Phase ratio Φ for an high performance liquid chromatography (HPLC) column is a parameter defined as the ratio between the volume of the stationary phase Vs and the void volume of the column Vm. Together with the equilibrium constant K of the separation process, phase ratio is part of the retention factor k (k = KΦ). Although a considerable number of studies have been dedicated to the evaluation of Vs and Vm with the goal of obtaining the value for Φ, there are still debatable results regarding the true value of Φ, which for a column with a specific stationary phase may vary with the composition of the mobile phase. One route for the evaluation of the value of Φ uses the measurements of retention factors k on a specific column and mobile phase for two or more hydrocarbons for which the octanol/water partition coefficients log Kow are known. This procedure has been applied in the present study for the evaluation of Φ for three commercially available C18 columns and two mobile phase compositions water/acetonitrile, in the temperature range 20 °C to 50 °C. It was found that phase ratio does change depending on the temperature, its "effective value" decreasing as the temperature increases which is in accordance with the decrease of retention times in reversed phase HPLC when the temperature increases. Besides other factors that may affect the correct calculation of thermodynamic functions, the change of phase ratio with temperature has implications regarding the possibility to calculate the enthalpy and entropy values from van't Hoff plots of the variation of log k as a function of 1/T, even when the retention process is dominated by unique hydrophobic type interactions.

中文翻译：

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反相高效液相色谱法中的相比是否随温度变化？

高效液相色谱（HPLC）柱的相比Φ是定义为固定相Vs的体积与柱Vm的空隙体积之比的参数。相比与分离过程的平衡常数K一起，是保持因子k（k =KΦ）的一部分。尽管已经进行了大量的研究来评估Vs和Vm的目的是获得Φ值，但对于Φ的真值仍有争议性的结果，对于具有特定固定相的色谱柱，该值可能会随流动相的组成。评估Φ值的一种方法是，使用已知的辛醇/水分配系数log Kow的两种或多种烃，在特定色谱柱和流动相上测量保留因子k。在本研究中，该程序已用于在20°C至50°C的温度范围内评估三种市售C18色谱柱和两种流动相组合物水/乙腈的Φ。已经发现，相比确实随温度而变化，其“有效值”随温度升高而降低，这与温度升高时反相HPLC中保留时间的减少相一致。除了可能影响热力学函数正确计算的其他因素外，相比随温度的变化还可能影响根据log k随1 /的函数的van't Hoff图计算焓和熵值的可能性。即使保留过程以独特的疏水型相互作用为主导，也是如此。