Interaction Mechanism Between Environmental Microorganisms And Environmental Contaminants And Environmental Behavior

Though the effects of environmental contaminants on environmental health have been paid more and more attention, the effects are not still solved as yet. The environmental contaminants include not only the conventional heavy metals, pesticides and other organic pollutants, but also the novel contaminants (e.g. carbon nanoparticles). Once they enter the environment, the toxic effect will not be avoided. Additionally, the interaction of various contaminants occurs due to their various physical-chemical properties, so that the environmental behavior will change, such as carbon nanoparticles with large surface area can adsorb the organic chemicals, and then changed their environmental behavior. Because environmental microorganisms respond quickly to environmental stress compared to higher organisms, hence, the microbial activities can be regarded as an important indicator to evaluate the levels of environmental health. To determine the environmental microbial activity, microcalorimetry has unique advantages to monitor in situ, real-time, and continuously the interaction of environmental contaminants with microorganisms, and can observe the small changes of microbial metabolic processes due to its sensitivity.On the basis of research background, this thesis illustrates the interaction mechanism of environmental contaminants with environmental microorganism in the point of view in microbial metabolic activities, and reveals the interaction mechanism of carbon nanoparticles with organic chemicals. The main contents are following:Heavy metals (As, Cu, Cd, Cr, Co, Pb and Zn) and pesticides (chlorpyrifos (CPF) and its oxon derivative (CPO)) were selected to evaluate their toxic effects on soil microbial community. In order to stimulate the microbial activities,5.0 mg glucose (carbon source) and 5.0 mg ammonium sulfate (nitrogen source) was spiked into about 1 g soil samples. The microbial activities were recorded as power-time curves, and their indices, microbial growth rate constant k, total heat evolution QT, metabolic enthalpyΔHmet and mass specific heat rate JQ/s, were calculated. Comparing these thermodynamic parameters associated with growth yield, a general order of toxicity to the soil was found to be Cr>Pb>As>Co>Zn>Cd>Cu. When soil was exposed to heavy metals, the amount of bacteria and fungi decreased with the incubation time, and the bacterial number diminished sharply. It illustrates that fungi are more tolerant, and bacteria-fungi ratio would be altered under metal stress. For the effect of CPF and CPO, the linear correlations, k vs biomass C andΔHmet vs biomass C, elucidated that k andΔHmet were growth yield dependent. In this work,20% inhibitory ratio IC20 was obtained with 9.8μg g-1 for CPF and 0.37μgg-1 CPO, meaning that the acute toxicity of CPO was 26 times that of CPF, since the CPO had more potent toxicity to living organism due to its active functional group. Comparing the change tendency of AHmet and other parameter, the values almost kept constant when exposure to CPF (< 5.0μg g-1). It illustrates that individual reacted to stress resulted from environment change by shifting resources from other biological activities (such as reproduction or growth) toward survival to some extent. Urease activity responses in relation to the CPF and CPO exposure were observed and consistent with above thermodynamic parameters.The values of AHmet are depended on the amount of glucose consumed in soil samples. Therefore we developed a novel biosensor to determine the concentration of glucose in soil. A glucose biosensor comprising a glucose oxidase/O-(2-hydroxyl) propyl-3-trimethyl ammonium chitosan chloride nanoparticle (O-HTCC NP)-immobilized onion inner membrane and dissolved oxygen (O2) sensor successfully. The detection scheme is based on the depletion of dissolved O2 content upon exposure to glucose. The decrease in O2 level was monitored and related to the glucose concentration. The biosensor shows linear response to glucose from 0.0 to 0.60 mM with a detection limit of 50μM (S/N=3). The effect of O-HTCC NP and enzyme loading, pH, temperature, and phosphate buffer concentration on the sensitivity of the biosensor were studied in detail. The biosensor exhibits fast response time (70 s), good repeatability (3.2%, n=10) and storage stability (90% of initial sensitivity after 3-week storage). Common interferents including acetic acid, lactic acid, propionic acid, butyric acid, folic acid, methanol, glycine, DL-a-alanine and DL-cysteine do not cause significant interferences on the biosensor. The proposed biosensor method was successfully applied to determine the glucose content in real samples such as orange juice, red wine and tea drink and the results were comparable to that obtained from a spectrophotometric method. The glucose recovery test demonstrates that the proposed glucose biosensor offers an excellent, accurate and precise method for determination of glucose in real samples.Iron as an essential metal is the major scientific and medical interest, but toxicological considerations are also important in terms of accidental acute exposures and chronic iron overload. Furthermore, iron exsits commonly in natural environment. Therefore, we studied systematically the intraction of different iron species with different pure microorganisms, the interaction of bacteria-fungi with the stress of iron, and the function of reducing the toxicity of As(III) by Fe(II). Microcalorimetry was applied to assess the toxic effect of EDTA-chelated trivalent iron on the single and mixed microbes in sterilized soil that was inoculated with the Pseudomonas putida (P. putida) (bacterium), Candida humicola (C. humicola) (fungus) or their mixed species. The microbial activity was stimulated by the addition of 5.0 mg glucose and 5.0 mg ammonium sulfate under a 35% controlled humidity in 1.2 g of the studied soil samples. Results showed that the microcalorimetric indexes decreased with the increasing dose of Fe(Ⅲ)-EDTA complex. The equation Y=A×exp(-x/t)+y0 could simulate the change tendency of these parameters. Comparing the single and mixed strains, the effect of Fe(III) on bacterium-fungus interaction was dominant at lower dose of Fe(III), while, the metal toxicity observed at higher dose of Fe(III) was the main factor affecting the microbial activity. Thus, mixed-strains have moderate tolerance to the iron overload, comparing with single species, and exhibit synergistic interaction in exponential growth phase (0-0.3 mg g-1).In order to evaluate the toxic effect of different iron species on microorganism, the microcalorimetric technique based on bacterial heat-output was explored to evaluate the toxic effect of iron species (Fe2+(FeCl2), Fe3+(FeCl3) and Fe3+(FeC6H5O7)) on Escherichia coli. With the increase of the concentration of the iron ions, lag phase, became longer and peak height declined gradually. These suggest that E. coli was inhibited to a different extent with the increase of concentrations of iron ions. By comparing the growh rate constants, k, Fe2+(FeCl2), Fe3+(FeCl3) and Fe3+(FeC6H5O7) at low concentration (2-10μg mL-1 (k,0.02862→0.03055 min-1); 2-10μg mL-1 (k,0.02890→0.03056 min-1); 2-4μg mL-1 (k,0.02213→0.02607 min-1), respectively) can promote the growth of E. coli, but inhibit the growth of E. coli at high concentration (10-80μg mL-1 (k,0.03055→0 min-1); 10-80μg mL-1 (k,0.03056→0 min-1); 4-20μg mL-1 (k,0.02607→0 min-1), respectively). The total heat released, Qtotal, for there iron forms, have good relationships with concentrations of individual iron. Addtionally, the obtained values of half-inhibitory concentration (IC50) were in the order of Fe3+(ferric citrate) (9.45μg mL-1)> Fe2+(ferrous chloride) (45.23μg mL-1)> Fe3+(ferric chloride) (47.12μg mL-1), indicating the chelated iron have strongest toxicity to microorganisms.In nature, some chelators are known as siderophores, which hold an extremely high affinity for Fe3+ but very low affinity for Fe2+. Hence, ammonium ferric sulfate (AFS) was used to investigate toxic action of on Bacillus subtilis, Pseudomonas putida and Candida humicola by a series of calorimetric experiments. Candida humicola, Bacillus subtilis and Pseudomonas putida were inhibited completely when the concentrations were up to 320.0,160.0 and 160.0μg mL-1 respectively. The relationships between growth rate constant (k) and doses of AFS were approximately linear for three microbes, Pseudomonas putida for 10.0-160.0μg mL-1 (R＝-0.9746), Bacillus subtilis for 0-160.0μg mL-1 (R=-0.9868), Candida humicola for 10.0-320.0μg mL-1 (R=-0.9955). The total heat dissipated per milliliter (QT) for three microbes remained balance approximately during the lower doses, Pseudomonas putida and Bacillus Subtilis less than the dose of 20.0μg mL-1,0.56±0.01 and 0.26±0.01 J mL-1, respectively, Candida humicola less than the dose of 40.0μg mL-1,0.58±0.03 J mL-1. Comparing the half-inhibitory concentration (IC50) of three microorganisms to the AFS, their tolerance was followed the order of Candida humicola (90.1μg mL-1)> Bacillus Subtilis (65.94μg mL-1)> Pseudomonas putida (58.79μg mL-1). The biomass and OD600 of three microorganism's growth in the absence of AFS also were obtained. The power-time curve of Candida humicola' growth coincided with its turbidity curve. It elucidates that microcalorimetric method agreed with the routine microbiologic method.In the anaerobic aquatic environment, especially underground water, As(Ⅲ), Fe(Ⅱ), and phosphate co-exsit in water. In order to evaluate their interaction and reduce the toxicity of As(Ⅲ), we used the metabolic activities of Pseudomonas fluorescens in different incubation conditions as index to evaluate the reduce of toxicity of As(Ⅲ). The effect of mixed As(Ⅲ), Fe(Ⅱ) and P were moderate, compared with control, single As(Ⅲ) or Fe(Ⅱ), such as: Pmax, 328.4±21.5μW for control,291.6±17.5μW for As(Ⅲ)+Fe(Ⅱ)+P and 253.5±20.3μW for Fe(Ⅱ), 208.3±14.9μW for As(Ⅲ); k2, (6.3±0.4)×10-3 min-1 for control, (4.8±0.4)×10-3 min-1 for Fe(II), (3.9±0.2)×10-3 min-1 for As(Ⅲ) and (5.4±0.3)×10-3 min-1 for As(Ⅲ)+Fe(Ⅱ)+P; QT,8.7±0.4 J for control,7.0±0.4 J for Fe(Ⅱ),5.4±0.5 J for As(Ⅲ) and 7.8±0.6 J for As(Ⅲ)+Fe(Ⅱ)+P. This may be mainly due to physical-chemical properties of As(Ⅲ) and Fe(Ⅱ) and their interaction in aquatic environment. Coprecipitation or sorption of As on to Fe precipitates can result in lower concentrations of dissolved arsenic. This reactions can achieve the aim of reducing the As toxicity. In addition, FT-IR spectra of dry P. fluorescens after the adsoption of As(Ⅲ) and Fe(Ⅱ) and their mixture showed that Fe influenced the C-H bonds of the functional groups on the cellwall, the As(Ⅲ) caused litter effect.In order to evaluate the interaction between different contaminants, in particular for comtaninants with adsorption function, so we selected dialkyl phthalate esters (DPEs) and Carbon nanotubes (CNTs). DPEs, with endocrine disrupting functions, are widely used and categorized as priority pollutants. CNTs as strong adsorbents could influence the fate, transport and availability of DPEs in the environment. Understanding adsorptive interactions between CNTs and DPEs is critical to the environmental applications of CNTs. Adsorption of DPEs by one single-walled CNTs (SWCNT) and three multi-walled CNTs (MWCNT) was evaluated. For a given CNT, the adsorptive affinity correlated well with hydrophobicity of DPEs with an order of dimethyl phthalate (DMP)< diethyl phthalate (DEP)< dibutyl phthalate (DBP). Normalized adsorption coefficient (K/KHW) of DPEs indicates thatπ-πelectron-donor-acceptor (EDA) interaction was also important for adsorption of DPEs (π-acceptor) on CNTs (π-donor). A charge-transfer band ofπ-πEDA complexes (mixed pyrene (PYR) asπ-donor and DPEs asπ-acceptor) showed their interaction strength in the order of DMP-PYR> DEP-PYR> DBP-PYR. Calculated monolayer adsorption capacities (log Q) were bigger (for DMP and DEP) than or approximately equal to (for DBP) the estimated adsorption capacities (log Q0), implying that the DPEs were adsorbed on the surface area of CNTs. For a given DPE, the adsorptive capacities decreased with the increasing outer diameters in the order of SWCNT> MWCNT10> MWCNT20> MWCNT40.Through the studies of this thesis, the interaction mechanisms of heavy metals and pesticide with soil microbial community and different iron species with different single pure environmental microorganisms were understood, and these studies could provide an available approach to evaluate the impact of environmental contaminants on environmental health. Meanwhile, a novel biosensor was developed successfully to measure the concentration of glucose in real samples. Finally, Understanding the interaction mechanisms of carbon nanoparticles with dialkyl phthalate esters is to establish a platform to further study the environmental behavior of mixed environmental contaminants, and also offer a theoretical principle to assess the mixed impact of environmental contaminants on environmental microorganisms.