Synthesis of Saccharide Chain by Using Cells and the Conversion to Functional Polymer

为了建立生物生产大量含gm3型寡糖有价值物质的最佳条件，合成了两种不同位置有叠氮基的乳苷引物，并将其引入B16黑色素瘤细胞。引物I和II分别由12-叠氮-1-十二烷醇和2-叠氮-1-十二烷醇与过乙酸乳糖糖基化和随后的去乙酰化完成。两种引物经糖基化得到gm3型低聚糖衍生物，释放到培养基中。引物II（2-叠氮十二烷基β-乳苷）的糖基化产物量几乎是引物I（12-叠氮十二烷基β-乳苷）的两倍。研究了种子细胞数、引物浓度和培养时间对糖基化效率的影响。本研究结果对利用叠氮多十二烷基乳苷引物和B16细胞批量生产gm3型低聚糖的工艺条件进行了优化。含氟标记的糖引物可作为合成低聚糖的有效支架。本研究表明，含氟糖衍生物实际上可以被细胞吸收，糖链被细胞酶延长，延长的产物被细胞释放到培养基中。用化学方法合成了一种含氟标记的乳糖苷引物6-（全氟己基）己基-4- o -（β- d -半乳糖酰基）-β- d -葡萄糖吡喃苷，并将其引入小鼠B16细胞中，用于低聚糖的合成。B16细胞对引物的吸收导致末端半乳糖残基的唾液化，从而产生与神经节苷脂GM3具有相同聚糖结构的低聚糖。许多氟原子的存在对细胞没有任何不利影响。此外，氟原子的数量并没有形成位阻屏障，相反，它们的存在可能增加了引物的疏水性，增强了膜的渗透性。这种使用含氟标记的引物和细胞的策略可以为通过一种环境友好的方法更容易地制备低聚糖铺平道路，从而消除了大量有机溶剂的使用。少

In order to establish the optimum condition for the bio-production of a large amount of valuable materials containing GM3-type oligosaccharides, two kinds of lactoside primers having azido group at different positions were synthesized and introduced into B16 melanoma cells. The synthesis of primers I and II was accomplished by glycosylation of 12-azido-1-dodecanol and 2-azido-1-dodecanol, respectively, with lactose peracetate and subsequent deacetylation.Both primers were glycosylated to give GM3-type oligosaccharide derivatives which were released to the culture medium. The amount of glycosylated product from primer II (2-azidododecyl β-lactoside) was almost twice as compared with that from primer I (12-azidododecyl β-lactoside). The effects of seeded cell number, primer concentration and length of incubation time on the glycosylation efficiency were also investigated. The results are necessary to optimize the conditions for the mass-production of GM3-type oligosaccharide using azido … More dodecyl lactoside primer and B16 cells. Fluorous-tagged saccharide primers could be viable scaffolds for the synthesis of oligosaccharides. This research demonstrates that a fluorine-containing saccharide derivative could actually be taken up by the cell, the saccharide chain elongated by cellular enzymes, and the elongated product released by the cells to the culture medium.A fluorous-tagged lactoside primer, 6-(perfluorohexyl)hexyl-4-O-(β-D-galactopyranosyl)-β-D-glucopyranoside, was chemically synthesized and introduced in mouse B16 cells to prime oligosaccharide synthesis. Uptake of the primer by B16 cells resulted in the sialylation of the terminal galactose residue to afford an oligosaccharide with the same glycan structure as ganglioside GM3. The presence of many fluorine atoms did not have any adverse effects to the cells. Moreover, the number of fluorine atoms did not pose a steric barrier and instead, their presence possibly increased the hydrophobicity of the primer and enhanced membrane permeability. This strategy of using a fluorous-tagged primer and cells can pave the way for an easier way of preparing oligosaccharides via an environment-friendly approach that eliminates the use of large amounts of organic solvents. Less