Individual working solutions of each carotenoid were injected in the HPLC system on
the day of the preparation for purity check (ratio of peak area of the individual carotenoid to sum of all the chromatogram peak areas)and the concentrations of the standard solutions were corrected accordingly. Six levels of working calibration solutions with a mixture of the six carotenoids (concentrations 0.05–5g/mL, evenly distributed) were injected daily to build the calibration curves by the weighted least squares method