Separations on reversed-phase media are usually done in mixtures of aqueous and organic solvents, which are often denaturing conditions. Samples are loaded onto the matrix in a high-salt buffer and elution is by a descending salt gradient. Separations on HIC matrices are usually done in aqueous salt solutions, which generally are nondenaturing conditions. Hydrophobic interaction chromatography (HIC) and reversed-phase chromatography (RPC) are two separation methods based on the interactions between the hydrophobic moieties of a sample and an insoluble, immobilized hydrophobic group. The nine hydrophilic amino acids are listed below, with the remaining two amino acids tyrosine (Tyr) and cysteine (Cys) defying categorization at this time. In addition, many hydrophobic amino acids are exposed on the surface and it is these that give a native protein its degree of hydrophobicity. The nine hydrophobic amino acids are alanine (Ala), glycine (Gly), valine (Val), leucine (Leu), isoleucine (Ile), phenylalanine (Phe), proline (Pro), methionine (Met), and tryptophan (Trp). This makes them hydrophobic (water fearing). Hydrophobic interactions stabilize the tertiary and quaternary structure of proteins. Group I amino acids are glycine, alanine, valine, leucine, isoleucine, proline, phenylalanine, methionine. Certain amino acids are hydrophobic in order of decreasing hydrophobicity, they are tryptophan, norleucine, phenylalanine, tyrosine, leucine, valine, methionine, and alanine. The degree of hydrophobicity of a protein is dependent on its amino acid sequence. The nine amino acids that have hydrophobic side chains are glycine (Gly), alanine (Ala), valine (Val), leucine (Leu), isoleucine (Ile), proline (Pro). It is the structure of the water that creates hydrophobic interactions. while the hydrophilic side- chains are on the exterior where they can. Hydrophobic interactions are forced on nonpolar compounds by the polar environment. of water-soluble proteins are predominantly composed of hydrophobic amino acids.

Hydrophobic amino acids in water