Natural Phytotherapeutic Antioxidants in the Treatment of Mercury Intoxication-A Review

Heavy metals taken into the organism can make the toxic effects on the metabolism in various ways. For example, they may interact with proteins to alter and inhibit their enzymatic and structural functions. Mercury is one of the toxic elements that are widely distributed in nature. Mercury toxicity poses a serious threat to human health. It is an element that causes oxidative stress to increase in individuals, leading to tissue damage. Oxidative stress is the result of the imbalance between the production of oxidative species and cellular antioxidant defense. Phytotherapy continues to play an important role in health care. Natural phytotherapeutic antioxidants, exhibit a broad sequence of biological impacts, including anti-oxidative stress, anti-aging, anti-toxicicity and anticancer. Many studies have also shown that the phytotherapeutic agents play an important role in the removal of mercury from the tissue and in reducing oxidative stress. Our goal in this review was to investigate alternative ways of extracting the mercury in the tissue.


Hg (Mercury)
Mercury (Hg) is a silver, fluid, bright, odorless heavy metal. The symbol is "Hg" and the atomic number is 80. The symbol Hg "hydrated silver" comes from the Latin term hydrargyrum. Mercury is a stable element with a valence of +2. 11,12 Metallic or elemental mercury (Hg°) are the naturally occurring main forms of mercury. In nature, elemental mercury is found in the form of organic and inorganic compounds. The industry is mainly used in medical devices such as mercury fluorescent lamps, blood pressure monitors and thermometers used in many areas. It is widely used as filler material in dentistry, mine melting, cement making and paper production. 13,14 In a study conducted in the United States, mercury was reported to be the third most common environmental metal. 15 Mercury exposure can occur through respiration, feeding and food chain. Mercury is a heavy metal that is extremely toxic, which can have multiple adverse effects, and ultimately leads to cell death. Mercury, dysfunction in the skeleton of the cell and in the endoplasmic reticulum, significant cytoplasmic acidosis causes loss of mitochondrial function. 16 Chemically, mercury and its compounds can be examined in 3 different categories, (As shown in Table 1). Elemental (metallic) Mercury; Elemental mercury may evaporate at room temperature. And steam is rapidly absorbed from the lungs and it spreads to the central nervous system. 17 Inorganic mercurials include ammoniated mercury(ClH2HgN), mercuric chloride (HgCl2), mercuric oxide (HgO), mercuric sulfide, mercuric iodide (HgI2) , and the phenylmercuric salts. (C8H8HgO2) Organic mercurials include ethylmercury (C2H5Hg + ), methylmercury (CH3Hg + ), thimerosal (merthiolate), and merbromin (C20H9Br2HgNa2O6 -mercurochrome). 18,19 Methyl and phenyl mercuric compounds, such as metallic mercury, erythematosus and pruritic skin diseases. 18,44,45 In people with tattoos containing red pigment of the origin of mercuric sulfur (cinnabar-vermilion, Chinese red), they may experience inflammation that is limited to this region within 6 months of tattooing. 18,46 Inflammation findings: Inflammation caused by the influence of heavy inorganic mercury causes the tendency to bleed in gingiva and oral mucosa. It increases the salivary secretion, causing sensation of metallic taste in the mouth. Gingiva, a gray line is formed, especially when the oral hygiene is bad. 29 Birth defects and reproductive system: There is also evidence that mercury poisoning leads to Young syndrome (bronchiectasis, low sperm count, impairs sperm quality.). [47][48][49] It has been determined that all forms of the mercury can pass to the placenta at varying gauges 50 Mercury exposure is dangerous for the baby, because the baby's neurological tissues develop during early gestation. 51 Immune system findings: Mercury compounds reduce the number of T lymphocytes. In addition, mercury exposure causes a decrease in T cell GSH content. Mercury is an immunotoxic agent. [52][53][54] In the case of inorganic mercury exposure, elimination occurs via urine and faeces. Organic mercury compounds predominantly excrete in humans. 11 Mercury, promotes the formation of reactive oxygen species (ROS) such as hydrogen peroxides. 55,56

ROS (Reactive Oxygen species)
Free radicals are highly active atoms or molecules that can be produced in many physiological and pathological processes, carrying one or more unpaired electrons in their orbit. These highly unstable atoms or molecules tend to react with molecules in their environment and to share these electrons. 57,58 Free radicals can be positively charged, negatively charged, or neutral and are most often formed by electron transfer in biological systems. The most important free radicals in biological systems are the oxygen radicals. In addition, another source of free radicals is the nitrogen molecule. There is no toxic effect of O2, but it becomes free oxygen radicals during aerobic cell metabolism. By partial reduction of O2, OH and O2are formed. [58][59][60] Superoxide Radical (O2 •-) Superoxide is the first radical to appear in living organisms. In almost all aerobic cells, reduction of oxygen by an electron takes place. 61 The superoxide radical plays an important role in the formation of other reactive oxygen species, such as H2O2, HO2or 1 O2. 58,62 The superoxide radical is produced either directly in mitochondria during oxidation or enzymatically by xanthine oxidase (XO), cytochrome p450 and other oxidases. Superoxide dismutase (SOD) enzyme or in H2O2 is spontaneously inactivated. 63,64 Combined with superoxide (O2 •-) and the free radical NO -, comes the reactive nitrogen derivative ONOO . (Peroxynitrite). ONOO-has harmful effects on direct proteins. 65

Hydroxyl radical (•OH)
The hydroxyl radical is the most reactive radical. It reacts with lipids, polypeptides, proteins, DNA and other molecules (such as thiamine and guanosine). 62,66,67 Singlet oxygen ( 1 O2) Singlet oxygen is a nonradical and induced status. Compared with other ROS, singlet oxygen is rather mild and non toxic for mammalian tissue. 68 1 O2 is a cell signal and messenger; redox active agents regulate ion channel activity in animals and plants. In the human organism, singlet oxygen is both a signal and a weapon, with therapeutic potency against various pathogens such as microbes, viruses, and cancer cells. 62

Hydrogen peroxide (H2O2)
There is no unpaired electron in the hydrogen peroxide molecule, and so it is not a radical. Hydrogen peroxide can be generated through a dismutation reaction from superoxide anion by superoxide dismutase (SOD). 58,62,63 Enzymes such as amino acid oxidase (AAO) and xanthine oxidase (XO) also produce hydrogen peroxide from the superoxide anion. H2O2 is the least reactive molecule among ROS and is stable under physiological pH and temperature in the lack of metal ions. H2O2 can produce singlet oxygen thanks to react with superoxide anion or with HOCl or chloramines in living systems. 62,68 Free radical species (ROS) affect all important structures of cells such as proteins, carbohydrates, lipids, DNA and enzymes. 58,69 Effects of ROS on lipids and proteins Free radicals have to cross the cell membrane in order to interact with the cell components. Inasmuch as cell membranes are rich in polyunsaturated fatty acids (PUFAs) and cholesterol, they are easily affected by oxidant radicals. 70 Free radicals move away the hydrogen atom from the fatty acid chain. Lipid peroxidation is oxidation of polyunsaturated fatty plural form with free radicals. The main primary products of lipid peroxidation are lipid hydroperoxides (LOOH). This peroxidation results in products such as MDA, 4-hydroxynoneal (HNA), 8-iso-prostaglandin F2α (8-iso-PGF2α), alcohols, ethane and pentane. MDA is mutagenic since DNA can react with nitrogen bases. 58,71 Additionally it is genotoxic and carcinogenic on cell cultures. Membrane damage caused by lipid peroxidation is irreversible. MDA is the most mutagenic product of lipid peroxidation. 4-HNE is the most toxic. 4-HNE is considered as the second toxic messengers of free radicals, one of the major generators of oxidative stress and a major lipid peroxidation product. 58,70,72 8-Iso-prostaglandin F2α (8-iso-PGF2α), a major F2-isoprostane, is biosynthesized in vivo through nonenzymatic free radical-catalysed peroxidation of arachidonic acid. 21,22,58,70,73,74 Proteins are defined as the major targets of oxidative damage. The products of cellular metabolism or environmental induced ROS changes in the amino acids of proteins and cause loss of activity of protein function / enzymatic activity as well. 75,76 Oxidative protein modifications may take place in different ways. ROS directly interacts with protein or Interaction of compounds such as carbohydrates, lipids, and nucleic acids with ROS can interact with proteins with the resulting products. 77 In this way, reactive and non-protein compounds react with proteins to form a wide array of structures. 78,79 Protein oxidation reactions are usually divided into modifying the protein construct and modifying the amino acid side chains. 80,81 In addition to the modification of the protein in proteins, amino acid side chains are target for ROS. Sulfide containing amino acids in the structure are highly sensitive to cysteine and methionine. 82 Aromatic structures are also the main targets for ROS. The oxidatively modified tyrosine, phenylalanine and tryptophan are usually oxidative damage a demonstration. 76 Oxidation of lysine, arginine, proline or threonine may result in the formation of carbonyl derivatives. 83,84 Protein carbonyls are among the most commonly used products for determination of the proteins of the oxides. 58,85 Protein carbonyl levels area well-used marker for oxidative stress. The toxic effects of mercury can be prevented by antioxidant defense mechanisms to a certain extent.

Reactive nitrogen species (RNS)
NO reacts with the superoxide radical or molecular oxygen, leading to the formation of various reactive intermediates called reactive RNS. RNS consist of nitrite derivatives such as NO, NO2 -and OONO -. 86

Nitric oxide (NO•)
Nitric Oxide (NO) is highly stable at high concentrations in an oxygen-free environment and stable at low concentrations in the presence of oxygen. NO is a signal molecule of low molecular weight known to be biologically active in mammalian cells. The NO radical is synthesized from L-arginine by nitric oxide synthase (NOS) enzyme catalysis in vascular endothelial cells. 87 NO is an important effector and messenger molecule that plays a role in various biological processes such as immune response, smooth muscle tone, apoptosis, angiogenesis and nervous system. 88 In addition, NO is a molecule that regulates numerous pathological and physiological states. 89 NO has very important physiological functions at low concentrations. NO binds with molecular oxygen to form nitrogen dioxide (NO2). Another important effect of NO is to produce a strong oxidant peroxynitrite (ONOO -. ). 90 NO is a mediator with autocrine and paracrine effects in hemostatic events and in the defense mechanisms of the organism. The most important function of NO is to function in parallel with the effects of interleukin-1 (IL-1) and cytokines in various tissues of the body. It is produced by macrophages, neutrophils, hepatocytes and endothelial cells. 91 However, at high concentrations it shows toxic effects on normal cells. Spontaneously decomposes to form nitrogen dioxide. 92 In mammals, NO can be produced with three different isoforms of enzymes NO synthase. NOS enzymes are found in two basic isoform. These; is called constitutive or structural (cNOS) and inducible (iNOS). Structural NOS enzymes have two isoforms, endothelial NOS (eNOS) and neuronal NOS (nOS). eNOS is located on the membrane and is synthesizing the endothelium-induced relaxation factor; nNOS produces NO2, the messenger molecule in the central nervous system and neurons. Structural NOS is Ca + 2 / calmodulin dependent as cofactor and produces small amounts of NO at intervals with low activity. 93,94 Peroxynitrite (OONO-) Peroxynitrite is an important biological oxidant formed by the reaction of nitric oxide and superoxide radicals. Peroxynitrite can cause oxidative damage, nitration, and S-nitrosylation of biomolecules including proteins, lipids, and DNA. 93 Peroxidation of lipids in the membrane distorts membrane integrity by reducing the fluidity, elasticity and permeability of the cell membrane. These radicals constantly increase the level of Ca 2 + in the cell and cause cytotoxic effect on the cell by inhibition of mitochondrial respiration and electron transport chain, decrease of ATP production and activation of radical generating enzymes. 95,96 Antioxidants Although free radical reactions are necessary for the defense mechanism of neutrophil, macrophage and other immune system cells, they result in overproduction of free radicals, tissue damage and cell death. The half life of ROS is short. However, they initiate free radical chain reactions that cause tissue damage. For this reason, defensive mechanisms against oxidative damage triggered by free radicals act. These are preventive mechanisms, repair mechanisms, physical defenses and antioxidant defenses. 57 Antioxidant defense is the prevention or delay of oxidation of oxidizing agents such as proteins, lipids, carbohydrates and DNA in living cells. The substances that play a role in this process are called 'antioxidants'. 97,98 Enzymatic antioxidants are Superoxide dismutase (SOD), Catalase (CAT), Glutathione peroxidase (GSH-Px), Glutathione reductase (GR), Glutathione S-Transferase (GST) ,enzymes. The SOD structure contains copper (Cu), zinc (Zn) and manganese (Mn). GSH-Px contains selenium ions. For this reason, these enzymes are called metalloenzymes. 99,100 In contrast to the intracellular environment, E and C vitamins, transferrin, ceruloplazmin, albumin, bilurubin, β-carotene are responsible for the non-enzymatic antioxidant defense in the extracellular environment. In addition, E and C vitamins have antioxidant properties within the cell. 58,65,72,101 Enzymatic antioxidants Superoxide dismutase (SOD): By catalyzing the conversion of O2•radical to H2O2, protects the cells from harmful effects of O2•radical. It also inhibits lipid peroxidation. SOD plays a role in controlling the levels of O2 •in the parts of the cell Superoxide dismutase activity varies with tissues. It is mainly intracellular and 10% is located outside the cell. 58

-Manganese superoxide dismutase (Mn SOD):
Mitochondrial Mn-SOD is a homotetramer containing one Manganese atom per subunit. Mn-SOD has the same reaction as Cu-Zn-SOD. However, it is a completely different enzyme in its structure. It contains Mn in its active site and is not stable. This form of SOD is not inhibited by cyanide. 105

Catalase (CAT):
Catalase is a hemoprotein that has four groups in its structure. Catalase converts hydrogen peroxide (H202) to water (H20). Catalase's effect is similar to SOD. 106 H2O2 + H2O2→H2O+1/2O2 Glutathione peroxidase (GSH-Px): GSH-Px can be found in two forms, selenium-bound and seleniumbound. Selenium based group, reducing hydrogen peroxide and other organic peroxides. It consists of four members. These are GSH-Px1 (celluler-GSH-Px), GSH-Px2 (GSH-Px-gastrointestinal), GSH-Px3 (plasma-GSH-Px) and GSH Px4 (PH-GSH-Px), respectively. 107 -GSH-Px1 or cellular GSH-Px (cGSH-Px), tetrameric in structure is a cytosolic enzyme. GSH-Px1 is active against organic hydroperoxides and H2O2. 108 -GSH-Px 2, or gastrointestinal GSH-Px (GSH-Px-GI) found in the gastrointestinal tract, but not in the kidney, heart and lung. 109 112 Phase-II detoxification is a member of the enzyme family. In addition, it prevents oxidative products or foreign toxic substances from merging with other macromolecules in the body and provides them to be removed without harming the cell components. Therefore, GSTs are one of a group of enzymes that are very important guardians. 113 GSTs are divided into three families as mitochondrial, cytosolic and microsomal. 114 Mercapturic acid plays an important role in the initial reactions of biosynthesis. The mercapturic acid formation process catalyzed by GSHconjugation of GST is generally described as detoxification reactions. The ability to reduction feature GSTs protects membrane components from lipid peroxidation. In addition, 4-hydroxy alkenals, products of lipid peroxidation in aldehyde structure, are conjugated with GSH. 115 GSTs, also considered as one of the natural protective systems, have an important role in the detoxification of electrophilic xenobiotics such as herbicides, pesticides, anticancer drugs, chemical carcinogens and environmental pollutants. 116 Glutathione reductase (GR): Glutathione reductase is an antioxidant enzyme that converts oxidized glutathione (GSSG) to reduced glutathione (GSH). GR uses NADPH as the coenzyme when performing catalysis. 117 The physiological GSH-GSSG ratio in the cells is of great importance. In the absence of GCSG, the level of intracellular NADPH is reduce and GR is inactivate. As the intracellular level of GSSG increases due to the oxidative stress, GR re-activates. 118

Nonenzymatic antioxidants
Nonenzymatic antioxidants; It is examined in two parts as natural antioxidants and synthetic antioxidants. This review will discuss natural antioxidants. For this reason, synthetic antioxidants were excluded from the discussion. 119 Glutathione (GSH): GSH is made from three amino acids: glycine, cysteine and glutamic acid. This tripeptide exists in reduced (GSH) and oxidized (GSSG) forms. The relative amounts of every form determine the cellular redox status (GSH/GSSG ratio) which is often used as a sign of antioxidative capacity of cells. Glutathione (GSH) has vital importance in fighting oxidative stress. It is a strong free radical and reactive oxygen species scavenger. 120,121 Vitamin E: Vitamin E has eight isoforms, α-, β-, γ-, and δ-tocopherol and α-, β-, γ-, and δ-tocotrienol. Vitamin E is a lipophilic radical-scavenging antioxidant. 122 Vitamin C: Vitamin C is a potent antioxidant protecting the body against endogenous and exogenous oxidative challenges. 123 Uric acid: Uric acid demonstrated its ability to scavenge reactive radicals resulting from harmful process, such as autoxidation of hemoglobin, or peroxide generation by macrophages. it is an efficient scavenger of singlet oxygen, peroxyl and hydroxyl radicals and protects erythrocyte membrane from lipid peroxidation. 124 Retinoids and carotenoids: Retinoids and carotenoids take place in the structure of lipids and cell membranes. In the singlet oxygen suppression to prevent the harmful effects of flavin and porphyrin, they work in preventing peroxide radicals. 101

Discussion
Plants are an exogenous source of antioxidants taken in the diet. It is believed that two thirds of the plant species in the world have medical prescription, and almost all of them have excellent antioxidant potential. 125 Increased exogenous antioxidant supplementation or endogenous antioxidant defense has been found to be effective in combating undesirable effects of oxidative stress. 126 The main natural antioxidants present in vitamins and protecting the human body from harmful free radicals are mainly vitamins (C, E and A vitamins), flavonoids, carotenoids and polyphenols. 127 Phenolic compounds exhibit physiological properties such as anti-allergic, anti-atherogenic, antimicrobial, anti-inflammatory, antioxidant, anticancer, antithrombotic, cardiovascular and vasodilatory effects. [128][129][130] In addition, Fruits, spices and many medicinal herbs are rich sources of pharmacological properties. These agents have antioxidants, free radical scavengers and anti-toxic properties. 16,23,[131][132] As shown in Table 2, many natural antioxidants against mercury poisoning have been tested for detoxification.

Conclusion
In summary, this study provides evidence that some natural antioxidants play a protective role against Mercury-derived toxicity. It provides foundation of studies of natural phytotherapeutic agents on mercury treatment. This study also provided information for candidate antidote, pharmaceutical agents in the treatment of mercury-induced toxicity. However, high doses of antioxidant supplements often do not work well or can be harmful. More research is needed for effective and safe antioxidant doses against mercury poisoning.

Ethical Issues
Not applicable.