INSTITUTE OF PHARMACY

BUNDELKHAND UNIVERSITY, JHANSI (U.P.)

SESSION: 2023-2024

AN ASSIGNMENT
On
“REACTIONS INVOLVED IN THE BIOTRANSFORMATION OF
DRUGS”

Submitted to: Submitted by:

Dr. Nirmala Devi Prajapati Adarsh Yadav
Assistant Professor- B.Pharm 4th Sem
Institute of Pharmacy Roll No.: 221251016005
B.U. Jhansi

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 Table of Contents

Sr. No. Particulars

1 Introduction

2 Importance of biotransformation

3 Fundamentals

4 Mechanism

5 Drug metabolism

6 Factors affecting Drug Metabolism

7 Clinical Significance

8 Pharmacology & Drug Therapy

9 Overview

10 References

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 Table of Figures

Sr. No. Name of the figure

1 Phases of chemical process

2 Importance of biotransformation

3 Mechanism

4 Reaction of Mechanism

5 Metabolism of drugs

6 Process of metabolism

7 Factor affecting metabolism

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 INTRODUCTION
Drug biotransformation reactions are essential chemical processes, mainly mediated by
enzymes, which lead to the formation of drug metabolites that are excreted from the body.
Historically, this metabolism was known as a detoxication mechanism. However, these
reactions which govern the pharmacological efficacy of the drugs, may in many cases, also be
responsible for toxic unwanted effects. Due to the large variety of chemical structures, drug
biotransformation involves numerous and sophisticated reaction mechanisms and metabolic
pathways that are catalysed by a large number of enzyme families. The aim of this chapter is
to give medicinal chemists the essential information to understand, from a chemical point of
view, the fate of drugs in human. This information is a prerequisite for predicting drug
metabolism and reactivity, and therefore for designing safer and more efficient drugs. Besides
drugs, xenobiotics present in our environment (pollutants, pesticides, food additives, etc.) also
undergo similar biotransformation reactions.

Drugs are generally lipophilic substances. This property allows them to penetrate easily into
the different cell compartments through the hydrophobic membrane barriers. In that context,
hydrophilic substances (saccharine, carbohydrate-derived substances) that cannot enter are
generally not subject to metabolic transformations. The general principle of drug metabolism
is to convert the lipophilic drugs via several chemical steps, into hydrophilic, water-soluble
derivatives which can easily be eliminated into urine. However, very highly lipophilic drugs,
polyhalogenated xenobiotics, such as some insecticides that can concentrate in the membrane
compartment are sterically shielded from metabolic attack.

(Figure 1: Phases of chemical process)

The metabolism of xenobiotics involves two sequential steps known as phase I and phase II
reactions. During phase I, a functionalization reaction of the xenobiotic is achieved. New polar
groups such as CO2H, OH or NH2 are introduced or unveiled from pre-existing functions
through oxidative, reductive or hydrolytic reactions. The polar group created serves then as an
anchor point for the second metabolic step. The phase II reactions, known as conjugation
reactions, link an endogenous, generally hydrophilic moiety, either to the original drug (if polar
functions are already present) or to the phase I metabolite. Common endogenous groups are
glucuronic acid, various amino acids, the tripeptide glutathione, or sulfate.

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 Importance of Biotransformation
Biotransformation is vital to survival because it transforms absorbed nutrients (food, oxygen,
etc.) into substances required for normal body functions. For some pharmaceuticals, it is a
metabolite that is therapeutic and not the absorbed drug. For example, phenoxybenzamine, a
drug given to relieve hypertension caused by pheochromocytoma, a kind of tumor, is
biotransformed into a metabolite, which is the active agent. Biotransformation also serves as
an important defense mechanism since toxic xenobiotics and body wastes are converted into
less harmful substances and substances that can be excreted from the body.

(Figure 2: Importance of biotransformation)

Toxicants that are lipophilic, non-polar, and of low molecular weight are readily absorbed
through the cell membranes of the skin, GI tract, and lung. These same chemical and physical
properties control the distribution of a chemical throughout the body and its penetration into
tissue cells. Lipophilic toxicants are hard for the body to eliminate and can accumulate to
hazardous levels. However, most lipophilic toxicants can be transformed into hydrophilic
metabolites that are less likely to pass through membranes of critical cells. Hydrophilic
chemicals are easier for the body to eliminate than lipophilic substances. Biotransformation is
thus a key body defense mechanism.

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 Fundamentals
The pathways of biotransformation are divided into phase I, phase II, and phase III. These
reactions may occur simultaneously or sequentially.

Phase I: Yields a polar, water-soluble, metabolite that is often still active. Many of the products
in this phase can also become substrates for phase II.

• Oxidation with cytochrome P450 (most common)

• Reduction
• Hydrolysis

Phase II: Yields a large polar metabolite by adding endogenous hydrophilic groups to form
water-soluble inactive compounds that can be excreted by the body.

• Methylation
• Glucuronidation (most common)
• Acetylation
• Sulfation
• Conjugation with glutathione
• Conjugation with amino acids (glycine, taurine, and glutamic acid)

Phase III: Occurs post-phase II, where a chemical substance can undergo further metabolism
and excretion. It classifies into the following superfamilies:

• ATP-binding cassette (ABC)

• Solute carrier (SLC) transporters

Cellular Level
The majority of biotransformation takes place within the liver in cells called hepatocytes.
However, several of the enzymes for phase I, phase II, and phase III reactions can also occur
in extrahepatic tissues, such as adipose, intestine, kidney, lung, and skin. The transformation
process takes place as a result of the interaction between the substrate and enzymes found
primarily in these cells' cytoplasm, endoplasmic reticulum, and mitochondria.

Molecular Level
Phase I reactions are mainly catalyzed by the cytochrome P450 system, which is a family of
membrane-bound enzymes found within the endoplasmic reticulum of hepatocytes. The unique
protein structures that make up the P450 enzymes will exhibit significant differences in their
substrate and product selectivity. Both constitutive and inducible regulations control these
enzymes. For example, there are mechanisms of transcriptional gene activation, such as those
that are stimulated by drugs or other exogenous substances.

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Similar gene sequences classify cytochrome P450 pathways; they are assigned a family number
and a subfamily letter and then become differentiated by a number for the isoform or individual
enzyme. CYP subtypes include CYP1A1/2, CYP1B1, CYP2A6, CYP2B6, CYP2C8, CYP2C9,
CYP2C19, CYP2D6, CYP2E1, CYP3A4/5/7. The most common pathways involve
CYP3A4/5, CYP2C9, CYP2D6, and CYP2C19, where CYP3A4 is the most abundant
isoenzyme in the human liver and accounts for about 50% of all CYP450 activity.

Mechanism :
Reactions classify as Phase I, Phase II, or Phase III. It is important to note that these reactions
do not have to take place sequentially and can even take place in reverse, Phase II, and then
Phase I, or as a single reaction. Phase I metabolism consists of reduction, oxidation, or
hydrolysis reactions. These reactions serve to convert lipophilic drugs into more polar
molecules by adding or exposing a polar functional group such as -NH2 or -OH. These
reactions also often create active metabolites and which is beneficial in activating prodrugs into
their active and therapeutic state. Phase I includes the use of the cytochrome P450 system found
with the membrane of the endoplasmic reticulum. If the CYP450 system becomes inhibited in
any way the drug level will increase, and vice versa if induced, the drug level will decrease.

(Figure 3: Mechanism)

Phase I reaction using the CYP450 system includes both an oxidative and reductive step using
NADPH and not ATP: Drug + O2+ NADPH -> Drug*+ H20 + NADP+.

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Drugs that have already have -OH, -NH2 or COOH groups can bypass Phase I and enter Phase
II directly to become conjugated. Phase II reactions consist of adding hydrophilic groups to the
original molecule, a toxic intermediate or a nontoxic metabolite formed in phase I, that requires
further transformation to increase its polarity. These reactions include conjugation reactions,
glucuronidation, acetylation, and sulfation. The ultimate goal of phase II reactions is to form
water-soluble products that can be excreted by the body.
A drug can ultimately undergo further metabolism during Phase III. In this phase, the drug that
gets transported via an ABC transporter, which will require energy (ATP) consumption to
actively uptake or efflux a compound from one side of the cell membrane to another. On the
other hand, it can get transported via an SLC transporter which facilitates the passage of
specific solutes across the membrane and actively transports other solutes against their
electrochemical gradients by merging this action with another solute or ion. The function of an
uptake transporter is to transfer a molecule into the cell, and the function of an efflux transporter
is to transfer a molecule outside the cell.
In general, these reactions serve to deactivate substances. However, they may also bioactivate
some substances by transforming them from nontoxic and/or inactive to the toxic and/or active
form. If the process produces a toxic intermediate, its effect on the body is dependent on a few
different factors. Some of these factors include how rapidly the intermediate can undergo
further metabolism to a less toxic metabolite, the amount of toxic byproduct that is produced
and then accumulated in cells, the type and amount of damage caused by the toxic intermediate,
and the factors that would prolong excretion of the toxic metabolite.

(Figure 4: Reaction of mechanisam)

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 Drug Metabolism

The liver is the principal site of drug metabolism,Although metabolism typically inactivates
drugs, some drug metabolites are pharmacologically active—sometimes even more so than the
parent compound. An inactive or weakly active substance that has an active metabolite is called
a prodrug, especially if designed to deliver the active moiety more effectively.

(Figure 5: Metabolism of drugs)

Drugs can be metabolized by oxidation, reduction, hydrolysis, hydration, conjugation,

condensation, or isomerization; whatever the process, the goal is to make the drug easier to
excrete. The enzymes involved in metabolism are present in many tissues but generally are
more concentrated in the liver. Drug metabolism rates vary among patients. Some patients
metabolize a drug so rapidly that therapeutically effective blood and tissue concentrations are
not reached; in others, metabolism may be so slow that usual doses have toxic effects.
Individual drug metabolism rates are influenced by genetic factors, coexisting disorders
(particularly chronic liver disorders and advanced heart failure), and drug interactions
(especially those involving induction or inhibition of metabolism).

For many drugs, metabolism occurs in 2 phases. Phase I reactions involve formation of a new
or modified functional group or cleavage (oxidation, reduction, hydrolysis); these reactions are
nonsynthetic. Phase II reactions involve conjugation with an endogenous substance (eg,
glucuronic acid, sulfate, glycine); these reactions are synthetic. Metabolites formed in synthetic
reactions are more polar and thus more readily excreted by the kidneys (in urine) and the liver
(in bile) than those formed in nonsynthetic reactions. Some drugs undergo only phase I or phase
II reactions; thus, phase numbers reflect functional rather than sequential classification.

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Hepatic drug transporters are present throughout parenchymal liver cells and affect a drug’s
liver disposition, metabolism, and elimination (for review, see The 2 primary types of
transporters are influx, which translocate molecules into the liver, and efflux, which mediate
excretion of drugs into the blood or bile. Genetic polymorphisms can variably affect the
expression and function of hepatic drug transporters to potentially alter a patient's susceptibility
to drug adverse effects and drug-induced liver injury. For example, carriers of certain
transporter genotypes exhibit increased blood levels of statins and are more susceptible to
statin-induced myopathy when statins are used for the treatment of hypercholesterolemia.

(Figure 6: Process of metabolism)

Factors affecting drug metabolism

The duration and intensity of pharmacological action of most lipophilic drugs are determined
by the rate they are metabolized to inactive products. The Cytochrome P450 monooxygenase
system is a crucial pathway in this regard. In general, anything that increases the rate of
metabolism (e.g., enzyme induction) of a pharmacologically active metabolite
will decrease the duration and intensity of the drug action. The opposite is also true, as
in enzyme inhibition. However, in cases where an enzyme is responsible for metabolizing a
pro-drug into a drug, enzyme induction can accelerate this conversion and increase drug levels,
potentially causing toxicity.

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Various physiological and pathological factors can also affect drug metabolism. Physiological
factors that can influence drug metabolism include age, individual variation
(e.g., pharmacogenetics), enterohepatic circulation, nutrition, sex differences or gut
microbiota. This last factor has significance because gut microorganisms are able to chemically
modify the structure of drugs through degradation and biotransformation processes, thus
altering the activity and toxicity of drugs. These processes can decrease the efficacy of drugs,
as is the case of digoxin in the presence of Eggerthella lenta in the microbiota. Genetic
variation (polymorphism) accounts for some of the variability in the effect of drugs.
In general, drugs are metabolized more slowly
in fetal, neonatal and elderly humans and animals than in adults. With N-acetyltransferases
(involved in Phase II reactions), individual variation creates a group of people who acetylate
slowly (slow acetylators) and those who acetylate quickly, split roughly 50:50 in the population
of Canada. This variation may have dramatic consequences, as the slow acetylator are more
prone to dose-dependent toxicity.
Dose, frequency, route of administration, tissue distribution and protein binding of the drug
affect its metabolism Pathological factors can also influence drug metabolism,
including liver, kidney, or heart diseases.
In silico modelling and simulation methods allow drug metabolism to be predicted in virtual
patient populations prior to performing clinical studies in human subjects. This can be used to
identify individuals most at risk from adverse reaction.

( Figure 7 : Factor affecting metabolism )

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 Clinical Significance
Biotransformation is affected by age, sex, nutritional status, disease state, medications, and
genetics of a patient. There is variance in expression and activity level of the CYP450 system
throughout prenatal development and from the neonatal period to adulthood. A limited amount
of studies have shown that cytochrome P450, specifically the activity of CYP3A4 is higher in
females than in males, in comparison to the activity of many other systems involved in drug
metabolism which may be higher in males than in females. A balanced diet, as compared to a
protein-deficient diet, will provide the necessary protein as well as essential metals and
minerals such as copper, zinc, and calcium needed for normal cellular enzymatic activities.
Also of note, patients with liver disease, such as cirrhosis, have damage to hepatocytes that can
then be replaced by fibrous connective tissue that is unable to carry out metabolic processes.

Most importantly, drug interactions should undergo checking for modulators of the CYP450
systems. Drugs with CYP activity may be inhibitors, inducers, or substrates for a specific CYP
enzymatic pathway which can lead to alteration of the metabolism of concurrently administered
agents. Some common substrates for the cytochrome P450 system include anti-epileptics,
theophylline, warfarin, oral contraceptive pills, and vitamin D. Therefore, when exposed to
inhibitors drug toxicity can ensue due to the decreased metabolism of the substrate, whereas
inducers would speed up the process by which these substrates become metabolized

PHARMACOLOGY AND DRUG THERAPY

Drug biotransformation or metabolism principally occurs in the liver, kidney, skin, and GI tract.
In the liver, biotransformation involves hydrolysis, oxidation, reduction, or demethylation and
conjugation of the metabolite with glycine, glucuronide, sulfate, or hippurate with subsequent
secretion into the bile. In the kidney, drugs may be filtered, filtered and secreted, filtered and
passively reabsorbed (e.g., acetaminophen), or filtered or actively secreted and passively
reabsorbed (e.g., salicylates).3 Many drugs used to treat rheumatic diseases are active in the
form in which they are administered; exceptions include sulindac, salsalate, prednisone,
leflunomide, azathioprine, mycophenolate mofetil (MMF), and cyclophosphamide, which
require biotransformation before they exert their principal effects.

Because the liver and kidney play such key roles in drug metabolism, dysfunction of these
organs may require alteration in drug dose. Generally, additional toxicity caused by drugs that
totally depend on the kidney for their elimination is not a danger, however, unless renal
function is diminished by more than 50%.2 In patients with significant renal or hepatic disease,
monitoring of drug levels and attention to the potential of drug toxicity become more critical.

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 Overview
Pharmaceutical substances known as xenobiotics are predominantly lipophilic and nonionized.
This enables them to permeate lipid bilayers, such as cell membranes, and interact with
intracellular target receptors. Lipophilic drugs have an advantage in crossing biological barriers
and reaching their intended sites of action. However, lipophilic drugs often have a restricted
capacity for renal expulsion or elimination from the body. When these drugs enter the kidneys
and undergo glomerular filtration, they can be quickly reabsorbed from the glomerular filtrate
back into the bloodstream through tubular reabsorption processes. This rapid reabsorption leads
to an accumulation of the drug in the body, extending its effects and potentially triggering toxic
reactions. To counteract this issue, lipophilic drugs undergo a biochemical transformation
known as biotransformation or metabolism. Lipophilic drugs are chemically modified during
metabolism into hydrophilic (water-soluble) derivatives. This transformation increases their
solubility in water and facilitates their efficient elimination from the body via urine. While
various bodily tissues can metabolize drugs, the liver is primarily responsible for this
biotransformation. Within the liver, specialized enzymes catalyze the biotransformation
process. Drug biotransformation involves two main enzymatically catalyzed reactions that
occur sequentially. Phase I reactions are catabolic reactions that involve the functionalization
of the drug molecule. These reactions introduce or expose functional groups, such as hydroxyl
(-OH), amino (-NH2), or carboxyl (-COOH) groups, to the drug molecule. This
functionalization often results in the formation of metabolites that are more polar than the
parent drug. Phase II reactions are anabolic and involve the conjugation of a drug or its phase
I metabolite. Conjugation reactions usually involve the addition of a larger, water-soluble
molecule to the drug or metabolite, such as glucuronic acid, a sulfate, or glutathione. This
conjugation further increases the drug's water solubility and prepares it for elimination from
the body. Generally, this metabolic process terminates the biological activity of the drug.
However, in the case of prodrugs, this process enhances their activity, leading to improved
therapeutic outcomes.

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