3.8 Required Practicals: Organisation and the Digestive System

A free GCSE Biology lesson for AQA (8461) from OBA School: required practical 3: food tests. Required practical 4: the effect of pH on the rate of reaction of amylase. Watch the video, read the explanation and worked examples, then try the revision questions. You can work through the full lesson, with every video, practice quizzes and printable worksheets with mark schemes, in our free online course.

What you will learn

• Carry out and describe the food tests for starch, reducing sugars, protein and lipids
• Plan an investigation into the effect of pH on amylase, identifying the variables
• Calculate rates (1000 ÷ time) and means from results
• Identify hazards, sources of error and improvements

Key words

• Qualitative test: A test that shows whether a substance is present (by a colour change), not how much.
• Reagent: A chemical used to test for something, e.g. Benedict’s solution.
• Independent variable: The variable you change on purpose (here, pH).
• Dependent variable: The variable you measure (here, time for starch to disappear).
• Control variable: A variable kept the same so the test is fair.
• Buffer solution: A solution that keeps the pH at a fixed value.
• Repeatable: Similar results are obtained when the same person repeats the method.

Required practical 3: food tests

Aim: to use chemical reagents to test foods for carbohydrates, lipids and proteins. Preparing a sample: (1) grind a small piece of food with distilled water using a pestle and mortar; (2) transfer to a beaker, stir, then filter so you have a clear solution; (3) divide it into test tubes, one for each test.

• Starch; Method Add a few drops of iodine solution.; Positive result Orange-brown → blue-black
• Reducing sugars (e.g. glucose); Method Add Benedict’s solution (equal volume). Heat in a water bath at about 75 °C for 5 minutes.; Positive result Blue → green, yellow or brick-red (more sugar → further along)
• Protein; Method Add Biuret reagent (or sodium hydroxide then copper sulfate solution).; Positive result Blue → purple / lilac
• Lipids; Method Add a few drops of Sudan III stain and shake.; Positive result A red-stained oily layer forms on top

Risk assessment: wear eye protection. Biuret reagent contains sodium hydroxide (irritant/corrosive in higher concentrations) and copper sulfate (harmful). Iodine solution is an irritant. Sudan III is dissolved in ethanol, which is flammable – keep away from flames. Use a water bath (not a Bunsen flame) to heat Benedict’s test and do not point tubes at anyone. Do not eat food in the lab.

Exam tip: Benedict’s test needs heat; Biuret does not. Sucrose (table sugar) is not a reducing sugar, so it gives a negative Benedict’s result.

Required practical 4: effect of pH on amylase

Aim: to find how pH affects the rate at which amylase breaks down starch. Equipment: spotting tile, iodine solution, amylase solution, 1% starch solution, buffer solutions (e.g. pH 4–9), water bath at 35 °C, test tubes, syringes / measuring cylinders, stop clock, dropping pipette, thermometer.

• Step 1: Put one drop of iodine solution in each well of a spotting tile.
• Step 2: Put 2 cm³ of amylase and 1 cm³ of pH 5 buffer in one tube, and 2 cm³ of starch solution in another. Stand both in the water bath for 5 minutes to reach 35 °C.
• Step 3: Mix the starch into the amylase/buffer and start the stop clock.
• Step 4: Every 30 seconds, put one drop of the mixture onto a fresh well of iodine.
• Step 5: Record the time when the iodine stays orange-brown (no starch left).
• Step 6: Repeat with each buffer, and repeat each pH three times. Calculate a mean.

• Independent; In this practical pH (set with buffer solutions)
• Dependent; In this practical Time for starch to be fully broken down (iodine stays orange-brown)
• Control; In this practical Temperature (water bath); volume and concentration of amylase and of starch; volume of buffer; sampling interval

Risk assessment: wear eye protection – iodine is an irritant and amylase can cause an allergic reaction (wipe spills, avoid skin contact). Take care with hot water.

Processing results

A shorter time means a faster rate. Use rate = 1000 ÷ time (units s⁻¹). Identify any anomalous repeat and leave it out of the mean.

Worked example: mean and rate

At pH 6 the three times were 60 s, 90 s and 60 s. The 90 s result is not anomalous here (it is within one sampling interval), so use all three.
Mean = (60 + 90 + 60) ÷ 3 = 70 s. Rate = 1000 ÷ 70 = 14.3 s⁻¹ (3 s.f.).

Answer: mean 70 s; rate 14.3 s⁻¹

Errors and improvements

• Sampling only every 30 s; Effect End point only known to within 30 s; Improvement Sample every 10 s
• Judging the colour by eye; Effect Subjective; people disagree on the end point; Improvement Use a colorimeter, or compare with a colour standard
• Solutions not at the bath temperature; Effect Rate changes during the test; Improvement Pre-warm all solutions; check with a thermometer
• Gaps between pH values; Effect Optimum not found precisely; Improvement Repeat with smaller pH steps near the optimum (e.g. 6.5, 7.0, 7.5)

Common mistakes

• Forgetting to heat in the Benedict’s test, or heating Biuret.
• Writing “clear” when you mean “colourless”. Clear means see-through; it can still be coloured.
• Saying temperature is the independent variable in the pH practical. It is a control variable.
• Calculating rate as time ÷ 1000. The rate is 1000 ÷ time, so shorter times give bigger rates.
• Including an anomalous result in the mean.

Revision questions

Try these, then check your answers below each one.

Question 1. True or false? The Biuret test must be heated in a water bath.

Answer: False

Working: False. Only Benedict’s test needs heating (water bath at about 75 °C).

Question 2. In the investigation of pH on amylase, what is the dependent variable? (Choose from: pH, Temperature of the water bath, Time taken for the starch to be broken down, Volume of amylase.)

Answer: Time taken for the starch to be broken down

Working: pH is changed (independent); the time for iodine to stay orange-brown is measured (dependent).

Question 3. Why are the amylase and starch put in the water bath for 5 minutes before mixing? (Choose from: So they start at the set temperature (a control variable), To denature the amylase, To dissolve the iodine, To change the pH.)

Answer: So they start at the set temperature (a control variable)

Working: Pre-warming keeps temperature the same for every pH, so it is a fair test.

Related lessons

• Previous lesson: 3.7 Making Digestion Efficient
• Next lesson: 4.1 The Blood
• In this chapter: 3.1 Tissues and Organs
• In this chapter: 3.2 The Human Digestive System
• In this chapter: 3.3 The Chemistry of Food

Continue in the full course

This lesson is section 3.8 (ARPBH0308) of our free GCSE Biology course for AQA. In the full course you get every video for this section, a self-marking practice quiz, a printable worksheet with a mark scheme, chapter tests, and your progress saved. Go to the full GCSE Biology course.