Hydrolysis of cellobiose by β-glucosidase in the presence of soil minerals - Interactions at solid-liquid interfaces and effects on enzyme activity levels

Extracellular enzymatic activities in soils are essential for the cycling of organic matter. These activities take place in multiphase environments where solid phases profoundly affect biocatalytic activities. Aspergillus niger is ubiquitous in soils; its β-glucosidase plays an important role in the degradation of cellulose, and therefore in the global carbon cycle and in the turnover of soil organic matter. However, the information on the interactions of this protein with soil minerals is very limited, and even less is known about their consequences for the hydrolysis of the natural substrate cellobiose. We therefore characterised the sorptive interactions of this enzyme with the soil minerals montmorillonite, kaolinite and goethite and quantified the resulting changes in the hydrolysis rate of cellobiose. Fractions of adsorbed protein, and the resulting catalytic activity loss, were lower for montmorillonite than for kaolinite and goethite at given experimental conditions; adsorption was 9.7 ± 7.3% for montmorillonite, 70.3 ± 3.1% for kaolinite and 71.4 ± 1.8% for goethite, respectively. Adsorption of the protein to the minerals caused a total decrease in the catalytic activity of 18.8 ± 3.4% for kaolinite and 17.9 ± 4.7% for goethite whereas it was not significant for montmorillonite. The average catalytic activity lost by the pool of adsorbed molecules was 26.8% for kaolinite and 25.0% for goethite. Both the amount of adsorbed protein and the resulting loss of catalytic activity were found to be independent of the specific surface areas yet were influenced by the electrical properties of the mineral surfaces. Under the experimental conditions, montmorillonite and kaolinite are negatively charged whereas goethite is positively charged. However, because of the adsorption of phosphate anions from the buffer, a charge reversal took place at the surface of goethite. This was confirmed by zeta (ζ)-potential measurements in phosphate buffer, revealing negative values for all the tested minerals. Indeed goethite interacted with the enzyme as a negatively charged surface: the amount of adsorbed protein and the resulting catalytic activity loss were very similar to those of kaolinite. Our results show that, even if an important fraction of β-glucosidase is adsorbed to the minerals, the catalytic activity is largely retained. We suggest that this strong activity retention in presence of soil minerals results from a selective pressure on A. niger, which benefits from the activity of the adsorbed, and thus stabilized, enzyme pool.     

供铁水平对不同苹果基因型生长及根系特性的影响

以小金海棠 (MalusxiaojinensisChengetJiang)和山定子 (MalusbaccataL .Borkh)为试材 ,研究供铁水平对其生长及根系特性的影响 ,结果表明 ,在幼苗生长量、新叶叶绿素含量、营养液pH日变化、根系活力、根系ATP酶活性诸方面 ,存在基因型差异。同一供铁水平 ,小金海棠有明显的铁胁迫反应 ,而山定子铁胁迫反应较弱。低铁胁迫处理 ,小金海棠叶片叶绿素含量、根系活力、根系ATP酶活性皆明显高于山定子。根系活力及根系ATP酶活性可用作铁高效基因型植物的筛选指标     

Verbesserungsvorschlag zur gefahrlosen Beheizung brennbarer Lösungsmittel zur Fettextraktion (Kurzmitteilung)

A crop rotation field study with manure application was established at Tartu in 1985. Biological and chemical properties were evaluated on fine sandy loam Podzoluvisol in May 1989. The treatments included unmanured (N_o and N₈₀) controls, two peat based composts and five manures of different origin. The procedures of the most probable number (MPN) and spread plate counts were used for microbiological investigation, but also enzymatic activities, nitrogen forms, total‐C and pH were simultaneously estimated in plough layer soil. The most variable i.e. the most clearly differentiated physiological groups within manures were cellulolytic and ammonifying bacteria followed by Azotobacter spp. together with actinomycetes. Abundance of aerobic cellulolytic and ammonifying bacteria correlated positively with the number of soil algae and fungi, and negatively with nitrate‐ and nitrite‐reductase. The number of actinomycetes correlated positively with urease and catalase activity. Soil enzymatic activity was mainly modified by nitrite‐reductase. Peat composts had the highest C‐content and highest pH value compared with other soils. Pig slurry and NH₄NO₃ (N80) treatment had the highest level of fixed NH⁺ ₄ ‐ions in soil. Nine months after manure application no differences were found in the unstable NO^– ₃content of soil. Variation in the number of the studied microbial physiological groups between treatments remained insignificant.     

Assessment of the effect of silver nanoparticles on the relevant soil protozoan genus Acanthamoeba

Silver nanoparticles (AgNP) are used in a broad range of consumer products and industrial applications. During the regular product life cycle and disposal, AgNP are continuously released into the environment. Hence, the aim of this study was to investigate the potential ecotoxicological effects of AgNP exposure on amoebae. The Acanthamoeba castellanii ATCC 30234 strain and environmental isolate Acanthamoeba strain C5/2, which are both affiliated with genotype T4, were chosen as representatives of ecologically important soil protozoan organisms. The amoebae were exposed to citrate‐stabilized AgNP (30 and 70 nm in size) for 24 h and 96 h at concentrations ranging from 600 µg L⁻¹ to 20 mg L⁻¹. A newly adopted cell culture based microscopic assay was applied to assess the adherence ability of the amoeba trophozoites. The general metabolic activity of Acanthamoeba was determined to be a second independent endpoint by means of intracellular reduction of the redox dye AlamarBlue®. The fate of AgNP within the amoebae and test solutions was visualized by light‐ and transmission electron microscopy (TEM). Both Acanthamoeba strains showed a significant dose‐dependent decrease of adherence ability (p  < 0.04) and metabolic activity (p < 0.01) after 96 h of AgNP exposure. The environmental Acanthamoeba strain C5/2 lost both its adherence ability and metabolic activity at lower AgNP concentrations than the type strain, indicating a higher sensitivity to ionic silver. This was confirmed by the application of AgNO₃, provoking a higher effect level in strain C5/2. AgNP was visualized intracellularly by transmission electron microscopy within the cytoplasm of Acanthamoeba . This is the first report to show the ecotoxicological effects of short‐term AgNP exposure on the soil protist Acanthamoeba , causing both changes in the adherence ability and metabolic activity of this amoeba. This combined approach may be a powerful tool in the future for predicting potential harmful ecotoxicological effects of AgNP exposure using soil protozoans.     

动力学光度法测定小麦过氧化氢酶活动度

过氧化氢酶与四甲基联苯胺和过氧化氢反应生成蓝色物质,通过分光光度法检测蓝色产物进而可测定小麦中过氧化氢酶活动度.结果表明,光度法测得的蓝色反应的初始速率与小麦中过氧化氢酶的活动度呈线性关系,线性方程为:y=110.26x +44.06,相关系数为R=0.993.采用此方法测定小麦中的过氧化氢酶活动度准确高,重复性好,简便快捷,是测定粮食中的过氧化氢酶活动度的一种较好的方法.     

柚皮提取物对植物病原真菌的抑菌活性测定

通过测定柚皮的乙醇提取物以及各萃取部分对植物病原真菌的抑菌活性,寻找柚皮中的农用活性物质,为其综合开发利用提供科学依据。采用菌丝生长速率法测定了柚皮乙醇提取物对8种植物病原菌的抑制作用。在供试浓度为5.0mg/ml时,柚皮乙醇提取物对苹果腐烂病菌(Valsa mali)、辣椒炭疽菌(Colletotrichum capsici)和棉花枯萎菌(Fusarium oxysporum vasinfectum)的抑菌活性较高,抑菌率分别为80.7%、72.9%、62.5%。柚皮乙醇提取物的乙酸乙酯层的抑菌活性较高,乙酸乙酯层对苹果腐烂病菌的抑菌作用最好,EC50达0.45mg/ml,柚皮乙醇提取物的乙酸乙酯层是进一步研究的重点。     

芸苔链格孢菌毒素对白菜细胞膜透性、SOD酶和POD酶活性的影响

 本文以白菜幼苗为材料,研究了芸苔链格孢菌毒素(AB-毒素)对白菜叶片细胞膜透性及叶片内SOD酶、POD酶活性变化的影响。结果表明,白菜叶片用AB-毒素处理后,细胞膜透性增大,且随处理时间的延长和毒素浓度的增加而增高。不同白菜品种用毒素处理后膜透性的改变与病菌接种的发病程度呈正相关,即电导率大的白菜品种,叶片发病也重。试验结果还发现,芸苔链格孢菌及其毒素处理白菜后,体内SOD酶和POD酶活性的变化基本趋于一致,即处理后12 h内急剧下降,然后(12~48 h)活性上升。而对照处理基本变化不大。可见,芸苔链格孢菌毒素表现了与其病原菌一致的致病作用。     

水稻根面争论产碱菌1AG4的分离、鉴定及其固氮和对病原菌的拮抗作用

从水稻(Oryza sativa L.)越光根面分离到一株固氮菌株 1AG4,经生理生化特性鉴定,归类为争论产碱菌(Alcaligenes paradoxus)。1AG4 的固氮酶乙炔还原活性(ARA)为13.06 C2 H4 nm ol/m l·h,固定15N2 活性(N2 FA)为 2.052 15N a.e.% ,表明 1AG4 具有较强的固氮能力。平皿拮抗试验证明 1AG4可拮抗包括水稻三大病害病原菌在内的 14 种植物病原菌(Xanthom onas oryzae pv. oryzae ZHE173、Thanatephorus cucum eris、Piricularia oryzae Cavara、Alternaria longipes(Ellis etEverhart) Tisdale etW adkins、Cochliobolus sativus、Colletotrichum agaves Cavara、Colletotrichum lagenarium (Passerini)Ellis et Halsted、Colletotrichum panacicola Uyeda et Takim oto、Fusarium oxysp orum f. niveum 、Helm inthosporium turcicum Passerini、Phytophthora parasitica Destur、Verticillium albo-atrum Reinkeet Berthold、Phytophthora m elonis Katsura、Pseudom onas solanacearum Sm ith)的生长     

A Novel and Reliable Spectrophotometric Method for Determination of Lactoperoxidase Activity in Raw Milk

With 3, 3'5, 5'-tetramethylbenzidine(TMB) as the detection substrate, a reliable and highly selective method was established and optimized for the determination of Lactoperoxidase(LP) activity in raw milk. The method was based on the enzymatic reaction principle, where hydrogen peroxide oxidated TMB in the presence of LP. The optimized conditions of this assay system were obtained, consisting of 20 mmol · L~(-1) TMB solution, 0.6 mmol · L~(-1) hydrogen peroxide and 0.1 mol · L~(-1) Citric Acid(CA)/0.2 mol · L~(-1) disodium hydrogen phosphate(Na P) buffer(pH 4.8). TMB detection method was applied to the analysis of LP in milk samples with a practical working concentration range from 2 to 14 mg · L~(-1). The intra-and inter-batch variation coefficients were all below 5%, indicating a good repeatability. Confirmation test between TMB method and 2, 2-azinobi(3-ethylbenzothiazoline-6-sulphonate) diammonium salt(ABTS) method was carried out, and the results of TMB assay were in accordance with that of ABTS method.