Search Results (35)

C-Phycocyanin (C-PC) is a photosynthetic pigment found in cyanobacteria, notably in Arthrospira species. The extraction of phycocyanobilin (PCB), the chromophore of C-PC, is a common approach to address the instability of C-PC under light, heat, and acidic conditions. Methanol is typically used for PCB extraction. However, its use poses challenges for industrial applications owing to the need for solvent removal, extensive purification, and safety validation. Therefore, this study proposes ethanol as an alternative to methanol, optimizing the ethanol extraction conditions through response surface methodology (RSM) using a central composite design (CCD) and techno-economic analysis. The parameters evaluated were the extraction temperature, time, and C-PC/solvent ratio. Optimal conditions—68.81 °C, 14.91 h, and a C-PC to solvent ratio of 1:95 (w/v)— yielded a predicted PCB yield of 29.18%, closely aligning with the actual value of 29.67 ± 1.33%. A techno-economic analysis for pilot-scale PCB production showed that optimized ethanol extraction could yield 147.13 kg/year with 506 batches, compared with 84.31 kg/year standard methanol extraction with 317 batches. Furthermore, it was evaluated to have a unit production cost of USD 1,413,588/kg, an internal rate of return (IRR) of 53.36%, and a payback time of 1.6 years with increased yields and reduced toxic solvent disposal costs. This study supports scalable PCB production with a natural blue pigment suitable for the food, beverage, and cosmetics industries. Full article

Damage to intestinal epithelial cells is present in obesity and other diseases because of inflammatory and oxidative processes. This damage compromises the gastrointestinal barrier, killing enterocytes, altering intestinal permeability, and eliciting abnormal immune responses that promote chronic inflammation. Recent evidence shows that spirulina is a potent natural agent with antioxidant and anti-inflammatory properties. Objectives: This study was conducted to evaluate the effect of spirulina aqueous extract (SPAE) on the alterations of the intestinal epithelium induced by lipid micelles (LMs) and/or inflammation induced by lipopolysaccharides (LPSs) in the Caco-2 cell line. Methods: In the current research, we assessed the protective actions of SPAE against cytotoxicity, oxidative stress, inflammation, and epithelial barrier perturbation by using an in vitro model, the intestinal Caco-2 cells, treated with LPSs and/or LMs. We also performed an in silico molecular docking analysis with spirulina’s bioactive compound, phycocyanobilin. Results: Our results showed that SPAE has no cytotoxic effect on Caco-2 cells. On the contrary, it improved cell viability and exhibited anti-inflammatory and antioxidant actions. SPAE also protected against endoplasmic reticulum stress and tight junction proteins, thus improving the epithelial barrier. The in silico study revealed a strong binding affinity of the phycocyanobilin compound with human SOD and NADPH oxidase and a good binding affinity towards COX-2 and iNOS. Conclusions: Taken together, these findings demonstrate the beneficial actions of SPAE on Caco-2 cells, suggesting it may be useful in preserving the epithelial intestinal barrier in human conditions involving oxidative stress and inflammation such as obesity. Full article

Phycocyanobilin was computationally investigated by means of DFT calculations in combination with implicit solvation starting from X-ray data. Different conformations and degrees of protonation were considered, and the acidity constants were estimated. The computed data suggest a syn-syn-syn conformation for the molecule, with the two carboxylic groups deprotonated under physiological conditions and weak acidic behavior of one of the pyrrolone heterocycles. The absorption transitions in the visible range were studied by means of TD-DFT calculations, focusing on the molecular orbitals involved. The frontier orbitals have a dominant role in the lowest energy absorption. Full article

Despite the many beneficial effects of phycocyanobilin (PCB) on human skin, its cosmetic applications have not been extensively investigated owing to its light and temperature sensitivity. This is the first report of PCB extract (SP) derived from marine Arthrospira maxima having skin anti-wrinkling effects associated with antioxidant efficacy and reduction of intracellular reactive oxygen species (ROS) production. We obtained 46.63 ± 1.72 mg PCB/g dry weight of A. maxima in SP through an ethanol extraction process. PCB extracts showed strong effects in increasing collagen synthesis and decreasing matrix metalloproteinases (MMP-1) production. Interestingly, skin anti-wrinkling effects of the PCB extracts were significantly increased by the addition of wheat bran extracts (WB), up to 20–30% of the effects of PCB at all concentrations, possibly due to the synergistic effects of soluble globulins and other active substances in WB. Moreover, the mixture of SP and WB (SPWB) greatly reduced cell cytotoxicity to approximately 15% of that of PCB. SPWB upregulated and downregulated the expression of collagen type I α1 (Col1A1) and MMP-1, respectively, although the downregulation of MMP-1 was higher than that of Col1A1. The optimal SPWB concentration for maintaining the highest skin anti-wrinkling effects was 0.5 mg/mL. We show that SPWB holds promise as a vegan cosmaceutical. Full article

Phycocyanobilin (PCB) is a small chromophore found in certain phycobiliproteins, such as phycocyanins (PCs) and allophycocyanins (APCs). PCB, along with other phycobilins (PBs) and intermediates such as biliverdin (BV) or phycoerythrobilin (PEB), is attracting increasing biotechnological interest due to its fluorescent and medicinal properties that allow potential applications in biomedicine and the food industry. This study aims to optimize PCB synthesis in Escherichia coli BL21 (DE3) and scale the process to a pre-industrial level. Parameters such as optimal temperature, inducer concentration, initial OD₆₀₀, and stirring speed were analyzed in shake flask cultures to maximize PCB production. The best results were obtained at a temperature of 28 °C, an IPTG concentration of 0.1 mM, an initial OD₆₀₀ of 0.5, and an orbital shaking speed of 260 rpm. Furthermore, the optimized protocol was scaled up into a 2 L bioreactor batch, achieving a maximum PCB concentration of 3.8 mg/L. Analysis of the results revealed that biosynthesis of exogenous PBs in Escherichia coli BL21 (DE3) is highly dependent on the metabolic burden of the host. Several scenarios, such as too rapid growth, high inducer concentration, or mechanical stress, can advance entry into the stationary phase. That progressively halts pigment synthesis, leading, in some cases, to its excretion into the growth media and ultimately triggering rapid degradation by the host. These conclusions provide a promising protocol for scalable PCB production and highlight the main biotechnological challenges to increase the yields of the process. Full article

Arthrospira platensis, commonly known as Spirulina, is a photosynthetic filamentous cyanobacterium (blue–green microalga) that has been utilized as a food source since ancient times. More recently, it has gained significant popularity as a dietary supplement due to its rich content of micro- and macro-nutrients. Of particular interest is a water soluble phycobiliprotein derived from Spirulina known as phycocyanin C (C-PC), which stands out as the most abundant protein in this cyanobacterium. C-PC is a fluorescent protein, with its chromophore represented by the tetrapyrrole molecule phycocyanobilin B (PCB-B). While C-PC is commonly employed in food for its coloring properties, it also serves as the molecular basis for numerous nutraceutical features associated with Spirulina. Indeed, the comprehensive C-PC, and to some extent, the isolated PCB-B, has been linked to various health-promoting effects. These benefits encompass conditions triggered by oxidative stress, inflammation, and other pathological conditions. The present review focuses on the bio-pharmacological properties of these molecules, positioning them as promising agents for potential new applications in the expanding nutraceutical market. Full article

Phycocyanobilin (PCB) is a natural blue tetrapyrrole chromophore that is found in phycocyanin and plays an essential role in photosynthesis. Due to PCB’s antioxidation, anti-inflammatory and anti-cancer properties, it has been utilized in the food, pharmaceutical and cosmetic industries. Currently, the extraction of PCB from Spirulina involves complex processes, which has led to increasing interest in the biosynthesis of PCB in Escherichia coli. However, the PCB titer remains low because of the poor activity of key enzymes and the insufficient precursor supply. Here, the synthesis of PCB was firstly improved by screening the optimal heme oxygenase (HO) from Thermosynechococcus elongatus BP-1(HO^T) and PCB: ferredoxin oxidoreductase from Synechocystis sp. PCC6803 (PcyA^S). In addition, based on a rational design and the infrared fluorescence method for high-throughput screening, the mutants of HO^T(F29W/K166D) and PcyA^S(D220G/H74M) with significantly higher activities were obtained. Furthermore, a DNA scaffold was applied in the assembly of HO^T and PcyA^S mutants to reduce the spatial barriers, and the heme supply was enhanced via the moderate overexpression of hemB and hemH, resulting in the highest PCB titer (184.20 mg/L) obtained in a 5 L fermenter. The strategies applied in this study lay the foundation for the industrial production of PCB and its heme derivatives. Full article

Phycocyanobilin (PCB) is the bioactive chromophore attached to Phycocyanin (PC) that is of special interest for nutraceutical and therapeutic applications. However, the production of PCB from the heterologous host Escherichia coli is still very low. To facilitate subsequent application of PCB, improving its production in microbial hosts is still a challenge to be solved. In this paper, a strategy involving fusion-expression of apo-proteins with signal peptides was adopted to improve PCB production in E. coli. First, we reconstructed the PCB biosynthesis pathway in E. coli and then optimized its culture media. Subsequently, one PC α (CpcA) subunit and one PC β (CpcB) subunit, which can capture free PCB, were introduced and increased the yield of PCB. Finally, CpcA was fused with seven signal peptides to generate recombinant proteins, among which, the signal peptide N20 fused with CpcA protein drastically improved PCB production in E. coli, providing a maximum flask output of 8.47 ± 0.18 mg/L. The results of this study demonstrate that PCB distribution and transporting manners in E. coli could affect the heterologous production efficiency. By fusing apo-proteins with signal peptides, the secretion of phycocyanin was refined and the production of PCB was successfully enhanced by 3.7-fold, compared with the starting strain (1.80 ± 0.12 mg/L). This work provided an alternative method for improving the production of PCB and other phycobilins. Full article

Due to their unique biochemical and spectroscopic properties, both heme and phycocyanobilin are widely applied in the medical and food industries. Synechocystis sp. PCC 6803 contains both heme and phycocyanin, and is capable of synthesizing phycocyanin using heme as a precursor. The aim of this study was to uncover viable metabolic targets in the porphyrin pathway from Synechocystis sp. PCC 6803 to promote the accumulation of heme and phycocyanin in the recombinant strains of microalgae. A total of 10 genes related to heme synthesis pathway derived from Synechococcus elongatus PCC 7942 and 12 genes related to endogenous heme synthesis were individually overexpressed in strain PCC 6803. The growth rate and pigment content (heme, phycocyanin, chlorophyll a and carotenoids) of 22 recombinant algal strains were characterized. Quantitative real-time PCR technology was used to investigate the molecular mechanisms underlying the changes in physiological indicators in the recombinant algal strains. Among the 22 mutant strains, the mutant overexpressing the haemoglobin gene (glbN) of strain PCC 6803 had the highest heme content, which was 2.5 times higher than the wild type; the mutant overexpressing the gene of strain PCC 7942 (hemF) had the highest phycocyanin content, which was 4.57 times higher than the wild type. Overall, the results suggest that genes in the porphyrin pathway could significantly affect the heme and phycocyanin content in strain PCC 6803. Our study provides novel crucial targets for promoting the accumulation of heme and phycocyanin in cyanobacteria. Full article

The utilization of natural blue pigments in foods is difficult as they are usually unstable during processing and the commonly applied pH. The current study focuses on natural blue pigment, possessing antioxidant properties, found in Arthrospira platensis (spirulina), and phycobiliproteins (PBP). These pigments are a complex of conjugated protein and non-protein components, known as phycocyanobilin. PBP has low stability during pasteurization (high-pressure or heat treatments), resulting in protein denaturation and color deterioration that limits the application. The phycocyanobilin pigment might also be liable to oxidation during pasteurization and storage, resulting in color deterioration. Yet, the instability of the pigment phycocyanobilin during the pasteurization process and storage conditions was never studied before, limiting the comprehensive understanding of the reasons for PBP instability. In this study, the stability of phycocyanobilin under high-pressure and high-temperature conditions was compared to the stability of phycobiliproteins. We revealed that phycobiliproteins have a higher color deterioration rate at 70–80 °C than at high-pressure (300–600 MPa) whereas phycocyanobilin remained stable during high-pressure and heat processing. During storage at pH 7, phycocyanobilin was oxidized, and the oxidation rate increased with increasing pH, while at lower pH phycocyanobilin had low solubility and resulted in aggregation. Full article

Phytochromes are biological red/far-red light sensors found in many organisms. The connection between photoconversion and the cellular output signal involves light-mediated global structural changes in the interaction between the photosensory module (PAS-GAF-PHY, PGP) and the C-terminal transmitter (output) module. We recently showed a direct correlation of chromophore deprotonation with pH-dependent conformational changes in the various domains of the prototypical phytochrome Cph1 PGP. These results suggested that the transient phycocyanobilin (PCB) chromophore deprotonation is closely associated with a higher protein mobility both in proximal and distal protein sites, implying a causal relationship that might be important for the global large-scale protein rearrangements. Here, we investigate the prototypical biliverdin (BV)-binding phytochrome Agp1. The structural changes at various positions in Agp1 PGP were investigated as a function of pH using picosecond time-resolved fluorescence anisotropy and site-directed fluorescence labeling of cysteine variants of Agp1 PGP. We show that the direct correlation of chromophore deprotonation with pH-dependent conformational changes does not occur in Agp1. Together with the absence of long-range effects between the PHY domain and chromophore pK_a, in contrast to the findings in Cph1, our results imply phytochrome species-specific correlations between transient chromophore deprotonation and intramolecular signal transduction. Full article

Phycobilisomes in cyanobacteria and red algae are large protein complexes that absorb light and transfer energy for use in photosynthesis. The light energy absorbed by chromophores binding to phycobiliproteins in the peripheral rods can be funneled to the core through chromophores at very high efficiency. The molecular mechanism of excitation energy transfer within a phycobilisome is an example of a higher and unique function in a living organism. However, the mechanism underlying the high efficiency remains unclear. Thus, this study was carried out as a step to resolve this mechanism theoretically. The three-dimensional structure of phycobilisomes containing the linker proteins of the red alga Porphyridium purpureum was determined by cryoelectron microscopy at 2.82 Å resolution in 2020. Using these data, the absorption wavelength of each β82 chromophore in the phycocyanin hexamer located next to the core was calculated using quantum chemical treatment, considering the electric effect from its surrounding phycocyanin proteins and two linker proteins. In addition to unaffected chromophores, chromophores that were redshifted and blueshifted under the electrical influence of the two linker proteins were found. Namely, the chromophore serving as the energy sink in the rod was determined. Full article

Despite the enormous environmental damage caused by plastic waste, it makes up over one-third of globally produced plastics. Polyethylene (PE) wastes have low recycling but high production rates. Towards the construction of ionic solar cells from PE, the present work describes the loading of a bioactive photoacid phycocyanobilin (PCB) dye from the pigment of Spirulina blue–green algae (as a natural resource) on low-density polyethylene (LDPE) plastic film. Dyeing was confirmed by X-ray photoelectron spectroscopy (XPS). Upon excitation of the Soret-band (400 nm), the photoluminescence (PL) spectra of PCB in neat solvents revealed two prominent emission peaks at 450–550 and 600–700 nm. The first band assigned to bilirubin-like (PCB_BR) species predominated the spectral profile in the highly rigid solvent glycerol and upon loading 0.45 % (w/w) of the dye on plastic. The photoluminescence excitation (PLE) spectra of PCB for the second region (Q-band) at 672 nm in the same solvents confirmed the ground state heterogenicity previously associated with the presence of PCB_A (neutral), PCB_B (cationic), and PCB_C (anionic) conformers. Time-resolved photoluminescence (TRPL) measurements induced via excitation of all PCB species at 510 nm in methanol revealed three-lifetime components with τ₁ = ~0.1 ns and τ₂ = ~2 ns associated with PCB_BR species and τ₃ = ~5 ns pertinent to the long-living photoproduct X*. Decay-associated spectra (DAS) analysis of the photoluminescence transient spectra of the final dyed films in the solid-state confirmed the improved generation of the long-living photoproduct as manifested in a significant increase in the PL intensity (~100-fold) and lifetime value (~90 ns) in the Q-region upon loading 6.92 % (w/w) of the dye on plastic. The photoproduct species were presumably assigned to the deprotonated PCB species, suggesting improved ionic mobility. The potential implementation of the PCB-sensitized PE solid wastes for the fabrication of ionic solar cells is discussed. Full article

Phycocyanin is an excellent antioxidant with anti-inflammatory effects on which recent studies are growing; however, its specific target remains unclear. Linear tetrapyrrole compounds such as bilirubin have been shown to lead to the induction of heme oxygenase 1 expression in vivo, thus achieving antioxidant and anti-inflammatory effects. Phycocyanin is bound internally with linear tetrapyrrole phycocyanobilin in a similar structure to bilirubin. We speculate that there is probably a way of inducing the expression of heme oxygenase 1, with which tissue oxidative stress and inflammation can be inhibited, thus inhibiting pulmonary fibrosis caused by oxidative damage and inflammation of lung. By optimizing the enzymatic hydrolysis process, phycocyanobilin-bound phycocyanin peptide were obtained, and its in vitro antioxidant, anti-inflammatory, and anti-pulmonary fibrosis activities were investigated. The results show that the phycocyanobilin peptide was able to alleviate oxidative and inflammatory damage in cells through the Keap1-Nrf2-HO-1 pathway, which in turn relieved pulmonary fibrosis symptoms. Full article

The effect of four different extraction methods on physicochemical characteristics and functionalities of chloro-phycocyanin (CP) was investigated. Swelling (S-CP), freezing and thawing (4FT-CP), ultrasonication with freezing and thawing (4FT+U-CP), and the high-pressure cell disruption (HPCD-CP) process affected CP differently, thus resulting in different levels of solubility, DPPH scavenging activity, ABTS scavenging activity, and reducing power. Among the four CPs, HPCD-CP had the highest CP content (15.3%), purity (1.66 ± 0.16), and ∆E value but the lowest ∆b value. The ζ potential of HPCD-CP (−38.8 mV) was the highest, but the average particle size of 4FT+U-CP (719.1 nm) was the highest. UV-Vis absorption spectra and fluorescence spectra illustrated that high-pressure cell disruption-assisted extraction had more profound impacts on the microenvironment of tetrapyrrole chromophores, the environment of aromatic amino acids, and the phycocyanobilin of CP. Furthermore, HPCD-CP and 4FT-CP showed higher solubility and antioxidant activities than S-CP, especially 4FT+U-CP. The results obtained in this study demonstrate that HPCD technology could obtain a food-grade C-phycocyanin product with higher CP concentration, purity, solubility, and antioxidant activity. Full article