PLANT POLYSACCHARIDES AS POTENTIAL BIOACTIVE THERAPEUTICS: NOVEL APPROACHES TOWARDS PRODUCTION, STRUCTURAL CHARACTERIZATION, IMMUNOMODULATION AND QUALITY CONTROL

Abstract

Historically, plants have been a tremendous source of novel bioactive small molecules or secondary metabolites. However, botanical macromolecules have not been as thoroughly studied. Among the different types of macromolecules, plant polysaccharides have shown potential in the biomedical field due to their immunomodulatory properties along with minimal toxicity. Novel immunomodulatory polysaccharides may be important for immunotherapies and discovery of adjuvants. Additionally, the use of polysaccharides as nutraceuticals and functional foods is becoming popular. Despite the interest in bioactive polysaccharides there are challenges to overcome. Purification costs, structural variability, bacterial endotoxin contamination, and laborious quality control procedures have hindered commercialization. In this study, novel methodologies have been employed in an attempt to address some of these challenges. A plant tissue culture-based system was used to produce polysaccharides with consistent structural characteristics and reduced endotoxin content. The structural features of four immunomodulatory polysaccharide fractions from Panax quinquefolius (American ginseng) suspension cultures were analyzed to begin to elucidate structure-function relationships. Structural analyses revealed the presence of arabinogalactan type II polysaccharides in three fractions and a pectic (rhamnogalacturonan I) polysaccharide in one fraction. The fractions activated RAW 264.7 murine macrophage cells and primary murine splenocytes by enhancing the production of several proinflammatory mediators such as nitric oxide, IL-6, TNF-α, MCP-1, and GM-CSF in vitro and ex vivo. Macrophage activation signaling pathways stimulated by treatment with purified polysaccharides were: iNOS, NF-κB (p65/RelA), and MAPK (p38). The polysaccharides also stimulated mouse splenocyte proliferation, another major indicator of immunostimulation. Finally, this study not only explored novel strategies for production of bioactive polysaccharides using tissue culture, but also developed methods for rapid quality control of carbohydrate polymers using HPLC coupled with charged aerosol detector (CAD). Overall, these studies have shown that American ginseng suspension culture polysaccharides have the potential to be developed into potent immunomodulatory therapeutics.