Comprehensive Toxicological screening with Symbiosis Pico and Q-TOF mass spectrometry

The Institute of Toxicology in the Clinical Toxicology and Poison Information Centre Berlin (BBGes) has developed a novel system for routine toxicological screening in close collaboration with Spark Holland.

Toxicological screening is challenging because fast and accurate identification of the intoxicant is required in complex biological matrices. HPLC combined with photo diode array detection (DAD) such as the REMEDi ™ HS drug profiling system from BioRad, has been a popular platform for many years, but the combination of liquid chromatography and quadrupole-time-of-flight mass spectrometry instruments with library searching is of increasing importance as a alternative or additional technology. On the sample prep side, on-line Solid Phase Extraction has emerged as a technology that improves both the level of automation and the level of confidence for toxiclogical screening, reducing human error and improving robustness, speed and sample tracking. Using such state-of-the-art technology, the Institute of Toxicology in the Clinical Toxicology and Poison Information Centre Berlin (BBGes) has developed a novel system for routine toxicological screening in close collaboration with Spark Holland. The objective was to develop a reliable and fully-automated on-line extraction screening method (XLC-QqTOF) for the detection of unknown xenobiotics in human plasma/serum samples and to employ platform-independent low/high resolution mass spectral reference libraries and high mass resolution spectra to produce a comprehensive report. For compound identification a new library search and identification algorithm for Q-TOF instruments was developed based on SmileMS library search software from GeneBio. Geneva.

From sample intake to reporting within half an hour

The workflow of the final method consists of 5 minutes (manual) sample pre-treatment, 17 minutes SPE-LC-MS/MS run time and 5 minutes data analysis and reporting. More than one hundred intoxications have been analyzed using the developed method. Besides the detection of basic compounds, neutral and acidic compounds were also identified. The use of a fresh SPE-cartridge for every analytical run effectively prevented cross-contamination between samples. The analytical column with 5µm particle size of offers stable and reproducible chromatography (RT`s, batch-to-batch RT`s(CV): <5%. Using the ABSciex QStar, the mass accuracy was <+/- 5 ppm and LOD`s ranged from 0.002 mg/L (e.g. buspirone, mirtazapine) to 0.050 mg/L (e.g. bromazepam, benzoylecognine, oxazepam). After the extraction of wiff-files from Analyst in SmileMS-compatible mzXML-files, all matches of the library search are tabulated together within a reconstructed and color-coded chromatogram, spectrum match viewer and spectra annotation details. The HR-MS library was constituted by using the iMethod™Forensic MS/MS spectra library from ABSciex for the identification of top analytes observed in intoxication samples and infusing reference standards (ACE and CES libraries with > 450 entries). As a very useful feature, the deactivation of pre-set filters in the comprehensive report can show metabolites not listed in the library, but identified by their exact mass deviations to proof the analytical findings and to expand the library. BBGes is continuesly working on expansion of the libray.

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