Synthesis And Biochemistry Of Ascorbic Acid Analogues

我们继续进行6-halo-6-脱氧抗坏血酸的合成工作，这些类似物已被证明是运输研究的有用工具，由Mark Levine博士合作完成。6-halo抗坏血酸类似物的还原形式在结构上与抗坏血酸相似，预测它们将由维生素C转运体SVCT1和SVCT2运输。13C核磁共振光谱法检测到的抗坏血酸（DHA）水溶液中氧化的唯一形式是水合双环半晶体，其形成需要C6羟基的存在。这个分子的整体几何形状和多个羟基的存在表明它的结构与葡萄糖的结构相似。这种结构上的相似性在一定程度上导致了近30年前的假设，即DHA和葡萄糖具有相同的运输机制，这一假设随后在几种促进的葡萄糖转运体中得到了验证。当6-溴-6-脱氧- l -抗坏血酸被氧化成6-溴-6-脱氧抗坏血酸（BrDHA）时，由于C-6卤素取代了6-OH基团，水合双环半晶不能形成。2,3-二酮内酯，或者更可能是水合形式，将代表BrDHA在溶液中的结构。这种化合物与葡萄糖缺乏结构相似性，我们预测它不会被GLUTs运输或结合。

We have continued our synthetic work with 6-halo-6-deoxyacorbic acids, analogues that have proven to be useful tools for transport studies, done in a collaborative project by Dr. Mark Levine. The reduced forms of the 6-halo ascorbic acid analogs are structurally similar to ascorbic acid, and it was predicted they would be transported by the vitamin C transporters SVCT1 and SVCT2. The only form of oxidized ascorbic acid (DHA) in aqueous detected by 13C NMR spectroscopy solution is a hydrated bicyclic hemiketal, formation of which requires the presence of the C6 hydroxyl group. The overall geometry of this molecule and the presence of multiple hydroxyl groups has suggested a similarity of this structure to that of glucose. This structural similarity in part led to the hypothesis nearly 30 years ago that DHA and glucose would share the same transport mechanisms, a hypothesis that was subsequently verified for several facilitated glucose transporters. When 6-bromo-6-deoxy-L-ascorbic acid is oxidized to 6-bromo-6-deoxyacorbic acid (BrDHA), the hydrated bicylcic hemiketal cannot form since the C-6 halogen has replaced the 6-OH group. The 2,3-diketolactone, or more likely the hydrated form, would represent the structure of BrDHA in solution. This compound lacks structural similarity to glucose, and we predicted that it would not be transported or bound by GLUTs. When GLUT1 or GLUT3 were expressed in Xenopus oocytes, BrDHA was neither transported nor bound. This contrasted to robust transport of DHA by Xenopus oocytes expressing glucose transporters GLUT1 and GLUT3. In addition, using activated human neutrophils, predicted to have ascorbate accumulation mediated predominantly by DHA and GLUT transporters, 6-bromo-6-deoxy-L-ascorbic acid accumulation was < 1% of accumulation compared to ascorbic acid. We conclude that 6-bromo-6-deoxy-L-ascorbic acid is the first transport substrate identified as completely specific for SVCTs, but not GLUTs, and provides a new strategy to determine the contribution of each pathway to ascorbate accumulation. We are continuing synthetic work to prepare new flavonoid analogues to study sturtural parameters of these inhibitors of ascorbate and glucose transport. Inculded will be potential affinity labels for the transport protein(s). Effects of fluorine substitution biolgical begavior of the flavonoids also will be examined. A series of fluorinated chalcones have been prepared and will be cyclized under conditions that produce the flavonoid structures.