Pharma Science

Practical HPLC for Biopharmaceuticals – Training Module 1: Isocratic HPLC Basics

Although “small molecule” and “biopolymer” separations have traditionally been considered as separate activities, analysts in the biopharmaceutical industry regularly have to deal with both. Fortunately, the underlying principles of chromatography apply equally well in both situations when interpreted appropriately.

You should take this course if you use LC on the job and would like a better understanding of how its various works techniques work and how and when to apply them to the analysis of biopharmaceuticals, “Practical HPLC for Biopharmaceuticals” is the course for you. It’s designed for pharmaceutical scientists who use HPLC as a regular part of their jobs and would like a better grounding in the technique, as well as enhancing knowledge and skills in alternative analytical solutions, and how and when to use them.

Analytical Training Solutions

The course covers:

  • What is HPLC?
    A summary of the history of HPLC, how it developed into the ubiquitous technique we use today, and some of the standard vocabulary and terminology.
  • Separation Chemistry
    An overview of the different chemical mechanisms used in HPLC:
    • Reversed phase
    • Normal phase & HILIC
    • Ion exchange
    • Ion pair
    • Size Exclusion (GFC & GPC)
  • Key Measurements: Retention and Selectivity
    A number of parameters are used to describe an HPLC separation. In this section we will define and explore the significance of k’ (retention) and alpha (selectivity).
  • Key Measurements: Efficiency and Resolution
    A number of parameters are used to describe an HPLC separation. In this section we will define and explore the significance of N (efficiency) and Rs (resolution).
  • Controlling Resolution
    Resolution (Rs) is the key parameter that describes the quality of a separation. In this section we will look at how Rs can be controlled by using k’, alpha, and N
  • Tailing and Fronting
    The Gaussian peak shape is an idealization that is seldom seen in practice. We will explore the standard measurements used to describe deviations from that ideal: the USP “Tailing Factor” and the ASTM “Asymmetry Factor”.

By attending this online training course, You will acquire a good understanding of the practicalities of HPLC and how it can be used in the analysis of biopharmaceuticals. What you learn will demystify the role of liquid chromatography in the analysis of biopharmaceuticals. You’ll find out the various roles that the different separation techniques play in the determination, characterization and quantitation of both small molecules and biopolymers. As well as conventional reversed phase HPLC you’ll will learn about ion-exchange, HILIC, size-exclusion chromatography and hydrophobic interaction chromatography - providing you with an analytical toolbox to apply to your own analytical challenges.

You will be able to answer these questions upon completion of the course:

  • What is HPLC?
  • When was liquid chromatography invented ?
  • What is the basis of Liquid chromatography?
  • What is band broadening in chromatography?
  • What is the key controlling band broadening in chromatography?
  • What control the differential migration in chromatography?
  • What is the basic principle of reversed-phase chromatography?
  • What does retention time used to characterize?
  • What is the symbol for deadtime?
  • How to calculate the deadtime?
  • How to calculate the k' value for the peak?
  • How to calculate the alpha value (relative retention) within two peaks?
  • How to calculate the plate number for a peak?
  • How to calculate the resolution between the peaks?
  • What is the typical "baseline" resolution?
  • what is the resolutions that acceptable for a Tailing Factor?
  • Will the average k' affect the resolution?
  • What is the "rule of three"?
  • How to double resolutions by changing the plate number?
  • What affect selectivity (alpha)?

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