Microecological characteristics of water bodies/sediments and microbial remediation strategies after 50 years of pollution exposure in ammunition destruction sites in China.

The effects of long-term ammunition pollution on microecological characteristics were analyzed to formulate microbial remediation strategies. Specifically, the response of enzyme systems, N/O stable isotopes, ion networks, and microbial community structure/function levels were analyzed in long-term (50 years) ammunition-contaminated water/sediments from a contamination site, and a compound bacterial agent capable of efficiently degrading trinitrotoluene (TNT) while tolerating many heavy metals was selected to remediate the ammunition-contaminated soil. The basic physical and chemical properties of the water/sediment (pH (up: 0.57-0.64), nitrate (up: 1.31-4.28 times), nitrite (up: 1.51-5.03 times), and ammonium (up: 7.06-70.93 times)) were changed significantly, and the significant differences in stable isotope ratios of N and O (nitrate nitrogen) confirmed the degradability of TNT by indigenous microorganisms exposed to long-term pollution. Heavy metals, such as Pb, Zn, Cu, Cd, Cs, and Sb, have synergistic toxic effects in ammunition-contaminated sites, and significantly decreased the microbial diversity and richness in the core pollution area. However, long-term exposure in the edge pollution area induced microorganisms to use TNT as a carbon and nitrogen sources for life activities and growth and development. The Bacteroidales microbial group was significantly inhibited by ammunition contamination, whereas microorganisms such as Proteobacteria, Acidobacteriota, and Comamonadaceae gradually adapted to this environmental stress by regulating their development and stress responses. Ammunition pollution significantly affected DNA replication and gene regulation in the microecological genetic networks and increased the risk to human health. Mg and K were significantly involved in the internal mechanism of microbial transport, enrichment, and metabolism of TNT. Nine strains of TNT-utilizing microbes were screened for efficient TNT degradation and tolerance to typical heavy metals (copper, zinc and lead) found in contaminated sites, and a compound bacterial agent prepared for effective repair of ammunition-contaminated soil significantly improved the soil ecological environment.

Analysis of environmental biological effects and OBT accumulation potential of microalgae in freshwater systems exposed to tritium pollution.

The ecological risk of tritiated wastewater into the environment has attracted much attention. Assessing the ecological risk of tritium-containing pollution is crucial by studying low-activity tritium exposure's environmental and biological effects on freshwater micro-environment and the enrichment potential of organically bound tritium (OBT) in microalgae and aquatic plants. The impact of tritium-contaminated wastewater on the microenvironment of freshwater systems was analyzed using microcosm experiments to simulate tritium pollution in freshwater systems. Low activity tritium pollution (105 Bq/L) induced differences in microbial abundance, with Proteobacteria, Bacteroidota, and Desulfobacterota occupying important ecological niches in the water system. Low activity tritium (105-107 Bq/L) did not affect the growth of microalgae and aquatic plants, but OBT was significantly enriched in microalgae and two aquatic plants (Pistia stratiotes, Spirodela polyrrhiza), with the enrichment coefficients of 2.08-3.39 and 1.71-2.13, respectively. At the transcriptional level, low-activity tritium (105 Bq/L) has the risk of interfering with gene expression in aquatic plants. Four dominant cyanobacterial strains (Leptolyngbya sp., Synechococcus elongatus, Nostoc sp., and Anabaena sp.) were isolated and demonstrated good environmental adaptability to tritium pollution. Environmental factors can modify the tritium accumulation potential in cyanobacteria and microalgae, theoretically enhancing food chain transfer.

Defense mechanisms of alfalfa against cyclic tetramethylene tetranitramine (HMX) stress.

Much attention has been paid to the environmental toxicity and ecological risk caused by cyclic tetramethylene tetranitramine (HMX) pollution in military activity sites. In this study, the response mechanism of alfalfa plants to HMX was analyzed from the aspects of the photosynthetic system, micromorphology, antioxidant enzyme system, mineral metabolism, and secondary metabolism, in order to improve the efficiency of plant restoration. Exposure to 5 mg·L-1 HMX resulted in a significant increase in leaf N content and a significant increase and drift of the Fourier transform infrared protein peak area. Transmission electron microscopy images revealed damage to the root system subcellular morphology, but the plant leaves effectively resisted HMX pressure, and the photosynthetic parameters essentially maintained steady-state levels. The root proline content decreased significantly by 23.1-47.2 %, and the root reactive oxygen species content increased significantly by 1.66-1.80 fold. The roots regulate the transport/absorption of many elements that impart stress resistance, and Cu, Mn, and Na uptake is significantly associated with secondary metabolism. The metabolism of roots was upregulated in general by HMX exposure, with the main differences appearing in the content of lipids and lipid-like molecules, further confirming damage to the root biofilm structure. HMX causes an imbalance in the energy supply from oxidative phosphorylation in roots and generates important biomarkers in the form of pyrophosphate and dihydrogen phosphate. Interestingly, HMX had no significant effect on basic metabolic networks (i.e., glycolysis/gluconeogenesis and the tricarboxylic acid cycle), confirming that alfalfa has good stress resistance. Alfalfa plants apparently regulate multiple network systems to resist/overcome HMX toxicity. These findings provide a scientific basis for improving plant stress tolerance and understanding the HMX toxicity mechanism.

Physiological response mechanism of alfalfa seedlings roots to typical explosive cyclotrimethylene trinitramine (RDX).

This study explored the physiological response mechanism of alfalfa seedlings roots to a typical explosive, cyclotrimethylenetrinitramine (RDX), so as to improve the efficiency of phytoremediation. The response of plants to different levels of RDX were analyzed from the perspectives of mineral nutrition and metabolic networks. Exposure to RDX at 10-40 mg L-1 had no significant effect on root morphology, but the plant roots significantly accumulated RDX in solution (17.6-40.9%). A 40 mg L-1 RDX exposure induced cell gap expansion and disrupted root mineral metabolism, The key response elements, P, Cu, and Mg, were significantly increased by 1.60-1.66, 1.74-1.90, and 1.85-2.50 times, respectively. The 40 mg L-1 RDX exposure also significantly disturbed root basal metabolism, resulting in a total of 197 differentially expressed metabolites (DEMs). The main response metabolites were lipids and lipid-like molecules, and the key physiological response pathways were arginine biosynthesis and aminoacyl-tRNA biosynthesis. A total of 19 DEMs in root metabolic pathways, including L-arginine, L-asparagine, and ornithine, were significantly responsive to RDX exposure. The physiological response mechanism of roots to RDX therefore involve mineral nutrition and metabolic networks and are of great significance for improving phytoremediation efficiency.

Generic detection of organophosphorus nerve agent adducts to butyrylcholinesterase in plasma using liquid chromatography-tandem mass spectrometry combined with an improved procainamide-gel separation and pepsin digestion method.

Organophosphorus nerve agent (OPNA) adducts to butyrylcholinesterase (BChE) can be applied to confirm exposure in humans. A sensitive method for generic detection of G- and V-series OPNA adducts to BChE in plasma was developed by combining an improved procainamide-gel separation (PGS) and pepsin digestion protocol with ultra-high-pressure liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). Residual matrix interferences from prior PGS purification of OPNA-BChE adducts from plasma were found to be a critical cause of significantly reduced UHPLC-MS/MS detection sensitivity. In our developed on-column PGS approach, the matrix interference was successfully removed by adding an appropriate concentration of NaCl to the washing buffer, and it could capture ≥92.5% of the BChE in plasma. The lower pH value and the longer digestion time in all previous pepsin digestion methods were found to be a key accelerated aging factor of several adducts such as tabun (GA)-, cyclohexylsarin (GF)-, and soman (GD)-BChE nonapeptide adducts, making them difficult to detect. The aging event of several OPNA-BChE nonapeptide adducts was so successfully addressed that the formic acid level in enzymatic buffer and digestion time were lowered to 0.05% (pH 2.67) and 0.5 h, respectively, and the post-digestion reaction was immediately terminated. The improved condition parameters were optimal for pepsin digestion of all types of OPNA-BChE adducts into their individual unaged nonapeptide adducts with the highest yields, expanding the applicability of the method. The method had a nearly one-fold decrease in sample preparation time through the reduction of digestion time and removal of ultrafiltration procedure after digestion. The limit of identification (LOI) were determined respectively as 0.13 ng mL-1, 0.28 ng mL-1, 0.50 ng mL-1, 0.41 ng mL-1 and 0.91 ng mL-1 for VX-, sarin (GB)-, GA-, GF-, and GD-exposed human plasma, being low exposure value compared to previously documented approaches. The approach was utilized to fully characterize the adducted (aged and unaged) BChE levels of five OPNAs in a series of their individual exposed concentration (1.00-400 nM) of plasma sample, and successfully detect OPNA exposure from all unknown plasma samples from OPCW's second and third biomedical proficiency tests. The OPNA-BChE adducts, their aged adducts, and unadducted BChE from OPNA-exposed plasma can simultaneously be measured using the method. The study provides a recommended diagnostic tool for generic verification of any OPNA exposure with high confidence by detecting its corresponding BChE adduct.

Tritium and Carbon-14 Contamination Reshaping the Microbial Community Structure, Metabolic Network, and Element Cycle in the Seawater Environment.

The potential ecological risks caused by entering radioactive wastewater containing tritium and carbon-14 into the sea require careful evaluation. This study simulated seawater's tritium and carbon-14 pollution and analyzed the effects on the seawater and sediment microenvironments. Tritium and carbon-14 pollution primarily altered nitrogen and phosphorus metabolism in the seawater environment. Analysis by 16S rRNA sequencing showed changes in the relative abundance of microorganisms involved in carbon, nitrogen, and phosphorus metabolism and organic matter degradation in response to tritium and carbon-14 exposure. Metabonomics and metagenomic analysis showed that tritium and carbon-14 exposure interfered with gene expression involving nucleotide and amino acid metabolites, in agreement with the results seen for microbial community structure. Tritium and carbon-14 exposure also modulated the abundance of functional genes involved in carbohydrate, phosphorus, sulfur, and nitrogen metabolic pathways in sediments. Tritium and carbon-14 pollution in seawater adversely affected microbial diversity, metabolic processes, and the abundance of nutrient-cycling genes. These results provide valuable information for further evaluating the risks of tritium and carbon-14 in marine environments.

Assessing the ecological risk of tritium and Carbon-14 discharge on cyanobacteria through metabolic profiling.

The ecological risk posed by tritium (T) and carbon-14 (C-14) discharge from nuclear accidents has gained attention. This study evaluated the toxic impact of T and C-14 (at a concentration of 37 kBq/L for 15 days) on the cyanobacteria (Synechococcus elongatus). The results showed that the assimilation efficiency of cyanobacteria was significantly higher for C-14 than T, and the intracellular C-14 activity reached 30.62-40.58 kBq/kg. T and C-14 exposure had no significant effect on cell proliferation but impacted photosynthesis and respiration. T exposure increased the content of Ca, Mg, Na, P, K, and Mn, while C-14 exposure primarily affected trace element absorption in cyanobacteria. 31, 27, and 58 different metabolites (DEMs) were identified under T, C-14, and combined exposure conditions. These DEMs were enriched in the amino acid biosynthesis pathway, and nitrogen assimilation was one of the crucial pathways affected by T and C-14 exposure. The absorption of mineral elements by cyanobacteria was influenced by the variation in metabolites in the ABC transporter pathway caused by T and C-14 exposure. Our findings provide insights into the metabolic response of cyanobacteria to T and C-14 exposure and will help to guide the ecological risk evaluation of nuclear accidents.

Direct Acetonitrile-Assisted Trypsin Digestion Method Combined with LC-MS/MS-Targeted Peptide Analysis for Unambiguous Identification of Intact Ricin.

Ricin is a type II ribosome-inactivating protein toxin consisting of A and B chains linked by one interchain disulfide bond. Because of its high toxicity depending on both chains together, confirming the presence of both A and B chains of intact ricin is required during the investigation of the illegal production and application. Here, we report a novel and sensitive acetonitrile (ACN)-assisted trypsin digestion method for unambiguous identification of intact ricin by simultaneous detection of its marker peptides from A and B chains. Marker peptides were generated with a simple procedure by direct cleaving the native ricin at 45 °C for 4 h using Promega modified sequencing grade trypsin under the assistance of 10% ACN, and then directly analyzed by ultrahigh performance liquid chromatography tandem mass spectrometry. The type of trypsin was found to be one critical factor for cleavage of intact ricin based on a significant difference in the yields of specific peptides generated while using various types of trypsin. A low content of ACN in enzymatic buffer significantly reduced the digestion time from overnight to 4 h. There was commonly a better MS response of marker peptides when using the developed ACN-assisted trypsin digestion method than methanol-assisted trypsin digestion within the same 4 h. Totally, seven specific peptides with high sensitivity and specificity including three in the A-chain (TA7, TA11, and TA10) and four in the B-chain (TB6, TB14-ss-TB16, TB20, and TB18) were obtained as good marker peptides for unambiguous identification of intact ricin. The lowest concentration of native ricin for unambiguous identification was 20 ng/mL, in which three marker peptides from both the A-chain and B-chain could be measured with a minimum of three ion transitions. Combined with affinity enrichment, the developed approach was successfully applied for the measurement of intact ricin from the complicated matrix samples of the second, third, and fourth biotoxin exercises organized by the Organisation for the Prohibition of Chemical Weapons (OPCW). This study has provided a recommended detection method combined with one novel ACN-assisted trypsin digestion with MS for forensic unambiguous confirmation of trace ricin intact with high confidence.

[Metal Organic Framework MIL-53(Fe) as a Photocatalyst for Visible-light Catalytic Reduction of U(â ¥) in Aqueous Solution].

Uranium typically occurs in the hexavalent form U(Ⅵ) as the mobile, aqueous uranyl ion in radioactive wastewater. The reduction of soluble U(Ⅵ) to insoluble U(Ⅳ) oxide is an effective approach to eliminate uranium pollution. Herein, the metal organic framework material MIL-53(Fe) was successfully synthesized by a solvothermal method, and its application as photocatalyst in the reduction of U(Ⅵ) under visible light was studied in detail using various types and concentrations of hole trapping agents, solution pH values, and catalyst dosages. The results show that the use of formic acid as the hole trapping agent greatly accelerates the catalytic reaction rate by improving the charge separation efficiency. When 1 mmol·L-1 formic acid was used and the initial concentration of U(Ⅵ) was 50 mg·L-1, MIL-53(Fe) achieved a high reduction rate of 80% after 2 hours of visible light exposure. Photoelectron spectroscopy (XPS) clearly suggested that U(Ⅳ) was generated during the reaction process. A possible mechanism is that formic acid reacted with the photogenerated hole, resulting in the formation of·COO-, which can reduce U(Ⅵ) to U(Ⅳ). Accordingly, the elimination of the uranium pollution from wastewater was achieved.

Quantitation of camellianin A in HepG2 cells using a high performance liquid chromatography-electrospray ionization tandem mass spectrometric method.

The present study was designed to develop a sensitive and selective high performance liquid chromatography-tandem mass spectrometric method for the determination of Camellianin A in HepG2 cells. The extraction of Camellianin A was achieved using 15% trichloroacetic acid and then separated on a C18 column interfaced with a triple quadrupole tandem mass spectrometer in multiple reaction monitoring mode. The mobile phase was consisted of methanol-water (0.1% formic acid) (55 : 45, V/V). The total run time was 5.0 min. The method was linear in the concentration range of 0.25-250.0 ng·mL-1. The lower limit of quantification was 0.25 ng·mL-1. The intra- and inter-day relative standard deviations of entire concentration range were less than 9.3%. The proposed HPLC-MS/MS method was successfully applied to detect the intracellular concentration of Camellianin A in HepG2 cells.

Simultaneous quantification of four metabolites of sulfur mustard in urine samples by ultra-high performance liquid chromatography-tandem mass spectrometry after solid phase extraction.

Four HD urinary metabolites including hydrolysis metabolite thiodiglycol (TDG), glutathione-derived metabolite 1,1'-sulfonylbis[2-S-(N-acetylcysteinyl)ethane] (SBSNAE), as well as the ß-lyase metabolites 1,1'-sulfonylbis[2-(methylsulfinyl)ethane] (SBMSE) and 1-methylsulfinyl-2-[2-(methylthio) ethylsulfonyl]ethane (MSMTESE) are considered as important biomarkers for short-term retrospective detection of HD exposure. In this study, a single method for simultaneous quantification of the four HD metabolites in urine samples was developed using ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). The four urinary metabolites were simultaneously extracted from urinary samples using a solid phase extraction (SPE) method with high extraction recoveries for all four metabolites varied in the range of 71.1-103% followed by UHPLC-MS/MS analysis. The SPE is simple and high effective only requiring 0.1mL of urinary samples and 0.5h time consuming. The problem of previous co-elution of TDG and SBSNAE in UHPLC was well solved, and complete separation of TDG, SBSNAE, SBMSE and MSMTESE from SPE-processed urine matrix was obtained to increase specificity and sensitivity. A full method validation was performed for each analyte in urine matrix. The linear range of calibration curves for the four analytes were respectively from 0.50-500ngmL-1 for TDG and SBSNAE, 0.05-500ngmL-1 for SBMSE and MSMTESE with coefficient of determination value (R2) ≥0.990. The limit of detection was 0.25ngmL-1 for TDG and SBSNAE, 0.01ngmL-1 for SBMSE and MSMTESE spiked in normal urine. The intra/inter-day precision for each analyte at three QC levels had relative standard deviation (%RSD) of ≤10.3%, and the intra/inter-day accuracy ranged between 88.0-108%. This developed method allows for simultaneous and trace measurement of four HD urinary metabolites within one single determination with the lowest usage amount of urine samples over all previous methods This study provides a useful tool for early diagnosis and monitoring of HD poisoning for medical treatment with high confidence, avoiding the need for application of several analysis methods.

Simultaneous quantification of soman and VX adducts to butyrylcholinesterase, their aged methylphosphonic acid adduct and butyrylcholinesterase in plasma using an off-column procainamide-gel separation method combined with UHPLC-MS/MS.

This work describes a novel and sensitive non-isotope dilution method for simultaneous quantification of organophosphorus nerve agents (OPNAs) soman (GD) and VX adducts to butyrylcholinesterase (BChE), their aged methylphosphonic acid (MeP) adduct and unadducted BChE in plasma exposed to OPNA. OPNA-BChE adducts were isolated with an off-column procainamide-gel separation (PGS) from plasma, and then digested with pepsin into specific adducted FGES*AGAAS nonapeptide (NP) biomarkers. The resulting NPs were detected by UHPLC-MS/MS MRM. The off-column PGS method can capture over 90% of BChE, MeP-BChE, VX-BChE and GD-BChE from their respective plasma materials. One newly designed and easily synthesized phosphorylated BChE nonapeptide with one Gly-to-Ala mutation was successfully reported to serve as internal standard instead of traditional isotopically labeled BChE nonapeptide. The linear range of calibration curves were from 1.00-200ngmL-1 for VX-NP, 2.00-200ngmL-1 for GD-NP and MeP-NP (R2≥0.995), and 3.00-200ngmL-1 for BChE NP (R2≥0.990). The inter-day precision had relative standard deviation (%RSD) of <8.89%, and the accuracy ranged between 88.9-120%. The limit of detection was calculated to be 0.411, 0.750, 0.800 and 1.43ngmL-1 for VX-NP, GD-NP, MeP-NP and BChE NP, respectively. OPNA-exposed quality control plasma samples were characterized as part of method validation. Investigation of plasma samples unexposed to OPNA revealed no baseline values or interferences. Using the off-column PGS method combined with UHPLC-MS/MS, VX-NP and GD-NP adducts can be unambiguously detected with high confidence in 0.10ngmL-1 and 0.50ngmL-1 of exposed human plasma respectively, only requiring 0.1mL of plasma sample and taking about four hours without special sample preparation equipment. These improvements make it a simple, sensitive and robust PGS-UHPLC-MS/MS method, and this method will become an attractive alternative to immunomagnetic separation (IMS) method and a useful diagnostic tool for retrospective detection of OPNA exposure with high confidence. Furthermore, using the developed method, the adducted BChE levels from VX and GD-exposed (0.10-100ngmL-1) plasma samples were completely characterized, and the fact that VX being more active and specific to BChE than GD was re-confirmed.

[Advances in peroxide-based decontaminating technologies].

With the boosting demand for eco-friendly decontaminants, great achievements in peroxide-based decontaminating technologies have been made in recent years. These technologies have been applied in countering chemical/biological terrorist attacks, dealing with chemical/biological disasters and destructing environmental pollutants. Recent research advances in alpha-nucleophilic/oxidative reaction mechanisms of peroxide-based decontamination against chemical warfare agents were reviewed, and some classical peroxide-based decontaminants such as aqueous decontaminating solution, decontaminating foam, decontaminating emulsions, decontaminating gels, decontaminating vapors, and some newly developed decontaminating media (e.g., peroxide-based self-decontaminating materials and heterogeneous nano-catalytic decontamination systems) were introduced. However, currently available peroxide-based decontaminants still have some deficiencies. For example, their decontamination efficiencies are not as high as those of chlorine-containing decontaminants, and some peroxide-based decontaminants show relatively poor effect against certain agents. More study on the mechanisms of peroxide-based decontaminants and the interfacial interactions in heterogeneous decontamination media is suggested. New catalysts, multifunctional surfactants, self-decontaminating materials and corrosion preventing technologies should be developed before peroxide-based decontaminants really become true "green" decontaminants.

[Photocatalytic removing of a mustard gas analogue 2-CEES vapor over SO4(2-)/TiO2].

Disinfection with photocatalysis, compared to with the conventional cleanout, is both high efficient and non contaminative, but the simple TiO2 photocatalyst is showing to be of low activity and low active stability so to be hardly practical application. In the paper, SO4(2-)/TiO2 were papered by surface modification of TiO2 with dilute H2SO4, and the photocatalytic degradation of 2-chloroethyl ethyl sulfide (2-CEES) on the samples was examined in a fixed-bed microreactor. The examination show that the acidic modification enhanced both the activity and the active stability of TiO2, and the sample ST200 prepared by calcination at 200 degrees C was better than ST400 by calcination at 400 degrees C. The effect of water vapor content and reaction temperature on the photocatalytic degradation of 2-CEES was also tested, showing that the sample ST200 had high activity and stability at 90 degrees C, and kept a constant activity when adding 30.5 mL/L water vapor into the reactive system in which 2-CEES initial concentration was low to < 61 microL x L(-1). In addition, it was found that supporting SO4(2-)/TiO2 on gamma-Al2O3, SiO2 and active carbon could improve on the activity and stability of SO4(2-)/TiO2, and on supports SiO2 is the best one.