Isolation of Mycosporine-like Amino Acids from Red Macroalgae and a Marine Lichen by High-Performance Countercurrent Chromatography: A Strategy to Obtain Biological UV-Filters.

Marine organisms have gained considerable biotechnological interest in recent years due to their wide variety of bioactive compounds with potential applications. Mycosporine-like amino acids (MAAs) are UV-absorbing secondary metabolites with antioxidant and photoprotective capacity, mainly found in organisms living under stress conditions (e.g., cyanobacteria, red algae, or lichens). In this work, five MAAs were isolated from two red macroalgae (Pyropia columbina and Gelidium corneum) and one marine lichen (Lichina pygmaea) by high-performance countercurrent chromatography (HPCCC). The selected biphasic solvent system consisted of ethanol, acetonitrile, saturated ammonium sulphate solution, and water (1:1:0.5:1; v:v:v:v). The HPCCC process for P. columbina and G. corneum consisted of eight separation cycles (1 g and 200 mg of extract per cycle, respectively), whereas three cycles were performed for of L. pygmaea (1.2 g extract per cycle). The separation process resulted in fractions enriched with palythine (2.3 mg), asterina-330 (3.3 mg), shinorine (14.8 mg), porphyra-334 (203.5 mg) and mycosporine-serinol (46.6 mg), which were subsequently desalted by using precipitation with methanol and permeation on a Sephadex G-10 column. Target molecules were identified by HPLC, MS, and NMR.

Raman Microspectroscopic Analysis of Selenium Bioaccumulation by Green Alga Chlorella vulgaris.

Selenium (Se) is an element with many commercial applications as well as an essential micronutrient. Dietary Se has antioxidant properties and it is known to play a role in cancer prevention. However, the general population often suffers from Se deficiency. Green algae, such as Chlorella vulgaris, cultivated in Se-enriched environment may be used as a food supplement to provide adequate levels of Se. We used Raman microspectroscopy (RS) for fast, reliable, and non-destructive measurement of Se concentration in living algal cells. We employed inductively coupled plasma-mass spectrometry as a reference method to RS and we found a substantial correlation between the Raman signal intensity at 252 cm-1 and total Se concentration in the studied cells. We used RS to assess the uptake of Se by living and inactivated algae and demonstrated the necessity of active cellular transport for Se accumulation. Additionally, we observed the intracellular Se being transformed into an insoluble elemental form, which we further supported by the energy-dispersive X-ray spectroscopy imaging.

Selenium Incorporation to Amino Acids in Chlorella Cultures Grown in Phototrophic and Heterotrophic Regimes.

Microalgae accumulate bioavailable selenium-containing amino acids (Se-AAs), and these are useful as a food supplement. While this accumulation has been studied in phototrophic algal cultures, little data exists for heterotrophic cultures. We have determined the Se-AAs content, selenium/sulfur (Se/S) substitution rates, and overall Se accumulation balance in photo- and heterotrophic Chlorella cultures. Laboratory trials revealed that heterotrophic cultures tolerate Se doses ∼8-fold higher compared to phototrophic cultures, resulting in a ∼2-3-fold higher Se-AAs content. In large-scale experiments, both cultivation regimes provided comparable Se-AAs content. Outdoor phototrophic cultures accumulated up to 400 µg g-1 of total Se-AAs and exhibited a high level of Se/S substitution (5-10%) with 30-60% organic/total Se embedded in the biomass. A slightly higher content of Se-AAs and ratio of Se/S substitution was obtained for a heterotrophic culture in pilot-scale fermentors. The data presented here shows that heterotrophic Chlorella cultures provide an alternative for Se-enriched biomass production and provides information on Se-AAs content and speciation in different cultivation regimes.

In vitro bioaccessibility of selenoamino acids from selenium (Se)-enriched Chlorella vulgaris biomass in comparison to selenized yeast; a Se-enriched food supplement; and Se-rich foods.

Selenium (Se) is an indispensable microelement in our diet and health issues resulting from deficiencies are well documented. Se-containing food supplements are available on the market including Se-enriched Chlorella vulgaris (Se-Chlorella) which accumulates Se in the form of Se-amino acids (Se-AAs). Despite its popular uses, data about the bioaccessibility of Se-AAs from Se-Chlorella are completely missing. In the present study, gastrointestinal digestion times were optimized and the in vitro bioaccessibility of Se-AAs in Se-Chlorella, Se-yeast, a commercially available Se-enriched food supplement (Se-supplement) and Se rich foods (Se-foods) were compared. Higher bioaccessibility was found in Se-Chlorella (∼49%) as compared to Se-yeast (∼21%), Se-supplement (∼32%) and Se-foods. The methods used in production of Se-Chlorella biomass were also investigated. We found that disintegration increased bioaccessibility whereas the drying process had no effect. Similarly, temperature treatment by microwave oven also increased bioaccessibility whereas boiling water did not.

Antimicrobial, Cytotoxic, Anti-Inflammatory, and Antioxidant Activity of Culinary Processed Shiitake Medicinal Mushroom (Lentinus edodes, Agaricomycetes) and Its Major Sulfur Sensory-Active Compound-Lenthionine.

The antimicrobial, cytotoxic, anti-inflammatory, and antioxidant properties of aqueous extracts of raw and culinary processed shiitake mushrooms were evaluated and compared with those of lenthionine (1,2,3,5,6-penta-thiepane), the principal aroma-bearing substance of the shiitake medicinal mushroom (Lentinus edodes). Antimicrobial activity was tested using a panel of 4 strains of bacteria, 2 yeasts, and 2 fungi. Cytotoxic properties were evaluated against 3 cell lines (HepG2, HeLa, PaTu), whereas the anti-inflammatory activity of tested samples was assayed based on their ability to attenuate the secretion of the cytokine tumor necrosis factor-α. Antioxidant activity was measured using in vitro DPPH and ABTS assays. It was found that lenthionine possesses significant antimicrobial properties; it is remarkably effective in inhibiting the growth of yeasts and fungi (minimum inhibitory concentration, 2-8 µg/mL) and thus is comparable to standard antifungal agents. Lenthionine is also able to decrease significantly the production of tumor necrosis factor-a and thus could be at least partly responsible for the observed anti-inflammatory effect of shiitake. On the other hand, lenthionine does not seem to contribute significantly to the well-known anticancer and antioxidant effects of the mushroom.

Separation of Aeruginosin-865 from Cultivated Soil Cyanobacterium (Nostoc sp.) by Centrifugal Partition Chromatography combined with Gel Permeation Chromatography.

Aeruginosin-865 was isolated from cultivated soil cyanobacteria using a combination of centrifugal partition chromatography (CPC) and gel permeation chromatography. The solubility of Aer-865 in different solvents was evaluated using the conductor-like screening model for real solvents (COSMO-RS). The CPC separation was performed in descending mode with a biphasic solvent system composed of water-n-BuOH-acetic acid (5:4:1, v/v/v). The upper phase was used as a stationary phase, whereas the lower phase was employed as a mobile phase at a flow rate of 10 mL/min. The revolution speed and temperature of the separation column were 1700 rpm and 25 degrees C, respectively. Preparative CPC separation followed by gel permeation chromatography was performed on 50 mg of crude extract yielding Aer-865 (3.5 mg), with a purity over 95% as determined by HPLC. The chemical identity of the isolated compound was confirmed by comparing its spectroscopic data (UV, HRESI-MS, HRESI-MS/MS) with those of an authentic standard and data available in the literature.