The complex of cobalt(II) with the ligand 2,12-dimethyl-3,7,11,17-tetraazabicyclo-[11.3.1]heptadeca-1(17)-2,11,13,15-pentaene (CoN4[11.3.1]) has been shown to bind two molecules of cyanide in a cooperative fashion with an association constant of 2.7 (±0.2) × 105. In vivo, irrespective of whether it is initially administered as the Co(II) or Co(III) cation, EPR spectroscopic measurements on blood samples show that at physiological levels of reductant (principally ascorbate) CoN4[11.3.1] becomes quantitatively reduced to the Co(II) form. However, following addition of sodium cyanide, a dicyano Co(III) species is formed, both in blood and in buffered aqueous solution at neutral pH. In keeping with other cobalt-containing cyanide-scavenging macrocycles like cobinamide and cobalt(III) meso-tetra(4-N-methylpyridyl)-porphine, we found that CoN4[11.3.1] exhibits rapid oxygen turnover in the presence of the physiological reductant ascorbate. This behavior could potentially render CoN4[11.3.1] cytotoxic and/or interfere with evaluations of the antidotal capability of the complex toward cyanide through respirometric measurements, particularly since cyanide rapidly inhibits this process, adding further complexity. A sublethal mouse model was used to assess the effectiveness of CoN4[11.3.1] as a potential cyanide antidote. The administration of CoN4[11.3.1] prophylactically to sodium cyanide-intoxicated mice resulted in the time required for the surviving animals to recover from “knockdown” (unconsciousness) being significantly decreased (3 ± 2 min) compared to that of the controls (22 ± 5 min). All observations are consistent with the demonstrated antidotal activity of CoN4[11.3.1] operating through a cyanide-scavenging mechanism, which is associated with a Co(II) → Co(III) oxidation of the cation. To test for postintoxication neuromuscular sequelae, the ability of mice to remain in position on a rotating cylinder (RotaRod test) was assessed during and after recovery. While intoxicated animals given CoN4[11.3.1] did recover ~30 min more quickly than controls given only toxicant, there were no indications of longer-term problems in either group, as determined by continuing the RotaRod testing up to 24 h after the intoxications and routine behavioral observations for a further week.

There are currently no FDA-approved antidotes for H2S/sulfide intoxication. Sodium nitrite, if given prophylactically to Swiss Webster mice, was shown to be highly protective against the acute toxic effects of sodium hydrosulfide (~LD40 dose) with both agents administered by intraperitoneal injections. However, sodium nitrite administered after the toxicant dose did not detectably ameliorate sulfide toxicity in this fast-delivery, single-shot experimental paradigm. Nitrite anion was shown to rapidly produce NO in the bloodstream, as judged by the appearance of EPR signals attributable to nitrosylhemoglobin and methemoglobin, together amounting to less than 5% of the total hemoglobin present. Sulfide-intoxicated mice were neither helped by the supplemental administration of 100% oxygen nor were there any detrimental effects. Compared to cyanide-intoxicated mice, animals surviving sulfide intoxication exhibited very short knockdown times (if any) and full recovery was extremely fast (~15 min) irrespective of whether sodium nitrite was administered. Behavioral experiments testing the ability of mice to maintain balance on a rotating cylinder showed no motor impairment up to 24 h post sulfide exposure. It is argued that antagonism of sulfide inhibition of cytochrome c oxidase by NO is the crucial antidotal activity of nitrite rather than formation of methemoglobin.

Four cobalt-containing macrocyclic compounds previously shown to ameliorate cyanide toxicity have been comparatively evaluated with an acute sublethal toxicity model in conscious (unanesthetized) adult male Swiss-Webster mice. All of the compounds (the cobalt-corrins cobalamin and cobinamide, a cobalt-porphyrin, plus a cobalt-Schiff base macrocycle) given 5 min prior to the toxicant dose significantly decreased the righting-recovery time of cyanide-intoxicated mice, but the doses required for maximal antidotal effect varied. Additionally, all of the compounds tested significantly reduced the righting-recovery time when administered at either 1 or 2 min after cyanide intoxication, but none of the compounds tested significantly reduced the righting-recovery time when delivered 5 min after the toxicant dose. Using the lowest effective dose of each compound determined during the first (prophylactic) set of experiments, neuromuscular recovery following cyanide intoxication in the presence/absence of the cobalt-based antidotes was assessed by RotaRod testing. All the compounds tested accelerated recovery of neuromuscular coordination, and no persistent impairment in any group, including those animals that received toxicant and no antidote, was apparent up to 2 weeks postexposures. The relative effectiveness of the cobalt compounds as cyanide antidotes are discussed and rationalized on the basis of the cyanide-binding stoichiometries and stability constants of the Co(III) cyano adducts, together with consideration of the rate constants for axial ligand substitutions by cyanide in the Co(II) forms.

Three cobalt-containing macrocyclic compounds previously shown to antagonize cyanide toxicity have been comparatively evaluated for the amelioration of sublethal azide toxicity in juvenile (7−8 weeks) Swiss-Webster mice. The lowest effective doses were determined for hydroxocobalamin, a cobalt porphyrin, and a cobalt-Schiff base macrocycle by giving the antidotes 5 min prior to the toxicant, 27 mg (415 μmol) /kg sodium azide. Both male and female mice were evaluated for their response to the toxicant as well as the antidotes, and no significant differences were noted once weight differences were taken into account. Two of the three compounds significantly decreased the recovery time of azide-intoxicated mice at 10 min after the administration of sodium azide, as determined by a behavioral test (pole climbing). Additionally, azide was determined to cause a several degree drop (∼3 °C) in measured tail temperature, and warming the mice led to a more rapid recovery. The mice were also shown to recover more rapidly when given sodium nitrite, 24 mg (350 μmol)/kg, 5 min after the toxicant; this treatment also suppressed the azide-induced tail temperature decrease. Electron paramagnetic resonance (EPR) measurements of mouse blood treated with sodium azide demonstrated the presence of nitrosylhemoglobin at levels of 10−20 μM which persisted for ∼300 min. The presence of the methemoglobin azide adduct was also detected by EPR at a maximum level of ∼300 μM, but these signals disappeared around 200 min after the administration of azide. The treatment of mice with 15 N sodium azide proved that the nitrosylhemoglobin was a product of the administered azide by the appearance of a two-line hyperfine (due to the 15 N) in the EPR spectrum of mouse blood.■ EXPERIMENTAL SECTION Chemicals. Gases were purchased from Matheson, and all nongaseous reagents (ACS grade or better), including Na 15 N 3 (98%

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There is presently no antidote available to treat azide poisoning. Here, the Schiff-base compound Co(II)-2, 12-dimethyl-3,7,11,17-tetraazabicyclo-[11.3.1]heptadeca-1(17)2,11,13,15-pentaenyl dibromide (Co(II)N 4 [11.3.1]) is investigated to determine if it has the capability to antagonize azide toxicity through a decorporation mechanism. The stopped-flow kinetics of azide binding to Co(II)N 4 [11.3.1] in the absence of oxygen exhibited three experimentally observable phases: I (fast); II (intermediate); and III (slow). The intermediate phase II accounted for ∼70% of the overall absorbance changes, representing the major process observed, with second-order rate constants of 29 (±4) M −1 s −1 at 25 °C and 70 (±10) M −1 s −1 at 37 °C. The data demonstrated pH independence of the reaction around neutrality, suggesting the unprotonated azide anion to be the attacking species. The binding of azide to Co(II)N 4 [11.3.1] appears to have a complicated mechanism leading to less than ideal antidotal capability; nonetheless, this cobalt complex does protect against azide intoxication. Administration of Co(II)N 4 [11.3.1] at 5 min post sodium azide injection (ip) to mice resulted in a substantial decrease of righting-recovery times, 12 (±4) min, compared to controls, 40 (±8) min. In addition, only two out of seven mice "knocked down" when the antidote was administered compared to the controls given toxicant only (100% knockdown).

Phosphine (PH3) poisoning continues to be a serious problem worldwide, for which there is no antidote currently available. An invertebrate model for examining potential toxicants and their putative antidotes has been used to determine if a strategy of using Au­(I) complexes as phosphine-scavenging compounds may be antidotally beneficial. When Galleria mellonella larvae (or wax worms) were subjected to phosphine exposures of 4300 (±700) ppm·min over a 20 min time span, they became immobile (paralyzed) for ∼35 min. The administration of Au­(I) complexes auro-sodium bisthiosulfate (AuTS), aurothioglucose (AuTG), and sodium aurothiomalate (AuTM) 5 min prior to phosphine exposure resulted in a drastic reduction in the recovery time (0–4 min). When the putative antidotes were given 10 min after the phosphine exposure, all the antidotes were therapeutic, resulting in mean recovery times of 14, 17, and 19 min for AuTS, AuTG, and AuTM, respectively. Since AuTS proved to be the best therapeutic agent in the G. mellonella model, it was subsequently tested in mice using a behavioral assessment (pole-climbing test). Mice given AuTS (50 mg/kg) 5 min prior to a 3200 (±500) ppm·min phosphine exposure exhibited behavior comparable to mice not exposed to phosphine. However, when mice were given a therapeutic dose of AuTS (50 mg/kg) 1 min after a similar phosphine exposure, only a very modest improvement in performance was observed.

Bovine pulmonary artery endothelial cells (BPAEC) respond in a dose-dependent manner to millimolar (0-10) levels of sodium sulfide (NaHS). No measurable increase in caspase-3 activity and no change in the extent of autophagy (or mitophagy) were observed in BPAEC. However, lactate dehydrogenase levels increased in the BPAEC exposed NaHS, which indicated necrotic cell death. In the case of galactose-conditioned BPAEC, the toxicity of NaHS was increased by 30% compared to that observed in BPAEC maintained in the regular glucose-containing culture medium, which indicated a link between mitochondrial oxidative phosphorylation and the mechanism of toxicant action. This is consistent with the widely held view that cytochrome c oxidase (complex IV of the mitochondrial electron-transport system) is the principal molecular target involved in the acute toxicity of "sulfide" (HS/HS). In support of this view, elevated NO (which can reverse cytochrome c oxidase inhibition) ameliorated the toxicity of NaHS and, conversely, suppression of endogenous NO production exacerbated the observed toxicity. Respirometric measurements showed the BPAEC to possess a robust sulfide oxidizing system, which was able to out-compete cytochrome c oxidase for available HS/HS at micromolar concentrations. This detoxification system has previously been reported by other groups in several cell types, but notably, not neurons. The findings appear to provide some insight into the question of why human survivors of HS inhalation frequently present at the clinic with respiratory insufficiency/pulmonary edema, while acutely poisoned laboratory animals tend to either succumb to cardiopulmonary paralysis or fully recover without any intervention.