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Institute of Medicine (US) Committee on R&D Needs for Improving Civilian Medical Response to Chemical and Biological Terrorism Incidents. Chemical and Biological Terrorism: Research and Development to Improve Civilian Medical Response. Washington (DC): National Academies Press (US); 1999.

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Chemical and Biological Terrorism: Research and Development to Improve Civilian Medical Response.

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4Detection and Measurement of Chemical Agents

Rapid identification of the chemical or biological agents involved in any hazardous material (Hazmat) incident is vital to the protection of first responders and emergency medical personnel at local medical facilities as well as to the effective treatment of casualties. This chapter of the report deals with devices for detecting and identifying chemical agents and is followed by two chapters focusing on biological agents. However, a potential complication that can easily be overlooked is the possibility that a terrorist attack may involve the use of more than a single agent. Therefore, detection of one agent should not bring identification efforts to a premature halt. Instead, detection of any agent should be taken as an indication of an imminent threat and should therefore provoke more extensive testing.


When addressing the requirement for chemical agent detection at the scene of a terrorist incident, it is important to consider who really has the responsibility for detection operations. Is it the police, the fire department, the Hazmat team, or the EMS units? If the responsibility falls on medical assets, how will the use of detection equipment increase the efficiency of the health care team in preserving life and preventing further injury? Furthermore, will the absence of more sensitive equipment somehow inhibit their performance? These are important questions that must be addressed by policymakers and incident response planners.

Because of their physical properties, CWA use in a domestic terrorist incident may not be associated with a high-explosive event. Rather, these agents may well be dispersed in a manner that would involve a vapor hazard within a confined space. The type of incident seen in Tokyo, while minimized by the inefficient release of the CWA, is an excellent example of the type of incident expected as a result of terrorist use of CWA. The highest probability of detecting the presence of CWA occurs in cases in which there is a continuing source of vapor. By the time emergency medical personnel arrive at an incident, inevitably the agent will have dispersed significantly. In the case of cyanide and phosgene, and most nerve agents, detection in the environment may not be possible by the time monitoring equipment is in place at the scene. In fact, once casualties of a vapor CWA incident are outside the area of the attack and accessible to medical personnel, the signs and symptoms of the patients may be the only detection method available, and the threat of spread of the CWA hazard from casualties may be minimal. However, in the case of the Tokyo sarin attack, it has been reported that up to 9 percent of EMS workers and a significant number of hospital staff experienced acute symptoms of nerve agent toxicity due to exposure to casualties in unventilated areas (Okumura et al., 1998b). Given the increased threat of CWA terrorism and the various CW agents that can be used, emergency responders must have accurate and timely detection information or the ability to detect and identify a CWA at the time of their response. Again in the case of the Tokyo subway attack, the first identification made was inaccurate, and it was not until three hours post incident that accurate detection of sarin was made and the information disseminated. The medical personnel on site will require equipment capable of detecting the widest range of chemical agents. For medical personnel, detection equipment may include rapid, minimally invasive or noninvasive clinical assays for various chemical agents or for the effects of the chemical agents, that is, cholinesterase inhibition. Without this ability, more individuals may be exposed, including emergency response and hospital personnel attempting to care for casualties. Chemical agent detection will be an essential part of both medical crisis and consequence management. Detection and identification of the chemical agent or agents at the scene of a terrorist incident must not be accomplished at the expense of rapid and appropriate medical treatment of chemical casualties.

An emergency response incident that involves the release of any chemicals or toxic materials will typically be categorized as a hazardous materials (Hazmat) incident. The response to a Hazmat incident is somewhat standardized across the country. Specialized Hazmat teams are normally called in to address these situations. Hazmat teams are typically part of the fire services and will possess a majority of the locality's chemical detection equipment. The emergency responders who arrive on the scene first, however, must be capable of determining that a Hazmat incident has occurred. These first responders will be the individuals responsible for determining whether the Hazmat team should be called for assistance. Most emergency response vehicles do not carry any chemical detection equipment.

Chemical detection equipment currently used by Hazmat teams varies considerably by locality. For large metropolitan areas, current detectors range from adequate instrumentation to absolutely no capability for CWA detection. Hazmat teams are routinely equipped with a variety of chemical detectors and monitoring kits, primarily chemical-specific tests indicating only the presence or absence of the suspected chemical or class of chemical. A negative response of the test means only that a specific substance is not present in significant quantity; a positive response says nothing about the possible presence of other hazardous agents. Colorimetric tubes, designed for the detection of known and unknown gases, are commonly used by Hazmat teams. There are over 200 different tubes available that can detect a variety of chemicals. The tubes used by responders are sold in basic detection sets typically consisting of the tubes and a hand or mechanical pump. The pump, used to draw the air sample through the tube, will simultaneously carry out a volume measurement with each stroke. Direct reading detector tubes can be used for both short-term measurement and long-term measurement. The long-term tubes consist of two types: one requiring a constant flow pump, and the other a diffusion detector tube. Hazmat team analytical capabilities commonly include tests for chlorine, cyanide, phosgene gas, and organophosphate pesticides. The last of these tests may respond to the military nerve agents, but the requisite validation studies have not been conducted. Colorimetric tubes for the detection of CWA are not standard issue items for Hazmat teams and rarely include a means of detecting the chemical vesicants, such as sulfur mustard or Lewisite.

Many modern detection devices used by Hazmat teams have not been thoroughly tested for their utility and reliability to detect CWA. There is an ongoing effort under the sponsorship of the U.S. Army-managed Domestic Preparedness Program to test currently used Hazmat detection systems against classical CW agents. Preliminary analysis, for example, has shown that combustible gas indicators and pH paper, which are available to most Hazmat teams, will not serve as CWA detectors. The combustible gas detector was designed to detect and measure concentrations of combustible gases and/or vapors in the air, such as carbon monoxide, oxygen, and hydrogen sulfide, while pH paper is simply too generic to be useful as an indicator of CWA.

Currently Available Detection Technologies

A wide variety of detection equipment is available commercially and through the Department of Defense (DoD). Tests, detectors, and monitors of varying sensitivity (lowest level detectable) and specificity (ability to distinguish target from similar compounds) have been developed and/or used by the armed forces to identify the nerve agents and vesicants (Table 4-1). A comparison of the column labeled “Sensitivity” with the data of Table 4-2 reveals that for all but the most expensive of these devices, the sensitivity of most currently available Army systems is adequate for detection of the presence of immediately dangerous concentrations of chemical agents, but too low for them to be appropriate to ensure the complete health and safety of victims and responders. Many currently fielded Army chemical agent detection systems also suffer from excessive false positive alarms, a characteristic which is highly undesirable in a domestic civilian situation, especially in monitoring applications (as opposed to testing for the cause of signs and symptoms of poisoning in on or more patients). Current chemical detection technologies have been incorporated into mobile or stationary detection platforms and can be used as a point-source detector or as a remote (stand-off) detector. The primary differences between mobile detectors and stationary detectors are size, weight, portability, and logistical support requirements. The following section briefly describes the technological basis of this equipment, and Appendix B provides an extensive but perhaps not all-inclusive list of manufacturers that employ these technologies in their instruments.

TABLE 4-1. Military Detection and Monitoring Equipment.


Military Detection and Monitoring Equipment.

TABLE 4-2. Estimated Human Exposure Guidance for Selected Chemical Warfare Agents.


Estimated Human Exposure Guidance for Selected Chemical Warfare Agents.

Ion Mobility Spectrometry (IS) operates by drawing air at atmospheric pressure into a reaction region where the constituents of the sample are ionized. The ionization is generally a collisional charge exchange or ion-molecule reaction, resulting in formation of low-energy, stable, charged molecules (ions). The agent ions travel through a charged tube where they collide with a detector plate and a charge (current) is registered. A plot of the current generated over time provides a characteristic ion mobility spectrum with a series of peaks. The intensity (height) of the peaks in the spectrum, which corresponds to the amount of charge, gives an indication of the relative concentration of the agent present. This IMS technology is mainly used in mobile detectors to detect nerve, blister, and blood agents.

Electrochemical sensors function by quantifying the interaction between an analyte's molecular chemistry and the properties of an electrical circuit. Fundamentally, electrochemistry is based on a chemical reaction that occurs when the CWA enters the detection region and produces some change in the electrical potential. This change is normally monitored through an electrode. A threshold concentration of agent is required, which corresponds to a change in the monitored electrical potential. This sensor technology provides a wide variety of possible configurations. Electrochemical detectors are used in mobile detectors to detect blister, nerve, blood, and choking agents.

In flame photometry, an air sample is burned in a hydrogen-rich flame. The compounds present emit light of specific wavelengths in the flame. An optical filter is used to let a specific wavelength of light pass through it. A photosensitive detector produces a representative response signal. Since most elements will emit a unique and characteristic wavelength of light when burned in this flame, this device allows for the detection of specific elements. Flame photometric detectors are commonly used in gas chromatographs.

Thermoelectric Conductivity. The electrical conductivity of certain materials can be strongly modulated following surface adsorption of various chemicals. Heated metal oxide semiconductors and room-temperature conductive polymers are two such materials that have been used commercially. The change in sensor conductivity can be measured using a simple electronic circuit, and the quantification of this resistance change forms the basis of sensor technology. Thermoelectric conductivity detection technology has only recently been applied to chemical agent detection.

Infrared Spectroscopy. The infrared (IR) region of the electromagnetic spectrum between 2.5 and 25 micrometers has proven valuable for the identification and quantification of gaseous molecular species. When infrared radiation passes through a gas, adsorption of radiation occurs at specific wavelengths that are characteristic of the vibrational structure of the gas molecules. Infrared detectors are used in mobile detectors to detect blister and nerve agent vapors.

Photoacoustic IR Spectroscopy. As in infrared spectroscopy, PIRS uses selective adsorption of infrared radiation by the CWA vapors to identify and quantify the agent present. A specific wavelength of infrared light is pulsated into a sample through an optical filter. The light transmitted by the optical filter is selectively adsorbed by the gas being monitored, which increases the temperature of the gas as well as the pressure of the gas. Because the light entering the cell is pulsating, the pressure in the cell will also fluctuate, creating an acoustic wave in the cell that is directly proportional to the concentration of the gas in the cell. Two microphones mounted inside the cell monitor the acoustic signal produced and send results to the control station. PIRS technology is fairly new, and it is expected that most agents can be detected with this technology.

Photo Ionization Detectors (PIDs) operate by passing the air sample between two charged metal electrodes in a vacuum that are irradiated with ultraviolet radiation, thus producing ions and electrons. The negatively charged electrode collects the positive ions, thus generating a current that is measured using an electrometer-type electronic circuit. The measured current can then be related to the concentration of the molecular species present. PID's are used in mobile detectors to detect nerve, blister, and mustard agents.

Surface Acoustic Wave (SAW) sensors detect changes in the properties of acoustic waves as they travel at ultrasonic frequencies in piezoelectric materials. The basic transduction mechanism involves interaction of these waves with surface-attached matter. Multiple sensor arrays with multiple coatings and pattern recognition algorithms provide the means to identify agent classes and reject interferant responses that could cause false alarms. Acoustic wave sensors are used in mobile detectors to detect nerve and blister agents.

Color-Change Chemistry. This technology is based upon chemical reactions that occur when CW agents interact with various solutions and substrates. The most common indicator (for a positive response) is a color change. Detection tubes, papers, or tickets use some form of surface or substrate to which a reagent solution is applied. Many of these kits are complex and include multiple tests for specific agents or families of agents. Color change detectors can detect nerve, blister, and blood agents.

Raman Spectroscopy is based upon the observation that when radiation is passed through a transparent medium, chemical species present in that medium scatter a portion of the radiation beam in different directions. The wavelength of a very small fraction of the radiation scattered differs from that of the incident beam. The difference between the scattered radiation and incident beam corresponds to wavelengths in the mid-infrared region. The degree of wavelength shift is dependent upon the chemical structure of the molecules causing the scattering. During irradiation, the spectrum of the scattered radiation is measured with a spectrometer. Raman spectroscopy appears not to be applicable for detecting CWA precursors and degradation products in soil samples but has applicability in air samples.

Mass Spectrometry (MS). A sample is introduced into the instrument, a charge is imparted to the molecules present in the sample, and the resultant ions are separated by the mass analyzer component. MS instruments are actually measuring the mass to charge ratio of the ions. A mass spectrum appears as a number of peaks on a graph. This technique only requires a few nanomoles of sample to obtain characteristic information regarding the structure and molecular weight of the analyte. Many mass spectrometers are specifically designed to detect various CW agents and have enormous applicability in detecting agents in most types of samples.

Gas Chromatography (GC) detectors are used to detect a variety of CW agents. Samples are subjected to a volatile solvent extraction. A small sample of the mixture is then injected through a rubber septum into a heated injection port that vaporizes the sample. The vaporized sample is then swept onto the column by the inert carrier gas and serves as the mobile phase. After passing through the column the solutes of interest generate a signal for a recording device to read. The detector is universal in nature in that it can respond to any change in the column effluent or only to solutes possessing a specific characteristic. Like mass spectroscopy, this method also offers high sensitivity and specificity in detecting CWA in many sample forms.

Fourier Transform Infrared (FTIR) Spectrometry. FTIR is a technique that can identify compounds that are separated by gas chromatography. After the separation of the compounds, the sample passes through a light pipe where an infrared (IR) beam is passed through it. The adsorption of the IR energy is monitored as the signal is continuously scanned. Scans are collected on each peak and the signals are then manipulated with a Fourier transform that enhances the signal to noise ratio of the spectra taken. FTIR detectors are used to detect a variety of CW agents.

MMST Equipment

The Metropolitan Medical Strike Teams (MMSTs) being organized and equipped by the Public Health Service (PHS) have purchased M8 and M9 detection paper, M256A1 Detection Kits, M18 Detection Kits, Draegar kits, portable surface-acoustical-wave (SAW) chemical agent detectors (SAW MiniCAD), and chemical agent monitors (CAM).

The M8 and M9 detection papers provide rapid (<1 minute), inexpensive tests for the presence of liquid mustard or nerve agents. Use of the paper is a screening test only, and results must be verified with more accurate methods of detection, particularly because of the paper's propensity to show false positive results for substances such as petroleum products and antifreeze. False positives are especially undesirable in a civilian context, where the mere rumor of “nerve gas” may cause hysteria.

M8 paper is supplied in the M256A1 Kit and the M18A2 Chemical Agent Detection Kit. M8 paper is a preliminary detection technique best suited for detection of liquid CWA on non-porous materials. M8 paper is tan in color and comes in a booklet containing twenty-five 2.5 inch × 4 inch perforated sheets. There are three sensitive indicator dyes suspended in the paper matrix. The paper is blotted on a suspected liquid agent and observed for a color change. V-type nerve agents turn the M8 paper dark green, G-type nerve agents turn it yellow, and blister agents turn it red.

M9 paper is an adhesive-backed, tape-like material designed to be worn on the outside of clothing or placed on vehicles, equipment, or supplies that may be exposed to liquid CWA droplets. The detector responds with a marked, contrasting color change, turning from the original green to red or pink when it comes in contact with a liquid CWA droplet.

The M256A1 kit includes enzyme-based detector “tickets,” which change color to indicate low concentrations of cyanide, vesicant, and nerve agents in vapor form. The tests take approximately 15 minutes. Sensitivity is such that the tickets may provide a negative reading at concentrations below that immediately dangerous to life and health (IDLH) but still as much as 500 times greater than the acceptable exposure limit (AEL). Occupational Safety and Health Administration (OSHA) rules call for the use of maximum personal protection until concentrations can be shown to be less than 50 times the AEL. The IDLH is the maximum concentration of a contaminant to which a person could be exposed for 30 minutes without experiencing any escape-impairing or irreversible health effects. The AEL is a general term indicating a level of exposure that is unlikely to result in adverse health effects.

The M18 detection kit, like the M256A1 kit, is a military item. In fact it might be termed a chemical weapons Draeger tube kit—a colorimetric device for measuring the concentration of selected airborne chemicals. The M18 comes with disposable tubes for cyanide, phosgene, Lewisite, sulfur mustard, and nerve agents GA (tabun), GB (sarin), GD (soman), and VX. Tests for each take only 2 to 3 minutes but must be conducted serially.

The SAW MiniCAD is a commercially available pocket-sized instrument that can automatically monitor for trace levels of toxic vapors of both sulfur mustard and the G nerve agents with a high degree of specificity. The instrument is equipped with a vapor-sampling pump and a thermal concentrator to provide enriched vapor sample concentration to a pair of high-sensitivity coated SAW microsensors. All subsystems are designed to consume minimal amounts of power from onboard batteries. Optimal use of the SAW MiniCAD requires that a suitable compromise be made among the conflicting demands of response time, sensitivity, and power consumption. Maximum protection requires high sensitivity and a rapid response. The SAW MiniCAD is able to achieve a high sensitivity with an increased vapor sampling time. However, a faster response can be achieved at a lower sensitivity setting. Testing of the SAW MiniCAD has been performed with chemical warfare agents GD, GA, and HD. These tests were performed at a variety of concentrations and humidity levels. There were no significant effects noted due to the changes in the humidity levels for any of the chemical agents tested.

The CAM uses ion mobility spectrometry to provide a portable handheld point detection instrument for monitoring nerve or vesicant agent vapors. It provides a graduated readout (low, medium, high). Response time is dependent on concentration but generally takes from 10 to 60 seconds. Minimum levels detectable are about 100 times the AEL for the nerve agents and about 50 times the AEL for vesicants. An obvious drawback to this relative insensitivity to low concentrations is an inability to fully check the efficacy of decontamination efforts, both in the field and subsequently at treatment facilities.

Few local governments or private medical facilities or organizations have invested in CWA detection equipment to date. This may change as the Army's Domestic Preparedness Program provides the training it has promised to 120 of the nation's largest metropolitan areas, but it seems likely that it is not simply information on availability but also the cost of these devices that has limited their procurement. For example, the CAM, a highly specific device designed to detect nerve and vesicant vapors only, costs almost $7,500. The equipment needs of early civilian responders to a domestic incident in which CWA may have been used are also different from those of military personnel in that the military has the advantage of intelligence information that enables the users of the equipment to predict a probable threat agent and the likely area of impact from the chemical agent. For first responders to a domestic terrorist incident, there are currently no such benefits of intelligence. Without such knowledge, first responders will be unlikely to use CWA-specific detection equipment immediately.

Much of today's technology has been developed into commercially available detection equipment, however, and this equipment should allow first responders, whether they be police, fire, Hazmat, or EMS units, to detect the presence or absence of CWA. This equipment is available, reasonably priced, and will detect a wide array of chemical agents. The M9 paper and the M256 kit are simple and inexpensive devices that enable responders to rapidly detect classical CW agents. The photo-ionization detector, the ion mobility detector, the surface acoustic wave detector, and the colorimetric tubes give medical personnel an ability to deal with a wider array of chemicals. As a market evolves for these items of detection equipment, modifications for the civilian community will be made to simplify their usage and the costs associated with their acquisition and maintenance should decrease.

Potential Advances

Current R&D in chemical agent detector technology is focused on increasing the speed and sensitivity of the instruments, while at the same time bringing down their size and cost. The vast majority of next-generation chemical detectors are based on the application of technology previously discussed. In some cases, the improvement will often be utilization of multiple technologies to simultaneously increase the sensitivity and specificity of the instrument. New CWA detector platforms with near-term successful development prospects include:

  • Automatic Chemical Agent Detection Alarm (ACADA-M22). The M22 is an advanced, point-sampling, chemical agent alarm system employing ion mobility spectrometry. It is designed to detect standard nerve and vesicant agents.
  • Improved Chemical Agent Point Detection System (IPDS). The IPDS also employs ion mobility spectrometry and is an improved version of a point detection system. In addition to G nerve agents and VX, the IPDS is designed to detect vesicant agent vapors. It is to be a shipboard instrument, and therefore will be much larger and need more power than portable IMS devices.
  • Joint Chemical Agent Detector (JCAD). The JCAD will employ surface acoustic wave (SAW) technology to detect nerve and blister agents. It is designed to be lightweight and portable and will reduce false alarms. The JCAD will also allow detection of new forms of nerve agents.
  • Joint CB Agent Water Monitor (JCBAWM). The JCBAWM will be a portable device to detect, identify, and quantify CB agents in water. It will allow the user to sample water and receive a digital readout of the contents. The technology to be employed in this monitor is still under review.
  • Joint Service Lightweight Standoff Chemical Agent Detector (JSLSCAD). The JSLSCAD is a passive, infrared detection unit employing Fourier Transform Infrared (FTIR) Spectrometry. The device is designed to detect nerve and blister vapor clouds at a distance of up to 5 km.
  • Shipboard Automatic Liquid Agent Detector (SALAD). Technologies to be used in the SALAD have recently been reviewed, but no decision has been made on the final selection. The instrument is designed to be an automated, externally mounted liquid agent detector capable of detecting G nerve agent and VX and vesicant chemical agents.
  • The Special Operations Forces (SOF) Nonintrusive Detector and the Swept Frequency Acoustic Interferometry (SFAI) detector are portable, hand-held acoustic instruments developed specifically to enable rapid detection and identification of CW agents within munitions, railcars, ton containers, etc.

Various organizations are touting the advantages of new technology that offers the user the ability to determine the composition of potential CWA material within various containers without the need for direct sampling. Prototypes of the SFAI instrument employs a piezoelectric transducer that creates standing waves of sound in the container's contents. Software algorithms utilize the resonant peaks to extract the speed of sound through the contents, the density of the contents, and the attenuation of the returned sound signal. This type of instrument is highly sought after by explosive ordnance units, but offers little utility to medical personnel responding to a chemical release.

Tremendous efforts, primarily sponsored by the DoD, are under way to improve chemical agent detectors. The advances that will be of greatest benefit to the first responding medical teams will be increased portability, greater ease of use, and increased reliability of the detector technology. Where the application of new CWA detection technologies could be of greatest potential benefit to the medical community is in fixed medical facilities and patient transport vehicles for monitoring air samples for low levels of CWA that may cause occupational hazards. Additionally, standoff detection equipment can also assist medical planners in obtaining preincident intelligence so critical to providing the appropriate emergency response.

Medical personnel must rely on accurate and timely information provided by the earliest responders on the scene. If medical teams are expected to be the earliest responders to the scene of a mass casualty incident involving chemical agents, then they should be provided with reliable detection equipment as well as training on the use of the equipment. There should be continued support for the Public Health Service efforts to equip Metropolitan Medical Strike Teams with effective and currently available chemical agent detection equipment. These detectors are reliable, relatively inexpensive, and provide for the detection of all classical chemical agents that may be utilized in a domestic terrorist incident. Furthermore, efficient and cost-effective portable hand-held CWA detectors employing photo ionization detectors, surface acoustic wave microsensors, or ion mobility spectrometry should be readily available to all Hazmat units expected to respond to a potential CWA incident.

Information must flow along clear lines of communication and there must be standard procedures for relaying vital detection information. Rapid and sensitive assays for CWA could assist in limiting further exposures as well as providing verification and justification for initiation of appropriate therapy. Most importantly, medical response teams must be educated as to the nature and properties of CWA, be trained in recognizing the signs of CWA exposure, and be prepared to treat the symptoms caused by their toxicity. Detection and identification of the CWA is critical for legal and forensic purposes and for minimizing the transfer of contamination to unprotected personnel. However, detection and identification of the agent must not be the primary goal of the early medical response units; rather, it must be seen only as an aid to them in providing rapid and appropriate medical services to the victims of the incident.


Rapid diagnosis of patients who have been exposed to a chemical agent will be important to saving lives and preventing further injury. The signs and symptoms of the patient will provide the most important information on which to base emergency treatment. Generally, instead of detecting an agent in the body, the clinician must look for some byproduct of the agent or a particular biochemical interaction within the patient that would indicate that an exposure to an agent has taken place. The specific biochemical interactions (such as, cholinesterase inhibition, thiodiglycol in urine) then lead the clinician to a determination of the likely agent. This section will examine methods for clinical analysis of four categories of agents: nerve agents, vesicating agents, respiratory agents, and cyanide.

Nerve Agents

Persons exposed to high concentrations of organophosphorus nerve agents usually develop signs and symptoms within a matter of minutes after exposure. Therefore, initial patient diagnoses and treatment are likely to be based on observations of signs and symptoms by the paramedic or other health care professional on the scene. Emergency medical personnel are, in any case, not equipped, trained, or encouraged to attempt clinical chemistry.

At the hospital level, treatment will continue to be guided by vital signs and clinical symptoms and monitored at this level by electrocardiogram, pulse oximetry, chest X-ray, arterial blood gas measurement, and other measures of physiologic status. Because nerve agents inhibit cholinesterase activity, laboratory tests estimating the level of this activity in red blood cells or plasma are sometimes used in estimating the degree of acute exposure. However, many hospitals cannot perform this test on site. Enzyme inhibition may only be loosely correlated with clinical signs and symptoms, and, because of high interindividual variability, only comparison between a baseline level of inhibition and the level just after exposure to a nerve agent will provide unambiguous evidence of a small or moderate exposure to nerve agents. A good example of this was reported in a recent study of 66 Japanese victims exposed to sarin (Masuda et al., 1995); patients exhibiting moderate symptoms of intoxication had serum values ranging from 300–750 IU/L. Normal serum cholinesterase activity ranges from 182 to 804 International units per liter (IU/L). These patients had red blood cell (RBC) ChE activity ranging between 0.3 and 2.0 IU as compared to 1.2–2.0 for patients not showing symptoms. Plasma ChE recovers in 30 to 40 days and RBC ChE recovers in 90 to 100 days after exposure to organophosphorus nerve agents (Grob et al., 1953).

Methods for Measuring Cholinesterase Inhibition

The standard methodology for determining blood ChE inhibition is based on the measurement of the enzymatic products derived from either acetylcholine or acetylthiocholine as substrates. The simplest and most convenient method is a colorimetric procedure (Ellman et al., 1961) in which thiocholine, the product of substrate acetylthiocholine hydrolysis, is detected by reacting with 5,5′-dithio-bis (2-nitrobenzoic acid) (DTNB). The time course of the reaction, monitored spectrophotometrically at 410 nm with and without various concentrations of the agent, is used to calculate the concentration of agent in the sample. A recently developed portable device utilizing this method, the Test-Mate™ OP Kit (EQM Research Incorporated, 2585 Montana Avenue, Cincinnati, OH 45211), provides a rapid, reasonably sensitive and reliable assay for ChE inhibition from potential OP exposure. The kit may prove suitable for use in a wide range of contingencies. It is recommended that the kit be used to establish ChE levels when exposure to nerve agents is suspected; it can be utilized at the site of an incident or in a hospital setting. This kit can determine RBC AChE and plasma butyrylcholinesterase (BChE) activities within minutes, requiring 10 µl of blood per determination.

Other procedures for determining serum ChE include liquid chromatography (Miller and Blank, 1991), an amperometric method using a hydrogen peroxide electrode, and the enzyme choline oxidase immobilized on a nylon net (Palleschi et al., 1990). Perhaps the most promising developments for screening kits for field use are immunochemical methods utilizing various murine monoclonal antibodies to acetylcholinesterase and ELISA (Novales-Li and Priddle, 1995).

Other Methods for Detection of Nerve Agent Exposure

A number of chemical methods have been used for the direct measurement of nerve agents and their metabolites in plasma and other body fluids. Among the most sensitive and reliable techniques are capillary gas chromatography (GC) (Bonierbale et al., 1997), gas chromatography-mass spectrometry, and gas chromatography-tandem mass spectrometry (Black et al., 1994). To date the most sensitive methods of retrospective detection of organophosphorus anticholinesterases are described by Polhuijs et al. (1997), who analyzed blood samples obtained from victims of the Tokyo subway incident. Fluoride ions are used to reactivate the inhibited enzyme, thus converting the OP moiety into the corresponding phosphofluoridate. It is suggested that this sensitive assay can be of benefit in biomonitoring of exposure for health surveillance, in cases of suspected use of nerve agents or pesticides, in medical treatment of OP intoxication, and in forensic cases against individuals suspected of handling anticholinesterases. Of practical utility are the GC/MS approaches of the U.S. Army (TB MED 296) for the determination of metabolites of sarin, cyclosarin (GF), and soman in urine. In animals exposed to the toxic organophosphorus nerve agents, substantial amounts of the parent compounds are hydrolyzed to their corresponding phosphonic acids (the rest is covalently bound to enzymes and tissue proteins) (Harris et al., 1964; Reynolds et al., 1985; Lenz et al., 1984, 1987). These methods are designed to detect these polar acids for verification of exposure. Urinary excretion of the metabolite is the primary elimination route for these three compounds. The major differences among them are primarily the extent and rate of excretion. Nearly total recoveries of the given doses for sarin and GF in metabolite form were obtained from urine, while soman was excreted at a slower rate with a recovery of only 62 percent. In animal studies, the acid metabolites can be detected in urine for 4 to 7 days postexposure. The development of ELISA and monoclonal-antibody-based detection systems holds great promise in simplifying and hastening the detection of nerve agents in biological samples. Efforts have been made to develop immunoassays to chemical warfare agents (Lenz et al., 1992, 1997). Monoclonal antibodies were developed against a structural analog of soman, and, when employed in a competitive inhibition enzyme immunoassay, were capable of detecting the nerve agent at a level of 80 ng/ml.

Vesicating Agents

Methods for the detection of mustard have been described using GC/ MS, based on the selective release by Edman degradation of the N-terminal valine adduct of hemoglobin with the agent (Fidder et al., 1996a; Noort et al., 1997) or by the determination of N7-(2-hydroxyethylthioethyl)-guanine, a novel urinary metabolite of sulfur mustard (Fidder et al., 1996b). The most abundant adduct, N1/N3-(2-hydroxyethylthioethyl)-L-histidine, is analyzed by LC-tandem MS, enabling the detection of exposure of human blood to 10 mM sulfur mustard in vitro. Verification of exposure to sulfur mustard in casualties of the Iran-Iraq conflict was conducted using these methods (Benschop et al., 1997). Gas chromatography-tandem mass spectrometry (GC-MS-MS) is also used to analyze urinary metabolites of sulfur mustard, derived from the beta-lyase pathway and from hydrolysis (Black et al., 1994). Another procedure utilizes GC-MS with ion chemical ionization to detect alkylated valine and histidine adducts of hemoglobin from casualties of sulfur mustard poisoning (Black et al., 1997a,b). Methods currently employed by the Theater Army Laboratory are those described in TB MED 296 (U.S. Army, 1995). In general, mustard cannot be simply assayed from urine because of its reactive nature. Thiodiglycol (TDG) is one of the in vivo degradation products of HD and can be used to confirm an exposure (Jakubowski et al., 1990; Davison et al., 1961; Roberts et al., 1963), although TDG is itself subject to chemical and enzymatic transformations. In this method, detection of TDG after derivatization with heptafluorobutyric anhydride is achieved by using a gas chromatograph coupled with a mass selective detector. The lowest quantifiable concentration is 5.0 ng/ml. Thiodipropanol is used as a stabilizer and octa-deuterated thiodiglycol as an internal standard. Again the advent of immunoassays for the detection of CW agents in body fluids holds the greatest promise in simplifying and reducing the time and cost to detect the presence of agents or their metabolites (Lenz et al., 1997).

Respiratory Agents

For sensitive and relatively specific detection of phosgene, a spectrophotometric GC-MS technique has been used that utilizes a reagent consisting of 0.4% of nitrobenzyl pyridine and 0.5% of sodium acetate in ethanol (Dangwal, 1994). The method is so sensitive as to detect phosgene at a concentration of 0.1 microgram/ml in the sampling solution with the coefficient of variation (CV) of 4.5%.


Four main laboratory findings are indicative of cyanide exposure: (1) an elevated blood cyanide concentration is the most definitive (most medical centers are unable to perform this measurement); (2) metabolic acidosis with a high concentration of lactic acid (lactic acidosis may result from a variety of conditions and is not in itself evidence of cyanide poisoning); (3) oxygen content of venous blood greater than normal (very difficult to measure and not specific for cyanide poisoning); and (4) presence in blood of cyanohemoglobin, which shows a characteristic absorption spectrum by UV spectrophotometry. As in the case of the nerve agents, however, the effects of cyanide (syncope, seizures, coma, respiratory arrest) occur so rapidly that treatment must begin long before laboratory findings are available.

Various methods have been used for CN detection in the blood (Feldstein et al., 1954; Lundquist et al., 1985). Most of them involve prolonged specimen preparation using diffusion or bubbling procedures, both of which require large blood volumes to achieve desired sensitivity. A sensitive and simple method for determining cyanide and its major metabolite, thiocyanate, in blood involved derivatization and determination by gas chromatography and mass spectrometry (Kage et al., 1996). The detection limits of cyanide and thiocyanate were 0.01 and 0.003 mmol/ml, respectively, while the gross recovery of both compounds was 80 percent. In another gas chromatographic procedure, cyanide was converted to cyanogen chloride by reaction with chloramine T and the product analyzed by electron-capture with a detection limit of 5 mg/L (Odoul et al., 1994). Cyanide determination in whole blood can be performed by spectrophotometry after using diffusion coupled with coloration by hydroxycobalamin in a Conway dish. The technique may be accelerated by the use of a heating sheet at 45°C. The method proved to be specific, sensitive, and fast, thus permitting measurements in emergency situations. A possible alternative to the GC-MS approach described above is an automated fluorometric measurement described in TB MED 296 (Groff et al., 1985). The CN assay methods provide direct measurement of plasma-free CN and the stabilization of total CN in blood. Samples for both plasma-free CN and total-blood CN are assayed directly without prior isolation of CN by a completely automated method requiring only 16 minutes from sampling to readout.


R&D needs in chemical detection vary from simply evaluating what already exists to developing better procedures and methods for better detecting exposure levels. The ultimate goal of chemical detection is to rapidly and inexpensively detect and perhaps identify toxic substances that threaten to endanger both responders and victims. To this end, the committee has identified the following list of research needs:

4-1 Conduct a thorough evaluation of all industrial chemical detection equipment in the inventory of Hazmat and EMS units for its sensitivity and specificity for detecting CWA.

4-2 Continue research efforts to miniaturize and reduce the acquisition costs of GC/MS technology that would monitor the environment within fixed medical facilities and patient transport vehicles.

4-3 Develop standard operating procedures for communication of CWA detection information from first responders to Hazmat teams, emergency medical services, and fixed medical facilities.

4-4 Direct research efforts towards the development of simplified, rapid, and inexpensive methods of determining exposure to and level of intoxication from chemical agents in clinical samples. Give highest priority to research focusing on immunoassays for such detection, but research should also be conducted to determine the suitability of currently available portable instrumentation for rapid determination of cholinesterase inhibition by EMS units and in fixed medical facilities.

Copyright 1999 by the National Academy of Sciences. All rights reserved.
Bookshelf ID: NBK230668


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