Cell-associated retinoids were extracted from the squamous cell carcinoma line SCC12b grown in submerged culture and were analyzed by gradient reverse-phase HPLC with photodiode array detection. In contrast to normal human keratinocytes (NHK) SCC12B have a 5 fold reduced retinoid concentration, the fractional composition of which is influenced by the degree of differentiation. In cells which have not undergone differentiation there is little ester detectable by mass. In cells allowed to remain at confluence for 10-14 days, the ester mass increases to about a third that of NHK. Assessment of the steady state concentration of cellular retinoids derived from medium 3 H-retinol added at 25nM revealed that the total retinoid load is about 1.8 pm/mg and 2.4 pm/mg in nondifferentiated and differentiated cultures, respectively. The nondifferentiated cells contain nearly equivalent amounts of retinol and retinyl esters (.8pm/mg and .5pm/mg, respectively). The differentiated cells show an increase in retinoid esters to an average of 1.6 pm/mg. In all cases the major retinoid species is retinyloleate/retinylpalmitate. Active metabolite concentrations are not altered significantly during differentiation and are similar to those found in confluent NHK. The impact of retinoic acid (RA) on retinol metabolism was also found to be similar to that of NHK; addition of RA (5 10 - 7 M) to the growth media increased retinyl ester concentrations. Three fold increases were seen in a 24 hour period and are consistent with an increase in the rate of esterification and/or a decrease in retinylester utilization.