Abstract: The present invention relates to a process for preparing human proinsulin which is represented as a following chemical formula(I): ##STR1## wherein, R is an amino acid residue or a peptide which is degradable enzymatically or chemically; and, X is a linkage of an amino group of A-1 in insulin A chain and a carboxyl group of B-30 in insulin B chain which can be separated from the A chain or the B chain enzymatically or chemically, provided that a region from A-1 to A-21 is the insulin A chain and a region from B-1 to B-30 is the insulin B chain. In accordance with the present invention, human recombinant insulin precursor can be simply manufactured with a good reproducibility, since dissolution, sulfonation, concentration, desalting and purification are remarkably simplified, while increasing the yield of refolding reaction.