HPLC Separation of Dihydroxybenzoic Acid

Separation of hydrophilic isomers is very difficult when only one mechanism of retention, like reversed-phase, is available. Isomers have very similar hydrophobic properties. In case of mixed-mode chromatography, small difference in hydrophobic and ionic properties allows to separate isomers. Isomers of dihydroxybenzoic acids are separated on a Primesep D column with good selectivity and peak shape. This method can be used for separation of other hydrophilic acidic compounds as well as hydrophobic basic compound.


Column Primesep D, 4.6x150 mm, 5 µm, 100A
Mobile Phase MeCN
Buffer TFA
Flow Rate 1.0 ml/min
Detection UV, 250 nm


Class of Compounds Drug, Acid, Hydrophilic, Ionizable, Vitamin, Supplements
Analyzing Compounds 3.4-dihydroxybenzoic acid, 3.5-dihydroxybenzoic acid,2,4-dihydroxybenzoic acid, 2,5-dihydroxybenzoic acid, 2,3-dihydroxybenzoic acid
SIELC Technologies usually develops more than one method for each compound on our website. 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 of interest.
Application Analytes:
2,3-Dihydroxybenzoic Acid
2,4-Dihydroxybenzoic Acid
2,5-Dihydroxybenzoic Acid
3,4-Dihydroxybenzoic Acid
3,5-Dihydroxybenzoic Acid

Application Detection:
UV Detection