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HPLC Application Sheet
Steroid Mixture (1) - Cogent UDC-Cholesterol™

Chromatograms
Steroid Compound Type

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printable Application Sheet
Method Conditions


Mobile Phase  Chromatograms A, B & C
50% Aqueous (0.1% TFA)
50% Acetonitrile (ACN)
Chromatograms D, E & F
45% Aqueous (0.1% TFA)
55% Methanol
Column Cogent UDC-Cholesterol™
Cat. No. 69069-75R
Dimensions 75 x 4.6mm id
Flow Rate 1.0ml/mn
Detection UV, 240nm


Discussion of Chromatograms

Chromatograms A, B & C
Using a Cogent UDC-Cholesterol™ column at 3 different temperatures and with Acetonitrile as an organic modifier, produced similar results (shown) as achieved on 17 different, available C18 columns. Little effect on selectivity and retention time occurred.
Chromatograms D, E & F
When using Methanol as the modifier, the relative retention times differ so much that the elution order is reversed to: Peak 2 eluting first, then Peak 1. A temperature change between 15°C and 45°C has a profound effect on both selectivity and retention times.
Implications
The Cogent UDC-Cholesterol™ column, when used with methanol, has a fundamental selectivity mechanism based on shape recognition. The same column when used with Acetonitrile has the fundamental selectivity mechanism based on polarity recognition (ARP).
Advantages
Using different selectivity mechanisms with the same column can achieve both shape and polarity recognition of some analytes.
Uses
One column can be used to confirm compound purity and identify degradation products.



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