A nonsaponification method for the determination of carotenoids, retinoids, and tocopherols in solid human tissues

There has been an increasing interest in the measurement of carotenoids, retinoids, and tocopherols in human tissues because some of these micronutrients have been shown to have chemopreventive activity. Since clinical tissue samples obtained for analysis are usually small in quantity, a sensitive analytical procedure that can simultaneously measure all the micronutrients of interest in one small piece of human tissue is necessary. Moreover, some solid tissues, such as skin, are very difficult to homogenize unless they are first saponified in alcoholic KOH, but the saponification often causes substantial destruction of some micronutrients. Thus, a nonsaponification procedure using collagenase to facilitate homogenization was developed. Solid tissues are first incubated in a collagenase solution, homogenized, then incubated in a protease solution, followed by precipitation of tissue proteins and extraction with hexane. Collagenase digestion facilitates homogenization, and protease digestion increases the extractable amounts of the micronutrients from certain tissue samples. In this study, the recovery and precision (coefficient of variation) of the new procedure was determined. In addition, the extracted amounts of seven carotenoids, two retinoids, and two tocopherols from human skin, cervical/ovarian tissue, as well as sarcoma and kidney tumors using the new procedure and a commonly applied saponification procedure were compared.