Case Studies and Publications

Applied Photophysics has supplied innovative stopped-flow solutions for over three decades contributing to thousands of peer-reviewed references.

Determination of enzyme kinetics

This study describes the use of an SX20 spectrometer to measure the presteady state kinetics of a well-studied enzymatic reaction: hydrolysis of p-Nitrophenyl acetate catalyzed by α-chymotrypsin. Kinetic parameters such as rate constants and the Michaelis-Menten constant are evaluated.

Since it was first studied over 50 years ago, extensive research has established the detailed mechanism of this reaction.

Investigating carbonic anhydrase inhibition properties of potential drug molecules for treatment of disease

Dr Claudiu Supuran and his team at the University of Florence use an SX stopped-flow system to assay carbonic anhydrase inhibition properties of potential drug molecules for treatment of a number of diseases including cancer, glaucoma and malaria.

The technique involves taking a complex of potential inhibitor and carbonic anhydrase and rapidly mixing with a carbon dioxide solution in the presence of a phenol red indicator to measure the kinetics of carbon dioxide hydration.

The assay is effectively a high-throughput stopped-flow application as each potential drug candidate is investigated complexed with carbonic anhydrase at four different concentrations before rapid mixing with the carbon dioxide solution and repeating three times.

During a typical 8-hour day, 48 candidate compounds can be investigated, generating a total of 576 kinetic traces. Since installation in 2001, stopped-flow data has contributed to more than 500 publications from Dr Supuran and his team.

Selected references:

Supuran CT "Carbonic anhydrases: novel therapeutic applications for inhibitors and activators” Nat Rev Drug Discov. 2008 Feb;7(2):168-81. Bozdag M, Ferraroni M, Carta F, Vullo D, Lucarini L, Orlandini E, Rossello A, Nuti E, Scozzafava A, Masini E, Supuran CT.

“Structural insights on carbonic anhydrase inhibitory action, isoform selectivity, and potency of sulfonamides and coumarins incorporating arylsulfonylureido groups” J Med Chem. 2014 Nov 13;57(21):9152-67.