Drug metabolism (Biotransformation) for GPAT preparation

• The onset of pharmacological response depends upon 2 pharmacokinetic processes: Absorption & Distribution.
• The duration & intensity of action depends upon the rate of drug removal from the body or rate of elimination & tissue redistribution of the drug.
• Biotransformation (metabolism) of drug is defined as the conversion from one chemical form to another.
• The chemical changes are usually affected enzymatically in the body & thus this definition excludes chemical instability. for example:

• Conversion of penicillin to penicilloic acid (Bacterial/mammalian enzymes) is metabolism.
• Conversion of penicillin to penicilleinic acid by stomach acid is chemical instability. 

Xenobiotics:  All chemical substances that are not nutrients for the body & enter the body through ingestion, inhalation or absorption are called xenobiotics.

Biotransformation often results in:

a) Pharmacological inactivation of the drug:

Inactivation of drug with biotransformation

b) No change in pharmacological activity:

No change in activity of the drugs after metabolism

c) Toxicological activation:

Toxic metabolites of metabolism

d) Pharmacological activation:

Activation of drugs after metabolism

e) Change in pharmacological activity:
i) Iproniazid (antidepressant) -------> Isoniazid (Anti-Tb)
ii) Diazepam (Tranquilizer) --------> Oxazepam (Anticonvulsant).

Drug metabolizing organs: Liver is the major site for the drug metabolism. Other sites for the drug metabolism in the decreasing order includes: Lungs > Kidney > Intestine > Placenta > Skin.

Drug metabolizing enzymes: 

a) Microsomal enzymes: Catalyzes a majority of drug biotransformation reactions. They are located in the microsomes of rough endoplasmic reticulum (RER). Some important characteristics of these enzymes include:

• Intact nature of lipoidal membrane bound enzymes of the microsomes is essential for the activity. 
• No. of lipid soluble substances can interact wih these enzymes.
• Lipid soluble substance is biotransformed to water soluble substances.

Chemical Pathways of drug biotransformation

RT Williams divided the pathways of drug metabolism reactions into two general categories:
a) Phase I reactions: (Functionalization/ Asynthetic reactions):

• It includes oxidative, reductive & hydrolytic reactions.
• A polar function group is either introduced or unmasked if already present on the otherwise lipid soluble substrate (OH, COOH, NH2 & SH).
• The resulting product of phase I reaction is susceptible to phase II reactions.

b) Phase II reactions: These reactions generally involve covalent attachment of small polar endogenous such as glucouronic acid, sulfate, glycine etc. to either unchanged drugs or phase I products having suitable functional groups (OH, COOH, NH2 & SH) & form highly water soluble conjugates which are readily excreted by the kidneys (or bile).
Also known as conjugation or true detoxification reactions.

Various reactions occurring in metabolism:
a) Phase I reactions:
1) Oxidative reactions:

• Oxidation of aromatic carbon atom.
• Oxidation of olefins (C=C).
• Oxidation of benzylic, allylic carbon atoms & carbon atoms alpha to carbonyl & imines.
• Oxidation of aliphatic carbon atoms.
• Oxidation of alicyclic carbon atoms.
• Oxidation of carbon-heteroatom

• Carbon Nitrogen system

• N-dealkylation.
• Oxidative deamination
• N-Oxide formation
• N-hydroxylation

• Carbon-sulphur system

• S-dealkylation
• Desulfuration
• S-oxidation 

• Oxidation of OH, C=O & COOH groups.
• Miscellaneous oxidative reactions 

2) Reductive reactions: 

• Reduction of carbonyl functions (Aldehydes/ketones).
• Reduction of alcohols & C=C bonds.
• Reduction of N compounds (nitro, azo & N-oxide)
• Miscellaneous reductive reactions.

3) Hydrolytic reactions: 

• Hydrolysis of esters & amides.
• Hydrolysis of amides.
• Hydrolytic cleavage of nonaromatic heterocyclics.
• Hydrolytic dehalogenation.
• Miscellaneous hydrolytic reactions.

b) Phase II reactions:

1. Conjugation with glucouronic acid.
2. Conjugation with sulfate moieties.
3. Conjugation with alpha-amino acids.
4. Conjugation with glutathione & mercaptopuric acid formation.
5. Accetylation reaction
6. Methylation reaction
7. Miscellaneous reaction

Phase I reactions of drug metabolism

1) Oxidation reactions in biotransformation of drugs:

• Oxidative reactions are the most important and most common metabolic reactions.
• Almost all drugs in Phase I biotransformation undergo oxidation at some stage or the other.
• Oxidative reactions increase hydrophilicity of xenobiotiocs by introducing polar functinal groups (OH).
• Oxidation of xenobiotics is non sppecifically catalyzed by a number of enzymes located in the microsomes. Such enzymes require both molecular oxygen (O2) & the reducing agent NADPH to effect reactions. They are therefore referred as mixed function oxidases. The overall stoichometry of the reaction is:

                              RH + O2 + NADPH + H+ -----------> ROH + H2O + NADP+

• Since only one oxygen atom from the molecular oxygen is incorporated in the product formed hence mixed function oxidases are also known as mono-oxygenases.
• The multienzyme function oxidase system located in the endoplasmic reticulum of hepatic cells is composed of an electron transfer system chain consisting of three components:

• A heme protein known as Cyt. P450, which is actually a family of enzymes.
• A second enzyme, the flavoprotein known as Cyt. P450 reductase which is NADPH dependent and function as an electron carrier, catalyzing  the reduction of cyt. P450 to the ferrous form.
• Heat stable lipid component known as phosphatidyl  choline (Facilitate  electron transfer from NADPH).

                   Mg2+ ions are required for maximum action 

Different types of oxidation reaction reactions occurring in Phase I biotransformation include:

• Oxidation of aromatic carbon atoms:

Oxidation of aromatic carbon atoms

• Oxidation of olefins:

Oxidation of olefins

• Oxidation of benzylic carbon atoms:

Oxidation of benzylic carbon atoms:

• Oxidation of Diazepam:

Oxidation Oo Diazepam

• Oxidation of Aliphatic carbon atoms

Oxidation of Valproic Acid

• Oxidation of Alicyclic Carbon Atoms 

Oxidation of Minoxidil 

• Oxidation of Carbon-Heteroatom system:

       In most of the drugs, we usually see C-O, C-S, and C-N heteroatoms. Their oxidation follows either the hydroxylation of the Carbon atom attached to the heteroatom or through the oxidation of the heteroatom itself. Here are a few examples of the biotransformation of the Carbon-heteroatom system:

Metabolism of Imipramine 

Metabolism of Trimethoprim

I1) Reductive reactions in biotransformation of drugs:

• Reductive reactions also play an important role in generating polar groups in the drugs which can further undergo Phase II reactions and further elimination of the drug from the body. 
• Reduction of  Carbonyl and Nitrogen-containing drugs undergo bioreduction reaction in Phase I of the drug metabolism.
• Following are the examples of bioreduction reactions of some important drugs:

BIOREDUCTION IN METABOLISM OF DRUGS

1II) Hydrolytic reactions in the biotransformation of drugs:

• Hydrolytic reactions involve the cleavage of larger molecules into smaller ones.
• These hydrolytic biotransformation phase I reactions are not confined to the liver only instead, it occurs in all major organs such as the intestine, lungs, kidney, etc.
• All the endogenous substances are also metabolized by these hydrolytic enzymes.
• Examples of hydrolytic phase I metabolism reactions are:

HYDROLYTIC PHASE I METABOLISM REACTIONS

Phase II reactions of Drug Metabolism

• Phase II reactions of the drug metabolism are considered as a true detoxification pathway of the biotransformation as large molecular moieties combine with the drug or its metabolite to make it more polar & non-toxic.
• High molecular weight conjugates ( More than 350 Da) are excreted through bile whereas low molecular weight conjugates excrete through urine ( less than 250 da).
• Glutathione conjugates always get excreted through bile.
• Phase II metabolism reactions are always capacity limited. 
• The table below summarises the Phase II metabolism reactions:

Name of Conjugation reaction	Conjugating agent	Enzyme	Intermediate used
Glucouronidation	Glucouronic acid	UDP-Glucouronyl transferase	UDPGA
Sulphation	Sulphate	Sulphotransferase	PAPS
Amino acid Conjugation	Glycine	Avyl transferase	Acyl CoA
Glutathione	Glutathione	Glutathione S transferase	----
Acetylation	Acetyl-CoA	N-acetyl Transferase	Acetyl-CoA
Methylation	L-Methionine	Methyl Transferase	S-adenosyl methionine