Excellent Retention and Peak Shape with Amide Column

Fluoxetine can have a tendency to tail in some HPLC methods due to its secondary amine group. However, peak shape with the Cogent Amide Column was found to be highly symmetrical without the aid of ion pairing agents. This allows the method conditions to be adapted to LC-MS if needed. With more complex fluoxetine analyses such as those of plasma extracts, LC-MS may be required.

Lot to lot reproducibility of the Cogent Amide Column is highly consistent and reliable. The chromatogram overlay shows results from three different synthesis batches, with a retention time %RSD of 1.2%.


Method Conditions
Column: Cogent Amide™, 4μm, 100Å
Catalog No.: 40036-05P
Dimensions: 4.6 x 50mm
Mobile Phase:
A: 90% DI Water/ 10% Acetonitrile / 0.1% Formic Acid (v/v)
B: Acetonitrile / 0.1% Formic Acid (v/v)

Time (Minutes) %B
0 93
2 93
6 60
7 93

Flow rate: 0.8 mL/minute
Detection: UV 228nm
Injection vol.: 0.5μL
Sample Preparation:
20 mg strength Fluoxetine capsule contents were added to a 50 mL volumetric flask with a portion of 50:50 solvent A:B diluent, sonicated 10 minutes, and diluted to mark. Then an aliquot was filtered through a 0.45 µm Nylon syringe filter (MicroSolv Technology Corp.) and used for injections.

Note: Fluoxetine is a widely prescribed antidepressant which acts by selective inhibition of presynaptic serotonin reuptake. In addition, Fluoxetine can also act as a noncompetitive antagonist of nicotinic acetylcholine receptors. Sold as a racemic mixture, Fluoxetine’s R and S forms show similar efficacy in vivo, and its binding affinity has been shown to be largely stereo independent.


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