William M. Walter

Elimination of color quench in liquid scintillation counting of 14C-carotenoids

Abstract Methods of eliminating color quench in measuring radioactivity of carotenes by liquid scintillation spectrometry were studied. Solutions containing internal 14 C standards and 1 mg of lycopene or beta carotene were counted with about 0.5% efficiency. The pigments were decolorized in a counting vial using benzoyl peroxide and light. After 1–3 hours of decolorization of the pigments, counting efficiencies of over 80% were obtained in PPO-POPOP phosphor. Similar results were obtained in counting 14 C carotenes using an external standard to determine efficiency.

[245] Preparation of14C-β-Carotene

Publisher Summary Most of the systems used for study of carotenogenesis are deficient in either yield of β-carotene or incorporation of label. This chapter discusses method for the preparation of 14 C-β-carotene. It is found that multiple crystallization gives higher yield and greater purity than any other method. Evaporate the chromatographic fraction to dryness in vacuum and redissolve in a minimum volume of boiling hexane, about 1 ml of hexane per milligram of carotene. The minimum volume can be determined by removing hexane vapors with a stream of nitrogen while warming the flask in a water or steam bath until an insoluble ring of carotene begins to form. Specific activity can be conveniently determined by measuring the amount of β-carotene spectrophotometrically and counting a known amount in suitable 14 C counting systems. Spectrophotometric determination in hexane at 450 nm using an absorption coefficient of 0.250 mg per liter gives excellent agreement with gravimetric determination.