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Nannochloropsis oceanica is an industrially relevant marine microalga rich in eicosapentaenoic acid (EPA, a valuable ω-3 polyunsaturated fatty acid), yet the algal production potential remains to be unlocked. Here we engineered N. oceanica to synthesize the high-value carotenoid astaxanthin independent of high-light (HL) induction for achieving multifaceted benefits. By screening β-carotenoid ketolases and hydroxylases of various origins, and strategically manipulating compartmentalization, fusion patterns, and linkers of the enzyme pair, a remarkable 133-fold increase in astaxanthin content was achieved in N. oceanica. Iterative metabolic engineering efforts led to further increases in astaxanthin synthesis up to 7.3 mg g -1 , the highest reported for microalgae under nonstress conditions. Astaxanthin was found in the photosystem components and allowed the alga HL resistance and augmented EPA production. Besides, we achieved co-production of astaxanthin and EPA by the engineered alga through a fed-batch cultivation approach. Our findings unveil the untapped potential of N. oceanica as a robust, light-driven chassis for constitutive astaxanthin synthesis and provide feasible strategies for the concurrent production of multiple high-value biochemicals from CO 2 , thereby paving the way for sustainable biotechnological applications of this alga., (© 2024 The Author(s). New Phytologist © 2024 New Phytologist Foundation.)

This study investigated the technical feasibility of using electrogermination to activate dormant cysts as an inoculum for subsequent 14-d photosynthetic astaxanthin production in Haematococcus lacustris. Electrotreatment affected the cell viability, surface charge, and morphology of H. lacustris cysts. At an optimal voltage of 2 V for 60 min, the cyst germination rate peaked at 44.6 % after 1 d, representing a 2.2-fold increase compared with that of the untreated control. Notably, electrogermination significantly enhanced both the astaxanthin content (44.9 mg/g cell) and productivity (13.2 mg/L/d) after 14 d of photobioreactor cultivation, corresponding to 1.7- and 1.5-fold increases compared with those in control, respectively. However, excessive electrotreatment, particularly at voltages exceeding 2 V or for durations beyond 60 min, did not enhance the astaxanthin production capability of H. lacustris. Proper optimization of renewable electrogermination can enable sustainable algal biorefinery to produce multiple bioactive products without compromising cell viability and astaxanthin productivity., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The acclimation of the green algae Chlamydomoas reinhardtii to high light (HL) has been studied predominantly under continuous illumination of the cells. Here, we investigated the impact of fluctuating HL in alternation with either low light (LL) or darkness on photosynthetic performance and on photoprotective responses. Compared to intervening LL phases, dark phases led to (1) more pronounced reduction of the photosystem II quantum efficiency, (2) reduced degradation of the PsbS protein, (3) lower energy dissipation capacity and (4) an increased pool size of the xanthophyll cycle pigments. These characteristics indicate increased photo-oxidative stress when HL periods are interrupted by dark phases instead of LL phases. This overall trend was similar when comparing long (8 h) and short (30 min) HL phases being interrupted by long (16 h) and short (60 min) phases of dark or low light, respectively. Only the degradation of PsbS was clearly more efficient during long (16 h) LL phases when compared to short (60 min) LL phases., (© 2024 The Author(s). Plant, Cell & Environment published by John Wiley & Sons Ltd.)

In high light, the antenna system in oxygenic photosynthetic organisms switches to a photoprotective mode, dissipating excess energy in a process called non-photochemical quenching (NPQ). Diatoms exhibit very efficient NPQ, accompanied by a xanthophyll cycle in which diadinoxanthin is de-epoxidized into diatoxanthin. Diatoms accumulate pigments from this cycle in high light, and exhibit faster and more pronounced NPQ. The mechanisms underlying NPQ in diatoms remain unclear, but it can be mimicked by aggregation of their isolated light-harvesting complexes, FCP (fucoxanthin chlorophyll-a/c protein). We assess this model system by resonance Raman measurements of two peripheral FCPs, trimeric FCPa and nonameric FCPb, isolated from high- and low-light-adapted cells (LL,HL). Quenching is associated with a reorganisation of these proteins, affecting the conformation of their bound carotenoids, and in a manner which is highly dependent on the protein considered. FCPa from LL diatoms exhibits significant changes in diadinoxanthin structure, together with a smaller conformational change of at least one fucoxanthin. For these LL-FCPa, quenching is associated with consecutive events, displaying distinct spectral signatures, and its amplitude correlates with the planarity of the diadinoxanthin structure. HL-FCPa aggregation is associated with a change in planarity of a 515-nm-absorbing fucoxanthin, and, to a lesser extent, of diadinoxanthin. Finally, in FCPb, a blue-absorbing fucoxanthin is primarily affected. FCPs thus possess a plastic structure, undergoing several conformational changes upon aggregation, dependent upon their precise composition and structure. NPQ in diatoms may therefore arise from a combination of structural changes, dependent on the environment the cells are adapted to., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

This study investigated the effects of oil phases on the encapsulation rate, storage stability, and bioavailability of astaxanthin (ASTA) in Pickering emulsions (PEs). Results showed PEs of mixed oils (olive oil/edible tea oil) had excellent encapsulation efficiency (about 96.0%) and storage stability of ASTA. In vitro simulated gastrointestinal digestion results showed the mixed oil PE with a smaller interfacial area and higher monounsaturated fatty acid content may play a better role in improving ASTA retention and bioaccessibility. In vivo absorption results confirmed the mixed oil PE with an olive oil/edible tea oil of 7:3 was more favorable for ASTA absorption. Molecular dynamics simulation showed ASTA bound more strongly and stably to fatty acid molecules in the system of olive oil/edible tea oil of 7:3; and van der Waals force was the main binding force. NMR further proved there really were interactions between ASTA and four main fatty acids., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Zeaxanthin (Zea) is a key component in the energy-dependent, rapidly reversible, nonphotochemical quenching process (qE) that regulates photosynthetic light harvesting. Previous transient absorption (TA) studies suggested that Zea can participate in direct quenching via chlorophyll (Chl) to Zea energy transfer. However, the contamination of intrinsic exciton-exciton annihilation (EEA) makes the assignment of TA signal ambiguous. In this study, we present EEA-free TA data using Nicotiana benthamiana thylakoid membranes, including the wild type and three NPQ mutants ( npq1 , npq4 , and lut2 ) generated by CRISPR/Cas9 mutagenesis. The results show a strong correlation between excitation energy transfer from excited Chl Q y to Zea S 1 and the xanthophyll cycle during qE activation. Notably, a Lut S 1 signal is absent in the npq1 thylakoids which lack zeaxanthin. Additionally, the fifth-order response analysis shows a reduction in the exciton diffusion length (L D ) from 62 ± 6 nm to 43 ± 3 nm under high light illumination, consistent with the reduced range of exciton motion being a key aspect of plants' response to excess light., Competing Interests: Competing interests statement:The authors declare no competing interest.

Microalgae play an important role in aquatic ecosystems, but the widespread presence of micro- and nano-plastics (MNPs) poses significant threats to them. Haematococcus pluvialis is well-known for its ability to produce the antioxidant astaxanthin when it experiences stress from environmental conditions. Here we examined the effects of polystyrene nanoplastics (PS-NPs) at concentrations of 0.1, 1, and 10 mg/L on H. pluvialis over an 18-day period. Our results show that PS-NPs caused a significant, dose-dependent inhibition of H. pluvialis growth and a reduction in photosynthesis. Furthermore, PS-NPs severely damaged the morphology of H. pluvialis, leading to cell shrinkage, collapse, content release, and aggregation. Additionally, PS-NPs induced a dose-dependent increase in soluble protein content and a decrease in the production of extracellular polymeric substances. These findings indicate that PS-NPs has the potential to adversely affect both the physiology and morphology of H. pluvialis. An increase in reactive oxygen species and antioxidant enzyme activities was also observed, suggesting an oxidative stress response to PS-NPs exposure. Notably, the synthesis of astaxanthin, which is crucial for H. pluvialis's survival under stress, was significantly inhibited in a dose-dependent manner under strong light conditions, along with the down-regulation of genes involved in the astaxanthin biosynthesis pathway. This suggests that PS-NPs exposure reduces H. pluvialis's ability to survive under adverse conditions. This study enhances our understanding of the toxic effects of PS-NPs on microalgae and underscores the urgent need for measures to mitigate MNP pollution to protect aquatic ecosystems., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Carotenoids belonging to the class of tetraterpenoids have extensive applications in medicine, food, nutrition, cosmetics, and feed. Among them, lutein and zeaxanthin can prevent macular degeneration in the elderly, which is very important for protecting vision. Here, we introduce the first metabolomic analysis of Sphingopyxis sp. USTB-05, aiming to shed light on the biosynthesis of carotenoids. Sphingopyxis sp. USTB-05 has the complete methylerythritol 4-phosphate (MEP) pathway and carotenoid biosynthesis pathway, especially involved in the bioconversion of zeaxanthin, violaxanthin, and astaxanthin. Metabolomic profiling identified seven carotenes and six xanthophylls synthesized by Sphingopyxis sp. USTB-05. Zeaxanthin, in particular, was found to be the most abundant, with a content of 37.1 µg/g dry cells. Collectively, the results presented herein greatly enhance our understanding of Sphingopyxis sp. USTB-05 in carotenoids biosynthesis, and thus further accelerate its fundamental molecular investigations and biotechnological applications.

Astaxanthin is a naturally occurring xanthophyll with powerful: antioxidant, antitumor, and antibacterial properties that are widely employed in food, feed, medicinal and nutraceutical industries. Currently, chemical synthesis dominates the world's astaxanthin market, but the increasing demand for natural products is shifting the market for natural astaxanthin. Haematococcus pluvialis (H. pluvialis) is the factory source of natural astaxanthin when grown in optimal conditions. Currently, various strategies for the production of astaxanthin have been proposed or are being developed in order to meet its market demand. This up-to-date review scrutinized the current approaches or strategies that aim to increase astaxanthin yield from H. pluvialis . We have emphasized the genetic and environmental parameters that increase astaxanthin yield. We also looked at the transcriptomic dynamics caused by environmental factors (phytohormones induction, light, salt, temperature, and nutrient starvation) on astaxanthin synthesizing genes and other metabolic changes. Genetic engineering and culture optimization (environmental factors) are effective approaches to producing more astaxanthin for commercial purposes. Genetic engineering, in particular, is accurate, specific, potent, and safer than conventional random mutagenesis approaches. New technologies, such as CRISPR-Cas9 coupled with omics and emerging computational tools, may be the principal strategies in the future to attain strains that can produce more astaxanthin. This review provides accessible data on the strategies to increase astaxanthin accumulation natively. Also, this review can be a starting point for new scholars interested in H. pluvialis research.

Natural astaxanthin is synthesized by diverse organisms including: bacteria, fungi, microalgae, and plants involving complex cellular processes, which depend on numerous interrelated parameters. Nonetheless, existing knowledge regarding astaxanthin biosynthesis and the conditions influencing astaxanthin accumulation is fairly limited. Thus, manipulation of the growth conditions to achieve desired biomass and astaxanthin yields can be a complicated process requiring cost-intensive and time-consuming experiment-based research. As a potential solution, modeling and simulation of biological systems have recently emerged, allowing researchers to predict/estimate astaxanthin production dynamics in selected organisms. Moreover, mathematical modeling techniques would enable further optimization of astaxanthin synthesis in a shorter period of time, ultimately contributing to a notable reduction in production costs. Thus, the present review comprehensively discusses existing mathematical modeling techniques which simulate the bioaccumulation of astaxanthin in diverse organisms. Associated challenges, solutions, and future perspectives are critically analyzed and presented.

Haematococcus lacustris-derived natural astaxanthin has significant commercial value, but stressful conditions alone impair cell growth and reduce the total productivity of astaxanthin in industrial settings. This study used gamma-aminobutyric acid (GABA) to increase biomass, astaxanthin productivity, and tolerance to salinity. GABA under NaCl stress enhanced the biomass to 1.76 g/L, astaxanthin content to 30.37 mg g -1 , and productivity to 4.10 mg/L d -1 , outperforming the control. Further analysis showed GABA enhanced nitrogen assimilation, Ca 2+ level, and cellular GABA content, boosting substrate synthesis, energy metabolism, osmoregulation, autophagy, and antioxidant defenses. GABA also activated signaling pathways involving phytohormones, cAMP, cGMP, and MAPK, aiding astaxanthin synthesis. The application of biomarkers (ethylene, salicylic acid, trans-zeatin) and an autophagy inhibitor cooperated with GABA to further enhance the total astaxanthin productivity under NaCl stress. Combining GABA with 25 μM salicylic acid maximized astaxanthin yield at 4.79 mg/L d -1 , offering new strategies for industrial astaxanthin production., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Vegetable waste, rich in bioactive compounds, offers a promising resource for producing value-added products. This study explored the use of tomato waste, containing glucose (40 mg/g), lycopene (95.12 μg/g), and β-carotene (24.31 μg/g), for cultivating fucoxanthin-rich Isochrysis galbana. Water-soluble lycopene (2.0 μg/mL) and β-carotene (0.4 μg/mL) effectively upregulated key carotenoid synthesis genes and boosted cell growth and fucoxanthin production (3.64 and 3.60 pg/cell, respectively) within 10 days in a mixotrophic culture. Optimized tomato waste hydrolysate achieved a high cell density of 1.21 × 10 7 cells/mL, 2.13 g/L biomass, and 21.02 mg/g fucoxanthin. This study highlights the potential of combining tomato waste with microalgae for a novel and innovative approach towards waste management and resource utilization., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Background: Avocado fruit is rich in xanthophylls, which have been related to positive effects on human health. Xanthophyl acetyltransferases (XATs) are enzymes catalyzing the esterification of carboxylic acids to the hydroxyl group of the xanthophyll molecule. This esterification is thought to increase the lipophilic nature of the xanthophyll and its stability in a lipophilic environment. Studies on XATs in fruits are very scarce, and no studies had been carried out in avocado fruit during postharvest. The objective of this work was to investigate the changes in the expression of genes encoding XAT, during avocado fruit ripening., Results: Avocado fruits were obtained from a local market and stored at 15 °C for 8 days. The fruit respiration rate, ethylene production, and fruit peel's color space parameters (L*, a*, b*) were measured during storage. Fruit mesocarp samples were taken after 1, 3, 5, and 7 days of storage and frozen with liquid nitrogen. Total RNA was extracted from fruit mesocarp, and the quantification of the two genes designated as COGE_ID: 936743791 and COGE_ID: 936800185 encoding XATs was performed with real-time quantitative reverse transcription polymerase chain reaction using actin as a reference gene. The presence of a climacteric peak and large changes in color were recorded during postharvest. The two genes studied showed a large expression after 3 days of fruit storage., Conclusions: We conclude that during the last stages of ripening in avocado fruit there was an active esterification of xanthophylls with carboxylic acids, which suggests the presence of esterified xanthophylls in the fruit mesocarp. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Pigments and other secondary metabolites originating from marine microbes have been a promising natural colorants and drugs for multifaceted applications. However, marine actinobacteria producing such natural molecules are least investigated in terms of their taxonomy, chemical diversity and applications in biomedical, textile, and food industries. In this study, sioxanthin pigment-producing Gram-positive actinobacteria, Micromonospora sp. strain SH-82 was isolated from a marine sponge, Scopalina hapalia, and its whole genome was analyzed. Strain SH-82is a prolific producer of diverse chemical molecules as it produced more compounds on A1 medium with different culture conditions. The genome size of SH-82 is 6.24 Mb (6,246,890 bp) carrying 23 identified biosynthetic gene clusters. A total of 5415 CDS, 60 tRNA, 9 rRNA, and 1 tmRNA are identified from SH-82 genome. The GC content (%) of whole genome was 71.6%. Strain SH-82 harbors genes encoding type I, type II, and type III polyketide synthases. Based on the multi-locus sequence analysis and fatty acid methyl ester (FAME) composition, strain SH-82 is confirmed as a novel species. The genetic information of Micromonospora sp. SH-82 has been deposited to NCBI under the BioProject ID PRJNA1087320, with corresponding identifiers in the Sequence Read Archive (SRA) as SAMN40439676 and the Genome accession as CP148049., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Neoporphyra haitanensis, a red alga harvested for food, thrives in the intertidal zone amid dynamic and harsh environments. High irradiance represents a major stressor in this habitat, posing a threat to the alga's photosynthetic apparatus. Interestingly, N. haitanensis has adapted to excessive light despite the absence of a crucial xanthophyll cycle-dependent photoprotection pathway. Thus, it is valuable to investigate the mechanisms by which N. haitanensis copes with excessive light and to understand the photoprotective roles of carotenoids. Under high light intensities and prolonged irradiation time, N. haitanensis displayed reduction in photosynthetic efficiency and phycobiliproteins levels, as well as different responses in carotenoids. The decreased carotene contents suggested their involvement in the synthesis of xanthophylls, as evidenced by the up-regulation of lycopene-β-cyclase (lcyb) and zeaxanthin epoxidase (zep) genes. Downstream xanthophylls such as lutein, zeaxanthin, and antheraxanthin increased proportionally to light stress, potentially participating in scavenging reactive oxygen species (ROS). When accompanied by the enhanced activity of ascorbate peroxidase (APX), these factors resulted in a reduction in ROS production. The responses of intermediates α-cryptoxanthin and β-cryptoxanthin were felt somewhere between carotenes and zeaxanthin/lutein. Furthermore, these changes were ameliorated when the organism was placed in darkness. In summary, down-regulation of the organism's photosynthetic capacity, coupled with heightened xanthophylls and APX activity, activates photoinhibition quenching (qI) and antioxidant activity, helping N. haitanensis to protect the organism from the damaging effects of excessive light exposure. These findings provide insights into how red algae adapt to intertidal lifestyles., (© 2024 Phycological Society of America.)

Carotenoids perform multifaceted roles in life ranging from coloration over light harvesting to photoprotection. The Orange Carotenoid Protein (OCP), a light-driven photoswitch involved in cyanobacterial photoprotection, accommodates a ketocarotenoid vital for its function. OCP extracts its ketocarotenoid directly from membranes, or accepts it from homologs of its C-terminal domain (CTDH). The CTDH from Anabaena (AnaCTDH) was shown to be important for carotenoid transfer and delivery from/to membranes. The C-terminal tail of AnaCTDH is a critical structural element likely serving as a gatekeeper and facilitator of carotenoid uptake from membranes. We investigated the impact of amino acid substitutions within the AnaCTDH-CTT on echinenone and canthaxanthin uptake from DOPC and DMPG liposomes. The transfer rate was uniformly reduced for substitutions of Arg-137 and Arg-138 to Gln or Ala, and depended on the lipid type, indicating a weaker interaction particularly with the lipid head group. Our results further suggest that Glu-132 has a membrane-anchoring effect on the PC lipids, specifically at the choline motif as inferred from the strongly different effects of the CTT variants on the extraction from the two liposome types. The substitution of Pro-130 by Gly suggests that the CTT is perpendicular to both the membrane and the main AnaCTDH protein during carotenoid extraction. Finally, the simultaneous mutation of Leu-133, Leu-134 and Leu-136 for alanines showed that the hydrophobicity of the CTT is crucial for carotenoid uptake. Since some substitutions accelerated carotenoid transfer into AnaCTDH while others slowed it down, carotenoprotein properties can be engineered toward the requirements of applications., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Astaxanthin is a red xanthophyll with high economic and industrial value in the pharmaceutical, nutraceutical, cosmetic and food industries. In recent years, the biotechnological production of astaxanthin has attracted much attention as a sustainable alternative to the predominating petrochemical-dependent chemical synthesis. In this regard, Xanthophyllomyces dendrorhous is regarded as a promising microorganism for industrial production of astaxanthin. Unfortunately, biotechnological production of the carotenoid is currently expensive. The present study investigated soy molasses (SM) and residual brewers' yeast as cheap fermentation feedstocks for the cultivation of X. dendrorhous and astaxanthin production. Yeast extract was obtained from residual brewers' yeast using various techniques and then combined with SM to formulate a two-component growth medium which was subsequently used to cultivate X. dendrorhous. Generally, the yeast extract produced from residual brewers' yeast supported X. dendrorhous growth and astaxanthin production at levels comparable to those seen with commercial yeast extract. Overall, cultivating X. dendrorhous in an SM-based medium containing 5% SM and 0.2% yeast extract obtained from residual brewers' yeast resulted in significantly higher (> 20% more) biomass accumulation compared to the control media (YPD). A similar slightly higher astaxanthin output (up to 14% more) was recorded in the SM-based medium compared to YPD. The formulated cultivation medium in this study provides an opportunity to reduce the production cost of astaxanthin from X. dendrorhous while simultaneously reducing the environmental impact related to the disposal of the industrial waste used as feedstock. KEY POINTS: • Cheap culture media were formulated from soy molasses and brewers' spent yeast • The formulated medium resulted in at least 20% more biomass than the control • Up to 14% more astaxanthin was produced in molasses-based medium., (© 2024. The Author(s).)

In this work, a multivariate analysis was carried out, using a Plackett-Burman (PB) design involving seventeen growth parameters, on carotenoids production of Pavlova gyrans (p < 0.10). Each assay was analysed regarding its content (mg g -1 ) of fucoxanthin (Fx), diatoxanthin, diadinoxanthin, β-carotene (βCar), α-carotene, and the sum of all carotenoids analysed individually (TCar). According to the statistical analysis, modified medium formulations were developed for the particular cases of Fx, βCar, and TCar. The study showed that Fx content was positively affected by nitrogen supplementation and lower light intensities. Higher concentrations of nitrogen and iron increased the final content of βCar as well. Similarly, salinity, light intensity, nitrogen, iron, and cobalt were identified as key factors in TCar production. The PB-based formulations showed significant improvements (p < 0.05) for TCar (11.794 mg g -1 ) and Fx (6.153 mg g -1 ) when compared to the control conditions (Walne's medium-2.010 mg g -1 ). Furthermore, effective control of key variables (e.g., light intensity) throughout P. gyrans growth proved successful (p < 0.05), increasing the productivity of Fx (0.759 mg L -1  d -1 ) and TCar (1.615 mg L -1  d -1 )., (© 2024. The Author(s).)

It is well established that Staphylococcus aureus can incorporate exogenous straight-chain unsaturated fatty acids (SCUFAs) into membrane phospho- and glyco-lipids from various sources in supplemented culture media and when growing in vivo during infection. Given the enhancement of membrane fluidity when oleic acid (C18:1Δ9) is incorporated into lipids, we were prompted to examine the effect of medium supplementation with C18:1Δ9 on growth at low temperatures. C18:1Δ9 supported the growth of a cold-sensitive, branched-chain fatty acid (BCFA)-deficient mutant at 12°C. Interestingly, we found similar results in the BCFA-sufficient parental strain, supported by the fact that the incorporation of C18:1Δ9 into the membrane increased membrane fluidity in both strains. We show that the incorporation of C18:1Δ9 and its elongation product C20:1Δ11 into membrane lipids was required for growth stimulation and relied on a functional FakAB incorporation system. Lipidomics analysis of the phosphatidylglycerol and diglycosyldiacylglycerol lipid classes revealed major impacts of C18:1Δ9 and temperature on lipid species. Growth at 12°C in the presence of C18:1Δ9 also led to increased production of the carotenoid pigment staphyloxanthin. The enhancement of growth by C18:1Δ9 is an example of homeoviscous adaptation to low temperatures utilizing an exogenous fatty acid. This may be significant in the growth of S. aureus at low temperatures in foods that commonly contain C18:1Δ9 and other SCUFAs in various forms., Importance: We show that Staphylococcus aureus can use its known ability to incorporate exogenous fatty acids to enhance its growth at low temperatures. Individual species of phosphatidylglycerols and diglycosyldiacylglycerols bearing one or two degrees of unsaturation derived from the incorporation of C18:1Δ9 at 12°C are described for the first time. In addition, enhanced production of the carotenoid staphyloxanthin occurs at low temperatures. The studies describe a biochemical reality underlying membrane biophysics. This is an example of homeoviscous adaptation to low temperatures utilizing exogenous fatty acids over the regulation of the biosynthesis of endogenous fatty acids. The studies have likely relevance to food safety in that unsaturated fatty acids may enhance the growth of S. aureus in the food environment., Competing Interests: The authors declare no conflict of interest.

Variances in the biological functions of astaxanthin geometric isomers (i.e., all- E , Z ) are related to their intestinal absorption, but the mechanism of isomer absorption mediated by transporters remains unclear. Here, models of in vitro cell overexpression, in situ intestinal perfusion, and in vivo mouse inhibition were employed to investigate the impact of cluster of differentiation 36 (CD36) on the absorption of astaxanthin isomers. Cells overexpressing CD36 notably enhanced the uptake of Z -astaxanthin, particularly the 9- Z -isomer (47.76%). The absorption rate and permeability of Z -astaxanthin surpassed that of the all- E -isomer by the in situ model. Furthermore, the addition of the CD36-specific inhibitor sulfo- N -succinimidyl oleate significantly reduced the absorption of Z -astaxanthin in the mouse duodenum and jejunum, especially the 9- Z -isomer (57.66%). Molecular docking and surface plasmon resonance techniques further validated that 9- Z -astaxanthin binds to more amino acids of CD36 with higher affinity and in a fast-binding, fast-dissociating mode, thus favoring transport. Our findings elucidate, for the first time, the mechanism of the CD36-mediated transmembrane transport of astaxanthin geometric isomers.

Astaxanthin is a powerful antioxidant known to enhance skin, cardiovascular, eye, and brain health. In this study, the genome insights and astaxanthin production of two newly isolated astaxanthin-producing yeasts (TL35-5 and PL61-2) were evaluated and compared. Based on their phenotypic and genotypic characteristics, TL35-5 and PL61-2 were identified as basidiomycetous yeasts belonging to Rhodotorula paludigena and Rhodotorula sampaioana, respectively. To optimize astaxanthin production, the effects of cultural medium composition and cultivation conditions were examined. The optimal conditions for astaxanthin production in R. paludigena TL35-5 involved cultivation in AP medium containing 10 g/L glucose as the sole carbon source, supplemented with 1.92 g/L potassium nitrate, pH 6.5, and incubation at 20°C for 3 days with shaking at 200 rpm. For R. sampaioana PL61-2, the optimal medium composition for astaxanthin production consisted of AP medium with 40 g/L glucose, supplemented with 0.67 g/L urea, pH 7.5, and the fermentation was carried out at 20°C for 3 days with agitating at 200 rpm. Under their optimal conditions, R. paludigena TL35-5 and R. sampaioana PL61-2 gave the highest astaxanthin yields of 3.689 ± 0.031 and 4.680 ± 0.019 mg/L, respectively. The genome of TL35-5 was 20,982,417 bp in length, with a GC content of 64.20%. A total of 6,789 protein-encoding genes were predicted. Similarly, the genome of PL61-2 was 21,374,169 bp long, with a GC content of 64.88%. It contained 6,802 predicted protein-encoding genes. Furthermore, all essential genes involved in astaxanthin biosynthesis, including CrtE, CrtYB, CrtI, CrtS, and CrtR, were identified in both R. paludigena TL35-5 and R. sampaioana PL61-2, providing evidence for their ability to produce astaxanthin., Competing Interests: The authors declare no competing interests., (Copyright: © 2024 Hoondee et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Snow (cryotolerant) algae often form red (pink) spots in mountain ecosystems on snowfields around the world, but little is known about their physiology and chemical composition. Content and composition of pigments in the cells of the cryotolerant green microalgae Chloromonas reticulata have been studied. Analysis of carotenoids content in the green (vegetative) cells grown under laboratory conditions and in the red resting cells collected from the snow surface in the Subpolar Urals was carried out. Carotenoids such as neoxanthin, violaxanthin, anteraxanthin, zeaxanthin, lutein, and β-carotene were detected. Among the carotenoids, the ketocarotenoid astaxanthin with high biological activity was also found. It was established that cultivation of the algae at low positive temperature (6°C) and moderate illumination (250 μmol quanta/(m 2 ⋅s) contributed to accumulation of all identified carotenoids, including extraplastidic astaxanthin. In addition to the pigments, fatty acids accumulated in the algae cells. The data obtained allow us to consider the studied microalgae as a potentially promising species for production of carotenoids.

Maximal sunlight intensity varies diurnally due to the earth's rotation. Whether this slow diurnal pattern influences the photoprotective capacity of plants throughout the day is unknown. We investigated diurnal variation in NPQ, along with NPQ capacity, induction, and relaxation kinetics after transitions to high light, in tomato plants grown under diurnal parabolic (DP) or constant (DC) light intensity regimes. DP light intensity peaked at midday (470 μmol m -2 s -1) while DC stayed constant at 300 μmol m -2 s -1 at a similar 12-hour photoperiod and daily light integral. NPQs were higher in the morning and afternoon at lower light intensities in DP compared to DC, except shortly after dawn. NPQ capacity increased from midday to the end of the day, with higher values in DP than in DC. At high light Φ PSII did not vary throughout the day, while Φ NPQ varied consistently with NPQ capacity. Reduced Φ NO suggested less susceptibility to photodamage at the end of the day. NPQ induction was faster at midday than at the start of the day and in DC than in DP, with overshoot occurring in the morning and midday but not at the end of the day. NPQ relaxation was faster in DP than in DC. The xanthophyll de-epoxidation state and reduced demand for photochemistry could not explain the observed diurnal variations in photoprotective capacity. In conclusion, this study showed diurnal variation in regulated photoprotective capacity at moderate growth light intensity, which was not explained by instantaneous light intensity or increasing photoinhibition over the day and was influenced by acclimation to constant light intensity., (© 2024 The Author(s). Physiologia Plantarum published by John Wiley & Sons Ltd on behalf of Scandinavian Plant Physiology Society.)

Objectives: Aggregation and misfolding of amyloid beta (Aβ) and tau proteins, suggested to arise from post-translational modification processes, are thought to be the main cause of Alzheimer's disease (AD). Additionally, a plethora of evidence exists that links metabolic dysfunctions such as obesity, type 2 diabetes (T2D), and dyslipidemia to the pathogenesis of AD. We thus investigated the combinatory effect of T2D and human glutaminyl cyclase activity (pyroglutamylation), on the pathology of AD and whether astaxanthin (ASX) treatment ameliorates accompanying pathophysiological manifestations., Methods: Male transgenic AD mice, APPxhQC, expressing human APP751 with the Swedish and the London mutation and human glutaminyl cyclase (hQC) enzyme and their non-transgenic (NTG) littermates were used. Both APPxhQC and NTG mice were allocated to 3 groups, control, T2D-control, and T2D-ASX. Mice were fed control or high fat diet ± ASX for 13 weeks starting at an age of 11-12 months. High fat diet fed mice were further treated with streptozocin for T2D induction. Effects of genotype, T2D induction, and ASX treatment were evaluated by analysing glycemic readouts, lipid concentration, Aβ deposition, hippocampus-dependent cognitive function and nutrient sensing using immunosorbent assay, ELISA-based assays, western blotting, immunofluorescence staining, and behavioral testing via Morris water maze (MWM), respectively., Results: APPxhQC mice presented a higher glucose sensitivity compared to NTG mice. T2D-induced brain dysfunction was more severe in NTG compared to the APPxhQC mice. T2D induction impaired memory functions while increasing hepatic LC3B, ABCA1, and p65 levels in NTG mice. T2D induction resulted in a progressive shift of Aβ from the soluble to insoluble form in APPxhQC mice. ASX treatment reversed T2D-induced memory dysfunction in NTG mice and in parallel increased hepatic pAKT while decreasing p65 and increasing cerebral p-S6rp and p65 levels. ASX treatment reduced soluble Aβ38 and Aβ40 and insoluble Aβ40 levels in T2D-induced APPxhQC mice., Conclusions: We demonstrate that T2D induction in APPxhQC mice poses additional risk for AD pathology as seen by increased Aβ deposition. Although ASX treatment reduced Aβ expression in T2D-induced APPxhQC mice and rescued T2D-induced memory impairment in NTG mice, ASX treatment alone may not be effective in cases of T2D comorbidity and AD., Competing Interests: Declaration of competing interest T.L., I.S., S.S., S.F., J.N., M.P. and B.H.P. are presently or formerly affiliated with QPS Austria GmbH. S.F. is further affiliated with BioDoks e. U., (Copyright © 2024 The Authors. Published by Elsevier GmbH.. All rights reserved.)

The determination of physiological tolerance ranges of photosynthetic species and of the biochemical mechanisms underneath are fundamental to identify target processes and metabolites that will inspire enhanced plant management and production for the future. In this context, the terrestrial green algae within the genus Prasiola represent ideal models due to their success in harsh environments (polar tundras) and their extraordinary ecological plasticity. Here we focus on the outstanding Prasiola antarctica and compare two natural populations living in very contrasting microenvironments in Antarctica: the dry sandy substrate of a beach and the rocky bed of an ephemeral freshwater stream. Specifically, we assessed their photosynthetic performance at different temperatures, reporting for the first time g nsd values in algae and changes in thylakoid metabolites in response to extreme desiccation. Stream population showed lower α-tocopherol content and thicker cell walls and thus, lower g nsd and photosynthesis. Both populations had high temperatures for optimal photosynthesis (around +20°C) and strong constitutive tolerance to freezing and desiccation. This tolerance seems to be related to the high constitutive levels of xanthophylls and of the cylindrical lipids di- and tri-galactosyldiacylglycerol in thylakoids, very likely related to the effective protection and stability of membranes. Overall, P. antarctica shows a complex battery of constitutive and plastic protective mechanisms that enable it to thrive under harsh conditions and to acclimate to very contrasting microenvironments, respectively. Some of these anatomical and biochemical adaptations may partially limit photosynthesis, but this has a great potential to rise in a context of increasing temperature., (© 2024 The Authors. The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.)

Astaxanthin is a high-value natural antioxidant, and can be accumulated in Microcystis aeruginosa. To enhance astaxanthin accumulation in the microalgae by using salt stress, the cell growth, photosynthetic abilities, reactive oxygen species (ROS) levels, astaxanthin and its precursor content, and gene expression were investigated under NaCl and KCl stresses. The two salt stresses inhibited the cell growth by lowering photosynthetic abilities and raising ROS levels. During the 6-day treatment, the two salt stresses improved the levels of astaxanthin, precursors (β-carotene and zeaxanthin) and carotenoids, which might be caused by the raised ROS up-regulating expression of 7 related genes. At the same concentration, KCl stress showed stronger inducing effect on astaxanthin and its precursor production than NaCl stress, due to higher expression of related genes. Therefore, NaCl and KCl stresses have obvious ion differences on astaxanthin accumulation, of which KCl stress is more suitable for the high-value antioxidant production from microalgae., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Monogalactosyldiacylglycerol (MGDG) is the main lipid constituent of thylakoids and a structural component of photosystems and photosynthesis-related proteo-lipid complexes in green tissues. Previously reported changes in MGDG abundance upon stress treatments are hypothesized to reflect mobilization of MGDG-based polyunsaturated lipid intermediates to maintain extraplastidial membrane integrity. While exchange of lipid intermediates between compartmental membranes is well documented, physiological consequences of mobilizing an essential thylakoid lipid, such as MGDG, for an alternative purpose are not well understood. Arabidopsis seedlings exposed to mild (50 mM) salt treatment displayed significantly increased abundance of both MGDG and the extraplastidial lipid, phosphatidylcholine (PC). Interestingly, similar increases in MGDG and PC were observed in Arabidopsis fad3 mutant seedlings defective in endoplasmic reticulum (ER)-localized linolenic acid formation, in which compensatory plastid-to-ER-directed mobilization of linolenic acid-containing intermediates takes place. The postulated (salt) or evident (fad3) plastid-ER exchange of intermediates concurred with altered thylakoid function according to parameters of photosynthetic performance. While salt treatment of wild-type seedlings inhibited photosynthetic parameters in a dose-dependent manner, interestingly, untreated fad3 mutants did not show overall reduced photosynthetic quantum yield. By contrast, we observed a reduction specifically of non-photochemical quenching (NPQ) under high light, representing only part of observed salt effects. The decreased NPQ in the fad3 mutant was accompanied by reduced activity of the xanthophyll cycle, leading to a reduced concentration of the NPQ-effective pigment zeaxanthin. The findings suggest that altered ER-located fatty acid unsaturation and ensuing inter-organellar compensation impacts on the function of specific thylakoid enzymes, rather than globally affecting thylakoid function., (© The Author(s) 2023. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Phaeodactylum tricornutum a prominent source of industrial fucoxanthin production, faces challenges in its application due to its tolerance to high-temperature environments. This study investigates the physiological responses of P. tricornutum to high-temperature stress and its impact on fucoxanthin content, with a specific focus on the role of cis -zeatin. The results reveal that high-temperature stress inhibits P. tricornutum 's growth and photosynthetic activity, leading to a decrease in fucoxanthin content. Transcriptome analysis shows that high temperature suppresses the expression of genes related to photosynthesis (e.g., psbO , psbQ , and OEC ) and fucoxanthin biosynthesis (e.g., PYS , PDS1 , and PSD2 ), underscoring the negative effects of high temperature on P. tricornutum . Interestingly, genes associated with cis -zeatin biosynthesis and cytokinesis signaling pathways exhibited increased expression under high-temperature conditions, indicating a potential role of cis -zeatin signaling in response to elevated temperatures. Content measurements confirm that high temperature enhances cis -zeatin content. Furthermore, the exogenous addition of cytokinesis mimetics or inhibitors significantly affected P. tricornutum 's high-temperature resistance. Overexpression of the cis-zeatin biosynthetic enzyme gene tRNA DMATase enhanced P. tricornutum 's resistance to high-temperature stress, while genetic knockout of tRNA DMATas e reduced its resistance to high temperatures. Therefore, this research not only uncovers a novel mechanism for high-temperature resistance in P. tricornutum but also offers a possible alga species that can withstand high temperatures for the industrial production of fucoxanthin, offering valuable insights for practical utilization.IMPORTANCEThis study delves into Phaeodactylum tricornutum 's response to high-temperature stress, specifically focusing on cis -zeatin. We uncover inhibited growth, reduced fucoxanthin, and significant cis -zeatin-related gene expression under high temperatures, highlighting potential signaling mechanisms. Crucially, genetic engineering and exogenous addition experiments confirm that the change in cis -zeatin levels could influence P. tricornutum 's resistance to high-temperature stress. This breakthrough deepens our understanding of microalgae adaptation to high temperatures and offers an innovative angle for industrial fucoxanthin production. This research is a pivotal step toward developing heat-resistant microalgae for industrial use., Competing Interests: The authors declare no conflict of interest.

Biofilm formation by methicillin-resistant Staphylococcus aureus (MRSA) on indwelling medical devices complicates the treatment of infection. Tetrabromobisphenol A (TBBPA), a synthetic, lipophilic, halogenated aromatic compound widely used as an additive in plastics and electronic products, has raised environmental concerns due to its potential for bioaccumulation. This study investigated the impact of sub-inhibitory concentrations of TBBPA on MRSA biofilm formation. Crystal violet staining and confocal laser scanning microscopy analysis demonstrated that 1/8 MIC (0.5 µg/mL) of TBBPA significantly stimulated MRSA biofilm formation (P < 0.0001). MTT assays indicated that the metabolic activity within the biofilms increased by 15.60-40.85% compared to untreated controls. Dot blot immunoassay, autolysis assay, and extracellular DNA (eDNA) quantification further revealed TBBPA enhanced the production of polysaccharide intercellular adhesin (PIA) and eDNA, which are key biofilm components. Additionally, TBBPA was found to enhance the production of staphyloxanthin, facilitating MRSA survival under oxidative conditions and in human whole blood. RT-qPCR analysis showed that TBBPA significantly upregulated genes associated with biofilm formation (icaA, atlA, sarA), staphyloxanthin biosynthesis (crtM and sigB), and oxidative stress responses (sodA and katA). These findings suggest that TBBPA promotes MRSA biofilm development and enhances bacterial resistance to adverse conditions, thereby potentially exacerbating risks to human health., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Dunaliella salina, a microalga that thrives under high-saline conditions, is notable for its high β-carotene content and the absence of a polysaccharide cell wall. These unique characteristics render it a prime candidate as a cellular platform for astaxanthin production. In this study, our initial tests in an E. coli revealed that β-ring-4-dehydrogenase (CBFD) and 4-hydroxy-β-ring-4-dehydrogenase (HBFD) genes from Adonis aestivalis outperformed β-carotene hydroxylase (BCH) and β-carotene ketolase (BKT) from Haematococcus pluvialis counterparts by two-fold in terms of astaxanthin biosynthesis efficiency. Subsequently, we utilized electroporation to integrate either the BKT gene or the CBFD and HBFD genes into the genome of D. salina. In comparison to wild-type D. salina, strains transformed with BKT or CBFD and HBFD exhibited inhibited growth, underwent color changes to shades of red and yellow, and saw a nearly 50% decline in cell density. HPLC analysis confirmed astaxanthin synthesis in engineered D. salina strains, with CBFD + HBFD-D. salina yielding 134.88 ± 9.12 μg/g of dry cell weight (DCW), significantly higher than BKT-D. salina (83.58 ± 2.40 μg/g). This represents the largest amount of astaxanthin extracted from transgenic D. salina, as reported to date. These findings have significant implications, opening up new avenues for the development of specialized D. salina-based microcell factories for efficient astaxanthin production., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

The production of compact vectors for gene stacking is hindered by a lack of effective linkers. Here, we report that a 26-nt nucleic acid linker, NAL1, from the fungus Glarea lozoyensis and its truncated derivatives could connect two genes as a bicistron, enabling independent translation in a maize protoplast transient expression system and human 293 T cells. The optimized 9-nt NAL10 linker was then used to connect four genes driven by a bidirectional promoter; this combination was successfully used to reconstruct the astaxanthin biosynthesis pathway in transgenic maize. The short and efficient nucleic acid linker NAL10 can be widely used in multi-gene expression and synthetic biology in animals and plants., (© 2024 Society for Experimental Biology and John Wiley & Sons Ltd.)

This study established and investigated continuous macular pigment (MP) production with a lutein (L):zeaxanthin (Z) ratio of 4-5:1 by an MP-rich Chlorella sp. CN6 mutant strain in a continuous microalgal culture module. Chlorella sp. CN6 was cultured in a four-stage module for 10 days. The microalgal culture volume increased to 200 L in the first stage (6 days). Biomass productivity increased to 0.931 g/L/day with continuous indoor white light irradiation during the second stage (3 days). MP content effectively increased to 8.29 mg/g upon continuous, indoor white light and blue light-emitting diode irradiation in the third stage (1 day), and the microalgal biomass and MP concentrations were 8.88 g/L and 73.6 mg/L in the fourth stage, respectively. Using a two-step MP extraction process, 80 % of the MP was recovered with a high purity of 93 %, and its L:Z ratio was 4-5:1., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Engineering Yarrowia lipolytica to produce astaxanthin provides a promising route. Here, Y. lipolytica M2 producing a titer of 181 mg/L astaxanthin was isolated by iterative atmospheric and room-temperature plasma mutagenesis and diphenylamine-mediated screening. Interestingly, a negative correlation was observed between cell biomass and astaxanthin production. To reveal the underlying mechanism, RNA-seq analysis of transcriptional changes was performed in high producer M2 and reference strain M1, and a total of 1379 differentially expressed genes were obtained. Data analysis revealed that carbon flux was elevated through lipid metabolism, acetyl-CoA and mevalonate supply, but restrained through central carbon metabolism in strain M2. Moreover, upregulation of other pathways such as ATP-binding cassette transporter and thiamine pyrophosphate possibly provided more cofactors for carotenoid hydroxylase and relieved cell membrane stress caused by astaxanthin insertion. These results suggest that balancing cell growth and astaxanthin production may be important to promote efficient biosynthesis of astaxanthin in Y. lipolytica., (© 2024 John Wiley & Sons Ltd.)

Effective metabolic regulators play an essential role in regulating astaxanthin biosynthesis in Phaffia rhodozyma. In this study, it was found that 5 mM glutamate increased the astaxanthin yield and biomass of P. rhodozyma D3 to 22.34 mg/L and 6.12 g/L, which were 1.22 and 1.33 times higher than the control group, respectively. Meanwhile, glucose uptake was increased and the level of reactive oxygen species (ROS) was reduced with 5 mM glutamate. To further explore the interrelationship between glutamate and astaxanthin synthesis, the energy metabolism of P. rhodozyma D3 with and without glutamate was analysed. Glutamate promoted the Embden-Meyerhof-Parnas pathway (EMP) metabolic flux, modulated the tricarboxylic acid (TCA) cycle and the pentose phosphate pathway (PPP), activated the ornithine cycle and purine metabolism, and provided more ATP and NADPH for astaxanthin accumulation. This study clarified the possible mechanism by which glutamate promoted astaxanthin accumulation in P. rhodozyma., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Microalgae are the preferred species for producing astaxanthin because they pose a low toxicity risk than chemical synthesis. Astaxanthin has multiple health benefits and is being used in: medicines, nutraceuticals, cosmetics, and functional foods. Haematococcus pluvialis is a model microalga for astaxanthin biosynthesis; however, its natural astaxanthin content is low. Therefore, it is necessary to develop methods to improve the biosynthesis of astaxanthin to meet industrial demands, making its commercialization cost-effective. Several strategies related to cultivation conditions are employed to enhance the biosynthesis of astaxanthin in H. pluvialis. However, the mechanism of its regulation by transcription factors is unknown. For the first time, this study critically reviewed the studies on identifying transcription factors, progress in H. pluvialis genetic transformation, and use of phytohormones that increase the gene expression related to astaxanthin biosynthesis. In addition, we propose future approaches, including (i) Cloning and characterization of transcription factors, (ii) Transcriptional engineering through overexpression of positive regulators or downregulation/silencing of negative regulators, (iii) Gene editing for enrichment or deletion of transcription factors binding sites, (iv) Hormonal modulation of transcription factors. This review provides considerable knowledge about the molecular regulation of astaxanthin biosynthesis and the existing research gap. Besides, it provides the basis for transcription factors mediated metabolic engineering of astaxanthin biosynthesis in H. pluvialis .

Fucoxanthin is a versatile substance in the food and pharmaceutical industries owing to its excellent antioxidant and anti-obesity properties. Several microalgae, including the haptophyte Pavlova spp., can produce fucoxanthin and are potential industrial fucoxanthin producers, as they lack rigid cell walls, which facilitates fucoxanthin extraction. However, the commercial application of Pavlova spp. is limited owing to insufficient biomass production. In this study, we aimed to develop a mixotrophic cultivation method to increase biomass and fucoxanthin production in Pavlova gyrans OPMS 30543X. The effects of culturing OPMS 30543X with different organic carbon sources, glycerol concentrations, mixed-nutrient conditions, and light intensities on the consumption of organic carbon sources, biomass production, and fucoxanthin accumulation were analyzed. Several organic carbon sources, such as glycerol, glucose, sucrose, and acetate, were examined, revealing that glycerol was well-consumed by the microalgae. Biomass and fucoxanthin production by OPMS 30543X increased in the presence of 10 mM glycerol compared to that observed without glycerol. Metabolomic analysis revealed higher levels of the metabolites related to the glycolytic, Calvin-Benson-Bassham, and tricarboxylic acid cycles under mixotrophic conditions than under autotrophic conditions. Cultures grown under mixotrophic conditions with a light intensity of 100 µmol photons m -2 s -1 produced more fucoxanthin than autotrophic cultures. Notably, the amount of fucoxanthin produced (18.9 mg/L) was the highest reported thus far for Pavlova species. In conclusion, the use of mixotrophic culture is a promising strategy for increasing fucoxanthin production in Pavlova species. KEY POINTS: • Glycerol enhances biomass and fucoxanthin production in Pavlova gyrans • Metabolite levels increase under mixotrophic conditions • Mixotrophic conditions and medium-light intensity are appropriate for P. gyrans., (© 2024. The Author(s).)

Physiological and environmental cues prompt microbes to synthesize diverse carotenoids, including dihydroxy xanthophylls, facilitating their adaptation and survival. Lutein and its isomeric counterpart, zeaxanthin, are notable dihydroxy xanthophylls with bioactive properties such as antioxidative, anti-inflammatory, anticancer, and neuroprotective effects, particularly beneficial for human ocular health. However, global natural resources for co-producing lutein and zeaxanthin are scarce, with zeaxanthin lacking commercial sources, unlike lutein sourced from marigold plants and microalgae. Traditionally, dihydroxy xanthophyll production primarily relies on petrochemical synthetic routes, with limited biological sourcing reported. Nonetheless, microbiological synthesis presents promising avenues as a commercial source, albeit challenged by low dihydroxy xanthophyll yield at high cell density. Strategies involving optimization of physical and chemical parameters are essential to achieve high-quality dihydroxy xanthophyll products. This overview briefly discusses dihydroxy xanthophyll biosynthesis and highlights recent advancements, discoveries, and industrial benefits of lutein and zeaxanthin production from microorganisms as alternative biofactories., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Violaxanthin is a plant-derived orange xanthophyll with remarkable antioxidant activity that has wide applications in various industries, such as food, agriculture, and cosmetics. In addition, it is the key precursor of important substances such as abscisic acid and fucoxanthin. Saccharomyces cerevisiae , as a GRAS (generally regarded as safe) chassis, provides a good platform for producing violaxanthin production with a yield of 7.3 mg/g DCW, which is far away from commercialization. Herein, an integrated strategy involving zeaxanthin epoxidase (ZEP) source screening, cytosol redox state engineering, and nicotinamide adenine dinucleotide phosphate (NADPH) regeneration was implemented to enhance violaxanthin production in S. cerevisiae . 58aa-truncated ZEP from Vitis vinifera exhibited optimal efficiency in an efficient zeaxanthin-producing strain. The titer of violaxanthin gradually increased by 17.9-fold (up to 119.2 mg/L, 15.19 mg/g DCW) via cytosol redox state engineering and NADPH supplementation. Furthermore, balancing redox homeostasis considerably improved the zeaxanthin concentration by 139.3% (up to 143.9 mg/L, 22.06 mg/g DCW). Thus, the highest reported titers of violaxanthin and zeaxanthin in S. cerevisiae were eventually achieved. This study not only builds an efficient platform for violaxanthin biosynthesis but also serves as a useful reference for the microbial production of xanthophylls.

Fucoxanthin, a carotenoid exclusively derived from algae, exerts its bioactivities with the modulation of the gut microbiota in mice. However, mechanisms through which fucoxanthin regulates the gut microbiota and its derived metabolites/metabolism in humans remain unclear. In this study, we investigated the effects of fucoxanthin on the gut microbiota and metabolism of non-obese individuals using an in vitro simulated digestion-fermentation cascade model. The results showed that about half of the fucoxanthin was not absorbed in the intestine, thus reaching the colon. The gut microbiota from fecal samples underwent significant changes after 48 or 72 hours in vitro fermentation. Specifically, fucoxanthin significantly enhanced the relative abundance of Bacteroidota and Parabacteroides , leading to improved functions of the gut microbiota in its development, glycan biosynthesis and metabolism as well as in improving the digestive system, endocrine system and immune system. The recovery of fucoxanthin during fermentation showed a decreasing trend with the slight bio-conversion of fucoxanthinol. Notably, fucoxanthin supplementation significantly altered metabolites, especially bile acids and indoles in the simulated human gut ecosystem. Correlation analysis indicated the involvement of the gut microbiota in the manipulation of these metabolites by fucoxanthin. Moreover, all these altered metabolites revealed the improvement in the capacity of fucoxanthin in manipulating gut metabolism, especially lipid metabolism. Overall, fucoxanthin determinedly reshaped the gut microbiota and metabolism, implying its potential health benefits in non-obese individuals.

Our goal was to determine whether anthocyanin-producing species (red) use different photoprotective strategies to cope with excess light during fall senescence compared with non-anthocyanin-producing species (yellow). In a previous study, we found that a yellow species retained the photoprotective PsbS protein in late autumn, while a red species did not. Specifically, we tested the hypothesis that red species make less use of zeaxanthin and PsbS-mediated thermal dissipation, as they rely on anthocyanins for photoprotection. We monitored four red (Acer ginnala, Rhus typhnia, Parenthocissus quinquefolia, Viburnum dentatum) and four yellow species (Acer negundo, Ostrya virginiana, Vitis riparia, Zanthoxylum americanum) throughout autumn senescence and analyzed pigments, protein content, and chlorophyll fluorescence. We found yellow species retained the PsbS protein at higher levels, and had higher dark retention of zeaxanthin in late autumn relative to red species. All species retained lutein and the pool of xanthophyll cycle pigments in higher amounts than other carotenoids in late autumn. Our data support the hypothesis that red species use anthocyanins as a photoprotective strategy during autumn senescence, and therefore make less use of PsbS and zeaxanthin-mediated thermal dissipation. We also found species-specific variation in the particular combination of photoprotective strategies used., (© 2024 Scandinavian Plant Physiology Society.)

Soil cadmium (Cd) contamination poses a significant threat to global food security and the environment. Astaxanthin (AX), a potent biological antioxidant belonging to the carotenoid group, has been demonstrated to confer tolerance against diverse abiotic stresses in plants. This study investigated the potential of AX in mitigating Cd-induced damage in wheat seedlings. Morpho-physiological, ultrastructural, and biochemical analyses were conducted to evaluate the impact of AX on Cd-exposed wheat seedlings. Illumina-based gene expression profiling was employed to uncover the molecular mechanisms underlying the protective effects of AX. The addition of 100 μM AX alleviated Cd toxicity by enhancing various parameters: growth, photosynthesis, carotenoid content, and total antioxidant capacity (T-AOC), while reducing Cd accumulation, malondialdehyde (MDA), and hydrogen peroxide (H 2 O 2 ) levels. RNA sequencing analysis revealed differentially expressed genes associated with Cd uptake and carotenoid metabolism, such as zinc/iron permease (ZIP), heavy metal-associated protein (HMA), 3-beta hydroxysteroid dehydrogenase/isomerase (3-beta-HSD), and thiolase. These findings suggest that AX enhances Cd tolerance in wheat seedlings by promoting the expression of detoxification and photosynthesis-related genes. This research offers valuable insights into the potential use of AX to address Cd contamination in agricultural systems, highlighting the significance of antioxidant supplementation in plant stress management., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Masson SAS. All rights reserved.)

Carotenoids are pigments that have a range of functions in human health. The carotenoid diatoxanthin is suggested to have antioxidant, anti-inflammatory and chemo-preventive properties. Diatoxanthin is only produced by a few groups of microalgae, where it functions in photoprotection. Its large-scale production in microalgae is currently not feasible. In fact, rapid conversion into the inactive pigment diadinoxanthin is triggered when cells are removed from a high-intensity light source, which is the case during large-scale harvesting of microalgae biomass. Zeaxanthin epoxidase (ZEP) 2 and/or ZEP3 have been suggested to be responsible for the back-conversion of high-light accumulated diatoxanthin to diadinoxanthin in low-light in diatoms. Using CRISPR/Cas9 gene editing technology, we knocked out the ZEP2 and ZEP3 genes in the marine diatom Phaeodactylum tricornutum to investigate their role in the diadinoxanthin-diatoxanthin cycle and determine if one of the mutant strains could function as a diatoxanthin production line. Light-shift experiments proved that ZEP3 encodes the enzyme converting diatoxanthin to diadinoxanthin in low light. Loss of ZEP3 caused the high-light-accumulated diatoxanthin to be stable for several hours after the cultures had been returned to low light, suggesting that zep3 mutant strains could be suitable as commercial production lines of diatoxanthin., Competing Interests: The authors declare no conflicts of interest.

Astaxanthin, a versatile C40 carotenoid prized for its applications in food, cosmetics, and health, is a bright red pigment with powerful antioxidant properties. To enhance astaxanthin production in Corynebacterium glutamicum, we employed rational pathway engineering strategies, focused on improving precursor availability and optimizing terminal oxy-functionalized C40 carotenoid biosynthesis. Our efforts resulted in an increased astaxanthin precursor supply with 1.5-fold higher β-carotene production with strain BETA6 (18 mg g -1  CDW). Further advancements in astaxanthin production were made by fine-tuning the expression of the β-carotene hydroxylase gene crtZ and β-carotene ketolase gene crtW, yielding a nearly fivefold increase in astaxanthin (strain ASTA**), with astaxanthin constituting 72% of total carotenoids. ASTA** was successfully transferred to a 2 L fed-batch fermentation with an enhanced titer of 103 mg L -1 astaxanthin with a volumetric productivity of 1.5 mg L -1  h -1 . Based on this strain a pathway expansion was achieved towards glycosylated C40 carotenoids under heterologous expression of the glycosyltransferase gene crtX. To the best of our knowledge, this is the first time astaxanthin-β-D-diglucoside was produced with C. glutamicum achieving high titers of microbial C40 glucosides of 39 mg L -1 . This study showcases the potential of pathway engineering to unlock novel C40 carotenoid variants for diverse industrial applications., (© 2024. The Author(s).)

Diatoms, a major group of algae, account for about a quarter of the global primary production on Earth. These photosynthetic organisms face significant challenges due to light intensity variations in their underwater habitat. To avoid photodamage, they have developed very efficient non-photochemical quenching (NPQ) mechanisms. These mechanisms originate in their light-harvesting antenna - the fucoxanthin-chlorophyll protein (FCP) complexes. Spectroscopic studies of NPQ in vivo are often hindered by strongly overlapping signals from the photosystems and their antennae. Fortunately, in vitro FCP aggregates constitute a useful model system to study fluorescence (FL) quenching in diatoms. In this work, we present streak-camera FL measurements on FCPa and FCPb complexes, isolated from a centric diatom Cyclotella meneghiniana, and their aggregates. We find that spectra of non-aggregated FCP are dominated by a single fluorescing species, but the FL spectra of FCP aggregates additionally contain contributions from a redshifted emissive state. We relate this red state to a charge transfer state between chlorophyll c and chlorophyll a molecules. The FL quenching, on the other hand, is due to an additional dark state that involves incoherent energy transfer to the fucoxanthin carotenoids. Overall, the global picture of energy transfer and quenching in FCP aggregates is very similar to that of major light-harvesting complexes in higher plants (LHCII), but microscopic details between FCPs and LHCIIs differ significantly., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The health benefits of astaxanthin (AST) are related to its geometric isomers. Generally, functional activity is realized by the interactions between active substances and transporters. Hereto, bovine serum albumin (BSA), as a model-binding protein and transporter, is able to recognize and transport isomers of active substances through binding with them. However, differences in the binding mechanism of isomers to BSA may affect the functional activities of isomers through the "binding-transport-activity" chain reaction. Thus, this study sought to elucidate the interactions between AST geometrical isomers and BSA using multi-spectroscopy, surface plasmon resonance and molecular docking. The results showed that Z-AST displayed more interacting amino acid residues and lower thermodynamic parameters than all-E-AST. Meanwhile, the order of binding affinity to BSA was 13Z-AST (1.56 × 10 -7 M) > 9Z-AST (2.70 × 10 -7 M) > all-E-AST (4.01 × 10 -7 M), indicating that Z-AST possessed stronger binding ability to BSA. Moreover, AST isomers were located at the junction between subdomains ⅡA and ⅢA of BSA, and showed the same interaction forces (hydrogen bond and van der Waals force) as well as kinetic processes (slow combination, slow dissociation). These interaction parameters provide valuable insights into their pharmacokinetics in vivo, and it was of great significance to explain the potential differences among AST isomers in functional activities., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

The demand for astaxanthin has been increasing for many health applications ranging from pharmaceuticals, food, cosmetics, and aquaculture due to its bioactive properties. Haematococcus pluvialis is widely recognized as the microalgae species with the highest natural accumulation of astaxanthin, which has made it a valuable source for industrial production. Astaxanthin produced by other sources such as chemical synthesis or fermentation are often produced in the cis configuration, which has been shown to have lower bioactivity. Additionally, some sources of astaxanthin, such as shrimp, may denature or degrade when exposed to high temperatures, which can result in a loss of bioactivity. Producing natural astaxanthin through the cultivation of H. pluvialis is presently a demanding and time-consuming task, which incurs high expenses and restricts the cost-effective industrial production of this valuable substance. The production of astaxanthin occurs through two distinct pathways, namely the cytosolic mevalonate pathway and the chloroplast methylerythritol phosphate (MEP) pathway. The latest advancements in enhancing product quality and extracting techniques at a reasonable cost are emphasized in this review. The comparative of specific extraction processes of H. pluvialis biological astaxanthin production that may be applied to large-scale industries were assessed. The article covers a contemporary approach to optimizing microalgae culture for increased astaxanthin content, as well as obtaining preliminary data on the sustainability of astaxanthin production and astaxanthin marketing information., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The economical way of Haematococcus pluvialis farming is to simultaneously achieve biomass, astaxanthin and lipid using less expensive chemicals. This paper explores the role of exogenous arginine in promoting growth and astaxanthin accumulation under stressful conditions. The application of arginine exerts a synergic effect on biomass, astaxanthin and lipid by improving carbon utilization, activating the arginine pathway and regulating carotenoid and lipid-related genes. Genes related to arginine catabolism, such as ADC, OCT, ASS1, NOS, and OAT, were up-regulated at both the cultivation and astaxanthin induction stages, signifying their importance in both growth and astaxanthin synthesis. Furthermore, transcriptome analysis revealed that arginine up-regulated transcription levels of genes involved carbon fixing, lipid biosynthesis, pyruvate metabolism, carotenoid, tricarboxylic acid cycle, and arginine and proline metabolism. The results provide a significant mechanism and applicability of using exogenous arginine and high light to stimulate bioproducts from Haematococcus pluvialis., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Fucoxanthin, a natural carotenoid that has substantial pharmaceutical value due to its anticancer, antioxidant, antiobesity, and antidiabetic properties, is biosynthesized from glyceraldehyde-3-phosphate (G3P) via a series of enzymatic reactions. However, our understanding of the transcriptional mechanisms involved in fucoxanthin biosynthesis remains limited. Using reverse genetics, the med8 mutant was identified based on its phenotype of reduced fucoxanthin content, and the biological functions of MED8 in fucoxanthin synthesis were characterized using approaches such as gene expression, protein subcellular localization, protein-protein interaction and chromatin immunoprecipitation assay. Gene-editing mutants of MED8 exhibited decreased fucoxanthin content as well as reduced expression levels of six key genes involved in fucoxanthin synthesis, namely DXS, PSY1, ZDS-like, CRTISO5, ZEP1, and ZEP3, when compared to the wild-type (WT) strain. Furthermore, we showed that MED8 interacts with HSF3, and genetic analysis revealed their shared involvement in the genetic pathway governing fucoxanthin synthesis. Additionally, HSF3 was required for MED8 association with the promoters of the six fucoxanthin synthesis genes. In conclusion, MED8 and HSF3 are involved in fucoxanthin synthesis by modulating the expression of the fucoxanthin synthesis genes. Our results increase the understanding of the molecular regulation mechanisms underlying fucoxanthin synthesis in the diatom P. tricornutum., (© 2023 The Authors New Phytologist © 2023 New Phytologist Foundation.)

Astaxanthin is a red-colored secondary metabolite with excellent antioxidant properties, typically finds application as foods, feed, cosmetics, nutraceuticals, and medications. Astaxanthin is usually produced synthetically using chemicals and costs less as compared to the natural astaxanthin obtained from fish, shrimps, and microorganisms. Over the decades, astaxanthin has been naturally synthesized from Haematococcus pluvialis in commercial scales and remains exceptional, attributed to its higher bioactive properties as compared to synthetic astaxanthin. However, the production cost of algal astaxanthin is still high due to several bottlenecks prevailing in the upstream and downstream processes. To that end, the present study intends to review the recent trends and advancements in astaxanthin production from microalgae. The structure of astaxanthin, sources, production strategies of microalgal astaxanthin, and factors influencing the synthesis of microalgal astaxanthin were discussed while detailing the pathway involved in astaxanthin biosynthesis. The study also discusses the relevant downstream process used in commercial scales and details the applications of astaxanthin in various health related issues., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Ltd.)

This work explores astaxanthin (AXT), a valuable xanthophyll ketocarotenoid pigment with significant health benefits and diverse applications across various industries. It discusses the prevalence of synthetic AXT, and the development of natural-based alternatives derived from microorganisms such as microalgae, bacteria, and yeast. The chapter examines the potential of microbial AXT production, highlighting the advantages and challenges associated with natural AXT. Key microorganisms like Haematococcus pluvialis, Paracoccus carotinifaciens, and Phaffia rhodozyma are emphasized for their role in commercially producing this valuable ketocarotenoid. The narrative covers the complexities and opportunities in microbial AXT production, from cell structure implications to downstream processing strategies. Additionally, the chapter addresses current applications, commercialization trends, and market dynamics of natural microbial AXT, emphasizing the importance of cost-effective production, regulatory compliance, and technological advancements to reduce the market cost of the final product. As demand for natural microbial-based AXT rises, this chapter envisions a future where research, innovation, and collaboration drive sustainable and competitive microbial AXT production, fostering growth in this dynamic market., Competing Interests: Conflicts of interest There are no conflicts to declare., (Copyright © 2024. Published by Elsevier Inc.)