Analysis was performed at the ARC-Irene Analytical Services,
using a method accredited according to ISO/IEC 17025:2005. The
accreditation body is the South African National Accreditation System
(SANAS). The concentration of vitamin A was determined as
described by Hulshof (2002). The basic method entails the alkaline
saponification of the test material to eliminate the fat and to liberate
the natural retinol in the cells. This is followed by ether extraction
of the retinol and determination by high performance liquid
chromatography (HPLC). The HPLC system (Shimadzu) consisted
of a Quaternary gradient pump (model LC-20AD), a solvent degasser
(model DGU-20A5), an auto-injector (model SIL-20A, 230 V), a
Photodiode Array Detector (DAD) with a thermostatted standard
cell (model SPD-M20A) and control and integration software
(LCsolution Ver. 1.1). A Nucleodur 250 4 mm reverse phase C18
column (5 lm particle size) with guard column was used. Separation
was achieved using a mobile phase of 97% methanol in deionised
water and a flow rate of 1.0 ml/min. Separation was performed
at 325 nm for the identification and quantification of retinol.