Detection, absorbance visible

A distinction is normally made between the visible and ultraviolet regions of the spectrum when detecting absorbing substances. Detection in the visible part of the spectrum can be carried out with the eye or with a photomultiplier. 

HPLC is the main separation technique for quantification of water-soluble, nonvolatile, thermally labile anthocyanins (Table 3.4). The columns most commonly used are reversed-phase C18. The pH of mobile phase is generally kept acidic with either formic, acetic, or trifluoroacetic acid. The order of elution follows the decreasing polarity of the anthocyanidins and is as follows delphinidin, cyanidin, petunidin, pelargonidin, peonidin, and malvidin. Disaccharide moieties in the C-3 position influence polarity less than the presence of the same two monosaccharides in C-3 and C-5 positions. Anthocyanins absorb visible light at 510-528 nm and UV light at 270-280 nm. DAD is the most common technique used for detection and quantification of anthocyanins, with the quantification occurring around the absorption... 

Trivalent chromium (Cr3+ ion) is not as toxic as the hexavalent form and can be directly detected by visible spectrophotometry, its UV absorption being nonspecific. Figure 25 presents the UV-visible spectra of the trivalent form and shows the great difference with one of the hexavalent form. Two peaks of absorbance are noted at 433 and 600 nm, giving a green colour to the solution. 

Scintillation Counter. Photomultiplier detectors, discussed in Chapter 5, are very sensitive to visible and UV light, but not to X-rays, to which they are transparent. In a scintillation detector the X-radiation falls on a compound that absorbs X-rays and emits visible light as a result. This phenomenon is called scintillation. A PMT can detect the visible light scintillations. The scintillating compound or phosphor can be an inorganic crystal, an organic crystal or an organic compound dissolved in solvent. 

The reagents used in post column derivatization will be specific for the particular solutes to be detected but will often be the same or similar to those used in precolumn derivatization. Reagents that produce derivatives that absorb visible light are popular as simple, relatively inexpensive detecting systems can be employed (43). In general, derivatization, particularly post column derivatization, is only used as a last resort to provide sensitivity or selectivity which is difficult or impossible to obtain by other means. Post column reactors complicate an already complex instrument, render it more difficult to operate, make it more expensive and unfortunately, however well designed, tends to denegrate the separation that has been achieved (often with difficulty) by the column. 

Note UV, ultraviolet absorbance detection Vis, visible absorbance detection FL, fluorescence detection MS, mass spectrometric detection CL, chemiluminescence detection. 

The polarization dependence of the photon absorbance in metal surface systems also brings about the so-called surface selection rule, which states that only vibrational modes with dynamic moments having components perpendicular to the surface plane can be detected by RAIRS [22, 23 and 24]. This rule may in some instances limit the usefidness of the reflection tecluiique for adsorbate identification because of the reduction in the number of modes visible in the IR spectra, but more often becomes an advantage thanks to the simplification of the data. Furthenuore, the relative intensities of different vibrational modes can be used to estimate the orientation of the surface moieties. This has been particularly useful in the study of self-... 

A UV/Vis absorbance detector can also be used if the solute ions absorb ultraviolet or visible radiation. Alternatively, solutions that do not absorb in the UV/Vis range can be detected indirectly if the mobile phase contains a UV/Vis-absorbing species. In this case, when a solute band passes through the detector, a decrease in absorbance is measured at the detector. 

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