AGP:1970/M/12/1 WHO/FOOD ADD/71.42 1970 EVALUATIONS OF SOME PESTICIDE RESIDUES IN FOOD THE MONOGRAPHS 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 Rome, 1971 CHLORMEQUAT IDENTITY Chemical name 2-chloroethyltrimethyl-ammonium ion (usually as the chloride) Synonyms chlorcholine chloride, Cycocel(R), CCC Formula [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 effectiveness. 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 Use pattern 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. TABLE I Chlormequat residues in grain from crops treated to prevent lodging Chlormequat Interval - Country Type of Crop kg/ha treatment to Residues harvest-weeks ppm 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 Sweden 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 1965 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 harvest-weeks ppm 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 1969 Wheat 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. Rye 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 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. Grapes 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 were detected. 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 sacrificed rats. 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 pathway. 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 Pasarela, 1967). 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 carried out. 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. NATIONAL TOLERANCES Australia grapes and dried vine fruit - 0.75 ppm APPRAISAL 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. RESIDUES 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 established) Full reports on the biochemical and toxicological studies conducted on chlormequat DESIRABLE 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 international trade 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. REFERENCES Annand. (1969) Residues of chlormequat in grapes and dried vine fruits. Submission to National Health and Medical Research Council, Australia Blinn, R.C. (1967) Plant Growth Regulant - Biochemical Behaviour of Chlormequat in Wheat and Rats. J. Agric. and Fd. Chem., 15 (6): 984-988 Cathey, H.M. and Stuart, N.W. (1961) Comparative plant growth retarding activity of Amo 1618, Phosphon and CCC; Botan. Gaz., 123: 51 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, LUFA, Speyer Jung, J. (1969b) Results of the residue examination of CCC treated oat from the vegetation period 1968. Internal BASF-Information No. 617, Limburgerhof 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 BIBLIOGRAPHY 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
See Also: Toxicological Abbreviations Chlormequat (WHO Pesticide Residues Series 2) Chlormequat (Pesticide residues in food: 1976 evaluations) Chlormequat (Pesticide residues in food: 1994 evaluations Part II Toxicology) Chlormequat (Pesticide residues in food: 1997 evaluations Part II Toxicological & Environmental) Chlormequat (JMPR Evaluations 1999 Part II Toxicological)