Characterization of the Pale Yellow Petal/Xanthophyll Esterase gene family in citrus as candidates for carotenoid esterification in fruits.

In orange-pigmented citrus fruits, the xanthophyll esters are the predominant carotenoids, but their biosynthetic origin is currently unknown. In this work, seven PYP/XES (Pale Yellow Petal/ Xanthophyll esterase) genes were identified in Citrus genomes, but only PYP1-4 and 6 contained the structural domains essential for activity. The PYP/XES expression profiles in sweet orange and in other Citrus species such as lemon, mandarin and pummelo with marked differences in fruit pigmentation and content of xanthophylls esters, showed the upregulation of PYP1,2 and 6 genes during ripening only in orange-pigmented fruits. Moreover, transcript levels of PYP1, 2 and 6 genes in peel and pulp of sweet orange were accompanied by the accumulation of xanthophyll esters during ripening. This work reports for the first time the PYP/XES gene family in Citrus and strongly suggests its involvement in xanthophyll esterification in citrus fruit tissues and its influence in carotenoid accumulation and fruit pigmentation.

HPLC-DAD-APCI-MSn analysis of the genuine carotenoid pattern of pineapple (Ananas comosus [L.] Merr.) infructescence.

The genuine carotenoid pattern of pineapple infructescence was assessed by HPLC-DAD-APCI-MSn analysis. Prevailing pigments in the shell of 'MD2' (syn. "Extra Sweet") fruit were (all-E)-lutein and (all-E)-ß-carotene, in addition to chlorophylls a and b. The edible flesh contained (all-E)-violaxanthin, (all-E)-ß-carotene, and diverse esters of (9Z)-violaxanthin with caprylic, capric, lauric, and myristic acid. The latter esters have been reported for the first time as pineapple constituents. Total carotenoid concentrations in the edible fractions of the four varieties 'Sugar Loaf', 'Smooth Cayenne', 'MD2', and 'Queen Victoria' cultivated in Ghana ranged between 29 and 565 µg/100 g of fresh weight (FW). Total carotenoids in the flesh of fully ripe 'MD2' fruit exported by air freight amounted to 302 µg/100 g of FW, those in green ripe samples dispatched by sea freight to 359-432 µg/100 g of FW. All yellow fleshed cultivars exhibited a highly similar qualitative carotenoid profile.

Genuine Carotenoid Profiles in Sweet Orange [Citrus sinensis (L.) Osbeck cv. Navel] Peel and Pulp at Different Maturity Stages.

The carotenogenesis in the endocarp and flavedo of Navel oranges over four consecutive maturity stages was assessed by high-performance liquid chromatography-diode array detection-atmospheric pressure chemical ionization-multistage mass spectrometry. After optimization of the extraction method, 77 carotenoids, including 26 monoesters and 33 diesters of violaxanthin, ß-citraurin, and antheraxanthin, were characterized. Whereas chloroplast-specific pigments, such as (all-E)-lutein and (all-E)-ß-carotene, predominated in the flavedo of green-ripe fruit, a highly complex pattern of xanthophyll esters was found in the mature oranges. Total carotenoid contents of flavedo were approximately 9-fold higher [12 605 µg/100 g of fresh weight (FW)] than those in the endocarp (1354 µg/100 g of FW) at the fully mature stage. The mature endocarp abundantly contained violaxanthin mono- and diesters, in addition to diverse antheraxanthin esters, which were exclusively detected in this fruit fraction. Likewise, ß-citraurin esters were found to be unique flavedo constituents of mature fruit. Therefore, they may support the detection of fraudulent use of peel fractions during orange juice production.

Life history shapes variation in egg composition in the blue tit Cyanistes caeruleus.

Maternal investment directly shapes early developmental conditions and therefore has long-term fitness consequences for the offspring. In oviparous species prenatal maternal investment is fixed at the time of laying. To ensure the best survival chances for most of their offspring, females must equip their eggs with the resources required to perform well under various circumstances, yet the actual mechanisms remain unknown. Here we describe the blue tit egg albumen and yolk proteomes and evaluate their potential to mediate maternal effects. We show that variation in egg composition (proteins, lipids, carotenoids) primarily depends on laying order and female age. Egg proteomic profiles are mainly driven by laying order, and investment in the egg proteome is functionally biased among eggs. Our results suggest that maternal effects on egg composition result from both passive and active (partly compensatory) mechanisms, and that variation in egg composition creates diverse biochemical environments for embryonic development.

Impact of Cultivation Method and Cultivar Selection on Soluble Carbohydrates and Pungent Principles in Onions ( Allium cepa L.).

The composition of soluble carbohydrates such as fructooligosaccharides (FOS) in onions ( Allium cepa L.) plays a role regarding their digestibility, long-term storability, and processability. Qualitative and quantitative profiles of soluble carbohydrates were determined in 23 different onion samples comprising 20 cultivars grown at two different locations in 2014 and 2015. FOS concentrations were 1.1-fold higher in set grown onions than in seed grown onions ( p = 0.001). FOS levels of dehydrator cultivars were higher (overall average: 130.8 ± 42.4 g/L FOS) than those of common set and seed (61.8 ± 20.0 and 29.4 ± 14.7 g/L FOS) grown cultivars. Consequently, cultivation method (seed vs. set planting) and cultivar selection were crucial when aiming at onions with defined FOS contents. Besides FOS and other carbohydrate-related parameters, levels of alk(en)yl cysteine sulfoxides, indicating onion oil yield and pungency of onions, were determined to be different in dehydrator onions (13.1 ± 2.6 µmol/mL), seed (8.4 ± 1.3 µmol/mL), and set grown onions (7.5 ± 1.6 µmol/mL).

Carotenoids and xanthophyll esters of yellow and red nance fruits (Byrsonima crassifolia (L.) Kunth) from Costa Rica.

Carotenoid profiles, by means of HPLC-PDA-MSn, and CIE-L*C*h° colour values of yellow and red nance fruits from Costa Rica were elucidated. Among 16 carotenoids detected, (all-E)-lutein was the most abundant accounting for >80% of the total carotenoids, followed by (all-E)-zeaxanthin (9-11%) and (all-E)-ß-carotene (2-9%). Minor constituents were (Z)-isomers of lutein and ß-carotene, as well as diverse lutein diesters. Among the esters, lutein dimyristate was the most abundant as substantiated by the comparison with a marigold flower extract. Total carotenoids in the peel (616.2 µg/100 g of FW in yellow nance and 174.2 µg/100 g of FW in red nance) were higher than in the pulp (39.4 µg/100 g of FW in yellow nance and 31.4 µg/100 g of FW in red nance). Since carotenoid profiles of yellow and red varieties were qualitatively similar, although the colour values showed significant differences (77.2 and 88.6 h° in peel and pulp of yellow nance, versus 32.7 and 67.3 h° in peel and pulp of red nance, respectively), pigments other than carotenoids may impart the colour of red nance. High lutein content renders nance fruit as a nutritionally relevant source of this micronutrient.

Screening of critical factors influencing the efficient hydrolysis of zeaxanthin dipalmitate in an adapted in vitro- digestion model.

As hydrolysis of carotenoid esters is believed to be highly efficient in vivo, their insufficient hydrolysis in in vitro-digestion models, particularly, regarding zeaxanthin diesters, is a current issue. Therefore, in this study, several factors related to the enzymatic hydrolysis were investigated in an adapted version of the standardized INFOGEST in vitro-digestion model, using zeaxanthin dipalmitate (ZDP) as a substrate. The results showed that pancreatic lipase was able to hydrolyze ZDP, whereas carboxyl ester lipase (CEL) substantially contributed to ZDP cleavage. Replacement of commonly used porcine with bovine bile extracts and the substitution of coffee creamer for soybean oil at identical fat contents both significantly improved hydrolysis efficiency and bioaccessibility of total zeaxanthin to better mimic in vivo conditions. Thus, bile and lipids selection for in vitro digestion of carotenoid esters was crucial. The combined use of coffee creamer, pancreatin, CEL, and bovine bile led to the highest hydrolysis efficiency of 29.5%.

Perspective on the Ongoing Replacement of Artificial and Animal-Based Dyes with Alternative Natural Pigments in Foods and Beverages.

This perspective highlights current trends, advances, and challenges related to the replacement of artificial dyes and the insect-based carmine with alternative natural pigments. Briefly reviewing the history of food coloration, key publications and public events leading to diverse concerns about artificial dyes and carmine will be summarized. An overview about promising alternatives in the market and those under development is provided, including a separate section on coloring foodstuffs. The perspective aims at supporting readers to keep abreast with the enormous efforts undertaken by the food and beverage industry to replace certain food dyes.

Subcritical water extraction of phenolic and antioxidant constituents from pistachio (Pistacia vera L.) hulls.

Pistachio hulls, important by-products of pistachio processing, were extracted using an environmentally friendly process with subcritical water (SCW) at a pressure of 6.9 MPa in the range of 110 and 190 °C, and a flow rate of 4 ml/min. Detailed HPLC-DAD-ESI/MSn analyses allowed the identification of 49 phenolic compounds in the SCW extracts. Total gallotannin yields up to 33 g/kg were reached at 150-170 °C, where gallic acid (22.2 g/kg) and penta-O-galloyl-ß-d-glucose (9.77 g/kg) levels were 13.2- and 10.6-fold higher than those in the aqueous methanol extracts. Flavonols were also effectively extracted at 110-150 °C (4.37-5.65 g/kg), while anacardic acid recovery was poor (1.13-2.77 g/kg). Accordingly, high amounts of anacardic acids (up to 50.7 g/kg) were retained in the extraction residue, revealing that SCW extraction allowed selective extraction of gallotannins and flavonols. Antioxidant capacities ranged from 0.68 to 1.20 mmol Trolox equivalents (TE)/g for SCW extracts increasing with temperature up 190 °C.

HPLC-DAD-APCI/ESI-MSn analysis of carotenoids and α-tocopherol in Costa Rican Acrocomia aculeata fruits of varying maturity stages.

Carotenoids and tocopherols were characterised in the meso- and exocarp of wild-growing Costa Rican Acrocomia aculeata fruits. Comprehensive profiling of these lipophilic micronutrients in fruits of three varying maturity stages was conducted for the first time. A method for the simultaneous extraction and quantitation of carotenoids and α-tocopherol was developed and validated. Detailed HPLC-DAD-APCI/ESI-MSn analyses enabled the identification of α-tocopherol and 25 carotenoids. The latter comprised antheraxanthin, ß-carotene, lutein, luteoxanthin, neoxanthin, phytoene, phytofluene, violaxanthin, zeaxanthin, and several (Z)-isomers of the aforementioned compounds. Quantitation by HPLC-DAD/FLD revealed total carotenoid concentrations of 872±178 and 3075±407µg/100g fresh weight in the meso- and exocarp of fully ripe fruits, respectively. In both fruit fractions, progressing maturation resulted in the accumulation of phytoene, phytofluene, (all-E)-zeaxanthin, (all-E)-antheraxanthin, and (all-E)-violaxanthin. Carotenoid profiling was supported by multivariate data analysis. Carotenoid precursors and xanthophyll cycle pigments characterised Macauba fruits of full maturity.

Effect of chlorogenic acid on spectral properties and stability of acylated and non-acylated cyanidin-3-O-glycosides.

Spectral characteristics and heat stability (90°C, 5h) of six isolated, structurally related cyanidin-3-O-glycosides from black carrot were investigated in aqueous solutions (pH 3.6 and 4.6) supplemented with chlorogenic acid (molar anthocyanin:co-pigment ratios 1:62.5-1:250). Chlorogenic acid supplementation generally increased absorbance of non-acylated anthocyanins by up to 97.9 and 122.9% at pH 3.6 and 4.6, respectively, being mainly attributed to the formation of intermolecular anthocyanin:co-pigment complexes. The effect significantly decreased when the chain length of the 3-O-linked non-acylated anthocyanins was increased from mono- to di- to triglycosyl moieties, possibly due to steric interferences of bound sugars and co-pigments. Intermolecular co-pigmentation was investigated for the first time for feruloylated and sinapoylated cyanidin-3-O-triglycosides. They exhibited weaker effects (10.8-16.0%) at pH 4.6, and no spectral response was observed at pH 3.6. Chlorogenic acid addition evoked weak enhancement of thermal pigment stability at pH 3.6, while it was highly detrimental for all anthocyanins at pH 4.6.

Effect of aggregation form on bioavailability of zeaxanthin in humans: a randomised cross-over study.

Carotenoid bioavailability from plant and animal food is highly variable depending on numerous factors such as the physical deposition form of carotenoids. As the carotenoid zeaxanthin is believed to play an important role in eye and brain health, we sought to compare the human bioavailability of an H-aggregated with that of a J-aggregated deposition form of zeaxanthin encapsulated into identical formulation matrices. A randomised two-way cross-over study with sixteen participants was designed to compare the post-prandial bioavailability of an H-aggregated zeaxanthin and a J-aggregated zeaxanthin dipalmitate formulation, both delivering 10 mg of free zeaxanthin. Carotenoid levels in TAG-rich lipoprotein fractions were analysed over 9·5 h after test meal consumption. Bioavailability from the J-aggregated formulation (AUC=55·9 nmol h/l) was 23 % higher than from the H-aggregated one (AUC=45·5 nmol h/l), although being only marginally significant (P=0·064). Furthermore, the same formulations were subjected to an internationally recognised in vitro digestion protocol to reveal potential strengths and weaknesses of simulated digestions. In agreement with our human study, liberation of zeaxanthin from the J-aggregated formulation into the simulated duodenal fluids was superior to that from the H-aggregated form. However, micellization rate (bioaccessibility) of the J-aggregated zeaxanthin dipalmitate was lower than that of the H-aggregated zeaxanthin, being contradictory to our in vivo results. An insufficient ester cleavage during simulated digestion was suggested to be the root cause for these observations. In brief, combining our in vitro and in vivo observations, the effect of the different aggregation forms on human bioavailability was lower than expected.

Carotenoids from gac fruit aril (Momordica cochinchinensis [Lour.] Spreng.) are more bioaccessible than those from carrot root and tomato fruit.

Using a simulated digestion procedure in vitro, liberation and bioaccessibility of ß-carotene (29.5±1.7% and 22.6±0.9%, respectively) and lycopene (51.3±2.6% and 33.2±3.1%, respectively) from gac fruit aril were found to be significantly higher than from carrot root (ß-carotene, 5.2±0.5% and 0.5±0.2%, respectively) and tomato fruit (lycopene, 15.9±2.8% and 1.8±0.5%, respectively). Gac fruit aril naturally contained significantly more lipids (11% on fresh weight base) than carrot root and tomato fruit (<1%). However, when test meals were supplemented with an O/W emulsion to match the content of gac fruit aril, carotenoid bioaccessibility was still considerably lower than that from genuine gac fruit aril. Carotenoids in gac fruit aril were found to be stored in small, round-shaped chromoplasts. Despite the high lipid content, these carotenoids are unlikely to occur in a lipid-dissolved state according to simple solubility estimations, instead being possibly deposited as submicroscopic crystallites. In contrast, carotenoids of carrot root and tomato fruit were stored in large, needle-like crystallous chromoplasts. Consequently, we hypothesized the natural deposition form to be majorly responsible for the observed differences in bioaccessibility. A favorable surface-to-volume ratio of the deposition form in gac fruit aril might have allowed a more rapid micellization during digestion, and thus, an enhanced bioaccessibility. Irrespective of the ultimate reason, gac fruit aril provided a highly bioaccessible form of both lycopene and provitamin A (ß-carotene), thus offering a most valuable dietary source of both carotenoids. Currently, gac is majorly grown in Southeast Asia, where its consumption might help to diminish the 'hidden hunger' namely the insufficient supply with vitamin A. Ultimately, gac fruit might thus contribute to alleviating most severe health implications of vitamin A deficiency, such as anaemia and xerophthalmia, the prevailing cause of preventable childhood blindness, as well as mortality from infectious diseases.

Carotenoids and Carotenoid Esters of Red and Yellow Physalis (Physalis alkekengi L. and P. pubescens L.) Fruits and Calyces.

Carotenoid profiles of fruits and calyces of red (Physalis alkekengi L.) and yellow (P. pubescens L.) Physalis were characterized by HPLC-DAD-APCI-MSn. Altogether 69 carotenoids were detected in red Physalis, thereof, 45 were identified. In yellow Physalis, 40 carotenoids were detected and 33 were identified. Zeaxanthin esters with various fatty acids were found to be the most abundant carotenoids in red Physalis, accounting for 51-63% of total carotenoids, followed by ß-cryptoxanthin esters (16-24%). In yellow Physalis, mainly free carotenoids such as lutein and ß-carotene were found. Total carotenoid contents ranged between 19.8 and 21.6 mg/100 g fresh red Physalis fruits and 1.28-1.38 mg/100 g fresh yellow Physalis fruits, demonstrating that Physalis fruits are rich sources of dietary carotenoids. Yellow Physalis calyces contained only 153-306 µg carotenoids/g dry weight, while those of red Physalis contained substantially higher amounts (14.6-17.6 mg/g dry weight), thus possibly exhibiting great potential as a natural source for commercial zeaxanthin extraction.

Anthocyanins from purple sweet potato (Ipomoea batatas (L.) Lam.) and their color modulation by the addition of phenolic acids and food-grade phenolic plant extracts.

Anthocyanin profiles and contents of three purple sweet potato provenances were investigated by HPLC-DAD-MSn. In contrast to widely uniform profiles, the contents of total (558-2477mg/100gDM) and individual anthocyanins varied widely. Furthermore, quantitative and qualitative effects of intermolecular co-pigmentation were studied by adding chlorogenic and rosmarinic acids, and food-grade phenolic apple and rosemary extracts at various dosages to a diluted purple sweet potato concentrate at pH 0.9, 2.6, 3.6, and 4.6. Addition of co-pigments generally increased pKH estimate-values of anthocyanins from 3.28 (without co-pigments) to up to 4.71, thus substantially broadening the pH range wherein colored forms prevail. The most pronounced hyperchromic shift by up to +50.5% at the absorption maximum was observed at pH 4.6. Simply by blending the co-pigments with purple sweet potato anthocyanins at pH-values ranging from 2.6 to 4.6, purplish-blue, light pink, magenta, brick-red, and intense red hues were accessible as expressed by CIE-L∗a∗b∗ color values.

Characterization of Mesocarp and Kernel Lipids from Elaeis guineensis Jacq., Elaeis oleifera [Kunth] Cortés, and Their Interspecific Hybrids.

Morphological traits, total lipid contents, and fatty acid profiles were assessed in fruits of several accessions of Elaeis oleifera [Kunth] Cortés, Elaeis guineensis Jacq., and their interspecific hybrids. The latter featured the highest mesocarp-to-fruit ratios (77.9-78.2%). The total lipid contents of both E. guineensis mesocarp and kernel were significantly higher than for E. oleifera accessions. Main fatty acids comprised C16:0, C18:1n9, and C18:2n6 in mesocarp and C12:0, C14:0, and C18:1n9 in kernels. E. oleifera samples were characterized by higher proportions of unsaturated long-chain fatty acids. Saturated medium-chain fatty acids supported the clustering of E. guineensis kernels in multivariate statistics. Hybrid mesocarp lipids had an intermediate fatty acid composition, whereas their kernel lipids resembled those of E. oleifera genotypes. Principal component analysis based on lipid contents and proportions of individual fatty acids permitted clear-cut distinction of E. oleifera, E. guineensis, and their hybrids.

Carotenoids and carotenoid esters of orange- and yellow-fleshed mamey sapote (Pouteria sapota (Jacq.) H.E. Moore & Stearn) fruit and their post-prandial absorption in humans.

Although different genotypes of mamey sapote with distinct pulp colors are consumed in countries from Central to South America, in-depth knowledge on genotype-related differences of their carotenoid profile is lacking. Since the fruit was found to contain the potentially vitamin A-active keto-carotenoids sapotexanthin and cryptocapsin, we sought to qualitatively and quantitatively describe the carotenoid profile of different genotypes by HPLC-DAD-MSn. Sapotexanthin and cryptocapsin were present in all genotypes. Keto-carotenoids such as cryptocapsin, capsoneoxanthin, and their esters were most abundant in orange-fleshed fruit, whereas several carotenoid epoxides prevailed in yellow-fleshed fruit. Differing carotenoid profiles were associated with different color hues of the fruit pulp, while the widely variable carotenoid content (3.7-8.0mg/100gFW) was mainly reflected by differences in color intensity (chroma C∗). Furthermore, the post-prandial absorption of sapotexanthin to human plasma was proven for the first time. Besides sapotexanthin, cryptocapsin was found to be resorbed.

Ultrastructural deposition forms and bioaccessibility of carotenoids and carotenoid esters from goji berries (Lycium barbarum L.).

Goji berries (Lycium barbarum L.) have been known to contain strikingly high levels of zeaxanthin, while the physical deposition form and bioaccessibility of the latter was yet unknown. In the present study, we associated ripening-induced modifications in the profile of carotenoids with fundamental changes of the deposition state of carotenoids in goji berries. Unripe fruit contained common chloroplast-specific carotenoids being protein-bound within chloroplastidal thylakoids. The subsequent ripening-induced transformation of chloroplasts to tubular chromoplasts was accompanied by an accumulation of up to 36mg/100g FW zeaxanthin dipalmitate and further minor xanthophyll esters, prevailing in a presumably liquid-crystalline state within the nano-scaled chromoplast tubules. The in vitro digestion unraveled the enhanced liberation and bioaccessibility of zeaxanthin from these tubular aggregates in goji berries as compared to protein-complexed lutein from spinach. Goji berries therefore might represent a more potent source of macular pigments than green leafy vegetables like spinach.

Urinary excretion of Citrus flavanones and their major catabolites after consumption of fresh oranges and pasteurized orange juice: A randomized cross-over study.

SCOPE: Orange juice contains flavanones including hesperidin and narirutin, albeit at lower concentrations as compared to orange fruit. Therefore, we compared bioavailability and colonic catabolism of flavanones from orange juice to a 2.4-fold higher dose from fresh oranges. METHODS AND RESULTS: Following a randomized two-way cross-over design, 12 healthy subjects consumed a test meal comprising either fresh oranges or pasteurized orange juice, delivering 1774 and 751 µmol of total Citrus flavanones, respectively. Deglucuronidated and desulfated hesperetin, naringenin, and the flavanone catabolites 3-(3'-hydroxy-4'-methoxyphenyl)propionic acid, 3-(3'-hydroxyphenyl)hydracrylic acid, 4-hydroxyhippuric acid, and hippuric acid were quantitated in 24-h urine by UHPLC-MS/MS. Differences in urinary hesperetin excretion were found to be nonsignificant (p = 0.5209) both after consumption of orange fruit (21.6 ± 8.0 µmol) and juice (18.3 ± 7.2 µmol). By analogy, postprandial flavanone catabolite excretions were highly similar between treatments. Excretion of 3-(3'-hydroxy-4'-methoxyphenyl)propionic acid was inversely related to that of hesperetin, illustrating the catabolite/precursor relationship. CONCLUSION: Despite 2.4-fold higher doses, excretion of flavanones from ingested fresh orange fruit did not differ from that following orange juice consumption, possibly due to a saturation of absorption or their entrapment in the fiber-rich matrix of the fruit.