HPLC-MS/MS定性分析大菱鲆不同组织中多糖成分

Qualitative Analysis of Polysaccharides from Different Tissues of Turbot (Scophthalmus maximus) by HPLC-MS/MS

为开发大菱鲆功能性多糖成分,采用胰蛋白酶和木瓜蛋白酶双酶法结合碱浸提法对大菱鲆鱼骨、鱼肉、鱼皮3个部位的多糖进行提取与检测,同时将所得多糖使用三氟乙酸水解、1-苯基-3-甲基-5-吡唑啉酮柱前衍生化,利用高效液相色谱-串联质谱对水解产物中的二糖衍生物片段进行分析。结果表明,采用先酶后碱法提取获得鱼骨粗多糖中性糖含量最高为6.79 mg/g干质量,鱼骨硫酸多糖含量最高为3.81 mg/g干质量。通过与标准品的保留时间和质谱数据对比,确定大菱鲆鱼骨中含糖醛酸多糖(uronic acid-containing polysaccharides,UACPs)主要为硫酸软骨素(chondroitin sulfate,CS);鱼皮中主要为透明质酸(hyaluronic acid,HA)和CS;鱼肉UACPs主要有HA、CS、肝素、Glc A(1→2)-Man聚糖以及一种由糖醛酸与己糖连接的重复片段构成的未知多糖UACP1。

Polysaccharides from bone,muscle,and skin of turbot(Scophthalmus maximus)were extracted by sequential hydrolysis with trypsin and papain followed by alkali extraction or the reverse sequence for functional food application.The disaccharide fragments from turbot polysaccharides were analyzed using high performance liquid chromatographytandem mass spectrometry(HPLC-MS/MS)after trifluoroacetic acid(TFA)hydrolysis and1-phenyl-3-methyl-5-pyrazolone(PMP)derivatization.The crude polysaccharides from fish bone produced by enzymatic pretreatment and alkali extraction contained the highest amounts of neutral polysaccharides(6.79mg/g dried weight)and sulfated polysaccharides(3.81mg/g dried weight).The uronic acid polysaccharides(UACPs)from fish bone were identified mainly as chondroitin sulfate(CS)by comparing their retention time and mass spectral data with those of standards,the UACPs from fish mainly included hyaluronic acid and chondroitin sulfate,and the UACPs from fish muscle mainly consisted of hyaluronic acid,heparin,chondroitin sulfate,a polysaccharide comprised of GlcA(1→2)-Man repeated units(AGSP),and an unknown polysaccharide UACP1with repeats of uronic acid linked to hexose.