HPLC Columns and Trends

5 Some General Guidelines for Bonded Phase Selection 56 

Modern HPLC for Practicing Scientists, by Michael W. Dong Copyright 2006 John Wiley Sons, Inc. 

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Monolithic column — The trend to use shorter columns in liquid chromatography means that the resultant lower separation efficiency is of concern. One way to improve HPLC separation efficiency on a shorter column is to reduce the size of the packing material, but at the cost of increased backpressure. Another approach to improve performance is increasing permeability with a monolithic column. Such a column consists of one solid piece with interconnected skeletons and flow paths. The single silica rod has abimodal pore structure with macropores for through-pore flow and mesopores for nanopores within a silica rod8182 (Figure 12.1). 

The most common HPLC column diameter is 4.6 mm. There is a trend toward narrower columns (2 mm, 1 mm, and capillary columns down to 25 pm) for several reasons. Narrow columns are more compatible with mass spectrometers, which require low solvent flow. Narrow columns require less sample and produce less waste. Heat generated by friction of solvent flow inside the column is more easily dissipated from a narrow column to maintain isothermal conditions. Instruments must be specially designed to accommodate column diameters <2 mm or else band broadening outside the column becomes significant. 

Different developed analytical method are discussed in this chapter related to the determination of illicit substances in blood (either whole blood, plasma, or serum), OF, urine, and hair. These methods take into consideration the particular chemical and physical composition of the matrix and applies each time a suitable pretreatment to remove interfering and matrix effect, to maximize recoveries and to achieve a suitable enrichment if necessary. For liquid matrices the applications of the most common techniques are considered from simple PPT to SPE and LLE the results of recent works from literature are reported and new trends as online SPE, pSPE, automated LLE (SLE) or MAE are examined. Several stationary phases have been shown to be suitable for determination of illicit drugs Cl8, pentafluorophenyl, strong cation-exchange, and HILIC columns. The trend toward fast chromatography is investigated, both UHPLC and HPLC with appropriate arrangements moreover, results obtained with different ion sources, ESI, A PCI, and APPI are compared. 

This chapter provides an overview of modern HPLC equipment, including the operating principles and trends of pumps, injectors, detectors, data systems, and specialized applications systems. System dwell volume and instrumental bandwidth are discussed, with their impacts on shorter and smaller diameter column applications. The most important performance characteristics are flow precision and compositional accuracy for the pump, sampling precision and carryover for the autosampler, and sensitivity for the detector. Manufacturers and selection criteria for HPLC equipment are reviewed. 

Low-dispersion HPLC systems are necessitated by the increasing trend of using shorter and narrower HPLC columns, which are more susceptible to the deleterious effects of extra-column band-broadening. HPLC manufacturers are designing newer analytical HPLC systems with improved instrumental bandwidths compatible with 2-mm i.d. columns by using micro injectors, smaller i.d. connection tubing, and detector flow cells. A new generation of ultra-low dispersion systems dedicated for micro and nano LC is also available. 

The diameter of HPLC columns may vary from several tens of micrometers to several tens of centimeters. Depending on the size of the separation column - nano-, capillary-, narrow-bore - analytical and preparative separations may be distinguished. A current trend in HPLC instrument development provides the hope that in time it may become feasible to use a column of any size with a single instrument. 

These trends toward the use of shorter columns with narrower diameter mean that the overall volume of today s UHPLC columns can be much smaller than the HPLC columns of the past. Consequently, the extra-column band-broadening effect on efficiency can become especially significant in UHPLC as the effect is closely related to column dimensions (14,15). Recently, extra-column band-broadening has been extensively investigated in UHPLC (16-19), conventional HPLC (20), capillary HPLC (21), and modified convenfional HPLC (22). Additionally, the effects of extracolumn volume on other chromatographic parameters including retention factor, selectivity, and pressure drop also become more significant in UHPLC. 

An equally important trend is a reduction in column size and a concomitant increase in analytical speed. Short columns can reduce analysis time, and efficiency may be as high as 5000 theoretical plates since the particle size of the packing material is 3 /u.m. The analysis time is about 1 minute. A standard HPLC column has a diameter of 4.6 mm as compared to 2, 1, or even 0.5 mm for the new columns. The small size of these columns allows the use of more expensive solvents deuterated solvents might be used when it is necessary to analyze the collected samples by NMR. The use of ultramicrobore (50-/u.m diameter) columns allows feeding of the eluted peaks directly into a mass spectrometer of FTIR instrument. 

Another current trend in HPLC development is the use of mini- and microbore columns with small diameters, as well as packed capillaries that require... 

Section II covers the latest trends in reducing sample preparation time, including direct sample infusion/injection and on-line solid phase extraction (SPE). In Section III, we focus on newer trends in stationary phases and how these phases hope to offer different selectivities compared to current CIS-based phases. Section IV briefly provides a few observations on how new detectors are increasing the versatility of HPLC. Finally, in Section V we examine monolithic columns, small particles packed in short columns, high-temperature LC, ultra high-pressure LC, and parallel injection techniques. 

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