The split-and-pool combinatorial synthesis method enables the efficient solid-phase preparation of libraries consisting of a single library member per resin bead. Miniaturized assays are used to screen the library and determine the activity of the library members. A means of determining the structure of the library members is required and a variety of methods have been proposed for this purpose. For libraries of peptides and peptide-like compounds single bead Edman degredation is a possibility, although due to the requirement for an N-terminus it does not work for all classes of compounds (e.g. capped peptides, cyclic peptides). Sequencing by mass spectrometry is an attractive option due to the requirement for only a small sample size and the high speed of analysis. Various strategies for chemical encoding and ladder preparation to aid mass spectrometric structure determination have been described, although many of these require additional complications to the library synthesis, and may result in contamination of the library products with the encoding molecules.

Direct sequencing of peptides by collision induced dissociation (CID) or post source decay (PSD) has commonly been applied in proteomics applications. We wished to employ this methodology for sequence determination of library peptides as it does not require any encoding/capping steps during the library synthesis. Using automated spectral acquistion and data analysis this technique has the potential for great speed, permitting sequencing of both library "hits" and inactive compounds for construction of preliminary QSAR.

The program Sequence V1.5 was developed specifically for the sequencing of peptides from split-and-pool combinatorial libraries using CID mass spectrometry.