Peak identification absorbance ratios

Table II shows the substances identified and amounts determined in the various dye samples. Resorcinol and 2,5-TDA were found in most samples. The identification of a-naphthol in sample (V) is somewhat uncertain since it was found that the absorbance ratios were not independent of the concentration for this substance. In general, few peaks could be seen on chromatographing all the samples and the same simple pattern showed up with several dyes.

Photodiode array detection has three major advantages for HPLC analysis (26) (a) multiple-wavelength detection, (b) peak identification, and (c) peak-purity determination. Since PDA can record the characteristic UV spectra of the different phenolics as they elute from the column, characterization and peak-purity information can be facilitated through comparison of the spectra at the front, the apex, and the tail of each peak. Furthermore, the rapid calculation of absorbance ratios between different wavelengths is possible, which can be used to classify the spectra by functional groups or by other criteria (Table 1). 

EDTA in vancomycin by liquid chromatography with absorbance ratioing for peak identification, J. Pharm. Biomed. Anal., 5 513 (1990). 

Positive identification of the suspected THC peak requires future isolation of the species from larger milk volumes, followed by off-line spectral analysis (NMR, IR, MS). At this time, the retention match, absorbance ratio match and spike test in a high resolution HPLC separation support a tentative identification of A9-THC in the milk. 

Figure 10.9 Identification of diamorphine (peak X) in extracts obtained from a scale pan and a syringe by the use of retention time and absorbance ratio data.

Baker et al. reported the identification of various drugs by means of their relative retention times in combination with the absorbance ratio, calculated from the peak heights observed with 254 and 280 nm UV-detection. 

As noted in the preceding table, elution of PAHs is detected by UV absorbance at two different wavelengths 280 nm and 365 nm. Fluorescence detectors are also applicable to the HPLC analysis of PAHs (9, 19). The UV detector monitors the sample simultaneously at two wavelengths, aiding in compound identification. For a specific compound, the ratio of absorbances at two different wavelengths is an intrinsic physical characteristic. Therefore, it is possible, in principle, to identify a sample analyte by this characteristic ratio. The chromatographic retention time of each of the specific peaks observed in the sample eluate is compared with those of known standard compounds for tentative analyte identification. For quantitation, peak areas of each standard, at each of six... 

The analyst can improve the identification of solutes or discrimination between samples by studying the multi-wavelength plots or spectra of the eluted components. Transformation of spectral data to yield derivative spectra have been used to improve sample discrimination, whilst a plot of the ratio of absorbances for two selected wavelengths is an excellent method for confirming the homogeneity of chromatographic peaks. However, there is a trend in forensic laboratories to use and develop techniques whereby the spectral data are reduced to a smaller number of numerical values. Results in this format at easier to compare and present in court and furthermore can be evaluated statistically. 

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