Purificacao e caracterizacao do pigmento vermelho R-Ficoeritrina da macroalga marinha Solieria filiformis (Kutzing) P.W. Gabrielson

Abstract

R-Phycoerythrin (R-PE) is a red photosynthetic pigment (phycobiliprotein) present in Rhodophyta and it has wide application in biotechnology as a fluorescent probe in immunology, cell biology, and natural colorant in foods and cosmetics. Therefore, this study aimed to purify and to characterize partially the fluorescent photosynthetic pigment R-PE from red marine macroalgae Solieria filiformis, abundant in the Brazilian northeastern coast with high potential for cultivation in the sea. A purification scheme with protein precipitation (ammonium sulfate 90% saturation) followed by chromatography on matrices DEAE-Sephacel and Sephacryl S-300, produced a purified pigment that have three absorption peaks (495, 540 and 564 nm), one fluorescence emission peak (575 nm when excited at 495 nm) and a purity index of 4.5, enabling it’s application as a fluorescent marker in biotechnology. The purification process of the R-PE was a recovery of 12.78% and a yield of 0.17 mg/g of dry seaweed. Electrophoresis (SDS-PAGE) of R-PE showed one protein band in the absence of reducing agent and three protein bands with molecular weight of 18, 20, 37 kDa, regarding the α, β and γ subunits, in the presence of a reducing agent. R-PE showed 60% of α-helix in their secondary structure, while maintaining stability of the structure up to 80 °C. The spectroscopic properties of the R-PE showed stability up to 70 °C, however, a reduction of 72.1% and 98.5% in their absorption and fluorescence emission spectra were observed, respectively, only at 80 °C for 1 hour . The absorption capacity of R-PE, stored for 30 days, remained stable at temperatures of -20 °C and 4 °C, but at 25 °C, a significant reduction of the absorption peak was observed at 5 days (81.6%, 90.6% and 92.7% compared to the peaks 495, 540 and 564, respectively). It was observed reduction of fluorescence capability of 42.5%, 33.3% and 100% when stored for 5 days at 20 °C, 4 °C and 25 °C, respectively. In accord with these results was established a purification protocol for R-phycoerythrin from S. filiformis, with a high purity index and a high molecular weight made up of monomeric subunits (α, β and γ), with predominance of secondary structure in α-helix thermostable, spectroscopic properties (absorption and fluorescence) of high stability, and especially in appropriate storage conditions.