[Effect of PFOA on Oxidative Stress and Membrane Damage of Escherichia coli].

Perfluorooctanoic acid (PFOA) is widely used in industrial production because of its strong chemical stabilities and good hydrophobic and oleophobic properties. It was considered to be a widespread persistent organic pollutant in environment in recent years. The oxidative stress and membrane damage of Escherichia coli exposed to PFOA were measured by flow cytometry (FCM) and the toxic mechanism of PFOA was also preliminarily explored. The results showed that, under the stress of PFOA, the intracellular reactive oxygen species (ROS) content of E. coli increased, the unsaturation degree of fatty acid decreased, the malondialdehyde (MDA) content increased, the membrane permeability increased, the membrane potential decreased, and the activities of Na+K+-ATPase and Ca2+Mg2+-ATPase showed a compensatory increase first and then decreased. Therefore, owing to the stress of PFOA, the higher intracellular ROS in E. coli reacted with membrane unsaturated fatty acids by peroxidation,and then reduced cell membrane fatty acid saturation, accumulated MDA in cells, and further caused damage to cell membrane, reduced the ATPase activity, and eventually resulted in inactivation or apoptosis of E. coli. This study provided more evidence for the further study on environmental ecological toxicology of PFOA.

Complete degradation of the endocrine disruptor di-(2-ethylhexyl) phthalate by a novel Agromyces sp. MT-O strain and its application to bioremediation of contaminated soil.

A newly isolated strain Agromyces sp. MT-O could utilize various phthalates and efficiently degraded di-(2-ethylhexyl) phthalate (DEHP). Response surface methodology was successfully employed for the optimization of culture conditions including pH (7.2), temperature (29.6), and inoculum size (OD600 of 0.2), resulting in almost complete degradation of DEHP (200mgL(-1)) within 7days. At different initial concentrations (50-1000mgL(-1)), DEHP degradation curves were fitted well with the first-order kinetic model, and the half-life of DEHP degradation ranged from 0.83 to 2.92days. Meanwhile, the substrate inhibition model was used to describe the special degradation rate with qmax, Ks, and Ki of 0.6298day(-1), 86.78mgL(-1), and 714.3mgL(-1), respectively. The GC-MS analysis indicated that DEHP was degraded into mono-ethylhexyl phthalate and phthalate acid before its complete mineralization. Bioaugmentation of DEHP-contaminated soils with strain MT-O has greatly enhanced DEHP disappearance rate in soils, providing great potential for efficiently remediating DEHP-contaminated environment.

[Characteristics and functional protein analysis of an effective decabromodiphenyl ether-degrading strain].

An effective decabromodiphenyl ether (BDE-209) degrading strain was isolated and identified as Enterococcus casseliflavus based on the 16S rRNA gene sequence analysis. The optimal conditions for strain growth were pH 7 and culture time of 48 h, respectively. E. casseliflavus has a good ability to degrade BDE-209. The biodegradation rate of 1 mg.L-1 BDE-209 by 1 g.L-1 E. casseliflavus reached the highest of 56. 7% after 4 days degradation with 5 mg.L-1 glucose as the additional carbon source. During the degradation process of BDE-209, SDS-PAGE demonstrated that some new extracellular proteins were induced under 2 mg.L-1 and 5 mg.L-1 BDE-209. As for the intracellular proteins, the quantity of protein expression varied, and some proteins even disappeared compared with the blank control. Two-dimensional electrophoresis steps for protein analysis detected 31 different protein points, demonstrating that during the degradation process, the conformation of some proteins which were related with degradation was changed, and resulted in the variation of type and content of the proteins.

[Effect of heavy metal ions on triphenyltin enzymatic degradation].

The influence of different metal ions and different forms of addition on triphenyltin enzymatic degradation was investigated under conditions using enzyme obtained from Klebsiella pneumoniae. The objective of this study is to illuminate the mechanism of enzymatic degradation of triphenyltin (TPhT). The results demonstrated that the strain was able to tolerate K+, Mg2+, CU2+, Ca2+ and Fe3+ at high concentrations. High concentrations of Zn2+ and Fe2+ had some toxic effects on the strain, thus affecting its growth. The endoenzyme activity was enhanced by metal ions such as K+, Mg2+, Zn2+, Cu2+ and Fe2+ at certain concentrations. In the presence of 30 mg/L of Mg2+, the removal percentage of TPhT was up to 77.22%. Fe3+ restrained the enzyme activity at certain concentrations. Adding K+, Mg2+, Zn2+, Cu2+ into medium can promote the production of enzyme, among which Mg2+ demonstrated up to 85.66% of removal percentage of TPhT, suggesting some metal ions at the appropriate concentration range can be used as enzyme activator for the enzymatic degradation of triphenyltin. Metal ions showed no relevant impact on the cell growth and enzyme production. Certain metal ions can only serve as activators of endoenzyme and exhibited no similar effect towards exoenzyme.

Biodegradation of anthracene by Aspergillus fumigatus.

An anthracene-degrading strain, identified as Aspergillus fumigatus, showed a favorable ability in degradation of anthracene. The degradation efficiency could be maintained at about 60% after 5d with initial pH of the medium kept between 5 and 7.5, and the optimal temperature of 30 °C. The activity of this strain was not affected significantly by high salinity. Exploration on co-metabolism showed that the highest degradation efficiency was reached at equal concentration of lactose and anthracene. Excessive carbon source would actually hamper the degradation efficiency. Meanwhile, the strain could utilize some aromatic hydrocarbons such as benzene, toluene, phenol etc. as sole source of carbon and energy, indicating its degradation diversity. Experiments on enzymatic degradation indicated that extracellular enzymes secreted by A. fumigatus could metabolize anthracene effectively, in which the lignin peroxidase may be the most important constituent. Analysis of ion chromatography showed that the release of anions of A. fumigatus was not affected by addition of anthracene. GC-MS analysis revealed that the molecular structure of anthracene changed with the action of the microbe, generating a series of intermediate compounds such as phthalic anhydride, anthrone and anthraquinone by ring-cleavage reactions.

[Cation exchanges during the process of Cd(2+) absorption by Alfalfa in aqueous solutions].

A hydroponic experiment was conducted to investigate the cation exchanges during the process of Cd2+ absorption by Alfalfa in aqueous solution. The absorption efficiency of Alfalfa plants with 0-10 mg x L(-1) Cd2+ treatments, changes of Na+, K+, Mg2+, Ca2+ and NH4(+) concentration, and the variation of pH values at different absorption time (0, 1, 2, 4, 8, 12, 24 and 72 h) were studied separately. The multiple linear regressions between Cd2+ absorption and cation variation were analyzed. The results indicated that when Cd2+ concentrations were 0.1, 1, 5, 10 mg x L(-1), the absorption efficiencies of Cd2+ by Alfalfa after 72 h were 85.86%, 52.14%, 15.97% and 7.81%. Cation exchange was involved in the removal of Cd2+ by Alfalfa in aqueous solution. Except for NH4(+), the concentrations of cationic metals Na+, K+, Mg2+ and Ca2+ in aqueous solution increased over time, which increased 11.30% - 61.72%, 21.44% - 98.73%, 24.09% - 8.90% and 37.04% - 191.96%, respectively. Kinetic studies illuminated that the release of Na+, K+, Mg2+ and Ca2+ by Alfalfa in Cd2+ solution with initial concentrations of 0, 0. 1, 1, 5, 10 mg x L(-1) best fitted pseudo-second-order equation,while the NH4(+) release fitted this model when Cd2+ concentrations were 1, 5, 10 mg x L(-1). The gradual decrease of pH during adsorption of Cd2+ by Alfalfa was observed. As the competition ion of Cd2+, H+ might affect the capacity of Alfalfa root system to absorb Cd2+. The ternary linear equation results demonstrated that the content of Cd2+ absorption by Alfalfa strongly related with the release of Ca2+, Mg2+, Na+. And this exchange mainly occurred among Cd2+ and divalent cations.

[Characteristics of biodegradation of triphenyltin by enzyme obtained from Klebsiella pneumoniae].

The objective of this study is to illuminate the mechanism of biodegradation of triphenyltin (TPhT). The removal of TPhT by Klebsiella pneumoniae was, therefore, investigated through characteristics studies. The influences of the various parameters were also discussed. The results demonstrated that the cell, extracellular secretion and intracellular enzyme were the effective biomasses for the biodegradation of TPhT. At initial concentration of 3 mg x L(-1), 10.9%, 5.3% and 47.9% of TPhT could be degraded by these biomasses respectively at 30 degrees C within 2 hours under an rotary shaker at 120 r x min(-1). The experimental results also showed that the enzyme activity could be affected by the buffers, pH, temperature, metals and the concentration of TPhT. The degradation efficiency would reach the highest point at pH 8, and at the optimal temperature of 50 degrees C. Metals including Mg2+, Mn2+, Fe2+ and Fe3+ improved the enzyme activity at certain concentrations. In the presence of 15 mg x L(-1) of Mg2+, the removal percentage of TPhT was up to 73.8%. It suggested that the metals activated the enzyme and interacted with the TPhT enabling its removal during the biodegradation process. Linear plots of removal ratios versus concentrations of TPhT meant that the biodegradation fitted the Michaelis-Menten model. The Vmax and Km of this biodegradation were 0. 15 mg x (L x min)(-1) and 47.1 mg x L(-1), respectively.

[Isolation of an anthracene-degrading strain Aspergillus fumigatus A10 and its degradation characteristics].

An anthracene-degrading strain (A10) was isolated from contaminated environment and identified as Aspergillus fumigatus. The experimental results showed that the biodegradation rate of anthracene increased with the increasing time. Between 12-84 h interval, the biodegradation performed rapidly, while after this, the increase of biodegradation rate tended to become slow, and ultimately the biodegradation rate could achieve approximately 83%. The degradatinn rate of anthracene reached 79.37% within 5 days when the initial concentration of anthracene in mineral salts medium (MSM) was 10 mg/L, the inoculum dosage was 50 g/L (wet weight) and the cell age was 36 h. The concentration of anthracene had notable influence on degradation function of strain A10 and the highest degradation rate (92.17%) was achieved when anthracene concentration was 5 mg/L. The degradation rate could maintain about 60% with initial pH of MSM in the range of 5.0-7.5, and also, the anthracene could be better broken down when the temperature was 30 degrees C and dissolved oxygen was 4.30 mg/L. Certain amount of nutrition salts promoted the biodegradation of anthracene to some extent. Addition of lactose as co-metabolic substrate most favorably accelerated degradation of anthracene by about 37.15%. The mechanism research revealed that the biodegradation by strain A10 was a dynamic process in which extracellular sorption and intracellular degradation were included. FT-IR analysis exhibited that the structure of anthracene changed with the action of microbe, generating a series of metabolites, such as aromatic acid, aromatic ketone, aromatic aldehyde with one or two benzene rings, as well as saturated hydrocarbons.

[Isolation and characteristics of triphenyltin-biosorption and biodegradation strain].

The biosorption and biodegradation of triphenyltin (TPhT) from aqueous solutions by isolated strains were investigated through microbial separation and characteristic studies. The results illuminated that Klebsiella pneumoniae was an effective strain for the biosorption and removal of TPhT. 70% to 97.9% of TPhT with initial concentration of 3 mg x L(-1) could be absorbed within 2 h, and 26.4% to 54.6% of this TPhT was biodegraded within 5 d using 0.3 g x L(-1) to 3.0 g x L(-1) biomass of K. pneumoniae. TPhT could be more effectively degraded by separated endoenzyme than by cell biomass,and 28.1% to 77.8% of TPhT would be degraded by this way within 2.0 h.The biodegradation experiments also showed that the degradation of TPhT by K. pneumoniae mainly occurred intracellularly and the increase of degradation rate gradually slowed down with time. Separated endoenzyme gave rise to the highest biodegradation of TPhT at the contact time of 2.0 h,while the intact cell achieved the highest rate of biodegradation within 1 d, and then reached a plateau among 2 d to 5 d. The experiments also revealed that the biodegradation process of TPhT included biosorption by cell wall, active transportation across cell membrane, and biodegradation within cytoplasm, in which TPhT bio-adsorbed oh the cell wall increased linearly with time,TPhT inside the cell wall decreased rapidly from 55.9% to 17.0% during the first 3 d, and then turned to smooth; while TPhT in the supernate changed reversely with that occurred inside the cell wall.

[Effects of low concentration heavy metals on biodegradation of BDE209 by Bacillus cereus].

In view of joint contamination of heavy metals and polybrominated diphenyl ethers (PBDEs) caused by electronic-wastes, analysis measures of GC-MS, ICP, UV scanning, fTIR, SEM, etc. were used to research on the debromination of deca-brominated diphenyl ethers (BDE209) and biodegradation capability under aerobic condition by combined Bacillus cereus XPB and XPC, and the effects of low concentrations of heavy metals on the biodegradation of BDE209 were also studied. The experimental results showed that combined Bacillus cereus efficiently debrominated and degraded BDE209 to hydroxybenzenes, and the highest debromination capability of 1.18 mg x L(-1) with the efficiency of 14.16% at least was achieved after 1 d reaction. Although biodegradation process was delayed at presence of low concentrations of heavy metals, satisfying degradation effect was still achieved with debromination efficiency of not less than 13.92%. Hydroxy, a minoacyl, and alkyl were confirmed to be the key functional groups for combined Bacillus cereus to biodegrade BDE209 and adsorb heavy metals. Obvious release of K+ and Na+ was observed and the release quantity rose up from 148.867 micromol x g(-1) and 225.835 micromol x g(-1) respectively, when only biodegradation was involved, to 156.482 micromol x g(-1) and 261.217 micromol x g(-1) individually when biodegradation and biosorption acted simultaneously. During the process of BDE209 biodegradation, the highest adsorption rates for Pb2+, Zn2+, and Cu2+ by combined Bacillus cereus were 89.47%, 72.22% and 39.83% respectively.

[Characteristics and pathway of naphthalene degradation by Pseudomonas sp. N7].

The biodegradation characteristics of a typical polycyclic aromatic hydrocarbon, naphthalene by the strain (Pseudomonas sp. N7) were investigated by using HPLC and UV analytical techniques. The results showed that the addition of nutritious salt and microelements accelerated the degradation of naphthalene by 23.65%. Degradation efficiency increased with increasing dissolved oxygen and reached 95.66%, then remained stabilized when dissolved oxygen was over 4.3 mg/L, yet decreased with increasing naphthalene concentration. Neutral and weak alkaline condition favored the biodegradation with degradation capacity all over 82.88%. Pseudomonas sp. N7 had a maximum degradation capability of 95.66% when dealing with 100 mg/L naphthalene at 30 degrees C and pH 7.5 with 165 r/min rotary shaking for 72 h. By measuring the absorbance, pH and degradation of substrates during treatment of different substrate with strain N7, it was demonstrated that Pseudomonas sp. N7 could also degrade other aromatic hydrocarbons, such as toluene, dimethylbenzene, phenol, 2,4-nitrophenols, benzyl acid, 1-naphthol and salicylic acid, utilizing each of them as sole carbon and energy source for growth and breeding, thus showing its good biodegradation diversity. The pathway of naphthalene degradation was explored through analyzing metabolic intermediates at different degradation stages by using UV-Vis and GC-MS. The result revealed that there were two possible degradation pathways for naphthalene: one was phthalic acid pathway, and the other was that naphthalene was first oxidized to 1,2-dihydroxynaphthalene, and then the cleavage of rings caused the formation of salicylic acid, catechol, and 2-hydroxymuconic semial-dehyde. Finally these metabolites entered the tricarboxylic acid cycle (TCA).

[Cloning and expression of the nickel/cobalt transferase gene in E. coli BL21 and bioaccumulation of nickel ion by genetically engineered strain].

1 053 bp of the nickel/cobalt transferase gene, NiCoT gene, from Staphylococcus aureus ATCC6538 was amplified by PCR and ligated into vector pET-3c. The recombined plasmid was constructed and transferred into E. coli BL21 at appropriate temperature. The recombined strain was isolated and identified by restriction enzyme digestion and PCR amplification. Nucleotide sequence analysis showed that the identity was more than 97% between the nickel/cobalt transferase gene from S. aureus ATCC6538 and the reported gene sequence of other species or subspecies of S. aureus at GenBank. There was a characteristic protein strip near the relatively molecular weight of 39 000 in the SDS-PAGE picture, which was identical to the expected value. The result demonstrated that the NiCoT gene of S. aureus had been successfully expressed in E. coli BL21. The E. coli BL21 containing the NiCoT gene had the highest bioaccumulation quantity when induced with 1.00 mmol x L(-1) IPTG for 4 h. The quantity of equilibrium accumulation of the genetically engineered E. coli BL21 was 11.33 mg x g(-1), which was 3 times more than that of the original E. coli BL21 at different nickel concentrations. The NiCoT of S. aureus ATCC6538 was a highly selective and accumulative nickel transporter and belonged to the class III of the nickel/cobalt transferase.