Carotenoid Formation Research Articles

The underlying molecular mechanisms of hormonal regulation of fruit color in fruit-bearing plants.

Fruit color is a key feature of fruit quality, primarily influenced by anthocyanin or carotenoid accumulation or chlorophyll degradation. Adapting the pigment content is crucial to improve the fruit's nutritional and commercial value. Genetic factors along with other environmental components (i.e., light, temperature, nutrition, etc.) regulate fruit coloration. The fruit coloration process is influenced by plant hormones, which alsoplay a vital role in various physiological and biochemical metabolic processes. Additionally, phytohormones play a role in the regulation of a highly conserved transcription factor complex, called MBW (MYB-bHLH-WD40). The MBW complex, which consists of myeloblastosis (MYB), basic helix-loop-helix (bHLH), and WD40 repeat (WDR) proteins, coordinates the expression of downstream structural genes associated with anthocyanin formation. In fruit production, the application of plant hormones may be important for promoting coloration. However, concerns such as improper concentration or application time must be addressed. This article explores the molecular processes underlying pigment formation and how they are influenced by various plant hormones. The ABA, jasmonate, and brassinosteroid increase anthocyanin and carotenoid formation, but ethylene, auxin, cytokinin, and gibberellin have positive as well as negative effects on anthocyanin formation. This article establishes the necessary groundwork for future studies into the molecular mechanisms of plant hormones regulating fruit color, ultimately aiding in their effective and scientific application towards fruit coloration.

The Application of Natural Carotenoids in Multiple Fields and Their Encapsulation Technology: A Review.

Carotenoids, which are inherent pigments occurring in plants and microorganisms, manifest a diverse array of vivid hues. Owing to their multifarious health advantages, carotenoids have engendered substantial interest among scholars and consumers alike. Presently, carotenoids are extensively employed in the realms of food, nutrition and health commodities, pharmaceuticals, and cosmetics, rendering them an indispensable constituent of our quotidian existence. Therefore, the objective of this review is to present a succinct and methodical examination of the sources, constituents, and factors influencing formation of carotenoids. Particular attention will be given to encapsulation strategies that maintain intrinsic characteristics, as the growing desire for carotenoids is propelled by individuals' escalating standards of living. Moreover, the applications of natural carotenoids in multiple fields, including pharmaceutical, food and feed, as well as cosmetics, are discussed in detail. Finally, this article explores the main challenges hindering the future advancement of carotenoids, aiming at facilitating their effective integration into the circular economy.

Carotenoid Production of Rhodosporodimum Toruloides under Light Irradiation

Carotenoids are widely used as an antioxidant, a precursor to vitamin A, and food colorants in the medical, cosmetic, chemical, food, and feed industries. They were distributed in diverse microorganisms including bacteria, algae, and fungi. Among them, the yeast Rhodosporidium toruloides is convenient for large-scale fermentation due to its unicellular nature and high growth rate. However, many factors influence the biotechnological synthesis of carotenoids. Light is a crucial consideration while making microbial carotenoids. Carotenogenic is a photo-protective technique used by microorganisms to defend themselves from the light that causes oxidative damage. R. toruloides raised their carotenoid productivity under light conditions in this study. Various lights were used to compare the effect of color light in carotenoid productivity. Results showed that R. toruloides produced more carotenoid content when cultured in blue or white light as compared to a red light or in the dark. Besides, results from microarray showed that this light influence was in the transcription gene level, the light irradiation encouraged the formation of antioxidants such carotenoids, causes a protective mechanism against DNA damage and oxidative stress.

Fruits from tomato carotenoid mutants have altered susceptibility to grey mold

The necrotrophic fungus Botritys cinerea takes advantage of the oxidative burst to facilitate tissue infection, leading to substantial losses during tomato postharvest. Tomato fruit is a source of carotenoids, pigments with a wide variety of isomeric configurations that determine their antioxidant capacity. Here, fruit susceptibility to B. cinerea was assessed in Micro-Tom Near Isogenic lines harboring mutations that alter the profile of carotenoids. Wound-inoculated fruit of the mutants Delta carotene (Del) and tangerine (t), which show large variety of carotenoids rather than the major accumulation of trans-lycopene, were less susceptible to the pathogen. Differences in susceptibility between the mutants were only observed in ripe fruit, after the formation of carotenoids, and they were associated with attenuation of damage caused by reactive oxygen species. The greater variety of carotenoid isomers, which in turn contributed to the greater lipophilic antioxidant capacity of fruit, was associated with the less susceptible mutants, Del and t. Together, our data reveals a potential activity of carotenoids in fruit defense, in addition to the well-known and widespread ecological role as attractors of seed dispersers.

High temperature elevates carotenoid accumulation of banana fruit via upregulation of MaEIL9 module

Banana is a good source of carotenoids, which are bioactive metabolites with health beneficial properties for human. However, the molecular mechanism of carotenoid accumulation in banana fruit is largely unclear. In this study, we found that high temperature elevated carotenoid production in banana pulp, which is presumably due to upregulation of a subset of carotenogenic genes as well as a carotenoid biosynthesis regulator MaSPL16. Moreover, an ethylene signaling component MaEIL9 was identified, whose transcript and protein contents were also induced by high temperature. In addition, MaEIL9 positively regulates transcription of MaDXR1, MaPDS1, MaZDS1 and MaSPL16 through directly targeting their promoters. Overexpression of MaEIL9 in tomato fruit substantially increased the expression of carotenoid formation genes and elevated carotenoid content. Importantly, transiently silencing MaEIL9 in banana fruit weakened carotenoid production caused by high temperature. Taken together, these results indicate that high temperature induces carotenoid production in banana fruit, at least in part, through MaEIL9-mediated activation of MaDXR1, MaPDS1, MaZDS1 and MaSPL16 expression.

Medium development and production of carotenoids and exopolysaccharides by the extremophile Rhodothermus marinus DSM16675 in glucose-based defined media

BackgroundThe marine thermophilic bacterium Rhodothermus marinus can degrade many polysaccharides which makes it interesting as a future cell factory. Progress using this bacterium has, however, been hampered by limited knowledge on media and conditions for biomass production, often resulting in low cell yields and low productivity, highlighting the need to develop conditions that allow studies of the microbe on molecular level. This study presents development of defined conditions that support growth, combined with evaluation of production of carotenoids and exopolysaccharides (EPSs) by R. marinus strain DSM 16675.ResultsTwo defined media were initially prepared: one including a low addition of yeast extract (modified Wolfe’s medium) and one based on specific components (defined medium base, DMB) to which two amino acids (N and Q), were added. Cultivation trials of R. marinus DSM 16675 in shake flasks, resulted in maximum cell densities (OD620 nm) of 2.36 ± 0.057, cell dry weight (CDW) 1.2 ± 0.14 mg/L, total carotenoids 0.59 × 10–3 mg/L, and EPSs 1.72 ± 0.03 mg/L using 2 g/L glucose in DMB. In Wolfe’s medium (supplemented by 0.05 g/L yeast extract and 2.5 g/L glucose), maximum OD620 nm was 2.07 ± 0.05, CDW 1.05 ± 0.07 mg/L, total carotenoids 0.39 × 10–3 mg/L, and EPSs 1.74 ± 0.2 mg/L. Growth trials at 5 g/L glucose in these media either failed or resulted in incomplete substrate utilization. To improve reproducibility and increase substrate utilization, a screening of macroelements (e.g. phosphate) in DMB, was combined with use of trace elements and vitamins of the modified Wolfe’s medium. The resulting defined minimal R. marinus medium, (DRM), allowed reproducible cultivations to a final OD620nm of 6.6 ± 0.05, CDW 2.85 ± 0.07 mg/L, a maximum specific growth rate (µmax) of 0.26 h−1, total carotenoids 0.77 × 10–3 mg/L and EPSs 3.4 ± 0.17 mg/L in cultivations supplemented with up to 5 g/L glucose.ConclusionA minimal defined medium (DRM) was designed that resulted in reproducible growth and an almost doubled formation of both total carotenoids and EPSs. Such defined conditions, are necessary for systematic studies of metabolic pathways, to determine the specific requirements for growth and fully characterize metabolite production.

Formation of Lutein, β-Carotene and Astaxanthin in a Coelastrella sp. Isolate.

In this study, the effect of media composition, N/P ratio and cultivation strategy on the formation of carotenoids in a Coelastrella sp. isolate was investigated. A two-stage process utilizing different media in the vegetative stage, with subsequent re-suspension in medium without nitrate, was employed to enhance the formation of carotenoids. The optimal growth and carotenoid content (β-carotene and lutein) in the vegetative phase were obtained by cultivation in M-8 and BG11 media. Use of a N/P ratio of 37.5 and low light intensity of 40 μmol m−2 s−1 (control conditions) led to optimal biomass production of up to 1.31 g L−1. Low concentrations of astaxanthin (maximum of 0.31 wt. %) were accumulated under stress conditions (nitrogen-deficient medium containing 1.5 % of NaCl and light intensity of 500 μmol m−2 s−1), while β-carotene and lutein (combined maximum of 2.12 wt. %) were produced under non-stress conditions. Lipid analysis revealed that palmitic (C16:0) and oleic (C18:1) constituted the main algal fatty acid chains (50.2 ± 2.1% of the total fatty acids), while esterifiable lipids constituted 17.2 ± 0.5% of the biomass by weight. These results suggest that Coelastrella sp. could also be a promising feedstock for biodiesel production.

Banana MaERF124 negatively modulates carotenoid accumulation during fruit ripening through repression of carotenogenesis genes

Banana is an important source of carotenoids, and the fruit accumulate carotenoids during the ripening process. However, the regulatory mechanisms underlying carotenoid production in banana fruit have not been well studied. Here, we report that MaERF124 is a negative transcriptional regulator of carotenogenesis genes in banana. The reduced accumulation of MaERF124 was accompanied with the increased contents of carotenoids during the ripening of the four banana varieties, suggesting a negative role of MaERF124 in carotenoid production. Further assays showed that MaERF124 acts as a transcriptional repressor and directly targets the promoters of carotenoid biosynthetic genes including MaDXR, MaGGPS, MaCRTISO and MaLCYB1.2. Moreover, ectopic expression of MaERF124 in tomato fruit repressed the transcript levels of carotenoid formation genes to inhibit carotenoid production. Taken together, these findings reveal that MaERF124 negatively modulates carotenoid formation via transcriptional repression of key genes involved in carotenoid synthesis, which provides a regulatory machinery for improving the contents of carotenoids in banana fruit.

Changes in carbon allocation and subplastidal amyloplast structures of specialised Ipomoea batatas (sweet potato) storage root phenotypes

Vitamin A deficiency (VAD) in Low and Medium Income countries remains a major health concern. Ipomoea batatas, orange sweet potato (OSP), is one of the biofortification solutions being implemented by the World Health Organisation (WHO) to combat VAD. However, high provitamin A (β-carotene) content has been associated with a reduction in dry matter, reducing calorific value and having adverse effects on consumer traits. Both starch and carotenoid formation are located in amyloplasts and could potentially compete for the same precursors. Hence, five different sweet potato storage root phenotypes were characterized through spatial metabolomics and proteomics at the sub-plastidal level. The metabolite data suggested an indirect correlation of starch and carotenoids through the TCA cycle and pentose phosphate pathway. Furthermore, a change in lipid composition was observed to accommodate the storage of carotenoids in the hydrophilic environment of the amyloplast. The data suggests an alteration of cellular ultra-structures and perturbation of metabolism in high β-carotene producing sweet potato roots. This corroborates with previous gene expression analysis through biochemical analysis of sweet potato root tissue.

Ascorbic acid prevents yellowing of fresh-cut yam by regulating pigment biosynthesis and energy metabolism

The fresh-cut yam would turn yellow under 25 ℃ for 36 h, which could reduce consumer’s acceptance. This study aimed to investigate the mechanism by which 2% ascorbic acid inhibits yellowing of fresh-cut yam by detecting enzyme activities, gene expressions and metabolites. Ascorbic acid treatment was found to decrease ATPase activities, ATP content, and energy charge. The transcriptional expression levels of citrate synthase, acetyl-CoA carboxylase, and acetyl-CoA synthetase, which are involved in the tricarboxylic acid cycle, were also decreased by ascorbic acid treatment, thus blocking the supply of precursors and energy for pigment biosynthesis. In addition, ascorbic acid effectively inhibited the formation of carotenoids, flavonoids, and bisdemethoxycurcumin, as indicated by lower metabolic levels, decreased enzyme activities, and downregulated transcriptional expressions. Thus, ascorbic acid prevents yellowing in fresh-cut yam by reducing the energy metabolism level as well as inhibiting pigment (carotenoids, flavonoids, bisdemethoxycurcumin) biosynthesis pathways. Accordingly, ascorbic acid treatment is a safe, effective, and cheap method for inhibiting fresh-cut yam yellowing.

Transcriptomics Integrated with Metabolomics Unveil Carotenoids Accumulation and Correlated Gene Regulation in White and Yellow-Fleshed Turnip (Brassica rapa ssp. rapa).

Turnip (Brassica rapa ssp. rapa) is considered to be a highly nutritious and health-promoting vegetable crop, whose flesh color can be divided into yellow and white. It is widely accepted that yellow-fleshed turnips have higher nutritional value. However, reports about flesh color formation is lacking. Here, the white-fleshed inbred line, W21, and yellow-fleshed inbred line, W25, were profiled from the swollen root of the turnip at three developmental periods to elucidate the yellow color formation. Transcriptomics integrated with metabolomics analysis showed that the PSY gene was the key gene affecting the carotenoids formation in W25. The coding sequence of BrrPSY-W25 was 1278 bp and that of BrrPSY-W21 was 1275 bp, and BrrPSY was more highly expressed in swollen roots in W25 than in W21. Transient transgenic tobacco leaf over-expressing BrrPSY-W and BrrPSY-Y showed higher transcript levels and carotenoids contents. Results revealed that yellow turnip formation is due to high expression of the PSY gene rather than mutations in the PSY gene, indicating that a post-transcriptional regulatory mechanism may affect carotenoids formation. Results obtained in this study will be helpful for explaining the carotenoids accumulation of turnips.

A DUF4281 domain-containing protein (homologue of ABA4) of Phaeodactylum tricornutum regulates the biosynthesis of fucoxanthin

Although fucoxanthin is one of the most valuable carotenoids, its biosynthesis pathway is still poorly understood. Neoxanthin has been regarded as the intermediate of fucoxanthin in recent years, however, multiple details still need to be explored. Previous study performed in Arabidopsis thaliana demonstrated a DUF4281-domain containing protein (ABA4) was necessary for the biosynthesis of neoxanthin, and the mutation of ABA4 led to a huge alteration of pigment composition. In Phaeodactylum tricornutum, a function unknown protein (designated as PtABA4 in this study) displayed high homology to ABA4, and also contained DUF4281 domain. To investigate its function on neoxanthin synthesis and fucoxanthin accumulation, we conducted endogenous overexpression and knockdown (RNA interference) of PtABA4 individually, as well as in vitro activity assay using recombinant protein. The overexpression and knockdown of PtABA4 lead to sharp increases and decrease of fucoxanthin, respectively, demonstrating a key role of PtABA4 on fucoxanthin accumulation. Unexpectedly, the knockdown of PtABA4 results in the decreases of upstream carotenoids of neoxanthin, rather than compensatory increases. In terms of in vitro activity assay, neither the recombinant protein produced by Escherichia coli nor the overexpressed transformant displays neoxanthin synthase activity. Taking all results into consideration, we speculate that the PtABA4 may constitute one multienzyme complexes with other membrane associated enzymes and participate in the formation of several carotenoids, rather than neoxanthin only. The ubiquity of DUF4281 domain across all taxa and the absence of xanthophylls in bacteria provide indirect evidence for our assumption. Above all, this study here provides a target gene for enhancing the fucoxanthin content by genetic engineering methods, and deeps the understanding about the function of DUF4281 domain.

Promiscuous activity of β-carotene hydroxylase CrtZ on epoxycarotenoids leads to the formation of rare carotenoids with 6-hydroxy-3-keto-ε-ends.

Carotenoids with rare 6-hydroxy-3-keto-ε-end groups, such as piprixanthin, vitixanthin, or cochloxanthin, found in manakin birds or plants, are rare carotenoids with high antioxidant activity. The same chemical structure is found in abscisic acid or blumenol, apocarotenoids found in plants or fungi. In this study, we serendipitously discovered that the promiscuous activity of the β-carotene hydroxylase CrtZ, a diiron-containing membrane protein, can catalyze the formation of 6-hydroxy-3-keto-ε-end by using epoxycarotenoids antheraxanthin or violaxanthin as substrate. We suggest that the reaction mechanism is similar to that of a rhodoxanthin biosynthetic enzyme. Our results provide a further understanding of the reaction mechanism of diiron-containing β-carotene hydroxylases, as well as insight into the biosynthesis of natural compounds with 6-hydroxy-3-keto-ε-end carotenoid derivatives.

Metabolomic approaches for the characterization of carotenoid metabolic engineering in planta.

Carotenoid biosynthesis has now been subjected to metabolic engineering for over two decades. The outputs clearly show that carotenoid formation is an integral component of metabolism. Perturbations can affect intermediary metabolism and other isoprenoids. The advances in omic technologies have enabled the quantitative assessment of changes in the transcriptome, proteome and metabolome in response to altered carotenoid biosynthesis. In the present article, the approaches and procedures relating to the capture of the metabolome in response to modulation of the carotenoid biosynthetic pathway are described. These data will contribute to the fundamental understanding of metabolic biology, underpinning future rationale design of New Plant Breeding Techniques (NPBTs) and associated regulatory affairs.

Live diatoms as potential biocatalyst in a microbial fuel cell for harvesting continuous diafuel, carotenoids and bioelectricity

Microbial fuel cell (MFC) with live diatoms (Nitzschia palea) displacing bacteria in the anodic chamber generated electrical potential. Unlike other microalgae, diatoms fix 25% of atmospheric CO2, thus releasing O2. They perform photolysis of water by photosynthesis in the plastid during light photoperiod and cellular respiration in the mitochondria during dark, producing electrons and protons, respectively. The electrogenic property of diatom was explored and evaluated by comparing the potential changes with reference fuel cell without diatoms and that operated with diatoms in the anodic chamber. Such photosynthetic diatom microbial fuel cell (PDMFC) employed f/2 media rich in nitrates, phosphates, metasilicates, trace metals and vitamins as the anolyte and potassium permanganate as catholyte enhanced the output voltage by 3rd day. The maximum power density for PDMFC was 12.62 mWm−2 and coulombic efficiency of 22.95%. Besides this, the fixed diatom cells at anode showed about 64.28% increase in lipid production on 15th day compared to that on 1st day along with the increment in formation of complex fatty acid methyl esters and carotenoids during its operation. Hence, diatoms can be envisaged to substitute bacteria in the anodic chamber of MFC to simultaneously produce bioelectricity and other valuable compounds. Further their silica nanoporous architecture serve as good absorbents for heavy metal removal found in many wastewaters.

Metabolic engineering of Synechocystis sp. PCC 6803 for the photoproduction of the sesquiterpene valencene

Cyanobacteria are extremely adaptable, fast-growing, solar-powered cell factories that, like plants, are able to convert carbon dioxide into sugar and oxygen and thereby produce a large number of important compounds. Due to their unique phototrophy-associated physiological properties, i.e. naturally occurring isoprenoid metabolic pathway, they represent a highly promising platform for terpenoid biosynthesis. Here, we implemented a carefully devised engineering strategy to boost the biosynthesis of commercially attractive plant sequiterpenes, in particular valencene. Sesquiterpenes are a diverse group of bioactive metabolites, mainly produced in higher plants, but with often low concentrations and expensive downstream extraction. In this work we successfully demonstrate a multi-component engineering approach towards the photosynthetic production of valencene in the cyanobacterium Synechocystis sp. PCC 6803. First, we improved the flux towards valencene by markerless genomic deletions of shc and sqs. Secondly, we downregulated the formation of carotenoids, which are essential for viability of the cell, using CRISPRi on crtE. Finally, we intended to increase the spatial proximity of the two enzymes, ispA and CnVS, involved in valencene formation by creating an operon construct, as well as a fusion protein. Combining the most successful strategies resulted in a valencene production of 19 mg/g DCW in Synechocystis. In this work, we have devised a useful platform for future engineering steps.

A light‐mediated study of carotenoids in carrots (Daucus carota) using resonance Raman spectroscopy

AbstractIn photosynthetic systems, carotenoids play significant roles, and the prime role is light harvesting, then transferred to the photosynthetic apparatus for photosynthesis. Understanding carotenoids is a significant concern as it has a vital role in photosynthesis, and it also acts as a protective pigment in the biosystem. Carotenoids are widely distributed in almost all photosynthetic biosystems but highly accumulated in carrots, especially in (Daucus carota). Therefore, carrots can be a simpler model to understand the formation of carotenoids and their regulation with the help of exposure to sunlight. Raman spectroscopy is a noninvasive method to study molecular signature; it can be an ideal tool to understand carotenoids pigments in the growing stages of carrots. In this work, using resonance Raman spectroscopy with 532‐nm wavelength laser excitation, we were able to detect predominant carotenoid bands such as β‐carotene, α‐carotene, and lutein, at 1517–1519 cm−1 and 1521–1524 cm−1 with high peak intensities. This work gives a clear description and evidence of how light plays a profound effect on the activation and regulation of carotenoids. We observed β‐carotene in the nonphotosynthetic region of the plant, as a photoprotection of developing life is one of the significant roles of carotenoids, and Raman bands of carotenoids can be considered as the biomarker of photoprotection of developing life.

Components interactions and changes at molecular level in maize flour-based blends as affected by the extrusion process. A multi-analytical approach

The aim of this work was to evaluate the influence of partial replacement of maize flour by sub-valuated whole grain flours on the interactions and spatial distribution of the main components of extruded snacks. Non-destructive, fast spectroscopic and microscopic complementary tools were employed. All blends (composed by maize + 25% of pearl millet, red sorghum, quinoa or hairless canary seed) presented lower carbohydrate and lipid contents and higher protein, ash and dietary fiber levels than the control maize extrudates. These compositional changes were reflected in FT-MIR and FT-Raman spectra which also allowed detecting modifications at molecular level, such as protein agglomeration, amylose-lipids complexes formation and surface exposure of carotenoids. Tridimensional distribution and spatial arrangements of the main components in the extruded samples were studied through CLSM and XPS. These properties are related to quality aspects such as lipid oxidation sensitivity and textural properties. The combined FT-MIR, FT-Raman, CLSM and XPS were helpful tools to understand thermo-mechanical modifications of starch, proteins and lipids as a consequence of the extrusion process.

A Four-Level Maturity Index for Hot Peppers (Capsicum annum) Using Non-Invasive Automated Mobile Raman Spectroscopy for On-Site Testing

A handheld Raman spectrometer was used to determine the ripeness of peppers. Raman spectra were recorded non-invasively on the fruit surface. The spectroscopic data were transformed into a classification scheme referred to as the maturity index which allowed for attribution of the fruit stadium to four levels from immature to fully mature. Hot pepper and tomato ripening includes pectic polysaccharide depolymerization, chlorophyll degradation and carotenoid formation, among others. The latter were followed non-invasively by Raman spectroscopy. Two portable systems and one benchtop system were compared for their applicability and robustness to establish a suitable maturity index. Spectral acquisition, data treatment and multivariate data analysis were automated using a Matlab script on a laptop computer. The automated workflow provided a graphic visualization of the relevant parameters and results on-site in real time. In terms of reliability and applicability, the chemometric model to determine the maturity of fruits was compared to a univariate procedure based on the average intensity and ratio of three characteristic signals. Portable Raman spectrometers in combination with the maturity index or a chemometric model should be suitable to assess the stage of maturing for carotenoid-containing fruits and thus to determine ripeness on-site or during a sorting process in an automated manner.

Tetraedron minimum, First Reported Member of Hydrodictyaceae to Accumulate Secondary Carotenoids.

We isolated a novel strain of the microalga Tetraedron minimum in Iceland from a terrestrial habitat. During long-term cultivation, a dish culture turned orange, indicating the presence of secondary pigments. Thus, we characterized T. minimum for growth and possible carotenoid production in different inorganic media. In a lab-scale photobioreactor, we confirmed that nitrogen starvation in combination with salt stress triggered a secondary carotenoid accumulation. The development of the pigment composition and the antioxidant capacity of the extracts was analyzed throughout the cultivations. The final secondary carotenoid composition was, on average, 61.1% astaxanthin and 38.9% adonixanthin. Moreover, the cells accumulated approx. 83.1% unsaturated fatty acids. This work presents the first report of the formation of secondary carotenoids within the family Hydrodictyaceae (Sphaeropleales, Chlorophyta).