Jh. Choe et al., PARAMETERS AFFECTING THE EFFICIENCY OF AFFINITY-BASED REVERSE MICELLAR EXTRACTION AND SEPARATION (ARMES) IN GLYCOPROTEIN PURIFICATION, Biotechnology progress, 13(4), 1997, pp. 440-445
Affinity-based reversed micellar extraction and separation (ARMES) is
an effective method for purifying both low and high molecular weight g
lycoproteins via liquid-liquid extraction. A range of extraction condi
tions were examined to gain insight into the mechanism of ARMES. Conca
navalin A (Con A) was used as the model affinity ligand to bind soybea
n peroxidase (SEP) and beta-galactosidase as model glycoproteins. Fact
orial design was used to investigate the effect of various system vari
ables on the extraction of SEP via ARMES. A quadratic model described
the system well, resulting in a standard deviation of 7% between calcu
lated and experimental extraction efficiencies. Sensitivity analysis s
uggested that the key criteria in ARMES were the NaCl concentration an
d pH of the aqueous feed phase. Extraction of both glycoproteins decre
ased above pH 7 but fell to zero only at pH values significantly above
the pi of the model glycoproteins and the Con A affinity ligand. It i
s proposed that the complex of the affinity lectin with the glycoprote
in results in a sufficiently hydrophobic species that can be extracted
into a reversed micellar organic phase even at pH's far above the pI'
s of the individual proteins that comprise the complex. This finding h
as practical considerations for the use of ARMES in the resolution and
purification of protein glycoforms.