Discovery and development of drugs — AQA GCSE Combined Science
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Discovery and development of drugs explained
New medicines begin with a potential drug, often found by screening thousands of compounds or by modifying a known active molecule.
Read the full explanation
Preclinical testing happens before any human exposure: laboratory studies on cells and tissues establish whether the substance works, and tests on animals such as mice or rats assess toxicity, dosage and side effects. If results are promising, clinical testing follows in three stages. Small groups of healthy volunteers check safety and dose; larger groups of patients check effectiveness and side effects; then many patients in trials compare the new drug with existing treatments or a placebo to confirm efficacy. Regulators review the evidence before a licence is granted.
Traditionally drugs were extracted from plants and microorganisms.
Many medicines used today originated from natural sources. For centuries, people extracted active substances from plants and microorganisms, often guided by traditional remedies. A classic example is digitalis, obtained from foxglove leaves, which affects heart muscle and is still used to treat some heart conditions. Aspirin was developed from salicylic acid found in willow bark. Penicillin is extracted from the mould Penicillium, a microorganism, and was discovered when Alexander Fleming noticed that mould killed bacteria on a plate. These examples show that plants and microorganisms are important sources of drugs, and modern scientists still screen natural products to find new medicines.
The heart drug digitalis originates from foxgloves.
Digitalis is a medicine used to strengthen the force of heart contractions, and it was originally obtained from the foxglove plant, Digitalis purpurea. Traditional herbal knowledge showed that foxglove extracts affected the heart; later scientific work isolated active compounds called cardiac glycosides, such as digoxin, and established safe doses. This illustrates a common drug-development pathway: a natural source provides a starting compound, which is then purified, tested for efficacy and toxicity, and formulated into a medicine. Students should link the plant source to the drug's medical use, and recognise that the active substance is a chemical produced by the plant rather than the whole plant being the drug. Foxgloves remain the biological origin even though modern production may use extraction or chemical synthesis.
The painkiller aspirin originates from willow.
Aspirin is a painkiller and anti-inflammatory medicine whose discovery traces back to willow trees, particularly species such as Salix alba. Willow bark contains salicin, a compound that the body converts into a pain-relieving substance. Chemists modified salicin to make acetylsalicylic acid, the active ingredient in aspirin, which is more effective and gentler on the stomach than raw willow preparations. This example shows how a traditional remedy from a plant can be investigated, its active chemical identified, and a related medicine developed through chemical modification and testing. Students should connect willow as the original source with aspirin as the modern drug, and recognise that the medicine is a purified or modified chemical rather than simply powdered bark.
Penicillin was discovered by Alexander Fleming from the Penicillium mould.
In 1928 Alexander Fleming noticed that a mould contaminating a culture plate of bacteria had killed the bacteria around it. He identified the mould as Penicillium and named the antibacterial substance it produced penicillin. This was the first antibiotic discovered. Fleming did not purify penicillin or turn it into a usable medicine; that required later work by Florey and Chain, who mass-produced it during the 1940s. The discovery shows how a chance observation, followed by careful scientific investigation, can lead to a life-saving drug. Penicillin works by interfering with bacterial cell wall formation, so it kills bacteria without directly damaging human cells. It is effective only against bacteria, not viruses, and some bacteria have evolved resistance, so antibiotics must be used carefully.
Most new drugs are synthesised by chemists in the pharmaceutical industry. However, the starting point may still be a chemical extracted from a plant.
Modern drug discovery usually begins with a target molecule that chemists design and synthesise in a laboratory. Pharmaceutical chemists make many related compounds, test them for useful activity and modify structures to improve effectiveness and reduce side effects. Even so, nature remains an important source of starting points. A chemical extracted from a plant may be purified, its structure determined, and then used as a template for synthesis or chemical modification. For example, willow bark contains salicin, which inspired aspirin, and the rosy periwinkle yields alkaloids used in some cancer treatments. The process involves identifying a lead compound, testing it, and then developing a safe, effective medicine through trials.
New medical drugs have to be tested and trialled before being used to check that they are safe and effective.
Before a new medical drug can be prescribed to patients, it must pass through testing and clinical trials. Testing first uses laboratory studies, including cells and tissues, to look for harmful effects. If results are promising, trials move to healthy volunteers at low dose to check safety, then to patients to see whether the drug actually treats the condition. Safety means the drug does not cause unacceptable harm; effectiveness means it produces the intended therapeutic benefit at a suitable dose. For example, a new antibiotic might kill bacteria in culture, then be given to volunteers to check side effects, then to patients to compare recovery with an existing treatment. Only after evidence from all stages is reviewed may the drug be approved for medical use.
New drugs are extensively tested for toxicity, efficacy and dose.
Testing a new drug covers three linked questions. Toxicity asks whether the substance harms the body, for example by damaging the liver or kidneys. Efficacy asks whether the drug produces the intended therapeutic effect, such as lowering blood pressure or killing a pathogen. Dose asks what amount gives the desired effect with acceptable side effects, because too little may not work and too much may be toxic. Preclinical laboratory work, often using cells or tissues, can reveal toxicity and suggest a starting dose. Clinical trials then compare groups receiving the drug with controls, using placebos where appropriate, to measure efficacy and refine dose. For example, a new painkiller might be tested at several doses to find the lowest dose that relieves pain without causing unacceptable side effects.
Preclinical testing is done in a laboratory using cells, tissues and live animals.
Preclinical testing happens before any medicine is given to people. It takes place in a laboratory and uses cells, tissues and live animals. Scientists first test a new drug on cultured cells to see whether it harms them or changes how they grow. They then test on tissues, such as a piece of heart or liver tissue, to see how the drug affects a whole group of cells working together. Finally, live animals are used, for example mice or rats, because a whole living body shows how the drug moves around, how it is broken down and whether it causes harm to organs. This stage checks safety and effectiveness before human trials, and it also helps decide a safe starting dose. It cannot prove a drug is safe for humans, so clinical trials must follow.
Clinical trials use healthy volunteers and patients.
Clinical trials happen after preclinical testing and involve people. Healthy volunteers are used first, usually in small numbers, to check that the drug is safe and to find out how the body handles it, including any side effects. If the drug seems safe, patients who have the illness are then given it to see whether it actually treats the condition. Some patients may receive a placebo so results can be compared, and trials are often blind or double-blind to reduce bias. The dose is also checked so that the benefits outweigh the risks. This stage is essential because effects in animals do not always match effects in humans, so clinical trials provide evidence that a treatment is safe and effective before it is approved for general use.
Very low doses of the drug are given at the start of the clinical trial.
Clinical trials test a new drug on humans only after laboratory and animal testing. At the start, volunteers receive very low doses. This is a safety precaution: humans may react differently from animals, and a low dose limits any harm while still allowing researchers to observe the drug's effects. For example, a trial might begin with a dose of a few milligrams and then increase it gradually in later stages. Starting low also helps scientists gather early data on how the body absorbs and removes the drug. If unexpected side effects appear, the trial can be stopped before anyone receives a larger dose.
If the drug is found to be safe, further clinical trials are carried out to find the optimum dose for the drug.
Once early clinical trials show that a drug is safe, further trials are carried out to find the optimum dose. The optimum dose is the amount that gives the best balance between treating the condition and causing acceptable side effects. Researchers test different doses in groups of patients and compare how well each dose works and how many side effects occur. For example, a trial might compare 10 mg, 20 mg and 40 mg doses. The optimum dose is not simply the largest dose; a higher dose may cause more harm without extra benefit. Finding the optimum dose allows the drug to be prescribed at a level that is effective and as safe as possible.
In double blind trials, some patients are given a placebo.
A double blind trial is a way of testing a new medicine fairly. Patients are split into groups: one group receives the new drug, while some patients receive a placebo — a treatment that looks identical but contains no active drug. Neither the patients nor the doctors assessing them know who received which treatment until the trial ends. This removes bias: patients cannot report feeling better simply because they expect the drug to work, and doctors cannot unconsciously favour one group when judging symptoms. For example, in a trial of a new painkiller, some patients swallow tablets containing the drug and others swallow identical dummy tablets. Comparing the two groups shows whether the drug itself, rather than expectation, caused any improvement.
Your focus
- Sequence preclinical and clinical testing accurately.
- State the purpose of each testing stage.
- Explain why regulation and licensing are needed before a medicine is used.
Show all 39 objectives
- Identify plants and microorganisms as traditional sources of drugs.
- Give named examples of drugs from natural sources.
- Describe how a natural source led to a useful medicine.
- Identify foxgloves as the natural source of the heart drug digitalis.
- Describe the effect of digitalis on heart muscle contraction.
- Explain that the active compound is extracted and developed into a safe medicine.
- Identify willow as the natural source linked to the painkiller aspirin.
- Describe how salicin from willow led to the development of aspirin.
- Explain that aspirin is a purified or modified active compound rather than raw willow bark.
- Identify Alexander Fleming as the scientist who discovered penicillin from the Penicillium mould.
- Describe how the mould produced a substance that killed bacteria on a culture plate.
- Explain that penicillin is an antibiotic used against bacterial infections and not viruses.
- State that most new drugs are synthesised by chemists in the pharmaceutical industry.
- Describe how a chemical extracted from a plant can be a starting point for a new drug.
- Explain how synthesis and testing allow chemists to develop effective and safe medicines.
- State that new medical drugs are tested and trialled before use.
- Describe how testing checks both safety and effectiveness.
- Explain why human trials follow laboratory testing in the development of a new drug.
- Define toxicity, efficacy and dose in the context of drug testing.
- Describe how each of toxicity, efficacy and dose is investigated during drug development.
- Explain why all three must be assessed before a new drug is approved for use.
- State that preclinical testing is carried out in a laboratory using cells, tissues and live animals.
- Describe the order in which cells, tissues and live animals are used.
- Explain why preclinical testing is needed before clinical trials begin.
- State that clinical trials use healthy volunteers and patients.
- Describe the different purposes of testing healthy volunteers and patients.
- Explain how placebos and blind trials help produce valid results.
- State that very low doses are given at the start of a clinical trial.
- Explain why starting with a very low dose improves safety for volunteers.
- Describe how the dose is increased gradually during a clinical trial.
- State that further clinical trials find the optimum dose after safety is established.
- Explain what is meant by the optimum dose of a drug.
- Describe how different doses are compared in clinical trials to identify the optimum dose.
- State what a placebo is and why some patients in a double blind trial receive one.
- Explain how a double blind trial reduces bias from both patients and assessing doctors.
- Compare outcomes for patients given the new drug with those given the placebo to judge whether the drug works.
Discovery and development of drugs exam tips
Marking Points
- Preclinical testing occurs before human trials and uses cells, tissues and whole animals to assess efficacy, toxicity and safe dosage.
- Clinical testing involves healthy volunteers first to check safety and tolerance, then patients to assess effectiveness and side effects.
- Later clinical trials compare the new medicine with existing treatments or a placebo, often using double-blind methods to reduce bias.
- Regulatory approval and licensing follow successful trials before a medicine can be prescribed.
- The whole process is long, costly and strictly regulated to protect participants and patients.
- Drugs have traditionally been obtained from plants, such as digitalis from foxglove and aspirin from willow.
- Microorganisms are also traditional sources, for example penicillin from the mould Penicillium.
- Natural sources provide active compounds that can be extracted, purified and later modified to make medicines.
- Traditional knowledge and observation of natural effects often guided early drug discovery.
- Digitalis is a drug that increases the strength of heart muscle contractions and is used to treat certain heart conditions such as heart failure.
- Its original natural source is the foxglove plant, Digitalis purpurea.
- The active compounds are cardiac glycosides, for example digoxin, which are extracted from foxglove tissue.
- This is an example of a drug discovered from a natural source, later developed by purifying and testing the active compound.
- Students should distinguish the plant source from the purified medicine: the drug is the active chemical, not the whole plant.
- The example shows how traditional plant knowledge can lead to modern pharmaceutical development.
- Aspirin is a painkiller used to relieve pain and reduce inflammation and fever.
- Its origin is the willow tree, whose bark contains the compound salicin.
- Salicin is converted in the body to a pain-relieving substance, and chemists developed acetylsalicylic acid as the active ingredient of aspirin.
- This illustrates drug development from a traditional plant remedy through identifying, modifying and testing an active compound.
- Students should distinguish willow bark as the natural source from aspirin as the purified or synthesised medicine.
- The example shows that chemical modification can improve a natural lead compound, for example making it more effective or reducing side effects.
- State that Alexander Fleming discovered penicillin in 1928 from the Penicillium mould.
- Describe that the mould produced a substance that killed bacteria growing on the culture plate.
- Name penicillin as an antibiotic that kills bacteria and is used to treat bacterial infections.
- Recognise that Fleming's discovery was a chance observation followed by scientific investigation.
- Explain that penicillin was later developed into a usable medicine by other scientists, such as Florey and Chain.
- Distinguish between discovery (Fleming) and development or mass production (later scientists).
- State that most new drugs are synthesised by chemists in the pharmaceutical industry.
- Describe that chemists design and make new compounds, often by chemical synthesis in a laboratory.
- Explain that a starting point for a new drug may be a chemical extracted from a plant.
- Give a named example, such as salicin from willow bark leading to aspirin, or an alkaloid from the rosy periwinkle.
- Outline that the extracted chemical can be purified, tested and then modified or used as a template for synthesis.
- Recognise that synthesis allows many similar compounds to be made and tested to find the most effective and safest drug.
- Testing before medical use identifies harmful effects such as organ damage or severe side effects, so safety can be judged.
- Trials establish whether the drug works against the target condition, which is what effective means in this context.
- Laboratory testing on cells, tissues or animals can screen a candidate drug before any human exposure.
- Clinical trials in humans normally begin with healthy volunteers at low dose to assess safety and tolerance.
- Later trials use patients who have the condition so that therapeutic effect can be compared with existing treatments or a placebo.
- Evidence from all stages is reviewed before a drug is approved for prescribing to the general public.
- Toxicity testing looks for harmful effects of the drug on the body, including damage to organs or dangerous side effects.
- Efficacy testing shows whether the drug produces the intended therapeutic effect in patients.
- Dose testing finds an amount that gives benefit while keeping side effects at an acceptable level.
- Preclinical laboratory studies can indicate toxicity and help choose a safe starting dose before human trials.
- Clinical trials compare treated groups with controls, often using placebos, to judge efficacy fairly.
- Testing several doses allows the relationship between dose, benefit and harm to be evaluated.
- Preclinical testing occurs in a laboratory before any testing on humans.
- Cells are used first, often grown in culture, to check for harm to individual cells.
- Tissues are used to see the effect on groups of cells working together.
- Live animals are used to study effects on a whole living body, including organs and how the drug is broken down.
- The purpose is to test safety and effectiveness and to help choose a safe starting dose for human trials.
- Results from preclinical testing cannot guarantee that a drug is safe for humans.
- Clinical trials take place after preclinical testing and involve human participants.
- Healthy volunteers are used first to test safety, side effects and how the body processes the drug.
- Patients with the illness are then used to test whether the drug actually works as a treatment.
- Control groups, placebos and blind or double-blind methods may be used to make results valid and reduce bias.
- Trials help establish a suitable dose and show whether benefits outweigh risks.
- Clinical trials are needed because animal results do not always predict human responses.
- Very low doses are used at the start of a clinical trial to reduce the risk of harm to human volunteers.
- The first human doses are given only after earlier laboratory and animal testing has been completed.
- Starting with a low dose allows researchers to observe effects and side effects before increasing the dose.
- Dose escalation is gradual, so any unexpected reaction can be detected and the trial stopped.
- Low starting doses help establish how the human body handles the drug, including absorption and removal.
- Further clinical trials are carried out only after the drug has been shown to be safe in earlier trials.
- The purpose of these further trials is to find the optimum dose of the drug.
- The optimum dose gives the best balance between effectiveness and acceptable side effects.
- Different doses are compared in groups of patients to see which works best with fewest problems.
- The optimum dose is not always the highest dose, because larger doses can cause more side effects.
- A placebo is a dummy treatment with no active drug, made to look, taste and be given in the same way as the real medicine.
- In a double blind trial, neither the patients nor the doctors or researchers assessing them know which treatment each patient received.
- Some patients receive the placebo so their outcomes can be compared with those of patients receiving the new drug.
- The placebo group controls for the psychological effect of expecting to improve, so any real effect of the drug can be identified.
- Keeping patients and assessors unaware of group allocation reduces bias in how symptoms and side effects are reported and judged.
- Ethical trials still allow placebos because patients are informed and consent, and no proven effective treatment is withheld.
Examiner Tips
- 💡Use the words preclinical and clinical correctly and in order, because examiners look for the sequence.
- 💡Give a reason for each stage, such as checking toxicity before human exposure or comparing with a placebo to judge effectiveness.
- 💡Link the stages to the purpose of protecting patients, which shows understanding rather than a list.
- 💡Name at least one plant and one microorganism source with the drug obtained, because specific examples earn credit.
- 💡Link the source to the drug's effect, such as foxglove affecting the heart, to show understanding.
- 💡Use the term active substance or compound when describing what is extracted.
- 💡Name both the drug and its plant source in one clear sentence, for example 'digitalis, from foxgloves'.
- 💡Link the drug to its effect on the heart rather than only naming the plant.
- 💡Use the term 'active compound' when explaining that a chemical from the plant is responsible for the medical effect.
- 💡State the plant source and the drug together, for example 'aspirin, originally from willow'.
- 💡Mention salicin as the compound in willow that led to aspirin.
- 💡Explain one advantage of the developed drug, such as being more effective or better tolerated, rather than only naming the plant.
- 💡Link the scientist to the specific action: Fleming observed, identified the mould and named penicillin.
- 💡Use the word antibiotic correctly and state that it treats bacterial infections.
- 💡If asked about drug development, separate discovery from later purification, testing and mass production.
- 💡Check that you have not written that Fleming made penicillin a usable medicine; credit usually depends on the correct role.
- 💡Use the phrase 'starting point' to show that a plant chemical may lead to a synthesised drug rather than being the final medicine.
- 💡Name one plant example accurately, such as willow bark and aspirin, to support your answer.
- 💡Distinguish clearly between synthesis in the pharmaceutical industry and extraction from a plant.
- 💡If asked to compare, mention that synthesis allows many variations to be made and tested.
- 💡Link each stage of testing to its purpose: safety, effectiveness or finding a suitable dose.
- 💡Use the terms safe and effective precisely; safe means absence of unacceptable harm, effective means the drug treats the condition.
- 💡When describing a sequence, make clear why healthy volunteers are used before patients rather than simply listing stages.
- 💡Define toxicity, efficacy and dose clearly, then link each to a specific stage or purpose of testing.
- 💡Use comparative language when describing trials, such as treated group compared with control group.
- 💡Explain why dose matters by referring to the balance between therapeutic benefit and unwanted effects.
- 💡Name all three types of preclinical test — cells, tissues and live animals — because questions often award a mark for each.
- 💡Link each test to its purpose, such as cells for cell damage and animals for whole-body effects.
- 💡Use the phrase 'before human trials' to show the correct order of drug development.
- 💡Distinguish clearly between healthy volunteers, used for safety, and patients, used to test effectiveness.
- 💡Mention placebos or blind trials when asked how bias is reduced, but do not invent exact numbers of participants.
- 💡Use the order preclinical testing, then clinical trials, then approval when describing drug development.
- 💡Link the low starting dose directly to safety, using words such as 'reduces risk of harm' or 'limits side effects'.
- 💡State clearly that the dose is increased gradually rather than jumping to the therapeutic dose.
- 💡Use the phrase 'clinical trial' accurately to mean testing on humans, not animals or cells.
- 💡Define the optimum dose in terms of both effectiveness and side effects, not just one of these.
- 💡Use comparative language such as 'different doses are compared' when describing how the optimum dose is found.
- 💡Make the sequence clear: safety first, then further trials for the optimum dose.
- 💡Define placebo and double blind separately, then link them in one sentence about fair comparison.
- 💡When asked why a placebo is used, refer to controlling the expectation effect and reducing bias, not just to keeping patients happy.
- 💡Use the phrase 'neither the patients nor the doctors know who received which treatment' to show precise understanding.
Common Mistakes
- Thinking clinical testing starts with patients: the first clinical stage normally uses healthy volunteers to check safety, so correct this by stating the sequence safety first, then effectiveness.
- Confusing preclinical and clinical stages: preclinical means laboratory and animal work before humans, so correct this by linking each stage to whether humans are involved.
- Assuming a drug is approved once it works in cells: regulators need evidence from trials, so correct this by adding that licensing follows successful clinical testing.
- Saying all drugs are made synthetically: many were originally extracted from plants or microorganisms, so correct this by naming a plant or microorganism source.
- Confusing the source with the scientist: Fleming discovered penicillin, but the drug comes from the mould Penicillium, so correct this by separating discoverer from source.
- Believing traditional extraction is the same as modern synthesis: extraction obtains a natural compound, while synthesis can make or modify it, so correct this by distinguishing the two.
- Saying digitalis is made by the heart; correction: it acts on the heart but originates from foxgloves.
- Confusing the source with the drug; correction: foxglove is the plant source, while digitalis or digoxin is the active medicine.
- Claiming all foxglove material is safe medicine; correction: foxglove plants are toxic, and only controlled purified doses are used as a medicine.
- Saying aspirin is extracted directly as salicin from willow; correction: willow contains salicin, and aspirin is acetylsalicylic acid developed from it.
- Confusing the source with the final medicine; correction: willow is the plant source, while aspirin is the manufactured active compound.
- Assuming all willow bark is a safe medicine; correction: raw plant material can be variable and harmful, so controlled purified doses are used.
- Error: saying Fleming invented or mass-produced penicillin. Correction: Fleming discovered penicillin; Florey and Chain later developed it into a medicine.
- Error: claiming penicillin kills viruses. Correction: penicillin is an antibiotic and acts only on bacteria, not viruses.
- Error: stating that Fleming deliberately set out to find an antibiotic. Correction: the discovery followed an accidental contamination of a culture plate.
- Error: writing that penicillin was discovered from a bacterium. Correction: it was produced by the Penicillium mould.
- Error: saying all new drugs come directly from plants. Correction: most are synthesised by chemists, but plants can provide starting chemicals.
- Error: confusing synthesis with extraction. Correction: synthesis means making a compound by chemical reactions; extraction means obtaining it from a natural source.
- Error: claiming that a plant chemical is automatically a safe medicine. Correction: it must be tested and developed before use.
- Error: writing that pharmaceutical chemists only copy plant chemicals exactly. Correction: they often modify the structure to improve the drug.
- Thinking that a drug only needs to be tested for safety: correction — effectiveness must also be demonstrated, because a safe drug that does not treat the condition is not useful.
- Assuming human trials always start with patients: correction — early human trials usually use healthy volunteers to check safety before patients are exposed.
- Believing that laboratory testing alone is enough for approval: correction — human trials are required because effects in cells or animals do not always predict effects in people.
- Confusing toxicity with efficacy: correction — toxicity is about harm, while efficacy is about whether the drug works.
- Treating dose as a single fixed value rather than a range that must be optimised: correction — different doses are tested to find one that balances benefit and side effects.
- Assuming a placebo is always used even when an effective treatment exists: correction — where a proven treatment exists, it may be unethical to withhold it, so trials may compare the new drug with the existing treatment.
- Thinking preclinical testing uses human volunteers: correct this by stating that humans are only used in clinical trials, after laboratory and animal testing.
- Believing animal testing proves a drug is safe for humans: correct this by explaining that animal results only give an indication, so human trials are still needed.
- Confusing cells with tissues: correct this by saying cells are individual units grown in culture, while tissues are groups of similar cells working together.
- Saying patients are always tested before healthy volunteers: correct this by stating healthy volunteers come first to check safety.
- Thinking a placebo is a real medicine: correct this by explaining a placebo contains no active drug and is used for comparison.
- Assuming clinical trials prove a drug works for everyone: correct this by noting trials provide evidence of safety and effectiveness for a defined group, with possible side effects.
- Thinking clinical trials begin with the full intended dose: correct this by stating that very low doses are given first and increased gradually.
- Confusing clinical trials with animal testing: correct this by noting that clinical trials involve human volunteers and follow earlier laboratory and animal tests.
- Assuming a low dose proves the drug works: correct this by explaining that early low doses mainly check safety and effects, not effectiveness.
- Believing the optimum dose is the maximum dose: correct this by explaining that the optimum dose balances benefit against side effects.
- Thinking safety testing and optimum-dose testing happen at the same time: correct this by stating that optimum-dose trials follow safety trials.
- Assuming all patients need the same dose: correct this by noting that trials compare doses to find the best overall dose for the drug.
- Thinking a placebo contains a low dose of the drug: it contains no active drug at all, so any change in the placebo group is not caused by the medicine.
- Confusing double blind with single blind: in a single blind trial only the patients do not know their treatment, whereas in a double blind trial the assessors also do not know.
- Assuming everyone in the trial gets a placebo: only some patients do, and the rest receive the treatment being tested so the groups can be compared.