The aroma of a particular food consists of volatile organic compounds (VOCs) of different functional groups (alcohols, aldehydes, esters, ketones, carboxylic acids, nitrogen compounds and sulphur compounds) in various combinations and concentrations. Compounds essential to its overall flavour are often used in the food industry as additives that either define the flavour of a product or merely enhance it. As the demand for natural flavourings increases and the cost of raw materials rises, so does the potential for adulteration.
According to the European Flavour and Fragrance Association (EFFA), Gas Chromatography-Combustion Isotope Ratio Mass Spectrometry (GC-C-IRMS) is the most specific and sophisticated method that can distinguish between natural and synthetic VOCs present in a complex sample based on their isotopic values. All laboratories involved in determining stable isotope ratios of light elements are required to use reference materials (RMs) that comply with the Principle of Identical Treatment (PIT) for method validation, quality control and quality assurance. The critical factors when selecting a suitable RM include isotopic homogeneity, a stable isotopic composition over a long period, and ease of handling and storage, which makes finding suitable RMs challenging and explains why, currently, there is a shortage of suitable RMs for daily quality control and accurate isotopic measurements of VOCs.
The main goal of the master's thesis was to determine the suitability of 43 VOCs for use as RMs with either Elemental Analyser - Isotope Ratio Mass Spectrometry (EA-IRMS) or GC-IRMS. Their suitability was evaluated by monitoring the stability of their isotopic composition regarding storage temperature and time (2-8 °C, 18-20 °C, 10 months), functional groups (alcohols, aldehydes, esters, ketones, carboxylic acids, nitrogen compounds and sulphur compounds) and origin (natural/synthetic).
The first part of the thesis included the measurement of the carbon isotope ratios (13C/12C) of each of the selected VOCs using EA-IRMS. It was found that storage temperature did not affect isotopic composition, while at the same time, compounds were sensitive to the measurement conditions irrespective of their functional group. Based on these results, 20 VOCs were proposed as suitable RMs. The selection included, among others, ethanoic acid with the most negative δ13C value (-41.90 ± 0.48 ‰) and 2-methyl-1-butanol with the most positive δ13C value (-7.09 ± 0.32 ‰).
The second part describes the preparation of three laboratory VOC mixtures that, based on literature data, best describe fruit, tuber and fish matrices. The mixtures are intended for data normalisation and identifying the compounds in authentic samples by GC C IRMS. The suitability of the prepared VOC mixtures was monitored with respect to chromatographic separation, VOC interactions in individual mixtures and the stability of the VOC isotopic composition with storage time. The stability results for fruit and fish indicate a RM shelf life of up to one month. In the case of tubers, stability was not confirmed within a time frame of one month. We suggest monitoring stability at shorter time intervals. It was also found that impurities in the mixtures could interfere chromatographically with the target compounds, affecting their isotopic composition. It is suggested that compounds with a purity level of at least 98 % should be used in the mixture.
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