The Studies On Purification Of Astaxanthin From Phaffia Rhodozyma By High-speed Counter-current Chromatography And Properties Of Astaxanthin

Astaxanthin?3,3?-dihydroxy-?,??-carotene-4,4?-dione?,a kind of non-vitamin A source ketone type carotenoid,has a lot of biological activities such as antioxidant,anti-aging,radiation protection and immunity enhancement.It has broad application prospects in food additives,functional food,cosmetics industry,medicine industry and other fields.Phaffia rhodozyma which is an astaxanthin over-producing strain is used to produce astaxanthin.Nowadays the traditional extraction method of astaxanthin from P.rhodozyma has some defects,such as longer separation time,heavy workload and so on.In this study,astaxanthin over-producing strain P.rhodozyma as raw material,a new rapid,efficient separation and purification method was researched which called high-speed counter-current chromatography.In addition to the research of the separation and purification method,a series of studies were done for the stability and safety assessment of astaxanthin.At the same time,other carotenoids of P.rhodozyma were separated to obtain more bioactive substances.The main results were shown as the following:?1?By single factor and orthogonal test,the best extracting conditions of astaxanthin from P.rhodozyma were obtained:acetone as extraction solvent;the dosage of DMSO,1:2;the wall temperature,45 ^oC;material to liquid,1:20?w/v?;extraction time,40 min;extraction times,twice.?2?With the solvent system of n-hexane-acetone-ethanol-water?1:1:1:1,v/v/v/v?,separation temperature of 25 ^oC,rotate speed of 850 rpm and the flow rate of 3 mL/min,20.6 mg of astaxanthin at 92.0%purity were gained from 100 mg crude extract of P.rhodozyma by high-speed counter-current chromatography By further silica gel column chromatography,the purity reached 99.0%.?3?Some factors had been done to confirm the certain impact on the stability of astaxanthin,such as the light,temperature,pH,metal ion and potassium sorbate.When the light was stronger and the temperature was higher,the stability of astaxanthin was lower.Therefore,preserve food should be in low temperature and avoid light as far as possible.Under the condition of neutral slightly alkaline astaxanthin is relatively stable.Astaxanthin is sensitive to the factors of Fe³⁺and Cu²⁺,so the packaging material should avoid using iron and copper.Meanwhile,a certain amount of potassium sorbate could be added to enhance the stability of astaxanthin products.?4?The acute oral toxicity test was used to measure the toxicity of astaxanthin crude extract?purity of 78%?.The mice tolerance dose of astaxanthin was greater than 15 mg/g,and its LD₅₀was greater than 15 mg/g.The acute toxicity was classified for non-toxic.?5?Further separation of crude extract from P.rhodozyma had been researched using high-speed counter-current chromatography and column chromatography technology.Except astaxanthin,P.rhodozyma may also contain?-carotene,astaxanthin esters,canthaxanthin,echinenone,etc.This study provided the theoretical basis and technical support for the development and utilization,a large number of preparation of astaxanthin and the preparation of other carotenes.