Which substrate is used in the Bessey-Lowry-Brock method for determining alkaline phosphatase activity?

Which substrate is used in the Bessey-Lowry-Brock method for determining alkaline phosphatase activity?

• A. Monophosphate
• B. Phenylphosphate
• C. Disodium phenylphosphate
• D. Para-nitrophenylphosphate

Answer: (D)

The test relies on a substrate that yields a color change when the enzyme acts on it, allowing simple spectrophotometric measurement of alkaline phosphatase activity. In the Bessey-Lowry-Brock method, the substrate is para-nitrophenyl phosphate. Alkaline phosphatase cleaves off the phosphate, producing para-nitrophenol. In alkaline conditions, para-nitrophenol turns yellow and has a strong absorbance at around 405 nm, so the rate of color development directly reflects enzyme activity. The other substrates listed either aren’t hydrolyzed to form a chromogenic product or don’t produce a measurable color change under the assay conditions, making them unsuitable for straightforward quantification.

The test relies on a substrate that yields a color change when the enzyme acts on it, allowing simple spectrophotometric measurement of alkaline phosphatase activity. In the Bessey-Lowry-Brock method, the substrate is para-nitrophenyl phosphate. Alkaline phosphatase cleaves off the phosphate, producing para-nitrophenol. In alkaline conditions, para-nitrophenol turns yellow and has a strong absorbance at around 405 nm, so the rate of color development directly reflects enzyme activity. The other substrates listed either aren’t hydrolyzed to form a chromogenic product or don’t produce a measurable color change under the assay conditions, making them unsuitable for straightforward quantification.