Novel methoxy-carotenoids from the burgundy-colored plumage of the Pompadour Cotinga Xipholena punicea

Recent advances in the fields of chromatography, mass spectrometry, and chemical analysis have greatly improved the efficiency with which carotenoids can be extracted and analyzed from avian plumage. Prior to these technological developments, Brush (1968) [1] concluded that the burgundy-colored plumage of the male pompadour Cotinga Xipholena punicea is produced by a combination of blue structural color and red carotenoids, including astaxanthin, canthaxanthin, isozeaxanthin, and a fourth unidentified, polar carotenoid. However, X. punicea does not in fact exhibit any structural coloration. This work aims to elucidate the carotenoid pigments of the burgundy color of X. punicea plumage using advanced analytical methodology. Feathers were collected from two burgundy male specimens and from a third aberrant orange-colored specimen. Pigments were extracted using a previously published technique (McGraw et al. (2005) [2]), separated by high-performance liquid chromatography (HPLC), and analyzed by UV/Vis absorption spectroscopy, chemical analysis, mass spectrometry, nuclear magnetic resonance (NMR), and comparison with direct synthetic products. Our investigation revealed the presence of eight ketocarotenoids, including astaxanthin and canthaxanthin as reported previously by Brush (1968) [1]. Six of the ketocarotenoids contained methoxyl groups, which is rare for naturally-occurring carotenoids and a novel finding in birds. Interestingly, the carotenoid composition was the same in both the burgundy and orange feathers, indicating that feather coloration in X. punicea is determined not only by the presence of carotenoids, but also by interactions between the bound carotenoid pigments and their protein environment in the barb rami and barbules. This paper presents the first evidence of metabolically-derived methoxy-carotenoids in birds.     

Somatic instability of carotenoid biosynthesis in the tomato ghost mutant and its effect on plastid development

The tomato (Lycopersicon esculentum (L.) Mill.) ghost plant is a mutant of the San Marzano cultivar affected in carotenoid biosynthesis. ghost plants exhibit a variable pattern of pigment biosynthesis during development. Cotyledons are green but true leaves are white. Green sectors, which appear to be clonal in origin, are frequently observed in the white tissue. Because of the lack of photosynthesis ghost plants have a very low viability in soil. We have developed a strategy for propagating ghost plants that employs organ culture to generate variegated green-white plants which, supported by the photosynthetic green areas, develop in soil to almost wild-type size. These plants were used to analyze the pigment content of the different tissues observed during development and plastid ultrastructure. Cotyledons and green leaves contain both colored carotenoids and chlorophyll but only the colorless carotenoid phytoene accumulates in white leaves. the plastids in the white tissue of ghost leaves lack internal membrane structures but normal chloroplasts can be observed in the green areas. The chromoplasts of white fruits are also impaired in their ability to form thylakoid membranes.     

Ultrastructure of carotenoid deprivation in photoreceptors of Manduca sexta: myeloid bodies and intracellular microvilli

Summary The ultrastructural effects of carotenoid (vitamin A) deprivation were studied in the adult photoreceptors of the tobacco hornworm moth Manduca sexta. Moths were reared on a deprived diet, which lacked the carotenoid sources of the photopigment chromophore, 3-hydroxy retinal, or on a control, fortified diet, containing ample carotenoid. The latter supported normal levels of visual function, whereas visual pigment and sensitivity were reduced by at least 3 log units in moths reared on the deprived diet. Myeloid bodies, massed cisternae of hypertrophied smooth endomembrane, filled the cytoplasm in the receptors of deprived animals. The myeloid bodies assumed various configurations that included lamellate stacks of parallel cisternae, and tubular networks in a paracrystalline form. Freeze-fracture preparations of myeloid membranes revealed a high density of P-face particles. Vacuoles containing microvilli similar to those of the rhabdomere were also present in deprived photoreceptors. We suggest that the elaboration of smooth endoplasmic reticulum as myeloid bodies in chromophore-deprived photoreceptors may stem from the hypertrophy of a biochemical system for processing the chromophore or the interruption of the intracellular pathway that normally carries visual pigment to the rhabdomere.     

Linear dichroism and orientational studies of carotenoid Langmuir-Blodgett films

The linear dichroism of single monolayers of lutein, zeaxanthin and a mixture of lutein and synthetic phosphatidylcholine has been measured. The angle of orientation of the carotenoid molecules was found to lie between 45° and 51° relative to the plane of the solid support. Although the adsorbed monolayers were mostly in a monomeric state, microscopic observations, as well as the II-A isotherms, indicated the existence of crystalline islets. The results have been interpreted in connection with Haidinger's polarization brushes.     

Carotenoid composition in milo (Sorghum vulgare) shoots as affected by phytochrome and chlorophyll

The composition of coloured carotenoids in the milo shoot was investigated quantitatively (high performance liquid chromatography) during light-mediated plastidogenesis, including the time span of photodelay as caused by medium and high light fluxes. It was found that as long as only the far-red-absorbing form of phytochrome operates, the carotenoid pattern remains virtually the same as in complete darkness (violaxanthin and lutein as major constituents, traces of -carotene). On the other hand, the pattern changes dramatically in white or red light with increasing amounts of chlorophyll (lutein and -carotene dominate, -carotene showing the strongest relative increase). Photodelay during the early phase of plastidogenesis affects the carotenoid composition strongly. Increase of neoxanthin, violaxanthin and -carotene contents are diminished while lutein accumulation proves resistant towards chlorophyll-mediated photoinhibition. The photodelay can be diminished by an appropriate light pretreatment. The data indicate that light-mediated control over carotenoid accumulation is exerted at three levels: i) a coarse control through phytochrome, ii) fine tuning in connection with chlorophyll accumulation, iii) stabilization of holocomplexes against photodecomposition.Abbreviations GG14 high fluence rate green-yellow light - HPLC high-performance liquid chromatography - Chl chlorophyll - WL_w weak white light (1200 lx) - WL_m medium flux white light (12000 lx)     

The carotenoid and abscisic acid content of viviparous kernels and seedlings ofZea mays L.

Carotenoid and abscisic acid (ABA) levels were determined in endosperm, embryos and seedlings of wild-type and viviparous (vp) mutants ofZea mays L. Carotenoid concentrations were determined by absorption spectrometry following purification by high-performance liquid chromatography and ABA concentrations by combined gas chromatography-mass spectrometry. Lutein and zeaxanthin were the terminal carotenoids in wild-type tissue. The carotenoid profiles ofvp-1 andvp-8 tissue were similar to that of the wild type; invp-2, vp-5, vp-7 andvp-9 carotenogenesis was blocked at early stages so that xanthophylls were absent. Except forvp-1, where the ABA content was similar to the wild type, the ABA content ofvp embryos was substantially reduced, to 6–16% of the corresponding wild type. Thus, the absence of xanthophylls was associated with reduced ABA content, which was in turn correlated with vivipary. Kernels ofvp-8 had a reduced ABA content although xanthophylls were present. Seedlings of carotenoid-deficient mutants rescued from viviparous kernels contained less ABA than did wild-type seedlings grown in the same way. Furthermore, the ABA concentration of such seedlings did not increase in response to water deficit. Conversely,vp-1 seedlings contained normal levels of carotenoids and ABA. Carotenoid-deficient seedlings did not contain appreciable amounts of chlorophyll so that chloroplast development was not normal. Thus ABA-deficiency could be associated with abnormal plastid development rather than the absence of carotenoids per se.Abbreviations ABA abscisic acid - DAP days after pollination - i.d. internal diameter - FW fresh weight - GC-MS combined gas chromatography-mass spectrometry - HPLC high-performance liquid chromatography - MS mass spectrometry - vp viviparous     

Comparative biochemical studies of carotenoids in sea urchins—III. Relationship between developmental mode and carotenoids in the Australian echinoids Heliocidaris erythrogramma and H. tuberculata and a comparison with Japanese species

The sea urchin Heliocidaris tuberculata is typical of most echinoids in having a small egg and a feeding larva, while H. erythrogramma has a large egg and modified development through a non-feeding larvae. The carotenoids in the gonads of these two species were investigated from the comparative biochemical points of view. The carotenoid content of the buoyant eggs of H. erythrogramma was approximately 60 times that of the negatively-buoyant eggs of H. tuberculata. With respect to cytoplasmic volume, however, the carotenoid concentration in the eggs of H. tuberculata was approximately twice that in the eggs of H. erythrogramma. In both species β-echinenone was the principal carotenoid found and their carotenoid patterns were similar. It is very interesting from a functional point of view that carotenoid levels per cytoplasmic volume are conserved across most of the species we have examined irrespective of phylogeny and egg size. In light of this result we suggest that carotenoids may play an important role in developing stage in all echinoids including indirect and direct developers.     

Carotenoids in the eyespot apparatus of the flagellate green alga Spermatozopsis similis: Adaptation to the retinal-based photoreceptor

Isolated intact eyespot apparatuses, the photoreceptive organelles involved in blue-light-mediated photoresponses of flagellate green algae, were analyzed regarding their carotenoid composition. Carotenoids from the eyespot apparatuses of Spermatozopsis similis were identified by high-performance liquid chromatography, visible-light absorption spectra, mass spectroscopy and by ¹H-nuclear magnetic resonance spectroscopy (carotenes), and compared with those of whole-cell extracts. Both extracts contained ,-carotene, ,-carotene (formerly -carotene), lycopene, lutein, zeaxanthin, violaxanthin and all-E-and 9-Z-neoxanthin. The relative carotenoid compositions, however, differed significantly. A twofold relative increase in the total carotene level was evident in the fraction enriched in eyespot apparatuses. This was mainly due to an increase in the monocyclic ,-carotene and the aliphatic lycopene, whereas the relative content of ,-carotene remained unchanged. On the other hand a relative decrease in the total xanthophyll content, especially of lutein and the epoxidic carotenoid neoxanthin, was observed in the eyespot apparatuses compared with the whole-cell extracts. The decrease of the latter resulted almost solely from a reduction of the 9-Z-rather than the all-E-isomer. The bulk of the carotenes is thought to be localized in the highly organized eyespot lipid globules, which act as a combined quarter-wave interference reflector and absorption screen for the photoreceptor in green algae. The enrichment of ,-carotene and lycopene in the eyespot apparatuses, extending the range of visible light absorption to longer wavelengths, represents an adaptation of the screen to the retinal-based photoreceptor of flagellate green algae and is one of the prerequisites for maximal directional sensitivity of the eyespot apparatus.Abbreviations ¹H-NMR nuclear magnetic resonance - IUPAC International Union of Pure and Applied Chemistry - VIS visible absorption spectra This work was supported by the Deutsche Forschungsgemeinschaft (G.K. and M.M.). M.G. was supported by a fellowship from the Norwegian Research Council of Science and Humanities.     

Seasonal changes in photosystem II organisation and pigment composition in Pinus sylvestris

Conifers of the boreal zone encounter considerable combined stress of low temperature and high light during winter, when photosynthetic consumption of excitation energy is blocked. In the evergreen Pinus sylvestris L. these stresses coincided with major seasonal changes in photosystem II (PSII) organisation and pigment composition. The earliest changes occurred in September, before any freezing stress, with initial losses of chlorophyll, the D1-protein of the PSII reaction centre and of PSII light-harvesting-complex (LHC II) proteins. In October there was a transient increase in F₀, resulting from detachment of the light-harvesting antennae as reaction centres lost D1. The D1-protein content eventually decreased to 90%, reaching a minimum by December, but PSII photochemical efficiency [variable fluorescence (F_v)/maximum fluorescence (F_m)] did not reach the winter minimum until mid-February. The carotenoid composition varied seasonally with a twofold increase in lutein and the carotenoids of the xanthophyll cycle during winter, while the epoxidation state of the xanthophylls decreased from 0.9 to 0.1 from October to January. The loss of chlorophyll was complete by October and during winter much of the remaining chlorophyll was reorganised in aggregates of specific polypeptide composition, which apparently efficiently quench excitation energy through non-radiative dissipation. The timing of the autumn and winter changes indicated that xanthophyll de-epoxidation correlates with winter quenching of chlorophyll fluorescence while the drop in photochemical efficiency relates more to loss of D1-protein. In April and May recovery of the photochemistry of PSII, protein synthesis, pigment rearrangements and zeaxanthin epoxidation occurred concomitantly. Indoor recovery of photosynthesis in winter-stressed branches under favourable conditions was completed within 3 d, with rapid increases in F₀, the epoxidation state of the xanthophylls and in light-harvesting polypeptides, followed by recovery of D1-protein content and F_v/F_m, all without net increase in chlorophyll. The fall and winter reorganisation allow Pinus sylvestris to maintain a large stock of chlorophyll in a quenched, photoprotected state, allowing rapid recovery of photosynthesis in spring.Abbreviations Elips early light-induced proteins - EPS epoxidation state - F₀ instantaneous fluorescence - F_m maximum fluorescence - F_v variable fluorescence - LHC II light-harvesting complex of PSII - LiDS lithium dodecyl sulfate This research was supported by the Swedish Natural Science Research Council. We wish to thank Dr. Adrian Clarke¹ (Department of Plant Physiology, University of Umeå, Sweden) for advice on electrophoresis, valuable discussion and providing antibodies. Dr. Stefan Jansson¹ and Dr. Torill Hundal (Department for Biochemistry, University of Stockholm, Sweden) provided antibodies. Jan Karlsson¹ helped with the HPLC, Dr. Marianna Krol gave advice on green gels and Dr. Vaughan Hurry (Cooperative Research Centre for Plant Sciences, Australian National University, Canberra, Australia) provided valuable discussion.     

Studies on well-coupled Photosystem-I-enriched subchloroplast vesicles. Discrimination of flash-induced fast and slow electric potential components

This work aimed at the resolution of the multi-component electric potential changes induced by single-turnover flash illumination of Photosystem-I-enriched subchloroplast vesicles. If supplemented with ferredoxin and under carefully adjusted redox poising, these vesicles show a pronounced slow-rising and -decaying electric potential component, as monitored by endogenous and exogenous field-sensitive probes, carotenoids and oxonol VI, respectively. The fast and slow potential components can be easily discriminated without the need for computer-assisted deconvolution after selective presaturation of the slow component by preillumination or a transmembrane ΔpH, after selective suppression of the slow component by low valinomycin or uncoupler concentrations or in the absence of ferredoxin. The slow electric potential component, as compared to the fast one, is relatively sensitive to low concentrations of ionophores and uncouplers, detergent, ageing and lower temperatures (4–12°C), is associated with electrogenic proton displacements and is interpreted to respond to a field that is more located on the membrane-bulk interface. Temperature effects show transition temperatures around 20°C for both the rise and decay of the slow potential component. The results provide further evidence that the carotenoids and oxonol VI sense the same (slow) electric field, but may be differently located in the thylakoid membrane.