Fast protein separation by reversed-phase high-performance liquid chromatography on octadecylsilyl-bonded nonporous silica gel. II. Improvement in recovery of hydrophobic proteins.
Recovery of hydrophobic proteins from an RP-HPLC column was improved using a fast-separation RP-HPLC system operated at room temperature. Hydrophobic proteins such as ovalbumin could be adequately eluted from a nonporous octadecylsilyl (C18) spherical silica gel with a particle diameter of 20 microns using steep gradient elution with a 0.1% aqueous trifluoroacetic acid-acetonitrile system at a constant flow rate of 4 ml/min. Recoveries improved under fast separation since the protein sample suffered only a slight amount of irreversible denaturation on the hydrophobic surface of the stationary phase. The fast-separation system was also applied to the separation of larger proteins such as apo-ferritin (443 kDa) and thyroglobulin (669 kDa) as well as egg white proteins.
Itoh
H
H
School of Pharmaceutical Sciences, Kitasato University, Tokyo, Japan.
Nimura
N
N
Kinoshita
T
T
Nagae
N
N
Nomura
M
M
D000818:Animals; D001052:Apoferritins / Q000302:isolation & purification; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D009113:Muramidase / Q000302:isolation & purification; D010047:Ovalbumin / Q000302:isolation & purification; D011506:Proteins / Q000302:isolation & purification*; D012822:Silicon Dioxide; D013954:Thyroglobulin / Q000302:isolation & purification; D013997:Time Factors
D011506:Proteins; C026679:octadecylsilica; D012822:Silicon Dioxide; D010047:Ovalbumin; D013954:Thyroglobulin; D001052:Apoferritins; D009113:Muramidase