Substrates and enzyme activities related to biotransformation of resveratrol from phenylalanine by Alternaria sp. MG1
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BIOTECHNOLOGICAL PRODUCTS AND PROCESS ENGINEERING
Substrates and enzyme activities related to biotransformation of resveratrol from phenylalanine by Alternaria sp. MG1 Jinhua Zhang & Junling Shi & Yanlin Liu
Received: 23 July 2013 / Revised: 19 August 2013 / Accepted: 21 August 2013 / Published online: 26 September 2013 # Springer-Verlag Berlin Heidelberg 2013
Abstract To identify the substrates and enzymes related to resveratrol biosynthesis in Alternaria sp. MG1, different substrates were used to produce resveratrol, and their influence on resveratrol production was analyzed using high performance liquid chromatography (HPLC). Formation of resveratrol and related intermediates was identified using mass spectrum. During the biotransformation, activities of related enzymes, including phenylalanine ammonia-lyase (PAL), trans cinnamate 4-hydroxylase (C4H), and 4-coumarate-CoA ligase (4CL), were analyzed and tracked. The reaction system contained 100 mL 0.2 mol/L phosphate buffer (pH 6.5), 120 g/L Alternaria sp. MG1 cells, 0.1 g/L MgSO4, and 0.2 g/L CaSO4 and different substrates according to the experimental design. The biotransformation was carried out for 21 h at 28 °C and 120 rpm. Resveratrol formation was identified when phenylalanine, tyrosine, cinnamic acid, and p-coumaric acid were separately used as the only substrate. Accumulation of cinnamic acid, p-coumaric acid, and resveratrol and the activities of PAL, C4H, and 4CL were identified
Electronic supplementary material The online version of this article (doi:10.1007/s00253-013-5212-3) contains supplementary material, which is available to authorized users. J. Zhang : J. Shi College of Food Science and Engineering, Northwest A&F University, 28 Xinong Road, Yangling, Shaanxi Province 712100, China J. Shi (*) School of Life Sciences, Northwestern Polytechnical University, 127 Youyi Xilu, Xi’an, Shaanxi Province 710072, China e-mail: [email protected] Y. Liu College of Enology, Northwest A&F University, 23 Xinong Road, Yangling, Shaanxi Province 712100, China
and changed in different trends during transformation with phenylalanine as the only substrate. The addition of carbohydrates and the increase of phenylalanine concentration promoted resveratrol production and yielded the highest value (4.57 μg/L) when 2 g/L glucose, 1 g/L cyclodextrin, and phenylalanine (4.7 mmol/L) were used simultaneously. Keywords Alternaria sp . Resting cell . Bioconversion . Phenylalanine
Introduction Resveratrol (3,5,4′-trihydroxystilbene) is a plant stilbene having multiple functions and is in high demand in food and medical industries (Bradamante et al. 2004; Baur and Sinclair 2006; Dudley et al. 2009; Gresele et al. 2011; Howitz et al. 2003; Lee et al. 2011). Biosynthesis of resveratrol using plant cells and genetically modified microorganisms is recently explored to overcome the difficulties caused by shortage of plant sources from which resveratrol is extracted (Beihadj et al. 2008; Kiselev et al. 2013; Donnez et al. 2011). However, these techniques normally need long ti
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