Comparing capillary electrophoresis-mass spectrometry fingerprints of urine samples obtained after intake of coffee, tea, or water.

Metabolomic fingerprinting is a growing strategy for characterizing complex biological samples without detailed prior knowledge about the metabolic system. A two-way analysis system with liquid separation and mass spectrometric detection provides detail-rich data suitable for such fingerprints. As a model study, human urine samples, obtained after intake of coffee, tea, or water, were analyzed with capillary electrophoresis electrospray ionization time-of-flight mass spectrometry (CE-ESI-TOF-MS). In-house-developed software (in Matlab) was utilized to manage and explore the large amount of data acquired (230 CE-MS runs, each with 50-100 million nonzero data points). After baseline and noise reduction, followed by suitable binning in time and m/z, the data sets comprised 9 and 14 million data points in negative and positive ESI mode, respectively. Finally, a signal threshold was applied, further reducing the number to about 100 000 data points per data set. A set of interactive exploratory tools, utilizing principal component analysis (PCA) and analysis of variance (ANOVA) results based on a general linear model, facilitated visual interpretation with score plots (for group assessment) and differential fingerprints (for "hot spot" detection). In the model study highly significant differences due to beverage intake were obtained among the 10 first principal components (p < 10(-6) for two of the components in both ESI modes). Especially, the contrasts between "coffee" and "tea or water" indicated several "hot spots" with highly elevated intensities (e.g., for uncharged masses 93, 94, 109, 119, 123, 132, 148, 169, 178, 187, 190, and 193) suitable for further analysis, for example, with tandem MS.

Multivariate comparison between peptide mass fingerprints obtained by liquid chromatography-electrospray ionization-mass spectrometry with different trypsin digestion procedures.

Peptide mass fingerprints were obtained for three different proteins using three different digestion procedures in triplicates with liquid chromatography coupled to electrospray ionization mass spectrometry. For each protein the results were compared with multivariate data analysis (cluster analysis, kernel principal component analysis) and pair-wise contrast evaluation. Clear systematic differences between the digestion procedures were established for all the proteins. The visual presentation of the pair-wise differences between procedures could to some extent be related to the protein fragments, although the main objective was to identify m/z and retention regions in the original peptide maps that should be subject to further exploration.

Qualitative screening for basic drugs in autopsy liver samples by dual-plate overpressured layer chromatography.

An overpressured layer chromatography (OPLC) method was evaluated for broad-scale screening of basic drugs in 5g autopsy liver samples using two parallel OPLC systems. Sample preparation included enzymatic digestion with trypsin and liquid-liquid extraction with butyl chloride. Chromatographic separation was performed as dual-plate analysis, with mobile phases composed of trichloroethylene-methylethylketone-n-butanol-acetic acid-water (17:8:25:6:4, v/v) (OPLC1), and butyl acetate-ethanol (96.1%)-tripropylamine-water (85:9.25:5:0.75, v/v). Identification was based on automated comparison of corrected R(f) values (hR(f)c) and in situ UV spectra with library values by dedicated software. The identification limit was determined for 25 basic drugs in liver ranging from 0.5 to 10mg/kg. The OPLC method proved to be well suited for routine screening analysis of basic drugs in post-mortem samples of varying quality, combining the benefit from moderately high separation power with the ease of disposable plates.

Screening for biomarkers in plasma from patients with gangrenous and phlegmonous appendicitis using CE and CEC in combination with MS.

Today a high degree of "false" appendicitis diagnoses are occurring. In this study, a screening experiment of biomarkers of two different kinds of appendicitis, gangrenous and phlegmonous, were conducted with CE and CEC coupled to MS. Plasma samples were obtained from patients pre- and post-surgery. A large amount of data was generated to be able to compare them, and chemometrics tools were utilized to visualize the differences. Indicative patterns were found for both pre- and post-surgery of the two types of inflammation as well as between them. The divergences were traced back to the MS peaks obtained in the CE- and CEC-MS setups as possible biomarkers for the two forms of appendicitis.

Urine profiling using capillary electrophoresis-mass spectrometry and multivariate data analysis.

This work presents the development of a general and fast method for metabolic profiling of urine, using capillary electrophoresis-electrospray ionisation mass spectrometry (CE-ESIMS) and multivariate data analysis (DA). Human urine samples collected before and after ingestion of paracetamol were analysed at acidic and basic CE conditions, using both positive and negative ESI-MS detection. Analysis of the entire resulting data set, with no prior knowledge of the target compounds, using pair-wise 'fuzzy' correlation and eigenvalue analysis enabled the samples to be discriminated between on the basis of blank urine and urine collected after drug intake. By generating two-dimensional loadings plots, it was also possible to identify the m/z values of the substances responsible for the differentiation between control and dosed samples.