Planta Med 2019; 85(18): 1546-1547
DOI: 10.1055/s-0039-3400085
Main Congress Poster
Poster Session 2
© Georg Thieme Verlag KG Stuttgart · New York

Screening for active substances with TLC bioautography: challenges with artefacts

R Merki
1   ICBT, Phytopharmacy and Natural Products Research Group, Zürich University of Applied Sciences,, Wädenswil, Switzerland
,
A Schönborn
2   IUNR, Ecological Engineering Research Group, Zürich University of Applied Sciences,, Wädenswil, Switzerland
,
E Wolfram
1   ICBT, Phytopharmacy and Natural Products Research Group, Zürich University of Applied Sciences,, Wädenswil, Switzerland
› Author Affiliations
Further Information

Publication History

Publication Date:
20 December 2019 (online)

Also available at
 

TLC, and its modern form HPTLC, is a standard method for identification and quality control of herbals [1]. Its combination with bioassays offers simultaneous separation and screening of plant extracts and other multi-compound mixtures for active substances. The intriguing simplicity and rapidness of the method might lead to underestimation of potential artefacts, first reported by Taibon et al. for HPTLC-Tyrosinase Inhibition [2]. In assays based on cells or enzymes in hydrophilic media, high lipophilic substances prevent contact with the potentially active substance and might lead to false-positive or false-negative results. Additionally, diffusion problems from the layer to the biological assay agents can also lead to wrong interpretation of results, due to difference in the physico-chemical properties of the molecules in the sample mixture.

The planar Yeast Estrogen Screen Assay (pYES) is used for detection of human estrogen receptor alpha (hERα) agonists in mixtures [3]. The hERα is expressing β-galactosidase in presence of estrogenic substances. Substances with a strong blue autofluorescence at 366 nm detection and low estrogenic activity are difficult to differentiate from real estrogenic activity, when using the blue fluorescent dye 4-methylumbelliferyl-β-D-galactopyranosid (MUG) as substrate for β-galactosidase. Results of blue fluorescing phenolic acid rich plant extracts with known estrogen activity are demonstrated and a control assay using resorufin β-D-galactopyranoside (RGP) as substrate is proposed.

The poster summarizes potential artefacts from pYES and other antimicrobial assays and reviews the theoretical considerations and suggests further investigation to overcome the challenges and improve bioautographic methods.

 

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