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Separation and quantitation of phycobiliproteins using phytic acid in capillary electrophoresis with laser-induced fluorescence detection.
J Chromatogr A. 2002 Oct 04; 972(2):269-76.JC

Abstract

The similar electrophoretic mobilities and sizes of several of the phycobiliproteins, which are derived from the photosynthetic apparatus of cyanobacteria and eukaryotic algae, render their separation and quantitation a challenging problem. However, we have developed a suitable capillary electrophoresis (CE) method that employs a phytic acid-boric acid buffer and laser-induced fluorescence (LIF) detection with a single 594 nm He-Ne laser. This method takes advantage of the remarkably high quantum yields of these naturally fluorescent proteins, which can be attributed to their linear tetrapyrrole chromophores covalently bound to cysteinyl residues. As such, limits of detection of 1.18 x 10(-14), 5.26 x 10(-15), and 2.38 x 10(-15) mol/l were obtained for R-phycoerythrin, C-phycocyanin, and allophycocyanin proteins, respectively, with a linear dynamic range of eight orders of magnitude in each case. Unlike previously published CE-LIF methods, this work describes the separation of all three major classes of phycobiliproteins in under 5 min. Very good recoveries, ranging from 93.2 to 105.5%, were obtained for a standard mixture of the phycobiliproteins, based on seven-point calibration curves for both peak height and peak area. It is believed that this development will prove useful for the determination of phycobiliprotein content in naturally occurring cyanobacteria populations, thus providing a useful tool for understanding biological and chemical oceanographic processes.

Authors

No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, Non-P.H.S.

Language

eng

PubMed ID

12416885

Citation

Viskari, Pertti J., and Christa L. Colyer. "Separation and Quantitation of Phycobiliproteins Using Phytic Acid in Capillary Electrophoresis With Laser-induced Fluorescence Detection." Journal of Chromatography. A, vol. 972, no. 2, 2002, pp. 269-76.
Viskari PJ, Colyer CL. Separation and quantitation of phycobiliproteins using phytic acid in capillary electrophoresis with laser-induced fluorescence detection. J Chromatogr A. 2002;972(2):269-76.
Viskari, P. J., & Colyer, C. L. (2002). Separation and quantitation of phycobiliproteins using phytic acid in capillary electrophoresis with laser-induced fluorescence detection. Journal of Chromatography. A, 972(2), 269-76.
Viskari PJ, Colyer CL. Separation and Quantitation of Phycobiliproteins Using Phytic Acid in Capillary Electrophoresis With Laser-induced Fluorescence Detection. J Chromatogr A. 2002 Oct 4;972(2):269-76. PubMed PMID: 12416885.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Separation and quantitation of phycobiliproteins using phytic acid in capillary electrophoresis with laser-induced fluorescence detection. AU - Viskari,Pertti J, AU - Colyer,Christa L, PY - 2002/11/6/pubmed PY - 2003/3/28/medline PY - 2002/11/6/entrez SP - 269 EP - 76 JF - Journal of chromatography. A JO - J Chromatogr A VL - 972 IS - 2 N2 - The similar electrophoretic mobilities and sizes of several of the phycobiliproteins, which are derived from the photosynthetic apparatus of cyanobacteria and eukaryotic algae, render their separation and quantitation a challenging problem. However, we have developed a suitable capillary electrophoresis (CE) method that employs a phytic acid-boric acid buffer and laser-induced fluorescence (LIF) detection with a single 594 nm He-Ne laser. This method takes advantage of the remarkably high quantum yields of these naturally fluorescent proteins, which can be attributed to their linear tetrapyrrole chromophores covalently bound to cysteinyl residues. As such, limits of detection of 1.18 x 10(-14), 5.26 x 10(-15), and 2.38 x 10(-15) mol/l were obtained for R-phycoerythrin, C-phycocyanin, and allophycocyanin proteins, respectively, with a linear dynamic range of eight orders of magnitude in each case. Unlike previously published CE-LIF methods, this work describes the separation of all three major classes of phycobiliproteins in under 5 min. Very good recoveries, ranging from 93.2 to 105.5%, were obtained for a standard mixture of the phycobiliproteins, based on seven-point calibration curves for both peak height and peak area. It is believed that this development will prove useful for the determination of phycobiliprotein content in naturally occurring cyanobacteria populations, thus providing a useful tool for understanding biological and chemical oceanographic processes. SN - 0021-9673 UR - https://www.unboundmedicine.com/medline/citation/12416885/Separation_and_quantitation_of_phycobiliproteins_using_phytic_acid_in_capillary_electrophoresis_with_laser_induced_fluorescence_detection_ DB - PRIME DP - Unbound Medicine ER -