Proton transport by bacteriorhodopsin through an interface film

Summary Interface films of purple membrane and lipid containing spectroscopically intact and oriented bacteriorhodopsin have been used as a model system to study the function of this protein. Small positive charges in surface potential (<1 mV) are detected upon illumination of these films at the air-water interface. These photopotentials, are not affected by overlaying the interface film with a thin layer (0.3 mm) of decane. However, they are dramatically increased when lipid soluble proton carriers FCCP or DNP are added to the decane. The polarity of the photopotential indicates that, in the light, positive charges are transported through the interface from the aqueous to the organic phase. The action spectrum of the photopotential is identical to the absorption spectrum of bacteriorhodopsin. Since bacteriorhodopsin molecules are oriented with their intracellular surface towards the aqueous subphase, the characteristics of the photopotential indicate that in the light bacteriorhodopsin translocates protons from its intracellular to its extracellular surface. The kinetics of the photopotential reveal that the rate and extent of proton transport are proportional both to the fraction of bacteriorhodopsin molecules excited and to the concentration of proton acceptor. The photopotentials result from changes in the ionic distribution across the decane-water interface and can be cancelled by lipid soluble anions.     

Light-induced potential and current across a large bacteriorhodopsin-asolectin planar membrane stabilized on a polyacrylamide gel surface

A phospholipid bilayer membrane was spread from an organic solvent solution between a polyacrylamide gel surface and an aqueous buffer solution. The membrane was quite similar to the conventional black lipid membrane, but was of a large size and was stable since it was supported on the gel surface. Bacteriorhodopsin, impregnated into the membrane, generated membrane potential and current upon illumination. The induced current was large, and this was attributed to the large area of the present membrane. Remarkable responses of the light-induced potential and current were also observed with a thick layer of organic solvent containing phospholipids. The effects of applied membrane potential, carbonylcyanide-m-chlorophenyl hydrazone (CCCP) and gramicidin were examined on these photoresponses. Steady-state current, which is due to protons flowing through the membrane, was enormously enhanced by applying membrane potential opposite to the photopotential or by adding gramicidin to the membrane-forming solution.     

Photovoltaic effect in pigment films in contact with an electrolytes. V. Theory of the Becquerel photovoltaic effect in porous films of phthalocyanine]

In accordance with literature and our experimental data a theoretical analysis of the model system metal-porous film of the pigment-electrolyte was carried out at stationary illumination in the regimes of photocurrent and photopotential. The main properties and positions of the model are supported experimentally. Specific behaviour of the photovoltaic system considered resides in the discovered dependence of the transfer mechanism on the value of catode potential of the electrode.     

Optical and photoelectrochemical properties of Langmuir films of a Zn-complex of ethioporphyrin II

The optical and photoelectrochemical properties of Zn ethioporphyrin II (ZnEP) Langmuir films deposited on an SnO2-optically transparent electrode were investigated. Absorption and fluorescence spectra were utilized to control the film structure. The main three factors, in our opinion, act on the photovoltage, namely, film thickness, ZnEP aggregate formation and the total ZnEP concentration in the monolayer. It was shown that the photopotential decreases as the amount of ZnEP aggregates in the film decreases, and the photopotential increases with surface pressure transferred to the SnO2 layer; the maximum photopotential is registered in films consisting of three and five layers.     

Health benefits of fucoxanthin in the prevention of chronic diseases

Fucoxanthin is a xanthophyll carotenoid abundant in macroalgae, such as brown seaweeds. When fucoxanthin is consumed, it can be esterified or hydrolyzed to fucoxanthinol in the gastrointestinal tract and further converted into amarouciaxanthin A in the liver. It has a unique chemical structure that confers its biological effects. Fucoxanthin has a strong antioxidant capacity by scavenging singlet molecular oxygen and free radicals. Also, it exerts an anti-inflammatory effect. Studies have demonstrated potential health benefits of fucoxanthin for the prevention of chronic diseases, such as cancer, obesity, diabetes mellitus, and liver disease. Animal studies have shown that fucoxanthin supplementation has no adverse effects. However, investigation of the safety of fucoxanthin consumption in humans is lacking. Clinical trials are required to assess the safety of fucoxanthin in conjunction with the study of mechanisms by which fucoxanthin exhibits its health benefits. This review focuses on current knowledge of metabolism and functions of fucoxanthin with its potential health benefits.This article is part of a Special Issue entitled Carotenoids recent advances in cell and molecular biology edited by Johannes von Lintig and Loredana Quadro.     

Photopotential during the photochemical reaction of chlorophyll with duroquinone]

Under the illumination of ethanol solutions of chlorophyll "alpha" and "beta" in the presence of duroquinone positive photopotential (PP) appears, the sign of which is not changed with an increase of pH. However, if duroquinone is used, which had been illuminated with white light in the absence of air, in ethanol, which results in an increase of concentration of its reduced forms, at neutral weakly alkaline pH the PP sign gets negative. This fact supports the assumption that the change of PP sign at the change of pH when parabenzoquinone is used as an oxidizer results from the change of the direction of electron transfer during photoreaction. During photoreaction of pheophytin incapable of photooxidation with duroquinone and disposed to photoreduction, PP at all pH is negative.     

Differential responses in EPA and fucoxanthin production by the marine diatom Stauroneis sp. under varying cultivation conditions

There has been an increasing drive toward better valorising raw biological materials in the context of the sustainability of bio-based industries and the circular economy. As such, microalgae hold the ability to biosynthesise valuable metabolites, which are sought after within the bioenergy, pharmaceuticals, cosmetics or nutrition sectors. Owing to their bioactivities, the xanthophyll pigment fucoxanthin and the omega-3 polyunsaturated fatty acid eicosapentaenoic acid (EPA) have fostered increasing interests in terms of sustainably refining them from natural sources, such as microalgae. Together with the suitability of individual species to industrial cultivation, a key challenge resides in optimizing the yields of these compounds within the microalgal biomass they are retrieved from. The marine diatom Stauroneis sp. LACW24 was batch cultivated into its stationary phase of growth prior to being subjected at high cell density (1 × 10⁶ cells mL⁻¹) to seven different regimes of light exposure in replenished medium and under nutritional limitation (silica and nitrate) for 12 days. The highest EPA proportions and yields were obtained under blue LED in f/2 medium (16.5% and 4.8 mg g⁻¹, respectively), double the values obtained under red LED illumination. The fucoxanthin yield was the highest when cells were subjected to blue LEDs (5.9 mg g⁻¹), a fourfold increase compared to the nitrogen-limited treatment under white LEDs. These results indicate that a two-stage approach to the batch cultivation of this diatom can be used for enhancing the production of the high-value metabolites fucoxanthin and EPA post-stationary phase.     

FUNCTIONAL BINDING OF DIATOM PIGMENTS TO A RHODOPHYTE LIGHT HARVESTING POLYPEPTIDE

A close relationship of light harvesting polypeptides (LHC) of rhodophytes, chromophytes and chlorophytes is inferred from the amino acid sequence similarity in three transmembrane helices, and from the conservation of 8 putative chlorophyll (Chl)-binding sites (Durnford et al. 1999, J. Mol. Evol. 48:59). Differences in Chl and carotenoid pigments have been a major classification feature. Thus, it was of interest to ascertain whether pigments from a diatom ( Thallasiosira fluviatilis ) could be functionally inserted into a red algal ( Porphyridium cruentum ) polypeptide. A recombinant polypeptide, LHCaR1, was reconstituted with pigment extracts from the diatom (Chls a and c , fucoxanthin, diadinoxanthin and β-carotene). The pigments were found attached to protein upon separation on sucrose gradients, and on non-denaturing gels. Absorption and fluorescence excitation spectra revealed individual peaks corresponding to the absorption maxima of Chl a at 438/672 nm; Chl c at 463/638 nm; and fucoxanthin at 493/540 nm. Fluorescence emission and CD spectra showed functional binding and suitable orientation for energy transfer from Chl c and carotenoids to Chl a. The LHCaR1 successfully folded in the presence of the heterologous pigments and bound 7 Chl a , 1 Chl c , 8 fucoxanthin, and 1.9 diadinoxanthin per polypeptide. By comparison, this polypeptide with P. cruentum pigments binds 8 Chl a , and 4 zeaxanthins, thus revealing its capability of functionally binding 8 Chls with variations in carotenoid numbers. Such a trait may have favored the diversification of a large family of LHCs and the successful radiation of photosynthetic eukaryotes into different light environments.     

高产岩藻黄素的海洋硅藻筛选及光照条件优化

为提高岩藻黄素(Fucoxanthin)提取效率, 对16株海洋硅藻岩藻黄素的检测含量和实际含量进行了分析, 并以一株牟氏角刺藻(Chaetoceros muelleri)为对象, 研究了岩藻黄素与另外两种关键光合色素(即叶绿素a和β-胡萝卜素)的含量及其消长受光强和光质影响的特征。研究结果显示, 不同种或同种不同株的海洋硅藻, 在相同的培养条件、收获时期、提取方法下, 其提取率差异大, 可从不足1%到89.78%; 岩藻黄素的检出含量各异, 从0.03到5.02 mg/g。受不同光照强度[低: 50 μmol/(m2·s); 中: 100 μmol/(m2·s); 高: 200 μmol/(m2·s)和光质(红光、蓝光)]的影响, 岩藻黄素产量呈现不同特征。在低光照强度下, 单位细胞岩藻黄素的含量相对较高。在红光条件下, 岩藻黄素的产量高于蓝光。在相同光强条件下, 岩藻黄素含量随增殖周期变化; 在单色光质条件下, 几种光合色素均在生长平台期后期含量增加。岩藻黄素的含量变化与叶绿素a和β-胡萝卜素的含量及其消长关系密切。研究为筛选藻株、优化硅藻培养条件以累积岩藻黄素提供了参考。     

Enrichment of the light-harvesting complex in diadinoxanthin and implications for the nonphotochemical fluorescence quenching in diatoms

The pigment composition of diatoms differs from that of green algae and plants. Diatoms contain chlorophyll (Chl(1)) c, fucoxanthin, and diadinoxanthin (DD). An intermittent light regime during growth induced a large increase in the DD content in the marine planktonic diatom Phaeodactylum tricornutum. Light-harvesting complex containing fucoxanthin (LHCF) subunits were purified on a sucrose gradient after treatment of thylakoid membranes with a mild detergent. DD was found in all the LHCF fractions: a "major" composite LHCF fraction and the two fractions where some LHCF was associated with photosystem centers. For cells enriched in DD, most of the additional DD molecules were bound to the major LHCF fraction. The DD enrichment of the major LHCF fraction was accompanied by a decrease in the fucoxanthin to Chl a ratio. Either some fucoxanthin molecules were replaced by DD or there could be a relative enrichment of subunits rich in DD at the expense of fucoxanthin/Chl c rich subunits. Under high light illumination, a higher degree of de-epoxidation of DD into DT was observed for the major LHCF of cells enriched in DD. This fraction has the higher DD content and the higher degree of de-epoxidation. These results show that the distal antennae, probably mostly isolated as the major LHCF fraction, play a crucial role in the formation of NPQ, its amplitude depending on the amount of DD bound and on the degree of de-epoxidation (Lavaud et al. (2002) Plant Physiol. 129, 1398-1406).     

不同干燥方式、存储温度对羊栖菜中岩藻黄素稳定性的影响

为探究不同干燥方式、不同存储温度和存储时间对羊栖菜中岩藻黄素稳定性的影响,本研究考察了三种不同干燥方式,包括自然阴干、低温烘干和冷冻干燥,对羊栖菜中岩藻黄素稳定性的影响。结果表明采用冷冻干燥所得羊栖菜,含水量低,岩藻黄素含量高,达到699.2μg/g,岩藻黄素保留率为94.7%,远高于自然阴干及低温烘干。因此,冷冻干燥是最佳干燥方法。考察了四个不同温度(-20、4、25、30℃)存储对冻干羊栖菜中岩藻黄素稳定性的影响,结果表明随着存储温度的升高,岩藻黄素的降解率明显增高,因此,-20℃为最佳储存温度。     

Phylogeny of nuclear-encoded plastid-targeted GAPDH gene supports separate origins for the peridinin- and the fucoxanthin derivative-containing plastids of dinoflagellates

Although most photosynthetic dinoflagellates have plastids with peridinin, the three dinoflagellate genera Karenia, Karlodinium, and Takayama possess anomalously pigmented plastids that contain fucoxanthin and its derivatives (19′-hexanoyloxy-fucoxanthin and 19′-butanoyloxy-fucoxanthin) instead of the peridinin. This pigment composition is similar to that of haptophytes. All peridinin-containing dinoflagellates investigated so far have at least two types of glyceraldehyde-3-phosphate dehydrogenase (GAPDH): cytosolic and plastid-targeted forms. In the present study, we cloned and sequenced genes encoding cytosolic and plastid-targeted GAPDH proteins from three species of the fucoxanthin derivative-containing dinoflagellates. Based on the molecular phylogeny, the plastid-targeted GAPDH genes of the fucoxanthin derivative-containing dinoflagellates were closely related to those of haptophyte algae rather than to the peridinin-containing dinoflagellates, while one of several cytosolic versions from the peridinin- and the fucoxanthin derivative-containing dinoflagellates are closely related to each other. Considering a previously reported theory that the plastid-targeted GAPDH from the peridinin-containing dinoflagellates originated by a gene duplication of the cytosolic form before the splitting of the dinoflagellate lineage, it is highly likely that the plastid-targeted GAPDH gene of the peridinin-containing dinoflagellates is original in this algal group and that in the fucoxanthin-containing dinoflagellates, the original plastid-targeted GAPDH was replaced by that of a haptophyte endosymbiont during a tertiary endosymbiosis. The present results strongly support the hypothesis that the plastids of the peridinin- and the fucoxanthin derivative-containing dinoflagellates are of separate origin.     

Protective effect of Fucoxanthin against UVB-induced skin photoaging in hairless mice

Fucoxanthin, a major carotenoid in brown algae, has various beneficial effects. In this study, we evaluated the effect of topical fucoxanthin on UVB-induced skin photoaging in hairless mice. The dorsal skins were treated topically with a 0.001% fucoxanthin solution 2 h each time before UVB irradiation (5 times a week) for 10 weeks. The formation of wrinkles in UVB-irradiated skin treated with vehicle alone significantly increased, as compared with the non-irradiated control. Treatment with fucoxanthin tended to suppress UVB-induced wrinkle formation, but there was no significant difference between wrinkle formation in the control group and the fucoxanthin treatment group. However, topical treatment with fucoxanthin significantly lessened UVB-induced epidermal hypertrophy, VEGF, and MMP-13 expression in the epidermis and thiobarbituric acid reactive substances (TBARS) in the skin. These results indicate that topical treatment with fucoxanthin prevents skin photoaging in UVB-irradiated hairless mice, possibly via antioxidant and antiangiogenic effects.     

Capacity of fucoxanthin for scavenging peroxyl radicals and inhibition of lipid peroxidation in model systems

Abstract

Carotenoids act as physiological antioxidant by scavenging reactive-free radicals as well as quenching singlet oxygen. Fucoxanthin is one of the abundant carotenoids found in edible brown seaweeds. The assessment of radical scavenging capacity of carotenoids has been the subject of extensive studies, which, however, gave inconsistent results. In the present study, the capacity of fucoxanthin for scavenging peroxyl radicals, chain carrying species of lipid peroxidation, was assessed quantitatively by measuring the effect of α-tocopherol on the decay of fucoxanthin induced by peroxyl radicals. It was found that α-tocopherol was 7.1 times more reactive than fucoxanthin in heptane solution, but interestingly fucoxanthin exerted 1.6 times higher reactivity than α-tocopherol in methanol solution. In SDS micelles, the relative reactivity of fucoxanthin and α-tocopherol depended on the site of peroxyl radical formation. The efficacy of lipid peroxidation inhibition by fucoxanthin was much less than that of α-tocopherol.     

The harmful alga Aureococcus anophagefferens utilizes 19'-butanoyloxyfucoxanthin as well as xanthophyll cycle carotenoids in acclimating to higher light intensities

Aureococcus anophagefferens is a picoplanktonic microalga that is very well adapted to growth at low nutrient and low light levels, causing devastating blooms ("brown tides") in estuarine waters. To study the factors involved in long-term acclimation to different light intensities, cells were acclimated for a number of generations to growth under low light (20μmolphotonsm(-2)s(-1)), medium light (60 or 90μmolphotonsm(-2)s(-1)) and high light (200μmolphotonsm(-2)s(-1)), and were analyzed for their contents of xanthophyll cycle carotenoids (the D pool), fucoxanthin and its derivatives (the F pool), Chls c(2) and c(3), and fucoxanthin Chl a/c polypeptides (FCPs). Higher growth light intensities resulted in increased steady state levels of both diadinoxanthin and diatoxanthin. However, it also resulted in the conversion of a significant fraction of fucoxanthin to 19'-butanoyloxyfucoxanthin without a change in the total F pool. The increase in 19'-butanoyloxyfucoxanthin was paralleled by a decrease in the effective antenna size, determined from the slope of the change in F(0) as a function of increasing light intensity. Transfer of acclimated cultures to a higher light intensity showed that the conversion of fucoxanthin to its derivative was a relatively slow process (time-frame of hours). We suggest the replacement of fucoxanthin with the bulkier 19'-butanoyloxyfucoxanthin results in a decrease in the light-harvesting efficiency of the FCP antenna and is part of the long-term acclimative response to growth at higher light intensities.     

铜藻岩藻黄素提取及纯化工艺研究

为了对岩藻黄素的提取、纯化进行系统研究,进而为高纯度岩藻黄素的工业化生产提供研究基础,筛选了适用于提取铜藻（Sargassum horneri）鲜藻中岩藻黄素的有机溶剂,并通过单因素实验和正交实验确定了最佳的提取溶剂浓度、提取温度、提取时间、料液比等工艺参数。随后采用硅胶柱层析法进行纯化,并通过单因素实验确定了最佳的硅胶柱床高度、上样量和洗脱流速。最后采用制备液相法对经层析纯化的岩藻黄素进一步纯化。结果表明,有机溶剂萃取的最佳工艺条件为：甲醇浓度90%,提取温度50 ℃,提取时间1 h,料液比1∶10,此条件下岩藻黄素提取率达到(0.258 9±0.003 6) mg·g-1鲜重（FW）［(1.078 8±0.015 0) mg·g-1干重（DW）］。硅胶柱层析的最佳工艺条件为：硅胶柱床高度10 cm,上样量6 g,洗脱流速10 mL·min-1,此条件下岩藻黄素得率为0.176 5 mg·g-1FW（0.735 3 mg·g-1 DW）,纯度为87.01%±0.88%。经制备液相进一步纯化后,岩藻黄素得率为0.127 1 mg·g-1 FW（0.529 4 mg·g-1 DW）,纯度为99.27%±0.22%。研究所用工艺简单,岩藻黄素得率高,为高纯度岩藻黄素的制备提供了实验基础。     

Photoelectric properties of chlorophyll and carotene solutions in nematic liquid crystal located between semiconducting electrodes

The photopotential and photocurrent generation for chlorophyll a, beta-carotene and a mixture of these pigments dissolved in nematic liquid crystal and located between transparent semiconducting electrodes were measured. Both pigments exhibit photopotential and photocurrent generation. From the photocurrent amplitudes it follows that the efficiency of electron transfer to a semiconducting electrode from beta-carotene is higher than from chlorophyll alpha. The photocurrent amplitude of the pigment mixture is slightly lower than that calculated as a sum of amplitudes of pigments located in separated cells. This difference can be explained by secondary effects, such as competition between carotene and chlorophyll molecules in a process of adsorption on a semiconducting electrode. Therefore it seems that no charge transfer complexes of chlorophyll and carotene are formed in the investigated model system.     

A marine carotenoid, fucoxanthin, induces regulatory T cells and inhibits Th17 cell differentiation in vitro

Fucoxanthin is a non-provitamin A carotenoid contained in brown seaweeds. We found that it suppressed interleukin-17 secretion from CD4(+) T cells under IL-17-producing T (Th17) cell development conditions. By evaluating T cell differentiation in vitro, fucoxanthin and its metabolite fucoxanthinol inhibited T cell differentiation into Th17 cells. This suggests that fucoxanthin can improve inflammatory diseases due to Th17 cells.     

Marine algal fucoxanthin inhibits the metastatic potential of cancer cells

Metastasis is major cause of malignant cancer-associated mortality. Fucoxanthin has effect on various pharmacological activities including anti-cancer activity. However, the inhibitory effect of fucoxanthin on cancer metastasis remains unclear. Here, we show that fucoxanthin isolated from brown alga Saccharina japonica has anti-metastatic activity. To check anti-metastatic properties of fucoxanthin, in vitro models including assays for invasion, migration, actin fiber organization and cancer cell–endothelial cell interaction were used. Fucoxanthin inhibited the expression and secretion of MMP-9 which plays a critical role in tumor invasion and migration, and also suppressed invasion of highly metastatic B16-F10 melanoma cells as evidenced by transwell invasion assay. In addition, fucoxanthin diminished the expressions of the cell surface glycoprotein CD44 and CXC chemokine receptor-4 (CXCR4) which play roles in migration, invasion and cancer–endothelial cell adhesion. Fucoxanthin markedly suppressed cell migration in wound healing assay and inhibited actin fiber formation. The adhesion of B16-F10 melanoma cells to the endothelial cells was significantly inhibited by fucoxanthin. Moreover, in experimental lung metastasis in vivo assay, fucoxanthin resulted in significant reduction of tumor nodules. Taken together, we demonstrate, for the first time, that fucoxanthin suppresses metastasis of highly metastatic B16-F10 melanoma cells in vitro and in vivo.