Gatschelhofer Christina, Magnes Christoph, Pieber Thomas R, Buchmeiser Michael R, Sinner Frank M
Institute of Medical Technologies and Health Management, Joanneum Research, Auenbruggerplatz 20/3, A-8036 Graz, Austria.
J Chromatogr A. 2005 Oct 7;1090(1-2):81-9. doi: 10.1016/j.chroma.2005.06.098.
Novel monolithic capillary HPLC columns were prepared via ring opening metathesis polymerization (ROMP) within the confines of fused silica columns with 200 microm i.d. using norborn-2-ene (NBE), 1,4,4a,5,8,8a-hexahydro-1,4,5,8, exo, endo-dimethanonaphthalene (DMN-H6) as monomers, 2-propanol and toluene as porogens and RuCl2(PCy3)2(CHPh) as initiator. Using the monolithic capillary HPLC columns, different sets of analytes (i.e. standard systems) were used for the evaluation of the monolithic columns: (i) a protein standard consisting of six proteins in the range of 5000-66 000 g/mol, (ii) an insulin-albumin standard, and (iii) a peptide standard obtained from a tryptic digest of cytochrome C. With these three different standard systems the reproducibility of synthesis in terms of separation performance proved to be 1-2% relative standard deviation in tR. Variation of polymerization parameters had a significant influence on the monolithic morphology and therefore separation efficiency and back pressure. The maximum analytical loading capacity of ROMP-derived monolithic capillary columns for albumin was found to be 30-125 ng, depending on the monomer content. Long-term stability studies showed no alteration in separation performance.
通过开环易位聚合（ROMP），在内径为200微米的熔融石英柱内，以降冰片-2-烯（NBE）、1,4,4a,5,8,8a-六氢-1,4,5,8,外,内-二亚甲基萘（DMN-H6）作为单体，2-丙醇和甲苯作为致孔剂，RuCl2(PCy3)2(CHPh)作为引发剂，制备了新型整体式毛细管高效液相色谱柱。使用这些整体式毛细管高效液相色谱柱，用不同的分析物组（即标准体系）来评估整体柱：（i）一种由六种分子量在5000 - 66000 g/mol范围内的蛋白质组成的蛋白质标准品，（ii）一种胰岛素-白蛋白标准品，以及（iii）一种由细胞色素C胰蛋白酶消化得到的肽标准品。对于这三种不同的标准体系，就分离性能而言，合成的重现性在保留时间方面的相对标准偏差为1 - 2%。聚合参数的变化对整体形态有显著影响，因此对分离效率和背压也有显著影响。发现ROMP衍生的整体式毛细管柱对白蛋白的最大分析负载量为30 - 125 ng，这取决于单体含量。长期稳定性研究表明分离性能没有改变。
