DRAFT CRITERIA  DOCUMENT
                         FOR CARBON  TETRACHLORIDE
                              FEBRUARY 1984
                               Prepared  by
                           JPB Associates,  Inc.
                       Contract No.  2-813-03-644-09
                          HEALTH  EFFECTS  BRANCH
                         OFFICE OF  DRINKING WATER
                         WASHINGTON, D.C.   20460


This draft document has not been released formally by the
Office of Drinking Water, U.S. Environmental Protection
Agency, and  should not at this stage be construed to represent
Agency policy.  It is being circulated for comments on its
technical merit and policy implications.


     The objective of this criteria document is to assess the
available data on health effects associated with exposure to
carbon tetrachloride in drinking water and to recommend a
maximum contaminant level.  To achieve this objectivet data
on pharmacokinetics, human exposure, acute and chronic health
effects in animals and humans, mechanisms of toxicity were
evaluated!/  Only the reports which were considered pertinent
for the derivation of the maximum contaminant level are cited
in the document.  Particular attention .was paid to the utiliza-
tion of primary references for the assessment of carbon tetra—
chloride-induced health effects.  Secondary references were
used rarely.  For comparison purposes, standards and criteria
developed by other organizations are included and discussed
in Section IX (Quantification of Toxicological Effects of
Carbon Tetrachloride).


  I.  SUMMARY                                            1-1


      Analysis of Carbon Tetrachloride  in              II-4
      Drinking Water

      Treatment of Carbon Tetrachloride in             II-5
      Drinking Water

      Summary                                          II-6

III.  PHARMACOKINETICS                                 III-l

      Absorption                                       III-l

      Distribution                                     III-5

      Metabolism                                       III-8

      Excretion                                       111-13

      Summary                                         III-15

IV.  HUMAN EXPOSURE*                                   IV-1

 V.  HEALTH EFFECTS IN ANIMALS                           V-l

     Acute Effects                                       V-l

     Chronic Effects                                   V-17

     Teratogenicity                                    V-21

     Reproductive Effects                              V-25

     Mutagenicity                                      V-28

     Carcinogenicity                                   V-34

     Summary                                           V-43

VI.  HEALTH EFFECTS IN HUMANS                          VI-1

     Case Studies—Acute  Effects                      VI-2

     Acute Effects                                     VI~4

*Prepared by the  Science  and Technology Branch


                     CONTENTS (Contined)

        Case Studies — Long-Term Effects

        Controlled Studies



        Formation of Carbonyl Chloride  (Phosgene)

        Dimerization to Hexachloroethane

        Free Radical Binding to Proteinds

        Lipid Peroxidation



        Quantification of Carcinogenic  Risk

        Sensitive Populations

        Interaction of Carbon Tetrachloride with
        Other Chemicals



       Non-Carcinogenic Effects

       Quantification of Non-Carcinogenic Effects

       Carcinogenic Effects

       Quantification of Carcinogenic Effects

       Special Considerations

  VI- 8

 VI- 13


 VI I- 1

 VI I- 1

 VI I- 2

 VI I- 2

 VI I- 8




VII I- 3



  IX- 1

  IX- 3


  IX- 9


  IX- 15

                          I. SUMMARY
          Carbon tetrachloride, also known as tetrachloro-

methane and perchloromethane, is a haloalkane with a wide

range of industrial and chemical applications.  In 1980,

322,000 kkg were produced in the United States.  Most of the

chemical produced is used in the manufacture of fluorocarbons,

which are used as refrigerants, foam blowing agents, and

solvents.  Carbon tetrachloride is also used as a solvent

in metal cleaning and in the manufacture of paints and plas-

tics as well as in fumigants.  It is being largely replaced

in grain fumigation by other registered pesticide products.

          Inhalation is the most important route of carbon

tetrachloride exposure.  There are no major gradients in the

atmospheric distribution of carbon tetrachloride; the

concentrations are similar in  the continental  and marine air

masses ambient (approximately  O.JD0070-0.00084  mg/m3 or

111-133 ppt).  The maximum level of carbon tetrachloride

reported was 0.113 mg/m3  (18,000 ppt) in Bayonne, New Jersey.

          Carbon tetrachloride has been found  in many sampled

waters  (including  rain, surface^ potable, and  sea)  at the ppb

 level.  The National Organics  Monitoring Survey  (NOMS)  sampled

 113 public water systems  and found carbon tetrachloride at

 very  low concentations, relative to  levels of  chloroform

 and other organics.  Positive  results were noted  in about  10


percent of the samples, with mean values ranging from 2.4-6.4

ppb depending on sampling and analytical procedures.  An

accidental discharge of an estimated 70 tons of carbon

tetrachloride into the Kanawha River resulted in levels of

carbon tetrachloride as high as 300 ppb in raw Ohio river

water; the drinking water levels were found to be as high as

100 ppb.  Chlorination of raw waters with chlorine gas contami-

nated with carbon tetrachloride may account for the presence

of the compound in some finished drinking waters.

          Carbon tetrachloride has been detected in a variety of

foodstuffs other than fish and shellfish in levels ranging from

1-20 ppb.  Carbon tetrachloride contaminations of food categories

has been reported as follows:  Dairy products (0.2-14.0- ppb);

meat  (7.0—9.0 ppb); oils and fats  (0.7-18.0 ppb); beverages

(0.2-6.0 ppb); fruits and vegetables (3.0-8.0 ppb); black grapes

(19.9 ppb); and fresh bread  (5-1-0 ppb).  The amount of carbon

tetrachloride residue in some foodstuffs depends on the

fumigant dosage, storage conditions, length of aeration, and

extent of processing.

          No information was found on levels or frequencies of

human dermal exposures.  Carbon tetrachloride is no longer on

the market as a component of hair  shampoo or cleaning solvents,

and because of .strict OSHA safety  regulations, dermal exposure

in the workplace has been minimal.

          Carbon tetrachloride is absorbed into circulation

through the lungs and the skin.  Absorption from the gas-

trointestinal tract also readily occurs but at a slower

rate.  Following a single exposure, high concentrations

of carbon tetrachloride were detected in the blood, brain,

kidney, and liver.  With repeated exposures, carbon tetra-

chloride is present mostly in the liver and, because of

its lipophilic nature, in the body fat and bone marrow.

          In both animals and humans exposed to carbon tetra-

chloride, the liver is the first organ to be affected.  Minor

liver damage, as evidenced by increased levels of liver enzymes

in the- serum, progresses through early necrotic changes, in-

creased vacuolization and fat droplets, to necrosis, degen-

eration, and fatty liver.  Most acute liver damage is rever-

sible with time.  Chronic necrotic and degenerative changes,

however, appear not to be reversed as readily.  Exposure to

other hepatotoxins especially ethanol and drugs such as

barbiturates, greatly exacerbates the hepatic toxicity of

carbon tetrachloride.  Pulmonary damage occurs, but mostly

at an ultrastructural level and to a much lesser extent

than liver damage.  Renal and pancreatic effects also have

been observed, but only following massive doses of carbon


          In "humans, acute toxic effects of carbon tetrachlo-

ride include gastrointestinal disturbances, nausea,  and

vomiting.  Hepatic injury occurs with all of the signs of

severe cellular damage.  Serious renal injury usually does

not become evident until 24-48 hours or several days after

the onset of initial symptoms of poisoning.  In severe

cases, death from hepatic coma and uremia occurs within

1-2 weeks.  After repeated or prolonged inhalation of less

toxic concentrations of carbon tetrachloride, gastrointesti-

nal and central nervous system symptoms occur such as

nausea, vomiting, visual disturbances, and giddiness.

After repeated exposure, severe hepatorenal damage can

occur.  Treatment and cessation of exposure usually result

in disappearance of toxic symptoms and reversal of the

liver and kidney damage.

          Carbon tetrachloride has been shown to be carcino-

genic in rats, mice, and hamsters.  Although a number of

cases of liver tumors following exposure to carbon tetrachlo-

ride have been reported, the  chemical has  not been established

to be a human carcinogen.  Although it is  not mutagenic in

the Salmonella (Ames) assay,  carbon tetrachloride has

elicited a mutagenic response in the yeast Saccharomyces

cerevisiae.  Carbon tetrachloride has also been shown to

be fetotoxic and to inhibit male fertility in  rats.

          The mechanism of toxicity of carbon tetrachloride has

been extensively studied with special emphasis on the hepatic

effects.  Carbon tetrachloride is metabolized by the liver

mixed function oxidase system to trichloromethyl (and other)

free radicals, which can either react with hepatic macromolecules

or be further metabolized via chloroform and phosgene to carbon

(mono/) dioxide.  The reaction of the trichloromethyl radical

with cellular lipids initiates a lipid peroxidation resulting

in the progressive and exponential destruction of membranes.

Membrane permeability also is greatly increased as a result

of lipid peroxidation.  Although some investigators still

maintain that direct reaction with hepatic macromolecules result

in hepatic  damage, the recent ultrastructural  evidence  tends to

substantiate the lipid peroxidation  theory.

          By extrapolation from animal studies, the National

Academy of  Sciences  (NAS) and EPA's  Carcinogen Assessment

Group  (CAG) have calculated  projected incremental excess

 cancer risk associated with  the consumption  of specific

 chemicals in drinking water  over a  70-year  lifetime.  Using

 the  multistage  model, NAS estimated that  comsumption  of 2

 liters of water per  day  over a  lifetime at  carbon tetrachloride

 concentrations  of  450, 45, or 4.5/liter would result  in the

 induction of  one excess  case of cancer per  10,000,  100,000,

 or 1,000,000  people  exposed, respectively.

          Using the "improved" multistage model, CAG esti-

mated that consumption of 2 liters of water per day at

carbon tetrachoride concentrations 42.2 ug/1, 4.2 ug/1, or

0.4 ug/1 would result in the induction of one excess case

of cancer per 10,000, 100,000, or 1/000,000 people exposed,


          The differences between NAS and CAG risk esti-

mates are partly explained by the fact that the extrapolation

models used by the two groups, although similar, are not

identical.  The NAS risk estimate was arrived at using the

multistage model; whereas, the CAG derived their risk esti-

mate using the "improved" version of the multistage model.

In addition, the models also differ in the data selected and

other parameters.  The NAS model used rats as the comparison

species; the CAG model used mice.

          These modeling methods share the assumption that

there is no threshold level for the action of a carcinogen.

However, no one method can accurately predict the absolute

numbers of excess cancer deaths that will be attributable  to

carbon tetrachloride in drinking water.  None of the methods

presently used to quantify carcinogenic risk accounts  for

increased carcinogenic risk from the interaction of carbon

tetrachloride with other environmental contaminants to

which humans can be exposed.

                             Ir 7
          It is noteworthy that in assessing CCl4~induced
toxicity, carcinogenicity or any other harmful effect,
compounds that react synergistically or antagonistically
with CC14 must be considered.  Identified synergistic
substances include ethanol, kepone, PCB, and PBB.  Antago-
nistic effects have been demonstrated with such compounds
as chloramphenicol and catechol.  Sensitive populations are
subgroups within the general population which appear at
higher than average risk upon exposure to CC14.  Some of
the populations that may be at greater risk include human
fetusesr alcohol consumers, and males of reproductive age.

          The quantification of non-carcinogenic effects in
humans could not be undertaken at this juncture due to lack
of acceptable chronic exposure data for this compound.  Be-
cause of positive results  in animal carcinogenicity studies,
carbon tetrachloride can be considered a suspect human
carcinogen.  Thus,  the recommended maximum contaminant level
(RMCL) for carbon tetrachloride should be based on  its
carcinogenic potential.  According to the Safe Drinking Water
Act (42  USC 300F, SDWA, 1974), this level should be zero
(0 mg/L).


          Carbon tetrachloride, or tetrachloromethane, is a

colorless liquid with a molecular weight of 154 and a boiling

point of 76.5"C (Weast, 1972).  It is a relatively nonpolar

compound that is miscible in alcohol, acetone, and other

organic solvents (Weast, 1972), but is only minimally soluble

in water (0.8 g/liter at 25°C)  (Johns, 1976).  The octanol/

water partition coefficient of  carbon tetrachloride is 2.64

(Johns, 1976).

          The properties of carbon tetrachloride favor volati-

lization of the compound from water to air.  Carbon tetrachlo-

ride has a high vapor pressure  (115.2 mm Hg at 25 *C)  (Johns,

1976).  The air/water partition coefficient of carbon tetra-

chloride at 20°C is 1.1 by volume, and about 1,000 by weigh t.

(Johns, 1976).  The rapid vaporization predicted from these

properties has been confirmed by Billing et al. (1975), who

reported a haIf-life of carbon  tetrachloride evaporation of

29 minutes from a  dilute aqueous solution  at about 25 *C.

          The density of carbon tetrachloride is 1.59 g/ml at

20°C  (Weast,  1972).  Because  its density is greater than the

density of water,  some  carbon  tetrachloride  from large  spills

in water might tend to  settle  before  it  is totally dispersed,

emulsified or volatilized.

          Carbon tetrachloride is produced industrially by the

chlorination of methane, propane, ethane, propylene, or carbon

disulfide (Rams et al., 1979).  In 1980, 322,000 kkg were syn-

thesized (USITC, 1981).  Carbon tetrachloride is also produced

indirectly during the production of compounds such as vinyl

chloride and'perchloroethylene (Rams et a_l., 1979).

          TJhe major use of carbon tetrachloride is in the pro-

duction of chlorofluorocarbons, which are used as refriger-

ants, foam-blowing agents, and solvents.  Carbon tetrachlo-

ride is also used in fumigants, and has a variety of minor

uses, including those as a solvent in metal cleaning and in

manufacture of paints and plastics (Rams et_ a_l., 1979).  It is

being replaced in grain fumigation by other registered pesti-

cide products (USEPA, 1980a), and its registration for use in

fumigants is presently under  review by USEPA (USEPA, 1980b).

          Carbon tetrachloride present in the environment

appears to be of anthropogenic origin (Singh et al., 1976).

It can enter natural waters through industrial and agricul-

tural activities.  Carbon tetrachloride may be carried to

surface waters through run-off from agricultural, industrial,

and dumping sites, and through industrial effluents.  Indus-

trial emissions may also contribute carbon tetrachloride

to the air, from which the compound may reach surface water

through rainfall.  Carbon tetrachloride may also reach

groundwater through leaching from solid waste sites.

          Once in the environment, carbon tetrachloride is

relatively stable.  Its half-life for hydrolytic breakdown

in water at pH 1.0-7.0 is estimated to be 70,000 years,

but hydrolysis appears to be accelerated in the presence

of metals such as iron and zinc  (Johns, 1976).  The high

.stability in water has little practical significance; how-

ever, since carbon tetrachloride vaporizes readily to air.

The atmospheric  lifetime of carbon tetrachloride appears

to be on the order of 30-100 years (Singh et al., 1976).

          The presence of carbon tetrachloride in the en-

vironment is of  concern  for two  reasons.  First, carbon

tetrachloride may contribute to  ozone-destroying photochemi-

cal reactions in the stratosphere, which might cause in-

creases in the incidence in human  skin cancers and  animal

cancers, affect  terrestrial and  aquatic ecosystems,  and

bring about climatic changes  (NAS, 1978).

          Although  levels of carbon  tetrachloride  in the  envi-

 ronment are generally  in the  low ppb range  or below (NAS,

 1979),  this chemical may pose  a  long-term danger because  of

 its possible  carcinogenic potential.   In urban and industrial

areas where higher concentrations of carbon tetrachloride

occur,  other toxic effects may result (e.g., liver and renal

damage).  The following chapters discuss the evidence for

health effects attributed to carbon tetrachloride, with

particular emphasis on ingestion via drinking water.

Analysis of Carbon Tetrachloride in Printing Water

          Carbon tetrachloride (and 47 other halogenated organ-

ics) in water can be analyzed by a purge and trap method

{Method 502»1) described by the EPA Environmental Monitoring

and Support Laboratory  (USEPA, 1980d). 'This method can be

used to measure purgeable organics at low concentrations.

Purgeable organic compounds are trapped on  a Tenax GC-contain-

ing trap at 22*C using  a purge gas rate of  40 ml/min  for 11

minutes.  The trapped material is then heated rapidly to

ISO'C  and backflushed with helium at a flow rate  of 20-60

ml/min for 4 minutes into the gas chromatographic analytical

column.  The programmable gas chromatograph used  is capable

of operating at 40"+1*C.  The primary analytical  column  is

 stainless steel packed  with 1% SP-1000 on Carbopack B (60/80)

mesh  (8  ft x 01 in. I.D.) and is  run at a. flow  rate of 40

ml/min.  The temperature program  sequence begins  at 45 °C for

 3 tninutes, increases B*C/min to 22Q°C, and  is then held

 constant for 15"minutes or until  all compounds  have eluted.

A halogen-specific detector with a sensitivity to 0.10 ug/

liter a relative standard deviation of 10% must be used.

The optional use of GC/MS techniques of comparable accuracy

and precision is acceptable.

Treatment of Carbon Tetrachloride in Drinking Water

          The information available on the removal of carbon

tetrachloride from drinking water is limited.  However, as

judged from data obtained for industrial waste treatment,

conventional treatment processes are not very effective in

the removal of this compound.  An isotherm study of carbon

tetrachloride on Filtration 400 activated carbon (GAG) showed

that at an equilibrium concentration range of 3 x 10~9 to 2.6

x 10~"7 moi/liter-, a maximum surface concentration of 2.6 x

10~5 mol/g was obtained  {NAS, 1979).  Aeration and adsorption

processes have also been evaluated  for removal of this

compound.  Powdered activated carbon  (PAC) at 2 to 4 mg/liter

was not effective in  treating contaminated river water  contain-

 ing 16.3 mg/liter of  carbon tetrachloride.   After PAC,  coag-

 ulation, settling, and filtration,  the finished water  still

 contained 16.0 mg/liter  (USEPA,  1980d).  Aeration by diffused

 air aerator  in a  laboratory study was  found  to be more suc-

 cessful.  At 4:.l  air-to-water  ratio,  a 91  percent  removal

 efficiency for carbon tetrachloride was  achieved  (USEPA,

1980d).  Adsorption by GAG in a pilot scale study revealed

that carbon tetrachloride at an average concentration of 12

rag/liter (Cincinnati tap water) was reduced to less than 0.1

ug/liter for 3 weeks with a 5-minute empty bed contact-time

(EBCT) and for 14-16 weeks with a 10-minute EBCT.


          Carbon tetrachloride is a colorless liquid at am-

bient temperature.  Its high vapor pressure favors rapid

volatilization from water to air.  Carbon tetrachloride is

produced commercially from the chlorination of methane, pro-

pane, ethane propylene or carbon disulfide and its major use

is in the production of chloroflurocarbons.  Carbon tetra-

chloride is present in the enviroment by anthropogenic means

and once in the environment appears relatively stable.  Its

presence is of concern because of a possible contribution to

ozone  destroying  chemical reaction in the atmosphere and

because ingestion via drinking water may present a human health

hazard.  Carbon tetrachloride in water  may be detected by

the purge and trap method described by  the USEPA Enviro-

nmental Monitoring and Support Laboratory.  Removal of  this

 chemical from water may proceed by aeration and  filtration

 through appropriate media.

                     III.  PHARMACOKINETICS
  Coef ficients.  Partition coefficients for

 various chlorinated solvents, including carbon tetrachloride,

 were determined by several experimenters  {Morgan et al. , 1972;

 Sato and Nakajima, 1979; Powell, 1945).  The partition  coeffi-

 cient is a measure of the relative solubility of a substance in

 two media.  The oil/air and oil/water partition coefficients

 can be used as indicators of solubility in  lipids.  The values

 of these and other partition coefficients for carbon tetrachlo-

 ride, listed in Table III-l, show this chemical to be  lipophi-

 lic.  Because of  its lipophilic  nature, one would predict that

 carbon tetrachloride, could be absorbed by  ingestion,  inhala-

 tion, and skin contact.  This prediction  is borne out  by

 results of the experimental  studies  described below.

 TABLE III-l  Partition Coefficients  for Carbon Tetrachloride
     Olive  oil/air
     Olive  oil/air

     Blood  serum/air


     Olive  oil/water

     Olive  oil/serum
     Olive  oil/blood


Adapted from Morgan et al.  (1972),  Sato and Nakajima (1979), and
Powell (1945).

          Absorp-tion from the Gas~troint.est.inal Tract..  Absorp-

tion of carbon tetrachloride through the gastrointestinal

tract of dogs was studied by Robbins (1929).  In a series of

experiments, the author determined the amount of carbon tetra-

chloride absorbed from the ligated stomach, small intestine,

and colon by measuring the concentration of carbon tetrachlo-

ride found in the exhaled breath.  The greatest concentration

of carbon tetrachloride in exhaled air was seen after  injec-

tion of the chemical into the small intestine.  Direct injec-

tion of carbon tetrachloride into the colon resulted in a

lower concentration of the chemical in exhaled air.  After

introduction directly into the  stomach by  intubation,  no

carbon tetrachloride was detected in exhaled  air.  The method

of detection in these experiments was thermal conductivity,

with stated detection limits of one part in ten.  Thus, the

results of the experiment can be viewed as a  qualitative  indi-

cation of relative absorption from the various components  of

the  gastrointestinal tract, rather than as quantitatively

accurate results.

          Absorption by Inhalation.  Von Oettingen  et  al.

 (1950)  studied the absorption of carbon tetrachloride  by

 inhalation  in Beagle dogs.  The sex was unspecified, but  the

 authors  stated that at  least  five dogs were used in each

 experiment.  The  dogs  inhaled  carbon  tetrachloride  (purity

                            II I-3
unspecified) at a concentration of 94,500 mg/m3 (15,000
ppm) for 475 minutes through a two-way valve attached to the
cannulated trachea.  Blood samples were taken at unspecified
intervals and analyzed for carbon tetrachloride.  Data pre-
sented graphically showed that the concentration of carbon
tetrachloride in blood reached a maximum of 31.2-34.3 mg/100
cc  (0.20-0*22 raillimole percent) after approximately 300
minutes of exposure and remained at  that level  for the dura-
tion of the exposure.
          McCollister ejt a_l. (1951)  investigated the absorp-
tion of carbon  tetrachloride by  inhaltion using rhesus mon-
keys.  Three female monkeys inhaled  99.9% 14Olabeled
carbon tetrachloride at an average concentration of 290
rag/m3  (46 ppm)  for 139, 344, and 300 minutes,  respectively.
The authors calculated by difference between  inhaled and
exhaled air that  the monkeys absorbed  an average of 30.4%  of
the total amount  of carbon tetrachloride inhaled.  Analysis
of blood drawn  after 270 minutes of  exposure  showed that  the
14C radioactivity was equal to  0.297 mg of  carbon  tetrachlo-
ride/ 100 g of  blood, distributed  as follows:  56.2% as  carbon
tetrachloride,  16.5% as  "acid-volatile" carbonates, and 27.3%
as nonvolatile  material.  No attempt was made to  characterize
metabolites  in  this  study.

          Absorption Through the Skin.  McCollister et al. (1951)

exposed the skins of one male and one female rhesus monkey

to l^c—labeled carbon tetrachloride vapor.  To determine

the amount of absorption, blood and exhaled air were analyzed

for l^C radioactivity.  After a skin exposure of 240 minutes

to carbon tetrachloride vapor at 3,056 mg/m3 (485 ppm), the

blood of the female monkey contained carbon tetrachloride at

0.012 rag/lOOg and the exhaled air contained 0.0008 mg/liter.

After exposure to 7,345 mg/m3 (1,150 ppm) for 270 minutes,

blood of the male monkey contained carbon tetrachloride at

0.03 mg/lOOg and the exhaled air contained 0.003 mg/liter.

          Three human volunteers, sex unspecified, immersed

their thumbs in carbon tetrachloride  for 30 minutes in an

experiment to measure skin absorption (Stewart and Dodd, 1964).

Carbon tetrachloride was analyzed by  infrared spectroscopy

and was found to contain no detectable impurities-  The

concentration of carbon tetrachloride in alveolar air was

used as the indicator of absorption and measured at 10, 20,

and 30 minutes after  the start  of exposure  and at 10,  30,

60, 120,  and 300 minutes after  cessation of exposure.  Carbon

tetrachloride was present in the alveolar air at each  time

interval,  reached a maximum concentration range of 2.8-5.7

mg/m3  (0.45-0.79 ppra) 30 minutes after exposure, and decreased

                              II I-5
 exponentially  thereafter.   The  authors  concluded that  carbon
 tetrachloride  could  be  absorbed by  the  skin  in toxic amounts
 if  the  chemical  came in contact with  arms  and  hands.


            Robbins  (1929) administered 159  g  (100 cc) of  carbon
 tetrachloride, purity unspecified,  to three  anesthetized dogs by
 stomach tube.  The dogs were  sacrificed at 6,  23, and  24 hours
 after treatment  and  blood  and various tissues  were analyzed for
 carbon  tetrachloride by converting  the  organic chloride  to
  inorganic  chloride and  titrating the  inorganic chloride  by the
 Volhard method,  which is accurate to  0.1-0.2%.  The results of
  the blood  and  tissue analysis are shown in Table IIT-2.

TABLE III-2   Carbon Tetrachloride Distribution at Various  Times
      After  Administration by Stomach Tube (rag/100 g of  tissues)

Blood, portal
Blood f arterial
Bone marrow
6 hrs
— _
23 hrs
24 hrs
— —

          From the experimental data, it appears that the

limit of detection was in the range of 4-5 mg of carbon tetra-

chloride/ 100 g of tissues.  In addition, it appears that the

liver, bone marrow, blood, and muscle retained the most carbon

tetrachloride for the longest time.

          Von Oettingen e_t al. (1950) reported the tissue dis-

tribution of carbon tetrachloride in Beagle dogs, each weighing

about 10 kg, exposed to carbon tetrachloride in air at 94,500

mg/m3 (15,000 ppm) for 475 minutes.  The dogs were sacrificed

at the end of the exposure.  Tissue and blood samples were

taken and analyzed for carbon tetrachloride.  The concentration

of carbon tetrachoride (per  100 g of tissue) was 65 mg/100 g

in the brain, 50 mg/100 g  in the heart, 36 mg/100 g in the

liver, and 34 mg/100 g in  the blood; the concentratiion  in

the  fat was not determined.  The investigators stated  that

the  accumulation of carbon tetrachloride in  the brain  was

consistent with its high oil/water partition coefficient and

resulted in its strong narcotic action.

          McCollister e_t a_l. (1951)  reported the  tissue  distri-

bution  of carbon  tetrachloride  in  rhesus monkeys  exposed to

290  mg/m3  (46 ppm) of  [14C]  carbon  tetrachloride  for  300

minutes.  The tissue distribution,  as  calculated  from the

14C  radioactivty,  is  shown in Table  III-3.   The  concentration

of carbon tetrachloride was greatest in the fat, followed by

the liver and bone marrow.

TABLE III-3  Tissues Distribution of [14C] Carbon Tetrachloride
                   Inhaled by Rhesus Monkeys

Bone marrow
Carbon tetrachloride
(mg/100 g of tissue)

 Souce:  McCollister et al.  (1951)

           Fowler  (1969) studied  the distribution  of  carbon

 tetrachoride In the tissues of rabbits given  the  chemical

 by stomach tube.   Five rabbits were given  carbon  tetrachloride


(1 ml/kg bw) as a 20% (v/v) solution in olive oil.  Analysis

of the carbon tetrachloride by gas chromatography showed

not more than 125 ppb of hexachloroethane.  The rabbits were

sacrificed 6, 24, and 48 hours after treatment and the tissues

analyzed for carbon tetrachloride by gas chromatography

equipped with an electron capture detector.  Six hours after

carbon tetrachloride was administered, the tissue concentrations

(per kg of tissue) were 787 +_ 289 (Mean ± SEM) mg/kg in fat,

96 +_ 11 mg/kg in liver, 21 +_ 12 mg/kg im muscle, and 20 +_ 13

mg/kg in kidney.  By 48 hours, these concentrations had

dropped to 45 +_ 12 mg/kg in fat, 4 + 0.1 mg/kg in liver, and

0.5 ^0.3 mg/kg in kidney and muscles.  These data indicate

that most of the dose of carbon tetrachloride is eliminated

from rabbit tissues within 48 hours.


          Chloroform was one of the first carbon tetrachloride

metabolites to be described (Butler, 1961).  Eight dogs were

exposed to carbon tetrachloride by tracheal cannula at the rate

of 8,000 mg/hr for 3 hours.  At the cessation of exposure, the

exhaled air from the dogs was collected and analyzed by both

gas  chromatography and the Fujiwara reaction, a colorimetric

procedure for- the identification of chloroform.  Chloroform

was  detected in the exhaled breath by both of these methods.

The  total amount of chloroform exhaled in 2 hours by each dog

was estimated at 0.10.5 mg by analysis of gas chromatographic

data.  Tissue homogenates were also shown to metabolize

carbon tetrachloride to chloroform.

          Evidence of metabolism to a free radical was sug-

gested by studies showing hexachloroethane to be a carbon

tetrachloride metabolite  (Bini et al., 1975).  Five Wistar

rats were administered 160-800 mg of  carbon tetrachloride

diluted in liquid paraffin by gavage  following a 24-hour

fast.  The animals were sacrificed 15 minutes to 8 hours

after treatment.  A graph displaying  carbon tetrachloride

concentrations in rat liver  versus time  showed the chemical

at approximately of tissue  after 15 minutes and at

maximal concentration (1.7 mg/kg) after  120 minutes.  Analysis

of the gas chromatographic data showed that chloroform, was

maximal at 0.037 mg/kg after 15.minutes;  after 4 hours it

had declined to 0.007 mg/kg.  Hexachloroethane was also

 present after 4 hours 0.005  mg/kg.  The  authors explained

 the formation of both chloroform  and  hexachloroethane as

 carbon tetrachloride metabolites  by proposing  that the tri-

 chloromethyl  free  radical was the primary metabolite of

 carbon tetrachloride.

           14C-labeled carbon dioxide  was detected in the

 exhaled  air of  rhesus monkeys after a 344-minute exposure to


E14C] carbon tetrachloride by inhalation (McCollister at

al. ,  1951).-  The amount of C14C] carbon dioxide exhaled

during the 7-day period following exposure was reported to

be 10-20% of the total radioactivity expired.   The authors

integrated the resulting equation from 18 to 1,800 hours (75

days) and estimated that 4.4 mg or 11% of the total amount of

radioactivity eliminated was excreted as carbon dioxide.

          Shah et al. (1979) studied the metabolism of [14C]

carbon tetrachloride by rat liver in vitro^  Samples of liver

homogenate equivalent to 0.167 g of tissue were incubated

for 30 minutes at 37.5*C with 10 umole of 14C-labeled carbon

tetraichloride alone, and with either NADH or NADPH or both.

[i4C3 carbon dioxide was detected by scintillation counting.

The results are shown in Table 1II-4.  The addition of NADPH

appeared to result in substantial conversion of carbon tetra-

chloride to carbon dioxide.  Addition of NADH and NADPH did

not increase the conversion over that seen with NADPH alone.

          Shah et al. (1979) tested for the possible formation

of carbonyl chloride in hepatic carbon tetachloride metabolism

by adding L-cysteine to the in vitro rat liver system described

above.  Carbonyl chloride and L-cysteine are known to react

chemically to form a condensation product, 2-oxothiazolidine-

4-carboxylic acid.  The presence of the condensation product

                            I IT- 11
was confirmed by thin-layer chroma tography and mass spectro-

metry.  The author inferred from the presence of 2-oxothiazoli-

dine-4-carboxylic acid that carbonyl chloride was  formed  in

the metabolism of carbon tetrachloride by rat liver microsomes,

The authors postulated a mechanism of biotransformation for

carbon tetrachloride which involves a sequential oxidation of

carbon tetrachloride while bound to a heme  (see Figure III-l).

Release of bound intermediates  then gives rise to  different

unrelated metabolites at the  site of release.

TABLE III-4    Conversion of  [14C] Carbon Tetrachloride to
                     Carbon Dioxide by Rat  Liver Homogenate
  Nucleotide  added   [C}C02 (mole/g  liver,  mean +_ SEM)



                                        27  + 5

                                       373  + 17

                                       464  + 33

                                       472  + 21
 Source:  Shah e_t al^.  (1979)

           Fowler (1969)  detected hexachloroethane and chloroform

 in the tissues of rabbits orally administered carbon tetrachlo-

 ride.  A total of 15 rabbits were given carbon tetrachloride

                                t ft
                                  •       CT ^

                                  -CSj-Oj*] —

     Figure III-l  Pathways of Carbon Tetrachloride Metab-
     olism.  Products identified as carbon  tetrachloride
     metabolites are underlined.  The electrons  utilized  in
     the reactions are assumed to come  from NADH or NADPH via
     the flavoprotein cytochrome reductases.  Fe^4" and Fe^"*"
     denote the respective ferro— and f erricytochromes .
     Redrawn from Shah et al. .(1979).

at 1 ml/kg bw and sacrificed in groups  of five at 6, 24,  and

48 hours after exposure.  Samples of fat, liver, kidney,  and

muscle tissues were analyzed for chloroform and  hexachloro-

ethane by gas chroma tography.  The results  of the analysis are

in Table III-5.  The fat contained the  highest amounts of hexa

chloroethane at each sampling time, but the highest concentra-

tions of chloroform appeared in the liver.
          McCollister et al. (1951)  studied  the  elimination of

            carbon tetrachloride  from  rhesus monkeys  exposed by

TABLE ITI-5  Chloroform and Hexachloroethane in Tissues of Rats
                 Given Carbon Tetrachloride Orally

Sample time
6 hours

24 hours

48 hours

(ug/g tissues)
4.7 + 0.5
4.9 + 1.5
1.4 + 0.6
0.1 + 0.1
1.0 + 0.2
1.0 + 0.4
0.4 + 0.2
0»1 + 0.1
0.4 + 0.1
0.8 + 0.2
0.2 -I- 0
0.1 4- 0.1
C13 CCC13
(ug/g tissues)
4.1 + 1.2
1.6 -t- 0.5
0.7 -t- 0.2
0.3 Hr 0.2
16.5 + 1.5
4.2 + 1.8
2.2 + 1.1
0.5 + 0.2
6.8 4- 2.4
1.0 + 0.3
   Sources  Fowler  (1969)

   inhalation for 344 minutes.  The  total  14C  radioactivity  in the

   blood decreased  12% during the  first  10 minutes  after exposure.

   Graphs of data from the analysis  of blood samples  obtained

   periodically  for 10-12 days  following exposure  showed that the

   level of carbon  tetrachloride  in  the  blood  decreased exponen-

   tially with time.  At 10  days,  the level of carbon tetrachlo-

   ride in blood was approximately 0.009 mg/100 g.   The authors

   estimated that 21% of the total amount  of  carbon tetrachloride

   absorbed was  eliminated  in  expired air  during the first 18


days.  By extrapolation of these data, the authors concluded
that over 1,800 hours (75 days) approximately 51% of the carbon
tetrachloride initially absorbed would be eliminated in exhaled
breath either as carbon tetrachloride or carbon dioxide.  Analy-
sis of urine and feces showed measurable amounts of radioacti-
vity after 15 and 12 days, respectively.  The authors inter-
preted these findings as indicating that significant quantities
of carbon tetrachloride and/or metabolites may be excreted by
these routes (breath, urine, and feces).


        Carbon tetachloride is readily absorbed from the lungs
and the gastrointestinal tract, as expected from  its partition
coefficients.  Although few quantitative data are available on
the amount of carbon tetrachloride absorbed through the lungs,
the chemical and its metabolites have been reported in blood,
many tissues, exhaled air, urine, and feces after administration
by  inhalation.  Carbon tetrachloride has also been absorbed
through the skin, but the  reported rate of absorption was
much slower than that of  inhalation.

          In published studies, carbon  tetrachloride has  appeared
to  be distributed to all major organs following  absorption.   The
highest concentrations have been  found  in  the  liver,  fat,  blood,
brain, kidney, spleen, and pancreas.


          Carbon tetrachloride metabolism has been reported to

occur primarily in the liver.  Carbon tetrachloride has been

postulated to be metabolized to a trichlororaethyl radical

bound to an iron atom in the cytochrome heme moiety.  This

trichloromethyl radical was reported to be either further

metabolized or released as a free radical.  The trichloro-

methyl free radical was reported to undergo a variety of

reactions; including hydrogen abstraction to form chloroform,

and dimerization to form hexachloroethane.  Further metabolism

of the heme-bound trichloromethyl radical was postulated to

result in the eventual formation of carbonyl chloride (phos-

gene) .

          Carbon tetrachloride and its metabolites have been

reported in many studies to be excreted primarily in exhaled

air,  and also in the urine and-feces.

                              IV.   HUMAN  EXPOSURE

     Humans may be  exposed to carbon  tetrachloride  in drinking  water,  food,
and air.   Detailed  information concerning  the occurrence of  and exposure to
carbon  tetrachloride  in  the  environment  is  presented  in  another  document
entitled "Occurrence of Carbon Tetrachloride in Drinking Water, Food, and Air"
(Letkiewicz et al.  1983).   This  section  summarizes the  pertinent information
presented in that document in order to assess the relative source contribution
from drinking water, food, and air.

Exposure Estimation
     This analysis  is  limited to  drinking  water,  food,  and  air, since these
media are  considered  to be  general  sources common to all  individuals.   Some
individuals may be exposed to carbon tetrachloride from sources other than the
three considered  here,  notably in  occupational  settings and  from  the  use of
consumer  products  containing - carbon  tetrachloride.    Even  in  limiting  the
analysis to  these three sources,  it must be  recognized  that individual expo-
sure will vary widely based  on many personal  choices  and several factors over
which there  is little control.  Where  one lives, works, and travels, what one
eats, and  physiologic  characteristics  related to  age.,  sex,  and health status
can  all  profoundly  affect daily  exposure and intake.   Individuals living in
the  same neighborhood  or even  in the  same  household  can  experience vastly
different exposure patterns.
     Unfortunately,  data  and methods  to  estimate  exposure  of identifiable
population  subgroups  from  all   sources  simultaneously  have  not yet  been
developed.   To the extent  possible,  estimates are provided  of the number of
individuals  exposed to  each  medium at  various carbon  tetrachloride  concentra-
tions.   The  70-kg  adult male is  used for estimating dose,  which  takes into
account  the amount of the medium contacted  (i.e.,  water and food  ingested; air
breathed)  and the amount  of carbon tetrachloride actually  absorbed into the

a.   Water
      Cumulative  estimates of  the U.S.  populations exposed  to various  carbon
tetrachloride levels  in drinking  water from public drinking water  systems are

presented in Table IV-I.   The  values  in the table were obtained using Federal
Reporting Data Systems data (FRDS 1983) on populations served by primary water
supply systems and the  estimated  number of these  water systems that contain a
given  level  of  carbon  tetrachloride.    An  estimated  26,810,000  individuals
(12.5%  of the  population  of  214,419,000  using  public  water  supplies)  are
exposed  to  levels  of carbon tetrachloride in drinking water  at or  above 0.5
ug/1 , while  2,087,000 individuals (1.0%) are exposed  to  levels above 5 ug/1.
It is estimated that 655,000 individuals are exposed to levels greater than 20
ug/1; none are estimated to be exposed  to levels exceeding 30 ug/1.

       Table IV-I.  Total  Estimated Cumulative Population  (in Thousands)
               Exposed to  Carbon  Tetrachloride in  Drinking Water
                     Exceeding  the Indicated Concentration
                  Number of       Cumulative population  (thousands) exposed
                people served   	to concentrations  (ug/1) of
in U.S.
System type (thousands)
Surface water
(% of total )
_>_ 0.5
      No data  were  obtained  on  regional  variations  in  the concentration  of
 carbon  tetrachloride  in  drinking  water.    The  highest  concentrations  are
 expected to occur near sites of  production and use  of  carbon tetrachloride and
 also, in the case of groundwater,  near waste disposal  sites.
      Little information  was available on gastrointestinal absorption rates for
 carbon  tetrachloride.   Because  of  its  lipophilic  nature, ingested  carbon
 tetrachloride is expected  to be readily absorbed.   In  one  study,  rats orally
 exposed to carbon  tetrachloride  were  found  to  excrete at  least   80%  of the
 administered dose within 10  hours  via the lungs,  indicating that at least 80%
 of the dose was absorbed (Marchland et  al.  1970 cited in USEPA 1982).  Another
 study  gave  an absorption factor of 50% for ingestion, but the report has been
 criticized  for not including substantiating information  or citing the specific

literature (Stokinger and Woodward  1958 cited in  USEPA 1982).  A conservative
estimate for the rate of gastrointestinal absorption based on the limited data
available is 100% (USEPA 1982).
     Daily  intake levels  of  carbon  tetrachloride  from  drinking  water were
estimated using various exposure levels and the assumptions presented in Table
IV-II.   The  data  in the table  suggest  that  the majority  of the persons using
public drinking water  supplies would be exposed  to  intake levels below 0.014

     Table IV-II.   Estimated Drinking Water Intake of Carbon Tetrachloride
Persons using supplies
exposed to indicated levels
Exposure level
% of Total
Intake (ug/kg/day)
70-kg man,  2 liters  of water/day,  gastrointestinal  absorption
rate of 100% (USEPA 1982).
      An  indication  of the overall  exposure  of the total population  to  carbon
 tetrachloride can be  obtained  through  the calculation  of population-concentra-
 tion  values.   These values are a summation  of  the  individual  levels  of  carbon
 tetrachloride  to  which each member of the population  is exposed.   An explana-
 tion  of the derivation  of these values  is  presented  in Appendix  C.   Popula-
 tion-concentration  estimates  for  carbon  tetrachloride  in drinking water  were
 3.3 x  107  ug/1  x  persons (best case),  9.7 x 107 ug/1  x  persons (mean  best
 case),  1.1  x  108  ug/1  x  persons (mean  worst case),  and  2.5  x 108  ug/1  x
 persons  (worst case).
      Assuming  a  consumption rate  of  2 liters  of water/day  and a gastrointes-
 tinal  absorption rate  of  100%, population-dose  values  of  6.6 x  107 ug/day x
 persons (best case),  1.9  x 108 ug/day x persons  (mean best  case),  2.2 x 10
 ug/day x  persons   (mean worst case),- and  5.0 x  108  ug/day  x  persons  (worst
 case) were derived.

b.  Diet
     The  dietary  intake  of  carbon  tetrachloride  in  the  United  States  was
estimated using data on concentrations  of carbon  tetrachloride in food compo-
sites  in  the  TEAM  study  (Pellizzari  et  al.  1982) and  data  on  the average
intake  of food by  food  class   (FDA  1980) (Table  IV-III).    Since  levels  of
carbon  tetrachloride  in  the composites  for  the  TEAM  survey  were  below the
quantitation  limit, daily   intakes  were  calculated by  assuming  that  these
levels  were equal  to  zero  (minimum estimate) or to the  quantitation  limit
{maximum  estimate).    Dietary intakes  of carbon  tetrachloride for  the food
classes  studied,  estimated  by  this  method,  varied  between  0-1.27 ug/day.

     Table IV-III.  Estimated Adult Dietary Intake of Carbon Tetrachloride
                     by Food Class3 Using TEAM Survey Data
                  Average intake
                  of food class
  Average level  of
carbon tetrachloride
  Average intake of
carbon tetrachloride
Food class
I- Dairy
II. Meat, fish,
and poultry
X. Oils and fats
XII. Beverages
0.1 28e

Minimum0 Maximum
0 1.0
0 0.9
._0 3.0
Q 0.5

 aEight food  classes  not analyzed:   grains and cereals  (III),  potatoes (IV),
  leafy legume and  root vegetables  (V,  VI, VII), garden  fruits  (VIII), fruits
  (IX), and sugars and adjuncts (XI).
 bFrom FDA 1980.
 CA11 nonquantifiable values assumed to be  equal to zero.
 dA11  nonquantifiable  values  assumed  to  be equal  to  the  quantitation limit
  (value  reported  is the  average of  those composites with  known quantitation
 Calculated  by  subtracting  14-day  drinking  water  consumption  from  14-day
  beverage consumption  (FDA  1980) and  dividing  by  14.
 Several problems arose in the use of the TEAM data:
      1)   The data  are  limited, since only  five  samples  from each food group
           were analyzed,  and they may not  be  representative of normal carbon
           tetrachloride levels  in foods.

     2)    Only  four  of twelve food classes, those suspected of containing the
          highest levels  of  volatile  organics, were analyzed.
     3)    Composite  samples  generally contained lower levels of carbon tetra-
          chloride   than  expected  from  levels  in  the  spiked   subcomposite
          samples  from  which they  were  obtained  (i.e.,  some  carbon tetra-
          chloride appeared  to be  lost  during compositing).
     4)    The grains and cereals  class,  which  may contain significant carbon
          tetrachloride  as a result of  grain  fumigation,  was not analyzed.
     Because of these  data  limitations,  the dietary intake values  presented
above are considered to  be approximations.
     Gastrointestinal  absorption of  carbon tetrachloride was discussed in the
previous section.   The  estimated  absorption  rate was  100%.   If the  average
adult male  weighs  70 kg and  has  a  daily  intake  of  1.27 ug of carbon tetra-
chloride  (the  maximum  estimated  in the  TEAM  study),  the  estimated  adult
dietary  intake  is  0.018 ug/kg/day.
     It  is expected that dietary levels  of carbon  tetrachloride  vary somewhat
with geographical  location,  with  higher  levels occurring in foods  from  areas
near  sources of carbon  tetrachloride  exposure.    However, no   estimates  of
variations in  intake  by  geographical region  could be made  from  the available
data.  Variances in individual  exposure  due  to differences in  diet also  could
not be assessed.

c.  Air
     Exposure to carbon  tetrachloride  in the atmosphere varies  from one  loca-
tion  to  another.    The  highest  level of carbon tetrachloride  reported in the
atmosphere was  69,000 ng/m3 (69 ug/m3)  (Battelle  1977 cited in Brodzinsky and
Singh 1982).   High  levels, averaging greater  than  10,000 ng/m3 (10 ug/m3),
have  been  detected in  other   areas.    Normal   levels,  however,  are somewhat
lower.   Brodzinsky and  Singh  (1982)  calculated  median air levels  of carbon
tetrachloride  for  rural/remote  areas, urban/suburban  areas,  and source  domi-
nated areas of  820 ng/m3  (0.82  ug/m3),  1,200  ng/m3  (1.2 ug/m3),  and  3,700
ng/m3 (3.7  ug/m3),  respectively.
     The  monitoring data available  are not sufficient  to determine  regional
variations  in  exposure  levels  for  carbon tetrachloride.   However, urban and
industrial  areas appear  to  contain higher  levels,  as expected.

     Pulmonary absorption rates for carbon tetrachloride have been reported in
several  studies.   In one,  rhesus monkeys  inhaled  ^C-labeled carbon  tetra-
chloride vapor  at  an average  concentration  of 290 mg/m  ,  and  absorption  was
measured as the difference between the  concentrations  of  carbon tetrachloride
in inhaled and exhaled air.   An  average absorption  rate of 30.4% was obtained
(McCollister et al.  1951 cited in  USEPA 1982).  In  another study, the absorp-
tion of carbon tetrachloride  by  humans  was  studied  by  the difference in quan-
tity of carbon tetrachloride in inhaled and exhaled  air.  The reported absorp-
tion range was  57-65%  (Lehmann and Schmidt-Kehl  1936  cited in USEPA 1982).  A
further study reported an  absorption factor for inhalation  of  30% (Stokinger
and  Woodward  1958 cited in  USEPA 1982).   The last study,  however,  has been
criticized  for not  including  substantiating  information  or  citing  litera-
ture.   From  these  data,  a  pulmonary  absorption  rate of 40%  was  estimated
(USEPA 1982).
     The daily  respiratory  intake of carbon tetrachloride from air was esti-
mated using the assumptions  presented in Table IV-IV and the median and maxi-
mum  levels  for carbon tetrachloride reported  above.    The estimates in Table
IV-IV indicate  that  the  daily carbon tetrachloride  intake from air for  adults
in  source  dominated areas  is approximately 0.5 ug/kg/day.   In  contrast,  the
intake calculated using the maximum  carbon tetrachloride  level  reported  is  9.1
ug/kg/day;  few  if any persons are believed to be  exposed at that level.   The
values presented do  not account  for  variances  in individual  exposure  or  uncer-
tainties in the assumptions  used to  estimate exposure.

      Table  IV-IV.   Estimated Respiratory Intake of Carbon Tetrachloride

      "Exposure  (ug/m3)                       	Intake (ug/kg/day)	
     Rural/remote  (0.82)                                     0.11
     Urban/suburban  (1.2)                                    °-16
     Source  dominated (3.7)                                  O*49
     Maximum (69)                      	9**	
 Assumptions-   70-kg  man,  23  m3  of air inhaled/day  (ICRP  1975),  pulmonary
               absorption rate of 40% (USEPA 1982).

     In addition to the available monitoring data, Systems Applications (1982)
has provided estimates of atmospheric levels of carbon  tetrachloride  by apply-
ing  air  dispersion  models  to  carbon  tetrachloride  emission  sources.    The
computed average  concentrations  of carbon  tetrachloride and  the number  of
individuals estimated to  be  exposed to these  concentrations  are  presented  in
Table  IV-V.   Specific  point  sources are individually  identified  sources  with
known  locations and  modes and rates of emissions.   These are generally manu-
facturing plants.   General point  sources are  those  which  are numerous, small,
or  of  uncertain   location.    However,  these  sources can  produce  isolated
patterns of  significant concentration;.   Area  sources  are numerous  and  emit
only  small   concentrations  of  the  chemical  (e.g.,  home chimneys,  automo-
biles).  These  estimates  indicate that less  than 600,000 persons are exposed
to  airborne  carbon tetrachloride  at concentrations greater  than  2,500 ng/m3
(2.5 ug/m3).
     Table  IV-V also presents  a  total  population-concentration  estimate for
carbon tetrachloride  of 6.45 x  107 ug/m3 x  persons.   Assuming an inhalation
rate  of  23  m3  of  air/day and a 40% pulmonary  absorption rate, a population-
dose of 5.93 x  10° ug/day x persons was calculated.

     Table IV-VI presents a  general  view of the total  amount of carbon tetra-
chloride received  by  an adult male  from air, food, and drinking water.  Four
separate exposure  levels  in  air, five  exposure levels in drinking water, and
one exposure level  from foods are shown in the  table.
     The data presented have  been selected from an  infinite number of  possible
combinations  of concentrations  for  the three  sources..  The actual   exposures
encountered  would  represent  some  finite  subset of  this infinite  series of
combinations.   Whether  exposure occurs  at  any specific combination of levels
is  not known; nor  is it possible  to  determine the number  of  persons  that  would
be  exposed  to  carbon  tetrachloride at any  of the combined exposure  levels.
The  data  presented represent possible  exposures based on the  occurrence data
and the estimated  intakes.
      The  relative  source  contribution data  for carbon  tetrachloride  account
for  differential   absorption  rates  for the  chemical  by  the respiratory  and
gastrointestinal routes.  Thus,  relative doses of the  chemical  directly enter-

                       Table  IV-V.   Exposure and Dosage Summary for Airborne Carbon Tetrachloride
3.8 x 10"5
Specific General
point point
source source
exposed (persons)
Area source
U.S. total
Dosage (ug/m x persons)
Specific General
point point
source source
Area source
U.S. total
Note:  The use of "--" as an entry indicates that the incremental
       not significant (relative to the last entry in that  column
       that the exposure of the same population may be counted in

Source:  Systems Applications 1982
increase In the dosage or
or to an entry in another
another column.
the population exposed Is
column at the same row) or

ing the  body  are compared.   However,  it should be  noted that the  relative
effects of the chemical  on the body may vary by different  routes of exposure.
     Brodzinsky and Singh  (1982) calculated  a  median  urban/suburban  air  level
of carbon tetrachloride of 1.2  ug/m3 based  on  air monitoring  data.   Assuming
an air  level  of  1.2  ug/m3 and the  estimated  carbon  tetrachloride  intake  of
0.018 ug/kg/day  in  foods, drinking  water would be the predominant  source  of
carbon tetrachloride exposure in the adult male at drinking water levels  above
6.3  ug/1.   An  accurate  assessment  of  the  number of  individuals for  which
drinking water is the predominant source of exposure  cannot be determined from
the  data since  specific  locations  containing  high  concentrations  of  carbon
tetrachloride in drinking water and  low  concentrations of carbon tetrachloride
in ambient air and food are unknown.
     Population-dose estimates  for carbon tetrachloride in drinking water and
air were presented previously.  Estimates  for drinking water ranged from 0.66-
5.0  x  108 ug/day  x  persons;  the estimate  for ambient air was  5.9 x 109 ug/day
x  persons.   These estimates suggest  that ambient  air may be a greater source
of exposure to carbon  tetrachloride  than  drinking water  on a  general popula-
tion  basis.   Comparison  of  these  estimates,  however,  may be  deceiving since
the  same population-dose  level  can occur if:   1) a whole  population  is exposed
to moderate  levels of  a  chemical  or  2X some  segments  of the  same  population
are  exposed  to  high  levels  and  others  to  low levels.   The  population-dose
values  presented  give  no  indication  of the relative  predominance of drinking
water  and air as  specific sources  of carbon  tetrachloride on  a site-by-site or
subpopulation basis.

        Table IV-VI.  Estimated Dose of Carbon Tetrachloride Absorbed
              from  the  Environment  by  Adult Males in ug/kg/day
                           (X from Drinking Water)
Concentration in Concentration in air
drinking water Rural /remote Urban/suburban
(ug/1) (0.82 ug/m3) (1.2 ug/m3}
0 0.13 (0%) 0.18 (0%)
0.5a 0.14 (9.9%) 0.19 (7.3%)
5.0b 0.28 (50%) 0.32 (44%)
10C 0.42 (69%) 0.47 (62%)
20d 0.71 (80%) 0.75 (76%)
Intake from each source (see Sections 5.1-5.3):
Water: 0.5 ug/1: 0.014 ug/kg/day
5.0 ug/1 : 0.14 ug/kg/day
10 ug/1 : 0.29 ug/kg/day
20 ug/1: 0.57 ug/kg/day
Air: 0.82 ug/m;*: 0.11 ug/kg/day
1.2 ug/m;*: 0.16 ug/kg/day
3.7 ug/m;*: 0.49 ug/kg/day
69 ug/m3: 9.1 ug/kg/day
Source dominated
(3.7 ug/m3)
0.51 (0%)
0.52 (2.7%)
0.65 (22%)
0.80 (36%)
1.1 (52%)

(69 ug/m3)
9.1 (0%)
9.1 (0.2%)
9.3 (1.5%)
9.4 (3.1%)
9.7 (5.9%)

Food:                        0.018 ug/kg/day

a26,810,000 individuals  using public drinking water  systems  are estimated to
 be exposed  to  levels   >_  0.5  ug/1  (12.5% of  population  using  public  water
b2,087,000 individuals using public drinking water systems are estimated to be
 exposed to levels > 5.0 ug/1 (1.0% of population using public water supplies).

C698,000 individuals  using  public drinking water systems  are estimated to be
 exposed to levels > 10 ug/1  (0.3% of population using public water supplies).

d655,000 individuals  using  public drinking water systems  are estimated to be
 exposed to levels > 20 ug/1  (0.3% of population using public water supplies).


 Battalia.    1977.   Environmental  monitoring near  industrial  sites:   Methyl -
chloroform.    Prepared by  Battelle Columbus  Laboratories,  Columbus,  OH,  for
 U/S.  Environmental Protection Agency.   EPA-560/6-77-025.   Cited in Brodzinsky
 and Singh  1982.

 Brodzinsky R, Singh HB.   1982.   Volatile organic chemicals in the atmosphere:
 An assessment of  available  data.  Prepared by  SRI  International, Menlo Park,
 CA, for  Environmental Sciences  Research  Laboratory,  Office of  Research  and
-Development,  U.S.  Environmental  Protection Agency,  Research  Triangle  Park,
 NC.  Contract No. 68-02-3452.

 FDA.    1980.   Food and  Drug  Administration.    Compliance  program  report of
 findings:    FY  77 total   diet  studies  — adult  (7320.73).    Washington,  DC:
 Industry Programs Branch, Food and Drug Administration.

 FRDS.   1983.  Federal  Reporting  Data System.  Facilities and population served
 by primary  water  supply  source  (FRDS07),  April  19,  1983.   U.S.  Environmental
 Protection Agency, Washington, DC,

 ICRP.   1975.  International  Commission on Radiological Protection.  Report of
 the task group on reference man.  New  York:  Pergamon  Press.   ICRP  Publication

 Lehmann KB,  Schmidt-Kehl  L.   1936.    The thirteen most important  chlorinated
 aliphatic  hydrocarbons  from the  standpoint  of  industrial  hygiene.   Arch.
 Hyg.  116:132-200.  Cited in USEPA 1982.

 Letkiewicz  F,  Johnston   P,  Macaluso   C, Elder  R,  Yu U,  Bason  C.   1983.
 Occurrence of carbon  tetrachloride in  drinking  water,  food,  and air.   Prepared
 by  JRB   Associates,   McLean,   VA,   for  Office   of   Drinking  Water,   U.S.
 Environmental Protection  Agency, Washington,  DC.   EPA  Contract  No.  68-01-6388.

 Marchland  C, McLean  S,  Plaa   GL.    1970.   The  effect  of  SKF  525A on   the
 distribution  of carbon tetrachloride in rats.   J.  Phar.  Exper.  Ther.   174:232.

 McCollister  OD,  Beamer WH,  Atchison  GJ,  Spencer  HC.   1951.   The  absorption,
 distribution, and elimination  of  radioactive carbon  tetrachloride  by monkeys
 upon exposure to  low  vapor concentrations.  J.  Pharmacol.  Exp.  Ther.   102:112-
 124.  Cited  in  USEPA  1982.

 Pellizzari  ED,   Hartwell  T,  Zelon  H, Leninger  C, Erickson  M,  Sparacino  C.
 1982.    Total  exposure   assessment  methodology  (TEAM):    Prepilot  study  -
 Northern  New Jersey.    Prepared by Research  Triangle Institute  for  Office of
 Research  and Development,  U.S.  Environmental  Protection Agency,  Washington,
 DC.  EPA  Contract 68-01-3849.

Stokinger HE,  Woodward  RL.    1958.    Toxicological  methods  for establishing
drinking water  standards.   J. Am.  Water Works Assoc.   52:515-529.   Cited in
USEPA 1982.

Systems Applications.  1982.   Human exposure to atmospheric concentrations of
selected chemicals.  Prepared  by  Systems Applications,  Inc. for Office of Air
Quality Planning and Standards, U.S. Environmental Protection Agency, Research
Triangle Park, NC.  Contract No.  68-02-3066.

USEPA.    1982.    U.S.  Environmental   Protection  Agency.    Health  assessment
document for  carbon  tetrachloride.   Washington,  DC:    Office  of Research and
Development, U.S. Environmental Protection Agency.  EPA-600/8-82-001.

                 V.  HEALTH EFFECTS IN ANIMALS

          This section discusses the acute and chronic effects of

carbon tetrachloride exposure* with emphasis on studies in which

dose-response relationships for minimal adverse effects have been

developed.  The subsection on teratogenicity, rautagenicity, and

carinogenicity outline the various study designs in more detail

and elaborate on their deficiencies where appropriate.

Acute Effects

         The acute toxicity of carbon tetrachloride has been

extensively documented.  This subsection will concentrate on

those studies that  (i) describe nonlethal effects and  (ii)

provide data on a  range of doses from which dose-response

relationships can  be determined.   For this reason a number of

studies referring  to LDso's will not be discussed.  However,

Table V-l summarizes some  of  the lethal dose data reported for

carbon tetrachloride in various species.

          Liver Effects.   Functional changes  in mouse  liver as

a  result  of carbon tetrachloride exposure were measured  by

increases in the activity  of  the enzyme serum glutamic-pyruvic

transaminase (SGPT)  and  in bromsulfophthalein  (BSP) retention

(Klaassen and Plaa,  1966).  Male Swiss-Webster mice were admin-

istered various amounts of analytical  grade  carbon  tetrachloride

intraperitoneally  in corn  oil at  a final  volume  of  10 ml/kg

TABLE v-1  Toxic Doses and Effects of Carbon Tetrachloride
             in Animals
Route of
Animal administration Effect*
Rat Oral LD50
Mouse LD50
Rabbit LD50
Rat Intraperitoneal ^-^50
Mouse ^^50
Dog ^DLQ
Rabbit LDLO
Rat Inhalation ^50
Mouse LC5Q
Guinea pig L^LO
Cat Subcutaneous LDLO
Rabbit LDLO
2,800 mg/kg
12,800 mg/kg
1,000 mg/kg
6,380 mg/kg
1,500 mg/kg
4,675 mg/kg
1,500 mg/kg
478 mg/kg
4,000 ppm/4 hrs
9,526 ppm/8 hrs
38,110 ppm/2 hrs
20,000 ppm/2 hrs
300 mg/kg
3,000 mg/kg

aLD50, dose  lethal  for  50%  of  animals

 LC50' concentration  lethal for  50%  of  animals

 LDi,O' lowest  lethal  dose

 Source: NIOSH1 (1978)

of body weight (bw).   Mice treated only with corn oil were

used to establish the normal range of values for BSP retention

and SGPT activity, which were determined 24 hours after treat-

ment.  The authors reported the median effective doses .of

carbon tetrachloride as 15.9 mg/kg bw for elevation of SGPT

activity and 94 mg/kg bw for BSP retention.  The authors did

not specify the range of carbon tetrachloride doses used or the

number of animals used at each dose.

          In another study, Klaassen and Plaa (1967) further

defined a dose-response relationship for carbon tetrachloride

exposure and elevated SGPT  levels in mice.  They used the "up

and down" method  in which one dose of the compound was given

to an animal and  the animal's SGPT activity 24 hours after

the dose was noted.  If the enzyme was elevated, the dose

was decreased 40% and the experiment repeated in another

animal.  If no effect was noted, the dose was increased 40%

and the experiment repeated in another animal.  This series

was repeated three times after one positive and one negative

response had been obtained.  The results  for mice  are  shown

in Table V-2.  The authors  concluded that  13 mg/kg bw was

the median effective dose of carbon tetrachloride  in mice as

measured by elevated SGPT values.

Table V-2  SGPT Values of Mice Administered Carbon Tetrachloride
              Intraperitoneally in "Up and Down" Experiment

(rag/kg faw)


aE, = Elevated SGPT after 24 hours

 N = Normal SGPT after 24 hours

 Source:  Klaassen and Plaa (1967)

          Sein and Chu (1979) studied the effect of carbon

tetrachloride on the level of the liver enzyme glucose-6-phos-

phatase in mice.  Groups of six male LAC strain mice were

treated intraperitoneally with 795, 1,590, or 3,180 mg/kg bw

of carbon tetrachloride  (purity unspecified) in paraffin

oil.  The animals were sacrificed 24 hours after treatment.

Control animals, number  unspecified, were given paraffin oil

and sacrificed on the same schedule.  The livers were removed

and analyzed for glucose-6-phosphatase.  The results of the

analysis showed that after treatment with carbon tetrachloride

at 795 or 1,590 mg/kg, the enzyme level fell to 40% of the

control value.  At a dose of 3,18'0 rag/kg, the enzyme level

had decreased to 20% of the control value.

          A series of experiments to determine the effects of

single carbon tetrachloride exposures on rats were performed

by Murphy and Malley (1969).  Adult male Holtzman rats (250-

350 g) were orally administred various doses of undiluted

carbon tetrachloride by gavage.  Control animals' were admin-

istered equal volumes of water.  At 2-20 hours after treatment,

animals were sacrificed and liver enzyme activities and

liver weights were measured.  The results are shown in Table

V-3.  The animals receiving carbon tetrachloride at 1,600

mg/kg bw were sacrificed 20 hours after treatment and the

livers examined histopathologically.  The examination showed

extensive fatty infiltration, inflammation, and some centrolo—

bular necrosis.  The liver-to-body weight ratios were also


          Murphy and Malley  (1969) also determined the effect

of single exposures to carbon tetrachloride on the activities

of the corticosterone-inducible  liver enzymes tryptophan pyr-

rolase and tyrosine-«*«-ketoglutarate transaminase.  Groups of

rats  (4-6 in each group, 8 untreated controls) were treated

with  carbon tetrachloride  (0, 400, 800, and 1,600 mg/kg bw)

and sacrificed 5 hours after treatment.   Data showed that

the enzyme levels were increased roughly in proportion to

the dose.
          Similar studies on the effect of carbon tetrachloride

administration on serum activity of liver enzymes in rats were

performed by Drotman and Lawhorn (1978).  Groups of four male

Cox rats were admministered carbon tetrachloride intraperito—

neally at 60, 120, 240, or 480 mg/kg bw in a total volume of

1 ml in corn oil and exsanguinated at specified time intervals.

Serum activities were determined for the enzymes sorbitol

dehydrogenase (SSDH), ornithine carbamyl transferase (SOCT),

aspartate aminotransferase (SAST), and isocitric dehydrogenase

(SICDH).  Liver specimens were taken from each animal and scored

for histopathologial changes.  The results of the enzyme analyses

and histopathology are tabulated., in Table V-4 by dose and

hours after dose.  The SOCT activities showed the best corre-

lation with liver histopathology in time of appearance as

well as extent of damage.  The authors concluded that SOCT

levels are a sensitive indicator of liver damage.

          Effects of acute exposure to  low levels of carbon

tetrachloride were also  reported by Korsrud et  al.  (1972).

Male Wistar rats  (260-400 g;  8-10  animals per treatment  group)

were administered single oral doses of  carbon tetrachloride

          Table V-3   Effects  of  Oral Carbon Tetrachloride on Liver Weight and
                     Liver  and Plasma  Enzyme Activities  in Male Rats

Dose Time
(rag. kg bw) (hr)
Number of Plasma
animals AKTa
7 2.6 +
4 2.1 +
5 13.2 +
5 35.2 +
4 35.2 +
5 18.1 +
4 9.3 +

b 2360 + 182
1813 ± 331
; 1174 + 559
1585 + 148
1596 + 194
2120 + 182
TKTa APa Weight (g/lOOg bw)
93 + 11 14.9 + 1.0 2.75 + 0.06
170 + 12 14.5 + 1.0 2.94 + 0.04
330 +36 14.2+1.3 3.26+0.10
361 + 42 34.6 + 5.8 4.36 + 0.06
305 + 21 37.1 + 1.6 3.95 + 0.07
294 + 61 33.3 + 3.6 3.90 + 0.05
138 + 5 29.1 + 6.5 3.47 + 0.07

aAKT =
AP =
b =
tyros ine
means +
SE in micromoles


product formed

per gram of fresh liver or
       milliliter of plasma per hour
Adapted from Murphy and Malley (1969)

(0-4,000 mg/Tcg bw) in corn oil (5 ml/kg bw).   The rats were

fasted for 6 hours before treatment and for 18 hours after-

ward, and then sacrificed.  Assays included liver weight and

fat content, serum urea and arginine levels,  and levels of

nine serum enzymes produced mainly in the liver.  At 20

mg/kg bw there was histopathologic evidence of toxic effects

on the liver.  These changes included a loss of basophilic

stippling, a few swollen cells, and minimal cytoplasmic

vacuolation.  At 40 mg/kg bw, liver fat, liver weight, serum

urea, and levels of five of the nine liver enzymes were

increased while serum arginine decreased.  At higher doses

the remaining four enzyme levels were also elevated.

          Alumot et £l.  (1976) reported the effects of sub-

chronic exposure to feed that had been fumigated with carbon

tetrachloride.  Groups of six weaning rats 4 weeks old were

fed a diet  containing carbon tetrachloride at  150, 275, or

520 mg/kg of feed for 5 weeks  (females) or 6 weeks  (males).

The fumigated feed was stored  in airtight containers; carbon

tetrachloride loss during the  storage period  of 7-10  days

was determined to be 5%.  The  animals were allowed  access  to

the feed only at  set time intervals  to minimize loss  of

carbon  tetrachloride by  volatilization.  The  authors  calcu-

lated that  the amount of carbon  tetrachloride remaining in

the consumed feed was 60-70%  of  the  amount  initially  present;

Table V-4
Effects of Carbon Tetrachloride on Liver
Histopathology and Serum Enzyme Levels
     Dose   after
   (mg/kg)  dose  Histologya
                   Serum enzyme concentrations
                relative to pretreatment levels
              SOCT     SSDH     SAST     SICDH





  aO = No observable changes.

   1 = Minimal changes.  Large central vein, swelling of hepato-
         cyte, etc.

   2 = Mild degenerative change.  Loss of cord arrangement.

   3 = Moderate degenerative change.  Pale cytoplasm, spindle cell.

   4- = Marked degenerative change.  Centrilobular fatty degeneration,

  * Significantly different from zero time as determined by one-way
    analysis of variance of the log-transformed data  (P £.01).

    Adapted from Drotman and Lawhorn (1978)

the total decrease reflected amounts lost during storage and

after removal from storage to feeding troughs.   From these

data and the weights of the animals, the authors calculated

that 275 rag/kg of feed represented a daily dose of 40 rag/kg

bw.  (By assuming that all parameters were the same and that

the delivered dose was proportional to the concentration in

feed, diets of 150 and 520 rag/kg of feed were calculated by

JRB to represent daily doses of 22 and 76 mg/kg bw, respec-

tively. )  At the end of the experiment the animals were

weighed and sacrificed.  Of the three doses, only the highest,

76 mg/kg bw (520 mg/kg of feed), caused significantly depressed

weight gain in males.  Weight gain in females appeared to be

unaffected by all doses.  Total lipid and triglyceride levels

in the liver were significantly higher in animals fed carbon

tetrachloride at 40 and 76 mg/kg bw than in controls or animals

fed 22 mg/kg bw.  Levels of liver phospholipids  (measured in

females) were not affected at any dose.  Of the  three doses

used in this experiment, the lowest, 22 mg/kg bw, failed to

produce effects on the measured parameters.

          In addition to the study of hepatic effects of carbon

tetrachloride in mice, described earlier in this section,

Klaassen and Plaa  (1967) also investigated the hepatic  effects

of carbon tetrachloride exposure in dogs.  Male  and  female mon-

grel dogs were treated intraperitoneally with carbon tetrachlo-

ride at 22-38 mg/kg bw in an "up and down"  experimental design.

  Blood samples were taken for measurement of SGPT  24 hours  after

  administration of carbon tetrachloride.   Control  dogs had  serum

  SGPT activity of 36+7 units.  Therefore,  36+2  standard  devia-

  tions or 50 units were chosen as the upper limit  of the normal

  value.  The results of the analysis are  shown  in  Table  V-5.

            The SGPT values returned to normal in 17-18 days.

  Animals were then sacrificed and the livers were  examined  his-

  topathologically.  They showed moderate  vacuolation of  the

  centrolobular and midzonal well as traces of

  brown material in the cytoplasm of centrolobular Kupffer cells.

Table V-5  SGPT Activity in Dogs 24 Hours  After Intraperitoneal
           Administration of Carbon Tetrachloride in "Up  and
           Down" Experiment



  aN = normal SGPT after 24 hours

   E = elevated SGPT after 24 hours

  Adapted from Klaassen and Plaa  (1967)

          Kidney Effects*  Carbon tetrachloride,  even at

high doses, failed to induce renal failure as measured by

phenolsulfonphtalein (PSP) excretion in mice although patho-

logical kidney alterations were present (Plaa and Larson,

1965) .  Male Swiss mice  (18-30 g) were given intraperitoneal

injjections of carbon tetrachloride (1,6006,400 mg/kg bw)

dissolved in corn oil at a final volume of 0.1 ml/10 g bw.

The animals were then hydrated with tap water (50 ml/kg bw)

by gavage and placed on  a urinary collection unit for 2

hours.  Even carbon tetrachloride doses lethal in some

animals (>^ 6,400 mg/kg bw) failed to cause renal dysfunction,

measured as excretion of PSP, urinary protein, and glucose,

in the majority of survivors.  At a high  nonlethal dose

(3,260 mg/kg bw) minimal renal dysfunction was observed

after 96 hours.  Histologic  examination of kidney sections

from five mice  that had  been administered this dose  showed

necrosis of proximal convoluted  tubules  (n=l) and swelling

of the  tubules  (n=4).

           Carbon  tetrachloride decreased  the activity of

glucose-6-phosphatase  in the kidney (Sein and Chu,  1979).

Male mice  (40-50  days  old,  weighing 24-28 g) were  injected

intraperitoneally with carbon tetrachloride at  795,  1,590,  or

3,180 mg/kg bw  in paraffin oil.   Twenty-four hours  after injec-

tion  of 795 or  1,590  mg/kg bw,  the renal glucose-6-phosphatase

activity decreased to 77% or 65% of the control value,  respec-

tively.  Increasing the dose to 3,180 rag/kg bw had no further

effect on the kidney enzyme level.

          These results were in contrast to the liver glucose-

6-phosphatase level discussed earlier, which decreased to 40%

of the control value at the two lower doses and decreased

further to 20% of the control value at 3,180 rag/kg bw.   The

authors attributed these differences to the limited meta-

bolic capacity of the kidneys.

          Klaassen and Plaa (1967) studied the effect of carbon

tetrachloride exposure on kidney  function in dogs.  PSP excre-

tion of less than 39% of control  values was considered indica-

tive of renal dysfunction.  An unspecified number of male

and female mongrel dogs were treated intraperitoneally with

carbon tetrachloride at 22-38 mg/kg bw and the 24 hour excre-r

tion rate for PSP was determined.  Control dogs were used to

determine a normal range for PSP  excretion.  None of the

dogs treated with carbon tetrachloride exhibited decreased

PSP excretion.  However, on histological examination of the

kidneys from the treated dogs, the Bowman's capsules appeared

dilated with some contraction of  glomerular tufts and calcifi-

cation of a small number of tubules  in the medulla.

          Lung Effects.  Boyd et al. (1980) investigated the

effect of ingestion and inhalation of carbon tetrachloride

on pulmonary Clara cells in Swiss mice.  For the ingestion

study, the mice were treated with carbon tetrachloride (4,000

m9/k9 fcw) in a 50% sesame oil solution and sacrificed 16

hours after treatment.  The lungs were removed and examined

by electron microscopy.  Clara cells exhibited massive

dilation of vesicles of smooth endoplasmic reticulum, increased

mitochondrial staining density, ribosomal disaggregation, nuclear

condensation, and occasional cellular necrosis.  Additional

experiments with oral carbon tetrachloride doses of less

than  1,600 ing/Kg bw did not produce any pulmonary lesions

visible by light microscopy.  Doses of 2,400-4,800 mg/kg bw

produced Clara cell lesions similar on electron microscopic

examination to those previously.-described.  The extent of

damage was proportional to the dose administered.  Boyd et

al.  (1980) also studied the time  course of the Clara  cell

damage caused by  ingestion of  carbon  tetrachloride  (4,000

mg/Tcg bw).  Pulmonary  tissue was  evaluated by light micros-

copy  at  12, 24, 36, 48, 96, and  168 hours.  The  lesions were

present  at 12 hours,  maximal at  24 hours,  and less  intense

at 36 hours.  By  48 hours, the lesions were seen infrequently

and  at 96 and 168 hours the pulmonary bronchioles appeared


          The pulmonary toxicity of inhaled carbon tetrachlo-

ride was also studied by Boyd et al. (1980).   Swiss mice were

exposed to carbon tetrachloride vapor at 71,800,  144,000,

287,000 or 574,000 mg/m3 for 60, 60, 12, or 2 minutes,  respec-

tively.  The animals were sacrificed 24 hours after exposure,

and the lungs examined.  Marked Clara cell lesions similar to

those seen after oral exposure were seen at all exposure levels

and necrosis was reported to be more frequent after inhalation

than after oral exposure, but no effort to quantify this find-

ing was reported.

          Gould and Smuckler (1971) investigated the structural

alterations in rat lungs following carbon tetrachloride inges-

tion.  Male Sprague-Dawley rats  (200-250 g) were fasted 16

hours prior to administration of carbon tetrachloride (4,000

mg/kg bw) by gavage.  The animals exhibited piloerection

and lassitude 3-4 hours  after treatment.  They were sacrificed

1, 4, 8, 12, or 24 hours following  treatment.  Necropsies were

performed on all animals.  Microscopic  examination of the

lungs of treated rats  revealed  perivascular  edema  and mono-

nuclear infiltration in  the  first 4 hours  after treatment.

These areas were local but were estimated  to involve 10% of

the parenchyma.  Areas of atelectasis  and  intraalveolar hemor-

rhage involving  15-20% of the parenchyma were observed  8-12

hours after treatment.

          Electron micrographs of rat lungs after carbon tetra-

chloride ingestion showed granular pneumocytes containing

swollen inclusions with decreased osraiophilia and attenuated

lamellae 1 hour after treatment (Gould and Smuckler,  1971).

These changes were more severe 4 hours following treatment.

By 4—8 hours after treatment, cytoplasmic edema, dislocation

of dense ribosomal aggregates, and mitochondrial disruption

were apparent.  Multivesicular bodies were "conspicuously

decreased" within the granular pneumocytes.  Necrosis was

evident 12-24 hours after treatment.  One hour after administra-

tion, endothelial cells displayed markedly increased pinocytotic

vesicles.  Severe disruption of endothelial cells was evident

from 8 hours onward.  Ultrastructural damage was seen in all

components of the alveolar wall, and fibrin was observed

within alveoli.  The authors  interpreted these  findings as

indicative of significant alterations in vascular permeability.

          Lesions of the Clara  cells in the  lungs of male

Sprague-Dawley rats orally treated with carbon  tetrachloride

were observed by Boyd et al.  (1980).  The  carbon tetrachloride

was administered by gavage at doses  of 3,816, 5,088, and

7,155 mg/kg as a 50% solution in  sesame oil.  Control animals

received  sesame oil only.  Clara  cell  lesions occurred  at

the two highest doses.   The  authors  stated that the  lesions

were less pronounced than those, in mice exposed to comparable

amounts of carbon te^rachloride.

Chronic Effects

          Smyth et al.  (1936) studied the chronic effects of

carbon tetrachloride inhalation exposure in rats.  Groups of

24 Wistar rats were exposed to carbon tetrachloride concentra-

tions of 315, 630, 1,260, or 2,520 mg/m3 (50, 100, 200, or

400 ppm) for 8 hours a  day, 5 days a week for 10.5 months.

The carbon tetrachloride was found to contain less than

0.003% carbon disulfide.  Control rats were used, but the

number was unspecified.  Growth retardation was observed

at 2,520 mg/m3.  At 630 and 1,260 mg/m3, growth was the

same as in controls, and at 315 mg/m3 growth was stimulated.

Cirrhosis developed in  rats exposed to 630, 1,260, and 2,520

mg/m3 after 173, 115, and 54 exposures,  respectively.  When

exposure was stopped, fatty liver degeneration  resolved within

50 days.  Surface  alterations- (hobnail  liver) did not  resolve

until 156 days  after cessation  of exposure.  Unspecified

renal damage was observed after 52^  exposures  to 315 mg/m3  and

after 12-20 exposures at the higher  concentrations.

          In a  chronic  oral  exposure study  (Alumot  et  aj..,

1976),  groups erf 36 rats  (18  male and 18 female littermates)

were  fed mash containing carbon tetrachloride at 0,'80,  or

200 mg/kg of feed.  The feed  was  stored in  airtight containers,

assayed for carbon tetrachloride content, and consumed soon

after removal to feeding troughs.  The authors calculated

that 200 mg/kg of feed represented a daily dose of 10-18

m9/fcg bw.  After 2 years, the surviving animals were sacri-»

ficed.  In these animals, serum values for glucose, protein^

albumin, urea, uric acid, cholesterol, SCOT, and SGPT in jfche

treated animals did not differ from those £n controls.  No

fatty livers were detected in the treated animals.  Thus,

the authors found no biochemical abnormalities attributable

to carbon tetrachloride exposure.  However, interpretation

of the results was complicated by the widespread incidence

of chronic respiratory disease in the animals.  More than

half the animals were dead at 21 months, although at 18

months the survival ranged from"¥l-89%.  The authors

indicated that 10-18 mg/kg bw (200 mg/kg of feed) is a no-

adverse-effect level of carbon tetrachloride over 2 years.

However, this conclusion may be questioned because of the

poor survival rate and widespread respiratory infection of

experimental animals.

          Groups of 24 guinea pigs were exposed to carbon tetra-

chloride vapor at 315, 635, 1,260, 2,500 mg/m3 in a study by

Smyth et al. (1936).  The frequency of exposure was 8 hours per

day, 5 days per week for up to 10.5 months.  Marked mortality

occurred in exposed animals:  9/24 at 315 mg/m3 after a median

of 44 exposures (exposure terminated at 135 days),  16/24 at

630 mg/m3 after a median of ten exposures, 13/24 at 1,260

mg/m3 after a median of three exposures, and 19/24 at

2,520 mg/m3 after a median of three exposures.

          Guinea pigs exposed to the 315 mg/m3 dose in the Sroyth

et a_l., (1936) study developed cirrhosis and hobnail surface

alterations of the liver in 105 exposures.  The authors

concluded that survival of guinea pigs at higher doses was

of insufficient duration to allow development of cirrhosis.

In addition, granular swelling was observed in adrenal glands

of guinea pigs exposed to carbon tetrachloride at 315, 630,

and 260 mg/m3  for 8, 7, and 17 exposures,  respectively.

Exposure to higher concentrations  (1,260  or 2,520 mg/m3) or

continued exposure to lower concentrations resulted  in marked

damage to the  sciatic nerves.  Dense  clumps of black granules

 (osmic acid stain) were observed paralleling  the  large majority

of  fibers.

          Prendergast et  al.  (1967)  repeatedly  exposed  15  guinea

pigs  to carbon tetrachloride  (purity unspecified)  at 515 mg/m3

 (82 ppm) over  a period  of 6 weeks  and observed  hepatic  changes.

Three guinea pigs died  on days 20,  22,  and 30,  respectively.

All  the animals showed  a  body weight loss.  The surviving

 animals were  sacrificed at 6  weeks and the livers examinined

histopathologically.  The examiniation revealed fatty infiltra-
tion, fibrosisr bile duct proliferation, hepatic cell degenera-
tion and regeneration, focal inflanunatory cell infiltration,
alteration of lobular structure, and early portal cirrhosis.
The hepatic lipid content of carbon tetrachloride-treated
animals (35.4 _+ 10.7%) was higher than that of the controls
(11.0 + 3.6%) .
          In addition, Prendergast e_t a_l. (1967) exposed
continuously guinea pigs to carbon tetrachloride vapor at 61
mg/m3 (10 ppm) for 90 days.  Three of the 15 guinea pigs
died: on days 47, 63, and 74 respectively.  All the exposed
animals showed a depressed weight gain.  A  "high incidence"
of enlarged and discolored livers was reported on gross
pathological  examination.  Histopathologic  examination of
the  livers revealed fatty changes,  fibroblastic proliferation,
collagen deposition,  hepatic  cell degenertion  and  regeneration,
and  structure alteration of  the liver  lobule.  Enzymatic
studies showed that only  the  succinic  dehydrogenase  (SDH)
activity was  moderately reduced as  compared to that  in  controls

          In addition to their studies on guinea pigs, Pren-

dergast et al. (1967) studied the effects of both repeated

and continuous exposure to carbon tetrachloride on three

squirrel monkeys, three New Zealand rabbits, and two beagle

dogs for each exposure regimen.  The experimental designs

were the same as those described for the guinea pigs.  All

the animals showed a weight loss during repeated exposure to

515 mg/ra3 (82 ppm).  Fatty changes were noted  in the liver

of all species; they were most severe in rabbits, followed

by dogs and monkeys.  In the continuous exposure to 61 mg/m3

(10 ppm) for 90 days, all species exhibited a  depressed

weight gain, as did guinea pigs.  Liver changes were also

noted, but enzyme activities  (as measured by NADH, NADPH,

SDH, LDH G6PI) were within the normal range.   At a continuous

exposure of 6.1 mg/m3  (1 ppm)-r no toxic signs  were noted.


          In two studies on  the  teratogenic and prenatal  toxi-

cologic effects  of  carbon  tetrachloride,  the chemical  was

reported to produce  prenatal  toxicity  following  inhalation


          Schwetz et al. (1974) exposed pregnant rats to car-

bon tetrachloride at 1,800 or 6,300 rag/m3 (300 or 1,000

ppm) for 7 hours per day on days 6-15 of gestation.  Statisti-

cally significant decreases in fetal body weight and crown-

rump length were observed.  Other parameters examined were

not significantly different from those of controls.  The

authors concluded that carbon tetrachloride was not terato—

genie at the concentrations used in this experiment.  Two

other statistically significant fetal effects were noted:

an increased incidence of sternebral anomalies in the 6,300

mg/m3 group and an increased incidence of subcutaneous edema

in the 1,800 mg/m^ group.  Edema was not seen in the 6.300

mg/m3 group.  The dams exposed to both concentrations of

carbon tetrachloride showed a statistically  significant

decrease in weight gain and also decreased food consumption

compared to control animals.  Hepatotoxicity as measured by

significantly increased SGPT activity was also seen  in the

dams.  However, the authors did not  establish any  consistent

pattern between fetal toxicity and maternal  toxicity at  the

subanesthetic levels of carbon tetrachloride used  in this


          Another study reported  no  teratogenic  effects  fol-

lowing exposure of pregnant  rats  to  carbon  tetrachloride at

1,575 mg/m3 (250 ppm) 8 hours per day for 5 consecutive
days between days 10-15 of pregnancy (Oilman, 1971).  Concomit-
ant exposure to 15% ethanol in drinking water also did not
result in teratogenic effects.  Carbon tetrachloride exposure,
however, did decrease the viability index to 83% as compared
to 99% for controls.  The lactation index was also decreased
to 83% as compared to 98% for controls.  A decrease in the
number of pups per litter also occurred:  9.2 as compared
to 10.3 for controls.  Concomitant ethanol exposure exacerbated
this effect:  8.48 pups per  litter as compared to  10.3 for con-
          No teratogenic effects were noted  in rats fed diets
containing carbon tetrachloride at either 80 or 200 mg/kg of
feed for up to  2 years  (Alumot et al.,  1976).  Groups  of 18
female rats were mated, at  3  months of  age  (6 weeks of  exposure)
with untreated  males.   Thereafter, they were mated every 2
months with groups  of  treated males.

           Exposure  to  carbon tetrachloride in  utero has been
 reported to result  in  liver damage  in rat  fetuses  and  neonates
 (Bhattacharyya,  1965).   In one  case,  subcutaneous  administra-
 tion of carbon  tetrachloride at 1,600 mg/kg bw to  a pregnant
 rat on  day 20 of gestation resulted in small areas of  focal
 hepatic necrosis in a  fetal liver 24 hours later.   Similar
 treatment  resulted  in  focal hepatic necrosis in neonates born
 48 or 72 hours  after treatment of dams on day 19 or 20 of

gestation.  Histologic findings generally included a sharply

demarcated area of centrolobular necrosis and proliferative

changes in nonnecrotic lobes.

          In addition, fetuses were directly treated with

carbon tetrachloride by subjecting the mother to a laparotomy

and either injecting the chemical directly into the fetus or

into the amniotic sac through the uterine wall (Bhattacharyya,

1965).  Liver changes following injection of 6 mg of carbon

tetrachloride were variable:  cells generally became extremely

pale in centrolobular and midzonal areas, indicating fatty

infiltration.  Livers remained abnormal until at least 4

days after birth.  No necrosis, hemorrhage,  or regeneration

was observed.

          Sensitivity of neonate rats  to carbon tetrachloride

was reported  to  be  low  1 hour  after birth, then to  rise

above the adult  level at 19  hours and  to decline by 3-7 days

after birth.  Thus,  only two of  ten 1-hour-old neonates

receiving carbon tetrachloride  (1,600  mg/kg)  subcutaneously

showed centrolobular necrosis  after 24 hours.  In  addition,

hepatic portal areas contained numerous neutrophils,  but  no

bile duct proliferation could  be observed,  in contrast to

findings  in adult animals.   Following  the  same treatment,

19-hour neonates showed more pronounced hepatic  damage than

1-hour neonates. Damage declined  in  3-and 4-day  old  neonates;

that in 5-,6-, and 7-day-olds was similar in appearance to

that of adults.

          Carbon tetrachloride can apparently be transferred

to the neonates through mother's milk  (Bhattacharyya, 1965).

Subcutaneous -administration of carbon  tetrachloride at 1,600

or 3,200 mg/kg bw to four- nursing rats resulted in hepatic

damage in the neonates 24 or 48 hours  later.  A dose of 800

mg/kg bw to dams did not produce any hepatic damage to off-


Reproductive Effects

          Testicular degeneration was  observed in rats receiv-

ing carbon tetrachloride  at 4,800 mg/kg bw  intraperitoneally

 (frequently unstated)  (Chatterjee,  1966).   One group of six

male rats  received  carbon tetrachloride as  a 1:1 mixture  in

 coconut oil;  a  control group  received  only  an equal volume

 of  coconut oil.  On day  15  all  animals were sacrificed.

 Body weights  were  similar for treated  and control animals.

 However, the  relative  testes  weight decreased  from  15.5  (+

 0.4) g/kg  bw  in controls  to 9.8 (+ 1.2)  g/kg bw  in  exposed

 animals.   Relative weight of seminal vesicles  showed  an  even

 more pronounced decrease:   1.27 (+0.171) g/kg bw  in  treated

 as  compared to 3.10 ( + 0.059) g/kg bw in control animals.

 Relative pituitary weight was also increased:   50  (  +_ 1.4)

mg/kg bw in treated as compared to 32.4 (j^ 0.9) mg/kg bw in

control animals.

          Histological examination of testes showed testicular

atrophy and "some abnormality" in spermatogenesis in carbon

tetrachloride-treated animals.  The authors proposed a mechan-

ism for carbon tetrachloride-induced testicular atrophy in

which blockage of pituitary hormone release results in atrophy

of Leydig cells within the seminal vesicles, followed by an

abnormal spermatogenesis.

          Intraperitoneal administration to male rats of carbon

tetrachloride (4,800 mg/kg bw as a 1:1 mixture of coconut oil)

for 10, 15, or 20 days (Group I, II, or III) led to impairments

in spermatogenesis as indicated by histological examination

(Kalla and Bansal, 1975).  Controls were administered equal

volumes of coconut oil.  Weights of testes, seminal vesicles,

epididyrais and prostates were decreased in exposed animals,

whereas the weight of adrenal glands  increased  (see Table V-6).

The gonadosomatic index  (GSI)  (equal  to body weight x testes

weight/100) was also decreased  in  treated  animals.  A slight

decrease in pituitary weight  was observed  following a 10-day

treatment but not after  either  the  15- or  20-day  treatment.   As

Table V-5 shows, the ratio of germinal to  nongerminal area

steadily decreased from  Group I  to  Group  III  and  was  always

higher in treated than control  animals.   Significant  differences

in total germinal area between treated and control animals,

however, were observed only at 20 days.  Histological examination

did not reveal any abnormalities in testes from Group I.

Clusters of mature sperm were present in the lumen.  In Group

II, slight testicular damage was observed; a decrease in

spermatogenic cells and increased lumen size.  In Group III,

shrinkage of the tubules and increased area of the lumen were

observed.  Arrangement of the germ cells was disrupted; early

gonadal cells were present in the lumen of many of the tubules.

No spermatids were observed.  Intersititial material was "dam-

aged" and in many places the basement membrane was detached

from the epithelium.

          Thus, carbon tetrachloride, at  a total dose of 48

g/kg bw over 10 days, had a distinct but  minor effect on male

rat reproductive physiology, whereas a total dose  of 96 g/kg

bw over 20 days resulted in severe disturbances of spermato


           Table V-6  Weight Changes  in Male Reproductive Organs After Carbon Tetrachloride Treatment

Treatment Body weight (g)
and Before After
Period treatment treatment
Group I
10 days
Control 257+5* 269+4.5
Treated 257+13.71 247+10.76
Group II
15 days
Control 230+15.49 235+5.0
Treated 230U5.49 173+13.42
Group III
20 days
Control 234+5.5 235+6.5
Treated 230+3 .5 2 10+0 . 5
Test is
(gAg bw)

8 .84+1 .04;


(gAg bw)









1. 29+J). 01







aMean +_ standard deviation.

Adapted from Kalla and Bansal (1975).

          There have been no reports of rautagenic activity
associated with carbon tetrachloride in any of the various
Salmonella (Ames) assays.  However, mutagenic activity asso-
ciated with carbon tetrachloride has been reported in a eukary-
otic test system using the yeast Saccharomyces cerevisiae.
This recent report has not been confirmed and should not be
accepted as conclusive evidence of carbon tetrachloride rauta-
genicity.  Nevertheless,  these eukaryotic test systems can be
used to screen for mutagenic chromosomal effects  that cannot
be detected in prokaryotic systems.  Gallon et al.  (1980)
have suggested that  the yeast system is more  sensitive than
the Salmonella assay system because active metabolites are
produced in much closer priximity  to the  nucleus  than they
are with liver S-9  in vitro activation systems used  in bac-
terial assays*   Proximity to  the  nucleus  would be advantageous
if the active metabolites were"particularly unstable or  reac-
tive, as is likely  to be  the  case  for  those  of halogenated
hydrocarbons  such  as carbon tetrachloride.
           Gallon et_ a_l.  (1980)  reported that carbon tetrachlo-
ride,  in addition to six other hydrocarbons,  induced a  mutagenic
response in S_.  cerevisiae strain D7.   Unlike the Salmonella
system,  in which promutagens must be treated with exogenous
 liver S-9  activation systems, the D7 strain of S_. cerevisiae
contains an endogenous cytochrome P-450 dependent mono-oxy-
genase activation systems.  In addition, strain D7 can be
 used  to  detect  both gene crossover and mitotic recombination.
 Three  concentrations of carbon tetrachloride (21, 28, and 34
 mM)  were  incubated at 37°C for either 1 or 4 hours  in 3-ml


aliquots of the yeast cell suspension (3 x 10^ eelIs/ml).

Treatment was terminated by the addition of 40 ml of ice-cold

buffer followed by centrifugation to remove the cells from

the media.  The resuspended cells were then plated on appro-

priate media to allow for estimation of mutagenic activity.

A spectral analysis of the cell suspension demonstrated that

carbon tetrachloride was gradually altered during the incuba-

tion periods; the authors concluded that it was absorbed and

metabolized by the cells.

          The 1-hour treatment of cells with carbon tetrachlo-

ride resulted in significant increases in gene crossover and

mitotic recombination.  Results for the different loci are

presented in Table V-7.  When cells were treated with increasing

carbon tetrachloride concentrations there was decreased  cell

survival and increased incidence of gene conversion and  mitotic

recombination.  There was a nonlinear correlation between  cell

survival and the concentration of carbon tetrachloride in  the

incubation mixture.  Only marginal mutagenic  activity was  noted

when incubation was  continued  for 4 hours  (data  not  reported

by  authors).  This observation  could have  resulted  from  toxicity

masking the  genetic  effects,  the  continued metabolism  of active

metabolites  to inactive products, or  the  destruction of  the

metabolic system.  Gallon  et  al.   (1980)  noted that carbon

 tetrachloride had been reported to  be inactive in test systems

 which used an exogenous mammalian activation system and  suggested

 that their yeast system was more  sensitive than some of  the

 other in vitro test  systems.

      TABLE V-7     Mutagenic Effects of Carbon Tetrachloride
                    on Strain D7 of Saccharomyces Cerevisiae3-
                                          Concentration (ing/liter)
                                          0     3234   4312    5128

     Total colonies                     1454    1252   1120     152

     % of control                        100      86     77      10

trp 5 locus (gene conversion)

     Total convertants                   285     331    350     506

     Convertants/I05 survivors             2.0     2.6    3.1    61.7

ade 2 locus (mitotic recombination)

     Total twin spots                      1       3      3      10

     Mitotic recombinants/104 survivors    1.6     5.3    5.8    40.1

     Total genetically altered
     colonies                             11      19     16      65

     Total genetically altered
     colonies/10^ survivors                1.7     3.4    3.1    33.3
ilv 1 locus (gene reversion)
Total revertants
Revertants/106 survivors

38 41 57 11
2.6 3.3 5.1 7.2

 a  The total number of colonies  in  the different  classes  represent  total
 counts of colonies from five plates in the  case of  survival,  conversion,
 and revertant-frequency estimations.  Mitotic  recombination was  estimated
 from counts of colonies growing  on  a total  of  30  plates,  20 plates  con-
 taining medium of which all surviving cells grew  and  10 plates containing
 medium on which only trp 5 convertants grew.

 Adapted from Callen et al. (1980)


          Studies to determine the rautagenic activity of carbon

tetrachloride in the Salmonella typhimurium system have been

uniformly negative (see reviews:  McCann et_ a^., 1975; Fishbein,

1976; and Rinkus and Legator, 1979).  Carbon tetrachloride has

also been void of mutagenic activity when tested in Escherichia

coli (Uehleke et al., 1976).

          Carbon tetrachloride did not induce chromosome damage

(i.e., chromatid gaps, deletions, or exchanges) during an in

vitro chromosomal assay using cultures rat-liver cells (Dean

and Walker, 1979).  Mirsalis and Butterworth  (1980) found that

treatment of adult male Fischer-344 rats  (200-250 g) with car-

bon tetrachloride  (10 or  100 mg/kg, po) produced no increase  in

unscheduled DNA  synthesis in cultures of  primary rat hepato-

cytes.  According  to the  authors these results  indicate  that

carbon tetrachloride does not act through a genotoxic mechanism

and confirm the  report of Craddock  and Henderson  (1978)  that

this  chemical does  not induce DNA repair  in hepatocytes  immedi-

ately  following  treatment.

          Uehleke  et  al.  (1977)  studied  the  interaction  of  car-

bon  tetrachloride  in  a  liver microsome  system used in conjunc-

tion  with assays in S_. typhimuriuro  strains TA 1535 (test for

base  pair substitutions)  and TA 1538 (tests  for frame shift

mutations).   The authors  incubated 3-4C-labeled carbon tetrachlo-

ride (1  mM) with rabbit  liver microsomes pretreated with pheno-

barbital (5  mg  of protein/ml)  and a NADPH regenerating system

at 37°C  for  60  minutes.   A total of 10% of the radioactivity


from carbon tetrachloride was irreversibly (covalently) bound

to endoplasraic portein and more than 30% was bound to microsomal

lipid.  No mutagenic activity was observed in S3. typhimurium

strains TA 1535 or TA 1538 which were incubated with 8 mM carbon

tetrachloride and microsomal suspensions.  The authors concluded

that a reactive species generated in the biological system may

not distribute into the incubation medium and thus may be

inaccessible to the test bacteria.  They also speculated that

the active products of carbon tetrachloride may have very short



          The carcinogenic effects of carbon tetrachloride have

been well documented (IARC, 1979).  IARC has judged the evidence

from animal studies demonstrating that carbon tetrachloride in-

duced hepatic neoplasms as conclusive for experimental animal

carcinogenesis  (IARC, 1979).

          In a NCI bioassay program, carbon tetrachloride was

used as a positive control in the bioassays of  chloroform and

1,1,1-trichloroethane  (NCI 1976, 1977).  The positive  control

groups described  in both bioassays were  of the  same strain and

source as the treated animals and were housed identically.

Groups of 50 Osborne-Mendel rats of each sex were  administered

carbon tetrachloride in corn oil by gavage  five times  weekly

for 78 weeks at two dose  levels:  47 and 94 rag/kg  bw  for males,

80 and 160 mg/kg  bw for females.  This  treatment resulted  in

toxicity: at 110  weeks at  the highest dose, only 7 of 50 males


  and 14  of 50  females survived as  compared  to 26  of  100 males and

  51  of 100 females  for controls.   The incidence of hepatocellular

  carcinomas was  increased in  animals  exposed .to carbon tetrachlo-

  ride as compared to controls (see Table V-8).  Absolute  incidence

  of  hepatic neoplasms was low.  The apparent  decrease in  the

  incidence of  hepatocellular  carcinomas in  female rats at the

  high dose was attributed to  increased lethality,  i.e., females

  died before tumors could be  expressed.  The  incidence of other

  neoplasms in  these rats was  acknowledged but not quantified.

  Table V-8.         Incidence of  Liver Tumors in  Carbon
               Tetrachloride-Treated Rats and  Colony  Controls

  Animal  group         Hepatocellular carcinoma     Neoplastic  nodule


Low dose
High dose
Low dose
High dose

Sources NCI (1976)

            Groups of 50 B6C3F1 mice of each sex were administered

  carbon tetrachloride in corn oil by gavage five times weekly for

  78 weeks at two doses, 1,250 and 2,500 mg/kg bw, for both males

  and females.  High exposure-related lethality occurred in all

  groups.  At 78 weeks at the high dose, only 2 of 50 males and 4

  of 50 females survived as compared to 53 of 77 V-3& males and 71

  of 80 females for controls.  At 91-92 weeks at the high dose,


  none of 50 males and 1 of 50 females survived as compared

  to 38 of 77 male and 65 of 80 female controls.   An exposure-

  related increase in the incidence of hepatocellular carcinomas

  was observed (see Table v-9).  The average latency for appearance

  of liver tumors was also significantly decreased in carbon

  tetrachloride exposed animals.   In high-dose males, the latency

  was 26 weeks, as compared to 48 weeks in low-dose males and 72

  weeks in control males.  In high-dose females,  the latency was

  19 weeks, as compared to 16 weeks in low-dose and 90 weeks in

  control female mice.
  TABLE V-9 Comparison of Hepatocellular Carcinoma Incidence
  in Carbon Tetrachloride-Treated Mice Vehicle-Treated Controls


group Hepatocellular carcinoma
Low dose
High dose
Low dose
High dose

Source: NCI (1976)

            The comparative carcinogenicity of carbon tetrachlo-

  ride has been studied in five rat species:  Japanese, Osborne-

  Medel, Wistar, Black, and Sprague-Dawley  (Reuber and Glover

  1970).  Groups of 12-17 male rats of each strain were given

twice weekly subcutaneous injections of carbon tetrachloride

(2,080 rag/kg bw as a 50% solution in corn oil).  Treated animals

were sacrificed when moribund; controls for each strain were

sacrificed at the time as the last experimental animal.  Incidence

of hepatic lesions is given in Table V-10.  The data indicate

that:  (i) sensitivity to carbon tetrachloride-induced neoplasms

varies widely among strains; and (ii) the trends in incidence of

neoplasms and cirrhosis run exactly opposite.  Varying amounts

of toxicity occurred,  all experimental animals of the Black rat

strain were dead at 18 weeks, and those of Sprague-Dawley Strain

at 16 weeks; the failure to find carcinomas in those strains may

have been caused in part by an insufficient latency time.  in all

three other strains, significant toxicity  (i.e., lethality)

occurred.  Toxicity decreased in the same  order as carcinogenicity

increased.  Thus, it appears that there is no causal connection

between the degree of toxicity and  carcinogenicity.

          Other neoplasms were also observed,  all in the Osborne—

Mendel and Japanese strains.  (It is unclear  from the  text whether

these were in experimental or control  animals.)  Heman-giomas of

the  spleen were seen in three rats:  two Japanese and  one Osborne-

Mendel.   Six carcinomas of the thyroid gland  were observed:  Three

in Japanese and three in Osborne-Mendel rats.  Multicystic kidneys

were observed in  two Osborne-Mendel and three Japanese rats.  One

rat  of the Japanese  strain had a  subcutaneous leiomyosarcoma.

          Relative organ weights  were  decreased  for  testes  and

increased for liver, spleen,  and  kidneys  of  all  experimental


 animals' as compared to strain controls.  The extent of atrophy

 of the testes, prostate, and seminal vesicles was correlated

 with the degree of cirrhosis.

           The carcinogenicity of carbon tetrachloride in hams-

 ters has also been described (Delia Porta et al., 1961).  Groups

 of 10 Syrian golden hamsters of each sex were administered weekly

 by gavage 20 mg of carbon tetrachloride (5% solution in corn
         TABLE V-10.   Evidence of the Most Advanced Lesions
              In Rats Administered Carbon Tetrachloride
                 Japanese   Osborne-   Wistar    Black    Sprague-
                              Mendel                     Dawley
No hyperplasia
Hyperplastic nodule
Small carcinoma
Large carcinoma
Total 'Carcinoma
No cirrhosis
Mild cirrhosis
Moderate cirrhosis"
Severe cirrhosis
Adapted from Reuber and Glover (1970)


oil) for 7 weeks, followed by 10 mg for 23 weeks (equivalent

to 200 and 100 mg/kg bw).  Survivors were sacrificed at 55

weeks.  Postnecrotic cirrhosis was observed in all animals that

died in week 41.  Postnecrotic cirrhosis was described by the

authors to involve "regenerative hypexplastic nodules."  In

current experiments, these probably would have been described

as neoplastic nodules in view of the following micropathological

findings:  nodules had obliteration of normal lobular architec-

ture and were surrounded by fibrpus tissues; cells were irregular

in shape and size? nuclei and cytoplasm stained abnormally with

uneven distribution of glycoge^i.  Each of the other 10 animals

had one or more hepatic carcinomas.  A total of 22 neo-plasms

was observed:  12 in five females and 10'in five males.  Some

were sizeable, measuring 5-30 mnu

          Thus,  Syrian golden hamsters appear sensitive to the

carcinogenic effects of carbon tetrachloride.  Although, the

number of animals in this study was small, the authors consider-

ed  the results  to be significant because the reported  control

rate of hepatic  tumors  in hamsters were 0/254.

          Carbon tetrachloride was also reported to be carcino-

genic in C3H mice  (Andervont,  1958).  Groups of 30-77  female or

male C3H mice were  administered by gavage  6.46 mg  of  carbon

tetrachloride once  weekly  for  2 weeks,  followed by administra-

tion of 9.6 mg  once weekly  for 17 weeks  {equivalent of 213  and

320 mg/kg bw).   Pathogen-free  or  normal  C3H mice were used.  For

no  difference in the  incidence of hepatomas was observed between

pathogen-free and normal mice:  79% as compared to 49% in con-

trols^  The average number of hepatoraas per animal was 1.8 in

treated animals and 1.3 in controls.  In females, a difference

between the incidence and hepatomas in pathogen-free and normal

rats was observed:  46% and 29%, respectively as compared to 3%

in controls.  The average number of hepatomas per mouse was 1.5,

1.2, and 1.0, respectively, indicating that both the incidence

as well as the average number of tumors per animal increased in

the order:  controls 

Table V-ll  Susceptibility of Strain A Mice to Liver Necrosis and the Incidence of Hepatones 30 Days
            After 120 or 30 Doses of Carbon Tetrachloride3
Carbon tetrachloride dose15

9600 mg/kg 4800 mg/kg 2400 mgAg 1200 mg/kg (olive oil)

23 > 0) H
•H H
r-j r-H C "j
10 fl) fll
2.31.13 1 2.
ri 50 -H to q q
•c-l p M-l O •" •"
|I|I j i
Sex Dose Conditions "o 

were detected by microscopic examination in two males.  The

authors concluded that repeated liver necrosis and its associated

chronic regenerative state are probably not necessary for the

induction of tumors with carbon tetrachloride.

          A third study also reported the induction of hepato-

mas in mice by exposure to carbon tetrachloride (Edwards, 1941).

Of 143 female C3H mice administered 64 mg of carbon tetrachloride

two or three times weekly for 36-55 doses, 126 or 88.1% developed

hepatomas.  In similar experiments, a 100% incidence of hepatomas

was observed in 54 male and female strain A mice having received

23—58 doses.  The first hepatoma was observed following 23 doses.


          Carbon tetrachloride-induced hepatic effects have been

reported after both acute and chronic exposure.  The degrees of

toxicity and of hepatic damage have appeared to be dose-related,

as measured by liver enzyme activity  in at least four  species:

mouse, rat, guinea pig, and dog.   In parallel acute studies  in

mice and dogs in which dose-response relationships for minimal

toxic effects were developed, mice have appeared more  sensitive

to the toxic effects of carbon tetrachloride  than have dogs.

Liver damage from acute exposure  to carbon tetrachloride  has

been reported to be reversible.   At 36 hours  after acute

exposure to various doses of  carbon tetrachloride, histopatho-

logical examination has shown no  abnormalities  in liver  speci-

mens.  In chronic exposure, liver damage  has  been reported  to


be reversible if the damage did not advance to the necrosis


          The degenerative liver changes observed in both

acute and chronic exposure have been increased serum/plasma

levels of hepatic enzymes, progressing through more pronounced

cellular degenerative changes such as fatty liver and pro-

liferation of rough endoplasmic reticulum, to fatty liver

degeneration and necrosis.  Lung damage has included similar

cellular effects, with the majority of changes occurring

within the Clara cells.  Degenerative kidney effects have

also been observed, but appeared significant only after high

doses of carbon tetrachloride.

          Carbon tetrachloride has produced prenatal toxic

effects, which could not be well correlated with extent of

maternal exposure.  Rats exposed to carbon tetrachloride in

utero have shown hepatic abnormalities at birth, but the

lifetime effects of these changes were not reported.

          Carbon tetrachloride has produced distinct degenera-

tive changes in testicular histology, eventually resulting

in aspermatogenesis and functional male  infertility.  These

effects occurred at medium to high doses.

          Carbon tetrachloride has elicited a mutagenic response

in a Saccharomyces cervisiae test system, but has consistently

tested nagative in the Salmonella  (Ames)  assay.  Investigators

have attributed the failure of carbon tetrachloride to induce


a mutagenic response in Salmonella to the absence of an in

vivo activating system in the prokaryote.  No other mutagenic

responses to carbon tetrachloride have been reported in the


          Carbon tetrachloride has been reported to be carcino-

genic in numerous animal studies.  Hepatocellular carcinomas

have been the neoplasm induced in all species.  Hamsters have

been the most sensitive species studied, followed by mice and

then rats.  A significant strain difference has been observed

in rats.  Females have appeared less sensitive to the chronic

toxic effects and more sensitive to the carcinogenic effects

of carbon tetrachloride in  both rats and mice.


          Considerable human exposure to carbon tetrachlo-

ride through inhalation has come through its use as an

industrial solvent and dry cleaning fluid.  Ingestion of

carbon tetrachloride or mixtures containing carbon

tetrachloride has also been documented in various case

reports.  Ingestion has occurred under different circum-

stances (e.g., suicide attempts, medical use) by persons

of diverse occupations and ages.  These acute exposures

have been followed by hepatoxic effects accompanied by

acute nephrosis.

          In the following section the effects of carbon  tetra-

chloride exposure are presented as reported  in case studies and

in controlled studies for humans.  The case  studies are divided

into reports of  acute and long-term  effects.  The human case

studies are often anecdotal,  with missing or imcomplete medical

descriptions of  clinical  signs  of poisonings.   In  the  controlled

studies using human  volunteers, changes .in  serum and urine

chemistry were measured  after exposure  to carbon tetrachloride,

but no histologic specimens  were  taken.   Because of  the

limitations  in  these studies of humans,  they are presented

as supporting evidence  for  the  harmful  effects  of  carbon

tetrachloride' in humans.


Case Studies-—-Acute Effects

          Ingestion.  Lamson et al. (1928) studied the lethal

effects of carbon tetrachloride in patients receiving carbon

tetrachloride and magnesium sulfate orally as a treatment for

hookworms.  The authors reported the treatment of thousands

of patients with a single dose of 2.5-15 ml of carbon tetra-

chloride without ill effects.  One man was reported to have

safely ingested  40 ml of carbon tetrachloride.  However, an

"extremely small" population of adults died after receiving

1.5 ml of carbon tetrachloride; doses of 0.18-0.92 ml were

reported to be fatal to children.  Susceptibility in adults

was cprrelated with alcoholic  intake  (chronic alcoholism or

exposure to alcohol shortly after treatment), the presence of

ascarid worms, and the intake  of foods, particularly of high

fatty content.

          A fatality attributed to ingestion of  carbon tetra-

chloride was  reported by  Smetana (1939).   The  victim, a photo-

grapher described as having  "a history of chronic alcoholism,"

died 10 days  after consuming an unknown amount of "some  fluid

containing carbon tetrachloride."  He presented  symptoms includ-

ing nausea, vomiting,  jaundice, anuria, and  semistupor.  In  the

final clinical diagnosis,  death was  attributed to carbon tetra-

chloride poisoning.

           A  case of  attempted suicide  by ingestion  of  carbon

tetrachloride was reported by  Stewart et  al.  (1963).  The


victim, a 29-year-old female who ingested 1 pint of a carbon

tetrachloride: methanol solution (2:1), experienced ringing

in the ears immediately after ingestion and lost consciousness.

She was hospitalized for 3 weeks.  Three hours after ingestion,

carbon tetrachloride in the exhaled breath and blood was con-

firmed by infrared analysis.  The exhaled breath was then

monitored throughout the hospitalization, and was reported to

decrease exponentially.  Because of the toxicity of the methanol

and the possibility of synergistic reactions with the carbon

tetrachloride, hemodialysis was performed soon after admission.

Mannitol solution was given by continuous intravenous infusion.

Clinical laboratory analyses during hospitalization showed some

elevation of SCOT, which reached a maximum of 75 units at day 6,

and an elevation of urinary urobilinogen to a maximum of 7.8

Ehrlich units at day 10.  Other  laboratory findings included

elevation of serum iron and depression of serum protein concen-

tration and albumin fractions.   The retention time of bromo-

sulfophthalein was increased.  These  finding were interpreted

as evidence of minimal hepatocellular injury.  Acute renal

dysfunction was not observed; the authors credited the mannitol

treatment with preventing renal  damage.

Acute Effects

          Inhalation.  Bilateral peripheral constriction of  the

ocular color  fields, resulting  in symptoms of toxic amblyopia

in three males, was attributed  to the inhalation of carbon

tetrachloride vapors  (Wirtschafter, 1933).  Five male  employees


of dry cleaning establishments who had been exposed to carbon

tetrachloride (of unknown concentration) from 8-10 hours daily

for 1-6 months were examined.  Two men also had signs' of con-

junctivitis.  Three of the men complained of visual disturbances

characterized by blurred vision or spots before the eyes. The

author concluded that toxic amblyopia may result from exposure

to carbon tetrachloride vapor.

          One fatality occurred in two cases of carbon tetra-

chloride poisoning reported by Smetana  (1939).  In the fatality,

a dry cleaner and interior decorator described as being "a steady

and heavy drinker" was exposed for several hours to carbon tetra-

chloride vapors during work.  Upon returning from work, he noted

dyspnea.  Several hours after the exposure, headache, dizziness,

and malaise developed, accompanied by nausea and repeated vomit-

ing that persisted for several days.  The patient also suffered

labored breathing and cough with bloody sputum before he died 9

9 days following exposure.

          The second inhaltion case  reported by Smetana was  a

housemaid also described as having a history of chronic  alcoho-

lism.  Three days before hospitalization, the patient  cleaned

dresses with carbon tetrachloride  for  3 hours in  a poorly

ventilated  room.  Soon after  exposure,  she  began  to  vomit.   She

suffered symptoms similar  to  those described  for  the other case.

After approximately 1-1/2  months  of  hospitalization,  this

patient was released  from  the hospital her  condition several

weeks later was  described  as  "much improved."


          Seven cases of carbon tetrachloride poisoning reported

by Norwood et al. (1950) resulted from both occupational and non-

occupational inhaltion exposures.  In the three cases described

as "severe" poisonings* there was a history of chronic alcoholism;

two fatalities occurred in this group.  In one case, the victim

had been exposed for about 15 minutes to an atmosphere containing

carbon tetrachloride at an estimated 1,575 mg/m^  (this estimate

was made by duplicating the conditions).  Histopathologic exami-

nation of liver and kidney tissue from the fatalities revealed

liver necrosis and degenaration of the renal tubules.  The four

remaining cases were characterized as "mild industrial"

exposures.  After exposure to carbon tetrachloride, all subjects

suffered varied symptoms including nauseas, vomiting, diarrhea,

headache, muscular ache, pain, or numbness, labored breathing,

and dizziness.

          In another case, a 31-year-old  janitor  suffered ma-

laise, back and lower  abdominal  pain, nausea,  and vomiting the

morning after working  for  5 hours in  a  closed  room with carbon

tetrachloride  (Kittleson and Borden,  1956).  He reportedly

consumed two bottles of beer during the  exposure  period.  The

patient required 2 months  of hospitalization  for  treatment of

acute renal insufficiency  as a  result of  carbon tetrachloride


        Elevated serum glutamic  oxaloacetic  transaminase  (SCOT)

activities with  concomitant  liver  changes were reported  in  two

men occupationally exposed to  unreported concentrations  of

carbon tetrachloride (Lachnit and Pietschmann, 1960).  One
became ill after exposure to carbon tetrachloride for 3 hours
in a relatively well-ventilated room.  He was hospitalized 3
days after exposure.  His liver was slightly enlarged, with
the (SGOT) value elevated by 6,000 units.  This value rapidly
decreased and returned to normal by the 10th day.  A biopsy of
the liver taken on the 8th day showed necrosis in the centers
of the lobuli, but the surrounding tissue was undamaged.  An
additional needle biopsy of the liver taken at the 28th day
showed that the cells had almost returned to normal.  In the
second case a male similarly exposed to carbon tetrachloride
entered the hospital 12 days after exposure.  The SGOT had
increased to 80 units.  A liver needle biopsy on the 22nd day
showed only moderate changes, some of a degenerative nature.

          In a chemical packing plant, use of carbon tetra-
chloride by two workers for equipment cleaning, as a substitue
for the customarily used acetone, resulted in the hospitaliza-
tion of 4 of 43 workers at the plant  (Folland e_t al., 1976).
Ten additional workers also became ill.  Eight of the 43 work-
ers fell ill within 12 hours following the start of  the 2-hour
exposure? six others followed within  the next 36 hours.  The
four hospitalized workers showed evidence of severe  disruption
of liver function:  one case had an  SCOT level of 13,390 units.
All patients recovered within 90 days.  All hospitalized work-
ers as well as most of the others taken  ill had worked  near a
bottle-filling operation for  isopropyl alcohol at the northern
end of the plant, adjacent to the carbodn  tetrachloride cleaning


          Carbon tetrachloride concentrations at the time of expo-
sure were not ascertained; acetone was normally used for cleaning.
Isoporopyl alcohol concentrations at the northern end of the plant
average 410 ppm.  Acetone in alveolar air samples of workers in the
northern area averaged 19 ppm.  The authors described the toxic
episode to carbon tetachloride toxicity potentiated by isopropyl
alcohol.  Because carbon tetrchloride concentrations were unknown
and isopropyl alcohol (and possibly other chemicals) were present,
the health effects reported in this study cannot be attributed
to carbon tetrachloride exposure alone.

Case Studies Long-Term Effects

          Straus  (1954) suggested  a possible causal relationship
between carbon  tetrachloride  exposure and aplastic anemia.  Three
males had been  exposed to  carbon tetrachloride  at unknown concen-
trations for 2  months to 3 years.  Autopsy  findings included hypo-
plasia of the bone marrow.  However,  a  causal  relationship between
carbon tetrachloride and  aplastic  anemia suspected by  the author
in these cases  is not supported adequately.  One of the  men had
also been exposed to kerosene for  3 years.   Another was  an  auto
mechanic who worked in a  garage.   The occupation of the  third
was not  specified although his exposure to  carbon tetrachloride
was occupationally related.   Thus  the effects  of other chemicals
cannot be  discounted.  The autopsy findings of two  of the  patients
included no  liver or kidney  damage of the type that would  be  ex-
pected  in  carbon tetrachloride poisoning.   In  one  case the liver
was reported  to have toxic hepatitis.which was considered  to  be
the result of carbon tetrachloride poisoning.   The information


reported in these case studies tends not to substantiate the

author's suggestion that the patients' illnesses may have been

caused by carbon tetrachloride.

          Carcinogenicity The possibility of carcinogenic

effects of carbon tetrachloride in humans has been raised in a

number of case reports.  These reports do not establish a causal

link between carbon tetrachloride exposure'xand the incidence of

neoplasms (heptomas)^  Thus, the suggestion that carbon tetra-

chloride is carcinogenic in humans remains purely speculative.

          A 59-year-old man with a history of moderate alcohol

consumption returned from a cocktail  party and noticed the vapor

of carbon tetrachloride used to clean a rug in his apartment

earlier that evening.  Five days later he developed nausea, vomit-

ing, and diarrhea and within 10 days  of exposure he developed

jaundice (Tracey and Sherlock, 1968K The patient recovered

following'a long and complex hospitalization and was discharged

after 9 weeks.  Four years  after hospitalization for jaundice,

he was found to have a smooth, enlarged, nontender liver.  He

denied alcohol consumption  within the intervening period.  Three

years after this checkup, the  patient was readmitted with  a

history of nausea, vomiting,  and diarrhea.  A  liver biopsy was

diagnosed as hepatocellular carcinoma.  No treatment was admin-

istered until readmission 5 months  later when  he received  X-ray

radiation dose of 3000 R.   He  died  2  weeks after discharge.

Postmortem examination revealed  the liver to be extensively

involved with the tumor.  Little  normal  liver  tissue remained.


          The connection between carbon tetrachloride exposure/

potentiated by prior alcohol use, and the induction of jaundice

appears well established.  In contrast, as the authors state, no

causal relationship between the carbon tetrachloride exposure and

hepatocellular carcinoma can be drawn from this report.  Aside.

from the acute exposure to carbon tetrachloride 7 years before

diagnosis of cancer, the patient's possible additional exposure

to this and other toxic chemicals -was not reported.  Ho medical

history was given for the 3 years before the final diagnosis.

          A. study of residents of an area surrounding a solvent

recovery plant in rural Maryland found a great increase in the

incidence of lymphatic cancer  (Capurro, 1979).  The mortality

experience of residents of a 1.5 Icm^ area around the plant—which

emitted at least 31 chemicals  (identified by gas chromatography),

including carbon tetrachloride—-was  followed from October 1968 to

October 1974.  Six deaths, one due  to  cancer, were expected over

this period.  Fourteen deaths were  observed, including seven

from cancer.  Four of seven malignancies were lymphomas, more

than 60 times the expected incidence.  These deaths were not

attributed to any particular chemical.

          Other case reports of  human  neoplasms  developing after

exposure to carbon tetrachloride have  appeared.   In  one case a

woman  developed modular  cirrhosis  of the  liver  followed by cancer

of the liver after exposure to  carbon  tetrachloride,  and died 3

years  after the first exposure (Johnstone,  1948).   However,  she

had suffered from periodic  jaundice for  5  years  prior to exposure


to carbon tetrachloride.  In a second case, a fireman developed

cirrhosis and an "epithelioma" of the liver 4 years after acute

carbon tetrachloride intoxication (Sintler et al., 1964).  In

none of the cases could a causal link between carbon tetrachlo-

ride exposure and development of neoplasms be established.

          Because of concomitant exposure to other chemicals

this study does establish a  causative association between carbon

tetrachloride exposure and increased mortality.

          Epidemiology.  A retrospective study  of laundry and dry

cleaning workers was conducted by Blair, ejt a!U  (1979) to determine

if occupational exposure to  carbon  tetrachloride, trichloroethy-

lene, tetrachloroethylene, and petroleum solvents resulted  in

increased morbidity or mortality.   Data for  cases were  obtained

from union records benefits  lists.   Sex, race,  age  at death, and

and underlying and contributing  cause  of death  were abstracted

from death certificates.  The age,  race, sex, and cause distribu-

tion for all deaths in  the United States  from 1957-1970 served

as the  control standard.  Causes of death  were  analyzed using

the proportionate mortality  method.  The  results of analysis

demonstrated an excess  of  lung and  cervical  cancer, and slight

excesses of  leukemia  and  liver cancer.

          Because  of  the  multiple historical exposures  experi-

enced by this  population,  it is difficult to establish  a causal

association  between  specific substances and increased mortality


 trends.  This  study is presented as  evidence of a possible  link

 between  carbon tetrachloride exposure and increased mortality

 rather than as a study in which quantitative extrapolations of

 the chemical's effects on human health are possible.

           A cross-sectional epidemiologic study (Sonich et  al.,

 unpublished) examined health effects of CC14 ingestion in humans.

 Seventy  tons of carbon tetrachloride were spilled in the Kanawha

. and Ohio River in 1977.  Measurements of raw water revealed maxi-

 mum concentration of 0.340 mg/1.  Twenty-one cities situated along

 the river were involved in the study.  These cities represent

 areas that obtained their drinking water directly from the  river

 and/or area that obtained their drinking water from sources not

 influenced by the quality of the river water.  By using river

 volumes  and flow rates/ periods of high exposure  (1977) and low

 exposure (1976) to carbon tetrachloride were estimated for each

 city along the river.  The results' of routine tests measuring

 serura chemistries reflecting liver and kidney function along with

 basic epideraiologic information were abstracted from approximated

 6,000 medical records.  The results obtained for  creatinine show

 a positive and statistically significant  (p<.05)  relationship

 between the carbon tetrachloride exposure  and the frequencly of

 elevated levels of serum  creatinine  in  exposed patients.  No

 similar results were  found  for the other parameters analyzed.

 Controlled Studies

           Inhalation.  Human volunteers were exposed  to  known

 concentrations of  carbon  tetrachloride  vapor in  an effort  to


correlate physiological and/or biochemical changes to the mag-

nitude of exposure  (Stewart et a_l., 1961).  Eight healthy male

volunteers were exposed to carbon tetrachloride vapors in a

series of three separate experiments performed 1 month apart.

Prior to exposure,  data on blood pressure, SCOT, and urinary

urobilinogen were obtained for each subject.  Samples of pre-

exposure exhaled breath, urine, and blood showed no detectable

carbon tetrachloride.  The volunteers were seated in a closed

room  (11 x 12 x 7.5 feet) where 99% pure carbon tetrachloride

was poured into a dish and covered with a towel.  An exhaust

system grill and door were closed during the  experiment but an

air supply grill was left open.   A fan circulated air across

the dish.  Carbon tetrachloride  ambient concentrations were

monitored with a Davis halide meter  and an  infrared  spectra-

meter.  The carbon  tetrachloride  concentration  ranges and

exposure times are  given below in Table VI-1.

TABLE VI-1  Exposure Times and Concentrations of  Carbon  Tetra-
            chloride Vapor  in a Study by  Stewart  et.  al.  (1961)

Average concentration,
time-weighted (mg/m^)
(minutes )



          Carbon tetrachloride was detected in exhaled breath in

all three experiments.  Graphs showed an exponential decrease in

concentration of carbon tetrachloride versus time.   The exact

values were not given.

          The serum iron showed an initial decrease in three of

six subjects at the 309 mg/m3 exposure level but had returned

to normal in two of these  subjects 68 hours after exposure.  The

remaining subject showed a 31% depression in serum iron at 68

hours, but the value was within the normal range.  Serum iron was

not analyzed in the other  two experiments.  Of the six subjects

exposed to carbon tetrachloride at 309 mg/m3, the serum trans-

minase level was slightly  elevated in some and depressed in

others, but remained within  the normal range.  Carbon tetrachlo-

ride was not detected  in the blood or urine at any  exposure  time

or dose, but the analytical  technique used was not  a  sensitive

one.  The authors concluded  that  no  ill  effects  were  observed

from exposure  to carbon tetrachloride at 63 mg/m3  for 180  minutes,

although  the small  changes in  serum  iron at the  309 mg/m3  dose

might have been  an  indication  of  liver  insult.

          Absorption  through Skin.   The absorption of carbon

tetrachloride  through human skin was measured by immersion of

the  thumbs  of  three male  and female  volunteers in a sample of

this  compound  for 30  minutes (Stewart and Dodd,  1964).   The car-

bon  tetrachloride was analyzed by infrared spectroscopy,  and no

 impurities  were detected.   Sequential sensations of burning and

 cooling were experienced by all volunteers during the immersion.


Burning ceased about 10 minutes after removal from the solvent.

The thumbs of all volunteers appeared scaly and red, a condition

that improved within several hours after exposure.  Carbon tetra-

chloride was detected in the alveolar air of each subject within

10 minutes of immersion of their thumbs.  The concentration in

the expired breath rose continuously to a maximum of 4.0 mg/rn-*

10 to 30 minutes after the exposure period ended, and then

decreased exponentially.  The mean concentration of carbon tetra-

chloride was 2.0 mg/m3, 2 hours after the end of exposure; at 5

hours after exposure, the alveolar air concentration was still

greater than 0.6 mg/m^.  The authors concluded that carbon tetra—

chloride could be absorbed through the skin  in toxic quantities.

          Reproductive Effects.  No  teratogenic  effects in humans

caused by carbon tetrachloride  exposure have been reported.  How-

ever, human fetuses in one study appeared to have selectively

accumulated carbon tetrachloride from  the mother's  circulation

 (Dowty et al., 1976).  Maternal blood  samples were  taken  from 11

women either before or directly after  (vaginal)  delivery.   (Prior

exposure of the women to  toxic  chemicals was not reported.)

Paired cord blood samples were  obtained  immediately after de-

 livery.  All volatiles were  analyzed by  gas  chromatography and

mass spectrometry.  Carbon  tetrachloride, benzene,  and chloroform

were present  in higher  concentratation in  cord  blood  as compared

 to maternal blood.



          Hepatic necrosis and renal pathology appear to be

characteristic effects of acute human exposure to carbon

tetrachloride.  If exposure is terminated, the liver shows

regeneration in most cases.  In cases of acute renal dysfunc-

tion, Sidney function returns to normal after exposure to

carbon tetrachloride is terminated and medical treatment is


          The possibility of an association between carbon

tetrachloride exposure and cancer or aplastic anemia is not

substantiated in epidemiological and case  studies.  These

studies are limited in number, and all suffer from the con-

founding factor of exposure to multiple chemicals.


          The toxicity of carbon tetrachloride to an organism

depends upon the ability of the organism to metabolize the com-

pound.  Thus, unmetabolized carbon terachloride does not appear

to be significantly toxic (Rechnagel and Glende, 1973).  In mam-

mals, carbon tetrachloride  is  thought to be metabolized in the

endoplasmic reticulum of the liver by the mixed-function oxidase

system of enzymes.  The reaction sequence proposed  in the litera-

ture for carbon tetrachloride  metabolism was outlined in Section

III, Pharmacokinetics.  Two free radicals have been postulated as

metabolic intermediates:  the  trichloromethyl radical and the

chlorine radical.  The  toxicity of carbon tetrachloride has been

attributed to subsequent reactions of  the trichloromethyl radical,

These  reactions include formation of carbonyl chloride  (phogene),

dimerizatiion to hexachloroethane,  free radical  binding protein,

and  lipid peroxidation.  In this  section each of these  proposed

pathways will be presented  in  conjuction with  the  toxic effects

attributed to it.

  Formation of Cabonyl  Chloride (phosgene)

          From  the results  of  an  ir± vitro study of carbon tetra-

chloride metabolism,  Shah  e_t  al^.  (1979) postulated the  formation

of  carbonyl  chloride from  the  trichloromethyl radical.   The

authors  incubated  L-cysteine  and  f^C]  carbon tetrachloride with

rat liver homogenate and  looked for the formation of 2-oxothio-

zolidine-4-carboxylic acid.  This compound is formed from the

reaction of  L-cysteine and carbonyl chloride.  Analysis of the

metabolic products by mass spectroscopy showed a fragmentation

                             VI I-2

pattern consistent with 2-oxothiozolidine-4-carboxylic acid.
The authors inferred from these analytical data that carbonyl
chloride was formed as a metabolic product of carbon tetrachlo-
ride.  Although carbonyl chloride (phosgene) is not reported
to be a carcinogen, the authors pointed out that the compound
is highly toxic and that the reactive chlorines could react
with macromolecules in ways similar  to alkylating agents.

Dimerization to Hexachloroethane

          Hexachloroethane has been  identified as a metabolite
of carbon tetrachloride by Fowler (1969).  The formation of this
compound is believed to take place by the dimerization of the
trichloromethyl radical.  Although hexachloroethane is a hepato-
toxin, its toxicity is less than that seen in carbon tetrahloride
poisoning.  Therefore, other mechanisms probably account for the
severity of the toxicity associated  with carbon tetrachloride.

Free Radical Binding to Proteins

          Free radical binding to proteins had been postulated as
one cause of toxicity associated with carbon  tetrachloride  (Rech-
nagle and Glended, 1973).  The binding was reported to  involve
reactions with cellular proteins, particularly  those with sulf-
hydryl groups.  Experimental results have  not confirmed  this
theory of carbon  tetrachloride toxicity,  nor is  it  supported
by the pathological changes seen  in  the  liver resulting  from
carbon tetrachloride exposure.   In  one  study,  [14C] carbon
tetrachloride has been observed  to  bind  irreversibly  to rabbit


microsomal proteins at a rate of approximately 20 mole per mg

of protein per hour (Uehleke and Werner, 1975).  Binding of

carbon tetrachloride  (or its metabolites)  to hepatic macromole-

cules was enhanced in the absence of oxygen, consistent with

the proposal that the trichloromethyl  radical is the reactive


          Although it has been  shown that  14C from carbon tetra-

chloride binds to proteins,  the question of carbon tetrachloride

binding to polynucleotides  is  still open.  The question is  impor-

tant  because of  its  implications for the mechanism of  carbon

tetrachloride carcinogenicity  and mutagenicity.  Rocchi et  al.

(1973) examined  the  possible binding of carbon tetrachloride with

nucleic acids  in Wistar  rats and Swiss mice  in vivo,  and with

DNA and polynucleotides  rn  vitro.   !4C-labelled  carbon tetra-

chloride  (367  umol/kg)  binds jjn vivo  to DNA of mouse  liver  and

to ribosomal RNA of  rat  liver if the animals  have  been pretreated

with  3-methylchloanthrene  (MCA).  The  pretreatment with MCA

 increases  the  amount of  the binding due to a higher activity of

 the microsomal system activating carbon tetrachloride.  In_ vitro

carbon  tetrachloride (0.218 umol)  is activated  by  microsomes and

pH 5  enzymes of  MBA-treated animals to a metabolite which  can

 react with DNA and  polynucleotides.  No binding  of carbon  tetra-

 chloride  metabolites to hepatic DNA from control mice or rats  was

detected.   On  the  other hand, Uehleke and Werner (1975) incubated

 [14C] carbon  tetrachloride with either isolated liver microsomes

 (rat  or mouse,  species not  identified) or with soluble RNA, they

observed no [14C] binding to ribosomal RNA or exogenous RNA.
Experimental details were not presented.
          Diaz Gomez and Castro (1980) reported that 1*C from
carbon tetrachloride irreversibly binds in vivo to hepatic
nuclear DNA from strain A/J mice and Sprague-Dawley rats (Table
VII-1).  Also binding of 14C from carbon tetrachloride to DNA
was observed in vitro in incubation mixtures containing micro—
somes and a NADPH generating system as well as in tissue slices
(Table VII-2) .  Liver nuclear proteins (Table VII-3) and lipids
(Table VII-4) irreversibly bind carbon tetrachloride metabolites.
The authors concluded that:  (a) the differences between the
results of Rocchi et al. (1973) (pretreatment with MCA required)
and theirs are possibly related to the use of different strains
of mice (Swiss vs A/J) and rats (Wistar vs Sprague-Dawley);
(b) the interaction of carbon tetrachloride metabolites with DNA
and nuclear proteins could be relevant to carbon tetrachloride
induced liver tumors and hepatoxic effects; and (c) the
epigenetic mechanisms for chemical induction of cancer, not
involving carbon tetrachloride-DNA interactions as shown in
their study, could also be relevant.

Lipid Peroxidation
          A number of the hepatic effects resulting from carbon
tetrachloride exposure,  including the  fatty  liver  syndrome, are
believed to arise as a result of  lipid peroxidation  (Rechnagel
and Glende, 1973).  The mechanism proposed  for  the  peroxidation

                Covalent Binding of 14C from 14CC14
                to Rat and Mouse Liver DNA In Vivo**
              pmol/mg + SD
                                  14C from 14CC11 in DNAb
                                 mol nucleotide/mol of CC14
                                 metabolite  (x
A/J mice
                0.72 + 0.05
                0.52   0.05
a 14CC14(27 mCi/mmol) was administered ip as an olive oil solu-
tion <2§ uCi/ml) at a dose of 10 ml of solution/kg bw.  Animals
were sacrificed 6 hr after 14CC14 and DNA was isolated and coun
ted.  For calculations it was 'considered that 1 mg of DNA con-
tained 3,237 umol of nucleotides.
b Results are the mean of 3 samples.  Each sample was a pool
of 10 livers in the case of mice and one liver for rats.
Values for rats were significantly lower than those of mice

Adapted from Diaz Gomez and Castro (1980)

  TABLE VIII-2     Covalent Binding of 14C from 14CC14 to
                  DNA In Vitro
                            14« From 14CCl/lb
                               DNA         mol nucleotide/mol Of
Experimental               pmol/mg +_ SD      CC14 metabolite

Microsomal activation     1.81 ± 0.13          1.75 X 106

Microsomal activation     4302 £ 300           7.5  X 102
     (33 mM CC14)

Chemical activation        826 + 250           3.84 X 103
aAnaerobic mixtures containing microsomes, NADPH, and mouse liver
 DNA were incubated for 30 min at 370 °C with 10 ul of an ethanol
 solution of I4CC14 (27 mCi/mmol) at a concentration of 13.63
 uCi/ml.  DNA was isolated and counted.  In the second experiment
 on the microsomal activation, the incubation system was as before
 except that cold CC14 was added (final concentration, 33 mM)
 and specific activity was 0.0137 mCi/mmol.  When chemical activa-
 tion was studied DNA/acetyltrimethyl ammonium bromide salts (3 mg)
 were heated for 16 hr at 80 °C in a sealed vial under N? atmosphere
 with 3 ml of an alcoholic solution of 14CC14 (1.82 uCi/ml) in the
 presence of benzoyl peroxide.  DNA was isolated and counted.
     results are the mean of triplicate  simultaneous experiments

 Adapted from Diaz Gomez and Castro  (1980).

   TABLE VI1-3
Covalent Binding of 14C from 14CC14 to Nuclear
Protein Fractions From Rat Livera
Nuclear Protein Fraction
           14C from 1
           in protein10
Percentage total
of label in each
- 7.5
- 10.1
- 5.3
- 28.5
- 56.8
- 63.5
18.25 -
29.73 -
7.76 -
12.14 -
6.55 -
7.13 -
18.44 -

a 14CC14 administration and  time  of  treatment as in Table VII-1.
Isolated nuclear proteins were  dissolved in formic acid and counted.
Fraction I and II correspond to nuclear  sap proteins.   Fraction  III
and IV are deoxuribonucleoproteins,   °	-'— TT n'~ *"** i-sv.™™r-i«»rw.
                   Fraction V is acid ribonucleo-
b  Pooled livers  from  4  rats  were used in each experiment.   These
results are 2 separate experiments .

Adapted from Diaz Gomez  and Castro (1980).

   TABLE VI1-4     Covalent Binding of 14C from 14CC14 to
                   Different. Lipid Fraction From Rat Livera
                            Percentage of total label

Lipid Fraction                   in each fraction*3

Phospholipids                       75.69-65.11

Diglycerides                        16.73-24.07

Cholesterol esters                   0.36 - 0.24

Triglycerides                        0.22 - 0.37

Free fatty acids                     1.19 - 0.64

Cholesterol                          5.70 - 9.50

Undetermined or loss                 0.11 - 0.07

a 14CC14 administration as in Table VII-1. Animals were sacrificed
3 hr after 14CC14 administration.  Total lipid  fractions were used
for counting.  Covalent binding to total nuclear  lipids was 113.5

k Pooled livers from 4 rats were used  in each experiment.  These
results are two separate experiments.

Adapted from Diaz Gomez and Castro  (1980).

                             VI I-9

is presented below, followed by a discussion of the evidence

that it's biochemical sequence results in the hepatic lesions

associated with carbon tetrachloride poisoning.

          The first step in the reaction sequence proposed for

lipid peroxidation is the production of free radicals, especially

the trichloromethyl free radical.  The radical initiates a chain

reaction by reacting with the hydrogen atom of a -CH2~group in an

unsaturated fatty acid, generating a fatty acid free radical.  On

reaction with molecular oxygen, the fatty acid-free radical is

converted into an unstable organic peroxide.  The peroxide disin-

tegrates in two fashions:  (i) intramolecular cyclization to form

malonic dialdehyde and two new free radicals, or  (ii) simple

homolytic fission that also yields two free radicals.  This whole

process occurs autocatalytically:  each free radical gives rise to

two new free radicals.  Figure "VII-1 summarizes this hypothesis

(Rechnagel and Glende, 1973).

          A number of indicators have been used in  in vivo and in

vitro assays of lipid peroxidation:  pentane and  ethane  levels in

exhaled air (arising from fatty acid decomposition) and-  malonic

dialdehyde concentrations in hepatocytes  (arising from intramolec-

ular cyclization).  Pentane production in male rats increased by

factors of 4.6,. 13.2, and 26.4 over that  in mineral oil  controls

within 30 minutes  following intraperitoneal administration of

carbon tetrachloride doses of  160, 430, and 1,440 mg/kg  bw,

respectively (Sagai and Tappel,  1979).


          A mechanism for the pathogenesis of^carbon tetrachlo—

rideinduced hepatic lesions based on lipid peroxidation has been

proposed (Pasquali-Ronchetti e_t ad., 1980).  According to this

hypothesis, lipid peroxidation  is suggested  to affect primarily

unsaturated acyl chains of membrane phospholipids, resulting in

breakage of the hydrocarbon and loss of phospholipids from the

membrane.  Lipid peroxidation would therefore produce progressive

degenerative changes in the assembly of membranous structures

such as microsomes and of (rat) liver endoplasmic reticulum.

          This hypothesis is supported by studies showing that

treatment with carbon tetrachloride produced lipid peroxidation

in rat liver endoplasmic reticulum  at a concentration of 0.5

ml/100 g bw (Pasquali-Ronchetti et  a_l., 1980); caused disinte-

gration of endoplasmic reticulum  in vitro within 10 minutes at

a concentration of 636 mg/liter (Pasquali-Ronchetti et al.,

1980); and was incorporated predominantly into liver phospho-

lipids in rats (Table VII-5) (Ciccoli e_t al_., 1978).

          Two pieces of contrary  evidence have been presented by

Diaz Gomez, e_t al. (1975).  One was that the order of species

susceptibility to liver necrosis  from carbon tetrachloride more

closely parallels the species order for  [14C] carbon tetrachlo-

ride binding to cellular components than the species order for

lipid peroxidation:

                  H        H       H
            c=c-c- c=c-c- c=o-oc=c-
                  H        H       H
                              HCC1-  »CCi-Trichisraeehyl Fres
                                      J        Sadical
            R2S03ANCZ (All   / * Organic Frse Radical
            Possible Foras Not

            J  ft  *    * a  fi   .
          - c=c-c-c-oc-c=c-c-c=c-
            Peroxide    v       *N*
            formation  OjT   diene coajuge.ian) ^_ »233cu
                      V    Organic Psroxide (Uasrabla)
    Inrraaolectilar cyclizaclon     Decanpasitioa co yield cvo free

    and deconposirion to yield     radicals. Evericuai stable decan-

    nalonic dialdehyde and cvo     posicion products highly organo-

    new organic free radicals.     leptic.
Figure VII-1.  Free Radical Initiated, Autocatalytic
             Peroxidation of Polyenoic Long-Chain
             Fatty Acids
             (Adapted from Recknagel and Glende 1973).


 Liver necrosis      mouse> guinea pig.= hamster> rat> chicken

 [  C]CC1^ binding   mouse = hamster> guinea pig> chicken = mouse

 Lipid peroxidation  rat> hamster = guinea pig> chicken = mouse

           A second result of their experiments was that for mice,

 necrosis proceeded for 24 hours in the absence of lipid peroxida-

 tion.  Together, these pieces of evidence imply that caution is

 required in accepting the lipid peroxidation mechanism for carbon

 tetrachloride toxicity.


           The metabolism of carbon tetrachloride is thought to

 proceed through the formation of the trichloromethyl free radi-

 cal.  The hepatotoxicity of carbon tetrachloride has been attri-

 buted to subsequent lipid peroxidation initiated by this radical

 in the following manner:  a hydrogen atom is abstracted by the

 free trichloromethly radical  from a long-chain  fatty acid to form

 chloroform and a. fatty acid-free radical.  Molecular oxygen,

 because of its triplet ground state, binds with the unparied

 electron on the fatty acid radical to form an organic peroxide.

 The peroxide is unstable and  decomposes to form more organic-

 free radicals, which in turn  form more organic  peroxides.  This

 process appears to lead to fatty acid chain decomposition, with

 the resulting breakdown of membrane structure.  This breakdown

 may lead to a halt in lipid excretion via the Golgi  apparatus,

 with fatty liver occurring as a consequence.  Cell necrosis would

 also follow directly from lipid destruction.  The mechanism by

which lipid peroxidation could lead to cell transformation is
not explained yet, and the molecular events leading to carbon
tetrachloride careinogenicity remain unknown.

          In addition to this proposed lipid peroxidation
mechanism, which produces chloroform, two minor metabolic path-
ways have been; postulated:  dimerization to two trichloromethyl
free radicals to form hexachloroethane and the formation of a
trichloromethyl peroxy radical which may result in production
of phosgene 'and carbon dioxide.  Both hexachloroethane and
phosgene are toxic, but the extent of their contribution to
observed hepatotoxicity is unknown.   A fourth possible
mechanism of hepatotoxicity is the binding of trichloromethyl
radical to cellular proteins.

                  VIII.  RISK ASSESSMENT

          The possibility that carbon tetrachloride is carcino-

genic in humans has been a subject of concern, and has resulted

in the development of several human risk assessments.  This

section of the report summarizes carcinogenic risk assessments

prepared by the National Academy of Sciences  (NAS) and the U.S.

Environmental Protection Agency (EPA).

          Current methods used to estimate carcinogenic risk

have in common the assumption that there is no threshold level

for the action of a carcinogen.  The  state-of-the-art and data

now available are such  that  no one method can accurarely predict

and/or model the absolute numbers of  excess cancer deaths attri-

butable to carbon tetrachloride in drinking water.  Because of

biological variability  and  the assumptions used,  each of the

methods used to quantify carcinogenic risk leads  to a different

value.  The estimates  and their interpretation may vary widely.

In addition, none of the method's  now  used  to  quantify carcino-

genic  risk can account for  the  increased  risk of  carbon  tetra-

chloride exposure to sensitive populations.

Quantification of Carcinogenic Risk

          Because of positive results in  animal  carcinogenicity

studies, carbon  tetrachloride can be  considered  a suspect  human

carcinogen.  Data from these animal  studies  have been used by

the NAS and  EPA's Carcinogen Assessment Group (CAG)  to calcu-

late  the number  of  additional cancer cases that  may occur  when


carbon tetrachloride is consumed in drinking water over a 70-

year lifetime.  The results of these calculations are shown in

Table VIII-1.

           The criteria for the CAG (USEPA 1980c, 1983) and NAS

(1977) risk calculations differ in several respects: (1) NAS

used the multistage model, while CAG used an "improved" multi-

stage model.   (2) NAS used the data set from the National Cancer

Institute  (NCI) negative study in male rats while CAG used the

data set from  NCI's positive  study in male mice.  Because of

these differences, the carbon tetrachloride levels reported by

CAG for cancer risk levels are approximately 1/10th those

reported by NAS.

           The U.S. Environmental Protection Agency has  also

developed  Ambient Water Quality Criteria  based  on estimates

of the increased lifetime cancer  risk resulting from a  life-

time consumption of both drinking water  (2  liters per  day) and

aquatic life  (6.5 g of fish  and shellfish per day).  These risk

estimates  differ from  those  in  the previous risk assessments,

which are  based on consumption  of drinking  water alone.   These

criteria  for  lifetime  cancer risks of  10~5,  and 10~6,  and 10~7

are 4.0,  0.40, and 0.04  ug/liter,  respectively  (USEPA  1980a).

Sensitive  Populations

           Sensitive populations are  subgroups  within the general

population which appear  at  higher than average  risk upon exposure

to carbon  tetrachloride.   Some  of the  populations  that may be

Table VIII-1 of Additional Carcinogenic Risk
Following Exposure of Humans to Carbon Tetra-
chloride in Drinking Watera
                           Carbon tetrachloride concentrations
 Excess cancer
    (upper  95%
(upper 95%
(point estimate)
(95%  confidence

aAn average  daily  drinking water consumption of 2  liters  per day
was assumed.

at greater risk  include human fetuses,  alcohol consumers,  and

males  of  reproductive age.  This section will deal with the

possible  effects of age,  sex, and nutritional status on toxicity

of carbon tetrachloride.

          The studies of Reuber and Glover (1968)  with rats

suggest that sensitivity to the effects of carbon tetrachloride

may vary  with age  and sex.  In these studies, inbred Buffalo

rats 4, 12,  24,  or 52 weeks of age were injected subcutaneously

twice  weekly for 12 weeks with carbon tetrachloride (1000 mg/kg

bw  in  corn oil).  Cirrhosis of the liver following exposure

increased with age in male rats, whereas female sensitivity


increased up to 24 weeks and declined thereafter.  Female

sensitivity exceeded male sensitivity in the 4- and 12-week

groups; this pattern reversed after 24 weeks.

          Nutritional status may also affect the degree of

toxicity following exposure to carbon tetrachloride in rats.

Gyorgy et al.  (1946) exposed young rats on various diets to

approximately  300 ppm of carbon tetrachloride  in a gas chamber

7 hours per day, 5 days per week for 5 months.  Animals were

then sacrificed and histopathological effects  on the liver and

kidneys were determined.  Compared to these  signs of toxicity

in animals fed standard chow,  these effects  were more severe

in animals fed a diet high  in  lipid and low  in carbohydrate,

or a diet low  in protein.  Methionine appeared to protect

against the  increased toxicity (particularly kidney damage)

caused by low-protein diets.

Interaction  of Carbon Tetrachloride with  Other Chemicals

          The  interaction  of  carbon  tetrachloride with  certain

chemicals has  resulted  in  an  enhancement  of  the toxic effects

produced  in  animals  by  either chemical  alone.   Exposure of

animals  to certain  environmental  carcinogens in combination with

carbon tetrachloride has  resulted in an increase in carcinogenic

efficacy.  In. addition,  certain chemicals appear to increase the

toxic  effects  of  carbon tetrachloride on the liver and other

organs of  experimental  animals.  There  is also clinical evidence

that two of  these chemicals,  isopropanol and ethanol,  may poten-

tiate  carbon tetcachloride toxicity in humans  (see Section VII).


       Carcinogenic Effects.   The interaction of carbon tetra-

chloride with several carcinogens has been studied.  The effects

of carbon tetrachloride on dimethylnitrosamine-induced carcino-

genicity were studied in male Sprague-Dawley rats treated with a

single dose of carbon tetrachloride (4,000 mg/kg bw) by gavage

42 or 60 hours prior to a single intraperitoneal dose of dimethyl-

nitrosamine (20 or 40 mg/kg bw).  This treatment resulted in a

greater than additive increase in incidence of tumors or tumor-

like lesions of the liver and kidney at 12 months as compared to

rats treated with carbon tetrachloride or dimethylnitrosaraine

along (Pound e_t al.r 1973).  If the pretreatment took place more

than 60 hours before injection of dimethylnitrosamine, the

incidence of kidney neoplasms decreased, while that of liver

neoplasms increased further.

          Similar results were- obtained with N-butylnitrosurea

(Takizawa et al., 1975).  Experimental male ICR/JCL mice were

treated with 80 mg of carbon tetrachloride subcutaneously 1 day

before administration of 10 or 20 mg of N-butylnitrosurea in 50%

ethanol by gavage.  This treatment  resulted in the  induction of

hepatomas after 15 months  in 12 of  23^mTce"(dose groups  combined)

as compared to 1 of 18 mice given N-butylnitrosurea but  not car-

bon  tetrachloride.  However, since  the ethanol vehicle was not

given to all'controls receiving carbon tetrachloride  alone, the

possible ethanol potentiation of  carbon  tetrachloride effects

could not be fully evaluated  (see  the  following  discussion of

ethanol effects on carbon  tetrachloride  toxicity).


          The effects of carbon tetrachloride on the carcinogeni-

city of 2,7-bis-(acetoamido)fluorene (2,7-AAF) were also studied.

Carbon tetrachloride (1,4000 mg in corn oil) was given by gavage

once weekly for 8 weeks to male SMA/Ms mice fed chow containing

0.025% of 2,7-AAF.  A greater than additive effect on the incidence

of hepatomas was observed in these animals as compared to controls,

which received carbon tetrachloride or 2,7-AAF for 8 weeks, or

carbon tetrachloride for one 8-week period and 2,7-AAF for another

8-233k period (Kozuka and Sassa, 1976).

          Toxicologic Effects.   Many chemicals have been reported

to potentiate noncarcinogenic effects of carbon tetrachloride.

Hewitt et al. (1980"t, in their review of studies on potentiation

of hepatotoxic effects of carbon tetrachloride by various agents,

have postulated that ketones (e.g., acetone, methyl ethyl ketone)

or chemicals that can be metabolized to ketones (e.g., isopropa-

nol, 2-butanol, 1,3-butanediol) potentiate  the effects of carbon

tetrachloride and other haloalkanes.

          Traiger and Plaa  (1971) administered isopropanol  (2,000

mg/kg bw by gavage) to male Sprague-Dawley  rats 18 hours before

intraperitoneal injection with carbon tetrachloride  (160 mg/kg

bw in corn oil).  The SGPT activity of these  animals was 22 times

higher than that in controls receiving carbon tetrachloride alone.

In addition, SGPT activities in animals  receiving  isopropanol and

carbon tetrachloride at 160 mg/kg bw were higher than  those in

animals receiving carbon tetrachloride at  1,600 mg/kg  bw without



          Later studies on isopropanol potentiation (reviewed by
Hewitt et al., 1980) using an inhibitor of isopropanol metabolism
demonstrated that potentiation probably depended on the metabolism
of isopropanol to acetone.  When the blood concentration of iso-
propanol was kept high and that of acetone was low, the potenti-
ating capacity of isopropanol was significantly reduced.  Acetone
administered directly (2,000 mg/kg bw) potentiated the hepato-
toxic effects of carbon tetrachloride (160 mg/kg bw) given 18
hours later.  Therefore, acetone rather than isopropanol appeared
to be responsible for the potentiation observed.

          Isopropanol was also implicated as the potentiating
factor in carbon tetrachloride toxicity (renal and hepatic)
in humans after accidental exposure to carbon tetrachloride
in an isopropanol packaging plan  (Folland et al., 1976).  Ele-
vated levels of acetone were found in samples of the expired
air of these workers.

          Traiger and Bruckner (1976) studied the effects of 2-
butanol and  its ketone metabolite 2-butone  (methyl ethyl ketone)
on the potentiation of carbon tetrachloride hepatotoxicity.
2-Butanol (1,800 mg/kg bw) or 2-butanone  (1,500  mg/kg  bw) were
administered orally to male Sprague-Dawley  rats  16 hours prior
to intraperitoneal  injection of  carbon tetrachloride  (160 mg/kg
bw in corn oil) .  SGPT activity  24 hours  after  carbon  tetrachlo-
ride exposure was 30-fold higher in both  groups than  in controls
given carbon  tetrachloride alone.  Glucose-6-phosphatase acti-
vity decreased  and  triglyceride  levels  increasd in experimental


animals as compared to controls.  2-Butanol had an apparent

oral half-life of 2.5 hours; by the time carbon tetrachloride

was administered, most of the 2-butanol present had been

metabolized to 2-butanone or eliminated.  The dose of 2-butanone

used approximated the level of 2-butanone in the blood after

administration of 2-butanol.  Because the effect of both treat-

ments were similar, the authors proposed that 2-butanone was

responsible for carbon tetrachloride potentiation in both

treatments.  Therefore, a similar mechanism was implicated

for 2-butanol as for  isopropanoli  metabolism to the ketone.

          Administration of 1,3-butanediol to rats has been

associated with a rapid rise  in the blood concentration of ketone

bodies.  This compound was  therefore used to test the hypothesis

that matabolic ketosis results  in an increase in carbon tetra-

chloride hepatotoxicity (Hewitt and Plaa, 1979).  Male Sprague-

Dawley rats received  1,3-butariediol (5  g/kg bw) three times daily

for 3 days.  On day 3, the  rats received an intraperiotoneal dose

of carbon tetrachloride (160  mg/kg bw  in corn oil), and were

sacrificed 24 hours later.  SGPT  activity  in experimental animals

incrased 18-fold over that  in controls  receiving carbon tetra-

chloride alone, while hepatic glucose-6-phosphatase activity

in experimental animals decreased compared  to controls.  These

results, therefore, also  implicated ketone's  in the potenti-

ation of carbon  tetrachloride toxicity.


          Several other chemicals that potentiate carbon tetra-

chloride toxicity, although complex in structure, also contain

ketone groups (i.e., phenobarbital, pentobarbital, triamcinolone,

progesterone, and Kepone).  Phenobarbital has been demonstrated

to markedly enhance carbon tetrachloride liver damage in mice and

rats.  Cans et al. (1976) detected an increase in liver weight,

protein synthesis, and DMA synthesis in male Swiss mice given both

phenobarbital and carbon tetrachloride as compared to those given

either compound alone.  Tuchweber and Kovacs (1971) found that

pretreatment of female ARS/Sprague-Dawley rats with phenobarbital

resulted in increased mortality, liver damage, and triglyceride

accumulation after carbon tetrachloride exposure as compared to

animals given carbon tetrachloride alone.  Pani et al.  (1973)

also demonstrated an increase  in mortality of male Wistar rats

given phenobarbital in conjunction with carbon tetrachloride as

compared to those given carbon  tetrachloride alone.   In addition,

serum enzyme activities were increased in animals given both

phenobarbital and carbon tetrachloride compared to those given

only carbon tetrachloride.  Lindstrom and Anders  (1978) detected

an increase in diene conjugation in hepatic microsomal  lipids  in

male Sprague-Dawley rats after  administration of phenobarbital

and carbon tetrachloride compared  to  that  in controls receiving

carbon tetrachloride or phenobarbital alone.  In  further studies,

isolated rat hepatocytes were  prepared from male  Sprague-Dawley

rats that had been treated with phenobarbital; control  hepatocytes

were prepared from untreated rats  (Lindstrom et  al.,  1978).   In

cultures containing carbon tetrachloride,  release of  lactic


dehydrogenase was potentiated in hepatocytes from treated animals

compared to controls.  Chang-Tsui and Ho (1980), in an experiment

with pentobarbital, demonstrated potentiation of carbon tetrachlo-

ride toxicity in male ICR mice with increases in SGPT and SGOT

following carbon tetrachloride exposure.

         Triamcinolone and progesterone have also been shown to

potentiate carbon tetrachloride hepatotoxicity  (Tuchweber and

Kovacs 1971)»  Female ARS/Sprague-Dawley rats were treated orally

twice daily with 5 mg of progesterone or 2 mg of triamcinolone.

After 4 days, the animals received carbon tetrachloride at either

16 or 4,000 ing/kg bw.  These treatments resulted in increases in

mortality, liver damage, and triglyceride accumulation in compar-

ison to controls given only carbon tetrachloride.

          Another Tcetone, Kepone, also  appears  to potentiate the

hepatotoxic effects  of carbon tetrachloride  (Curtis et a_l., 1979).

After being fed chow containing 0-10 ppm of  Kepone for 15 days,

male Sprague-Dawley  rats received a single intraperitoneal dose

of carbon tetrachloride  (0-320 mg/kg bw in corn oil).  Compared

to controls given  carbon tetrachloride  but no Kepone, Kepone-fed

animals showed increased impairment of  biliary  excretion at carbon

tetrachloride doses  of 80 mg/kg bw or above, elevated SGPT and

SGOT  (carbon tetrachloride  doses  of 160 mg/kg bw  and above), and

morphological changes in the  (carbon  tetrachloride doses of SO

mg/kg bw and higher).  Some animals in  these studies were  also

exposed to pentobarbital during  anesthesia  and  possibly  to  acetone

residue in the food, since  acetone was  used  as  a  vehicle  for


Kepone.  The possible effects of these exposures on the results

could not be determined.  Hewitt et_ _al. (1980) reported that

Kepone potentiated the toxic effects of chloroform, a compound

similar to carbon tetrachloride (i.e., 1 a haloalkane3.

          In addition to the alcohols previously discussed,

ethanol and methanol have been shown to potentiate the effects

of carbon tetrachloride.  In one study, hepatotoxic effects of

carbon tetrachloride were potentiated by ethanol and methanol in

male Swiss-Webster mice and by ethanol in male Sprague-Dawley

rats (Traiger and Plaa, 1971).  SGPT activity was  increased in

mice exposed to both ethanol and carbon tetrachloride compared

to controls receiving carbon tetrachloride  alone.  Rats treated

with both ethanol and carbon tetrachloride  also showed increased

levels of hepatic triglycerides and  decreased activity of  hepatic

glucose-6-phosphatase.  Cornish and  Adefuin (1966) found that a

single oral dose of ethanol followed by exposure to  carbon tetra-

chloride vapors resulted  in increased  levels  of SGPT,  SCOT, and

serum  isocitric dehydrogenase  in male  Sprague-Dawley rats. Con-

trol animals receiving  carbon  tetrachloride or ethanol alone had

lower  levels of these enzymes, although some  effect  occurred with

carbon tetrachloride alone.  Maling,  et a_l. (1975) also  found an

increase in SGPT activity in male  Sprague-Dawley rats  treated both

with ethanol and carbon tetrachloride  compared to  rats treated

with carbon tetrachloride alone.   Increases were also  noted in

liver  triglycerides, hepatic necrosis,  and the  covalent  binding

of  carbon  tetrachloride to liver protein  and lipid.


          Strubelt et al. (1978) also reported increases in

serum enzyme activities and changes in liver morphology in male

Wistar rats exposed to ethanol plus carbon tetrachloride compared

to controls given either chemical alone.  Cantilena et al. (1979)

demonstrated the methanol potentiation of carbon tetrachloride

hepatoxicity in male Sprague—Darley rats.  Plasma alanine amino-

transferase activity and hepatic triglyceride levels were in-

creased and hepatic glucose-6-phosphotase activity was decreased

compared to controls given carbon tetrachloride alone.

          Ethanol and methanol have been judged to be less effec-

tive than isopropanol in potentiating the toxic effects of carbon

tetrachloride in mice and rats  (Traiger and Plaa, 1971) .  In

humans, the effects of potentiation of ethanol and methanol on

carbon tetrachloride toxicity have not been quantified.  Never-

theless, case reports appear to indicate ethanol potentiating

effects in humans qualitatively similar to those in animals.

          Another class of chemicals which have been reported to

interact with carbon tetrachloride to produce toxic effects  is

polychlorinated biphenyls.  Carlson (1975) reported the potenti-

ation of carbon tetrachloride hepatotoxicity in male albino  rats

by the polychlorinated biphenyls Arochlor 1254, 1221, and 1260.

The animals received intraperitoneal injections of one of these

compounds in corn oil for 6 days.  Twenty-four hours after the

last injection, the rats were exposed to carbon tetrachloride

vapors (3,700-26,400 mg/m3; 590-4,200 ppm) for 2 hours.   Changes


were demonstrated in various serum and liver enzyme activities

after these treatments as compared to activities after carbon

tetrachloride treatment or Arochlor treatment alone.


          Carbon tetrachloride is a known animal carcinogen and

a suspect human carcinogen.  The risk of cancer from human expo-

sure to carbon tetrachloride in drinking water has been estimated

by the NAS and the CAG.  These quantification efforts differed

in methodology and in animal data. used.  An excess cancer risk

of 10~5 over a 70-year lifetime would result from exposure to

drinking water containing tetrachloride at 4.2 ug/liter in the

CAG estimate or 45 ug/liter in the NAS estimate.

          In addition to the quantitative estimation of risk, two

factors that can be considered in evaluating risk of exposure to

a chemical are the variation in sensitivity among populations and

the interaction of the chemical with others.  In the case of

carbon tetrachloride, studies in rats have suggested that young

females (less than 12 weeks old) are more sensitive to the

chemical's cirrhotic effects, with the sensitivity pattern revers-

ing at older ages.   Furthermore, dietary changes  (high-lipid/

low-carbohydrate diets, or  low protein diets) were  reported  to

render rats more sensitive  to liver and kidney  damage  following

carbon tetrachloride exposure.  These conclusions  can  be only


          Greater than additive toxicity  resulting from  exposure

to carbon tetrachloride concomitantly with other  chemicals has


been reported in many animal studies.  The interactions have
been of two forms.

          1) Enhanced carcinogencity when carbon tetrachloride
was given before a carcinogen.  Greater than additive numbers
of liver and kidney tumors were seen when dimethylnitrosamine,
n-butylurea, or 2-AAF was given after carbon tetrachloride.
          2)  Enhanced hepatotoxic effects when carbon tetra-
chloride was given along with a second chemical/ usually a
ketone or an alcohol that is metabolized to a ketone.  Poten-
tiating effects on carbon tetrachloride toxicity have been
best documented for isopropanol and butanol.  Kinetic and
metabolic inhibition studies have been consistent with the
hypothesis that these alcohols must be metabolized to ketones
(acetone and methyl ethyl ketone, respectively) before they
exert their potentiating effects.  Among the other alcohols
reported to potentiate carbon tetrachloride toxicity are 1,3-
butanediol (which is associated with a rapid rise in ketone
bodies), ethanol, and methanol.   In addition, potentiating
effects have been reported  for several chemicals with ketone
groups  (phenobarbital, pentobarbital, triamcinolone,
progesterone, and Kepone),  and for several polychlorinated


     The quantification of toxicological effects of a chemical

consists of an assessment of the noncarcinogenic and carcinogenic

effects.  In the quantification of noncarcinogenic effects,

an Acceptable Daily Intake (ADI) is calculated.  From this an

Adjusted Acceptable Daily Intake (AADI) and Health Advisory

(HA) values for the chemical are calculated to define the

appropriate drinking water concentrations to limit human

exposure.  For ingestion data,  this approach is illustrated

as follows:
     ADI = (NOAEL or LOAEL in mg/kg/day) (Body weight in kg) = mg/day
                      Uncertainty/Safety factor

     AADI »  	ADI	 = mg/L
             Drinking water volume  in L/day


     NOAEL = no-observed-adverse-effeet level.

     LOAEL = lowest-obvserved-adverse-effeet level.

     Body weight = 70 kg for adult  or 10 kg for child.

     Drinking Water volume = 2  L per day for adults or  1 L
                             per day for children.

     Uncertainty/Safety  factor  = 10, 100 or 1,000.

     Utilizing these equations, the following drinking  water

concentrations are developed for noncarcinogenic  effects:


     1.  A one-day HA for 10-kg child.

     2.  A one-day HA for 70-kg adult.

     3.  A ten-day HA for 10-kg child.

     4.  A ten-day HA for 70-kg adult.

     5.  A lifetime AADI for a 70-kg adult.

     The distinctions made between the HA calculations (items

1 through 4) are associated with the duration of anticipated

exposure.  Items 1 and 2 assume a single acute exposure to

the chemical.   Items 3 and 4 assume a limited period of

exposure (possibly 1 to 2 weeks).  The HA values will not be

used in  establishing a drinking water standard for the

chemical.  Rather, they will be used as informal scientific

guidance to municipalities and other organizations when

emergency spills or contamination situations occur.  The AADI

value (item 5} is intended to provide the scientific basis

for establishing a drinking water standard based upon

noncarcinogenic effects.

     A NOAEL or LOAEL is determined from animal toxicity data

or human effects data.  For animal data, this level  is divided

by an uncertainty factor because there is no universally

acceptable quantitative method to extrapolate from animals to

humans.   The possibility must be- considered that humans are

more sensitive to the toxic effects of chemicals than are


animals.   For human data, an uncertainty factor is also used

to account for the heterogeneity of the human population in

which persons exhibit differing sensitivity to toxic chemicals.

A modification of the guidelines set forth by the National

Academy of Sciences (NAS 1977, 1980) are used in establishing

uncertainty factors as follows:

o An uncertainty factor of 10  is used when good acute- or

  chronic human exposure data  are available and supported by

  acute or chronic toxicity data in other species.

o An uncertainty factor of 100 is used when good acute or

  chronic toxicity data  identifying NOEL/NOAEL are available

  for one or more species, but human data are not available.

o An uncertainty factor of 1,000 is used when limited or

  incomplete acute or chronic  toxicity data  in all species  are

  available or when the  acute  or chronic  toxicity data  identify

  a LOAEL (but not NOEL/NOAEL) for one or more species, but

  human data are not available.

     If toxicological evidence requires the  chemical  to be

classified as a potential carcinogen  (i.e.,  carbon  tetrachloride),

mathematical models are  used  to  calculate  the estimated excess

cancer risks associated  with  the ingestion of the chemical

via drinking water.  The bioassay  data  used  in  these  estimates

are from  animal experiments.   In order  to predict the risk


for humans from these data, it must be converted to an

equivalent human dose.  This conversion includes correction

for non-continuous animal feeding, non-lifetime studies and

for the difference in size.  The factor that compensates for

the size difference  is the cube root of the ratio of the

animal and human body weights.  It is assumed that the average

human body weight  is  70 kg and that the average human consumes

2 liters of water  per day.  The multistage model is then fit

to the equivalent  human data  to estimate  the risk at low

doses.  The upper  95% confidence limit of this estimate is

used.  Excess cancer  risks can also be estimated using other

models such as the one-hit model,  the Weibull model, the

logit model and the  probit model.  There  is no basis in the

current understanding of  the  biological mechanisms  involved

in cancer to choose  among these models.   The esitmates of

low doses for these models can differ  by  several orders of

magnitude.  The multistage model  does  not necessarily give

the highest or lowest risk estimates  at  low doses.  Whether-'

it is the most conservative,  least conservative  or  predicts a

risk  in the middle of the range of risks  predicted  by other

'models  is chemical specific.  The  multistage model  is used

because CAG and NAS  use  it.

      The scientific  data  base used to  calculate  and support

the setting of risk  rate  levels  has  an inherent  uncertainty.

This  is because the  tools of  scientific  measurement,  by  their


very nature,- involve both systematic and random error.  In

most cases, only studies using experimental animals have been

performed.  There is thus uncertainty when the data are

extrapolated to humans.  When developing risk rate levels,

several other areas of uncertainty exist, such as  (1) incomplete

knowledge concerning the health effects of contaminants

in drinking water,  (2) the impact of test animal age, sex and

species and the nature of target organ systems examined on

the toxicity study  results and  (3) the actual rate of exposure

of internal targets in test animals or humans.  Dose-response

data are usually only  available  for high  levels of exposure,

not for the lower levels of exposure for  which a standard is

being set.  When there  is exposure to  more  than one  contaminant,

additional uncertainty results  from a  lack  of information

about possible  synergistic or  antagonistic  effects.

A.  Noncarcinogenic Effects

     Varying degrees of  carbon  tetrachloride  (CC14)-induced

toxicity have been  reported  in  humans  and animals  following  acute

and chronic exposures  via  ingestion,  inhalation or dermal admini-

stration.  The  following paragraphs  discuss the  results of  perti-

nent studies to be  considered  for  the  derivation  of  the RMCL at

a  later date.


     Effects of acute exposure to low levels of CCl$ in rats

were reported by Korsrud ejt a_l. (1972).  Male rats (260-400 g)

were administreed single oral doses of CC14 (0 to 4,000 mg/kg bw)

in corn oil.  The rats were fasted for 6 hours before dosing and

for 18 hours afterward, and then sacrificed.  Assays included

liver weight and fat content, serum urea and arginine levels, and

levels of nine serum enzymes, produced mainly in the liver.  At

20 mg/kg bw, there was histopathologic evidence of toxic effects

on the liver.  At 40 mg/kg bw, liver  fat,  liver weight, serum

urea, serum arginine, and  levels of six of  the nine liver enzymes

were increased.  At higher doses the  remaining three enzyme  levels

were also elevated.  The histologic changes seen at the minimum

effect level, 20 mg/kg bw, included a loss  of basophilic stippling,

a few swollen cells, and minimal cytoplasmic vacuolation.  This

study was used by EPA's Office -of  Drinking Water to derive the

existing one-day and ten-day  health advisories for CC14  (USEPA,


     Murphy and Malley  (1969)  investigated the effects  of

single oral doses of CC14  on  the  corticosterone-inducible  liver

enzymes, tyrosine-«c-ketoglutarate transaminase,  alkaline  phos-

phatase, and  tryptophan  pyrrolase  in  rats.  Specifically,  groups

of  4-7 male rats were  administered by gavage  400,  800,  1600, 2400,

or  3200 mg/kg  undiluted  CC14.   Single doses of  400  mg/kg or  greater


of CC14 increased liver tyrosine-^c-ketoglutarate transaminase

acid and alkaline phosphatase, but not tryptophan pyrrolase

activity within 5 hours.  The National Academy of Sciences

(NAS) used this experiment to calculate one-day and seven-day

suggested no-adverse-response levels  (SNARLs)  for CC14

(NAS, 1980).

     In a chronic oral exposure study (Alumot et al., 1976),

groups of 36 rats (18 males and  18 females) were fed

mash containing CC14 at 0, 80, or 200 mg/kg of feed.  The

authors calculated that the 200 mg/kg of feed represented a

daily dose of  10-18 mg/kg  bw.  After  2 years, the surviving

animals were sacrificed.   Serum  values for glucose, protein,

albumin, urea, uric acid,  cholesterol, SCOT,  and SGPT in  the

treated animals did not differ  from  those  in  controls.  No

fatty  livers were detected in the  treated  animals.  The

authors found  no biochemical  changes  attributable  to  CC14

exposure.  However,  interpretation  of the  results  was compli-

cated  by the widespread  incidence  of  chronic  respiratory

disease in  the animals.   At  18  months,  the survival ranged

from 61-89%, and more  than half  the  animals were dead at  21

months.  Although  the  authors indicated  that  10-18 mg/kg  bw

 (200 mg/kg  of  feed)  is  a  no-adverse-effect level of CC14

over 2  years,  this  conclusion may be questioned  because  of

 the poor  survival  and  chronic respiratory  infection of

experimental  animals.


     Recently, Bruckner et al. (manuscript in preparation)

investigated the oral toxicity of CC14 in male Sprague

Dawley rats.  In Study I, rats weighing 300-350 g were randomly

divided into groups of 5 animals each.  The animals were

administered by gavage 0, 20, 40, or 80 mg CCl4/kg bw (in

corn oil) daily for 5 days, allowed 2 day without dosing,

and then dosed once daily for 4 additional days.  One group

of animals at each dosage level was sacrificed at 1, 4, and

11 days following the initiation of the dosing.  The following

observations were made:  (1)  One-day treatment:  At 20 and

40 mg/kg, there were no significant changes in blood urea

nitrogen (BUN), glutamic-pyruvic transaminase  (GPT) activity,

sorbitol dehydrogenase (SDH) activity, ornithine-carbamy1

tranferase (OCT) activity or histopathological changes in

the liver or kidneys.  At 80 mg/kg, increased GPT  (p<0.05)

activity was observed.  Furthermore,  there was vacuolization

of centrilobular cells adjacent to  the central vein in each

liver.  (2)  Four-day treatment: Animals exposed to 20 mg/kg

did not exhibit any significant alterations in enzymatic

activities.  At 40 and 80 mg/kg, there was an  increase  (p<0.05)

in SDH activity. GPT activity was elevated (p<0.05) in rats

given  80 mg/kg.  Centrilobular vacuolization  was  noted  in

groups receiving 20 and  40 mg/kg.   Midzonal vacuolization was

observed in rats exposed  to  80 mg/kg.   (3)  Eleven-day

treatment: The lowest dose  (20 mg/kg) produced an  increase


(p<0.01) in SDH activity whereas 40 and 80 mg/kg increased

(p<0.05 and <0.001) serum levels of all three enzymes.   The

extent of morphological changes in the liver was dose-related.

     In Study II, rats weighing 200-250 g were randomly

divided into groups of 15 to 16 animals each.  The animals were

given by gavage 0, 1, 10, or 33 mg CCl4/kg bw (in corn oil).  The

animals were dosed on a daily basis, 5 times weekly, for a total

period of 12 weeks.  Blood samples were obtained from alternate

animals at the following  intervals: 2, 4, 6, 8, 10, and 12 weeks

post-treatment.  The serum was  analyzed for BUN, GPT, SDH and

OCT.  At 1 mg/kg,  there were no significant biochemical/histo-

pathological changes at any time during the study.  SDH, the

most sensitive index of hepatotoxicity, was elevated (p<0.05)

in rats receiving  10 mg/kg throughout the study.  Also,

these rats exhibited mild hepatic  centrilobular vacuoli-

zation.  At 33 mg/kg, levels of GPT, SDH  and OCT were markedly

increased  (p<0.01) and severe  hepatic  lesions were  apparent.

Prominent  fibrosis, bile  duct  hyperplasia,  and  hepatocellular

vacuolization were seen  in  the  portal  and periportal regions

of hepatic  lobules.  Nuclear pleomorphic  and  severe cytoplasmic

degenerative changes were commonly present  in mid-  and  centrilobular

hepatocytes.  There was  no  evidence  that  CC14 was  nephrotoxic.

Studies I  and  II  will  be  used  for  the  derivation  of one-day

and  ten-day Health Advisories;  and lifetime AADI.


     A cross-sectional epidemiclogic study (Sonich et al.,

unpublished) examined health effects of CC14 ingestion in humans.

Seventy tons of CC14 were spilled in the Kanawha and Ohio Rivers

in 1977.  Measurements of raw water revealed maximum concentra-

tions of 0.340 mg/1.  Twenty-one cities situated along the river

were involved in the study.  These cities represented areas that

obtained their drinking water directly from the river and/or

areas that obtained their drinking water from sources not in-

fluenced by the quality of  the river water.  By using river

volumes and flow rates, periods of high exposure (1977) and low

exposure (1976) to CC14 were estimated -for each city along the

river.  The results of routine tests measuring serum chemistries

reflecting  liver and kidney function along with basic epidemiologic

information were abstracted from approximated 6,000 medical records,

The results obtained for  serum creatinine  show a positive and

statistically significant  (p<0.65) relationship between  the CC14

exposure and the frequency  of elevated  levels of serum creatinine

in exposed patients.  No  similar results were found  for  the other

parameters  analyzed.

     Stewart ejt al.  (1961)  reported  the  toxic effects of

experimental exposure of  human volunteers  to CC14  vapor.  Healthy

males,  30-59 years  of age,  were  exposed  to concentrations of  63,

69, and 309 mg/m3 of CC14  (99% pure)  in  an exposure  chamber for

180 minutes at  the  two  lower doses  or  70 minutes at  the  highest

dose.   All  subjects  had  undergone  periodic physical  examinations;


some participated in more than one of the exposure experiments,

which were conducted more than 4 weeks apart.  Six subjects

exposed to the highest concentration experienced no nausea or

light-headedness, and CC14 was not detected in blood and urine

during or after exposure.  One of these six subjects had an

increased level of urinary urobilinogen 7 days after exposure.

In addition, two of four subjects exposed to the highest concen-

tration and monitored for serum  iron showed a decrease within 48

hours after exposure.  CC14 was  also not detected in the blood

or urine of volunteers exposed at 63 or 69 rng/m-^, and the

volunteers reported no physiologic effects.  No  changes in blood

pressure, serum transaminase  levels, or urinary  urobilinogen

levels were noted.

B. Quantification of Noncarcinogenic Effects

     Korsrud et_ al.  (1972)  reported  that  the  lowest  acute

oral dose of CC14 inducing  an adverse  effect  in  rats was  20

mg/kg bw  (lowest-observed-adverse-effect  level)  (Korsrud  et

al., 1972).  The  length  of  this  study  was  only  18  hours.   Alumot

et al.  (1976) proposed that 10 mg  CCl4/kg  is  the acceptable

daily  intake or  no-adverse-effect-level for  rats of  both  sexes

fed CC14  for a period of 2  years.   However,  as  discussed  earlier

this study  could  not  be  considered at  this time  due  to  high

morbidity/mortality  rate among experimental  rats and their

respective  controls.   It is noteworthy that  the  results for  the


first year of this study was used by ECAO-Cin. (USEPA, 1982a)

to derive the ambient water level for CC14 of 3.3 mg/1 for pro-

tection against CCl4~induced noncarcinogenic effects.  It should

be noted that values for consumption of contaminated water/fish

(2 I/day and 0.0065 kg/day) and bioconcentration factor for CC14

(18.75 I/kg) were taken  into consideration during the calculation

of this protective level.  The corresponding "drinking water

only" value  is 3.5 mg/1.

     EPA's existing health advisories  andi NAS' SNARLs for

CC14 could be summarized as follows:

                    EPA-health advisory  '  -  NAS-SNARL

One-day                    0.2 mg/1             14 mg/1
Seven-day                    -                 2 ^g/1
Ten-day                    0.02 mg/1
Longer-term                None3                None0

a EPA did not calculate  a  longer-term health advisory  for
CC14 due to  lack of acceptable chronic exposure  data.

.b NAS did not determine  a  longer-term SNARL  for  CC14
because  this chemical  is a carcinogen in  animals.

     The assessment of  human health  risks,  that  is,  the

likelihood  of certain  adverse  effects from given exposure

scenarios,  is hampered  by  the  paucity of  good dose-response

data  in  humans.   A  human no-observed-adverse-effect level

 (NOAEL)  for  oral  ingestion was  reported as 0.2 mg/day (  ss  0.1


ppm) for L-day exposure (Sonich et al. ,  198-1, unpublished).  The

observed effect was a dose-related increase in the frequency of

elevated serum creatinine levels in the exposed population.

However, these findings have not been published at this time.  A

human no-observed-effect level (NOEL) for inhalation was reported

as 63 mg/m^ ( fs 10 ppm) for 3-hour exposure (Stewart et al.,

1961).  The monitored effects were changes in serum enzyme and

iron levels.

     At this juncture, the two studies of Bruckner et al.

manuscript in preparation provide us with acceptable data

(dose-response relationship, length of exposure, etc.) to  revise

the existing health advisories (USEPA, 1981d) and to derive  a

lifetime AADI as follows:

     Study I (Bruckner £t_ al_. , manuscript in preparation)

showed  that one-day exposure to 20 or 40 mg CCl4/kg did

not produce any significant changes  in BUN, GPT, SDH, OCT,

or histopathological  changes in the  liver and kidneys.  At

80 mg/kg,  increased GPT activity and  vacuolization of

centrilobular cells adjacent to the  central vein in each

liver were observed.   Therefore, the  largest dose with no

significant biochemical or histopathological effects  is 40 mg

CCl4/kg  (NOAEL).


                   One-Day Health Advisory (HA)

a.   One-Day Health Advisory for Child

    (40 mg/kg/day) (10 kg) = 4 mg/1
    (100)  (1 I/day)

    where: 40 mg/kg/day = NOAEL following one-day exposure

           10 kg = weight of child

           100 = uncertainty factor based upon a good
                animal study revealing NOAEL

           1 I/day = assumed water consumption by a
                 10-kg child

b.   One-day Health Advisory for Adult

    (40 mg/kg/day) (70 kg) = 14 mg/1
    (100)  (2 I/day)

     where:  40 mg/kg/day = NOAEL following one-day exposure

             70 kg = weight of adult human

            100 = uncertainty factor based upon a good animal
                  study revealing NOAEL

               1 I/day = assumed water consumption by a
                         70-kg adult human

     Study I (Bruckner et_ al_. , manuscript in preparation)

found that eleven-day exposure to the lowest dose  (20 mg

CCl4/kg)  induced an increase in SDH whereas 40 and 80 mg

CCl4/kg increased serum levels of GPT, SDH, and OCT.  The

extent of morphological changes  in the liver was dose-related

Thus, the lowest dose of  20 mg/kg should  be considered as

the NOAEL.


                   Ten-Day Health Advisory

a.  Ten-day Health Advisory for Child

    (20 mg/kg/day) (5 days) (10 kg) = 0.142 mg/1 or 142 ug/1
    (1000) {7 days) (1 I/day)

    where:  20 mg/kg/day - NOAEL following 11-day exposure

            5/7 days = fraction converting from 5 to 7-day
                       oral exposure

            10 kg = weight of child

            1000 = uncertainty factor based upon a good
                   animal  study revealing NOAEL

             1 I/day = assumed water consumption by a
                       10-kg child

    b.  Ten-day Health Advisory for Adult

        (20 mg/kg/day) (5  days)  (70 kg) = 0.5 mg/1 or  500 ug/1
        (1000) (7 days)  (2 I/day)

         where:  20 mg/kg/day = NOAEL following 11-day exposure

                 5/7 days  = fraction converting from 5 to 7-day
                            oral exposure

                 70 kg = weight of adult  human

                  1000 =  uncertainty  factor based upon  a good
                         animal  study revealing  NOAEL

                  2 I/day = assumed water  consumption by a
                           70-kg adult  human

      Study II  (Bruckner  et_ aA.,  manuscript  in preparation)

 showed  that, following 90-day  exposure  to 1 mg  CCl4/kg,

 there were no  significant  biochemical/histopathological

 changes in rats.   At  10  mg/kg/  SDH,  the most  sensitive index

 of  hepatotoxicity, was elevated,  and mild hepatic  centrilobular

 vacuolization  was  detected.


  Lifetime Adjusted Acceptable Daily Intake (AADI) For Adult

ADI = (1 mg/kg/day) (5 days) (70 kg) = 0.050 mg/day or 50 ug/day
      (100) (10) (7 days)

      where:  1 mg/kg/day = NOAEL following 90-day exposure

              5/7 days = fraction converting from 5 to 7 day
                         oral exposure

              70 kg = weight of adult human

              100 = uncertainty factor based upon a good
                    animal  study revealing NOAEL

                10 = uncertainty factor to take into account
                    the  length of exposure  (i.e., convert
                    90-day  to lifetime exposure)

AADI = 50 ug/day  = 25 ug/day
       2 I/day

       where:   50 ug/day =  ADI

                2 I/day = assumed water consumption by a  70-kg
                         adult human

C. Carcinogenic Effects

     The carcinogenic effects of CC14 have  been  well

documented.  Oral administration of CC14  have  been shown to

be carcinogenic in  rats, mice,  and  hamsters.   In all  three

species,  liver  neoplasms developed  although  hamsters  appeared

to be the  most  sensitive.

     The  International Agency  for Research  on  Cancer  (IARC)

concluded  that  the  evidence from animal  studies  demonstrating

CCl4-induced hepatic  neoplasms  was  sufficient  to indicate

experimental animal carcinogenesis  (IARC, 1979).  The National


Cancer Institute (MCI) also identifies CC14 as an animal

carcinogen and has used it as the positive control  in three' of

its bioassays.  The following paragraphs will focus on pertinent

studies demonstrating  the carcinogenicity of CC14.

          In an NCI  (1976) bioassay  for  trichloroethylene, CC14

was used as the positive  control.  Rats; The positive control

groups of 50 Osborne-Mendel rats  of  each sex were administered

CC14  in corn oil by  gavage five  times  weekly for  78 weeks  at

two dose levels: 47  and 94 mg/kg  bw  for  males,  80 and  159

mg/kg bw for females.  The incidence of  hepatocellular carci-

nomas was increased  in animals  exposed to  CC14  as compared

with  pooled colony  controls.   However, this  was statistically

significant only for females  given  the low dose as  compared

with  the colony  controls  and  not the matched controls.   Absolute

incidence of hepatic neoplasms  was  low (5% in  the animals exposed

to CC14).  This  may be attributed to the resistance by this

rat strain to  CC14.   The  incidence  of other neoplasms was

acknowledged  but  not quantified.  This study was used by NAS

 (1978)  in  determining the carcinogenic risk estimate for CC14

due to  the dose levels used and the appropriate length of the

 study.   Mice;  B6C3F1 male and female mice (35 days of age, 50

 per group)  were given CC14 (1,250 or  2,500 mg/kg bw) in corn

 oil by  gavage five times weekly for 78 weeks.  Surviving mice

 were sacrificed at 92 weeks from the  start of  the  study.  There


were 20 control mice af each sex that were given corn oil only.

A necropsy was performed on all mice along with complete histo-

logical examinations.

     Most male and female mice treated with CC14 were dead

by 78 weeks.  Hepatocellular carcinomas were  found  in practically

all mice receiving CCl^, including  those dying before termination

of the test.  The first carcinomas  were observed in  low dose

female mice at 16 weeks, in high dose  female  mice at 19 weeks,

in high dose males at  26 weeks and  in  low  dose males at 48 weeks,

compared to 72 weeks for pooled control males and 90 weeks for

pooled control females.  Cystic endometrial hyperplasia occurred

in both control  and  treated female  mice.   Thrombosis of the

atrium of the heart was seen  in 9  of  41 high  dose female  mice

(22%), all  of which  died with  carcinomas  of  the  liver.   In summary,

this study  found CC14  to be high-ly carcinogenic  for liver in

mice and  is used by  the World  Health  Organization  (WHO,  1981)  in

ascertaining  the carcinogenic risk estimates  for CC14.

     Edwards  et_ a.1.  (1942) investigated the carcinogenic  potential

of  CC14  in  mice.  The  mice used were  inbred strain  L with

extremely low incidence of spontaneous hepatomas,  2.5-3.5 months

or  3.5-7.5  months of age at the onset of the experiment.   The

 number of mice  varied  from 8-39 per group.  Carbon tetrachloride

was administered in olive oil by stomach tube usually three,   but

 occasionally  two, times weekly.  Each treatment consisted of

 0.1 cc of a 40% solution or 0.04 ml of CC14.  Mice were given


 46 administrations  of  CC14  over  a  4-month  period  and  were

 sacrificed and  necropsied  3-3.5  months  after the  last treatment.

 The mice  varied from 8.5-14 months of  age  at necropsy.   The  liver

.was examined  histologically.

     Hepatomas  developed  in 34/73  mice  (47%) given CC14.

 Ftepatomas were  observed in 7/15  younger male mice (47%),  21/39

 older  male mice (54%), 3/8 younger females (38%), and 3/11 older

 females  (27%).   Cirrhosis  of the liver was not mentioned.  Histori-

 cally, the incidence of spontaneous  hepatomas in  strain L  mice  is

 extremely low:   2/152 (1%)  in untreated mice.  One of 23 untreated

 virgin male mice (4%)  and  0 of 28  females  (0%),. necropsied at 15

 months of age,  had  tumors  of the liver.  Tumors were  not present

 in 22  males and 28  females 18 months of age or in 27  female

 breeders  12-23  months of age.  One of 24 male breeders (4%)  had a

 tumor.   In summary, strain L male  and female mice were highly

 susceptible  to  the  induction of  hepatomas  by CC14, and male  mice

 were slightly more  susceptible than female mice.

     Delia Porta e_t_ al. (1961) exposed Syrian golden  hamsters

 to CC14  in order to investigate the response of  this  species to

 carcinogens  that induced liver neoplasms in other species.   Ten

 female and  10 male  Syrian golden hamsters,  12 weeks old, were

 used.  At the onset of the experiment, males weighed  an average

 of 99  g  and  females weighed an average of 109 g.   At  the end of

 the experiment, the average weight was 104 g for both sexes.


'The treatment consisted  of weekly  administration  by  stomach

tube of a 5% solution of CC14 in corn  oil  for  30  weeks.  Controls

cited were historical controls  kept  by the investigators in  the

same laboratory  for  a  lifespan.  A total of 145  female  and 109

male hamsters of  the same  strain,  fed  the  same diet,  did not

develop hepatic  tumors.  The authors also  cited  controls for  a

different study  they conducted.   In  this latter  study,  30  female

and 50 male  hamsters fed the same  diet but given 0.5 ml corn  oil

via stomach  tube  twice  weekly for  45 weeks also  did  not develop

hepatic tumors.   During the  first  7  weeks  of the former experiment,

0.25 ml of  the solution containing 12.5 ul CC14  was  given  each

week.  This  dose was then  reduced  to 0.125 ml and contained  6.25

ul of CC14.   After this treatment, the survivors were kept

under observation for  25 additional weeks  and then sacrificed.

Detailed  histopathological examinations of all hamsters were

conducted,  except for  one  female lost through cannabalism  at

 the  28th  week.

     Weights of  the hamsters varied irregularly during the

 period  following treatment.   In general,  the weights increased.

 Females weighed  an average of 114 g and males 113 g.  One  female

 died  at  the 10th week of treatment; three  females and  five males

 died  or  were sacrificed between the 17th  and  the 28th week.   Three

 females  died at weeks 41, 43 and  54.  The  surviving  three females

 land  five males  were sacrificed at  the end  of the 55th week.


     Hamsters dying during the treatment and at the 41st week

manifested cirrhosis, as well as hyperplastic nodules that were .two

to several layers thick.  The cells showed  irregularities in the

shape, size and staining qualities of their cytoplasm and nucleus,

with an uneven distribution of glycogen.

     All of the animals, five males and  five females, dying or

sacrificed 13-25 weeks  after  the end of  the treatment, had one or

more hepatic carcinomas  (a total of 22 tumors:  12  in the females

and 10 in the males).   No mention  was made  of toxicity in these

animals.  Hepatic carcinomas  were  not found in  the  other animals

dying before week 43.

     In summary, Syrian  golden hamsters  appear  sensitive to

the carcinogenic effects of 0014.  Although the number of animals

in this study was small, the  authors considered the results  to  be

significant because  the reported historical control incidence of

hepatic tumors in hamsters was 0/254.  Hyperplastic nodules  ap-

peared during treatment, and  carcinomas  appeared  after CC14  admin-

istration had been discontinued, which suggests that the  nodules

or benign tumors were  precursor  lesions  for carcinomas.   It  should

be noted  that this study is. the  only  report found in the  available

literature of CC14 induction  of  tumors  in hamsters.

     The  above studies by NCI (1976),  Edwards  e_t  al_.  (1942),

and Delia Porta  et al. (1961) were used  by EPA's  Office  of  Health


and Environmental Assessment (OHEA) to calculate the unit risk

estimates for CCl^ (USEPA, 1983).

     Although several investigators noted that toxic effects

of CC14 are concurrent with liver tumors (hepatomas), it has

not been established that tissue damage (i.e., necrotic cirrhosis)

is a necessary precursor  to CCl4-induced carcinogenesis.

     Despite a wealth of  data on its toxic effects, there is

little definitive information on its metabolism or  its mode of

carcinogenic action.  Among reported metabolic reactions in liver

are conversion to carbon  dioxide, chloroform, hexachloroethane,

carbonyl chloride (phosgene), and binding to  lipids and proteins.

Diaz Gomez and Castro  (1980) reported  that L4C from 4CC14

irreversibly binds in vivo to hepatic  nuclear DNA  from mice and

rats.  Also, binding of 14C from 14CC14 to DNA was  detected

in vitro in  incubation mixtures  containing microsomes and a

NADPH-generating system as well  as  in  tissue  slices.  Liver

nuclear proteins and  lipids  irreversibly bind CC14  metabolites.

The authors  concluded  that  (a)  the  interaction of  CC14  metabolites

with DNA and nuclear  proteins could be relevant  to  CCl4-induced

liver  tumors and hepatoxic  effects; and  (b)  the  epigenetic  mechan-

isms for chemical induction  of  cancer, not  involving CC14-DNA

interactions could also be  relevant.   There  have  been  no  reports

of mutagenic activity  associated with  CC14  in any  of  the  various


Salmonella (Ames) assays.  However, mutagenic activity associated

with CC14 has been observed in a eukaryotic test system using

the yeast Saccharomyces cerevisiae (Gallon el: al., 1980).  This

report has not been confirmed and should not be accepted as

conclusive evidence of CC14 mutagenicity.  Carbon tetrachloride

did not cause chromosome damage (i.e., chromatid gaps, deletions,

or exchanges) during an  in vitro chromosome assay using cultured

rat-liver cells  (Dean and Walker, 1979).  Mirsalis and Butterworth

(1980) found that treatment of male rats with CC14 (10 or 100

ing/kg administered by gavage) produced no increase in unscheduled

DNA synthesis in cultures of primary  rat hepatocytes.  According

to the authors,  this observation indicates that CC14 does not

act through a genotoxic  mechanism.  Thus, the genotoxic potential

of CC14 obviously needs  further investigation.

D. Quantification of Carcinogenic'Risk

     Because of  positive results in animal carcinogenicity

studies, CC14 can be considered a suspect human  carcinogen.

Data from the animal studies  have been used  by NAS  (1977) and

OHEA  (USEPA, 1980a;  1983)  to  calculate  the upper  bound  on the

number of additional cancer cases  that may occur  when CC14  is

consumed  in drinking water over  a  70-year  lifetime.   As  shown  in

Table IX-I, using the OHEA and NAS  data,  estimates  of additional

carcinogenic risk following  the  exposure  of  humans  to CC14  may

be derived.

                          Table IX-I
Estimates of Additional Carcinogenic Risk Following Exposure to
                   CC14 in Drinking Water3
                               CC14 Concentrations  (ug/1)
Excess Cancer
(Lifetime )

(USEPA, 1980a)J

(USEPA, 1983)e
NAS ( 1 9 7 7 ) <-"
a assuming an average  daily  drinking  water  consumption  of  2  liters

b lower 95% confidence  limit

c based on NCI  (1976)  (rats)

d based on NCI  (1976)  (mice)

e based on NCI  (1976)  (rats  and  mice),  Edwards e_t al_.  (1942)  (mice),
  and Delia Porta  et al.  (1961)  (hamsters)


     The criteria for the OHEA and NAS risk calculations

differ in several respects: (1) NAS used the multistage model,

while OHEA used an "improved" multistage model,  (2) NAS used the

data set from the National Cancer Institute (NCI) study in male

rats while OHEA used the data set from NCI's study  in male mice

(USEPA, 1980a), and used a geometric mean of four studies  (NCI,/

1976 - mice; NCI, 1976  - rats; Edwards e_t a_l. ,  1942 - mice; and-

Delia Porta et_ al_. , 1961 - hamsters)  (USEPA, 1983).

     EPA's Ambient Water Quality Criteria for CC14  (USEPA,

1980a) were based on increased lifetime  cancer  risk estimates

of 10-5 (4.0 ug/1), 10~6 (0.40 ug/1), and 10~7  (0.04 ug/1)

calculated by--OHEA.  It is noteworthy that  these estimates were

derived by assuming a lifetime consumption  of both  drinking

water  (2 I/day) and aquatic life  (6.5 g  fish and shellfish/day)

grown  in waters containing the corresponding CC14  levels.

Specifically, OHEA's daily CC14 exposure assumptions were  as

follows: 94% from ingesting drinking  water  and  6%  from  consuming

seafood "fish factor."  The corresponding  "drink water  only"  con-

centrations are 4.4, 0.44, and 0.04  ug/1, respectively.

     Using the same data set  as OHEA and a  linear  multistage

model, WHO (1983) derived  a recommended  tentative  limit for CC14

of 3 ug/1.  This  level  should give  rise  to  less than  1  additional

cancer per 100,000 population for  a  lifetime of exposure  assuming

a 2-liter daily consumption of drinking  water.


     In addition to MAS', OHEA's and WHO'S estimates of addi-

tional carcinogenic risk following exposure of humans to CC14 in

drinking water, OHEA calculated a unit risk estimate for humans

from exposure to CC14 in water as follows: 0.37 x 10~5 for a

person continuously exposed to 1 ug CC14 per liter of water,

(USEPA, 1983).  Since no single study was entirely adequate for

risk assessment, this estimate is based upon the geometric

mean of four studies discussed above and correspond to drinking

water concentrations presented in Table IX-I.  It should be noted

that EPA's Science Advisory Board approved OHEA's approach for

calculating unit risk estimates for 0014.

E. Special Considerations

     It is noteworthy that  in assessing CCl4~induced  toxicity,

carcinogenicity or any  other harmful effect,  compounds  that

react synergistically or antagonistically with CC14 must be

considered.  Identified synergistic  substances  include  ethanol,

kepone, PCB, and PBB.   Antagonistic  effects  have  been  demonstrated

with such  compounds  as  chloramphenicol  and  catechol.

     Sensitive  populations  are  subgroups  within  the  general

population which appear at  higher than  average  risk  upon  exposure

to CC14.   Some  of  the populations that  may  be at  greater  risk

include human  fetuses,  alcohol  consumers,  and males  of repro-

ductive age.


F.  Summary

     The recommended values for one-day, ten-day for both

children and adult humans, the lifetime AADI for adult humans, and

the estimated lifetime cancer risks are summarized in Table IX-II.

                           Table IX-II

    Summary of Quantification of Toxicological Effects of CC14
                                        Drinking Water
                              10-kg Child               70-kg Adult
 One-Day Health Advisory         4 mg/1                  14 mg/1

 Ten-Day Health Advisory       142 ug/1                 500 ug/1

 Lifetime AADI                                          25 ug/1

 Excess Cancer Risk

        10-4                                            27    ug/1

        10-5                                              2.7  ug/1

        10-6                                              0.3  ug/1

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