CDOM From Heterosigma Akashiwo (Hada) Culture Solution: Chemical Characterization And Its Potential Allelopathic Effects

The chromophoric dissolvable organic matter (CDOM) , one of an important components in the marine dissolvable organic matter storage, play an important role in influencing ocean color, primary productivity and ecosystem structure, as well as the air-sea exchange. Therefore, study on CDOM has became a hot topic in recent years. As its composition is too complex to fractionate, researches at home and abroad are mainly carried on optics behavior of CDOM in natural waters, such as oceans and lakes, while isolation and identification of CDOM are still in an exploratory stage.In this study, the CDOM excreted by Raphidophyte microalga Heterosigma akashiwo (Hada) was used for detailed characterization. The CDOM was selectively absorbed on HP-20 macroporous resin, and eluted gradiently by methyl alcohol/water to obtain six fractions, which was used for further discussion on photochemistry nature, chemical analysis by UPLC-Q-TOF-MS, and assessment of hematolysis activeness. The results were summarized described as follows:Firstly, the photochemistry nature of six fractions, such as absorption spectrum, fluorescence spectrum and photodegradation was studied. The results showed that as eluent polarity reduced, the average molecular weight of eluate decreased, aromatic substance lessened. There were eight fluorophores in all: three humic-like fluorophores, four protein-like and one pigment-like fluorophores, besides, comparative content of humic-like fluorescence was down, while protein-like's was up. The photodegradation processes of absorption coefficient followed the first-order dynamic equation, and the kinetic constant was about 0.2~0.35.Secondly, six fractions were all analyzed by using Ultra Performance Liquid Chromatography-Electrospray ionization-Quadrupole-Time of Flight Mass Spectrometry (UPLC-ESI-Q-TOF-MS). Method that can isolate various components effectively was established and their average m/z was calculated at 300~700. Efforts were also focused on identifying the main ions of CDOM from heterosigma akashiwo culture, corresponding fragment ions besides. All of these results had provided support for further chemical constitution research.Finally, hemolytic activity of different eluates was determined. The experiment demonstrated that the low polar materials are main factors responsible for hemolysis excreted by heterosigma akashiwo. Utilizing Highly Performance Liquid Chromatography to further separate eluate a5, the fraction had the highest hemolytic degree, finally found two groups of hemolytic compounds in eluate a5: one was homolog with a core structure (m/z 183.01), and it had been speculated as 3-dodecylphenyl hydrogen sulfite; the other was unsaturated fatty acid (UFA), containing octadecenoic C18:1(n-9), octadecadienoic C18:2(n-6), octadecatrienoic C18:3(n-6) and dodecenoic C20:1(n-9) acids.In conclusion, CDOM excreted by Raphidophyte microalga Heterosigma akashiwo (Hada) were mainly low molecular weight materials, and the low polar materials, such as fulvic acids (FA), were lower than the high polar materials of which majority were humic acids (HA). While the hemolytic toxin mainly appeared in the low polar elution group, the structure were respectively speculated as 3-aklyl benzenes hydroxyl sulfurous acid and the unsaturated fatty acid, hemolytic compounds and ultraviolet absorption materials had no correlativity.