Evaluation and Characterization of a Commercial Immunosorbent Cartridge for the Solid-Phase Extraction of Phenylureas from Aqueous Matrices

Journal of Chromatographic Science, Vol. 41, October 2003 Evaluation and Characterization of a Commercial Immunosorbent Cartridge for the Solid-Phase Extraction of Phenylureas from Aqueous Matrices Departamento de Química Analítica, Facultad de Química, Universidad Nacional Autónoma de México, 04510 México D.F., México Abstract The behavior and main characteristics of a commercial immunosorbent (IS) cartridge for the solid-phase extraction of phenylureas are determined in this work. The measured capacity for the analyte–antigen (isoproturon) in a new cartridge is 215 ng and, after more than 100 adsorption–desorption cycles, the remaining capacity still is approximately 70 ng, demonstrating the good stability of the bonded antibody and the interesting possibility of extensive cartridge reuse. Only isoproturon and diuron are specifically retained in this sorbent. The weak nonspecific retention of other pesticides, including other phenylureas, can be avoided by increasing the sample volume during the loading step. Thus, a very selective and sensitive method for the determination of isoproturon and diuron in natural and potable waters is developed by loading a 50-mL sample adjusted to pH 7.4 in the IS cartridge, eluting with methanol–water (60:40, v/v), and analyzing the eluate by highperformance liquid chromatography with UV detection. The clean chromatograms, low detection limits (~ 0.1 µg/L), and good precision (< 5%) obtained with this rapid and simple method demonstrate that immunoaffinity extraction can be an excellent alternative for sample preparation in the environmental monitoring of particular pesticides in water matrices. Introduction The development of new selective sorbents for the extraction, preconcentration, and cleanup of pesticides, toxins, and other pollutants from complex samples has been the object of numerous research works in recent years. The interest has been particularly directed toward materials that allow the structure recognition of the target analytes, such as the immunosorbents (IS) (1–7) and molecularly imprinted polymers (MIP) (8–10). Thus, considerable efforts have been made to optimize all stages of the sorbent preparation process and to study and define the relevant parameters affecting the extraction/elution of the analytes (1,4–7,9). Until the Author to whom correspondence should be addressed: email . 480 present, the IS technology has reached a higher degree of maturity, as testify the reported applications of these sorbents in the solid-phase extraction (SPE) of trace pollutants from environmental samples and food extracts (11–14). Indeed, analytical methods for the determination of herbicides in water, using experimental antiphenylurea or antiatrazine IS precolumns coupled to high-performance liquid chromatography (HPLC) with various detection modes, have already been validated (15–16), and the first commercial IS cartridges for the SPE of aflatoxins, phenylureas, and triazines have appeared in the market in the last few years. Besides, some excellent reviews dealing with the online or offline SPE of low-molecular-mass analytes using IS have been published (17–19). Comparatively, the preparation and application of MIP for environmental analyses is still at a research stage, but because of their potential advantages (concerning reproducibility, sample capacity, stability, and cost), great improvements are expected in this area in the near future (20,21). The basis of immunoaffinity extraction is the highly selective antigen–antibody interaction. Antibodies raised against a specific compound can be immobilized by covalent bonding to the surface of an adequate support (1,3,4,12) or, alternatively, they can be encapsulated into the pores of a solid matrix (5–7). These biomaterials will selectively retain the analyte–antigen present in, for example, a surface water sample, thus effecting extraction, preconcentration, and cleanup in the same step (17). However, because of the unavoidable cross-reactivity of antibodies, especially when they are raised against a small molecule with few determinant groups (as is the case for most pesticides), other structurally related compounds may also be retained by the IS. This cross-reactivity has been exploited to develop class-selective sorbents that can be used for the simultaneous SPE of several members of the same chemical family (12,14–18). It has been stated that sample preparation methods using IS also result in enhanced sensitivity of analysis for complex samples because, as the obtained extracts are in principle free of most matrix interferences, it is possible to use highly sensitive detection conditions (11,13). Despite the interesting advantages of immunoaffinity extraction, its application in the environmental field still is restricted to Reproduction (photocopying) of editorial content of this journal is prohibited without publisher’s permission. Luz E. Vera-Avila*, Laura Rangel-Ordoñez, and Rosario Covarrubias-Herrera Journal of Chromatographic Science, Vol. 41, October 2003 Experimental Reagents and materials LC-grade acetonitrile and methanol were from Prolabo (Paris, France). Type 1 reagent water was obtained from a Nanopure (Barnstead International, Dubuque, IA) deionizer. All other chemicals (sodium azide; sodium chloride; potassium chloride; phosphoric acid; sodium hydroxide; and perchloric, formic, and acetic acids) were analytical-grade reagents from various furnishers; they were used without further purification. The phenylurea herbicides (isoproturon, diuron, fluometuron, monuron, linuron, and neburon) and two other pesticides (methiocarb and parathion-methyl) were obtained from Chem Service (West Chester, PA) with certified purity of 99%. Stock solutions (1000 mg/L) of each herbicide were prepared in acetonitrile and stored at –20°C when not in use. Working standards were prepared by dissolving appropriate aliquots of the stock solutions in acetonitrile–water (40:60, v/v) or in phosphate-buffered saline solution (PBS), depending on the experiments. PBS was 0.02M (in phosphate), unless otherwise indicated. It was prepared by dilution of the adequate phosphoric acid volume in reagent water, addition of NaCl (0.137M) and KCl (0.0027M), and adjustment to pH 7.4 with a sodium hydroxide solution. For some experiments requiring other buffer concentrations, the PBS was prepared keeping the same phosphate–NaCl–KCl molar ratio. The immunosorbent cartridges were purchased from Abkem (Ottawa, Canada). A package of ten ImmunoSep phenylurea cartridges (same lot) was used throughout this work. The cartridges are guaranteed for retention of 450 ng of isoproturon and for three uses without alteration. According to information given in the furnisher catalog, polyclonal antibodies raised against isoproturon were covalently bonded to a modified silica support to prepare the IS. The specified cross-reactivity of this sorbent for the phenylureas considered in this work, with respect (...truncated)