BCH 318 - Xenobiotics - Dr. Adegbesan 2022-2023 PDF

BCH 318: XENOBIOTICS (2 UNITS; STATUS: C) COURSE LECTURERS: DR. B.O ADEGBESAN AND DR E.O ADESANYA  COURSE CONTENTS 1. Allergy and its biochemical basis - Dr. Adegbesan 2. Detoxification mechanism...

BCH 318: XENOBIOTICS (2 UNITS; STATUS: C) COURSE LECTURERS: DR. B.O ADEGBESAN AND DR E.O ADESANYA  COURSE CONTENTS 1. Allergy and its biochemical basis - Dr. Adegbesan 2. Detoxification mechanism - Dr. Adegbesan 2. Toxicological effects of expired drugs - Dr. Adesanya 3. Side-effects of drugs as a result of use and or misuse - Dr. Adesanya 4. The biochemical modes of action of pesticides and herbicides - Dr. Adegbesan 6. Toxins in processed foods and beverages - Dr. Adesanya 7. Biochemistry of Aflatoxins and mycotoxins - Dr. Adesanya 8. Screening for toxic pollutants in foods, water etc. - Dr. Adesanya 9. Toxicological studies of food additives and food preservatives - Dr. Adegbesan  INTRODUCTION Xenobiotic or Xenobiochemistry as a course introduces and describes the various types of xenobiotics or chemicals including pesticides and agrochemicals, their biochemical mode of action and their effects on foods and water; biochemistry of Aflatoxins and mycotoxins and toxicological studies of food additives and food preservatives. The course also describes the various screening techniques used in identification and screening of toxic pollutants in foods and beverages industries. The biochemical basis of allergy resulting from drugs use and misuse is also within the scope of this course.  WHAT ARE XENOBIOTICS?  Xenobiotics or toxicants or drugs or pollutants are substances that are foreign to the body which upon introduction to the body can cause change in structure and functions of such an organism.  They get into the body from external sources i.e they are not usually produced within the body but there exist some few endogenous ones. TYPES OF XENOBIOTICS 1. All therapeutic drugs 2. drug metabolites 3. Food additives which include colouring, flavours, sweetners (sucrose, saccharine); food preservatives 4. Environmental compounds such as pollutants and industrial chemicals (chloroform, acetone) 5. Agrochemicals: herbicides, pesticides (insecticides, rodenticides, e.t.c) 6. Cosmetics Xenobiotics can be -(a) Exogenous - The foreign molecules which are not normally ingested or utilized by the organism but they gain entry through dietary food stuffs, or in the form of certain medicines/ drugs used for a therapeutic cause or are inhaled through environment. Examples- Drugs, food additives, pollutants, insecticides, chemical carcinogens etc. Xenobiotics can be -(b) Endogenous – Though they are not foreign substances but have effects similar to exogenous xenobiotics. These are synthesized in the body or are produced as metabolites of various processes in the body. Examples- Bilirubin, Bile acids, Steroids, Eicosanoids and certain fatty acids. ALLERGY AND ITS BIOCHEMICAL BASIS  ALLERGY AND ITS BIOCHEMICAL BASIS  Allergy can be described as an altered or abnormal tissue reaction. It is an exaggerated response by certain tissues to some foreign substances. This occurs only in hypersensitive people but in normal people, the same amount of the same foreign substance which is usually a protein produces no reaction at all.  Allergic reaction results from contact between a foreign protein called ALLERGEN and body tissues that are sensitive to it. Allergy reactions to substances in food are sometimes spontaneous, peculiar to certain humans and rarely demonstrable in lower animals. There are therefore no good animal model for evaluating such reactions.  The most common afflictions caused by a food allergy is of the skin and respiratory tracts. The GIT is less frequently involved. This is because GIT has a defense system.  THE GASTRO INTESTINAL DEFENSE SYSTEM  The GIT is served by defense barrier which prevents the penetration of a great variety of potentially harmful antigenic substances and micro- organisms. Substances which survive the effect of digestive juices and the protective effect of mucous secretion including the effects of immunoglobulin antibodies may still be able to penetrate the epithelial barrier of the gut.  The gut is a major immunological organ and it contains about 1010 lymphocytes. When these lymphocytes are activated, they reach circulation via the mesenteric thoracic ducts. Occasionally, food allergy affects the Central Nervous System (CNS) and produces a manifestation similar to migraine headache and in some severe forms, it may predispose to convulsive seizures. The allergic response of individuals may be evaluated by skin test.  A Graminaceae family of food commonly produce allergy response in people and these include barley, corn, rice, oats and wheats. Of the legumes, soya bean has been described as an Allergen when it is used as milk substitute.  A single allergen may cause reactions at several sites. Ingested proteins do not usually enter the blood stream unchanged but rather they are split into their components’ polypeptides and amino acids in the guts; This after absorption have no allergic properties but sometimes small number of proteins may be absorbed and changed, then allergy manifestation occurs.  The detailed nature of allergy reactions has not been clearly understood but there is little doubt that it results from an abnormal effect of the combination of the combination of antigenic allergen with antibodies within the tissue cells. An antigen is a chemical compound present in living or non-living proteins that is capable of stimulating a defensive response while An antibody is a defensive substance that neutralizes an antigen by combining with it. Cell damage apparently results which leads to the release of histamine which probably contribute to the clinical effect. Decarboxylation of histidine produces histamine which after production is stored in parts of the body. However, it is released during ANAPHYLAXIS and as a result of allergy. ANAPHYLAXIS is a shock reaction to an injection of an antigen which are animals that have been previously injected with the same antigen shows. Normal immune reaction between antigen and antibody does not evoke any allergy effects. Allergic reactions are of two main types and these are: 1.IMMEDIATE REACTION TYPE: In this type, the allergic manifestations occur within 4hrs of the patient coming in contact with the allergen. It may even be within secs or mins. Skin tests are nearly all positive in this type of allergy. 2.DELAYED REACTION TYPE: Here, manifestations may not appear for many hours or even for many days. In this type, skin tests are usually negative. Once the allergic reaction has developed, the resulting manifestation in the gastrointestinal, respiratory and nervous systems are the same in both types. Allergic reaction may be trivial such as in URTICARIA or nettle rash which is a widespread itching or it may be so serious to result in sudden death.  TREATMENTS The treatment of allergic patient is not always easy. It is possible to inject small amount of foreign substances into the skin and so discover which substances are likely to produce the allergic response. 1. A vaccine may then be prepared, if this is injected regularly in increasing amount, it may often reduce the state of allergy. 2. Another treatment type is the elimination of the causative foods i.e the causative food factor is identified and eliminated to prevent the occurrence of symptoms. 3. Substitution of alternative foods 4. The use of drugs such as adrenaline, epinephrine, corticosteroid, aminophylline etc. Note: The prescription of tranquillizers and sedative is often indicated because of connection between emotion and allergy. Detoxification mechanism  DETOXIFICATION OR BIOTRANSFORMATION OF XENOBIOTICS All biochemical reactions involved in the conversion of foreign, toxic or water insoluble molecules to non-toxic, water soluble and excretable forms are referred to as DETOXIFICATION or BIOTRANSFORMATION reactions. These reactions  convert lipophilic to hydrophilic and  Facilitate excretion. Consequently, they  change the solubility characteristics of compounds  detoxify and  induce metabolic activation  DETOXIFICATION OR BIOTRANSFORMATION OF XENOBIOTICS The liver is the main organ involved in the biotransformation or detoxification reactions due to the fact that  Wide variety of enzymes involved in the detoxification processes of xenobiotics are contained in the hepatocytes. These enzymes are also present in the cytosol, endoplasmic reticulum and to lesser extent in other organelles. The overall purpose of these two phases of reactions is to  increase the water solubility (polarity) of xenobiotics and consequently  aid their excretion from the body  ENTOXIFICATION  This is a situation whereby the reactions of biotransformation of some xenobiotics conversely increase the toxicity of these compounds.  An example of this reaction is found in methanol toxicity.  ROLES OF BIOTRANSFORMATION REACTIONS Phase 1 reactions can  limit the toxicity of a drug  Convert xenobiotics from inactive to biologically active compounds (metabolic activation). The original xenobiotics are referred to as prodrug or procarcinogens. Phase 2 / conjugation reactions can  Convert the active product from phase 1 reactions to less active or inactive species which are consequently excreted in the urine or bile.  In some few cases, conjugation may also increase metabolic activation of xenobiotics PHASE 1 REACTIONS Phase 1 reactions comprise of  Oxidation  Reduction  Hydrolysis reactions Phase 1 reactions are also called hydroxylation reactions because they introduce or expose a functional group (e.g., -OH) that serves as the active center or point of conjugation for sequential conjugation in a phase II reaction. Methanol toxicity – Methanol possesses relatively low toxicity and it is metabolized in the liver. In the first step of degradation, methanol is transformed to formaldehyde by the enzyme alcohol dehydrogenase (ADH). Formaldehyde is transformed to Formic acid in a faster reaction requiring an enzyme aldehyde dehydrogenase. The metabolism of formic acid is very slow and this results in its accumulation in the body and this condition is referred to as metabolic acidosis. The major damage occurs to the optic nerve. Ethanol is given as an antidote since it is the true substrate of ADH and methanol is spared. PHASE 2 REACTIONS Phase 2 reactions are generally known as conjugation reactions. There are about 10 different phase ii reactions out of which 8 commonly occur in man. They are:  Glucuronidation reaction/glucuronide synthesis – UDP-Glucuronic acid (conjugating agent)  Sulphuric acid ester synthesis - Sulphate  Hippuric acid synthesis/ Glycine conjugation – Glycine  Glutamine conjugation – Glutamine  Glutathione conjugation – Glutathione  Acetylation – Acetyl group  Methylation – Methyl group  Thiocyanate synthesis – Thiol group  Ornithine conjugation (in birds)  Glucoside conjugation in insects – Glucoside a. GLUCURONIDATION REACTION/GLUCURONIDE SYNTHESIS  Compounds with free -SH, -COOH, -OH and -NH2 groups undergo glucuronidation after phase 1 reactions.  The conjugating agent is UDP- glucuronic acid synthesized from glucose.  The joining of the xenobiotic to the conjugating agent is achieved by the enzymes transferases found in the endoplasmic reticulum.  The products of glucuronidation reaction are usually in Beta conformation.  (Find the reaction pathway in your lecture note) b. SULPHATE CONJUGATION  This reaction is performed by Sulphotransferase, a cytosolic enzyme.  The reaction requires an endogenous substrate 3’- phosphoadenosine- 5’- phosphosulphates (PAPs).  Sulphate conjugation is limited by the availability of the endogenous substrate (PAPs) rather than the transferase enzyme  Two types of transferases exist for sulphate conjugation and these are: 1. Transferase for steroids 2. Transferase for other xenobiotics  The reaction starts with the synthesis of PAPs which conjugates with xenobiotic to yield resulting conjugated product.  (Find the reaction pathway in your lecture note). C. GLUTATHIONE CONJUGATION  Compounds with free -SH, -COOH, -OH and -NH2 groups undergo glutathione conjugation reaction after phase 1 reactions.  An important criterion for compounds to undergo this reaction is that the compound must be an electrophile.  The conjugating agent here is a tripeptide, glutathione (L-γ-glutamyl cysteinyl glycine).  Glutathione conjugation is catalysed by a family of transferase enzymes collectively known as ‘’Glutathione transferases’’.  (Find the reaction pathway in your lecture note). Cytochrome P450 system is the most important of all enzymes involved in biotransformation of drugs. Cytochrome P450 is a super-family of membrane bound enzyme systems which are present in the smooth endoplasmic reticulum of the cell and therefore are located in the microsomal fraction following homogenization and centrifugation. Apart from cytochromes P450, Flavin-containing Monooxygenases (FMOs) is another important microsomal enzyme. These two enzymes are involved in the process of metabolism of xenobiotics and their main function is to add molecular oxygen to lipophilic compounds, making them soluble to ensure rapid excretion.  FACTORS AFFECTING BIOTRANSFORMATION OF XENOBIOTICS/DRUGS  Prior administration of the drug or co-administration with other drugs  Diet  Hormonal status  Genetics  Diseases  Age and developmental status  Functional status of liver and kidney Reference 1. BIOTRANSFORMATION/DETOXIFICATION REACTIONS METABOLISM OF XENOBIOTICS Biochemistry for Medics The biochemical modes of action of pesticides and herbicides DEFINITIONS OF PESTICIDES  A pesticide is a substance or mixture of substances intended for preventing, destroying, repelling, or lessening the damage of any pest. The pest can be insects, plant pathogens, weeds, mollusks, birds, mammals, fish, nematodes (roundworms) and microbes that compete with humans for food, destroy property, spread or help carry or spread diseases or are seen as a nuisance.  Pesticides are substance or mixture of substance which differ in their physical, chemical and identical properties from one to other.  The most common used pesticides include insecticides, herbicides, fungicides and rodenticides.  The other less well-known pesticides comprise growth regulators, plant defoliants, surface disinfectants and some swimming pool chemicals.  Most commonly, pesticides are used in health sector and agricultural crops.  They are useful in public health for killing vectors of the disease, such as mosquitoes while, pests damaging agricultural crops are killed by pesticides.  Naturally, pesticides are potentially toxic to other non-target organisms, including humans. Hence, it is necessary to use them safely and dispose properly. CLASSIFICATIONS  Hence, they are classified based on these physical, chemical and identical properties. Some pesticides are also categorized into various classes depending on the needs.  Presently, three most popular classifications of pesticides which are widely used is. (i) classification based on the mode of entry, (ii) classification based on pesticide function and the pest organism they kill and (iii) classification based on the chemical composition of the pesticide  Based on toxicity of pesticides, WHO classified them into four classes: extremely dangerous, highly dangerous, moderately dangerous and slightly dangerous.  Improper application of pesticides can cause severe harmful effect to living system and the environment.  Most pesticides do not distinguish between pests and other similar incidental lifeform and kill them all.  The toxicity of insecticides to an organism is usually expressed in terms of the LD50 (lethal dose 50 percent) and LC50 (50 percent lethal concentration).  I. CLASSIFICATION BASED ON MODE OF ENTRY The ways pesticides come in contact with or enter the target are called modes of entry. These include systemic, contact, stomach poisons, fumigants, and repellents. 1. Systemic pesticides: Systemic pesticides are pesticides which are absorbed by plants or animals and transfer to untreated tissues. Systemic herbicide moves through the plant and can reach to untreated areas of leaves, stems or roots. They are capable in killing of weeds with partial spray coverage. They can effectively penetrate in the plant tissues and move through plant vascular system to kill specific pests. Some systemic insecticides are also applied and move through animals to control pests such as warble grubs, lice, or fleas. The movement of pesticides in plant tissues may be unidirectional or multidirectional. Some pesticides may only move in one direction either up or down within the plant while other pesticides may only move upwards in plants. If applied to the root zone, it will travel throughout the plant, but if applied to the leaves it will not move throughout the plant. Furthermore, few pesticides are considered locally systemic and move only to a short distance in a plant from the point of contact. Examples of systemic pesticides include 2, 4-Dichlorophenoxyacetic acid (2, 4-D) and glyphosate. 2.Non-systemic (Contact) pesticides: The non-systemic pesticides are also called contact pesticides as it acts on target pests when they come in contact. Pesticides must come into physical contact with the pest to be effective. The pesticide enters the body of pests via their epidermis upon contact and causes death by poisoning. These pesticides do not necessarily penetrate the plant tissues and consequently not transported through the plant vascular system. Examples of contact pesticides are paraquat and diquat dibromide. 3. Stomach poisoning and stomach toxicants: Stomach poisoning pesticide enters the pest’s body through their mouth and digestive system and causes death by poisoning. Stomach poisons are acquired during feeding of pests, when they ingest the insecticide applied in the leaves and other parts of the plant. Stomach toxicants may also enter the body of insects through the mouth and digestive tract, where they are absorbed into the insect’s body. This is more appropriate especially in vector control including bacteria, or their toxins, applied to the water where filter-feeding mosquito or black fly larvae will consume the poison. These insecticides kill the vector by destroying the midgut (or stomach) of the larvae. Example: Malathion. 4. Fumigants: Fumigants are such pesticides which acts or may kill the target pests by producing vapor. These pesticides form poisonous gases when applied. These pesticides in vapor form enter the body of pests via their tracheal system (respiratory) through spiracles and causes death by poisoning. Some of their active ingredients are liquids when packaged under high pressure but change to gases when they are released. Other active ingredients are volatile liquids when enclosed in an ordinary container and are not formulated under pressure. Fumigants are used to remove stored product pests from fruits, vegetables and grains. They are also very useful in controlling of pests in soil. 5. Repellents: Repellents do not kill but are distasteful enough to keep pests away from treated areas/commodities. They also interfere with pest’s ability to locate crop II. CLASSIFICATION BASED ON PESTICIDE FUNCTION AND PEST ORGANISM THEY KILL Under this method, pesticides are classified based on target pest’s organism and pesticides are given specific names to reflect their activity. The group names of these pesticides arise from the Latin word cide (meaning kill or killer) that are used as suffix after corresponding name of pests they kill (Table 1). Not necessarily, all pesticides end with word-cide. Some pesticides are also classified according to their function. For examples: growth regulators, which stimulate or retard the growth of pests; defoliants, which cause plants to drop their leaves; desiccants, which speed the drying of plants for mechanical harvest or cause insects to dry out and die; repellents which repel pests; attractants, which attract pests, usually to a trap; and chemosterilants, which sterilize pests.  Also, there are pesticides that control more than one class of pests and may be considered in more than one pesticide class. Aldicarb, which is widely used in Florida citrus production, may be considered an acaricide, insecticide, or nematicide because it controls mites, insects, and nematodes, respectively. Another common example is 2, 4-D, which is used as an herbicide for broadleaf weed control, but it is a plant growth regulator at low rates.  Attractants and repellents are considered pesticides because of their use in pest control. III. CLASSIFICATION BASED ON CHEMICAL COMPOSITION OF PESTICIDES The most common and useful method of classifying pesticide is based on their chemical composition and nature of active ingredients. It is such kind of classification that gives the clue about the efficacy, physical and chemical properties of the respective pesticides. The information on chemical and physical characteristics of pesticides is very useful in determining the mode of application, precautions that need to be taken during application and the application rates. Based on chemical composition, pesticides are classified into four main groups namely; organochlorines, organophosphorus, carbamates and pyrethrin and pyrethroids. The chemical-based classification of pesticides is rather complex. In general, modern pesticides are organic chemicals. They include pesticides of both synthetic and plant origin. However, some inorganic compound is also used as pesticides. Insecticides are important pesticides that can be further classified into several sub-classes. The sub-classification of insecticides is given in Fig. 1. OTHER MINOR CLASSES OF PESTICIDES IV. CLASSIFICATION BASED ON MODE OF ACTION Based on mode of action, pesticides are classified as: 1. Physical poison: These classes of pesticides bring about killing of one insect by exerting a physical effect. For example: Activated clay 2. Protoplasmic poison: These pesticides are responsible for precipitation of protein. Example of this pesticide is Arsenicals 3. Respiratory poison: Respiratory poisons are chemicals which inactivate respiratory enzymes. Example: Hydrogen cyanide 4. Nerve poison: Chemicals inhibit impulse conduction. Example: Malathion 5. Chitin inhibition These classes of chemicals inhibit the chitin synthesis in pests. Example: Diflubenzuron  TOXICOLOGICAL STUDIES OF CERTAIN FOOD ADDITIVES (TERM PAPER) Several studies have reported toxicology of different food additives. Identify any four studies of your choice and briefly describe them for your records. You may also want to explore the reference given on food additives. REFERENCES 1. Evaluation of certain food additives: World Health Organization technical report series 617; the twenty-first report of the joint FAO/WHO Expert Committee on Food additives (1978). 2. Katerina Mastovska (2013); Modern Analysis of Chemical Contaminants in Food; Food safety magazine. 3. Alekseyev, Y., Hamm, M., & Essigmann, J. (2004). Aflatoxin B1 formamidopyrimidine adducts is preferentially repaired by the nucleotide excision repair pathway in vivo. Carcinogenesis, 25 6, 1045-51