A model mechanism for protein precipitation by caprylic acid: Application to plasma purification

Abstract

A model for the mechanism of protein precipitation by caprylic acid (CA) is developed on the basis of quantitative assays of precipitation with bovine serum albumin (BSA) and CA at different concentrations. It was found that the effect of CA is due to direct interaction with the precipitating protein. Maximum precipitation was achieved when the mass ratio of CA-BSA was close to 1, equivalent to about 450 CA molecules per molecule of BSA. This value was confirmed by optimizing the CA purification of immunoglobulins from equine blood plasma. With a sample diluted 1:1, it was found that CA at a final concentration of 3.5% is optimal to obtain immunoglobulins essentially free of albumin by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It is proposed that CA binds to specific sites of the protein, thereby inducing partial unfolding of the protein, which exposes additional binding sites. More CA molecules incorporate into all sites in the form of mixed micelles. Thus, the interfacial protein surface becomes highly hydrophobic and increases protein-protein attraction, causing association and precipitation of the macromolecular complexes.

Immunoglobulin Purification by Caprylic Acid

“…Caprylic acid precipitates albumin and other plasma proteins by direct interaction [ 20 ]. The protocol can be easily scaled up to larger volumes.…”

“…Morais and Massaldi [ 20 ] have recently proposed a mechanistic model for the action of caprylic acid to precipitate plasma proteins, where direct interaction between protein and caprylic acid takes place. They also found a mass ratio of caprylic acid to albumin about 1 to be optimal for maximum precipitation and purity of the remaining IgGs at any dilution.…”

“…Several articles have shown that precipitation at pH 5 gives the best purity results for the supernatant immunoglobulins after precipitation of the non-immunoglobulin fraction [ 20 ]. At pH above 8 or below 3, precipitation is minimal [ 20 ].…”

“…Several articles have shown that precipitation at pH 5 gives the best purity results for the supernatant immunoglobulins after precipitation of the non-immunoglobulin fraction [ 20 ]. At pH above 8 or below 3, precipitation is minimal [ 20 ]. The optimal caprylic acid concentration is 7 % v/v for undiluted plasma [ 23 ] or 3.5-4 % v/v for a 1:1 plasma dilution [ 20 ].…”

Optimization of purification procedure for horse F(ab´)2 antivenom against <i>Androctonus crassicauda</i> (Scorpion) venom

“…In addition, it is generally used alone or in combination with salt precipitation in order to purify immunized plasma from both equine and ovine origins [13]. Furthermore, caprylic acid is an unsaturated fatty acid and a good precipitating agent for undesired proteins from animal plasma essentially albumin without any effect on immunoglobulin fraction [14].…”