Biochemistry Lab Guide OBE

Name:______________________________________________________ Date Performed:_______________
EXPERIMENT NO. 1
PHYSICAL CHEMISTRY
Learning Outcomes
At the end of the experiment the student must be able to:
1. perform the procedures of titration and pH determination.

2. compute for the pH of a given sample.
3. differentiate the properties of emulsoids and suspensoids.
4. investigate the physical properties of a substance by determining their diffusability,
osmosis and adsorption property, surface tension, etc.
I. Discussion:
A. Titration
Titration is a process of ascertaining the quality of a constituent present in a compound by
using a standard solution.
Two Types of Titration
A. Direct Method – this is a method in which the standard acid solution is added to an
alkali solution until the end is reacted.
B. Residual or Indirect Method – a solution in which a definite concentration of the

solute is dissolved in the solution.
Standard Solution – A solution in which a definite concentration is dissolved in the solution.
II. PROCEDURE:
a. Measure 10 ml of sodium hydroxide of unknown strength and place in a 250 ml
Erlenmeyer flask.
b. Add 2 drops of phenolphthalein
c. Mix by shaking
d. Titrate with 0.1N HCl until the solution becomes colorless.
e. Take note of the volume of the standard acid used and determine the normality of the
solution using the equation:
N1 V 1 = N 2 V 2 or Na V a = N b V b
Write your results and observations on a space below:
1
B. DETERMINATION OF pH
The pH of a solution is compound logarithm of the reciprocal of the hydrogen ion concentration
expressed as:
pH = Log 1/H+
Pure water is slightly ionized and at 25°c contains 0.00000001 or 10 -7 per liter. The pH of water
is therefore:
pH = Log 1/10-7
pH = -Log 10-7
pH = 7
If the hydrogen ion concentration (H +) of a solution is greater than in pure water, the pH is a
smaller number than 7, such solution is acidic. Conversely, if the (H +) is less than in pure water
the solution is basic. The range of pH values fall between 0 – 14
pH is detected and measured by using a pH meter, but the most common detectors are the color
changes of acid base indicators. An acid base indicator is a weak organic acid. The ionized and
unionized forms of the indicators have different colors. The colors of indicators solution is
therefore a measure of pH.
pH Determination
Procedure:
1. Prepare 1 ml in four clean test tubes of the following samples below and label:
A. 0.1N HCl C. Fresh Milk E. Urine

B. 0.1N NaOH D. soft drink (soda) F. Orange juice
2. Determine the approximate pH of the sample by adding 1 drop in each test tube using
the following indicators:
A. Phenolphthalein C. Phenol Red

B. Congo Red D. Methyl Orange
2
Observed the color product in each test tube. Tabulate your results by recording the color of each
Indicator.
SAMPLES INDICATORS
Phenolphthalein Congo Red Phenol Red Methyl Orange
1. 0.1N HCl
2. 0.1N NaOH
3. Fresh Milk
4. Urine
5. Orange
Juice
6. Soft drink
Determine the accurate pH of the samples using the pH meter by following these steps.
Operatory Procedure:
1. Set the temperature control knob at about 25°C.
2. Dip the electrode of the pH meter into the standard buffer solution.
3. Turn the knob to position “pH” and check if the pointer gives the correct reading of pH of the
buffer solution.
4. If the reading is not correct, turn the standardization knob until the accurate reading is
obtained.
5. Remove the buffer solution and rinse the electrode with distilled water. Wipe the electrode with
tissue paper.
6. Dip the electrode into the first solution and turn the knob to the position “pH”. Get the pH
reading on the scale of the instrument and record it.
7. Turn back the knob to off position and removed the electrode from the test solution.
8. Rinse the electrode with distilled water and wipe it dry.
9. Repeat the test with the other samples.
Results and Observation
Samples pH meter
1. 0.1N HCl
2. 0.1N NaOH
3. Fresh Milk
4. Urine
5.Orange Juice
6. Soft drink
Activity Sheet Number 1
3
Name:_______________________________________________________Section__________________ Date _______________
Instruction: Based on the results of your performed experiments, discuss among your group mates
all of your observations and answer the following questions:
1. What is the biochemical relevance of pH?
2. How does the pH of your assigned samples as obtained from the pH values (indicators)
compared with the pH obtained from the pH meter?
3. What indicator/s is/are used when titrating a weak acid against a strong base? A weak base
against the strong acid?
4
C. BUFFERS
A buffer is one that resists a change in pH when a small amount of acid or base is added. A buffer
solution contains a weak Bronsted acid and its conjugate base.
If a small amount of strong acid is added to the buffer solution, some of the anions from the buffers
salt will combine with the added protons to form more dissociated acid.
H+ (from outside source) + A- (buffer salt) HA
Similarly, if the small amount of the base is added to the buffer solution, some of the buffer acid
reacts with the pH- to form more buffer salt.
PROCEDURE:
1. Prepare duplicate samples of the following:

a. 10ml freshly boiled and cooled distilled water
b. 10ml 0.1N sodium acetate solution
c. 10ml egg albumin solution
2. Determine the original pH of each sample.
3. To the first set add 0.1ml of 0.1N NaOH to each sample. Determine the pH using the pH meter.
Record your result.
4. To the second set add 0.1ml of 0.1N HCl to each sample. Determine the pH using the pH meter.
Record your results.
Results and Observations:
Buffer pH meter reading 0.1ml of 0.1N NaOH 0.1ml of 0.1N HCl

(original) (read with ppt) (read with ppt)
1. Distilled water
2. Cooled distilled H2O
3. Egg albumin
4. Sodium acetate
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Instruction: Based on the results of your performed experiments, discuss among your group mates
1. Which sample exhibited buffer actions? Explain
2. How would you correlate the buffer systems involved in whole blood and plasma?
3. What are the two factors that determine the effectiveness or capacity of buffer solutions?
4. Give examples of buffer solutions.
D. COLLOIDS
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The word colloids include any dispersion of very small particles that are larger than simple
molecules. Colloidal size lies between those of true solution which are homogenous and coarse
mixtures which separate on standing. This includes particles with dimensions in teh range of 10 oA
to 10,000 oA or .001 micron to 1 micron.
A natural colloidal substance which composes the protoplasm of the living cells is called biocolloids.
Biocolloids include nearly all of the energy foods such as starches, proteins, fats and biocatalysts
such as enzymes, hormones and vitamins.
Preparation of an Emulsoid: (Gelatin)
1. Prepare 5% solution of gelatin with the aid of heat.
2. Cool under the tap water and observe. What is produced?
3. Heat again and observe the result.
Write your results and observations below:
Preparation of Suspensoids: (FeCl3 solution saturated)
1. Place 100ml of boiling water in a beaker.
2. Add 2ml of saturated ferric chloride solution.
3. Observed the product formed.
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Precipitation with Electrolytes:
1. Place 5ml 5% gelatin solution in a test tube.
2. Add saturated solution of ammonium sulfate drop by drop counting the number of drops until a
permanent precipitate is formed.
3. Repeat the procedure using colloidal ferric.
REVERSABILITY
1. Decant the supernatant liquid from each of the two test colloids prepared in letter B procedure.
2. Add an excess of water and note whether the two colloids are reversible.
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Instruction: Based on the results of your performed experiments, discuss among your groupmates
1. What is meant by a protective colloid?
2. On a tabular form compare Emulsoid from Suspensoid.
3. Discuss by differentiating sols and gels.
4. Explain, why is Suspensoid is readily precipitated by salts?
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E. DIALYSIS
Dialysis is a common biochemical method of separation and purification by selective

passage of ions and small molecules through a semi-permeable membrane but will not
allow colloids to pass through.
The rate of dialysis depends on many factors: the area of the dialyzer, the size of the pores,
the temperature, the electric charges and the relative concentration of solution on the two
sides of the membrane.
Procedures:
1. Pour some colloid on solution (5% in equal parts of ether and alcohol) into a clean and dry
hard test tube taking care not to form air bubbles.
2. Wet completely the inner side of the test tube and pour the excess of colloidion back to the
bottle.
3. Allow to drain and rotate the tube slowly in horizontal position until the ether has partly
separated.
4. Invert the tube and allow to dry for 15 minutes.
5. When dry, fill with water and stand for 10 minutes.
6. Loosen the film around the tip of the tube and allow cold water to run between membranes.
If necessary, loosen the membrane from the side of the tube with a smooth glass stirring
rod.
7. Rinse the colloidion bag thus prepared with distilled water several times.
8. Take 3ml of distilled water and treat it with 2 drops of silver nitrate t.s., If negative use
about 2/3 of a beaker with distilled water
9. By means of stirring, suspend the dialyzing bag containing equal volumes of 1% starch and
1% sodium chloride solutions.
10. Let this stand for 30 minutes.
11. Test the dialyzing by using silver nitrate t.s. and iodine solution.
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1. Which of the substances inside the bag diffuses out of the colloidion mixtures? Explain why?
2. What other membranes beside colloidion might be used in dialysis?
3. What are the factors that affect the rate of dialysis? Explain why?
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F. DIFFUSION THROUG GELS
All solutes tend to diffuse through solutions until the composition is homogenous
throughout. Small molecules and ions move with sufficient velocity to distribute themselves
throughout the solvent rapidly. On the other hand, macromolecules move slowly because of
their latge molecule size.
Procedure:
1. Put 2ml of 10% copper sulfate solution in a test tube. Incline the test tube and carefully add 3ml
of distilled water. Take note the time for the entire body of fluid to have the same color.
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Name:_______________________________________________________ Date Performed:_______________
EXPERIMENT NO. 2
ELEMENTS FOUND IN PHYSIOLOGICAL COMPOUNDS
Learning outcomes
1. identify the elements, present in physiological compounds.

2. perform the different test procedures in the determination of physiologic
elements
3. differentiate their properties and components.
Test for Carbon

Procedure:
1. Place small amount of sugar in a test tube.

2. Heat gently over the Bunsen flame.
3. Note the change undergone by sugar.
Test for Hydrogen and Oxygen
Procedure:
1. Repeat the procedure above.

2. Observe the upper cooler portion of the test tube.
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1. What are the elements contained in the sugar molecules?
2. Give 5 substances or compounds containing elements of C, H and O
3. Discuss the role of these elements in physiologic compounds.
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Test for Nitrogen
Procedure A
1. Burn hair, feather or horn and note the odor produced.
2. Describe the odor produced and account for it.
Procedure B
1. Place about 0.5 gm of egg albumin in a porcelain dish.
2. Place about 1 gm of soda lime. Mix and grind the mixture.
3. Place the mixture in dry test tube and heat gently over a Bunsen burner lame. Note the odor
of the gas evolved.
4. Test the gas evolved by:
a. Exposing previously moistened red litmus paper over the fumes. Take note of the
changes in color.
b. Observed the odor of the fumes.
c. Exposing a glass rod previously dipped in concentrated hydrochloric acid.
Test for Sulfur
Procedure:
1. Place 10ml of sodium hydroxide t.s. in a test tube.

2. Add 2 drops of neutral lead acetate solution.
3. Add a small amount of egg albumin (0.3 gm) into the mixture.
4. Heat the mixture and take note of the changes in color. Observe.
15
Test for Phosphorous
Procedure:
1. Place in a porcelain crucible about 0.5 gm of casein
2. Add equal amount each of potassium hydroxide and potassium nitrate.
3. Heat the mixture strongly. Cool, dissolve in water and filter.
4. Place 2 ml of the filtrate in a test tube.
5. Acidify by adding 5-10 drops of concentrated nitric acid.
6. Add about 15 drops of ammonium molybdate.
7. Note the change produced.
OSMOSIS
Osmosis is the passage or movement of the solvent molecules from more dilute solution to
the concentrated when these are separated by a semi-permeable membrane. In the
following procedure, the stroma of the erythrocytes acts as a semi-permeable membrane.
Osmotic Pressure of the Red Blood Cells
1. Prepare 3 clean and dry test tube labeled 1, 2 and 3 each containing 5ml each
of the following solutions 0.3, 0.9 and 5% NaCl.
2. To each test tube, add 2 drops of blood
3. Allow the test tube to stand for 3-5 minutes.
4. Examine each suspension under the microscope
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1. What are the factors that play the dominant role in this experiment?
2. In a tabular form differentiate Osmosis from Diffusion
3. What is an NSS?
4. What is the effect of the following solution on the red blood cells; Draw the cells found in each
tube.
a. Hypotonic;
b. Hypertonic;
c. Isotonic salt?
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ADSORPTION
Adsorption occurs when a layer of ions, molecules or aggregates of molecules are condensed on the
surface of the solid. This arises from the valence forces or other attractive forces of the atoms or
molecules in the outermost layer of the solid. The amount of adsorption depends upon the content
of the surface and the surface adsorbed.
Procedure:
1. Place 10ml of 0.1% aqueous solution of methylene blue in a flask or a beaker.
2. Add about 2 rams of animal charcoal.
3. Mix by shaking vigorously
4. Filter and note the color of the filter and set it aside. Then allow the residue left in the filter
paper to dry.
5. Pour 10ml of the 95% ethyl alcohol over the residue in the filter paper and collect the filtrate in
another clean test tube.
6. A very faint blue color of the second filtrate should be considered.
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SURFACE TENSION
The surface of the liquid possesses special properties which are due to the unbalanced forces of
molecular attraction at the surface. The molecules at the surface are pulled inward by then other
molecules of the liquid and the liquid tends to adjust itself to give the minimum surface area.
When the two partially miscible liquids are placed in contact, each dissolve to a certain extent in the
other causing marked changes in surface tension. The addition of the third component may lower
the surface tension considerably. Soaps, detergents and salts of bile acids are specially considered.
Procedure:
1. Prepared 2 test tubes labeled no. 1 and no. 2. To each test tube, place 1ml distilled water and
1ml CHCl3. Add 1ml of soap solution to test tube no. 2 Shake both test tube then let stand for a
few minutes (Note the time it takes for the drop to coalesce in each test tube)
2. Put 5ml of bile solution in a dry evaporating dish, sprinkle a p[inch of sulfur powder on the
surface of the solution. Observe. Repeat the test tube using distilled water.
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1. Explain the behavior of the methylene blue toward animal charcoal.
2. What is the role of bile salts during fat digestion?
3. Explain how soap lowers the surface tension of fats.
4. Define the following terms:

a. Adsorption
b. Absorbent
c. Elution
d. Adsorbate
e. Eluate
5. What is surface tension?
6. What is its importance?
DONNAN’S EQUILIBRIUM
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The presence of a non-diffusable ion in one side of a semi-permeable membrane wills at
equilibrium, produce an unequal distribution of the diffusible ion’s in both sides of the
membrane.
Procedure:
1. Place 25cc of 2% gelatin into a 50cc beaker.

2. In another 50cc beaker place 25cc water.
3. Add 0.5cc of phenolphthalein to each.
4. To the water add 0.1N NaOH drop by drop until a red color develops. (about pH 10.0)
5. Add 0.1N NaOH to the gelatin solution drop by drop until a similar shade of color is
obtained. The two solutions have row approximately the same hydroxyl ion concentration.
6. Transfer the gelatin solution to a dialyzing bag tie one end with a string and suspend it in a
beaker containing the water.
7. Let it stand for 2 hours and observe the difference in color that develops in both solutions
by taking aliquot portions from each and placing these in two clean test tubes for
comparison.
8. Repeat the procedure using 0.5cc of congo red indicator instead of phenolphthalein.
9. To the water add 0.1N HCl instead of 0.1N NaOH drop by drop until blue color develops.
(about pH of 3.0)
10. Add 0.1N HCl to the gelatin solution drop by drop until a similar shade of color is obtained.
11. Transfer the gelatin solution in a dialyzing bag and proceed as before.
12. It is suggested that the one half of the group use phenolphthalein as indicator and the other
half use congo red.
Reagents:
Albumin solution
Dilute one fresh egg white 5 times with distilled water
Hydrochloric Acid 0.1N
Add 0.83ml of concentrated HCl to 50ml distilled water and dilute to 100ml
Sodium Acetate 0.1N
Dissolve 0.82 g NaC2H3O2 in distilled water to make 100ml solution
Sodium Hydroxide 0.1N
Dissolve 0.10 g NaOH in distilled water to make 100ml solution
Buffer Phosphates (sorenses)
a. M/15 Potassium Dihydrogen phosphates dissolve 9.00 grams of KH 2PO4 in distilled
water to make 1 liter of solution.
b. M/15 Disodium hydrogen phosphates dissolve 11.88 grams of Na 2HPO42H2O in distilled
water to make 1 liter solution
Bile Solution
Dilute bile from fresh milk fish with sufficient distilled water
Name:_______________________________________________________ Date Performed:_______________
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EXPERIMENT NO. 3
CHEMISTRY OF CARBOHYDRATES
CARBOHYDRATES
Learning outcomes
1. Discuss the properties of carbohydrates
2. Differentiate the types of carbohydrates based on their physical and chemical
characteristics
3. Perform the tests in classifying the different types of carbohydrates
Carbohydrates are organic compounds containing carbon, hydrogen and oxygen in which
the ration of hydrogen to oxygen is 2:1 the same molecule as a molecule of water. They are
polyhydric alcohols containing potential aldehyde or ketone groups. The reaction described
here are some of the chemical reactions attributed either to the carbonyl group or the
carbinol group.
- Physical Properties -
Take a small amount of the following sugars and explain its forms, color, odor and shapes.
SUGARS SYNONYMS FORM COLOR ODOR TASTE SHAPE

Glucose
Fructose
Lactose
Sucrose
Starch
Galactose
Maltose
Cellulose
Solubility
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Prepare seven (7) clean and dry test tubes labeled as to content. Observe and record the
results obtained.
SOLVENTS GLUCOSE FRUCTOSE SUCROSE LACTOSE STARCH

Water
105 NaCl
0.5% Na2CO3
Dilute HCl
Conc. HCl
Conc. NaOH
Ethyl Alcohol
Color Test for Carbohydrates
General Principle: Sugars upon addition of mineral acids such as HCl or H 2SO4 undergo
dehydration to form furfural or furfural derivatives which in turn condenses to form characteristic
colors.
A. Molisch Test or Alpha Napthol Test
Procedure:
1. To 5 ml of the test solution in a test tube, add 5 drops of Molisch reagent.
2. Mix thoroughly by inversion
3. Incline the tube and pour about 5 ml concentrated acid down the sides of the test tube forming
a layer of acid beneath the sugar solution.
4. A reddish-violet ring at the junction is a positive reaction.
B. Seliwanoff’s Test for Resorcinol-HCl test – this is a test for the ketone sugars.
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Procedure:
1. To 2.5 ml of Seliwanoff’s reagent in a test tube add 5 drops of the unknown sugar solution and
boil.
2. The appearance of a red color is a positive reaction.
3. Note the time of appearance of the red color.
4. Do the test also with sucrose, glucose and maltose.
To confirm whether the red color is due to precipitate formed with fructose, add 1 ml of methyl
alcohol. If the precipitate dissolves, then the result is definitely due to fructose.
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1. Is Seliwanoff’s test this test specific for fructose?
2. Why do all sugars give positive Molisch Test? Does the test confirm the presence of sugars?
Explain.
3. Is glucose normally positive with this test? Account for the result obtained with glucose.
4. How could this test be used to distinguish fructose from sucrose?
Mucic Acid Test – test for Galactose and lactose
25
Procedure:
1. Place about 50 mg of Galactose in a clean and dry test tube.
2. Add 1 ml of distilled water and 1 ml of concentrated nitric acid.
3. Heat on a boiling water bath for 1 ½ hours.
4. Allow it to stand overnight.
5. Observe the crystalline precipitate formed and draw the crystals seen under the
microscope.
Phenylhydrazine Test or Osazone Test
Procedure:
1. To 1 ml of 9% sugar solution, add 0.5 ml of phenylhydrazine mixture.
2. Shake well and heat on a boiling water for ½ hour.
3. Allow to cool slowly and examine the crystals under the microscope.
4. Perform the test with the following solutions: glucose, maltose, fructose and lactose.
Reduction Test for Carbohydrates
26
Principle Involved:
Sugars having a free or a potentially free aldehyde or ketone group are capable of reducing certain
oxidizing agents such as Cu++, Hg+, Bi+, Ag+ are oxidized to the corresponding sugar acids. The
metallic ions are reduced to a lower valence state.
The reagents used are solution of metallic salts either in acid or alkaline medium, certain organic
compounds containing alcoholic groups such as Rochelle salt and sodium citrate or glycerol maybe
added.
A. Trommer’s Test
Procedure:
1. To 1 ml of 5% glucose solution add 0.5 ml of NaOH and mix.
2. Add drop by drop dilute solution of CuSO 4 until there is a slight permanent precipitate of
Cu(OH)2.
Illustrate the chemical reaction Involved:

Repeat the test using water instead of sugar solution. What difference do you observe? Explain.
27
B. Fehling’s Test
Procedure:
1. Mix 1 ml of Fehling’s A and B in a test tube.
2. Add 5 ml of water and boil. If a precipitate forms, the solution must not be used.
3. To the warm Fehling’s solution, add the unknown solution drop by drop and heat the
mixture after each addition.
4. A positive reaction test is indicated by the production of a yellow to brick red
precipitate.
5. Do the test with at least 3 sugars and starch.
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1. Why is it important that the Fehling’s solution for use as reagent be freshly mixed?
2. Why are the 2 Fehling’s solution kept in separate container
3. Give the chemical composition of the two Fehling’s reagent and the role played by each.
4. Differentiate reducing from non-reducing sugars.
5. Give the use of the test solution used in carbohydrate identification.
C. Benedict’s Test
29
Procedure:
1. Place about 2.5 ml of Benedict’s reagent in the test tube.
2. Boil. The solution must remain clear. If precipitate is formed, the reagent must be
changed.
3. Add 1 drop of the sugar solution and boil over direct flame for 2 minutes or place in a
water bath for 5 minutes.
4. Reduction is manifested by the formation of colored precipitate like the one in Fehling’s
test.
5. Perform the test using the following sugar solution.
a. Glucose c, Lactose e. Starch

b. Fructose d. Sucrose
Show results and observations:

Sugar Solution Results and Observations
1. Glucose
2. Fructose
3. Sucrose
4. Lactose
5. Starch
6. Galactose

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1. Give the difference between Fehling’s test and Benedict’s test.
2. Which is more sensitive of these two reagents? Why?
3. Name the other sugar that showed a positive result and explain why?
4. Explain, what makes some sugar positive for Benedict’s tests?
D. Barfoed’s Test
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Procedure:
1. To about 3 ml of the Barfoed’s test reagent add 0.5 ml of the sugar solution in a test
tube.
2. Place on a warm water bath for 5 minutes.
3. Remove from the water and cool.
4. Observe at once and after 5 minutes of cooling.
5. Reduction is indication of a formation of colored precipitates similar to the one seen in
Fehling’s and Benedict’s test.
6. Perform this test using the sugars used in the Benedict’s test.
Result and Observation:
Sugar Solution Results

1. Glucose
2. Fructose
3. Sucrose
4. Lactose
5. Starch
6. Galactose

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1. How does the condition of Barfoed’s test different from the Benedict’s test?
2. Why is the element strictly followed in the performance of this test?
3. What is the use of the test?
4. Can the Barfoed’s test be used in place of the Benedict’s test for the detection of sugar in the
urine?
5. What precipitate results when reducing sugars are heated with Fehling’s, Benedict’s and
Barfoed’s reagents.
E. Nylander’s Test
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Procedure:
1. Place 1 ml of sugar solution in a clean test tube. Use at least 5 sugar solutions.
2. Add 0.5 ml of the Nylander’s solution.
3. Heat for 5 minutes in boiling water bath.
4. Note the change in color while boiling and after cooling for a few minutes.

34
1. Give the importance of the reagents and the role of each component.
2. Write the equations involved.
E. Picric Acid
35
Procedure:
1. Place 5 ml of the sugar solution to be tested in a test tube.
2. Add 2 ml of aqueous solution of picric acid.
3. Add 2 ml of 10% sodium hydroxide.
4. Heat in a boiling water bath.
5. Reduction is indicated by the formation of a mahogany red color.
D. Inversion of Sucrose
Procedure:
1. Take about 25 ml of sucrose solution in a beaker or flask; add about 1 ml of
concentrated sulfuric acid.
2. Boil for 15 minutes.
3. Cool and neutralized with 15% NaOH using litmus paper as indicator.
4. Take a portion and perform the phenylhydrazine test.
5. In another portion, perform the Benedict’s test.

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1. What is responsible for the end color produced?
2. Write the chemical reaction involved.
Fermentation Test
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Fermentation is the process of converting hexone sugars to alcohol and carbon dioxide
through the action of a mixture of enzymes called zymase which is present in yeast. Hexoses
are readily fermented while pentoses are not fermented by yeast. For dissacharides to be
fermented they must be hydrolyzed to their monossacharide component by the enzyme
found in yeast. The chemical step in the fermentation of lactose by yeast juice is very similar
to the aerobic breakdown in muscle extracts.
Procedure:
1. Mix 10 ml of sugar solution with an equal volume of yeast solution in a beaker and
transfer the mixture to fermentation tube or saccharometer.
2. See to it that the long arm is filled with the solution and no bubbles are present.
3. Set aside in warm place in about 12 hours. If the sugar is fermentable, carbon dioxide
gas will be collected in the upper portion of the tube.
4. At the end of the incubation period, add 1.2 ml of 10% NaOH solution in the long arm by
means of a pipette.
5. Fill the bulb portion with water, place the thumb tightly over the opening of the
apparatus and invert the saccharometer.
6. Filter some of the mixture.
7. To 5.5 ml of the filtrate, add several drops (enough to give a yellow color of the whole
mixture) of a strong solution of I and II.
8. Warm gently. Note the iodoform odor and examine the iodoform crystals under the
microscope. What does a positive test here indicate?
9. Draw the crystals.
10. Tabulate the result obtained in all sugars.

38
1. What is the gas collected?
2. What organic compound is produced by fermentation?
POLYSACCHARIDES
39
A. Starch
Preparation of Starch:
Prepare a raw potato, comminute and grind. Mix with a little of water. Strain through gauze
to remove coarse particles. Wash by repeated decantation. Drain the starch and dry in the
air. This starch maybe used for experiment.
Macroscopic Examination of Starch

Examine some potato starch grains as to form, color and taste.
Microscopic Examination of Starch

Suspend some granules of different samples of starch in a drop of water and examine under
the microscope. Draw them.
Draw your observation:
Iodine Test on Starch Paste
Procedure:
1. To 5 ml of dilute starch paste solution, add a drop of iodine solution. Observe the color
2. Heat the mixture and observe what happens.
3. Cool again and observe the changes.
Hydrolysis of Starch
40
Procedure:
1. Place 20 ml of starch paste solution in a small beaker. Add 5 ml concentrated HCl and
boil the mixture. At interval of 1 minute transfer a drop of a mixture in a watch glass
over a white paper and test with iodine solution. Continue the test until it gives a
negative color with iodine. Then test it with Benedict’s reagent. Outline the hydrolysis of
starch.

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1. In the iodine test, what difference was observed using plant and animal starch? Explain the
difference.
2. Account for the disappearance of the color upon heating the iodo-starch complex.
3. Name 2 chemical components of starch
B. Cellulose
42
Solubility of Cellulose
Procedure:
1. Take a few shreds of cotton and test the solubility in water, dilute and concentrated
acids and alkaline.
2. What can you conclude with regard to the solubility of cellulose?
Cross and Bevan’s Test (ZnCl2-HCl)

Procedure:
1. Place a few shreds of cotton in a test tube and add 1 ml of Cross and Bevan’s reagent.
2. Stir until the solution is affected.
3. Add an equal amount of 9% alcohol.
Formation of Amyloid
Formation of Amyloid
Procedure:
1. Add by little 2.5 ml of sulfuric acid to 1.5 ml of distilled water, cooling the solution under
the water after each addition.
2. Place about 0.5 of filter paper
3. Stir for 5 to 10 minutes until it is dissolved. Pour 0.5 ml of the solution to 2.5 ml of water
and note the production of flocculated precipitate, which is the Amyloid.
4. Add a drop of iodine to a small portion of the precipitate.
5. Pour the remaining portion in 7.5 ml of water and boil for 5 to 15 minutes.
6. Cool and neutralized with NaOH.
7. Test with Benedict’s and other reducing agents.
Glycogen
43
Procedure:
1. Cut the fresh crystals into small pieces.
2. Mix with sand and grind in mortar.
3. Place in a beaker and add 25 ml of water.
4. Heat for 20 minutes, adding water from time to time to maintain the volume.
5. Acidify with 5% acetic acid.
6. Boil for 1 minute, cool and filter.
7. Add four volumes of 95% alcohol.
8. Allow the precipitate to settle, pour off the supernatant fluid and filter.
9. Dissolve the precipitate in hot water to make a glycogen solution.
10. Test a small portion of the solution of glycogen with benedict’s reagent and iodine t.s.
Acidify the remaining solutions with HCl and boil for 10 minutes. Cool and neutralize
solutions with HCl and boil for 10 minutes. Cool and neutralize with 10% NaOH and test
with Benedict’s reagent.
Compare the results of your reduction test with that was obtained previous to hydrolysis.
C. Hemicellulose
Gum Arabic
Physical Appearance
Examine the sample as to form, color, odor, and taste.
Solubility of Gum Arabic

Test the solubility of Gum Arabic in
A. Cold water
B. Hot water
C. Alcohol
Hydrolysis of Gum Arabic

Procedure:
44
1. To 20 ml of gum Arabic solution, add 5 ml of concentrated HCl solution. Heat for 5
minutes, cool and neutralize with NaOH solution.
2. Test with Benedict’s reagent.
Galactam
Agar
Physical Properties of Agar

Examine the microscopic characteristic of agar.
Solubility Test for Agar

Test the solubility in
45
A. Cold water
B. Hot water
C. Alcohol
Observe the imbibing capacity of agar.

Pectin
Preparation of Pectin:
1. Take the inner white ring of orange and chop into fine pieces.
2. Place in a beaker and cover with distilled water.
3. Allow to stand until the next laboratory period.
4. Drain through cheese cloth and squeeze out of the liquid.
5. Boil the pulp with water for 2 hours maintaining the volume by adding more water.
6. After 6 hours reduce the volume to just sufficient amount to cover the pulp.
7. Filter through cheese cloth.
8. Cool the filtrate and add twice its volume of 95% alcohol.
9. Filter and dry the precipitated pectin.

46
1. What substance is responsible for the starch iodine complex color formation?
2. Give the color reaction produced when starch is subjected to Benedicts test and Iodine tests.
I. Discussion:
47
Chromatography of Sugars
A mixture of sugar in the solution can be separated by the use of paper chromatography if they
posses different partition coefficients between the mobile phase and the water saturated cellulose.
The solvent is stationary usually water bound to cellulose. The other usually organic, moves over
the filter paper by capillary actions. When the partition is in favor of the aqueous phase, the solute
tends to remain near the point of application. On the other hand, if the partition favors the organic
phase, the solute will move with solvent phase.
II. Procedure:
1. Cut a piece of whatman no. 1 filter paper 1.5 x 8.0 cm.
2. Draw a light pencil line across the width of the paper 2.5 cm from the end of the strip.
3. With a pointed end of applicator stick, place a small drop of a solution containing a mixture of
glucose.
4. Allow the spot to dry in air.
5. When dry, put the other end of the strip in a razor slit in a rubber stopper.
6. Insert the stopper tightly into a 25 x 200 mm test tube containing 5 ml of developing solvent (a
mixture of n-butanol, glacial acetic acid and water 4:1:5 parts by volume)
7. The end of the strips should be placed into the solvent 1-1.5 but the solvent level must be below
the carbohydrates spot in the starting line. The paper should not touch the sides of the test
tubes.
8. The solvent will rise by capillary action. When it reaches within a few centimeters, remove the
strip, mark the solvent boundary with a pencil and allow the strip to dry in air.
9. When the strip is dry, spray with CuSO4 – Na2CO3 – Sodium Citrate spray reagent.
10. Place the moist strip in an oven at 105°c for 5-10 minutes. The carbohydrates will appear as
yellow spots which will turn blue when exposed to light.
11. Draw a pencil around each spot and place a dot at the point where the color is most dense.
12. Calculate the R.f. value for each carbohydrate by measuring the distance. The carbohydrate
moved from the starting line and the distance the solvent boundary moved from the starting
line.
R.f. = distance traveled by sugar

distance traveled by solvent
The R.f. value in this solvent has two, found to be Glucose 0.18, fructose 0.23, ribose 0.31
Name:_______________________________________________ Date Performed:_______________
48
EXPERIMENT NO. 4
CHEMISTRY OF LIPIDS
Extraction, Fractionation, and Identification of brain lipids
The brain has a high content of lipids. It contains complex lipids such as the phosphatides,
cerebrosides, spinosides. This contain fatty acids component of the C-16 to C-24 carbon chains,
chiefly in the form strearic, linoleic, oleic and arachidoic acids. The steroid, alcohol, cholesterol is
abundantly accuring in the brain.
The experiment maybe divided essentially into four main parts:

1. Extraction of the total lipids
2. Isolation of the triglycerides saponification of the triglycerides make way for the study of soaps,
fatty acid and tryglycerol
3. Isolation and tests for lecithin
4. Isolation and tests for cholesterol
Precautions:
Keep flammable solvents and extracts away from open flames.
Procedure:
Extraction of the lipids from the brain materials

1. Cut about 40 gm of pig’s brain and homogenized the material in a Waring Blender with ether-
alcohol mixture (5:2 prepared by mixing 150 ml ether with 60 ml ethyl alcohol)
2. Stir the mixture for 10-15 minutes
3. Filter the material, the ether alcohol extracts contain the total lipids
4. Divide the ether-alcohol extract into two parts
a. 2/3 portion
b. 1/3 portion
Isolation of Triglycerides
1. Take the first 2/3 portion (from A) and evaporate the extract over hot water bath (without
flame) or over hot plate to a syrupy consistency.
2. Add 30 ml of alcoholic KOH, stir and transfer the mixture into an Erlenmeyer flask
3. Place a few glass beads to prevent bumping during heating. Cover the mouth of the flask with
funnel and put the flask over low heat for 30 minutes with occasional shaking.
4. Add distilled water if necessary to replace the lost volume. Continue heating until saponification
is complete. Add now the saponified mixture and 30 ml distilled water. Remove any insoluble
portion that may separate out.
5. Transfer the saponified solution into an evaporating dish. Heat again until the alcohol is driven
off.
6. Evaporate to a thicker consistency. Finally, add 50 ml of water and divide the solution into:
a. 20 ml portion
b. 30 ml portion
Test for Soap
49
(Use the 20 ml portion)
Salting-out
Take the 5 ml of the solution and add a little NaCl. The salt will dissolve; add some more until no
more dissolves. Note the precipitate formed. The soap has been salted out. The precipitate should
dissolve in water.
Insoluble Soaps
To 2.5 ml portion of the soap solution in two tubes, add to the point of saturation, 0.5N CaCl 2 to the
first and 0.5M MgSO4 to the second tube. Test the solubilities of the precipitate in water.
Test for Fatty Acids (Use the 30 ml portion from b)

Heat the solution to boiling. Add concentrated HCl drop by drop until no more precipitate forms
(Excess HCl dissolves the precipitate). The precipitate is fatty acids. The fatty acids may appear as
an oil layer while the solution is still hot but will precipitate upon cooling (Reserve the aqueous
portion for test for glycerol)
Solubility Test: Test the solubility of the fatty acids in water chloroform.
Translucent Tests
Place a small amount of the precipitate in 5 ml of CHCl 3. Add Hubi’s iodine reagent drop by drop and
shake between additions. The iodine reagent will decolorize if unsaturated fatty acids are present.
This is due to the adsorption of the iodine by the double bonds of the fatty acids. A central test
should be done by shaking CHCl3 with the iodine reagent but to which no fatty acids has been added.
Test for Glycerol

Place the aqueous layer obtained from the test for fatty acids in an evaporating dish. Heat until the
solution is reduced to about 20 ml. use this concentrate for the following tests:
50
Solubility Test: Add a few drops of the solution into 3ml portion of water and chloroform.
Acrolein Test: Place about 0.5 gm of powdered KHSO 4 in clean, dry hard test tube. Add about 10
drops of the glycerol solution. Heat the solution, cautiously at the first then more strongly. Note the
odor produced.
Benedict’s Test: Test your solution with litmus paper for any activity if acidic, neutralize with a
few drops of dilute sodium carbonate solution. Add 5 drops of the glycerol solution into a test tube
containing 5 ml of Benedict’s reagent. Boil the mixture for 2 to 3 minutes. Observe the results,
compare with the results obtained from the tests for sugars
Isolation and test for Lecithin

Place the remaining 1/3 portion from (b) in an evaporating dish and heat until concentrated to a
syrup consistency. Add about 15 ml of acetone and stir again. The precipitate is mostly lecithin.
Filter off the precipitate and reserve the filtrate for cholesterol.
Test for the presence of Phosphorous in Lecithin

Procedure:
1. To a small portion of lecithin precipitate in an evaporating dish, add about 5 ml of 10% HCl.
2. Boil for 10 minutes making sure that it does not dry up.
3. Add an equal volume of concentrated nitric acid and a few ml of ammonium phosphomolybdate
solution.
4. Warm the mixture continuously until fine yellow crystals ammonium phosphomolybdate are
produced. The crystal may appear upon cooling.
Isolation test for Cholesterol

The test for cholesterol maybe done in two ways:
51
A. Directly from the acetone-ether filtrate from the isolation of lecithin. The filtrate, however, contains
saponifiable lipids.
Procedure:
Evaporate the ether-acetone filtrate over a hot water bath or hot plate until syrupy. The residue
should not carbonize. Cool and dissolve the residue in 8 ml of CHCl 3. Filter and get a clean solution. The
chloroform solution is ready for the color reaction.
B. Treating the ether-acetone filter with alcoholic KOH to remove the

saponifiable lipids. The cholesterol by this method will be in a more or less pure form.
Procedure:
Evaporate the ether-acetone filtrate from the isolated lecithin (b) to a syrupy consistency. Cool and
add alcoholic KOH and follow the procedure. When the saponification is complete, add 30 ml of ether. The
ether extracts contain cholesterol, the aqueous portion contain soaps and glycerol.
Separate the ether layer, transfer into an evaporating dish and heat over a hot plate or water bath
until dry. Dissolve the residue in 8 ml CHCl3. Filter if necessary.
The CHCl3 extracts of cholesterol from the isolation of cholesterol may now be used for the color
reactions.
Note: use dry test tubes, the CHCl3 solution must be free from water.
Salkowski Test
Place the 4 ml of the extracts in a test tube and add equal volume of concentrated sulfuric acid. Note
the colors produced in the CHCl3 layer and I acid layer.
Liebermann-Burchard Test
To the remaining 4 ml of the extract, add about 10 drops of acetic acid anhydride and 2 drops of
concentrated sulfuric acid. Note the final color produced.
This is a very sensitive test for cholesterol and is utilized for the quantitative termination of
cholesterol.
52
1. Using the general formula for triglycerides, write the equation involved in saponification.
2. What is the principle involved in the cleaning action of soaps?
3. Differentiate insoluble soaps from soft soaps and hard soaps.
4. Compare the properties of glycerol and fatty acids as performed in the laboratory.
5. Write the chemical reaction involved in acrolein test.
6. Why will glycerol gives positive Benedict’s test after being heated?
7. Write the chemical composition of lecithin. Relate the composition with the test performed.
8. To what class of lipids does lecithin belong?
9. Give some functions of lecithin in the body.
53
10. Why is alcoholic medium used in the alkaline hydrolysis of lipids?
11. What is the structural formula of cholesterol?
12. Explain the reactions involved in the Liebermann-Burchard test.
54
EXPERIMENT NO. 5
NUCLEIC ACIDS
Learning outcomes
1. Perform qualitative test for RNA
2. Investigate the products of hydrolysis of Nucleic acid.
I. Discussion:
Nucleic Acid
Nucleic acids are polyanionic molecules of high molecular weight. These polymers are composed of
a sequence of polynucleotide.
There are two types of nucleic acids, the ribonucleic acid (RNA) and deoxyribonucleic acid (DNA)
which on hydrolysis yields the sugar ribose and deoxyribose respectively. Nucleic acids are found in the
chromatin granules of the nucleus (DNA) and cytoplasm (RNA). They are responsible for the storage and
transmission of genetic information.
Nucleic acids are mostly conjugated with proteins such as histines to form nucleoproteins which on
hydrolysis yields protein and nucleic acids, DNA can be isolated from thymus or muscle tissue, RNA from
yeast
The main difference between DNA and RNA lies in their sugar components and in one of their
pyrimidine bases, DNA contains d-deoxyribose and thymine while RNA has d-ribose and uracil instead of
thymine.
Various physio-chemical studies on DNA show that aqueous solution of DNA has very large
molecular weight and very rigid structure.
Isolation of Ribonucleic Acid

Mix 1 gm of yeast with 1 gm of white sand. Grind the mixture in a mortar. Then add 15 ml of freshly
prepared 0.8% NaOH to make a smooth creamy paste. Pour the mixture in a 250 ml beaker and make-up
the volume to 50 ml by adding enough 0.2% NaOH. Cover with a watch glass to prevent evaporation. Heat
the beaker in a water bath controlled at 90°c for 30 minutes. Filter 3 times through cheesecloth and once
through filter paper. Cool the filtrate. Perform the following test on the filtrate.
Procedure:
Qualitative Test for RNA

1. Test for Nucleoprotein
To 1 ml of the filtrate add 1 ml of 10% NaOH solution and 5-10 drops of 0.5% CuSO 4 solution.
2. Mild Acid Hydrolysis

Add 20 ml of 10% H2SO4 to the remaining filtrate. Boil gently for a few minutes. Divide the solution
to 3 parts and perform the following tests:
55
A. Test for the presence of Purines
Add 3 ml of 10% NH4OH to 2 ml of the solution. Mix 2-3 drops of
AgNO3 solution.
B. Test for the presence of Ribose

Mix 3 ml of Bial’s orcinol reagent to 1.5 ml of the following solutions:
No.1 = solution from acid hydrolysis
No. 2 = 0.1% ribose
No. 3 = 0.1% glucose
Place the test tubes on a boiling water bath until a color develops.
C. Test for the presence of Phosphates

To 1 ml of the solution add 2 ml of 10% HNO3 and 2 ml of 5%
(NH4)2MoO4. Heat to boiling. Let it stand for a few minutes.
Isolation of Deoxyribonucleic Acid

To 10 gm of fresh ground beef muscle tissue add 50 ml of 10% TCA. Mix thoroughly. Heat in water
bath controlled at 90-95°c for 15 minutes. Stir the contents occasionally. Filter and cool.
56
Physical Property of DNA
1. Determination of Viscosity (at room temperature)
Use the DNA solution prepared above. Draw the solution into a 1 ml pipette. Record the time in
seconds required to drain the pipette. Perform three (3) trials.
Note: Hold the end of the pipette into the solution during the determination. Compare the time requires for
distilled water to fall the same distance. Perform three (3) trials also
2. Effect of Heat on Viscosity

Device a procedure to show the effect of temperature on the viscosity of DNA solutions.
Qualitative test for DNA

Repeat the test for nucleoproteins, purine and phosphates of the DNA solution obtained above
1. Test for the presence of Deoxyribose

1. Add 5 ml of diphenylamine to 2 ml of the following solutions
No. 1 = filtrate from the DNA solutions
No. 2 = 0.1% ribose
No. 3 = 0.2% glucose
Place on boiling water bath until a color develops.

57
A. Test for Nucleoproteins

1. What type of molecule is present in the filtrate?
2. What are the nucleoproteins found?
B. Mild Acid Hydrolysis

1. What are the products of hydrolysis of nucleoproteins?
2. Would the proteins be hydrolyzed to amino acids by the procedure? Why?
C. Test for Purines

Name the purines in the nucleic acid yeast
D. Test for Ribose

What components of RNA react with Bial’s Orcinol Reagent?
58
EXPERIMENT NO. 6
PROTEINS
Learning outcomes
At the end of the experiment the student must be able to::
1. To be able to test for the presence of different elements in protein sample.
2. To perform the color reaction test for proteins to determine other elements
present in the sample.
I. Discussion
Proteins
Proteins are nitrogen-containing compounds composed of amino acids joined covalently by peptide
bonds. Each amino acid has a basic group and acidic group plus a characteristic radical. The chemical
properties of these amino acids are due to these groups and the chemical properties of proteins in turn are
dependent upon these amino acids. Upon hydrolysis with acids, alkali or enzymes finally to alpha amino
acids.
II. Procedure
A. Elements found in Proteins

1. Boil 0.2 gm of hair with 3 ml of sodium hydroxide. Boil for 3 minutes. Dilute with 5 ml of water, cool
and filter. Acidify the filtrate with acetic acid and a few drops of lead acetate.
What element is present?
2. Mix 0.2 gm of casein with 1 gm of soda lime and heat strongly. Note the odor of the vapor. Take a
confirmatory test for the gas evolved.

What element is present?

59
1. Give the chemical equation involved.
2. What is the principle involved in the test?
3. What is the black precipitate formed in the above test?
B. Isoelectric Precipitation of Casein

1. Place 20 ml of skimmed milk in a 250 ml beaker
60
2. Add slowly 0.1M HCl from a buret with constant mixing until the pH of the solution is 4.6 (use pH
meter or an appropriate indicator). The acid should be added at a rate of about 5 ml per minute while
mixing the solution in order to avoid momentary acidity.
3. Allow the suspension to stand until the precipitated casein settles. Filter the precipitate.
4. Decant and discard the supernatant.
5. Wash the precipitate with water until the washing is no longer acid to litmus paper.
6. Dry the precipitate by pressing between filter papers.
7. Weigh the dried precipitate, using the density of milk as 1.03 g/ml, calculate the % by weight of casein
isolated.

61
1. What is meant by the isoelectric point of a protein?
2. At what pH is a protein least soluble? Why?
C. Isolation of Bean Proteins

1. Sort and wash 1 cup of beans
62
2. Soak in water for 1 day.
3. Grind the beans finely with water and blend with more water using the waring blender.
4. Extract the milk through cheesecloth and discard the insoluble residue.
5. Add 1N HAC until precipitate is complete. Allow the mixture to stand for sometime.
6. Strain through cheesecloth and squeeze the water out as much as possible.
7. Weigh the dry precipitate.

63
1. What is the purpose of soaking the bean in water?
2. What are the predominant proteins present on beans?
3. How are these proteins precipitated by an acid?
D. Color Reactions of Proteins
64
These tests are based on the production of colored end products as a result of the reaction between
chemical reagents with one or more radicals of the reaction between chemical reagents with one or more
radicals or groups present in the complex protein molecule. The test yield varying degrees or intensities of
colors depending on the nature and the amount of groups contained in the particular protein examined. It
should be emphasized that these color tests are not specific for proteins because other substance may
respond to these tests as long as they have the radicals or group similar to proteins.
1. Million’s Test
To 5 ml of protein in a test tube, add 3-4 drops of Million’s reagent. Mix and bring the mixture
gradually to boiling point over a low flame.

65
1. Name an amino acid that responds to this test.
2. Why is this test never used in detecting protein in urine?
3. Why do all proteins give a positive Biuret test?
2. Xanthoproteic Reaction
66
To 3 ml of protein solution in a test tube, add 1 ml of concentrated HNO 3. Mix and warm the
mixture, observe. Cool the solution and add an excess of concentrated NaOH solution.
Principle involved in this test?
67
1. Why is the test not needed for urinary examination?
2. What amino acids give a positive reaction with this test?
3. Why does nitric acid stain the skin a yellow color?
3. Hopkin’s Cole Reaction
68
Mix 2 ml of protein solution and 2 ml of Hopkin’s Cole reagent. Incline the tube and allow 3 ml of
concentrated sulfuric acid to flow down the sides of the tube to form a layer of acid beneath the protein
mixture.
69
1. What is the cause of this color reaction?
2. Name the amino acids that respond to this test.
4. Biuret Test
70
To 3 ml of protein solution in a test tube, add an equal volume of 10% NaOH solution, mix
thoroughly and add 0.5% Copper sulfate drop by drop until a purplish violet color is observed.
Repeat the test on UREA. Fill the round bottom of the test tube with urea. Heat until it melts. What
fumes are given off/ what do you call the white residue? Perform the test on the residue. What color is
produced?
71
1. What is responsible for color reaction?
2. If the protein is completely hydrolyzed, will the hydrolysate yield a positive result? Why?
5. Test for Glycoprotein
72
Add 2 drops of 1% solution of alpha Napthol in alcohol to 3 ml of protein solution in a test tube. Mix
and run 2-3 ml of concentrated sulfuric acid down the sides of the test tube containing the mixture.
6. Sakaguchi test
To 3 ml of protein solution, add 1 ml of 10% NaOH solution in a test tube. Add about 15 drops of
alkaline hypobromite. A color develops which fades immediately. However it can be prevented by addition
of 40% urea solution.
7. Erlich’s Diazo Reaction

1. Mix equal volume of sulfanilic acid and 0.5% sodium nitrate solution.
2. Allow to stand for 1 minute.
3. Add 1 ml of protein solution and make alkaline with 10% sodium carbonate.
4. Run a control using water and compare the result with that obtained with protein solution.
8. Ninhydrine Test
1. To 2 ml of protein solution, add 0.5 ml of a 0.1% Ninhydrine solution.
2. Heat to boiling for about 3 minutes and allow cooling.
73
1. What chemical groupings give a positive test?
2. What color will be produced by protein with Ninhydrine?
9. Tryptophan Reaction (Adamkiewie’s)
74
1. To 1 ml of glacial acetic acid, add 2 drops of protein solution.
2. Mix well and in an inclined position pour concentrated sulfuric acid down the sides of the test tube
until a colored layer forms at the junction.
10. Liebermann’s Reaction

Mix 1 ml of egg albumin with 6 drops of 1% sucrose solution. Layer with concentrated sulfuric acid
Principle Involved in this test?
75
EXPERIMENT NO. 7
PRECIPITATION REACTIONS OF PROTEINS
Learning outcomes
1. To be able to conduct different methods of testing to precipitate protein in identifying the
presence of the different substances present in the sample.
2. Identify the different protein precipitate reaction as to color and solubility.
I. Discussion
Precipitation Reactions of Proteins

Proteins are precipitated from their solutions by a variety of reagents namely neutral salts, acids,
salts of heavy metals, dehydrating agents, dyes, etc. Many proteins are coagulated by heat. The reactions
and mechanisms involved are generally still uncertain and difficult to rationalize considering the variety
complex nature of proteins. Proteins show greatr differences in behavior with the reagents. However the
reactions are presumably explained on the basis of their physio-chemical properties and colloidal nature.
II. Procedure.
Fill 6 test tubes with 2 ml of dilute egg albumin solution, add the following reagents drop by drop.
Reagents Color of precipitate

a. Mercuric chloride
b. Lead acetate
c. Silver nitrate
d. Copper sulfate
e. Ferric chloride
f. Barium chloride
76
1. Why is the egg albumin in a good antidote for lead and mercuric poisoning?
2. Is it equally a good antidote for other metallic salts tested? Why?
Salting-out Process
77
Add solid ammonium sulfate to the pint of saturation to 10 ml of saturation to 10 ml of egg albumin
solution, keeping the temperature below 40°c. Filter and test the precipitate with Million’s reagent and the
filtrate with Biuret test.
Precipitated by Alcohols
Place in each of the three test tubes, 5 ml of 95% alcohol. To the first add 1 drop of dilute HCl. To
the second, add 2 drops of 10% NaOH. Leave the third neutral. Add to each tube drops of egg albumin
solution.
78
1. Why alcoholic solution used to disinfect areas of the skin before surgery?
2. Which is more effective disinfectant, 95% alcohol or 70% alcohol? Why?
Precipitation by Alkaloidal Reagents
79
Place 12 ml of dilute egg albumin solution in each of the 6 test tubes. Add the following reagents
drop by drop at first until an excess is added. Observe the changes.
Reagents Color of precipitate

a. Picric acid
b. Trichloroacetic acid
c. Tannic acid
d. Phosphomolybdic acid
e. Potassic-mercuric acid
f. Phosphotungstic acid
Give the practical application of picric acid and tannic acid.
Heller’s Test
Place the 5 ml of concentrated nitric acid in a test tube. Incline the tube and allow dilute egg
albumin in solution to flow slowly on the sides of the tube. At the point of contact, what do you
observed?
Robert’s Test
Place 5 ml of Robert’s reagent in a test tube. Allow the albumin solution to flow slowly on the sides
of the tubes. What is observed at the junction of the two liquid?
80
1. What is common between the two tests? Give the principle involved in each tests.
2. What is the basis of the Heller’s test?
Denaturation of Alkali Metaproteins
81
Preparation of Alkali Metaproteins
1. Place the white of an egg in an evaporating dish. Add 10% NaOH solution drop by drop with
constant stirring. The mass gradually thickens and finally assumes the consistency of a jelly. Do not
add an excess of the alkali or the jelly will dissolves.
2. Cut into small pieces and wash with running water until free from adherent alkali.
3. Add a small amount of water and dissolve with the aid of gently heat.
4. Cool and divide into two parts.
5. To the first part, neutralize with dilute HCl. Note the odor of the gas liberated and the precipitate
formed.
6. Filter off the precipitate and test as follows:
Solubility Result
Precipitate in water
Precipitate in HCl
Precipitate in NaCl
Precipitate in NaOH
Test for Sulfur

To a solution of metaprotein, add an equal amount of KOH solution. Add 2-3 drops of lead acetate
t.s. and boil. Note the color produced. Add HCl solution and note the characteristic, odor evolved from the
solution.
Solubility Test
Use coagulated egg white prepared by boiling the egg for 10-15 minutes with the following solvents
Solvents Observation
1. Water
2. NaCl Solution
3. Dilute HCl
4. Dilute NaOH
Differentiate coagulation and Denaturation.
82
EXPERIMENT NO. 8
MILK EXAMINATION
Learning outcomes
1. To test the properties of milk using different qualitative analysis.
2. Discuss the significance of the reaction produced in different procedures
I. Procedure
1. Reaction of Milk: Test a sample of milk with red and blue litmus papers, congo red and
phenolphthalein.
Results and observations
2. Place a drop of fresh whole milk on a side and cover with cover slip. Examine under the microscope.
Illustrate below:
3. Determination of Specific Gravity: Determine the specific gravity of both fresh whole milk and
skimmed milk. Which has higher specific gravity? Why?
Results and observations:
4. Coagulation Test: Place 2 ml of fresh milk in a test tube, acidify with dilute acetic acid and heat to
boiling. Observe.
5. Film Formation: Place 10 ml of the fresh milk in a small evaporating dish and boil. Note the
formation of a film on the surface. Remove the first film and heat again. Try to obtain a film.
Question:
What do you think is the film made of?
Test for the presence of Inorganic Salts
83
To about 5 ml, ad 6-8 drops of dilute HNO 3 to form a precipitate. Filter and test the filtrate for the
presence of the following salts:
Inorganic Salts Observation

1. Chloride
2. Sulfates
3. Phosphates
Test for the presence of Enzymes
A. Guiac Test
To 3 ml of water and a drop of milk and enough alcoholic solution of Guiac until turbidity is
produced. Mix thoroughly and note any change in color. If no color appears within 5 minutes add
hydrogen peroxide until color is produced.
B. Benzidine Test
To 2 ml of dilute milk, add 2 drops of saturated solution of Benzidine in glacial acetic acid and 1
drop of hydrogen peroxide.
Saturation of Magnesium Sulfate

Place 2 ml of milk and saturated with solid magnesium sulfate. What is the precipitate produced?
84
EXPERIMENT NO. 9
ENZYMES
Learning outcomes
1. Determine different properties of enzymes
2. Determine the biological activities of enzyme on varied conditions
I. Discussion
Enzymes
Enzymes are proteins which catalyze biological reactions. The activity of the enzymes is
altered to a large extent by changes in the condition of the reaction medium. The change maybe a
deviation from the optimum condition or the addition of a substance that can influence activity. In
either case, the result would be an enhancement or a decrease in the activity of the enzymes.
The activity of the enzymes is affected by several factors:
a. Substance concentration
b. Enzyme concentration
c. Temperature
d. pH of the reaction
e. Presence of activators and inhibitors
Enzyme action is usually demonstrated by the increased formation of the products and the
decrease in concentration of the substrate. In the digestion of starch by salivary amylase there is a
decrease in the substrate concentrate as determined by change in the starch-iodine reaction.
II. Procedure:
1. Determination of Salivary Amylase

To 1 ml of 1% starch paste in a small beaker, add 25 ml of saliva and stir the mixture
thoroughly. Remove a drop of the mixture at intervals of 5 minutes and test by the iodine test. Note
the color change produced using iodine test solution.
Observation:
When the solution gives no color reaction with iodine, test with Benedict’s test. Explain changes
observed.
2. Action of Heat on Enzymes
85
To 5 ml of water, add half a volume of saliva and boil the mixture for 2-3 minutes. Add to the
mixture a small amount of starch and shake thoroughly the final mixture.
Repeat Iodine test at intervals of 3 minutes. Repeat the same using the Benedict’s test.
Results:
3. Determination of Oxides
Put slices (10) from Chico, guava and potato. Put them on a watch glass and examine at
intervals. Note the changes.
Results: (Give the principle involved)
4. Effect of pH
Prepare 2 test tubes containing equal amounts of egg white. Place the test tubes in a hot
water bath. When the egg white is coagulated, cool and mark the height of the following solidified
egg white. Then add the following reagents to each test tube.
Test Tube 1 = 5 ml of 2% pepsin + 10 drops of 0.4% HCl
Test Tube 2 = 5 ml of 2% pepsin + 10 drops of 0.4% Na2CO3
Keep the test tube for 1 hour in a beaker of water controlled at 40°c. Determine the extent
of digestion in each test tube by noting the amount of protein dissolved or disintegrated. Perform
Biuret test on 1 ml of the supernatant liquid from each test tube. Compare the colors obtained.
86
1. What is the optimum pH of pepsin?
2. In what test tube has enzyme action occurred? Why?
5. Preparation of Catalase
87
Peel a potato and grate into 100 ml of distilled water. Let it stand for 10 minutes. Stir
occasionally, strain through cheesecloth and finally filter and extract through filter paper. Use the
extract in the following tests:
A. Biuret Test = to 10 ml of the extract add 2 ml of 10% NaOH solution. Mix thoroughly and
add 2-3 drops of 1% Copper sulfate. Observe the appearance of a violet color.
B. Test for Catalase Activity = Prepare 3 test tubes labeled 1, 2 and 3 respectively. Place 5 ml
of potato extract into tube no. 1, 5 ml of boiled extract in a test tube no. 2 and 5 ml of water
in test tube no. 3. Now add to each test tube 5 drops of an 8% solution of water and add 1 ml
of Benzidine solution. Note the formation of a blue to green solution.
Results and Observations:
88
1. What type of enzyme is Catalase?
2. Give the importance of the enzyme reaction in analysis.
3. How is Catalase prepared?
4. Give the positive test to identify this type of enzyme.
89
EXPERIMENT NO. 10
SALIVARY DIGESTION
Learning outcomes
1. Discuss the role of salivary enzymes in digestion
2. Determine the properties of salivary enzyme using test experiments
Procedure:
1. Microscopic examination
Stain unfiltered saliva with methylene blue and examine a drop under the microscope. Illustrate
below:
2. Reaction
Test the reaction of saliva on the following:
Red litmus paper

Blue litmus paper
Phenolphthalein
Congo Red
3. Specific gravity
The urinometer cylinder is partially filled with saliva and urinometer is introduced. Make own
reading.
Readings:
What are the different organic and inorganic solids present in saliva?
4. Test for inorganic constituents

Place 25 ml of filtered saliva in beaker, acidify it with acetic acid and heat to boiling. Filtrate is used
for the determination of inorganic constituents.
For Chloride: acidify with nitric acid, heat and then add silver nitrate solution
90
For Phosphates: acidify with nitric acid, heat and then add ammonium molybdate solution
For Sulfates: acidify with HCl, than add NaCl solution and warm
For Calcium: acidify with acetic acid, then add ammonium oxalate solution
5. Test for Nitrates

Take a small amount of saliva in a test tube and add 2 drops of concentrated sulfuric acid. Shake
well the tube and add a few drops of KI solution and a little potato starch paste
6. Test for thiocyanates

Add a few drops of dilute ferric chloride solution to a small amount of saliva. Acidify the mixture
slightly with HCl. Note the result.
7. Test for mucin
a. Precipitation of mucin
Add 1-2 drops of dilute acid to a small amount of saliva in a test tube. What precipitate is
formed? Write your observation:
b. Biuret test
Place a small amount of saliva in a test tube and render it alkaline with a 3 drops of
KOH. To the mixture, add a few drops of dilute copper sulfate solution. Write your result:
91
c. Million’s test
To a small amount of saliva in a test tube, add a few drops of Million’s reagent. Note the color of
the precipitate formed. Gradually heat the mixture and note the changes in color of the
precipitate.
8. Influence of dilution
Set on your test tube rack, 6 test tubes each containing 9 ml of water. To test tube 1, add 1
ml of saliva then mix thoroughly. Transfer 1ml of solution from test tube 1 to test tube 2. Mix the
content of test tube2 and transfer 1 ml of this to test tube 3 and so on and so forth until you have
dilutions of decreasing strength. To each tube, add a small and equal amount starch paste, shake
well and allow them to remaining in the water bath at 40°c for 20 minutes. Test the mixture with
iodine and Fehling’s solution. Tabulate your result.
Test tube Fehling’s test Iodine test

1
2
3
4
5
6
9. Influence of temperature
Prepare 4 test tubes and place 5ml of starch paste in each test tube. Immerse the first tube
in cold running water. Keep the second tube at room temperature.
Place the third tube in water bath set at 40°c. Add to the contents of these three tubes, a
small amount of saliva (2ml) and to the content of the 4 th tube add a small amount of boiled saliva.
Shake to mix the contents of the four tubes. Note in which test tube there is more rapid
digestion in terms of unit of time (in terms of minutes)
Test tube Benedict’s test Iodine test

1
2
3
92
EXPERIMENT NO. 11
URINE
Learning outcomes
1. Describe the physical and chemical properties of urine
2. Perform the physical, chemical and microscopic examination of urine
3. Relate the findings of urine tests to actual clinical conditions
Discussion:
Urine
The study of the composition of urine is important in determining the physiological or

pathological state of an individual.
The composition of urine carries from time to time within 24-hour period and is dependent
on several factors such as the physiological state of the individual and the nature of diet. Therefore,
quantitative estimation of the constituents should be done on a 24 hour sample rather than on
samples collected at random preferably before breakfast is satisfactory although it is more accurate
to do a quantitative and qualitative determination on a 24-hour urine sample. Samples voided at
random should be examined within 1-2 hour after voiding
Procedure
Physical Examination
Color -
Volume/unit of time -
Odor -
Transparency -
Specific gravity -
93
Specific Determination of Gravity
Procedure:
1. Fill the cylinder nearly full with urine sample
2. Place the urinometer to float
3. Determine the reading at an eye level
4. Take care that the urinometer does not touch the cylinder
5. Make 3 readings and get the average
Results:
Chemical Tests
A. TEST FOR GLUCOSE
Benedict’s Test
a. Place 5 ml. Of Benedict’s reagent in a test tube.
b. Add o.5 ml. (or 10 drops) of urine.
c. Mix thoroughly by shaking.
d. Place in a pan of boiling water for 5 minutes.
e. Remove from boiling water boiling and read results immediately.
INTERPRETATION:
Negative clear blue solution

Trace green opacity but no precipitate
+ green opacity with yellow precipitate
++ Yellow to green solution with yellow precipitate
+++ Muddy orange solution with yellow precipitate
++++ Orange to red precipitate
Result and Observation: (Illustrate)
94
B. TEST FOR BLOOD
Guaiac’s Test
a. Place 1 ml. Of urine in a test tube.

b. Add 1 ml. of glacial acetic acid. Mix.
c. Ass 2 ml. of fresh 3% hydrogen peroxide. Shake.
d. Pour the guaiac solution down the side of the tube.
e. A green to blue color will be formed at the zone of contact if blood is present.
Result and observations
C. TEST FOR PROTEIN

Test for Albumin
Heat and Acetic acid test
1. Place 2 ml. Of clear urine in a test tube.

2. Boil the upper portion of urine by direct heating using the alchol lamp.
3. Add 2 – 4 drops of 10 – 20 % acetic acid.
4. Take note of cloudiness after addition of acetic acid.
NOTE: Albumin is present if the cloudiness in the boiled portion persists after
addition of acetic acid.
Report result as follows:
Negative - no turbidity observed

Trace - turbidity is barely visible
+ - distinct turbidity
++ - turbidity with granulation
+++ - heavy turbidity
++++ - heavy and flocculent turbidity
95
D. Test for Indican
Obermeyer’s test
1. Place 2 ml of urine in a test tube
2. Add equal amount of Obermeyer’s reagent
3. Add I ml of chloroform
4. Shake by inverting the tube
Results and Observation:
E. TEST FOR BILE PIGMENT
Gmelin Test
a. Place 2 ml. of concentrated nitric acid in test tube.

b. Overlay with 5 ml. of urine
c. In the presence of bile pigments, play of colors (green, violet, yellow, red, blue)
will be observed
96
1. What are the substances that are responsible for the normal color of urine?
2. Why is a 24-hour urine specimen alkaline in reaction?
3. Define the following terms:
a. Polyuria
b. Anuria
c. Nocturia
d. Oliguria
e. Isestinuria
f. Hypotenuria
g. Hyperstenuria
97
EXPERIMENT NO. 12
STOOL ANALYSIS
Learning outcomes
1. Describe the physical and chemical properties of stool
2. Perform the physical, chemical and microscopic examination of stool
3. Relate the findings of stool exams to actual clinical conditions
Procedure:
Macroscopic Appearance
Color –
Consistency –
Odor –
What is responsible for the normal color of the stool?
Microscopic Examination
1. Place a small amount of feces in a clean slide
2. Add a drop of water and emulsify
3. Break the mass into finer particles by means of a glass rod
4. Examine under LPO and HPO
Results and Observation:
Reaction
1. Mix thoroughly a small amount of feces with distilled water
2. Test the reaction with red and blue litmus paper, congo red and phenolphthalein.
98
Chemical Examination of Stool
Determination of Inorganic Constituents

1. Incinerate a small amount of feces in a crucible
2. Dissolve the ash in a small amount of diluted nitric acid
3. Dilute with water
4. Filter and test the filtrate for the following:
A. Chloride
Acidify with nitric acid and then add silver nitrate solution. Note the precipitate formed
B. Phosphates
Acidify with nitric acid, heat and then add few drops of ammonium molybdate solution
C. Sulfates
Acidify with HCL and add barium chloride solution and then heat. Note the precipitate
formed
D. Calcium
Acidify with acetic acid and then a few drops of ammonium oxalate. Note the production of
a precipitate.
Detection for Occult Blood
A. Benzidine reaction
1. Make a thin suspension of feces in 2 ml of water
2. Shake with 2 ml of ether to remove the fat. Then discard this ethereal extract
3. Add 1-2 drops of acetic acid to the residue and extract again with 2 ml of ether
4. Evaporate the ether extract in hot water bath
5. Dissolve the residue with 2 drops of water
6. Add a drop of a saturated solution of benzene in glacial acetic acid and drop of hydrogen
peroxide
Results:
B. Ortho-toluidine test
1. Place1 ml of fecal suspension in a test tube
2. Add 1 ml of ortho-toluidine in a glacial acetic acid
3. Then add 1 ml of hydrogen peroxide and observe
Results:
99
C. Guiac test
1. Place 1 ml of fecal suspension on a watch glass
2. Add a solution of Guiac in glacial acetic acid drop by drop
3. Gradually add hydrogen peroxide or old turpentine. Observe
Results:
Test for Bile Pigments
A. Gmellin’s Test
1. Using an evaporating dish or mortar, mix 7 ml of 10% barium chloride with a piece of
stool about a size of marble
2. After an emulsion is made, filter the mixture
3. When the funnel is completely drained, add a few drops of concentrated nitric acid to the
filter paper
4. Note the play of colors produced
Results:
B. Schmidt Sublimate test (test for stercobilin)

1. Place a portion of fresh feces the size of a marble in a mortar and rub with a small
quantity of mercuric chloride solution
2. Allow to stand for 24 hours in a covered petri dish
3. If stercobilin is present, a deep red color develops in a few minutes. If only traces are
present, it may take 24 hours to develop this color
Results:
100
C. Schlesinger test (urobilin)
1. Shake a sample of feces with ether to remove fat.
2. Extract the residue with 5% acid alcohol
3. Neutralize the yellow fluid that results with ammonia or sodium hydroxide solution
4. Mix with equal quantity of 1% zinc acetate in alcohol
5. Remove the precipitate, if any by filtration
6. Examine the clear filtrate against a dark background for the green fluorescence that
indicates the presence of urobilin
Results:
Detection of Fats - Microscopic Examination

1. A small aliquot of stool suspension is placed on a slide and mixed with 2 drops of 95%
alcohol
2. Add 2 drops of alcoholic solution of Sudan III and mix thoroughly
3. Place a cover slip and examine under microscope
Result:
Microscopic Examination of the Stool
Draw all microscopic elements that you may also see from fres stool specimen under the
microscope:
101
1. What is the significance of testing the presence of occult blood in the stool?
2. What is occult blood? Give the significance of its presence in feces
3. What is meant by steatorrhea?
4. What are the substances that contribute to the normal color of the stool?
5. Enumerate the substances which when ingested may alter the color of the feces.
102
EXPERIMENT NO. 13
BLOOD
Learning outcomes
1. Discuss the physical characteristics of blood.
2. Differentiate the solid, liquid and gaseous components of blood.
3. Perform blood tests that aids in diagnosis of disease.
Discussion:
BLOOD
Blood is the circulation tissue of the body. It consists of solid elements (RBC, WBC, Platelets)
suspended in a liquid medium, the plasma. Because of its numerous functions, its composition is
complex. It has access to all cells of the body, thus it contains countless substances in dispressed or
dissolve state, nutrients, enzymes, hormones, vitamins, metabolites, electrolytes, etc. as well as
intermediary and waste products of cell metabolism
Microscopic Blood Examination
Cellular Elements
Stain prepared blood smear with Wright’s stain and examine the cellular elements of the blood
under the microscope.
Draw the following:
RBC:
WBC:
Neutrophils:
Eosinophils:
Basophils:
103
Plasma Preparation
1. Make a venipuncture and withdraw 5ml of blood
2. Transfer the blood to a test tube containing the specified and quantity of anticoagulant
3. Mix the blood and the anticoagulant by completely inverting the test tube several times. Do
not shake
4. Centrifuge the blood at full speed for about 10 minutes
5. Collect the clear fluid and transfer it into a clean test tube
Drawing:
Serum preparation
1. Make a venipuncture and withdraw about 5 ml of blood
2. Transfer the blood to a serological tube
3. Place the tube containing the blood in a beaker or test tube rack
4. Let it stand for at least 30 minutes. This allows the blood to clot
5. Centrifuge at full speed for 10 minutes
6. Remove the test tube from the centrifuge
7. With a quick deliberate action, pour the serum into another test tube
Drawing:
Blood Collection Determination
A. Capillary Method
Sterilize the finger with alcohol and puncture as for blood counts. Fill a capillary tube with
the blood and note the time. After 2-3 minutes, carefully break off the end if the tube a small
piece at a time, as half minute interval and ends are slowly separated
When threads of fibrin are seen, coagulation has taken place.
B. Drop Method
Sterilize the finger with alcohol, dry and puncture as in blood count. Place several drops on
a clean slide and note the time
Draw a needle through another of the drops at half minute intervals and note the time when
threads of fibrin clining to the needle. Note the time
104
Test for the Extraction of Blood
A. Guiac test
To a solution of blood in attest tube, add solution of Guiac in glacial acetic acid (1.50) drop
until the fluid becomes turbid
Gradually add hydrogen peroxide and note the color produced.
Result:
B. Benzidine reaction
Place 3 ml of a dilute solution of blood in a test tube. Then add 3 ml of saturated solution of
benzidine in glacial acetic acid
Add about 1 ml of hydrogen peroxide and note the color produced
105
TABLE OF CONTENTS
Preface
Experiment 1 - Physical Chemistry 1
Experiment 2 – Elements Found in Physiologic Compounds 11
Experiment 3 – Chemistry of Carbohydrates 35
Experiment 4 – Chemistry of Nucleic Acid 43
Experiment 5 – Proteins 47
Experiment 6 – Precipitation Reaction of Protein 54
Experiment 7 – Milk Examination 58
Experiment 8 – Enzymes 60
Experiment 9 – Salivary Digestion 63
Experiment 10 – Urine Analysis 66
Experiment 11 – Stool Analysis 68
Experiment 12 – Blood Analysis 72
106
PREFACE
Biochemistry Laboratory Manual for Medical Technology Students has primarily been
prepared to provide students with basic concepts about the principle and skills involved in
procedures, and also to provide them with enough review materials in preparation for their future
professional subjects
It is with this ides in mind that the laboratory manual has been prepared to provide a brief
and concise presentation on the fundamental knowledge and principles of clinical and laboratory
procedures. Efforts have been exerted to present the topics in the simplest and clearest possible
way, so that the students may not find much difficulty in grasping and absorbing the ideas that the
lesson would like to impart
The author does not claim originality of the ideas, particularly of the procedures presented
in the manual. These have merely been taken from various authorities in the field, compiled and
reconstructed to provide the students with a simplified knowledge of the subject matter
AUTHOR
107

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Name:______________________________________________________ Date Performed:_______________

1. perform the procedures of titration and pH determination.

Two Types of Titration

B. Residual or Indirect Method – a solution in which a definite concentration of the

Standard Solution – A solution in which a definite concentration is dissolved in the solution.

A. 0.1N HCl C. Fresh Milk E. Urine

A. Phenolphthalein C. Phenol Red

8. Rinse the electrode with distilled water and wipe it dry.

9. Repeat the test with the other samples.

Results and Observation

1. What is the biochemical relevance of pH?

H+ (from outside source) + A- (buffer salt) HA

1. Prepare duplicate samples of the following:

2. Determine the original pH of each sample.

Results and Observations:

Buffer pH meter reading 0.1ml of 0.1N NaOH 0.1ml of 0.1N HCl

1. Which sample exhibited buffer actions? Explain

4. Give examples of buffer solutions.

Preparation of an Emulsoid: (Gelatin)

1. Prepare 5% solution of gelatin with the aid of heat.

2. Cool under the tap water and observe. What is produced?

3. Heat again and observe the result.

Write your results and observations below:

Preparation of Suspensoids: (FeCl3 solution saturated)

1. Place 100ml of boiling water in a beaker.

2. Add 2ml of saturated ferric chloride solution.

3. Observed the product formed.

Write your results and observations below:

1. Place 5ml 5% gelatin solution in a test tube.

3. Repeat the procedure using colloidal ferric.

Write your results and observations below:

Write your results and observations below:

Name:_______________________________________________________Section__________________ Date _______________

1. What is meant by a protective colloid?

2. On a tabular form compare Emulsoid from Suspensoid.

3. Discuss by differentiating sols and gels.

4. Explain, why is Suspensoid is readily precipitated by salts?

Dialysis is a common biochemical method of separation and purification by selective

4. Invert the tube and allow to dry for 15 minutes.

5. When dry, fill with water and stand for 10 minutes.

10. Let this stand for 30 minutes.

Write your results and observations below:

2. What other membranes beside colloidion might be used in dialysis?

Write your results and observations below:

1. identify the elements, present in physiological compounds.

Test for Carbon

1. Place small amount of sugar in a test tube.

Write your results and observations below:

Test for Hydrogen and Oxygen

1. Repeat the procedure above.

Write your results and observations below:

1. What are the elements contained in the sugar molecules?

2. Give 5 substances or compounds containing elements of C, H and O

3. Discuss the role of these elements in physiologic compounds.

Write your results and observations below:

Test for Sulfur

1. Place 10ml of sodium hydroxide t.s. in a test tube.

Name:________________________________________ Date Performed:_

Name:_______________________________________Section Date _