HPLC Analysis of Insulin (Species Variants) on BIOshell A160 Peptide C18
Materials
analytical column
Ascentis® Express Peptide ES-C18, 2.7 μm HPLC Column
2.7 μm particle size, L × I.D. 15 cm × 2.1 mmCONDITIONS
column
BIOshell A160 Peptide C18, 15 cm x 2.1 mm I.D., 2.7 μm particles (66905-U)
mobile phase
[A] 70:30, water with 0.1% TFA:acetonitrile with 0.09% TFA; [B] 55:45, water with 0.1% TFA:acetonitrile with 0.09% TFA
gradient
0 to 100% B in 7.5 min
flow rate
0.3 mL/min
pressure
4040 psi (278 bar)
column temp.
30 °C
detector
UV, 215 nm
injection
0.5 μL
sample
200 μg/mL insulin (bovine, human, porcine) in 0.1% formic acid
Description
Analysis Note
These three insulins have more than 76% identical positions in a primary structure alignment. The human and porcine proteins are most similar with 85% identity. Baseline resolution of these insulins in achieved on BIOshell A160 Peptide C18, with relative band spacing of these three peaks approximating their relative sequence similarities. BIOshell A160 Peptide C18 is ideally suited for RP chromatography of small proteins or large peptides.
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