Preparation Of Galactan Oligosaccharides Chip And Interaction Of Oligosaccharides With Lectins

In this thesis, seven series of marine galactan-derived oligosaccharides were obtained fromκ-,τ-,λ-carrageenan, agarose, sulfated agarose,desulfatedκ-,λ-carrageenan by mild acid hydrolysis and low pressure gel permeation chromatography （LPGPC）. The sequence of the thirty seven oligosaccharides were determined by electrospray ionization mass chromatography （ESI-MS）, in which nine oligosaccharides were firstly reported. Then, all galactan oligosaccharides were covalently linked to 1,2-dihexadecyl-sn- glycero-3-phospho-ethanolamine （DHPE） to acquire neoglycolipids by a reductive animation reaction. Forty neoglycolipids （NGLs） were prepared, and forteen NGLs were firstly reported.Four methods of preparing glycochip, including micro-glycochip by automatic TLC sampler, micro-glycochip by 8-pin hand-held micoarrayer, mini-glycochip by automatic TLC sampler and glycochip by high-density microarrayer were established. Mini-glycochip was fit for establishing marine glycan chip, while micro-glycochip was fit for optimizing the content of oligosaccharides probes and lectins. Specificity of RCA₁₂₀ and galactan oligosaccharides was estimated by min-glycan chip and ELISA. We firstly found that RCA₁₂₀ can strongly bind to the oligosaccharides who have the sturcture of Galβ1,4anGal-R, and we further confirmed that: 1) the binding ability to RCA₁₂₀ was significantly strengthen by adding a sulfate group at the 6-O- or 2-O-position of non-reducing end galactose; 2) the binding ability to RCA₁₂₀ was abolished by the addition of a sulfate group at the 4-O-position of non-reducing end galactose; 3) the binding ability to RCA₁₂₀ wasn’t affected by substitution ofα-D-Gal at the 3-O-position of non-reducing end galactose, but can be interrupted by adding α-L-anGal. The binding ability of galactan oligosaccharides with RCA₁₂₀ was as following: Gal6Sβ1,4anGal（L）-R > Gal2Sβ1,4Gal2S6S-R > Galβ1,4anGal（L）-R > Galβ1,4GlcNAc-R > Galβ1,4anGal（D）-R > Galβ1,4Gal-R≈Galα1,3Galβ1,4Gal-R > Gal4Sβ1,4anGal（D）-R≈Galβ1,3GlcNAc-R≈Neu5Acα2,6Galβ1,4GlcNAc-R≈anGal（L）α1,3Gal6Sβ1,4anGal（L）-R （R stands for monosaccharide or oligosaccharides）. ECL strongly bound to Galβ1,4GlcNAc structure, but displayed weak affinity with other sulfated oligosaccharides such asκ-,ι-,λ-carrageenan oligosaccharides. ECL only showed weak affinity to Galβ1,4anGal（L or D） structure.Furthermore, the affinity differences of thirty marine galactan oligosaccharides to Galectin-3 was also studied by mini-glycochip technique. Galectin-3 displayed strong binding with odd-numbered agaro-pentasaccharide （Galβ1,4anGal（L）-R） and 6-O-sulfated agarose tri- and pentasaccharides （Gal6Sβ1,4anGal（L）-R）, except for recognize regular Galβ1,4GlcNAc structure.The results indicated that sulfated agarose oligosaccharides could be used as Galectin-3 inhibitors, which was applied to the development of new anticancer drugs.