The Extraction And Purification Of Astaxanthin In Antarctic Krill (Euphausia Superba)

Background The Antarctic Krill (Euphausia superba) is a kind of marineinvertebrate belonging among class Malacostraca. It provides with the biggestbiomass energy in the nature，and it is the principal prey of many marine predatorssuch as Cetacea, Phocidae, by reason of its low station in the food chain. Theexploitation of Antarctic Krill has been popular in many countries on the earth, ourcountry also officially began to catch and study it from December, 2009. It is not onlythe key organism in the southern ocean, but also the biggest fishery resource in theworld which urgently to await development and utilization. Meanwhile, the AntarcticKrill is one of the best resource of natural astaxanthin with high content of astaxanthinand other carotenoids, and astaxanthin esters are main components in the carapace ofEuphausia superba.The ketocarotenoid astaxanthin is a kind of molecule with lively hydroxyketonegroup, its molecular formula is C40H52O4. Astaxanthin is one of most importantcarotenoid in nature in virtue of its ultra oxidative stability. Moreover, it still has othergood physiological functions such as natural coloring (in fish muscles, squamas, birdfeathers and yolks, etc.), immunity enhancement, antitumor effect and protection fromultraviolet radiation. Overall, astaxanthin can play an important part in healthcare andcosmetic manufacture area with enormous exploitative and applicable prospect.Intention This paper aims to investigate some easy, fast and high-efficiency methodsfor extraction, purification and routine detection of astaxanthin in Euphausia superba ,the high performance thin layer chromatography(HPTLC) and the normal pressuresilica gel column chromatography were mainly selected to separate and purify theastaxanthin，the thin layer scanning method and the LC-MS method were used toverify the purity. High efficiency, rapidness and reagent-saving should be drived for inthe experiments, and the methods were also environmentally friendly with lesspoisonous organic solvent residue. Technique Red oil of Euphausia superba was extracted from dry Antarctic Krill withliposoluble organic solvents, the high performance thin layer chromatography(HPTLC) method was used to separate and analyze the astaxanthin species, theastaxanthin species were composed of free astaxanthin and different kinds ofastaxanthin esters. The methanol solution of KOH was used in the saponification ofastaxanthin esters, which were hydrolyzed to release the free astaxanthin. The sampleswere concentrated after saponification. The traditional normal pressure silica gelcolumn chromatography was selected to purify it immediately. Every kind of pigmentfrom elution was collected and concentrated, then they were contrasted to theastaxanthin standard by TPTLC to find out which sample was the free astaxanthin,and at last the degree of purity was checked up by HPTLC and LC-MS.Result After the HPTLC analysis of the extrated sample of astaxanthin esters,three constituents was mainly separated. The Rf Value of the free astaxanthin,themonoester and the diester was 0.32, 0.63, 0.91, respectively. The degree of purity ofthe free astaxanthin was from 5%±1.0% to 85%±5.0% after saponification inmethanol solution of KOH. The free astaxanthin that was purified by this experimentwas analyzed by LC-MS and was get a relative molecular mass of 596.3, which wasthe same with the natural astaxanthin, which is 596.86, and the UVspectrodensitometric analysis of it by the TLC scanner 3 spectrodensitometer was 476nm ofλmax, which was the same with the standard of astaxanthin. On the whole, themolecule which was purified by the traditional silica gel column chromatographycould corroborate to be the natural free astaxanthin.Conclusion and Innovation In this paper, a series of experiments were carriedout to extract and purify the astaxanthin in the Antarctic Krill, and the purity of theastaxanthin after purification can be 99.36% The innovation: A high performance thinlayer chromatography (HPTLC) method was established and validated for analyzingthe astaxanthin and astaxanthin esters in the Antarctic Krill for the first time, and themethod was proved to be stable, dependable and with good reproducibility, anon-pressurized column chromatography was selected for the purification of the freeastaxanthin with a fine effect, the effect of purification was proved to be well after theanalysis by HPTLC and LC-MS, with good specificity, succinctness and efficiency.