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Hydrophilic interaction liquid chromatography (HILIC) coupled to inductively coupled plasma mass spectrometry (ICPMS) utilizing a mobile phase with a low-volatile organic modifier for the determination of cisplatin, and its monohydrolyzed metabolite
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Umeå University, Faculty of Medicine, Department of Surgical and Perioperative Sciences.
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2008 (English)In: Journal of Analytical Atomic Spectrometry, Vol. 23, no 7, 948-54 p.Article in journal (Refereed) Published
Abstract [en]

This study demonstrates the on-line coupling of hydrophilic interaction liquid chromatography (HILIC) to inductively coupled plasma mass spectrometry (ICPMS). A low volatile organic solvent, dimethylformamide (DMF), was used as organic modifier of the mobile phase to minimize the solvent loading of the ICP and thereby improve analyte sensitivity and method robustness. The concept was illustrated by the selective determination of cisplatin (cis-DDP), and its mono-hydrolyzed metabolite (MH-DDP). Species were separated on a 2.1 × 150 mm ZIC-HILIC column and detected on-line by selective platinum (m/z 194 and 195) monitoring, giving a detection limit of 2 pg Pt (0.2 ng ml-1). Compared to the use of the conventional solvent acetonitrile (AcN), with DMF, cis-DDP sensitivity was enhanced as much as 36 times and no addition of oxygen to the plasma was needed to avoid carbon depositions on instrumental parts. Furthermore, several non-identified platinum containing compounds were observed when using AcN as a result of unwanted reactions between this solvent and the analytes. No such species were observed when DMF was used. The molecular structures of eluting compounds were verified by electrospray ionization mass spectrometry. Combined with a simple sample treatment protocol, the HILIC-ICPMS system allowed determination of free intracellular cis-DDP in in-vitro grown T289 human malignant melanoma cells up to 60 min after exposure to 50 g ml-1 cis-DDP for 1 h. This work shows that HILIC-ICPMS is a potent hyphenated technique for the analysis of hydrophilic metal compounds and that the use of chromatographic mobile phases with low-volatile organic solvents may be a generic approach to improve analyte sensitivity and system robustness of HPLC-ICPMS when mobile phases with high amount of organic solvent are used.

Place, publisher, year, edition, pages
2008. Vol. 23, no 7, 948-54 p.
URN: urn:nbn:se:umu:diva-10266DOI: doi:10.1039/b716093cOAI: diva2:149937
Available from: 2008-08-05 Created: 2008-08-05 Last updated: 2010-11-05Bibliographically approved
In thesis
1. Advances in analytical methodologies for studies of the platinum metallome in malignant cells exposed to cisplatin
Open this publication in new window or tab >>Advances in analytical methodologies for studies of the platinum metallome in malignant cells exposed to cisplatin
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Alternative title[sv]
Förbättrade analytiska metodologier för studier av platina-metallomet i maligna celler exponerade för cisplatin.
Abstract [en]

The scientific progress about the important chemotherapeutic drug substance cisplatin (CDDP) and its function has often been rendered by data difficult to interpret, and still many questions about its mode of action remains to be clarified by the scientific community. However, studies of CDDP possess a high complexity due to; i) low intracellular concentration, ii) many potential biomolecule targets, iii) poor or unknown stability of the intact drug and its biomolecule adducts and iv) complex and varying sample matrices. Metallomic studies, using advanced analytical techniques may contribute to clarify the interactions between CDDP and intracellular biomolecules. For a successful outcome sample preparation conditions as well as separation and detection techniques must be carefully selected and optimized to achieve accurate results and correct interpretation of data.

        This thesis describes some new and improved analytical methodologies for characterizing the Pt metallome in CDDP-exposed malignant cells. The developed methods are based on powerful liquid chromatography (LC) methods hyphenated to sensitive detection by inductively coupled plasma- (ICP) and electrospray ionization mass spectrometry (ESIMS). Consideration has also been taken about sample preparation conditions.

        By selecting “chemically inert” sample preparation (cell lysis by osmosis) and separation (using only nonreactive or no additatives) conditions we could avoid the formation of platinum artifact compounds previously described in the literature (Paper I and II). Using oxygen containing organic solvents with high boiling points (dimethylformamide; DMF, 1,4-dioxane, n-propanol and ethanol) as alternatives to acetonitrile in the LC separations, significant improvements were achieved in ICPMS sensitivity and robustness. When evaluated in combination with chromatographic performance and ESIMS detection the overall best performance was achieved with n-propanol (Paper II, III and IV). From the studies in Paper II we could show that free intact CDDP can be found in malignant cells, as supporting evidence for passive or endocytotic uptake of the drug and further estimate a half-life for intracellular CDDP to about 15 minutes. Such data has not been shown before. In Paper V, the above improved LC methods were used to demonstrate differences in the platinum and cupper metallome from sensitive and resistant T289 melanoma cells exposed to CDDP at near clinical levels.

        In a wider perspective we have shown the potential of using hydrophilic liquid interaction chromatography (HILIC) hyphenated to ICPMS detection as a general approach for analysis of hydrophilic metallo-compounds (Paper II). Taking advantage of the superior ICPMS performance using n-propanol gradients for reversed phase liquid chromatography (RPLC) possess a true alternative and /or complimentary technique to size exclusion chromatography (SEC) commonly applied within metallomic studies of biomolecules (Paper V). Using n-propanol in HILIC as well as in RPLC enables parallel detection by ICP- and ESIMS using only one set of chromatographic parameters (Paper III and IV), something commonly called for by scientists in the field.

Place, publisher, year, edition, pages
Umeå: Kemiska Institutionen, Umeå universitet, 2010. 8+44 p.
Method development, Metallome, Inductively coupled plasma mass spectrometry, Electrospray ionization mass spectrometry, Liquid chromatography, Hyphenation, Organic modifier, Cisplatin, Platinum
National Category
Analytical Chemistry
Research subject
Analytical Chemistry
urn:nbn:se:umu:diva-37400 (URN)978-91-7459-085-2 (ISBN)
Public defence
2010-11-26, KBC-huset, Stora Hörsalen, KB3A1, Umeå Universitet, Umeå, 13:00 (English)
Available from: 2010-11-05 Created: 2010-11-02 Last updated: 2010-11-05Bibliographically approved

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Nygren, YvonneNaredi, PeterBjörn, Erik
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Department of ChemistryDepartment of Surgical and Perioperative Sciences

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