Botryococcus as an alternative source of carotenoids and its possible applications – an overview

The enhanced interest in carotenoid research arises partly because of their application in the food and health industries and partly because of the necessity to find a commercially viable natural source for their mass production. The bottlenecks in finding a natural source of carotenoids which can compete with the synthetic products is the mass production of the organism that produces carotenoids, cell harvesting and extraction methods of carotenoids. The microalga Botryococcus braunii is an interesting organism for its commercial value as a rich source of carotenoids. It contains lutein as major carotenoid which is considered to be one of the beneficial carotenoids in human health applications. The current paper reviews the status of B. braunii as an alternative source of carotenoid production on the commercial scale addressing aspects like cultures of algae, factors that enhance the production and accumulation of carotenoids, cell harvesting methods, and carotenoid extraction. The paper also presents an overview of identification, characterization and structural elucidation of carotenoids from B. braunii and their bioactivity.     

Enhancing the carotenoid content of <Emphasis Type="Italic">Brassica napus</Emphasis> seeds by downregulating lycopene epsilon cyclase

The accumulation of carotenoids in higher plants is regulated by the environment, tissue type and developmental stage. In Brassica napus leaves, beta-carotene and lutein were the main carotenoids present while petals primarily accumulated lutein and violaxanthin. Carotenoid accumulation in seeds was developmentally regulated with the highest levels detected at 35-40 days post anthesis. The carotenoid biosynthesis pathway branches after the formation of lycopene. One branch forms carotenoids with two beta rings such as beta-carotene, zeaxanthin and violaxanthin, while the other introduces both beta- and epsilon-rings in lycopene to form alpha-carotene and lutein. By reducing the expression of lycopene epsilon-cyclase (epsilon-CYC) using RNAi, we investigated altering carotenoid accumulation in seeds of B. napus. Transgenic seeds expressing this construct had increased levels of beta-carotene, zeaxanthin, violaxanthin and, unexpectedly, lutein. The higher total carotenoid content resulting from reduction of epsilon-CYC expression in seeds suggests that this gene is a rate-limiting step in the carotenoid biosynthesis pathway. epsilon-CYC activity and carotenoid production may also be related to fatty acid biosynthesis in seeds as transgenic seeds showed an overall decrease in total fatty acid content and minor changes in the proportions of various fatty acids.     

Phytoene desaturase is localized exclusively in the chloroplast and up-regulated at the mRNA level during accumulation of secondary carotenoids in Haematococcus pluvialis (Volvocales, chlorophyceae)

The unicellular green alga Haematococcus pluvialis Flotow is known for its massive accumulation of ketocarotenoids under various stress conditions. Therefore, this microalga is one of the favored organisms for biotechnological production of these antioxidative compounds. Astaxanthin makes up the main part of the secondary carotenoids and is accumulated mostly in an esterified form in extraplastidic lipid vesicles. We have studied phytoene desaturase, an early enzyme of the carotenoid biosynthetic pathway. The increase in the phytoene desaturase protein levels that occurs following induction is accompanied by a corresponding increase of its mRNA during the accumulation period, indicating that phytoene desaturase is regulated at the mRNA level. We also investigated the localization of the enzyme by western-blot analysis of cell fractions and by immunogold labeling of ultrathin sections for electron microscopy. In spite of the fact that secondary carotenoids accumulate outside the chloroplast, no extra pathway specific for secondary carotenoid biosynthesis in H. pluvialis was found, at least at this early stage in the biosynthesis. A transport process of carotenoids from the site of biosynthesis (chloroplast) to the site of accumulation (cytoplasmatic located lipid vesicles) is implicated.     

Carotenoid biotechnology in plants for nutritionally improved foods

Carotenoids participate in light harvesting and are essential for photoprotection in photosynthetic plant tissues. They also furnish non-photosynthetic flowers and fruits with yellow to red colors to attract animals for pollination and dispersal of seeds. Although animals can not synthesize carotenoids de novo , carotenoid-derived products such as retinoids (including vitamin A) are required as visual pigments and signaling molecules. Dietary carotenoids also provide health benefits based on their antioxidant properties. The main pathway for carotenoid biosynthesis in plants and microorganisms has been virtually elucidated in recent years, and some of the identified biosynthetic genes have been successfully used in metabolic engineering approaches to overproduce carotenoids of interest in plants. Alternative approaches that enhance the metabolic flux to carotenoids by upregulating the production of their isoprenoid precursors or interfere with light-mediated regulation of carotenogenesis have been recently shown to result in increased carotenoid levels. Despite spectacular achievements in the metabolic engineering of plant carotenogenesis, much work is still ahead to better understand the regulation of carotenoid biosynthesis and accumulation in plant cells. New genetic and genomic approaches are now in progress to identify regulatory factors that might significantly contribute to improve the nutritional value of plant-derived foods by increasing their carotenoid levels.     

Carotenoid content in growing cells of Haematococcus pluvialis during a sunlight cycle

Under certain culture conditions, cells of the chlorophyte Haematococcus pluvialis accumulate significant amounts of astaxanthin. This study describes biomass and carotenoid production during a sunlight cycle in a continuous culture of growing cells of H. pluvialis and shows that these two parameters are under the control of irradiance. The hourly carotenoid production increases with light intensity and, in our culture conditions, carotenoid accumulation occurs in a few hours and without any morphological change in the algae. These carotenoids seem to be efficient in protecting algal cells against photoinhibition damage if their content is greater than 1% dry biomass. Below this concentration, that is to say in the early hours of high light intensity, dry biomass decreases due to cell lysis. The results demonstrate that secondary carotenoid accumulation in H. pluvialis may occur in the active growth phase and is stimulated from the first hours of sunlight illumination.     

Metabolic engineering of carotenoid accumulation by creating a metabolic sink

Carotenoids are highly beneficial for human nutrition and health because they provide essential nutrients and important antioxidants in our diets. However, many food crops, especially the major staple crops contain only trace to low amounts of carotenoids. Although significant progress has been made in developing food crops rich in carotenoids by altering the expression of carotenoid biosynthetic genes, in many cases it has proved to be difficult to reach the desired levels of carotenoid enrichment. The recent identification and characterization of a novel gene mutation in cauliflower reveals that creating a metabolic sink to sequester carotenoids is an important mechanism to control carotenoid accumulation in plants. The successful demonstration of increased carotenoid accumulation in association with the formation of sink structures in transgenic crops offers a new and alternative approach to increase carotenoid content. Manipulation of the formation of metabolic sink along with the catalytic activity of the pathway may represent a promising strategy for maximally improving the nutritional quality of food crops.     

枸杞子中类胡萝卜素的组成及含量

枸杞子(ＦｒｕｃｔｕｓＬｙｃｉｉ)为宁夏枸杞(ＬｙｃｉｕｍｂａｒｂａｒｕｍＬ.)的成熟果实,有补肾养肝、润肺明目的功效。枸杞子中的类胡萝卜素已有一些研究[1,2],但只对其中个别类胡萝卜素提取液皂化后进行了鉴定,不能真正代表其组成。本文采用ＨＰＬＣ法对6种不同产地...     

Sequestration of host plant carotenoids in the larval tissues of Helicoverpa zea

To determine the cause of the unique yellow coloration in mandibular glands of soybean-fed Helicoverpa zea larvae, the accumulation of carotenoids in various tissues of last instar larvae fed soybean, cotton and tomato foliage was quantified. Five carotenoids were detected in the foliage of all host plants but at significantly different concentrations. Xanthophylls rather than carotenes were most likely to accumulate in larval tissues. Carotenoids accumulated at different rates and some were significantly affected by larval diet. Highest levels of carotenoid accumulation, notably lutein, were detected in the testes, followed by midgut epithelium, fat body and integument. The midgut epithelium contained the greatest and the testes the least diversity of carotenoid types. Low levels of lutein were detected in both labial and mandibular glands. Tomato foliage had the highest carotenoid content and caterpillar tissues fed these leaves often had the highest amounts of carotenoid. However, the accumulation of carotenoids did not protect larvae from antibiotic effects of tomato foliage because these caterpillars had the highest mortality and slowest growth rates of all the three host plants. Transport and absorption of lipid and oxidative stress may be some reasons for differential carotenoid accumulation.     

Carotenoids of Marine Sponges

The 42 identified carotenoids isolated from 36 different marine spontes may, from structural considerations, be divided into four groups; (a) native phytoplankton-type carotenoids; (b) intact carotenoids of possible zooplankton origin, 9c) intact carotenoids of probable bacterial of fungal origin; and (d) sponge metabolized carotenoids. Groups (a) and (d) are the major categories, group (d) comprises several mono- and diaryl carotenoids and some oxygenated carotenoids so far peculiar to the Demospongiae. Chemosystematic considerations suggest that highest capacity for carotenoid accumulation and transformation is to be found within the orders Poecilosclerida and Axinellida, which exhibit similar carotenoid patterns. The screening of carotenoids in 34 coloured species from the Australian RRIMP collection showed a total carotenoid content of 0.1–90 × 10^?3% of the ry wt. individual carotenoids were characterized for 16 species including 11 previously known carotenoids, two new partly characterized methoxylated carotenoids and some phenolic carotenoids.     

Isolation of a β-Carotene Over-Producing Soil Bacterium, Sphingomonas sp.

A carotenoid-accumulating bacterium isolated from soil, identified as a Sphingomonas sp., grew at 0.18 h(-1) and produced 1.7 mg carotenoids g(-1) dry cell, among which beta-carotene (29% of total carotenoids) and nostoxanthin (36%). A mutant strain, obtained by treatment with ethyl methanesulfonate, accumulated up to 3.5 mg carotenoids g(-1) dry cell. Accumulation of beta-carotene by this strain depended on the oxygenation of the growth medium, with maximal accumulation (89%) occurring under limiting conditions. Beta-carotene accumulation could be further enhanced by incubating the cells in the presence of glycerol (either not or only slowly assimilated) and yeast extract resulting in an accumulation of 5.7 mg beta-carotene g(-1) dry cell wt. The strain used lactose as carbon source with similar biomass and carotenoid production, providing a viable alternative use for cheese whey ultra-filtrate.     

苹果类胡萝卜素研究进展

类胡萝卜素是苹果果实色泽形成的一个重要影响因子,其种类和含量决定果实是否具有良好的外观和丰富的营养。本文综述了近年来有关苹果果实类胡萝卜素方面的研究进展,并对苹果类胡萝卜素的种类和含量,苹果发育和贮藏过程中类胡萝卜素含量的变化规律,生物合成途径中相关基因的表达,以及环境因子对类胡萝卜素积累的影响等方面进行了阐述。     

Optimization of carotenoid production from hyper-producing Rhodotorula glutinis mutant 32 by a factorial approach

AIMS: Optimization of carotenoid production by a mutant of Rhodotorula glutinis. METHODS: The growth and carotenoid production was optimized in shake flasks using a two-level, three-variable factorial design. RESULTS: The volumetric carotenoid production could be increased to 129 +/- 2 mg x 1(-1) in a medium containing (g x l(-1)) yeast extract 11.74, glucose 46 and threonine 18 along with other micronutrients, wherein, beta-carotene yield was 102 +/- 2 mg x l(-1), accounting for 80% of the total carotenoids. SIGNIFICANCE AND IMPACT OF THE STUDY: The medium optimization resulted in a fourfold increase in volumetric production and a twofold increase in the cellular accumulation of carotenoids. In view of such high yields, the mutant of Rhodotorula glutinis can be a potential source of beta-carotene.     

The effects of dietary carotenoid intake on carotenoid accumulation in the retina of a wild bird, the house finch (Carpodacus mexicanus)

Carotenoid pigments accumulate in the retinas of many animals, including humans, where they play an important role in visual health and performance. Recently, birds have emerged as a model system for studying the mechanisms and functions of carotenoid accumulation in the retina. However, these studies have been limited to a small number of domesticated species, and the effects of dietary carotenoid access on retinal carotenoid accumulation have not been investigated in any wild animal species. The purpose of our studies was to examine how variation in dietary carotenoid types and levels affect retinal accumulation in house finches (Carpodacus mexicanus), a common and colorful North American songbird. We carried out three 8-week studies with wild-caught captive birds: (1) we tracked the rate of retinal carotenoid depletion, compared to other body tissues, on a very low-carotenoid diet, (2) we supplemented birds with two common dietary carotenoids (lutein + zeaxanthin) and measured the effect on retinal accumulation, and (3) we separately supplemented birds with high levels of zeaxanthin - an important dietary precursor for retinal carotenoids - or astaxanthin - a dominant retinal carotenoid not commonly found in the diet (i.e. a metabolic derivative). We found that carotenoids depleted slowly from the retina compared to other tissues, with a significant (∼50%) decline observed only after 8 weeks on a very low-carotenoid diet. Supplementation with lutein + zeaxanthin or zeaxanthin alone significantly increased only retinal galloxanthin and ε-carotene levels, while other carotenoid types in the retina remained unaffected. Concentrations of retinal astaxanthin were unaffected by direct dietary supplementation with astaxanthin. These results suggest highly specific mechanisms of retinal carotenoid metabolism and accumulation, as well as differential rates of turnover among retinal carotenoid types, all of which have important implications for visual health maintenance and interventions.     

Interspecific variation in dietary carotenoid assimilation in birds: links to phylogeny and color ornamentation

Many birds use carotenoid pigments to acquire rich red, orange, and yellow coloration in feathers and bare parts that is used as a signal of mate quality. Because carotenoids are derived from foods, much attention has been paid to the role of diet in generating color variation both within and among avian species. Less consideration has been given to physiological underpinnings of color variability, especially among species. Here, I surveyed published literature (e.g. captive feeding studies) on carotenoid assimilation in six bird species and completed additional controlled carotenoid-supplementation experiments in two others to consider the ability of different taxa to extract carotenoids from the diet in relation to phylogeny and coloration. I found that, for a given level of carotenoids in the diet, passerine birds (zebra finch, Taeniopygia guttata; house finch, Carpodacus mexicanus; American goldfinch, Carduelis tristis; society finch, Lonchura domestica) exhibit higher levels of carotenoids in circulation than non-passerines like gamebirds (domestic chicken, Gallus domesticus; red junglefowl, Gallus gallus; Japanese quail, Coturnix coturnix; red-legged partridge, Alectoris rufa). This difference in carotenoid accumulation is likely due to interspecific variation in micelle, chylomicron, or lipoprotein concentrations or affinities for xanthophyll carotenoids. Passerine birds more commonly develop carotenoid-based colors than do birds from ancient avian lineages such as Galliformes, and the physiological differences I uncover may explain why songbirds especially capitalize on carotenoid pigments for color production. Ultimately, because we can deconstruct color traits into component biochemical, physical, and physiological parts, avian color signals may serve as a valuable model for illuminating the proximate mechanisms behind interspecific variation in signal use in animals.     

Carotenoid production and change of photosynthetic functions in Scenedesmus sp. exposed to nitrogen limitation and acetate treatment

Under stress conditions, some microalgae up-regulate certain biosynthetic pathways, leading to the accumulation of specific compounds. For example, changing nutrient composition can induce stress in algae’s physiological activities, which may trigger an intense increase in carotenoid production. In this study, the change of photosynthetic functions and carotenoid production in the green microalga Scenedesmus sp. was investigated when algal cultures were exposed to conditions including limited nitrogen content with the addition of sodium acetate. Microalgal cultures were treated for 18 days under higher irradiance conditions. We observed a decrease of chlorophyll content induced concomitantly with a decline of photosystem II and I photochemistry. At the same time, an important increase in carotenoid content was detected. By using high-performance liquid chromatographic analysis, we found that the secondary carotenoids astaxanthin and canthaxanthin were accumulated compared to controls. During the process of carotenoid accumulation, chlorophyll degradation was found in addition to a strong decrease in photosynthetic electron transport. Such changes may be associated with the structural reorganization of the photosynthetic apparatus and can be a useful indicator of secondary carotenoid accumulation in algal cultures.     

Chromoplast biogenesis and carotenoid accumulation

Chromoplasts are special organelles that possess superior ability to synthesize and store massive amounts of carotenoids. They are responsible for the distinctive colors found in fruits, flowers, and roots. Chromoplasts exhibit various morphologies and are derived from either pre-existing chloroplasts or other non-photosynthetic plastids such as proplastids, leucoplasts or amyloplasts. While little is known about the molecular mechanisms underlying chromoplast biogenesis, research progress along with proteomics study of chromoplast proteomes signifies various processes and factors important for chromoplast differentiation and development. Chromoplasts act as a metabolic sink that enables great biosynthesis and high storage capacity of carotenoids. The formation of chromoplasts enhances carotenoid metabolic sink strength and controls carotenoid accumulation in plants. The objective of this review is to provide an integrated view on our understanding of chromoplast biogenesis and carotenoid accumulation in plants.     

A novel gene mutation that confers abnormal patterns of beta-carotene accumulation in cauliflower (Brassica oleracea var. botrytis)

The Or gene of cauliflower (Brassica oleracea var. botrytis) causes many tissues of the plant to accumulate carotenoids and turn orange, which is suggestive of a perturbation of the normal regulation of carotenogenesis. A series of experiments to explore the cellular basis of the carotenoid accumulation induced by the Or gene was completed. The Or gene causes obvious carotenoid accumulation in weakly or unpigmented tissues such as the curd, pith, leaf bases and shoot meristems, and cryptically in some cells of other organs, including the roots and developing fruits. The dominant carotenoid accumulated is beta-carotene, which can reach levels that are several hundred-fold higher than those in comparable wild-type tissues. The beta-carotene accumulates in plastids mainly as a component of massive, highly ordered sheets. The Or gene does not affect carotenoid composition of leaves, nor does it alter color and chromoplast appearance in flower petals. Interestingly, mRNA from carotenogenic and other isoprenoid biosynthetic genes upstream of the carotenoid pathway was detected both in orange tissues of the mutant, and in comparable unpigmented wild-type tissues. Thus the unpigmented wild-type tissues are likely to be competent to synthesize carotenoids, but this process is suppressed by an unidentified mechanism. Our results suggest that the Or gene may induce carotenoid accumulation by initiating the synthesis of a carotenoid deposition sink in the form of the large carotenoid-sequestering sheets.