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Mobile phase selection for the combined use of liquid chromatography-inductively coupled plasma mass spectrometry and electrospray ionisation mass spectrometry
Umeå University, Faculty of Science and Technology, Department of Chemistry.
Umeå University, Faculty of Science and Technology, Department of Chemistry.
2010 (English)In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1217, no 30, 4980-4986 p.Article in journal (Refereed) Published
Abstract [en]

Four different organic solvents: dimethylformamide, 1,4-dioxane, n-propanol and ethanol were evaluated as alternative organic modifiers to acetonitrile for liquid chromatography (LC) separations. The aim was to establish common sets of chromatographic conditions that could be applied for LC hyphenation to inductively coupled plasma mass spectrometry (ICPMS) as well as to electrospray ionization MS (ESIMS). The approach was to evaluate candidate solvents that, compared to acetonitrile, potentially could give improved analytical performance (low solvent vapor loading, maximized analyte sensitivity and minimized carbon depositions on instrumental parts) in ICPMS analysis while retaining chromatographic and ESIMS performances. The study showed that dimethylformamide, 1,4-dioxane, n-propanol and ethanol all can be advantageous chromatographic modifiers for LC-ICPMS analysis, giving superior performance compared to acetonitrile. For the combined use of LC-ICPMS and LC-ESIMS with a common set of chromatographic conditions, n-propanol gave the best overall performance. The 195Pt+ signal in ICPMS was continuously monitored during a 0-60% organic solvent gradient and at 25% of organic modifier, 100% of the signal obtained at the gradient start was preserved for n-propanol compared to only 35% of the signal when using acetonitrile. Platinum detection limits were 5-8 times lower using n-propanol compared with acetonitrile. Signal-to-noise ratio in continuous ESIMS signal measurements was 100, 90 and 110 for a 100 microg/ml solution of leucine-enkephaline using acetonitrile, ethanol and n-propanol, respectively. Chromatographic efficiency in reversed phase separations was preserved for n-propanol compared to acetonitrile for the analysis of the whole protein cytochrome C and the peptide bacitracin on a column with particle and pore sizes of 5 microm and 300 A, but slightly deteriorated for the separation of the peptides leucine-enkephaline and bacitracin on a 3 microm and 90 A column as the peak width at half height for both peptides increased by a factor of two. The performance on the smaller dimensioned column could however be improved by running the separations at 40 degrees C.

Place, publisher, year, edition, pages
2010. Vol. 1217, no 30, 4980-4986 p.
Keyword [en]
ICPMS, ESIMS, liquid chromatography, mobile phase, hyphenation
URN: urn:nbn:se:umu:diva-35285DOI: 10.1016/j.chroma.2010.05.062ISI: 000280019300013PubMedID: 20573350OAI: diva2:342878
Available from: 2010-08-11 Created: 2010-08-11 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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