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This article is part of the supplement: 30th International Symposium on Intensive Care and Emergency Medicine

Poster presentation

Extraction and purification of 1-hydroxymidazolam glucuronide from ultrafiltrate

C Mckenzie1*, D Naughton2, G Davies3 and D Treacher1

  • * Corresponding author: C Mckenzie

Author Affiliations

1 Guy's and St Thomas' NHS Foundation Trust, London, UK

2 Kingston University, London, UK

3 King's College, London, UK

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Critical Care 2010, 14(Suppl 1):P512  doi:10.1186/cc8744

The electronic version of this article is the complete one and can be found online at: http://ccforum.com/content/14/S1/P512


Published:1 March 2010

© 2010 BioMed Central Ltd.

Introduction

We developed a technique for detection of MDZ and 1-hydroxymidazolam glucuronide (1-OHMG). No commercial source of 1-OHMG prevented assay development. 1-OHMG had pharmacological activity and is renally excreted. We postulated that ultrafiltrate (UFR) would be rich in 1-OHMG. We describe a method to purify 1-OHMG from UFR.

Methods

Ethics approval was granted. To purify, UFR was extracted on a C-18 column, it was washed and eluted. HPLC-MS separation was performed. The 1-OHMG (mol. wt 517/518 in MS) was desiccated and re-dissolved four times to maximise purity. Electrospray (ESI) mass spectrometry (MS) characterised 1-OHMG. The purity was calculated using NMR. A calibration plot of 1-OHMG, MDZ and DZP (internal standard) for HPLC-MS was performed.

Results

1-OHMG identity was confirmed using MS/MS (Figure 1). Two milligrams of 1-OHMG was purified from 5,000 ml UFR. The 1-OHMG was 98% pure (NMR). The extinction coefficient was identical to MDZ. The calibration plot resulted in correlation of 0.912. The assay was applied into clinical practice, to report sera and UFR levels.

thumbnailFigure 1. Centroided MS of purified 1-OHMG.

Conclusions

We were able to extract and purify an active drug metabolite from UFR. Five litres of UFR resulted in 2 mg 1-OHMG. This is a potentially rich source of drugs or drug metabolites, allowing pharmacokinetic studies greatly required in critical illness.

References

  1. McKenzie , et al.: Differentiating MDZ oversedation from neurological damage in the intensive care unit.

    Crit Care 2005, 9:32-33. BioMed Central Full Text OpenURL