Ultra Performance Liquid Chromatography (UPLC): An Introduction
نویسنده
چکیده
Michael E. Swartz, Ph.D. Principal Scientist, Waters Corporation, Milford, Massachusetts, e-mail [email protected]. igh performance liquid chromatography (HPLC) is a proven technique that has been used in laboratories worldwide over the past 30-plus years. One of the primary drivers for the growth of this technique has been the evolution of packing materials used to effect the separation. The underlying principles of this evolution are governed by the van Deemter equation, which is an empirical formula that describes the relationship between linear velocity (flow rate) and plate height (HETP or column efficiency). Since particle size is one of the variables, a van Deemter curve can be used to investigate chromatographic performance. According to the van Deemter equation, as the particle size decreases to less than 2.5 mm, not only is there a significant gain in efficiency, but the efficiency does not diminish at increased flow rates or linear velocities. By using smaller particles, speed and peak capacity (number of peaks resolved per unit time in gradient separations) can be extended to new limits, termed Ultra Performance Liquid Chromatography, or UPLC. The technology takes full advantage of chromatographic principles to run separations using columns packed with smaller particles and/or higher flow rates for increased speed, with superior resolution and sensitivity. Figure 1 shows a stability indicating assay of five related substances accomplished in under one minute, proving that the resolving power of UPLC is not compromised even at high speed. The current USP lists multiple HPLC methods for the analysis of these same compounds with run times approaching 20 min, with broad, tailed peaks.
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