Elevated Temperatures in Liquid Chromatography, Part I: Benefits and Practical Considerations (original) (raw)
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Journal of Separation Science, 2006
Elevated temperature and temperature programming in conventional liquid chromatography -fundamentals and applications Temperature, as a powerful variable in conventional LC is discussed from a fundamental point of view and illustrated with applications from the author's laboratory. Emphasis is given to the influence of temperature on speed, selectivity, efficiency, detectability, and mobile phase composition (green chromatography). The problems accompanying the use of elevated temperature and temperature programming in LC are reviewed and solutions are described. The available stationary phases for high temperature operation are summarized and a brief overview of recent applications reported in the literature is given.
Journal of Chromatography A, 2007
Theoretical aspects of temperature in liquid chromatography (LC) have mostly been studied to elucidate changes in retention behavior of small and large molecules in various solvents. That temperature also plays a significant role in chromatographic performance is less known. Kinetic plots are an established tool to predict chromatographic performance in terms of speed and efficiency that can be obtained with a certain particle size at the maximum attainable column pressure. In this paper, temperature effects on mobile phase viscosity and analyte diffusion are incorporated in these plots to prove that superior performances are within experimental reach for conventional LC columns and equipment. Verification of the modified kinetic plots with experimental data points is presented.
Enhanced Fluidity Liquid Chromatography for Biological Applications
The following work describes the successful application of enhanced fluidity liquid (EFL) mobile phases to improving isocratic chromatographic separation of biological molecules in hydrophilic interaction liquid chromatography (HILIC) mode. The mobile phase was buffered methanol/water with carbon dioxide added to create the enhanced fluidity liquid. Nucleosides (adenosine, uridine, cytidine, guanosine) were employed as the test sample for this method comparison. Using UV detection at 262 nm, the separation of the sample molecules was studied under each mobile phase condition. Increases in peak resolution between all four were observed as a function of increasing additions of carbon dioxide to create the enhanced fluidity liquid.
Journal of Chromatography A, 2011
In the present work it is shown that the linear elution strength (LES) model which was adapted from temperature-programming gas chromatography (GC) can also be employed for systematic method development in high-temperature liquid chromatography (HT-HPLC). The ability to predict isothermal retention times based on temperature-gradient as well as isothermal input data was investigated. For a small temperature interval of T = 40 • C, both approaches result in very similar predictions. Average relative errors of predicted retention times of 2.7% and 1.9% were observed for simulations based on isothermal and temperature-gradient measurements, respectively. Concurrently, it was investigated whether the accuracy of retention time predictions of segmented temperature gradients can be further improved by temperature dependent calculation of the parameter S T of the LES relationship. It was found that the accuracy of retention time predictions of multi-step temperature gradients can be improved to around 1.5%, if S T was also calculated temperature dependent. The adjusted experimental design making use of four temperature-gradient measurements was applied for systematic method development of selected food additives by high-temperature liquid chromatography. Method development was performed within a temperature interval from 40 • C to 180 • C using water as mobile phase. Two separation methods were established where selected food additives were baseline separated. In addition, a good agreement between simulation and experiment was observed, because an average relative error of predicted retention times of complex segmented temperature gradients less than 5% was observed. Finally, a schedule of recommendations to assist the practitioner during systematic method development in high-temperature liquid chromatography was established.
Some specific problems in the practice of preparative high-performance liquid chromatography
Journal of Chromatography A, 1994
The practice of preparative high-performance liquid chromatography (PLC) is reviewed. Special attention is paid to problems with the use of this method in research and development which are insignificant or unfamiliar on an analytical scale. PLC cohnnn concepts, stability and related packing procedures are discussed. Guidelines to column size selection and optimum use are presented. The paramount importance of high resolution for successful PLC separation is stressed and the effect of friction heat generated by viscous flow on the column performance is described. The significance of sufficient sample solubility in the mobile phase is discussed. Possible deleterious effects of the use of strong solvents with viscosities different from that of the mobile phase are considered. The packing solubility is shown to influence product purity; various product isolation procedures are d&cussed and the use of solid-phase extraction is recommended.
Journal of Pharmaceutical and Biomedical Analysis, 2011
In the pharmaceutical industry fast and efficient separation techniques play an increasing role among analytical methods because the samples to be investigated grow both in complexity and number, and there is an increasing time pressure to complete the analysis. Reducing the analysis time without decreasing the efficiency is possible using higher pressures, elevated temperatures, smaller particle sizes, or a combination of these approaches. Recently developed chromatographic techniques such as the UHPLC (ultra high performance liquid chromatography) and HTLC (high temperature liquid chromatography) are highly promising in meeting these demands. In this study, high temperature liquid chromatography (HTLC) with a zirconia-based column and temperatures elevated up to 150 • C was used. We investigated the chromatographic behaviour of a steroid active pharmaceutical ingredient (levonorgestrel) and its structurally related impurities as model compounds. The effect of the temperature in the range of 50-150 • C and the flow-rate in the range of 0.5-3.0 ml/min, and using methanol as an organic modifier, were studied for optimisation of the separation method.
Journal of Chromatography A, 2008
To fulfil the increasing demand for faster and more complex separations, modern HPLC separations are performed at ever higher pressures and temperatures. Under these operating conditions, it is no longer possible to safely assume the mobile phase fluid properties to be invariable of the governing pressures and temperatures, without this resulting in significantly deficient results. A detailed insight in the influence of pressure and temperature on the physico-chemical properties of the most commonly used liquid mobile phases: water-methanol and water-acetonitrile mixtures, therefore becomes very timely. Viscosity, isothermal compressibility and density were measured for pressures up to 1000 bar and temperatures up to 100 • C for the entire range of water-methanol and water-acetonitrile mixtures. The paper reports on two different viscosity values: apparent and real viscosities. The apparent viscosities represent the apparent flow resistance under high pressure referred to by the flow rates measured at atmospheric pressure. They are of great practical use, because the flow rates at atmospheric pressure are commonly stable and more easily measurable in a chromatographic setup. The real viscosities are those complying with the physical definition of viscosity and they are important from a fundamental point of view. By measuring the isothermal compressibility, the actual volumetric flow rates at elevated pressures and temperatures can be calculated. The viscosities corresponding to these flow rates are the real viscosities of the solvent under the given elevated pressure and temperature. The measurements agree very well with existing literature data, which mainly focus on pure water, methanol and acetonitrile and are only available for a limited range of temperatures and pressures. As a consequence, the physico-chemical properties reported on in this paper provide a significant extension to the range of data available, hereby providing useful data to practical as well as theoretical chromatographers investigating the limits of modern day HPLC.
Analytical Chemistry, 2002
The improvement in the analysis of telechelic polymer matrixes continues to be a pursuit for many scientists of varying disciplines. This quest for a new technique has led to the continued development of liquid chromatography at the critical condition (LCCC) or liquid chromatography at the critical adsorption point (LC-CAP). LCCC allows for the isolation of one area of the polymer matrix so that other areas of the polymer can be probed with sizeexclusion or adsorptive chromatographic modes. Although this technique has been successfully applied to the analysis of telechelic polymers, the practice of LCCC can be difficult. These difficulties include finding and maintaining a solvent system appropriate for the practice of LCCC as well as deterioration of peak shape once the system is operating at the LCCC mode. Because of the specificity of the mobile phase required for the practice of LCCC, the work is routinely practiced by premixing solvents. Previous work with enhanced-fluidity liquid mobile phases demonstrated that these mobile phases removed many of the aforementioned challenges associated with working at the LCCC mode. These mobile phases utilize both pressure and temperature variation in order to maintain the specific solvent strength necessary for the LCCC work. This work studies the coupling and optimization of enhanced-fluidity, EF, liquid mobile phases for LCCC. Several EF-LCCC systems, differing in mobile phase composition, temperature, and pressure, were routinely established, resulting in the effective practice of critical chromatography. The practice of LCCC with on-line mobile phase preparation is demonstrated using commercially available instrumentation. Finally, EF-LCCC is used to analyze triblock and diblock copolymers.