Improvenmet and applications of nano-electrospray LC/MS for the microscale structural analysis of glycolipids.

为了简化糖脂的糖类结构分析，对纳米电喷雾LC/MS进行了改进。利用FABMS或SIMS对糖脂糖链进行分析已成为无需MS/MS即可快速获得糖链信息的常用方法。然而，近年来随着离子阱MS等的出现，我们已经能够使用低成本的MS/MS设备。将微尺度LC/MS/MS与纳米电喷雾这一高灵敏的电离方法相结合，将使糖脂含量少、纯度低的糖脂结构分析变得容易。负电喷雾适合于糖脂的高灵敏度分析，因为只有去质子化的分子出现在光谱中。在正离子模式下，分子相关离子分为质子化分子和钠加成离子。卤化物溶剂体系去除了雾滴中的电子，有利于在负极模式下实现稳定的电喷雾。尤其是氯仿是一种方便的溶剂，因为它经常被用作糖脂的层析溶剂。将氨和一甲胺等碱性化合物加入到溶剂中也是有效的，以便有效地形成负离子。在本研究中，精确的LC流量控制很难与纳米电喷雾相结合。直径只有几微米的喷嘴也很难避免堵塞。在实际应用中，直径约50微米的喷嘴适合于微型LC/MS。也证明了微型HPLC需要微型排泄物输送泵，但在目前的情况下，考虑到成本，将使用传统的泵。将喷雾芯片放置在靠近柱出口的位置，以减少洗脱带的扩散。但用硅酸钾可重复形成柱端熔体并非易事。目前，我正在努力克服这个问题。

I tried improvement of nano-electrospray LC/MS with the aim of simplification of the carbohydrate structure analysis of glycoipid. Glycolipid sugar chain analysis by FABMS or SIMS becomes the common method to obtain quickly sugar-chain information without MS/MS.However, we have being able to use low-cost MS/MS equipment by the appearance of ion trap MS, etc. recently. With using the microscale LC/MS/MS combining with nano-electrospray which is the high-sensitive ionization method, the carbohydrate structure analysis will become easy with limited amount and low purified glycolipid. Negative electrospray was suitable for the high sensitive analysis of glycolipid, because only the deprotonated molecule appeared in the spectrum. In the positive mode, the molecular related ions have been divided into protonated molecule and sodium adducted ion. Halide solvent-containing solvent system which removes the electrons in the spray droplet was effective for stable electrospray in the negative mode. Especially, the chloroform was convenient solvent, because it is often used as a chromatography solvent for glycolipids. The addition of the basic compounds such as ammonia and monomethylamine to the solvent was also effective in order to the efficient formation of negative ions. In this study, the precise LC flow control was difficult to attach with the nano-electrospray. There was also difficult to avoid the plugging of the spray nozzle in diameter of few micrometers. The spray nozzle about 50 micrometers in diameter was suitable for micro LC/MS in practice. It was also proven that the micro litter-delivering pump was required for the micro HPLC.In present circumstances, however, the conventional pump would be used concerning the cost. The spray chip was placed close to the column exit for reducing the spread of elution band. But the reproducible formation of column end frit using potassium silicate was not easy. At present time, I am trying to overcome this problem.