The Study On The Isolation, Purification, Identification And Antioxidant Activity Of Mycosporine-like Amino Acids (MAAs) In Eucheuma

Mycosporine-like amino acids (MAAs) are widely distributed in Marineorganisms, especially in algae. With strong ultraviolet absorption characteristics, it isa natural high-efficient ultra-violet absorber. To date many researchers have found thatMAAs existed in a variety of marine organisms. Meanwhile, light induction of MAAssynthesis as well as its type and content in a variety of marine organisms have alsobeen reported. However, there were few studies paid attention to the activity andapplication of MAAs from Eucheuma. China is rich in marine resources, particularlyin seaweed Eucheuma, however the utilization is still very low. Therefore, it hasbecome a trend in future research to improve the utilization of algae.In this study, Hainan Eucheuma was selected as material, the contents are asfollows.(1) the classes of the MAAs compounds in Hainan Eucheuma wereconfirmed by analyzing the extracting solution of Hainan Eucheuma using liquidchromatographic and mass spectrometry;(2) the extraction process of the MAAscompounds by water bath extraction and ultrasonic-assisted extraction were optimized;(3) the purity of MAAs compounds in the samples was improved by ion exchangechromatographic.(4) MAAs were confirmed the antioxidant activity by vitroantioxidant experiments. The details are listed as below:(1)In this paper, the major MAAs ingredients from Eucheuma were analyzed andidentified using the UV spectrum scanning techniques, liquid chromatographic, liquidchromatographic-mass spectrometry. There were totally eight kinds of MAAs fromEucheuma. The maximum absorption wavelength, liquid chromatography-massspectrometry results showed that eight kinds of MAAs were identified, namely:Mycosporine-Gly, Palythine, Mycosporine-NMA:Ser, Mycosporine-2Gly, Shinorine,Usujirene, Porphyra-334, E-Palythenic Acid, of which the major is Shinorine and Usujirene, accounting for more than95%of the identified MAAs.(2) Two kinds of methods were establisted to determine the total MAAs content,including UV spectrometry and high performance liquid chromatographic (HPLC).MAAs content can be determined using HPLC due to the high accuracy. A fittingequation y=1.04727x2+(2.33413E-4)x+15.74627can also be used to determine MAAscontent.(3) On basis of orthogonal experiments, the optimum extraction process used inwater bath extraction method was as follows: the extraction time was2h, theextraction temperature was40℃, the ratio of liquid and material was60(g/mL),methanol concentration was30%. In order to overcome the shortcomings oftraditional water bath extraction methods, including the amount of solvent and longprocessing time, ultrasonic assisted extraction method had been introduced to improvethe extraction ratio of MAAs. The experiments not only determined the maininfluncing factors and optimal extraction conditions in terms of the MAAs extractionratio of the ultrasonic assisted extraction, but also reduced to liquid-solid ratio andorganic solvents such as methanol,and improved the extraction ratio. Therefore, theoptimal extraction conditions for ultrasonic assisted extraction were determined asfollow: ultrasonic time was20min, methanol concentration was10%, the ratio ofliquid and material was30(g/mL). Under optimal conditions, the extraction ratio ofultrasonic assisted extraction method was5.52mg/g. Compared with the water bathextraction, the extraction ratio increased by0.51mg/g.(4)001×7ion exchange resins was filtered out by static experiments,which it hadthe maximum ratio of adsorption and resolution. The the best purification conditionsof001×7ion exchange resins in static experiments were used as: concentration ofsample was5mg/mL, pH values was6, adsorption time was80min, ammonia waterconcentration was3%. In dynamic experiments study, the optimal adsorptionconditions of001×7column were used as: the sample flow ratio was2BV/h (columnvolume/h), the sample volume was6BV. Optimal elution conditions were as follows:elution ratio was1BV/h, elution dose was10BV. Under optimum purificationconditions, yield of MAAs was89.2%, purity of MAAs was92.3%. (5) The results confirmed that MAAs possessed the ability of scavenginghydroxyl radicals and superoxide anion, so MAAs had the ability of antioxidantactivity.