Free xanthophylls

Conversion of xanthophyll esters to free xanthophylls in marigold using methanol... 

Free xanthophylls, both endogenous and present in the saponified samples, are more polar and extract less efficiently into lipophilic solvents. Frequently, the addition of a polar organic solvent (tetrahydrofuran, methylene chloride, diethyl ether) is required to thoroughly extract them from the sample matrix and aqueous phase. 

Red fruit bodies of Phillipsia carminea collected in the Central African Republic contain the diester derivative (447) of phillipsiaxanthin (445) as the predominant pigment 47). The free xanthophyll was identified by chemical and physical methods, including direct comparison with material synthesised previously by Isler and coworkers (572). 

Note Free xanthophylls, both endogenous and present in the saponified samples, are more polar and extract less eficiently into lipophilic solvents. 

In processed food, the maceration reduces the particle size and removes some barriers, increases the contact superficies for interaction with digestive. Effectors, such as the presence of oil, can also have an influence on the bioaccessibUity and/or bioavailability of carotenoids these compounds are lipophilic molecules, and they have to be incorporated in mixed micelles in the duodenum before they can be absorbed in the mucosa [31]. Certain structural differences may alter fat solubility and modify the efficiency of the micellization. One of them is the esterification of xanthophyll with fatty acids. Esterified xanthophylls exhibit increased fat solubility relative to their corresponding free xanthophylls and even against carotenes. 

Wisniewska, A. and Subczynski, W.K., Accumulation of macular xanthophylls in unsaturated membrane domains. Free Radio. Biol. Med., 40, 1820, 2006. 

The degree of linkage of a compound may also affect its bioaccessibility in the gut. It is generally admitted that a compound linked with other molecules (e.g., via esterification, glycosylation, etc.) is not absorbed as well as its free form and thus it must be hydrolyzed in the gut in order to be taken up by enterocytes. Due to the presence of hydroxyl or keto groups on their molecules, the xanthophylls (lutein, zeaxanthin, and P-cryptoxanthin) are found in both free and esterified (monoester or diester) forms in nature, but few studies have been conducted to date to assess the bioavailabilities of these esters. 

Alkaline hydrolysis (saponification) has been used to remove contaminating lipids from fat-rich samples (e.g., pahn oil) and hydrolyze chlorophyll (e.g., green vegetables) and carotenoid esters (e.g., fruits). Xanthophylls, both free and with different degrees of esterification with a mixture of different fatty acids, are typically found in fruits, and saponification allows easier chromatographic separation, identification, and quantification. For this reason, most methods for quantitative carotenoid analysis include a saponification step. 

However, complete hydrolysis of carotenoid esters sometimes is not achieved in 1 to 3 hr. The saponification degree can be verified easily by the presence of carotenol ester peaks eluting later than the peaks of P-carotene on reversed phase columns. Retinol palmitate, added as an internal standard to orange juice, also serves to indicate whether saponification is complete, since it is converted to retinol which elutes at lower retention time. The mixture is subsequently washed with water until free of alkali in a separatory funnel. Other more polar solvents such as CH2CI2 or EtOAc, and diethyl ether alone or mixtured with petroleum ether can be used to increase the recovery of polar xanthophylls from the water phase. 

APCl in positive mode ionization and triple quadrupole detection was used for determination of free and bound carotenoids in paprika, obtaining the [M + H]+ and losses of fatty acids as neutral molecules from the [M + H]+ with MeOH, MTBE, and H2O as eluent from the C30 column. The positions of the fatty acids on unsymmetrical xanthophylls could not be established by the MS data. 

Taking into consideration that antenna xanthophylls not only possess original absorption but also resonance Raman spectra, and the fact that the Raman signal is virtually free from vibrational spectroscopy artifacts (water, sample condition, etc.), it seemed of obvious advantage to apply the described combination of spectroscopies for the identification of these pigments. 

FIGU RE 10.13 Schematic drawing of the distribution of xanthophyll molecules between raft domain (DRM) and bulk domain (DSM) in lipid bilayer membranes. For this illustration, the xanthophyll partition coefficient between domains is the same as obtained experimentally for raft-forming mixture. However, to better visualize the observed effect in the drawing, the number of lipid molecules was decreased and the total number of xanthophyll molecules was increased about 10 times. (From Wisniewska, A. and Subczynski, W.K., Free Radio. Biol. Med., 40, 1820, 2006. With permission.)... 

Wisniewska, A. and W. K. Subczynski. 2006b. Distribution of macular xanthophylls between domains in model of photoreceptor outer segment membranes. Free Radic. Biol. Med. 4 1257-1265. 

Mortensen, A. and L. H. Skibsted. 1997a. Free radical transients in photobleaching of xanthophylls and carotenes. Free Rad. Res. 26 549-563. 

Johnson, E. J., M. Neuringer et al. (2005). Nutritional manipulation of primate retinas. III. Effects of lutein or zeaxanthin supplementation on adipose and retina of xanthophyll-free monkeys. Invest. Ophthalmol. Vis. Sci. 46(2) 692-702. 

It has been shown in many studies that protective effects of carotenoids can be observed only at small carotenoid concentrations, whereas at high concentrations carotenoids exert pro-oxidant effects via propagation of free radical damage (Chucair et al., 2007 Lowe et al., 1999 Palozza, 1998, 2001 Young and Lowe, 2001). For example, supplementation of rat retinal photoreceptors with small concentrations of lutein and zeaxanthin reduces apoptosis in photoreceptors, preserves mitochondrial potential, and prevents cytochrome c release from mitochondria subjected to oxidative stress induced by paraquat or hydrogen peroxide (Chucair et al., 2007). However, this protective effect has been observed only at low concentrations of xanthophylls, of 0.14 and 0.17 pM for lutein and zeaxanthin, respectively. Higher concentrations of carotenoids have led to deleterious effects (Chucair et al., 2007). 

Xanthophyll esters are common in fruits and vegetables. Few data exist regarding the effect of carotenoid esterification on carotenoid bioavailability. Xanthophyll esters are readily broken in the human intestine (West and Castenmiller 1998 Breithaupt and others 2003 Faulks and Southon 2005). Chitchumroonchokchai and Failla (2006) demonstrated that hydrolysis of zeaxanthin esters increases zeaxanthin bioavailability. Wingerath and others (1995) did not find (3-cryptoxanthin esters in chylomicrons from humans fed with tangerine juice. Herbst and others (1997) demonstrated that lutein diesters are more bioavailable than free lutein. However, the question of whether the free or the esterified form is more bioavailable to humans is still an ongoing discussion. 

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