Study On Extracting Technology And Analyzing Methods Of Lutein In Marigold Flower

This paper relates to a process for the preparation of xanthophylls crystals containing at least 96% trans-lutein. Fist, HPLC method was established for the quantitative determination of lutein which prepared from Marigold Flower. The HPLC used Diamonsil C18 column, a mixture of methanol,acetonitrile,ethyl acetate and water which volume followed this rate: methanol: acetonitrile: ethyl acetate: water= 30:29:40:1 as mobile phase, with a flow rate of 0.75 mL·min^-1 and UV detection at 450nm.The temperature of column is 25℃.Each experiment determined 20μL of the sample. The detection limit was 1×10^-7g·mL^-1 ,The RSD was 0.96%;the recovery of added lutein was 99.49 %～100.22%. Various experiments were performed to evaluate the current Association of Official Analytical Chemists (AOAC) method used for the determination of carotenes and xanthophylls. Incorporation of nitrogen gas and VC during extraction and saponification and combination of extraction time and saponification time for 16 h increased carotene and xanthophylls contents. Respectively, cold saponification also resulted in higher xanthophylls contents than hot saponification. A 1:1 mixture of magnesia and diatomaceous earth was found suitable to replace silica gel as the adsorbent to separate major carotenoids by open-column chromatography. the extraction proportion of which is 12:8:5:5; the xanthophylls eluant was adjusted as n-hexane: acetonitrile: methanol equals to 70:25:5; the total content of xanthophylls was added about 33.32% in all.In addition, the effects of the supercritical CO₂ extractive process parameters on total output of xanthophylls resin were investigated to confirm a better output. To optimize the extracting process orthogonal test was taken and variance analysis were used.The best supercritical CO₂ extractive parameters are as follows: 45 MPa of extracting pressure, 50℃of extracting temperature, 8 MPa and 55℃for the separating kettleâ… , 5MPa and 20℃for the separating kettleâ…¡.CO₂ flow rate 18 L·h-1,extractive time 3h. Then ultrasonic-assisted process was investigated for the mixed solvents petroleum ether and acetone(1:1,v/v) extraction of xanthophylls resin from the remainder of the supercritical CO₂ extractive process. And The optimal ultrasonic-assisted extraction conditions were 25:l for ratio liquid of to material.450W for ultrasonic power and 25 min for ultrasonic time. Extracting xanthophylls resin from Marigold Flower can improve the total output of extraction from Marigold Flower from 10% to 24%.Then, the oleoresin of xanthophylls is mixed with the methanol and isopropanol in which alkali is dissolved. The ratio of the constituents of the saponification reaction mixture are about 1 part oleoresin (by weight) and 12 parts isopropanol(by volume), preferably methanol and 0.25 parts of alkali(by weight). The saponification is performed by heating the reaction mixture preferably to a temperature of 65℃for a period of 4 h. Removing the aliphatic alcohol from the mixture to get saponified crude concentrate; admixing the saponified crude concentrate obtained with water; extracting the diluted oily mixture obtained with dichloromethane to get xanthophylls crystals. Collect the aqueous phase and added calcium chloride to deposit Potassium fatty acid and separate the xanthophylls. The concentrate of xanthophylls obtained is admixed with a mixture of ethyl acetate and petroleum ether mixture at room temperature with stirring. The ratio of the mixture used 1:4 volume/volume. Finally, these crude crystals are further purified by washing with ethanol.At length, stabilities of lutein during the process of its preparation and storage .Extracting xanthophylls resin from Marigold Flower with supercritical CO₂ the extracting temperature should be under 55℃. The saponification process using excessive KOH under high temperature condition could make Lutein degradation, the best mass of KOH is 1/4 of the mass of xanthophylls resin. The temperature must be under 80℃,the Stability was better with N2 and VC as protector, The High purity Lutein should be vacuum packed in aluminum foil bag and preserved in dark place and the temperature should be -20℃.In solvent, the stability of Lutein is better.