C7030 Separation methods

Faculty of Science
Autumn 2012
Extent and Intensity
2/0/0. 2 credit(s) (plus extra credits for completion). Recommended Type of Completion: zk (examination). Other types of completion: k (colloquium).
Teacher(s)
doc. Mgr. Jan Havliš, Dr. (lecturer)
Guaranteed by
doc. Mgr. Jan Havliš, Dr.
Department of Biochemistry – Chemistry Section – Faculty of Science
Contact Person: doc. Mgr. Jan Havliš, Dr.
Supplier department: Department of Biochemistry – Chemistry Section – Faculty of Science
Prerequisites
basic knowledge of physical (C4660 basic physical chemistry, C4020 advanced physical chemistry) and analytical chemistry (C3100 analytical chemistry I, C4050 analytical chemistry II) is prerequisited.
Course Enrolment Limitations
The course is offered to students of any study field.
Course objectives
main objectives of lecture: introduction into analytical separation methods; mastering their principles; acquainting with the wide scope of their contemporary implementation and utilisation; definition of terms pre-analytical separation, separations analytical and preparative; choice and optimisation of separation system, evaluation of its efficiency, with validation of separation method and evaluation of the separation results.
Syllabus
  • 1. introduction; separation methods: preparative and analytical; theoretical foundations of separation: molecular equilibrium, intermolecular interactions, influence of entropy.
  • 2. extraction: liquid-liquid extraction (LLE), description of extraction equilibria, influence of analyte-solvent interaction, choice of extraction system, carry-out of extraction; extraction liquid-solid (LSE); description of one component behaviour, repeated extraction, description of multiple component behaviour.
  • 3. preanalytical sample preparation: CCE (counter-current extraction); SFE (supercritical fluid extraction); ASE (accelerated solvent extraction); MASE (microwave assisted extraction), SPE (solid phase extraction), DSPE/MSPE (disperse solid phase extraction), SPME (solid phase microextraction), HSE (head-space extraction).
  • 4. analytical separation; chromatography – theoretical foundations; physico-chemical description of chromatographic processes, graphic presentation of separation, thermodynamic aspects of separation; kinetic aspects of separation (van Deemter equation).
  • 5. study and description of separation; quantitative relations between structure and selectivity in regard to retention; liquid chromatography – history; classification of basic separation mechanisms in LC (LLC, LSC, IC); LC arrangement: mobile phase delivery, elution and elution force (isocratic and gradient); injection devices; separation column, types of stationary phases (particle, monolithic, on-chip), overview of sorbents.
  • 6. detectors (diode array detectors, refractometric, fluorescence, amperometric, conductivity, light-scattering, mass spectrometric); description and evaluation of separation results: defining the chromatographic system, analytical information in chromatogram (qualitative, quantitative); precision, accuracy, limits of analysis (MDL, LOD, LOQ); evaluation of separation efficiency, testing of the system and its “good” behaviour.
  • 7. basic LC modes: CCC (counter-current chromatography), NP-HPLC (normal phase liquid chromatography), RP-HPLC (reverse phase liquid chromatography), TLC (thin-layer chromatography), HPTLC (high performance thin-layer chromatography), UPLC (ultra-high performance chromatography), HTLC (high temperature liquid chromatography), HT-UPLC (elevated-temperature ultra-high performance chromatography), HILIC (hydrophilic interaction chromatography), HIC (hydrophobic interaction chromatography).
  • 8. IEC (ion exchange chromatography) – principles and carry-out, chromatofocusation, affinity chromatography – principles, IMAC (immobilised-metal affinity chromatography); SFC (supercritical fluid chromatography); PC (perfusion chromatography); multi-dimensional chromatography; hyphenated techniques; preparative chromatography – principles and carry-out.
  • 9. development of chromatographic method, criteria and optimisation of separation system; chiral separation: definition of chirality; optical rotatory dispersion; chirality depiction, chiral selectors and their application in separation methods (HPLC).
  • 10. gas chromatography (GC) – history, differences from LC (isotherms, gas compressibility, retention), GC arrangement, mobile phase delivery; injection (splitless, with split); separation column, stationary phase types (WCOT, FSOT, SCOT, PLOT), detectors (flame ionisation, chemiluminiscence, mass spectrometric, etc.).
  • 11. separation of macromolecules: history, theoretical base – description of macromolecule, molecular mass; modes of macromolecule separation – GPC (gel permeation chromatography), carry-out of GPC – columns, detectors (light scattering), HC (hydrodynamic chromatography), FFF (force field fractionation) – principles, instrumentation, modes (SdFFF, ThFFF, EFFF, FFFF).
  • 12. electromigration methods – history, theoretical foundations - separation in external field, separation description – ion mobility, electroosmotic flow, efficiency and influencing factors (heat, diffusion, dispersion); instrumentation – injection, voltage sources, separation channel (capillary, membrane, gel, chip).
  • 13. detectors (photometric, laser induced fluorescence, conductivity detection, mass spectrometry etc.); preparative modes; electrophoretic system definition and analytic information in electrophoretogram; basic modes of electromigration methods – capillary zone electrophoresis (CZE), capillary gel electrophoresis (CGE) slab gel electrophoresis (GE), capillary isoelectric focusation (CIEF), capillary isotachophoresis (CITF), micellar electrokinetic chromatography, thin-layer elektrochromatography, capillary electrochromatography; affinity and non-aqueous electrophoreses; validation of analytical separation method; why and how, elemental terms –method identity, method precision, calibration, selectivity, robustness.
Literature
    recommended literature
  • ŠTULÍK, Karel. Analytické separační metody. 1. vyd. Praha: Karolinum, 2004, 264 s. ISBN 8024608529. info
  • DOLNÍK, Vladislav. Úvod do kapilární elektroforézy. 1994. info
  • CHURÁČEK, Jaroslav. Úvod do vysokoúčinné kapalinové kolonové chromatograrie. Edited by Pavel Jandera. Vyd. 1. Praha: SNTL - Nakladatelství technické literatury, 1984, 188 s. info
Teaching methods
the lecture is based on ppt presentation and its explication. presentation it-self will be available as a study material (black-and-white printable pdf with high resolution and restricted access rights). it is recommended to attend the lecture, because of the explication, which significantly extends the presentation and because there are no available textbooks in czech language covering certain parts of the subject.
Assessment methods
oral examination; students are required to understand and be familiar with the principles and its applications. examination consists of three basic questions, which would be during the examination expanded to let the student demonstrate the extent of topic understanding.
Language of instruction
Czech
Further Comments
The course can also be completed outside the examination period.
The course is taught annually.
The course is taught: every week.
The course is also listed under the following terms Autumn 2010 - only for the accreditation, Autumn 2008, Autumn 2009, Autumn 2010, Autumn 2011, Autumn 2011 - acreditation, Autumn 2013, Autumn 2014, Autumn 2015, Autumn 2016.
  • Enrolment Statistics (Autumn 2012, recent)
  • Permalink: https://is.muni.cz/course/sci/autumn2012/C7030