Kunitz (KTI) and Bowman-Birk (BBI) trypsin inhibitors were characterized in soybean seeds. Cultivars having KTI/BBI (Pannónia Kincse, PK) or lacking KTI (Aries; Hilario; Bahia) were assessed with well-characterized soybean varieties having Ti-a or ti types of KTI mobility. The TIA values of Pannónia Kincse (9.8 ± 0.48 mg/g) were not significantly different (p ≤ 0.05) from Ti-a samples (10.07 ± 1.86 mg/g), while of Aires, Bahia, Hilario (6.19 ± 1.89) were identical (p ≤ 0.05) with ti samples (6.63 ± 1.99). Radiofrequency heat treatment (RF) decreased TIA values (p ≤ 0.05) at ≥ 100 °C. However, in the traditional soybean variety, the RF at 110 °C was more effective in eliminating the residual KTI activity. The remaining or the disapperaing bioactive form of trypsin inhibitors were succesfully characterized by the means of a standardized in vitro digestion model. It showed that residual BBI-originated trypsin inhibitor activity was in the stomach even after RF at 110 °C, whereas its chymotrypsin inhibitor activity was not detectable at all. Although PK and KTI null types of soybean seeds still required an energy-saving, gentle heat treatment to inactive the trypsin inhibitors before using them as food or feed, the physicochemical properties and processing quality of soybean products were protected, improved.

The inhibition of the alpha-amylase digestive enzyme impedes starch digestion by blocking access to the active site of the enzyme, thereby playing a role in the prevention of obesity and type 2 diabetes. Plant-derived alpha-amylase inhibitors (αAIs) are promising nonpharmacological alternatives for the prevention of these diseases. Alpha-amylase inhibitor-1 (αAI-1) present in common bean (Phaseolus vulgaris) is derived from a precursor protein. In this study, the effect of digestion on the digestibility, immune reactivity, and bioactivity of αAI-1 was assessed from four varieties of Hungarian common bean (Phaseolus vulgaris), with special regard to the precursor protein. For this purpose, αAI-1 was tested in both matrix (native flour and cooked flour) and purified forms under in vitro and acute rat in vivo digestion experiments. The effect of digestion on αAI-1s was monitored by lab-on-a-chip (LOC) electrophoresis, SDS-PAGE/immunoblot, and inhibitory activity analyses by native PAGE. After both in vitro and in vivo digestion, we established that αAI-1 was not degraded even after 60 min gastric digestion and showed immune-reactive properties as well. Although the activity of the purified αAI-1 was lost, that of αAI-1 in the flour matrix (noncooked and cooked) was retained in the stomach. Presumably, in the beans, αAI-1 polypeptides became active due to the pepsin digestion of the precursor protein. The latter samples were also tested in vivo in the small intestine and their resistance and immune reactivity were observed, but αAI-1 did not show activity, as αAI-1 polypeptides were probably complexed by pancreatic amylases. From these results, we can assume that the αAI-1-rich bean protein preparation can affect the carbohydrate metabolism; thus, it could be a promising ingredient for weight loss purposes.

Chickpea (Cicer arietinum L.) generally contains lower levels of these compounds than many other legumes, yet information on Hungarian chickpea cultivars is scarce. This study aimed to characterize protein-based antinutritional factors in twenty chickpeas grown under different agroclimatic conditions over three consecutive years (15 samples from seven Hungarian cultivars from three cultivation areas, and five commercially available foreign genotypes). Protein profiles were examined by SDS-PAGE and native PAGE, while trypsin inhibitor activity (TIA) was quantified spectrophotometrically according to ISO 14902, and lectin activity was determined using a hemagglutination assay. SDS-PAGE revealed highly similar protein patterns among samples, indicating comparable overall protein composition. Native PAGE combined with activity staining confirmed the presence of Kunitz-type trypsin inhibitors, with multiple isoforms detected, but no Bowman–Birk-type inhibitor activity was observed. TIA values were low (0.49–4.07 mg inhibited trypsin/g), and lectin activities were generally low (1–2.5 HU/mg flour; only one sample reached 5 HU/mg) or undetectable. Neither cultivation area nor growing year had a significant effect on TIA or lectin activity, confirmed by statistical analyses. Overall, Hungarian chickpea varieties exhibited low and stable levels of antinutritional proteins, supporting their favorable nutritional quality and suitability for human consumption and expanded cultivation under Hungarian agroclimatic conditions.