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Unit: Control and Regulation (H): Genetic control
Title: The effects of lactase (ß galactosidase) enzyme on lactose in milk
Background Information
The enzyme lactase (ß galactosidase) is widely used in the dairy industry to convert the naturally
occurring disaccharide sugar lactose into glucose and galactose. This conversion takes place for a
variety of reasons:
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Many adults, particularly in the developing world, are unable to digest lactose.
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When making yoghurt or cottage-cheese lactase-treated milk reduces the time required to reach
the desired low pH and in the case of yoghurt produces a sweeter product which has a longer
shelf life. (in the case of fruit yoghurt this reduces the amount of sugar which has to be added to
the product)
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In the production of condensed milk lactase conversion is required to avoid the problems of
lactose crystallisation.
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In ice-cream production lactose conversion prevents lactose crystallisation which makes icecream gritty. The ice-cream product of lactose treated milk will therefore be more digestible,
creamier and more naturally sweet.
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Whey is a waste product from the cheese industry – if whey is treated with the lactase enzyme
the product can be purified and concentrated and used as a sweetener in various food products
Further information about these processes can be found in the BMB fact-files and the data sheet for
lactozym.
In this practical you are going to convert milk with lactase and test the glucose content of converted
milk you will then freeze converted and non-converted milk samples in freezer compartment of a
fridge and compare the products.
If possible work in a group and give each person in the group a different type of milk for example:
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Full fat milk
Semi-skimmed milk
Skimmed milk
Unit: Control and Regulation (H): Genetic control
Title: The effects of lactase (ß galactosidase) enzyme on lactose in milk
Equipment and materials
Materials required by each student/group:
2 cm3 of lactase enzyme
50 cm3 calcium chloride
8 cm3 of sodium alginate
25 cm3 of milk
2 x 10 cm3 syringe
1 x 250 cm3 beaker
2 x 100 cm3 beaker
circle of gauze
stirring rod
tea strainer
3 way tap
2 diastix
paper towel
Instructions
Step 1 Immobilising the lactose enzyme (ß galactosidase)
1. Use a 10 cm3 syringe to draw up 2 cm3 of lactase
2. Collect a piece of paper towel and then add 8 cm3 of sodium alginate to the same syringe
(use the paper towel to catch any alginate drips)
3. Rock the syringe until the two liquids mix completely (this may take about 5 minutes)
4. Collect a 250 cm3 beaker and add 100 cm3 of calcium chloride
5. Add the alginate/enzyme mixture to the calcium chloride one drop at a time.
6. Leave the beads in the calcium chloride for 3 minutes to allow them to set.
7. Filter the immobilised enzyme beads from the calcium chloride solution and then rinse the
beads in distilled water
Step 2 Setting up a Continuous Flow System
1. Collect a 3 way tap and check that you know how to use it. Practise with water.
2. Place a piece of nylon gauze at the base of a 10 cm3 syringe and attach a 3 way tap to the
end of the syringe.
3. Add the beads to the syringe.
4. Collect 25 cm3 of milk in a beaker.
5. Test the glucose content of the milk using diastix.
6. Slowly add the milk to the syringe and alter the 3 way tap to ensurethat the milk passes
through at a slow but steady rate. Collect the treated milk in a small beaker
7. Test the glucose content of the treated milk
Unit: Control and Regulation (H): Genetic control
Title: The effects of lactase (ß galactosidase) enzyme on lactose in milk
8. Record your result
9. Collect a freezer ice-cube tray and add 10 cm3 of the treated and non-treated milks to the ice
indents in the tray. Make sure that you label the ice tray carefully and then place in the
freezer for at least 24 hours.
10. Remove the ice tray from the freezer and note the appearance of each cube.
11. Place each block of iced milk on a grid fixed to a filter funnel and measure the drip rate from
each sample.
12. Record your results.
In addition a digital thermometer probe can be placed in the milk samples before freezing allowing the
temperature of the sample to be recorded every 30 seconds as it melts.
Results and Conclusions
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Present your results in a summary table
Draw a graph to show the melt rate of the iced milk samples
Relate your results back to the aim of the experiment.
Evaluating your experiment
In your evaluation of the experiment you should discuss:
1.
2.
3.
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6.
the effectiveness of the tests which you used
the limitations of the equipment
sources of error
possible improvements
ideas for further work
the economic importance of the process which you are studying.
Unit: Control and Regulation (H): Genetic control
Title: The effects of lactase (ß galactosidase) enzyme on lactose in milk
Questions
1. What does the enzyme ß galactosidase convert lactose into?
2. What effect does lactose conversion have on yoghurt and cottage cheese making?
3. What advantages does lactose conversion have for ice-cream manufacturers?
4. Describe the effect of temperature on conversion of lactase (mention optimum
temperature, time and pH)
5. Use table 1. Estimated Lactozym dosages, to list the factors which would be used
to determine the optimum dosage and indicate, with reasons, what dosage you would
recommend.
6. Name the organism used to produce lactozym.
Unit: Control and Regulation (H): Genetic control
Title: The effects of lactase (ß galactosidase) enzyme on lactose in milk
Ideas for Projects
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End point inhibition – when lactose is converted into glucose and galactose within a cell the
glucose and galactose will be utilised as respiratory substrates. Within this experiment if they are
allowed to build up within the reaction then they inhibit the enzyme reaction. When trying to reuse the immobilised ß galactosidase beads they will not convert the lactose if they have had
prolonged exposure to glucose, similarly if the flow rate of the reaction is too slow glucose and
galactose will build up and inhibit the ß galactosidase enzyme. Students can investigate this
reaction and relate it to the underpinning biochemistry of catabolite repression and the reactions
involved with Diauxic growth.
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Specific Heat Capacity – treating the milk with lactase lowers the freezing point and this can be
illustrated if a thermometer is placed within the ice block as suggested in the experiment. When
the frozen samples are removed from the freezer and melted at room temperature both the
texture of each frozen sample and the drip rate from each sample can be measured as it melts.
The thermometer allows the temperature of the sample to be recorded every 30 seconds as it
melts. The melting rates of the treated and non-treated milks can be measured and the graphs
drawn show evidence of alteration of the specific heat capacity