Beginner's Guide to UPLC

Beginner's Guide to Convergence Chromatography

Beginner's Guide to Size-Exclusion Chromatography

Beginners Guide to Liquid Chromatography

Preparative Liquid Chromatography Primer

Beginner's Guide to Preparative SFC

Practical Approaches to Peptide Isolation

Size-Exclusion Chromatography (SEC) Optimization Guide

The Mass Spectrometry Primer

Beginner's Guide to SPE - Solid-Phase Extraction

SPE - Sample Enrichment and Purification using Solid-Phase Extraction

Beginner's Guide to Preparative SFC

GLOSSARY

Additive – acids, bases and other chemicals added to the co-solvent portion of the mobile phase to improve the chromatography

Back pressure – pressure after the column at the back pressure regulator, it is also the pressure set by the user to control system pressure

Back pressure regulator (BPR) – instrument module used in SFC to control the back pressure on the system at a user determined set point

Chiral – a compound is considered chiral if it has one or more chiral centers resulting in optical isomers or enantiomers that are “mirror images” of each other

Chiral center – typically a carbon (C) atom within a molecule that has four different bonded functional groups

Chiral columns – specific column chemistries required to separate optical isomers (chiral compounds)

Chromophore – molecular group which absorbs light at a particular frequency

Collection timing delay – the time between when the peak exits the detector and reaches the collection selector valve or solenoid

Column oven – oven used in SFC systems for temperature control and column selection

Column volume – volume inside the column “cylinder” based on length and ID minus the approximate volume of the stationary phase

Conditioning solvent – solvent used with the splitter to enhance signal in the MS detector (QDa)

Co-solvent – in SFC, the organic portion of the mobile phase (solvent B)

Critical point – the specific temperature and pressure point for a fluid above which it exists as a supercritical fluid

Density – mass per volume (grams per liter)

Detector channels – single channels that can be monitored independently and in parallel for detection or collection

Diastereomers – compounds that differ in the optical orientation of two chiral centers

Diffusion coefficient – the rate at which a diffusing substance is transported between opposing systems when there is a concentration difference between them

Diffusivity – in chromatography, the ability of a substance to diffuse into and out of the stationary phase particles of the column based on the diffusion coefficient

Dwell volume – the volume from the point of the gradient mixing to the head of the column in a chromatographic system

Efficiency – chromatographic efficiency is measured by the number of theoretical plates or plate height, a low plate height indicates high efficiency and results when a peak has good retention and narrow width

Enantiomers – compounds that differ in the optical orientation of a single chiral center

Extra-column volume – volume of the system between the injector and the detector without the column; impacts peak quality

Extraction – to separate or obtain a product (extract) from a mixture (or matrix) by force

Focused gradient – shallow gradients generated to optimize the separation of a target compound based on the elution percentage from a scouting gradient

Fraction analysis – qualitative and/or quantitative analysis of the collected fractions

Front pressure – pressure before the column in SFC (system pressure)

Gradient methods – chromatography method in which the mobile phase composition changes over the course of a defined period of time

Gradient slope – the rate of change in organic solvent composition per column volume during a gradient

Hydrophilic – miscible with or soluble in water, polar

Hydrophobic – is not miscible with or soluble in water, non-polar

Inlet pressure – pressure of the CO2 before it enters the pump

Isocratic methods – chromatography methods in which the mobile phase composition remains constant over the course of the separation

Joule-Thomson cooling – the cooling of a real gas or liquid when it expands through a valve or opening

Loading – the amount of sample injected onto the chromatographic system

Loading capacity – the amount (mass) of compound that can be loaded based on the dimensions of the column

Lypophilic – lipid soluble, typically used to indicate low polarity

Make-up solvent – solvent used to aid in collection in SFC systems after the CO2 has evaporated

Mass spec detection (MS) – a detection technique that separates compounds based on mass

Mass-flow control – CO2 pumps that control flow based on mass of CO2 (g/min)

Matrix – the substance containing the target compound or isolate

Matrix effects – detection and chromatographic interferences caused by components in the matrix

Mixed-stream injection – injections are performed into the pre-mixed mobile phase

Mobile-phase – the solvent or solvents used to elute compounds from the stationary phase (column)

Modifier – in SFC this is synonymous with co-solvent or the organic portion of the mobile phase

Modifier-stream injection – injections are performed into the co-solvent (modifier) stream before mixing with CO2

Multi-step purification – when purification is accomplished using multiple methods with differing selectivity, whether using two techniques like LC with SFC, or different columns achiral with chiral

Non-polar – describes compounds or solvents that have low polarity, or are hydrophobic and have high LogP values

Normal-phase liquid chromatography (NPLC) – a separation technique in which the mobile phase is typically non-polar and the separation occurs on a stationary phase that is polar

Open-bed collection – collection systems that deposit fractions into tubes or bottles placed in racks in a bed open to the atmosphere

Orthogonal – describes mutually independent or well separated capabilities that span the entire capability range

Phase separation – when a single substance or composition of substances separate into two or more distinct phases

Photodiode array detection (PDA) – a two dimensional UV detection technique where one axis shows time and the other shows UV spectral scans

Polar – describes compounds or solvents that have high polarity, or are hydrophilic and have low LogP values

Pressure drop – the drop in pressure across the column measured by the difference between the front pressure and the back pressure

Productivity – the rate at which the final product can be generated or purified

Purity – a measure of how pure a fraction is based on analytical analysis

Recovery – a measure of the amount of product collected compared to the amount injected or the amount of starting material

Resolution – a measure of the width of two peaks relative to the distance between those peaks

Reversed-phase chromatography (RPLC) – a widely used separation technique in which the mobile phase is polar and the separation occurs on a stationary phase that is non-polar

Scale-up – transferring an analytical scale method to a preparative scale method

Shallow gradient – chromatography method in which the rate of change of organic solvent per column volume is low

Solvating capacity – the amount of compound that can be dissolved per volume of solvent

Split ratio – the ratio of the total preparative flow that is directed to the detectors by the splitter

Splitter – device used send a small ratio of the preparative flow stream to the detectors, accompanied by make-up or conditioning solvent to enhance signal

Stationary phase – the active particles in the column where chromatographic separation occurs

Supercritical fluid – the result when a fluid is above its critical pressure and critical temperature and there is no longer an interface between liquid and gas phases.

System pressure – typically stated as the pump pressure in liquid chromatography, same as front pressure in SFC

Throughput – amount of sample processed and collected per unit time

Van Deemter curves - A plot of plate height vs. average linear velocity of the mobile phase

Viscosity – The degree to which a fluid resists flow under an applied force

Volume-flow control – CO2 pumps that control flow based on volume of CO2 (mL/min)

 

REFERENCES

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    4. M. Rantakyla, Particle Production by Supercritical Antisolvent Processing Techniques, Helsinki University of Technology, Plant Design Report, Series No. 76
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    12. J.D. Pinkston, D.Wen, K.L. Morand, D.A. Tirey, D.T. Stanton, Comparison of LC/MS and SFC/MS for Screening of a Large and Diverse Library of Pharmaceutically Relevant Compounds, Analytical Chemistry, Vol. 78, 2006 (7467-7472)
    13. F. G. Denardin, S. A. B. Vierra de Melo, R. Mammucari, N.R. Foster, Phase transition and volume expansion in CO2 expanded liquid systems, Chemical Engineering Transactions, Vol. 23, 2013 (529-534)
    14. M.K. Tekuri, Solubility & Partition Coefficient: Detailed Review, http://www. slideshare.net/manoj2205/solubility-and-partition-coefficient, July 2014
    15. J. Runco, R. Chen, Diastereoselective separation of permethrin using the ACQUITY UPC2 System, Technology Brief, August 2012, 720004246EN
    16. J.Runco, L. Yang, K. Yu, R. Wang, Y. Li, Z. Wang, F.Li, A. Millar, R. Chen, Qualitative and Quantitative Analysis of Enantiomers (Epigoitrin/Goitrin) from Isatis Indigotica Fort Root Extract Using SFC-MS, Application Note, July 2011, 720004049EN
    17. J. Runco, L. Subbarao, R. Chen, Qualitative and Quantitative Analysis of ß-carotene using UPC2, Application Note, January 2013, 720004550EN
    18. A. Aubin, J. Jablonski, Extraction and Isolation of a Natural Product from Schisandra Berry extract using SFE and SFC, Application Note, July 2015, 720005448EN
    19. A. Aubin, J. Jablonski, Extraction, Purification and Analysis of Dang Gui Root using Supercritical Fluid Techniques, Pittcon Poster, March 2015
    20. J. Jablonski, A. Aubin, Prep 150 LC System: Considerations for Analytical to Preparative Scaling, Application Note, July 2015, 720005458EN
    21. J. Runco, J. McCauley, Chiral purification of Volatile Flavors and Fragrances by SFC, Application Note, August 2014, 720005150EN
    22. C.J. Hudalla, A. Tarafder, J. Jablonski, R. Roshchin, K. Fountain, M. Patel, M. Hardink, 
    23. For more information of useful Application Notes and References, please visit www.waters.com and put the blue reference numbers above in the search box.
    24. T. Yan, F. Riley, UPC2 Strategy for Scaling SFC Methods: Applications for Preparative Chromatography, Application Note, May 2014, 720005064EN