Ascorbic acid

HPLC Analysis of Ascorbic and Citric Acids on Amaze TH Mixed-Mode Column
Application description

Ascorbic acid is a vitamin and preservative. Citric acid is a preservative. Both are widely used in food industry. Both acids are polar and acidic in nature. Retention on a regular C18 column can be achieved with addition of ion-pairing reagent. We have developed a simple and robust method for separation of ascorbic and citric acid in HILIC/anion-exchange mode on Amaze TH HPLC column. Method is using volatile mobile phase which is compatible with MS spectrometry. Method can be used for analysis of ascorbic and citric acids in various sample matrices including biofluids and complex food food matrices. Amaze TH column can retain compounds by combination of HILIC, cation- and anion-exchange. Amount of ACN, buffer concentration and buffer pH will affect retention of polar and ionizable compounds.

Conditions of Experiment
Column: Amaze TH
Separation Modes: HILIC, anion-exchange
Column Dimenstions: 4.6 x 150 mm, 5 um, 100A,
Mobile Phase: 75% ACN with 25 mM AmAc pH 4.2
Detection: ELSD
Sample: 0.3 mg/ml
Injection: 3 uL
Flow rate: 1 ml/min
Analytes
Class of compounds: Aromatic acid, Organic acid, Preservative, Supplement, Vitamin
Nature of compounds: Acidic, Hydrophilic, Polar
Compounds: Ascorbic acid, Citric acid
HPLC Analysis of Ascorbic, Methylmalonic and Succinic Acids on Heritage N Column
Conditions of Experiment
Column: Heritage N
Separation Modes: HILIC, anion-exclusion
Column Dimenstions: 4.6 x 150 mm, 5 um, 100A
Mobile Phase: 75% ACN with 15 mM AmAc pH 7
Detection: ELSD
Sample: 0.3 mg/ml
Injection: 5 uL
Flow rate: 1 ml/min
Analytes
Class of compounds: Aromatic acid, Organic acid, Supplement, Vitamin
Nature of compounds: Acidic, Hydrophilic, Polar
Compounds: Ascorbic acid, Methylmalonic acid, Succinic acid
HPLC Analysis of Ascorbic and Dehydroascorbic acids on Coresep SB Column in reversed-Phase Anion-Exchange Modes
Application description

Ascorbic acid, or Vitamin C, is a vitamin found in plants, fruits and other food-related sources. It is one of the main dietary supplements. Vitamin C is an essential nutrient involved in repair of tissues. It is available as generic over-the-counter medication. Dehydroascorbic acid is an oxidized form of ascorbic acid. Both ascorbic and dehydroascorbic acids have similar biological activity as antiviral medications, but dehydroascorbic acid also has neuroprotective effects. Both compounds are very polar, with limited or no retention in reversed-phase chromatography. Both acids were analyzed and retained by weak reversed-phase and anion-exchange mechanisms on a core-shell mixed-mode anion-exchange column. The method and column can be used for analysis of these and other hydrophilic acids by mixed-mode chromatography. The method is compatible with LC/MS, ELSD and CAD and can be used for quantitation of ascorbic and dehydroascorbic acids in various foods, drug formulations and biofluids. Coresep SB column combines unique selectivity of mixed-mode with ultra-high efficiency of core-shell particles technology. HPLC method is reproducible and robust.

Conditions of Experiment
Column: Coresep SB
Separation Modes: reversed-phase and anion-exchange
Column Dimenstions: 3.2 x 100 mm, 2.7 um, 90A
Mobile Phase: 5% ACN with 0.05% of formic acid
Detection: UV 250 nm
Sample: 0.5 mg/ml
Injection: 1 uL
Flow rate: 0.8 ml/mim
Analytes
Class of compounds: Aromatic acid, Organic acid, Supplement, Vitamin
Nature of compounds: Acidic, Hydrophilic, Polar
Compounds: Dehydroascorbic acid, Ascorbic acid