Phase I reactions

Describe oxidation, reduction and hydrolysis, name the enzymes and cofactors involved, and list the five possible outcomes of Phase I.

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Why this matters

Phase I is where most drugs are switched off, and occasionally switched on. It explains why a medicine can be inactive until the liver touches it, why the same reaction that clears a drug can also create a toxic product, and which enzyme or cofactor another drug might compete for. Without it, enzyme induction and inhibition in section 3.4 would be pure memorisation.

What you need to know first

You need section 3.1, especially the difference between microsomal and non-microsomal enzymes and the names of the four functional groups (–OH, –NH₂, –SH, –COOH). No organic chemistry is assumed: the reactions are described in words.

Words to know (6)

Oxidation: A reaction in which a drug gains oxygen or loses hydrogen, usually catalysed by CYP450 with oxygen and NADPH; the commonest Phase I reaction. Ethanol is oxidised to acetaldehyde and then to acetic acid.
Reduction: A reaction in which a drug gains electrons or hydrogen, catalysed by non-microsomal reductases; it is less common than oxidation and hydrolysis. Nitro-reductase reduces nitro groups on a small number of drugs.
Hydrolysis: A reaction that splits a drug molecule by adding water across a bond, carried out by amidases and esterases. Cholinesterase hydrolyses acetylcholine, the parasympathetic transmitter.
Functional group: A small chemical group attached to a molecule that gives it particular properties; Phase I introduces or unmasks groups such as –OH and –NH₂. A drug that gains an –OH group becomes much more water-soluble.
NADPH: A cofactor that carries electrons and acts as the reducing agent for CYP450 oxidation; without it, oxidation cannot proceed. NADPH is converted to NADP⁺ as the drug is oxidised.
Bioactivation: The conversion of a substance into a pharmacologically active or toxic product by metabolism. A prodrug is useless until bioactivation turns it into the active drug.

One idea at a timeIdea 1 / 6

In one sentence Phase I reactions make small chemical changes that add or expose a polar group on the drug molecule.

Phase I introduces or unmasks a functional group

The major pathways of hepatic biotransformation are Phase I and Phase II. The main Phase I reactions are oxidation, reduction and hydrolysis, and they are catalysed by microsomal cytochrome P450 enzymes in the smooth endoplasmic reticulum of the liver. Their shared result is that the molecule gains or exposes a functional group such as –OH, –NH₂, –SH or –COOH. Phase I is therefore called functionalisation: it does not usually make the drug leave the body by itself, but it prepares a handle that Phase II can use. A drug can pass through Phase I alone, through Phase II alone, or through both in either order.

For example

A drug with no polar group at all may enter Phase I; after hydroxylation it carries –OH and is a suitable substrate for the conjugation reactions of Phase II.

Think of it like

Phase I is like fitting a handle to a heavy crate: the handle does not carry the crate out of the building by itself, but it gives the next step something to grip. The comparison has a limit, because the new group is not a neutral fitting — it changes the molecule’s chemistry, so the same reaction can switch a prodrug on or create a toxic metabolite.

Pause and check yourself

What do all three Phase I reactions have in common?

Show the answer

They introduce or unmask a functional group such as –OH, –NH₂, –SH or –COOH, making the drug more polar and ready for Phase II.

Read detailed explanations

The oxidation reaction in detail: cofactor, iron and three ways to slow it: Oxidation consumes a reducing agent as well as oxygen. The CYP450 enzyme holds an iron atom at the centre of a haem ring. In the simplified equation of the lecture notes, Drug + O₂ + NADPH + H⁺ → Drug modified + H₂O + NADP⁺: the drug gains oxygen, molecular oxygen is split so that one atom joins the drug and one forms water, and NADPH supplies the electrons that keep the iron in its working state. Because the reaction needs a cofactor and an intact enzyme, it can be slowed in three different ways. Another drug can occupy the active site, which is competitive inhibition. The enzyme can be destroyed, which is irreversible inactivation. Or the supply of NADPH can be exhausted, which is cofactor depletion. Oxidation also covers chemically different jobs: amines lose a carbon chain, alcohols are dehydrogenated, and aromatic rings gain a hydroxyl group. Each product carries a new polar group, which is exactly what Phase II needs in order to attach its large water-soluble partner. The practical lesson is that any statement about ‘faster’ or ‘slower’ metabolism should name which of these three mechanisms is at work.
Hydrolysis and esterases: fast reactions outside the CYP450 system: Hydrolysis splits a molecule by adding water across a bond. Amidases break amides and esterases break esters, and both are non-microsomal enzymes that work in the cytoplasm rather than in the smooth endoplasmic reticulum. Because they are widely distributed and do not need NADPH or oxygen, hydrolysis can happen in plasma as well as in the liver, and it can be very fast. Cholinesterase, for example, inactivates acetylcholine, the transmitter of the parasympathetic nervous system, within milliseconds; the same family of enzymes hydrolyses ester-linked drugs. This speed is used deliberately in drug design. An ester bond can be built into a molecule so that plasma esterases destroy it soon after it enters the blood, which is the basis of short-acting local anaesthetics and of some prodrugs that must be converted only after absorption. It is also a warning: a drug that contains an ester may be broken down before it reaches its target, which is one reason chemical instability and the choice of route matter. Hydrolysis adds water and produces two smaller, more polar fragments, so like oxidation it usually leaves the molecule easier to excrete. Unlike oxidation, it is not affected by a CYP450 inhibitor, so it is always worth asking which enzyme system a drug actually uses.

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Phase I: three reactions

Oxidation is the commonest Phase I reaction, followed in frequency by hydrolysis and then reduction. All three introduce or unmask a polar functional group on the drug molecule.

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01 / 6Oxidation

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Oxidation

The commonest Phase I reaction, catalysed by CYP450 oxidase with oxygen and the reducing agent NADPH. It includes amine oxidation and alcohol dehydrogenation, such as the oxidation of ethanol to acetaldehyde.

Schematic summary, not to scale. Relative box sizes do not indicate how often each reaction occurs; oxidation is by far the most frequent.

Key points to remember

Phase I reactions are oxidation, reduction and hydrolysis; they introduce or unmask –OH, –NH₂, –SH or –COOH.
Oxidation is the commonest: CYP450 + O₂ + NADPH + H⁺ modify the drug, producing water and NADP⁺.
Reduction uses non-microsomal reductases such as nitro-reductase and is the least common reaction.
Hydrolysis uses amidases and esterases; cholinesterase inactivates acetylcholine.
Phase I outcomes: prodrug activation, activation to a more active or toxic metabolite, deactivation, increased hydrophilicity, and a substrate for Phase II.

Go deeper

Worked example

Classifying three reactions, then predicting their outcomes. Three descriptions are given. (1) Ethanol loses hydrogen and becomes acetaldehyde. (2) A drug containing an ester bond is split into two fragments in the plasma. (3) A nitro group on a drug gains hydrogen. Name each reaction and state which outcome of Phase I it illustrates.

Description 1 is oxidation, catalysed by CYP450 with oxygen and NADPH; the alcohol is dehydrogenated. Its outcome is bioactivation, because acetaldehyde is more toxic than ethanol.
Description 2 is hydrolysis, carried out by a plasma esterase. One outcome is increased hydrophilicity, because the two fragments are smaller and more polar than the parent molecule.
Description 3 is reduction, carried out by a non-microsomal reductase such as nitro-reductase; this is the least common of the three reactions and concerns few drugs.
In all three cases the product carries a new polar group, so it either is excreted more easily or offers a point of attachment for Phase II conjugation.

The name of a reaction tells you the enzyme class and the cofactor it needs; the outcome tells you whether the effect will rise, fall or become toxic. Always ask both questions.

Common misconception

Many beginners believe Phase I always makes a drug inactive. It often does, but it can also activate a prodrug or convert an active drug into a metabolite that is more active or toxic. Metabolism is a change, not a guarantee of safety.

Check your reasoning

A drug is described as a prodrug that is activated by CYP450 oxidation. If a patient also takes a strong CYP450 inhibitor, would you expect the effect of the prodrug to rise or fall? Explain your reasoning.

Reveal an answer

The effect should fall, because the prodrug must be oxidised before it can act. An inhibitor slows that oxidation, so less active metabolite is formed. This is the opposite of what happens with an ordinary active drug, whose effect would rise when its metabolism is inhibited.

Where this comes from

BMS 123 General Pharmacology II lecture notes · Lecture 3: Phase I pathways — oxidation, reduction, hydrolysis and outcomes of Phase I
Rang and Dale’s Pharmacology · Chapters on drug metabolism and the cytochrome P450 system

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About this course and its sources

Bilingual lessons organised from the supplied BMS 123 General Pharmacology II lecture notes (Principles of Pharmacokinetics and Principles of Pharmacodynamics). Diagrams are original educational schematics; pharmacology, nursing and French-language review are pending.