Isinglass/collagen: denaturation and functionality

Isinglass is widely used commercially to clarify alcoholic beverages by agg regation of the yeast and other insoluble particles. It is derived from swi m bladders of tropical fish by solubilisation in organic acids and consists predominantly of the protein collagen. The low content of intermolecular c ross-links allows ready dissolution of swim bladder compared to bovine hide which is cross-linked by a high proportion of stable bonds and requires en zymic digestion to solubilise. Isinglass is no longer effective as a clarif ying agent if thermally denatured hence the collagenous triple helical stru cture must be maintained. Thermal denaturation of isinglass occurs at 29 de grees C, compared to 40-41 degrees C for mammalian collagens, primarily due to the lower hydroxyproline content. The hydroxyproline is essential for t he formation of H-bonded water-bridges through the hydroxyl group and the p eptide chain thereby stabilising the triple helix. Based on the lower entha lpy determined by differential scanning calorimetry we have calculated that the thermally labile domain of the isinglass molecule was 41 residues comp ared to 66 for mammalian collagen. The fining efficiency was unaffected by pH, chelating agents, detergents and removal of surface proteins from yeast cells. Studies on the mechanism of action of isinglass have shown that hig her molecular weight aggregates that increase the length of the collagen mo lecules (trimers, tetramers, etc.) increase efficiency and that their surfa ce charge are important in the clarification process. By chemical modificat ion, we have shown that blocking positively charged groups had no effect on the fining process, whilst negative charges are clearly essential and that increasing the negative charge by succinylation increases its efficacy. So lutions of bovine hide collagen were shown to be equally effective in refin ing beers and standard yeast preparations. The higher thermal denaturation temperature, ready availability and reproducibility of bovine collagen prep arations gives it considerable advantages over isinglass. (C) 2000 Elsevier Science B.V. All rights reserved.