Feature

Reducing Free Radicals

We hear a lot about free radicals, but what are they?

Dr Steven Gieseg

Over the past few years, free radicals have been implicated in many different diseases. Every health supplement seems to include some protection against them -- but what are they and more importantly, what do they do?

A free radical is a chemical with a very reactive chemical bond. The chemical bonds which hold atoms together to make molecules contain pairs of electrons. For example there are two electrons in each of the bonds holding the hydrogens to the oxygen in the water molecule. The two electrons stabilise the bond between the atoms. However, some molecules, especially those containing oxygen, can easily gain only one of a pair of these bonding electrons. In a sense we have a molecule with a free chemical bond.

Essentially a free radical is a molecule containing unpaired electrons. An unpaired electron makes the molecule very reactive. The molecule will "steal" an electron from other molecules in order to gain a pair for its lone electron. The structure of the molecule that loses the electron is subsequently changed and can often become a free radical itself.

The most reactive free radical molecule known is the hydroxyl radical. It can be made by X-rays or gamma rays splitting water molecules. The hydroxyl radical (written HO to show the lone electron) is so reactive that it only takes one billionth of a second to react with a neighbouring molecules, usually by stealing hydrogen atoms from other molecules. The hydroxyl radical regains the lost electron in this manner and forms a water molecule (H₂O). To regain its electron pair, the hydroxyl radical has changed another molecule by removing a hydrogen electron from it. This causes further reactions leading to a major chemical change in the molecule. If the molecule the hydroxyl radical steals the hydrogen from is a piece of DNA, a genetic mutation may occur, resulting in a cancer gene being switched on.

Superoxide

Thankfully, exposure to gamma rays is not an everyday event for most of us. Superoxide, rather than the hydroxyl radical, is the most commonly encountered free radical in biology. Superoxide is an oxygen molecule with an extra unpaired electron and is usually written as O_2-. Superoxide is formed in the body either deliberately by white blood cells to kill invading bacteria and viruses, or as a leakage of energy when cells burn food molecules. In both situations it appears the superoxide formed can cause changes to the biological molecules of our bodies, which results in various types of damage. Some of this damage can appear as clinical diseases.

A major source of superoxide is the specialised white blood cells called macrophages and neutrophils. An enzyme, (NADPH oxidase) is found on the surfaces of these cells. This enzyme is activated when the macrophages and neutrophils encounter a foreign invading molecule such as from a bacteria. The enzyme adds an electron to the oxygen molecules around it, creating superoxide. Superoxide is not very reactive even though it is a radical. It usually reacts with itself to form hydrogen peroxide. Hydrogen peroxide is often referred to as a reactive oxygen species (ROS) and is toxic to many bacteria. It destroys the bacteria by oxidising various metabolic control molecules and generates further radicals within the bacteria by reacting with copper and iron.

The neutrophils can enhance the destructive power of hydrogen peroxide by reacting it with the salt in our body using an enzyme called myeloperoxidase. This enzyme is an amazing green colour in the test-tube but most people are familiar with it as the green phlegm in their handkerchief during a bad head cold. This enzyme converts the hydrogen peroxide to hypochlorite, also known as chlorine bleach. Chlorine bleach is a potent and lethal killer of bacteria. Unfortunately it also kills anything else it encounters, such as your healthy cells. When unleashed on a cell it destroys the enzymes and protein structures by adding chlorine atoms to them. This causes cell metabolism to grind to a halt and the cell dies. Hypochlorite production, at the wrong time, wrong place or in excess has been implicated in a number of diseases, including those of the lungs and possibly the heart.

How white blood cells produce hypochlorite (chlorine bleach) to destroy invading organisms.
The enzyme NADPH oxidase in the neutrophils (a type of white blood cell) adds electrons to the surrounding oxygen molecules, converting them to superoxide, which then reacts to form hydrogen peroxide. The enzyme myeloperoxidase combines the hydrogen peroxide with chloride.

We like to think of our body as a perfect machine but it is not. Five percent of the oxygen we breathe ends up as superoxide. The final stage of generating energy from the foods we eat involves a series of reactions where electrons are passed from one molecule to the next, forming a type of electric current. This process is known as the respiratory chain, and occurs in the cells' mitochondria. One time out of twenty, one of the electron carriers, called Coenzyme Q, passes the electron to oxygen instead of the next electron carrier, so producing superoxide.

The superoxide generated within the cell will react with a number of important molecules if not removed quickly. The most significant of these vulnerable molecules is nitric oxide, one of the central blood pressure-controlling molecules in the body. To prevent this superoxide mediated damage, all cells contain the enzyme superoxide dismutase. This enzyme speeds up the reaction between superoxide molecules that forms hydrogen peroxide. Ironically, hydrogen peroxide is, as mentioned earlier, toxic to cells and must also be removed.

The majority of the hydrogen peroxide is broken down to oxygen and water by the cellular enzyme catalase. In addition to catalase, the body also has a group of selenium-containing enzymes collectively called glutathione peroxidases. These enzymes break down hydrogen peroxide and any peroxides which form on fats and oils within the body. They are called glutathione peroxidases because they transfer the energy of the reactive peroxides to a very small sulphur-containing protein called glutathione. The selenium contained in the enzymes acts as the reactive centre, carrying reactive electrons from the peroxide to the glutathione. It is the glutathione that is the antioxidant in the reaction, not the selenium as many health food companies would lead us to believe. Selenium by itself is a potent oxidant which can be very toxic if too much is taken.

Antioxidants

The body creates free radicals and oxygen reactive species relentlessly and continuously. Antioxidants are compounds which provide our body with protection against the harmful effects of damaging free radicals and other reactive oxygen species. By definition, an antioxidant is a compound that is able to react with free radicals, forming harmless unreactive molecules and protecting other biological molecules from damage. Antioxidants are either reactive chemicals such as vitamin E or specialised enzymes such as catalase. The body produces enzymatic antioxidants but it cannot make antioxidant chemicals such as vitamin E, C and flavanoids. These antioxidant chemicals protect the sites in the body which the enzymatic systems cannot reach. We obtain these antioxidant chemicals from our diet but they are rapidly turned over in the body and need to be constantly replenished.

Vitamin E is an antioxidant which dissolves in our body's fats and oils. Any radicals formed in the fats will react with vitamin E to form a vitamin E radical. This vitamin E radical lacks the energy to cause any further damage but will react with vitamin C in the blood to regenerate vitamin E. The breakdown product of vitamin C is then removed by the kidneys. In this way, radicals formed in fats are removed from the body by transfer to vitamin E then to vitamin C and out through the kidneys as urine. Vitamin C also reacts with a number of water soluble radicals formed in the blood.

Flavanoids are another antioxidant which may be of great importance to our health. Flavanoids are ring-shaped compounds found in most plant tissues and usually have a reddish colour. They are compounds that give red wine its colour and, possibly, its beneficial health effects. It has been suggested by many scientists that one of the beneficial effects of eating fresh fruit and vegetables may be the intake of the flavanoid antioxidants. Commercial interest in these compounds as dietary supplements has made them the focus of a considerable amount of research both here and overseas.

Free Radicals

It appears that, for at least the first 20-30 years of our lives, our bodies are well protected from free radical damage. Assuming one has a healthy diet containing fresh fruit and vegetables, the levels of antioxidant molecules and antioxidant enzymes is usually high enough to absorb most of the free radicals produced in the body.

As we get older, the effectiveness of these protective systems appears to slowly decrease. It seems not all the free radicals produced are neutralised, so there is a slow build up of damaged molecules. The enzymatic antioxidant defence systems also appear to be vulnerable to free radical damage. As we age, there is a slow decrease in the amount of active radical-removing enzymes in our bodies. As friction wears out a machine, free radicals wear out the body. This is why severe complications of free radical damage appear as the diseases we associate with aging.

It is believed by many scientists and medical practitioners that increasing the dietary intake of antioxidants, either by increased consumption of fresh fruit and vegetables, or dietary supplements of vitamin E, C and possibly flavanoids, the processes of free radical damage and the associated diseases can be slowed. Proving this is a very slow and difficult process because free radicals are very reactive and therefore very short lived. Scientists cannot measure these radicals directly but must look for the damage they cause as an indication they are present.

Unfortunately, free radical reactions are, at best, described as being messy. The hydroxyl radical can form over a hundred different products when it reacts with a protein. The situation is even more complex with fats. Many of the products of free radical damage to fats are unstable and break down into even more complex compounds. The measurement of one of these compounds in the blood does not always mean that body is being damaged permanently as the body may be dealing with the damage successfully by removing it. To make the task more complex, diseases such as heart disease develop very slowly, over many years, at rates impractical to model in the test tube.

To overcome these problems scientists continue to develop model systems which show how the biological chemistry reacts to free radicals. New markers of free radical damage are also being developed to monitor what actually happens within the body. The goal of this research is to produce a non-invasive way of measuring the level of free radical damage in various diseases and to deliver the appropriate amount of antioxidant therapy.

Diet Best Source of Antioxidants

To date the best source of antioxidants, based on scientific evidence, is to consume a healthy well balanced diet, says Dr Carolyn Lister who heads a research project investigating which fruit and vegetables provide levels of antioxidants that will improve health and ward off disease. In some cases, the compounds present in supplements are not those with the highest antioxidant activity. More research is being done to determine these.

Some studies looking at the effect of beta-carotene and vitamin E on cancers and heart disease have shown no benefit, and some studies showed increased incidence of disease by those taking the supplements. More recently some supplements have been produced from fruit and vegetables or other plant extracts, and the initial results are more positive. However, the results of larger and longer trials are required before definite conclusions can be drawn.

Lister says the quantities of antioxidants present in supplements do not always relate to the amounts present in a healthy diet. There may be no additional benefit from having high doses of single compounds. Conversely some supplements may not give high enough levels of some compounds to reach desirable levels.

The array of antioxidants present in food, once ingested, interact in a complex way with one another and with other cell components. There is indirect evidence that antioxidant nutrients act synergistically, in combination offering more efficient protec- t ion than the sum of parts.

The mix of foods we eat and the interaction of antioxidants with other food components may play a role in how they are absorbed. Some antioxidants are fat soluble and others are water soluble, so what they are eaten with may influence how readily they are absorbed.

Antioxidants are compounds generally derived from our diet that provide protection against the harmful effects of free radicals in our bodies. Free radicals are very reactive with, and cause damage to our DNA, lipids and proteins. Our normal metabolic function produces free radicals and they occur in pollutants such as cigarette smoke and UV light. The main sources of antioxidants in the diet are fruit, vegetables and whole grains. Some drinks, especially tea and red wine, make significant contributions. Herbs, nuts and seeds are also rich in antioxidants but we eat them in smaller amounts.

Studies of population groups have produced interesting results. The "French Paradox" is the most well known of these. Many French people eat large amounts of fat, yet have a comparatively low rate of heart disease. It is thought that the natural antioxidants in wine, especially red wine, are responsible for the protective effects. Other studies of vegetarians and groups with higher than normal intake of fruit and vegetables have shown these people have a lower incidence of diseases such as cancer, heart disease and stroke. While the observational studies are interesting, good scientific evidence is needed so decisions about supplementing the diet are made on an informed basis.

Dr Steven Gieseg lectures in biochemistry and free radical biochemistry at Canterbury University.