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Metabolic fingerprinting of rat urine by LC/MS. Part 1. Analysis by hydrophilic interaction liquid chromatography-electrospray ionization mass spectrometry
Stockholm University, Faculty of Science, Department of Analytical Chemistry.
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2005 (English)In: Journal of chromatography. B, ISSN 1570-0232, E-ISSN 1873-376X, Vol. 828, no 1-2, 9-13 p.Article in journal (Refereed) Published
Place, publisher, year, edition, pages
2005. Vol. 828, no 1-2, 9-13 p.
Identifiers
URN: urn:nbn:se:su:diva-24518DOI: 10.1016/j.jchromb.2005.07.031OAI: oai:DiVA.org:su-24518DiVA: diva2:197694
Available from: 2007-11-01 Created: 2007-10-16 Last updated: 2017-12-13Bibliographically approved
In thesis
1. Analysis of Metabolites in Complex Biological Samples Using LC/MS and Multivariate Data Analysis: Metabolic Fingerprinting and Detection of Biomarkers
Open this publication in new window or tab >>Analysis of Metabolites in Complex Biological Samples Using LC/MS and Multivariate Data Analysis: Metabolic Fingerprinting and Detection of Biomarkers
2007 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

To facilitate early diagnosis of diseases and elucidation of the processes involved in their development and progression, various specific compounds or ‘biomarkers’ are often monitored. The first step is to decide which compounds to analyze. An untargeted approach within the field of metabolomics, can provide a unique chemical fingerprint representing the biological state of the studied organism. Fingerprints can be used for classification and thus facilitate the identification of biomarkers and investigation of drug metabolism.

In this work a method was developed for the general detection of metabolites using liquid chromatography/mass spectrometry (LC/MS) to obtain data for metabolic fingerprinting. One part of the project focused on generating the data and the other on analyzing the acquired data. First part: Solid phase extraction was applied to rat urine samples and protein precipitation to human plasma samples. Several LC/MS variants were used, i.e. reversed phase LC, hydrophilic interaction LC and ultrahigh pressure LC (UHPLC) coupled to a triple quadrupole MS or time of flight (ToF) MS. Second part: Methods for handling LC/MS data and extracting information, e.g. curve resolution, were evaluated. In addition data fusion methods were investigated. Principal Component Analysis (PCA) and partial least squares (PLS), were applied for pattern recognition. Furthermore, 3-way classification methods, such as parallel factor analysis (PARAFAC) and N-way PLS, were also explored.

The developed method was applied to the early detection of phospholipi-dosis in drug development and to search for indicators of successful treatment in breast cancer therapy. Potential biomarkers were suggested, but have not yet been fully evaluated. In addition, new metabolites of an antidepressant drug were discovered and identified using this approach.

Place, publisher, year, edition, pages
Stockholm: Institutionen för analytisk kemi, 2007
Keyword
Metabolomics, Metabonomics, Metabolic Fingerprinting, Drug Metabolism, Biomarkers, Human Plasma, Rat Urine, LC/MS, ZIC-HILIC, UPLC, UHPLC, ToF, Curve Resolution, Peak Detection, Pattern Recognition, Classification, PCA, PLS, PARAFAC, N-PLS, Data Fusion, Hierarchical Modeling, Batch modeling, Data Concatenation, Breast Cancer, Phospholipidosis
National Category
Analytical Chemistry
Research subject
Analytical Chemistry
Identifiers
urn:nbn:se:su:diva-7123 (URN)978-91-7155-421-5 (ISBN)
Public defence
2007-11-23, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 12 A, Stockholm, 13:00
Opponent
Supervisors
Available from: 2007-11-01 Created: 2007-10-16Bibliographically approved
2. Methods for structural studies of an antibody, screening metabolites in rat urine and analysis of spent cell cultivation media using LC/ESI-MS and chemometrics
Open this publication in new window or tab >>Methods for structural studies of an antibody, screening metabolites in rat urine and analysis of spent cell cultivation media using LC/ESI-MS and chemometrics
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes bioanalytical methods for generating fingerprints of biological systems for extracting relevant information with (protein) drugs in focus. Similarities and differences between samples can reveal the hidden relevant information, which can be used to optimize the production and facilitate the quality control of such protein drugs during their development and manufacture. Metabolic fingerprinting and multivariate data analysis (MVDA) can also facilitate early diagnosis of diseases and the effects and toxicity of drugs.

Currently, several protein drugs are available on the global market. Nevertheless, despite, the success of such biotherapeutics significant challenges remain to be overcome in maintaining their stability and efficacity throughout their production cycle and long-term storage. The native structure and functional activity of therapeutic proteins is affected by many variables from production to delivery, incl. variables assoc. with conditions in bioreactors, purification, storage and delivery. Thus, part of the work underlying this thesis focused on structural analysis of a protein drug using chemical labeling, peptide mapping, and evaluation of the charge state distributions of the whole protein generated by ESI. The other part focuses on non-targeted metabolomics with a view to optimizing the cell cultivation process and assessment of the drug’s toxicity. A combination of appropriate analytical methods and MVDA is needed to find markers that can facilitate optimization of the cultivation system and expression of the target proteins in early stages of process development. Rapid methods for characterizing the protein drugs in different stages of the process are also required for quality control.

In order to obtain high quality fingerprints analytical separation techniques with high resolution (such as HPLC or UHPLC) and sensitive analytical detection techniques (such as ESI, quadrupole or TOF MS) have been used, singly or in combination.

Place, publisher, year, edition, pages
Stockholm: Department of Analytical Chemistry, Stockholm University, 2009. 74 p.
Keyword
Screening, Fingerprinting, metabolomics, Protein drugs, Mammalian Cell lines, LC/ESI-MS, chemometrics
National Category
Analytical Chemistry
Research subject
Analytical Chemistry
Identifiers
urn:nbn:se:su:diva-28921 (URN)978-91-7155-897-8 (ISBN)
Public defence
2009-09-16, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 12 A, Stockholm, 10:00 (English)
Opponent
Supervisors
Note
At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 3: Manuscript.Available from: 2009-08-25 Created: 2009-07-20 Last updated: 2009-08-27Bibliographically approved

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