Biotransformation and elimination

metabolism and excretion

1. Biotransformation and elimination By M aheshwari Hireholi Mpharm 1 st year

2. OVERVIEW Definition Consequences Types Phase I and II reactions Enzyme induction/inhibition First pass metabolism Excretion Types Factors affecting excretion Kinetics of elimination

3. Definition Chemical alteration of drug in the body . Non polar lipid soluble compounds are made polar lipid insoluble, so that they are easily excreted. Biotransformation is needed for detoxification & protect the body from ingested toxins.

4. SITES Primary site – Liver Others – Kidney, Intestine, Lungs, Plasma…

5. Biotransformation normally leads in pharmacological inactivation of drugs Ex: conversion of phenytoin to p- hydroxy phenytoin, occasionally yields metabolites with equal activity ex: conversion of phenylbutazone to oxyphenbutazone

6. Rarely leads to toxicological activation of drugs Ex : conversion of paracetamol to reactive metabolites that cause hepatic necrosis. Activation of inactive drug Ex : D igitoxin to Digoxin, Primidone to Phenobarbitone.

7. Absorbed drugs – undergo mainly three changes Metabolic changes by Enzymes ( Microsomal, Cytoplasmic, Mitochondrial) Spontaneous Molecular rearrangement – HOFMANN ELIMINATION ( This refers to inactivation of the drug in the body fluids by spontaneous molecular rearrangement without the agency of any enzyme, e.g. Atracurium ) Excreted unchanged (highly polar drugs) - Aminoglycosides , Methotrexate,Neostigmin

8. CONSEQUENCES A ) Drug inactivation - inactive or less active Propranolol, Pentobarbitone , Chloramphenicol, Paracetamol, Ibuprofen, lignocaine B) Active drug to Active metabolite- active metabolite Ex: P henacetin to Paracetamol

9. C) Inactive drug (Prodrug) - Active drug Prodrugs are inactive drugs which need BT in the body to form active metabolites. ADVANTAGE: More stable Better BA Less toxicity

10. Active drug Active metabolite Chloral hydrate Trichloroethanol Morphine Morphine-6-glucuronide Cefotaxime Desacetyl cefotaxime Allopurinol Alloxanthine Procainamide N-acetyl procainamide Primidone Phenobarbitone Diazepam Desmethyl -diazepam, oxazepam Imipramine Desipramine Amitriptyline Nortriptyline Codeine Morphine Spironolactone Canrenone

11. TYPES Phase I / Non synthetic / Functionalization A functional group is generated Catabolic reaction Metabolite – active or inactive Phase II / Synthetic / Conjugation An endogenous radical is conjugated Anabolic Metabolite is usually inactive

12. Phase I Reactions Oxidation Reduction Hydrolysis Cyclization Decyclization

13. Phase II Reactions Glucuronide conjugation Acetylation Methylation Sulfate conjugation Glycine conjugation Glutathione conjugation Ribonucleotide / Ribonucleoside synthesis

14. a) OXIDATION Addition of Oxygen / negatively charged radical or removal of Hydrogen / Positively charged radical Oxidation is the main process of metabolism Produces unstable intermediates - Epoxides, Superoxides, Quinones Oxidation – 9 types

15. 1.OXIDATION AT NITROGEN ATOM RNH2 o RNHOH Chlorpheniramine Dapsone Meperidine

16. 2.OXIDATION AT SULPHUR ATOM Chlorpromazine Chloramphenicol

17. 3.ALIPHATIC HYDROXYLATION Hydroxyl group added to drug RCH2CH3 O RCHOHCH3 Salicylic acid to Gentisic acid Ibuprofen Tolbutamide, Chlorpropamide,

18. 4.AROMATIC HYDROXYLATION Phenytoin Phenobarbitone Propranolol

19. 5.DEALKYLATON AT OXYGEN ATOM ROCH3 O ROH + CH2O Phenacetin to Paracetamol

20. 6.DEALKYLATON AT NITROGEN ATOM RNHCH3 O RNH2 + CH2O Amitriptyline to Nortriptyline

21. 7.DEALKYLATON AT SULPHUR ATOM 6-Methyl thiopurine to Mercaptopurine

22. 8.OXIDATIVE DEAMINATION Amphetamine

23. 9.DESULFURATION Parathion to Paraoxon The main enzymes involved in phase I reaction are ooxygenases.

24. b) REDUCTION Addition of Hydrogen / positively charged radical or removal of Oxygen / negatively charged radical MICROSOMAL REDUCTION by Monooxygenases need NADPH & cytochrome c reductase. A.NITRO Reduction RNo2 RNH2 Chloramphenicol to aryl amine metabolite B.KETO Reduction Cortisone to Hydrocortisone

25. c. Azo reduction Prontosil to Sulfanilamide NON MICROSOMAL REDUCTION Chloral hydrate to Trichloro ethanol,

26. c) HYDROLYSIS Drug is split combining with water Ester + water Esterases Alcohol & Acid Microsomal hydrolysis Pethidine to meperidinic acid Non microsomal hydrolysis Esterases,Amidases & Peptidases Atropine to Tropic acid

27. d) CYCLIZATION Formation of ring structure from a straight chain compound. Eg : Proguanil

28. e) DE CYCLIZATION Ring structure is opened Phenytoin, Barbiturates

29. PHASE II REACTIONS /Synthetic reactions Involve transfer of a suitable moiety such as glucuronic acid, sulfate, glycine , etc, in presence of enzyme transferases to drugs or metabolites of phase I reaction having suitable functional groups to form highly polar n readily excretable conjuagtes . Phase II- need energy

30. 1.CONJUGATION WITH GLUCURONIC ACID Carried out by UDP glucuronyl transferases Compounds with OH & COOH are easily conjugated with glucuronic acid derived from glucose Drug + UDPGA Microsomal Glucuronyl transferase Drug glucuronide + UDP Drugs - Aspirin,Paracetamol,PABA, Metronidazole, Morphine, Diazepam

31. ↑Mol. Weight – favours biliary excretion Drug glucuronides excreted in bile are hydrolyzed by intestinal microfloral enzymes - parent drug released - reabsorbed into systemic circulation- ↓excretion ↑duration of action - Oral contraceptives, Phenolphthalein Endogenous substrates - Steroid, Thyroxin, Bilirubin

32. 2. ACETYLATION Drugs with Amino or Hydrazine groups - INH, PAS,Hydralazine, Sulfonamides ,Procainamide, Dapsone. Genetic polymorphism Acetylation- Rapid / Slow

33. 3. CONJUGATION WITH SULFATE Drug groups-Amino, Hydroxyl Cytoplasmic Enzymes - Sulfotransferases / Sulfokinases . Methyl dopa, Steroids, Chloramphenicol, Warfarin The phenolic compounds and steroids are sulfated by sulfotransferases (SULTs).

34. 4. CONJUGATION WITH GLYCINE Drugs having Carboxylic group Salicylic acid , Benzoic acid

35. 5. Conjugation with glutathione Drug groups- Epoxide , Quinone Toxic metabolites of Paracetamol, Ethacrynic acid Cytoplasmic Enzyme - Glutathione S- Transferase

36. 6. METHYLATION Drugs with Amino & Phenol groups Histamine, Adrenaline, Nicotinic acid, Dopamine, Methyl dopa, Captopril Enzyme- Methyl transferase Endogenous substance- Cysteine , Methionine

37. 7. RIBONUCLEOTIDE/RIBONUCLEOSIDE SYNTHESIS Action of Purine & Pyrimidine antimetabolites 6 Mercaptopurine

38. Enzymes involved Microsomal enzymes located on SER, primarily in liver, also in kidney, intestinal mucosa and lungs. Ex : monooxygenases, cytchrome P450, glucuronyl transferase etc,. Catalyse most of the oxidation, reduction, hydrolysis and glucuronide conjugation. Induced by drugs, diet and agencies.

39. Non- microsomal enzyme Present in cytoplasm and mitochondria of hepatic cells, as well as in other tissues including plasma. Ex : flavoprotein oxidases, esterases, amidases and conjugases. Reaction catalysed are oxidation, reduction, many hydroytic and all conjugation except glucuronidation . These are not inducible but many show genetic polyphorism ( acetyl transferases)

40. Microsomal enzyme induction Drugs, insecticides, carcinogens will induce the synthesis of microsomal enzyme proteins. Accelerated metabolism and reduced pharmacological response.

41. Consequences 1. Decreased intensity and/or duration of action of drugs that are inactivated by metabolism, e.g. failure of contraception with oral contraceptives. 2. Increased intensity of action of drugs that are activated by metabolism. Acute paracetamol toxicity is due to one of its metabolites—toxicity occurs at lower doses in patients receiving enzyme inducers. 3. Tolerance—if the drug induces its own metabolism (auto induction), e.g. carbamazepine , rifampin . 4. Some endogenous substrates (steroids, bilirubin ) are also metabolized faster. 5. Precipitation of acute intermittent porphyria : enzyme induction increases porphyrin synthesis. 6. Intermittent use of an inducer may interfere with adjustment of dose of another drug prescribed on regular basis, e.g. oral anticoagulants, oral hypoglycaemics , antiepileptics , antihypertensives . 7. Interference with chronic toxicity testing in animals

42. FIRST PASS (PRESYSTEMIC) METABOLISM This refers to metabolism of a drug during its passage from the site of absorption into the systemic circulation. ↓ed BA ↓ed therapeutic response SITES * Gut wall *Gut lumen *Liver (major site) * Lungs *Skin

43. Excretion It is the process where by drug or their metabolite are irreversibly transferred from internal to external environment through renal or non renal route. The principle organ of excretion are kidneys.

44. Types of excretion RENAL EXCRETION NON RENAL EXCRETION Biliary excretion Pulmonary excretion Salivary excretion Mammary excretion Skin/ dermal excretion Gastrointestinal excretion

45. RENAL EXCRETION The kidney is responsible for excreting all water soluble substances. Net renal excretion =(Glomerular filtration + tubular secretion) – tubular reabsorption

46. GLOMERULAR FILTRATION It Is non selective , unidirectional process Ionized or unionized drugs are filtered, except those that are bound to plasma proteins. Driving force for GF is hydrostatic pressure of blood flowing in capillaries. Glomerular filtration rate ( g.f.r .), normally ~ 120 ml/min, declines progressively after the age of 50, and is low in renal failure.

47. ACTIVE TUBULAR SECRETION This mainly occurs in proximal tubule. It is carrier mediated process which requires energy for transportation of compounds against conc. gradient Two secretion mechanisms are identified. System for secretion of organic acids/anions E.g. Penicillin, salicylates etc System for organic base / cations E.g. morphine, mecamylamine hexamethonium Active secretion is Unaffected by change in pH and protein binding. Drug undergoes active secretion have excretion rate values greater than normal GFR e.g. Penicillin.

48. TUBULAR REABSORPTION It occurs after the glomerular filtration of drugs. It takes place all along the renal tubules. Reabsorption of drugs indicated when the excretion rate value are less than the GFR 130ml/min. e.g. Glucose TR can be active or passive processes. Reabsorption results in increase in the half life of the drug.

49. Active Tubular Reabsorption Its commonly seen with endogenous substances or nutrients that the body needs to conserve e.g. electrolytes, glucose, vitamins. Passive Tubular Reabsorption It is common for many exogenous substances including drugs. The driving force is Conc. Gradient which is due to re-absorption of water, sodium and inorganic ions. I

50. FACTORS AFFECTING RENAL EXCRETION Physicochemical properties of drug Urine pH Blood flow to the kidney Biological factor Drug interaction Disease state

51. KINETICS OF ELIMINATION There are three fundamental pharmacokinetic parameters, viz. bioavailability (F), volume of distribution (V) and clearance (CL) which must be understood. The first two have already been considered.

52. Clearance Is defined as the hypothetical volume of body fluids containing drug from which the drug is removed/ cleared completely in a specific period of time. Expressed in ml/min. CL = Rate of elimination/C

53. TOTAL BODY CLEARANCE Is defined as the sum of individual clearances by all eliminating organs is called total body clearance/ total systemic clearance Total body clearance= CL liver + CL kidney + CL lungs +CL x

54. Plasma half-life The Plasma half-life (t½) of a drug is the time taken for its plasma concentration to be reduced by half of its original value. Mathematically, elimination t½ is t½ = ln2 /t½ Where ln2 is the natural logarithm of 2 (or 0.693) and k is the elimination rate constant of the drug, i.e. the fraction of the total amount of drug in the body which is removed per unit time.

55. Half life of some representative drugs Aspirin 4 hr Digoxin 40 hr Penicillin-G 30 min Digitoxin 7 days Doxycycline 20 hr Phenobarbitone 90 hr

56. Repeated drug administration When a drug is repeated at relatively short intervals, it accumulates in the body until elimination balances input and a steady state plasma concentration ( Cpss ) is attained, Cpss =dose rate/CL After oral administration, often only a fraction (F) of the dose reaches systemic circulation in the active form. In such a case— dose rate=target Cpss × CL/F

57. Loading dose The loading dose is one or a series of doses that may be given at the onset of therapy with the aim of achieving the target concentration rapidly. The appropriate magnitude for the loading dose is Loading dose=target Cp × V L/ F loading dose is governed only by V and not by CL or t½.

58. Maintenance dose This dose is one that is to be repeated at specified intervals after the attainment of target Cpss so as to maintain the same by balancing elimination. The maintenance dose rate is computed by equation dose rate=target Cpss × CL/F and is governed by CL (or t½) of the drug .

59. Reference Goodman and Gilman’s.(2011) ‘The Pharmacological B asis of Therapeutics’.26-30. Tripati.K.D .(2013) ‘ Essentials of medical Pharmacology’ , 22-36. Brahmankar.D.M , Jaiswal.B . Sunil.(2008) ‘ Biopharmaceutics and Pharmacokinetics’ ,111-157.