IA 3 - HPLC & RP-HPLC at Hochschule Bonn-Rhein-Sieg

Flashcards and summaries for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg

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Study with flashcards and summaries for the course IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Advantages of silica based packing materials (SiO2)

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Disadvantages of silica based packing materials (SiO2)

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Factors influencing mainly k and α (phase equilibrium of analyte)

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Factors influencing N (“column conditions”)

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Systematic HPLC method development

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Totally porous particles (TPP)

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Superficially porous particles (SPP) or core-shell particles

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Perfusion particles

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Ultra high pressure liquid chromatography

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Reducing particle size makes it possible to ...

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

How to prepare a hydrophobic stationary phase for RP-HPLC?

Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

Endcapping of surface silanol groups

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Exemplary flashcards for IA 3 - HPLC & RP-HPLC at the Hochschule Bonn-Rhein-Sieg on StudySmarter:

IA 3 - HPLC & RP-HPLC

Advantages of silica based packing materials (SiO2)
- High mechanical strength allows high operating pressures for long periods of time - Very high column efficiency due to tailor designed totally porous silica particles with a defined range of pore sizes and particle diameters - Chemical surface modification with a wide variety of functional groups - Compatible with aqueous and organic solvents without swelling

IA 3 - HPLC & RP-HPLC

Disadvantages of silica based packing materials (SiO2)
- Limited stability at pH < 3 and at pH > 9 (silica degradation by hydrolysis) Alternative: Polymer based packing materials are compatible with a pH range of 1-13 - Surface silanol groups are acidic (pKa ≈ 4): Basic analytes produce peaks with significant tailing

IA 3 - HPLC & RP-HPLC

Factors influencing mainly k and α (phase equilibrium of analyte)
- Composition of mobile phase - Composition of stationary phase - Temperature (small effect)

IA 3 - HPLC & RP-HPLC

Factors influencing N (“column conditions”)
- Flow rate - Column length - Particle size of stationary phase

IA 3 - HPLC & RP-HPLC

Systematic HPLC method development
- Reduction of k decreases the resolution - Increase of k improves the resolution (but requires higher analysis times) - Increase of α improves the resolution by shifting relative position of peak maxima - Increase of N improves the resolution by producing narrower peaks

IA 3 - HPLC & RP-HPLC

Totally porous particles (TPP)
- most common - most versatile

IA 3 - HPLC & RP-HPLC

Superficially porous particles (SPP) or core-shell particles
- Limited sample load - High plate numbers due to shorter diffusion paths - Sharper peaks provide for higher detection sensitivity

IA 3 - HPLC & RP-HPLC

Perfusion particles
- Large throughpores (400-800 nm) - Diffusion pores (30-100 nm) - Faster mass transfer kinetics for high MW analytes

IA 3 - HPLC & RP-HPLC

Ultra high pressure liquid chromatography
The same column efficiency N can be achieved by shorter columns! Faster separations with smaller particles!

IA 3 - HPLC & RP-HPLC

Reducing particle size makes it possible to ...
... use shorter columns having the same column efficiency

IA 3 - HPLC & RP-HPLC

How to prepare a hydrophobic stationary phase for RP-HPLC?
Reaction of surface silanol groups with chlorosilanes (X = Cl) carrying long alkyl residues R (C8 or C18):

IA 3 - HPLC & RP-HPLC

Endcapping of surface silanol groups
''Endcapping” of accessible unreacted free silanol groups in the bonded phase with short alkyl residues (R = CH3) to reduce surface silanol population

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