title: Novel substrates for the measurement of endo-1,4-β-glucanase (endo-cellulase).
abstract: A specific and sensitive substrate for the assay of endo-1,4-β-glucanase (cellulase) has been prepared. The substrate mixture comprises benzylidene end-blocked 2-chloro-4-nitrophenyl-β-cellotrioside (BzCNPG3) in the presence of thermostable β-glucosidase. Hydrolysis by exo-acting enzymes such as β-glucosidase and exo-β-glucanase is prevented by the presence of the benzylidene group on the non-reducing end d-glucosyl residue. On hydrolysis by cellulase, the 2-chloro-4-nitrophenyl-β-glycoside is immediately hydrolysed to 2-chloro-4-nitrophenol and free d-glucose by the β-glucosidase in the substrate mixture. The reaction is terminated and colour developed by the addition of a weak alkaline solution. The assay procedure is simple to use, specific, accurate, robust and readily adapted to automation. This procedure should find widespread applications in biomass enzymology and in the specific assay of endo-1,4-β-glucanase in general.
authors.affiliation: Megazyme International Ireland, Bray Business Park, Southern Cross Road, Bray, County Wicklow, Ireland.
authors.forename: Barry V
authors.initials: BV
authors.lastname: McCleary
authors.affiliation: Megazyme International Ireland, Bray Business Park, Southern Cross Road, Bray, County Wicklow, Ireland. Electronic address: david@megazyme.com.
authors.forename: David
authors.initials: D
authors.lastname: Mangan
authors.affiliation: Megazyme International Ireland, Bray Business Park, Southern Cross Road, Bray, County Wicklow, Ireland.
authors.forename: Robin
authors.initials: R
authors.lastname: Daly
authors.affiliation: Centre de Recherches sur les Macromolécules Végétales (CERMAV, UPR-CNRS 5301), Affiliated with Grenoble Alpes University, Member of Institut de Chimie Moléculaire de Grenoble (ICMG, FR-CNRS 2607) and Institut Carnot PolyNat, BP53, 38041 Grenoble Cedex 9, France.
authors.forename: Sébastien
authors.initials: S
authors.lastname: Fort
authors.affiliation: Megazyme International Ireland, Bray Business Park, Southern Cross Road, Bray, County Wicklow, Ireland.
authors.forename: Ruth
authors.initials: R
authors.lastname: Ivory
authors.affiliation: Megazyme International Ireland, Bray Business Park, Southern Cross Road, Bray, County Wicklow, Ireland.
authors.forename: Niall
authors.initials: N
authors.lastname: McCormack
mesh_terms: D002480:Cellulase / Q000032:analysis / Q000378:metabolism*; D003124:Colorimetry / Q000379:methods; D009596:Nitrophenols / Q000737:chemistry; D014312:Trisaccharides / Q000138:chemical synthesis / Q000378:metabolism*; D001617:beta-Glucosidase
chemical_list: D009596:Nitrophenols; D014312:Trisaccharides; D001617:beta-Glucosidase; D002480:Cellulase; C548837:2-chloro-4-nitrophenol
keywords: 2-Chloro-4-nitrophenyl; Assay procedure; Cellulase; Colourimetric; Oligosaccharides; endo-1,4-β-Glucanase