Evaluation of alginate purification methods: Effect on polyphenol, endotoxin, and protein contamination

Abstract

Alginate, a polysaccharide extracted from brown seaweed, is widely used for the microencapsulation of islets of Langerhans, allowing their transplantation without immunosuppression. This natural polymer is known to be largely contaminated. The implantation of islets encapsulated using unpurified alginate leads to the development of fibrotic cell overgrowth around the microcapsules and normalization of the blood glucose is restricted to a very short period if it is achieved at all. Several research groups have developed their own purification method and obtained relatively good results. No comparative evaluation of the efficiencies of these methods has been published. We conducted an evaluative study of five different alginate preparations: a pharmaceutical-grade alginate in its raw state, the same alginate after purification according to three different published methods, and a commercially available purified alginate. The results showed that all purification methods reduced the amounts of known contaminants, that is, polyphenols, endotoxins, and proteins, although with varying efficiencies. Increased viscosity of alginate solutions was observed after purification of the alginates. Despite a general efficiency in decreasing contamination levels, all of the purified alginates contained relatively high residual amounts of protein contaminants. Because proteins may be immunogenic, these residual proteins may have a role in persisting microcapsule immunogenicity.

Immunological Challenges Facing Translation of Alginate Encapsulated Porcine Islet Xenotransplantation to Human Clinical Trials

“…The three methods were specifi cally aimed at lowering concentrations of polyphenols, endotoxins and proteins in crude alginate, of which they were all successful, albeit to varying degrees. However, the most convincing results were obtained by Klock [ 10 ]. Upon further modifi cation to the technique devised by Klock et al incorporating size exclusion preparative chromatography and further dialysis, subsequent endeavors resulted in protein concentrations that were reduced by up to 90 %.…”

“…However, the subsequent development of multiple, albeit unstandardized, purifi cation techniques have warranted further comparative investigations. In an evaluation of the effectiveness of three in-house alginate purifi cation procedures designed by Prokop and Wang, de Vos et al, and Klock et al as opposed to using crude, unpurifi ed, and commercially available purifi ed alginate, all three groups reported varying results [ 10 ]. The three methods were specifi cally aimed at lowering concentrations of polyphenols, endotoxins and proteins in crude alginate, of which they were all successful, albeit to varying degrees.…”

“…Upon further modifi cation to the technique devised by Klock et al incorporating size exclusion preparative chromatography and further dialysis, subsequent endeavors resulted in protein concentrations that were reduced by up to 90 %. In vivo effects of these purifi cation techniques in relation to PFO were quantitatively measured via cell adhesion scores upon retrieval from implantation sites [ 10 ]. Subsequent to the treatment of alginate by in-house purifi cation protocols, many involving chemical extraction procedures [ 11 ], the properties of the polymer were also susceptible to change.…”

Functionalization of Alginate via Phosphorization: Preparation, Characterization.

“…A shoulder was observed around 250-290 nm. Absorbance within the 250-290 nm range is commonly attributed to π − π* electron transitions in aromatic and poly-aromatic compounds [22], which could connect with the impurities such as humic acid, phenol, DNA or proteins [23]. The decrease of the …”

A Study on Proliferation and Behavior of Retinal Pigment Epithelial Cells on Purified Alginate Films

“…Among them, Alginate is quickly gelatinized with divalent cations, such as calcium chloride and barium chloride and has possibility for using on diverse parts due to including cells and drugs (25). However, because of contaminants, such as protein and endotoxin in the Alginate, some recent studies have shown the results that proliferation and adhesion of cells on alginate films were reduced (26)(27)(28).…”