HPLC Separation of Antibiotics

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Antibiotics are widely used for treatment and prevention of bacterial infections.  Various antibiotics, particularly those with a sulfanilamide structure, were separated in HPLC using mixed-mode columns with varying strengths of ion-pairing groups.  Primesep 200 has weak acidic ion-exchange pairing groups while Newcrom A has strong acidic ion-exchange groups.  In addition, methanol can be used in the mobile phase on Newcrom A column.  The antibiotics were resolved on both columns with a gradient mobile phase consisting of acetonitrile (ACN) or methanol (MeOH), water and sulfuric acid (H2SO4) buffer.  UV detection at 275nm.

Condition 1

Column Primesep 200, 4.6×100 mm, 5 µm, 100A
Mobile Phase MeCN Gradient
Buffer H2SO4 – 0.5%
Flow Rate 1.0 ml/min
Detection UV 275 nm

Condition 2

Column Newcrom A, 3.2×100 mm, 5 µm, 100A
Mobile Phase MeOH Gradient
Buffer H2SO4 Gradient
Flow Rate  0.5 ml/min
Detection UV 275 nm



Class of Compounds Drugs, Antibiotics
Analyzing Compounds Sulfanilamide, Sulfadiazine, Sulfaguanidine, Dapsone, Sulfaquinoxaline

Application Column

Primesep 200

The Primesep family of mixed-mode columns offers a wide variety of stationary phases, boasting unprecedented selectivity in the separation of a broad array of chemical compounds across multiple applications. Corresponding Primesep guard columns, available with all stationary phases, do not require holders. SIELC provides a method development service available to all customers. Inquire about our specially-tailored custom LC-phases for specific separations.

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Newcrom A

The Newcrom columns are a family of reverse-phase-based columns. Newcrom A, AH, B, and BH are all mixed-mode columns with either positive or negative ion-pairing groups attached to either short (25 Å) or long (100 Å) ligand chains. Newcrom R1 is a special reverse-phase column with low silanol activity.

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Application Analytes:
SIELC Technologies usually develops more than one method for each compound. Therefore, this particular method may not be the best available method from our portfolio for your specific application. Before you decide to implement this method in your research, please send us an email to research@sielc.com so we can ensure you get optimal results for your compound/s of interest.