Immobilization of urease using Amberlite MB-1

Amberlite MB-1 was used to immobilize urease (EC.3.5.1.5). The thermal stability of the immobilized urease was better than that of the free urease. Its highest activity was obtained at 75?°C and at pH 6.5 while the optimum temperature for the free urease was found to be 25?°C. Urease immobilized on Amberlite MB-1 retained 65% of the original activity after 5 repeated uses and 62% of the activity after 60 days when stored at 4?°C.     

Urease immobilized on nylon: preparation and properties

Urease (EC 3.5.1.5) was covalently attached through glutaraldehyde to partially hydrolysed nylon 6/6 tubes. The highest activity of immobilized enzyme was obtained at 65?°C and pH 6.5, while the optimum temperature for free urease was found to be 25?°C. Immobilized urease showed an improved thermal stability in comparison to free urease. It retained 76% of the original activity after 60 days when stored at 4?°C and 78% of the activity after 5 repeated uses.     

Immobilization of urease on vermiculite

Urease (EC 3.5.1.5) of high activity was obtained when the enzyme was immobilized on vermiculite crosslinked with 2.5% glutaraldehyde in chilled EDTA-phosphate buffer (pH 5.5). The highest activity of the immobilized enzyme was at 65°C and pH 6.5 while the optimum temperature for free urease was found to be 25°C. The thermal stability of immobilized urease was observed to be much better than that of the free urease. When stored at 4°C, urease immobilized on vermiculite retained 69 to 81% of its activity after 60 days and 61 to 75% of its original activity was retained after 4 repeated uses.     

Activity and stability of urease entrapped in thermosensitive poly(N-isopropylacrylamide-co-poly(ethyleneglycol)-methacrylate) hydrogel

Urease was entrapped in thermally responsive poly(N-isopropylacrylamide-co-poly(ethyleneglycol)-methacrylate), p[NIPAM-p(PEG)-MA], copolymer hydrogels. The copolymer membrane shows temperature-responsive properties similar to conventional p(NIPAM) hydrogels, which reversibly swell below and de-swell above the lower critical solution temperature of p(NIPAM) hydrogel at around 32 °C. The retained activities of the entrapped urease (in p[NIPAM-p(PEG)-MA]-4 hydrogels) were between 83 and 53 % compared to that of the same quantity of free enzyme. Due to the thermo-responsive character of the hydrogel matrix, the maximum activity was achieved at around 25 °C with the immobilized urease. Optimum pH was the same for both free and entrapped enzyme. Operational, thermal and storage stabilities of the enzyme were found to increase with entrapment of urease in the thermoresponsive hydrogel matrixes. As for reusability, the immobilized urease retained 89 % of its activity after ten repeated uses.     

Immobilization of watermelon (<Emphasis Type="Italic">Citrullus vulgaris</Emphasis>) urease in agarose gel for urea estimation

Urease from dehusked seeds of watermelon was immobilized in 1.5% agarose gel with 53.9% entrapment. There was negligible leaching (<10% at 4°C) and the same gel membrane could repeatedly be used for seven days. The immobilization exhibited no apparent change in the optimum pH but there was a significant decrease in the optimum temperature (50°C as compared to 65°C for soluble urease). The immobilized urease revealed an apparentK _m of 9.3±0.3 mM; 1.2 times lower than the soluble enzyme (11.4±0.2 mM). Unlike soluble enzyme which was inhibited at 200 mM urea, the immobilized urease was inhibited at 600 mM of urea and above, and about 47% activity was retained at 2 M urea. The time-dependent thermal inactivation kinetics at 48 and 52°C was found to be biphasic, in which half of the initial activity was destroyed more rapidly than the remaining half. These gel membranes were also used for estimating the urea content of the blood samples from the University hospital. The results obtained matched well with those obtained by the usual method employed in the clinical pathology laboratory. The significance of these observations is discussed.     

Assay of urease activity in marine sands — its use as an indicator of sewage contamination of beaches

Two methods were evaluated to determine urease activity in marine sands; in the first the amount of urea lost during the assay was determined, while in the second the amount of NH⁺₄-N formed was used as a measure of urea hydrolysis. Urease activity was detected and characterized in unpolluted coastal sands, and was found to be particularly high in sands colonized by higher plants. The pH and temperature optimum for the enzyme in sand was 6.2 and 55°C, respectively. Urease activity was very high in sands sampled close to an outfall releasing sewage out to sea, but decreased with increasing distance from the point of contamination. The possibility of using urease activity measurements as an indicator of sewage pollution on beaches is discussed.     

Changes in microclimate induced by experimental warming and clipping in tallgrass prairie

In order to facilitate interpretation and comparison of warming effects on ecosystems across various habitats, it is imperative to quantify changes in microclimate induced by warming facilities. This paper reports observed changes in air temperature, soil temperature and soil‐moisture content under experimental warming and clipping in a tallgrass prairie in the Great Plains, USA. We used a factorial design with warming as the primary factor nested with clipping as the secondary factor. Infrared heater was used in order to simulate climatic warming and clipping to mimic mowing for hay or grazing. The warming treatment significantly increased daily mean and minimum air temperatures by 1.1 and 2.3 °C, respectively, but had no effect on daily maximum air temperature, resulting in reduced diurnal air‐temperature range. Infrared heaters substantially increased daily maximum (2.5 and 3.5 °C), mean (2.0 and 2.6 °C) and minimum (1.8 and 2.1 °C) soil temperatures in both the unclipped and clipped subplots. Clipping also significantly increased daily maximum (3.4 and 4.3 °C) and mean (0.6 and 1.2 °C) soil temperatures, but decreased daily minimum soil temperature (1.0 and 0.6 °C in the control and warmed plots, respectively). Daily maximum, mean and minimum soil temperatures in the clipped, warmed subplots were 6.8, 3.2 and 1.1 °C higher than those in the unclipped, control subplots. Infrared heaters caused a reduction of 11.0% in soil moisture in the clipped subplots, but not in the unclipped subplots. Clipping reduced soil‐moisture content by 17.7 and 22.7% in the control and warmed plots, respectively. Experimental warming and clipping interacted to exacerbate soil‐moisture loss (26.7%). Overall, infrared heaters simulated climate warming well by enhancing downward infrared radiation and by reducing the diurnal air‐temperature range.     

Tracking Soil Temperature and Moisture in a Multi-Factor Climate Experiment in Temperate Grassland: Do Climate Manipulation Methods Produce their Intended Effects?

Passive open-top chambers (OTCs) and rainout shelters (RSs) have been used for over two decades to manipulate temperature and water availability in experiments on plant communities. These types of manipulations have been independently evaluated; however, as experiments become more complex and multiple factors are evaluated the potential for unknown or undesirable treatment effects increases. We present the effects of temperature manipulations (with OTCs), water manipulations (with RSs and water additions), and a clipping treatment, implemented in a fully factorial design, on soil moisture and temperature over 2 years in a temperate grassland. Temperature was increased 0.2°C by OTCs. Soil volumetric water content was reduced 3% by RSs and increased 2% by watering. However, clipping vegetation, treatment interactions, and weather conditions also affected soil temperature and moisture. For example, in OTCs RSs increased the temperature an additional 0.4°C, watering lowered it 0.4°C, and clipping raised temperature 2°C. Similarly, changes in soil moisture due to the RSs decreased VWC by 3% and increased 1% by clipping whereas soil moisture due to watering was reduced 1% by the OTCs and clipping. We also found that OTCs are more effective at raising temperatures on cooler days when soil temperatures are below 16.3°C. Our results suggest that all treatment types generally affect soil variables in predicable ways, but use of such devices should be adopted with caution, as they do not act independently, or exclusively, on the target variables.     

Effects of constant and fluctuating soil temperature on growth,development and nutrient uptake of maize seedlings

Summary The effect of constant and fluctuating soil temperature and two soil moisture regimes on the growth, development, transpiration and nutrient uptake by maize seedlings was studied in a greenhouse investigation. The constant root temperatures were maintained at 30, 34, 35, 36, 37, and 38°C for both 250 and 750 cm of soil moisture suctions. The fluctuating root temperature, for 250 cm of soil moisture suction only, of 30–35, 30–39, 30–40, 30–45 and 30–48°C were maintained to simulate the soil temperature regime under field conditions. The constant root temperature of 35°C and fluctuating temperature between 30–40°C significantly decreased the shoot and root growth and transpiration rate. On the average, there was 1.3 and 0.7 g decrease in fresh shoot weight and 0.36 and 0.30 g in fresh root weight per degree increase in root temperature for 250 and 750 soil moisture suction, respectively. In general, the effect of high soil moisture suction on maize seedlings was more severe when at high root temperature. The shoot and root concentration of N, P, and K decreased while that of B increased with increase in root temperature. The root concentration of Zn also decreased with increase in root temperature.     

高寒草甸土壤脲酶活性的研究

高寒草甸是青藏高原分布广、面积大的主要草场类型。我们于1984年6—10月在海北高寒草甸生态系统定位站,对四种植被类型土壤的脲酶活性进行了测定,数据列于表1。试验结果表明:(1)脲酶活性有明显的季节性动态,脲酶活性出现的高峰期均在7月和8月份,9月以后随温度的下降而逐渐降低;(2)脲酶活性也有明显的层次性差异,0—10厘米深土壤脲酶活性最高,并随土壤深度的加深而递减;(3)脲酶活性与某些氮素代谢微生物的数量有不同程度的相关性;(4)脲酶活性与某些氮素代谢微生物的生化活性也有一定的相关性;(5)脲酶活性与土壤温度具有一定的相关性;(6)脲酶活性与月降水量也相关,高寒草甸的不同植被类型土壤脲酶活性的季节性变化有所不同,并且与氮素代谢微生物的数量及活性的相关性也有差异,与土壤温度和降水量的相关程度也不一样。 关于土壤脲酶活性的研究,国外已有不少报道。在国内也有一些学者对土壤脲酶活性进行过研究。但对青藏高原高寒草甸土壤的脲酶活性的研究,目前尚未见报道,因为脲酶是氮循环的一种关键酶,与土壤肥力有关。本文对高寒草甸四种植被类型土壤的脲酶活性进行了初步的研究。     

Variations in soil culturable bacteria communities and biochemical characteristics in the Dongkemadi glacier forefield along a chronosequence

The variations in the soil culturable bacterial communities and biochemical parameters of early successional soils from a receding glacier in the Tanggula Mountain were investigated. We examined low organic carbon (C) and nitrogen (N) contents and enzymatic activity, correlated with fewer bacterial groups and numbers in the glacier forefield soils. The soil pH values decreased, but the soil water content, organic C and total N significantly increased, along the chronosequence. The soil C/N ratio decreased in the early development soils and increased in the late development soils and it did not correlate with the soil age since deglaciation. The activities of soil urease, sucrase, protease, polyphenol oxidase, catalase, and dehydrogenase increased along the chronosequence. The numbers of culturable bacteria in the soils increased as cultured at 25°C while decreased at 4°C from younger soils to older soils. Total numbers of culturable bacteria in the soils cultured at 25°C were significantly positively correlated to the soil total N, organic C, and soil water content, as well as the activities of soil urease, sucrase, dehydrogenase, catalase, and polyphenol oxidase. We have obtained 224 isolates from the glacier forefield soils. The isolates were clustered into 28 groups by amplified ribosomal DNA restriction analysis (ARDRA). Among them, 27 groups and 25 groups were obtained from the soils at 25°C and at 4°C incubation temperatures, respectively. These groups are affiliated with 18 genera that belong to six taxa, viz, Actinobacteria, Gammaproteobacteria, Bacteroidetes, Firmicutes, Alphaproteobacteria, and Betaproteobacteria. The dominant taxa were Actinobacteria, Gammaproteobacteria, and Bacteroidetes in all the samples. The abundance and the diversity of the genera isolated at 25°C incubation temperature were greater than that at 4°C.     

Factors influencing soil enzyme activity in China’s forest ecosystems

Enzyme activity (EA) mediates soil organic matter (SOM) degradation, transformation, and mineralization, thereby maintaining the biogeochemical cycles and energy flow of ecosystems. To determine the main factors explaining EA variations in China’s forest ecosystems, we created a database of soil EAs and relevant variables using data from the literature and analysed relationships between EAs and both climatic and edaphic variables. Catalase, phenol oxidase, acid (alkaline) phosphatase, and protease activities differed significantly among different types of forests. Catalase and urease activities were generally higher in primosols, cambisols, and argosols than in ferrosols. EA largely decreased with soil depth and increased with SOM. Phenol oxidase and urease activities were negatively correlated with mean annual temperature (MAT); in contrast, catalase, invertase, and protease activities first decreased (< 2.5 °C), increased (2.5–17.5 °C), and then decreased (> 17.5 °C) with increasing MAT. Although protease activity was slightly positively correlated with mean annual precipitation (MAP), catalase, phenol oxidase, and urease activities were all negatively related to MAP. Catalase, invertase, acid (alkaline) phosphatase, urease, and protease activities first increased (< 2000 m.a.s.l.) and then decreased (2000–4100 m.a.s.l.) with increasing elevation. Principal component analysis revealed most EAs to be correlated with climate conditions and soil pH. These findings suggest that climatic and edaphic variables directly and indirectly correlate with forest type and greatly impact soil EA.     

Immobilization of urease on vapour phase stain etched porous silicon

Porous silicon layers fabricated by the reaction-induced vapor phase stain etch method were coated with 5% polyethylenimine. Urease from Canavalia brasiliensis beans was immobilized on this support through covalent linking with 2.5% glutaraldehyde. The pH and temperature profile of the immobilized and free urease exhibited higher activity at pH 6.5 and 37 °C. After being stored for 30 days at 4 °C, the immobilized enzyme had 75% of the initial activity. The maximum apparent Michaelis constant for free urease (K_m) was 94.33 mM whereas for immobilized urease was 53.04 mM. The maximum reaction velocity (V_max) for free urease was 3.51 mmol/min and for immobilized urease was 1.57 mmol/min.     

黄土丘陵区子午岭不同植物群落下土壤氮素及相关酶活性的特征

以黄土丘陵区子午岭林区裸露地为对照,选择撂荒地、白羊草草地、油松、山杨和辽东栎林地五种典型植被群落下0—10cm和10—20 cm土层的土壤为研究对象,对土壤无机氮、有机氮、微生物量氮含量和脲酶、蛋白酶以及硝酸还原酶的活性进行了研究。结果表明,土壤中各种氮素基本表现为乔木林,尤其是辽东栎和油松下含量最高,而有机氮则在白羊草地富集明显。铵态氮为子午岭林区速效氮的主要形式。土壤铵态氮与微生物氮极显著正相关;有机氮和亚硝态氮、矿化氮、微生物氮均显著正相关。脲酶和硝酸还原酶活性在辽东栎群落下最高,蛋白酶在白羊草地下较高,且脲酶活性在土壤上层高于下层,而蛋白酶和硝酸还原酶并没有表现出明显规律。脲酶活性和铵态氮、有机氮含量显著正相关,与微生物量氮极显著正相关;硝酸还原酶活性与铵态氮含量显著正相关;蛋白酶活性和土壤各种氮素含量无相关性。     

Changes in Urease Activity in Apple Trees as Related to Urea Applications

Changes in urease (E.C.3.5.1.5.) were followed during the growth of 1-year-old MM 106 and 9-year-old Golden Delicious apple trees (Malus pumila Rehd.). Urease was found in leaves, roots, and bark with actively growing tissues containing more activity than senescing tissues. The urease activity in the leaves declined steadily during leaf senescence but abscised leaves still contained about half of their initial urease activity. In the bark the urease activity changed only slightly. Urease activities in the leaves and bark of apple trees were always greater in those trees which had received an application of urea. In senescing apple leaves, urea induced a rapid increase in urease activity. The changes in total activity and specific activity of urease were parallel and suggests that urease was synthesized de novo. After urease activity reached a maximum, a rapid decline occurred. Urease was inhibited by low concentrations of ammonia and this decline may be due to product inhibition.     

Rapid laboratory measurement of the temperature dependence of soil respiration and application to changes in three diverse soils through the year

Determining the temperature dependence of soil respiration is needed to test predictive models such as Arrhenius-like functions and macro-molecular rate theory (MMRT). We tested a method for rapid measurement of respiration using a temperature gradient block, cooled at one end (~2 °C) and heated at the other (~50 °C) that accommodated 44 tubes containing soil incubated at roughly 1 °C increments. Gas samples were taken after 5 h incubation and analysed for CO₂. The temperature gradient block allowed rapid assessment of temperature dependence of soil respiration with the precision needed to test models and explore existing theories of how temperature and moisture interact to control biochemical processes. Temperature response curves were well fitted by MMRT and allowed calculation of the temperature at which absolute temperature sensitivity was maximal (T_inf). We measured temperature response of three soils at seven moisture contents and showed that the absolute rate and sensitivity of respiration was partly dependent on adjusted moisture content. This result implied that comparisons between soils need to be made at a common moisture content. We also measured potential changes in the temperature dependence (and sensitivity) of respiration for three different soils collected at one site throughout a year. T_inf ranged from 43 to 51 °C for the three soils. T_inf and temperature sensitivity were not dependent on soil type collected but was partly dependent on time of year of collection. Temporal changes in temperature response suggested that the microbial communities may tune their metabolisms in response to changes in soil temperatures.     

水肥处理对黄瓜土壤养分、酶及微生物多样性的影响

以津优1号黄瓜为试材,设3个土壤相对含水量水平（50%～60%、70%～80%、90%～100%）和2个肥料追施量（600 kg N·hm^-2和420 kg P₂O₅·hm^-2,420 kg N·hm^-2和294 kg P₂O₅·hm^-2）处理,研究了不同水肥供应对日光温室黄瓜土壤养分、酶活性及微生物多样性的影响.结果表明：土壤中NH₄⁺-N含量随施肥量的增加而提高,随土壤相对含水量的增加而降低;水肥供给的增加有利于提高土壤中速效磷含量和蔗糖酶活性;肥料增加使土壤中蛋白酶活性降低,而水分降低使土壤中脲酶活性提高.土壤中微生物多样性与土壤中养分含量无显著相关性,与土壤脲酶活性呈显著正相关,与蔗糖酶活性呈显著负相关.土壤相对含水量70%～80%、氮肥追施量600 kg N·hm^-2和420 kg P₂O₅·hm^-2处理的土壤养分含量、蔗糖酶、磷酸酶和脲酶活性较高,且土壤中微生物多样性和均匀度显著高于其他处理,土壤生产潜力最优.     

Carbon exchange in biological soil crust communities under differential temperatures and soil water contents: implications for global change

Biological soil crusts (biocrusts) are an integral part of the soil system in arid regions worldwide, stabilizing soil surfaces, aiding vascular plant establishment, and are significant sources of ecosystem nitrogen and carbon. Hydration and temperature primarily control ecosystem CO₂ flux in these systems. Using constructed mesocosms for incubations under controlled laboratory conditions, we examined the effect of temperature (5–35 °C) and water content (WC, 20–100%) on CO₂ exchange in light (cyanobacterially dominated) and dark (cyanobacteria/lichen and moss dominated) biocrusts of the cool Colorado Plateau Desert in Utah and the hot Chihuahuan Desert in New Mexico. In light crusts from both Utah and New Mexico, net photosynthesis was highest at temperatures >30 °C. Net photosynthesis in light crusts from Utah was relatively insensitive to changes in soil moisture. In contrast, light crusts from New Mexico tended to exhibit higher rates of net photosynthesis at higher soil moisture. Dark crusts originating from both sites exhibited the greatest net photosynthesis at intermediate soil water content (40–60%). Declines in net photosynthesis were observed in dark crusts with crusts from Utah showing declines at temperatures >25 °C and those originating from New Mexico showing declines at temperatures >35 °C. Maximum net photosynthesis in all crust types from all locations were strongly influenced by offsets in the optimal temperature and water content for gross photosynthesis compared with dark respiration. Gross photosynthesis tended to be maximized at some intermediate value of temperature and water content and dark respiration tended to increase linearly. The results of this study suggest biocrusts are capable of CO₂ exchange under a wide range of conditions. However, significant changes in the magnitude of this exchange should be expected for the temperature and precipitation changes suggested by current climate models.     

碳添加下黑钙土胞内、胞外脲酶活性变化及其机制

土壤脲酶作为能够催化尿素水解的最重要酶类,对草地生态系统氮素供应具有重要作用。目前探讨不同碳添加对草地土壤胞外脲酶影响的研究报道相对较多,但碳添加对土壤胞内脲酶的影响,以及胞内和胞外脲酶对碳添加的响应是否一致等尚需深入研究。本研究依托额尔古纳森林草原过渡带生态系统研究站开展的碳添加野外试验平台(以葡萄糖为碳源),选取无碳添加(C₀)、250(C₂₅₀)和500(C₅₀₀) kg C·hm^-2·a^-1处理为供试对象,探讨碳添加下黑钙土胞内、胞外脲酶活性响应及其与土壤性质的关系。结果表明: 碳添加显著提高了土壤胞内脲酶活性,增加了土壤胞内脲酶活性占总脲酶活性的比例,但对土壤胞外脲酶活性没有显著影响。土壤胞内脲酶活性与微生物生物量具有显著正相关关系,表明胞内脲酶活性增加主要是由微生物生物量增加引起的。结构方程模型(SEM)分析表明,碳添加通过影响土壤微生物生物量间接提高了土壤胞内脲酶活性。     

Effect of Environment on Wild Oat (Avena fatua) Control with Imazamethabenz or Fenoxaprop Tank-Mixed with Additives or MCPA

Greenhouse and growth chamber experiments were conducted to determine the effect of soil moisture and temperature on the phytotoxicity in wild oat of imazamethabenz or fenoxaprop tank-mixed with certain additives or MCPA. The surfactants Agral 90 at 0.5% and Enhance at 0.5% increased imazamethabenz phytotoxicity under both moist and drought conditions. These surfactants had no significant effect on fenoxaprop phytotoxicity regardless of the soil moisture regimes. Fenoxaprop activity was increased by ammonium sulfate [(NH₄)₂SO₄] at 1% but only under well watered conditions. Wild oat control with imazamethabenz was also slightly enhanced in a well watered regime by the addition of sodium bisulfate (NaHSO₄) at 0.13%. At high temperature (30/20°C) and low temperature (10/5°C), the phytotoxicity of imazamethabenz was increased when tank-mixed with Agral 90 at 0.25% or NaHSO₄ at 0.13% compared with that when imazamethabenz was applied alone, if soil moisture was adequate. There was no such increase under conditions of drought and high temperature. (NH₄)₂SO₄ at 1% did not significantly affect imazamethabenz performance irrespective of temperature/soil moisture conditions. The phytotoxicity to wild oat of imazamethabenz or fenoxaprop was not changed by tank-mixing with MCPA isooctyl ester at 300 g a.i./ha, regardless of soil moisture levels. The reduced fenoxaprop phytotoxicity in wild oat due to moisture stress was not readily alleviated by the inclusion of selected additives or MCPA in the tank mixture. Received May 10, 1996; accepted January 10, 1997