BioProtocols--Met10

Urea

Urea derivative: Urea-TFA

Reagents:

1. Trifluoroacetic anhydride (TFA Supelco 33165-U)

2. Methylene chloride (Dichloromethane, Fisher D151-4)

Procedure:

1. Add 200 ml of methylene chloride and 50 ul of TFA to the urea sample.

2. Incubate at 70 degree centigrade C for 1 hour.

3. Dry sample under nitrogen

4. Add methylene chloride as solvent for GC/MS analysis.

GC/MS:

Instrument: HP 5890 series II GC system is connected to the HP 5971A mass selective detector.

GC column: A 2m x 2mm-glass column was packed with custom packing from Supelco. The specifications for the packing are ethylene glycol adipate (EGA) 0.65% on 80-100 mesh chromosorb WAW.

Inlet Temperature: 200 degree centigrade.

Oven Temperature: 80 degree centigrade for 2 min, then 5 degree centigrade/min ramp to 130 degree centigrade and hold for 1 min. Then 50 degree centigrade/min ramp to 200 degree centigrade and hold for 2 min. Retentiom time: 10.3 min.

Reference: Nissim, I., Yudkoff, M., Yang, W., Teriwilliger, T., Segal, S.(1981) Rapid
Gas Chromatographic-Mass of 15N Urea: Application to Human Metabolic
Studies. Clinica Chimica Acta. Vol. 109. pp295-304.