Inability of GSTT1 to activate iodinated halomethanes to mutagens in Salmonella.

Drinking water disinfection by-products (DBPs), including the ubiquitous trihalomethanes (THMs), are formed during the treatment of water with disinfectants (e.g., chlorine, chloramines) to produce and distribute potable water. Brominated THMs (Br-THMs) are activated to mutagens via glutathione S-transferase theta 1 (GSTT1); however, iodinated THMs (I-THMs) have never been evaluated for activation by GSTT1. Among the I-THMs, only triiodomethane (iodoform) has been tested previously for mutagenicity in Salmonella and was positive (in the absence of GSTT1) in three strains (TA98, TA100, and BA13), all of which have error-prone DNA repair (pKM101). We evaluated five I-THMs (chlorodiiodomethane, dichloroiodomethane, dibromoiodomethane, bromochloroiodomethane, and triiodomethane) for mutagenicity in Salmonella strain RSJ100, which expresses GSTT1, and its homologue TPT100, which does not; neither strain has pKM101. We also evaluated chlorodiiodo-, dichloroiodo-, and dibromoiodo-methanes in strain TA100 +/- rat liver S9 mix; TA100 has pKM101. None was mutagenic in any of the strains. The I-THMs were generally more cytotoxic than their brominated and chlorinated analogues but less cytotoxic than analogous trihalonitromethanes tested previously. All five I-THMs showed similar thresholds for cytotoxicity at ~2.5 µmoles/plate, possibly due to release of iodine, a well-known antimicrobial. Although none of these I-THMs was activated by GSTT1, iodoform appears to be the only I-THM that is mutagenic in Salmonella, only in strains deficient in nucleotide excision repair (uvrB) and having pKM101. Given that only iodoform is mutagenic among the I-THMs and is generally present at low concentrations in drinking water, the I-THMs likely play little role in the mutagenicity of drinking water.

Formation of iodinated trihalomethanes during chlorination of amino acid in waters.

Chlorination is essential to provide safe drinking water. However, this process leads to the formation of disinfection byproducts (DBPs). In this study, tryptophan (Trp) has been selected as a precursor to conduct the chlorine disinfection. Moreover, the factors that affect the formation of trihalomethanes (THMs) and iodinated trihalomethanes (I-THMs) are investigated. The formation pathway of Trp chlorination is proposed based on the intermediate products identified. According to the experimental results, the formation of THMs and I-THMs during Trp chlorination fitted a new first-order kinetic model. The dosage of chlorine, temperature, pH and the ratio of bromide and iodide had major influence on the formation of THMs and I-THMs during chlorination. In addition, the inhibition of luminescent bacteria Vibrio fischeri in the water sample increased during Trp chlorination.

In Vitro Response of Rumen Microbiota to the Antimethanogenic Red Macroalga Asparagopsis taxiformis.

The red macroalga Asparagopsis taxiformis has been shown to significantly decrease methane production by rumen microbial communities. This has been attributed to the bioaccumulation of halogenated methane analogues produced as algal secondary metabolites. The objective of this study was to evaluate the impact of A. taxiformis supplementation on the relative abundance of methanogens and microbial community structure during in vitro batch fermentation. Addition of A. taxiformis (2% organic matter) or the halogenated methane analogue bromoform (5 µM) reduced methane production by over 99% compared to a basal substrate-only control. Quantitative PCR confirmed that the decrease in methane production was correlated with a decrease in the relative abundance of methanogens. High-throughput 16S ribosomal RNA gene amplicon sequencing showed that both treatments reduced the abundance of the three main orders of methanogens present in ruminants (Methanobacteriales, Methanomassiliicoccales and Methanomicrobiales). Shifts in bacterial community structure due to the addition of A. taxiformis and 5 µM bromoform were similar and concomitant with increases in hydrogen concentration in the headspace of the fermenters. With high potency and broad-spectrum activity against rumen methanogens, A. taxiformis represents a promising natural strategy for reducing enteric methane emissions from ruminant livestock.

Purinergic receptor X7 is a key modulator of metabolic oxidative stress-mediated autophagy and inflammation in experimental nonalcoholic steatohepatitis.

Recent studies indicate that metabolic oxidative stress, autophagy, and inflammation are hallmarks of nonalcoholic steatohepatitis (NASH) progression. However, the molecular mechanisms that link these important events in NASH remain unclear. In this study, we investigated the mechanistic role of purinergic receptor X7 (P2X7) in modulating autophagy and resultant inflammation in NASH in response to metabolic oxidative stress. The study uses two rodent models of NASH. In one of them, a CYP2E1 substrate bromodichloromethane is used to induce metabolic oxidative stress and NASH. Methyl choline-deficient diet feeding is used for the other NASH model. CYP2E1 and P2X7 receptor gene-deleted mice are used to establish their roles in regulating metabolic oxidative stress and autophagy. Autophagy gene expression, protein levels, confocal microscopy based-immunolocalization of lysosome-associated membrane protein (LAMP)2A and histopathological analysis were performed. CYP2E1-dependent metabolic oxidative stress induced increases in P2X7 receptor expression and chaperone-mediated autophagy markers LAMP2A and heat shock cognate 70 but caused depletion of light chain 3 isoform B (LC3B) protein levels. P2X7 receptor gene deletion significantly decreased LAMP2A and inflammatory indicators while significantly increasing LC3B protein levels compared with wild-type mice treated with bromodichloromethane. P2X7 receptor-deleted mice were also protected from NASH pathology as evidenced by decreased inflammation and fibrosis. Our studies establish that P2X7 receptor is a key regulator of autophagy induced by metabolic oxidative stress in NASH, thereby modulating hepatic inflammation. Furthermore, our findings presented here form a basis for P2X7 receptor as a potential therapeutic target in the treatment for NASH.

Mutagenicity and genotoxicity of water treated for human consumption induced by chlorination by-products.

Water used for human consumption may contain mutagens and carcinogens generated during the disinfection process with chlorine. In the study described in this article, the mutagenicity and genotoxicity of water samples taken from the San Cristobal treatment plant in Medellin, Colombia, were evaluated. Short-term mutagenic and genotoxic assays using the Ames test and comet assay, respectively, were employed to examine the genotoxic activity of the extracts of these water samples. Two samples were taken before and after the chlorination process. The treated water samples without chlorination did not show mutagenic effects using the Ames test, while the chlorinated samples produced mutagenic activity in both strains. A dose-response relationship for the comet assay was obtained only in the chlorinated samples. MX (3-chloro-4-[dichloromethyl]-5-hydroxy-2[5H]-furanone), E-MX ([E]-2-chloro-3-[dichloromethyl]-4-oxobutenoic acid), and some trihalomethanes were detected at low concentrations. These concentrations were enough, however, to cause detectable mutagenic and genotoxic activity in the extracts of chlorinated water samples.

Multisite binding of a general anesthetic to the prokaryotic pentameric Erwinia chrysanthemi ligand-gated ion channel (ELIC).

Pentameric ligand-gated ion channels (pLGICs), such as nicotinic acetylcholine, glycine, γ-aminobutyric acid GABA(A/C) receptors, and the Gloeobacter violaceus ligand-gated ion channel (GLIC), are receptors that contain multiple allosteric binding sites for a variety of therapeutics, including general anesthetics. Here, we report the x-ray crystal structure of the Erwinia chrysanthemi ligand-gated ion channel (ELIC) in complex with a derivative of chloroform, which reveals important features of anesthetic recognition, involving multiple binding at three different sites. One site is located in the channel pore and equates with a noncompetitive inhibitor site found in many pLGICs. A second transmembrane site is novel and is located in the lower part of the transmembrane domain, at an interface formed between adjacent subunits. A third site is also novel and is located in the extracellular domain in a hydrophobic pocket between the ß7-ß10 strands. Together, these results extend our understanding of pLGIC modulation and reveal several specific binding interactions that may contribute to modulator recognition, further substantiating a multisite model of allosteric modulation in this family of ion channels.

Trihalomethanes and haloacetic acid species from the chlorination of algal organic matter and bromide.

Bromide and algal pollution are important factors influencing disinfection byproduct (DBP) formation and speciation in reservoir water in coastal areas. In this study, the chlorination of model algal cellular compounds (bovine serum albumin, fish oil and starch), Microcystis aeruginosa and its extra-cellular organic matter (EOM) were conducted in the absence and presence of bromide. The main aim of the present study is to explore their potential as precursors for trihalomethanes (THMs) and haloacetic acid (HAAs) speciation upon chlorination in the presence of bromide. The results showed that all brominated THMs species were generated, whereas only bromochloroacetic acid (BCAA) or/and dibromoacetic acid (DBAA) was/were produced as for brominated HAAs (Br-HAAs) from the three model compounds in the presence of bromide. The effect of bromide on Br-HAAs speciation upon fish oil chlorination was more evident than with BSA and starch. There was a good correlation between the species predicted from the model compounds and those obtained from specific algal species. Br-HAAs and Br-THMs species from Microcystis aeruginosa cells or EOM were the same as those from bovine serum albumin in the presence of bromide.

Disinfection by-product formation and mitigation strategies in point-of-use chlorination with sodium dichloroisocyanurate in Tanzania.

Almost a billion persons lack access to improved drinking water, and diarrheal diseases cause an estimated 1.87 million deaths per year. Sodium dichloroisocyanurate (NaDCC) tablets are widely recommended for household water treatment to reduce diarrhea. Because NaDCC is directly added to untreated water sources, concerns have been raised about the potential health impact of disinfection by-products. This study investigated trihalomethane (THM) production in water from six sources used for drinking (0.6-888.5 nephelometric turbidity units) near Arusha, Tanzania. No sample collected at 1, 8, and 24 hours after NaDCC addition exceeded the World Health Organization guideline values for either individual or total THMs. Ceramic filtration, sand filtration, cloth filtration, and settling and decanting were not effective mitigation strategies to reduce THM formation. Chlorine residual and THM formation were not significantly different in NaDCC and sodium hypochlorite treatment. Household chlorination of turbid and non-turbid waters did not create THM concentrations that exceeded health risk guidelines.

Development of normal human colon cell cultures to identify priority unregulated disinfection by-products with a carcinogenic potential.

Research was initiated to develop an in vitro system to identify disinfection by-products with a potential to transform normal human colonocytes into malignant cells. Tribromomethane and bromochloroacetic acid, rodent colon carcinogens, dibromonitromethane and tribromonitromethane, recently identified in drinking water, and azoxymethane, a classic colon carcinogen, were tested for the ability to transform NCM460 cells. The chronic toxicity was determined for the series of trihalomethanes, haloacetic acids and halonitromethanes as well as NCM460 cell enzymatic capabilities. The order of cytotoxicity was halonitromethanes > haloacetic acids > trihalomethanes. Cytotoxicity within a series increased with the degree of bromination and decreased with the molecular weight. The genotoxicity profile was similar to that for cytotoxicity. Enzymatic analysis demonstrated that NCM460 cells possess glutathione-S transerase-1-1 and CYP450 activity similar to that measured in the large intestine. NCM460 cells were exposed to 10(-6) M of the test chemicals for three days. While NCM460 cells from all treatments had the ability to grow in soft agar to some extent, only cells exposed to azoxymethane or tribromomethane were able to grow in media lacking serum and growth factors. When sub cultured, NCM460 cells exposed to 10(-9) M azoxymethane for three weeks formed colonies with morphology distinct from untreated cells.

Molecular volume determines the activity of the halogenated alkane bromoform at wild-type and mutant GABA(A) receptors.

The GABA(A) receptor is an important target for a variety of general anesthetics, including halogenated ethers such as isoflurane and halogenated ethers such as chloroform. Bromoform is a halogenated alkane that exhibits anesthetic properties and has been shown by X-ray crystallography to bind to model anesthetic targets. In this study we report the ability of bromoform to potentiate GABA-induced current in recombinant GABA(A) receptors composed of alpha(1)beta(2)gamma(2s) subunits. Recent studies have shown that specific point mutations in the transmembrane (TM) region of the GABA(A) receptor alpha-subunit can selectively abolish the modulatory activity of specific general anesthetics, and that molecular volume is a key determinant of anesthetic activity. The action of bromoform was examined in a series of mutant receptors and compared with the activity profiles of three other volatile anesthetics. The pharmacological profile of bromoform at the mutant receptors used in this study was similar to that seen with halothane, and distinct from that observed for isoflurane and chloroform. The molecular volume of bromoform is closest to that of halothane, and therefore our data are consistent with the idea that molecular volume is an important determinant of inhaled anesthetic activity at the GABA(A) receptor.

[Aggravation by halomethanes of damages in isolated erythrocyte membranes photosensitized by porphyrins]. / Usilenie galogenmetanami fotosensibiliziruemykh porfirinami povrezhdenii izolirovannykh éritrotsitarnykh membran.

It was found that halogen methanes (CCl4, CHBr3) enhance the destruction of protein tryptophanyls and lipid peroxidation photosensitized by Zn-tetraphenylporphin and Zn-tetramethylpyridylporphyrin in isolated erythrocyte membranes. It was shown that this effect is due to photoinduced electron transport from Zn-porphyrins to halogen methanes with the formation of highly reactive halogenmethyl radicals. The hydrophilic Zn-tetramethylpyridylporphyrin is more active at the photosensition of damage to membrane proteins, whereas the hydrophobic Zn-tetraphenylporphin is more effective in lipid peroxidation.