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Pherogram N ormali z ation for CE and MEKC analyses of bio-matrices PowerPoint Presentation
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Pherogram N ormali z ation for CE and MEKC analyses of bio-matrices

Pherogram N ormali z ation for CE and MEKC analyses of bio-matrices

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Pherogram N ormali z ation for CE and MEKC analyses of bio-matrices

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  1. Pherogram Normalization for CE and MEKCanalyses of bio-matrices Jetse Reijenga1*, Han Martens2, Andrea Giuliani3 and Marcella Chiari3 1. Department of Chemical Engineering and Chemistry, Eindhoven University of Technology, The Netherlands 2.Martens Engineering BV, Heerlen, The Netherlands 3.Institute of Biocatalysis and Molecular Recognition, C.N.R., Milano, Italy

  2. migration time reproducibility, limitations due to: • Different injection volumes • Stacking differences • Non-uniform fieldstrengths • EOF variations • Coating deterioration

  3. reference migration time tiR = LdLt / V(ui + ueof) sample migration time ti = LdLt /  V(ui + ueof + )

  4. calculation of , () Peak 1. LdLt /Vt1 - ueof = LdLt / Vt1R - ueof -  Peak 2. LdLt /Vt2 - ueof = LdLt / Vt2R - ueof -  • = (1/t1 - 1/t2)/(1/t1R - 1/t2R) 2V/LdLt =  = (1/t1 + 1/t2)/ - (1/t1R + 1/t2R)

  5. migration time correction tiR = 1/(1/ti - /2) FIELD shift factor: 1/ EOF shift factor: ti/tiR

  6. CEqualizer Freeware, download from http://www.ceyork.f2s.com

  7. what CEqualizer does: • Load data files (ref + new) • Select identification peaks • Calculate correction factors, • Display FIELD and EOF shift factors • Normalize with respect to reference • Save normalized new data file

  8. Example: DNA ladder

  9. Example: Steroids with sweeping MEKC J.-B. Kim, J.P. Quirino, K. Otsuka, S. Terabe, J. Chromatogr. A 916 (2001) 123

  10. Example: Opioids in urine A.B.Wey, W. Thormann, J. Chromatogr. A 916 (2001) 225

  11. Example: inorganic anions with twin CE detection system R. Bodor, D. Kaniansky, M. Masár, J. Chromatogr. A 916 (2001) 31

  12. Example: β-blockers in large volume stacking CE J.P. Quirino, S. Terabe, Electrophoresis 21 (2000) 355

  13. Example: aromatic amines in large volume stacking CE J.P. Quirino, S. Terabe, Electrophoresis 21 (2000) 355

  14. Conclusions • Diagnose peak shifts samples vs standards • Distinguish Fieldstrength and EOF effects • Normalise sample Electropherograms

  15. Acknowledgements data files of published material were provided by: D. Kaniansky (Bratislava), J.-B. Kim and S. Terabe (Hyogo), J.P. Quirino (Sunnyvale) and W. Thormann (Berne) CEqualizer on http://www.ceyork.f2s.com