Practical Protein Chemistry - A. Darbre 1989

Separation of protein and peptide mixtures by high-performance liquid chromatography
Reversed-phase separation of peptides

RP-HPLC was first used for the Separation of peptide mixtures in 1976 simultaneously and independently in several laboratories. Since then, numerous reports have been published on The Use of various Supports and mobile phases to develop Methods for the rapid, high-efficiency separation of biologically active Peptides or protein fragments. Because many similar sorbents exhibited only minor differences in resolving power, the literature became flooded with largely overlapping methodologies.

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FIG. 6.9. Separation of native Proteins (~20 μg each) by RP-HPLC using wide-pore sorbents with short alkyl chains. Proteins were fractionated in 0.01 M TFA with an acetonitrile concentration gradient (20–70%) at a flow rate of 0.2 mL/min. An Ultrapore RPSC Column (4.6 × 75 mm) was used. Detection was performed by optical density measurement at 280 nm.

The peptide separation process is influenced by The Nature of the organic solvent, Water content, Ionic strength, pH, Temperature, and the capacity of the system for ion-pairing and ion-exchange interactions.



Last update: 06/08/2026

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