产品资料

Megazyme/Malt β-Glucanase/Lichenase Assay Kit (MBG4 Method)/K-MBG4/100 assays (manual) / 400 assays (auto-analyser)

Maltβ-glucanase:
100assays(manualformat)/ 400assays(auto-analyserformat)
Lichenase:
100/200assays(manualformat)/ 330assays(auto-analyserformat)

TheMBG4reagentcontainsasinglesubstrate,namely4,6-O-benzylidene-2-chloro-4-nitrophenyl-β-(3¹-β-D-cellotriosyl-glucoside)(BCNPBG4).Thebenzylideneacetalgrouppreventsanyhydrolyticactionbyexo-actinghydrolyticenzymessuchasβ-glucosidaseorcellobiohydrolase.Mixedlinkage
β-glucanase(endo-1,3:1,4-β-glucanase)/lichenase(EC3.2.1.73)actsspecificallytorelease2-chloro-4-nitrophenol(CNP)fromthissubstrate.TherateofreleaseofCNPisdirectlyrelatedtotheβ-glucanase/lichenaseactivityinasample.ThereactionisterminatedandthephenolatecolourisdevelopedonadditionofTrisbuffersolution(pH=10.0).

Notethatthesubstrateisnothydrolysedbyβ-glucosidaseorcellobiohydrolase.Thesubstratecanbehydrolysedbycertainendo-cellulases(e.g.Trichodermasp.)butthisdoesnotresultinanincreaseinabsorbance.

Datacalculatorsarelocatedinthe Documentationtab.

Novelapproachestotheautomatedassayofβ-glucanaseandlichenaseactivity.

Mangan,D.,Liadova,A.,Ivory,R.&McCleary,B.V.(2016).CarbohydrateResearch,435,162-172.

LinktoArticle

ReadAbstract

Wereporthereinthedevelopmentofanovelassayprocedureforthemeasurementofβ-glucanaseandlichenase(EC3.2.1.73)incrudeenzymeextracts.Twoassayformatsbasedona)adirectcleavageorb)anenzymecoupledsubstratewereinitiallyinvestigated.The‘directcleavage’substrate,namely4,6-O-benzylidene-2-chloro-4-nitrophenyl-β-3¹-cellotriosyl-β-glucopyranoside(MBG4),wasfoundtobethemoregenerallyapplicablereagent.Thissubstratewasfullycharacterisedusingacrudemaltβ-glucanaseextract,abacteriallichenase(Bacillussp.)andanon-specificendo-1,3(4)-β-glucanasefromClostridiumThermocellum(EC3.2.1.6).Standardcurveswerederivedthatallowtheassayabsorbanceresponsetobedirectlyconvertedtoβ-glucanase/lichenaseactivityonbarleyβ-glucan.ThespecificityofMBG4wasconfirmedbyanalysingtheactionofcompetingglycosylhydrolasesthataretypicallyfoundinmaltonthesubstrate.Manualandautomatedassayformatsweredevelopedfortheanalysisofa)β-glucanaseinmaltflourandb)lichenaseenzymeextractsandtherepeatABIlityoftheseassayswasfullyinvestigated.

Colourimetricmethodforthedeterminationof endo-1,3:1.4-β-glucanase/lichenaseincrudemaltextractsand enzymepreparations

Principle:
(endo-1,3:1,4-β-glucanase/lichenase)
(1) Benzylidene-BGTETB-β-CNP+H₂O→Benzymidene-BGTETB +CNP

(trisbuffer)
(2)CNP →phenolateion(yellowcolour)
Note: CNP=2-Chloro-4-nitrophenol

Kitsize:
Maltβ-glucanase100assays(manual)/400(auto-analyser)
or
Lichenase100/200assays(manual)/330(auto-analyser)

Method:                     Spectrophotometricat400nm
Totalassaytime:
Maltβ-glucanase20min(manual)/10min(auto-analyser)
                                       or
Lichenase10min(manual)/10min(auto-analyser)
Detectionlimit:
Maltβ-glucanase4.3x10^-4U/mL
                                       or
Lichenase9.1x10^-5U/mL
Applicationexamples:
Crudemaltextracts,industrialenzymepreparations
Methodrecognition:   Novelmethod

Advantages

Verycosteffective
Allreagentsstablefor>2years
Specificfor endo-1,3:1,4-β-glucanase/lichenase
Simple, convenient,rapidassay
Wellsuitedtoautomation
Maltflourstandardandlichenasestandardincluded

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Megazyme/Malt β-Glucanase/Lichenase Assay Kit (MBG4 Method)/K-MBG4/100 assays (manual) / 400 assays (auto-analyser)

Novelapproachestotheautomatedassayofβ-glucanaseandlichenaseactivity.