Maximizing Polysaccharides and Phycoerythrin in Porphyridium purpureum via the Addition of Exogenous Compounds: A Response-Surface-Methodology Approach.

Phycoerythrin and polysaccharides have significant commercial value in medicine, cosmetics, and food industries due to their excellent bioactive functions. To maximize the production of biomass, phycoerythrin, and polysaccharides in Porphyridium purpureum, culture media were supplemented with calcium gluconate (CG), magnesium gluconate (MG) and polypeptides (BT), and their optimal amounts were determined using the response surface methodology (RSM) based on three single-factor experiments. The optimal concentrations of CG, MG, and BT were determined to be 4, 12, and 2 g L-1, respectively. The RSM-based models indicated that biomass and phycoerythrin production were significantly affected only by MG and BT, respectively. However, polysaccharide production was significantly affected by the interactions between CG and BT and those between MG and BT, with no significant effect from BT alone. Using the optimized culture conditions, the maximum biomass (5.97 g L-1), phycoerythrin (102.95 mg L-1), and polysaccharide (1.42 g L-1) concentrations met and even surpassed the model-predicted maximums. After optimization, biomass, phycoerythrin, and polysaccharides concentrations increased by 132.3%, 27.97%, and 136.67%, respectively, compared to the control. Overall, this study establishes a strong foundation for the highly efficient production of phycoerythrin and polysaccharides using P. purpureum.

Modulation of the metabolite content of the unicellular rhodophyte Porphyridium purpureum using a 2-stage cultivation approach and chemical stressors.

There have been growing interests in microalgal biotechnology for the biorefining of bioactive compounds such as carotenoid pigments, ω-3 fatty acids, antioxidants or antimicrobials for sectoral applications in the pharmacology, nutraceutical and cosmetic fields. This study focused on the unicellular marine rhodophyte Porphyridium purpureum CCAP 1380/1 A, which was cultivated via a two-stage batch growth mode for 10 days using hydrogen peroxide (H2O2), the phytohormone methyl jasmonate (MJ) and three plant extracts (Passiflora incarnata, Panax ginseng and Valeriana officinalis). The microalgal biomass was then analysed for its protein, phycoerythtin, carbohydrate and pigment composition together with its pigment content and antioxidant activity. Of note, MJ increased the protein and phycoerythtin content (up to 225 µg BSA eq./mg DW and 15 mg/ml, respectively) while both the MJ and H2O2 treatments increased carotenoid pigment yields (ß-carotene and zeaxanthin, up to 5 and 4 mg/g, respectively). Carbohydrates were enhanced ∼10 fold by the Valeriana officinalis treatment (up 192 µg starch eq./mg). Overall, neutral lipids and antioxidants were mostly negatively affected by the plant extracts. The greatest antioxidant activity registered was obtained with the H2O2 treatment (15 µmol Trolox eq./g DW with TEAC assay). P. purpureum contains multiple valuable compounds of commercial interest. These results indicate that they can be favorably modulated using specific cultivation regimes and chemical enhancers, thereby facilitating the exploitation of the biomass by applying a suitable co-refinery pipeline.

Therapeutic Potential of Microalgae-Derived Bioactive Metabolites Is Influenced by Different Large-Scale Culture Strategies.

Microalgae have been identified as one of the most promising sources of novel bioactive compounds for biomedical applications, the food industry, and cosmetics. In the last decade, several biotechnological developments have facilitated the identification of a growing number of compounds as well as the study of optimal microalgae culture conditions for the production of biomass enriched in specific molecules of interest. In this study, two common commercial marine microalgae (Nannochloropsis oculata and Porphyridium purpureum) were cultured in standard and nutrient-stressed conditions and the obtained biomass extracts were assessed for their potential to inhibit cancer cell proliferation and migration as well as their antioxidant activity. Results from viability in 2D and 3D cancer cell models showed an enhancement of the antitumour activity of P. purpureum in the 3D model compared to 2D, together with a greater capacity to reduce the migration capacity of cancer cells with the biomass from nutrient-stressed conditions, whereas the antioxidant activity of N. oculata decreased when exposed to nutrient-stressed conditions. To date, this is one of the few studies that proves that controlled changes in large-scale culturing conditions such as nutrient depletion have a relevant impact in the bioactivity of the biomass on cancer cells.

Photobiological Effects on Biochemical Composition in Porphyridium cruentum (Rhodophyta) with a Biotechnological Application.

This study describes the relation of photosynthetic capacity, growth and biochemical compounds in the microalgae Porphyridium cruentum under saturated irradiance (200 µmol m-2  s-1 ) by white light (WL) and low-pressure sodium vapor lamps (SOX lamps-control) and supplemented by fluorescent lamps (FLs) with different light qualities (blue: λmax = 440 nm; green: λmax = 560 nm; and red: λmax = 660 nm). The maximum photosynthetic efficiency (Fv / Fm ) showed a positive correlation with the light quality by saturating light SOX in mixture with stimulating blue light than the white light (WL) at the harvest day (10 days). The production, that is maximal electron transport rate (ETRmax ), and energy dissipation, that is maximal nonphotochemical quenching (NPQmax ), had the same pattern throughout the time (3-6 days) being the values higher under white light (WL) compared with SOX and SOX plus supplemented different light qualities. Total protein levels increased significantly in the presence of SOX light, while phycoerythrin (B-PE) showed significant differences under SOX+ blue light. Arachidonic acid (ARA) was higher under SOX and SOX plus supplemented different light qualities than that under WL, whereas eicosapentaenoic acid (EPA) was the reverse. The high photomorphogenic potential by SOX light shows promising application for microalgal biotechnology.

Using a B-Phycoerythrin Extract as a Natural Colorant: Application in Milk-Based Products.

Nowadays, there is a growing interest in finding new coloring molecules of natural origin that can increase and diversify the offer of natural food dyes already present in the market. In the present work, a B-phycoerythrin extract from the microalgae Porphyridium cruentum was tested as a food colorant in milk-based products. Using spectroscopy and colorimetry, the extract was characterized and gave evidence of good properties and good stability in the pH range between 4.0 and 9.0. Coloring studies were conducted to demonstrate that samples carrying the pink extract could be used for simulating the pink color of marketed milk-based products. The staining factors, representing the amount of pink protein to be added to reproduce the color of strawberry commercial products, ranged between 1.6 mg/L and 49.5 mg/L, being sufficiently low in all samples. Additionally, color stability during a short period of cold storage was studied: it demonstrated that the three tested types of dairy products remained stable throughout the 11-day analysis period with no significant changes. These results prove the potential of the B-phycoerythrin extract as a natural colorant and alternative ingredient to synthetic coloring molecules.

Rheological characterization of Porphyridium sordidum and Porphyridium purpureum exopolysaccharides.

Porphyridium exopolysaccharides (EPSs), which contain sulfate and methyl groups, have a similar potential for use in multiple industrial applications as macroalgae counterparts but lack detailed characterization. For this reason, we produced 0.21 g L-1 of P. sordidum EPS and 0.17 g L-1P. purpureum EPS, followed by a thorough rheological characterization in respect to their differences in monomer composition, sulfate concentrations and methyl patterns. Furthermore, the effect of NaCl and CaCl2 was evaluated, and the effect of high salinity media on the rheological properties of the biopolymers was analyzed. Both Porphyridium EPSs show a remarkable stability at high temperature and under the effect of mono- and divalent cations, and high salinity cultivation medium, which was evidenced by the rheological properties of the EPS. This feature is not displayed by many carbohydrate polymers, making it possible to enrich current applications in which EPS are used.

Effects of Different Environmental Factors on the Growth and Bioactive Substance Accumulation of Porphyridium purpureum.

Genus Porphyridium is a primitive single-celled red algae widely distributed in seawater, freshwater, and moist soil. It can synthesize bioactive substances such as phycoerythrin, extracellular polysaccharides and polyunsaturated fatty acids during the growth process. In this paper, the culture and bioactive substance yield of Porphyridium purpureum were studied by setting salinity, nitrogen-to-phosphorus ratio, and pH at different gradient levels. The results showed that the optimal conditions for the growth of P. purpureum were salinity 34 ppt, nitrogen-to-phosphorus ratio 169:1, and pH 8; the optimal conditions for obtaining the polysaccharides were salinity 17 ppt, nitrogen-to-phosphorus ratio 14:1, and pH 8; the optimal conditions for obtaining phycoerythrin were salinity 17 ppt, nitrogen-to-phosphorus ratio 68:1, and pH 8; the optimal conditions for obtaining the lipids were salinity 34 ppt, nitrogen-to-phosphorus ratio 1:1, and pH 8. In actual production applications, culture conditions should be set according to different product accumulation purposes in order to achieve the optimal production efficiency.

Lipid and unsaturated fatty acid productions from three microalgae using nitrate and light-emitting diodes with complementary LED wavelength in a two-phase culture system.

In this study, Pavlova lutheri, Chlorella vulgaris, and Porphyridium cruentum were cultured using modified F/2 media in a 1 L flask culture. Various nitrate concentrations were tested to determine an optimal nitrate concentration for algal growth. Subsequently, the effect of light emitted at a specific wavelength on biomass and lipid production by three microalgae was evaluated using various wavelengths of light-emitting diodes (LED). Biomass production by P. lutheri, C. vulgaris, and P. cruentum were the highest with blue, red, and green LED wavelength with 1.09 g dcw/L, 1.23 g dcw/L, and 1.28 g dcw/L on day 14, respectively. Biomass production was highest at the complementary LED wavelength to the color of microalgae. Lipid production by P. lutheri, C. vulgaris, and P. cruentum were the highest with yellow, green, and red LEDs' wavelength, respectively. Eicosapentaenoic acid production by P. lutheri, C. vulgaris, and P. cruentum was 10.35%, 10.14%, and 14.61%, and those of docosahexaenoic acid were 6.09%, 8.95%, and 11.29%, respectively.

Impact of different sequences of mechanical and thermal processing on the rheological properties of Porphyridium cruentum and Chlorella vulgaris as functional food ingredients.

Microalgae are a promising and sustainable source for enhancing the nutritional value of food products. Moreover, incorporation of the total biomass might contribute to the structural properties of the enriched food product. Our previous study demonstrated the potential of Porphyridium cruentum and Chlorella vulgaris as multifunctional food ingredients, as they displayed interesting rheological properties after applying a specific combination of mechanical and thermal processing. The aim of the current study was to investigate the impact of a different sequence of high pressure homogenization (HPH) and thermal processing on the thickening and gelling potential of these microalgal biomasses in aqueous suspensions. Thermal processing largely increased the gel strength and viscosity of both microalgae, which was ascribed to larger and stronger aggregates as a result of partial solubilization of polymers, while subsequent HPH generally reduced the rheological properties. Interestingly, large amounts of intact cells were still observed for both microalgae when HPH was performed after a thermal treatment, irrespective of the applied homogenization pressure, implying that cell disruption was hindered by the preceding thermal treatment. Although thermal processing was regarded as the most effective processing technique to obtain increased rheological properties, the combination with a preceding HPH treatment should still be considered when cell disruption is desired, for instance to increase the bioavailability of intracellular components. Finally, biomass of P. cruentum showed the largest potential for use as a structuring agent, as the gel strength and viscosity in thermally treated suspensions of this microalga were about 10 times higher than for C. vulgaris.

Effects of nitrogen and phosphorous stress on the formation of high value LC-PUFAs in Porphyridium cruentum.

This study systematically examined the effect of nitrogen and phosphorous stress on the formation of linoleic acid (LA), arachidonic acid (ARA), and eicosapentaenoic acid (EPA) in Porphyridium cruentum gy-h56. P. cruentum was cultivated in six different media conferring different conditions of nitrogen (N) sufficiency/deprivation and phosphorous (P) sufficiency/limitation/deprivation. Over a 16-day cultivation process, the dry-weight content, proportion of total fatty acids (TFAs), and the concentration in the medium of linoleic acid (LA) were greatly improved by a maximum of 2.5-, 1.6-, and 1.1-fold, respectively, under conditions of N or P deprivation compared with N and P sufficiency. In contrast, levels of EPA or ARA were not enhanced under N or P stress conditions. Additionally, the results showed that N deprivation weakened the impact of P deficiency on the content and proportions of LA and EPA, while P deprivation enhanced the impact of N starvation on the content and proportions of LA and EPA. The conditions of N sufficiency and P deprivation (N+P-) were the optimal conditions for the production of LA, while the optimal conditions for EPA, ARA, and TFAs production were N sufficiency and P limitation (N+P-lim). This study suggests the potential application of combining N removal from saline wastewater with the production of LA, ARA, EPA, and biodiesel.

Salt Effect on the Antioxidant Activity of Red Microalgal Sulfated Polysaccharides in Soy-Bean Formula.

Sulfated polysaccharides produced by microalgae, which are known to exhibit various biological activities, may potentially serve as natural antioxidant sources. To date, only a few studies have examined the antioxidant bioactivity of red microalgal polysaccharides. In this research, the effect of different salts on the antioxidant activities of two red microalgal sulfated polysaccharides derived from Porphyridium sp. and Porphyridium aerugineum were studied in a soy bean-based infant milk formula. Salt composition and concentration were both shown to affect the polysaccharides' antioxidant activity. It can be postulated that the salt ions intefer with the polysaccharide chains' interactions and alter their structure, leading to a new three-dimensional structure that better exposes antiooxidant sites in comparison to the polysaccharide without salt supplement. Among the cations that were studied, Ca(2+) had the strongest enhancement effect on antioxidant activities of both polysaccharides. Understanding the effect of salts on polysaccharides' stucture, in addition to furthering knowledge on polysaccharide bioactivities, may also shed light on the position of the antioxidant active sites.

UPLC-MSE profiling of Phytoplankton metabolites: application to the identification of pigments and structural analysis of metabolites in Porphyridium purpureum.

A fast and high-resolution UPLC-MSE analysis was used to identify phytoplankton pigments in an ethanol extract of Porphyridium purpureum (Pp) devoid of phycobiliproteins. In a first step, 22 standard pigments were analyzed by UPLC-MSE to build a database including retention time and accurate masses of parent and fragment ions. Using this database, seven pigments or derivatives previously reported in Pp were unequivocally identified: ß,ß-carotene, chlorophyll a, zeaxanthin, chlorophyllide a, pheophorbide a, pheophytin a, and cryptoxanthin. Minor amounts of Divinyl chlorophyll a, a chemotaxonomic pigment marker for prochlorophytes, were also unequivocally identified using the database. Additional analysis of ionization and fragmentation patterns indicated the presence of ions that could correspond to hydroxylated derivatives of chlorophyll a and pheophytin a, produced during the ethanolic extraction, as well as previously described galactosyldiacylglycerols, the thylakoid coenzyme plastoquinone, and gracilamide B, a molecule previously reported in the red seaweed Gracillaria asiatica. These data point to UPLC-MSE as an efficient technique to identify phytoplankton pigments for which standards are available, and demonstrate its major interest as a complementary method for the structural elucidation of ionizable marine molecules.

Influence of sulphate on the composition and antibacterial and antiviral properties of the exopolysaccharide from Porphyridium cruentum.

AIMS: The influence of two culture media and three different concentrations of sulphate in the medium on the growth of two strains of Porphyridium cruentum and on the production, composition and viscoelastic characteristics, and antimicrobial properties of the sulphated exopolysaccharide (EPS) were studied. MAIN METHODS: A Bohlin C50 rheometer was used to evaluate the viscosity and elasticity of the EPS solutions. HSV virus, types 1 and 2, Vaccinia virus and Vesicular stomatitis virus were used along with two Gram-negative (Escherichia coli and Salmonella enteritidis) and one Gram-positive (Staphylococcus aureus) bacteria, for testing the antimicrobial activity of EPS. KEY-FINDINGS: The growth of microalgae was higher in NTIP medium and the production of EPS was enhanced by sulphate 21mM. The protein content of the EPS was enhanced by the addition of sulphate 52mM and 104mM; this concentration also induced an increase in sulphate content of the EPS. However, neither the contents of EPS in carbohydrates and uronic acids were affected by the culture medium supplementation in sulphate. In general, the EPS from the Spanish strain presented a higher antiviral activity than the EPS from the Israeli strain. All EPS extracts revealed a strong activity against V. stomatitis virus, higher than the activity of all chemical compounds tested. The EPS from the Israeli strain also presented antibacterial activity against S. enteritidis. SIGNIFICANCE: Enrichment of the culture medium with sulphate improved protein and sulphate content of EPS. EPS extracts presented a relevant activity against V. stomatitis virus and S. enteritidis bacterium.

Uptake of uranyl ions from uranium ores and sludges by means of Spirulina platensis, Porphyridium cruentum and Nostok linckia alga.

In this paper was studied the uranyl ions biosorption on three types of alga: Nostok linckia, Porphyridium cruentum and Spirulina platensis. These ions were supplied either from a pure solution of uranyl nitrate, or after leaching process of uranium ore, or from the sludge resulting in the output of pure UO(2) technology. It was investigated the retention degree versus contact time and afterwards the Langmuir and Freundlich biosorption isotherms of uranyl ions on the three alga types. The retention of UO(2)(2+) ions on alga was proved through FTIR spectra plotted before and after biosorption processes. From the experimental data it was found that regardless of origin of uranyl ions, the retention degree on alga decreased in the series. Spirulina platensis > Porphyridium cruentum ≥ Nostok linckia.

Overcoming recalcitrance in Porphyridium aerugineum Geitler employing encapsulation-dehydration cryopreservation methods.

Cultures of the recalcitrant microalga Porphyridium aerugineum were cryopreserved. A two-step, uncontrolled rapid freezing protocol, using methanol as cryoprotectant resulted in 23.8 percent viable cells. Cultures in the exponential growth phase, grown under low light intensity to prevent vacuole formation in cells, cryopreserved using a passive freezer, showed 22.4 percent viability. This value was enhanced to 31.5 percent when a controlled-rate freezer was employed. Optimized cultures in the exponential growth phase, cultivated in medium supplemented or not with vitamin B12, were then tested for freezing using the encapsulation-dehydration protocol. High cell loss was observed early during the sorbitol dehydration steps, but 63.6 percent of the remaining encapsulated cells were viable after thawing. This study confirmed the potential of encapsulation-dehydration as a method allowing to improve the low viability obtained with two-step freezing protocols. It also showed the importance of monitoring the response of algal cells to bead osmotic and evaporative dehydration pretreatments before freezing.

Effect of dietary administration of Porphyridium cruentum on the respiratory burst activity of sole, Solea senegalensis (Kaup), phagocytes.

The stimulatory effect of the red microalga Porphyridium cruentum on respiratory burst activity of sole phagocytes was evaluated in vivo. Oral administration of a diet supplemented with lyophilized P. cruentum cells (10 g kg(-1)) stimulated respiratory burst activity after 4 weeks feeding in sole vaccinated with Photobacterium damselae subsp. piscicida bacterin.

Effect of drying on the biological activities of a red microalgal polysaccharide.

The red microalga Porphyridium sp. produces a polysaccharide exhibiting a variety of biological activities with potential for medical and cosmetic uses. For this reason, it is important that the drying process, which is the end point of production, should not destroy the natural characteristics of the material. The objective of this study was to evaluate the effect of drying at temperatures ranging from 40 to 140 degrees C on the bioactivities of the polysaccharide. Drying the polysaccharide at temperatures above 90 degrees C caused a significant decline in its biological activities (antiviral and anti-cell proliferation) and reduced elasticity, viscosity, and intrinsic viscosity relative to lyophilized polysaccharide and to the starting product. The relationship between molecular weight and intrinsic viscosity indicated that the polysaccharide takes a rigid coil conformation, which stiffens as a result of drying. FTIR analysis revealed that drying caused both significant conformational alterations in the polymer chains and changes in the interaction between the polysaccharide and the glycoprotein to which it is noncovalently associated. Differential scanning calorimetry analysis of the water adsorbed on the charged groups of the polysaccharide showed that drying at higher temperatures increased the bound water content due to dissociation of the polymer chains. Thus, it is recommended that the polysaccharide be dried in a two-step process in which free water is removed by convection and bound freezing water is removed by lyphophilization.