A simple activity staining protocol for lipases and esterases.
A simple activity staining protocol for rapid detection and differentiation of lipases and esterases was developed based on pH drop due to fatty acids released following lipolysis. Though the detection of lipolysis as a function of drop in pH is not new, the present method has been made more sensitive by the judicious selection of the initial pH of the chromogenic substrate, which has been set near the end point of the dye so that even a slight drop in pH results in immediate color change. In the present case, the dye phenol red was taken, which has the end point at pH 7.3-7.4 where the color is pink. A slight drop due to fatty acid release results in yellow coloration. The assay has high reproducibility and can detect as low as 0.5 p-NPP enzyme units within 15 min. In addition, this method can be used for various lipidic substrates such as oils and tributyrin, making it suitable for both lipases and esterases.
Singh
Rajni
R
Department of Microbiology, University of Delhi South Campus, Benito Juarez Road, New Delhi, 110 021, India.
Gupta
Namita
N
Goswami
Vineet Kumar
VK
Gupta
Rani
R
D019117:Burkholderia / Q000201:enzymology; D004950:Esterases / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000378:metabolism*; D008828:Microbiological Techniques; D009089:Mucor / Q000201:enzymology; D010637:Phenolsulfonphthalein / Q000378:metabolism*; D011549:Pseudomonas / Q000201:enzymology; D013194:Staining and Labeling / Q000379:methods*; D014280:Triglycerides / Q000378:metabolism; D014304:Triolein / Q000378:metabolism
D014280:Triglycerides; D014304:Triolein; D004950:Esterases; D008049:Lipase; D010637:Phenolsulfonphthalein; C005830:tributyrin