Fluorescence of pesticides

Initial Measurements. The data in Table 2 for the fluorescence of pesticides in hexane and methanol were obtained with a single-beam spectrofluorometer. No attempt was made to adjust these values for either the intensity distribution of the excitation source or the relative sensitivity of the emission unit with wavelength. However, several observations can be drawn from this data that can be useful when applied to a HPLC fluorescence detector. 

Robert J. Argauer, "Fluorescence Methods for Pesticides." Chapter 4 in Analytical Methods for Pesticides and Plant Growth Regulators, Volume IX, "Spectroscopic Methods of Analysis," Academic Press, Inc., New York, N.Y., 1977. (contains over 100 literature references to fluorescence of pesticides)... 

Fluorescence of Pesticides by the Direct Approach. The fluorescence obtained by the methods just mentioned comes from the reagents, i.e., fluorogenic sprays and labelling compounds. There is sometimes a lack of selectivity since the fluorescence is nearly always the same for all the compounds on a particular chromatogram whether they are pesticides or impurities. In some cases there is also background fluorescence in addition to interferences from co-extractives which fluoresce under the experimental conditions. But for many pesticides there is no other choice. 

Type of Layer. The influence of the type of thin-layer was investigated by Caissie and Mallet (51) and it was shown to have an important effect on the fluorescence of pesticides. For instance, the spectral data vary from one layer to another (Table VII) some pesticides are fluorescent on basic aluminium oxide layers but they are not fluorescent on acidic layers. All these experiments demonstrate the flexibility of using fluorometric procedures and the selectivity that could be achieved if desired. 

Amplification of the natural fluorescence of some pesticides and bathochromic shift of the excitation and emission maxima detection limits 5-100 ng. 

Note A range of pesticides can be detected on cellulose layers using 3-hydroxyflavones without prior bromination. Thus, the natural fluorescence of robinetin or fisetin, which is weak in a non-polar environment, is significantly enhanced by the presence of polar pesticides [2, 5, 7, 8],... 

The principal limitation in the use of electrophoretic techniques is the lack of availability of suitable detection systems for quantitative analysis and unequivocal identification of pesticide analytes. Traditionally, either ultraviolet/visible (UVA IS) or fluorescence detection techniques have been used. However, as with chromatographic techniques, MS should be the detection system of choice. A brief comparison of the numbers of recent papers on the application of GC/MS and LC/MS with capillary elec-trophoresis/mass spectrometery (CE/MS) demonstrates that interfaces between CE... 

Carbamates and substituted ureas are a numerous group of pesticides widely used to control weeds, pests, and diseases in fruit trees, vegetables, and cereals. Carbamate residues in foods are commonly extracted with water-miscible solvents and determined by using a liquid chromatograph equipped with a sensitive detector, frequently a UV detector. In addition, to obtain adequate detection selectivity, the postcolumn fluorimetric labeling technique is used for methyl carbamates. Substituted ureas are normally extracted from foods with organic solvents, and they can be determined directly by HPLC-UV or after postcolumn derivatization by fluorescence determination of their derivatives. 

The initial studies on fluorimetry of pesticides were made in solution. The fluorescence behaviour of compounds such as Guthion (azinphosmethyl), Potsan, Warfarin and piperonyl butoxide was investigated for possible uses in the analysis of their residues [145]. Much work has been done by Sawicki and his co-workers in connection with fluorescent air pollutants and this has recently been reviewed [146]. However, most of the fluorimetric analyses of pesticides require pre-treatment of the compounds in order to convert them into fluorescent species. 

Most of the older methods of fluorimetric analysis of pesticides involved hydrolysis to form fluorescent anions. Co-ral (coumaphos) [147] was hydrolyzed in alkali to the hydroxybenzopyran, which was subsequently determined by means of its fluorescence. Guthion (azinphosmethyl) was hydrolyzed to anthranilic acid for fluorimetric analysis [148,149]. A method was developed [150] for Maretin (N-hydroxynaphthalimide diethyl phosphate) in fat and meat which involved hydrolysis in 0.5 M methanolic sodium hydroxide followed by determination of the fluorescence of the liberated naphthalimide moiety. Carbaryl (1-naphthyl N-methylcarbamate) and its metabolites have been determined by a number of workers using base hydrolysis and the fluorescence of the resulting naphtholate anion [151-153]. Nanogram quantities of the naphtholate anion could be detected. Zectran (4-dimethylamino-3,5-xylyl N-methylcarbamate) has been determined by the fluorescence of its hydrolysis product [154]. The fluorescence behaviour of other carbamate insecticides in neutral and basic media has been reported [155]. Gibberellin spray used on cherries has been determined fluorimetrically after treatment with strong acid [156]. Benomyl (methyl N-[l-(butylcarbamoyl)-2-benzimidazolyl]carbamate) has been analyzed by fluorimetry after hydrolysis to 2-aminobenzimidazole [157]. 

A number of 3-hydroxyflavones, unsubstituted in the S position, were found to be practically non-fluorescent in non-polar media while exhibiting intense fluorescence in polar environments [161]. The use of such compounds for analysis of pesticide residues has been evaluated for polar compounds separated by TLC [162]. The developed plates are sprayed with fisetin and a fluorescent spot results where the pesticide is located on a weakly fluorescent or non-fluorescent background. 

Preliminary results using a fluorescent tracer, which was added to the spray tank at the same time as the pesticide (Guthion WP), indicated that the distribution of the tracer, and presumably the pesticide, was not uniform, emphasizing the difficulty in the placement of the patches (2) This tracer technique is currently being evaluated as a tool for quantitative exposure estimation. This could result in a more realistic measurement of pesticide contact on the skin and minimize the reliance on extrapolation from the patch data. 

Fluorescence and Ultraviolet Absorbance of Pesticides and Naturally Occurring Chemicals in Agricultural Products After HPLC Separation on a Bonded-CN Polar Phase... 

Figure 1. Fundamental approaches generally used in the analysis of pesticides (or other chemicals) by fluorescence (or some other physical property)...

Pesticide-solvent interactions affect the fluorescence of the pesticides. 

Variation of Fluorescence Signal with Polarity of the Mobile Phase. A change in polarity of the mobile phase often aids the resolution of the pesticide from interfering co-elutants. However, the relative retention area for several chromatographic peaks obtained in the fluorescence mode of detection was found to vary as the mobile phase in the HPLC was changed. The data in Table 3 show the effects of common chromatographic solvents on the fluorescence of two pesticides. For maretin and o-phenyl-phenol, the relative fluorescence intensity, as measured in a spectrofluorometer, increased substantially as the polarity of the solvent increased. However, as shown for pyrazophos in... 

Pesticides Possessing Native Fluorescence. The number of pesticides that possess enough natural fluorescence to enable their quantitative determination directly on thin-layer chromatograms is rather limited. Typical examples are given in Table X which includes their spectral characteristics [9]. With the exception of diphacinone, most fluoresce in the blue region of the spectrum. The limit of detection for each compound is in the low nanogram per spot range. 

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