Development and Validation of SPE-HPLC-MS/MS Method for Determining Cyclophosphamide in Surface Waters
Radosław Czernych1, Jarosław Jan Halkiewicz2, Agata Kot-Wasik3, Jacek Namieśnik3
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1Medical University of Gdansk, Department of Health Sciences, Faculty of Health Sciences with Subfaculty of Nursing
and Institute of Maritime and Tropical Medicine, Department of Environmental Toxicology,
Debowa 23, 80-204 Gdańsk, Poland
2PROLAB, Pharmaceutical Analytical Production,
Przemysłowa 3, Paterek, 89-100 Nakło n/Notecią, Poland
3Department of Analytical Chemistry, Chemical Faculty, Gdańsk University of Technology (GUT),
Narutowicza 11/12 G, 80-952 Gdańsk, Poland
Pol. J. Environ. Stud. 2014;23(5):1537–1545
A rapid and selective method for trace amounts determination of cyclophosphamide in surface water samples has been developed. A solid phase extraction SPE method for extraction and clean-up procedure has been optimized for determination by reversed-phase high-performance liquid chromatography with tandem mass spectrometry. The analyses proceed in the positive ion mode by means of the electrospray ionization method (ESI). Clean up was accomplished using a polimeric surface modified styrene – divinylobenzene SPE column.
The final method was validated according to international chemical harmonization (ICH) standards. High selectivity of assay procedure was obtained by choosing optimal columns (most columns contained a bed modified with C18 groups but varied in the amount of bed: 50-500 mg, and grain size: 33-80 μm) and setting separation conditions, such as flow rate during sorption (2 and 6 mL/min), sorbent type, pH of the sample (samples at pH 3 and 7 were examined), solvent strength during desorption (methanol and dichloromethane), time of evaporation (10, 20, and 30 minutes), type of solvent used for injection (HPLC) (water, methanol and methanol-water mixture, 1:1, v/v), matrix effect (tap water and river water), and time “sample preparationanalysis” on recovery (3, 6, and 9 days).
After optimization of sample preparation procedure and analytical conditions environmental water samples were collected from five sampling sites situated in Gdańsk (Pomerania, Poland) and its outskirts and subjected to validated methodology. In four samples cyclophosphamide has been quantified.