The analysis of oligopeptides in samples of food, tissues, and body fluids attracts considerable attention. The complexity of such samples requires efficient sample preparation (i.e., concentration and cleanup) procedures to remove interfering endogenous compounds and inorganic or organic salts. The methods of sample pretreatment that enable effective and selective isolation and/or preconcentration of oligopeptides from complex sample matrices have been reviewed. In each case, examples of application were presented and discussed, taking into account selectivity, enrichment, method automation, cleanup, and environmental aspects of the developed methods.