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Non Derivatized Amino Acids
from Synthetic Urine
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printable Application Sheet
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Notes: The determination of amino acids in biological fluids is important in clinical biochemistry and analytical
chemistry. For example measuring methionine concentrations in retinal venous occlusive disease (RVO) is essential.
In addition, increased methionine concentrations have been implicated in a variety of other clinical conditions,
including neural tube defects, spontaneous abortion, placental abruption, osteoporosis, diabetic angiopathy, and
neurological disorders. Methionine is an important amino acid involved in protein synthesis and transmethylation
reactions. It is also the precursor of homocysteine and cysteine, two important risk factors for cardiovascular
diseases. As homocysteine research has gained interest, the evaluation of methionine concentrations in physiological
fluids has acquired importance. Glutamic acid is a major excitatory neurotransmitter in the mammalian central nervous
system. Studies have shown that the changes in the amount of Glu in special brain regions are an indication of
Parkinson’s disease (PD).
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Simple and Easy Analysis of amino acids (underivatized): L–methionine (Met) and L–glutamic acid (Glu)
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% formic acid + 0.005% TFA
B: acetonitrile + 0.1% formic acid + 0.005% TFA |
| Mobile Phase |
Gradient Inverse (ANP) t0 = 1.44 min
| Time |
%B |
Time |
%B |
| 0.0 |
95 |
7.0 |
90 |
| 5.0 |
95 |
9.0 |
90 |
| 6.0 |
90 |
10.0 |
80 |
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| Flow Rate |
0.4 mL/min. |
| Sample |
Synthetic urine |
| Compounds |
1. L – methionine 150 m/z (M+H)+
2. L – glutamic acid 148 m/z (M+H)+ |
| Detection |
ESI – pos - Agilent 6210 MSD TOF mass spectrometer |
Discussion
Amino acid analysis is generally performed by HPLC along with a time consuming pre column derivatization process that is very
tedious and adds cost. Using this new method, the derivatization step is eliminated and the compounds are easily quantified.
L- Glutamic acid is normally an extremely difficult amino acid to analyze due to the strong adsorption by the smallest amount
of residual silanols present on the surface of the columns. And therefore L–glutamic acid is often seen as a very broad peak.
Although we primarily focused on the analysis of amino acids in urine (synthetic or human), the method could also be applied
to the determination of these compounds in other physiological fluids.
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