MILITARY MEDICINE, 167, 8:628, 2002

Laboratory Conditions and Safety in a Chemical Warfare Agent

Analysis and Research Laboratory
Guarantor: COL Turan Karayilanoglu
Contributors: MAJ Levent Kenar; COL Turan Karayilanoglu: Songul Kose

Toxic chemicals have been used as weapons of war and also as gene); (4) cyanides (hydrogen and cyanogen chloride); (5) inca-
means of terrorist attacks on civilian populations. Research pacitants (3-quinoclinidinyl benzoate, lysergic acid diethylam-
focusing on chemical warfare agents (CWAs) may be associ- ide); (6) riot-control agents (lacrimators and vomiting agent);

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ated with an increased risk of exposure to and contamination and (7) herbicides (cacodylic acid, picloram).
by these agents. This article summarizes some of the regula- Chemical agents, likeall other substances, may existas sol-
tions concerning designation and safety in a CWAanalysis and
research laboratory and medical countermeasures in case of ids, Itquids, or gases, depending on temperature and pressure.
an accidental exposure. The design of such a laboratory, cou- Chemical agents in preparations and military munitions are
pled with a set of safety guidelines, provides for the safe con- generally liquids. After detonation ofthe container, the agentis
duct of research and studies involving CWAs. Thus, a discus- primarily dispersed as Iiquld oran aerosol, soa collection ofvery
sion of decontamination and protection means against CWAs smallsolid particles orliquid droplets can be suspended in gas.2
is also presented. For many years, attention has centered on chemical com-
pounds with a military application. The need for research con-
cerning CWAs has also posed a potential risk to scientists and
Introduction toxicologists ofa laboratory accident. However, given the phys-
he Chemical Weapons Convention, signed bymore than 130 icalproperties ofthe agentslistedin Table I, reducing potential
T participants in Paris in January 1993, enteredinto force in risks is a great concern.'
January 1995. To create confidence in the analytical results
under the Chemical Weapons Convention, the standardization Designation and Safetyin a CWAARL
ofequipment and procedures shouldbe established.
The analytical procedures can be conducted to perform (1) Designation
screening and preliminary identification of the chemicals, (2) Aresearchlaboratory assigned to study chemical weapons is
confirmation ofthe preliminary identification, (3) elucidation of designed to support a variety of chemical activities involving
the structures of unknown agents, (4) quantitation of well-de- hazardousmaterials. In such a laboratory, chemical wastestor-
fined agents, and (5) research activities covering all of these ageshouldbe controlled, techniques forhandling and storage of
analytical procedures. These procedures shouldbe followed by chemical toxins and decontamination facilities should be well
competent technicians and investigators familiar withthe basic defined and controlled, and protective systems, including per-
principles ofanalytical methods and working in a well-equipped sonal protective devices, shouldbe established. 3
laboratory. 1 The features of a laboratory designed to accommodate the
Chemical warfare agents(CWAs) under analysis and research safe handling and working with CWAs and like substances are
have some their own specifications requiring a different ap- described below.
proach. Sampling, sample preparation, cleaning of glassware,
handling of the samples, transport of the samples, analytical
Working Area
conditions, protective measures, decontamination and antidotal
therapy, and casualty management need to be specialized for Thelaboratory room shouldbe separatedfrom otherworking
these toxic warfare substances. Thus, laboratory safety rules areas.TheCWMRL has to consistofthree mainareas: a dress-
and other instructions are to be strictly obeyed during studies ingroom, an enter room, and the mainlaboratory. Because the
performed in a CWA analysis and research laboratory majority of studies in the laboratory are performed in contain-
(CWMRL).l ment devices, personnel usually do not take offtheir everyday
clothes; rather, they wear protective laboratory suits and
Overview of CWAs
masks. The main laboratory area contains general working
CWAs are toxic substances that are capable of producing space, spectal containment devices, auxiliary instrumentation,
incapacitation, serious Injury, and death. They may be used and storage areas. There should also be at least two large ob-
either on the battlefield or in a terroristattack.2 servation windows. The location of the observation windows
These agents can be classified according to their mecha- shouldallow researchers inside the laboratory to be in view ofa
nisms: (1) nerve agents (tabun, sarin, soman, VX); (2) vesicants coworker outside.
(sulfur mustard, lewisite); (3) choking agents (chlorine, phos- Additionally, an interlaboratory communication system,
which is always to be on when studies are being performed,
Department ofNuclear, Biological, andChemical, Gulhane Military Medical Acad- permits convenient communication among laboratory person-
emy, 06018 Ankara, Turkey.
This manuscript wasreceived for review inMay 2001. The revised manuscript was nelworking insideand outside the mainlaboratory.' Laboratory
accepted for publication in February 2002. bench space, eye wash stations, emergency showers, fire extin-
Reprint & Copyright © byAssociation ofMilitary Surgeons ofU.S., 2002. guishers, gas connection, a glove box, a gas chromatography

Military Medicine, Vol. 167, August 2002 628

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PHYSIOCHEMICAL CHARACTERISTICS OF SOME COMMONLY STUDIED CWAs IN RESEARCH LABORATORIES ~
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Tabun Sarin Soman VX Sulfur Mustard Lewisite HCN CN ~
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Chemical name 0- Ethyl
dimethyl Isopropyl Pinacolyl 0- Ethyl
S-diisopropyl Bis(2-chloroethyl) Dichloro(2- Hydrogen cyanide I-Chloro- "'1
amldophosphoryl methyl- methyl- aminomethyl sulfide chloro(vinyl)arsine acetophenone ~
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cyanide phosphono phosphono ~
fluoridate fluoridate
Vapor Pressure 0.037 2.9 0.40 0.0007 0.72 0.39 740 0.0041
(mmHgat
20°C)
Liquid density 1.08 1.10 1.02 1.008 1.274 1.89 0.68 1.187
(g/mL at
25°C)
Freezing point -5 -56 -42 <-51 14.4 -18 -13.3 54
(OC)
Boiling point 230 158 198 298 217 190 25.7 248
(OC)
Volatility (mg/ 610 22.000 3900 10.5 610 4480 1.1 X 10 6 34.3
m" at 25°C)
LD so (mg/min/ 15,000 by skin (vapor) 10,000 by skin 2,500 by skin 150 by skin (vapor) 900 (inhaled); 1,200-1,500 (inhaled); Varies widely with 7,000 to 14,000
m'') or 1,500 liquid: 70 (vapor) or (vapor) or 350 or 5 liqutd: 15 5,000 by skin 100,000 (skin) concentration
inhaled 1,700 Iiquid: ltquid: 35 inhaled (vapor) or
35 inhaled inhaled 1,400 liquid
~ Physical Fairly fruity No odor Fruity, oil of Rotten fish odor Onion, garlic Amber to dark brown Odor of bitter Fragrant like
recognition camphor odor to black almonds or apple
f peach kernels blossoms
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('l) LD5o, lethal dose for 50% of the population.
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630 Safety in a Chemical Warfare Agent Laboratory

area, a high performance liquid chromatography apparatus, Storage

and a fume hood should also be available in the laboratory
area."
Decontamination System
A separate shower system, decontamination system, and
dressingarea should be provided.
Alarm System
There should be an alarm system available to provide safety
against spills, accidents, and systemfailure detection.
Theproperstorage and use ofchemicals is necessary to avoid
dangerssuch as bums, explosions, fires, and toxic fumes. Thus,
knowledge of the properties of the chemicals in use and of
proper handling procedures greatly reduces dangerous situa-
tions." Afew importantsafetyitemsare described briefly below."
Handling and Transportation
Careful attentionin a CWAARL must be paid to the appropri-
ate collection, handling, and transport offorensic and pathology
specimens7 and to the disposal of chemical/biological hazard-

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ous materials, including allbiological and otherrelated(soil, air,
Air-Lock System water) specimens." Toxic agents or samples contaminated by
Chemical agents and related equipment should be treated them also must be transported by technicians in special con-
through a well-organized air-lock system. tainers having a card alarm system, thereby creating additional
physical workas well as potentially hazardous situations."
Laminar-Flow Cabinet Equipment
Exhaust air is filtered via a high-efficiency particulate air Experimental Animals
filter, and wastewatershouldbe disposed ofaccording to safety Experimental animalsare commonly used in investigations of
rules. the effects, actions and mechanisms, and diagnostic and ther-
apeutic faculties of CWAS.8,9 The most commonly used labora-
Personal Protective Clothing tory animals are guinea pigs, rabbits, and rats.'" Various an-
Personal protective clothing and equipment, including a typ- esthetic methods via different routes are used to sedate the
icalfull-face respirator, are required in this laboratory. However, animals to keep them stable. The most commonly performed
somehealth problems can arise because ofthe use ofprotective methods are ketamine injection (0.5 mg/kg) and administration
clothing and equipment, Full-face respirators can createvisual of halothane/oxygen/nitrous oxide by mask.'? Experimental
distortion, cause breathingto be more difficult, and impairver- animalsshould be kept in the hood until the end ofthe experi-
bal communication. Mobility can alsobe inhibited by protective ment; after they are killed, they should be packed in a closed
suits." polyethylene nylon sack and immediately burned.'! The incin-
erator located in the destruction facilities is one of the integral
Destruction Facility units ofa CWAARL and shouldbe located in a separate area far
from laboratory.
All contaminated materials with CWAs and other laboratory
tools shouldbe destroyed by appropriate techniques or burning Decontamination Facilities and Shower System
systems such as incinerators.
Eye washers or face washers that connectto existing plumb-
Ventilation-Exhaust System
ing or to a specially filled container should be available in a
position next to the working area. They must be located at the
A ventilation-exhaust system is one of the most important entrance of the laboratory, and first-aid materials and medical
design features ofany laboratory. An emergency-purge ventila- drugs as well as an assigned physician must be stationedin the
tion system is also necessary, which would be activated in a decontamination facility. Each CWAARL should have at least
toxic leakage alertedbyan alarmsystem. Thebalanceofair flow one shower system permanently established for the manage-
is adjusted to provide a negative pressure. Most of the air is ment ofexposed personnel. Drainage from the shower must be
supplied through the main building service. collected in signed containers and disposed ofsafely so that the
wastedoesnot enter the community sewer system." Flammable
Fire Exiinquishet
solvents and chemical hazardousmaterials must be located and
Afire extinguisher is necessary to put out fires in the labora- treated in convenient sites such as safetycabinetsand laminar
tory. Everyone in the laboratory shouldbe instructed in the use flow hoodsin the laboratory." Polyethylene pumps,pipettes, and
of these extinguishers or any other fire-fighting equipment. 5 pipettebulbs and filters should be handled to pump and spill.
Each employee should know the location ofavailable fire-fight- All contaminated glassware and disposable materials shouldbe
ing equipment and how to use it and where the evacuation neutralized for at least 48 hours in solutions containing 1:1 N
routes are." NaOH/ethanol. l l

Safety in a Study with CWAs Laminar Flow Hood

A large number of safety items should be available for a
CWAARL. Implementation of these standards should be based
on improving researchers'health, safety, productivity, and work
quality. Tooptimize the design ofa laboratory, designers should
workclosely withchemists, technicians, and safetypersonnel to
best evaluate the needs.'
A laminar flow hood is a necessity for every CWA research
laboratory. It is the only safe place to work with any material
that gives offtoxic and harmfulvapors, to prepare reagents, to
apply toxins to cell cultures and experimental animals, and to
heat flammable solvents" (Fig. 1). All stages of any studies and
animal experiments should be performed in a permanently op-

Military Medicine, Vol. 167, August 2002

Safety in a Chemical Warfare Agent Laboratory 631

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Fig. 2. AIllaboratory staff members need to use protective equipment during all
stages of any study related to CWAs.
Fig. 1. AIl samples. including experimental animals. should be treated within a
laminar flow hood cabinet. Chemical Hygiene and Safety Plan
Ahygiene and safety plan must be established in a CWMRL.
erated fume hood with air flow. The hoods are constructed of Elements that should be covered in a chemical hygiene and
stainless steel and are of the auxiliary air type. If the hood safety plan are listedin Table 11. 6
malfunctions, an audible alarmsounds and the hood air supply
fans automatically shut off.' What to Do in Case of an Exposure to CWAs
Ifindividuals are accidentally exposed toan agent, rapididen-
Medical Countermeasures before Working tification mayassist in determining earlymedical interventions.
Clinical signs and symptoms in exposed personnel may be the
Protective Equipment most useful indicators of the likely agent and will be critical in
Spattering from acids, caustic materials other than CWAs, guiding emergency medical care.? Therefore, laboratory staff
represents the greatesthazardto body, eyes, and clothing and is should be aware of the clinical signs and symptoms of CWA
a potential sourceofchemical bums. Heat-resistant gloves and exposure (Table III).l3
rubber gloves with high resistance to chemicals and organics One of the most difficult problems faced in laboratories that
should be worn when handling agents and other chemicals. handle hazardous compounds is contamination and decontam-
They also should be tear resistant. Safety goggles should be ination. Iflaboratory staffand equipment are exposed to a CWA
available in many sizes and shapes." Gloves and boots are al- oranysolution containing the agent, decontamination is needed
most always made ofbutyl rubber or similar impermeable ma- to eliminate the hazard. Decontamination is defined as the re-
terial that is resistant to CWAs.2 Laboratory staff members duction or removal ofchemical (or biological) agentssothat they
wearing contact lenses should be aware of the danger of toxic are no longer hazardous. Agents may be removed by physical
chemicals. becausewearing such lensescan make it difficult to means or be neutralized chemically (detoxification).
irrigate the eye properly. Safety face shields made of polypro- Three basic methods of decontamination are physical re-
pylene should be used for efficient eye and face protection." moval, chemical deactivation, and biological deactivation. Bio-
Protective masks usually consist of filters with canisters filled logical deactivation has not beendeveloped to the pointofbeing
with activated charcoal, which filters out CWAs as air passes practical.14
through."
TABLEII
Proper storage of CWAs and flammable organic solvents is
important for the safety ofthe laboratory. Organic solvents such TOPICS OF A CHEMICAL HYGIENE AND SAFElY PLAN THAT MUST
as methanol, dichloromethane, ethyl acetate, and chloroform BE SPECIFIED FOR A CWAARL
are generally used for extraction procedures ofCWAs in soil, air,
Glossary of terms and identification
and watersamples. Theiruse in a CWMRL represents a poten- Standard operating procedures
tial hazard via the inhalation of toxic vapors and skin contact. Criteria used to determine and implement control measures
These solutions should be used in a laminar flow hood and Chemical labeling and storage
stored in a specially designed cold room inside the laboratory Environmental monitoring and maintenance of safety devices
that is equipped witha magnetic card alarmsystem. Toprevent Hazard ous waste
skinand mucousmembrane contamination from materials con- Educ ation and training of laboratory staff
taminated with the agents and solutions containing them or Personal prot ective equipment
their vapors, personal protective equipment, including protec- Protocol for working with particular CWAsand simulants
tive suits, gloves, face shields, masks, mouth pieces, pocket Decontamination and first-aid in a chemi cal agent accident
Medical consulta tion and examination
masks, and resuscitation bags, must be used by laboratory
Record keeping and database
workers't" (Fig. 2).

Military Medicine, Vol. 167, August 2002

632 Safety in a Chemical Warfare Agent Laboratory

TABLEm
EACH STAFFMEMBER MUSTBE AWARE OF THE PATHOLOGICAL EFFECTSAND THERAPEUTIC APPROACH AND MEDICAL CARE INTHE
CASE OF EXPOSURE TO CWAS

Agent Toxicity Signs and Symptoms Treatment

Nerve agents: tabun, Immediately life-threatening Miosis, excessive sweating, increased
sarin, soman, VX bronchial secretion, drooling,
vomiting, diarrhea, in large dose;
immediate loss of consciousness,
convulsions, flaccid paralysis
Atropine sulfate (2-8 mg intramuscular
or intravenous), pralidoxime chloride
(14-28 mg/kg intravenous) in 100-
150 mL of saline given slowly over
15-30 minutes, 10 mg of diazepam,
ventilation and cardiac support if
available

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Vesicants: sulfur mustard, Delayed effects; in large dose, Erytheme, vesication, bums, eye, lung, No antidote for mustards;
nitrogen mustard, life-threatening if untreated and skin damage, respiratory effects, decontamination within 2 minutes,
lewisite leukopenia, thrombocytopenia, bum care, eye therapy, and
decrease in RBCs red blood cells, respiratory support; intramuscular
sepsis administration of bronchoalveolar
lavage and its analogs for lewisite
Choking gases: rhosgene Delayed effects; in large dose, Ocular and respiratory irritation, No antidote; close observation for at
life-threatening if untreated dyspnea, massive pulmonary edema, least 4 hours, symptomatic treatment
hypotension, bronchospasm,
increased bronchial secretion
Blood agents: hydrocyanic Immediately life-threatening Dizziness, nausea, vomiting, weakness, Administration of a cyanide antidote kit
acid, cyanogen chloride respiratory distress, loss of containing amyl nitrite, sodium
consciousness, convulsions, apnea, nitrite, sodium thiosulfate, and
death supplemental oxygen
Riot-control agents: Rarely life-threatening Nose and eye irritation, tearing, Symptomatic treatment
chloroacetophenone, blepharospasm, coughing, mild
o-chlorobenzlidene dyspnea
malononitrile
Incapacitating agents
Tranquilizers Possibly life-threatening Hypotension, paralysis, loss of Observation, symptomatic treatment
consciousness
3-Quinoclinidinyl Rarely life-threatening Sedation, confusion, hallucinations, As antidote, physostigmine salicylate
benzoate disorientation, impaired memory, intramuscular or oral, observation
incoherent speech, mydriasis with and reassurance in a quiet place
paralysis of accommodation, dry
mouth and skin, tachycardia

Themostimportantand mosteffective decontamination after
anychemical orbiological exposure is that performed within the
first 2 minutes after exposure to prevent further absorbp-
tion. 14 ,15 This may be accomplished by washing with soap and
frequent changes ofwater. However, decontamination is oflittle
value againstvaporexposure." Decontamination ofskin is the
primary concern, but decontamination of eyes and wounds
must alsobe done whennecessary. Many substanceshavebeen
evaluated for their usefulness in skindecontamination. An ideal
decontaminant is one that rapidly and completely removes or
detoxifies all known CWAs. Because chemical agentsmaybegin
penetrating the skin before complete reactive decontamination
(detoxification) takes place, early physical removal is most im-
portant."
Washing the area with oil, kerosene, or gasoline followed by
copious washing with soap and water has also been recom-
mended, as has usingneutralhypochloride (1%chlorine). Some
resins, such as Ambergard XE-555 and Ambergard XE-556,
which readily absorband detoxify the agent, can alsobe used for
decontamination ofskin and eyes. 15 Thechloramide compound
1,3,4,6-tetrachloro-7,8-diphenyl-2,5-diiminoglycoluril (S-330)
was also found to be a strong reactant in dermal formulations
for the decontamination of sulfur mustard." Some household
products such as flour, talcum powder, salad oil, and dryIwet
tissue paper are also available to protectworkers. Adry decon-
tamination (e.g., Fuller's Earth or other absorbant powder) can
be used if no water is available. Clothing must be changed
immediately. 15
Contamination ofthe eyes by agents or solutions containing
them requires immediate decontamination by the individual
who has been splashed. Eyes may be irrigated with alkaline
solutions such as 2 to 5% NaHC0 3 orwithneutralizing solutions
such as 0.50/0 chloramine T dissolved in chlorinated paraffin or
chlorinated diphenylether. Flushing the eyes withchloramine T
for the first 15 minutes after contact may prevent further eye
damage." Simple water and soap can provide effective decon-
tamination in many instances. When needed, 5% hypochlorite
or household bleach is commonly used for skin decontamina-
tion.?
At the time of any accident, the building air supplyventing
inside the laboratory should be automatically closed, and all
external air supply fans on fume hoods should be turned off.
This air is introduced into the laboratory through two supply
units mounted near the ceiling. In this emergency-purge mode,
air in the main laboratory can be exhausted very rapidly (ap-

Military Medicine, Vol. 167, August 2002

Safety in a Chemical Warfare Agent Laboratory
proximately 12 to 15 air changes per hour); thus, within a few
minutes, the room air is exchanged almost completely.'
After an accidental spillage ofthe chemical agentand related
materials, the spillarea shouldbe cleaned withdecontaminants
and cleaning solutions until no droplets remain. All containers
and bottles shouldbe kept well stoppered." During an accident
with the chemicals, the respiratory system must be protected
against aerosol and gases in the air, and at the same time, the
rest of the body must be protected against direct contact with
CWAs in the form of Itquid or solid particles. 3
Decontamination generally involves using a water-based
bleach material to neutralize any agent present. Soaps and
detergents are effective in physically removing contamination. A
0.5°;6 sodium hypochlorite solution (diluted household bleach) is
the mosteffective method fordecontamination ofindividuals. A
5.0°;6 solution may also be used to decontaminate equipment.
However, becausethesemethods will only physically remove the
agents and not neutralize them, casualty-producing levels of
contamination may remainin the runoffwater, which must be
considered contaminated. Sodium hydroxide and sodium hypo-
chlorite are two common constituents of decontaminant solu-
tions."
Liquids are best for decontaminating large or irregular sur-
face areas. For hypochlorite to be most effective, it has to be
relatively fresh (made daily or more frequently, particularly in a
warm environment in which evaporation occurs) and have a
concentration of 0.5% at an alkaline pH (pH 10-11). Hypochlo-
rite solutions are for use on skin and soft tissue wounds only.
Surgical irrigation solutions shouldbe used in liberal amounts
in the abdomen and chest. Only copious amounts of water,
normal saline, or eye solutions are recommended forthe eye. 14
Most importantly, in a CWA accident in a CWAARL, rescue
staff must also protect themselves from contamination by
means ofmasks, gloves, and overgarments or by ensuringthat
the exposed personhas beenthoroughly decontaminated." After
the patientis completely decontaminated, antidote therapy(de-
fined in Table III) and symptomatic treatmentshouldbe admin-
istered. The intensityof treatment can be varied based on the
patient'spresenting signsand symptoms and the severity ofthe
exposure.
Conclusion
Laboratory staffworking in a CWAARL shouldbe aware ofthe
basicconcepts needed to recognize the clinical signsand symp-
toms of a chemical agent exposure and the appropriate thera-
peutic approaches to manage chemical agent casualties. Al-
though the chemical agents discussed here are unique to the
633
military, the hazards to personnel working with these sub-
stances must be taken into consideration in designing such a
laboratory. Aspecified system, consisting ofa decontamination
unit, alarm unit, laminarflow hood, protective equipment, de-
structionfacility, and ventilation-exhaust unit, must be estab-
lished. Moreover, in the case of a laboratory accident, proper
treatment and adequate decontamination must be provided as
quickly as possible to avoid serious injuries. Therefore, a
CWAARL must be designed to protect againstthe mostdanger-
ous effects ofCWAs; these measuresare essential to protect the
health and life ofthe working staff.
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Military Medicine, Vol. 167, August 2002