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MIDLANDS STATE UNIVERSITY
FACULTY OF SCIENCE AND TECHNOLOGY
DEPARTMENT OF FOOD SCIENCE AND NUTRITION
FSN 107 & 204 FOOD CHEMISTRY LABORATORY
MANUAL
A. Physical properties of food
1. Specific Gravity and Refractive index
To illustrate the use of hydrometers and refractometers in determining the absolute or
relative concentration of a substance in a liquid.
Materials
Hydrometer (scale 1-2 or lactometer)
Refractometer
Skim milk
52% w/w sucrose solution
Saccharometer (Brix scale 30-60%)
Whole Milk
42% w/w sucrose solution
Procedure
a. Bring whole egg and skim milk to the temperature at which the hydrometer is
calibrated.Transfer samples to appropriate cylinders. Gently lower clean dry
hydrometer into each sample. Read scale when bubbles are gone and hydrometer
is at rest.
b. Use saccharometer to determine the percentage sucrose in a 42% sugar solution
and a 52% sugar solution.
c. Use a refractometer to determine the soluble solid content of the same sucrose
solutions as in step 2.
Questions
I. What is the effect of butterfat concentration on the specific gravity of milk?
II. What kind of dispersion is milk?
III. When would it be advisable to use a saccharometer Vs a refractometer.
Dispersion Of Matter
Solutions
To illustrate the effects of solutes on two colligative properties.
Materials
Sucrose
NaCl (reagent grade)
Procedure
1. Calibrate your themometer by determining the boiling point of deionised (D.I) water .
2. a) Prepare a solution with 30g NaCl and 300ml DI water in a 400ml beaker.
b) Place beaker on burner over an asbestos wire pad. Suspend thermometer in beaker.
Heat to a boling point.
c) Remove approx. 2ml into premarked tubes at each of the following temperatures,
101, 105, 1060C. Do not undershoot temperatures. Calculate the expected salt
concentrations ( g solute/ g solvent) use the formula
T = Kb or f x wt of solute
x 1000
x of particles per molecule in a solution
wt of solvent
M.W solvent
Convert the concentration (wt solute/ wt solvent) to % by substituting wt solute/ wt
solvent in the formula with
X
x 100 where x = wt solute /wt solvent.
X+1
Plot the boiling point vs soluble solids measured on the refractometer and the calculated
% solids extrapolated from the figure on the same graph.
d) Determine the % soluble solids with a refractometer.
3. a) Prepare a sucrose solution with 30g sucrose and 300g DI water in a 400ml beaker.
b) Place beaker on burner over an asbestos wire pad. Suspend thermometer, heat to a
rolling boil.
c)Remove approx. 2ml solution into premarked tubes at the following temperatures
100.5, 101.5, 102oC . Do not undershoot temperatures.
d) Determine % solids with refractometer.
4. a. Prepare a sucrose solution with 150g sucrose, and 150 ml water in a 400ml beaker.
b) Repeat the above
c)Repeat the above at temperatures 103, 106.5, 112, 114 and 1300C
d) Repeat above
5. Calculate the expected sucrose concentration from 3 and 4 above and graph the boiling
point vs the expected and observed sucrose concentration on the same graph as
described for salt in 2c.
6.a) Prepare 50g of 10% (W/W) solution of sucrose and a 10% of NaCl .
b) Determine the freezing point of the 10% solutions by placing a thermometer in a test
tube containing enough of the solution to cover the bulb. Place the test tube in a 4: 1
ice to NaCl mixture and observe the temperature at which the solution solidifies.
QUES
1. Why do the calculated and observed concentrations differ for sucrose but not for
NaCl?
2. Why is it not possible to reach a boiling point higher than 1070C for NaCl.
3. What is the boiling point for the thread? Soft ball stage? Hard ball stage? Hard crack
stage? How does the final boiling point relate to the sucrose concentration and firmness
of fudge or toffee.
4. Enumerate the characteristics of a true solution.
Emulsions
To demonstrate the effectiveness of various substances as emulsifying agents.
Materials
Sudan red dye
Liquid detergent
Vegetable oil
Slides
Electric blender
Lecithin
Egg yolk
Microscope
Sucrose ester (D. K ester F. 160)
Test tubes
Sucrose ester (D.K ester F10
Bile
Polyoxyethylene Sorbitan mono palmitate (Tween 40)
Liqiud For Dissoving
Emulsifier
Emulsifier
Oil
Water
O/W emulsions
Control – no emulsifier
Lecithin
0.5g
Egg yolk
0.5ml
Detergent
0.5ml
Bile
0.5g
Polyoxyethylene Sorbitan 0.5g
Sucrose ester (D. K ester F. 160) 0.5g
Sucrose ester (D.K ester F10) 0.5g
W/O emulsions
Control – no emulsifier
Lecithin
0.5g
Egg yolk
0.5ml
Vegetable oil
0.5ml
Bile
0.5g
Polyoxyethylene Sorbitan
0.5g
Sucrose ester (D. K ester F. 160) 0.5g
Sucrose ester (D.K ester F10) 0.5g
ml
10
10
10
10
10
10
10
10
ml
40
40
40
40
40
40
40
40
-------oil
water
water
water
oil
Water
oil
40
40
40
40
40
40
40
40
10
10
10
10
10
10
10
10
---Oil
Water
Water
Water
Oil
Water
oil
Procedure
1. Colour oil with sudan red dye.
2. Mix emulsifier with liquid specified for your variation.
3. Put oil and water into sample cup of mixer.
4. Mix for 30sec on speed 5.
5. Pour into beaker and observe.
6. Observe emulsions under microscope to determine whether you made a O/W or W/O
emulsion.
QUES.
1. How does the type of emulsifier affect emulsion formation and stability for W/O or
O/W emulsions.
2. What chemical properties should a good emulsifier have?
3. How can emulsifiers be classified be classified by their hydrophilic Lipophylic balance
(HLB) number?
4. What kind of emulsion is butter? Cream? Margarine? Salad dressing?
5. Why are detergents good cleaning agents?
6. What is the role of bile in digestion and absorption of fats?
Foaming properties of Proteins
To compare the foaming abilities of various protein. To investigate the mechanism of
foam formation and stability. To determine the effect of other chemical substsnces and
temperatures on protein foams.
Materials.
Egg albumin
Caseinate
Soya albumen (soya Lacto)
Cornstarch
Whey
Sugar
Vegetable oil
NaCl
100ml graduated cylinders
Mixer
Procedure
1. Prepare 100ml of the following solutions;
a. 0.5% caseinate
b. 0.5% whey
c. 0.5% egg albumin
d. 0.5% Soy albumen
e. 0.5% soy albumen + 0.5 % corn starch
f. 0.5% soy albumen + 0.5% sugar
g. 0.5% soy albumen + 0.5% veg. oil
h. 0.5% soy albumen + 0.5% NaCl
2. Place 50ml into sample cup of electric mixer, mix at speed 5 for 30sec and place
into 100ml graduated cylinder. Repeat for each.
3. Place containers at room temperatures and in a 400C water bath.
4. Measure foam volume at 0.5 and 30sec after mixing.
QUES
1. Graph the loss of foam volume over time for each temperature for each solution.
2. Compare the volume and stability of the foams as a function of
a) nature of protein present
b) Type of ingredient in the presence of the protein
c) Temperature
Carbohydrates
1. Qualitative Tests For Carbohydrates In Solution
Materials
Various carbohydrates in powder or granules
Test tubes
Molisch Test (for soluble carbohydrates)
a. Take 2cm3 solution in a test tube, add a few drops Molisch reagent (alcoholic
solution of alpha napthol). Mix well.
b. Add 1-2cm3 concentrated sulphuric acid carefully, pouring down the inside of the
tube to form a lower layer
c. Leave to stand then observe.
Iodine Test (for carbohydrate polymers)
Take 2cm3 of solution in a test tube, add a few drops iodine solution. Observe.
Fehlings Test (given by all reducing sugars)
To 1cm3 solution in a test tube, add 1cm3 of Fehlings solutions l and ll. Mix and heat
in a boiling water bath for 2 mins.
Benedict’s Test (given by all reducing sugars. It is similar to Fehlings but more
specific and less sensitive)
- To 1cm3 of solution in a test tube, add 2cm3 Benedict’s solution. Mix and heat for
2mins in a boiling water bath.
- Yellow or orange precipitate indicates reducing carbohydrates present.
QUES
1.What colour changes have you observed in the above reactions? What conclusions can
you make out of these observations.?
2. What is a reducing sugar? What is the significance of the reducing ability of a sugar?
2. Draw the structure of each sugar tested and indicate the reducing potential of each
sugar.
2. Quantitative Determination Of Concentration Of A Solution Known To Contain One
Reducing Sugar
Benedict’s Method
Reagents
Benedict’s quantitative reagent
0.3% glucose solution
Solution of named reducing sugar in unknown concentrations
Sodium carbonate- anhydrous
Method
1. Pipette 10cm3 benedict’s quantitative reagent into a small conical flask and add 2g
anhydrous Na2CO3.
2. Add a piece of porous pot and heat the solution gently to boil.
3. Keeping the mixture just boiling, run in the sugar solution in small amounts from
a burette, boiling well after each addition until the blue or green colour
disappears.
NB. The end point is reached when further addition of solution after boiling fails to cause
any change in colour. When nearing the end point, boil gently for 30sec. After the
addition of sugar to avoid overshooting. Read the burette to the nearest 0.05cm3 and do at
least two careful titrations to reach an agreement.
10cm3 Benedict’s solution should be equivalent to:
- 20.0mg glucose
- 21.2mg fructose
- 20.6mg sucrose after hydrolysis
- 27.1mg lactose
- 28.2mg maltose
3. Microscopic Examination Of Starch Grains
Obtain specimens of starch extracted from different sources together with one from an
unknown source eg. Laundry starch. Disperse minute quantities of each known starch in
turn onto a microscope slide in a drop of water. Cover with a cover slip and observe
under low and high power. Draw each indicating comparative sizes and characteristic
features which allow for identification.
Examine unknown starches and make suggestions regarding their identity.
4. Gelatinisation Of Starch
Materials
180ml water
15g maize meal
15g wheat flour
15g rice flour
15g sorghum meal
Procedure
1. Place of starch in a beaker. Add water slowly mixing well. No lumps.
2. Heat slowly stirring all the time.
3. Pour mixture into a large wetted crucible or petridish.
4. Repeat with the other starches.
5. Leave to cool for several hours or overnight. Compare the gels including any
evidence of synerisis and record.
6. Place a sheet of A4 size on the table. Place a sheet of carbon paper face down on
the paper. Place a piece of grease proof paper on top of the carbon paper. Fix
them to the table so that they will not move.
7. Turn out the first mould on the left hand side of the grease proof paper. Continue
across the paper with the other three moulds. Leave for 5mins to allow time to for
any of the moulds to spread.
8. Draw around the perimeter of each mould with a pencil. Press firmly so that the
outline will appear on the plain paper.
9. Carefully remove the moulds and grease proof paper. Write the name of the starch
on the impression of the mould. Compare the impressions for size of spread.
QUES
Which starch would be suitable for thickening white sauce? Steak pie? Sauce to be
frozen? Explain your answers.
LIPIDS
1. Detection Of Lipids In Foods
Grease Spot Test
Wrap a fragment in filter paper and leave in a warm place long enough to drive out any
water. Observe stain.
Sudan lll Test for traces of lipids (use with fragments, powders, crumbs, gratings)
Sudan lll is a red dye used in alcoholic solution from which it is readily absorbed by
lipids.
Method
a. Place a standard amount (level spatula) of fragmented food in a test tube with 5cm3 of
Sudan lll solution and shake thoroughly.
b. Filter collecting the filtrate in another test tube. Repeat with an equivalent amount of
lipid free material eg. Starch and compare the depth of colour of the filtrates. Record your
observations.
2. Demonstration of Acrolein Formation
Heat 1 or 2 drops of glycerol in a dry hard glass test tube with an equal volume of
potassium hydrogen sulphate crystals, a dehydrating agent. Note the characteristic smell.
Suggest where the smell could be coming from.
3. Visual evidence of invisible fat from foods
Materials
Potato chips
Foil
Groundnuts
Acetone
Paper towel
100ml petri dishes
Acetone
Hammer
Microwave
100 and 600ml beakers
Balance or scale
Safety goggles
Rubber/ Latex gloves
Graduated cylinders
Procedure
Potato chips
i)
Measure out 2g of potato chips and place on a paper towel.
ii)
Microwave on hogh for 25sec.
iii)
Fold the paper towel over the potato chips and crush the chips with a hammer.
iv)
Allow it to sit for 5mins. Open up the paper towel. Record your results.
Groundnuts
i)
Measure out 2g of groundnuts and put on a paper towel.
ii)
iii)
iv)
Microwave for 25sec on high.
Fold the paper towel over the nuts and crush the nuts with a hammer
Allow it to sit for 5mins. Open up the paper towel. Record your results
4. Quantitative Determination Of Lipids In Food
Extraction of fat
Materials
Potato chips
Foil
Groundnuts
Acetone
Paper towel
100ml petri dishes
Acetone
Hammer
Microwave
100 and 600ml beakers
Balance or scale
Face musks
Rubber/ Latex gloves
Graduated cylinders
Glass rod
Procedure
1. Weigh out 5g of potato chips, crush with your fingures.
2. Label the beakers that you are using, one for chips the other for nuts. Record the
weights of the labeled beakers.
3. Place the potato chips in the labeled beaker . Record the weight of beaker + chips.
4. Add 10ml of acetone to the crushed potato chips in the beaker.
5. Stir with a glass rod in a well ventilated area.
6. Carefully decant the acetone into the petri dish making sure the chips remain in the
beaker.
7. Add 10ml of acetone to the chips and repeat steps 5 & 6.
8. Allow the acetone in the petri dish to dry overnight in a hood or well ventilated place
to visualize the lipid that was extracted.
9. Allow the beaker with the chips to dry overnight. Weigh the beaker with the chips.
Ground nuts
1.Weigh out 5g of groundnuts. Crush the nuts between 2 pieces of foil with a hammer.
2. Repeat steps 2-9 in chips.
Tabulate your results under the following headings
Food, wt of beaker, wt of beaker with raw food, wt of raw food, wt of beaker with dried
food, wt loss from food, % lipid extraction ie( wt lost from food/wt of raw food) x100
Ques.
Which lipid contained saturated and unsaturated fatty acids in this experiment?
Soxhlet Extraction
Weigh out 5g of the food into a dry Soxhlet thimble. Suspend the thimble in a beaker and
dry it to constant weight in a steam oven. Place it in the Soxhlet condenser attached to a
previously weighed flask containing sufficient ether to fill the thimble and half fill the
flask. Attach the reflux condenser to the extraction tube. Heat the flask by means of a
water bath or electric hot plate so as to keep the ether gently boiling. The ether vapour
passes up the side tube of the extractor to the reflux condenser where it is condensed and
runs back onto the food in the thimble. When the thimble is practically full, the ether is
returned to the flask by an automatic siphoning device, carrying with it some of the fat
from the food. In this way, although a comparatively small quantity of ether is used, th
food is being constantly extracted with fresh fat free solvent.
After siphoning over 24 times, stop the experiment just before the next lot of ether is on
the point of syphoning over. The Soxhlet flask will the contain all the fat in a substance
and little ether. The flask is then dried off in air and and weighed to constant weight. The
fat is then washed out of the Soxhlet flask by a fat solvent, and then flask dried and
weighed again. The percentage weight of fat in the food is then calculated.
Original weight of food sample = xg
Weight of flask + extracted liquid = yg
Weight of flask
=zg
Percent lipid in food
= y - z
x
x 100
PROTEINS
1.Thermal Decomposition Of Protein
Strongly heat some dry protein in an ignition tube held by tongs in a horizontal
Position. Note the typical smell and test the fumes emitted with:
a) Moist red litmus paper
b) Moist lead acetate paper
Note the colour changes and explain the reaction produced that colour.
What does the water vapour indicate? What causes the black residue?
2. Water Solubility of Protein
Materials ;- A variety of powdered or fragmented protein.
Add a small quantity of protein in powdered or fragmented form to half a test tube of
water. Shake thoroughly. Foaming is a sign of solubility, even if slight. If insoluble in
cold water, try heating.
a.) Classify the samples as
l. Cold water soluble,
ll. Hot water soluble and
lll. Insoluble
QUES. What types of dispersions are formed?
Use the solutions for the next experiment. Tabulate your results for each test.
3.Qaulitative Tests For Protein
Biuret Test (for compounds with two or more peptide linkages)
Mix 5cm3 2M Sodium Hydroxide and three drops 1% copper sulphate. Reserve some of
this mixture for colour comparisons. Add 1cm3 Biuret mixture to an equal volume of
solution/ suspension under test and leave to stand.
Millon’s Test ( for compounds containing the hydroxy–phenyl group, C6H4OH)
N.B Millon’s reagent (Mercury dissolved in nitric acid) is poisonous use minimum
quantities.
Add 2 drops Millon’s reagent to about 2cm3 of solution or suspension under test and heat
in a water bath. Observe results.
Xanthoproteic Test (For compounds containing the phenyl group, C6H5)
Add about 2 drops concentrated nitric acid to about 2cm3 of solution or suspension under
test and heat in a water bath. Cool and neutralize by adding an ample amount of
ammonium hydroxide solution.
Sakaguchi Test (for all known proteins)
To 3cm3 of solution/suspension under test, add 1cm3 2M sodium hydroxide, 2 drops 1%
alcoholic alpha naphthol and 3 drops 10% sodium hypochloride. Mix well and allow to
stand. Repeat on a blank, ie. a test tube containing water instead of test
solution/suspension.
Sulphide Test (For sulphur containing amino acids)
Add 2cm3 of dilute sodium hydroxide to to 1cm3 of solution or suspension under test.
Boil thoroughly in a water bath noting dissolution of insoluble protein. Either add a few
drops of lead acetate solution or dip in lead acetate test paper.
QUES
a) What are the specific reactions involved in producing the colour in each of the tests.
b) What difficulties might be encountered in the amount of protein using the same colour
reaction.
3.Denaturation of Protein
Experiment 1
Materials
Egg white
0.1M Sodium chloride solution
0.1M Calcium chloride solution
0.1M ferric chloride solution
0.1M sucrose solution
1.0M sucrose solution
0.01M Hydrochloric acid solution
0.1M Hydrochloric acid solution
Distilled water
Procedure
Dilute 1 egg white (slightly beaten) with 3 volumes distilled water, stir slowly but
thoroughly and filter. To each test tube add 10ml albumin solution and 5ml of each of the
solutions listed above, including distilled water. Record the pH of the solution containing
distilled water and 0.01M and 0.1M hydrochloric acid. Place all in a beaker of water, heat
slowly and note the temperature at which opalescence develops.
Experiment 2
Materials
2% egg albumin solution
4cm3 industrial spirit (alcohol)
2cm3 mercuric chloride solution (poisonous)
2cm3 tannin acid solution
NB. If instead, egg white is used, (albumen) make up a 10% solution and filter off
insoluble globulins.
Procedure
Place approx. 3cm3 of albumin solution in each of the three test tubes.
In the forth test tube, heat gently at first and then bring to the boil.
Record results describing any coagulum that forms
While these apply in particular to egg albumin, the effects of organic solvents, salts of
heavy metals, alkanoids, eg. Tannin acid and heat are similar for most soluble proteins.
Using this information, explain the following.
a. pickling of anatomical specimens in alcohol
b. swallowing egg white as an antedote for lead poisoning
c. applying tannic acid ointment to burns and why this is no longer advocated.
d. Destroying enzymes by heat.
Maillard Reaction
To evaluate the aroma and colour of heated amino acid – glucose solutions.
Materials
Glucose
Methionine
Aspartic acid
Leucine
Lysine
Proline
Phenylalanine
Arginine
Valine
Procedure
1. To 50g of glucose, add 50mg of an amino acid in a test tube, add 0.5ml distilled
water. Mix thoroughly.
2. Smell each mixture and record any sensations.
3. Cover each test tube with heavy aluminium foil, heat solutions in a water bath at
1000C for 45mins. Cool to about 250C in a water bath. Record the odour sensations
for each solution eg. choc-like, potato-like.
4. Record the colour as; 0=none, 1=light yellow, 2=deep yellow, 3=brown.
QUES
What factors influence the degree of maillard browning?