The root of Lithospermum erythrorhizon which produces shikonin pigment has long been used as a medicine for wounds, burns, hemorroids, and dyestuffs in Asia (Fujita et al., 1985). We have previously selected single cell clones (LE16, LE19, LE67, LE87, LE88) which produce shikonin pigment derivatives and another group of cell lines (LE1, LE11, LE18) which fail to produce shikonin pigment derivatives from the single cell culture of L. erythrorhizon. In this experiment, we analyzed in vivo and in vitro protein synthesis related to the shikonin production between shikonin-producing (LE16, LE87) and shikonin-nonproducing (LE1, LF11) cell lines in suspension cell culture. The cellular cytoplasmic proteins which were labeled with ^(35)S-methionine in vivo were fractionated by one-dimensional two-dimensional gel electrophoresis and visualized by auto radiography at different culture time. The variation of mRNAs was also analyzed from the autoradiograms of in vitro translation products of poly(A)^+ RNA isolated at different culture time. The results of one-dimensional electrophoresis were analyzed by densitometer in the visible light zone. Several polypeptides showed qualitative or quantitative changes during the shikonin production.