Steroids Retained and Separated with a Unique Separation Strategy

Polarity of Anti Inflammatory and Birth Control Steroids and their chemical shapes were used to analyze them with HPLC. Retention Time profiles with 3 different Mobile Phases were compared to assess the likely impact of varying Acetonitrile and Methanol ratios in the Mobile Phase. The Organic Solvent percentage and Temperature dependence were studied for optimal Separation.

Temperature changes above 15°C had little effect on the Polarity based, Reversed Phase Selectivity and negative effects on Shape Selectivity. As a result 15°C was used.

The Chromatograms below show that Relative Retention Times (RRT) and Elution Order can vary when the ratios of the Selectivity Mechanisms are modified with Acetonitrile and Methanol changes.

Peaks:
1. Prednisolone
2. Corticosterone
3. Estradiol
4. Ethinyl Estradiol
5. Estrone
6. Norgesterel
7. Progesterone

Method Conditions
Column: Cogent UDC-Cholesterol™, 4μm, 100Å
Catalog No.: 69069-75P
Dimensions: 4.6 x 75mm
Mobile Phase:
Chromatogram A. 56:20:24 DI Water with 0.1% Trifluoroacetic Acid (TFA) / Methanol / Acetonitrile
Chromatogram B. 53:27:20 DI Water with 0.1% Trifluoroacetic Acid (TFA) / Methanol / Acetonitrile
Chromatogram C. 56:24:20 DI Water with 0.1% Trifluoroacetic Acid (TFA) / Methanol / Acetonitrile
Temperature: 15°C
Flow rate: 1mL / minute
Detection: UV @ 240nm

Attachment

No 27 Combining Polarity and Shape Recognition in HPLC pdf 0.2 Mb Download File