Biochemistry and Antioxidant Effects of Melatonin

Abstract

Melatonin (N-acetyl-5-methoxy-tryptamine) is a hormone taking place in many biological and physiological processes, such as reproduction, sleep, antioxidant effect, and circadian rhythm (biological clock), and is a multifunctional indolamine compound synthesized mainly from the metabolism of tryptophan via serotonin in the pineal gland. Melatonin, which is a hormone synthesized from the essential amino acid tryptophan, is substantially secreted from the pineal gland between the cerebral hemispheres found in the mammalian brain. In addition to this, it is also produced in the cells and tissues, such as the gastrointestinal system, gall, epithelial hair follicles, skin, retina, spleen, testis, salivary glands, bone marrow, leukocytes, placenta, and thrombocytes. It plays a role in many physiological events, such as synchronizing circadian rhythms, reproduction, fattening, molting, hibernation, and change of pigment granules, preserving the integrity of the gastrointestinal system with an anti-ulcerative effect in tissues and organs from which it is produced. Melatonin is also a powerful antioxidant and anti-apoptotic agent that prevents oxidative and nitrosative damage to all macromolecules due to its ability to form in metabolic activities, directly excrete toxic oxygen derivatives, and reduce the formation of reactive oxygen and nitrogen species. In this book chapter, we will explain the structure, synthesis, metabolism, and antioxidant effects of the melatonin hormone.

1. Introduction

Melatonin (N-acetyl-5-methoxy-tryptamine) is a hormone taking place in many biological and physiological processes and is a multifunctional indolamine compound synthesized mainly from the metabolism of tryptophan via serotonin in the pineal gland [1]. Its molecular formula is C₁₃H₁₆N₂O₂ and its molecular weight is 232.278 g/mol (Figure 1) [2]. It has the capacity to be able to pass through all biological membranes due to its small molecular size and high lipophilicity and it is evenly distributed to all biological tissues and fluids by crossing the blood–brain barrier [3].

Figure 1.

Chemical structure of melatonin.

Although the existence of the pineal gland has been known since ancient times, the French philosopher Rene Descartes described the pineal gland as the “throne of the soul” about three hundred years ago [4]. Melatonin hormone was first described in 1958 by the American dermatologist Aaron Lerner by obtaining from the pineal gland of cattle [5]. Melatonin, which is produced in the cells called pinealocytes of the pineal gland, is a hormone that plays a role in the regulation of many physiological and biological functions, such as circadian rhythm, sleep/wake cycle, pubertal development-reproduction, locomotor activity, regulation of immunity and blood pressure, molting, and hibernation [6, 7, 8, 9, 10].

Cortisol and melatonin levels act in opposite directions. Immediately after cortisol levels drop at night, melatonin levels begin to increase. The balance between these two hormones is important for good health and various diseases can occur with low melatonin and high cortisol levels [11, 12].

While melatonin shows a stimulating effect on the gonads in animals, such as sheep, goats, and deer, it shows suppressive properties in animals, such as horses, hamsters, and camels [13]. Melatonin acts as a timer by providing to follow-up the changes in the light/dark ratio of the animal seasonally [14].

In sheep, melatonin secretion and plasma levels are low in daylight [15, 16]. After sunset, melatonin secretion increases 10–20 times and rises rapidly to reach a peak by the end of the night. Thus, melatonin signal reflects the duration of the dark phase [14]. Melatonin initiates a series of events that lead to the start of the reproduction season [17]. The decreasing light exposure time increases melatonin secretion in the autumn-winter months when the days start to get shorter. Increased melatonin secretion stimulates gonadotrophin-releasing hormone (GnRH) secretion and provides to initiate estrus by acting on the hypothalamus in sheep [18, 19].

2. Melatonin

2.1 Tryptophan synthesis

The shikimate pathway is found in bacteria, fungi, plants, and algae, as well as in some protozoans. However, this pathway does not occur in animals and therefore animals must obtain aromatic amino acids from their diets as essential nutrients. Phosphoenolpyruvate (PEP), sugar with 3-carbon, product of the glycolysis pathway and erythrose-4-phosphate (E4-P), sugar with 4-carbon, synthesized from the pentose phosphate pathway starts with its conversion to 2-keto-3-deoxy-D-arabinoheptulosonate 7-phosphate, 7-carbon compound, with the hydrolysis of phosphate by 2-keto-3-deoxy-D-arabino-heptulosonate 7-phosphate synthase enzyme [20, 21].

In the second reaction, 3-dehydroquinate, a cyclic product, occurs by elimination of phosphate from 2-keto-3-deoxy-D-arabinoheptulosonate 7-phosphate by the catalysis of 3-dehydroquinate synthase enzyme with the help of NAD⁺. 3-dehydroshikimate is formed from this product with the effect of 3-dehydroquinate dehydratase enzyme as a result of the loss of the H₂O molecule [20, 22].

The next reaction is the reduction of 3-dehydroshikimate to shikimate by the shikimate dehydrogenase enzyme, which is used as a cofactor with NADPH. It is converted to shikimate 3-phosphate by the shikimate kinase enzyme by using one ATP per molecule. 5-enolpyruvylshikimate-3-phosphate is formed from shikimate 3-phosphate by binding to phosphoenolpyruvate (PEP) and catalyzing with 5-enolpyruvylshikimate-3-phosphate synthase enzyme. In the last reaction, 5-enolpyruvylshikimate-3-phosphate is converted to chorismate by the enzyme chorismate synthase [21, 23].

It forms anthranilate from chorismate formed in the last reaction of the shikimate pathway by giving amino group, which is part of the indole ring, in subsequent reactions for glutamine amino acid by way of the anthranilate synthase enzyme, which catalyzes the initial reaction of tryptophan biosynthesis [22]. N-(5′-phosphoribosyl) anthranilate is formed as a result of the elimination of pyrophosphate from phosphoribosyl-pyrophosphate (PRPP) by the enzyme anthranilate phosphoribosyltransferase. N-(5′-phosphoribosyl) anthranilate isomerase is responsible for the isomerization of N-(5′-phosphoribosyl) anthranilate to enol-1-o-carboxyphenylamino-1-deoxy-ribulose phosphate [24, 25].

Indole-3-glycerol-phosphate synthase catalyzes its conversion to indole-3-glycerol-phosphate by decarboxylating enol-1-o-carboxyphenylamino-1-deoxy-ribulose phosphate. In the last reaction, tryptophan synthase catalyzes the formation of tryptophan from indole-3-glycerol-phosphate by using indole and serine amino acids (Figure 2) [21, 26].

Figure 2.

Tryptophan synthesis.

2.2 Synthesis of melatonin

Anabolism: Tryptophan is a nonpolar amino acid containing an indole ring [27]. Tryptophan amino acid, which is in the class of essential amino acids, is required to be taken from the diet through nutrition since it cannot be synthesized in humans and monogastric animals [20].

Melatonin, which is synthesized from the amino acid tryptophan, is synthesized in bacteria, unicellular eukaryotes, and plants. Melatonin is synthesized from retina, gastrointestinal system, kidney, liver, thyroid gland, bone marrow, leukocytes, membranous cochlea, placenta, Harderian gland, gonads, breast tissue, adrenal gland, lung, skin, adipose tissue, blood vessels, lymphocytes, neutrophils, lymphoid tissues, and some brain areas, is mainly synthesized from the pineal gland [28, 29].

The pineal gland plays an important role as a functional neuroendocrine transducer of photoperiodic changes that occur in environmental or seasonal events by activating N-acetyl transferase transfer [30]. Norepinephrine is the most important transmitter in the postganglionic sympathetic nerve endings in this gland. The suprachiasmatic nucleus (SCN), which is one of the nuclei that can receive signals from the retina through nerves during the day and in light, effectively inhibits the release of norepinephrine from these nerve endings [31, 32]. In the dark, the release of norepinephrine from the nerve endings begins. Norepinephrine then binds to β-adrenergic receptors on the pinealocyte membrane and causes an increase in intracellular cAMP. As a result, it increases the activity of the N-acetyltransferase enzyme, which is a rate-limiting enzyme in melatonin synthesis in increasing intracellular cAMP and melatonin synthesis increases from serotonin [30, 32].

Melatonin synthesis begins with the uptake of tryptophan from the circulatory system by pinealocytes [33]. While tryptophan is converted to 5-hydroxy-tryptophan by the tryptophan hydroxylase enzyme catalysis, 5-hydroxy-tryptophan is also converted to serotonin (5-hydroxy-tryptamine) via aromatic L-amino acid decarboxylase [34]. N-acetyl serotonin is formed with the catalysis of 5-hydroxy-tryptamine N-acetyltransferase enzyme. N-acetyl serotonin is converted into melatonin (N-acetyl-5-methoxy-tryptamine) by being methylated with the catalysis of 5-hydroxyindole-o-methyltransferase enzyme (Figure 3) [28, 35, 36].

Figure 3.

Anabolism and catabolism of melatonin.

Catabolism: Melatonin is metabolized by isoforms of cytochrome P450 mono-oxygenase enzymes (CYP1A2, CYP1A1, and CYP1B1) found in the liver. Of these isoforms, both CYP1A2 and CY2C19 enzymes can demethylate melatonin to N-acetyl serotonin or can convert melatonin to 6-hydroxymelatonin by hydroxylation [37]. The half-life of melatonin, of which 70% is transported to the liver depending on albumin, varies between 3 and 45 minutes [38]. Less than 1% of melatonin is reported to be excreted with urine as 6-sulphatoxymelatonin by conjugating lesser extent with glucuronic acid or mostly conjugating with sulfate for the rest (Figure 3) [39, 40].

3. Antioxidant effects of melatonin

Free radicals, which are low molecular weight, short-lived and unstable structures, are highly active chemical structures that have unpaired electrons in their final orbits and try to share the electrons of other compounds to make up for this gap [41, 42]. Free radicals, which cause oxidation, are mainly oxygen-derived metabolites, superoxide anions (O₂⁻), hydrogen peroxide (H₂O₂), hydroxyl radical (OH⁻), and lipid peroxides [43]. Oxidative stress occurs as a result of increased reactive oxygen species (ROS) for various reasons and insufficient antioxidant mechanisms. Free radicals, which are formed by natural metabolic pathways in the body, are normally eliminated by radical scavenging antioxidant systems. Antioxidants are the molecules that prevent cell damage by inhibiting the formation of free radicals or scavenging existing radicals [44, 45]. While antioxidants can be classified according to their structure as those being enzymatic [superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), and glutathione-S-transferase (GST)] and non-enzymatic [Reduced glutathione (GSH), vitamin A, vitamin C, vitamin E, and melatonin], they can also be classified according to their cell localization as (i) intracellular antioxidants (SOD, CAT, and GPx), (ii) extracellular antioxidants (albumin, vitamin C, and urate) and (iii) membrane antioxidants (vitamin A and vitamin E) [46].

Melatonin hormone, which is a non-enzyme antioxidant, helps to eliminate harmful conditions stimulated by oxidative stress by inhibiting protein oxidation, lipid peroxidation, mitochondrial damage, and DNA degradation due to both its direct free radical scavenging activity and its contribution to the antioxidant defense system [47]. Melatonin, which is an antioxidant and was first suggested in 1991 by Ianaş et al., keeps radicals before the membrane and detoxifies them by protecting the membrane by attaching them to the outer surface of the cell membrane [48]. Melatonin hormone removes hydroxyl and oxygen radicals and inhibits nitric oxide synthase [49] and this feature of melatonin [50] that prevents lipid peroxidation reactions, especially by scavenging the OH˙ radical, is due to the pyrrole ring in its structure [51]. Melatonin increases the levels of antioxidant enzymes, such as mitochondrial SOD, cytosolic SOD, GPx, and GSH in the cell [52]. In addition, it is a powerful antioxidant that prevents oxidative and nitrosative damage due to its ability to eliminate toxic oxygen derivatives formed in metabolic activities and reduces the formation of ROS and reactive nitrogen species [32, 53].

Melatonin shows its antioxidant effect in three ways. (i) Direct antioxidant effect: Blocking free radicals with the formation of N₁-Acetyl-N₂-formyl-5-methoxyquinuramine from the pyrrole ring of melatonin in the presence of free radicals [54, 55, 56, 57], (ii) Indirect antioxidant effect: Suppressing oxidative stress by increasing the activity of enzymes, such as SOD, GPx, and GSH [54, 55], and (iii) Effect via prooxidant enzyme: Reducing free radical formation as a result of suppressing some prooxidant enzymes [48].

Antioxidant properties of melatonin have been shown in the studies conducted [58, 59, 60, 61, 62]. It has been reported that melatonin is at least two times more effective antioxidant than vitamin E and five times more effective than glutathione [55]. In the studies conducted, it is known that when melatonin is applied together with vitamin E and vitamin C, better protection is obtained than when applied alone [63] and melatonin has a suppressive effect on the formation of free radicals formed during electron transport in mitochondria. Melatonin reduces the formation of destructive toxic hydroxyl radicals by chelating transition metals taking place in Fenton/Haber-Weiss reactions [64]. It protects the biomolecules found in the whole structure of the cell against free radical formation by reacting with toxic hydroxyl radicals. Melatonin forms a non-enzymatic defense mechanism against the destructive properties of hydroxyl radicals and is more effective than other known antioxidants in protecting the organism against oxidative damage. It terminates lipid peroxidation by capturing the peroxide radical unlike antioxidants, such as ascorbic acid, alpha-tocopherol, and GSH. It has been reported that liver, kidney, and brain tissue glutathione peroxidase activity in rats increased after the administration of melatonin. Significant decreases in liver, lung, brain tissue, and glutathione peroxidase activity were reported in rats for which pinealectomy is made [48].

Many studies examining the effects on the synthesis and circulating amount of nitric oxide (NO), which is an important molecule of melatonin that plays a role in many physiological and physiopathological events, have been done. The physiological effect of NO occurs when soluble guanylate cyclase is activated to form cGMP. Decreased melatonin level causes decreased guanylate cyclase activity in many tissues. As a result, the cGMP level decreases, and the cAMP level increases. Thus, cell membrane thickness and rigidity increase, and degenerative damage formation accelerates [65]. When the relationship between NO and melatonin is examined, it has been suggested that the peroxynitrite anion, which is formed as a result of the reaction between NO and nitrosomelatonin and the reaction of NO and O₂ in the presence of O₂, is also occupied by melatonin [66, 67].

Unlike other antioxidants, it does not have a toxic effect in excessive use. Melatonin differs from classical antioxidants in various aspects and turns into less harmful pro-oxidant substances from the oxidants whose effects they abolish. However, after melatonin effects oxidant substances, is also effective as antioxidants in the intermediate stages and the resulting products. This property is very valuable as an antioxidant agent and is characterized as a “suicidal or terminal antioxidant” [68]. Melatonin reduces the synthesis of adhesion molecules and proinflammatory cytokines [69, 70]. Melatonin prevents linoleic acid, which is an energy source and growth factor of melatonin, which promotes the repair of damaged DNA, from entering the cell and suppresses its metabolism [71, 72].

4. Conclusion

The shikimate pathway, used by bacteria, fungi, algae, and plants, is a seven-step metabolic pathway for the biosynthesis of aromatic amino acids. However, this pathway is absent in animals. For this reason, melatonin, a neurohormone synthesized from tryptophan, an essential amino acid that must be taken with food, is synthesized by the retina, bone marrow, and gastrointestinal system, mainly by the pineal gland. Melatonin plays a role in the regulation of many physiological and biological functions in animals, such as the regulation of sleep time, blood pressure, and breeding season. In addition, due to its small molecular size and high lipophilic, it can reach all organelles of the cell and cross the blood–brain barrier. In addition, melatonin, which is a powerful antioxidant, provides direct scavenging of hydroxyl and oxygen radicals with high toxicity and stimulates antioxidant enzymes.