Knowledge Tools used in Chromatograph
THE growth driver for chromatography/adsorption technology is the creation and availability of novel molecules. An other driver are more stringent conditions as regards food safety and new production methods.
We humans suffers about 7000 different forms of diseases of which approx 1000 have been addressed by pharmaceutical and medtech products. The remaining 6000 diseases may be described as rare or orphane diseases. (Rare diseases affect approx. 1 in 2000 persons). Thus the market volume for novel pharmaceutical is smaller, the opportunity for large profits declining and the pressure for cost reduction growing. Traditional operational managers try to reduce cost by reducing choice of materials to be procured and by automating processes. This strategy works in commodity industries but not in the highly complex live science and health product sector.
In the past 10 years we experienced a decline in output of novel small molecule drugs produced by organic chemistry. In contrast molecules produced by biological means experienced a healthy growth during the same period.. However, since about two years we observe a revival in discovery of small molecules. This should lead in future to a healthy growth in the pharmaceutical sector. This will lead to new job opportunities for chromatographers and pressures to reduce development costs. The most successful way to reduce cost is to increase speed to market. We can assist by providing help to Method Developers and to supply competitive process chromatography system. We are determined to make chromatography a competitive purification technology that can compete with re-crystallisation and distillation.
There are about 4000 different brand of columns/chromatography materials in the market. They survive because everyone of them has unique properties and applications. Since many years various groups are trying to quantifying the “chromatographic properties” of stationary phases. The Tanaka* test is established worldwide as industrial standard test which assesses selectivity and performance differences between HPLC columns. These column parameters should be known for effectively choosing the appropriate HPLC column for a particulate separation and allow comparing columns easily.
A set of seven selected substances is used to describe capacity, hydrophobicity, steric selectivity, and silanophilic properties. To facilitate the illustration and to recognize the quality of a sorbent at one glance, the values of these parameters are outlined on the six axes of a hexagon. The more symmetrical the hexagon appears and the larger its area, the more balanced the stationary phase is in the sum of its chromatographic properties. (* Prof. Tanaka, Kyoto Institute of Technology, J. of Chrom. Sci. 27, 721, 1989). After Tanaka a number of other researcher have advanced tests based on new discoveries in chromatography (See N.S. Wilson, M.D. Nelson, J.W. Dolan, L.R. Snyder, R.G. Wolcott, and P.W. Carr, J. Chromatogr. A 961, 171–193 (2002). and M.R. Euerby and P. Petersson, J. Chromatogr. A 994, 13–36 (2003).) Today the following parameters are considered to be vital in sationary phase characterisation
- HR = hydrophobic retention (retention based on a hydrophobic probe)
- HS = hydrophobic selectivity (ability to discriminate between probes of similar hydrophobicity (hydrophobic selectivity)
- SS = steric selectivity (ability to discriminate between analytes of different shape or hydrodynamic volume (shape or steric selectivity)
- HBC = hydrogen bonding capacity(extent of hydrogen bonding with acids or bases (typically via the silanol surface, polar end capping reagents, or functional groups within the bonded ligand)
- BA = base activity;
- C = chelation;
- IEX = ion-exchange capacity at pH 2.6 ( extent of ion-exchange interactions at low pH 2.8 is typical to differentiate between situations in which surface silanol species will be potentially ionized or ion suppressed)
- IEX = ion-exchange capacity at pH 7.6 (extent of ion-exchange interactions at mid pH 7.0 is typical to differentiate between situations in which surface silanol species will be potentially ionized or ion suppressed)
- AI = acid interaction
The problem today is that chromatography results are also dependent on application requirements (e.g. process vs analytical chromatography) and differences in equipment configurations.
seen as vital factors influencing a parameters Unfortunately, all tests were focussed on small molecules.
•ability to discriminate between probes of similar hydrophobicity (hydrophobic selectivity)
•extent of hydrogen bonding with acids or bases (typically via the silanol surface, polar end capping reagents, or functional groups within the bonded ligand)
The Tanaka Test has become the standart
The first step in Method Development is to
- Characterise your sample molecules (Types, Functionalities, Solubility etc. )
- Define what you aim to achieve.