A comprehensive course in the systematic development of liquid chromatography separations using QbD principles. Here we review some basic chromatographic measurements – N, α, k, As, TF, Rs – and how these play into a method development strategy based on the fundamental resolution equation. We see how we can quantitatively compare the influence on different separation variables on chromatographic selectivity.
This course is designed for practical workers in the laboratory who have the responsibility for designing new HPLC or UHPLC methods or transferring methods between laboratories. Experienced workers will get the most out of this class, but because it builds from the fundamentals, anyone with experience in HPLC or UHPLC will gain valuable knowledge.
This course covers:
Here we review some basic chromatographic measurements – N, α, k, As, TF, Rs – and how these play into a method development strategy based on the fundamental resolution equation. We see how we can quantitatively compare the influence on different separation variables on chromatographic selectivity. The tradeoffs are examined for prioritizing which variables to use when developing a method.
- Fundamental Measurements
- More About Resolution
- Controlling Selectivity
By attending this online training course you get full access to the 38 video modules and approximately 12.5 hours of instruction. You also get handouts containing copies of all of the approximately 520 PowerPoint slides used in the class. These are arranged for easy note-taking while you view each module and give you a valuable resource for future reference.
This module has a complementary quiz. By correctly answering the quiz questions for all of the modules as part of the course you will be able to download your certificate of completion. Types of questions to expect are:
- What is the difference between differential migration (DM) and band spreading (BS) in a chromatographic separation?
- What is the approximate dead volume of a 150 x 4.6 mm column packed with 5 µm particle?
- What is the approximate retention factor, k, or capacity factor, k’ (within ±0.5 units) if you observe a solvent peak at 2.1 min and a sample peak at 6 min?
- For isocratic separations, what range of k-values is desired?
- What is the closest estimate of the column plate number, N, for a 100 x 2.1 mm column packed with 3 µm diameter particles?
- If two peaks have retention times of 6 and 7 min and the average peak width at baseline is 30 sec, what is the best estimate of resolution?
- When peak tailing (TF or As) exceeds 2, what practical effects on the chromatogram are observed?
- When relating resolution, Rs, to the column plate number, N, how does doubling N affect Rs?
- How does resolution change when the retention factor, k (or capacity factor, k’), is increased?
- When k < 1, what happen to resolution?
- If the width of the front half of a peak is 0.2 min and the back half is 0.4 min, measured at 5% of the peak height, which is the best estimate of the tailing factor, TF?
- what best describes an “orthogonal” separation condition?
- What are the variables that can produce a change in the relative peak positions (selectivity) in a chromatogram for reversed-phase separations?
- What describes the change in separation with a change in the organic solvent composition (%B) in a reversed-phase separation?
- What best describes what you expect to see when the solvent type is changed, such as switching from methanol (MeOH) to acetonitrile (ACN)?
- What is the effect of a change in mobile phase pH in reversed-phase?
Analytical Training Solutions, brought to you by Separation Science, is the leading global portal for fundamentals, best practice, troubleshooting and method development training for chromatographic and mass spectrometric techniques. Comprehensive, self-paced online courses and validated learning provides a unique education resource for analytical chemists. Currently, we offer HPLC training, LC-MS training, GC training and GC-MS training.