The effect of various factors on the activity of rat liver sorbitol dehydrogenase was investigated. The activity of the enzyme (expressed per gram of tissue) was similar in the livers of adult male and female rats and was not affected by adrenalectomy, cortisone injection, or hypophysectomy. Different modes of administration of sorbitol also produced little change in activity. The enzyme activity declined significantly during 1 week of starvation. Activity was low in foetal liver and rose gradually during the postnatal period to reach adult levels by approximately 40 days after birth. Sorbitol dehydrogenase activity in the moderately well-differentiated Morris hepatoma 5123 T.C. was studied and found to be approximately 40% of that of normal liver. Using the technique of starch-gel electrophoresis in sodium borate, the rat liver enzyme was found to occur in multiple molecular forms. The electrophoretic pattern of the rat liver enzyme in sodium borate was not affected by a variety of metabolic conditions and was also identical in the hepatoma. From a study of various factors influencing the electrophoretic separation of rat liver sorbitol dehydrogenase, it is concluded that the multiple molecular forms are similar species of the enzyme and that interaction with a starch–borate complex may be important in their resolution.