Characterization of peptides in human cerebrospinal fluid

The report of endorphin-like activity in human CSF [7] has stimulated us to study peptides in various CSF specimens. At first we attempted to fractionate CSF by gel-filtration and paper-mapping procedures. By these techniques, we have isolated a peptide (Gly-Ala3-Val-Leu) contaminated with glutamic acid (or glutamine), which could not be removed by either of these methods. An HPLC method was developed for peptide fractionations on a RP-18 column with a gradient of 18 mM ammonium acetate and acetonitrile. This system has been evaluated with synthetic peptides of molecular weight up to about 1850, and has the advantages that the buffer salts may be removed by lyophilization and separations are performed at or near neutral pH thus minimizing alterations in the structure of the peptides. The retention characteristics of peptides on a C18 column is a function of the pH of the buffer. This facilitates the characterization of peptides since each may be identified by its unique chromatographic behavior at different pH values on one column.