The total carotenoid content was determined
according to the method of Lee and Castle (25)
that was modified for pumpkin. A sample of 2 ml
FPP was mixed with 38-ml extraction solvent
(hexane-acetone-ethanol (50:25:25 v/v/v)) with
the homogenizer at speed 4 (Ultra-Turrax
Ika-Werke, Staufen, Germany). After centrifugation
(4000 rpm, 10 min, 5 °C) (CFC free Universal
Hettich Zentrifugen), the absorption was
measured with a Varian Cary 50 Scan (Australia)
spectrophotometer at 450 nm. Total carotenoid
content was determined as milligrams of
E-carotene per gram of sample (db) as Eq. 1: