Malt β-glucanase:
100 assays (manual format) / 400 assays (auto-analyser format)
Lichenase:
100 / 200 assays (manual format) / 330 / 660 assays (auto-analyser format)
The MBG4 reagent contains a single substrate, namely 4,6-O-benzylidene-2-chloro-4-nitrophenyl-β-(31-β-D-cellotriosyl-glucoside) (BCNPBG4). The benzylidene acetal group prevents any hydrolytic action by exo-acting hydrolytic enzymes such as β-glucosidase or cellobiohydrolase. Mixed linkage
β-glucanase (endo-1,3:1,4-β-glucanase) / lichenase (EC 3.2.1.73) acts specifically to release 2-chloro-4-nitrophenol (CNP) from this substrate. The rate of release of CNP is directly related to the β-glucanase/lichenase activity in a sample. The reaction is terminated and the phenolate colour is developed on addition of Tris buffer solution (pH = 10.0).
Note that the substrate is not hydrolysed by β-glucosidase or cellobiohydrolase. The substrate can be hydrolysed by certain endo-cellulases (e.g. Trichoderma sp.) but this does not result in an increase in absorbance.
Colourimetric method for the determination of endo-1,3:1.4-β-glucanase/lichenase in crude malt extracts and enzyme preparations
Principle:
(endo-1,3:1,4-β-glucanase/lichenase)
(1) Benzylidene-BGTETB-β-CNP + H2O → Benzymidene-BGTETB + CNP
(tris buffer)
(2) CNP → phenolate ion (yellow colour)
Note: CNP = 2-Chloro-4-nitrophenol
Kit size:
Malt β-glucanase 100 assays (manual) / 400 (auto-analyser)
or
Lichenase 100 / 200 assays (manual) / 330 / 660 (auto-analyser)
Method: Spectrophotometric at 400 nm
Total assay time:
Malt β-glucanase 20 min (manual) / 10 min (auto-analyser)
or
Lichenase 10 min (manual) / 10 min (auto-analyser)
Detection limit:
Malt β-glucanase 4.3 x 10-4 U/mL
or
Lichenase 9.1 x 10-5 U/mL
Application examples:
Crude malt extracts, industrial enzyme preparations
Method recognition: Novel method
