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New procedures for autonomous analyses of dried blood spots by capillary electrophoresis
Profousová, Sylvie ; Glatz, Zdeněk (referee) ; Dvořák, Miloš (advisor)
Collection of capillary blood in the form of dry blood spot (DBS) samples on cellulose sampling cards has been popular in clinical analysis for some time. However, in the diagnosis and monitoring of metabolic disorders and diseases, target analytes (e.g. inorganic and organic ions) are examined from blood plasma or serum. Determination of the target analytes from capillary blood or DBS samples is not the standard and reference values are not available due to the different composition of plasma/serum vs. capillary blood. Moreover, DBS sorbents have been found to release several ions into DBS eluates resulting in their contamination and, together with the effect of blood hematocrit and retention of target analytes, can adversely affect the quantitative analysis of DBS samples. For these reasons, this thesis addresses the design of an alternative concept for DBS collection that would eliminate the use of sampling sorbents and ensure the collection of a defined and precise volume of blood. The experimental part focuses on the determination of selected inorganic and organic ions (NH4+, K+, Na+, Ca2+, Mg2+, Cl, NO3, NO2, SO42, formate, lactate, and acetate) by capillary electrophoresis with capacitively coupled contactless conductivity detection (CE-C4D) in various DBS samples. All these ions are present in blood at different concentrations and can be simultaneously released as contaminants from the DBS sorbents. A non-porous polypropylene vial was used for the novel sorbentless DBS collection. A small drop of blood of defined volume (e.g. 5 l) was transferred into this vial and then dried directly therein. The selected vial is suitable for both direct DBS elution and DBS eluate analysis and is fully compatible with autosamplers of commercial CE instruments. The proposed concept of the sorbentless blood collection eliminated unwanted contamination and proved suitable for the determination of the tested cations and anions. The stability of anions was generally lower than that of cations but was still significantly better compared to standard DBS sampling sorbents. Determination of some anions appeared to be acceptable only within 0–48 hours after drying of the sorbentless DBS samples (stored at laboratory temperature), as there was a continuous increase in their content. This was probably due to microbial degradation of the blood matrix. It was confirmed that this increase could be slowed down by storing sorbentless DBS samples at low temperatures. Different blood sampling procedures have been proposed for sorbentless DBS collection that are simple, economical, convenient, and repeatable and are therefore suitable for patient-centric blood sampling as well as for sampling in medical centers. This is a basic pilot study applied only to selected analytes that served as model examples, nevertheless, based on this thesis, more extensive scientific studies will follow up to further develop the new concept of sorbentless DBS collection.
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Determination of sweetener substitute content in chewing gum
Profousová, Sylvie ; Nábělek, Jakub (referee) ; Diviš, Pavel (advisor)
The bachelor thesis deals with the simultaneous determination of selected sweeteners in chewing gum by high-performance liquid chromatography with ELSD detection. The theoretical part is devoted to the characterization of carbohydrates and their effect on human health, classification, characterization of selected sweeteners and their use. Furthermore, a brief overview of analytical methods by which sweeteners can be determined is given. The experimental part focuses on the selection of a suitable chromatographic column for the simple identification of seven sweeteners (aspartame, acesulfame K, mannitol, maltitol, sucralose and xylitol) and the optimization of the mobile phase composition. It was found that it is not possible to determine all sweeteners by one universal method, however, using an Agilent Hi-Plex Ca column (300 x 7.7 mm) and the mobile phase of acetonitrile:water at 7:93 % v/v, it was possible to determine the concentration of (maltitol, mannitol, sorbitol, sucralose and xylitol). For the analysis of the individual sweeteners, different sample dilutions had to be used or the composition of the mobile phase had to be slightly changed. The concentration of artificial sweeteners in the chewing gum ranged from 0,35 mg/g (sucralose) to 508 mg/g (xylitol).
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Determination of sweetener substitute content in chewing gum
Profousová, Sylvie ; Nábělek, Jakub (referee) ; Diviš, Pavel (advisor)
The bachelor thesis deals with the simultaneous determination of selected sweeteners in chewing gum by high-performance liquid chromatography with ELSD detection. The theoretical part is devoted to the characterization of carbohydrates and their effect on human health, classification, characterization of selected sweeteners and their use. Furthermore, a brief overview of analytical methods by which sweeteners can be determined is given. The experimental part focuses on the selection of a suitable chromatographic column for the simple identification of seven sweeteners (aspartame, acesulfame K, mannitol, maltitol, sucralose and xylitol) and the optimization of the mobile phase composition. It was found that it is not possible to determine all sweeteners by one universal method, however, using an Agilent Hi-Plex Ca column (300 x 7.7 mm) and the mobile phase of acetonitrile:water at 7:93 % v/v, it was possible to determine the concentration of (maltitol, mannitol, sorbitol, sucralose and xylitol). For the analysis of the individual sweeteners, different sample dilutions had to be used or the composition of the mobile phase had to be slightly changed. The concentration of artificial sweeteners in the chewing gum ranged from 0,35 mg/g (sucralose) to 508 mg/g (xylitol).
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