The document discusses the effect of bonded-phase on the resolution and retention of polypeptides. It highlights that differences in bonded-phase selectivity can significantly improve the resolution in reversed-phase analysis of peptides and proteins. The separation between peaks 2 and 3, peaks 5 and 6, and peaks 8 and 9 illustrate these selectivity changes. Additionally, it mentions that temperature can also be used to achieve desired separations. The steric protection of ZORBAX SB packings makes them well suited for chromatography at very low pH.
The document highlights that selectivity changes between samples are often accomplished by using different bonded phases. It notes that sterically-protected bonded phases of ZORBAX SB packings are well suited to chromatography at very low pH.
A specific experiment is described comparing the separation of various polypeptides, including Met Enkephalin, Leu Enkephalin, Angiotensin II, Neurotensin, Insulin (BOV), Cytochrome C, Myoglobin, and Calmodulin. The analysis was performed using ZORBAX 300 SB-CN, SB-C3, SB-C8, and SB-C18 columns under specific mobile phase conditions.
The experiment uses ZORBAX 300 SB-CN, SB-C3, SB-C8, and SB-C18 columns.
The mobile phase consists of ACN:Water with TFA at varying percentages over 20 minutes.