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1,4-Benzodiazepines in Urine

Reference Number: AA-02318 Created: 05/01/2014 01:58 PM Last Updated: 03/12/2019 04:09 PM

LC-MS method with SPE

 1,4-Benzodiazepinein Urine

Peak: 1. Temazepam 301.0739 m/z [M+H]+

   2. Diazepam 285.0790 [M+H]+

    3. Nordiazepam 271.0633 [M+H]+

    4. Midazolam 326.0855 [M+H]+

     Method Conditions
      Column: Cogent Diamond Hydride 2.0™, 2.2µm, 120Å
      Catalog No.: 70200 -05P-2 
      Dimensions: 2.1 x 50 mm
      Solvents: A: DI  H20  /  0 .1% formic  Acid (v/v)
B: Acetonitrile/ 0.1% formic Acid (v/v)
time (min.)     %B
   0       85
  6       70
  7       20
  9       20
        10     85

      Injection vol.: 1 µL 
      Flow rate: 0.4 ml/min 
ESI – POS  Perki Elme AxIO 2 TOF mass spectrometer
Extraction method: Spike urine  sampl was loaded into SPcartridge I (Clean Screen Xcel purchased from UCT BristolPA, USA) and eluted with 0.78 mL of acetonitrile, 200 microL of 2-propanol, 20 microL of ammonia. After the elution, thsampl wadried unde N2 gas and dissolved in 100 microof 50% methanol / 50% DI H2O / 0.1% formic acid. 

      Before injection, the 10 ppm spiked sample was filtered through a 0 .45µm nylon syringe filter (MicroSolv Tech Corp).


The Cogent Diamond Hydride 2.o¯ column was successfully used in analysis  of 1,4-benzodiazepines in urine  samples  after SPE extraction. Four available compounds were well retained and separated.

The procedure could be used for determination of this class of compounds in urine  samples  and other body fluids.


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