Separation of carotenol fatty acid esters by high-performance liquid chromatography

doi:10.1016/S0021-9673(00)94373-5

Journal of Chromatography A

Volume 449, 1988, Pages 119-133

https://doi.org/10.1016/S0021-9673(00)94373-5 Get rights and content

Abstract

Employing isocratic and gradient-elution high-performance liquid chromatography HPLC a number of straight-chain fatty acid esters (decanoate, laurate, myristate, palmitate) of violaxanthin, auroxanthin, lutein, zeaxanthin, isozeaxanthin, and β-cryptoxanthin, prepared by partial synthesis, have been separated on a C₁₈ reversed-phase column. Several chromatographic conditions were developed that separated a mixture of di-fatty acid esters (dimyristate, myristate palmitate mixed ester, dipalmitate) of violaxanthin, auroxanthin, lutein, and zeaxanthin in a single chromatographic run. Hydroxycarotenoids such as lutein, zeaxanthin, and isozeaxanthin that are not easily separated by HPLC on C₁₈ reversed-phase columns, can be readily separated after derivatization with fatty acids and chromatography of their esters. Chromatographic conditions for optimum separation of carotenoids from various classes are discussed.

References (20)

T. Philip et al.
J. Chromatogr.
(1988)
W. Gau et al.
J. Chromatogr.
(1983)
F. Zonta et al.
J. Chromatogr.
(1987)
K. Tsukida et al.
Arch. Biochem. Biophys.
(1958)
G.R. Beecher et al.
J. Natl. Cancer Inst.
(1984)
F. Khachik et al.
J. Agric. Food Chem.
(1986)
F. Khachik et al.
J. Agric. Food Chem.
(1987)
G.R. Beecher et al.
M. Micozi
G. Noga et al.
Chromatographia
(1983)

There are more references available in the full text version of this article.

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Carotenoid esters in foods - A review and practical directions on analysis and occurrence
2017, Food Research International
Citation Excerpt :
Although not used for quantitative purposes, Schlatterer and Breithaupt (2005) also followed the method proposed by Khachik & Beecher's group to synthesize β-cryptoxanthin and zeinoxanthin esters with saturated fatty acids. Moreover, for studies on HPLC separation, violaxanthin esters were obtained by reaction of zeaxanthin fatty acid esters with m-chloroperbenzoic acid, followed by purification by HPLC (Khachik & Beecher, 1988b). Other protocol was used by mixing fatty acids and 1,3-dicyclocarbodiimide and 4-(dimethyamino)pyridine for synthesis of lutein esters (Abdel-Aal & Rabalski, 2015; Young et al., 2007).
Carotenoids are naturally found in both free form and esterified with fatty acids in most fruits and some vegetables; however, up to now the great majority of studies presents data on carotenoid composition only after saponification. The reasons for this approach are that a single xanthophyll can be esterified with several different fatty acids, generating a great number of different compounds with similar chemical and structural characteristics, thus, increasing the complexity of analysis compared to the respective saponified extract. This means that since UV/Vis spectrum does not change due to esterification, differentiation between free and acylated xanthophylls is dependent at least on elution order and mass spectrometry (MS) features. The presence of interfering compounds, especially triacylglycerides (TAGs), in the non-saponified extract of carotenoids can also impair carotenoid ester analyses by MS due to high background noise and ionization suppression since TAGs can be present in much higher concentrations than the carotenoid esters. This leads to the need of development of new and effective clean-up procedures to remove the potential interferents. In addition, only few standards of xanthophyll esters are commercially available, making identification and quantification of such compounds even more difficult. Xanthophyll esterification may also alter some properties of these compounds, including solubility, thermostability and bioavailability. Considering that commonly consumed foods are dietary sources of xanthophyll esters and that it is the actual form of ingestion of such compounds, an increasing interest on the native carotenoid composition of foods is observed nowadays. This review presents a compilation of the current available information about xanthophyll ester analyses and occurrence and a practical guide for extraction, pre-chromatographic procedures, separation and identification by liquid chromatography-tandem mass spectrometry (LC-MS/MS).
Physical and chemical characterization in fruit from 22 summer squash (Cucurbita pepo L.) cultivars
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The carotenoids were extracted from the rehydrated sample with 5 mL of ethanol containing 1 mg mL−1 butylated hydroxytoluene (BHT) using a Polytron homogenizer. Samples were saponified in order to hydrolyze esterified carotenoids that might complicate the chromatographic determinations (Khachik & Beecher, 1988). One millilitre of a 40 g KOH 100 mL−1methanolic solution was added to each tube, and the samples were saponified for 10 min at 85 °C.
Physical and chemical characters of 22 summer squash genotypes (traditional and commercial cultivars) were evaluated in fruit using correlation and principal component analysis. External and internal fruit quality was assessed by standard parameters (fruit weight and dimensions, firmness, color, dry matter, soluble solids, pH, total acidity) and nutritional quality (individual and total carotenoid content) in fruit epicarp and mesocarp. Some of the cultivars showed distinctive and interesting characters in mesocarp, such as high soluble solid (4.7 °Brix: Cu-8 traditional cultivar), dry matter (6.6 g 100 g⁻¹: Cu-36 commercial cultivar), β-carotene (45 mg kg⁻¹ dry matter: Cu-11 traditional cultivar) and total carotenoid contents (371 mg kg⁻¹ dry matter: Cu-29 commercial cultivar). Moderate and significant positive relation was found between epicarp and mesocarp color parameter (b*) with lutein and total carotenoid contents in mesocarp (r > 0.52***), suggesting that indirect selection for high carotenoid content will be successful. Multivariable analysis allowed the grouping of variables, the most important being the a* and b* parameters (in epicarp and mesocarp), dry matter, soluble solid, total acidity, lutein, β-carotene and total carotenoid contents (in epicarp). Some cultivars may serve as a source of desirable genes to develop improved varieties that respond to the new needs of the market.
Application of near-infrared reflectance spectroscopy for predicting carotenoid content in summer squash fruit
2014, Computers and Electronics in Agriculture
Citation Excerpt :
All steps were carried out in darkness or under gold fluorescent light to prevent possible photodegradation of products. Samples were saponified in order to hydrolyze esterified carotenoids that might complicate the chromatographic determinations (Khachik and Beecher, 1988). One millilitre of a 40% w/v KOH methanolic solution was added to each tube, and the samples were saponified for 10 min at 85 °C.
The potential of near-infrared reflectance spectroscopy (NIRS) for predicting total carotenoid, lutein and β-carotene contents in skin and flesh of Cucurbita pepo fruits was assessed. The carotenoid contents were performed by HPLC, and were regressed against different spectral transformations by modified partial least square (PLSm) regression. Coefficients of determination in the external validation varied from 0.81 to 0.96, which characterize those equations as having from good to excellent quantitative information. The standard deviation (SD) to standard error of prediction ratio (RPD) and range to standard error of prediction ratio (RER) were variable for the different fruit part and compounds, and showed values that were characteristic of equations suitable for screening purposes. PLSm loading plots corresponding to the first terms of the equations showed that effects of the C–H group of starch and lipids, O–H group of water, as well as protein and chlorophyll, were most important in modeling prediction equations. The use of NIRS represents an important breakthrough in breeding for improved nutritional quality of summer squash fruit.
Prooxidant activity of lutein and its dimyristate esters in corn triacylglyceride
2003, Food Chemistry
For the purpose of comparing the characteristics and utilities of carotenoid fatty acid esters (CFAEs) with their free forms, the pro-oxidant activities of free lutein (FL) and lutein dimyristate (LD), against purified corn-triacylglyceride (TAG), were studied at 40 °C in the dark. FL tended to promote the oxidation of corn-TAG, though at low concentration. The higher the amount of FL added, the higher the oxidation rate of corn-TAG. LD tended to have no significant effect on oxidation of corn-TAG at low concentration, but high concentration of LD promoted the oxidation. The degradation products of FL and LD might involve the cycle of corn-TAG oxidation, resulting in a higher rate than that of the control. Due to the higher stability of LD, the rate of corn-TAG oxidation induced by LD was slower than that by FL.
No effect of esterification with fatty acid on antioxidant activity of lutein
2001, Food Research International
Antioxidant activities of lutein and lutein myristate esters were evaluated by using both the methyl linoleate hydroperoxide method and colorimetry of the quenching activity against 1,1-diphenyl-2-picrylhydrazyl. The antioxidant activities of free lutein (FL), lutein monomyristate ester (LM) and lutein dimyristate ester (LD) were not significantly different in both methods. No synergetic effects among FL, LM and LD were obtained. These results suggest that esterification of OH groups with fatty acids might not affect antioxidant activity of lutein.
Chromatographic determination of carotenoids in foods
2000, Journal of Chromatography A
In recent years, there has been particular emphasis on obtaining more accurate data on the types and concentrations of carotenoids in foods for various health and nutrition activities. The analysis of carotenoids is complicated because of the diversity and the presence of cis–trans isomeric forms of this group of compounds. In addition, a wide variety of food products of vegetal and animal origin, vegetables and animal samples contain carotenoids, and a great range of carotenoids can be found in these samples. The characteristic conjugated double bond system of carotenoids produces the main problem associated with work and manipulation on carotenoids, that is their particular instability, especially towards light, heat, oxygen and acids. For this reason, several precautions are necessary when handling carotenoids. Another problem associated with analysis of carotenoids is the difficulty in obtaining standard compounds. High-performance liquid chromatographic methods for the determination of carotenoids in foods are reviewed. The sample extraction and treatment, carotenoid purification and standard manipulation are briefly commented on. We present a critical assessment of chromatographic methods developed for the determination of carotenoids in foods. Finally, some methods for carotenoid ester separation are reviewed.

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Separation of carotenol fatty acid esters by high-performance liquid chromatography

Abstract

J. Chromatogr.

J. Chromatogr.

J. Chromatogr.

Arch. Biochem. Biophys.

J. Natl. Cancer Inst.

J. Agric. Food Chem.

J. Agric. Food Chem.

Chromatographia