Effect of chemical structural properties of chitooligosaccharides on the immune activity of macrophages

Abstract

The immunological effects according to the difference in the degree of deacetylation (DD) and molecular weight (MW) of chitosan purified from Hermetia illucens larvae were investigated. The DD of purified high molecular weight chitosan (Hi-CTSN) from H. illucens larvae was found to be 87.2%. According to the substrate specificity of the enzyme, Hi-CTSN was treated with chitosanase and chitinase to prepare chitooligosaccharides (CTS) named CTS2 and CTS3, respectively. For comparison of relative activity, CTS1 obtained by treating crab shell-derived HMWC1 with chitosanase was used as a control. Matrix-assisted laser desorption/ionization (MALDI-TOF) mass spectrometry was subjected to determine the sugar components constituting CTS1, CTS2, and CTS3, respectively. By examining the biological activity of CTS and the immuno-stimulatory activity of CTS according to MW and DD, the expression of many inflammatory cytokines related to the peripheral immune system in macrophage RAW 264.7 cells was confirmed. As a result, all CTS showed similar effects on the expression of tumor necrosis factor-alpha (TNF-alpha) in all concentration ranges used. Among them, CTS1 was verified to have a significant effect on the secretion of inflammatory cytokines, such as interleukin-6 (IL-6) and interleukin-1 beta (IL-1 beta), at a high concentration of 250-500 mu g/mL. However, CTS3 showed significantly higher expression than CTS1 and CTS2 at low concentrations ranging from 50 to 100 mu g/mL. However, there was no significant difference in antioxidant activity against FRAP, ABTS, and DPPH in all CTS. Our results suggest that the presence or absence of the acetyl group of CTS in response to immune stimulation showed a very significant difference, but had a less substantial association with antioxidant activity. Therefore, our data suggested that the difference in DD had a more substantial effect on the immuno-stimulatory activity than the MW of CTS.