I am running an experiment to detect ammonia produced in a biological system die to the action of an enzyme using the Berthelot reaction. It seems like the addition of detergent NP-40 to my reaction buffer causes a precipitation when phenol nitroprusside is added, which starts to clear when alkaline hypochlorite is added. Will this interfere with my assay?
Dear Sir. Concerning your issue about whether the detergent NP-40 interfere with Berthelot test for ammonia. Ammonia is extracted using 2M KCl and determined colorimetrically (NH4-N by Berthelot colorimetry, NO3-N by Cd reduction and NED colorimetry) . Analysis is carried out on fresh samples, to avoid losses of NH4-N on drying. Results are reported on a dry weight basis however, by calculation using %moisture of the soil (measured as dry matter). Also, another procedure : sample can be hydrolyzed by the action of the urease to produce ammonia and carbon dioxide. The ammonia reacts with hypochlorite and phenol in the presence of nitroprusside to form indophenol, which in alkaline medium gives an intense blue colour. The intensity of the colour formed is directly proportional to concentration of urea in the sample. To avoid interference you can the above methods. The following below links may help you in your analysis:
http://www.hill-laboratories.com/file/fileid/33832
http://pdf.usaid.gov/pdf_docs/Pnadf783.pdf
Thanks