反相液相色谱中生物大分子保留自由能及相比的测定

Determination of the Changes of Free Energy and Column Phase Ratio of Biopolymers in Reversed-Phase Liquid Chromatography

依据液相色谱中溶质的计量置换保留模型 (SDM R)和反相液相色谱 (RPLC)中小分子溶质柱相比的热力学新定义 ,建立了准确测定生物大分子柱相比的理论模式和实验方法 ,准确测定了生物大分子的柱相比 .同时依据计量置换参数logI和Z具有热力学平衡常数的性质 ,并以logI和Z对 1/T作图呈线性为途径 ,解决了生物大分子色谱热力学研究中存在的三个难题 ,准确测定了七种标准蛋白质在RPLC保留过程中的自由能变 。

Based on the stoichiometric displacement model for retention of solute (SDM-R) and the new definition of column phase ratio valid for small solute in reversed-phase liquid chromatography (RPLC), a new definition of the column phase ratio (log(phi)) suitable for biopolymers was presented and determined accurately. In addition, it was found that the linear parameters of logI and Z of biopolymers also had the characteristics of thermodynamical equilibrium constant. The linearity of the plots of logI and Z vs. 1/T was found as good as that for small solutes. With the linear plots of logI and Z vs. 1/T instead of van't Hoff plot in order to solve the three problems of the chromatographic thermodynamics of biopolymers in RPLC, the changes of free energy of seven proteins in RPLC were measured. Compared to the experimental results, it was shown that the contribution of log(phi) to the whole Gibbs' free energy changes of biopolymers during chromatographic process was almost equal to the net changes of Gibbs' free energy of biopolymers.