Complete information on Bio-Transformation of Toxic Agents

A biological system is constantly exposed to a variety of foreign chemicals or xenobiotics, as they are sometimes called. These chemical agents may be natural or manmade and their absorption brings about plenty of interference in the system. A biological system is particularly defenseless against lipid soluble substances, which have rather a free access across the plasma membrane entering the cell simply by dissolving and diffusing through the lipid layer.

Though lipid solubility of a toxic agent is a property which facilitates efficient absorption, the solubility of these toxic agents in water appears to be an important factor in their excretion from a biological system. Thus, in general lipohilic chemicals are converted into hydrophilic ones if the biological system is to get rid of them quickly and efficiently.

In the process, chemical agents also undergo detoxification or lose their toxic properties. But at times these biochemical processes might cause such alterations in the chemical agent which result in their conversion to more toxic or active state. These biochemical reactions which alter and detoxify chemical agents are cumulatively known as biotransformation reactions. Although most of the chemicals which enter a biological system are subjected to the process of biotransformation, there are some which are excreted as such and do not undergo any change in their chemical structure.

1. Location of Biotransformation Reactions:

Liver is by far the most important organ in which most of the biotransformation reactions are carried out. Most of the enzymes which catalyse biotransformation reactions have been shown to be present in the cells of the liver on endoplasmic reticulum which is a net-work of interconnected structures made up of lipoproteins.

Tissues other than those of liver can also carry on biotransformation of foreign compounds. However, their capacity to do so is limited in as much as the variety and the amount of chemical agent metabolised, are concerned.

When liver cells are removed fragmented and homogenised, endoplasmic reticulum breaks up and its fragments round off to form small vesicles which are referred to as microsomes. These vesicles can be obtained in relatively pure state by centrifugation at a very high speed (105,000 g). This microsomal fraction of endoplasmic reticulum of cells of liver can carry out most of the biotransformation reactions in-vitro.

2. The Pattern of Biotransformation Reactions:

Toxic agents are metabolised in the liver cells by unique pathways which appear to exist mainly for the purpose and are capable of dealing with a tremendous variety of compounds which man’s ingenuity and synthetic skill can create. These reactions occur usually in the following two stages:

Reactions of Stage I:

Stage I reactions involve, hydrolysis, oxidation and reductions in which functional groups are altered, added or pre-existing functional groups are uncovered or exposed. These reactions result in products which have OH, COOH, NH₂ groups etc. Most of these reactions are catalysed by a common hydroxylating enzyme system located on smooth endoplasmic reticulum (microsomal fraction) of the liver cells and which is the most important enzyme system of Stage I reactions.

Due to its capacity to oxidise a number of substrates this enzyme is known as Mixed function oxidase (MFO) or Mono-oxygenase system. This enzyme system utilises a molecule of Oxygen per molecule of chemical agent oxidised and requires the presence of reduced cofactor, NADPH₂, which is a relatively non-specific primary eletrone donor. It shows the general pattern of functioning of this enzyme system. A major component of the MFO system is the haemoprotein, cytochrome P-450, so called because it forms a ligand with carbon monooxide which has a maximum absorbance at 450 nm.

It will be seen from the list of reactions given above that Mixed function oxidase system usually catalyses/general non-specific type of reactions which may result in activation, changes in activity or inactivation of the toxic agents. In general oxidative biotransformation results in detoxification while reductive biotransformation often results in formation of products more toxic than the original substrate. It is usually the lack of oxygen which promotes reductive transformations.

Reactions of Stage II:

Reactions of Stage II are Almost invariably synthetic or conjugation reactions which require energy. Original compounds or their metabolites are conjugated to various organic acids or other groups and this is accomplished by activating co-factors or substrates to high energy intermediates. These reactions usually result in the formation of highly water soluble compounds which are usually inert or harmless. Conjugation reactions occur at the site of reactive groups introduced during the Stage I reactions. If the required reactive sites are already present on the substrate molecules Stage II reactions may proceed without Stage I reactions.

Some of the important Stage II reactions are conjugation reactions with glucuronic acid, aminoacids (especially glycine), sulphates and acetates, all of which introduce strong acidic groups in the molecules concerned raising their water solubility. In addition to these methylation and conjugation reactions leading to the formation of mercapturic acid derivatives are also significant in detoxifying a variety of chemical toxicants.

1. Conjugation with Glucuronic Acid:

Various alcohols, especially those which are not readily oxidised, such as secondary and tertiary ones, phenolic, COOH and NH₂ groups may be conjugated with glucuronic acid resulting in the production of highly water soluble compounds. Glucuronic acid is a strong acid with a large number of OH groups due to which it becomes strongly hydrophilic substance.

2. Conjugation with Sulphates:

Usually phenols conjugate with sulphates to form sulphate esters which are highly water soluble and are quickly eliminated by active excretory mechanism. The ratio of organic and inorganic sulphates in urine is, therefore, an indicator of consumption of phenolic compounds or substances which are degraded to phenols.

3. Conjugation with Acetate:

This type of conjugation reaction occurs with toxic agents having amino-groups which are not suitable for oxidative domination. A common example of this type of reaction is that of aniline in which amino-group is attached directly to the aromatic nucleus. Acetylation although always results in detoxification of the chemical agent, it involves a decrease in the hydrophilicity of the substances which may at times causes some problems.

Methylation is an uncommon reaction taking place on nitrogen atoms and on unsatured heterocyclic compounds. It usually results in detoxification of toxic agents. Likewise the formation of mercapturic acid derivatives is rather a more complicated reaction involving organic chloro-and bromo-compounds (such as bromo-benzene), and results in replacement of halogen atom with a mercapturic acid group. The mercapturic acid derivatives are strongly hydrophilic substances which are readily excreted.

Important enzymes or enzyme-systems in Stage II reactions are glucuronosyl transferases, sulphotransferases, methyl transferases, acetyl transferases and glutathiones-transferases. These enzymes acting cumulatively with Stage I enzymes are capable of detoxifying most of chemical agents that may occur in nature or the synthetic skill and ingenuity of man can produce.

3. Hepatic Non-Microsomal Biotransformation:

In the sap of liver cells there are several enzymes which can perform several Stage II reactions. For example, N-acetyl-transferase, which is responsible for acetylation of drugs like sulphonamide and procainamide etc., is located in the liver cell sap. Thus, in addition to the enzyme system localised on the endoplasmic reticulum of liver cells, sap of these cells also plays a part in biotransformation activities.

4. Extra Hepatic Biotransformation:

Most of the foreign chemicals are biotransformed in liver. But there are many tissues in a biological system which are capable of carrying out the transformation reactions and regulate the fate of foreign chemicals. Tissues which are involved in absorption and excretion of chemical such as those of lungs, kidneys and gastro-intestinal track may metabolise toxic agents, though the rate and the degree of these biotransformation, may not be as fast and complete as those found in liver cells.

5. Induction of Biotransformation Enzymes:

Another feature of importance of the enzyme systems involved in biotransformation reactions is that these enzymes can be induced or enhanced by chemical treatment. Administration of various drugs and chemical agents increases the rate of production of these enzymes which often require de novo protein synthesis. Thus, exposure to a chemical or foreign agent sets into action the machinery which causes rapid production of biotransformation enzymes and this constitutes an important feature of defence against toxic agents with which an individual has to come up with.

Enzyme induction, however, also induces some morphologic and biochemical changes in the liver of the organism. Liver may undergo marked hypertrophy and there is a rise in the total functional protein content of the cell. The synthesis of phospholipids also goes up.

Endoplasmic reticulum enlarges and proliferates. All these reactions are part of a chain of activity which is set into motion by the presence of toxic agent in the system. It has been shown for certain cases that specific sites are available in the cytosol of the liver cells which upon being exposed to inducing agent form a ligand which is transported to nucleus of the cell and causes the derepression of the specific gene required. Derepression results in the transcription and protein synthesis is started which forms the enzymes necessary for the biodegradation of foreign agent.