UV resonance Raman spectroscopic study of six short proline-containing peptides with antioxidant activity isolated from human breast milk was performed. The amide II′ proline spectroscopic band was used to estimate relative cis trans isomerization state of proline amide bonds in the different peptides. Antioxidant activity of the peptides was determined using 1,1-diphenyl-2-picryl-hydrazyl (DPPH) assay and linoleic acid emulsion assay. Although no clear correlation between the amide II′p position and antioxidant activity of the peptides was observed, they both were found to be sensitive to the presence and/or relative position of proline and tyrosine residues in the peptide. Copyright © 2011 John Wiley & Sons, Ltd.