Natural pigments detection

Supercritical fluid chromatography is the name for all chromatographic methods in which the mobile phase is supercritical under the conditions of analysis and the solvating properties of the fluid have a measurable effect on the separation. SFC has some advantages over GC and HPLC it extends the molecular weight range of GC, thermally labile compounds can be separated at lower temperatures, compounds without chromophores can be sensitively detected, and the use of open-tubular and packed columns is feasible. SFC can be employed in both the analysis of natural pigments and synthetic dyes, however it has not been frequently applied in up-to-date analytical practice. 

Organic modifiers have been frequently employed in CE to increase the solubility of hydrophobic solutes in the aqueous buffer system. Unfortunately, many organic modifiers are UV absorbent and cannot be used without considerable loss of sensitivity of detection. A contactless conductivity detection system has been developed which extends the application range of UV-absorbing solvents [ 119]. As both natural pigments and synthetic dyes absorb in the visible part of the spectra, the application of UV-absorbing organic modifiers in their CE analysis does not cause detection problems. 

Although there are a number of important food components which are naturally fluorescent (e.g., cereal brans, lignified materials such as pea, soy and cotton fiber, and even proteins and pigments), detection of many food components requires application of specific fluorochromes or diachromes. Therefore, quantitative analysis using microscopic imaging also requires judicious use of sensitive dyes or stains suitable for visualization and rapid measurement. The dyes must be stable, non-toxic to liing cells, easily and inexpensively... 

Identification and Assignment of Structure The identity of a synthetic product is often established by comparison of its absorption curve with that of the natural product or another standard sample. The absorption curves of synthetic and natural vitamin A2 are shown in Figure 7.1. The identification pf coloured substances is easier since colour can almost always be correlated with the presence of certain types of chromo-phoric groups. Thus, visible absorption spectra are employed for the detection and identification of the natural pigments (e.g. carotenoids, anthocyanins md porphyrins), dyes and other colouring matters. Hie absorption spectra of these colouring principles will be discussed separately in Chapter 12. 

Carotenoids are an important group of natural pigments with pronounced antioxidant activity, provitamin A factor, and many health benefits. The structure with conjugated double bonds governs mainly the proprieties of color, stability, detection, and quantification. With an increase of namral carotenoid markets, more different sources will be necessary and new technologies will be developed greener and cleaner, showing how promissory and profitable industrial sector can be. 

Photometric detection, 208-210 Photomultipliers, 378-379 Physical methods of detection, 206-211 photometric detection, 208-210 visual detection, 206-208 Physical phenomena in TLC, 49-53 broadening of chromatographic spots, 50-53 capillary flow, 49-50 volatility of solvents, 53 Pigments. See Natural pigments Planar chromatography (instrumental TLC), 3, 129-148,373-385 automation in, 131,382-384 chromatogram development, 135-140 automated multiple development (AMD), 138-140... 

Neutralization indicators, or acid-base indicators or pH indicators, are auxiliary reagents added to the titrand solution in order to detect the equivalence point in acid-base titrations. They can also be used for an accurate quantitative measure of the pH. Tournesol, a natural pigment extracted from some blue-green lichens, was the first pH indicator to be used (1850). Phenolphthalein and methyl orange were introduced somewhat later (1877 and 1878, respectively). Undeniably, the chief interests in the use of acid-base indicators are their low cost and ease of handling. However, they give rise to less precise and less accurate endpoints than some instrumental methods. 

NMR spectroscopy is most effective in qualitative analysis when the samples examinated are substantially pure compounds and has been used to confirm the theoretically predicted low-energy conformations of the Af-acylated hindered amine light stabiliser Tinuvin 440 [210]. Trace amounts of PDMS (quantification limit 0.1 ppm) in plastic additives, dyes and pigments were determined by 111 NMR after Soxhlet extraction [211]. ll NMR was also used for the detection of octadecanol, an impurity in Irganox PS 802 (3,3 -dioctadecyl thiodipropionate). NMR has identified the nature of a supposedly UV stabiliser of empirical formula C17H18N3CIO [44] (Scheme 5.2). 

Numerous CE separations have been published for synthetic colours, sweeteners and preservatives (Frazier et al., 2000a Sadecka and Polonsky, 2000 Frazier et al., 2000b). A rapid CZE separation with diode array detection for six common synthetic food dyes in beverages, jellies and symps was described by Perez-Urquiza and Beltran (2000). Kuo et al. (1998) separated eight colours within 10 minutes using a pH 9.5 borax-NaOH buffer containing 5 mM /3-cyclodextrin. This latter method was suitable for separation of synthetic food colours in ice-cream bars and fmit soda drinks with very limited sample preparation. However the procedure was not validated for quantitative analysis. A review of natural colours and pigments analysis was made by Watanabe and Terabe (2000). Da Costa et al. (2000) reviewed the analysis of anthocyanin colours by CE and HPLC but concluded that the latter technique is more robust and applicable to complex sample types. Caramel type IV in soft drinks was identified and quantified by CE (Royle et al., 1998). 

Dichlorobenzidine does not naturally oceur in the environment (lARC 1982a). 3,3 -Diehlorobenzidine was not detected in ambient air of two dyestuff produetion plants at detection limits of 5 (Narang et al. 1982) and 0.1 ng/m (Riggin et al. 1983). More reeent data on occupational exposure levels indieate the presence of levels 0.6-2.5 g/m in 3,3 -diehlorobenzidine production and pigment manufaeturing plants in Germany (DCMA 1989). 

Chalcones (l,3-diaryl-2-propen-l-ones) are open-chain fiavonoids that are widely biosynthesized in plants. They are important for the pigmentation of flowers and, hence act as attractants to the pollinators. As fiavonoids, chalcones also play an important role in defending against pathogens and insects. A review of the literature concerning the evaluation of natural chalcones from medicinal plants reveals that many studies into their antimicrobial and antiviral activities have been carried out in recent years. Some of these compounds were isolated by bioassay-guided fractionation, after previously detecting activity on the part of the plants. 

Qualitative Tests.—Natural mineral colours are usually mixtures of various components in different proportions, whereas artificial ones mostly have definite compositions. Qualitative analysis is made with the object of ascertaining the composition and thus the nature of the pigment and of detecting impurities and adulterants to this end the general procedure of inorganic analysis may be followed or, more simply, certain tests and reactions suited to each particular case may be carried out. 

Pteridines are widely distributed in nature and function as pigments, biological markers, and cofactors of enzymatic reactions. The oxidized heteroaromatic forms show a characteristic fluorescence which allows easy detection even in low concentrations. However, the more active 5,6,7,8-tetrahydro derivatives are nonfluorescing and oxidizable and create experimental problems in handling this type of compound. So far, all naturally occurring pteridines have turned out to be derivatives of pterin (2) and lumazine (3) which are modified by different substituents and functional groups in the 6- and/or 7-position. 

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