Effect of l-phenylalanine on PAL activity and production of naphthoquinone pigments in suspension cultures of Arnebia euchroma Royle JohnstReport as inadecuate




Effect of l-phenylalanine on PAL activity and production of naphthoquinone pigments in suspension cultures of Arnebia euchroma Royle Johnst - Download this document for free, or read online. Document in PDF available to download.

In Vitro Cellular and Developmental Biology - Plant

, Volume 48, Issue 5, pp 555–564

First Online: 21 June 2012Received: 20 September 2011Accepted: 28 May 2012

Abstract

The effects of l-phenylalanine PHE on cell growth and production of shikonin and its derivatives, acetylshikonin ACS and isobutyrylshikonin IBS, in suspension cultures of Arnebia euchroma were examined. Supplementing media using PHE have been successfully utilized to enhance shikonin production in cell cultures of other species of Boraginaceae. l-Phenylalanine, the key compound in the phenylpropanoid pathway, is converted by phenylalanine ammonia lyase PAL to trans-cinnamic acid, which is the precursor of p-hydroxybenzoic acid PHB. Coupling of PHB and geranyl pyrophosphate derived from mevalonate pathway by p-hydroxybenzoate-m-geranyltransferase leads later to biosynthesis of shikonins. The addition of 0.01 or 0.1 mM PHE to the culture medium stimulated cell proliferation, where the highest observed increase in fresh cell biomass measured as a ratio of final weight to initial weight was 12-fold, in contrast to an eightfold increase in control cultures. Whereas, growth media supplemented with 1 mM PHE markedly reduced the rate of cell growth to only twofold. Precursor feeding had detrimental effects on both ACS and IBS production in all PHE-supplemented media. The highest total content intracellular + extracellular of the investigated red pigments 9.5 mg per flask was detected in the control culture without PHE. ACS was the major component of the naphthoquinone fraction determined in cells and post-culture media. Shikonin itself was found only in the post-culture media from cultures supplemented with 0.01 or 0.1 mM PHE. Increases in PAL activity corresponded well with the accumulation of investigated naphthoquinones in control culture. However, peak PAL activity did not directly correlate with maximum production of shikonin derivatives. Cytotoxicity of extracts, prepared from the cells cultivated in the presence of PHE or in control cultures, was tested on three cancer cell lines: HL-60, HeLa, and MCF-7. The extracts prepared from the untreated control cultures proved to be the most potent against the examined cancer cell lines. The mean inhibitory concentration values were 0.3, 13, and 8 μg ml for the HL-60, HeLa, and MCF-7 cells, respectively.

KeywordsArnebia euchroma l-phenylalanine PAL activity Shikonin derivatives Cytotoxic activity Secondary metabolites Editor: John W. Forster

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Author: Katarzyna Sykłowska-Baranek - Agnieszka Pietrosiuk - Marcin R. Naliwajski - Anna Kawiak - Małgorzata Jeziorek - Sylwia Wy

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