HEXACHLOROBENZENE JMPR 1974
Hexachlorobenzene was studied by the Joint Meeting in 1969
(FAO/WHO, 1970) and a "tentative negligible daily intake" of 0.0006
mg/kg bw was established. The Joint Meeting in 1973 (FAO/WHO, 1974a)
recommended that the term "tentative negligible daily intake" should
not be used and the figure of 0.0006 mg/kg bw was retained as a guide
for setting upper limits for residues until it was possible to
allocate an ADI based on the results of comprehensive toxicological
studies. The further work requested in 1969 included: Metabolic
studies in animals including identification of the toxic product or
products excreted in milk; short-term studies in non-rodent mammalian
species and long-term studies especially in relation to the effect on
bone-marrow; reproduction studies in rats.
The following recommendation was made in 1973 (FAO/WHO, 1974a):
In view of the potential toxicity of hexachlorobenzene and the lack of
adequate toxicological data to assess its safety, WHO and FAO should
promote and, where necessary, co-ordinate research needed on the
In view of the widespread occurrence of HCB residues in a wide
range of food commodities and in the environment the 1973 Meeting also
decided that this compound should be kept under continual review and
that it should be reconsidered by the 1974 Joint Meeting. Further
information was required to enable the Meeting to review proposals
The report and monographs of the 1973 Joint Meeting were not
available in time for governments and interested authorities to
consider requests for additional information. The present Meeting had
before it considerable information obtained by members from published
and unpublished scientific literature and this provided the basis for
reconsideration of the question of hexachlorobenzene residues.
EVALUATION FOR ACCEPTABLE DAILY INTAKE
The 1973 Joint Meeting reviewed the status of HCB and found that
very little toxicological data required by the 1969 Joint Meeting had
The Meeting examined recently published reports relating to tests
for mutagenic and teratogenic action (Khera, 1974), biochemical
effects (Grant et al., 1974a,b), histopathology (Kuiper-Goodman, 1974;
Kimbrough and Linder, 1974), tissue disposition studies (Villeneuve et
al., 1974; Villeneuve, 1975; Villeneuve and Hierlihy, 1975), and
reproduction studies (Somers et al., 1973). Although none of these
reports included long-term studies, the results that were reported
permitted the Meeting to reaffirm the value of 0.0006 mg/kg bw
suggested by the 1973 Meeting (FAO/WHO 1974a) as a guide for setting
upper limits for residues. Since the results of a long-term feeding
and carcinogenesis study, known to be in progress, were not available,
the Meeting deferred full evaluation for an ADI. The present Meeting
allocated a value of 0.0006 mg/kg bw as a conditional ADI.
This Meeting expressed concern that some effects attributed to
HCB might be due to impurities in the test samples (Villanueva et al.,
It was noted that the number of reports of residues in foods,
feeds and human tissues was increasing. The sources of these residues
are known to include disposal of industrial and municipal wastes,
contamination of other chlorinated pesticides, the approved use of HCB
as a seed-dressing, and misuse of HCB-treated seeds in animal feeds.
The meeting urges that:
(1) support be given to an international monitoring program to
identify the source and extent of contamination;
(2) the presence of HCB as an impurity in other pesticides be
monitored and minimized;
(3) the recommendations for use as a seed dressing (FAO/WHO, 1974b) be
carefully adhered to; and
(4) HCB should be used only as a seed-dressing and only when no
suitable substitute is available. Because of the persistent nature of
HCB and its general occurrence in the environment, it must be
recognized that even if all sources of emission could be stopped, some
food contamination with HCB will continue for many years and the
Meeting therefore urged that data required by the 1973 Joint Meeting
(FAO/WHO, 1974a) be promptly obtained.
RESIDUES IN FOOD AND THEIR EVALUATION
SOURCE OF RESIDUES
In the previous monograph (FAO/WHO, 1974b) reference was made to
various industrial as well as agricultural sources of HCB residues.
The Meeting was able to consider a number of publications
indicating the widespread distribution of hexachlorobenzene residues
derived, apparently, from many different sources. Siyali and Stricker
(1973) report the presence of hexachlorobenzene along with other
organochlorine pesticides in milk from a survey carried out in
Australia. Johnson et al. (1974) report finding hexachlorobenzene
residues in fish, fish eggs, fish fry and fish oil from a collection
of samples taken throughout the United States in 1971 and 1972. The
widespread occurrence points to the possibility that there are many
different sources of HCB contamination. Zitko (1971) reports finding
trace amounts of hexachlorobenzene together with PCB and other
organochlorine residues in fresh water and marine fishes during a
survey in Canada.
The U.S. Environmental Protection Agency reported (1973) that
whereas HCB residues had frequently been found in the fat of domestic
animals from several regions of the U.S.A., examination of 150 samples
of soil from 16 separate states revealed that there was no general
widespread contamination of crop land soils with HCB residues at
levels above the limit of determination (0.01 mg/kg). On the other
hand residues of other organochlorine pesticides were frequently
found, dieldrin being found in 30% of the samples examined.
Gilbertson and Reynolds (1972) report finding significant levels
of HCB in the eggs of common terns in Hamilton Harbour, Ontario,
Canada. The level of HCB residues is similar to that of dieldrin,
being noticeably lower than the level of DDT and PCB in the same eggs.
This information suggests that there is a general low level of HCB
contamination in the whole environment. HCB residues were reported to
occur in soil and crops after repeated soil treatments with quintozene
(Smelt and Leistra, 1974).
In reporting studies on the loss of chloronitrobenzene fungicides
from the soil, Caseley (1968) mentions that 400,000 lbs of quintozene
was used annually in the San Joaquin Valley in California, U.S.A. in
1963. The Joint Meeting has already referred (FAO/WHO, 1974b) to the
significant contamination of quintozene with hexachlorobenzene,
leading to residues of hexachlorobenzene in food produced with the aid
of quintozene fungicides. Beck and Hansen (1974) refer to the fate of
HCB in the soil following the use of quintozene.
RESIDUES IN MEAT RESULTING FROM EXPERIMENTAL FEEDING
A study of the tendency of broiler chickens to accumulate
organochlorine pesticide residues from contaminants in their feed was
carried out in The Netherlands (De Vos et al., 1972). Day old broiler
chickens were fed combinations of various organochlorine pesticide
residues at levels of 0.05, 0.15 and 0.3 ppm in the feed for a period
of about 7 weeks. Birds were killed at regular intervals and samples
of abdominal fat and other tissues examined for residues of the
organochlorine pesticides. At the end of the 7 week period heptachlor
epoxide residues were highest, followed by hexachlorobenzene,
dieldrin, DDT, endrin and lindane.
As shown in Table 1 the results of this experiment indicate a
direct relationship between the amount of HCB in the feed and the
amount in the fat of the chickens. The concentration in the abdominal
fat of chickens receiving 0.15 ppm of HCB in their rations is
approximately 3 times that of the corresponding birds receiving
rations containing 0.05 ppm. Analysis of livers of the same broiler
chickens indicate a similar relationship between the concentration of
HCB in the feed and in the liver fat.
TABLE 1 HCB residues, mg/kg, in chickens fed HCB in their diet (from DeVos et al., 1972.
Level (ppm) fed: 0.05 0.15 0.30
Days on diet: 15 29 52 15 29 52 15 29 52
(whole) 0.045 0.061 0.075 0.12 0.21 0.24 0.34 0.37 0.42
(fat basis) 0.76 0.62 0.68 2.3 2.0 1.8 6.2 3.5 3.1
Fat (abdominal) 0.68 0.55 0.66 2.2 1.8 1.8 3.9 3.1 3.2
(whole) - - 0.020 - - 0.043 - - 0.14
basis) - - 0.43 - - 1.1 - - 2.1
chickens - - 0.047 - - 0.15 - - 0.29
(fat basis) - - 0.47 - - 1.6 - - 2.8
RESIDUES OF HCB IN SOIL AND CROPS RESULTING FROM
TREATMENT OF SOIL WITH QUINTOZENE
In a study of the disappearance of quintozene and its technical
impurities and metabolites pentachlorobenzene, hexachlorobenzene and
pentachloroaniline from soil Beck and Hansen (1974) calculated that
hexachlorobenzene had a half-life of 969-2089 days when applied to
soil in laboratory experiments at the rate of 10 kg/ha. The same
authors studied samples of soil collected at two different locations
representing different soil types from fields which had previously
been treated with quintozene. They found HCB residues in 21 of the 22
samples at levels ranging from 0.17 to 0.94 mg/kg with an average of
0.38 mg/kg. Soil from fields which had not been treated for two or
more years showed only slightly less HCB than soil from fields which
had been treated more recently. They found no direct relationship
between the hexachlorobenzene and quintozene residue levels in the
soil. This is understandable because quintozene is metabolized whereas
HCB is degraded only very slowly. It is obvious from these studies
that soil treated with technical quintozene which contains
hexachlorobenzene as an impurity may be expected to carry appreciable
residues of hexachlorobenzene for a number of years. Other studies
(Bech and Hansen, 1973) have shown that such residues may be taken up
by potatoes and carrots.
The Meeting had available results of studies in The Netherlands
aimed at determining the HCB content of soils and crops following
repeated treatment with quintozene. Samples of soil were collected
from 22 fields which had been treated with quintozene from 1 to 12
times during more than 10 years. These samples were all found to
contain HCB residues at levels ranging from 0.001 to 0.41 ppm. There
appeared to be no direct relationship between the level of HCB
residues and the number of treatments which had been applied. However
the fields had been cultivated many times during the period of use of
quintozene and it is very likely that the residue would have been
distributed among the mass of soil to a considerable depth, depending
upon the form of cultivation (Smelt and Leistra, 1974). In the same
studies the content of HCB in crops growing in contaminated soil was
studied and the ratio of the plant residue to the soil residue was
determined. As seen from table 2 there is a distinct uptake of HCB by
root crops, and in the case of carrots the concentration in the root
was from 12-19 times as high as in the soil. There is an even higher
uptake by the roots of grass. There is evidence of a distinct
translocation into the leaves of some crops, particularly carrots,
grass and sugar beet. With the exception of carrots, grass and turnip
roots, the concentration of HCB in the dry plant material is lower
than in the corresponding soil.
Residues of HCB in dry mass of crops and soil, and ratio of
plant residue to soil residue (from Smelt and Leistra, 1974)
Crop, Residue (mg/kg) Ratio,
part of plant No crop soil crop/soil
Potato, 1 0.11 0.22 0.50
tubers 2 0.20 0.40 0.50
3 0.13 0.18 0.72
4 0.017 0.027 0.63
5 0.10 0.16 0.63
6a) 0.21 1.17 1.24
7a) 0.021 0.034 0.62
8a) 0.27 0.30 0.90
TABLE 2. (cont'd)
Crop, Residue (mg/kg) Ratio,
part of plant No crop soil crop/soil
Tulip, 1 0.12 0.30 0.40
bulbs 2 0.065 0.10 0.65
3 <0.002 0.003 <0.67
4 0.16 0.10 1.60
Shallot, 1 0.056 0.12 0.47
Sugar-beet, 1 0.095 0.41 0.23
beets below 2 0.012 0.027 0.44
surface 3 0.027 0.056 0.48
4 0.010 0.024 0.42
Sugar-beet, 1 0.014 0.41 0.03
crowns 2 0.004 0.027 0.15
3 0.005 0.056 0.09
4 0.002 0.024 0.08
Sugar-beet 1 0.022 0.41 0.05
leaves 2 0.010 0.027 0.37
3 0.017 0.056 0.30
4 0.006 0.024 0.25
Grass, 1b) 0.81 0.09 9.0
roots 2b) 0.56 0.14 4.0
3b) 0.19 0.07 2.7
4c) 0.25 0.027 9.2
5d) 0.76 0.033 23
6d) 0.039 0.001 39
Grass, 1b) 0.22 0.09 2.4
lower part of 2b) 0.20 0.14 1.4
blades (0-5 cm) 3b) 0.10 0.07 1.4
Grass, 1b) 0.028 0.09 0.31
upper part of 2b) 0.042 0.14 0.30
blades (above 5 cm) 3b) 0.016 0.07 0.23
4c) 0.021 0.027 0.78
5d) 0.003 0.033 0.09
6d) <0.003 0.001 -
7d) 0.011 0.12 0.09
Grass, 6d) <0.003 0.001 -
hay 7d) 0.011 0.12 0.09
Carrot, 1 1.25 0.065 19
roots 2 0.48 0.04 12
TABLE 2. (cont'd)
Crop, Residue (mg/kg) Ratio,
part of plant No crop soil crop/soil
Carrot, 1 0.44 0.065 6.8
leaves 2 0.25 0.04 6.2
Turnip, 1 0.18 0.062 2.9
Turnip, 1 0.014 0.062 0.23
a) sampled in early July.
b) young grass, one to two months after sowing, about 20 cm long.
c) as b), but about 10 cm long.
d) pasture, one year old.
EVIDENCE OF RESIDUES IN FOOD IN COMMERCE OR AT CONSUMPTION
A survey of 198 milk samples from a wide cross section of New
South Wales was conducted over three years for pesticide residue
assessment (Siyali and Stricker 1973). HCB residues were found in 12%
of the samples, the highest residue reported being 0.005 mg/kg.
METHODS OF RESIDUE ANALYSIS
The Meeting had before it a number of papers concerned with the
analysis of residues of hexachlorobenzene.
The international co-operative study of organochlorine pesticide
residues in terrestrial and aquatic wild life (Holden, 1970) included
test samples containing small concentrations of HCB. Only a few of the
17 co-operating laboratories were able to detect this residue and even
these reported wide variations. On one sample the results were
reported to range from a trace to 0.14 mg/kg and on another sample
only two of the 17 laboratories reported finding HCB.
A method of confirming the presence of HCB residues by chemical
reaction suitable for use in most residue laboratories has been
advanced by Baker (1973). When HCB in hexane solution is treated with
sodium ethoxide, monoethoxypentachlorobenzene is formed. The method is
useful in distinguishing HCB from various HCH (BHC) isomers.
A method of sampling and an analytical procedure for determining
HCB in air has been published by Mann et al. (1974). The method is
suitable for determining the distribution of HCB from industrial or
A new regulation has been announced by The Netherlands concerning
the fungicide pentachloronitrobenzene (quintozene), which normally
contains up to 2´% of hexachlorobenzene as an impurity. In view of the
high persistence of hexachlorobenzene and the fact that it is likely
to enter the food chain, thus causing unduly high unintentional
residues in animal products, the following restrictions are being
imposed. As from 1 August 1974 only quintozene containing no more than
0.1% of hexachlorobenzene and no more than 1% of pentachlorobenzene is
approved for use as a fungicide on lettuce and on seed-potatoes. As
from 1 May 1975 the same restriction will apply to the material for
use on flower-bulbs and for all other uses. Meanwhile, investigations
are being carried out by the Laboratory for Insecticide Research with
respect to accumulation of hexachlorobenzene in soil and its uptake in
The practical residue limits recommended in 1969 and 1973 are
Baker, B.E. (1973) Confirmation of hexachlorobenzene by chemical
reaction. Bull. environ. Contam. Toxicol., 10(5):279-284.
Beck, J. and Hansen, K.E. (1973) National Food Institute, DK 2860
Beck, J. and Hansen, K.E. (1974) The degradation of quintozene,
pentachlorobenzene, hexachlorobenzene and pentachloroaniline in soil.
Pestic. Sci., 5:41-48.
Caseley, J.C. (1968) The loss of three chloronitrobenzene fungicides
from the soil. Bull. environ. Contam. Toxicol., 3(3):180-193.
De Vos, R.H., Bouwman, J. and Engel, A.B. (1972) Residues of
organochlorine pesticides in broilers from feed fortified with known
levels of these compounds. Pestic. Sci., 3:421-432.
FAO/WHO. (1974a) 1973 Evaluations of some pesticide residues in food,
FAO/AGP/1973/M/9/1. WHO Pesticide residues series, No. 3.
FAO/WHO. (1974) The use of mercury and alternative compounds as seed
dressing. Report. FAO Agricultural Studies, No. 95; Wld. Hlth Org.
techn. rep. ser., No. 555.
Gilbertson, M. and Reynolds, L.M. (1972) Hexachlorobenzene (HCB) in
the eggs of common terns in Hamilton Harbour, Ontario, Bull. environ.
Contam. Toxicol., 7:371-373.
Grant, D.L., Hatina, G.V. and Munro, I.C. (1974) Hexachlorobenzene
accumulation and decline of tissue residues and relationship to some
toxicity criteria in rats. Preprint of paper presented at IUPAC
Conference, Helsinki, July 1974, submitted to Environ. Qual. and Saf.
Grant, D.L., Iverson, F., Hatina, G.V. and Villeneuve, D.C. (1974)
Effects of hexachlorobenzene on liver porphyrin levels and microsomal
enzymes in the rat. Environ. Physiol., 4:159-165.
Holden, A.V. (1970) International Co-operative Study of Organochlorine
Pesticide Residues in Terrestrial and Aquatic Wildlife, 1967/1968.
Pestic. Monit. J., 4:117.
Johnson, J.L., Stalling, D.L. and Hogan, J.W. (1974) Hexachlorobenzene
(HCB) residues in fish. Bull. environ. Contam. Toxicol.,
Khera, K.S. Hexachlorobenzene: Teratogenicity and dominant lethal
studies in rats. Food Cosmet. Toxicol. (In press)
Kimbrough, R.D. and Linder, R.E. (1974) The toxicity of technical
hexachlorobenzene in the Sherman strain rat. A preliminary study.
Chem. Path. Pharmacol., 8:653-664.
Kuiper-Goodman, T., Pontefroct, R.D. and Grant, D.L. (1974) X-Ray
(TEM) microanalysis of iron accumulated in liver parenchymal cells of
rats with porphyria. Proc. Microsc. Soc. Canada, 1:14.
Mann, J.B., Enos, H.F., Gonzalez, J. and Thompson, J.E. (1974)
Development of sampling and analytical procedures for determining
hexachlorobenzene and hexachloro-1,3-butadiene in air. Environ. Sci.
Siyali, D.S. and Stricker, P. (1973) Hexachlorobenzene and other
organochlorine pesticides in milk. Aust. J. Dairy Technol., 28:55-58.
Smelt, J.H. and Leistra, M. Contents of HCB in soil and crops after
repeated soil treatments with quintozene. Paper from Lab. for Research
on Insecticides, Wageningen, Neth. (in press)
Somers, E., Grant, D.L. and Phillips, W.E.J. (1973) Toxicodynamics of
organochlorine compounds, from Pesticides and the Environment: A
Continuing, Controversy, Symposia Specialists, North Miami, Florida.
U.S. Environmental Protection Agency. (1973) HCB residues in cropland
soils - Monitoring Note 3. Part of report presented to Hazardous
Materials Advisory Committee.
Villeneuve, D.C., Panopio, L.G. and Grant, D.L. (1974) Placental
transfer of hexachlorobenzene in the rabbit. Environ. Physiol.
Villeneuve, D.C. The effect of food restriction on the redistribution
of hexachlorobenzene (HCB) in the rat. Toxicol. Appl. Pharmacol. (in
Villeneuve, D.C. and Hierlihy, S.L. Placental transfer of
hexachlorobenzene in the rat. Bull. environ. Contam. Toxicol. (in
Villanueva, E.C., Jennings, R.W., Burse, V.W. and Kimbrough, R.D.
(1974) Evidence of chlorodibenzo-p-dioxin and chlorodibenzofuran in
hexachlorobenzene. J. agr. Food Chem., 22:916-917.
Zitko, V. (1971) Polychlorinated biphenyls and organochlorine
pesticides in some freshwater and marine fishes. Bull. environ.
Contam. Toxicol. 6:464-470.