ANP Retention of a Hydrophobic Compound 

This Application Note shows an example of a hydrophobic analyte retention in Aqueous Normal Phase (ANP / HILIC Like Gradient) HPLC. Although Cyclobenzaprine has a logP of approximately 4.6–4.9, it is well retained with a Symmetrical Peak Shape.

Data from two production lots of Columns is shown to illustrate the Reproducibility of the Method.


Method Conditions
Column: Cogent Diamond Hydride™, 4μm, 100Å
Catalog No.: 70000-7.5P
Dimensions: 4.6 x 75mm
Mobile Phase:
A: DI Water with 0.1% Formic Acid (v/v)
B: Acetonitrile with 0.1% Formic Acid (v/v)

Time (minutes) %B
0 95
1 95
6 40
7 95

Injection vol.: 1μL
Flow rate: 1.0mL / minute
Detection: UV @ 224nm
Sample Preparation: 10mg strength Cyclobenzaprine Tablet was ground and added to a 25mL volumetric flask. A diluent of 50:50 Solvent A / Solvent B was added and the flask was sonicated for 30 minutes. It was then diluted to mark, mixed, and filtered with a 0.45μm Nylon Syringe Filter (MicroSolv Tech Corp.). A portion of the filtrate was diluted 1:10 for HPLC Injections.
t0: 0.9 minutes

Note: Cyclobenzaprine is a muscle relaxant and used to treat Fibromyalgia. Cyclobenzaprine is structurally related to first-generation tricyclic antidepressants. It is marketed under the trade name Flexeril®.


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