Effect of different concentrations of calcium chloride and potassium chloride on egg white proteins during isoelectric precipitation of ovomucin

Abstract

The effect of various concentrations of CaCl2 and KCl on egg white proteins during isoelectric precipitation of ovomucin was investigated in this study. At low concentrations of CaCl2 (<50 mM), lysozyme was the major contaminant in the precipitated ovomucin, whereas ovalbumin was the predominant one at high concentrations (>or=100 mM). At 50 mM CaCl2 concentration, the concentrations of both lysozyme and ovalbumin were moderate. Ovomucin with a purity of 97.3% was prepared using a 2-step method: egg white was first precipitated in the presence of 50 mM CaCl2 followed by a second 500 mM CaCl2 extraction. The concentrations of other proteins in the precipitate were 1.3% of ovalbumin, 1.1% of lysozyme, and 0.4% of ovomucoid. Unlike CaCl2-treated samples, ovotransferrin was found to be the second major contaminant in all KCl-treated precipitates. Compared with the control, adding KCl at the lowest concentration of 2.5 mM increased significantly the content of ovalbumin (from 7.6 to 68.0%) and reduced significantly the content of lysozyme (from 25.5 to 6.4%) in the precipitates; however, increasing the concentrations of KCl up to 500 mM did not affect the content of ovalbumin, but the content of lysozyme showed a general reduction trend. Although KCl was used widely in literature as the last step of ovomucin washing, our results show that KCl is not an efficient salt in purifying ovomucin.

Role of magnesium chloride on the purity and activity of ovomucin during the isolation process

“…Ovomucin is usually present in egg white as a complex with lysozyme and other albumen proteins [1]. It was interesting that there was only a weak band co-responsible for lysozyme, which was greatly different from the results using NaCl or CaCl 2 in previous reports [10,14].…”

“…In particular, the content of lysozyme was below 1% when the MgCl 2 concentration was greater than 50 mM. Moreover, magnesium ion, a weak salt-in reagent, can be helpful to enhance the activity of proteins (Table 3), although the content of albumin could also be reduced to some extent by using the appropriate concentration of NaCl and CaCl 2 [10,14]. Thus, we attempted to isolate ovomucin using IEP in the presence of different types of salts.…”

“…Thus, solutions at pH 6 were used to precipitate ovomucin. Since there have been reports on the effects of NaCl and CaCl 2 on the ovomucin precipitation composition [10,14], we decided to investigate the effects of MgCl 2 concentration on the protein composition and antivirus activity of the ovomucin precipitates prepared using isoelectric precipitation.…”

“…It was reported that high salt concentration minimizes the complex formation between ovomucin and other egg white proteins like lysozyme [18]. The protein compositions of ovomucin precipitates are significantly affected by the NaCl or CaCl 2 concentrations [10,14]. Magnesium ion was also reported to decrease the interaction between ovomucin and lysozyme [19].…”

“…On the other hand, a higher purity of ovomucin separated via IEP can be obtained by KCl washing, yet KCl washing was reported to cause a loss of ovomucin [12,13]. Recently, modifications of the IEP method have been proposed that employ a limited number of salts such as NaCl or CaCl 2 to overcome the latter problem [10,14]. However, salt is well known to have some effects on the function or biological activity of proteins.…”