Abstract
Advancements in column technology resulted in smaller particles and more efficient phases. In parallel, the use of columns with reduced dimensions is becoming more common. This means the effective column volume is also decreased, thereby making the systems more susceptible to effects of band broadening due to extra-column volume. Despite these trends and the fact that a growing number of miniaturized liquid chromatography systems are being offered commercially, manufacturers often stick to the modular concept with dedicated units for pumps, column oven, and detectors. This modular design results in long connection capillaries, which leads to extra-column band broadening and consequently prevents the exploitation of the intrinsic efficiency of state-of-the-art columns. In particular, band broadening post column has a considerable negative effect on efficiency. In this study, mass flow and concentration-dependent detectors were examined for their influence on band broadening using a micro-LC system. A mass spectrometric detector, an evaporative light scattering detector, two UV detectors, and a previously undescribed fluorescence detector were compared. The influence on efficiency is compared using plate height vs linear velocity data and peak variance. It is shown that an increase in the inner diameter after the post-column transfer capillary leads to significant loss in plate height. Comparing the UV detectors, it could be shown that the dispersion was reduced by 38% by the reduction of the post-column volume. The largest variance was found for the evaporative light scattering detector, which was 368% higher compared to the variance of the detector with the least effect on band broadening.
Graphical abstract
Similar content being viewed by others
Abbreviations
- ECBB:
-
Extra-column band broadening
- CFD:
-
Computational fluid dynamics
- ELSD:
-
Evaporative light scattering detector
- FS:
-
Fused silica
References
Vargas Medina DA, Maciel EVS, de Toffoli AL, Lanças FM. Miniaturization of liquid chromatography coupled to mass spectrometry.: 2. Achievements on modern instrumentation for miniaturized liquid chromatography coupled to mass spectrometry. TrAC - Trends Anal Chem. 2020;128:115910. https://doi.org/10.1016/j.trac.2020.115910.
Mejía-Carmona K, da Silva Soares, Burato J, Borsatto JVB, de Toffoli AL, Lanças FM. Miniaturization of liquid chromatography coupled to mass spectrometry: 1. Current trends on miniaturized LC columns. TrAC - Trends Anal Chem. 2020;122:115735. https://doi.org/10.1016/j.trac.2019.115735.
Agrawal A, Keçili R, Ghorbani-Bidkorbeh F, Hussain CM. Green miniaturized technologies in analytical and bioanalytical chemistry. TrAC Trends Anal Chem. 2021;143: 116383. https://doi.org/10.1016/j.trac.2021.116383.
Napolitano-Tabares PI, Negrín-Santamaría I, Gutiérrez-Serpa A, Pino V. Recent efforts to increase greenness in chromatography. Curr Opin Green Sustain Chem. 2021;32: 100536. https://doi.org/10.1016/j.cogsc.2021.100536.
Bian Y, Zheng R, Bayer FP, Wong C, Chang YC, Meng C, Zolg DP, Reinecke M, Zecha J, Wiechmann S, Heinzlmeir S, Scherr J, Hemmer B, Baynham M, Gingras AC, Boychenko O, Kuster B. Robust, reproducible and quantitative analysis of thousands of proteomes by micro-flow LC–MS/MS. Nat Commun. 2020;11:1–12. https://doi.org/10.1038/s41467-019-13973-x.
Vehus T, Roberg-Larsen H, Waaler J, Aslaksen S, Krauss S, Wilson SR, Lundanes E. Versatile, sensitive liquid chromatography mass spectrometry-Implementation of 10 μm OT columns suitable for small molecules, peptides and proteins. Sci Rep. 2016;6:1–10. https://doi.org/10.1038/srep37507.
Hetzel T, vom Eyser C, Tuerk J, Teutenberg T, Schmidt TC. Micro-liquid chromatography mass spectrometry for the analysis of antineoplastic drugs from wipe samples. Anal Bioanal Chem. 2016;408:8221–9. https://doi.org/10.1007/s00216-016-9932-y.
Vanderlinden K, Broeckhoven K, Vanderheyden Y, Desmet G. Effect of pre- and post-column band broadening on the performance of high-speed chromatography columns under isocratic and gradient conditions. J Chromatogr A. 2016;1442:73–82. https://doi.org/10.1016/j.chroma.2016.03.016.
Broeckhoven K, Desmet G. The future of UHPLC: towards higher pressure and/or smaller particles? TrAC Trends Anal Chem. 2014;63:65–75. https://doi.org/10.1016/j.trac.2014.06.022.
Rahimi F, Chatzimichail S, Saifuddin A, Surman AJ, Taylor-Robinson SD, Salehi-Reyhani A. A review of portable high-performance liquid chromatography: the future of the field? Berlin Heidelberg: Springer; 2020.
Lam SC, Coates LJ, Hemida M, Gupta V, Haddad PR, Macka M, Paull B. Miniature and fully portable gradient capillary liquid chromatograph. Anal Chim Acta. 2020;1101:199–210. https://doi.org/10.1016/j.aca.2019.12.014.
Sharma S, Plistil A, Barnett HE, Tolley HD, Farnsworth PB, Stearns SD, Lee ML. Hand-portable gradient capillary liquid chromatography pumping system. Anal Chem. 2015;87:10457–61. https://doi.org/10.1021/acs.analchem.5b02583.
Lankelma J, van Iperen DJ, van der Sluis PJ. Towards using high-performance liquid chromatography at home. J Chromatogr A. 2021;1639: 461925. https://doi.org/10.1016/j.chroma.2021.461925.
Su C-K. Review of 3D-Printed functionalized devices for chemical and biochemical analysis. Anal Chim Acta. 2021;1158: 338348. https://doi.org/10.1016/j.aca.2021.338348.
Balakrishnan HK, Doeven EH, Merenda A, Dumée LF, Guijt RM. 3D printing for the integration of porous materials into miniaturised fluidic devices: a review. Anal Chim Acta. 2021;338796.https://doi.org/10.1016/j.aca.2021.338796.
Desmet G, Broeckhoven K. Extra-column band broadening effects in contemporary liquid chromatography: causes and solutions. TrAC - Trends Anal Chem. 2019;119: 115619. https://doi.org/10.1016/j.trac.2019.115619.
Hetzel T, Loeker D, Teutenberg T, Schmidt TC. Characterization of the efficiency of microbore liquid chromatography columns by van Deemter and kinetic plot analysis. J Sep Sci. 2016;39:3889–97. https://doi.org/10.1002/jssc.201600775.
Chester TL. Recent developments in high-performance liquid chromatography stationary phases. Anal Chem. 2013;85:579–89. https://doi.org/10.1021/ac303180y.
Werres T, Schmidt TC, Teutenberg T. The influence of injection volume on efficiency of microbore liquid chromatography columns for gradient and isocratic elution. J Chromatogr A. 2021;1641.https://doi.org/10.1016/j.chroma.2021.461965.
Taylor G. Dispersion of soluble matter in solvent flowing slowly through a tube. Proc R Soc London Ser A Math Phys Sci. 1953;219:186–203. https://doi.org/10.1098/rspa.1953.0139.
Aris R. On the dispersion of a solute in a fluid flowing through a tube. Proc R Soc London Ser A Math Phys Sci. 1956;235:67–77. https://doi.org/10.1098/rspa.1956.0065.
He B, Tait N, Regnier F. Fabrication of nanocolumns for liquid chromatography. Anal Chem. 1998;70:3790–7. https://doi.org/10.1021/ac980028h.
Filip B, Bochenek R, Baran K, Strzałka D, Antos D. Influence of the geometry of extra column volumes on band broadening in a chromatographic system. Predictions by computational fluid dynamics. J Chromatogr A. 2021;1653:462410. https://doi.org/10.1016/j.chroma.2021.462410.
Gunnarson C, Lauer T, Willenbring H, Larson E, Dittmann M, Broeckhoven K, Stoll DR. Implications of dispersion in connecting capillaries for separation systems involving post-column flow splitting. J Chromatogr A. 2021;1639: 461893. https://doi.org/10.1016/j.chroma.2021.461893.
Desmet G, Cabooter D, Broeckhoven K. Graphical data representation methods to assess the quality of LC columns. Anal Chem. 2015;87:8593–602. https://doi.org/10.1021/ac504473p.
Spaggiari D, Fekete S, Eugster PJ, Veuthey JL, Geiser L, Rudaz S, Guillarme D. Contribution of various types of liquid chromatography-mass spectrometry instruments to band broadening in fast analysis. J Chromatogr A. 2013;1310:45–55. https://doi.org/10.1016/j.chroma.2013.08.001.
Acknowledgements
The authors would like to thank the German Federal Ministry of Economic Affairs and Climate Action for the financial support within the agenda for the promotion of industrial cooperative research and development (IGF) based on a decision of the German Bundestag (IGF – Project No. 19144 N and 20666 N).
Author information
Authors and Affiliations
Contributions
Tobias Werres: conceptualization, investigation, visualization, project administration, writing—original draft; Thorsten Teutenberg: funding acquisition, supervision, writing—review & editing; Torsten C. Schmidt: supervision, writing—review and editing.
Corresponding author
Ethics declarations
Conflict of interest
The authors declare no competing interests.
Additional information
Publisher's note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Below is the link to the electronic supplementary material.
Rights and permissions
About this article
Cite this article
Werres, T., Schmidt, T.C. & Teutenberg, T. Peak broadening caused by using different micro–liquid chromatography detectors. Anal Bioanal Chem 414, 6107–6114 (2022). https://doi.org/10.1007/s00216-022-04170-9
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00216-022-04170-9