Carotenoids isomers

Emenhiser C., Sander L.C., and Schwartz, S.J., Capability of a polymeric C30 stationary phase to resolve cis-trans carotenoid isomers in reversed-phase liquid chromatography, J. Chromatogr. A, 101, 105, 1995. 

Emenhiser, C. et al.. Separation of geometrical carotenoid isomers in biological extracts using a polymeric Cjq column in reversed-phase liquid chromatography, J. Agric. Food Chem., 44, 3887, 1996. 

Sander, L.C. et ah. Separation of carotenoid isomers by capillary electrochromatography with C30 stationary phases. Anal. Chem., 71, 3477, 1999. 

Bioactive compounds, such as carotenoids have strong antioxidative properties and are used as efficient radical scavengers. In some natural sources several carotenoid isomers can be found, which differ in their biochemical activities such as bioavailability or antioxidation potency. Knowing the structure and concentration of each stereoisomer is crucial for an understanding of the effectiveness... 

NMR spectroscopy is essential for the structure determination of carotenoid isomers because the TI-NMR signals of the olefinic range are characteristic for the arrangement of the isomers. The stereoisomers of astaxanthin, as shown in Figure 4.16, can be separated on a shape-selective C30 capillary column with methanol under isocratic conditions. 

Sander, L. C., Epler Sharpless, K., Craft, N. E., and Wise, S. A. 1994. Development of engineered stationary phases for the separation of carotenoid isomers. Anal. Chem. 66 1667-1674. 

A second enzyme, BC02, was identified that cleaves carotenoids asymmetrically at the 9,10-double bond to produce the 10-apocarotenal (C27) and (3-ionone (C13), in a reaction similar to the Arabidopsis CCD7. Examples of BC02 have been cloned from mouse, zebra fish, ferret, and human (Kiefer et al. 2001, von Lintig et al. 2005, Hu et al. 2006). Substrate studies with different BC02s showed that these enzymes prefer acyclic carotenoids such as lycopene over cyclic carotenoids (Kiefer et al. 2001, von Lintig et al. 2005, Hu et al. 2006). These enzymes also seem to be selective for different carotenoid isomers. BC02 from ferret for example cleaves d,v-isomers of lycopene but not all-trans-lycopene (Hu et al. 2006). 

Fig. 2.25. (a-c) Comparison of HPLC separation of carotenoid isomers with a C30 column. Column temperature 25°C, flow rate lml/min, detection 450 nm. Mobile phase was acetone and water mixtures. Reprinted with permission from T. Glaser et al. [52]. 

Bell, C.M., Sander, L.C., Fetzer, J.C., and Wise, S.A., Synthesis and characterization of extended length aUcyl stationary phases for liquid chromatography with application to the separation of carotenoid isomers, J. Chromatogr. A, 753, 37, 1996. 

MH Saleh, B Tan. Separation and identification of cis/trans carotenoid isomers. J Agric Food Chem 39 1438-1443, 1991. 

As /3-carotene also has some vitamin A activity it is sometimes added to products for fortification as well as colourant purposes. If this is the case, the methods outlined above can be used. A more recent reference has used a similar method to look at carotenoid isomers in carrot juices and fortified drinks (Marx et al., 2000). 

Hyphenation of Modern Extraction Techniques to LC-NMR for the Analysis of Geometrical Carotenoid Isomers in Functional Food and Biological Tissues... 

ARTIFACT-FREE ISOLATION OF GEOMETRICAL CAROTENOID ISOMERS... 

Figure 5.2.1 Scheme for the extraction of geometrical carotenoid isomers from solid biological samples employing matrix solid phase dispersion (MSPD)... 

Figure 5.2.2 Scheme for the isolation and separation of geometrical carotenoid isomers from serum samples employing on-line solid phase extraction (SPE)-HPLC coupling... 

HPLC is commonly used to separate and quantify carotenoids using C18 and, more efficiently, on C30 stationary phases, which led to superior separations and improved peak shape.32 4046 An isocratic reversed-phase HPLC method for routine analysis of carotenoids was developed using the mobile phase composed of either methanol acetonitrile methylene chloride water (50 30 15 5 v/v/v/v)82 or methanol acetonitrile tetrahydrofuran (75 20 5 v/v/v).45 This method was achieved within 30 minutes, whereas gradient methods for the separation of carotenoids can be more than 60 minutes. Normal-phase HPLC has also been used for carotenoid analyses using P-cyclobond46 and silica stationary phases.94 The reversed-phase methods employing C18 and C30 stationary phases achieved better separation of individual isomers. The di-isomers of lycopene, lutein, and P-carotene are often identified by comparing their spectral characteristic Q ratios and/or the relative retention times of the individual isomers obtained from iodine/heat-isomerized lycopene solutions.16 34 46 70 74 101 However, these methods alone cannot be used for the identification of numerous carotenoids isomers that co-elute (e.g., 13-ds lycopene and 15-cis lycopene). In the case of compounds whose standards are not available, additional techniques such as MS and NMR are required for complete structural elucidation and validation. 

Retinyl esters Retinyl esters Retinyl esters Carotenoid isomers N-nitrosodiethanolamine in cosmetics Aloins, and related constituents of aloe Food colorants and aromatic glucoronides Azo and antraquinone textile dyes Alkaloids... 

Additional noteworthy applications of CEC include natural products such as the plant flavonoids hesperetin and hesperidin [160], anthraquinones extracted from rhubarb and from Chinese medicine [161], and heterocyclic compounds present in oils of bergamot, mandarin, and sweet orange [162], The CEC analysis of retinyl esters has been investigated by Roed et al. in nonaqueous mode for the separation of liver extracts of arctic seal [163]. Carotenoid isomers were also separated on C30 stationary phases by nonaqueous CEC [164]. It was found that CEC offered increased resolution compared to HPLC, and in CEC... 

Often, low levels of carotenoids in biological samples provide significant challenges in quantification by HPLC-PDA alone. Electrochemical detection (ECD) has been successful in quantifying low concentrations of carotenoids (MacCrehan and Schonberger, 1987 Finckh et ah, 1995 Yamashita and Yamamoto, 1997). More information about ECD can be found in Chapter 2. ECD has also been successful in quantifying carotenoid isomers in foods, plasma, prostate tissue, cervical tissue, and buccal mucosal cells (Ferruzzi et ah, 1998,2001 Allen et ah, 2003 Unlu et ah, 2007). Electrochemical array detection for all-irans - 3-carotene has been reported to be 10 fmol on column, which is approximately 100-1000 times more sensitive than UVA is detectors (Ferruzzi et ah, 1998). 

C30 columns have commonly been used in LC-NMR analyses of carotenoid isomers due to their greater shape selectivity compared to traditional C18 columns (Sander et al., 1994). This feature provides... 

Bdhm, V. 2001. Use of column temperature to optimize carotenoid isomer separation by C30 high-performance liquid chromatography. J. Sep. Sci. 24 955-959. 

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