Use of kinetic plots for the optimization of the separation time in ultra‐high‐pressure LC

Abstract

The kinetic-plot approach, in which experimental t(0) and N-values are extrapolated to the performance at maximum system pressure by increasing the column length, was validated by coupling 250×3 mm columns packed with 3 μm particles. The extra-column volume introduced by coupling columns could be neglected with respect to the peak volumes. Plate numbers of up to 132,000 were experimentally achieved by coupling four columns. The maximum deviation between the experimental and predicted plate numbers was 7% for two coupled columns, and decreasing to 0.1% for four coupled columns. Kinetic plots were used to find the conditions to separate a critical pair, with a preset value for the effective plate number, in the shortest possible time. For high-efficiency separations yielding 100,000 effective plates, the optimum critical-pair retention factor was around 4.5. Kinetic plots are presented to find the optimal column length to obtain the fastest possible 100,000 effective-plate separation, taking into account the effect of mobile-phase viscosity on column pressure, and consequently the optimum column length.

Maximizing the peak capacity using coupled columns packed with 2.6μm core–shell particles operated at 1200bar

“…The deviation in retention time was in the order of 2.3%, 5.2% and 0.7% for 2, 3 and 4 coupled respectively. This degree of agreement is in line with that observed in earlier studies [32,33]. Small deviations may be caused by effects of very high pressure, radial and longitudinal frictional heating, changes in viscosity, diffusion coefficients, molar volume, among others.…”

Resolving power in liquid chromatography: A trade‐off between efficiency and analysis time

“…Typical values for α range between 1 and 2, and this corresponds to a variation from 0.001 up to 1 in the term ( α – 1)/ α . The k of the critical pair is generally tuned to reach a value between 3 and 5 (for separations conducted in isocratic mode), which consequently leads to the shortest possible analysis time . This can either be achieved by optimization of the stationary‐phase chemistry, tuning the mobile‐phase composition with respect to elution strength, or a combination of both.…”

Kinetic performance optimisation for liquid chromatography: Principles and practice

“…Perhaps the greatest practical impediment to approaching kinetically optimised performance conditions is the restriction on available column lengths. However, columns can be easily coupled using zero‐dead‐volume connectors, provided the additional extra‐column dispersion (Section 5.1) can be neglected with respect to peak volumes 8. This is illustrated using a practical example of an isocratic separation of a test mixture of diuretic compounds (Fig.…”

“…Alternatively, increasing the pressure limit of the LC system may allow the use of short columns with smaller particles to achieve even faster separations maintaining resolution. For example, 8 describes an isocratic kinetic plot approach to find the conditions (i.e. operating pressure, particle size, column length, and critical pair retention factor) to separate a critical pair in the shortest possible time.…”