Efficient and harmless removal of insecticide diazinon via the stepwise combination of biodegradation and photodegradation.

Diazinon, an organophosphorus insecticide, is predominantly removed through photodegradation and biodegradation in the environment. However, photodegradation can generate diazoxon, a highly toxic oxidation byproduct, while biodegradation is hard to complete mineralize diazinon, showing limitations in both methods. In this study, we provided an efficient strategy for the complete and harmless removal of diazinon by synergistically employing biodegradation and photodegradation. The diazinon-degrading strain X1 was capable of completely degrading 200 µM of diazinon into 2-isopropyl-6-methyl-4-pyrimidinol (IMP) within 6 h without producing the highly toxic diazoxon. IMP was the only intermediate metabolite in biodegradation process, which cannot be further degraded by strain X1. Through RT-qPCR and prokaryotic expression analyses, the hydrolase OpdB was pinpointed as the key enzyme for diazinon degradation in strain X1. Photodegradation was further used to degrade IMP and a pyridazine ring-opening product of IMP was identified via high resolution mass spectrometry. The acute toxicity of this product to aquatic organisms were 123 times and 6630 times lower than that of diazinon and IMP, respectively. The stepwise application of biodegradation and photodegradation was proved to be a successful approach for the remediation of diazinon and its metabolite IMP. This integrated method ensures the harmless and complete elimination of diazinon and IMP within only 6 h. The research provides a theoretical basis for the efficient and harmless remediation of organophosphorus insecticide residuals in the environment.

In-vitro effect of diazoxon, a metabolite of diazinon, on proliferation, signal transduction, and death induction in mononuclear cells of Nile tilapia fish (Oreochromis niloticus).

The immune response of teleosts (bonefish) is altered by diazinon (DZN), an organophosphate pesticide. It has been suggested that such alteration is due to the extraneuronal cholinergic system in fish leukocytes that renders these cells a target of pesticides. Diazoxon (DZO), the oxon metabolite of DZN, has been attributed immunotoxic effects. Still, to date there are no reports on the effects of DZO upon parameters involved in the signaling cascade of immune response cells. Therefore, this work evaluated the effect of DZO on key parameters of cell signaling (intracellular Ca2+ flux, ERK 1/2 phosphorylation), cell proliferation, and antiproliferative processes (apoptosis, senescence, mitochondrial membrane potential) in spleen mononuclear cells of Nile tilapia fish. The results obtained show that DZO does not affect cell proliferation but causes a lack of response to stimulation with PMA and ionomycin to release intracellular calcium. In addition, it inhibits ERK 1/2 phosphorylation and causes loss of mitochondrial membrane potential, apoptosis, and senescence. These results suggest that the lack of cell response to release intracytoplasmic Ca2+ inhibits ERK which disrupts the mitochondrial membrane potential, leading to cell apoptosis and senescence. These findings prove that DZO significantly affects key parameters involved in the survival of immune response cells.

Diazinon and diazoxon impair the ability of astrocytes to foster neurite outgrowth in primary hippocampal neurons.

Evidence from in vivo and epidemiological studies suggests that organophosphorus insecticides (OPs) are developmental neurotoxicants, but possible underlying mechanisms are still unclear. Astrocytes are increasingly recognized for their active role in normal neuronal development. This study sought to investigate whether the widely-used OP diazinon (DZ), and its oxygen metabolite diazoxon (DZO), would affect glial-neuronal interactions as a potential mechanism of developmental neurotoxicity. Specifically, we investigated the effects of DZ and DZO on the ability of astrocytes to foster neurite outgrowth in primary hippocampal neurons. The results show that both DZ and DZO adversely affect astrocyte function, resulting in inhibited neurite outgrowth in hippocampal neurons. This effect appears to be mediated by oxidative stress, as indicated by OP-induced increased reactive oxygen species production in astrocytes and prevention of neurite outgrowth inhibition by antioxidants. The concentrations of OPs were devoid of cytotoxicity, and cause limited acetylcholinesterase inhibition in astrocytes (18 and 25% for DZ and DZO, respectively). Among astrocytic neuritogenic factors, the most important one is the extracellular matrix protein fibronectin. DZ and DZO decreased levels of fibronectin in astrocytes, and this effect was also attenuated by antioxidants. Underscoring the importance of fibronectin in this context, adding exogenous fibronectin to the co-culture system successfully prevented inhibition of neurite outgrowth caused by DZ and DZO. These results indicate that DZ and DZO increase oxidative stress in astrocytes, and this in turn modulates astrocytic fibronectin, leading to impaired neurite outgrowth in hippocampal neurons.

Subchronic neurotoxicity of diazinon in albino mice: Impact of oxidative stress, AChE activity, and gene expression disturbances in the cerebral cortex and hippocampus on mood, spatial learning, and memory function.

Diazinon (DZN) with prominent neurotoxic effects perturbs CNS function via multiple mechanisms. This investigation intends to explore mood, spatial learning, and memory dysfunction, acetylcholine esterase (AChE) activity, and neurodegeneration-related gene expression in the cortex and hippocampus regions of mice exposed to DZN for 63 consecutive days (subchronic exposure). Adult male albino mice were orally given sublethal DZN (DZNL = 0.1 mg/kg, DZNM = 1 mg/kg and DZNH = 10 mg/kg). All mice in the DZNH group died within 3 weeks postexposure. DZNL and DZNM caused body and brain weight loss (p < 0.05). Completing 9 weeks of DZN exposure, a marked decline in AChE activity and oxidative stress level was indicated in both brain regions (p < 0.05). Also, synaptophysin, vesicular acetylcholine transferase, and glutamate decarboxylase gene expressions were affected in both brain regions (p < 0.05). Furthermore, the present study revealed that DZN administration increased anxiety and depressive-like behaviors (p < 0.0001). Spatial learning and short- and long-memory were severely affected by DZNL and DZNM treatments (p < 0.0001). Taken together, subchronic exposure to low and medium doses of DZN can cause AChE inhibition, oxidative damage, and neurotransmitter disturbances in brain cells and induce neurodegeneration. These changes would impair mood, spatial learning, and memory function.

Unexpectedly higher diazoxon hydrolysis by serum paraoxonase-1 in coronary heart disease.

OBJECTIVES: Low serum PON1 activities (paraoxon, phenyl-acetate or lactone substrates) are associated with coronary heart disease (CHD). We investigated the rate of diazoxon hydrolysis by PON1 in a population with CHD. DESIGN & METHODS: Case- control study of 410 subjects with CHD and 274 controls. PON1 activity towards paraoxon and diazoxon, PON1 serum concentration and the PON1-55 and 192 polymorphisms were determined. RESULTS: There were no differences in the distribution of the PON1-55 or PON1-192 genotypes between the CHD and controls, however, PON1 activity towards diazoxon (DIAZ) was significantly (+160%) higher in CHD. In the control population, DIAZ was significantly different between the PON1-192 genotypes in the order QQ > QR > RR (P < .001). However, in CHD the order was QQ > QR = RR. In CHD DIAZ was significantly higher in all the PON1-192 and 55 genotypes compared to controls. In both populations DIAZ was significantly different between the PON1-55 genotypes in the order LL > LM > MM (P < .001). CONCLUSION: If this result can be replicated in other studies and/or with other PON1 substrates, there may be major diagnostic and mechanistic implications for the relationship of PON1 and CHD.

Astrocytes protect against diazinon- and diazoxon-induced inhibition of neurite outgrowth by regulating neuronal glutathione.

Evidence demonstrating that human exposure to various organophosphorus insecticides (OPs) is associated with neurobehavioral deficits in children continues to emerge. The present study focused on diazinon (DZ) and its active oxygen metabolite, diazoxon (DZO), and explored their ability to impair neurite outgrowth in rat primary hippocampal neurons as a mechanism of developmental neurotoxicity. Both DZ and DZO (0.5-10 µM) significantly inhibited neurite outgrowth in hippocampal neurons, at concentrations devoid of any cyototoxicity. These effects appeared to be mediated by oxidative stress, as they were prevented by antioxidants (melatonin, N-t-butyl-alpha-phenylnitrone, and glutathione ethyl ester). Inhibition of neurite outgrowth was observed at concentrations below those required to inhibit the catalytic activity of acetylcholinesterase. The presence of astrocytes in the culture was able to provide protection against inhibition of neurite outgrowth by DZ and DZO. Astrocytes increased neuronal glutathione (GSH) in neurons, to levels comparable to those of GSH ethyl ester. Astrocytes depleted of GSH by L-buthionine-(S,R)-sulfoximine no longer conferred protection against DZ- and DZO-induced inhibition of neurite outgrowth. The findings indicate that DZ and DZO inhibit neurite outgrowth in hippocampal neurons by mechanisms involving oxidative stress, and that these effects can be modulated by astrocytes and astrocyte-derived GSH. Oxidative stress from other chemical exposures, as well as genetic abnormalities that result in deficiencies in GSH synthesis and regulation, may render individuals more susceptible to these developmental neurotoxic effects of OPs.