草后对根际土壤酸性和碱性磷酸酶活性以及菌丝酶活性对土壤有机P的影响。结果表明,接种AM菌根菌 (9周) 对根际土壤酸性和碱性磷酸酶活性均有增强作用,但作用强度主要取决于菌丝在土壤中的生长状况,Glomus属菌根菌在整个菌丝室 (0~6cm) 都影响土壤磷酸酶的活性,其活性在整个菌丝室中都比Gigaspora的高。同一属不同种的根际土壤磷酸酶活性差异不大。AM菌根根际土壤磷酸酶对土壤有机P的降解有很强的促进作用。     

Phosphomonoesterase,phosphodiesterase, phosphotriesterase,and inorganic pyrophosphatase activities in forest soils in an alpine area: effect of pH on enzyme activity and extractability

We investigated different types of phosphatase activity (phosphomono-, phosphodi-, phosphotriesterase, inorganic pyrophosphatase) in five forest soils in Vorarlberg, Austria. Phosphatase activity was determined both in soils and in soil extracts prepared with different solutions (distilled water, 0.1M sodium pyrophosphate at pH 7, 0.1M sodium phosphate buffer/1M KCl at pH 6.5, and a modified universal buffer at pH 4, 6.5, 9, and 11). High phosphomonoesterase activity in these soils indicated a severe deficiency in available P. Acidic phosphomonoesterase prevailed over alkaline phosphomonoesterase activity. Phosphodiesterase was highest in the least acidic soil but no general trend towards an optimum pH was recognized. Phosphotriesterase activity was observed in only two of the five soils and favoured an alkaline optimum pH; this activity was not detected in strongly acid soils. Inorganic pyrophosphatase activity was high in soils with no phosphotriesterase. Phosphomonoesterase, phosphodiesterase and inorganic pyrophosphatase activities were much lower in soil extracts than in soils.     

Response of soil enzymes and microbial communities to root extracts of the alien Alternanthera philoxeroides

Alternanthera philoxeroides is an alien invasive species, which can cause substantial damage to the local ecosystem by secreting allelochemicals. The objective of the present study was to appraise the effects of root extracts of the invasive plant Alternanthera philoxeroides on soil enzyme and native microbial community. Urease activity was measured using the phenol sodium hypochlorite colorimetric method, sucrase activity was determined by 3, 5-dinitrosalicylic acid colorimetric method, catalase activity was determined using the KMnO₄ titration method and alkaline phosphatase activity was determined by the disodium phenyl phosphate colorimetric method. The soil microbial community was investigated by pyrosequencing of the 16S rRNA gene. Results showed that the alkaline phosphatase, sucrase and urease activities were all significantly lower than that of the control treatment. The structure of the soil microbial community in the treatments with root extracts addition clearly differed from that of the control treatment. The result showed that the relative abundance of microorganism decreased including Burkhholderia, Gp6, Gp1 and Gp4 in the applied treatments at genus level as compared to control treatment. Hence, we inferred that A. philoxeroides could inhibit the growth of native plant species through toxic effects on soil enzyme activities and the microbial community.     

Changes in soil biological activities under reduced soil pH during Thlaspi caerulescens phytoextraction

Phytoextraction of soil Cd and Zn may require reduction in soil pH in order to achieve high metal uptake. Reducing the pH of high metal soil, however, could negatively affect soil ecosystem function and health. The objectives of this study were to characterize the quantitative causal relationship between pH and soil biological activities in two Zn and Cd contaminated soils and to investigate the relationship between metals and soil biological activities under low pH. Soils were adjusted to five or six different pH levels by sulfur addition, followed by salt leaching. Thlaspi caerulescens was grown for 6 months, and both the rhizosphere and non-rhizosphere soil biological activities were tested after harvest. Reducing pH significantly lowered soil alkaline phosphatase activity, arylsulphatase activity, nitrification potential, and respiration. However, acid phosphatase activity was increased with decreasing pH. The relationship between soil biological activities and pH was well characterized by linear or quadratic regression models with R² values ranging from 0.57 to 0.99. In general, the three enzyme activities, nitrification potential, and the ratio of alkaline phosphatase to acid phosphatase activity were very sensitive indicators of soil pH status while soil respiration was not sensitive to pH change. The rhizosphere soil had higher biological activities than non-rhizosphere soil. The negative effects observed in the non-rhizosphere soil were alleviated by the rhizosphere influence. However, rhizosphere soil after 6 months phytoextraction showed lower nitrification potential than non-rhizosphere soil, probably due to substrate limitation in our study.     

Does salinity enhance Cd toxicity to soil alkaline phosphatase?

The aim of this study was to provide data to assess the additive effects of soil salinity on the toxicity of Cd to soil alkaline phosphatase (EC 3.1.3.1). Two soils (Langroud acid soil and Shervedan calcareous soil) were artificially salinized with NaCl. The natural and salinized soils were treated with CdSO₄ solutions to give a Cd concentration in the range 3–5000 mg kg^?1. Soil alkaline phosphatase activity was measured after 3 days of incubation. Salinity enhanced the extractable Cd concentration in both Langroud and Shervedan soils. The percentage of soil alkaline phosphatase activity inhibited by Cd was significantly increased from 27.8 to 45 in the Langroud acid soil as salinity increased from natural levels to 28 dS m^?1. An increase in the inhibition percentage was not observed in the Shervedan soil. Lower values for the ecological dose causing 50% inhibition (ED₅₀) under saline conditions in the Shervedan soil supported the hypothesis that Cd may be more toxic to soil alkaline phosphatase when the soil is more saline. We conclude that Cd toxicity to soil alkaline phosphatase is salinity dependent and that higher Cd concentrations under saline conditions are probably responsible for the enhanced Cd toxicity to soil alkaline phosphatase.     

不同母质类型水稻土酶活性及其与理化性质的关系

将选自湖南双季稻区6种典型成土母质发育的水稻土置于长沙市郊同一生态条件下,通过7年定位池栽试验,研究了不同母质类型对土壤酶活性的影响以及酶活性与土壤理化性质之间的关系。结果表明：不同母质类型对土壤蔗糖酶活性的影响达到显著水平(P<0.05),对酸性磷酸酶、脲酶及过氧化氢酶活性的影响达到极显著水平(P<0.01)。主成分分析表明不同母质发育的水稻土酸性磷酸酶及脲酶作为综合评价土壤酶活性的指标要优于蔗糖酶及过氧化氢酶。6种不同母质水稻土土壤酶综合得分排序为：板页岩>河流沉积物>花岗岩>第四纪红土>紫色页岩>石灰岩。相关性分析表明,土壤蔗糖酶活性与有机质、全氮含量呈显著的正相关关系(P<0.05)。脲酶活性与硝态氮、铵态氮含量呈显著的正相关关系(P<0.05),与pH值呈显著的负相关关系(P<0.05)。酸性磷酸酶活性与速效磷含量呈显著正相关关系(P<0.05),与pH值呈显著负相关关系(P<0.05)。     

Response of phosphomonoesterases in soils to chloroform fumigation

The chloroform fumigation technique has been successfully employed to quantify intracellular and extracellular urease and arylsulfatase activities in soil. In this study, the same approach was evaluated for its ability to differentiate between various pools of phosphomonoesterase activities in soils and reference proteins purified from plant and microbial sources. The activities of acid and alkaline phosphatases were assayed in 10 surface soils and reference proteins at their optimal pH values before and after chloroform fumigation and in the presence and absence of toluene. Chloroform fumigation decreased the activities of acid and alkaline phosphatases in soils, on average, by 6 and 8%, respectively. Similarly, the activities of two purified reference enzyme proteins were decreased after fumigation, with acid and alkaline phosphatase activities exhibiting a reduction of 17 and 8%, respectively. Toluene treatment caused an increase in the activities of acid and alkaline phosphatases by 8 to 18% in nonfumigated soils, but showed no effect in the fumigated soils. Average enzyme protein concentrations, calculated for the 10 soils based on the activity values of the soils and the specific activity of the purified enzymes (i.e., activity values per mg protein), were 22.5 and 2.1 mg protein (kg soil)^—1 for acid and alkaline phosphatase, respectively. The decrease in enzyme activity by the fumigant was either by direct denaturing of the periplasmic and extracellular portion of the particular protein after lysis of the microbial cell membrane, by absorption and/or inhibition of the released phosphomonoesterases by organic and inorganic constituents or by degradation of the protein by soil proteases. The ratios of acid phosphatase protein concentrations relative to organic C in six soils were significantly, but negatively correlated with soil organic C, suggesting differences in organic C quality. Comparison of the activity values of soil phosphatases with those of the protein concentrations present in soils indicated that alkaline phosphatase has greater catalytic efficiency than does acid phosphatase.