In vitro screening of natural extracts for microbialtrimethylamine-production inhibitory activity using capillary electrophoresis

Elevated level of trimethylamine N-oxide (TMAO) in plasma has been recently associated with the development or propagation of atherosclerosis and other adverse cardiovascular events. TMAO is generated via a metaorganismal pathway that starts with conversion of dietary precursors (choline, L-carnitine, etc.) into trimethylamine (TMA) by gut microbiota, followed by host hepatic enzymes of the flavin-containing monooxygenase family. The discovery of novel dietary compounds able to interfere with the microbial TMA formation would be an innovative approach for reduction of risk for cardiovascular diseases. To date there have been no reports of dietary compounds or natural extracts able to inhibit the activity of carnitine monooxygenase, a microbial enzyme that features the conversion of L-carnitine to TMA. In this work we present a methodology that combines resazurin-based microplate assay with a fast and cost-effective CE-UV method for the high-throughput assessment of carnitine monooxygenase inhibitory potential of a collection of natural extracts. For this purpose, a strain of Klebsiella pneumoniae, isolated from human gut, and capable of producing TMA from Lcarnitine in vitro, was used. Here, we demonstrate the versatile applicability of CE-UV by tracking TMA within a highly complex sample background to determine carnitine monooxygenase inhibitory potential of natural extracts.