Solid phase pegylation of hemoglobin.
A solid phase conjugation process was developed for attachment of polyethylene glycol to hemoglobin molecule. Bovine hemoglobin was loaded onto an ion exchange chromatography column and adsorbed by the solid medium. Succinimidyl carbonate mPEG was introduced in the mobile phase after the adsorption. Pegylation took place between the hemoglobin on the solid phase, and the pegylation reagent in the liquid phase. A further elution was carried out to separate the pegylated and the unpegylated protein. Analysis by HPSEC, SDS-PAGE, and MALLS demonstrated that the fractions eluted from the solid phase contained well-defined components. Pegylated hemoglobin with one PEG chain was obtained with the yield of 75%, in comparison to the yield of 30% in the liquid phase pegylation. The P(50) values of the mono-pegylated hemoglobin, prepared with SC-mPEG 5 kDa, 10 kDa and 20 kDa, were 19.97, 20.23 and 20.54 mmHg, which were much closer to the value of red blood cells than that of pegylated hemoglobin prepared with the conventional method.
Suo
Xiaoyan
X
National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing, People's Republic of China.
Zheng
Chunyang
C
Yu
Pengzhan
P
Lu
Xiuling
X
Ma
Guanghui
G
Su
Zhiguo
Z
D000327:Adsorption; D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006454:Hemoglobins / Q000032:analysis / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D007202:Indicators and Reagents / Q000737:chemistry; D007834:Lasers; D008970:Molecular Weight; D011092:Polyethylene Glycols / Q000737:chemistry*; D012542:Scattering, Radiation
D006454:Hemoglobins; D007202:Indicators and Reagents; D011092:Polyethylene Glycols