Effect of copper and salicylic acid on phenolic metabolites and free amino acids in Scenedesmus quadricauda (Chlorophyceae) Jozef Kova´cˇik a, *, Borˇivoj Klejdus b , Josef Hedbavny b , Martin Bacˇkor a a Department of Botany, Institute of Biology and Ecology, Faculty of Science, P.J. S ˇ afa´rik University, Ma´nesova 23, 041 67 Kosˇice, Slovak Republic b Department of Chemistry and Biochemistry, Mendel University of Agriculture and Forestry Brno, Zemeˇdeˇlska´ 1, 613 00 Brno, Czech Republic 1. Introduction Aquatic environments are often exposed to various pollutants including heavy metals owing to increasing industrial and/or agricultural wastes. Besides, copper (Cu) is widely used in chemicals for control of phytoplankton development and in agricultural fungicides [1]. Cu is an essential plant micronutrient playing an important role in both photosynthetic and respiratory electron transport, being a cofactor for many enzymes. In terms of Cu excess, this metal is redox-active and stimulates generation of reactive oxygen species (ROS) through Fenton–Haber–Weiss reactions [2]. This leads to damage of membranes, essential biomolecules and loss of important nutrients such as potassium in both vascular and non-vascular plants [3,4]. Regulated production of ROS and maintenance of ‘‘redox homeostasis’’ are essential for the physiological health of organisms. To minimize functional and structural damages, plants have developed different mechanisms enabling them to counteract negative effects caused by metals abundance in their tissue. Among them, phenolic metabolites represent substantial part of plant organic matter and are potent scavengers of ROS or radical-chain breakers thus extinguishing strongly oxidative free radicals such as the hydroxyl radical [5]. Despite high number of studies about accumulation of phenols in vascular plants (including our own with chamomile), there still exists deficit of information about phenols in non-vascular plants. In published papers, phenols in algae have been estimated as total soluble phenols or with respect to antioxidative properties of different extracts mainly from food algal species [6]. Recent papers have shown first comparison of selected phenolic metabolites (phenolic acids) in different microalgae and cyanobacterial species and new extraction methods [7,8]. Several other compounds may also alleviate stress symptoms. Salicylic acid (SA), an important signalling plant molecule, may reduce symptoms of environmental stress [9] but SA-ameliorating effects are closely related to applied concentration [10,11]. There exists very limited number of studies about SA effect on algae [12]. Accumulation of some amino acids such as proline may also contribute to amelioration of negative impacts owing to heavy metal excess [3,13]. Species of the genus Scenedesmus sp. (Chlorophyceae) are frequently used in studies focused on heavy metal toxicity [14]. We therefore tested responses of S. quadricauda to short-term effect of Cu, SA and their combined application in order to check possible preventive role of SA against Cu-induced metabolic changes. Concentrations of Cu and SA were selected based on the survey of Plant Science 178 (2010) 307–311 ARTICLE INFO Article history: Received 18 November 2009 Received in revised form 1 January 2010 Accepted 24 January 2010 Available online 1 February 2010 Keywords: Algae Copper toxicity Heavy metals Oxidative stress Water pollution ABSTRACT Species of the genus Scenedesmus are frequently used in studies focused on heavy metal toxicity and phenols represent important tool for stress protection. However, deeper knowledge about accumulation of phenolic metabolites in algae still absents. We therefore studied selected metabolic responses of widely distributed green algal species Scenedesmus quadricauda to short-term (24 h) effect of copper (Cu), salicylic acid (SA) and their combined application (25 mM for both). Exposure to Cu led to decrease in chlorophylls while SA stimulated increase in chlorophylls. Soluble proteins and total soluble phenols were depressed by Cu treatment while SA had no effect and did not reverse influence of Cu in combined variant. Endogenous SA decreased in Cu + SA treatment and this depletion had no direct effect on the accumulation of the majority of detected phenolic acids, while individual amino acids seemed to be affected. Considerable accumulation of benzoic acids and their noticeable responses to SA may suggest involvement in protection against Cu toxicity since intracellular Cu remained unchanged by exposure to Cu + SA. These observations are discussed in the context of biosynthesis of phenols and possible signalling mechanisms are also suggested. ß 2010 Elsevier Ireland Ltd. All rights reserved. * Corresponding author. Tel.: +421 905 678861. E-mail address: jozkovacik@yahoo.com (J. Kova´cˇik). Contents lists available at ScienceDirect Plant Science journal homepage: www.elsevier.com/locate/plantsci 0168-9452/$ – see front matter ß 2010 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.plantsci.2010.01.009