全面的二维液相色谱(LC×LC)可以为分离复杂样品提供增强的分辨能力和更高的峰容量。从第一维度转移洗脱液强度过高的馏分,然而,可能导致峰变宽。此过程与色谱柱尺寸有关,流速,流动相组成,和固定相兼容性。温度响应型LC(TRLC)使用与二氧化硅偶联的智能聚合物(聚(N-异丙基丙烯酰胺),pNIPAAm),在柱温适度变化时表现出极性变化。因此,保留率是通过温度而不是通过有机溶剂来调节的,允许使用纯水性流动相。由于这些水性流动相在反相柱上显示非常低的洗脱强度,它有助于在TRLC×RPLC的二维柱头处进行波段重新聚焦。TRLC的剩余障碍之一是分析时间长。在这项研究中,该柱组合在分析物重聚焦方面的潜力将被开发。首先,表明,可以在TRLC×RPLC的第一维中实现向上的流量梯度。当第二维中的流量保持在恒定水平时,一维流动梯度不会导致灵敏度受损,并且对所得峰大小没有负面影响。然后,将引入向下温度梯度与向上流动梯度的新颖组合以进一步加速分析。分析时间是,根据使用的方法,减少了36-54%,正如对食品添加剂混合物的分析表明的那样,酚类化合物,和小分子药物模拟杂质分析在0.05%的水平。
Comprehensive two-dimensional liquid chromatography (LC×LC) can provide enhanced resolving power and higher peak capacities for the separation of complex samples. The transfer of fractions of too high eluotropic strength from the first dimension, however, can lead to peak broadening. This process is related to the column dimensions, the flow rates, mobile phase compositions, and stationary phase compatibility. Temperature-responsive LC (TRLC) uses a smart polymer coupled to silica (poly(N-isopropylacrylamide), pNIPAAm), that exhibits a change in polarity upon modest variations in column temperature. Retention is thus modulated by temperature and not by organic solvents, allowing for the use of purely aqueous mobile phases. As these aqueous mobile phases depict a very low eluotropic strength on a reversed-phase column, it facilitates band refocusing at the second-dimension column head in TRLC×RPLC. One of the remaining obstacles of TRLC is the long analysis time. In this research, the potential of this column combination in terms of analyte refocusing will be exploited. First, it is shown that upwards flow gradients can be implemented in the first dimension of TRLC×RPLC. As the flow in the second dimension is maintained at a constant level, a first-dimension flow gradient does not lead to impaired sensitivity and has no negative effects on the resulting peak size. Then, the novel combination of a downwards temperature gradient with an upwards flow gradient will be introduced to speed up the analyses further. Analysis time was, depending on the method used, reduced by 36-54%, as demonstrated by the analysis of mixtures of food additives, phenolic compounds, and small molecule pharmaceuticals mimicking impurity analysis at a 0.05% level.