1、L/O/G/OStructural confirmation of oligosaccharides newly isolated from suger beet molassesOutlinesBackgroundAchievementExperimentalConclusion&DiscussionBackgroundWhy should we do this experi-ment?raffinose;other oligosaccharides;examined in detailWhat contributions could the study bring?new oligosac
2、charides;structure comfirmed;full assignment of the 1H,13C signals;2D-NMR Four oligosaccharides were isolatedThe structural of them were confirmedAchievement1st The first full assignment of 1H，13C and 2D-NMR of saccharide 14 were carried out2ndExperimental Diluted five times with waterMOLASSESPrepar
3、ative HPLCODS column35water3.5mL/min60 timesDILUTED MOLASSESMeasured with a Refractive Index DetectorConcentrated in vacuo then freeze-driedELUTEDSACCHARIDES POWDERED FRACTIONS I to XII ExperimentalbyHPAEC and TOF-MS analysesFractions I to XIIFractions XI(13.8 mg)Fractions IV(51.6 mg)Fractions IX(24
4、0.0 mg)FractionsII(41.1 mg)Experimentalcarbon-Celite column10%ethanol(1.8 2.4 L)FractionFractionIV IXFractionII SACCHARIDE1carbon-Celite column5%ethanol(0.3-1.1L)carbon-Celite column10%ethanol(2.52.9L)SACCHARIDE2SACCHARIDE3Preparative HPLCrepeated 60 timesrentention time 34.5-35.5FractionXI SACCHARI
5、DE4Experimental1st2nd3rdHPAECSaccharide14 were shown to be homogene-ous,but didnt correspond to any authentic saccharidesTOF-MSThe polymerization was established as 3Acid HydrolyzatesHPLCSaccharides 1&2:Glucose,Fructose,Galactose(1:1:1)3&4:Glucose,Fructose(2:1)Methylation and methanolysis GLCMethyla
6、tion was carried out by Hakomori methodMethanolyzed by heating at 96 for 30min with 1.5%methanolic hydrochloric acidresultsaccharide 31-Kestose1.10 1.32 1-Kestose1.471.06Raffinose2.633.741.453.68 1.29 1.082.55 Methyl-1,3,4,6-tetra-O-methyl-D-fructoside Methyl-2,3,4,6-tetra-O-methyl-D-glucosideMethyl
7、-2,3,4-tri-O-methyl-D-glucoside-D-glucopyranosyl-(1-6)-D-glucopyranosyl-(1 2)-D-fructofuranoside1.08ExperimentalGlcanomeric proton H-1(H 4.98)methylene proton H-6(C 61.25)exhibited correlation peaks to both H-1 and H-2 by HMBC C-3(C 73.8)gave correlation peak to H-1 by COSYH-2(H 3.57)exhibited a cor
8、relation peak to H-1 by HMBCC-5(C 72.63)The remaining methine proton included in the same spin-spinnetwork with H-1 H-4(H 3.44)FruThe characteristic doublet signal in the 1 H-NMR spectrum H-3(H 4.24)exhibited a correlation peak to H-3 by COSY H-4(H4.07)The methylene proton in the samespin-spin netwo
9、rk with H-3,H-4,and H-5 was assigned H-6(H 3.86)gave correlation peak to H-4 by COSYH-5(H 3.91)exhibited a correlation peak to H-3 by HMBC C-1(62.19)The quaternary carbon,which exhibited a correlation peak to H-1 by HMBC C-2(104.57)Saccharide 3ExperimentalThe arrangement of the sugar residuesThe ano
10、meric proton(H-1)of Glc exhibited a correlation peak to the quaternary carbon(C-2)of FruGlc(1-6)to Glc(1 2”)to FruThe anomeric proton(H-1)of Glc exhibited a correlation peak to the methylene carbon(C-6)of GlcSaccharide 3 C H JHHGlc198.994.98d3.8272.283.57dd 9.4,3.8373.83.76dd 10.0,9.4470.293.44dd 10
11、.1,1.0572.633.71ddd10.1,5.1,2.2661.253.86dd 12.4,2.23.78dd 12.4,5.1Glc 192.885.45d3.9271.723.59dd 10.2,3.9373.513.77dd 10.2,9.5470.113.56dd 9.5,8.6572.074.06m666.394.05m3.69dd 12.6,3.0ExperimentalSaccharide 3Conclusion&DiscussionIsolationStructureConfirma-tion Preparative HPLC carbon-Celite column chromatography methylation analysis MALDI-TOF-MS NMR measurementsL/O/G/OThank You!
