Reverse‐phase HPLC was used to fractionate 40S ribosomal proteins from human placenta. Application of a C4, reverse‐phase column allowed us to obtain 27 well‐resolved peaks. The protein composition of each chromatographic fraction was established by two‐dimensional polyacrylamide gel electrophoresis and N‐terminal sequencing. N‐terminally blocked proteins were cleaved with endoproteinase Lys‐C, and suitable peptides were sequenced. All sequences were compared with those of ribosomal proteins available from data bases. This allowed us to identify all proteins from the 40S human ribosomal subunit in the HPLC elution profile. By matrix‐assisted laser‐desorption ionization mass spectrometry the masses of the 40S proteins were determined and checked for the presence of post‐translational modifications. For several proteins differences to the deduced sequences and the calculated masses were found to be due to post‐translational modifications.