Modification of cellulose‐based packaging materials for enzyme immobilization

Abstract

SUMMARYActive cellulose-based packaging materials were prepared by binding lysozyme to paper modifi ed with anionic polyelectrolytes. Polyelectrolytes improve the paper binding capacity towards the positively charged lysozyme and play a protective role towards lysozyme activity during the paper-making process. The charge density of paper was increased by incorporating carboxymethylcellulose (CMC) or polygalacturonic acid (PGA). The presence of either CMC or PGA greatly increased the amount of bound lysozyme, and the stability of its binding towards buffers or towards non-ionic chaotropes which disrupt the three dimensional structure in macromolecules (urea, 8 M). Binding of lysozyme to modifi ed papers was sensitive to anionic detergents (sodium dodecyl sulphate, 1%) and to non-chaotrope salts (NaCl, 0.5M). These data provide information about the nature of the interactions between the protein and the various types of paper, and provide insights on how to preserve the activity of lysozyme-loaded paper. The polymers used for lysozyme immobilization were found to affect only marginally the structural and functional properties of the antimicrobial protein, and to facilitate the recovery of structural features of the protein after heat treatment. Presence of a negative polyelectrolyte (and of CMC in particular) improves the thermal stability of the protein, making it resistant to thermal inactivation, even under the conditions used for drying processes without compromising the paper's mechanical properties. The activity measurements showed that paperbound lysozyme retains its lytic (and therefore, antimicrobial) activity against the cell walls of Micrococcus lysodeikticus, the target microrganism used as standard for lysozyme bioassays.

Antimicrobial activity of lysozyme and lactoferrin incorporated in cellulose-based food packaging

“…This characteristic make lactoferrin a great candidate for being incorporated in a cellulose-based active packaging by using the same strategies developed in our previous work for lysozyme (Mascheroni et al, 2010).…”

“…Paper sheets were prepared according to the international standard ISO 5269-2:2004, using various ratios of long and short fibers in the presence/absence of 10% sodium carboxymethyl cellulose (CMC, WALOCEL CRT 60000 PPA07, DS 0.7e0.8, Wolff Cellulosic, Schkopau, Germany), retaining its physical and mechanical properties, as reported elsewhere (Mascheroni et al, 2010). Food grade lysozyme (Caglificio Clerici s.p.a., Codorago, Italy) or bovine lactoferrin (Fonterra, Cernusco Lombardone, Italy) were added directly to the slurry used for paper preparation.…”

“…The amount of total protein was 10% (w/w) of the fiber used for papermaking. The slurry was then processed into paper following the procedure described by Mascheroni et al (2010). Paper sheets were wet pressed and dried under restraint for 2 min at 90 C.…”

“…In previous work our group has produced and characterized a cellulose-based active packaging loaded with the well known antimicrobial enzyme lysozyme (Mascheroni et al, 2010). By modulating the anionic charge of the matrix it was possible to bind lysozyme onto (and into) the material via non-covalent interactions.…”

Study of cellulose-lysozyme interactions aimed to a controlled release system for bioactives

“…If the charge density of the substrate is not sufficient, it may be increased by introducing polyelectrolytes, such as carboxymethyl cellulose (CMC) (Horvath and Lindström 2007;Laurell 1952;Wiebe and Klug 1957;Laine et al 2000;Blomstedt et al 2007), that are also able to enhance paper strength (Duker and Lindstron 2008). A previous study of the same authors (Mascheroni et al 2010) shows that lysozyme absorption on the paper sheet surface and thermal stability greatly increased by incorporating CMC into the pergamyn paper.…”