Fruit Juice Production & Enzyme Recovery
Technical & Teaching Notes
Type and purpose of activity
This experiment can be used to:
 develop knowledge and understanding of the use of cellulases, pectinases and amylases
in the production of fruit drinks
 develop problem solving skills and in particular
o conclusions drawn are valid and explanations given are supported by evidence
o experimental procedures are planned, designed and evaluated appropriately.
Background information
Fruit juice companies use a variety of different treatments and enzymes to maximise their yield.
The enzymes which are used are designed to break down the cells walls within the fruits and
release the liquids and the sugars which make up the fruit. Pectinases, amylases and cellulases
all break down different structures of the fruit cells and so affect the extraction process in different
ways. During breakdown of the fruit cells a variety of polysaccharides are found within the juice
extract, these can cause the juice to become cloudy and reduce its market value. Pectinases and
amylases can both break down these insoluble compounds releasing soluble sugars which clarify
the juice producing a clearer, sweeter product.
In this experiment students will investigate the effectiveness of these different enzymes. Note that
the experimental procedure and recording of results in this practical is very straightforward. For
Outcome 3 purposes, emphasis should be placed on the evaluation in the written report.
Classroom management
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Students can work individually or in pairs for this experiment.
The experiment will take about 1 hour to complete.
The experiment can be developed further by using the Supplementary Student
Information Sheet to investigate different ways of re-using the enzymes.
Supply of materials
In order to satisfy the core skill in problem solving, students will be required to identify and obtain
resources required for themselves. Further advice on supply of material is given in the Technical
Guide.
Extension work
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Different fruits and vegetables can be used.
The enzyme data sheets can be used to identify the optimum pH and temperature of
these enzymes and this can be tested through experimentation.
The enzymes can be immobilised and continual reuse of the enzymes can be tested (see
supplementary sheet).
Different methods can be used to investigate re-using the enzymes (see supplementary
sheet).
Science & Plants for Schools: www.saps.org.uk
Fruit Juice Production: p. 1
This document may be photocopied for educational use in any institution taking part in the SAPS programme.
It may not be photocopied for any other purpose. Revised 2012.
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A comparison can be made between these enzymes and other amylases, pectinases and
cellulases.
Find out the optimum combination of enzymes for maximum yield.
Determine the effect that reducing the concentration of enzyme has on yield.
References
NCBE booklet “In a jam and out of juice”. ISBN: 0 7049 1373 9
Available free from NCBE’s website:
http://www.ncbe.reading.ac.uk/NCBE/PROTOCOLS/juice.html or from NCBE, Science and
Technology Centre, Earley Gate, University of Reading, Whiteknights, READING, RG6 6BZ. Tel:
0118 987 3743 Fax: 0118 975 0140.
This experiment was produced by the SAPS Biotechnology Scotland Project. Funding for the
project was provided by SAPS, Unilever and The Scottish Office. Support was also provided by
Edinburgh University, Quest International, Learning and Teaching Scotland, the Higher Still
Development Unit and SSERC.
Materials required
Materials required by each student/group:
 one apple
 knife
 Termamyl/Pectinex/Celluclast
 four 10 cm3 measuring cylinder
 filter papers
 4 small filter funnels
 timer
 four 100 cm3 beakers
 four weigh boats
 stirring rod
 5 cuvettes (or test tubes if suitable colorimeter is used)
Materials to be shared:
 balance (accurate to 0.1 g)
 colorimeter
For the SUPPLEMENTARY part of this activity in addition to the above materials:
Materials required by each student/group
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enzyme Immobilisation Instruction Cards
200 cm3 1.5% calcium chloride
10 cm3 4% sodium alginate
enzyme solution e.g. pectinex/celluclast
distilled water
250 cm3 beaker
10 cm3 syringe
20 cm3 syringe
tea strainer
iron fillings
bar magnet in a plastic bag
Science & Plants for Schools: www.saps.org.uk
Fruit Juice Production: p. 2
This document may be photocopied for educational use in any institution taking part in the SAPS programme.
It may not be photocopied for any other purpose. Revised 2012.
Preparation of materials
An ordinary eating apple should be used or tins of Heinz apple sauce.
Plastic measuring cylinders and filter funnels are best for this activity.
Termamyl/Pectinex/Celluclast can be obtained from NCBE, Science and Technology Centre,
Earley Gate, University of Reading, Whiteknights, READING, RG6 6BZ. Tel: 0118 987 3743 Fax:
0118 975 0140
Enzyme data sheets are automatically supplied with the order.
CARE!: Avoid direct skin and eye contact, wear eye protection and gloves. Enzyme powder can
cause allergies. Do not allow any spillages to dry up. Wipe up spillages immediately and rinse
cloth thoroughly with water.
Supply of materials
It is not appropriate to provide all equipment and materials in, for example, a tray system for each
student/group. Equipment and materials should be supplied in a way that students have to
identify and obtain resources. Normal laboratory apparatus should not be made available in kits
but should generally be available in the laboratory. Trays could be provided containing one type
of specialist equipment or materials.
Science & Plants for Schools: www.saps.org.uk
Fruit Juice Production: p. 3
This document may be photocopied for educational use in any institution taking part in the SAPS programme.
It may not be photocopied for any other purpose. Revised 2012.
Fruit Juice Production
Preparing for the activity
Read through the Student Activity Guide and consider the following questions.
Analysis of activity
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What is the aim of the activity?
What is being varied in the activity?
What measurements are you going to make?
Getting organised for experimental work
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Collect a data sheet for the enzymes Termamyl, Pectinex and Celluclast and familiarise
yourself with the action and commercial use of these enzymes.
In your group decide how the activity will be managed by allocating tasks to each
member. For Outcome 3 it is important that you play an active part in setting up the
experiment and in collecting results.
Recording of data
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Prepare tables to record your group results.
You should use a ruler, correct headings and appropriate units.
Evaluation
1.
2.
3.
4.
5.
6.
How effective were the tests which you used?
What were the limitations of the equipment?
What were the sources of error?
What possible improvements could be made to the experiment?
What factors other than the enzyme used might affect the volume of juice produced?
What factors, other than volume of juice produced, might be important in determining
effectiveness of the procedures?
7. What ideas do you have for further work?
8. What is the economic importance of the process which you are studying?
Science & Plants for Schools: www.saps.org.uk
Fruit Juice Production: p. 4
This document may be photocopied for educational use in any institution taking part in the SAPS programme.
It may not be photocopied for any other purpose. Revised 2012.
Fruit Juice Production
Student Activity Guide
Introduction
Fruit juice can be extracted from a wide variety of fruits. This can be done by simply squeezing
the fruits but it is more common to use enzymes to increase the volume of juice produced and the
speed of extraction. The enzymes which are used, both commercially and in this experiment,
break down the cell walls within the fruits and release liquids and sugars. Fruit cell walls are very
complex molecular structures and to get the maximum breakdown of the compounds found in
them, fruit juice companies use a variety of different treatments and enzymes to maximise the
yield of juice. Fruit is made up of cells linked by middle lamellae which contain insoluble protopectin. Pectinase breaks down the pectin chains and therefore reduces its binding action. The cell
walls are composed largely of cellulose and hemicellulose and cellulases weaken the cell walls
and make it easier to extract the juice. As the breakdown of the fruit cells continues, a variety of
polysaccharides are found within the juice extract. These can cause the juice to become cloudy
and reduce its market value. Pectinases and amylases can both break down these insoluble
compounds releasing soluble sugars which clarify the juice producing a clearer, sweeter product.
Different combinations of enzymes are used with each different type of fruit.
Enzymes are expensive products and clearly juice manufacturers would wish to minimise their
costs by using the enzymes at their optimum conditions and therefore maximising their
effectiveness and re-using the enzymes where possible.
You are going to set out an investigation to look at the effectiveness of these different enzymes.
Equipment and materials
Materials required by each student/group:
 one apple
 knife
 Termamyl/Pectinex/Celluclast
 four 10 cm3 measuring cylinder
 filter paper
 4 small filter funnels
 timer
 four 100 cm3 beakers
 four weigh boats
 stirring rod
 5 cuvettes (or test tubes)
Materials to be shared
 balance
 colorimeter
Science & Plants for Schools: www.saps.org.uk
Fruit Juice Production: p. 5
This document may be photocopied for educational use in any institution taking part in the SAPS programme.
It may not be photocopied for any other purpose. Revised 2012.
Instructions
1. Cut up the apple into small pieces.
2. Divide up the chopped apple into roughly 4 equal parts and use the balance to weigh
each part to equalise accurately and record the weight.
3. Place each quarter into a beaker and add 2 cm 3 of pectinase to one beaker, 2 cm 3 of
cellulase to another, 2 cm 3 of amylase to a third and 2 cm 3 of distilled water to the last
beaker.
CARE!: Avoid direct skin and eye contact, wear eye protection and gloves. Enzyme powder can
cause allergies. Do not allow any spillages to dry up. Wipe up spillages immediately and rinse
cloth thoroughly with water.
4.
5.
6.
7.
8.
9.
Stir each beaker and leave for 5 minutes.
Place beaker contents into a funnel and filter the apple juice into the measuring cylinders.
Record the volume of juice every 2 minutes and the final volume of juice.
Take a sample of each juice and measure its clarity by using a colorimeter.
Weigh the waste apple from each extraction.
Record your results in a table with suitable headings.
SUPPLEMENTARY STUDENT INFORMATION
Re-using enzymes
Introduction
You are going to try immobilising the cellulase or the pectinase enzymes, or a combination of the
two, to see if you can extract the enzymes from the waste apple and re-use them.
You will probably already be familiar with the technique of enzyme immobilisation. If you cannot
remember how to immobilise enzymes collect an Enzyme Immobilisation Instruction Card.
Ideally you should work in a group of about 4 people and share out the 4 tasks shown in the
following table.
Additional materials required
 1.5% calcium chloride
 4% sodium alginate
 distilled water
 250 cm3 beaker
 10 cm3 syringe
 20 cm3 syringe
 tea strainer
 iron filings
 bar magnet in a plastic bag
Enzyme/s
 Immobilise the enzyme/s
 Use the immobilised beads to extract juice and then remove the beads from the waste
fruit and re-use them
 Compare your results with the results from the initial experiment.
Science & Plants for Schools: www.saps.org.uk
Fruit Juice Production: p. 6
This document may be photocopied for educational use in any institution taking part in the SAPS programme.
It may not be photocopied for any other purpose. Revised 2012.
Enzyme/s plus air bubbles
 Immobilise the enzyme/s using a 20 cm 3 syringe adding air bubbles to the
enzyme/alginate mixture by passing it in and out of the syringe before placing into
calcium chloride
 Use the immobilised beads to extract the juice from the fruit
 Place the waste apple into a beaker of water and see if the beads float and are easier to
remove from the waste fruit
 Reuse the beads
 Compare your results with the results from the initial experiment.
Enzyme/s and iron filings
 Add 3 g of iron filings to the enzyme/s
 Immobilise the enzyme/s and iron filings
 Use the immobilised beads to extract the juice from the fruit
 Place the waste apple into a beaker of water and use a magnet to try to remove the
beads from the apple
 Reuse the beads
 Compare your results with the results from the initial experiment.
Enzyme/s plus air bubbles and iron fillings
 Add 3 g of iron filings to the enzyme/s
 Immobilise the enzyme/s and iron filings using a 20 cm 3 syringe adding air bubbles to the
enzyme/alginate mixture by passing it in and out of the syringe before placing into
calcium chloride
 Use the immobilised beads to extract the juice from the fruit
 Place the waste apple into a beaker of water and use a magnet to try to remove the
beads from the apple
 Reuse the beads
 Compare your results with the results from the initial experiment.
Now work as a team to discuss the results of the 4 different experiments and further work which
could be done to find out more about the action of these enzymes.
Science & Plants for Schools: www.saps.org.uk
Fruit Juice Production: p. 7
This document may be photocopied for educational use in any institution taking part in the SAPS programme.
It may not be photocopied for any other purpose. Revised 2012.
ENZYME IMMOBILISATION INSTRUCTION CARD
Introduction
Enzymes such as pectinase are often used in industrial processes. At the end of the process the
enzyme is often mixed up with the product and cannot be easily separated from it. Immobilisation
is a method which traps the enzymes in a bead which can be more easily separated from the
product. The method also means that it may be possible to reuse the enzyme once it has been
separated from the product. This method is often used in a continuous flow system.
Equipment and materials
Materials required by each student/group
 4% sodium alginate
 1.5% calcium chloride
 enzyme solution e.g. pectinase
 distilled water
 250 cm3 beaker
 10 cm3 syringe
 tea strainer
Instructions
The enzyme is mixed with sodium alginate and then fixed in a solution of calcium chloride.
1. Draw up 3 cm3 of enzyme solution into a syringe (if you are using a combination
treatment use 1.5 cm 3 of each enzyme).
2. Add 7 cm3 of sodium alginate to the same syringe and mix thoroughly in the syringe.
3. Gently drop by drop release the contents of the syringe to a beaker containing calcium
chloride solution and leave for 5 minutes to set.
4. Place the beads in a small tea strainer and thoroughly rinse them in distilled water.
5. Now place the beads in the apple puree, leave for 5 minutes and then place in the filter
funnel
Science & Plants for Schools: www.saps.org.uk
Fruit Juice Production: p. 8
This document may be photocopied for educational use in any institution taking part in the SAPS programme.
It may not be photocopied for any other purpose. Revised 2012.