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Resolution and signal-to-noise in analysis of carbohydrate isomers by graphitised carbon chromatography with charged aerosol detection
Journal of Chromatography A ( IF 3.8 ) Pub Date : 2018-06-30 , DOI: 10.1016/j.chroma.2018.06.068
Aleksander Lie , Lars Haastrup Pedersen

The effects of co-eluents and additives on separation and signal-to–noise ratio in analyses of monosaccharides by graphitised carbon chromatography (GCC) in combination with charged aerosol detection were studied. Design of experiments was used to model and predict the elution of two monosaccharide isomers, galactose and glucose and the corresponding amine at varying isocratic conditions, including concentration of water-soluble co-eluent, flow and temperature. The study confirmed the well-known order of eluent strength of the co-eluents investigated but showed that the eluent strength of MeOH was significantly lower than that of ACN, and at co-eluent concentrations ≥5% (v/v) IPA approached that of THF. Addition of NH3 increased retention and improved both peak shape and separation but the detector response decreased with increasing NH3 concentration lowering the signal-to-noise ratio. The best combination of response, retention and separation was obtained at 0.1% NH3. Increasing column temperature in the range of 15–50 °C resulted in decreased retention times and resolution. The corresponding Van’t Hoff correlations showed negative adsorption enthalpies indicating an exothermic adsorption process driven by a decrease in entropy minimising the surface energy of the system. Isocratic elution with MeOH as co-eluent offered limited possibilities for optimisation of resolution due to the opposite effects of changes in co-eluent concentration and changes in flow rate. Elution with acetonitrile as co-eluent showed possibilities for optimisation of the resolution within the range of flow rates of 0.6-0.95 mL/min and co-eluent concentrations of 0.1–0.3%, with the highest resolution predicted at 0.1% acetonitrile and a flow rate of 0.81 mL/min. Saccharides in the size range DP1–4, including amino, acetamido, and deoxy sugars, were separated using a binary gradient method. Higher retention was observed for increasing degree of polymerisation (DP) and N-acetylated saccharides were retained longer than non-substituted saccharides of corresponding DP. Partial resolution of two lacto-N-tetrasaccharide positional isomers was obtained.



中文翻译:

石墨化碳色谱和带电气溶胶检测技术分析碳水化合物异构体时的分离度和信噪比

研究了通过石墨化碳色谱(GCC)结合带电气溶胶检测分析单糖时共洗脱液和添加剂对分离和信噪比的影响。实验设计用于模拟和预测两种单糖异构体半乳糖和葡萄糖以及相应的胺在不同等度条件下的洗脱,包括水溶性共洗脱液的浓度,流量和温度。该研究证实了所研究的共洗脱液的洗脱强度众所周知的顺序,但表明MeOH的洗脱强度显着低于ACN,在共洗脱液浓度≥5%(v / v)时,IPA接近THF。NH 3的添加保留增加并改善了峰形和分离度,但检测器响应随NH 3浓度的增加而降低,从而降低了信噪比。在0.1%NH 3下获得响应,保留和分离的最佳组合。色谱柱温度在15–50°C范围内升高,导致保留时间和分离度降低。相应的Van't Hoff相关性显示出负的吸附焓,表明放热的吸附过程是由熵的降低驱动的,从而使系统的表面能最小化。由于共洗脱液浓度变化和流速变化的相反影响,以MeOH作为共洗脱液的等度洗脱为优化分离度提供了有限的可能性。用乙腈作为共洗脱液洗脱,显示了在0.6-0.95 mL / min的流速和0.1-0.3%的共洗脱液浓度范围内优化分离度的可能性,最高分离度预计为0.1%乙腈和流速速度为0.81 mL / min。大小为DP1-4的糖类,包括氨基,乙酰氨基,使用二元梯度法分离糖和脱氧糖。观察到较高的保留以增加聚合度(DP),并且N-乙酰化糖的保留时间比相应DP的未取代糖更长。获得了两种乳酸-N-四糖位置异构体的部分拆分。

更新日期:2018-06-30
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