Fractional Protein Precipitation Using Triazine Dyes

Abstract

Changes in temperature, pH and ionic strength, or the addition of organic solvents, may precipitate protein from aqueous solution. These effects have been widely used in protein fractionation (Green and Hughes, 1955), and before the introduction of chromatographic methods were virtually the only means of doing so (Kaplan, 1983). Other techniques for effecting protein precipitation include the use of heavy metal ions and ionic polymers. A range of compounds comprising a mixture of aromatic and charged groups are known to promote protein precipitation. Of particular interest for the purposes of the present discussion are those observations of protein precipitation in the presence of organic dyes. Thus, 0.4 per cent rivanol was observed to precipitate albumin, fibrinogen and many other plasma proteins under mildly alkaline (pH 8.0) conditions (Horejsi and Smetana, 1956). Fibrinogen was precipitated from plasma by use of high concentrations of tetrazolium blue (0.5 mg/ml) to yield active protein (Vila et al., 1984). Both these dyes are positively charged aromatic compounds at the pH used. Protein precipitation has also been obtained by use of polysulphonated aromatic compounds, including triazine dyes. Thus, polylysine is quantitatively precipitated from solution upon the addition of Methyl Orange (Itzhaki, 1972), or Trypan Blue (Shen et al., 1984).