Autalyst, July, 1968, Vol. 93, pp. 443444 443 A Rapid Method for the Determination of Malathion in Wheat Grains BY E. WEISENBERG, S. GERTNER AND J. SCHOENBERG (Institute of Control and Standardisation of Drugs, Ministry of Health, Jerusalem, Israel) A simple method for the determination of malathion in wheat grains is described. The insecticide was extracted with chloroform and the extract treated with a Celite - Nuchar - sodium sulphate mixture to absorb impur- ities selectively. The hydrolysis was carried out with ethanolic sodium hydroxide solution and the copper complex extracted into cyclohexane. The method is applied to amounts of 50 to 250 pg. THE use of malathion, which has been widely used for the control of pests in stored grains, has been based on its relatively strong pesticidal qualities and its low toxicity to mammals.The method described by Norris, Easter, Fuller and Kuchar,l based on the alkaline decom- position of malathion, has been recommended for the determination of residues in cereals and oil seeds.2 Bates, Rowlands and Harris3 used chromatography on Fuller’s earth for the purification of the plant extracts before analysis. In a later publication, Bates and Rowland9 found that acceptable recoveries were being obtained without the preliminary use of an absorbent column. Uphams reported a modification of this method for the determination of malathion in formulations, in which carbon tetrachloride was replaced by cyclohexane, and the colour stability was found to have improved. In an attempt to improve the accuracy and precision of the method, a critical study was made by Orloski.6 We adapted and modified this method in order to determine amounts as low as 50 pg.Chloroform was selected as the extracting solvent; according to Upham,s the descending order of solvent power for malathion was found to be chloroform, carbon tetrachloride and carbon disulphide. As a preliminary purification of the chloroform extract was found necessary, we effected selective absorption of the impurities by Celite - Nuchar - sodium sulphate as recommended by Koivistoinen, Karinpaa, Kononen and Roine.7 This treatment eliminated the need for repeated washings of the plant extract, which were found to be criticaL2 REAGENTS- METHOD Chloroform, B.P. quality. Acetonitrile, analytical-reagent grade.Cyclohexane, analytical-reagent gmde. Ethanolic sodium hydroxide solution, about 0.5 N-Dissolve 1 g of analytical-reagent grade Iron(II1) chloride - hydrochloric acid solutiow---Dissolve 0.2 g of iron(II1) chloride in Copper sulphate solution, 1 per cent. w/v. Absorbent for clean-up-This consisted of 1 part of Celite 545 (Johns-Manville) , plus 2 parts of Nuchar C-190-N (West Virginia Pulp and Paper Co., Covington, Virginia) plus 1 part of anhydrous analytical-reagent grade sodium sulphate. Ethanol, absolute, analytical-reagent grade. PROCEDURE- Shake mechanically 150 mg of coarsely ground wheat grains with 300 ml of chloroform for 24 hours. Filter the mixture through a Buchner funnel and wash the residue in the funnel with three 50-ml portions of chloroform.Evaporate the extract and washings to 200 rnl in a Rinco vacuum evaporator. Transfer a 100-ml aliquot into a 250-ml Erlenmeyer flask, add 4 g of the Celite - Nuchar - sodium sulphate mixture and shake it for 5 minutes. sodium hydroxide in 50ml of absolute ethanol by heating under reflux. 8 ml of hydrochloric acid (sp. gr. 1-18) and make up to 1 litre with water. 0 SAC and the authors.444 WEISENBERG, GERTNER AND SCHOENBERG Filter through paper, wash the first flask and the residue on the filter six times with 10-ml portions of chloroform and add these washings to the filtrate, then evaporate the filtrate in a vacuum evaporator to about 10 ml. Transfer it to a small beaker, evaporate to dryness with a current of hot air and dissolve the residue in 8 ml of ethanol; transfer to a 125-ml separating funnel and wash the beaker with 5 ml of cyclohexane and add to the ethanol.To the contents of the separating funnel add 0.2 ml of acetonitrile and 1 ml of ethanolic sodium hydroxide solution; swirl the funnel gently (do not shake) for 5 to 10 seconds and let it stand for 2 minutes. Add 25 ml of iron(II1) chloride solution (cooled to 10” C), mix well by swirling it for 10 seconds and let it stand for 5 minutes to allow the phases to separate; discard the cyclohexane. Add exactly 10 ml of cyclohexane and 1 ml of copper sulphate solu- tion and shake the separating funnel immediately for 1 minute. Allow the phases to separate and, as soon as separation occurs, discard the aqueous phase and filter the cyclohexane solution through a small filter containing 0.1 g of anhydrous sodium sulphate.Measure the optical density of the yellow solution, within 15 minutes from the beginning of the hydrolysis, at 420 mp in a cell of l-cm path length. Prepare a standard graph from pure malathion to cover the range of 50 to 250pg of malathion. The line has a slope of 1.48 optical density units for 1 mg of malathion. RESULTS AND DISCUSSION In the samples of wheat grains analysed the malathion content was found to vary The reliability of the method was studied by recovery tests performed on an untreated from 0 to 4 p.p.m. sample, and the results are presented in Table I. TABLE I RECOVERY OF MALATHION FROM WHEAT GRAINS Amount added, p.p.m. Amount found, p.p.m. Recovery, per cent. 1.04 0.96 92 2-05 2.03 97 4.16 3.96 95 6.24 5-73 92 8.67 8.33 96 From these results we found that satisfactory recoveries were obtained.We found that the hydrolysis with ethanolic sodium hydroxide in the presence of cyclo- hexane was advantageous. The last traces of coloured impurities remained in the cyclohexane layer after the addition of the iron(II1) chloride solution, and the aqueous solution remained colourless and clear. The stability of the copper complex in cyclohexane has been found satisfactory. It is possible that the stability of the colour is connected with the fact that in our method a clear solution free from impurities was obtained directly after the hydrolysis. The calibration graph was found to be reproducible and, therefore, it was unnecessary to run daily standards. It is proposed to study the applicability of the method to a range of foodstuffs in which malathion residues may be present. We gratefully acknowledge the contribution made by Mrs. S. Gershson, in adapting the Orloski method for the determination of micro amounts of malathion. REFERENCES 1. 2. 3. 4. 5. 6. Orloski, E. J., Ibid., 1964, 47, 248. 7. Norris, M. V., Easter, E. W., Fuller, L. T., and Kuchar, E. J., J . Agric. Fd Chem., 1958, 6, 111. Analytical Methods Committee, “The Determination of Malathion Residues in Cereals and Oil- Bates, A. N., Rowlands, D. G., and Harris, A. H., Ibid., 1962, 87, 643. Bates, A. N., and Rowlands, D. G., Ibid., 1964, 89, 286. Upham, S. D., J . Ass. Off. Agric. Chem., 1960, 43, 360. Koivistoinen, P., Karinpaa, A., Kononen, M., and Robe, P., J . Agric. Fd Chem., 1964, 12, 561. Received December 6th, 1967 seeds,” Analyst, 1960, 85, 915.