PhD conferral R. Ötvös



Aula, Vrije Universiteit Amsterdam

High resolution screening analytics for identification of natural bioactives

R. Ötvös

Prof. G.W. Somsen, Prof. A.B. Smit and Dr. J. Kool

Amsterdam Institute of Molecular and Life Sciences

Earth and Life Sciences

PhD conferral

Foto R. ÖtvösIt is our pleasure to invite you to the thesis defense of Réka Ötvös

Réka will defend her thesis entitled “High resolution screening analytics for identification of natural bioactives” on Tuesday, 11thOctober 2016 at 11:45 in the Aula of Vrije Universiteit Amsterdam.
Vrije Universiteit Amsterdam can be easily reached by public transport or by car. You can find the directions to reach the University on this link. The Aula is located on the first floor above the main entrance of the university.

High resolution screening analytics for identification of natural bioactives
This thesis describes the development and application of different hyphenated high resolution screening (HRS) methods in order to identify novel bioactive compounds acting on brain receptors. HRS is a technique in which high-performance liquid chromatography (HPLC) separations are coupled to mass spectrometry (MS) detection with in parallel an on-line biochemical assay. The first part of this thesis focuses on a microfluidic HRS system using nano-LC which allows analysis of biological extracts available only in low amounts, like animal venoms. In the microfluidic HRS system a fluorescence enhancement based bioassay for the acetylcholine binding protein (AChBP) was applied, and used for a snake venom screening campaign. Subsequently, development of a follow-up workflow allowed for MS guided purification and chemical identification of novel bioactives found in the venoms analyzed. Furthermore, the same methodology was applied to the development of a HRS setup for determination of affinity to the 5-hydroxytryptamine binding protein (5HTBP). To obtain real cellular biological data on bioactive compounds in venoms acting on brain receptors, subsequently an at-line cellular HRS setup was developed with a cell-based functional calcium-flux assay using α7-nicotinic-acetylcholine receptor over expressing neuronal cells. This at-line HRS system was finally transferred from plate reader format to a real on-line HRS methodology based on continuous cellular infusion with LC eluent followed by an on-line calcium flux assay measured with a flow cytometer. With the HRS screening methods developed new potential lead molecules can be identified from natural complex mixtures for medicines targeting central nervous system (CNS) diseases.

Figure 1. Schematic overview of the on-line cell-based bioactivity screening platform. The platform combines reversed-phase liquid chromatography (LC) with on-line flow cytometry (FC) and mass spectrometry (MS).