Antidotal Action Research Articles

Cyanide and Cyanogenic Compounds—Toxicity, Molecular Targets, and Therapeutic Agents

Cyanide (CN) is a well-known mitochondrial poison. CN poisoning may result from acute or long-term exposure to a number of CN compounds. Recent insight into the chemical affinities of the CN anion has increased our understanding of its toxicity and the mechanisms of antidotal actions, which, together with information on various exposure sources, are reviewed in the present article. A literature search in Scopus, Embase, Web of Science, PubMed, and Google Scholar for the period 2001–2024 revealed that the CN anion after exposure or degradation of CN compounds is distributed to vulnerable copper and iron-containing targets, especially in mitochondria, thus blocking the electron transport chain. Intake of cyanogenic compounds may exert subacute or chronic toxic effects, also because of the interaction with cobalt in vitamin B12. Antidotal agents exert their effects through the affinity of CN for cobalt- or iron-containing compounds. Research on CN interactions with metalloproteins may increase our insight into CN toxicity and efficient antidotal regimens.

Evaluating the Antidote Action of Nimbuk Swarasa and Peya in Jayapala Toxicity: A Critical Review

Jayapala (Myristica fragrans), a plant-based toxin, is recognized for its therapeutic importance despite its toxic properties. Improper administration or insufficient purification can lead to adverse effects, including gastrointestinal irritation, salivation, and even death. The management of Jayapala poisoning is well-documented in classical Ayurvedic texts, with specific antidotes such as Nimbuk Swarasa (lemon juice) and Peya (rice water) being highlighted. This review critically evaluates the mode of action of Nimbuk Swarasa and Peya in Jayapala poisoning, to establish their therapeutic efficacy and practical applicability as antidotes. The study highlights the antidotal potential of Nimbuk Swarasa and Peya in enhancing digestive function, alleviating symptoms, and providing a cost-effective solution for managing Jayapala toxicity. The findings indicate that these preparations not only mitigate harmful effects but also embody a practical approach to addressing accidental poisoning

Antidotal Effects of the Antihistamine Diphenhydramine Against Cholinesterase Inhibitor Poisoning: A Meta-Analysis of Median Lethal Doses in Experimental Animals.

The H1-antihistamine diphenhydramine antagonizes cholinesterase inhibitor poisoning in various animal species. One aspect of acute antidotal actions of diphenhydramine is increasing the median lethal doses (LD50) of toxicants. The objective of this meta-analysis was to assess the antidotal action of diphenhydramine against short-term toxicity (LD50) of cholinesterase inhibitors in experimental animals. The experimental studies selected were according to the Preferred Reporting Items for Systematic Reviews and Meta-Analysis (PRISMA) guidelines. They were conducted in laboratory animals (mice, rats, and chicks) to determine acute LD50 values of cholinesterase inhibitors (organophosphates, carbamates, and imidocarb) under the influence of diphenhydramine vs. controls. Twenty-eight records were selected from 12 studies on mice (n= 242), rats (n= 27), and young chicks (n= 128). The forest plot of randomized two-group meta-analysis assessed effect size, subgroup analysis, drapery prediction, heterogeneity, publication bias-funnel plot as well as one-group proportions meta-analysis of percent protection. Diphenhydramine significantly increased the combined effect size (i.e. increased LD50) in intoxicated experimental animals in comparison to controls (-3.71, standard error (SE) 0.36, 95%CI -4.46, -2.97). The drapery plot proposed a wide range of confidence intervals. The I2 index of heterogeneity of the combined effect size was high at 81.03% (Q= 142.3, p < 0.0001). Galbraith regression also indicated data heterogeneity; however, the normal quantile plot indicated no outliers. Subgroup analysis indicated significantly high heterogeneity with organophosphates (I2 = 63.72%) and carbamates (I2 = 76.41%), but low with imidocarb (I2 = 51.48%). The funnel plot and Egger regression test (t= -13.7, p < 0.0001) revealed publication bias. The median of the diphenhydramine protection ratio was 1.655, and the related forest plot of one group proportion meta-analysis revealed a statistically high level of protection (0.594, SE0.083, 95%CI 0.432, 0.756), withhigh heterogeneity (I2= 99.86). The risk of bias assessment was unclear, while the total score (16 out of 20) of each study leaned towards the side of the low risk of bias. In conclusion, the meta-analysis of LD50 values indicated that diphenhydramine unequivocally protected experimental animals from the acute toxicity of cholinesterase inhibitors. The drug could be an additional antidote against acute poisoning induced by cholinesterase inhibitors, but a word of caution: it is not to be considered as a replacement for the standard antidote atropine sulfate. Further studies are needed to examine the action of diphenhydramine on adverse chronic effects of cholinesterase inhibitors.

Pharmacognostical and Pharmaceutical Exploration of the traditional medicine Ipomoea carnea Jacq.

Background: Ipomoea carnea Jacq is an ethnopharmacological drug and its leaves, roots and latex are used for various indications by traditional healers. But seeds are not used in the treatment as they are more toxic. As the herbal medicines are becoming endangered, ethnopharmacological drug can be used efficiently as they are available abundantly. But the toxicity should be reduced for safer use in the treatment. Aim: Seeds of Ipomoea carnea Jacq are studied in the present study to observe the macroscopic, microscopic, physicochemical and phytochemical changes in seeds after detoxification in different media. Methods and Materials: Detoxification method described in Ayurveda was used. Ipomoea carnea seeds were subjected to boiling for three hours with Cow’s milk. The same procedure was followed for another sample separately with Cow urine, sour gruel, Triphala (Emblica officinalis, Terminalia chebula and Terminalia bellirica) decoction and Distilled water as media. One raw sample was kept for comparison. The transverse section of the raw seed was studied. Microscopy of powder, physicochemical and phytochemical analysis of all the six samples was conducted and the observations were compared among all the samples. Results: The toxic principle Swainsonine was estimated as 3% in all the samples at retention time 3 minutes. The numbers of peaks were reduced in all the treated samples except DWSIC in comparison with raw sample and 2-4 peaks are observed at retention time 1-3 minutes in all treated samples. A new phytoamine having concentration 12% was detected at retention time 17 minutes in all (crude and purified) samples. Conclusion: The morpho-anatomical characteristics of the seed remain same even after being treated with different liquid media. The changes observed in HPLC may be due to antagonist and antidotal action of media used for detoxification.

First meta-analysis study of cholinesterase inhibition in experimental animals by organophosphate or carbamate insecticides under the influence of diphenhydramine.

Diphenhydramine is an H1-antihistamine that counteracts the toxic effects of organophosphate and carbamate insecticides that inhibit cholinesterase (ChE) activity. This meta-analysis aimed to investigate the effects of diphenhydramine on ChE inhibition induced by these insecticides in the plasma, erythrocytes, or whole brain of experimental animals. A data search was performed on erythrocyte, plasma, and brain ChE inhibition caused by organophosphate and carbamate insecticides in experimental animals (mice, rats, and chicks) treated with the antihistamine diphenhydramine in accordance with preferred reporting items for systematic reviews and meta-analysis, which was done by the two-group random-effects model meta-analysis. The meta-analysis included 18 records extracted from six studies that, appeared from 1996 to 2022. Using the random-effects model, a two-group meta-analysis revealed that the combined effect size (ChE inhibition) was significantly more favorable in the control group than in the diphenhydramine intervention, as shown by a forest plot. The combined effect size (standardized mean difference) was 0.67, with a standard error of 0.3, a lower limit of 0.04, and an upper limit of 1.29 (p = 0.025). The heterogeneity was moderate, as I2 of the combined effect size was 74%, with a significant Cochrane Q-test result (Q = 65, p < 0.0001). Subgroup analysis indicated that, with brain ChE inhibition, the heterogeneity (I2) became 5%, which was lower than ChE inhibition in plasma (84%) and erythrocytes (78%). No publication bias was identified using the funnel plot and Egger's test. This meta-analysis suggests that, in addition to its documented antidotal action against ChE-inhibiting insecticides, diphenhydramine can also reduce the extent of ChE inhibition, especially in the brain, which is the main site of toxicity of these insecticides. There is a need for additional studies to assess such enzyme inhibition in different parts of the brain.

Recognition and Assessment of Antidotal Effects of Diphenhydramine against Acute Carbaryl Insecticide Poisoning in a Chick Model

Diphenhydramine antagonizes poisoning produced by cholinesterase (ChE) inhibiting insecticides. This study examines the effects of diphenhydramine against acute poisoning induced by the carbamate insecticide carbaryl in a chick model. The effects of diphenhydramine on the 24 h median Lethal Dose (LD50), and acute toxicity of carbaryl were assessed in chicks (7-15 days old). The plasma and whole brain ChE activities were measured electrometrically in vitro and in vivo. Diphenhydramine at 10mg/Kg Body wt. administered intramuscularly 15 min before carbaryl dosing increased the oral LD50 value of carbaryl (207 mg/Kg Body wt.) by 62%. Carbaryl at 250 mg/Kg Body wt. has orally produced toxidrome of cholinergic poisoning with 100% lethality in 24 h. Diphenhydramine (10mg/ Kg Body wt.) used 15 min before carbaryl (250mg/Kg Body wt., orally) was the most effective dose (vs 5 and 20mg/Kg Body wt.) in delaying carbaryl-toxicity and increasing survivals in chicks. The intramuscular median effective dose (ED50) of diphenhydramine which prevented 24 h carbaryl-death in chicks was 8.6mg/ Kg Body wt. The antidotal response to diphenhydramine was similar to that of the standard antidote atropine sulfate. Diphenhydramine at 10mg/Kg Body wt., given immediately after carbaryl (200mg/Kg Body wt.), reduced the percentages of plasma and whole brain ChE inhibitions in vivo by 12- and 13%, respectively. Carbaryl (10μmol/L) in vitro inhibited ChE activities in the plasma and brain by 53 and 77%, respectively; these inhibitions were reduced by 13- and 14%, respectively, when diphenhydramine (10μmol/L) was added to in vitro reactions. Diphenhydramine exerted antidotal action against a model of acute and lethal carbaryl intoxication in chicks.

Efficacy of Jambu (Seed) as an Antidote in Kuchala Poisoning an Experimental Study

Abstract: Kuchala (Strychnos nux-vomica) is spinal poison which is also classified as Upavisha in Ayurveda.Various poisons have been described in Ayurveda along with their antidotes. Mode of action of these antidotes is not mentioned in texts. In Ayurvedic literature According to Basavrajeeyam under the heading of Vish-Prativishani, Jambu (seed) have been described to be possessing antidote action which may act by some way to counter toxicity of Kuchala. It is necessary to verify the efficacy of these antidotes on scientific parameters so that it can be useful in emergencies. Objectives: To establish the action and mechanism of Jambu (seed) against toxic effects of Kuchala on albino wistar rats. Methods: albino wistar rats were selected as an animal model and antidote potential of Jambu (seed) against Kuchala is evaluated by measuring convulsions and histopathological reports of brain and spinal cord. Results: Jambu (seed) administration before Kuchala poisoning reduced the toxic effects of Kuchala such as convulsion and histopathological changes in brain and spinal cord. Conclusion: From this study, we can conclude that Jambu (seed) can resist the toxic effects of Kuchala up to some extent.

A review of Dhatura as Poison and Kamala Patra as Antidote

Dhatura plant has been noted for intoxicating, narcotic properties, they produce temporary insensibility (stupefying effects) in ordinary doses.&#x0D; Dhatura (Datura metel) is cerebrotoxic, Deleriant poison which is also classified as Upavisha in Ayurveda. &#x0D; In Ayurvedic literature according to Basavrajeeyam under the heading of Vish-Prativishani, Chincha Rasa and Kamal Patra churna has been described to be possessing antidote action which may act by some way to counter toxicity of Dhatura.&#x0D; It is necessary to verify the efficacy of these antidotes on scientific parameters so that it can be useful in emergencies.

Mechanisms of action of new oral coagulants

The article provides a brief description of drugs in the class of direct oral anticoagulants. The mechanisms of the therapeutic effect and the ways of excretion of drugs-inhibitors of blood coagulation factors Xa and IIa are considered. It was shown that all of them are characterized by high selectivity to the active centers of target molecules, successfully affect the coagulation cascade and have a quick therapeutic effect. A comparative analysis of drugs of the class of direct oral anticoagulants showed that rivaroxaban, apixaban, edoxaban, betrixaban (inhibitors of FXa) and dabigatran (inhibitor of factor IIa) do not have significant advantages over each other in terms of mechanism and speed of action. A comparison of the main characteristics of direct oral anticoagulants and the indirect anticoagulant warfarin is presented. It is shown that, first of all, the advantage of the former is associated with an immediate pharmacological response. The decision to prescribe this drug requires consideration of risk factors for the patient, such as age, the presence of chronic diseases of the gastrointestinal tract, liver and kidneys. The article presents the most frequent complications of anticoagulant therapy, as well as the mechanisms of action of specific and universal antidotes indicated for use in patients with life-threatening bleeding or in need of urgent surgery when using direct oral anticoagulants. However, such neutralizing agents have not yet found widespread use due to their high cost and low availability. Nevertheless, the emergence of direct anticoagulants on the market has undoubtedly led to significant progress in the treatment of thrombotic and cardiovascular diseases (1 table, bibliography: 28 refs).

Comparative phytochemical assessment of Ashodhita and Shodhita seeds of Ipomoea carnea Jacq (An ethno pharmacological drug).

Background: Ipomoea carnea Jacq is an ethnopharmacological drug and its leaves, roots and latex are used for various indications by traditional healers. But seeds are not used in the treatment as they are more toxic. As the herbal medicines are becoming endangered, ethnopharmacological drug can be used efficiently as they are available abundantly. But the toxicity should be reduced for safer use in the treatment. To study the effect of Shodhana of Ipomoea carnea seeds and to study the physicochemical and phytochemical changes in seeds after shodhana in different media, the present study was undertaken. Methods: Shodhana of 20 gm of Ipomoea carnea seeds was performed with Godugdha (Cow’s milk) with the help of method of swedana (boiling) in dolayantra for three hours. The same procedure was followed for another samples separately with Gomutra (Cow urine), Kanji (sour gruel), Triphala kwatha and Distilled water as shodhana media. One raw sample was kept as Ashodhita sample for comparison. Physicochemical and phytochemical analysis of all the six samples was conducted and the observations were compared among all the samples. Results: The toxic principle Swainsonine was estimated as 3% in all samples at retention time 3 minutes. The number of peaks were reduced in Godugdha, Gomutra, Kanji and Triphala Kwatha Shodhita samples in comparison with Ashodhita sample and 2-4 peaks are observed at retention time 1-3 minutes in all shodhita samples. A new phytoamine having concentration 12% was detected at retention time 17 minutes in all (crude and purified) samples. Conclusion: The number of peaks was reduced in shodita samples which may be due to antagonist action of shodhana media and 2-4 peaks are found in all shodhita samples which may be added due to the antidotal action of shodhana media. A new phytoamine detected at retention time 17 minutes in all samples needs to be identified and characterized using more advance tools like LC-MS or GC-MS.

Reaction Kinetics of Cyanide Binding to a Cobalt Schiff-Base Macrocycle Relevant to Its Mechanism of Antidotal Action.

The Co(II/III)-containing macrocycle, cobalt 2,12-dimethyl-3,7,11,17-tetraazabicyclo-[11.3.1]-heptadeca-1(17)2,11,13,15-pentaenyl cation, or CoN4[11.3.1], is a potential cyanide-scavenging agent. The rate of reduction of Co(III)N4[11.3.1] by ascorbate is reasonably facile under pseudo-first-order conditions; a second-order rate constant of 11.7(±0.4) M-1 s-1 was determined at 25 °C and pH 7.4, along with the activation parameters for the reaction (ΔH⧧ = 53.9(±0.8) kJ mol-1; ΔS -79(±3) J mol-1 K-1). It follows that any cyanide-decorporating capability of the cobalt complex should depend more on the cyanide-binding characteristics of Co(II)N4[11.3.1] than the oxidized form. The kinetics of the reaction of cyanide with Co(II)N4[11.3.1] under anaerobic pseudo-first-order conditions is rapid and resulted in a linear dependence on the cyanide concentration, kHCN = 8 × 104 M-1 s-1, with a nonlinear intercept of 420 s-1 at 10 °C, pH 7.6. The observed reaction rate increases significantly with increasing pH. A rate law is suggested, kobs = k'[X] + (kHCN + kCNKa/[H+])[HCN], where kCN is estimated to be ∼2 × 106 M-1 s-1. Activation parameters for the reaction with HCN (ΔH⧧ = 10.7(±0.4) kJ mol-1; ΔS⧧ = -153(±1) J mol-1 K-1) suggest an associative mechanism. In the presence of excess oxygen, i.e., at higher levels than free oxygen in vivo, the reaction rate was too fast to be measured, and the final product was the oxidized complex, Co(III)N4[11.3.1], where any cyanide ligands had been lost. This is much more rapid than the oxidation of the parent compound by oxygen, for which a second-order rate constant of 0.5(±0.02) M-1 s-1 at 25 °C was obtained. The study has gone some way toward enhancing our understanding of the reaction of Co(II)N4[11.3.1] with cyanide. The fast reaction rate implies a high efficacy of the cyanide-scavenging capability of the complex and further supports the suggestion stemming from our previous work that Co(II)N4[11.3.1] could prove to be a better and more cost-effective cyanide antidote than the FDA-approved hydroxocobalamin.

Assessing modulators of cytochrome c oxidase activity in Galleria mellonella larvae

Caterpillars of the greater wax moth, Galleria mellonella, are shown to be a useful invertebrate organism for examining mitochondrial toxicants (inhibitors of electron transport) and testing putative antidotes. Administration of sodium azide, sodium cyanide, or sodium (hydro)sulfide by intra-haemocoel injection (through a proleg) results in a dose-dependent paralysed state in the larvae lasting from <1 to ~40 min. The duration of paralysis is easily monitored, because if turned onto their backs, the larvae right themselves onto their prolegs once they are able to move again. The efficacy of putative antidotes to the three toxicants can routinely be assessed by observing shortened periods of paralysis with larvae given toxicant and antidote compared to larvae administered only the same dose of toxicant. The validity of the approach is demonstrated with agents previously shown to be antidotal towards cyanide intoxication in mice; namely, sodium nitrite and CoN4[11.3.1] (cobalt(II/III) 2,12-dimethyl-3,7,11,17-tetraazabicyclo-[11.3.1]-heptadeca-1(7)2,11,13,15-pentaenyl cation). These same compounds are shown to be antidotal towards all three toxicants in the G. mellonella caterpillars; findings that may prove important in relation to azide and sulfide poisonings, for which there are currently no effective antidotes available. The observation that sodium nitrite ameliorates cyanide toxicity in the larvae is additionally interesting because it unambiguously demonstrates that the antidotal action of nitrites does not require the involvement of methemoglobin, contributing to the resolution of an ongoing controversy.

THE VISHAGHNA PROPERTIES OF MANJISHTHA (Rubia cordifolia) IN AYURVEDIC AND CONTEMPORARY SCIENCE: AN OVERVIEW

The present concept is given to help for to establish as bridge between Ayurveda and contemporary science. The Concept of Ayurvedic vishaghna properties is near about detoxifying action (Antidote action) in modern era. It is may be beneficial integrated concept for metabolic toxicity, substance acquired acute and chronic toxicity, biological toxicity, cumulative toxicity etc. and it also help to understand the detoxifying phenomenon. The single and multiple preparations of manjistha are available which indicates its utility in many poisoning. All aspects of manjishtha are studied in detail especially in field of detoxification. Selection of all logical references are done and collection, correlation and explanation as per requirement. According to indication and detoxifying therapeutic use of manjishtha are highlights, which described especially in Ayurvedic and modern text. All ayurvedic and contemporary references regarding vishaghna are collected from Ayurvedic fundamental books and various textbooks, research article, international journals. Theoretically, it will breakdown the pathogenesis of toxicity and progress forward to denaturized any poison. It can be useful for disease which comes under area of any field of toxicity. We can prevent and treat much toxicological disorder. On the basis of concept of vishaghana properties of manjishtha, it can be broadly used in today’s era for preventive as well as curative disease free life from toxicological agent.&#x0D; Keywords; manjishtha, vishaghana properties, detoxifying uses

Efficacy of antidotes and their combinations in the treatment of acute carbamate poisoning in rats

BackgroundPhysostigmine and its analogues neostigmine, pyridostigmine and rivastigmine are carbamates nowadays used in many indications, including antidotal effects against antimuscarinic poisonings, reversal of competitive neuromuscular block, myasthenia gravis, Alzheimer’s disease and prophylaxis against nerve agent intoxications. Use of these medicinal carbamates, but also of carbamate insecticides, created need for research into the potential and mechanisms of action of several antidotes against carbamate poisonings, including anticholinergics and oximes. AimThe goal of this experimental study was to ascertain the life-preserving potential of anticholinergics atropine, hexamethonium and d-tubocurarine, oxime HI-6 and their combinations in rats poisoned with physostigmine or pyridostigmine. Materials and methodsExperiments were performed in Wistar rats. Carbamates were injected subcutaneously (sc) and antidotes intramuscularly (im). Median lethal dose (LD50) in animals treated with antidotes were compared to the ones in saline-treated rats and protective ratios (PRs) were calculated. Atropine (5, 10 and 20 mg/kg), hexamethonium (5, 10 and 20 mg/kg), d-tubocurarine (0.005, 0.010 and 0.020 mg/kg) and oxime HI-6 (25, 50 and 100 mg/kg) were used as monotherapies and in dual combinations, where atropine was the obligatory antidote. Biochemical experiments consisted in measuring of the cholinesterase activities in brain, whole blood and diaphragm in rats 5, 15, 30, 60, 120 and 240 min after poisoning with 0.8 LD50 of physostigmine or pyridostigmine. ResultsAll the tested antidotes assured some degree of protection against the two carbamates. Atropine and hexamethonium produced better protection in physostigmine-poisoned rats, while d-tubocurarine and HI-6 were more efficacious in pyridostigmine-intoxicated animals. Oxime HI-6 50 mg/kg reactivated acetylcholinesterase (AChE) in brain inhibited by physostigmine and in diaphragm inhibited by pyridostigmine. ConclusionsMechanism of physostigmine-induced lethal effect is predominantly central and it involves inhibition of brain AChE, while pyridostigmine produces the same effect exclusively outside the central nervous system, by inhibiting AChE in the respiratory muscles. As a consequence, increasing doses of atropine and their combination with hexamethonium assure excellent protection against physostigmine toxicity, while the best protection against pyridostigmine is provided by a strictly peripherally acting antinicotinic d-tubocurarine and bispyridinium oxime HI-6. The oxime acts as antidote against physostigmine and pyridostigmine poisoning by reactivating AChE in the brain and diaphragm, respectively.

Toxicodynamics of Organophosphates with Human Acetylcholinesterase Interaction at Novel Site Trp-86 for Antidote Action

Molecular Docking, site directed mutagenesis and molecular dynamic (MD) simulation approaches were used to explore mode of binding and inhibition for human acetylcholinesterase (hAChE) and organophosphates (OPs). More than 200 OPs molecules were investigated using Glide docking module of Schrodinger suite as co-crystal structure between two are not available in protein data bank. In initial screening Trp86 was found to be involved in maximum π-cation interaction on anionic subsite of hAChE other than Ser203 (catalytic site). With extra precision glide docking phoxim ethyl phosphonate (PEP) tops among 200 OPs based on glide docking score while interacted with Trp86, Gly121 and Ser203 whereas MMGBSA score shows less binding affinity than heptenophos and dichlorovos. Trp86 preferred π interaction with ring bearing OPs and hydrophobic interactions with smaller OPs without ring bearing structures. Site directed mutagenesis at Trp86 (Trp86 to Ala86) shown the deterioration of the binding site in terms of size reduction, loss of electrostatic and geometric stabilization in binding cavity and significant reduction in binding of OPs in preferred orientation. Dock score of both wild and mutated hAChE shows a perfect qualitative agreement (R 2 =64.1%) towards the study. Molecular dynamic simulation (GROMACS 4.5.5) of hAChE-PEP complex for 4×10 4 pico-second with SPC16 water system at 310K temperature explained the evident role of Trp86 in stabilizing the ligand at P-site of the enzyme. Asp74 and Tyr 124 were noticed in conveying H-bonds. Trp86 have shown consistent and stable distance between residues and ligand. Asp74 and Tyr124 appeared as important residues which contributed H bonds and Ser203 was expected to be closer for interaction to happen as it disappeared during simulation. Study suggests role of Trp86 on binding site is equally important to take consideration. As residue Trp86 plays significant role, it can be taken into consideration for the studies on development of more efficient antidotes to overcome the case of human poisoning.

Method development for detecting the novel cyanide antidote dimethyl trisulfide from blood and brain, and its interaction with blood.

The antidotal potency of dimethyl trisulfide (DMTS) against cyanide poisoning was discovered and investigated in our previous studies. Based on our results it has better efficacy than the Cyanokit and the Nithiodote therapies that are presently used against cyanide intoxication in the US. Because of their absence in the literature, the goal of this work was to develop analytical methods for determining DMTS from blood and brain that could be employed in future pharmacokinetic studies. An HPLC-UV method for detection of DMTS from blood, a GC-MS method for detection of DMTS from brain, and associated validation experiments are described here. These analytical methods were developed using in vitro spiking of brain and blood, and are suitable for determining the in vivo DMTS concentrations in blood and brain in future pharmacokinetic and distribution studies. An important phenomenon was observed in the process of developing these methods. Specifically, recoveries from fresh blood spiked with DMTS were found to be significantly lower than recoveries from aged blood spiked in the same manner with DMTS. This decreased DMTS recovery from fresh blood is important, both because of the role it may play in the antidotal action of DMTS in the presence of cyanide, and because it adds the requirement of sample stabilization to the method development process. Mitigation procedures for stabilizing DMTS samples in blood are reported.

Novel nucleophiles enhance the human serum paraoxonase 1 (PON1)-mediated detoxication of organophosphates.

Paraoxonase 1 (PON1) is a calcium-dependent hydrolase associated with serum high-density lipoprotein particles. PON1 hydrolyzes some organophosphates (OPs), including some nerve agents, through nucleophilic attack of hydroxide ion (from water) in the active site. Most OPs are hydrolyzed inefficiently. This project seeks to identify nucleophiles that can enhance PON1-mediated OP degradation. A series of novel nucleophiles, substituted phenoxyalkyl pyridinium oximes, has been synthesized which enhance the degradation of surrogates of sarin (nitrophenyl isopropyl methylphosphonate; NIMP) and VX (nitrophenyl ethyl methylphosphonate; NEMP). Two types of in vitro assays have been conducted, a direct assay using millimolar concentrations of substrate with direct spectrophotometric quantitation of a hydrolysis product (4-nitrophenol) and an indirect assay using submicromolar concentrations of substrate with quantitation by the level of inhibition of an exogenous source of acetylcholinesterase from non-hydrolyzed substrate. Neither NIMP nor NEMP is hydrolyzed effectively by PON1 if one of these novel oximes is absent. However, in the presence of eight novel oximes, PON1-mediated degradation of both surrogates occurs. Computational modeling has created a model of PON1 embedded in phospholipid and has indicated general agreement of the binding enthalpies with the relative efficacy as PON1 enhancers. PON1 enhancement of degradation of OPs could be a unique and unprecedented mechanism of antidotal action.

Pre-illumination of rice blast conidia induces tolerance to subsequent oxidative stress

Many environmental factors, alone or combined, affect organisms by changing a pro-/antioxidant balance. Here we tested rice blast fungus (Magnaporthe oryzae) for possible cross-adaptations caused by relatively intense light and protecting from artificially formed reactive oxygen species (ROS) and ROS-dependent fungitoxic response of the host plant. Spore germination was found to be suppressed under 4-h and, to larger extent, 5-h illumination. The effect was diminished by antioxidants and, therefore, suggests involvement of ROS. One-hour of light did not affect spore germination, but stimulated their chemically assayed superoxide production. The illuminated spores were more tolerant (than non-illuminated ones) to artificially generated H2O2, O2−, or OH or to toxic diffusate of rice leaf. They also caused more severe disease symptoms if applied to leaves of the susceptible rice cultivar at low concentration. Spore diffusates decomposed hydrogen peroxide. They detoxified exogenous H2O2 and superoxide radical as well as leaf diffusates. Spore illumination increased some of these protective effects. It is suggested that short-term light led to mild oxidative stress, which induced spore antioxidant capacity, enhancing spore tolerance to subsequent stronger oxidative stress and its aggressiveness in planta. Such tolerance depends partly on the antidotal action of spore extracellular compounds, which may also be light-stimulated. Therefore, a certain ROS-related environmental factor may adapt a fungus to other factors and so modulate its pathogenic properties.

Toxicologic Emergencies in the Intensive Care Unit

To review the most common drugs implicated in overdoses admitted to the intensive care unit focusing on antidotes and reversal agents used in their management. Poisonings and overdoses due to pharmaceutical agents result in more than 100 000 critical care unit admissions each year. Ingestion of toxic alcohols, calcium channel blockers, beta-adrenergic antagonists, benzodiazepines, opioids, acetaminophen, tricyclic antidepressants, and salicylates are associated with a high rate of morbidity and mortality. Reviewing the mechanism of toxicity due to specific agents along with the mechanism of action, dosing, and adverse effects of appropriate antidotes is important for the successful management of these patients within the critical care unit. Understanding the most prevalent overdoses and their management using reversal agents and antidotes is essential to the overall treatment of these critically ill patients.

P63: The antidotal action of nitrites toward cyanide intoxication: An example of medicine disguised as quackery

Sodium nitrite ameliorates sub-lethal cyanide toxicity in mice when given from ∼1 h before until 20 min after the toxic dose as demonstrated by the recovery of righting ability. In most experiments nitrite was given intraperitoneally, but may also be administered as an inhaled aqueous vapor-contrary to recommended EMR protocols, it is not necessary to infuse the antidote intravenously and the co-administration of adjuvants (specifically thiosulfate) is unnecessary. Intraperitoneally administered nitrite rapidly produces NO in the bloodstream as judged by the dose-dependent appearance of EPR signals attributable to nitrosylhemoglobin and methemoglobin. However, the commonly held belief that the antidotal mechanism of nitrites involves the scavenging of cyanide anion by methemoglobin is firmly contraindicated by recent evidence. Intriguingly, the FDA-approved and clinically tested cyanide scavenger cobalamin has, in fact, rather indifferent binding affinity for cyanide anion. It follows that the clinical usefulness of cobalamin may be dependent on endogenous nitric oxide displacing the cyanide from the active site of cytochrome c oxidase to facilitate the cyanide-scavenging action of cobalamin. Antagonism of cyanide inhibition of cytochrome c oxidase by NO appears to be the crucial antidotal activity rather than any methemoglobin-forming action of nitrite. Concomitant addition of sodium thiosulfate to nitrite-treated blood results in the production of sulfidomethemoblobin as detected by EPR spectroscopy. Sulfide is a product of thiosulfate hydrolysis and, like cyanide, is known to be a potent inhibitor of cytochrome c oxidase; the effects of the two inhibitors being essentially additive under standard assay conditions, rather than dominated by either one. The findings afford a plausible explanation for an observed detrimental effect in mice associated with the use of the standard nitrite-thiosulfate combination therapy at sub-lethal levels of cyanide intoxication. Isoamyl nitrite, given intraperitoneally in equimolar amounts to sodium nitrite, is comparatively less effective at restoring the righting recovery of cyanide-challenged mice. The results of investigations with the hydrolysis products of isoamyl nitrite, isoamyl alcohol and nitrite anion, suggest that the toxicity of isoamyl alcohol renders the organic nitrite a less desirable antidotal agent than sodium nitrite. Disclosure Supported by the CounterACT Program, National Institutes of Health Office of the Director (NIH OD), and the National Institute of Neurological Disorders and Stroke (NINDS), Grant No. NS063732 to J.P and L.L.P and Bruce R. Pitt.