Characteristic Sterols allow the Detection of Cereal Admixture to Amaranth and the Determination of Amaranth Levels
Simon Hammann, Katja Lehnert, Sabine Zikeli, Walter Vetter  
University of Hohenheim, Stuttgart/Germany

The pseudocereal amaranth is an old cultivated plant originating from Central America. Due to the presence of nutrionally valuable ingredients (protein, squalene, tocopherols) the consumption of amaranth has been associated with various beneficial health effects [1, 2]. This nutritional value made it staple food for many South American civilizations like Maya and Inca [1]. Nowadays, it is also consumed in countries not traditionally implementing amaranth in their cuisine. Likewise, it does not contain gluten and is therefore suited for persons suffering from celiac disease [1]. Accordingly, several products containing amaranth are also sold in Germany. The amount of amaranth in these products ranges, e.g. from 10% in corn waffles and 20% in crisp bread. In addition, pure amaranth flour as well as baking mixtures are an the market.
The goal of our study was to provide an easy and fast method to determine the amount of amaranth in processed food and detect admixture of cereals like wheat and spelt to amaranth flour. The method was based on the analysis of characteristic phytosterols. Phytosterols are a class of substances found in the unsaponifable matter of all plants.  
The sterol composition of amaranth is quite unusual, featuring high amounts of α-spinasterol and Δ7-sitosterol, which are scarcely found in other plants. Samples were directly subjected to alkaline saponification followed by extraction of the unsaponifable matter. After silylation, the phytosterols were analyzed by means of GC/ MS analysis in the selected-ion-monitoring (SIM) mode. By doing this, we were able to detect blends of wheat and amaranth flour from the ratio 99:1 to 1:99. However, this method failed when additional oil or fat were present in the processed food containing amaranth. For these samples we determined the amount of amaranth in the product by quantitative determination of the marker sterols.

References:
[1] V. M. Caselato-Sousa et al. J. Food Science, 2012(77) 
[2]  M. León-Camacho et al. Eur Food Res Technol. Anal., 2001(213): 349–355