Plant Science 183 (2012) 57–64 Contents lists available at SciVerse ScienceDirect Plant Science journal homepage: www.elsevier.com/locate/plantsci Light qualities and dose influence ascorbate pool size in detached oat leaves Linda Mastropasqua, Giuseppe Borraccino, Laura Bianco, Costantino Paciolla ∗ Dipartimento di Biologia, Università degli Studi di Bari Aldo Moro, via E. Orabona 4, 70125 Bari, Italy article info Article history: Received 4 July 2011 Received in revised form 10 November 2011 Accepted 11 November 2011 Available online 18 November 2011 Keywords: Ascorbic acid Enzyme activity Light qualities Low light Oat (Avena sativa L.) abstract In this work, we studied the mechanism of light influence on AsA pool size in Avena sativa L. under the effects of low intensity light at different wavelengths. Exposure to low intensity light of oat leaf segments incubated in water or in l-galactono-1,4-lactone (GL), resulted in an increase in AsA content compared with the dark control. This increase was due to modulation of l-galactono-1,4-lactone dehydrogenase (GLDH; EC 1.3.2.3) light-dependent activity and was dependent on the size of the endogenous GL pool. Both blue and red light were effective in increasing AsA, and this increase depended on both exposure time and light intensity. Protein biosynthesis, photosynthesis and calcium were involved in controlling the level of light-dependent AsA. We suggest that multiple checkpoints correlated to the presence of light underlie the ascorbate pool size. The presence of a light-activated switch for the maintenance of an adequate AsA level seems to be necessary for the various tasks of scavenging reactive oxygen species, in response to the dark-light cycle which plants experience under natural conditions. © 2011 Elsevier Ireland Ltd. All rights reserved. 1. Introduction In plant cells, the synthesis of ascorbic acid (AsA) is common in higher plants. AsA is considered one of the major soluble antiox- idant compounds actively synthesized in green tissues as well as in storage organs, and is formed in plants via several biosyn- thetic pathways. One such pathway, beginning from d-glucose [1], involves d-mannose and l-galactose as key intermediates, while the final step in this AsA biosynthetic pathway requires the oxidation of l-galactono-1,4-lactone (GL) by the enzyme GL dehydrogenase (GLDH) [2]. Another proposed pathway involves the conversion of d-galacturonic acid to l-galactonic acid by d-galacturonic acid reductase; this is then converted into GL [3]. An alternative pathway involves the conversion of glucose to d-glucosone, then to sorbosone and finally to AsA [4]. The biosynthesis of ascorbate from GL has been repeatedly observed in plant tissues [5,6], and the enzyme GLDH has been char- acterized and cloned [7]. Ascorbic acid is a component of the ascorbate–glutathione cycle [8]. In this cycle, ascorbate (ASC) is mainly oxidized by the enzymes ascorbate peroxidase (APX) and Abbreviations: AsA, ascorbate; APX, ascorbate peroxidase; BL, blue light; CAP, chloramphenicol; CHX, cycloheximide; DHA, dehydroascorbate; DHAR, dehydroascorbate reductase; GL, l-galactono-1,4-lactone; GLDH, l-galactono-1,4- lactone dehydrogenase; GR, glutathione reductase; MDHAR, monodehydroascor- bate reductase; RL, red light; WL, white light. ∗ Corresponding author. Tel.: +39 080 5443557; fax: +39 080 5443557. E-mail address: paciolla@botanica.uniba.it (C. Paciolla). ascorbate oxidase into monodehydroascorbate (MDHA), which is spontaneously disproportionated to ASC and dehydroascorbate (DHA). MDHA and DHA can be reconverted to ASC by MDHA reductase (MDHAR) and DHA reductase (DHAR), respectively; the former enzyme accepts electrons from NAD(P)H and the latter from reduced glutathione (GSH). The network of reactions in this cycle also includes reconversion of oxidized glutathione (GSSG) into GSH by glutathione reductase (GR). AsA is involved in many physiological processes of plant metabolism [8]. It is a quencher of reactive oxygen species and can also regenerate some membrane-bound radical quenchers, such as -tocopherol and zeaxanthin [9]. A number of hydroxylation reactions are affected by AsA [10], and it also eliminates hydrogen peroxide in a reaction catalysed by APX [11], whose activity seems to be correlated with cytosolic calcium [12]. AsA synthesis has been reported to be stimulated by calcium during calcium oxalate synthesis [13] and the intracellular calcium increase induced by beta-amyloid protein seems to be prevented by AsA [14]. In cells, the regulation of AsA levels depends on its turnover or synthesis and is tissue-dependent. In root meristems, its turnover is under the control of AsA oxidase activity and in the quiescent centre there may be an interaction between AsA oxidase and auxin [15]. Other hormones, such as gibberellin and cytokinins, have been associ- ated with increased AsA [16]. It is known that in detached leaves several changes characterizing senescence are triggered, such as decay of chlorophyll, onset of proteolysis and imbalance of hor- mones [17,18]. Ascorbate content has been shown to decrease in senescing leaves [6]; correlating with AsA levels, GLDH activity has been found to be high in young potato leaves and then decreased 0168-9452/$ – see front matter © 2011 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.plantsci.2011.11.009