Amylase Trypsin Inhibitors (ATIs) in a Selection of Ancient and Modern Wheat: Effect of Genotype and Growing Environment on Inhibitory Activities

Abstract

Wheat amylase-trypsin inhibitors (ATIs) are a family of plant defense proteins with an important role in human health for their involvement in allergies, celiac disease and non-celiac wheat sensitivity. Information about the differences in ATI activities among wheat genotypes and the influence of the growing environment is scarce. Therefore, ten selected wheat accessions with different ploidy level and year of release, previously characterized for their ATI gene sequences, were grown during three consecutive crop years at two growing areas and used for in vitro ATI activities. The contributions of the genotype and the crop year were significant for both activities. The hexaploid wheat genotypes showed the highest inhibitory activities. Einkorn had a peculiar behavior showing the lowest alpha-amylase inhibitory activity, but the highest trypsin inhibitory activity. It was not possible to observe any trend in ATI activities as a function of the release year of the wheat samples. The two inhibitory activities were differently affected by the growing conditions and were negatively correlated with the protein content. This information can be important in understanding the extent of variation of ATI inhibitory properties in relation to the wheat genotype and the growing environment and the impact of ATIs, if any, on human health and nutrition.

Screening of α-amylase/trypsin inhibitor activity in wheat, spelt and einkorn by high-performance thin-layer chromatography

“…These results were consistent with the findings of Simonetti et al , 36 which evaluated the highest trypsin inhibition for einkorn samples; however, contrary to the results of Geisslitz et al , 19 where einkorn had the lowest ATI amount. Unfortunately, the assay conditions were not comparable to those of other studies 24,36 since they used chromogenic substrates, which have a different affinity to trypsin compared to the used casein.…”

“…According to Geisslitz et al , 19 einkorn contained the lowest amount of ATIs compared to wheat, spelt, and emmer, resulting in the lowest α-amylase inhibition, as confirmed by Simonetti et al 36 Our results did not support this hypothesis. The flour extract of einkorn showed no significantly lower inhibition than all other investigated flours, keeping in mind the possible presence of other α-amylase inhibitors, the affinity of the enzyme regarding its origin (human or animal), 26 the different analytical approaches, and the natural variety of the flour composition.…”

“…According to Geisslitz et al, 19 einkorn contained the lowest amount of ATIs compared to wheat, spelt, and emmer, resulting in the lowest a-amylase inhibition, as conrmed by Simonetti et al 36 Our results did not support this hypothesis. The our extract of einkorn showed no signicantly lower inhibition than all other investigated ours, keeping in mind the possible Fig.…”

“…presence of other a-amylase inhibitors, the affinity of the enzyme regarding its origin (human or animal), 26 the different analytical approaches, and the natural variety of the our composition. Other studies on a-amylase inhibition 26,36 have also reported moderate to high standard deviations. Calculating the overall inhibition (Table S1 †) as a simulation of values obtained by spectrophotometric assays, rened spelt (15%) showed the strongest inhibition, and rened wheat the lowest (−40%).…”

No correlation between amylase/trypsin-inhibitor content and amylase inhibitory activity in hexaploid and tetraploid wheat species

“…In contrast to the tetraploid and hexaploid wheat species, diploid einkorn contains very low amounts of ATI or the ATI are even not present at all, because they are not expressed ( Afzal et al, 2023 ; Geisslitz et al, 2020 ; Sielaff et al, 2021 ; Zoccatelli et al, 2012 ). This is in accordance to the missing α-amylase inhibitory activity ( Bedetti et al, 1974 ; Sanchez-Monge et al, 1996 ; Simonetti et al, 2022 ), but in contrast to a very potent inhibitory activity towards trypsin ( Call et al, 2020 ; Simonetti et al, 2022 ), which might result from a specific einkorn trypsin inhibitor ( Sielaff et al, 2021 ; Taddei et al, 2009 ). The inhibitory activity towards trypsin is lower in emmer than in einkorn, whereas common wheat, spelt and durum wheat are in between ( Call et al, 2020 ).…”