1970 EVALUATIONS OF SOME PESTICIDE RESIDUES IN FOOD
Issued jointly by FAO and WHO
The content of this document is the result of the deliberations of the
Joint Meeting of the FAO Working Party of Experts and the WHO Expert
Group on Pesticide Residues, which met in Rome, 9-16 November, 1970.
FOOD AND AGRICULTURE ORGANIZATION OF THE UNITED NATIONS
WORLD HEALTH ORGANIZATION
2-chloroethyltrimethyl-ammonium ion (usually as the chloride)
chlorcholine chloride, Cycocel(R), CCC
[Cl CH2. CH2N (CH3)3]+Cl-
Other information on identity and properties
White crystalline solid, typical amine (fish like) odour. Soluble in
lower alcohols; insoluble in ether and hydrocarbons; water solubility
74 g/100 ml at 20°C.
Aqueous solutions are chemically stable and retain their biological
Purity of technical grade, 97 to 98 percent. Traces of 1,
2-dichloroethane and trimethylamine occur as impurities.
Commercial formulations include 10 percent, 40 percent and 50 percent
solutions and 65 percent dust.
EVALUATION FOR ACCEPTABLE DAILY INTAKE
Although considerable studies are reported to have been conducted on
the biochemistry and toxicology of chlormequat, including short and
long-term studies, much of this information was not available to the
Meeting. In the absence of full reports on this subject, it was not
possible to consider the toxicology of chlormequat. Therefore, no
acceptable daily intake (ADI) was established at this time.
RESIDUES IN FOOD AND THEIR EVALUATION
Chlormequat is registered in at least 17 countries as a plant growth
regulator particularly to promote sturdier growth in wheat, rye and
oats and thus reduce the risk of lodging. It is applied to wheat as a
single treatment at the rate of 0.6-3.0 kg per hectare and to oats at
the rate of 1-2 kg/ha when the cereal crop plants are 10-20 cm high
(wheat) and 25 to 30 cm high (oats and rye). It is also used to
improve the fruit set and yield of grapes (Coombe, 1965). For this
purpose, 300 grammes of active ingredient is applied per hectare of
vines one to three weeks before flowering.
Chlormequat is used in a number of countries as a growth regulator for
use on ornamentals to reduce vegetative growth and enhance flowering.
Many other uses have been evaluated. While a number of these
applications appear promising, there is as yet no acceptance of these
uses on commercial food crops other than wheat, rye, oats and grapes.
RESIDUES RESULTING FROM SUPERVISED TRIALS
Residue data assembled from controlled experiments in Sweden, France,
Netherlands, Germany, United Kingdom, Kenya and New Zealand show that
when chlormequat is used to regulate the growth of wheat, rye and oats
so as to give the plants added resistance to lodging, residues of the
parent compound occur in the grain at harvest.
Table I gives typical results obtained from supervised trials with
wheat and rye.
Chlormequat residues in grain from crops treated to prevent lodging
Chlormequat Interval -
Country Type of Crop kg/ha treatment to Residues
Sweden Rye A 2 13 1.0
1967 Rye A 2 16 3.2
Rye B 2 14 4.2
Rye B 2 17 1.5
1968 Rye C 2 14 0.57
Rye C 2 15 0.52
Rye C 2 17 0.36
Rye A 2 14 1.67
Rye A 2 15 1.2
Rye A 2 17 1.2
United Spring Wheat 1 12 0.75
Kingdom " " 1 15 0.1
1966 " " 2 10 0.68
" " 2 15 0.15
Winter Wheat 2.5 12 0.44
" " 2.5 17 0.18
United Spring Wheat 1.0 22 0.4
Kingdom Winter Wheat 1.5 20 0.34
TABLE I (cont'd)
Chlormequat residues in grain from crops treated to prevent lodging
Chlormequat Interval -
Country Type of Crop kg/ha treatment to Residues
Sweden Spring Wheat 2 12 0.23
1965 Spring Wheat 2 15 0.1
Winter Wheat 3 12 1.5
Winter Wheat 3 18 0.13
Netherlands Wheat 2 12 0.65
1965 Wheat 2 24 0.1
Kenya Wheat A 1 21 0.1
1965 Wheat A 1.5 21 0.19
Wheat B 1.5 21 0.1
France Wheat 2.25 13 0.3
1965 Wheat 4.5 13 0.4
New Zealand Wheat 2 17 0.4
Jung (1964) examined 150 wheat samples from more than 20 field trials
in West Germany during 1963 and established a distinct relationship
between residues and the quantity of chlormequat applied. Using an
analytical method sensitive to only 0.5 ppm, it was reported that no
residues could be detected in the grain of crops sprayed early with 3
kg/ha, and that only 23% of 22 samples from crops treated at the rate
of 4.5 kg/ha contained detectable residues ranging around 0.5 ppm with
a maximum of 1 ppm. Later trials (Jung, 1969a) on 12 crops and using
more sensitive analytical procedures showed maximum residues of 0.47
ppm when chlormequat was applied according to label recommendations.
At rates of application of 1.0-1.5 kg per hectare and recommended
harvest intervals, residues in the grain were most frequently in the
range of 0.1-0.3 ppm, with an occasional value approaching 1 ppm
(American Cyanamid Co., 1966b, 1968b; Cyanamid of Great Britain Ltd.,
1965, 1966; Jung and Henjes, 1964).
On the basis of these results, it appears that the residue level in
wheat gradually declines, so that early treatment or slow maturing
varieties are likely to have lower residues than those crops where
treatment is applied closer to harvest. The range of residue values
is, however, not great. Residues are not likely to exceed 1 ppm in
wheat at harvest under normal conditions.
Studies by Blinn (1967) using 14C-labelled chlormequat showed that
0.2 ppm of the parent substance remained as a residue in the grain
after the foliage had been treated 12 weeks previously.
Jung (1968) evaluated the residues in short-straw winter rye treated
as recommended in West Germany at 4 locations and found residues
ranging from 1.0 to 1.3 ppm following the application of 1.8 kg/ha.
Reports from other countries indicate that the residue levels in rye
are somewhat higher than in wheat and often exceed 3 ppm. Numerous
results appear in the range 3-4 ppm.
Oats present a somewhat different problem in that the most effective
results are obtained by treating when the crop has reached a height of
40-50 cm. These late season treatments lead to residues in the grain
of 3.8 ppm and 6.1 ppm from the use of 1.5 kg/ha and 3 kg/ha,
respectively (Jung 1968a, 1969b).
Residues in the whole grain have been reported (American Cyanamid Co.,
1967, 1968a) to range from 0.16-4.2 ppm, depending upon rate of
application, harvest interval and variety of plant.
Experiments which were carried out in Australia (Annand, 1968) with
the object of determining the residues resulting from the use of
chlormequat sprays on grapes at flowering time had a sensitivity of
0.75 ppm. No chlormequat residues could be found in grapes treated
with sprays containing 300 ppm of chlormequat 16 weeks prior to
harvest. Paper chromatography was used as an analytical method,
because it was the most sensitive of the several methods available.
Sultanas made by drying grapes grown with the aid of chlormequat
treatment were also analysed by the same laboratory, but no residues
Residues of 0.2-0.6 ppm have been found in the fruit at harvest, and
similar levels in the finished wine products (American Cyanamid Co.,
1969a, 1969b; Tafuri et al., 1970a).
Fate of residues
Blinn (1967) using 14C-labelled chlormequat showed the compound was
not metabolized in wheat plants or in rats. The unchanged compound was
the only radio labelled material found as residues in wheat foliage,
roots and grain and in rat urine.
It appears that chlormequat is absorbed by wheat foliage but undergoes
no metabolism and very little translocation to the roots. Therefore,
the residue analytical procedure developed by Mooney and Pasarela
(1967) which responds to the parent chlormequat should provide
realistic evaluation of the residual properties of chlormequat plant
growth regulant in wheat foliage and grain.
Blinn (1967) reported that over 96% of the compound was detected
unchanged in the urine and faeces within 48 hours when rats were fed
chlormequat in their diet. Less than 0.5% was respired as carbon
dioxide in the breath, and less than 1 ppm was detected in tissues of
Jung and El-Fouly (1969) applied high rates of chlormequat to wheat
plants and showed that there is a slight increase in the choline
chloride and betain content of the plant tissues 3 and 24 days after
treatment. The authors proposed a theoretical basis for the metabolic
In a study of factors affecting degradation of chlormequat by
wheat-plant extracts (El-Fouly and Jung, 1969), it was suggested that
an enzymatic system might be involved. Bier and Faust (1967), using
chlormequat labelled with nitrogen-15 applied foliarly to intact wheat
plants, showed that there is no metabolism of the compound to betaine,
choline, dimethylchloroethylamine, or trimethylamine. A rate of
disappearance study at a treatment level of 4 pounds of chlormequat
per acre of wheat showed a biological half-life of 13 days (Mooney and
Jung and El-Fouly (1969) showed that the residue in wheat grain
(1.0-2.5 ppm) declined to 0.5 ppm and below after the wheat was held
in store for 12 months. Residues are highest in grain grown under
particularly dry conditions. High rainfall can, apparently, almost
entirely eliminate the detectable residue in grain at harvest. The
residue of chlormequat remaining in the straw of treated cereals is
usually somewhat higher than the residue in the grain.
Straw from treated crops will possibly be used as fodder for sheep and
cattle. Jung (1969b) showed that when oat straw containing 5 ppm and
10 ppm chlormequat was fed to a lactating cow, no residue could be
found in milk. When fodder containing 20 and 40 ppm chlormequat (40
and 80 mg/day) was fed, only traces of less than 0.1 ppm chlormequat
could be traced in the milk. Examination of the urine of the test cow
established that chlormequat was excreted quantitatively in the urine.
Chlormequat residues in soil decompose relatively fast. Cathey and
Stuart (1961) consider the persistency in soil to be only three weeks.
Jung (1965) showed that soil applications were inactivated after 4
weeks at 20°C.
Evidence of residues in food in commerce or at consumption.
No information was available on residues of chlormequat in food
commodities moving in commerce nor have studies been conducted to
determine residues in foodstuffs as consumed. Studies of the effect of
processing and cooking on chlormequat residues appear not to have been
METHODS OF RESIDUE ANALYSIS
A thin layer chromatographic method developed by Jung and Henjes
(1964) was used in evaluating residues in field trials, but its
sensitivity was limited to 0.5 ppm.
Mooney and Pasarela (1967) have described a method for the
chromatographic separation and subsequent colorimetric determination
of chlormequat at residue levels in wheat grain and plants at various
stages of growth. The compound, after extraction, is removed from the
plant tissue background by adsorption on aluminum oxide and measured
colorimetrically as a complex with dipicrylamine at 415 mµ.
Based on both of these procedures, Jung and Henjes, (1969) developed a
further method involving extraction with methanol, isolation by column
chromatography using alumina and an acidic cation exchange resin and
measurement as the dipicrylamine complex. The method is suitable for a
wide variety of plant products, grain, wine and animal organs. The
sensitivity ranges from 0.1 to 0.3 ppm with recoveries of 90-100%.
This method is practically the same as used by Businelli et al. (1969)
for the determination of chlormequat residues in tomatoes and grapes.
Tafuri et al. (1970b) have reported a method for the gas
chromatographic estimation of chlormequat at residue levels based upon
a reaction with sodium benzenethiolate which converts chlormequat to
1-phenylthio-2-dimethylaminoethane. Tafuri et al. (1970a) have used
both the colorimetric and gas chromatographic methods for the
determination of chlormequat residues in grapes and wine products.
grapes and dried vine fruit - 0.75 ppm
Chlormequat has been used during the past five years as a plant growth
regulator to reduce the risk of lodging in wheat, rye and oats. It is
applied to the growing wheat and rye when the first node can be felt
in the majority of tillers and in oats when the second node can be
felt. It is also used on grape vines to aid in the setting of the
grape harvest and for increasing yields.
Chlormequat is sold as an aqueous solution containing from 12% to 40%
w/v active ingredient. In parts of Europe, a formulation containing
46% chlormequat and 32% choline chloride is marketed. The addition of
choline chloride reduced the acute toxicity of chlormequat to a range
of laboratory animals but does not appear to modify the mode of action
of chlormequat in plants.
Technical chlormequat used in preparing commercial formulations is 97
to 98% pure 2 chloroethyltrimethylammonium chloride.
The data available to the meeting was obtained from published
literature and from the major European and American manufacturers.
Residue data were available from supervised trials carried out in a
number of European countries, Kenya, New Zealand and Australia.
Available data indicate that residues of unchanged chlormequat may
occur in the straw and grain of treated small grain crops, especially
wheat, rye and oats. Residues in wheat appear to lie mostly below 1
ppm, but some samples, especially from crops grown under dry
conditions, have residues ranging up to 22 ppm. Residues in rye appear
somewhat higher, ranging up to 4 ppm. Residues in oats appear higher
still, because treatment is applied when the crop is in a more
advanced stage of development when the interval between application
and harvest is much less. Residues in oats may range as high as 6 ppm.
The feeding of straw from treated crops to dairy cows does not give
rise to detectable residues in milk, nor is it to be anticipated that
residues could occur in edible tissues of ruminants receiving such
plant materials as forage.
A specific method of analysis suitable for determining residues of
chlormequat in cereal grain, plant products, fruit and vegetables and
animal tissues at a sensitivity of about 0.2 ppm has been published.
This method appears suitable from regulatory purposes as the
extraction and cleanup procedure makes it highly specific.
The following is an indication the maximum residues which will occur
in specified food commodities following approved use of chlormequat.
Residues will occur at this level only in some treated crops. The
residue levels are not expected to decline significantly during
storage of treated crops following harvest.
Raw grains (rye and oats) 5 ppm
Raw grain (wheat) 2 ppm
Grapes and dried vine fruits 1 ppm
FURTHER WORK OR INFORMATION
REQUIRED (before an acceptable daily intake for man can be
Full reports on the biochemical and toxicological studies conducted on
1. Information on other registered uses for chlormequat
2. Further information on residues in new agricultural commodities
from a number of additional countries
3. Information on the effect of milling and preparation on the level
of residues in grain products
4. Information on chlormequat residues in commodities moving in
5. Analytical methods capable of recovering and determining
chlormequat residues in plant and animal products at levels down
to at least 0.1 ppm, to be established for regulatory purposes.
Annand. (1969) Residues of chlormequat in grapes and dried vine
fruits. Submission to National Health and Medical Research Council,
Blinn, R.C. (1967) Plant Growth Regulant - Biochemical Behaviour of
Chlormequat in Wheat and Rats. J. Agric. and Fd. Chem., 15 (6):
Cathey, H.M. and Stuart, N.W. (1961) Comparative plant growth
retarding activity of Amo 1618, Phosphon and CCC; Botan. Gaz., 123:
Coombe, B.G. (1965) Increase in fruit set in Vitis vinifera by
treatments with growth retardants, Nature, 205: 4968
Jung, J. (1964) Analytical research on wheat samples of CCC-trials.
Landw. Forschg., 17; 267
Jung, J. and Henjes, G. (1964) Proof and half-quantitative
determination of chlorcholinchloride (CCC) in wheat grain and straw.
Zeitschr. f. Pflanzenernährg., Düngg. u. Bodenkde., 106: 108
Jung, J. (1965) Behaviour of CCC in plants and soil. CCC-Symposium of
the BASF, Limburgerhof
Jung, J. (1968) CCC-residue in rye according to trial plan DIII and
DIIId. Internal BASF-Information
Jung, J. (1968a) Residue trials with CCC-treated oat. Internal
BASF-Information No. 581, Limburgerhof
Jung, J. and El-Fouly, M.M. (1969) On the decomposition of chlorine
choline chloride (CCC) in the plant. Zeitschr. f. Pflanzenernährg.,
Düngg. u. Bodenkde., 114: 128
Jung, J. (1969a) Personal information to Herrn Professor Dr. Siegel,
Jung, J. (1969b) Results of the residue examination of CCC treated oat
from the vegetation period 1968. Internal BASF-Information No. 617,
Jung, J. and Henjes, G. (1969) Determination of the growth regulators
CCC (chlorcholinchloride) and CMH
(N-dimethyl-B-ethyl-chloride-hydrazone chloride) in biological material
Mooney, R.P. and Pasarela, N.R. (1967) Determination of
chlorcholinchloride residues in Wheat grain, straw and green wheat
foliage. J. Agric. Fd. Chem., 15: 989
An extensive bibliography containing over 280 references to papers on
the properties, uses and mode of action of chlormequat has been
published by American Cyanamid Company Wayne, New Jersey, U.S.A.,
entitled "CYCOSTAT" Plant Growth Regulant