Two‐dimensional liquid chromatography analysis of all‐trans‐, 9‐cis‐, and 13‐cis‐astaxanthin in raw extracts from Phaffia rhodozyma

An effective two‐dimensional liquid chromatography method has been established for the analysis of all‐trans‐astaxanthin and its geometric isomers from Phaffia rhodozyma employing a C18 column at the first dimension and a C30 column in the second dimension, connected by a 10‐port valve using the photo‐diode array detector. The regression equation of astaxanthin calibration curve was established, and the precision and accuracy values were found to be in the range of 0.32–1.14% and 98.21–106.13%, respectively. By using two‐dimensional liquid chromatography, it was found that day light, ultrasonic treatment, and heat treatment have significant influence on the content of all‐trans‐astaxanthin in the extract from P. rhodozyma due to the transformation of all‐trans‐astaxanthin to cis‐astaxanthin. The day light and ultrasonic treatments more likely transform all‐trans‐astaxanthin to 9‐cis‐astaxanthin, and the thermal treatment transforms all‐trans‐astaxanthin to 13‐cis‐astaxanthin. These results indicate that the two‐dimensional liquid chromatography method can facilitate monitoring astaxanthin isomerization in the raw extract from P. rhodozyma. In addition, the study will provide a general reference for monitoring other medicals and bioactive chemicals with geometric isomers.     

Metabolism of carotenoids in salmonids. Part 2. Distribution and absolute configuration of idoxanthin in various organs and tissues of one atlantic salmon (Salmo salar,L.) fed with astaxanthin

The content of total carotenoids and the ratio astaxanthin/idoxanthin ( = 3,3′-dihydroxy-β,β-carotene-4,4′-dione/3,3′,4′-trihydroxy-β,β-caroten-4-one) in varoius organs and tissues of one Atlantic salmon (Salmo salar, L.) reared indoors in a tank were analyzed after feeding ‘racemic’ ((3R,3′R)/(3R,3′S; meso)/(3S,3′S) 1:2:) astaxanthin (90 mg/kg feed) during one yera. Configurational analysis of astaxanthin was carried out via the (?)-dicamphanate derivative and that of idoxanthin after reaction with (+)-(S)-l-(l-naphthyl)ethyl isocyanate. Separation of all eight optical isomers of idoxanthin-tricarbamate derivatives by HPLC is described. In salmon, enzymatic reduction of astaxanthin was found to be sterospecific leading to th (4′R)-hydroxy group irrespective of the configuration at C(3′), thus resulting in four different stereoisomers of idoxanthin formed from (3R,3′R), (3R,3′S; meso)-, and (3S3′S)-astaxanthin present in the diet.     

Metabolism of Carotenoids in Salmonids. Part 3. Metabolites of astaxanthin and canthaxanthin in the skin of atlantic salmon (salmo salar,L.)

Diets supplemented with astaxanthin and canthaxanthin, respectively, and a control diet without carotenoid additions, were fed to 1½-year-old Atlantic salmon (Salmo salar, L.) for one year. The integuments were investigated as to their quantitative and qualitative carotenoid composition. Astaxanthin and canthaxanthin deposited in the skin amounted to 20 and 14% of the total carotenoids only. Seventy % must be considered as metabolites of astaxanthin and canthaxanthin and 10% as basic xanthophylls also present in the control groups. Astaxanthin apparently underwent the following metabolic pathway: astaxanthin→idoxanthin→adonixanthin→zeaxanthin→zeaxanthin 5,6-epoxides. Reduction of the 4′-carbonyl group was stereospecific leading to the (4′R)-idoxanthin. Canthaxanthin was obviously converted to β,β-carotene via 4′-hydroxyechinenone, echinenone, and 4-hydroxy-β,β-carotene.     

Study of Growth Parameters for Phaffia rhodozyma Cultivated in Peat Hydrolysates

The yeastPhaffia rhodozyma, known for its ability to synthesize carotenoids, was adapted and cultivated in liquid-phase media using peat hydrolysates as the main substrate source. The hydrolysates were prepared using high-pressure treatment at 185°C, without the addition of acid. The growth of the yeast was studied as a function of the pH, temperature, culture time, and agitation speed. The best conditions for the growth of the yeast were: a pH of 7, a temperature of 18°C, 5-d culture time, and 200-rpm agitation. Under those conditions, P.rhodozyma produced a concentration of 1279.82 μg carotenoids g^-1 dry yeast, which compares well with other previously reported results.     

Production of Astaxanthin from Cellulosic Biomass Sugars by Mutants of the Yeast <Emphasis Type="Italic">Phaffia rhodozyma</Emphasis>

Astaxanthin is a potential high-value coproduct in an ethanol biorefinery. Three mutant strains of the astaxanthin-producing yeast Phaffia rhodozyma, which were derived from the parent strain ATCC 24202 (UCD 67-210) and designated JTM166, JTM185, and SSM19, were tested for their capability of utilizing the major sugars that can be generated from cellulosic biomass, including glucose, xylose, and arabinose, for astaxanthin production. While all three strains were capable of metabolizing these sugars, individually and in mixtures, JTM185 demonstrated the greatest sugar utilization and astaxanthin production. Astaxanthin yield by this strain (milligrams astaxanthin per gram of sugar consumed) was highest for xylose, followed by arabinose and then glucose. The kinetics of sugar utilization by strain JTM185 was studied in fermenters using mixtures of glucose, xylose, and arabinose at varied concentrations. It was found that glucose was utilized preferentially, followed by xylose, and lastly, arabinose. Astaxanthin yield was significantly affected by sugar concentrations. Highest yields were observed with sugar mixtures containing the highest concentrations of xylose and arabinose. Hydrolysates produced from sugarcane bagasse and barley straw pretreated by the soaking in aqueous ammonia method and hydrolyzed with the commercial cellulase preparation, Accellerase™ 1000, were used for astaxanthin production by the mutant strain JTM185. The organism was capable of metabolizing all of the sugars present in the hydrolysates from both biomass sources and produced similar amounts of astaxanthin from both hydrolysates, although these amounts were lower when compared to yields obtained with reagent grade sugars.     

Astaxanthinogenesis in the yeastPhaffia rhodozyma

Astaxanthin is a diketo-dihydroxy-carotenoid produced byPhaffia rhodozyma, a basidiomicetous yeast. A low-cost fermentation medium consisting of raw sugarcane juice and urea was developed to exploit the active sucrolytic/urelolytic enzyme apparatus inherent to the yeast. As compared to the beneficial effect of 0.1 g% urea, a ready nitrogen source, mild phosphoric pre inversion of juice sucrose to glucose and fructose, promptly fermentable carbon sources, resulted in smaller benefits. Corn steep liquor (CSL) was found to be a valuable supplement for both yeast biomass yield (9.2 g dry cells/L) and astaxanthin production (1.3 mg/g cells). Distillery effluent (vinace), despite only a slightly positive effect on yeast growth, allowed for the highest pigment productivity (1.9 mg/g cells). Trace amounts of Ni² (1 mg/L, as a cofactor for urease) resulted in controversial effects, namely, biomass decrease and astaxanthin increase, with no effect on the release (and uptake) of ammonium ion from urea. Since the synthesized astaxanthin is associated with the yeast cell and the pigment requires facilitated release for aquaculture uses (farmed fish meat staining), an investigation of the yeast cell wall was undertaken using detergent-treated cells. The composition of the rigid yeast envelope was found to be heterogeneous. Its partial acid or enzymatic depolymerization revealed glucose and xylose as common monomeric units of the cell-wall glycopolymers. Yeast cell-wall partial depolymerization with appropriate hydrolases may improve the pigment bioavailability for captive aquatic species and poultry.

     

Screening Factors Influencing the Production of Astaxanthin from Freshwater and Marine Microalgae

Astaxanthin, a carotenoid pigment found in several aquatic organisms, is responsible for the red colour of salmon, trout and crustaceans. In this study, astaxanthin production from freshwater microalga Chlorella sorokiniana and marine microalga Tetraselmis sp. was investigated. Cell growth and astaxanthin production were determined spectrophotometrically at 620 and 480 nm, respectively. Astaxanthin was extracted using acetone and measured subsequent to biomass removal. Aerated conditions favoured astaxanthin production in C. sorokiniana, whereas Tetraselmis sp. was best cultured under unaerated conditions. C. sorokiniana produced more astaxanthin with the highest yield reached at 7.83 mg/l in 6.0 mM in nitrate containing medium compared to Tetraselmis sp. which recorded the highest yield of only 1.96 mg/l in 1.5 mM nitrate containing medium. Production in C. sorokiniana started at the early exponential phase, indicating that astaxanthin may be a growth-associated product in this microalga. Further optimization of astaxanthin production was performed using C. sorokiniana through a 2³ full factorial experimental design, and a yield of 8.39 mg/l was achieved. Overall, the study has shown that both microalgae are capable of producing astaxanthin. Additionally, this research has highlighted C. sorokiniana as a potential astaxanthin producer that could serve as a natural astaxanthin source in the current market.     

Natural Occurrence of Enantiomeric and meso-Astaxanthin. 5. Ex wild salmon (Salmo salar and Oncorhynchus)

The configurational isomers of astaxanthin (3,3′-dihydroxy-β,β-carotene-4,4′-dione) from the flesh of salmon (Salmo salar and Oncorhynchus) caught at different places in Europe and Canada were isolated and analyzed as (?)-camphanic acid diesters by means of HPLC. The biological variation in the composition of the configurational isomers in seven fish was surprisingly similar: 78 to 85% of (3S, 3′S)-astaxanthin, 12 to 17% (3R, 3′R)-astaxanthin and 2 to 6% meso-astaxanthin.     

Carotenoids of the Carotenoprotein Asteriarubin. Optical Purity of Asterinic Acid

The carotenoid composition of the carotenoprotein asteriarubin ex the starfish Asterias rubens, determined by HPLC, comprised canthaxanthin ( 6 , 3% of total), all-trans-astaxanthin ( 1 , 14%), all-trans-7,8-didehydroastaxanthin ( 2 , 24%), all-trans-7,8,7′,8′-tetradehydroastaxanthin ( 3 , 43%) and 4-oxomytiloxanthin ( 7 , 10%). The previously unknown 4-oxomytiloxanthin was tentatively identified by the UV./VIS., 'H-NMR. spectra and MS. data. The optical purity was determined by HPLC. of the di-(?)-camphanates, by comparison with those of synthetic standards: 7,8,7′, 8′-tetradehydroastaxanthin (92% (3S, 3′S), 2% meso), 7,8-didehydroastaxanthin (89% (3S,3′S), 2% meso?), and astaxanthin (78% (3S,3′S), 14% (3R,3′S), and 5% (3R,3′R)). It is concluded that the acetylenic derivatives of astaxanthin in contrast to astaxanthin from marine animal sources are essentially pure (3S, 3′S)-isomers. This might reflect their probable metabolic formation by 4-oxo modification of acetylenic (3R,3′R)-carotenols ex Mytilus edulis in their diet.     

Enzymatic Fractionation of SAA-Pretreated Barley Straw for Production of Fuel Ethanol and Astaxanthin as a Value-Added Co-Product

Barley straw was used to demonstrate an integrated process for production of fuel ethanol and astaxanthin as a value-added co-product. Barley straw was pretreated by soaking in aqueous ammonia using the previously determined optimum conditions, which included 77.6 °C treatment temperature, 12.1 h treatment time, 15 wt% ammonia concentration, and 1:8 solid-to-liquid ratio. In the newly developed process, the pretreated barley straw was first hydrolyzed with ACCELLERASE® XY (a commercial hemicellulase product) to generate a xylose-rich solution, which contained 3.8 g/l glucose, 22.9 g/l xylose, and 2.4 g/l arabinose, with 96 % of the original glucan being left intact. The xylose-rich solution was used for production of astaxanthin by the yeast Phaffia rhodozyma without further treatment. The resulting cellulose-enriched solid residue was used for ethanol production in a fed-batch simultaneous saccharification and fermentation using ACCELLERASE® 1500 (a commercial cellulase product) and the industrial yeast Saccharomyces cerevisiae. At the end of the fermentation, 70 g/l ethanol was obtained, which was equivalent to 63 % theoretical yield based on the glucan content of the solid substrate.     

A peri-Xanthenoxanthene Centered Columnar-Stacking Organic Semiconductor for Efficient,Photothermally Stable Perovskite Solar Cells

Modulating the structure and property of hole-transporting organic semiconductors is of paramount importance for high-efficiency and stable perovskite solar cells (PSCs). This work reports a low-cost peri-xanthenoxanthene based small-molecule P1, which is prepared at a total yield of 82 % using a three-step synthetic route from the low-cost starting material 2-naphthol. P1 molecules stack in one-dimensional columnar arrangement characteristic of strong intermolecular π–π interactions, contributing to the formation of a solution-processed, semicrystalline thin-film exhibiting one order of magnitude higher hole mobility than the amorphous one based on the state-of-the art hole-transporter, 2,2-7,7-tetrakis(N,N′-di-paramethoxy-phenylamine 9,9′-spirobifluorene (spiro-OMeTAD). PSCs employing P1 as the hole-transporting layer attain a high efficiency of 19.8 % at the standard AM 1.5 G conditions, and good long-term stability under continuous full sunlight exposure at 40 °C.     

Synthesis and applications of 2,7‐carbazole‐based conjugated main‐chain copolymers containing electron deficient bithiazole units for organic solar cells

A series of low‐band‐gap (LBG) donor–accepor conjugated main‐chain copolymers ( P1 – P4 ) containing planar 2,7‐carbazole as electron donors and bithiazole units (4,4′‐dihexyl‐2,2′‐bithiazole and 4,4′‐dihexyl‐5,5′‐di(thiophen‐2‐yl)‐2,2′‐bithiazole) as electron acceptors were synthesized and studied for the applications in bulk heterojunction (BHJ) solar cells. The effects of electron deficient bithiazole units on the thermal, optical, electrochemical, and photovoltaic (PV) properties of these LBG copolymers were investigated. Absorption spectra revealed that polymers P1 – P4 exhibited broad absorption bands in UV and visible regions from 300 to 600 nm with optical band gaps in the range of 1.93–1.99 eV, which overlapped with the major region of the solar emission spectrum. Moreover, carbazole‐based polymers P1 – P4 showed low values of the highest occupied molecular orbital (HOMO) levels, which provided good air stability and high open circuit voltages (V_oc) in the PV applications. The BHJ PV devices were fabricated using polymers P1 – P4 as electron donors and (6,6)‐phenyl‐C₆₁‐butyric acid methyl ester (PC₆₁BM) or (6,6)‐phenyl‐C₇₁‐butyric acid methyl ester (PC₇₁BM) as electron acceptors in different weight ratios. The PV device bearing an active layer of polymer blend P4:PC₇₁BM (1:1.5 w/w) showed the best power conversion efficiency value of 1.01% with a short circuit current density (J_sc) of 4.83 mA/cm², a fill factor (FF) of 35%, and V_oc = 0.60 V under 100 mW/cm² of AM 1.5 white‐light illumination. © 2010 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem, 2010     

Photosensitization of the yeast phaffia rhodozyma at a low temperature for screening carotenoid hyperproducing mutants

Phaffia rhodozyma strain Ant-1 produces more carotenoids, known as antioxidants, but it was more sensitive to light plus toluidine blue O (TBO), a superoxide producer, than wild strain 67-385 at 20°C. Carotenoid hyper-producing mutants (CHMs), Ant-1 and 2A2N, exhibited decreased activity of superoxide dismutase (SOD) compared to 67-385, and this is in part responsible for hypersensitivity of the mutants to photosensitization. Light plus TBO at 2°C allowed carotenoid hyperproducing mutants to produce higher colony-forming units than the wild-type. Photosensitization with limited cell metabolism by a low temperature, provides an idea of selective conditions for carotenoid hyperproducers ofP. rhodozyma.     

Synthesis of Amino Acid Derivatives of Neamine and 2-Deoxystreptamine to be used as Mutasynthons

ABSTRACT

6′-N derivatives of neamine with alanine, phenylalanine and lysine were synthesized either using an active esters method in one step under controlled conditions, or using a mixed anhydride method after blocking every functional group of neamine and leaving the 6′-amino group free to react. Similarly N,N′-diamino acid and monoamino acid derivatives of 2-deoxystreptamine were synthesized.     

Temperature and Concentration Dependent Circular Dichroism of Mono- and Di-cis Isomers of (3S,3′S)-Astaxanthin Diacetate

The circular dichroism (CD.) spectra of all-trans-(3S, 3′S) astaxanthin diacetate and its 9-cis, 13-cis, 9,9′-di-cis, 9,13′-di-cis, and 9,13-di-cis isomers conform to the rules previously formulated for optically active carotenoids with a 4-oxo-β-end ring containing an asymmetric C-atom [1]. Thus the CD. bands of the all-trans and the di-cis isomers show the same signs whereas those of the mono-cis isomers have opposite signs. The CD. spectra of all the astaxanthin diacetate isomers invert sign upon cooling to ?180°. The CD. spectra of the 9-mono-cis and 9,9′-di-cis isomers and to a lesser extent also those of the 9, 13′-di-cis and 9, 13-di-cis isomers are concentration dependent at ?180°, with the longest wavelength band giving at the higher concentration a bisignate CD. curve under the main absorption characteristic of aggregation. This phenomenon has been observed only in isomers with a 9-cis linkage. It is suggested that steric hindrance prevents such aggregation taking place in the other isomers.     

trans/cis Isomerization of Astaxanthin Diacetate/Isolation by HPLC. and Identification by 1H-NMR. Spectroscopy of Three Mono-cis- and Six Di-cis-Isomers

Thermal and iodine-catalyzed photochemical trans/cis isomerization of synthetic, racemic astaxanthin diacetate (3,3′-dihydroxy-β,β-carotene-4,4′-dione diacetate) yielded multi-component mixtures of cis-isomers. Separation and isolation of 10 different cis-isomers in quantities between 10 and 70 μg was achieved by HPLC. Investigation of their 270-MHz-FT-¹H-NMR. spectra led to the identification of 9 of these isomers, namely the 9-, 13-, and 15-mono-cis-, the 9,9′-, 9,13-, 9,13′-, 9,15-, 13,13′-, and 13,15-di-cis-astaxanthin diacetate.     

Enzymatic Conversion of Cypridina Luciferyl Sulfate to Cypridina Luciferin with Coenzyme A as a Sulfate Acceptor in Cypridina (Vargula) hilgendorfii

In the luminous ostracod Cypridina (presently Vargula) hilgendorfii, Cypridina luciferyl sulfate (3‐enol sulfate of Cypridina luciferin) is converted to Cypridina luciferin by a sulfotransferase with 3′‐phosphoadenosine‐5′‐phosphate (PAP) as a sulfate acceptor. The resultant Cypridina luciferin is used for the luciferase–luciferin reaction of Cypridina to emit blue light. The luminescence stimulation with major organic cofactors was examined using the crude extracts of Cypridina specimens, and we found that the addition of coenzyme A (CoA) to the crude extracts significantly stimulated luminescence intensity. Further, the light‐emitting source in the crude extracts stimulated with CoA was identified as Cypridina luciferyl sulfate, and we demonstrated that CoA could act as a sulfate acceptor from Cypridina luciferyl sulfate. In addition, the sulfate group of Cypridina luciferyl sulfate was also transferred to adenosine 5′‐monophosphate (5′‐AMP) and adenosine 3′‐monophosphate (3′‐AMP) by a sulfotransferase. The sulfated products corresponding to CoA, 5′‐AMP and 3′‐AMP were identified using mass spectrometry. This is the first report that CoA can act as a sulfate acceptor in a sulfotransferase reaction.     

Synthese von optisch aktiven,natürlichen Carotinoiden und strukturell verwandten Naturprodukten. II. Synthese von (3 S, 3′S)-Astaxanthin

Synthesis of optically active natural carotenoids and structurally related compounds. II. Synthesis of (3S, 3′S)-astaxanthin The syntheses of rac. astaxanthin, (3 S, 3′S)-astaxanthin ( 1 ), its 15-cis isomer ( 21 ), its diacetate ( 22 ), and of (3 S, 3′ S)-15, 15′-didehydroastaxanthin ( 20 ) are reported.     

Gold(I)/Chiral Brønsted Acid Catalyzed Enantioselective Hydroamination–Hydroarylation of Alkynes: The Effect of a Remote Hydroxyl Group on the Reactivity and Enantioselectivity

The catalytic enantioselective hydroamination–hydroarylation of alkynes under the catalysis of (R₃P)AuMe/(S)‐3,3′‐bis(2,4,6‐triisopropylphenyl)‐1,1′‐binaphthyl‐2,2′‐diyl hydrogenphosphate ((S)‐TRIP) is reported. The alkyne was reacted with a range of pyrrole‐based aromatic amines to give pyrrole‐embedded aza‐heterocyclic scaffolds bearing a quaternary carbon center. The presence of a hydroxyl group in the alkyne tether turned out to be very crucial for obtaining products in high yields and enantioselectivities. The mechanism of enantioinduction was established by carefully performing experimental and computational studies.     

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.