Sandra A. Thomson

Inhalation toxicity of Cyclosarin (GF) vapor in rats as a function of exposure concentration and duration: potency comparison to sarin (GB).

The inhalation toxicity of cyclohexyl methylphosphonofluoridate (GF) was examined in male and female Sprague-Dawley rats exposed by whole body in a dynamic 750-L chamber. The objectives of this study were to (1) generate GF vapor in a dynamic inhalation chamber system, starting in the lethal to near-lethal concentration range, (2) examine dose-response effects of inhaled GF vapor and analyze the relationship between concentration (C) and exposure duration (T) in determining probability of lethality, and (3) establish a lethal potency ratio between GF and the more volatile agent Sarin (GB). Using a syringe pump, GF vapor concentrations were generated for exposure times of 10, 60, and 240 min. Dose-response curves with associated slopes were determined for each exposure duration by the Bliss probit method. GF vapor exposures were associated with sublethal clinical signs such as tremors, convulsions, salivation, and miosis. Concentration-exposure time values for lethality in 50% of the exposed population (LCT(50)) were calculated for 24-h and 14-day postexposure periods for 10-, 60-, and 240-min exposures. In general, LCT(50) values were lower in female rats than males and increased with exposure duration; that is, CT was not constant over time. The GF LCT(50) values for female rats were 253 mg min/m(3) at 10 min, 334 mg min/m(3) at 60 min, and 533 mg min/m(3) at 240 min, while the values for males were 371, 396, and 585 mg min/m(3), respectively. The GB LCT(50) values for female rats were 235 mg min/m(3) at 10 min, 355 mg min/m(3) at 60 min, and 840 mg min/m(3) at 240 min, while the values for males were 316, 433, and 1296 mg min/m(3), respectively. At longer exposure durations, the LCT(50) for GF was less than that found for GB but at shorter exposure durations, the LCT(50) for GF was more than that found for GB. Empirical models, consisting of the toxic load model plus higher order terms, were developed and successfully fit to the data.

Effects of Whole-Body VX Vapor Exposure on Lethality in Rats

Male and female rats were whole-body exposed to VX vapor in a 1000-L single-pass exposure chamber. Estimated exposure dosages producing lethal (LCT50) effects in 50% of exposed male and female rats were established for 10, 60, and 240 min exposure durations. A potency comparison with GB and GF shows that VX becomes increasingly more potent than these G agents with increasing exposure duration. VX is approximately 4–30 times more potent than GB and 5–15 times more potent than GF. Gender differences in the estimated median dosages were not significant at the 10, 60, and 240 min exposure durations. An empirical toxic load model was developed and the toxic load exponent for lethality (n) in the equation Cn× T = k was determined to be n = 0.92. The VX–G regeneration assay was successfully used as a biomarker for the presence of VX in the blood plasma and RBC fractions of the blood 24 h postexposure.

Gender difference in the miotic potency of soman vapor in rats

The present study was undertaken to investigate the miotic potency of soman vapor in the rat, as well as gender differences in the miotic response to soman vapor that have been reported previously for other nerve agents. The results of the present study demonstrate that the miotic potency of soman vapor is significantly less than that of other nerve agents, and that female rats are 2.5–3.0 times more sensitive to soman vapor than male rats. The results also demonstrate that ocular acetylcholinesterase and butyrylcholinesterase activities differ between males and females, although this difference is not likely large enough to account for the observed gender difference.

Generation, sampling, and analysis for low-level GB (Sarin) and GF (Cyclosarin) vapor for inhalation toxicology studies.

This study tested and optimized various methodologies to generate, sample, and characterize GB and GF test atmospheres in an inhalation chamber, particularly at low vapor levels. A syringe drive/spray atomization system produced vapor concentrations at a range of 1–50 mg/m3. A saturator cell was used to generate vapor at sub-lethal concentrations ranging from 1 mg/m3 down to low levels approaching the threshold limit value time-weighted average (TLV-TWA) of 0.0001 mg/m3 for GB. Both generation techniques demonstrated the ability to produce stable vapor concentrations over extended exposure periods. This capability was important to determine sublethal nerve agent effects, such as miosis, for inhalation toxicology studies. In addition, the techniques employed for producing and maintaining low-level agent vapor would lay the foundation for testing less volatile chemical warfare agents such as VX.

Biomarkers of low-level exposure to soman vapor: comparison of fluoride regeneration to acetylcholinesterase inhibition.

The nerve agent O-pinacolyl methylphosphonofluoridate, also known as soman or by its military designation GD, is a highly toxic organophosphorous compound that exerts its effects through inhibition of the enzyme acetylcholinesterase (AChE). In the present study, a fluoride ion based regeneration assay was developed to quantify the level of soman present in the blood of rats following a low-level whole-body inhalation exposure. It was hypothesized that the amount of regenerated nerve agent in the blood would be dose dependent in rats subjected to a whole-body inhalation exposure to a low-level dose of soman vapor, and that the fluoride ion-based regeneration method would be more sensitive for the detection of a low-level exposure to soman vapor than the measurement of whole blood AChE activity. Regenerated soman was dose-dependently detected in both the red blood cells (RBCs) and plasma of exposed rats at all concentrations tested (0.033–0.280 mg/m3 for a 240-min exposure). Significant inhibition of whole blood AChE activity did not occur below a concentration of 0.101 mg/m3, and was only depressed by approximately 10–25% at concentrations ranging from 0.101 mg/m3 to 0.280 mg/m3. This study is the first to utilize a fluoride ion-based regeneration assay to demonstrate the dose-dependent increases in soman in the blood following whole-body inhalation exposure to low levels of vapor. Additionally, the results of the present study demonstrate that the fluoride ion based regeneration assay was approximately threefold more sensitive than the measurement of AChE activity in the blood for the detection of exposure to soman, and also that miosis is a more sensitive marker of soman exposure than inhibition of AChE activity.

CHEMICAL AND TOXICOLOGICAL EVALUATION OF PYROTECHNICALLY DISSEMINATED TEREPTHALIC ACID SMOKE

The terephthalic acid (TPA) smoke obscurants (M-83 grenade and M-8 smoke pot) were developed by the U.S. Army for training purposes to replace the more toxic hexachloroethane (HC) smoke. Inhalation toxicity testing and chemical characterization of pyrotechnically generated TPA was conducted to assess the health hazard potential of TPA and its combustion products. Fisher 344 rats were subjected to acute and repeated exposures to TPA smoke generated from the M-83 grenade. Acute exposure levels ranged from 150-1,900 mg/m3 for 30 minutes and repeated dose exposures ranged from 128-1,965 mg/m3 for 30 min/day for 5 days. Exposed and control rats were evaluated for toxic signs, and histopathologic changes. During exposure, the rats exhibited slight to moderate lacrimation, rhinorrhea, lethargy and dyspnea, which reversed within 1-hr post-exposure. No deaths occurred, even at the highest smoke concentrations. Histopathological changes were confined to exposure related nasal necrosis and inflammation in both the acute and repeated dose exposures at levels above 900 mg/m3. Chemical characterization of the M-83 grenade and the M-8 smoke pot showed that formaldehyde, benzene and carbon monoxide were the major organic vapor by-products formed. These by-products were above their respective ACGIH threshold limit values at various concentrations, but should not pose a hazard if the smoke is deployed in an open area. Overall, TPA is a safer training smoke to replace the HC smoke.

Kinetics of sarin (GB) following a single sublethal inhalation exposure in the guinea pig.

To improve toxicity estimates from sublethal exposures to chemical warfare nerve agents (CWNA), it is necessary to generate mathematical models of the absorption, distribution, and elimination of nerve agents. However, current models are based on representative data sets generated with different routes of exposure and in different species and are designed to interpolate between limited laboratory data sets to predict a wide range of possible human exposure scenarios. This study was performed to integrate CWNA sublethal toxicity data in male Duncan Hartley guinea pigs. Specific goal was to compare uptake and clearance kinetics of different sublethal doses of sarin (either 0.1 × or 0.4 × LC50) in blood and tissues of guinea pigs exposed to agent by acute whole-body inhalation exposure after the 60-min LC50 was determined. Arterial catheterization allowed repeated blood sampling from the same animal at various time periods. Blood and tissue levels of acetylcholinesterase, butyrylcholinesterase, and regenerated sarin (rGB) were determined at various time points during and following sarin exposure. The following pharmacokinetic parameters were calculated from the graph of plasma or RBC rGB concentration versus time: time to reach the maximal concentration; maximal concentration; mean residence time; clearance; volume of distribution at steady state; terminal elimination-phase rate constant; and area under plasma concentration time curve extrapolated to infinity using the WinNonlin analysis program 5.0. Plasma and RBC t1/2 for rGB was also calculated. Data will be used to develop mathematical model of absorption and distribution of sublethal sarin doses into susceptible tissues.