HISTORICAL RESEARCH REPORT
Research Report TM/80/05
1980
The chronic toxicity of benzene: with
particular reference to its cytogenetic
effects. A review of the literature
Lloyd MH
HISTORICAL RESEARCH REPORT
Research Report TM/80/05
1980
The chronic toxicity of benzene: with particular
reference to its cytogenetic effects. A review of the
literature
Lloyd MH
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ii
Research Report TM/80/05
REPORT NO. TM/80/5
UDC 615.9 :
5^7.532
THE CHRONIC TOXICITY OF
BENZENE : WITH
PARTICULAR REFERENCE TO
ITS CYTOGENETIC EFFECTS
A Review of the
Literature
Margaret H. Lloyd
JULY 1980
Price:
£40.00 (UK)
£45.00 (Overseas)
I N S T I T U T E
OF
O C C U P A T I O N A L
M E D I C I N E
THE CHRONIC TOXICITY OF BENZENE : WITH PARTICULAR REFERENCE TO
ITS CYTOGENETIC EFFECTS
A Review of the Literature
by
Margaret H. Lloyd
Medical Branch,
Institute of Occupational Medicine,
Roxburgh Place,
EDINBURGH EH9 9SU.
(Tel. 031-667-5131)
July 1980
(ii)
CONTENTS
•
Page No.
SUMMARY
(iv)
1.
INTRODUCTION
1
2.
METABOLISM OF BENZENE
2
3.
BONE MARROW TOXICITY OF BENZENE
3
3.1 Pancytopenia
3
3.2 Leukaemia
4
4.
6
CHROMOSOMAL ABNORMALITIES AND BENZENE
4.1 Methods of detecting chromosomal damage
....
6
4.1.1
Chromosomal analysis
6
4.1.2
Sister chromatid exchanges
7
4.2 Animal chromosome studies
8
4.3 Human chromosome studies
8
4.3.1
4.3.2
Studies on individuals with a
history of benzene toxicity . . . . . .
8
Studies on workers showing no.
signs of benzene toxicity
9
4.4 The significance of chromosomal abnormalities
...
10
5.
SURVEILLANCE OF WORKERS EXPOSED TO BENZENE . . . .
12
5.1
The benzene standard
12
5.2
Monitoring of benzene exposure
12
5.2.1
Urinary phenol levels
12
5.2.2
Expired air benzene concentrations
5.3
5.4
. . . .
13
Screening of exposed workers for haematological
abnormalities
13
Screening of exposed workers for chromosomal
abnormalities
13
5.4.1
Indications for screening
13
5.4.2
Feasibility of chromosomal screening
...
14
(iii)
CONTENTS contd.
Page No
6.
CONCLUSIONS
6.1
Is there evidence of an increased frequency
of chromosomal aberrations in workers
exposed to benzene?
.
16
6.2
What is the significance of these abnormalities?
16
6.3
Should workers be screened for chromosomal
abnormalities?
•
•
*
•
•
•
•
•
•
•
*
•
•
•
16
16
ACKNOWLEDGMENT
19
REFERENCES
21
GLOSSARY
25
REPORT NO. TM/80/5
(iv)
I N S T I T U T E
OF
O C C U P A T I O N A L
M E D I C I N E
THE CHRONIC TOXICITY OF BENZENE : WITH PARTICULAR REFERENCE
TO ITS CYTOGENETIC EFFECTS
A Review of the Literature
by
Margaret H. Lloyd
SUMMARY
Benzene is toxic to the bone marrow. Exposure of
susceptible individuals to high levels causes pancytopenia
and leukaemia which may develop after a latent period of
many years. The haematological effects of low exposure
levels are less well documented.
There is substantial evidence that exposure to high levels
of benzene causes chromosomal abnormalities in the
peripheral blood and bone marrow which may persist for years.
There are conflicting reports that exposure to levels within
the present safety standard for benzene may cause chromosomal
abnormalities in the absence of clinical or haematological
signs of benzene toxicity.
The significance of the observed chromosomal abnormalities
in exposed workers is unknown. It is possible that they may
predispose to the development of leukaemia.
There is no
evidence at present to suggest that benzene causes lethal or
inherited mutations.
There is a need for further studies to measure the prevalence
of chromosomal abnormalities in workers exposed to low levels
of benzene, and for the follow-up of workers who have
chromosomal abnormalities as a result of high exposure in the
past. The results of these studies would indicate whether
there is a need for routine chromosomal screening and longterm follow-up of workers exposed to benzene.
1.
INTRODUCTION
Benzene is produced and used in enormous quantities throughout the
industrialised world.
In 197^ the National Institute of Occupational
Safety and Health (NIOSH) estimated that two million people in the USA
were potentially exposed to it.
Benzene and other aromatic hydrocarbons
are produced during the manufacture of coke and during the refining of
petroleum.
It has been widely used as a solvent in the rubber,
printing and chemical industries but because of its health hazards it
is now being replaced by other solvents.
Benzene is also important
in the synthetic chemical industry as a precursor of other organic
compounds.
The bone marrow toxicity of benzene was first recognized in 1897 by
Santesson and is now well established.
Its propensity to cause
chromosomal damage has only recently been recognized and the significance
of this has not yet been evaluated; with so many people at risk the
implications for this and future generations could be serious. The aim
of this paper is to review the literature on the cytogenetic effects of
benzene and attempt to answer two questions. Is there evidence of an
increased prevalence of chromosomal abnormalities in workers exposed to
benzene?
If there is, what is the clinical significance of these
abnormalities and should screening procedures be established?
A review of the metabolism and bone marrow toxicity of benzene is
relevant to a discussion of its effects at a chromosomal level.
2.
2.
METABOLISM OF BENZENE
Benzene may be inhaled or absorbed through the skin.
After
inhalation, about 12$ of the absorbed dose is eliminated unchanged
in the expired air.
The remainder is metabolised in the liver by
oxidative enzymes to phenol, catechol and quinol which are then
transformed mainly to phenylsulphuric and phenylglutaric acids and
excreted in the urine (RUSCH ^t al., 1977).
There is evidence from
animal experiments that bone marrow toxicity is caused by a metabolite
of benzene rather than by benzene itself. SNYDER & KOCSIS (197*0
studied benzene-treated mice and used the reduction in incorporation of
59Fe into their erythrocytes as a measure of the haemopoietic toxicity
of benzene;
they showed that toluene which is a competitive inhibitor
of benzene metabolism, increased 59Fe uptake and phenobarbitone, which
increases the activity of the hepatic oxidative enzymes and therefore
stimulates benzene metabolism, decreased 59Fe uptake. If a metabolite
is responsible for the toxic effects of benzene, then individual and
species differences in the metabolic handling of benzene could account
for the observed differences in individual susceptibility to benzene
and for the failure to induce leukaemia in benzene-treated animals.
3.
3.
BONE MARROW TOXICITY OF BENZENE
The serious effects of chronic exposure to benzene are limited to the
bone marrow. The fact that workers are often exposed to a mixture
of volatile compounds rather than to benzene alone has been a major
problem in establishing a causal relationship between observed bone
marrow toxicity and benzene exposure. However, there is now good
evidence to support such a relationship between benzene and both
pancytopenia and leukaemia (GOLDSTEIN, 1977).
There is a wide
variation in individual susceptibility the reason for which is not
well understood.
AKSOY (1979) from his observations on leukaemia
among Istanbul shoemakers >hasr suggested that a geme.tic,a;Lly;.-determine.d
host factor for leukaemia may be triggered by benzene exposure.
3.1
Pancytopenia
A decrease in the three formed elements of the blood is the most
frequent effect of chronic exposure to benzene, the leucocytes being
most commonly affected. The red blood cell (RBC) count may be
decreased but this change is usually preceded by the development of
macrocytosis. A proportion of exposed individuals develop a decrease
in all three elements (pancytopenia) and some have gone on to develop
fatal aplastic anaemia.
The bone marrow is usually hypoplastic but
hyperplasia has been described (GOLDSTEIN, 1977).
The true incidence of bone marrow toxicity is difficult to establish.
The majority of reports have been on individuals exposed to high levels
of benzene.
HERNBERG jet al. (1966) studied 1^7 shoe factory workers
who were exposed to high levels of benzene (400 parts per million (ppm))
for up to ten years. One hundred and seven of these workers were
found to have abnormal blood counts - of which the most common was
thrombocytopenia.
Ten of these men required hospital admission and of
these one man died. When studied again nine years later, some of the
workers still had subnormal cell counts; the severity of the original
changes did not correlate with eventual recovery.
Of greater relevance are the studies done on workers exposed to lower
concentrations of benzene.
An investigation of 282 Dow chemical workers
exposed to 2 - 30 ppm Time-Weighted Average (TWA) of benzene for 1 - 2 0
years found that the only abnormalities were a statistically significant
k.
decrease in the meam RBC count and total serum bilirubin when compared
with a control group (TOWNSEND et al., 1978). No correlation was
found between these changes and duration of exposure or estimated
career dosage of benzene.
A number of reports have been published of individuals who have
recovered from benzene-induced aplastic anaemia and who have
subsequently developed leukaemia, in some cases many years later (AKSOY,
1979).
3.2 Leukaemia
Evidence that chronic exposure to benzene causes leukaemia came
initially from isolated case reports and from studies of groups of
individuals working under poor conditions and exposed to high
concentrations of benzene.
AKSOY (1979) studied 35 cases of leukaemia
among Istanbul shoemakers between 1967 and 1975- He found
myeloblastic leukaemia was most common (37.500 but that the
of the usually rare preleukaemia and acute erythroleukaemia
The duration of exposure to benzene ranged from 4 months to
that acute
incidence
was also high.
20 years and
in some cases leukaemia developed years after exposure to benzene had
ceased.
Similar findings have been reported in workers in other
industries (GOLDSTEIN, 1977). In the Istanbul shoe factories the use
of benzene was stopped in 1969 and AKSOY (1979) reports that no new cases
of leukaemia presented in 1976.
Several epidemiological studies among rubber workers in the United States
have been carried out and all of them have reported an excess mortality
from leukaemia among exposed workers compared with control groups not
occupationally exposed to benzene (McMICHAEL eit al., 1975; MONSON &
NAKANO, 1976; INFANTE et al., 1977). No detailed information about
levels of benzene exposure is given.
Two of these studies carried out
by McMichael and Monson included workers who at one time used benzene for
cleaning purposes and therefore were exposed to high concentrations.
The third study was on a group of 7^8 men involved in the production of
natural rubber film (Pliofilm) in Ohio between the years 19*fO - 19^9.
During this time they were exposed to levels of benzene below the
benzene standard in operation at that time, i.e. 35 ppm TWA. When
studied retrospectively in 1975 it was found that the exposed workers
had an estimated fivefold risk of dying from leukaemia when compared
with two control populations (a group of fibrous glass workers in Ohio
who had not been exposed to benzene and a general population group)
(INFANTE £t al., 1977).
Conflicting results were produced by two other studies. THORPE (197*0
failed to produce evidence from a study of 38,000 petrochemical workers,
some of whom had been exposed to benzene, of an increased mortality from
leukaemia.
OTT et_ &L. (1978) studied a group of 335 men who had begun
working in benzene areas before 1950 in the Dow Chemical Group and were
known to have been exposed to low concentrations of benzene, i.e. never
greater than 35 ppm TWA. Although they concluded that no deaths could
be directly attributed to benzene exposure, three cases of myeloid
leukaemia were reported in this group (expected incidence 0.8) but they
observed that benzene was not the only solvent to which these men were
exposed.
So far it has proved impossible to induce leukaemia in animals by
exposing them to benzene (GOLDSTEIN, 1977). The reason for this is not
understood; it may be that there is a species difference in the
metabolic pathways of benzene and that a toxic metabolite is not
produced in the species which have been used experimentally;
alternatively benzene leukaemia may occur only in genetically susceptible
individuals as suggested by AKSOY (1979) or when benzene exposure occurs
in conjunction with other chemical or viral agents. This lack of an
animal model has contributed to our lack of understanding of the
mechanism by which benzene induces leukaemia.
The possible association
between leukaemia and chromosomal aberrations will be discussed later
6.
k.
CHROMOSOMAL ABNORMALITIES AND BENZENE
Genetic changes can be produced by a chemical compound in three ways:
(i) It may produce molecular changes in the DNA of the gene.
This is
termed a point mutation and the change is invisible under the microscope,
(ii) It may cause chromosome breaks which may lead to chromosomal
aberrations if the chromosome fragments rejoin abnormally, e.g. to form
ring chromosomes, translocations, etc. (iii) It may produce a change
in the number of chromosomes by affecting the distribution of
chromosomes at cell division.
^.1 Methods of detecting chromosomal damage
Evidence of damage may be sought directly by chromosomal analysis or, less
directly, by detection of an increased number of sister chromatid exchanges.
If.1.1 Chromosomal analysis
The effect of a chemical on chromosomes may be studied by in vitro or in vivo
methods. In the former-the chemical is added to the culture medium containthe cells, the chromosomal pattern of which is subsequently determined.
In vivo methods involve analysis of the cells of animals or individuals
who have been exposed to the chemical.
Few in vitro studies of the effect
of benzene on cells have been carried out and the subsequent discussion in
this paper will refer to in vivo methods.
Lymphocytes are the cells most commonly studied. They can be stimulated
to divide by incubation with phytohaemagglutinin for kS - 72 hours;
mitosis is stopped at metaphase by the addition of colchicine.
The
chromosomes are stained and viewed under a light microscope.
The banding
pattern produced by staining is specific for each chromosome, making
accurate identification of each chromosome and the detection of
abnormalities possible. Two types of chromosomal abnormalities are
recognized; unstable chromosomal aberrations (Cu) include fragmentation
of chromosomes, deletions, dicentric and ring chromosomes.
These
abnormalities do not usually survive cell division and are therefore
unlikely to have any long-term effect.
Stable chromosomal aberrations
(Cs) include abnormal monocentric chromosomes produced by deletions and
translocations.
These changes do survive cell division and may
therefore constitute a persistent mutation.
greater significance than Cu.
They are therefore of
7.
Two important facts must be borne in mind when evaluating results:
( i)
A large number of factors (including virus infections
and some drugs) are known to produce chromosomal
aberrations (LOEFFLER, 1973; PURCHASE, 1978).
There
is also evidence that chromosomal abnormalities increase
with age (TOUGH e£ al., 1970) and in a study of
irradiation-induced chromosomal damage a positive age
dose interaction was observed (EVANS ££ jQ., 1979)*
(ii)
Point to point mutations (i.e. changes in the DNA
composition of a gene) do not alter the appearance of
a chromosome.
A chemical compound may cause genetic
damage without altering the microscopic appearance of
chromosomes; other experimental techniques, e.g. using
microbial systems or the fruit fly, Drosophila, must be
used to test the mutagenicity of a compound (SOBELS, 1977).
4.1.2 Sjster-chromatid exchanges (SCE)
Genetic material (DNA) can be exchanged between sister chromatids
(i.e. the pair of chromatids which constitute a chromosome).
The
exchange sites can be detected in cultured cells by special staining
techniques after incubation of a cell culture for two rounds of mitoses
with bromodeoxyuridine which becomes incorporated into chromosomal DNA
(EMERY, 1979). A small number of SCEs may be detected in normal cells
but the number is increased on exposure to irradiation and certain
chemical carcinogens (including benzene). It is generally considered
that SCEs reflect DNA damage although the exact relationship between
SCEs and induced mutations is not fully understood (WOLFF & CARRANO,
1979).
The number of SCEs in cultured cells is also increased in two rare
clinical syndromes, xeroderma pigmentosa and Bloom's syndrome
(dwarfism and a photosensitive skin rash). It is of interest and may
be relevant to this discussion of benzene toxicity, that in both of
these conditions there is an increased risk of neoplasia; patients
with Bloom's syndrome are predisposed to develop leukaemia and those
with xeroderma are predisposed to skin cancers (EMERY, 1979).
8.
k.2 Animal chromosome studies
Experiments using rats and rabbits exposed to subcutaneous benzene
have demonstrated a greatly increased incidence of chromosomal
abnormalities in these groups compared with control animals (DEAN,
1978). Although there is experimental evidence that benzene produces
chromosomal damage, there is no evidence as yet that it produces point
mutations.
Experiments to test the mutagenicity of benzene using
microbial systems (e.g. Salmonella typhimurum) and Drosophila have
produced negative results (DEAN, 1978; RAY, 1979).
^.3 Human chromosome studies
These have provided the most convincing evidence that benzene exposure
can cause chromosomal damage (WOLMAN, 1977). As with bone marrow
toxicity, the fact that workers are often exposed to a mixture of volatile
compounds including benzene, may complicate the interpretation of the
results of chromosomal studies. Of the compounds with which benzene may
be mixed, the limited clinical evidence available suggests that neither
xylene nor toluene cause chromosomal damage (DEAN, 1978; FORNI et al.,
1971a); phenols have been shown to interfere with cell division in plant
tissues but there are no reports of their effect on animal tissues
(DEAN, 1978).
k.3.1
Studies on individuals with a history of benzene toxicity
Cytogenic studies on leucocytes from patients suffering from benzeneinduced pancytopenia and leukaemia have frequently shown an increase in
both stable and unstable chromosomal abnormalities (FORNI e£Lal^, 1971b;
ERDOGAN & AKSOY, 1973). In several cases cells containing V?
chromosomes (instead of the usual ^6) were consistently isolated from
the bone marrow and peripheral blood suggesting that the stable
chromosome abnormalities induced by benzene had given rise to the
development of abnormal clones of cells (DEAN, 1978).
Studies on groups of individuals with a past history of benzene toxicity
have also shown an increase in chromosomal aberrations when compared with
a group of age- and sex-matched controls (FORNI et al., 1971b). These
abnormalities were still present in some cases years after cessation
of exposure to benzene and return of the blood picture to normal. There
was no correlation between the severity of benzene poisoning and the
9.
persistence of chromosome changes.
In these reports very little
information about the level of benzene exposure is given but it is
very likely that the subjects were exposed to high concentrations of
benzene, in some cases for many years.
Some cases of acute benzene poisoning have been reported.
The
majority of these have a high number of unstable chromosomal abnormalities
which falls over subsequent years (FORNI ie_t al., 19?1b).
^.3.2 Studies on workers showing no signs of benzene toxicity
Studies on workers who have been exposed to low levels of benzene and
who have shown no clinical or haematological evidence of benzene toxicity
are highly relevant to the present discussion. Unfortunately few such
studies have been carried out.
TOUGH et_ jil. (1970) studied a group of
20 men who were working in a distillation plant where benzene was known to
have been present intermittently. The benzene levels were not
accurately known but they state that the atmospheric benzene level 'was
probably low, in the region of 12 ppm1. Amongst this group there was no
evidence of an increased frequency of chromosomal abnormalities when
compared with a general population control group.
A better study involving a larger group of workers and more detailed
estimations of benzene exposure was reported by PICCIANO (1979). He
carried out cytogenetic studies on a group of 52 workers said to have
been exposed to low levels of benzene (< 10 ppm) for periods ranging
from one month to 26 years. He compared the results with those obtained
from a group of kk men seen for pre-employment examinations.
He found
that the benzene exposed workers had twice the percentage of chromosome
breaks and three times the percentage of ring and dicentric chromosomes
compared with the control group. The major criticism of this study is
once again the failure to relate the prevalence of chromosomal
abnormalities to the degree and duration of benzene exposure.
Picciano's
statement that workers were exposed to 'low levels of benzene, less than
10 ppm' seems to be based on a single measurement of urinary phenol
immediately prior to blood collection and measurement of ambient benzene
by fixed air and personal monitors over the four-year period prior to
the study. No information is given about exposure prior to this or to
the relationship between the duration of employment and the incidence of
chromosomal aberrations.
10.
If it is assumed that exposures prior to the four-year sampling period
were equally low then these results do suggest that chromosomal
abnormalities occur in workers exposed to low concentrations of
benzene and who show no other signs of toxicity. There is obviously
a need for a larger, better designed study of such workers.
k.k The significance of chromosomal abnormalities
In general, damage to the genetic material of germ cells may cause
increased foetal loss, congenital abnormalities in the offspring of
affected workers or in the members of future generations if the mutation
is recessive.
No long-term family studies of workers with a past history of benzene
exposure have been reported so it is not known if benzene causes
increased foetal loss, congenital abnormalities or recessive mutations.
FORNI ert al_. d971b) describe the cases of two pregnant women with bone
marrow toxicity and both delivered normal children. One of the women
had chromosomal abnormalities in peripheral leucocytes suggestive of the
presence of an abnormal clone of cells; during this time she produced
two normal children and cytogenetic studies on one of them showed no
evidence of chromosomal damage. Animal experiments have shown that
exposure of pregnant rats and rabbits to benzene at a critical time in
pregnancy caused retarded foetal growth and minor skeletal defects (e.g.
fused ribs) but no serious abnormalities have been noted (HUDAK & UNGUARY,
1978).
Damage to somatic cells may predispose the individual to the development
of malignant tumours (JACKSON, 1973). The link between benzene and
leukaemia is established but there is no evidence that benzene exposure
causes malignant change in organs other than the bone marrow (OTT et al.,
1978).
The relationship between chromosomal aberrations and leukaemia is
uncertain.
As discussed previously, patients with benzene induced
leukaemia do have a high incidence of chromosomal abnormalities but so do
patients with leukaemia not attributed to benzene exposure. The
observed chromosome changes could therefore be the result of rather
than the cause of the leukaemia. However, several cases have been
11.
described where the chromosome changes have preceded the development
of leukaemia (DEAN, 1978). In these cases it seems likely that
benzene caused chromosomal damage which led to the development of
abnormal clones of leucocytes (FORNI ert al., 1971b).
Alternatively
benzene could produce leukaemia in susceptible individuals by activation
of a latent leukaemogenic virus or by damaging lymphocytes and thereby
impairing 'immune surveillance'.
There is a similarity between these observations and those linking
irradiation-induced chromosomal damage with an increased incidence of
neoplastic disease. There are also two rare, but well described
genetically determined syndromes, Fanconi's anaemia and ataxia
telangiectasia, which are characterised by an increased tendency to
spontaneous chromosomal aberrations and a predisposition to the
subsequent development of malignant disease (JACKSON, 1973)•
12.
5.
SURVEILLANCE OF WORKERS EXPOSED TO BENZENE
The recognition of the haematoxicity of benzene had led to the
successive tightening of the benzene standard over the years and the
development of methods of monitoring exposed workers.
5.1 The benzene standard
'' ii*.
This currently stands at 10 ppm Time-Weighted Average (TWA) for a
^0-hour working week (ACGIH, 1979).
Since 1920 when the standard was
100 ppm there have been successive reductions in the standard (THORPE,
1978).
The present standard was introduced in 197^ and has been
adopted by most countries including the United Kingdom. In 1976 NIOSH
recommended that the occupational exposure standard for benzene be
lowered to 1 ppm as determined by a two-hour air sample collected at
one litre per minute. Following the publication of studies on the
rubber workers of Ohio (INFANTE jet al., 1977) an emergency standard of
1 ppm was introduced in the United States. This is currently being
contested in the courts by the industries concerned (ZENZ, 1978). Some
authors have argued that such a reduction is unnecessary (THORPE, 1978).
Until the outcome of the legal battle has been decided the standard
remains at 10 ppm TWA for an eight-hour day in the United States and
most European countries.
5.2 Monitoring of benzene exposure
In addition to monitoring of ambient benzene concentrations, personal
exposure can be measured by estimation of phenol levels in urine or
of benzene in expired air.
5.2.1
Urinary phenol levels
This is the most widely used technique in industry. A large
proportion of inhaled benzene is metabolised to phenolic compounds
which are excreted in declining quantities over kS hours after
exposure to benzene.
The collection of a single urine specimen is
acceptable to workers and the assay method using gas chromatography is
sensitive and easy to perform (CARTER, 1979). There is a linear
relationship between urinary phenol and environmental benzene levels
(KUSNETZ & HUTCHISON, 1979). The main drawback of this technique is
its lack of specificity as other factors including drugs (e.g. mild
analgesics and caffeine) and food can influence urinary phenol levels.
Benzene exposure at levels of 5 - 10 ppm TWA but not below 5 PP"i are
13.
reflected in urinary phenol levels. Individuals with levels
exceeding 50 mg/litre (standardised to urine specific gravity (S.G.)
1.0l6)should be investigated and the cause established (CARTER, 1979).
Excretion of over 100 mg/litre of phenol (standardised to S.G. 1.016)
strongly indicates excessive industrial exposure (TREVETHICK, 1976).
5.2.2 Expired air benzene concentrations
Immediately after exposure, the concentration of benzene in exhaled
air falls rapidly but then the rate of fall flattens out. SHERWOOD
(1972) has suggested that benzene is distributed between at least two
compartments; the first, represented by circulating blood has a half
life of 2.5 hours and the second, represented by muscle and fat has a
half life of approximately 2k hours. Thus measurement of exhaled
benzene gives an indication of the timing and duration of exposure.
A rapid fall in concentration of benzene over a few hours indicates
recent exposure; a slow fall in concentration suggests greater exposure
in the more distant past. CARTER (1979) considered that this method
could be developed for use at exposure levels as low as 1 ppm.
5.3 Screening of exposed workers for haematological abnormalities
Pre-employment and regular (e.g. six-monthly) blood counts are usually
recommended with investigation and removal from benzene exposure if
indicated, of individuals showing abnormal counts (TREVETHICK, 1976).
There is evidence that qualitative changes in circulating blood may
precede quantitative changes. They include decrease in leucocyte
alkaline phosphatase, decrease in phagocytic function, aggregation of
platelets and change in serum levels of immunoglobulins (GOLDSTEIN, 1977;
COHEN e£ al., 1978). It is possible that these may be affected by very
low levels of benzene, i.e. < 5 ppra but their use in screening exposed
workers has not been evaluated.
5.^ Screening of exposed workers for chromosomal abnormalities
5.^.1 Indications for screening
As previously discussed there is some evidence to suggest that low levels
of benzene exposure may cause chromosomal abnormalities.
Further studies
of chromosomal aberrations in benzene workers are essential, but before
undertaking regular routine screening of all benzene workers it would
14.
be advisable to carry out studies designed specifically to investigate
the prevalence and attack rate of chromosomal abnormalities in workers
whose exposure to benzene is or has been measured. Such studies
would provide information on which to base decisions on routine screening
as well as information contributing to the choice of safety standards.
Some information of this nature may become available from regular cytogenic surveillance of workers exposed to possible clastogenic agents in
progress in the Dow Chemical Group in the USA (KILIAN & PICCIANO, 19?6).
Furthermore, nothing is known about the clinical significance and longterm effects of chromosomal abnormalities such as those observed by
PICCIANO (1979) in men working in supposedly low concentrations of
benzene. A follow-up study of benzene workers with and without
chromosomal abnormalities, with suitable control groups is necessary.
It would also be important to study the effects of variations in exposure
levels on workers. There is evidence from work on the clastogenic effects
of radiation and of vinyl chloride that high doses given over short periods
produce more chromosomal damage than an equivalent dose given at a low
dose rate (PURCHASE, 19?8). It may be that workers exposed to peak
concentrations of benzene are more at risk than workers with a constant
exposure level.
5.4.2 Feasibility of chromosomal screening
There are three possible techniques which could be used to screen exposed
workers:
(
i) Sjster-chromatid exchanges
This may be of value in screening the accidentally
over-exposed worker but is unlikely to be useful as
a widespread screening technique. It is known that
an increase in the number of SCEs indicates genetic
damage and that such changes are produced by irradiation
and chemical mutagens. However, its use as a screening
test in chronically exposed workers is limited by the
fact that the increase in SCEs is a transient one after
exposure (STETKA & WOLFF, 19?6). There are no reports
of SCE levels in workers exposed to low levels of
benzene.
15.
( ii) Traditional light microscopy methods
If screening were to be carried out in this country
the traditional method of karyotyping would be used.
For each individual screened, a blood sample has to
be cultured for 72 hours and then after suitable
staining, the karyotype of at least 100 cells has to
be individually determined by a technician; the
examination for each individual usually taking about
three hours.
If results are to be interpretable it is likely that
a Marge number-of,.karyotype-determinations on a
large number- of-individuals would ••be'necessary (TOUGH
et al., 1970). The workload imposed on a cytogenetics
unit by regular cytogenic surveillance of an industrial
population is well illustrated by the work of Evans and
his colleagues (EVANS et al., 1979). They studied a
group of 197 workers in a nuclear dockyard over a period
of 10 years. Samples of blood were taken from the men
before employment in irradiated areas and at their
annual medical checkups. This involved determining the
karyotype of at least 50,000 lymphocytes.
It was found
that there was a dose-dependent increase in unstable
chromosomal aberrations (Cu). The incidence of stable
chromosomal aberrations was also increased but not so
markedly as Cu and the change was not dose-dependent.
The increase in certain types of aberration (mainly of
the chromatids) increased with age and a positive age-dose
interaction was noted.
(iii)
Automated chromosome analysis
This is being developed in the United States by the Dow
Chemical Company (KILIAN & PICCIANO, 1976). Research
into the development of automated methods is also in
progress in this country but no method for routine use is
available at present (MASON & RUTOVITZ, 1978).
16.
6. CONCLUSIONS
It is now possible to answer the questions raised in the Introduction.
6.1 Is there evidence of an increased frequency of chromosomal
aberrations in workers exposed to benzene?
There is no doubt that exposure to high concentrations of benzene causes
bone marrow toxicity and chromosomal abnormalities in some exposed workers.
The effect of low concentrations of benzene is less clear. There is
certainly some evidence to suggest that exposure to low benzene levels
does cause chromosomal damage but further work in this field is needed.
It must be pointed out that all the reported studies have been carried
out on somatic cells (e.g. lymphocytes) and not on the germ cells of
exposed workers. However, it is generally considered that environmental
factors which cause genetic damage in somatic cells may produce similar
damage in germ cells.
6.2 What is the significance of these abnormalities?
Very little is known about the significance to the individual worker and
future generations of the observed chromosomal changes. It is possible
that the chromosomal abnormalities seen in workers exposed to high
concentrations of benzene predisposes them to leukaemia. Nothing is
known about the significance of chromosomal abnormalities seen in workers
exposed to low concentrations of benzene.
6.3 Should workers be screened for chromosomal abnormalities?
Routine screening of exposed workers for chromosomal abnormalities is
time-consuming and expensive and the results need careful evaluation in
view of the other known factors which may cause chromosomal damage. To
justify routine screening there should be good evidence that exposure to
a chemical at concentrations within the current safety standard causes
chromosomal damage. In the case of benzene there is insufficient
evidence on which to base recommendations on routine screening of exposed
workers.
Whilst the evidence linking low benzene concentrations with chromosomal
damage is insufficient to recommend routine screening, it is certainly
sufficient to recommend further research in this field. Other questions
remain unanswered. There are no reported studies on workers exposed to
17.
high concentrations of benzene in the past which compare the clinical
and family histories of those found to have chromosomal abnormalities
with the histories of those found to have a normal karyotype.
studies need to be carried out to answer these questions.
Further
A cross-sectional epidemiological study of chromosomal aberrations in
benzene workers, with inclusion of suitable controls, would indicate
whether there is an increased prevalence of chromosomal abnormalities
in the current workforce. In these studies it would be essential to
have measurements of benzene exposure using environmental and biological
monitoring techniques so that attempts could be made to correlate the
frequency of chromosomal aberrations with measurements of present, and
estimates of previous, exposure.
A prospective study would also provide valuable information on the
cytogenetic effects of benzene on exposed workers.
Such a study could
only be carried out on individuals who have no past history of benzene
exposure and who are about to enter occupations where they will become
exposed to benzene. Once again measurements of benzene exposure would
be essential. Their initial cytogenetic studies would provide built-in
controls and by taking medical histories at the time of blood sampling
it would be possible to identify other factors (e.g. viral infections)
which may cause chromosomal damage. Studies along these lines have
been carried out on nuclear dockyard workers (EVANS et_ al., 1979) and on
workers exposed to epichlorohydrin (KUCEROVA & ZHURKOV, 1977).
If a link between chromosomal aberrations and exposure to low
concentrations of benzene is established, then routine screening of
exposed workers and long-term follow-up of them and their families would
be advisable to determine whether the presence of chromosomal
abnormalities indicates an increased risk of clinical disease.
19.
ACKNOWLEDGMENT'
I should like to thank Dr. C.A. Soutar for his helpful suggestions
during the preparation of this paper.
<f
21.
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25.
GLOSSARY
Aplastic anaemia
The condition which results from failure
of the bone marrow to produce and release
into the blood stream mature red cells,
white cells and platelets.
In many
patients the cause is unknown but in others
the condition may be linked with exposure
to drugs, chemicals or irradiation.
Ataxia telangiectasia
An inherited condition characterised by
dilated blood vessels in the skin,
degeneration of the part of the brain which
controls balance, chromosomal aberrations
and a predisposition to develop tumours of
the lymphatic system.
Bloom's syndrome
An inherited condition; individuals
affected have stunted growth, a photosensitive skin rash, chromosomal
aberrations and an increased risk of
leukaemia.
Chromatids
When cells divide each chromosome splits
longitudinally into two strands which are
called chromatids.
A change in the number or structure of the
Chromosomal abnormality/
aberration
chromosomes.
Chromosomes
Thread-like structures present in the
nucleus of each cell. They are composed of
DNA and protein and carry genetic
information.
Human germ cells contain one
set of chromosomes, i.e. N = 23. All other
body cells contain two sets, i.e. 2N = 46,
one set derived from each parent.
26.
Clastogenic
An agent which causes genetic damage.
Clone
All the cells derived from a single cell
by repeated mitoses and all having the
same genetic constitution.
Congenital abnormality
An abnormality which is apparent at birth.
It may be of genetic origin or may be
caused by exposure of the developing foetus
to toxic agents.
Deletion
A type of chromosomal aberration when part
of a chromosome is lost.
Dicentric chromosome
An abnormal chromosome which has its
chromatids joined at two sites instead of
one.
DNA
Deoxyribonucleic acid: chromosomes are
largely composed of this nucleic acid which
carries the genetic code.
Fanconi's anaemia
An inherited condition; affected
individuals may have kidney and bone
abnormalities and develop aplastic anaemia
during childhood.
They have a high rate of
chromosomal aberrations and tend to
develop leukaemia and solid tumours.
Gene
This is the part of the DNA molecule which
controls protein synthesis.
Germ cells
Sperms or ova.
Karyotype
A photomicrograph of an individual's
chromosomes arranged in a standard way,
27.
Leucocytes
White blood cells.
Leukaemia
An abnormal and uncontrolled proliferation
of white cells which invade the bone
marrow and other tissues.
Leukaemia may
be classified as acute or chronic and by
the predominant type of white cell
involved, e.g. lymphatic leukaemia when
lymphocytes proliferate and myeloid
leukaemia when granulocytes
proliferate.
Lymphocytes
A type of white blood cell.
Macrocytosis
Enlargement of the red cells;
there are
many causes of this and it may be
associated with anaemia (macrocytic
anaemia).
Metaphase
This is one of five stages which occur
during cell division.
Mitosis
This term is used to describe the process
of division of somatic cells.
Monocentric chromosome
The normal appearance of a chromosome when
its two chromatids are joined at a single
site, i.e. they appear X-shaped under the
microscope.
Mutation
An alteration in the genetic material of a
cell brought about either by a change in
the number or structure of the chromosomes
or a molecular change in the DNA of a gene.
A mutation which occurs in the germ cells
(sperms or ova) is inherited; if it
produces a trait which is incompatible with
life it is termed a lethal mutation. A
mutation which occurs in somatic cells is
not inherited.
28.
Pancytopenia
A reduction in the number of white cells,
red cells and platelets in the peripheral
blood circulation.
One of the causes of
pancytopenia is aplastic anaemia.
Recessive
A recessive mutation only becomes apparent
in an individual if it is inherited from
both parents.
Ring chromosome
An abnormal chromosome which is ring-shaped.
Somatic cells
All cells of the body apart from those of
the reproductive organs (i.e. sperms or ova),
Thrombocytopenia
A reduction in the number of platelets in
the circulating blood.
Time-Weighted Average
(TWA)
This is one of the categories of Threshold
Limit Values for chemical substances in workroom air adopted by the American Conference
of Governmental Industrial Hygienists in
1978.
TWA is the time-weighted average
concentration for a ^0-hour work week to
which workers may be repeatedly exposed
without adverse effects.
Translocation
A chromosomal aberration when a part of one
chromosome is displaced and joins onto
another chromosome.
Xerodenna pigmentosa
Affected individuals have solar induced
skin damage and are predisposed to skin
cancers; the condition is inherited.
(A20115) IOM (R) ReportCov art
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