1-METHOXY-l,3,5-CYCLOHEPTATRIENE

Possible Long-Term Health Effects of Short-Term Exposure To Chemical Agents, Volume 2: Cholinesterase Reactivators, Psychochemicals and Irritants and Vesicants (1984)

CHARACTERISTICS

Riot-control agents - such as CS, CN, and CR - are solids. Edgewood scientists9 felt that dissemination and decontamination would be simpler with a volatile liquid agent. Research led to the production and testing of l-methoxy-l,3,5-cycloheptatriene (CHT, EA 4923), a liquid substance of high volatility (Table 4-40) with physiologic effects typical of riot-control agents (e.g., lacrimation, skin irritation, and mucous membrane irritation).

CHT, known as tropilidene in England,2 is pale yellow, with a molecular weight of 92, a boiling point of 117ºC at 749 mm Hg, and a freezing point of -79.5ºC. It is unstable when exposed to air or light. Early Edgewood samples were difficult to produce in a pure state, ranging from 78 to 89%, with EA 4922 (a relatively inert Isomer) forming most of the remainder.

CHT is less irritating than CS and CR to humans. It is roughly as toxic as CR and less toxic than CS by inhalation in animals, and it has the capacity to penetrate skin or rubber.9

TOXICITY IN ANIMALS

Brown et al.2 evaluated the safe handling of CHT, using only four males and four females per test. They used 96% pure CHT. In gavage studies, the LD50 of CHT in rats and mice was 57 and 171 mg/kg, respectively. Clonic convulsions suggested CNS involvement as a mechanism of toxic effects. Undiluted CHT applied to clipped back skin of rats under an impermeable bandage for 24 h produced an LD50 of 442-884 mg/kg. When 1 ml of CHT was applied to a 2 x 2-in. patch that was then placed on the clipped back skin of rabbits under an impermeable cover, and this procedure was repeated daily for 3 d for 6 h at a time, severe damage was caused, with necrosis and ulceration. Application of 1 ml of CHT to the clipped, uncovered backs of two rabbits and 10 guinea pigs daily for 23 d, produced gross skin irritation, with spongiosis, acanthosis, and necrosis. Attempts to sensitize guinea pigs yielded negative results. Undiluted CHT applied to rabbit eyes caused severe conjunctivitis that cleared in 24 h; there was swelling of lids, but no corneal involvement.

Early studies at Edgewood were conducted on dogs.9 Severe neurologic damage was observed in one animal dosed with degraded agent (a dark mixture with a nonvolatile component). This observation led to further studies, including the use of CHT stabilized with antioxidants. Percutaneous toxicity was tested by applying CHT to the backs of clipped dogs and covering the area with a Saran plastic sheet held in place with a cloth jacket. Jackets were removed after 24 h, and the dogs' skins were then decontaminated. Results (Table 4-41) showed that ataxia, tremors, and death can be produced by these procedures. Other tests involved topical applications of CHT at 0.5 and 1.0 ml/kg (86.7 mol% pure) on two sets of dogs; stabilized and non-stabilized material was used. Eight dogs (two per test) were tethered outdoors and exposed uncovered for 4 h at 75ºF. A second group of eight dogs was similarly tested at 100ºF. No dogs died, but one showed transitory mild tremors after application of the stabilized agent at 1.0 ml/kg. The percutaneous LD50 in dogs (occluded skin) was estimated at about 500 mg/kg. Results were similar for stabilized and nonstabilized CHT. Ocular instillation of 5 1 (2 d), 10 1 (1 d) and 50 1 (4 d) of freshly distilled, about 90% pure agent caused mild erythema, inflammation, and iritis in rabbits, which cleared in 4-6 d. Other samples and degraded fractions given in single instillations of 10-50 1 produced varied but increased eye injury, including permanent corneal opacity, and in some cases injury to skin.9

The combined effects of inhaled and topically applied CHT were tested on eight dogs on which stabilized and nonstabilized CHT was applied at 1.0 and 0.5 ml/kg.4 The dogs were placed in an unventilated 441-ft3 room whose atmosphere was saturated with CHT and exposed twice, once for 1.5 h and a second time for 6 h. Effects of 1.5-h exposures were long-lasting, but not lethal. Deaths occurred from the 6-h exposures, however (Table 4-42). The effects of inhaled CHT were studied in dogs by allowing CHT equivalent to 0.5 and 1.0 ml/kg to evaporate from absorbent cotton in a closed cubicle. No ill effects were seen after exposure for 1.5 h. The effectiveness of decontamination procedures with bleach or soap solution was tested (Table 4-43).

McNamara et al.7 used several species of animals and several routes of administration. Various samples of CHT were used, both controlled (relatively pure and stabilized) and uncontrolled (containing degradation products). They stated that toxic signs associated with controlled and uncontrolled samples were often the same. The intravenous doses of neat agent (uncontrolled samples) were 99, 88, and about 20 mg/kg in the mouse, rabbit, and dog, respectively. The percutaneous (covered-patch) LD50 in rabbits varied from 480 to 1,000 mg/kg for different samples. The acute inhalation LCt50 (controlled samples) were 184,000, 176,000, and 63,000 in the rat, guinea pig, and dog, respectively. Death usually occurred within 24 h of exposure to lethal doses. Neuromuscular weakness persisted for several months in some dogs after the cutaneous (covered) application of CHT at 500 mg/kg or intravenous administration at 10 mg/kg. Persistent weakness was seen in rabbits after cutaneous exposure. Some samples of CHT produced corneal lesions in rabbits. Necropsies showed no lesions after CHT was given intraperitoneally, intratracheally, ocularly, or cutaneously. Pulmonary lesions were found after intravenous administration to dogs and after inhalation by dogs, rats, and guinea pigs. In surviving dogs and guinea pigs, but not always in rats, the lesions were reversible.

Biskup et_ al.,1 also at Edgewood, compared the parenteral toxicities of six agents, including CHT. LD50 in the mouse, rat, and rabbit are shown in Table 4-44.

Toxicity studies were conducted at Stanford Research Institute (SRI) with CHT under contracts with Edgewood. The first part of the SRI work was concerned with the pulmonary and neurologic lesions caused by CHT in dogs and monkeys. Monkeys tolerated high concentrations of CHT for short times and low concentrations for periods up to 100 min without adverse, irreversible effects. Ct's of 985-105,100 mg-min/m3 were tested. No significant changes in blood pressure, heart rate, or respiration rate were observed. A comparison with CS indicated that CS can produce lung lesions at a Ct of 47,000 mg-min/m3 in dogs and 20,000 mg-min/m3 in monkeys. These are much lower than Ct's of CHT that produced no significant lesions. Neurologic lesions were produced by intravenous administration of CHT in dogs, but not in monkeys. This confirms the early results at Edgewood and suggests that dogs may be peculiarly susceptible to this type of treatment.4 14 -labeled CHT was used to study distribution and excretion. No indication of retention in any organ was observed.4 A summary of percutaneous toxicity of CHT in dogs with occluded dressings is in Table 4-45.3 In a later study, the inhalation of CHT in goats was studied at SRI. Eighteen goats were tested: six were controls, six were given a low dose (Ct, 3,000 mg-min/m3), and six were given a high dose (30,000 mg-min/m3). Three of the six goats in the high-dose test died before the exposure ended, so useful data were not obtained. All exposures, however, lowered the white-cell count and the hematocrit. No change in blood-gas contents was seen. No significant changes in biochemical or pathologic factors were found.10

TOXICITY IN HUMANS

During the chemical synthetic work that led to the development of CHT, several men were exposed to its vapors. The resulting lacrimation and irritation of the mucous membranes passed quickly with no after-effects. The compound was therefore judged to be a potent irritant, but relatively harmless.9 Because of these observations, human volunteers and animals were tested simultaneously at Edgewood.

Fourteen men were exposed to various concentrations of CHT to establish an ICt50. Nine of these men withstood exposures of 28-64 mg-min/m3. These tests were terminated before an ICt50 could be established, because some animal tests suggested that the compound might be toxic at higher concentration.7 Eventually, it was found that CHT had been degraded by exposure to air and light. Purified and stabilized agent was then prepared for further toxicity testing, but no further human tests were conducted at Edgewood. Accidental exposures reported by SRI involved ocular, cutaneous, and inhalation routes. Irritant effects developed at once and disappeared 15-30 min after removal of the agent. No after-effects were reported.3

MUTAGENICITY

CHT has not been tested thoroughly for mutagenicity. On the basis of unpublished studies conducted by Edgewood, it has been suggested that CHT is nonmutagenic in the Salmonella reverse-mutation test,11 the micronucleus test in mice.11 and a dominant-lethal test in rats6. Limitations in the available data base, however, make it impossible to reach clear conclusions regarding the genotoxicity of CHT.

The Salmonella reverse-mutation test reported for CHT does not seem to have been an adequate negative test. In strains TA1538 and TA100 with metabolic activation, the range of dosages was too limited, and there was no suitable positive control for strain TA1537. Moreover, the data on strain TA98 could suggest a positive response, in that there was a fourfold increase in numbers of revertants per plate. If this increase were reproducible, one could conclude that CHT la positive, rather than negative. The available Information on the size of the experiments, their reproducibility, and the variability of the data is also too limited to support the general conclusion that CHT is nonmutagenic in bacteria.

It was reported that CHT was nonmutagenic in a test for dominant lethal mutations in rats.11 Treatment was by inhalation at concentrations of 100 and 4,000 mkl/m3; treatment involved single 20-min exposures or five 20-min exposures on consecutive days. Not having reviewed the dominant-lethal test data, we can neither accept nor refute the claim that CHT was negative in this test. However, a negative result in dominant-lethal tests is not generally regarded as strong evidence of lack of genotoxicity.8

CHT was tested for its capacity to cause an increase in the frequency of mlcronuclei in polychromatic erythrocytes in mouse bone marrow. Micronucleus formation is an indicator of chromosomal damage, and CHT had no apparent effect in the test. It should be noted, however, that the procedures used may not have conformed to current standards for a sensitive micronucleus test. For example, Heddle et al.5 recommended examining at least 500 polychromatic erythrocytes from each of at least eight animals for every dosage and time for which results were analyzed; they also recommended that sampling be extended to at least 72 h after treatment and that the highest possible dosages (e.g., 80% of the 7-d LD50) be used. If the test is conducted by a less sensitive procedure, some mutagens may not be detected. In the tests conducted at Edgewood, the highest dosage was about 25% of the LD50 - Three mice were treated per group by single intraperitoneal injection, and they were killed to determine results after only 24 h.

In our view, it is premature to reach a conclusion regarding the genetic toxicity of CHT. Although the mutagenicity tests conducted at Edgewood provided no evidence that CHT is mutagenic, they were not adequate to support the general conclusion that it is nonmutagenic. One can therefore make no statement about the possibility that CHT could pose a mutagenic hazard for man. For a thorough discussion of bases for determining the likelihood that a given chemical is muta-genic In humans, see the recent report of the National Research Council Committee on Chemical Environmental Mutagens.8

EFFECTS ON HUMAN SUBJECTS AT EDGEWOOD

In late 1969 and early 1970, 16 subjects each underwent one experimental exposure to CHT in an aerosol chamber at Edgewood. The duration of exposure among the 16 subjects ranged from 30 s to 5 min. Ct's ranged from 15.4 to 64 mg-min/m3.

The effects of exposure on the subjects were transient, with complete resolution by 15 min after leaving the chamber. The predominant effects were lacrimatory, causing incapacitation due to eye closure and blurred vision lasting several minutes after exposure. Dermal irritation and rhinorrhea occurred. One subject had "chest congestion."

Laboratory analyses were performed 9 d after exposure to CHT. Two subjects had slight increases in SCOT after exposure (31.5 and 44.5 IU), representing slightly less than a doubling of their preexposure control values. SCOT was normal in both 1 mo later. One subject had a slight persistent increase in alkaline phosphatase 9 and 15 d after exposure (13.7, 14.3 KA), representing an increase to approximately 1.5 times his exposure control value. Other tests of liver function performed on the subject with increased alkaline phosphatase 21 d after exposure had normal results. One subject had a decrease in hemoglobin content, from 16.8 g before exposure to as low as 13 g after exposure. A decrease in hematocrit from 48% to 44% accompanied the decrease in hemoglobin, and reticulocytes were 0.8% after exposure. Results of hemoglobin analyses from 9 to 20 d after exposure ranged from 13 to 14.2 g in the subject whose hemoglobin decreased, but his white-cell count was normal. These subjects were exposed to no other chemicals at Edgewood before the postexposure laboratory analysis. The slight abnormalities in SCOT after exposure to CHT might have represented idiosyncratic hepatic reactions to the chemical. Complete recovery is likely. The increase in alkaline phosphatase and the decrease in hemoglobin after CHT exposure are difficult to relate to the exposure.

Given the available information on CHT and on the Edgewood subjects exposed to the chemical, long-term health effects of the exposure on the subjects are difficult to predict.

SUMMARY

CHT appears to be a powerful lacrimator and irritant with a high safety factor and a short recovery time. It is highly volatile and dissipates rapidly in open air or in ventilated enclosures.

CHT appears safer than CR or CN. Neuromuscular damage in one dog from degraded CHT led to further studies, which demonstrated severe eye and skin damage from some of the degraded products, including neurologic signs, but no permanent damage in surviving animals.

Data are insufficient to support a conclusion regarding the mutagenicity or carcinogenicity of CHT.

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URL: Невидимое оружие Copyright © Leshchinsky Igor, 2002-2005 Revised - 1 сентября 2004г.