ISC 2016 Short Courses

ISC 800x320 (3)

Short Course 1
– Development and Control of Robust HPLC Methods by Modeling

Imre Molnár, Hans-Jürgen Rieger, Santoso Idris,
Molnár-Institute, Berlin, Germany

13.00 – 13.30     Introduction to UHPLC method development
History of HPLC/UPLC
Fundamentals of RPC, HILIC and other techniques

13.30 – 14.00     Isocratic work, Gradient elution,
Critical Resolution, Principle of Equal Band Spacing
Influence of column dimensions, L, ID, dp,
Instrument parameter: Vd, Vext.col., tG, %B(start), %B(end)

14.00 – 14.30      Terms of Quality by Design: HPLC and Statistics
Column selection and Characterization, Snyder-Dolan-HS-Database
Characterization of Columns by the 3D-critical Resolution Cube’s

14.30 – 15.00       Coffee breaks

15.00 – 15.30      Design of Experiments (DoE) – Modeling UHPLC
1-D (OFAT): %B-, pH-, tG-modeling
2-D: tG-T, tG-pH-models

15.30 – 16.00      3-D-Extension to the Cube: most efficient model
pH-Cube: tG-T-pH-model, and the ternary Cube: tG-T-tC-model

16.00 – 16.30      Dealing with Multifactorial Variabilities:
Robustness studies: Instrument Variabilities
Automated generation of the Cube

16.30 – 17.00     Case Studies of Modeling Separations in the Pharmaceutical Industry
Reducing analysis time from 60 min to 6 min
Reducing analysis time from 160 min to 3 min
Modeling methods for large molecules: Therapeutic Proteins, MAbs, ADC’s
Knowledge Management Documentation


Method Modeling became novadays an important tool in analytical sciences. Since the introduction of QbD in analytical work, trial & error type of method development became obsolete. Robust HPLC methods are today required by regulatory agencies and communication about details of Robustness in HPLC methods need to be easily understandable for companies which produce products in pharma, food and biotech area.

With modeling method development timescale can be shortened by a factor of 5-10.

This tutorial session is dealing with column selection using the Snyder-Dolan Hydrophobicity Subtraction-Database. We will discuss 2- and 3-dimensional retention modeling, gradient elution and robustness issues in HPLC method modeling. Besides of several industrial case studies about pharmaceutical applications for small and large molecules we will talk about aspects of regulatory work and principles of quality by design (QbD) in HPLC method development.

isc button (5)




Short Course 2

– Fundamental and Practical Aspects of Ultra-High-Performance Liquid Chromatography (UHPLC)

Monika Dittmann & Tony Edge
Agilent Technologies


The increasing need for higher speed and resolution in analytical separations drives the trend towards moving from HPLC (ca. 400 bar max. pressure) to UHPLC (>1000 bar).

This tutorial will cover the basic principles of UHPLC and discuss various aspects that need to be considered to successfully apply this technology in method development and routine work.

In the first section the basic theory of mass transfer in columns packed with different particle sizes and morphologies (fully porous, core-shell, monoliths) will be discussed.  It will be demonstrated how the increase in operation pressure and/or temperature can result in enhanced resolution and/or speed of separation.

The second section will be dedicated to optimization strategies for UHPLC separations for different types of applications using the concept of kinetic plots. The kinetic plot (also known as Poppe plot) model uses column efficiency in combination with column permeability to determine the maximum achievable number of plates generated per unit time. This is a much more useful measure for comparing column performance than using column efficiency or plate number alone. The kinetic plot model will be explained in detail and examples for different particle sizes and morphologies will be shown.

The third section will focus on instrumental aspects of UHPLC such as dwell volumes and system dispersion, where the requirements are much more stringent as in HPLC instruments due to the smaller column diameters used in UHPLC. The impact of extra-column volume on the efficiency for small bore columns will be demonstrated for isocratic and gradient separations. Another topic is the transfer of methods between HPLC and UHPLC instruments where differences in system properties (dwell volume and mixing behavior) can lead to changes in retention times and selectivity.

The final section will investigate the effects of temperature on the chromatographic performance criteria. The effect of varying the temperature on the physical properties of the mobile phase will be discussed in relation to the mobile phase viscosity and also the dielectric constant. The session will then move into a discussion of the effects of temperature on the Gibbs free energy of binding. This allows for differences in selectivity for different compounds at different temperatures. The implications of this will be discussed in terms of method development. Finally, the effect that temperature has on the chromatographic efficiency will be discussed.
isc button (5)




Short Course 3
– SFC – Principles, Instrumentation, Method Development, and Applications

Abhijit Tarafdera, Isabelle Francoisb
a Waters Corporation, 34 Maple Street, Milford, MA 01757, USA

b Waters European Headquarter, 5 rue Jacques, Monod, 78280 Guyancourt, France


SFC uses CO2 as the principal mobile phase solvent at pressures generally higher than 100 bar. Apart from this, from operational point of view, there is hardly any difference between SFC and RPLC. Regarding chromatographic capabilities, SFC has a more diverse scope with the types of analytes and selectivities. The recent availability of highly robust analytical SFC systems render the technique more attractive than ever before to a wide range of application areas.

The objective of this short-course is to introduce the students to the capabilities of modern SFC as an analytical instrument and separation device. We will describe in detail – (1) How SFC works (apparatus, detectors and coupling to MS) (2) How SFC compares with LC – focusing on where SFC has unique advantages (high flow rates, coupling achiral + chiral columns etc) and where not (e.g. analyzing large biomolecules), (3) How SFC instrumentation compares with LC, (4) Method development and optimization of (a) achiral SFC, and (b) chiral SFC (5) SFC scale-up and method-transfer, and (6) Sample applications.

The pre-requisite is experience with any chromatographic technique and the principles of chromatography. At the end of the course it is expected that the student will be knowledgeable enough to start working with an SFC instrument and carry out an intended analysis.

isc button (5)