Comparative Chemical Characterization of Lunasin-enriched Preparations and Modifications of the Inflammasomes <i>In Vitro</i>

Soybean (Glycine max) is one of the most cultivated crops in the world providing the population with large amounts of protein and oil. In addition to its nutritional composition, soybean also contains biologically active compounds with potential health-promoting properties. The presence of these bioactives may be responsible for the lower incidence of chronic diseases in populations that consume a significant portion of soybeans in their diet. One group of soybeanderived bioactives are bioactive peptides and proteins including lunasin, Bowman-Birk inhibitor (BBI) and Kunitz-type trypsin inhibitor (KTI). The overall objective of this research was to develop a method of preparing lunasin-enriched material and evaluate the ability of lunasin-enriched material to inhibit activation of the inflammasomes in vitro. Lunasin-enriched materials were prepared using calcium chloride and pH precipitation methods and compared with two commercially-available lunasin-enriched products. The stability of lunasin against pepsin-pancreatin hydrolysis was evaluated in these materials and the effect of BBI and KTI concentrations were analyzed. Lunasin concentrations ranged from 8.5 to 71.0 μg [microgram]/g pre-hydrolysis and 4.0 to 13.2 μg/g after hydrolysis. In all products tested, lunasin concentration after pepsin-pancreatin hydrolysis (PPH) significantly correlated with BBI and KTI concentrations. One lunasinenriched preparation was evaluated for its ability to modify activation of the inflammasomes in vitro using THP-1 human macrophages. Aberrant activation of the inflammasomes is associated with development of human diseases such as cancer, diabetes and inflammatory bowel diseases. The activation of the inflammasomes in THP-1 human macrophages was accomplished by priming with lipopolysaccharide followed by adenosine triphosphate. Lunasin-enriched material was added during the priming step at concentrations ranging from 0.0625 to 0.25 mg/mL. Addition of lunasin-enriched preparation led to reduction of intracellular reactive oxygen species (ROS) which correlated with reduction in the amount of pro-inflammatory cytokines interleukin-1β(beta) and interleukin-18. These results indicate that ROSs play an integral role in lunasin’s ability to inhibit inflammation and inflammasomes’ activation. This research is the first to report on the role of Kunitz-type trypsin inhibitor on the stability of lunasin against PPH and potential of lunasin-enriched preparations as chemopreventive agent against diseases associated with aberrant activation of the inflammasomes.