| Summary: | 碩士 === 國立清華大學 === 生物科技研究所 === 102 === Carotenoids are yellow to red pigments that are notable for their wide distribution, structural diversity, and various functions. They are synthesized in plants and in some microorganisms.
The epidemiologic studies have revealed that there is an inverse relationship between the presence of degenerative disorders and dietary carotenoids. Moreover, anti-oxidative activities of carotenoid pigments have been notified in a number of studies. The prevention of these degenerative disorders by dietary carotenoids has been associated with capacity of carotenoids to protect cells and tissues from oxidative damages.
A powerful antioxidant can scavenge or trap free radical effectively. The common free radicals in human body are reactive oxygen species (ROS) such as hydroxyl radical and peroxyl radical that may cause oxidative stress. ROS are generated during normal cell aerobic respiration, which are harmful byproducts. Thus, people are beginning to attach importance to antioxidant for preventing free-radical-involved diseases. To date, the anti-oxidative activity of carotenoids has been measured by different methods; however, it is difficult to compare the results directly because of variation in free radicals, reaction conditions, and the types of cells used for analysis.
The aim of this research was to set up ABTS, FRAP, LPSC, and CAP-e assays to compare the anti-oxidative performance for a variety of carotenoids obtained from the specific microorganism and Haematococcus pluvialis (F1, F2, F3, P, Z and H, respectively) and commercial caroteinoids (astaxanthin, canthaxanthin, zeaxanthin, -carotene and lycopene) as well as from commonly used non-pigment antioxidants (BHA and Trolox). Carotenoids obtained from recombinant bacteria and algae were purified via Open Column Chromatography (OCC) followed by reprecipitation method. Among the four assays which were all modified for application in this study, LPSC and CAP-e were particularly adopted because they both use hydrogen peroxide (H2O2) as the source of oxidative stress.
In all four assays, Z and H (the carotenoids we purified) presented strong anti-oxidative activity in general. Zeaxanthin, Z and H had inhibition from 50 to 88%; and-carotene, lycopene and P also had over 33 percent inhibition in ABTS assay. The sample H and zeaxanthin had 6 times higher ferric reducing ability than other carotenoids in the FRAP analysis.
In LPSC and CAP-e assays, all analyzed carotenoids showed > 52% inhibition of H2O2-induced oxidation, indicating good anti-oxidative activity of carotenoids. Xantophylls presented significantly better antioxidative activities as compared with -carotene and lycopene at the same concentration.
We further compared natural carotenoids purified in this research with commercial carotenoids which were supposed to be in all-trans form, and found that their anti-oxidative activities had significant difference in CAP-e assays. Although commercial astaxanthin standard and the isolated F3 showed similar anti-oxidative activity in ABTS, FRAP and LPSC methods, F3 presented significantly higher anti-oxidative activities as compared with astaxanthin standard (P<0.05). Likewise, the commercial canthaxanthin showed significantly lower activities in comparison with F1 (P<0.001). At concentration of 5 μg/ml, natural carotenoids (F1 and F3) purified in this study showed 64% inhibition in average, while commercial carotenoids (astaxanthin and canthaxanthin) reached 32% only.
The possible reasons of the variation in anti-oxidative activities might be addressed to difference of chemical structures between stereoisoforms that probably affect the solubility and cellular uptake as well as distribution in the cell membrane.
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