Bioproduction of sulfated glycosaminoglycan oligosaccharides and their effect on non-heme iron uptake: Studies in human intestinal cell lines (Caco-2 cells)

• Author / Creator

  Wang, He Nan

• Glycosaminoglycans (GAGs) such as chondroitin sulfate (CS), dermatan sulfate (DS), heparin (HP), heparan sulfate (HS), and non-sulfated hyaluronic acid (HA) are a group of complex polysaccharides exhibiting a wide range of biological functions. These macromolecules are structurally heterogeneous depending on the animal source, and their functional properties may also be affected by the specific purification procedure. The overall thesis objective is to investigate the bioproduction of GAG-derived sulfated oligosaccharides and study their effect on iron uptake by Caco-2 cells. The first study aimed to develop a food-grade approach for the extraction of GAGs from chicken cartilage and skin, which are low-value poultry by-products. Purified sulfated GAGs were successfully extracted from chicken cartilage (CS) and skin (CS/DS) using porcine pancreatin (a food-grade proteolysis) followed by ethanol precipitation and purification with anion exchange chromatography. In the second stage of this study, the antioxidant capacity of these chicken sulfated GAG polysaccharides and their respective oligosaccharides (oligos) obtained by enzymatic digestion were evaluated. Bovine trachea CS and porcine intestinal DS were used as controls. Then, their effect on iron uptake was evaluated by ferritin formation using an in vitro digestion in Caco-2 cell model. The CS-oligos derived from both CS polysaccharides of chicken and bovine sources possessed the greatest DPPH scavenging and ferric reducing activities (p < 0.05) but had limited ferrous chelating activities. Both chicken sulfated GAG polysaccharides and their controls showed greater ferritin formation compared to the blank (p < 0.05). Depolymerisation of CS polysaccharides further improved ferritin formation by two-fold in both the sample and control. The enhanced iron uptake through enzymatic CS depolymerisation may be due to the combined effects of reduced molecular weight, increased amount of hydroxyl terminal groups, and ferric reducing activities. Based on these results, the second study focused on supplementing enzymatic depolymerised bovine trachea CS into skim bovine milk (SBM) to verify if its promoting effect on Fe uptake can remain pronounced. The effect of CS-oligos with various SBM fractions on Fe uptake by an in vitro digestion/Caco-2 cell model was evaluated. SBM was separated into casein, whey, lactose and oligosaccharides by ultrafiltration. Modified fractions representing the absence of either casein, whey, or lactose were then prepared. Extrinsic ferrous sulfate was added since the intrinsic iron content of SBM is relatively low. The effect of milk oligosaccharides on Fe uptake was evaluated for the first time, which was similar to that of whey. The ferritin synthesis rate was 1.5-fold greater in SBM after the addition of CS-oligos. Its enhancing effect was the most prominent with milk oligosaccharides and the least with casein. The results suggest the possibility of supplementing CS-oligos as a dietary intervention strategy to address iron deficiency anaemia. However, the costly enzymes would limit the possibility of implementing such a technology for commercial opportunities. The last study evaluated possible chemical depolymerisation methods. Bovine trachea CS was hydrolyzed under acid, alkaline and combined (acid and alkaline) conditions at 60°C. The combined acid- and alkaline-treated CS possessed the greatest antioxidant capacity. All chemical hydrolyzed CS samples showed greater ferritin formation compared to the blank (p < 0.05), where the greatest ferritin level was achieved by the combined treated CS. Overall, this thesis provides a research foundation for producing high-quality CS and CS-oligos from the animal sources, as an individual compound or in a food mixture, that can enhance Fe uptake in a Caco-2 cell model. It demonstrates the potential of animal CS and CS-oligos to act as a supplement with multiple bioactivities.

• Subjects / Keywords

  • sulfated glycosaminoglycan and oligosaccharides, iron uptake, Caco-2 cell line

• Graduation date

  Fall 2018

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/R3TB0ZB2S

• License

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