VALIDATION OF AN IMMUNOASSAY METHOD FOR THE DETERMINATION OF TRACES OF CARBARYL IN VEGETABLE AND FRUIT EXTRACTS BY LIQUID-CHROMATOGRAPHY WITH PHOTODIODE-ARRAY AND MASS-SPECTROMETRIC DETECTION

A competitive enzyme-linked immunosorbent assay (ELISA) method for car baryl quantitation in crop extracts was validated by liquid chromatogr aphy (LC) with diode array detection (DAD). For this purpose, six crop s (banana, carrot, green bean, orange, peach and potato) were chosen f or recovery and reproducibility studies. The general sample preparatio n included extraction with methanol followed by liquid-liquid partitio ning and clean-up on Celite-charcoal adsorbent column of the vegetable extracts. ELISA samples consisted of a diluted LC extract in assay ph osphate buffer (pH 7.5). The potential effect of methanol in these sam ples was evaluated. It was observed that a maximum content of 10% meth anol present in the assay buffer could be tolerated without expressive losses in the ELISA performance. Under these conditions, a IC50 simil ar to 1.48 mu g l(-1) was obtained. A minimum matrix effect with a 1:5 0 dilution of the methanolic extracts in assay buffer was noticed, exc ept for green bean samples that inhibited completely the assay. For th e vegetable extracts, the ELISA sensitivities varied from 3.9 to 5.7 m u g l(-1), and good recoveries (82-96%) with R.S.D.s ranging from 5.7 to 12.1% were found. An excellent correlation between the LC-DAD and E LISA techniques was obtained. The confirmation of the carbaryl in less concentrated samples was achieved by LC-mass spectrometry interfaced with atmospheric pressure chemical ionisation. The [M + H](+)= 202 and [M + H-57](+)=145 ions, equivalent to the protonated molecular and l- naphthol ions, respectively, were used to carbaryl identification in t hese samples. (C) 1998 Elsevier Science B.V. All rights reserved.