# Vitamin C in Plants: From Functions to Biofortification
## 2 Vitamin C as an Antioxidant  
Vitamin C is an essential element of plant and animal antioxidant systems, which can be defined as complex redox networks, including metabolites and enzymes, with mutual interactions and synergistic effects [36]. Antioxidants can spontaneously provide electrons to free radicals, alleviating the oxidative cellular environments caused by aerobic metabolism. Chemically, vitamin C is a dibasic acid with an enediol group on C2 and C3 of a heterocyclic lactone ring. At physiological pH, the hydroxyl group at C3 is deprotonated, giving a monovalent anion, which is indicated as ascorbate (ASC) [37]. The enediol group permits the donation of one or two electrons, forming monodehydroascorbate (MDHA) and dehydromacorbate (DHA), respectively [36]. The ASC redox potential ranges from +0.40 to +0.50 V [38; 39]; thus, the molecule can directly donate electrons to reactive oxygen species (ROS), such as singlet oxygen, superoxide anions, and hydroxyl radicals, as well as tocopheroxyl radicals (Figure 1) [36]. Being able to reduce toporeoxyl radicals, ASC is indirectly involved in the scavenging of lipid peroxides and radicals, contributing to the decrease of lipid peroxidation and, consequently, to the protection of membranes [40]. Due to the fast reduction of ROS by ASC, the damage of biomolecules can be prevented before the activation of antioxidant enzymes. Figure 1: Redox-dependent reactions of vitamin C. ASC can donate electrons directly to reactive oxygen species, metals, and toporeoxyl radicals. The reduction of H\({}_{2}\)O\({}_{2}\) by ASC occurs via APX. MDHA can undergo dismutation (green arrows), providing ASC and DHA. The reduction of oxidized forms occurs through the ASC-GSH cycle. MDHA and DHA are reduced by MDHAR and DHAR, respectively, whereas the reduced glutathione is recovered by GR (more details are provided in the text). Abbreviations: ASC, ascorbate; APX, ascorbate peroxidase; DHA, dehydromacorbate; DHAR, dehydromacorbate reductase; MDHA, monodehydroascorbate; MDHAR, monodehydroascorbate reductase; GR, glutathione reductase; GSH, glutathione; GSSG, glutathione disulphide. ASC can also reduce metals such as copper and iron, leading to the formation of ROS through the Haber-Weiss and Fenton reactions [41]. Thus, in some cases, ASC, acting as a reducing agent, will generate oxidants. This can occur in cell culture media, within the physiologic concentration of ASC, in presence of metals or in vivo in humans only when plasma and extracellular fluids contain millimolar concentrations of ASC [42]. Due to resonance stabilization of unpaired electrons, MDHA, derived from the loss of one electron, has very low reactivity with other radicals and, consequently, has very low toxicity. Two molecules of MDHA can spontaneously disimutate in ASC and DHA (Figure 1) [43]. DHA, if not rapidly reduced to ASC, will be permanently hydrolyzed to threonine, oxalate, oxalyl threonine, or tartrate [44]. ASC has low direct reactivity with H\({}_{2}\)O\({}_{2}\), but in plants, ASC works as a specific electron donor for ascorbate peroxidase (APX), a heme peroxidase which catalyzes the conversion of H\({}_{2}\)O\({}_{2}\) to H\({}_{2}\)O and O\({}_{2}\), giving MDHA (Figure 1). APX has a high affinity for H\({}_{2}\)O\({}_{2}\) and removes this ROS, even at low concentrations [36]. In plants, different APX isoenzymes have been identified in cytosol, mitochondria, peroxisomes, and chloroplasts, but all are coded by nuclear genes. The plant model system _Arabidopsis thaliana_ possesses six APX genes coding for six isoenzymes, two of which are targeted to the cytosol (APX1 and 2), two to peroxisomes (APX3 and 5), one to the thylakoid membrane (tAPX), and one that is dual-targeted to chloroplast stroma and mitochondrial matrix (sAPX) [45]. The crop species rice and tomato possess seven and eight APX isozymes, respectively [46]. APX is part of the ASC-glutathione (GSH) cycle (Figure 1), which is involved in ASC regeneration [47].