Atoms and molecules possess electrons in shells, with each shell containing two paired electrons. Free radicals are formed when atoms or molecules possess electrons that are unpaired, and this tends to make the molecules very reactive. In biological systems such as cells and tissues, these free radicals can react with cellular components and cause damage by rapidly taking electrons from the surroundings. This can result in cellular damage and if left unchecked can lead to chain reactions that cause chronic diseases such as cancer or heart disease. However, under normal circumstances, the body is able to protect itself from free radicals by use of a range of antioxidants. Antioxidants are effective at scavenging free radicals because they can donate the electrons needed by the free radical in a chemical process called reduction.
In the process of reducing free radicals by donating electrons, antioxidants become oxidised to free radicals themselves, because they now possess unpaired electrons. However, in this oxidised state antioxidants tend to be less reactive than the original radical. This is because they can delocalise the negative charge of the unpaired electron on account of their molecular structure. For example, flavonoids are able to scavenge free radicals and then stabilise their unpaired electron using their phenolic ring structures. Antioxidants do not work in isolation, but interact with each other extensively in complex recycling systems. Antioxidants that have been oxidised while scavenging free radicals can be scavenged themselves to their reduced state by other antioxidants. The cell therefore contains interconnected networks of oxidation reduction (redox) reactions as electrons are passed from molecule to molecule.
Glutathione should be considered the master antioxidant because it plays such an important part in the antioxidant defence of cells. Higher levels of glutathione are associated with reduced cellular damage because glutathione protects the cellular components free radicals. Genetic defects that reduce cellular glutathione are associated with increased cell damage and accelerated aging. Glutathione is tripeptide belonging to a chemical group called thiols which possess a sulfhydryl residue (-SH). Thiols are important antioxidants because chemically they can act as reducing agents. Reduced glutathione (GSH) is able to donate electrons to free radicals within cells and in the process becomes oxidised to a thiyl radical (C-S∙). Thiyl radicals are unstable and two will combine to form glutathione disulphide (GSSG). Glutathione disulphide can then be recycled back to reduced glutathione by the enzyme glutathione reductase using NAD(P)H as a donor for hydrogen.
Figure 1. Cellular antioxidant recycling. GPx = glutathione peroxidase which requires selenium. Adapted from Sen, C. K. and Packer, L. 2000. Thiol Homeostasis and supplements in physical exercise. American Journal of Clinical Nutrition. 72: 653S-669S
The reduction of free radicals by glutathione is dependent on the enzyme glutathione peroxidase, an enzyme which catalyses the reaction of reduced glutathione (GSH) to glutathione disulphide (GSSG). Glutathione peroxidase is a well researched selenoprotein that requires the trace mineral selenium to function properly. Selenium is therefore considered a dietary antioxidant because of its role in cellular antioxidant defences. Neutrophils are one cell type with high concentrations of glutathione and glutathione peroxidase. Neutrophils produce large amounts of free radicals in order to destroy the cellular components of invading pathogens. The neutrophils are protected from damage by the free radicals because of their cellular thiol antioxidant system. Without selenium, glutathione peroxidase and glutathione, the free radicals would destroy the neutrophils. As the number of neutrophils decreased the body would become more susceptible to infection.
Lipoic acid (lipoate) is another thiol compound (possess a sulfhydryl residue -SH) that acts as an important antioxidant in cells. Lipoic acid is produced naturally in the body but is also present in the diet in foods such as spinach, broccoli, tomatoes, brussel sprouts and peas. Dietary lipoic acid is rapidly taken up by the cells where it is reduced to dihydrolipoate by enzymes such as dihydrolipoamide dehydrogenase using NAD(P)H as a hydrogen donor. Dihydrolipoate is able to scavenge a wide range of free radicals making it one of the most powerful cellular antioxidants. Dihydrolipoate is also released to the extracellular fluid where it is able to reduce the amino acid cystine to cysteine. This increases cellular levels of cysteine which subsequently reacts with glutamate and glycine to produce more glutathione. Dihydrolipoate is thus able boosts cellular levels of glutathione.
One of the key functions of thiols such as glutathione and dihydrolipoate is the recycling of both vitamin C and vitamin E. Vitamin E (tocopherol and tocotrienol) is a fat soluble compound which accumulates in the phospholipid membranes that surround cells and protects them from free radicals. Vitamin E is able to donate electrons to the lipid peroxyl radical thus scavenging the radical. In the process the vitamin becomes oxidised to the tocopherol or tocotrienol radical but remains relatively unreactive because it delocalises the unpaired electron over its chromanol head. It is estimated that there is just 1 vitamin E molecule for every 2000 molecules of phospholipid in the cellular membrane, yet despite this vitamin E is able to effectively protect the membranes of cells because it is continually recycled back to its reduced form by cellular thiol compounds.
In contrast to vitamin E which accumulates in the phospholipid membranes, vitamin C is a water soluble compound and so it concentrated mainly inside the cells. Here it is an effective antioxidant against water soluble radicals. Vitamin C is able to react with the superoxide radical to form hydrogen peroxide, in the process becoming converted to dehydroascobic acid. Hydrogen peroxide can then react with vitamin C to form water and dehydroascorbic acid. Vitamin C can also donate electrons to the hydroxyl radical forming the fairly non-reactive semihydroascorbate radical in the process. In addition, vitamin C is able to donate electrons to the vitamin E radical and thus recycle vitamin E to its reduced form. Thiols are able to scavenge vitamin C radicals and reform reduced vitamin C so that it can continue to perform its important radical scavenging function within the cells.
Increasing cellular levels of glutathione is an important step towards optimum nutrition. Supplementation with oral glutathione has been shown to be ineffective in raising cellular levels of glutathione. However, research has demonstrated that supplemental lipoic acid and vitamin C both increase cellular levels of glutathione. Because selenium is required for glutathione peroxidase function it would also seem sensible to maintain adequate intake. While vitamin E is the most important antioxidant in the phospholipid membrane, carotenoid antioxidants are also able to prevent lipid peroxides. Therefore a wide range of coloured plant foods should be consumed. This will also provide a range of flavonoid antioxidants which make up the bulk of the antioxidant contents of fruits and vegetables. While co-enzyme Q10 is an important cellular antioxidant, it is produced within the body and so supplementation for antioxidant purposes in healthy people is not necessary.
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