Chromatographic separation and mass spectrometric identification of positional isomers of polyethylene glycol-modified growth hormone-releasing factor (1-29).
A one-step chromatographic method capable of separating all isomers of polyethylene glycol (PEG)-growth hormone-releasing factor (GRF) (1-29) conjugates was developed. The unmodified GRF (1-29) and seven different isomers of PEG-GRF (1-29) conjugates were separated by using a simple reversed-phase HPLC method depending on the differences of hydrophobicity due to the number and site of PEG attachment. The PEGylation sites of all isomers of PEG-GRF (1-29) conjugates were identified by determining the molecular masses of the Lys-C digested fragments with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This study is a first report for the separation of all PEG-conjugate isomers and would be useful for further studies to find the promising conjugate by evaluating biological activity and stability of each isomer.
Youn
Yu Seok
YS
Drug Targeting Laboratory, College of Pharmacy, SungKyunKwan University, 300 Chonchon-dong, Jangan-ku, Suwon City 440-746, South Korea.
Na
Dong Hee
DH
Yoo
Sun Dong
SD
Song
Soo-Chang
SC
Lee
Kang Choon
KC
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D013007:Growth Hormone-Releasing Hormone / Q000737:chemistry*; D007536:Isomerism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry*; D011092:Polyethylene Glycols / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D010446:Peptide Fragments; D011092:Polyethylene Glycols; D013007:Growth Hormone-Releasing Hormone; C059148:somatotropin releasing hormone (1-29)