The　structure　of　heparan　sulfate　determines　its　interaction　with　proteins,　therefore　the　analysis　of　heparan　sulfate　can　help　further　clarify　structure　and　function　relationships.　The　characteristic　fragmentation　pattern　of　twelve　heparan　sulfate　disaccharide　was　discussed　by　electrospray　ionization-ion　trap　mass　spectrometry　(ESI-MSn)　in　negative　mode.　The　results　show　that　the　position　and　quantity　of　sulfate　groups　in　disaccharide　determine　its　stability　in　MS,　moreover,　C-2sulfate　and　C-6　sulfate　in　disaccharide　are　more　stable　than　N-sulfate.　MS2　revealed　that　the　disaccharide　isomers　have　a　characteristic　fragment　pattern.　The　results　may　contribute　to　identify　the　disaccharides　isomer　of　heparan　sulfate,　which　can　provide　theoretical　knowleage　to　identify　the　structural　of　heparan　sulfate.　©,　2015,　Chinese　Society　for　Mass　Spectrometry.　All　right　reserved.