Immunoassay of pesticides: an update.

Measurement of levels of pesticides residues in foods and crops most often requires extensive cleanup and instrumental techniques such as gas chromatography. Immunoassay measurement techniques, on the other hand, may be used directly on the test portion or require only minimal cleanup. Further refinements of the common antibody-enzyme-based solid-phase assays, such as use of coated magnetic particles, antibody-coated crystals, and continuous-flow devices, have extended the measurement range and applicability of these assays. Likewise, new immunoassays for pesticides have been developed, and existing assays have been refined, optimized, and more completely characterized and validated. In addition to their ability to accurately and reliably measure amounts of residues present in food and crops, immunoassays can be readily used as rapid screening methods for contaminants in field samples. We have previously reviewed much of the work in the area of pesticide immunoassay; this report updates previous information and discusses some new immunoassay techniques.

Modification of the AOAC method for determination of fumigants in wheat.

The official first action AOAC method for determination of residues of fumigants in grains has been modified for use by 6 laboratories in an FDA pesticide surveillance program. A 15% OV-17 gas-liquid chromatographic column, installed in a chromatograph equipped with a constant current 63Ni electron capture detector, provides for improved resolution, multiresidue capability, and lower limits of detection. A study was made of the behavior of chloroform, carbon tetrachloride, ethylene dichloride, trichloroethylene, tetrachloroethylene, and ethylene dibromide through the method. Recoveries from fortified wheat samples averaged 105-115%. Experimental evidence is presented which suggests that part of the consistent trend toward high recovery can be attributed to selective sorption of acetone by the CaCl2 used in the final drying step.

Immunoassay of pesticides.

Determination of the presence and levels of pesticide residues in food is fundamental in monitoring and regulatory programs. Residues are separated from the food matrix by solvent extraction, followed by cleanup steps. The residues are most often identified and quantitated by instrumental analysis, usually liquid or gas chromatography. Extraction and cleanup are often laborious and time-consuming; determination requires expensive, sophisticated instrumentation. There is a need for rapid, easily performed tests, such as immunoassays, that could be used for screening under field conditions or for quantitation of residues in foods in the laboratory. Although a large number of immunoassays have been developed for pesticide chemicals, very few have been specifically applied to foods, and only a very small number are currently available commercially. The agencies charged with monitoring and regulatory responsibilities--Environmental Protection Agency, Food and Drug Administration, and U.S. Department of Agriculture--as well as professional societies such as AOAC and the International Union of Pure and Applied Chemistry, are investigating and developing guidelines for test kit evaluation and standards to be met before a kit can be accepted as a practical and useful method of analysis for use in their programs.

Method for determination of organohalogen pesticide residues in vegetable oil refinery by-products.

A method using gel permeation and Florisil column chromatographic cleanup techniques is described for determination of residues of nonpolar organohalogen pesticides and pesticide alteration products in vegetable oils and their refinery by-products. Supplemental Florisil separation and alkali cleanup techniques are used to facilitate determinations. Residues are determined with a 63Ni electron capture gas chromatographic detection system used in conjunction with 3 different gas chromatographic columns. Residue identities are confirmed by gas chromatography-mass spectrometry. Recoveries of 7 organohalogen pesticides, ranging from 90 to 103%, were determined by the supplemental Florisil separation technique to augment previously reported recovery data determined for initial GPC and Florisil cleanup steps. Soybean, peanut, and cottonseed deodorizer distillates and crude and refined oil, as well as additional refinery by-products, were analyzed. Nine to 13 organohalogen residues ranging from 0.5 to 6.3 ppm were determined in the 2 soybean deodorizer distillate samples used to develop and test the method. Identities of residues present at greater than or equal to 0.3 ppm were confirmed by gas chromatography-mass spectrometry. An intralaboratory trial of the method provided additional recovery and residue determination data as follows: Recoveries ranging from 102 to 116% were obtained for 4 pesticides added to peanut oil deodorizer distillate. Residues determined in 1 soybean deodorizer distillate sample supported previously obtained data for this sample.

Comparison of methodology for determination of ethylene dibromide in grains and grain-based foods.

Three commonly used methods for determination of ethylene dibromide (EDB) in grains and grain products have been compared. EDB residues were extracted by soaking in hexane, triple co-distillation with hexane from an aqueous sample solution, and soaking in acetone-water (5 + 1). Each method was used for triplicate analyses of 12 samples containing incurred residues of EDB ranging from about 10 to 1000 ppb and representing whole grains (wheat and oats) and intermediate grain-based products such as corn meal and flour. The 4-day hexane soaking method extracted the least EDB. In some cases, this was half of the amount determined by the other methods. Levels from soaking in acetone-water were equal to, or up to 25% greater than, those from distillation. Although soaking for 2 days is required for whole grains in the method, a period of only 16 h was found acceptable for ground products. Results were obtained faster with the distillation method, but more analyst time per sample was required. A single distillation recovered about 80% (40-60% from wheat) of total EDB extracted by triple distillation. Foaming was reduced by the addition of concentrated H2SO4 to the aqueous hexane-sample mixture, plus stirring during distillation, thereby allowing complete recovery of the hexane.

Effect of cooking on levels of ethylene dibromide residues in rice.

Two studies were conducted to determine the effect that cooking has on the level of residues of ethylene dibromide (EDB) in rice. In the first study, 4 samples of long and medium grain polished white rice containing 113, 295, 956, and 1568 ppb EDB were cooked according to typical label directions. Three batches of cooked rice were prepared from each sample of polished rice and frozen until analysis; each batch was analyzed in duplicate. EDB levels in all cooked rice samples were less than 10 ppb. In the second study, conducted jointly by the Food and Drug Administration (FDA) and the Environmental Protection Agency (EPA), a sample of medium grain polished white rice containing about 1600 ppb EDB was cooked by each laboratory. Overall average EDB levels in rice analyzed immediately after cooking were 16 and 37 ppb for FDA and EPA, respectively. The corresponding frozen samples contained 8 and 39 ppb EDB. The 2 laboratories exchanged these frozen samples and reanalyzed them to check variability in the analytical procedure. FDA found 49 ppb EDB in the sample cooked by EPA and EPA found 8 ppb EDB in the sample cooked by FDA, thus indicating that analytical methodology was not a major source of variability. The range of EDB levels was therefore attributed to minor differences in the way the rice was cooked or handled immediately after cooking.

Elevated TCDD in chicken eggs and farm-raised catfish fed a diet with ball clay from a Southern United States mine.

The U.S. Food and Drug Administration (FDA) terminated the use of ball clay from a mine in Mississippi as an additive in animal feed after discovering nanogram per gram concentrations of 2,3,7,8-tetrachlorodibenzo-p-dioxin (2,3,7,8-TCDD). The FDA collected chicken eggs and farm-raised catfish in affected areas and throughout the remaining continental United States to assess levels of 2,3,7,8-TCDD. A new method using quadrupole ion storage tandem-in-time mass spectrometry (QISTMS) measured the 2,3,7,8-TCDD levels in 42 catfish fillet composites, 3 Tilapia fillet composites, 46 chicken egg samples, and 6 chicken feeds. Six catfish composites and 20 egg samples had 2,3,7,8-TCDD concentrations significantly above 1.0 pg/g wet weight of fillet or whole egg. Farm-raised catfish not exposed to feed containing ball clay had a mean 2,3,7,8-TCDD concentration of 0.12 pg/g. The TCDD isomer pattern in ball clay differed from the TCDD isomer pattern in a fly ash sample and from the "chick edema factor" TCDD pattern in a sample of reference toxic fat used as a feed ingredient in the 1950s.