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Mass spectrometry in lipidomics
Cvačka, Josef
Current approaches of analytical chemistry in lipidomics based on mass spectrometry and chromatography are discussed in detail. Examples of qualitative, quantitative and imaging approaches are given.
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Determination of the fatty acids composition in selected hardened fats
JANEŠÍKOVÁ, Kristina
The aim of this work is to get oneself acquainted with the method of spectrum evaluation of fatty acids in hardened fats by means of gas chromatography and mass spectrometry. The theoretical part deals with information concerning the types of lipids, their compositions, properties of fatty acids especially in oils and hardened cooking fats and with basic information regarding the methods of oil hardening. This part is concentrated on the evaluation method of fatty acids by means of gas chromatography with the use of gualitative using the mass spectrometry. The fatty acids were evaluated in some commercially available fats (Albert, Ceres Soft, Iva, Lando, Lira, Omega, Varoma). Palmitic acid (hexadecane acid) was represented in the highest proportion in all fats, it was 36-45.5%, frequently represented was also oleic acid (cis-octadec-9-ane) with its 32.2-39%. The last of the more frequently represented acids is the linoleic acid (cis, cis-octadeca-9.12-diene) with its 7.7% to 10.5%. A lower amount of the linoleic acid is contained in the Albert fat, where it is represented with 4.5% only. Interesting is the content of acids being typical for milk fat of ruminants, it includes the caprylic acid (octane), capric acid (decane) and lauric acid (dodecane) and the content of trans-acids. The hardened Albert fat has the highest content of trans-acids, they occupy 23.4% of the product. On the base of this result it can be deducted that this product has probably a lower quality than other ones. In comparison with other fats, the content of acid typical for milk fakt is higher in the Lando and Iva fats. This results indicate the milk fat was probably added in these products during the production. In the time period of one year no considerable changes in the composition of selected hardened fats were registered, however, from the long termed point-of-view, there is an evident marked alternation in the composition of the Ceres Soft and Omega hardened fats being accompanied with a basic decrease of trans-acids.
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Analysis of lipids mobilized by adipokinetic hormones in the firebug Pyrrhocoris apterur (Heteroptera: Pyrrhocoridae)
BÁRTŮ, Iva
An effect of two adipokinetic hormones (Pyrap-AKH and Peram-CAH-II) on distribution of diacylglycerol (DG) molecular species and their fatty acid (FA) constituents were investigated by liquid chromatography (HPLC) - electrospray ionization mass spectrometry (ESI-MS) in haemolymph of the firebug Pyrrhocoris apterus. The results show that DGs with characteristic FAs are preferentially mobilized from fat body (FB) by the action of both AKHs produced by P. apterus. Both the macropterous and brachypterous morphs show very close DG and FA profiles. A separate action of the Pyrap-AKH and the Peram-CAH-II, however, exerts distinct differences in FA distribution at the macropterous morph. It seems that partially C16 and mainly unsaturated C18 FAs play a dominant role in the AKH action with an exclusive position of linoleic acid (18:2) which represents 50 - 60% of the total DG mobilization. The metabolically active C16 and C18 FAs are preferentially absorbed from the linden seeds and accumulated in the FB.
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LC-MS analysis based on probabilistic approach
URBAN, Jan
Liquid chromatography (LC) in tandem with mass spectrometry (MS) is a measurement tool for obtain information about the compounds in the investigated extracts. There were already developed methods for processing and analysis of measured data sets. However, only partial problems of processing/analysis task were handled independently. Therefore, the rst part describes existing methods and techniques commonly used in the LC-MS for the processing and analysis today. In this thesis an approach based on the theory of systems is used for description of abstract model above the measured data. This model encapsulated all processing/analysis steps into appropriate and consistent mathematical space. The creation of this model via description of the measurement device and data outputs is introduced. Abstract model of LC-MS data set is used to decompose the measurement into three partial contributions, the analyte signal, the random noise and the systemic noise. The separation process of the signal could be estimated using the probabilistic approach. That probabilistic approach to the LC-MS analysis was implemented in the developed software, which was published in the Bioinformatics Journal.
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Analysis of organic acids by a novel procedure of extractive derivatization in human plasma and serum by GC/MS
JAHODOVÁ, Lucie
The theme of the bachelor{\crq}s thesis was ``The Analysis of Organic Acids by New Extractive Derivatization Process in Blood Plasma and Serum Using the GC/MS Technique{\crqq}. The objective of the thesis was to develop a method for fast determination of organic acids by a new process of extractive derivatization, to identify and determine organic acids in body fluids, to compare their abundance and to evaluate the possibility of implementation of this method in clinical diagnostics. Body fluids contain hundreds of organic acids, which co-create a clinical picture of a patient{\crq}s health condition. The current research focuses on implementation of methods providing for comprehensive, metabolism-based analysis of these metabolites important from the point of view of diagnostics. Nowadays, an analyst can select from a wide range of methods and several effective analytical and separation techniques for separation, determination and identification of organic acids in complex biological matrices. In the introductory, theoretical part of the bachelor{\crq}s thesis, the knowledge of properties of organic acids and possibilities of their determination are summarized. In the practical part, the method of fast determination of organic acids in body fluids by the GC/MS techniques was developed. By means of this technique, it is possible to analyze large quantities of organic acids in minimum volume of body fluid. Derivatization of a set of several tens of organic acids with ethyl chloroformate in aqueous medium was carried out. The reaction proceeds very fast and at laboratory temperature. The applicability of the developed method was verified through analysis of organic acids in biological samples. In samples of blood serum and plasma, tens of organic acids were identified by means of the above-mentioned method, retention data and EI mass spectra. Experiments performed for the purpose of this thesis proved that derivatization with chloroformates in aqueous medium is a relatively simple, fast and promising method for analysis of organic acids. Follow-up experimental research is necessary to make the methodology applicable in clinical diagnostics and to define advantages and limitations of this new methodological procedure.
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Analyses of glycoproteins from the salivary glands of the tick \kur{Ixodes ricinus}
BUČINSKÁ, Lenka
I characterized several potential glycoproteins in salivary gland extracts from unfed and partially fed females of ticks Ixodes ricinus using enzyme deglycosylation and lectin labeling. Affinity-based (chromatografic) analysis was applied for isolations of glycoproteins with specificity for GNA (mannose), HPA (N-acetylgalactosamine) and MAA II (sialic acid) lectins. GNA specific 120 kDa glycoprotein was isolated from partially fed females and is modified with N-linked glycans containing {$\alpha$}1,3-mannose. Mass spectrometry analyses confirmed the presence carboxypeptidase M in elution fraction gain with GNA affinity chromatography. GNA specific proteins were purified from unfed female salivary gland extracts. MS analyses identified them as proteins similar to arylsulfatase B and cytoskeletal Sojo protein. Proteins (85 and 56 kDa) isolated with HPA affinity chromatography were characterized as Trappin 12, which is a host protein. MAA II lectin was used for labelling and isolation of 100 kDa protein. N-terminal sequence of the MAA II specific protein predicted similarity with a host protein, Siglec 1. Fucose in salivary gland extract was detected with the labelling of AAA, AAL, UEA I and LTL lectins. Results showed that salivary gland extracts contain {$\alpha$}1,2-; {$\alpha$}1,3- and {$\alpha$}1,6- N-linked fucose and O-linked fucose probably as well. GNA specific proteins were detected in partially fed salivary glands acini type II and III using electron transmission microscopy. Fucose was detected on gut and salivary gland structures using fucose-specific lectin AAL.
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