[Analytical Chemistry] Detection of enzyme inhibitors in crude natural extracts using droplet-based
Natural product screening for new bioactive compounds can greatly benefit from low reagents consumption and high throughput capacity of droplet-based microfluidic systems. However, the creation of large droplet libraries in which each droplet carries a different compound is a challenging task. A possible solution is to use an HPLC coupled to a droplet generating microfluidic device to sequentially encapsulate the eluting compounds. In this work we demonstrate the feasibility of carrying out enzyme inhibiting assays inside nano-liter droplets with the different components of a natural crude extract after being separated by a coupled HPLC column. In the droplet formation zone, the eluted components are mixed with an enzyme and a fluorogenic substrate that permits to follow the enzymatic reaction in the presence of each chromatographic peak and identify those inhibiting the enzyme activity. Using a fractal shape channel design and automated image analysis we were able to identify inhibitors of Clostridium perfringens neuraminidase present in a root extract of the Pelargonium sidoides plant. This work demonstrates the feasibility of bioprofiling a natural crude extract after being separated in HPLC using microfluidic droplets on-line and represents an advance in the miniaturization of natural products screening.
Abraham Ochoa, Enrique Álvarez-Bohórquez, Eduardo Castillero, and Luis F. Olguin Anal. Chem., Just Accepted Manuscript DOI: 10.1021/acs.analchem.6b04988 Publication Date (Web): April 4, 2017 Copyright © 2017 American Chemical Society