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Organic Acids
An Important Class of Compounds
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printable Application Sheet
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Notes: Chromatogram presented is adapted from: J.J.Pesek, M.T. Matyska, S. M. Fisher, T. R. Sana, Journal of Chromatography A,
1204 (2008) p53.
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Method Conditions
| Column |
Cogent Diamond Hydride™, 4µm, 100A |
| Catalog No. |
70000-15P-2 |
| Dimensions |
2.1 x 150 mm |
| Solvents |
A: DI water + 0.1% ammonium formate
B: 90% acetonitrile/10% DI water/0.1% ammonium formate |
| Gradient |
| Time (min) |
%B |
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Time (min) |
%B |
| 0.0-3.0 |
90.0 |
7.0-7.1 |
to 30 |
| 3.0-6.0 |
to 70 |
7.1-8.0 |
30 |
| 6.0-7.0 |
70 |
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| Post time |
5 min |
| Flow rate |
0.4 mL/min. |
| Sample Peaks |
1. Maleic acid, 115 m/z (M - H)-
2. trans Aconitic acid, 173 m/z (M - H)-
3. cis Aconitic acid, 173 m/z (M - H)-
4. Impurity 173 m/z (M - H)-
5. Fumaric acid, 115 m/z ((M - H)-
6. Citric acid, 191 m/z (M - H)-
7. Oxaloacetic acid, 131 m/z (M - H)-
Sample stock solutions were made in acidified DI water at a
concentration of 0.2 – 0.3 mg/mL. Samples for analysis were made by diluting the stock 1:100 in 50:50 solution A and B.
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| Detection |
ESI – neg - Agilent 6210 MSD TOF mass spectrometer. |
Discussion
Organic acids, an important class of metabolites, are very well retained and separated in this method. Figure A shows the combined EICs
for five small organic acids: maleic, aconitic, fumaric, citric and oxaloacetic acids. Two of these acids are isobaric compounds;
maleic and fumaric acids (115 m/z) and are separated very well. There are three peaks at 173 m/z. The peaks were assigned to trans
(peak 2) and cis (peak 3) isomers of the aconitic acid and an impurity (peak 4). Citric and oxaloacetic acids were not resolved using
the developed mobile phase and gradient, but can be very easy assigned using MS detection. Figure B shows the EIC at 131 m/z for
oxaloacetic acid for integration purpose. This application note is very reproducible for the retention times and also features very
fast equilibration when the gradient is required. This method was repeated for ten consecutive injections and the repeatability was
excellent (%RSD from 0.08 for fumaric acid to 0.26 for trans Aconitic acid).
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