Naturally and separately, after the fungus cultivation in synthetic and lignocellulose-containing media
There are a couple of publications dealing with the method. I am sending you one that I think would be able to help you.
Let me know if this was helpful. Have a wonderful day,
Maria
Biotechnology Techniques
1997, Volume 11, Issue 9, pp 701-706
Lignin and manganese peroxidase activity in extracts from straw solid substrate fermentations
M. del Pilar Castillo, P. Ander, J. Stenstrom
. Manganese peroxidase Assay
The activity of MnP can be measured by the method of Wariishi et al. (1992) . The assay was performed by the addition of 0.4 mL MnSO4 (1mM) in 1 mL sodium malonate buffer (50mM) in the presence of 0.4 mL H2O2 (0.1mM) and culture supernatant. Manganic ions Mn+3 form a complex with malonate and the reaction mixture was incubated at 25oC and absorbance of each sample was taken at 270 nm after 10 min interval (Є436=11590 M-1cm-1).
One unit of Manganese peroxidase activity corresponds to the change in absorbance per minute at 25 °C
I am grateful to all my colleagues. I know well these methods. However, there is no correlation between, for example, MnP activities measured using phenol red and Mn+2 oxidation. Moreover, in some cases (depending on fungus, substrate) the activity measured with phenol red reaches 1000 u/l and it is not detected with Mn+2; in other cases we obtain opposite results! What is a true MnP activity?
I would like to use the MBTH/DMAB assay (Castillo et al., 1997) for quantifying manganese peroxidase activity. Do you know if I can use 3-Methyl-2-benzothiazolinone hydrazone hydrochloride monohydrate instead of 3-Methyl-2-benzothiazolinone hydrazone?. Is the salt form of MBTH supposed to alter the reaction speed?
This is the paper from where it get the method
Biotechnology Techniques
1997, Volume 11, Issue 9, pp 701-706
Lignin and manganese peroxidase activity in extracts from straw solid substrate fermentations
M. del Pilar Castillo, P. Ander, J. Stenstrom
Regards
David
Hi, This method is from the following reference.
(Production of Manganese Peroxidase by Trametes villosa on Unexpensive Substrate and Its Application in the Removal of Lignin from Agricultural Wastes). Silva et al. 2014
The reaction medium was composed of 500 μL of the crude enzyme extract, 50 μL of manganese sulfate (2.0 mM), 200 μL of bovine albumin (0.5% w/v), 50 μL of hydrogen peroxide (2 mM) in sodium succinate buffer (0.2 M, pH 4.5), 100 μL of sodium lactate (0.25 M) and 100 μL of phenol red (0.01% w/v). The reaction was monitored by reading the absorbance at 610 nm in a UV-Vis spectrophotometer.
The activity of MnP (U/L) was calculated using Equation:
U/mL= [(ΔAbs/min)/ε*R*t] *10^6 (ml)
where ΔAbs is the difference between the absorbance of the boiled extract and non-boiled extract; ε is the extinction coefficient of oxidized phenol red at 610 nm = 4460 L, m−1·cm−1; R is the aliquot of crude enzyme extract (mL); and t is the reaction time (min).
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