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J Chromatogr A. 2019 Nov 8;1605:460371. doi: 10.1016/j.chroma.2019.460371. Epub 2019 Jul 15.

Effect-directed analysis by high-performance thin-layer chromatography for bioactive metabolites tracking in Primula veris flower and Primula boveana leaf extracts.

Author information

1
Chair of Food Science, Institute of Nutritional Science and Interdisciplinary Research Center, Justus Liebig University Giessen, Heinrich-Buff-Ring 26-32, 35392 Giessen, Germany; Institute of Pharmaceutical Chemistry, Philipps University Marburg, Marbacher Weg 6-10, 35032 Marburg, Germany; Pharmacognosy Department, Faculty of Pharmacy, Al-Azhar University, 11371 Cairo, Egypt.
2
Saint Catherine Protectorate, Nature Conservation Sector, Egyptian Environmental Affairs Agency, 64616 South Sinai, Egypt.
3
Institute of Pharmaceutical Chemistry, Philipps University Marburg, Marbacher Weg 6-10, 35032 Marburg, Germany.
4
Chair of Food Science, Institute of Nutritional Science and Interdisciplinary Research Center, Justus Liebig University Giessen, Heinrich-Buff-Ring 26-32, 35392 Giessen, Germany. Electronic address: gertrud.morlock@uni-giessen.de.

Abstract

The genus Primula (Primulaceae) comprises species with high medicinal as well as ornamental values. Plants belonging to this genus are well recognized for their richness in bioactive constituents. The huge variety of secondary metabolites make their complete analysis impossible. In order to cope with this challenge, effect-directed analysis (EDA) via HPTLC coupled to structure elucidation techniques was applied on Primula species for the first time. As straightforward non-target bioanalytical technique, HPTLC-UV/Vis/FLD-EDA-ESI-HRMS hyphenates three different orthogonal dimensions, i.e. chromatography with spectrometric detection, biological/enzymatic assays and HRMS. The bioactive metabolites were determined in the middle polar extracts of two Primula species, P. veris (flower) and P. boveana (leaf). The bioactivity profiling comprised the antibacterial activity against Aliivibrio fischeri and Bacillus subtilis bacterial strains and acetyl-/butyrylcholinesterase (AChE/BChE) inhibition assays. The compounds were characterized and identified via their recorded spectral data (HRMS and 1H NMR). The results showed that linoleic and linolenic acids were the principle bioactive compounds present in the studied P. veris flower extract. In the P. boveana leaf extract, flavone, 2'-methoxy-, 2'-hydroxy- and 5,6,2',6'-tetramethoxyflavone (zapotin) were determined as active metabolites. The identification of zapotin, which was previously undescribed in the investigated plant, was considered as the strength of the straightforward non-target bioanalytical technique. Flavone turned out to be the highest potent metabolite, and at the same time, a multipotent compound referring to its various bioactivities discovered. An equivalency calculation of the HPTLC-AChE inhibition by flavone was performed with reference to the well-known inhibitor rivastigmine. As a result, the amount of flavone contained in 10.0 μg dry powder of P. boveana (corresponding to 0.1 μL extract) inhibited as strong as 4.5 μg rivastigmine. In other words, the flavone contained in P. boveana leaf extract powder turned out to be half as strong as the well-known AChE inhibitor rivastigmine.

KEYWORDS:

Antibacterial assay; Bioluminescence; Cholinesterase inhibition; Equivalency calculation; Flavonoids; PUFAs

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