Strain-Dependent Carotenoid Productions in Metabolically Engineered <Emphasis Type="Italic">Escherichia coli</Emphasis>

Seven Escherichia coli strains, which were metabolically engineered with carotenoid biosynthetic pathways, were systematically compared in order to investigate the strain-specific formation of carotenoids of structural diversity. C30 acyclic carotenoids, diaponeurosporene and diapolycopene were well produced in all E. coli strains tested. However, the C30 monocyclic diapotorulene formation was strongly strain dependent. Reduced diapotorulene formation was observed in the E. coli strain Top10, MG1655, and MDS42 while better formation was observed in the E. coli strain JM109, SURE, DH5a, and XL1-Blue. Interestingly, C40 carotenoids, which have longer backbones than C30 carotenoids, also showed strain dependency as C30 diapotorulene did. Quantitative analysis showed that the SURE strain was the best producer for C40 acyclic lycopene, C40 dicyclic β-carotene, and C30 monocyclic diapotorulene. Of the seven strains examined, the highest volumetric productivity for most of the carotenoids structures was observed in the recombinant SURE strain. In conclusion, we showed that recombinant hosts and carotenoid structures influenced carotenoid productions significantly, and this information can serve as the basis for the subsequent development of microorganisms for carotenoids of interest.     

Characterization of Carotenoid Triplet States in the Light-Harvesting Complex B800–850from the Purple Bacterium Rubrivivax gelationsus

The carotenoid triplet states in the light-harvesting complex B800–850from purple bacterium Rubrivivax gelatinosus were characterized by absorption-detected magnetic resonance in zero magnetic field (ADMR) spectroscopy. Detailed HPLC analysis of carotenoids from B800–850demonstrated the presence of several carotenoids bound to the complex: the major ones are hydroxyspheroidene and spheroidene (together 80%), followed by neurosporene and hydroxyneurosporene (7%), spheroidenone and hydroxyspheroidenone (7.5%) and two other minor carotenoids that could be 3,4-dihydrospheroidenone and 3,4-dihydrohydroxyspheroidenone (5.5%). Three triplet states originating from carotenoids present in the B800–850were observed. The identical T-S spectra recorded at selectively chosen 2|E| transitions of carotenoids indicated that all these triplet states can be attributed to three different populations of one carotenoid family, probably to spheroidene and to hydroxyspheroidene, with different out-of-plane distortions of their polyene chain due to a different protein environment. Triplet states of the neurosporene and the spheroidenone families are probably not observed because of the low signal amplitude.     

Carotenoid and fatty acid profiles of bilberries and cultivated blueberries from Romania

The worldwide consumption of blueberries has increased due to their potential health effects. They contain micronutrients, various phytochemicals and anti-oxidant pigments, represented especially by anthocyanins. In addition to these, carotenoids and fatty acids are also present. The aim of this study was the identification and quantification of carotenoids (by HPLC) and fatty acids (by GC with flame ionisation detection) in two bilberry genotypes (Wild 1 and 2) and three cultivars (BlueCrop, Elliot, and Duke). The main carotenoids identified were lutein, ??-cryptoxanthin, and ??-carotene, with the average total carotenoid content of 266 ??g per 100 g of fruit. The ratio of saturated fatty acids (SFAs) to polyunsaturated fatty acids (PUFAs) was 0.098 for Wild 1 and 0.11 for Wild 2 genotypes and ranged from 0.14 to 0.28 for cultivars. The main fatty acids identified were linoleic (18:2) and linolenic (18:3) acids in all the samples analysed. In Vaccinium myrtillus the amount of carotenoids and fatty acids was higher than in Vaccinium corymbosum species.     

Isolation of a New Carotenoid and Two New Carotenoid Glycosides from Curtobacterium flaccumfaciens pvar poinsettiae

An efficient method for the extraction of the carotenoids from Curtobacterium flaccumfaciens pvar poinsettiae was developed. The glucosides of C.p. 450 (=(all‐E,2R,2′R)‐2‐[4‐(β‐D ‐glucopyranosyloxy)‐3‐methylbut‐2‐enyl]‐2′‐(4‐hydroxy‐3‐methylbut‐2‐enyl)‐β, β‐carotene; 4 ) and of C.p. 473 (=(all‐E,2R,2′S)‐2‐[4‐(β‐D ‐glucopyranosyloxy)‐3‐methylbut‐2‐enyl]‐2′‐(3‐methylbut‐2‐enyl)‐3′,4′‐didehydro‐1′,2′‐dihydro‐β,ψ‐caroten‐1′‐ol; 5 ) were isolated for the first time. In addition, the hitherto unknown 3′,4′‐dihydro derivative of C.p. 450, called C.p. 460 (=(all‐E,2R,2′R)‐2‐(4‐hydroxy‐3‐methylbut‐2‐enyl)‐2′‐(3‐methylbut‐2‐enyl)‐1′,2′‐dihydro‐β,ψ‐caroten‐1′‐ol; 6 ), was identified. The structures were established by UV/VIS, CD, ¹H‐ and ¹³C‐NMR, and mass spectra.     

Antioxidative Activities of Algal Keto Carotenoids Acting as Antioxidative Protectants in the Chloroplast

Very diverse carotenoid structures exist in the photosynthesis apparatus of different algae. Among them, the keto derivatives are regarded the most antioxidative. Therefore, four different keto carotenoids, peridinin, fucoxanthin, siphonaxanthin and astaxanthin fatty acid monoesters, were isolated and purified from Amphidinium carterae, Phaeodactylum tricornutum, Caulerpa taxifolia and Haematococcus pluvialis, respectively. The carotenoids were assayed as inhibitors of photosensitizer initiated reactions or scavengers of radicals in the early events generating reactive oxygen species as starters for peroxidation and as protectants against the whole reaction chain finally leading to lipid peroxidation. These in vitro studies demonstrated the substantial antioxidative properties as indicated by the IC₅₀ values of all four keto carotenoids with superior protection by astaxanthin fatty acid monoesters which were as effective as free astaxanthin and of peridinin against radicals. As an example, the in vivo relevance of fucoxanthin for protection of photosynthesis from excess light and from peroxidative agents was evaluated with intact cells. Cultures of P. tricornutum with decreased fucoxanthin content generated by inhibitor treatment were exposed to strong light or cumene hydroperoxyde. In each case, oxidation of chlorophyll as marker for damaging of the photosynthesis apparatus was less severe when the fucoxanthin was at maximum level.     

Improved Carotenoid Productivity and COD Removal Efficiency by Co-culture of Rhodotorula glutinis and Chlorella vulgaris Using Starch Wastewaters as Raw Material

Utilization of low-cost raw materials for the bio-based chemical production, such as carotenoids, by the co-culture of Rhodotorula glutinis and Chlorella vulgaris has recently become an attractive option. In this study, the primary nutrients of starch wastewater were analyzed, which were used for carotenoid production by the co-culture strategy in a 5-L fermenter around 4000 Lux light intensity. Synergistic effect of gas utilization revealed that the two species could build up the beneficial balance on mutualism. The maximum carotenoid productivity and COD removal efficiency were 12.34 mg/L and 79.6%, respectively, which were higher than those of monoculture yeast (8.31 mg/L and 54.1%). The organic acids, amino acids, and sugar removal efficiencies were increased by 85%, 31%, and 44%, respectively, and more than three kinds of carotenoids were identified compared with those of monoculture yeast. The results demonstrated that the co-culture strategy of two different nutritional microorganisms could significantly improve carotenoid productivity and COD removal efficiency.

     

Carotenoids from the ripening bacterium Brevibacterium linens impart color to the rind of the French cheese,Fourme de Montbrison (PDO)

Abstract

Rind color of some high-value PDO cheeses is related to the presence of carotenoids, but little is known about the structure of the pigmented compounds and their origin. Our objective was to describe the carotenoids extracted from the rind of a French cheese, Fourme de Montbrison, and to compare them with the pigments produced by a bacterial strain used as an adjunct culture in the cheese ripening process. Eleven carotenoids were detected in the cheese rinds or in the biomass of Brevibacterium linens. Most of the carotenoids from the rinds belonged to the aryl (aromatic) carotenoid family, including hydroxylated and non-hydroxylated isorenieratene. Chlorobactene, a carotenoid rarely found in food products, was also detected. Agelaxanthin A was identified in the cheese rinds as well as in the B. linens biomass. Occurrence of this compound was previously described in only one scientific publication, where it was isolated from the sponge Agela schmidtii.     

SURFACE-ENHANCED RESONANCE RAMAN SCATTERING SPECTROSCOPY AS A SURFACE TOPOGRAPHY PROBE IN PLANT PHOTOSYNTHETIC MEMBRANES

Strong resonance Raman (RR) and surface-enhanced resonance Raman scattering (SERRS) signals from carotenoids were detected from thylakoid (stromal-side out) vesicles and inside-out (lumenal-side out) vesicles isolated from spinach chloroplasts. The intensity of the signals from both types of membranes was comparable, indicating that plant carotenoids are exposed on or close to both surfaces or sides of the thylakoid membrane. This is in contrast to previous studies with bacterial photosynthetic membranes (Picorel et al., 1988, J. Biol. Chem. 263 , 4374–4380; and 1990, Biochemistry 29 , 707–712) that show carotenoids selectively located on the cytoplasmic side. In addition_; strong RR and SERRS signals were detected from stacked and unstacked photosystem-II-enriched membrane fragments, demonstrating that carotenoids are also exposed on both surfaces of the appressed region of the thylakoid membrane. Antibodies against the photosystem (PS) II extrinsic proteins blocked SERRS signals from stacked PS II membrane fragments, but only partially affected the SERRS signals from unstacked membranes. The results indicate that these antibodies, which preferentially cover the surface of the original lumenalside of the appressed region, act as spacers between the membrane and SERRS electrode surfaces. The original stromal-side of the appressed region is unaffected. These findings verify the distance sensitivity of the SERRS technique and underscore the above conclusion about the location of carotenoids in the appressed regions. Finally, SERRS signals are sensitive to membrane aging and storage temperature; caution is suggested to those applying SERRS spectroscopy to intact membrane systems.     

Aesculaxanthin,a New Carotenoid Isolated from Pollens of Aesculus hippocastanum

From the pollens of Aesculus hippocastanum, a new apocarotenoid was isolated as the main carotenoid and, based on the spectroscopic data, identified as (all-E,3R)-3-hydroxy-6′-apo-β-caroten-6′-al ( 4 , aesculaxanthin). In addition, (all-E)-lutein ( 3 ) and (all-E)-β-citraurin ( 5 ) were isolated. Furthermore, 6 (aesculaxanthol) was prepared by reduction of 4 with NaBH₄ and tentatively identified as natural carotenoid.     

Allylic Sulfones Containing Triene Moieties as Key Synthons for Carotenoid Synthesis

An efficient synthetic method for the allylic sulfone 2 containing a conjugated triene moiety has been proposed involving i) coupling of allylic sulfones 4 with the C₅ bromoallylic sulfide 5 , ii) base‐promoted dehydrosulfonation in the presence of allylic sulfide, and iii) selective oxidation of the resulting trienyl sulfide to the corresponding sulfone. Total synthesis of lycopene starting from the C₁₅ allylic sulfone 2b has been described, where the new C₁₀ bis(chloroallylic) sulfone 11 proved to be a useful substitute for the C₁₀ bis(chloroallylic) sulfide 3 , which did not require the problematic chemoselective sulfur oxidation in a conjugated polyene.     

The Chemistry of Rose Pigments

In this article we present a survey of the pigments found in the flowers and fruits of old and modern varieties of roses. The yellow colors are produced by carotenoids, the reds by anthocyanins, and the modern oranges by a mixture of the two. The great structural diversity of the carotenoids contrasts with a surprisingly small number of anthocyanins. For the carotenoids found in roses, a clear correspondence exists between the structure and the breeding partners used; the old yellow roses, which arose from crosses with Chinese varieties, mainly contain carotenoids from early stages in the biosynthesis, while in the modern yellow roses, which are descended from Central Asian foetida types, hydroxylations, epoxidations, and epoxide transformations readily occur. A recently elucidated carotenoid degradation sequence follows the scheme C₄₀ → C₁₃ + C₂₇ → C₁₃ + C₁₄. The C₁₃ compounds are odoriferous substances that contribute to the scent of roses. In the physiological pH region, copigmentation with flavonol glycosides is crucial for stabilization of the anthocyanin chromophores. Many roses, including the “apothecary's rose”, which was once used medicinally, contain large amounts of strongly astringent ellagitannins, monosaccharide esters of gallic acid.     

Preparation of Partially Acetylated Carotenoids

Partially acetylated carotenoids were prepared from fully acetylated carotenoids by reaction with NaBH₄, and were characterized by UV/VIS, CD, ¹H‐NMR and mass spectra. The 3,6′‐diacetate, 3′,6′‐diacetate, and 6′‐acetate 10 – 12 , respectively, of (6′R)‐capsanthol (=(3R,3′S,5′R,6′R)‐β,κ‐carotene‐3,3′,6′‐triol; 4 ) were obtained from (6′R)‐capsanthol‐3,3′,6′‐triacetate ( 9 ), and the 3‐ and 3′‐acetates 13 and 14 , respectively, of 4 from (6′R)‐capsanthol 3,3′‐diacetate ( 8 ). The utility of this method was also demonstrated by the preparation of zeaxanthin and lutein monoacetates 16, 19 , and 20 .     

Determination of carotenoids in two algae species from the saline water of Kapulukaya reservoir by HPLC

The local algae species, Chlorella vulgaris and Scenedesmus regularis, from a highly saline water body of Kapulukaya Reservoir were isolated to analyze their carotenoid composition and content using HPLC method. The gradient solvent system of methanol–acetonitrile–water (84:14:2, v/v/v) and methylene chloride (100%), used to resolve a range of carotenoids from the saponified cells, proved an acceptable separation as inferred from the retention factor (k) ranging between 0.75 and 7.76 and the separation factor (α) values greater than 1. Resolution peaks assigned to carotenoids, 21 for C. vulgaris extracts and 22 for S. regularis extracts, were reached within the duration time of 45?min. Main carotenoids identified either tentatively or positively were all-trans-lutein, 9- or 9′-cis-lutein, 13- or 13′-cis-lutein, cis-lutein, All-trans-α-carotene, 9- or 9′-cis-α-carotene, All-trans-β-carotene, 9- or 9′-cis-β-carotene in the species except for all-trans-β-cryptoxanthin found only in S. regularis. Auroxanthin, neochrome, neoxanthin, and cis-neoxanthin were identified as epoxy-containing compounds. Quantitatively, C. vulgaris was distinguished to have greater amount of lutein and cis-isomers (2.74?mg/g), 77.89% while S. regularis was predominated by β-carotene and cis isomers as major component, being 80.72% (5.76?mg/g) in total carotenoids (TC). In terms of total carotenoids, the species were considered to be efficient sources for further practical applications.     

Phenolic,Carotenoid and Saccharide Compositions of Vietnamese Camellia sinensis Teas and Herbal Teas

Tea (Camellia sinensis) and herbal tea have been recognized as rich sources of bioactive constituents with the ability to exert antioxidant actions. The aims of this study were to analyze phenolic, carotenoid and saccharide contents in a set of Vietnamese tea and herbal tea and compare the results with those of green and black teas marketed in the U.S. In total, 27 phenolics, six carotenoids and chlorophylls, and three saccharides were quantitatively identified. Catechins, quercetin glycosides and chlorogenic acid were the predominating phenolics in the teas, with the concentrations following the order: jasmine/green teas > oolong tea > black tea. Lutein was the dominant carotenoid in the teas and its concentrations were generally found to be higher in the jasmine and green teas than in the oolong and black teas. The study showed that the green teas originating in Vietnam had much higher levels of phenolics and carotenoids than their counterparts stemming from another country. The application of partial least squares discriminant analysis (PLS-DA) as a chemometric tool was able to differentiate phenolic profiles between methanolic extracts and tea infusions. Through principal component analysis (PCA), the similarities and dissimilarities among the jasmine, green, oolong, black teas and herbal teas were depicted.     

FT‐IR Study of Some Carotenoids

Some natural and semisynthetic carotenoids were examined by means of FT‐IR spectroscopy. The IR bands of the characteristic functional groups (CH₃, CH₂, CC, CO, OH, etc.) were assigned when possible. Some special functional groups – without H‐atoms – such as CCC, ‘cross epoxides', etc., which cannot be easily identified by ¹H‐NMR methods, were also detected in the FT‐IR spectra.     

Report of in vitro antileishmanial properties of Iberian macroalgae

Here is reported the anti Leishmania infantum activity of 48 hexane, CH₂Cl₂ and MeOH extracts from 16 macroalgae collected on the Iberian Coast. Seven hexane and CH₂Cl₂ Cystoseira baccata, Cystoseira barbata, Cystoseira tamariscifolia, Cystoseira usneoides, Dictyota spiralis and Plocamium cartilagineum extracts were active towards promastigotes (IC₅₀ 29.8–101.8 μg/mL) inducing strong morphological alterations in the parasites. Hexane extracts of C. baccata and C. barbata were also active against intracellular amastigotes (IC₅₀ 5.1 and 6.8 μg/mL, respectively). Fatty acids, triacylglycerols, carotenoids, steroids and meroterpenoids were detected by nuclear magnetic resonance (NMR), and gas chromatography in the Cystoseira extracts. These results suggest that Cystoseira macroalgae contain compounds with antileishmanial activity, which could be explored as scaffolds to the development of novel sources of antiparasitic derivatives.     

Carotenoid photoprotection in artificial photosynthetic antennas

A series of phthalocyanine-carotenoid dyads in which a phenylamino group links a phthalocyanine to carotenoids having 8-11 backbone double bonds were examined by visible and near-infrared femtosecond pump-probe spectroscopy combined with global fitting analysis. The series of molecules has permitted investigation of the role of carotenoids in the quenching of excited states of cyclic tetrapyrroles. The transient behavior varied dramatically with the length of the carotenoid and the solvent environment. Clear spectroscopic signatures of radical species revealed photoinduced electron transfer as the main quenching mechanism for all dyads dissolved in a polar solvent (THF), and the quenching rate was almost independent of carotenoid length. However, in a nonpolar solvent (toluene), quenching rates displayed a strong dependence on the conjugation length of the carotenoid and the mechanism did not include charge separation. The lack of any rise time components of a carotenoid S(1) signature in all experiments in toluene suggests that an excitonic coupling between the carotenoid S(1) state and phthalocyanine Q state, rather than a conventional energy transfer process, is the major mechanism of quenching. A pronounced inhomogeneity of the system was observed and attributed to the presence of a phenyl-amino linker between phthalocyanine and carotenoids. On the basis of accumulated work on various caroteno-phthalocyanine dyads and triads, we have now identified three mechanisms of tetrapyrrole singlet excited state quenching by carotenoids in artificial systems: (i) Car-Pc electron transfer and recombination; (ii)(1) Pc to Car S(1) energy transfer and fast internal conversion to the Car ground state; (iii) excitonic coupling between (1)Pc and Car S(1) and ensuing internal conversion to the ground state of the carotenoid. The dominant mechanism depends upon the exact molecular architecture and solvent environment. These synthetic systems are providing a deeper understanding of structural and environmental effects on the interactions between carotenoids and tetrapyrroles and thereby better defining their role in controlling natural photosynthetic systems.     

HPLC‐PAD‐atmospheric pressure chemical ionization‐MS metabolite profiling of cytotoxic carotenoids from the echinoderm Marthasterias glacialis (spiny sea‐star)

An HPLC‐PAD‐atmospheric pressure chemical ionization‐MS metabolite profiling analysis was conducted on the marine echinoderm Marthasterias glacialis (spiny sea‐star). Bio‐guided purification of the methanolic extract led to the isolation of several carotenoids, namely zeaxanthin, astaxanthin and lutein. These compounds were characterized using both UV–Vis characteristics and MS spectra interpretation. No previous works addressed the MS analysis of carotenoids present in this organism. The purified carotenoid fraction displayed a strong cell proliferation inhibition against rat basophilic leukemia RBL‐2H3 (IC₂₅=268 μg/mL) cancer cell line. Against healthy V79 (rat lung fibroblasts (IC₂₅=411 μg/mL)) cell line, however, toxicity was lower, as it is desired for anti‐cancer molecules. This study suggests that M. glacialis may constitute a good source of bioactive compounds that can be used as lead compounds for the pharmaceutical industry.     

Antioxidant Activity and Protective effect of Banana Peel against Oxidative Hemolysis of Human Erythrocyte at Different Stages of Ripening

Phytochemicals such as polyphenols and carotenoids are gaining importance because of their contribution to human health and their multiple biological effects such as antioxidant, antimutagenic, anticarcinogenic, and cytoprotective activities and their therapeutic properties. Banana peel is a major by-product in pulp industry and it contains various bioactive compounds like polyphenols, carotenoids, and others. In the present study, effect of ripening, solvent polarity on the content of bioactive compounds of crude banana peel and the protective effect of peel extracts of unripe, ripe, and leaky ripe banana fruit on hydrogen peroxide-induced hemolysis and their antioxidant capacity were investigated. Banana (Musa paradisica) peel at different stages of ripening (unripe, ripe, leaky ripe) were treated with 70% acetone, which were partitioned in order of polarity with water, ethyl acetate, chloroform (CHCl₃), and hexane sequentially. The antioxidant activity of the samples was evaluated by the red cell hemolysis assay, free radical scavenging (1,1-diphenyl-2-picrylhydrazyl free radical elimination) and superoxide dismutase activities. The Folin–Ciocalteu's reagent assay was used to estimate the phenolic content of extracts. The findings of this investigation suggest that the unripe banana peel sample had higher antioxidant potency than ripe and leaky ripe. Further on fractionation, ethyl acetate and water soluble fractions of unripe peel displayed high antioxidant activity than CHCl₃ and hexane fraction, respectively. A positive correlation between free radical scavenging capacity and the content of phenolic compound were found in unripe, ripe, and leaky ripe stages of banana peel.     

Biogeneration of C13-norisoprenoid compounds: experiments supportive for an apo-carotenoid pathway in grapevines

The first step of the proposed biogenetic pathway in grapes that leads from carotenoids to C₁₃-norisoprenoids involves the enzymatic degradation of carotenoids by regiospecific oxygenases. Chemical, photochemical and oxidase-coupled degradation of carotenoids to norisoprenoids have been studied in vitro and enzymatic systems shown to be involved in mammals. However, no enzymatic system has been shown to be involved in the formation of C₁₃-norisoprenoids in grapes, despite all recent studies carried out on grapevines supporting such a model. These findings include the preponderance of norisoprenoids possessing 13 carbon atoms that thereby indicates the specificity of cleavage, the configuration of the asymmetric centres and axes common to C₁₃-norisoprenoids and the corresponding carotenoids, the negative correlations observed between the levels of norisoprenoids and carotenoids during berry development, and the in vivo transfer of ¹³C markers from carotenoids to norisoprenoids in berries between véraison and berry maturity. All of these findings are major arguments in favour of the hypothesis that glycosylated C₁₃-norisoprenoids derive from carotenoids in grape berries. Carotenoids are mostly synthesised from the first stage of fruit formation until véraison, and then degrade between véraison and maturity to produce glycosylated C₁₃-norisoprenoids and other compounds.