Pyrite-assisted degradation of methoxychlor by laccase immobilized on Fe3S4/earthworm-like mesoporous SiO2.

Laccase immobilized and cross-linked on Fe3S4/earthworm-like mesoporous SiO2 (Fe3S4/EW-mSiO2) was used to degrade methoxychlor (MXC) in aqueous environments. The effects of various parameters on the degradation of MXC were determined using free and immobilized laccase. Immobilization improved the thermal stability and reuse of laccase significantly. Under the conditions of pH 4.5, temperature 40 °C, and reaction time 8 h, the degradation rate of MXC by immobilized laccase reached a maximum value of 40.99% and remained at 1/3 of the original after six cycles. The excellent degradation performance of Fe3S4/EW-mSiO2 was attributable to the pyrite (FeS2) impurity in Fe3S4, which could act as an electron donor in reductive dehalogenation. Sulfide groups and Fe2+ reduced the activation energy of the system resulting in pyrite-assisted degradation of MXC. The degradation mechanism of MXC in aqueous environments by laccase immobilized on Fe3S4/EW-mSiO2 was determined via mass spectroscopy of the degradation products. This study is a new attempt to use pyrite to support immobilized laccase degradation.

Characterization of enzyme-immobilized catalytic support and its exploitation for the degradation of methoxychlor in simulated polluted soils.

Chiral mesoporous silica (SiO2) with helical structure was synthesized by using anionic surfactants as template. Pre-prepared graphene oxide (GO) was then loaded onto SiO2 to synthesize composite carrier chial-meso-SiO2@GO for the immobilization of laccase. The enzyme activity, thermostability, acid stability, and repeatability of the immobilized enzyme were significantly improved after immobilization. The chial-meso-SiO2@GO-immobilized laccase was then used for the degradation of MXC in aqueous phase. The degradation conditions, including temperature, time, pH, MXC concentration, and the dose of immobilized enzyme for cellulosic hydrolysis, were optimized. The optimum conditions for degradation of methoxychlor were selected as pH 4.5, MXC concentration 30 mg/L, immobilized enzyme dose 0.1 g, the maximum MXC removal of over 85% and the maximum degradation rate of 50.75% were achieved after degradation time of six h at temperature of 45 °C. In addition, the immobilized cellulase was added into the immobilized laccase system to form chial-meso-SiO2@GO-immobilized compound enzyme with the maximum MXC degradation rate of 59.58%, higher than that of 50.75% by immobilized laccase. An assessment was made for the effect of chial-meso-SiO2@GO-immobilized compound enzyme on the degradation of MXC in soil phase. For three contaminated soils with MXC concentration of 25 mg/kg, 50 mg/kg, and 100 mg/kg, the MXC removals were 93.0%, 85.8%, and 65.1%, respectively. According to the GC-MS analyses, it was inferred that chial-meso-SiO2@GO-immobilized compound enzyme had a different degradation route with that of chial-meso-SiO2@GO-immobilized laccase. The hydrolysis by immobilized cellulase might attack at a weak location of the MXC molecule with its free radical OH and ultimately removed three chlorine atoms from MXC molecule, leading to generating small molecular amount of degradation product.

USPIO assisting degradation of MXC by host/guest-type immobilized laccase in AOT reverse micelle system.

The laccase and ultrasmall superparamagnetic iron oxide nanoparticles (USPIO) have been assembled inside the tubular mesoporous silica via co-adsorption technology to prepare host/guest-type immobilized laccase, which is applied to degrade methoxychlor (MXC) in aqueous and reverse micelle environments. The effects of various parameters on degradation of MXC were studied. Under the optimum conditions, the degradation rate could reach maximum value of 45.6 % and remain at 20.8 % after seven cycles. Moreover, the addition of small molecular compound 2, 2'-azinobis-(3-ethylbenzthiazoline-6-sulphonate) to the system could greatly improve the degradation efficiency. The MXC degradation process is a first-order reaction, and the activation energy of MXC degradation catalyzed by immobilized laccase (41.46 kJ mol(-1)) is relatively lower than that catalyzed by free laccase (44.91 kJ mol(-1)). Based on the degradation products measured by gas chromatograph-mass spectrometer (GC-MS) and nuclear magnetic resonance (NMR), the degradation mechanism of MXC has also been proposed.

Destruction of halogen-containing pesticides by means of detonation combustion.

Pesticides that contain a halogen functional group have been destructed by means of detonative combustion. The following compounds were examined: (1) atrazine-2-chloro-4-ethylamino-6-isopropylamino-1,3,5-triazine-herbicide; (2) bromophos-O,4-bromo-2,5-dichlorophenyl O,O-dimethyl phosphorothioate-insecticide; (3) chloridazon-5-amino-4-chloro-2-phenylopyridazin-3(2H)-one-herbicide; (4) linuron-3-(3,4-dichlorophenyl)-1-metoxy-1-methylurea-herbicide; (5) metoxychlor-1,1,1-trichloro-2,2-bis(4-metoxyphenyl)ethane-insecticide and acaricide; and (6) trichlorfon-dimethyl 2,2,2-trichloro-1-hydroxyethylphosphonate-insecticide. Explosive material has been produced on the basis of ammonium nitrate, which served as an oxidizer while the pesticides were used as fuels. Composition of the explosive was adjusted in such a way as to respect thermodynamic parameters. Detonative decomposition of the mixtures has been carried out in shot-holes pre-drilled in soil. Efficiency of the pesticide decomposition has been examined with gas chromatography in order to determine pesticides residues in the environment. It was found that for some, the amount of pesticides in some compounds in the analyzed samples after decomposition was below the determination threshold of the applied method.

Zinc(II) complexes with dithiocarbamato derivatives: structural characterisation and biological assays on cancerous cell lines.

Zinc is one of the most important trace elements in the body and it is essential as a cofactor for the structure and function of a number of cellular molecules including enzymes, transcription factors, cellular signalling proteins and DNA repair enzymes. On the other hand, recent studies have shown that zinc could play a role both in the development of various cancers and in the induction of apoptosis in some cell types, however, no established common relationships of zinc with cancer development and progression have been identified. To date, in our research group different metal-dithiocarbamato complexes have been designed that were expected to resemble the main features of cisplatin together with higher activity, improved selectivity and bioavailability, and lower side-effects. On the basis of the obtained encouraging achievements with other metals (such as gold and copper) we have decided to enlarge the studies to the complexes of zinc(II) using the same ligands. Hereby, we report the results on the synthesis and characterisation of ZnL(2) complexes with five different dithiocarbamato derivatives, such as dimethyl-(DMDT), pyrrolidine-(PyDT), methyl-(MSDT), ethyl-(ESDT) and tert-butyl-(TSDT) sarcosinedithiocarbamate. All the obtained compounds have fully been characterised by means of several spectroscopic techniques. In addition, the crystal structure of [Zn(MSDT)(2)](2) dinuclear complex is also reported. In order to evaluate the in vitro cytotoxic properties, some biological assays have been carried out on a panel of human tumour cell lines sensible and resistant to cisplatin. Some of the tested compounds show cytotoxicity levels comparable or even greater than the reference drug (cisplatin).

Pesticides removal from waste water by activated carbon prepared from waste rubber tire.

Waste rubber tire has been used for the removal of pesticides from waste water by adsorption phenomenon. By applying successive chemical and thermal treatment, a basically cabonaceous adsorbent is prepared which has not only a higher mesopore, macropore content but also has a favorable surface chemistry. Presence of oxygen functional groups as evidenced by FTIR spectra along with excellent porous and surface properties were the driving force for good adsorption efficiency observed for the studied pesticides: methoxychlor, methyl parathion and atrazine. Batch adsorption studies revealed maximum adsorption of 112.0 mg g(-1), 104.9 mg g(-1) and 88.9 mg g(-1) for methoxychlor, atrazine and methyl parathion respectively occurring at a contact time of 60 min at pH 2 from an initial pesticide concentration of 12 mg/L. These promising results were confirmed by column experiments; thereby establishing the practicality of the developed system. Effect of various operating parameters along with equilibrium, kinetic and thermodynamic studies reveal the efficacy of the adsorbent with a higher adsorption capacity than most other adsorbents. The adsorption equilibrium data obey Langmuir model and the kinetic data were well described by the pseudo-first-order model. Applicability of Bangham's equation indicates that diffusion of pesticide molecules into pores of the adsorbent mainly controls the adsorption process. Spontaneous, exothermic and random characteristics of the process are confirmed by thermodynamic studies. The developed sorbent is inexpensive in comparison to commercial carbon and has a far better efficiency for pesticide removal than most other adsorbents reported in literature.

Bioconcentration and biotransformation of [¹4C]methoxychlor in the brackish water bivalve Corbicula japonica.

To obtain basic information on the metabolic fate of xenobiotics in the brackish water, bivalve Corbicula japonica, bioconcentration and biotransformation experiments were performed using methoxychlor (MXC) as a model compound. Bivalves were exposed to [ring-U-¹4C]MXC (10 µg L⁻¹) for 28 days under semi-static conditions followed by a 14-day depuration phase. The ¹4C concentration in the bivalves rapidly increased and reached a steady state after exposure for 7 days (BCFss = 2010); however, it rapidly decreased with a half-life of 2.2 days in the depuration phase. Mono- and bis-demethylated MXC, and their corresponding sulphate conjugates, were identified as minor metabolites. No glycoside conjugates (including glucuronide and glucoside) were detected. Despite this biotransformation system, bivalves were found to excrete retained MXC mostly unchanged although its relatively hydrophobic nature.

Detection of thymocytes apoptosis in mice induced by organochlorine pesticides methoxychlor.

The thymus has long been known to be vulnerable to atrophy when exposed to variety of stimuli, including hormones, immunosuppressive pharmaceuticals, and environmental chemicals. The organochlorine pesticide methoxychlor (MXC) is an immunosuppressive agent thought to affect thymic atrophy by inducing apoptosis of thymocyte T cells. We sought to develop an experimental protocol to detect in vivo thymocyte apoptosis induced by MXC in Balb/c mice. We treated the mice with 150-400 mg/kg MXC. We then measured thymus weight, cell counts, caspase activity (3/7, 8, and 9), annexin V labeling of phosphatidylserine (PS) and DNA fragmentation. In MXC-treated mice we observed decreases in thymus weight and cell counts and increases in caspase activity (3/7, 8, and 9), annexin V PS labeling and DNA fragmentation. These results suggest that MXC induces thymic atrophy caused by thymocyte apoptosis, and that our protocol may be useful for detecting in vivo thymocyte apoptosis induced by environmental chemicals in short-time.

Metabolism of methoxychlor by the P450-monooxygenase CYP6G1 involved in insecticide resistance of Drosophila melanogaster after expression in cell cultures of Nicotiana tabacum.

Cytochrome P450 monooxygenase CYP6G1 of Drosophila melanogaster was heterologously expressed in a cell suspension culture of Nicotiana tabacum. This in vitro system was used to study the capability of CYP6G1 to metabolize the insecticide methoxychlor (=1,1,1-trichloro-2,2-bis(4-methoxyphenyl)ethane, 1) against the background of endogenous enzymes of the corresponding non-transgenic culture. The Cyp6g1-transgenic cell culture metabolized 96% of applied methoxychlor (45.8 microg per assay) within 24 h by demethylation and hydroxylation mainly to trishydroxy and catechol methoxychlor (16 and 17%, resp.). About 34% of the metabolism and the distinct formation of trishydroxy and catechol methoxychlor were due to foreign enzyme CYP6G1. Furthermore, methoxychlor metabolism was inhibited by 43% after simultaneous addition of piperonyl butoxide (458 microg), whereas inhibition in the non-transgenic culture amounted to 92%. Additionally, the rate of glycosylation was reduced in both cultures. These results were supported by the inhibition of the metabolism of the insecticide imidacloprid (6; 20 microg, 24 h) in the Cyp6g1-transgenic culture by 82% in the presence of piperonyl butoxide (200 microg). Due to CYP6G1 being responsible for imidacloprid resistance of Drosophila or being involved in DDT resistance, it is likely that CYP6G1 conveys resistance to methoxychlor (1). Furthermore, treating Drosophila with piperonyl butoxide could weaken the observed resistance phenomena.

Metabolomics approach to risk assessment: methoxyclor exposure in rats.

The primary objective of this study was to develop exposure biomarkers that "correlate with the endocrine-disrupting effects induced by methoxyclor (MTC), an organochlorine pesticide, using" urinary (1)H nuclear magnetic resonance (NMR) spectral data. Exposure biomarkers play an important role in risk assessment. MTC is an environmental endocrine disruptor with estrogenic, anti-estrogenic, and anti-androgenic properties. A new approach of proton nuclear magnetic resonance ((1)H NMR) urinalysis using pattern recognition was proposed for exposure biomarkers of MTC in female rats. The endocrine disruptor was expected to induce estrogenic effects in a dose dependent manner which, was confirmed by the uterotrophic assay. MTC [50, 100, or 200 m g/kg/d, orally (p.o.) or subcutaneously (s.c.)] was administered to ovariectomized female Sprague-Dawley (SD) rats for 3 d consecutively and urine was collected every 24 h. The animals were sacrificed 24 h after the last dose. All animals treated orally with MTC showed a significant increase in uterine and vaginal weight at all doses. However, in the s.c. route, only a high dose of 200 mg MTC/kg induced a significant increase in uterine and vaginal weight. (1)H NMR spectroscopy revealed evident separate clustering between pre- and post-treatment groups using global metabolic profiling through principal component analysis (PCA) and partial least square (PLS) discrimination analysis (DA) after different exposure routes. With targeted profiling, the endogenous metabolites of acetate, alanine, benzoate, lactate, and glycine were selected as putative exposure biomarkers for MTC. Data suggest that the proposed putative exposure biomarkers may be useful in a risk assessment of the endocrine-disrupting effects produced by MTC.

In silico description of differential enantioselectivity in methoxychlor O-demethylation by CYP2C enzymes.

Methoxychlor undergoes metabolism by cytochrome P450 (CYP) enzymes forming a chiral mono-phenolic derivative (Mono-OH-M) as main metabolite. In the current study, members of the CYP2C family were examined for their chiral preference in Mono-OH-M formation. CYP2C9 and CYP2C19 possessed high enantioselectivity favoring the formation of S-Mono-OH-M; CYP2C3 showed no enantioselectivity, whereas CYP2C5 slightly favored the formation of R-Mono-OH-M. Molecular modeling calculations were utilized in order to explain the observed differences in chiral preference of CYP2C enzymes. Molecular docking calculations could describe neither the existence of chiral preference in metabolism, nor the enantiomer which is preferentially formed. Molecular dynamic calculations were also carried out and were found to be useful for accurate description of chiral preference in biotransformation of methoxychlor by CYP2C enzymes. An in silico model capable of predicting chiral preference in cytochrome P450 enzymes in general can be developed based on the analysis of the stability and rigidity parameters of interacting partners during molecular dynamic simulation.

Ru(III)-based compounds with sulfur donor ligands: synthesis, characterization, electrochemical behaviour and anticancer activity.

In recent years, Ru(iii) complexes have emerged as a new class of effective anticancer agents against tumors that proved to be resistant to all other chemotherapeutic drugs currently in clinical use. To extend our previous studies on metal complexes containing sulfur-donor ligands, we report here on the synthesis and characterization, by means of several spectroscopic and analytical techniques, some [Ru(RSDT)(3)] and [Ru(2)(RSDT)(5)]Cl complexes with dithiocarbamato ligands derived from methyl/ethyl/tert-butyl esters of sarcosine. Their electrochemical behaviour was also studied by cyclic voltammetry. All the complexes were tested for their cytotoxicity on a panel of human tumor cell lines showing highly significant antitumor activity. The chemical and biological properties of the newly synthesized complexes, were compared with those of [Ru(DMDT)(3)] and [Ru(2)(DMDT)(5)]Cl species (DMDT = N,N-dimethyldithiocarbamate) whose chemical (not biological) characterization has been already reported in literature.

Interactions between CYP2C9 and CYP2C19 in reconstituted binary systems influence their catalytic activity: possible rationale for the inability of CYP2C19 to catalyze methoxychlor demethylation in human liver microsomes.

Previous studies in our laboratory showed that among cDNA-expressed human cytochrome P450 (P450) supersomes, CYP2C19 was the most active in methoxychlor-O-demethylation. However, based on the lack of inhibition of methoxychlor-O-demethylation by monoclonal anti-CYP2C19 antibodies in human liver microsomes (HLM), CYP2C19 did not seem to catalyze that reaction in HLM. By contrast, CYP2C9, much less active than CYP2C19 in supersomes, was the most active in HLM. The current study examines whether the lack of methoxychlor-O-demethylation by CYP2C19 in HLM was due to CYP2C19 exhibiting inferior competition for the NADPH-cytochrome P450 reductase (CPR) versus CYP2C9 and explores the interactions between CYP2C9 and CYP2C19 in a singular and binary complex of a reconstituted system. When reconstituted with CPR, cytochrome b(5), and lipid, purified CYP2C19 and CYP2C9 catalyzed methoxychlor-O-demethylation. However, whereas equimolar CPR to CYP2C9 supported maximal rates of methoxychlor demethylation and diclofenac hydroxylation, the rate of methoxychlor demethylation by CYP2C19 was not fully saturated, even with a 9-fold molar excess of CPR over CYP2C19. This behavior of CYP2C19 was also observed with S-mephenytoin as the substrate. When a binary reconstitution system was prepared by mixing CYP2C9 and CYP2C19 enzymes, methoxychlor-O-demethylation and S-mephenytoin hydroxylation by CYP2C19 were dramatically inhibited. Inhibition depended on the amount of CPR and substrate used. By contrast, in the incubation containing CYP2C9, diclofenac hydroxylation was activated by the presence of CYP2C19. These results show that interactions among P450 enzymes can modulate their catalytic rates, which depend on the substrate undergoing metabolism.

Phytoestrogen genistein and its pharmacological interactions with synthetic endocrine-active compounds.

Phytoestrogens are plant-derived compounds with estrogen-like activities. Certain foods such as soy-derived products are known to have high levels of phytoestrogens, and about 25% of commercial infant formulas used in the United States are soy-based. One of the most important phytoestrogens is the isoflavone genistein. Human exposures to genistein occur through normal dietary intake and through the use of genistein or other isoflavone extracts as nutritional supplements. Among the issues raising concerns about human exposure to phytoestrogens is how such exposure may affect responsiveness and sensitivity of the exposed subjects to other xenobiotics, particularly drugs and environmental chemicals with estrogenic or other endocrine activities. This article describes our recent studies on the developmental effects of dietary genistein in rats and its potential to interact with the toxicology of the endocrine-active pesticide methoxychlor. Data from our studies demonstrated that genistein is capable of altering the toxicological behaviors of methoxychlor and likely other endocrine active compounds as well. The complexities of such interactions are difficult to predict based on their in vitro steroid receptor reactivities.

A novel flexible approach for evaluating fixed ratio mixtures of full and partial agonists.

Assessing for interactions among chemicals in a mixture involves the comparison of actual mixture responses to those predicted under the assumption of zero interaction (additivity), based on individual chemical dose-response data. However, current statistical methods do not adequately account for differences in the shapes of the dose-response curves of the individual mixture components, as occurs with mixtures of full and partial receptor agonists. We present here a novel extension of current methods, which overcomes some of these limitations. Flexible single chemical concentration-effect curves combined with a common background parameter are used to describe an additivity surface along each axis. The predicted mixture response under the assumption of additivity is based on the constraint of Berenbaum's definition of additivity. Iterative algorithms are used to estimate mean responses at observed mixture combinations using only single chemical parameters. A full model allowing for different maximum response levels, different thresholds, and different slope parameters for each mixture component is compared to a reduced model under the assumption of additivity. A likelihood-ratio test is used to test the hypothesis of additivity by utilizing the full and reduced model predictions. This approach is useful for mixtures of chemicals with threshold regions and whose component chemicals exhibit differing response maxima (e.g., mixtures of full and partial agonists). The methods are illustrated with a combination of six chemicals in an estrogen receptor-alpha (ER-alpha) reporter gene assay.

Enantioselective recognition of mono-demethylated methoxychlor metabolites by the estrogen receptor.

Metabolites of methoxychlor such as 2-(p-hydroxyphenyl)-2-(p-methoxyphenyl)-1,1,1-trichloroethane (mono-OH-MXC) and 2,2-bis(p-hydroxyphenyl)-1,1,1-trichloroethane (bis-OH-MXC), have estrogenic activity. Mono-OH-MXC is a chiral compound in which the carbon atom bridging two benzene rings is the chiral centre. In previous studies the estrogenic activity of racemic mono-OH-MXC has been measured, and the activity of each enantiomer of this compound has not yet been elucidated. In this study, we evaluated the estrogen receptor-binding activity of each enantiomer of mono-OH-MXC to clarify the enantioselective recognition by the estrogen receptor. (S)-mono-OH-MXC showed 3-fold higher binding activity than that of the (R) enantiomer. The activity of bis-OH-MXC was only 1.7-fold higher than that of (S)-mono-OH-MXC. This result suggests that the one hydroxy group and the orientation of the CCl3 group of mono- and bis-OH-MXCs are important for the interaction with the estrogen receptor. The result also points out the estrogenic activity of methoxychlor after metabolic activation in vivo, which predominantly produces the (S)-mono-OH-MXC, may be higher than estimated from the in vitro activity of racemic mixtures.

In vitro metabolism of [14C]methoxychlor in rat, mouse, Japanese quail and rainbow trout in precision-cut liver slices.

1. The in vitro metabolism of [14C]methoxychlor (MXC) has been studied using precision-cut liver slices from the Sprague-Dawley male rat, CD-1 male mouse, WE strain male Japanese quail and juvenile rainbow trout (Oncorhynchus mykiss). The results demonstrated integrated phase I and II metabolism of MXC and species differences in the metabolic profiles were observed. 2. In rat liver slice preparations, MXC was rapidly metabolized to bis-OH-MXC by sequential O-demethylation followed by subsequent O-glucuronidation forming bis-OH-MXC glucuronide. No mono-OH-MXC glucuronide was detected. The doubly conjugated metabolite, bis-OH-MXC 4-O-sulphate 4'-O-glucuronide, was also detected as a rat-specific metabolite. 3. Formation of mono-OH-MXC and its glucuronide was the main metabolic pathway in the mouse and Japanese quail. In contrast to the rat, only minor amounts of bis-OH-MXC glucuronide were detected. A reductively dehalogenated metabolite, dechlorinated mono-OH-MXC glucuronide, was observed only in mouse preparations. 4. In rainbow trout, comparative amounts of both mono- and bis-OH-MXC glucuronide were formed as the major metabolites. Unconjugated forms of these metabolites were detected only as minor products. 5. The different metabolic profiles of MXC observed in the four animal species are possibly due to substrate specificity of contributing CYP450 monooxgenase enzyme(s) in different animal species.

Enantioselective metabolism of the endocrine disruptor pesticide methoxychlor by human cytochromes P450 (P450s): major differences in selective enantiomer formation by various P450 isoforms.

Methoxychlor, a currently used pesticide that in mammals elicits proestrogenic/estrogenic activity and reproductive toxicity, has been classified as a prototype endocrine disruptor. Methoxychlor is prochiral, and its metabolites 1,1,1-trichloro-2-(4-hydroxyphenyl)-2-(4-methoxyphenyl)ethane (mono-OH-M); 1,1,1-trichloro- 2-(4-methoxyphenyl)-2-(3, 4-dihydroxyphenyl)ethane (catechol-M); and 1,1,1-trichloro-2-(4-hydroxyphenyl)-2-(3, 4-dihydroxyphenyl)ethane (tris-OH-M) are chiral; whereas 1,1,1-trichloro-2, 2-bis(4-hydroxyphenyl)ethane (bis-OH-M) is achiral. These metabolites are formed during methoxychlor incubation with liver microsomes or recombinant cytochrome p450s (rp450s). Since methoxychlor-metabolite enantiomers may have different estrogenic/antiestrogenic/antiandrogenic activities than corresponding racemates, the possibility that p450s preferentially generate or use R or S enantiomers, was examined. Indeed, rCYP1A2 and r2A6 mono-demethylated methoxychlor primarily into (R)-mono-OH-M at 91 and 75%, respectively, whereas rCYP1A1, 2B6, 2C8, 2C9, 2C19, and 2D6 formed the (S)-enantiomer at 69, 66, 75, 95, 96, and 80%, respectively. However, rCYP3A4, 3A5, and 2B1(rat) weakly demethylated methoxychlor without enantioselectivity. Human liver microsomes generated (S)-mono-OH-M (77-87%), suggesting that CYP1A2 and 2A6 display only minor catalytic contribution. P450 inhibitors demonstrated that CYP2C9 and possibly 2C19 are major hepatic catalysts forming (S)-mono-OH-M, and CYP1A2 is primarily involved in forming the (R)-mono-OH-M. Demethylation rate of (S)-mono-OH-M versus (R)-mono-OH-M forming achiral bis-OH-M by rCYP1A2 was 97/3, compared with 15/85 and 17/83 for rCYP2C9 and 2C19, respectively, indicating opposite substrate enantioselectivity of rCYP1A2 versus 2C9 and 2C19. Also, rCYP1A2 preferentially O-demethylated (R)-catechol-M into (R)-tris-OH-M (at 80%), contrasting r2C9 and r2C19 that yielded (S)-tris-OH-M at 80 and 77%, respectively. Ortho-hydroxylation of mono-OH-M into catechol-M and bis-OH-M into tris-OH-M was primarily by 3A4 and was not enantioselective. In conclusion, enantiomeric abundance of methoxychlor metabolites depends on the relative catalytic activity of the hepatic p450 isoforms.

Interaction of methoxychlor and related compounds with estrogen receptor alpha and beta, and androgen receptor: structure-activity studies.

We previously demonstrated differential interactions of the methoxychlor metabolite 2,2-bis(p-hydroxyphenyl)-1,1, 1-trichloroethane (HPTE) with estrogen receptor alpha (ERalpha), ERbeta, and the androgen receptor (AR). In this study, we characterize the ERalpha, ERbeta, and AR activity of structurally related methoxychlor metabolites. Human hepatoma cells (HepG2) were transiently transfected with human ERalpha, ERbeta, and AR plus an appropriate steroid-responsive luciferase reporter vector. After transfection, cells were treated with various concentrations of HPTE or structurally related compounds in the presence (for detecting antagonism) and absence (for detecting agonism) of 17beta-estradiol and dihydrotestosterone. The monohydroxy analog of methoxychlor, as well as monohydroxy and dihydroxy analogs of 2, 2-bis(p-hydroxyphenyl)-1,1-dichloroethylene, had ERalpha agonist activity and ERbeta and AR antagonist activity similar to HPTE. The trihydroxy metabolite of methoxychlor displayed only weak ERalpha agonist activity and did not alter ERbeta or AR activities. Replacement of the trichloroethane or dichloroethylene group with a methyl group resulted in a compound with ERalpha and ERbeta agonist activity that retained antiandrogenic activities. This study identifies some of the structural requirements for ERalpha and ERbeta activity and demonstrates the complexity involved in determining the mechanism of action of endocrine-active chemicals that simultaneously act as agonists or antagonists through one or more hormone receptors.

Chloride ions formation during degradation of organochloride compounds using TiO2 deposited on glass microspheres.

Aqueous solutions containing 200 mg/dm3 of p,p'-DDT and methoxychlor were photodegraded for 60 min in UV/TiO2/O2 system and chloride ions concentration and pH were measured. From 60 to 80% of the investigated pesticides were eliminated after treatment. Over 27% of chlorine atoms were splitted off for methoxychlor and 10% for p,p'-DDT. The experimental data suggest, that chlorine atoms were removed from the -CCl3 moiety but the chlorine atoms bound to aromatic ring were left intact at this step of photodegradation.