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 language | English (US) |
|---|---|
| Pages (from-to) | 47-57 |
| Number of pages | 11 |
| Journal | Journal of Chromatography A |
| Volume | 1383 |
| DOIs | |
| State | Published - Feb 27 2015 |
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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 journal › Article
}
TY - JOUR
T1 - Efficiency of short, small-diameter columns for reversed-phase liquid chromatography under practical operating conditions
AU - Ma, Yan
AU - Chassy, Alexander W.
AU - Miyazaki, Shota
AU - Motokawa, Masanori
AU - Morisato, Kei
AU - Uzu, Hideyuki
AU - Ohira, Masayoshi
AU - Furuno, Masahiro
AU - Nakanishi, Kazuki
AU - Minakuchi, Hiroyoshi
AU - Mriziq, Khaled
AU - Farkas, Tivadar
AU - Fiehn, Oliver
AU - Tanaka, Nobuo
PY - 2015/2/27
Y1 - 2015/2/27
N2 - 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.
AB - 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.
KW - 1mm I.D. column
KW - Column efficiency
KW - Core-shell particles
KW - Extra-column effect
KW - Monolithic silica
KW - Weak-wash solvent
UR - http://www.scopus.com/inward/record.url?scp=84922926766&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84922926766&partnerID=8YFLogxK
U2 - 10.1016/j.chroma.2015.01.014
DO - 10.1016/j.chroma.2015.01.014
M3 - Article
C2 - 25648581
AN - SCOPUS:84922926766
VL - 1383
SP - 47
EP - 57
JO - Journal of Chromatography
JF - Journal of Chromatography
SN - 0021-9673
ER -