Browsing by Author "Salonen, Fanny"

The latest version of liquid chromatography is ultra-high performance (or pressure) chromatography (UHPLC). In the technique, short and narrow-bore columns with particle sizes below 3 µm are used. The extremely high pressure used results in very short analysis times, excellent separation, and good resolution. This makes UHPLC a good choice for steroidal analysis. Steroids are a highly interesting area of study; they can be recognized as biomarkers for several diseases and are a relevant topic in doping testing. In this thesis articles on the topic ‘steroid analysis with UHPLC’, published prior to April 2017, are reviewed. UHPLC is always combined with mass spectrometry (MS) for steroid analysis. The MS utilized is usually of multi-dimension: quadrupole time of flight (QTOF) or triple quadrupole (QqQ). The instrumentation is suitable for both untargeted and targeted analysis. In untargeted studies, the study of changes in the human metabolome has been especially interesting. The articles on targeted studies are usually focused on doping control and quantification of identified biomarkers. The analysis with UHPLC-MS/MS usually provide reliable results with fast analysis time, without complicated sample preparation. Typically, the sample preparation processes can include only protein precipitation, liquid-liquid extraction or solid-phase extraction. UHPLC is also a valuable tool in simple and routine analysis. The separation efficiency is increased by the small plate height and the analysis time can thus be reduced. In this thesis work the technique was utilized for the analysis of food additives. For validation of an UHPLC method the repeatability, trueness, bias, measurement uncertainty and other factors need to be assessed. The experimental part of the thesis is dedicated to describe the development and validation of a method for analysis of five food additives and caffeine. The developed method was partly validated, with the aim to fulfil the needs of the Finnish Customs Laboratory. The optimized method comprised of an injection volume of 2 µL and a flow rate of 1.0 mL/min. The buffer was a phosphate buffer at pH of 4.0 and the gradient elution program was from 6 % to 30 % of acetonitrile in 1.6 minutes, then 1.6-1.7 minutes with 6% acetonitrile. The total run time was only 1.7 minutes. The limit of detection values was between 0.02 µg/mL and 1.73 µg/mL. The limit of quantitation values was between 0.054 µg/mL to 5.78 µg/mL, which should be sufficient for the Customs needs in the sense of checking if a product is over a certain limit. Expanded measurement uncertainties were around 20 %.