Malcolm C. Bowman

Stressed bioassay systems for rapid screening of pesticide residues. Part I: Evaluation of bioassay systems.

An investigation of bioassay systems employingDaphnia, Hyalella, Culex, orPalaemonetes was initiated in response to the need for rapid, sensitive, and inexpensive screening procedures for toxicants in food and feed. The procedures may also be useful to determine whether farm workers may safely reenter fields treated with pesticides. The evaluation of inherent toxicities related to types and amount of organic solvents commonly used in such systems indicated that dimethyl sulfoxide (DMSO) and methanol (MeOH) were least toxic in the aqueous test media. These solvents were then used in 18-hr tests to determine sensitivities of the four organisms to a representative compound from six classes of pesticides,i.e., dieldrin, parathion, carbaryl, atrazine,o-phenylphenol, and maneb.Stress factors such as the amount of organic solvent and volume of test medium were adjusted to determine their effects on three of the organisms tested against dieldrin and parathion. The highest sensitivity obtained with dieldrin (50% mortality with 2 ng in a 25-ml test medium) was withCulex stressed with 2% of MeOH in a reduced test volume.Hyalella stressed with 2% of MeOH were most sensitive to parathion (50% mortality with 85 pg in a 100-ml test medium); further stress imposed by reducing the volume of test medium diminished sensitivity.Daphnia, which were also very sensitive to dieldrin and parathion, were most sensitive to the other four pesticides which exhibited low toxicities to all test organisms.Daphnia would, therefore, be the organism of choice if only one could be used.The amounts of carbaryl, atrazine,o-phenylphenol, and maneb required to yield 50% mortality withDaphnia in 25-ml test systems (0.4% MeOH) were about 0.05, 15, 47 and 2.4μg, respectively. Preliminary experiments with various extracts of animal feed indicate that an extensive effort will be required to develop a method that will provide extracts that are compatible with the bioassay systems.

Carcinogens and Analogs: Trace Analysis of Thirteen Compounds in Admixture in Wastewater and Human Urine

A gas chromatographic method is described for determining traces of 13 carcinogens and related compounds (aromatic amines and estrogens) in admixture in wastewater and human urine. This method was developed for use in toxicological research for monitoring the safe disposal of wastewater and to signal any accidental exposure of personnel to hazardous test substances. Salient elements of the procedure are: extraction of phenolic and neutral residues from the acidified sample, liquid--liquid partitioning cleanup and separation of neutral from phenolic residues at pH 14 and 10.2, acid hydrolysis of the neutral component, subsequent alkalinization of the sample and extraction of the basic residues as the free amines, conversion of all residues to the corresponding pentafluoropropionyl (PFP) derivatives and quantification by electron-capture gas chromatography. Residues were detectable in wastewater and urine at the 0.1 and 1 ppb levels, respectively. Additional information is provided concerning partition values for all PFP derivatives in five solvent systems, structure verification of the derivatives by mass spectrometry and the adaption of this method to the monitoring of surfaces and air in potentially contaminated work areas.

A toxicological risk assessment procedure: A proposal for a surveillance index for hazardous chemicals

Health-oriented government agencies responsible for the protection of the public from possible adverse effects, such as that posed by chemical residues in the food supply, must establish priorities on regulatory action as well as for manpower to conduct monitoring programs. A new procedure entitled Surveillance Index (SI) is presented as a technique to evaluate toxic materials that are potential candidates as residues in food supplies and to assign an index number that identifies a relative hazard to the public among the various residues. The SI numbers can be used as criteria by which chemical residues should be monitored whenever conditions such as time and available resources are limiting factors.

4-aminobiphenyl, 2-naphthylamine, and analogs: analytical properties and trace analysis in five substrates.

Methods for monitoring trace levels of 4-aminobiphenyl, 2-naphthylamine, and their hydrochloride salts in waste water, microbiological growth media, potable water, human urine, and mouse blood utilizing spectrophotofluorometry (SPF) are described. The salient elements of the methods are extraction of the residues as the free amine with benzene, rapid cleanup on an alumina column, and quantification of the free amine in methanol via SPF. Potable water solutions of the salts are diluted with 0.01 N aqueous HCL and quantified directly by SPF. Ancillary analytical information concerning gas chromatography of the free amines, partitioning properties of the compounds between solvent pairs, their solubility and stability in water, and thin-layer chromatographic data is presented. The compositions of various admixtures of 1- and 2-naphthylamine or their salts were determined by using SPF with calculations based on simultaneous equations.

Analysis of 2-acetylaminofluorene: Residues in laboratory chow and microbiological media

Abstract Analytical methods are described for determining residues of 2-AAF in lab chow and microbiological growth media by FID-GC and SPF. Solubility values for 2-AAF in 14 solvents were determined. Ancillary data concerning p-values, GC and SPF characteristics, and adsorption liquid chromatographic properties of 2-AAF, 2-AF, and fluorene are also presented.

Trace analysis of 3,3'-dichlorobenzidine in animal chow, wastewater and human urine by three gas chromatographic procedures.

Gas chromatographic methods are described for the trace analysis of 3,3′-dichlorobenzidine and its dihydrochloride salt in animal chow, wastewater, and human urine. Salient elements of the method for these known carcinogens in chow are: Extraction of the residues as the free amine and a cleanupvia acid-base liquid-liquid partitioning with benzene followed by a silica gel column. With wastewater and human urine, residues are adsorbed by percolating the sample through a column of XAD-2, eluted with acetone, and cleaned up with acid-base partitioning and a silica gel column. Residues are assayed by gas chromatography (GC) either as the free amine or after conversion to the pentafluoropropionyl (PFP) derivative by using an electron capture (EC) or a rubidium-sensitized thermionic-type (N/P) detector. Minimum detectable residues in chow, wastewater, and human urine are about 3 ppb, 18 ppt and 60 ppt, respectively, as determined by EC-GC of the PFP derivative.

Determination of sodium saccharin in animal feed, wastewater and human urine by high-pressure liquid chromatography.

Analytical methods are described for sodium saccharin in animal feed, wastewater and human urine at levels as low as 10, 0.1 and 10 ppm, respectively. Samples of animal feed and wastewater are subjected to liquid-liquid partitioning then the feed is further cleaned up on a column of silica gel prior to analysis by high-pressure liquid chromatography (HPLC) using a paired-ion mobile phase and an ultraviolet detector set at 230 nm. Samples of human urine require a cleanpu on a column of XAD-2 prior to the partitioning and silica gel steps as well as an adjustment in the composition of the mobile phase to quantify saccharin. Data concerning partition values and the stability of sodium saccharin in animal feed are also presented.

Removal of trace levels of 2-acetylaminofluorene (2-AAF) from wastewater

An adsorption system is described for the removal of part per billion levels of the chemical carcinogen, 2-acetylaminofluorene (2-AAF), from industrial wastewater. The system consists primarily of filters and activated carbon and non-ionic polymeric adsorbents arranged in tandem. It is highly efficient, operates at low cost, and requires minimal attention. The chemical monitoring of the raw and/or cleaned-up wastewater is based on a highly sensitive and specific spectrophotofluorometric method that allows acceptance or rejection of samples at the 0.2 part per billion level. The system is presented as a model for evaluating the removal of traces of organic chemicals from wastewater prior to recycling or discharging it into the environment. Results of laboratory evaluations of several other approaches to the purification of 2-AAF-containing wastewater are also presented.

Stressed bioassay systems for rapid screening of pesticide residues. Part II: Determination of foliar residues for safe reentry of agricultural workers into the field

A simple bioassay procedure employingDaphnia orHyalella that can be conducted in the field is described as an alternative to chemical analysis to determine safe reentry into crops treated with pesticides. Dislodgeable residues on the foliage of peach trees were determined chemically and with bioassays using the two organisms at various intervals after sequential sprays of azinphosmethyl, phosmet, and carbaryl. Results obtained by the parallel chemical and bioassay procedures were in excellent agreement. Ancillary chemical data are also presented on the percentage of total residues dislodgeable with water from foliage and soil and uniformity of residues on the foliage in a peach orchard.

Control of Test Substances

The efficient control of chemicals during a nonclinical laboratory study is of the utmost importance from three main points of view: (1) integrity of the experiment, (2) safety of personnel, and (3) protection of the environment. While the general principles involved in chemical control may be set forth in formal GLPs, detailed problems concerning individual test substances can only be resolved by utilizing the competency and judgment of a researcher who possesses adequate analytical chemical methods. Integrity of an experiment requires knowledge of the identity, purity, and stability of the test substance, as well as its proper handling and storage and a knowledge of its chemical properties. Assurance of homogeneity, stability, and proper concentration of the chemical in the dosage form is also of primary importance. The development and use of adequate analytical chemical procedures for determining the test agent in human samples, air, clothing, and on work surfaces is necessary for the initiation of surveillance procedures to assure safety of personnel from exposure to the test substances. The environment must be protected from hazardous test substances by using adequate disposal techniques for the chemical and all contaminated materials. This usually involves development of wastewater cleanup systems and continuous monitoring of the effluent to prevent discharge of hazardous substances. Incineration is the most useful method for disposal of solids and highly contaminated liquids. Work from our laboratory, done during the past 5 years, provides an illustration of the important principles, problems, and pitfalls encountered in chemical control.