We have established and validated a protocol for the peptidomic analysis of rat urine using CE coupled to MS (CE-MS). In the first experiments, the reproducibility of the CE-MS set-up and of the established preparation procedure were assessed. To establish a first rat urinary peptidome map, samples were also analyzed using CE-FT-ICR. The subsequent analysis of independent samples from two different strains (WISTAR and CD) indicated strain-specific differences, which were validated in a blinded assessment. MS/MS revealed the presence of specific fragments from well-known urinary rat peptides, such as collagens, alpha-1-antitrypsin, and serum albumin. The CE-MS-based peptidomics platform may provide novel insights into body fluids of animal models, such as rat or mice. Together with peptide identification, the technology is an excellent, complimentary, and non-invasive tool to analyze toxicological or other (patho)physiological effects of pharmaceutical compounds in animal models.

Figure 1: Reproducibility of urinary rat polypeptide evaluation. Examples of electropherograms obtained for 4 (of 19) consecutive measurements of a single urine sample. The m/z values of the 2-D raw data plots (upper panel) and the molecular weight (logarithmic scale) of the deconvoluted 3-D plots (lower panel) on the y-axis are plotted against CEmigration time on the x-axis. The arrangement of the analyzed peptides in distinct lines is obvious, and can be taken as a result of the number of positive charges at pH 2. The white arrow indicates 18.7-kDa signals probably representing major urine protein.