| Acidic polysaccharides are important nutritional components of animal derived seafood.In this paper,three kinds of acidic polysaccharides,namely abalone gonad sulfated polysaccharide(AGSP),chondroitin sulfate(CS)and sea cucumber polysaccharide were selected as the research objects,high performance liquid chromatography mass spectrometry(HPLC-MS)was used to analyze their acid degradation products,and the corresponding qualitative and quantitative detection methods were established.The main research contents and results were as follows:In order to analyze the effect of weak acid on the degradation of AGSP,hydrolysates obtained at different trifluoroacetic acid(TFA)concentrations were labelled with 1-phenyl-3-methyl-5-pyrazolone(PMP),and then analyzed by HPLC-MSn.Two disaccharides(DS1 and DS2)and a tetrasaccharide(TS)were found in the AGSP hydrolysates.The two disaccharides were composed of a hexuronic acid linked to a hexose thro ugh a glycosidic bond.DS2 was deduced as ?-GlcA(1?2)-Man,and TS was proposed as a dimmer of DS2.In hydrolysis reaction with 0.1?2.0 mol/L TFA at 100? for 1h,yields of the three oligosaccharides all increased with the raise of acid concentration.The known methods for the identification of CS in complex biological matrices typically time-consuming and labor intensive,requiring isolation and multistep purification.Therefore,a method based on acid hydrolysis followed by PMP-HPLC-MSn analysis has been explored in the present study.Firstly,acid hydrolysates of commercial CS reference standards from shark cartilage and porcine bone were elucidated using HPLC-MSn after PMP derivatization,and altogether 11 di-,tri-and tetra-saccharides with or without sulfate/acetyl groups were identified by their multi-stage mass spectra.Then the trends of reaction yields of these oligosaccharides alone with TFA concentrations(0.1?2.0 mol/L)were investigated,and 0.2-0.8 mol/L TFA was recommended.Finally,using these 11 oligosaccharide derivatives as markers,the identification of CS in sturgeon backbone,porcine trachea and sea cucumber was accomplished after a simple isolation procedure,proving its powerful potent of the present method to profile CS in a complex biological matrix within short time.Sea cucumber polysaccharide is an indispensable nutritional evaluation index of sea cucumber.In order to control and evaluate the quality of sea cucumbers and their related products,a method for quantitative analysis of sea cucumber polysaccharide was established.The quantitative analysis method of fucosylated chondroitin sulfate(FucCS)and fucoidan in sea cucumbers was developed by analyzing CS disaccharide and Fuc produced though acid hydrolysis using PMP-HPLC-MS/MS.The results illustrated that both of CS disaccharide and Fuc had excellent linearities in concentration range of 0.02?0.2 mg/mL.Their relative standard deviations(RSDs)were less than 10%,and the recoveries were 99.2%and 100.2%,respectively.Detected by this method,FucCS and fucoidan contents of fresh sea cucumber were 0.12 mg/g and 0.46 mg/g,while the counterparts of soaked sea cucumber were 0.07 mg/g and 0.29 mg/g,indicating that there was an acidic polysaccharide loss of sea cucumbers in the boiled and rehydrated process.Besides,polysaccharide contents from 9 different sources of fresh sea cucumbers were detected by chemical method,the aquaculture industry standard method and our demonstrated method.The results suggested that there was no significant correlation between the results of each method,and there were some differences in the polysaccharide contents of sea cucumbers from different areas and cultivation methods.This research provides information of acid degradation products from AGSP,CS and sea cucumber polysaccharide,and establishes a faster and more effective analysis method,offering the scientific basis and detection methods for the development and utilization of acidic polysaccharides from marine animals and the quality control of related products. |
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