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The CHROMacademy Essential Guide Webcast:
Critical Evaluation of HPLC Methods

Thursday 16th August 2018,
11:00am EDT/ 8:00am PDT/ 4:00pm BST/ 5:00pm CEST



This Essential Guide webcast will evaluate many of the aspects of the HPLC process giving you useful practical information on how to use these parameters to alter your HPLC method, resulting in better and more robust chromatography. 

Presented by

Dr. Dawn Watson (CHROMacademy Technical Expert, Crawford Scientific).

Topics Covered

  • The effect of mobile phase polarity on HPLC separations
  • The role of a buffer in an HPLC separation and how it should be chosen
  • The effect of temperature on analyte retention and selectivity
  • Additives for improved chromatography
  • Stationary phase and support material chemistry
  • The effect of column dimensions on gradient elution
  • Why and when you would use a guard column or in-line filter

Presenter Information »


Critical Evaluation of HPLC Methods tutorial

This month’s webcast focused on the critical evaluation of HPLC methods.  Every facet of the HPLC method can impact your chromatography, therefore, using the CHROMacademy material provided in this month’s tutorial you can fully understand how you can use each parameter of an HPLC method to improve resolution or produce alternate chromatographic selectivity and results.

Our Quick guides, CHROMacademy course content, and archived webcasts and tutorials will provide you with the knowledge you require to apply this powerful analytical technique to your applications.  Our content will provide you with the basic theory behind the separation mechanism, teach you how to alter mobile phases to control retention of ionizable analyte, give you an understanding of the effect of stationary phases on chromatography, and impart practical knowledge on aspects such as column selection.

Watch the webcast
eLearning Modules

Reversed Phase HPLC

Reversed phase HPLC is one of the most utilized modes of chromatography.  This comprehensive module will provide you with the knowledge and understanding you need to apply this method, design or improve mobile phases through solvent choice or pH adjustment, and which parameters can be altered to change selectivity and retention and improve resolution.

Analyte Retention in Reversed Phase HPLC | Changing the Organic Modifier
Reversed Phase HPLC of Ionizable Samples | Controlling the Extent of Ionization
Basic Analytes and Ion Suppression


Column Chemistry

Column stationary phase chemistry has the largest impact on selectivity which in turn affects resolution to the greatest degree.  Understanding the phase chemistries as well as the influence of the packing material can help to produce robust chromatography with optimum resolution.

Chemically Bonded Phases | Surface Treatment - End capping
Reverse Phase Stationary Phases | Silanol and Separation


Mobile Phase Considerations

The chemical nature of the mobile phase can be altered to affect the retention and separation of analyte molecules.  However, there are some physical parameters which must be considered such as, viscosity, UV cut off, and mobile phase preparation (degassing, filtering etc.) in order to obtain successful analyses.

Solvent Type and Selectivity | Optimising Selectivity using Retention
Solvent Miscibility | UV Cut Off


Quick Guides

Why Is pH Important for HPLC Buffers?

The ionization state of an analyte affects its retention in HPLC.  Ionization is controlled by buffers in the mobile phase pH, therefore, understanding the role of pH and how it affects analyte ionization and retention gives you an excellent tool for manipulating separations.

Why Is pH Important for HPLC Buffers? »


Is the Art of HPLC Method Development Dead?

What do you think?  Is there an art to method development?  Read this article and see if you agree or find out if there’s anything you could be doing to improve your method development.

Is the Art of HPLC Method Development Dead? »


How to Choose an HPLC Column

Tony Edge (Agilent Technologies) shares some great general hints and tips for HPLC column selection.

How to Choose an HPLC Column »


HPLC Column Dimensions

HPLC columns are manufactured in a variety of different internal diameter and length combinations, as well as having an assortment of particle sizes.  HPLC column dimensions will affect the efficiency sensitivity, and speed of an analysis. The choice of column dimensions will depend on the chromatographic application; analytical, semi-preparative, preparative, number of analytes in the mixture etc.  However, the dimensions can also be altered to improve chromatography by achieving more efficient, sensitive, and faster analyses. 

HPLC Column Dimensions »


HPLC Mobile Phases - 10 Bad Habits to Avoid

How many of our 10 bad habits are you guilty of?

HPLC Mobile Phases - 10 Bad Habits to Avoid »


What HPLC Operators Need to Know Part I

During this two part event we will look at the entire HPLC separation process from mobile phase selection and preparation, to injection, separation, and detection, from the perspective of the laboratory practitioner. We will provide practical hints and tips for the essential HPLC parameters in order to make your use of this powerful technique straightforward and successful.

What HPLC Operators Need to Know Part I »


What HPLC Operators Need to Know Part II

Part two starts with column selection probably one of the most important choices in any chromatographic method. We will share our insights on how to select the correct column each time covering particle size, morphology, physicochemical properties as well as stationary phase selection.
Finally, we will detail how to utilize and optimize HPLC gradients and review the various detectors available and when and how they are best employed.

What HPLC Operators Need to Know Part II »


Critical Choices in HPLC - Selecting Column Stationary Phase and Dimensions

How do you currently select a column for an HPLC application? Do you just use the one that is on the instrument, root around in an old drawer and hope that you pull out one that works, use your favorite C18 or ODS column, or borrow one from a colleague who’s chromatography is currently working particularly well? These are probably not ideal situations but there will be many of us who are guilty of these exact scenarios. Instead, why not join us for the CHROMacademy Essential Guide webcast on HPLC column selection in which we will look at the available column stationary phases and support materials (including conventional fully porous and core-shell/superficially porous material), column dimensions and the effect that they can have on your chromatography, and how to optimize and improve your current chromatographic separations. This practical guide will give you the tools required to make an informed choice every time you develop a new HPLC method.

Critical Choices in HPLC - Selecting Column Stationary Phase and Dimensions »


Developing Better HPLC (MS) Methods

Where do you start when you want to develop a better HPLC method? Whether you work in a regulated environment or not, setting specifications for your HPLC protocols is a good practice. In this Essential Guide we will set out guidelines for chromatographic parameters such as retention, resolution, and efficiency that can be used to improve your HPLC methods prior to method validation. We will explain how mobile phase design and instrument parameters can be used to achieve improvements in method robustness and what to look for when a method is failing. Practical tips for sample and eluent preparation, and the correct detector settings to use will also be discussed.

Developing Better HPLC (MS) Methods »

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Dr. Dawn Watson CHROMacademy
Technical Expert
Crawford Scientific

Dawn completed her PhD in synthetic inorganic chemistry at the University of Strathclyde, Glasgow, followed by postdoctoral research on small molecule reaction kinetics at Princeton University.

Following several years working for a major instrument manufacturer, she became a technical expert for CHROMacademy in 2013.

She has expertise in various analytical techniques, including HPLC, GC, GC–MS, LC–MS, NMR, and molecular spectroscopy.  As well as numerous wet chemistry and sample preparation techniques.