Amino acid replacements were used to probe the roles of 14 sites in two well-characterized intermediates in the folding pathway of bovine pancreatic trypsin inhibitor (BPTI). One of these intermediates contains one of the three disulfides found in the native protein (30--51). NMR studies have shown that approximately two-thirds of this polypeptide has a native-like conformation. The other intermediate contains two native disulfides (30--51 and 5--55) and has a fully folded conformation. The phi-values for a majority of residues were <1, indicating that the native protein was significantly more destabilized than either intermediate even when the altered residue was located in a well-ordered region of the intermediate. These observations suggest that folding intermediates and transition states may generally be more structured than indicated by phi-values alone.