title: Very high pressure liquid chromatography using fully porous particles: quantitative analysis of fast gradient separations without post-run times.
abstract: Using a column packed with fully porous particles, four methods for controlling the flow rates at which gradient elution runs are conducted in very high pressure liquid chromatography (VHPLC) were tested to determine whether reproducible thermal conditions could be achieved, such that subsequent analyses would proceed at nearly the same initial temperature. In VHPLC high flow rates are achieved, producing fast analyses but requiring high inlet pressures. The combination of high flow rates and high inlet pressures generates local heat, leading to temperature changes in the column. Usually in this case a post-run time is input into the analytical method to allow the return of the column temperature to its initial state. An alternative strategy involves operating the column without a post-run equilibration period and maintaining constant temperature variations for subsequent analysis after conducting one or a few separations to bring the column to a reproducible starting temperature. A liquid chromatography instrument equipped with a pressure controller was used to perform constant pressure and constant flow rate VHPLC separations. Six replicate gradient separations of a nine component mixture consisting of acetophenone, propiophenone, butyrophenone, valerophenone, hexanophenone, heptanophenone, octanophenone, benzophenone, and acetanilide dissolved in water/acetonitrile (65:35, v/v) were performed under various experimental conditions: constant flow rate, two sets of constant pressure, and constant pressure operation with a programmed flow rate. The relative standard deviations of the response factors for all the analytes are lower than 5% across the methods. Programming the flow rate to maintain a fairly constant pressure instead of using instrument controlled constant pressure improves the reproducibility of the retention times by a factor of 5, when plotting the chromatograms in time.
authors.affiliation: Department of Chemistry, University of Tennessee, Knoxville, TN, USA.
authors.forename: Joseph J
authors.initials: JJ
authors.lastname: Stankovich
authors.affiliation: Department of Chemistry, University of Tennessee, Knoxville, TN, USA.
authors.forename: Fabrice
authors.initials: F
authors.lastname: Gritti
authors.affiliation: Department of Chemistry, University of Tennessee, Knoxville, TN, USA.
authors.forename: Paul G
authors.initials: PG
authors.lastname: Stevenson
authors.affiliation: Department of Chemistry, University of Tennessee, Knoxville, TN, USA.
authors.forename: Lois Ann
authors.initials: LA
authors.lastname: Beaver
authors.affiliation: Department of Chemistry, University of Tennessee, Knoxville, TN, USA. Electronic address: guiochon@utk.edu.
authors.forename: Georges
authors.initials: G
authors.lastname: Guiochon
mesh_terms: D000097:Acetonitriles / Q000737:chemistry; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation* / Q000379:methods; D016062:Porosity; D015203:Reproducibility of Results; D013696:Temperature; D013997:Time Factors; D014867:Water / Q000737:chemistry
chemical_list: D000097:Acetonitriles; D014867:Water; C032159:acetonitrile
keywords: Constant flow rate gradient elution; Constant pressure gradient elution; Thermal equilibrium; Very high pressure liquid chromatography (VHPLC)