In chemical analyses, it is crucial to distinguish between chemical species. This is often accomplished via chromatographic separations. These separations are often pushed to their limits in terms of the number of analytes that can be sufficiently resolved from one another, particularly when a quantitative analysis of these compounds is needed. Very often, complicated methods or new technology is required to provide adequate separation of samples arising from a variety of fields such as metabolomics, environmental science, food analysis, etc. An often overlooked means for improving analysis is the use of chemometric data analysis techniques. Particularly, the use of chemometric curve resolution techniques can mathematically resolve analyte signals that may be overlapped in the instrumental data. The use of chemometric techniques facilitates quantitation, pattern recognition, or any other desired analyses. Unfortunately, these methods have seen little use outside of traditionally chemometrics focused research groups. In this dissertation, we attempt to show the utility of one of these methods, multivariate curve resolution-alternating least squares (MCR-ALS), to liquid chromatography as well as its application to more advanced separation techniques. First, a general characterization of the performance of MCR-ALS for the analysis of liquid chromatography-diode array detection (LC-DAD) data is accomplished. It is shown that under a wide range of conditions (low chromatographic resolution, low signal-to-noise, and high similarity between analyte spectra), MCR-ALS is able to increase the number of quantitatively analyzable peaks. This increase is up to five-fold in many cases. Second, a novel methodology for MCR-ALS analysis of comprehensive two-dimensional liquid chromatography (LC x LC) is described. This method, called two dimensional assisted liquid chromatography (2DALC), aims to improve quantitation in LC x LC by combining the advantages of both one-dimensional and two dimensional chromatographic data. We show that 2DALC can provide superior quantitation to both LC x LC and one dimensional LC under certain conditions. Finally, we apply MCR-ALS to an LC x LC analysis of fourteen furanocoumarins in three apiaceous vegetables. The optimal implementation of MCR-ALS and subsequent integration was determined. For these data, simply performing MCR-ALS on the two dimensional chromatogram and manually integrating the results proved to be the superior method. These results demonstrate the usefulness of these curve resolution techniques as a compliment to advanced chromatographic techniques.

Fumonisins, carcinogenic mycotoxins produced by various Fusarium species, occur naturally in maize and maize-based food products. They are hazards for animal and human health as they cause cancer in rodents and have been associated with oesophageal cancer and neural tube defects in humans. The most abundant naturally occurring fumonisins analogues in maize are fumonisin B1, B2 and B3 (FB1, FB2 and FB3). For analytical determination, they mostly require suitable extraction, clean-up and pre or post-column derivatization together with reversed-phase HPLC separation. o- Phthaldialdehyde (OPA) had been adopted as the most widely used derivatization reagent for fumonisins as they lack useful chromophores or fluorophores. Alternative derivatization reagents, naphthalene-2,3- dicarboxaldehyde (NDA) and dansyl chloride (DnS-Cl), were investigated in this study

Fumonisins, carcinogenic mycotoxins produced by various Fusarium species, occur naturally in maize and maize-based food products. They are hazards for animal and human health as they cause cancer in rodents and have been associated with oesophageal cancer and neural tube defects in humans. The most abundant naturally occurring fumonisins analogues in maize are fumonisin B1, B2 and B3 (FB1, FB2 and FB3). For analytical determination, they mostly require suitable extraction, clean-up and pre or post-column derivatization together with reversed-phase HPLC separation. o- Phthaldialdehyde (OPA) had been adopted as the most widely used derivatization reagent for fumonisins as they lack useful chromophores or fluorophores. Alternative derivatization reagents, naphthalene-2,3- dicarboxaldehyde (NDA) and dansyl chloride (DnS-Cl), were investigated in this study

Masters of Science / Fumonisins, carcinogenic mycotoxins produced by various Fusarium species, occur naturally in maize and maize-based food products. They are hazards for animal and human health as they cause cancer in rodents and have been associated with oesophageal cancer and neural tube defects in humans. The most abundant naturally occurring fumonisins analogues in maize are fumonisin B1, B2 and B3 (FB1, FB2 and FB3). For analytical determination, they mostly require suitable extraction, clean-up and pre or post-column derivatization together with reversed-phase HPLC separation. o- Phthaldialdehyde (OPA) had been adopted as the most widely used derivatization reagent for fumonisins as they lack useful chromophores or fluorophores. Alternative derivatization reagents, naphthalene-2,3- dicarboxaldehyde (NDA) and dansyl chloride (DnS-Cl), were investigated in this study. / South Africa

This diploma thesis is concerned with determination anthocyanins dyes in elderberry (Sambucus nigra L.). In theoretical part there is described general characterization of the plant, its chemical composition, short view of elderberry breeding and possibilities of usage in food and medical industry. Bigger attention is paid to anthocyanins dyes. There is described their characterization, occurrence, factors that have an impact on their stability. Biosynhesis of the pigments and metabolism, benefit effects and their usage in food chemistry are then shortly sumarized. Methods of determination of anthocyanins are discused too. Next chapter deals with high performance liquid chromatography. At the end of theoretical part, there is sumarized findings of anthocyan dyes analysis by expert scientists in various matrix. In experimental part, total of 16 species of elderberry were analyzed. These species are from research and breeding institute of pomology in Holovousy. The dyes were identification and quantification by high performace liquid chromatography coupled with a diode array detector. Detection wavelenght was 520 nm. Gradient of mobile phase was optimilized because of shortening analyses time and resolution improvement. Four dyes were identified by HPLC: cyanidin-3-sambubioside, cyanidin-3-glukoside, cyanidin-3,5-diglukoside a cyanidin-3-sambubioside-5-glukoside. Total amount of anthocyanins ranged from 33627167,0 to 5525284,0 mAU•min-1. The most anthocyanins dyes were determined in Mammut species (33627167,0 mAU•min-1), the lowest amount was measured in Aurea species (5525284,0 mAU•min-1). The most abundant anthocyanin in all species of eldeberry fruit except Haschebrg and Aurea species was cyanidin-3-sambubioside. The concentrations of cynidin-3-glucoside in all of the berries were quantified using regression equation of calibration curve. Mammut species had the most amount of this anthocyan (0,486 mg•ml-1) and the lowest amount had Aurea species (0,033 mg•ml-1).

Die Systematische Toxikologische Analyse (STA) stellt auf Grund der großen Vielfalt und der ständigen Zunahme an toxikologisch relevanten Substanzen eine der größten Herausforderungen in der chemischen Analyse dar. In der vorliegenden Arbeit wurde daher die Eignung der Flüssigchromatographie in Kombination mit der Hybrid-Quadrupol-Flugzeitmassenspektrometrie (LC-QTOF-MS) für diesen Zweck untersucht. Dazu wurden eine Datenbank mit über 7360 und eine CID-Spektrenbibliothek mit mehr als 2720 toxikologisch relevanten Substanzen erstellt und geeignete Probenvorbereitungsmethoden entwickelt. Die Erprobung der Methoden erfolgte an dotierten Blut- und Haarproben. Hierbei zeigte sich, dass die Analyse im Auto-MS/MS-Modus (Messzyklen von MS- und MS/MS-Spektren) eine Identifizierung basischer Substanzen mittels CID-Spektren zwischen 0,5 und 2 ng/ml im Blut ermöglichte. Die Nachweisgrenzen der für 24 Wirkstoffe validierten Methode in Haaren lagen bei 3 bis 15 pg/mg. Die Eignung der LC-QTOF-MS zur STA von Haarproben wurde an 30 Drogentodesfällen und 60 Todesfällen mit bekannter chronischer Medikamenteneinnahme zu Lebzeiten sowie an 77 Blutproben nachgewiesen. Für die Suche nach Metaboliten wurde ein Metaboliten-Tool entwickelt. In der praktischen Anwendung auf Datenfiles von Blut- und Haarproben erwies sich das Tool als wertvolles Hilfsmittel zur Identifizierung unbekannter Peaks und zur Bestätigung von Suchergebnissen in der Datenbank. Zur automatischen Konzentrationsabschätzung identifizierter Substanzen wurde ein Tool „Estimate Concentration“ geschaffen. Die Überprüfung des Verfahrens an realen Blut- und Haarproben durch Vergleich mit HPLC-DAD- und GC-MS-Ergebnissen wies eine gute Übereinstimmung der Konzentrationen auf. Insgesamt zeigten die Untersuchungen, dass die LC-QTOF-MS zurzeit die am besten geeignete Methode für die STA darstellt. Auch bei einem erst später aufkommenden Verdacht kann eine gezielte Suche in dem bereits gemessenen Datenfile durchgeführt werden. / Due to the large variety and the steady increase of toxicologically relevant substances, systematic toxicological analysis (STA) is one of the most difficult tasks in analytical chemistry and, therefore, a steady topic of research and methodical improvement. For this reason, the suitability of liquid chromatography in combination with hybrid quadrupole time-of-flight mass spectrometry (LC-QTOF-MS) for STA was investigated. For this purpose, a database of more than 7360 and a CID spectra library of more than 2720 toxicologically relevant substances and suitable methods for sample preparation were developed. The application was evaluated at spiked blood and hair samples. It was found that the analysis in Auto-MS/MS mode (alternating measurement cycles of MS and MS/MS spectra) allowed substance identification in blood using CID spectra between 0.5 and 2 ng/ml for basic substances. The detection limits of the validated method in hair ranged from 3 to 15 pg/mg for 24 drugs. The suitability of LC-QTOF-MS for STA was tested for hair samples from 30 drug-related death cases and from 60 death cases with known chronic medication as well as for 77 blood samples. For the search of metabolites, a metabolite tool was developed. In the practical application to data files from blood and hair samples, the tool proved to be very helpful for identification of unknown peaks and for confirmation of results obtained only from the database without CID spectra. A tool "Estimate Concentration" was created for automatic estimation of concentrations of identified substances. The application to real blood and hair samples and the comparison of the concentrations with results from HPLC-DAD and GC-MS showed good agreement. Overall, these investigations showed that LC-QTOF-MS is currently the most favorable method for STA. Because of the comprehensive registration of all substances in a sample, the data files can be checked for the presence of certain poisons even later without new measurements.