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Biochemistry Mar 2019Organophosphorus flame retardants are stable toxic compounds used in nearly all durable plastic products and are considered major emerging pollutants. The...
Organophosphorus flame retardants are stable toxic compounds used in nearly all durable plastic products and are considered major emerging pollutants. The phosphotriesterase from Sphingobium sp. TCM1 ( Sb-PTE) is one of the few enzymes known to be able to hydrolyze organophosphorus flame retardants such as triphenyl phosphate and tris(2-chloroethyl) phosphate. The effectiveness of Sb-PTE for the hydrolysis of these organophosphates appears to arise from its ability to hydrolyze unactivated alkyl and phenolic esters from the central phosphorus core. How Sb-PTE is able to catalyze the hydrolysis of the unactivated substituents is not known. To interrogate the catalytic hydrolysis mechanism of Sb-PTE, the pH dependence of the reaction and the effects of changing the solvent viscosity were determined. These experiments were complemented by measurement of the primary and secondary 18-oxygen isotope effects on substrate hydrolysis and a determination of the effects of changing the p K of the leaving group on the magnitude of the rate constants for hydrolysis. Collectively, the results indicated that a single group must be ionized for nucleophilic attack and that a separate general acid is not involved in protonation of the leaving group. The Brønsted analysis and the heavy atom kinetic isotope effects are consistent with an early associative transition state with subsequent proton transfers not being rate limiting. A novel binding mode of the substrate to the binuclear metal center and a catalytic mechanism are proposed to explain the unusual ability of Sb-PTE to hydrolyze unactivated esters from a wide range of organophosphate substrates.
Topics: Bacterial Proteins; Catalysis; Catalytic Domain; Deuterium; Hydrogen-Ion Concentration; Hydrolysis; Kinetics; Magnetic Resonance Spectroscopy; Organophosphates; Paraoxon; Phosphoric Triester Hydrolases; Solvents; Sphingomonadaceae; Viscosity
PubMed: 30730705
DOI: 10.1021/acs.biochem.9b00041 -
Biochemia Medica Feb 2024Paraoxonase 1 (PON1) is the enzyme that removes carcinogenic radicals from lipids. The aim of the study was to investigate the differences in PON1 activity and oxidation...
INTRODUCTION
Paraoxonase 1 (PON1) is the enzyme that removes carcinogenic radicals from lipids. The aim of the study was to investigate the differences in PON1 activity and oxidation stress parameters between patients with cervical intraepithelial neoplasia (CIN) and healthy controls.
MATERIALS AND METHODS
The study included 65 women with CIN and 109 healthy women. Lipid parameters were determined on Cobas Integra 400 plus (Roche, Mannheim, Germany). Tiols and reduced glutathione (GSH) were determined spectrophotometric using Eliman reagent. Activity of PON1 was assessed with two substrates, paraoxon and phenylacetate by spectrophotometric method. Malondialdehyde (MDA) was determined by high performance liquid chromatography (Shimadzu Corporation, Kyoto, Japan). Mann-Whitney-test, t-test, χ2-test, correlation and logistic regression was used in statistical analysis. P < 0.05 was considered statistically significant.
RESULTS
The basal (P = 0.929) and NaCl-stimulated (P = 0.985) PON1 activity and activities standardised on the concentration of high-density lipoprotein (HDL; P = 0.076; P = 0.065, respectively) and apolipoprotein AI (apo AI; P = 0.444; P = 0.499, respectively) as well as PON1 phenotypes (P = 0.842) did not differ significantly between the groups. The PON1 arylesterase activity (53±19 kU/L vs. 77±17 kU/L; P < 0.001) and HDL-standardized activity (37 (28-44) kU/mmol . 43 (37-50) kU/mmol; P < 0.001) and apoAI (29±11 kU/g . 44±11 kU/g; P < 0.001) was significantly reduced in the CIN group. The concentration of the thiol groups was similar (P = 0.519), of MDA was lower (0.39 (0.27-0.55) µmol/L . 0.76 (0.57-1.15) µmol/L; P < 0.001) and of GSH was higher (112.0 (66.0-129.6) µg/mL . 53.4 (34.8-134.4) µg/mL; P < 0.001) in the CIN group.
CONCLUSION
Reduced PON1 arylesterase activity, lower MDA and higher GSH concentration were observed in CIN patients.
Topics: Humans; Female; Aryldialkylphosphatase; Carboxylic Ester Hydrolases; Oxidative Stress; Uterine Cervical Dysplasia
PubMed: 38125616
DOI: 10.11613/BM.2024.010701 -
Pesticide Biochemistry and Physiology Mar 2022Although the toxic effects of organophosphorus (OP) pesticides have been classically attributed to inhibition of the acetylcholinesterase, other neurotoxic mechanisms,...
Although the toxic effects of organophosphorus (OP) pesticides have been classically attributed to inhibition of the acetylcholinesterase, other neurotoxic mechanisms, as oxidative stress can also occur. Here we evaluated if antioxidants prevent the excessive dopamine release induced by OP pesticides in conscious and freely moving rats, using cerebral microdialysis technique. Intrastriatal infusion of paraoxon (5 mM), glufosinate (10 mM) or glyphosate (5 mM) significantly increased the dopamine release (1006 ± 106%, 991 ± 142%, and 1164 ± 128%, relative to baseline, respectively). To evaluate if these increased dopamine release could be related to oxidative stress, we pretreated animals with antioxidants glutathione (GSH, 400 or 800 μM), dithiothreitol (DTT, 5 or 10 μM), trolox (1 or 3 mM), and α-lipoic acid (ALA, 400 or 800 μM) before administration of OP pesticides. Intrastriatal administration of the antioxidants GSH, DTT, trolox, and ALA was highly effective in preventing the glyphosate and glufosinate-induced dopamine overflow. However, only GSH (800 μM) significantly decreased the effect of paraoxon on dopamine levels. The high toxicity of this pesticide and the low concentrations used could explain this lack of effect in our experimental conditions. The fact that ROS scavengers prevent the excessive dopamine release induced by OP pesticides, further supports the view that dopamine overflow can cause neuronal damage mediated, at least in part, by oxidative stress.
Topics: Acetylcholinesterase; Animals; Antioxidants; Dopamine; Organophosphorus Compounds; Pesticides; Rats; Rats, Sprague-Dawley
PubMed: 35249645
DOI: 10.1016/j.pestbp.2022.105035 -
ACS Omega Feb 2022Graphene-enhanced Raman scattering (GERS) produces enhancement of the Raman signal, which is based on chemical rather than electromagnetic mechanism such as in the...
Graphene-enhanced Raman scattering (GERS) produces enhancement of the Raman signal, which is based on chemical rather than electromagnetic mechanism such as in the surface-enhanced Raman scattering. Graphene oxide, amino- and guanidine-functionalized graphene oxide, exfoliated graphene, and commercial graphene nanoplatelets have been used to investigate the GERS response with the change of graphene properties. Different graphene nanostructures have been embedded into organic-inorganic microporous films to build a platform for the fast and sensitive detection of pesticides in water. The graphene nanostructures vary in the number of layers, lateral size, degree of oxidation, and surface functionalization. The GERS performances of the graphene nanostructures cast on silicon substrates and embedded in the nanocomposite films have been comparatively evaluated. After casting a few droplets of the pesticide aqueous solution on the graphene nanostructures, the Raman band enhancements of the analytes have been measured. In the nanocomposite films, the characteristic Raman bands originating from pesticides such as paraoxon, parathion, and glyphosate could be traced at concentrations below 10, 10, and 10 M, respectively. The results show that the surface functionalization reduces the GERS effect because it increases the ratio between the sp carbon and sp carbon. On the other hand, the comparison among different types of graphenes shows that the monolayers are more efficient than the few-layer nanostructures in enhancing the Raman signal.
PubMed: 35224328
DOI: 10.1021/acsomega.1c04863 -
Advanced Science (Weinheim,... Jan 2020Organophosphate nerve agents rapidly inhibit cholinesterases thereby destroying the ability to sustain life. Strong nucleophiles, such as oximes, have been used as...
Organophosphate nerve agents rapidly inhibit cholinesterases thereby destroying the ability to sustain life. Strong nucleophiles, such as oximes, have been used as therapeutic reactivators of cholinesterase-organophosphate complexes, but suffer from short half-lives and limited efficacy across the broad spectrum of organophosphate nerve agents. Cholinesterases have been used as long-lived therapeutic bioscavengers for unreacted organophosphates with limited success because they react with organophosphate nerve agents with one-to-one stoichiometries. The chemical power of nucleophilic reactivators is coupled to long-lived bioscavengers by designing and synthesizing cholinesterase-polymer-oxime conjugates using atom transfer radical polymerization and azide-alkyne "click" chemistry. Detailed kinetic studies show that butyrylcholinesterase-polymer-oxime activity is dependent on the electrostatic properties of the polymers and the amount of oxime within the conjugate. The covalent coupling of oxime-containing polymers to the surface of butyrylcholinesterase slows the rate of inactivation of paraoxon, a model nerve agent. Furthermore, when the enzyme is covalently inhibited by paraoxon, the covalently attached oxime induced inter- and intramolecular reactivation. Intramolecular reactivation will open the door to the generation of a new class of nerve agent scavengers that couple the speed and selectivity of biology to the ruggedness and simplicity of synthetic chemicals.
PubMed: 31921563
DOI: 10.1002/advs.201901904 -
Archives of Medical Science : AMS Jun 2017Paraoxonase (PON1) is an enigmatic enzyme with multiple enzymatic properties including arylesterase and lactonase activities besides its ability to hydrolyze the toxic...
INTRODUCTION
Paraoxonase (PON1) is an enigmatic enzyme with multiple enzymatic properties including arylesterase and lactonase activities besides its ability to hydrolyze the toxic metabolite of parathion, paraoxon. The aim of this study was to determine the phenotype distribution of PON1 in patients with cardiac disease who were classified in coronary artery bypass grafting (CABG), heart valve disease (HVD), heart failure (HF) and ST elevation myocardial infarction (STEMI) groups and healthy subjects as a control group.
MATERIAL AND METHODS
A total of 300 people (100 cardiac surgery (70 CABG and 30 HVD), 70 HF, 30 STEMI patients and 100 healthy controls) were admitted to this study. Individual variations in PON1 were determined using the dual substrate (paraoxon and phenylacetate) method.
RESULTS
The following phenotype distributions were found in the cardiac disease and control groups: cardiac disease group ( = 200): 48.5% (QQ), 42.5% (QR), 9% (RR) and control group ( = 100): 58% (QQ), 39% (QR), 3% (RR). RR (high activity) phenotypic distribution was more common in the cardiac disease group than in controls ( = 0.04). In particular, the frequency of the RR phenotype was two- to three-fold higher in the STEMI and HF patients compared to the controls as well as CABG and HVD groups.
CONCLUSIONS
We found a higher percentage of RR phenotype in STEMI and HF patients compared to a large control group as well as compared to two other groups of cardiac disease patients.
PubMed: 28721150
DOI: 10.5114/aoms.2016.59674 -
Annals of the New York Academy of... Nov 2020Organophosphorus (OP) compounds are chemical threat agents and are irreversible inhibitors of the enzyme acetylcholinesterase that lead to a hypercholinergic response... (Comparative Study)
Comparative Study
Organophosphorus (OP) compounds are chemical threat agents and are irreversible inhibitors of the enzyme acetylcholinesterase that lead to a hypercholinergic response that could include status epilepticus (SE). SE particularly targets the heart and brain and despite existing therapies, it is still associated with significant mortality and morbidity. Here, we investigated the effect of intramuscular (i.m.) adjunct therapy consisting of atenolol (AT) and levetiracetam (LV) when administered after paraoxon (POX)-induced SE. The combination therapy was administered twice daily for 2, 7, or 14 days. POX exposure in rats produced rapid SE onset that was treated with atropine, pralidoxime chloride, and midazolam. Here, AT + LV therapy produced significant reductions in POX SE mortality assessed at 30 days post-SE. AT + LV therapy exhibited muscle pathology inflammation scores that were not significantly different from saline-treated controls. Pharmacokinetic analyses revealed that the i.m. route achieved faster and stabler plasma therapeutic levels for both AT and LV under OP SE conditions compared with oral administrations. Our data provide evidence of the safety and efficacy of i.m. AT + LV therapy for reducing mortality following POX SE.
Topics: Administration, Oral; Animals; Atenolol; Injections, Intramuscular; Levetiracetam; Male; Paraoxon; Rats; Rats, Sprague-Dawley; Status Epilepticus
PubMed: 32961584
DOI: 10.1111/nyas.14500 -
Journal of Hazardous Materials Oct 2015Organophosphorus insecticides have been widely used, which are highly poisonous and cause serious concerns over food safety and environmental pollution. A bacterial...
Organophosphorus insecticides have been widely used, which are highly poisonous and cause serious concerns over food safety and environmental pollution. A bacterial strain being capable of degrading O,O-dialkyl phosphorothioate and O,O-dialkyl phosphate insecticides, designated as G1, was isolated from sludge collected at the drain outlet of a chlorpyrifos manufacture plant. Physiological and biochemical characteristics and 16S rDNA gene sequence analysis suggested that strain G1 belongs to the genus Stenotrophomonas. At an initial concentration of 50 mg/L, strain G1 degraded 100% of methyl parathion, methyl paraoxon, diazinon, and phoxim, 95% of parathion, 63% of chlorpyrifos, 38% of profenofos, and 34% of triazophos in 24 h. Orthogonal experiments showed that the optimum conditions were an inoculum volume of 20% (v/v), a substrate concentration of 50 mg/L, and an incubation temperature in 40 °C. p-Nitrophenol was detected as the metabolite of methyl parathion, for which intracellular methyl parathion hydrolase was responsible. Strain G1 can efficiently degrade eight organophosphorus pesticides (OPs) and is a very excellent candidate for applications in OP pollution remediation.
Topics: Biodegradation, Environmental; Organophosphorus Compounds; Pesticides; RNA, Ribosomal, 16S; Sewage; Stenotrophomonas; Water Pollutants, Chemical
PubMed: 25938642
DOI: 10.1016/j.jhazmat.2015.04.052 -
Veterinary Clinical Pathology Mar 2018Paraoxonase-1 (PON-1) is an antioxidant compound that is considered a negative acute phase protein. No information on the analytic performance of the paraoxon method for...
BACKGROUND
Paraoxonase-1 (PON-1) is an antioxidant compound that is considered a negative acute phase protein. No information on the analytic performance of the paraoxon method for measuring PON-1 in horse serum is available.
OBJECTIVES
The aim of this study was to validate a paraoxon-based method to measure PON-1 in horses and to establish RIs in healthy horses and foals.
METHODS
Horses and foals classified as healthy after physical examination and routine biochemistry were used in the study. Serum PON-1 activity was measured with an automated spectrophotometer and an enzymatic method validated in other species. After the analytic validation (precision, accuracy, interference studies), RIs were determined using the Reference Value Advisor software. The possible sex-, age-, and breed-related differences were statistically investigated.
RESULTS
The healthy study population included 120 horses and 55 foals. The paraoxon-based method was precise (CVs < 4.0%) and accurate (P < .001 in linearity under dilution and spike-recovery testing) but was affected by interference from mild bilirubinemia, severe lipemia, and hemoglobinemia. The RIs recorded in the whole population were 38.1-80.8 U/mL. According to the Harris and Boyd test, it would be advisable to use separate RIs only for adult females and for Warmblood and Trotter adults.
CONCLUSIONS
This study demonstrated that the analytic performances of the paraoxon-based method for measurement of PON-1 in horses are acceptable. The PON-1 activity is lower in horses than in other domestic species. These results may provide a basis for further studies designed to establish whether healthy and sick horses can be correctly classified by using the PON-1 assay.
Topics: Animals; Aryldialkylphosphatase; Blood Chemical Analysis; Female; Horses; Male; Reference Values
PubMed: 29575140
DOI: 10.1111/vcp.12562 -
Laboratory Animal Research Jun 2024The aim of the study was to develop a technique for quantitative determination of rat urine metabolites by HPLC-MS/MS, which can be used to search for biomarkers of...
BACKGROUND
The aim of the study was to develop a technique for quantitative determination of rat urine metabolites by HPLC-MS/MS, which can be used to search for biomarkers of acute intoxication with organophosphates (OPs).
RESULTS
The content of metabolites in the urine of rats exposed to a single dose of paraoxon (POX1x); interval, twice daily administration of paraoxon (POX2x); exposure to 2-(o-cresyl)-4H-1, 3, 2-benzodioxaphosphorin-2-oxide and paraoxon (CBPOX) was investigated. New data were obtained on the content in the urine of intact rats as well as rats in 3 models of OP poisoning: 3-methylhistidine, threonine, creatine, creatinine, lactic acid, acetylcarnitine, inosine, hypoxanthine, adenine, 3-hydroxymethyl-butyrate and 2-hydroxymethyl-butyrate.
CONCLUSIONS
The proposed assay procedure is a simple and reliable tool for urine metabolomic studies. Within 1-3 days after OP exposure in all three models of acute intoxication, the concentration of metabolites in rat urine, with the exception of adenine, changes similarly and symmetrically, regardless of the method of poisoning modeling, in all three models of acute intoxication. Further studies are needed to determine the specificity and reliability of using urinary metabolite concentration changes as potential biomarkers of acute organophosphate intoxication.
PubMed: 38845041
DOI: 10.1186/s42826-024-00209-3