HPLC Analysis of Glucose and Gluconic Acid on Amaze TH Mixed-Mode Column

Application description

Several techniques are used for the retention and separation of polar and polar ionizable compounds. HILIC mixed-mode proves to be a powerful approach to independently control the elution of ionizable compounds by changing buffer concentration and buffer pH. Glucose is a six member monosaccharide which is polar and neutral in nature. Gluconic acid is polar and an acidic sugar-based compound which is a product of glucose oxidation in nature. Gluconic acid naturally occurs in fruits, honey and wine. It is also used as a food additive to regulate acidity. Gluconic acid mostly exists in a linear form. Gluconic acid is one of the fermentation products in Kombucha tea. We have developed a HILIC/anion-exchange mixed-mode approach for the separation of glucose and gluconic acid on the Amaze TH mixed-mode HPLC column. Glucose is retained by the HILIC mechanism and gluconic acid is retained by HILIC and anion-exchange mechanisms. The retention time is controlled by the amount of ACN, buffer pH and buffer concentration. The change in pH will affect ionization and the polarity of the stationary phase and ionizable analytes. This method is robust and reproducible. It can be used for the analysis of neutral acidic and basic sugar-like molecules in one run. This method is compatible with major detection techniques ( (ELSD, CAD, RI, UV and mass spec)

Conditions of Experiment
Column: Amaze TH
Separation Modes: HILIC, anion-exchnage
Column Dimenstions: 4.6x150 mm, 5 um, 100A
Mobile Phase: ACN/water/AmAc Buffer
Detection: ELSD, 50°C
Sample: 1 mg/ml
Injection: 3 uL
Flow rate: 1 ml/min
Class of compounds: Organic acid, Sugar
Nature of compounds: Acidic, Hydrophilic, Neutral, Polar
Compounds: Glucose, Gluconic acid