Open database design lists pollutants in the Rhine

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  • Published: Sep 4, 2019
  • Author: Ryan De Vooght-Johnson
  • Channels: Laboratory Informatics
thumbnail image: Open database design lists pollutants in the Rhine

River water analysis is challenging

Rivers can contain a vast range of small molecules, including pharmaceuticals, industrial chemicals and agrochemicals. Analysis is typically carried out by some form of LC-MS, but manual examination of the output from multiple samples is laborious, so automated matching to a database is usually preferred. However, problems caused by varying instrumentation and data handling protocols can make matching difficult, and not all systems allow for the retrospective examination of data for new compounds of interest.

Scientists from the Federal Institute of Hydrology (Koblenz, Germany) devised a new protocol for river water analysis using LC-MS/MS. A new, open database of ‘likely’ compounds was prepared, but new compounds could be added if required, and the data searched retrospectively. Database-assisted screening was used, where compounds in the database were searched for by filtering the raw data.

LC-MS/MS applied to river water samples

Samples of water from the Rhine were taken on a daily basis throughout a year (2016), filtered and stored at -20 ˚C prior to analysis. A mixture of three different internal standards was added to each sample. HPLC used an Agilent 1260 Infinity instrument, fitted with a Zorbax Eclipse Plus C18 column. Elution was carried out using a water/acetonitrile gradient, the eluent being buffered with 0.1% formic acid.

Mass spectrometry employed a Sciex TripleTOF 6600 instrument with electrospray ionisation (ESI). For each sample, one MS survey scan was run, followed by eight MS2 scans of the most intense ions from the initial scan. The data was converted from the Sciex format to the open mzXML format for subsequent processing. A database was prepared using 693 standard compounds, run under the same conditions as the river samples. Spectra were extracted using the RMassBank package and then transferred to a relational database system (SQLite) for use in the screening process.

A standard mixture of 70 compounds was prepared and used to spike river water samples, in order to test the effectiveness of the method. The number of false negatives and false positives were calculated. Using the three search criteria of m/z, tR (retention time) and MS2 peaks gave only 0.5% false negatives and 5.6% false positives. The latter could be identified by subsequent manual examination. Using only two of the three criteria gave significantly more false positive results, 12% when only m/z and MS2 were used and 15% when only m/z and tR were used. Thus it was decided to use the three search criteria for routine analysis. Other possible criteria, such as isotopic ratios, proved not to be generally useful at the concentrations typically found in river water.

The monitoring of Rhine water over a year led to 135 compounds being tentatively identified. The majority of these were pharmaceuticals or their metabolites, although industrial chemicals, agrochemicals and natural products were also present. Many compounds were noted at roughly the same levels throughout the year, exceptions were agrochemicals, some industrial chemicals and hay-fever pharmaceuticals, where seasonal variation was noted.

River water monitoring made easier by new pollutant database

The use of the new database and the three matching criteria of m/z, tR and MS2 signals made the routine analysis of river water samples more straightforward. It would be useful if other research groups started using and contributing to the new database. The results from the Rhine show the extent of pharmaceutical pollution, a problem that tends to be somewhat neglected compared to other pollutants such as agrochemicals.

Related Links

Jewell, K., Kunkel, U., Ehlig, B. et al. (2019). Comparing mass, retention time and MS2 spectra as criteria for the automated screening of small molecules in aqueous environmental samples analyzed by LC‐QToF‐MS/MS. Rapid Communications in Mass Spectrometry doi: 10.1002/rcm.8541.

Alygizakis, N., Samanipour, S., Hollender, J. et al. (2018). Exploring the potential of a global emerging contaminant early warning network through the use of retrospective suspect screening with high-resolution mass spectrometry. Environmental Science & Technology 52 (9): 5135-5144.

Rimayi, C., Chimuka, L., Gravell, A. et al (2019). Use of the Chemcatcher® passive sampler and time-of-flight mass spectrometry to screen for emerging pollutants in rivers in Gauteng Province of South Africa. Environmental Monitoring and Assessment 191: 388.

Wikipedia, Electron capture detector

Article by Ryan De Vooght-Johnson

The views represented in this article are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd.

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