Browsing by Author "Abderhalden, Sharon"

Avenanthramides are hydroxycinnamic acids unique to oat (Avena sativa L., Poaceae). They consist of an anthranilic acid part (anthranilic acid or hydroxylated and/or methoxylated derivative of anthranilic acid) that is conjugated to a cinnamic acid part via an amide bond. More than 25 different avenanthramides are found and identified in oat. However, the most common forms are esters of 5-hydroxyanthranilic acid with caffeic (2c), ferulic (2f) and p-coumaric (2p) acids. Avenanthramides have been shown to possess antioxidant and anti-inflammatory properties. Currently there is a great interest towards oat and bioactive compounds of oat, like avenanthramides. In the previous studies there has been a lot of diversity concerning the extraction method used for analysis of avenanthramides and usually quantitation methods were based on high-performance liquid chromatography (HPLC). The aim of this research was to develop a method based on ultra-high-performance liquid chromatography (UHPLC) for the analysis of the most common forms of avenanthramides (2c, 2f and 2p) in oat and oat products. In addition, also other forms of avenanthramides, like 2fd and 2pd, were identified and quantified using liquid chromatography-mass spectrometry (UHPLC-QTOF-MS). The extraction method of avenanthramides was optimized and the UHPLC-PDA method used for quantitation was validated. Finally, the study measured the differences in concentrations of avenanthramides in different oat products. In this study it was recognized that the concentrations and the extent of variation of the concentrations of avenanthramides were affected by the sample amount, the homogeneity of the samples and the extraction time used. Especially bigger sample amount of oat flour (0.5 g) led to larger and more reproducible results than smaller amount (0.1 g). The ratio of sample and solvent 1:10 worked excellently and as an extraction solvent ethanol:water (80:20) was more efficient than tested ethanol:water (80:20) with a phosphate buffer at pH 2.8. Smaller particle size of oat flour and extraction overnight led to better extractability of avenanthramides than a short extraction of sample with larger particle size. The UHPLC method was optimized using chromatographic parameters. The avenanthramides separated from each other acceptably and were eluted as follows during the 16-minute run: 2c, 2p, 2f. The response of the UHPLC instrument was linear in the tested concentration range for all three avenanthramides. The results were reproducible, and the accuracy of the UHPLC instrument was acceptable. The recovery % for avenanthramides were 99–117%. Also, other forms of avenanthramides, like 2fd, 2pd, 5p, 5f ja 3f, were identified in different oat samples using UHPLC-QTOF-MS technique. The total amount of avenanthramides in analyzed oat samples varied between 3–41.3 µg/g per fresh weight. Oat bran included them the most and oat snack the least. Avenanthramide 2c was dominant in oat flour which included 2p the least. In oat bran, in oat drink, in oat snack, in Nyhtökaura and in Muru the avenanthramide 2f was dominant over the two other forms. The UHPLC method developed in this study can be applied to the analysis of the most common forms of avenanthramides 2c, 2f and 2p in oat and in oat products and can be used in oat research in the future. The method can also be used to identify and quantitate more widely other forms of avenanthramides in different oat products. The optimized extraction was shown to be functional and reproducible to already homogeneous and processed samples, but the extraction of non-homogeneous samples could be optimized further in the future.