Efficiency of short, small-diameter columns for reversed-phase liquid chromatography under practical operating conditions

Yan Ma, Alexander W. Chassy, Shota Miyazaki, Masanori Motokawa, Kei Morisato, Hideyuki Uzu, Masayoshi Ohira, Masahiro Furuno, Kazuki Nakanishi, Hiroyoshi Minakuchi, Khaled Mriziq, Tivadar Farkas, Oliver Fiehn, Nobuo Tanaka

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

Prototype small-size (1.0mm I.D., 5cm long) columns for reversed-phase HPLC were evaluated in relation to instrument requirements. The performance of three types of columns, monolithic silica and particulate silica (2μm, totally porous and 2.6μm, core-shell particles) was studied in the presence of considerable or minimal extra-column effects, while the detector contribution to band broadening was minimized by employing a small size UV-detector cell (6- or 90nL). A micro-LC instrument having small system volume (<1μL) provided extra-column band variance of only 0.01-0.02μL2. The three columns generated about 8500 theoretical plates for solutes with retention factor, k>1-3 (depending on the column), in acetonitrile/water mobile phase (65/35=vol/vol) at 0.05mL/min, with the instrument specified above. The column efficiency was lower by up to 30% than that observed with a 2.1mm I.D. commercial column. The small-size columns also provided 8000-8500 theoretical plates for well retained solutes with a commercial ultrahigh-pressure liquid chromatography (UHPLC) instrument when extra-column contributions were minimized. While a significant extra-column effect was observed for early eluting solutes (k<2-4, depending on column) with methanol/water (20/80=vol/vol) as weak-wash solvent, the use of methanol/water=50/50 as wash solvent affected the column efficiency for most analytes. The results suggest that the band compression effect by the weak-wash solvent associated with partial-loop injection may provide a practical means to reducing the extra-column effect for small-size columns, while the use of an instrument with minimum extra-column effect is highly desirable.

Original languageEnglish (US)
Pages (from-to)47-57
Number of pages11
JournalJournal of Chromatography A
Volume1383
DOIs
StatePublished - Feb 27 2015

Fingerprint

Liquid chromatography
Reverse-Phase Chromatography
Silicon Dioxide
Methanol
Water
Liquid Chromatography
Ultraviolet detectors
High Pressure Liquid Chromatography
Pressure
Injections
Detectors

Keywords

  • 1mm I.D. column
  • Column efficiency
  • Core-shell particles
  • Extra-column effect
  • Monolithic silica
  • Weak-wash solvent

ASJC Scopus subject areas

  • Analytical Chemistry
  • Organic Chemistry
  • Biochemistry

Cite this

Efficiency of short, small-diameter columns for reversed-phase liquid chromatography under practical operating conditions. / Ma, Yan; Chassy, Alexander W.; Miyazaki, Shota; Motokawa, Masanori; Morisato, Kei; Uzu, Hideyuki; Ohira, Masayoshi; Furuno, Masahiro; Nakanishi, Kazuki; Minakuchi, Hiroyoshi; Mriziq, Khaled; Farkas, Tivadar; Fiehn, Oliver; Tanaka, Nobuo.

In: Journal of Chromatography A, Vol. 1383, 27.02.2015, p. 47-57.

Research output: Contribution to journalArticle

Ma, Y, Chassy, AW, Miyazaki, S, Motokawa, M, Morisato, K, Uzu, H, Ohira, M, Furuno, M, Nakanishi, K, Minakuchi, H, Mriziq, K, Farkas, T, Fiehn, O & Tanaka, N 2015, 'Efficiency of short, small-diameter columns for reversed-phase liquid chromatography under practical operating conditions', Journal of Chromatography A, vol. 1383, pp. 47-57. https://doi.org/10.1016/j.chroma.2015.01.014
Ma, Yan ; Chassy, Alexander W. ; Miyazaki, Shota ; Motokawa, Masanori ; Morisato, Kei ; Uzu, Hideyuki ; Ohira, Masayoshi ; Furuno, Masahiro ; Nakanishi, Kazuki ; Minakuchi, Hiroyoshi ; Mriziq, Khaled ; Farkas, Tivadar ; Fiehn, Oliver ; Tanaka, Nobuo. / Efficiency of short, small-diameter columns for reversed-phase liquid chromatography under practical operating conditions. In: Journal of Chromatography A. 2015 ; Vol. 1383. pp. 47-57.
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abstract = "Prototype small-size (1.0mm I.D., 5cm long) columns for reversed-phase HPLC were evaluated in relation to instrument requirements. The performance of three types of columns, monolithic silica and particulate silica (2μm, totally porous and 2.6μm, core-shell particles) was studied in the presence of considerable or minimal extra-column effects, while the detector contribution to band broadening was minimized by employing a small size UV-detector cell (6- or 90nL). A micro-LC instrument having small system volume (<1μL) provided extra-column band variance of only 0.01-0.02μL2. The three columns generated about 8500 theoretical plates for solutes with retention factor, k>1-3 (depending on the column), in acetonitrile/water mobile phase (65/35=vol/vol) at 0.05mL/min, with the instrument specified above. The column efficiency was lower by up to 30{\%} than that observed with a 2.1mm I.D. commercial column. The small-size columns also provided 8000-8500 theoretical plates for well retained solutes with a commercial ultrahigh-pressure liquid chromatography (UHPLC) instrument when extra-column contributions were minimized. While a significant extra-column effect was observed for early eluting solutes (k<2-4, depending on column) with methanol/water (20/80=vol/vol) as weak-wash solvent, the use of methanol/water=50/50 as wash solvent affected the column efficiency for most analytes. The results suggest that the band compression effect by the weak-wash solvent associated with partial-loop injection may provide a practical means to reducing the extra-column effect for small-size columns, while the use of an instrument with minimum extra-column effect is highly desirable.",
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AU - Morisato, Kei

AU - Uzu, Hideyuki

AU - Ohira, Masayoshi

AU - Furuno, Masahiro

AU - Nakanishi, Kazuki

AU - Minakuchi, Hiroyoshi

AU - Mriziq, Khaled

AU - Farkas, Tivadar

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AU - Tanaka, Nobuo

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