Esterified Xanthophylls Research Articles

Changes in the expression of genes encoding xanthophyl acyltransferases during the postharvest ripening of avocado (Persea americana) fruit.

Avocado fruit is rich in xanthophylls, which have been related to positive effects on human health. Xanthophyl acetyltransferases (XATs) are enzymes catalyzing the esterification of carboxylic acids to the hydroxyl group of the xanthophyll molecule. This esterification is thought to increase the lipophilic nature of the xanthophyll and its stability in a lipophilic environment. Studies on XATs in fruits are very scarce, and no studies had been carried out in avocado fruit during postharvest. The objective of this work was to investigate the changes in the expression of genes encoding XAT, during avocado fruit ripening. Avocado fruits were obtained from a local market and stored at 15 °C for 8 days. The fruit respiration rate, ethylene production, and fruit peel's color space parameters (L*, a*, b*) were measured during storage. Fruit mesocarp samples were taken after 1, 3, 5, and 7 days of storage and frozen with liquid nitrogen. Total RNA was extracted from fruit mesocarp, and the quantification of the two genes designated as COGE_ID: 936743791 and COGE_ID: 936800185 encoding XATs was performed with real-time quantitative reverse transcription polymerase chain reaction using actin as a reference gene. The presence of a climacteric peak and large changes in color were recorded during postharvest. The two genes studied showed a large expression after 3 days of fruit storage. We conclude that during the last stages of ripening in avocado fruit there was an active esterification of xanthophylls with carboxylic acids, which suggests the presence of esterified xanthophylls in the fruit mesocarp. © 2024 Society of Chemical Industry.

Xanthophyll esterases in association with fibrillins control the stable storage of carotenoids in yellow flowers of rapeseed (Brassica juncea).

Biosynthesis, stabilization, and storage of carotenoids are vital processes in plants that collectively contribute to the vibrant colors observed in flowers and fruits. Despite its importance, the carotenoid storage pathway remains poorly understood and lacks thorough characterization. We identified two homologous genes, BjA02.PC1 and BjB04.PC2, belonging to the esterase/lipase/thioesterase (ELT) family of acyltransferases. We showed that BjPCs in association with fibrillin gene BjFBN1b control the stable storage of carotenoids in yellow flowers of Brassica juncea. Through genetic, high-resolution mass spectrometry and transmission electron microscopy analyses, we demonstrated that both BjA02.PC1 and BjB04.PC2 can promote the accumulation of esterified xanthophylls, facilitating the formation of carotenoid-enriched plastoglobules (PGs) and ultimately producing yellow pigments in flowers. The elimination of BjPCs led to the redirection of metabolic flux from xanthophyll ester biosynthesis to lipid biosynthesis, resulting in white flowers for B. juncea. Moreover, we genetically verified the function of two fibrillin genes, BjA01.FBN1b and BjB05.FBN1b, in mediating PG formation and demonstrated that xanthophyll esters must be deposited in PGs for stable storage. These findings identified a previously unknown carotenoid storage pathway that is regulated by BjPCs and BjFBN1b, while offering unique opportunities for improving the stability, deposition, and bioavailability of carotenoids.

Heterologous expression of xanthophyll esterase genes affects carotenoid accumulation in petunia corollas

The majority of carotenoids in petals are xanthophylls and most of these xanthophylls are esterified with fatty acids. Although petunia (Petunia x hybrida) is an important ornamental plant, it cannot accumulate enough carotenoids to have deep-yellow flowers. Our previous study suggested that low esterification activity causes low carotenoid accumulation in petunia corollas. Here, we introduced xanthophyll esterase (XES) from the petals of Ipomoea obscura, tomato (Solanum lycopersicum), and marigold (Tagetes erecta) into a pale-yellow-flowered cultivar of petunia to see whether these affect carotenoid accumulation in petunia corollas. Carotenoid contents and the proportions of esterified xanthophylls were elevated in the corollas of XES-overexpressing (XES-OX) transformants. Expression analysis showed that the transcript levels of endogenous carotenoid biosynthetic genes, which included geranylgeranyl diphosphate synthase 2, ζ-carotene desaturase, and lycopene β-ring cyclase in corolla tubes were upregulated in XES-OX plants. In addition, we discovered a difference in the composition of esterified xanthophylls among XES-OX plants, which may be caused by differences in the substrate specificity of their respective XESs. We conclude that esterification is an important process for carotenoid accumulation and XES is a useful tool for the quantitative and qualitative control of carotenoid accumulation in petals.

The MYB Activator WHITE PETAL1 Associates with MtTT8 and MtWD40-1 to Regulate Carotenoid-Derived Flower Pigmentation in Medicago truncatula.

Carotenoids are a group of natural tetraterpenoid pigments with indispensable roles in the plant life cycle and the human diet. Although the carotenoid biosynthetic pathway has been well characterized, the regulatory mechanisms that control carotenoid metabolism, especially in floral organs, remain poorly understood. In this study, we identified an anthocyanin-related R2R3-MYB protein, WHITE PETAL1 (WP1), that plays a critical role in regulating floral carotenoid pigmentation in Medicago truncatula Carotenoid analyses showed that the yellow petals of the wild-type M. truncatula contained high concentrations of carotenoids that largely consisted of esterified lutein and that disruption of WP1 function via Tnt1 insertion led to substantially reduced lutein accumulation. WP1 mainly functions as a transcriptional activator and directly regulates the expression of carotenoid biosynthetic genes including MtLYCe and MtLYCb through its C-terminal acidic activation motif. Further molecular and genetic analyses revealed that WP1 physically interacts with MtTT8 and MtWD40-1 proteins and that this interaction facilitates WP1's function in the transcriptional activation of both carotenoid and anthocyanin biosynthetic genes. Our findings demonstrate the molecular mechanism of WP1-mediated regulation of floral carotenoid pigmentation and suggest that the conserved MYB-basic-helix-loop-helix-WD40 regulatory module functions in carotenoid biosynthesis in M. truncatula, with specificity imposed by the MYB partner.

Diversification of seed carotenoid content and profile in wild barley (Hordeum chilense Roem. et Schultz.) and Hordeum vulgare L.–H. chilense synteny as revealed by DArTSeq markers

The high carotenoid content and distinctive carotenoid profile of tritordeum are conferred by its wild progenitor, Hordeum chilense. Genetic studies on this wild barley could exploit the knowledge gained in Hordeum vulgare L. if the synteny between H. vulgare and H. chilense is established. DArTSeq markers were aligned to barley genome and used to inspect H. chilense-barley synteny. All chromosome pairs showed a good degree of collinearity with the exception of 7Hv–7Hch, where a reciprocal translocation in 7Hch was identified. Carotenoid analyses revealed a high diversity for total carotenoids, free and esterified lutein in a collection of H. chilense. Population structure analyses revealed the existence of two subgroups contrasting for total carotenoids, free lutein and esterified lutein in seeds. Lutein esters were produced with palmitic and linoleic acids as happens in tritordeum. However, tritordeum prefer palmitic acid for the synthesis of lutein esters but this preference is not maintained in H. chilense. This indicates the existence of diversity in the enzymes involved in the esterification which could be useful in tritordeum breeding. Furthermore, several accessions produced lutein monoesters but they lacked diesters which suggests that esterification is controlled by more than one enzyme in H. chilense. A total of 91 marker-trait associations were identified for carotenoid content and profile. These associations constitute a good starting point for future genetic analyses for the identification of candidate genes from H. vulgare genome.

Comparison of petunia and calibrachoa in carotenoid pigmentation of corollas

Petunia (Petunia hybrida) is an important ornamental plant with a wide range of corolla colors. Although pale-yellow-flowered cultivars, with a low amount of carotenoids in their corollas, are now available, no deep-yellow-flowered cultivars exist. To find why petunia cannot accumulate enough carotenoids to have deep-yellow flowers, we compared carotenoid profiles and expression of carotenoid metabolic genes between pale-yellow-flowered petunia and deep-yellow-flowered calibrachoa (Calibrachoa hybrida), a close relative. The carotenoid contents and the ratios of esterified xanthophylls to total xanthophylls in petunia corollas were significantly lower than those in calibrachoa, despite similar carotenoid components. A lower esterification rate of trans-xanthophylls than of cis-xanthophylls in petunia suggests that petunia xanthophyll esterase (XES) has low substrate specificity for trans-xanthophylls, which are more abundant than cis-xanthophylls in petunia corolla. The expression of genes encoding key enzymes of carotenoid biosynthesis was lower and that of a carotenoid catabolic gene was higher in petunia. XES expression was significantly lower in petunia. The results suggest that low biosynthetic activity, high cleavage activity, and low esterification activity cause low carotenoid accumulation in petunia corollas.

Lutein Esterification in Wheat Flour Increases the Carotenoid Retention and Is Induced by Storage Temperatures.

The present study aimed to evaluate the effects of long-term storage on the carotenoid pigments present in whole-grain flours prepared from durum wheat and tritordeum. As expected, higher storage temperatures showed a catabolic effect, which was very marked for free carotenoid pigments. Surprisingly, for both cereal genotypes, the thermal conditions favoured the synthesis of lutein esters, leading to an enhanced stability, slower degradation, and, subsequently, a greater carotenoid retention. The putative involvement of lipase enzymes in lutein esterification in flours is discussed, particularly regarding the preferential esterification of the hydroxyl group with linoleic acid at the 3′ in the ε-ring of the lutein molecule. The negative effects of processing on carotenoid retention were less pronounced in durum wheat flours, which could be due to an increased esterifying activity (the de novo formation of diesterified xanthophylls was observed). Moreover, clear differences were observed for tritordeum depending on whether the lutein was in a free or esterified state. For instance, lutein-3′-O-monolinoleate showed a three-fold lower degradation rate than free lutein at 37 °C. In view of our results, we advise that the biofortification research aimed at increasing the carotenoid contents in cereals should be based on the selection of varieties with an enhanced content of esterified xanthophylls.

Salt-stress and plant hormone-like responses for selective reactions of esterified xanthophylls in the aerial microalga Coelastrella sp. KGU-Y002

Esterification of xanthophylls is important for improving their bioactive potency. The effects of metal cations on esterified xanthophyll biosynthesis and on the accumulation and composition of salt stress-induced carotenoids were investigated in the aerial microalga Coelastrella sp. KGU-Y002 (Chlorophyta, Scenedesmaceae). Under nitrogen-deficient and high-light conditions, salt-supplemented cultures showed enhanced synthesis of esterified astaxanthin, adonixanthin, and zeaxanthin. In particular, KCl supplementation caused a remarkable increase in esterification, such that 73 % of xanthophylls were esterified. After 1 day of KCl treatment, the concentration of endogenous abscisic acid had increased to 200-fold its initial concentration. In algal cells treated with abscisic acid in nitrogen-deficient and high-light conditions, the reactive oxygen species concentration was almost the same as that in the KCl-supplemented culture. In these algal cells, abscisic acid enhanced esterification without affecting total carotenoid accumulation. These findings suggested that xanthophyll esterification is regulated during the response to salt stress by abscisic acid, which has plant hormone-like bioactivity in this aerial microalga. Further studies have been designed to examine large-scale production of esterified xanthophylls by this aerial microalga under salt-stress conditions, and to assess the bioactivity of esterified xanthophylls extracted from algal cells.

Effect of long-term storage on the free and esterified carotenoids in durum wheat (Triticum turgidum conv. durum) and tritordeum (× Tritordeum Ascherson et Graebner) grains

The effect of long-term storage on the carotenoid composition in durum wheat and tritordeum grains was studied. Total carotenoid (mainly lutein) content decreased according to a temperature dependent first-order degradative kinetic model. The carotenoid retention was similar in both genotypes at the lower temperatures (71–73% at −32°C, 70% at 6°C and 55–56% at 20°C), whereas at the higher temperatures the pigment retention was higher in tritordeum (42% at 37°C; 10% at 50°C) than durum wheat (23% at 37°C; 1% at 50°C), probably due to the greater proportion of esterified xanthophylls. A clear difference between free and esterified pigments was observed, with smaller losses and slower degradation (higher stability) observed for the latter. The xanthophyll esterification process was highly specific and had a significant effect on the carotenoid stability according to the fatty acids involved in the esterification and their position on the lutein molecule. The results were consistent with a degradation process in which the carotenoid behavior is influenced by the chemical structure. Xanthophyll esterification can be promoted by environmental conditions, regardless of the cereal genotype, and is a powerful tool to modulate the carotenoid profile in cereals.

Lutein, zeaxanthin and meso-zeaxanthin content of eggs laid by hens supplemented with free and esterified xanthophylls.

The xanthophyll carotenoids lutein (L), zeaxanthin (Z) and meso-zeaxanthin (MZ) are found at the macula, the central part of the retina, where they are referred to as macular pigment (MP). MP is studied in human subjects because of its proven role in enhancing visual function and its putative role in protecting against age-related macular degeneration. These benefits are probably due to the antioxidant and short-wavelength filtering properties of MP. It is known that eggs are a dietary source of L and Z. This experiment was designed to measure the egg yolk carotenoid response to hen supplementation with L, Z and MZ. A total of forty hens were used in the trial and were divided into eight groups of five hens. Each group was supplemented (with about 140mg active xanthophylls/kg feed) with one of the following oil-based carotenoid formulations for 6 weeks: unesterified L (group 1); L diacetate (group 2); unesterified Z (group 3); Z diacetate (group 4); unesterified MZ (group 5); MZ diacetate (group 6); L-MZ (1:1) diacetate mixture (group 7); L-MZ diacetate (1:3) mixture (group 8). Yolk carotenoid content was analysed weekly (in four randomly selected eggs) by HPLC. We found that hens supplemented with Z diacetate and MZ diacetate produced eggs with significantly greater carotenoid concentrations than their free form counterparts. This finding potentially represents the development of a novel food, suitable to increase MP and its constituent carotenoids in serum.

Chlorophyll and carotenoid pigments in the peel and flesh of commercial apple fruit varieties

Apple (Malus×domestica Borkh) has always been considered a fruit with low chlorophyll and carotenoid contents; however these pigments contribute also to the external (peel) and internal (flesh) fruit colourations, as well as to the health benefits associated with the regular consumption of this fruit. In the present work we studied the chlorophyll and carotenoid compositions of the peel and flesh of thirteen marketed apple varieties presenting different external colourations (green, yellow and red). All the varieties were characterised by a common pigment profile comprised by chlorophylls a and b, lutein, violaxanthin, neoxanthin, β-carotene, and esterified carotenoids (mainly violaxanthin and neoxanthin) as major compounds. Total pigment content was always higher in the peel (58.72–1510.77μg/g dry wt) than in the flesh (14.80–71.57μg/g dry wt) for each particular cultivar. In general, the green cultivars showed the highest pigment content both in the peel and in the flesh, which were followed in decreasing order by the peel of some red-skinned cultivars and the flesh of the yellow ones. In most cultivars, a characteristic chloroplastic pigment profile was found in the peel, with lutein as the main free carotenoid whilst a chromoplastic profile was usually observed in the flesh, with a prevalence of violaxanthin and neoxanthin and their corresponding acyl esters (mono- and diesters). A correlation between the carotenoid content and the amount of esterified xanthophylls was observed, in particular in the flesh, which reinforce the putative role of the esterification process in the accumulation of these lipophilic compounds within the chromoplasts.

Carotenogenesis and physico-chemical characteristics during maturation of red fleshed papaya fruit (Carica papaya L.)

Carotenoid development of red fleshed papaya fruit (Carica papaya L.) was investigated in the course of a complete pre- and postharvest period using HPLC-DAD coupled to mass spectrometry. Esterified xanthophylls such as β-cryptoxanthin laurate and caprate were the most abundant pigments during incipient carotenoid biosynthesis. Subsequent fruit maturation led to a gradual accumulation of carotenoids, whereas particularly β-cryptoxanthin laurate and total lycopene contents disproportionately increased, reaching maximum contents of up to 775 and 3168μg/100gof fresh weight (FW), respectively. Total carotenoid contents of fully ripe papaya ranged from 5414 to 6214μg/100gofFW, while corresponding biosynthetic precursors like phytoene, phytofluene, and ζ-carotene were only detected in trace amounts. Due to high contents of vitamin A precursors like β-carotene and β-cryptoxanthins, edible parts of the ripe fruit contained 132-166μg retinol equivalents per 100g of FW.Furthermore, the development of morphological and physico-chemical fruit traits was characterized revealing significant correlations to carotenoid accumulation. A ripening index derived from several parameters was developed to easily allow exact assignment of ripening stages in studies of carotenoid development during fruit ontogenesis. Additionally, on-tree ripened versus postharvest ripened fruits were compared revealing striking similarity of their physico-chemical parameters and contents of individual carotenoids.

Serum Lutein Response Is Greater from Free Lutein Than from Esterified Lutein during 4 Weeks of Supplementation in Healthy Adults

Background: Current data suggest great variability in serum response following lutein ingestion from various sources.Objective: To compare the relative serum response during supplementation with free lutein (fL) and lutein esters (Le).Methods: 72 volunteers (23–52 years; body mass index [BMI] >20 and <30 kg/m2; baseline serum lutein <20 µg/dL [<352 nmol/L]) were identified. Subjects, matched for gender, age, and BMI, were randomly assigned to the fL or Le group. fL and Le capsules contained 12.2 mg of free lutein or 27 mg of lutein ester (equivalent to 13.5 mg free lutein), respectively. Fasting blood was obtained at baseline and after 7, 14, 21, and 28 days of supplementation. Supplements were consumed with standard portions of dry, ready-to-eat cereal and 2% cow's milk.Results: Absolute changes in serum lutein, per mg daily dose, were significantly greater in fL vs. Le after 21 days (p = 0.0012) and remained so after 28 days (p = 0.0011) of supplementation. Serum lutein Area Under the Curve [AUC(day 0-28)] response was 17% greater for fL vs. Le (p = 0.0187). Regression models were used and determined that (1) baseline serum lutein levels and (2) the form of lutein ingested (fL > Le) influence the serum lutein response during supplementation, while subject age, gender, BMI, and serum lipids do not affect serum response.Conclusions: These results suggest that the relative serum lutein response will be significantly greater from supplements containing free lutein than from supplements containing lutein esters. These findings should be useful for future clinical trials exploring the effectiveness of lutein supplementation in the prevention of or protection against age-related macular degeneration and/or cataracts.

Overexpression of CrtR-b2 (carotene beta hydroxylase 2) from S. lycopersicum L. differentially affects xanthophyll synthesis and accumulation in transgenic tomato plants

Plant chloroplasts are enriched in xanthophylls which participate in photosynthesis as light-absorbing pigments and as dissipaters of excess light. In comparison, chromoplasts have evolved the capacity to synthesize and store brightly coloured carotenoid pigments to give flowers and fruits the power to attract pollinators and fruit dispersers. The best performing accumulator of xanthophylls in tomato is the petal chromoplast in contrast to the fruit chromoplast which only seems able to store carotenes. We have generated genetically engineered tomato lines carrying the tomato CrtR-b2 transgene with the aim of forcing the fruit to accumulate beta-xanthophylls. Both chloroplast- and chromoplast-containing tissues of hemizygous transgenic plants were found to contain elevated xanthophyll contents as a direct consequence of the increased number of CrtR-b2 transcripts. Hemizygous transgenic leaves contained fourfold more violaxanthin than control leaves. Developing fruits were yellow instead of green since they lacked chlorophyll a, and their violaxanthin and neoxanthin contents were seven- and threefold higher, respectively, than those of the control. Ripe fruits of hemizygous transgenic plants contained free violaxanthin and significant amounts of esterified xanthophylls. Esterified xanthophylls were present also in ripe fruits of control and homozygous plants. However, in transgenic homozygous plants, we observed a reduction in transcript content in most tissues, particularly in petals, due to a post-transcriptional gene silencing process. These findings demonstrate that tomato fruit chromoplasts can accumulate xanthophylls with the same sequestration mechanism (esterification) as that exploited by chromoplasts of the tomato petal and pepper fruit. This study on transgenic plants overexpressing an important carotenoid gene (CrtR-b2) provides an interesting model for future investigations on perturbations in beta-carotene-derived xanthophyll synthesis which in turn may provide insights into the molecular mechanisms controlling carotenoid metabolism in tomato.

Bioavailability comparison of free and esterified lutein for layer hens

Lutein supplements are often used to pigment and enrich layer chicken eggs. This experiment was conduced to compare the bioavailability of free and esterified lutein, by depletion method. Forty chickens were randomly divided into two groups. After 2 weeks of washout period, when all birds were fed the same low lutein basal diet, the two groups were fed for another 2 weeks with diets supplemented with free lutein or esterified lutein. Two experimental diets were supplemented with the same amount of 15 mg lutein/kg. On day 0 (baseline) and days 3, 7, and 14, birds fasting morning plasma samples were collected and stored for lutein analysis by high performance liquid chromatography method (HPLC). Results showed that: 1) Plasma lutein concentration increased dramatically after feeding free or esterified lutein; 2) Plasma lutein level in birds fed the diet supplemented with free lutein significantly differed from those fed esterified lutein on day 3. 3) There were no significant differences in plasma lutein levels between free lutein and esterified lutein on days 7 and 14. In conclusion, the lutein bioavailability from free lutein or esterified lutein supplements was comparable.

Bioavailability of Non-Provitamin A Carotenoids

The role of carotenoids in human nutrition has gained increased interest, especially due to their associated health-beneficial effects for a number of chronic diseases, including certain types of cancer and cardiovascular disease. Whereas data is available on the intake and presence of carotenoids in foods, limited information exists on factors influencing their bioavailability, especially for the non-provitamin A carotenoids. However, carotenoid absorption strongly depends on a number of factors which are not entirely understood. These include mainly the release of carotenoids from the food matrix, their incorporation into mixed bile micelles, the transfer of carotenoids from micelles to the mucosa for passive or facilitated absorption (via SR-BI proteins), and the sequestration into chylomicrons. Thus, dietary compounds influencing carotenoid micelle incorporation, e.g. the amount fat present in an ingested meal or components competing for uptake, such as phytosterols and other carotenoids, can have a considerable impact on carotenoid bioavailability. However, the effect of many dietary factors, including dietary fiber, type of fat, or minerals on carotenoid absorption is not well understood. In addition, bioavailability also depends on the carotenoid structure; in general, polar carotenoids are preferably incorporated into mixed micelles and tend to be of higher bioavailability, as may be the case for free vs. esterified xanthophylls, and cis-isomers vs. their trans-form, due to apparent shorter chain-lengths. Whereas their importance as part of a healthy diet warrants an improved understanding of carotenoids, their dietary fate following ingestion, including their stability, efficiency of micelle-incorporation, and pathway of absorption are still marginally understood.

Lutein and zeaxanthin uptake in cultured retinal pigmented epithelial cells.

The retinal pigment epithelium (RPE) has the essent ial role of providing oxygen and nutrients to the photoreceptors but also to removes their deb ris and metabolites. The RPE is subject to continuo usly and high level of oxidative stress. RPE use endogenous enzymatic and non-enzymatic antioxidants as a mechanism of reactive oxygen species (ROS) detoxification. RP E is also particularly rich in non-enzymatic antiox idants such are tocopherols, ascorbic acid, lutein and zea xanthin. In this study we investigated the ability of RPE cu ltured cells to incorporate lutein and zeaxanthin, in free and esterified forms, the carotenoid effect on cell culture viability and on cell antioxidant defense. Carotenoids were delivered to the cell culture using small amou nts of tetrahydrofuran added to the fetal calf seru m. The incorporation yield was determined by UV-Vis spectrophotometry, the cells viability by MTT assay and t he antioxidant effect by intracellular ROS assay (DCF- DA). Human RPE cultured cells can incorporate the major dietary carotenoids, lutein and zeaxanthin. The highest incorporation yield was found for zeaxanthi n, with 8.1 % of incorporation yield, while lutein incorporation yield was 7.12 %. Esterified xanthop hylls can not be incorporated into RPE cells. Lutei n and zeaxanthin administrated to RPE cells does not sign ificantly influence the cell viability. They showed a small positive effect on the cell viability in a concentr ation of 10 �M in the culture media. Lutein and zeaxanthin have an inhibitory effect on reactive oxygen species gen eration in the RPE cells.

Comparative study of the effect of the maturation process of the olive fruit on the chlorophyll and carotenoid fractions of drupes and virgin oils from Arbequina and Farga cultivars

Two varieties of olive fruit (Arbequina and Farga) have been characterized by their contents and types of chlorophyll and carotenoid pigments during the olive ripening. Independently of the initial content, the pigment concentrations decreased with fruit maturity. Chlorophyllides a and b, esterified xanthophylls and α-carotene were only detected in Arbequina fruits. Moreover, the synthesis of esterified xanthophylls, when the skin fruit colour changed from green to turning-colour only, was observed in this variety. The pigments transferred from fruit to the oil were also studied. All the pigments that were found in the fruit were transferred to the oils, in addition to derivative pigments associated with acidic medium in the oil extraction process. The destruction of the chlorophyll fraction was greater than of the yellow pigments during the olive oil extraction process.

Microstructure transitions derived from solubilization of lutein and lutein esters in food microemulsions

Solubilization and structural transformations in nonionic food microemulsions, characterized by large continuous isotropic region, containing solubilized lutein, and lutein ester were studied. The preparations are of oil-based concentrates composed of solubilized lutein or lutein ester in reverse micelles constructed from hydrophilic surfactant, alcohol and R(+)-limonene. The reverse micelles are diluted along a 60/40 surfactant/oil dilution line with aqueous phase (containing water and glycerol) up to the opposite corner of the diagram where o/w microemulsions are formed passing through a bicontinuous phase, without any visual phase separation. The dilution goes through two structural transitions that were determined in empty (reference) and solubilized systems. The solubilization capacity of both free lutein (FL) and lutein ester (LE) is considerably higher (as expected) in the reverse micelles (no aqueous phase), and in the W/O microemulsions than in the O/W microemulsions, but the highest solubilization was obtained within the bicontinuous phase. The solubilization was found to be affected by the lipophilicity of the surfactant and is also concentration-dependent throughout all the microemulsions structures, but the most pronounced surfactant effect was shown to be in the bicontinuous region. Structural transitions w/o→bicontinuous occurred at 30 wt.% aqueous phase in ‘empty’ (reference) microemulsions as well as in microemulsions containing solubilized free and esterified lutein (at the maximum solubilization levels). However, the transitions bicontinuous→o/w occurred at higher aqueous phase contents of 50 and 60 wt.% aqueous phase for FL and LE, respectively, in comparison to a transition at 40 wt.% for the empty microemulsion. As a result, the bicontinuous region in the presence of the guest molecules becomes much broader. The results indicate that in the reverse swollen micelles the guest molecules, being practically lipophilic will be easily accommodated at the concaved water interfaces (in the w/o region), which will enable high solubilization capacities, and without significant modifications of the interface. It becomes also evident that at zero curvature interfaces (bicontinuous) and in convex interfaces (oil/water) the lutein ester (the more lipophilic molecule) penetrates into the interface and swells both the bicontinuous interface and the droplets, similarly to most guest molecules. However, free lutein is poorly accommodated at the interface and interferes with the interfacial organization and modifies the flattened curvature, affecting the transformation from bicontinuous to o/w droplets.

Identification and quantification of zeaxanthin esters in plants using liquid chromatography-mass spectrometry.

It has been suggested that lutein and zeaxanthin may decrease the risk for age-related macular degeneration. Surprisingly, oleoresins rich in zeaxanthin are not yet available on the market. Several authors have reported enhanced stability of esterified xanthophylls, so plants containing zeaxanthin esters were investigated to establish valuable sources for the production of durable oleoresins. Liquid chromatography-atmospheric pressure chemical ionization mass spectrometry [LC-(APCI)MS] was used to unequivocally identify zeaxanthin esters of a standard mixture and in several plant extracts. Zeaxanthin esters were quantified on the basis of their respective molecular masses using zeaxanthin for calibration; total zeaxanthin was determined after saponification of aliquots of the extracts. Thus, dried wolfberries (Lycium barbarum), Chinese lanterns (Physalis alkekengi), orange pepper (Capsicum annuum), and sea buckthorn (Hippophae rhamnoides) proved to be valuable zeaxanthin ester sources. The present LC-MS method allows for an even more detailed analysis of zeaxanthin esters than reported previously.