Transformation of humic acids by two-domain laccase from Streptomyces anulatus

The properties of two-domain laccase of Streptomyces anulatus (SaSL) and its role in transformation of humic acids (HA) from chernozem, sod-podzolic soil and peat at alkaline pH values were studied. The SaSL was cloned, expressed in E. coli and obtained in an electrophoretically homogeneous state. SaSL is an oligomeric protein with a molecular weight of 235-300 kDa and six or eight subunits in the molecule. The molecular weight of the subunit is 37.7 kDa. Its catalytic properties are similar to those of the previously described two-domain laccase. The enzyme catalyzed oxidation of electron donors (K₄[Fe(CN)₆], ABTS) at acidic pH and phenolic substrates (2-methoxyphenol, 2,6-dimethixyphenol) at alkaline pH values. The efficiency of catalysis was higher in the case of electron donors than phenolic substrates. The enzyme showed a high thermal stability and was more stable at neutral and alkaline pH values. The enzyme effectively transformed humic acids at alkaline pH values. It was found that polymerization reactions took place during interaction of SaSL with HA, as well as with their high-molecular weight (>80 kDa) and low-molecular weight (<5 kDa) fractions. Our results suggest a possible involvement of the two-domaim laccases in humification processes in alkaline soils.     

Electrophoretic subfractionation of low and high molecular weight humic acids fractions

Summary Soil humic acid was fractionated on a molecular weight basis either using Sephadex gel filtration or electrophoresis on a discontinuous polyacrylamide gel. Low and high molecular weight fractions obtained by these two methods were choosen for subsequent subfractionation using electrophoretic methods. The high and low molecular weight fractions yielded several subfractions after separation by isotachophoresis or isoelectric focusing. Components of the high molecular weight fractions occupied the upper portion of the mobility train; components of the low molecular weight fractions lead the mobility train. Adsorption by Sephadex was avoided by using 4M urea as an eluent. The elution of the humic substances adsorbed to the polyacrylamide gel matrix was achieved by using a 0.1M Tris –0.025M EDTA solution.     

Decomposition of humic acids by incubation in a soil water-extract under various conditions of oxygen availability

Summary Decomposition of humic acids suspended in a soil water-extract under various conditions of oxygen availability (aerobic, anaerobic, alternating aerobic/anaerobic) led to the formation of fulvic acids, lower molecular weight compounds, and humin, a more complex mixture of insoluble compounds. The transformation of humic acids and formation of new humic compounds were higher under aerobic conditions than under either anaerobic or alternating aerobic/anaerobic ones.Although subjected to decomposition under greatly differing conditions of oxygen availability, the residual humic acids showed essentially the same types of chemical alterations, an increase of functional groups containing oxygen and a decrease of hydrolysable ternary and quarternary fractions.     

Gel filtration and absorption spectroscopic investigations on humic substances from organic fertilizers

Summary Three organic fertilizers: a farmyard manure (FYM), humified poplar barks (PBF) and spruce barks (SBF) were investigated by means of chemical analysis and by the study of humic substances extracted by 0.1M sodium pyrophosphate (pH=10.0). The organic carbon content and the C/N ratio were higher in SBF, probably as a consequence of the short fermentation period (5 months). Yields of organic substances removed by three consecutive extractions with Na₄P₂O₇ solution were in the order: PBF>FYM>SBF. Nominal molecular weight distribution of total extracts was studied using Sephadex G type gels; TRIS buffer (pH=9.0) was the eluant, since the use of water produced complex gel-solute interactions. It was shown that SBF had a higher content of small size particles than the other two products which exhibited quite similar elution curves. Slopes of log A(absorbance)vs log plots and ratio of absorbances at 465 and 665 nm (E₄/E₆) of fractions containing particle of lower size were the steepest and the highest, respectively.     

Degradation and transformation of humic substances by saprotrophic fungi: processes and mechanisms

Humic substances represent the main carbon reservoir in the biosphere, estimated at 1600 × 10¹⁵ g C. Due to their crucial role in reductive and oxidative reactions, sorption, complexation and transport of pollutants, minerals and trace elements, sustaining plant growth, soil structure and formation, and control of the biogeochemistry of organic carbon in the global ecosystem, humic substances are extremely important to environmental processes. Saprotrophic fungi active in the decomposition process of humic substances include mainly ascomycetes and basidiomycetes, which are both common in the upper layers of soils. White rot and litter decomposing fungi are the most important organisms in the degradation and mineralization of refractory organic matter (OM), whereas ascomycetes are mainly involved in the modification and polymerization of humic substances. The mechanisms of degradation probably involve mainly a variety of non specific oxidizing enzymes. This review provides an overview of the subject, while bridging two main disciplines: soil OM chemistry and fungal microbiology. It is aimed to highlight problems, unsolved questions and hypotheses.     

The porosity of soil aggregates from bulk soil and from soil adhering to roots

Summary Total porosity and pore-size distribution (p.s.d.) were determined in soil aggregates taken in plots planted with maize and treated with farmyard manure and three rates of compost. Soil aggregates were collected from the soil adherent to the maize roots (root soil aggregates) and from bulk soil (bulk soil aggregates). Mercury intrusion porosimetry was used to evaluate the total porosity and the p.s.d. Treatments did not affect the total porosity of the bulk soil aggregates. The same was observed for the root soil aggregates. However the total porosity of the root soil aggregates was always lower than that of the bulk soil aggregates. The loss of total porosity was found to be due to a decrease in the percentage of larger pores with respect to the total.     

Anin vitro effect of soil organic matter fractions and synthetic humic acids on the generation of superoxide radicals

Summary Humic acid, and the acid-extracted residue obtained from it, stimulated the production of superoxide radicals (O₂ ^.–) generated in the xanthine-xanthine oxidase system. Several synthetic humic acids, prepared by the oxidation of simple phenolic substances, also stimulated the production of O₂ ^.– but the degree of stimulation depended on the initial phenol. Fulvic acid and water-extractable soil organic matter were less effective in stimulating O₂ ^.– production than was humic acid. The activity of superoxide dismutase, an enzyme which destroys O₂ ^.–, was also enhanced by HA. In contrast, fulvic acid and water-extractable soil organic matter had little effect on the activity of the dismutase.     

Nature and properties of humic acid prepared from different sources and its effect on nutrient availability

Summary Effect of sources, moisture levels and extractants on extraction, analytical properties, IR-spectra, macromolecular characteristics, interaction with cations and physiological properties of the extracted material (humic acid) were studied. Among the three variables, extractant influenced the degree of humification, IR-spectra, nature and properties of humic acids to a greater extent though the other two had significant contribution. Sodium pyrophosphate found to be an ideal extractant for humic acid. Reduced viscosity of humic acids increased with dilution particularly below 0.1 g/dl and decreased with increasing H⁺ concentration between pH 6.5 and 8.5 irrespective of source, moisture level and extractant. The amount of proton released and drop in initial pH of humic acids on addition of different metal ions indicated formation of mono-, di-hydroxy metal-HA complexes which broke down at pH 8.0 and above. HA-Fe²⁺ and HA-Cu²⁺ complexes were found to be most stable. Humic acids from different sources varied significantly in their effect on available nutrient status of soils though, the latter is main determining factor particularly for calcium and magnesium.     

东北大骨节病病区主要土壤腐殖酸硒与大骨节病关系的研究

本文研究东北大骨节病病区土壤中硒的形态和含量与大骨节病的关系。研究结果表明,病区土壤中硒以有机结合态硒为主,又以富里酸态硒占主要比重。根据20个土壤样点的对比研究,显示土壤富里酸中硒含量与病情呈显著负相关。     

Effects of different organic waste amendments on soil microaggregates stability and molecular sizes of humic substances

Three soils which had been amended for several years with pig slurry, cattle slurry, and sewage sludge were dry-sieved to obtain microaggregates in the size range of 250–125, 125–50, and <50 μm. With amendments, aggregate size distribution of whole soils was shifted to larger sizes, especially for the most fragile soil, whereas percent content of microaggregates decreased except for the lower size aggregates of the fragile soil. Particle size distribution of microaggregates revealed an increase in percent sand and a reduction of percent silt and clay in the <50 μg size fraction for all soils. These results showed the aggregation effect induced by the organic waste additions. Aggregate stability of microaggregates revealed significant correlation with humic substances content (humic acids alone and humic plus fulvic acids) and non significant with total organic matter substantiating the belief that humic substances are the predominant binding agents in this aggregation range. Molecular weight distribution of humic acids extracted from microaggregates of unamended soils demonstrated that the lower the soil aggregate size distribution, the larger the contribution of the high molecular weight fraction. All microaggregates from amended soils showed a progressive increase of the high molecular weight humic acids with decreasing size, reaching a maximum in the <50 μm fraction. In this aggregate size a parallel enhancement of the aggregate stability was also evident. It is concluded that a close relationship exists between aggregate stability and high molecular weight humic substances. Additions to soils of organic material containing high molecular weight constituents would represent a useful management practice to improve aggregate stability.     

Complexation of trace metals with humic acids from soil,sediment and sewage

ABSTRACT

Metal—humate interactions in aqueous and terrestrial ecosystems control the speciation of trace metals and hence, their bioavailability and toxicity. The present study investigated the complexation interactions of copper, cadmium and lead, in their divalent states, with humic acids extracted from three different sources: the treated sewage from a treatment plant, Yamuna river bed sediment, and Yamuna river flood plain soil, all at Okhla, Delhi, employing ion selective electrode potentiometry. The conditional stability constants of the complexes were computed from Scatchard plots. The influences of the two important metal speciation factors, viz., pH and ionic strength of the reaction medium on the conditional stability constants were ascertained by investigating the reactions under three different pH: 5.0, 6.0 and 7.0 and three different ionic strengths: 0.01, 0.03 and 0.1. Stabilities of metal—humic acid (HA) complexes follow the order: Cu-HA > Pb-HA > Cd-HA for humic acids from any single source and are found to increase with a rise in pH and fall in a ionic strength of the medium. The humic acids extracted from the soil and the sediment emerged as stronger complexing agents, as compared to that extracted from sewage.     

施肥对日光温室黄瓜生长和土壤生物学特性的影响

采用田间试验研究了施肥对黄土高原日光温室黄瓜生长发育和产量,以及对土壤微生物区系和土壤酶活性的影响。结果表明,施用有机肥和沼肥明显促进了黄瓜生长发育,提高了黄瓜产量,叶面施肥可以降低无机化肥和有机肥用量,施肥对日光温室土壤生物学特性有明显影响,施用有机肥和叶面施肥增加了土壤细菌数量;施用无机化肥和沼肥增加了真菌的数量,施用有机肥降低了真菌的数量;施肥增加了放线菌的数量．同时施肥提高了土壤脲酶、磷酸酶、蔗糖酶活性,但对过氧化氢酶影响较小．施用有机肥提高了脲酶和磷酸酶活性,施用化肥和沼肥对土壤脲酶和土壤磷酸酶活性影响差异不大。     

腐殖酸对小麦抗旱性的生理效应

在干旱气候条件下,喷施腐殖酸钠（ＨＡ）、黄腐酸钠（ＦＡ）可降低土壤水分损耗,提高小麦叶片持水能力,叶片细胞超氧物歧化酶（ＳＯＤ）、过氧化氢酶（ＣＡＴ）活性明显提高,丙二醛（ＭＤＡ）含量和电解质渗出率明显降低,可减缓叶绿素降解,增强光合速率和光合产物积累,延缓植株衰老,小麦抗旱性增强,千粒重增加．     

Soil physical properties and carbon/nitrogen relationships in stable aggregates under legume and grass fallow

Short-season fallow with legumes and/or grasses can restore the soil organic C and nitrogen (N) and improve soil structure. In this study, we accessed the effects of 2-season legume and grass fallow on structural properties and C/N relationships in aggregates of a sandy loam soil. Two legumes (Calopogonium mucunoides and Centrosema pubescens), and two grasses (Guinea grass (Panicum maximum) and goose grass (Eleusine indica) were used. Results showed that Calopogonium and Centrosema increased soil total porosity and reduced soil bulk densities, while goose grass increased bulk density and reduced total porosity of the soils at 0–15 and 15–30?cm depths. Guinea grass significantly increased the saturated hydraulic conductivity (50.4?cm?h^?1) and water holding capacity of the soils. Aggregates, 4.75 to 0.5?mm were greater in Guinea grass and least in goose grass fallowed soils. Calopogonium increased macro-aggregates at 0–15?cm soils by 48%, and mean weight diameter (MWD) by 44%. Organic carbon in 0.5–0.25?mm and <0.25?mm aggregate sizes was higher in Guinea grass soils. Generally, grasses had 4-fold increases of C:N contents in dry aggregates. In conclusion, short-season fallow with Guinea grass, Calopogonium and Centrosema, increased soil C and N and protected them from losses in stable aggregates.     

Isolation of soil Streptomyces strains capable of degrading humic acids and analysis of their peroxidase activity

Abstract Fifteen Streptomyces strains capable of decolorizing humic acids in presence of glucose were isolated from soil samples using the dilute suspension technique and spread on agar plates. Six strains, displaying a significant and stable activity, were selected for further characterization. Some features of these isolates (carbon source utilization, enzyme production, antibiotic resistance) were compared with those of the reference strain Streptomyces viridosporus ATCC 39115. Degradation properties studied in batch cultures at pH 7.0 showed that the catabolic activity on humic acids was generally stimulated by incubation with 100% oxygen and was cell surface-associated. Peroxidase activity from cell-free extracts was analysed by using the oxidation of N,N,N′,N′-tetramethyl-phenylene-diamine. PAGE analysis revealed the existence of two major types of peroxidases (molecular mass: about 39.2 and 61.6 kDa), dividing the strains into two groups. The role of cell surface-associated peroxidase activity in the breakdown of humic acids is discussed.     

土壤水分对温室嫁接和非嫁接黄瓜生长与生理特性的影响

观测了黄瓜生长动态与生理特性对土壤水分状况的反应。结果表明,日光温室嫁接与未嫁接黄瓜分别在土壤相对含水量70％和80％时株高,叶面积和产量增长最快,土壤水分过高与过低均不利于黄瓜的生长;随土壤含水量降低;叶片光合速率,气孔导度,水势与根系活力的下降。     

土壤干旱条件下氮素营养对玉米内源激素含量影响

在田间持水量分别保持于35％、55％和75％±5％的土壤水分条件下,利用盆栽实验研究了土壤干旱和氮素营养对玉米内源激素和气孔导度的影响．结果表明,土壤干旱下氮素营养明显降低了玉米根系木质部汁液ABA浓度,而正常供水下施氮处理间则无显著差异(施氮处理仍较低),同时测定的叶片ABA浓度则呈相反的变化趋势,表现为干旱下施氮处理要高于不施氮处理;施氮处理木质部汁液中ZRs浓度应低于相应的不施氮处理,在调控气孔行为方面并未表现拮抗ABA作用;3种土壤水分条件下,施氮玉米叶片的气孔导度均高于不施氮处理,与木质部汁液ABA浓度呈负相关,说明施氮处理较低的根源ABA浓度是导致其气孔导度较大的主要原因．     

施肥对雷竹林土壤活性有机碳的影响

采用重施肥料试验研究了不同重施肥习惯对雷竹林土壤碳库产生的影响,结果表明,各有机肥、化肥混合处理土壤总有机碳(TOC)、水溶性碳(WSOC)、微生物量碳(MBC)、矿化态碳(MC)及WSOC/TOC、MBC/TOC和MC/TOC均显著或极显著高于单施化肥各处理.3个有机肥、化肥混施处理中,随着有机肥用量减少,TOC、WSOC、MBC和MBC/TOC显著下降,有机肥用量减少一半,上述各类碳分别下降10.75%、12.02%、30.94%和22.61%.单施化肥处理中,氮素用量超过1009.5 kg·hm^-2·年^-1会使土壤WSOC、MBC、MBC/TOC明显降低.雷竹土壤TOC、WSOC、MBC和MC两两之间相关性均达显著或极显著水平,进一步通过6个处理变异系数分析发现,土壤MBC、MBC/TOC是衡量雷竹土壤碳库质量的最佳指标.     

Amino acid distribution in some fungal melanins and of soil humic acids from Brazil

Summary Humic acids from four Brazilian topsoils of different origins and four soil fungal melanins, synthesized under two cultural conditions, were subjected to 6N HCl hydrolysis and their amino acid distribution patterns qualitatively and quantitatively determined. Both soil and fungal polymers showed similar patterns with aspartic acid, glutamic acid, glycine and alanine as the dominant amino acids. Some variations noted were more quantitative than qualitative, the similarities were more pronounced than differences, indicating that the fungal melanins may play a significant role in the formation of soil humic acid polymers. The humic acids of Brazilian soils had amino acid distribution patterns similar to those reported for humic acids of other tropical and temperate soils.     

不同管理模式对温室土壤螨群落结构的影响

为了解日光温室内不同农业管理模式对土壤螨群落的影响,以日光温室长期定位试验为研究对象,于2012年8月至2012年12月进行了5次取样,调查了有机(ORG)、无公害(LOW)和常规(CON)管理模式下土壤螨的丰度与群落结构。结果显示:管理模式对螨的丰度、各亚目的比例、菌食性隐气门螨和捕食性螨比例均无显著影响,仅对菌食性非隐气门螨和食线虫螨有显著影响。土壤螨的总数及各营养类群的数量呈现不同程度的时间波动,且波动程度在不同的管理模式下存在差异。粉螨是温室土壤中最为丰富的类群(平均37.8%)。尽管螨的类群数目呈现ORG(26)LOW(21)CON(18)的趋势,但香农指数和优势度指数没有显著变化。总之,在高投入、高扰动日光温室环境中,管理模式对土壤螨的数量和群落结构影响均较小。