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Analytical and Bioanalytical Chemistry

, Volume 400, Issue 1, pp 153–164 | Cite as

Reduction of temazepam to diazepam and lorazepam to delorazepam during enzymatic hydrolysis

  • Shanlin Fu
  • Anna Molnar
  • Peter Bowron
  • John Lewis
  • Hongjie Wang
Original Paper

Abstract

It has been previously reported that treatment of urinary oxazepam by commercial β-glucuronidase enzyme preparations, from Escherichia coli, Helix pomatia and Patella vulgata, results in production of nordiazepam (desmethyldiazepam) artefact. In this study, we report that this unusual reductive transformation also occurs in other benzodiazepines with a hydroxyl group at the C3 position such as temazepam and lorazepam. As determined by liquid chromatography-mass spectrometry analysis, all three enzyme preparations were found capable of converting urinary temazepam into diazepam following enzymatic incubation and subsequent liquid–liquid extraction procedures. For example, when H. pomatia enzymes were used with incubation conditions of 18 h and 50 °C, the percentage conversion, although small, was significant—approximately 1% (0.59–1.54%) in both patient and spiked blank urines. Similarly, using H. pomatia enzyme under these incubation conditions, a reductive transformation of urinary lorazepam into delorazepam (chlordesmethyldiazepam) occurred. These findings have both clinical and forensic implications. Detection of diazepam or delorazepam in biological samples following enzyme treatment should be interpreted with care.

Keywords

β-Glucuronidase Temazepam Diazepam Lorazepam Delorazepam Mass spectrometry 

Notes

Acknowledgement

We are indebted to the contribution of Associate Professor Michael Dawson who sadly passed away in December 2010.

Supplementary material

216_2011_4723_MOESM1_ESM.pdf (189 kb)
ESM 1 (PDF 188 kb)

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Copyright information

© Springer-Verlag 2011

Authors and Affiliations

  • Shanlin Fu
    • 1
  • Anna Molnar
    • 1
  • Peter Bowron
    • 2
  • John Lewis
    • 1
  • Hongjie Wang
    • 3
  1. 1.Centre for Forensic ScienceUniversity of Technology, Sydney (UTS)BroadwayAustralia
  2. 2.Toxicology Unit, Pacific Laboratory Medicine ServicesMacquarie HospitalNorth RydeAustralia
  3. 3.National Measurement InstitutePymbleAustralia

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