|
|
Determination of Xanthine, Uric acid & Hypoxanthine
From Human Fluids
Sample Peak:
- Xanthine (X) 153.04070 m/z
- Uric acid (UA)169.03560 m/z
Unknown A 153.06080 m/z
- Hypoxanthine (HX) 137.04580 m/z
Unknown B 153.06606 m/z
Click here to view
printable Application Sheet
|
Notes: Xanthine oxidase, an enzyme which catalyzes the oxidation of hypoxanthine (HX) to xanthine (X) to uric acid (UA) can be
inhibited by allopurinol and other drugs. Uric acid lowering drugs are used in the treatment of gout and the prevention of
tumor lysis syndrome. High concentrations of UA in blood (hyperuricemia) cause deposition of urate crystals, which could
ultimately result in chronic joint inflammation and renal impairment. The determination of UA has been one of the tests in the
clinical chemistry laboratory performed for patient diagnosis of gout.
|
|
Method Conditions
| Column |
Cogent Diamond Hydride™, 4µm, 100A |
| Catalog No. |
70000-10P-2 |
| Dimensions |
2.1 x 100 mm |
| Solvents |
A: DI water/0.1% formic acid
B: Acetonitrile + 0.1% formic acid |
| Gradient |
| Time |
%B |
Time |
%B |
| 0.0 |
95 |
9.0 |
80 |
| 0.2 |
95 |
10.0 |
50 |
| 8.0 |
80 |
12.0 |
50 |
|
| Post time |
5 min |
| Flow rate |
0.4 mL/min. Injection Volume: 2µL |
| Sample |
Human Urine |
| Sample Prep |
400 microL of acetonitrile was added to 100 microL of human
urine and sample was centrifuged (3000 g). Next 20 microL of
the supernatant was mixed with 10 microL of the 50% acetonitrile/50% DI water + 0.1% formic acid.
|
| Detection |
ESI – pos - Agilent 6210 MSD TOF mass spectrometer. |
Discussion
A simple ANP method was developed for the determination of xanthine (X), uric acid (UA), and hypoxanthine (HX) at concentrations in
human urine (can be used for human serum) to support pharmacodynamic (PD) studies of a novel xanthine oxidase inhibitor during its
clinical development. PD biomarkers (UA, X, and HX) were well separated from each other. In addition xanthine was separated from two
isobaric unknowns (unknown A and B) present in this particular urine sample. Current HPLC methods for UA/X/HX measurements suffer from
low sensitivity, poor selectivity, and/or inefficient sample throughput. The developed ANP method is fast and sensitive and it will
allow high sample throughput.
|
