NUCLEOPROTEINS

Experiment No. 4

WHAT ARE NUCLEOPROTEINS?

Conjugated proteins, the prosthetic group of which are

nucleic acids
Found abundant in tissues possessing closely packed
cells with big nuclei, like thymus, liver, spleen, kidney
and pancreas
Also found in bacteria, bacteriophages, chromosomes
and constitute the whole makeup of the simple
filterable viruses

WHAT ARE NUCLEOPROTEINS?

Not coagulated by heat but exhibit the precipitation
and color reactions characteristic of protein substances
Have been proven to be desoxyribonucleic acids

LABORATORY RESULTS: SOLUBILITY

Solvents

Results

H2 O

Insoluble

NaCl

Soluble

HCl

Insoluble

KOH

Soluble

Alcohol

Insoluble

LABORATORY RESULTS: TEST FOR

PHOSPHATES AND COLOR REACTION TEST
Tests

Results

Biuret Test

Purple color

Xanthoproteic Reaction

Yellow to orange color

Benedicts Test

Brick red precipitate

Molisch Test

Violet ring

Test for Phosphates

Yellow solution with yellow precipitate

CHEMISTRY OF ENZYMES
Experiment No. 5

LABORATORY RESULTS: TEST FOR THE

ACTIVITY OF CATALASE
Test tube 1

Test tube 2

Production of more
Production of less
bubbles
bubbles
Production of blue
No reaction with
flame with lighted
lighted matchsticks
matchsticks
The yellow color was
Production of blue to
intensified with
green precipitate with
benzidine
benzidine

LABORATORY RESULTS: ACTION OF

PANCREATIN AND AMYLASE
Sample
Test tube 1

Result and
Observations
Blue black color

(starch solution + iodine solution)

Test tube 2

Violet to Blue color

(starch solution + pancreatin solution + heat +

iodine solution)

Test tube 3

Violet to Blue color

(starch solution + amylase solution + heat +

iodine solution)

Test tube 4

Blue color

(starch + amylase + ethanol + heat + iodine

solution)

Test tube 5
(starch + pancreatin + heat + iodine solution)

Violet color

EXPERIMENT 6A 7
MONOSACCHAARIDES AND
DISACCHARIDES

CARBOHYDRATES:
ARE HYDRATES OF CARBON

(CH2O)n
-are polyhydroxyaldehydes or
polyhydroxyketone, or
substances that give these
compounds on hydrolysis.

Carbohydrates
They have the molecular formulas Cn(H2O)n

CLASSIFICATION OF CARBOHYDRATES
1.

According to type of carbonyl group

aldoses - aldehyde group
ketoses - keto group

D and L notations are used to describe the

configurations of carbohydrates

CLASSIFICATION OF CARBOHYDRATES
2. According to the number of carbon atoms

Name

Formula

Triose
Tetrose
Pentose

C3 H6 O3
C4 H8 O4

Hexose
Heptose
Octose

C5 H1 0 O 5
C6 H1 2 O 6
C7 H1 4 O 7
C8 H1 6 O 8

CLASSIFICATION OF CARBOHYDRATES
There

are only two trioses:

CHO

CH2 OH

CHOH

C= O

CH2 OH

CH2 OH

Glyceraldehyde
(an aldotriose)

Dihydroxyacetone
(a ketotriose)

Often aldo- and keto- are omitted and these compounds

are referred to simply as trioses.

Although triose does not tell the nature of the carbonyl

group, it at least tells the number of carbons.

CLASSIFICATION OF CARBOHYDRATES
3. According to the number of sugar units
Monosaccharide
Simple sugars

compose of one sugar units

Disaccharide-

compose of two
monosaccharide units

Oligosaccharides

Oligo = "a few" - usually

2 to 10 monosaccharide units

Polysaccharides are polymers of the simple

sugars usually more than 10
monosaccharide units

EXPERIMENT 6-A
MONOSACCHARIDES(1)

TEST FOR CARBOHYDRATES

Moores

Test Test for the presence of

carbohydrates in a substance in the
influence of a concentrated alkali
Molisch Test is a general test for
carbohydrates where sugars are mixed with
-naphthol, the test tube is inclined and about
1mL of concentrated sulphuric acid is added
along the sides of the tube.

The color formed is due to the reaction of alphanaphthol with furfural and/or its derivative
formed by the dehydration of sugars by
concentrated sulphuric acid. All carbohydrates
react positively with this reagent.

TEST FOR CARBOHYDRATES

Seliwanoffs

Test use to
differentiate ketohexoses from
aldohexoses.
In the concentrated HCl, ketones
undergo dehydration to yield furfural
derivatives more rapidly than do
aldoses. These derivatives form
complexes with resorcinol to yield deep
red color.
It is a timed color reaction specific for
ketohexoses.

TEST FOR CARBOHYDRATES

Phenylhydrazine

The ketoses and aldoses react with phenylhydrazine to produce a phenylhydrazone

which in turn reacts with another two
molecules of phenylhydrazine to form the
osazone.
Glucose, fructose and mannose produce
needle-shaped yellow osazone crystals.
Whereas lactosazone in mushroom-shaped.
Different osazones show crystals of
different shapes. Maltose produces starshaped (flower-shaped) crystals.

EXPT. NO. 6 CARBOHYDRATES: THE

MONOSACCHARIDES (1)
Data and Results

A. Macroscopic Appearance
Glucose

Fructose

Galactose

Color

White

White

White

Odor

Odorless

Odorless

Odorless

Form

Crystalline

Crystalline

Crystalline

Fructose

Galactose

B. Solubility
Glucose
Water

Freely
Soluble

Freely
Soluble

Freely
Soluble

10% NaCl

Soluble

Soluble

Soluble

0.25% HCl

Soluble

Soluble

Soluble

95%Ethyl
alcohol

Slightly
soluble

Slightly
soluble

Slightly
soluble

C. Moores Test
Glucose

Fructose

Galactose

Color Change

brown

brown

Brown

Odor produced

Caramel like

Caramel like

Caramel like

Glucose

Fructose

Galactose

Purple to red
violet

Purple to red
violet

Purple to red
violet

D. Molisch Test

Color produced
at the junction
of the two
liquids

PHENYLHYDRAZINE REACTION
Glucosazone

yellow osazone
crystals are
formed

Fructosazone

Galactosazone

POST LAB QUESTIONS

The three simple sugars are most soluble in water.
2. In Molisch Test. what is the principle behind the
production of color at the junction of two liquids?
- The color formed is due to the reaction of alphanaphthol with furfural and/or its derivative formed
by the dehydration of sugars by concentrated
sulphuric acid
3. In phenylhydrazine reaction, what is the color of the
crystals formed? - yellow
4. In Moores Test. what is responsible for production of
color at the junction of two liquids?
- presence of concentrated alkali (NaOH)
1.

EXPERIMENT 6-B
MONOSACCHARIDES(2)

TEST FOR CARBOHYDRATES

Fehlings

Test
The blue alkaline cupric hydroxide present
in solution, when heated in the presence
of reducing sugars, gets reduced to yellow
or red cuprous oxide and it gets
precipitated. Hence, formation of the
colored precipitate indicates the presence
of reducing sugars in the test solution.
Cu(OH)2
Cu2O + H2O

FEHLINGS TEST
Reaction:
Rochelle salt

CuSO4 + KOH

Cu(OH)2 + K2SO4

reducing sugar

2 Cu(OH)2

Cu2O + H2O + [O]

brick red ppt

Note:
Fehlings A and B have to be kept in separate containers to prevent the
formation of Cu(OH)2 immediately.

TROMMERS TEST
Reaction:
CuSO4 + 2 NaOH

Cu(OH)2 + Na2SO4

reducing sugar

2 Cu(OH)2

Cu2O + H2O + [O]

brick red ppt

Cu(OH)2 + H2O

CuO + H2O
black ppt

NYLANDERS TEST
Reaction:
Rochelle salt

Bi(OH)2NO3 + KOH

Bi(OH)3 + KNO3

reducing sugar

2Bi(OH)3

2Bi + 3H2O + 3[O]

black ppt

TEST FOR CARBOHYDRATES

Benedicts Test
As in Fehlings test, the reducing sugar
because of having potentially free
aldehyde or keto group reduce cupric
hydroxide in alkaline solution to red
colored cuprous oxide.
Formation of red, yellow or green
color/precipitate.

BENEDICTS TEST
Reaction:
Na2CO3 + 2H2O

2NaOH + CO2 + H2O

sodium citrate

NaOH + CuSO4

Cu(OH)2 + Na2SO4

reducing sugar

2 Cu(OH)2

Cu2O + H2O + [O]

brick red ppt

TEST FOR CARBOHYDRATES

Barfoeds

Test

Only monosaccharides answer this test.

Since Barfoeds reagent (Copper acetate)
is weakly acidic, it is reduced only by
monosaccharides.
Formation of brick-red precipitate.

TEST FOR CARBOHYDRATES

Mucic

Acid Test

Formation of crystals
The both end carbon groups are
oxidized to carboxylic groups. The
resultant saccharic acid of galactose is
called mucic acid which is insoluble in
water.

TEST FOR CARBOHYDRATES

Picric
test

Acid Test

for the presence of reducing sugars

Picric acid is a chemical compound formally
called 2,4,6-trinitrophenol (TNP).
a toxic yellow crystalline solid and one of the
most acidic phenols.
It reacts with metal to form metal picrates.
formation of MAHOGANY RED SOLUTION

TEST FOR CARBOHYDRATES

EXPT.6-B THE MONOSACCHARIDES

(PART 2)
A.

Data and Results

Reduction Tests
Glucose

Fructose

Galactose

Trommers Test

Brick red

Brick red

Brick red

Fehlings Test

Brick red

Brick red

Brick red

Benedicts Test

Brick red

Brick red

Brick red

Nylanders Test

Black

Black

Black

Barfoeds Test

Brick red

Brick red

Brick red

Picric acid Test

Mahogany red

Mahogany red

Mahogany red

B. Phloroglucinol-HCl Test

Color produced

Glucose

Fructose

Galactose

brown

brown

red

C. Mucic Acid Test

Glucose and Fructose- no crystals formed
Galactose- white sandy crystals

D. Seliwanoffs Test
Glucose

Fructose

Galactose

yellow

Red color

Yellow

Time required for the

development of color

Color of the dissolved

precipitate in alcohol

Post-Lab Questions
1.

a. What difference have you observed when

Trommers test was carried out using distilled
water instead of sugar solution?
- formation of brick red ppt. is not observed in
distilled water, carbohydrate is not present.

1.b) What is the role of the sugar solutions in the

changes that have taken place?
- sugars act as reducing agents; sugars reduced
alkaline metals present in the solution/reagent
2.a. What is responsible for the production of the
colored precipitate? Write the reaction
involved.
- free monosaccharides are responsible for the
production of the colored precipitate
b. Why are Fehlings solution kept in separate
containers?
- to avoid formation of complex compound from the
substances present in Fehlings A and B that may
interfere Fehlings test

3. a. What are the differences between Fehlings

and Benedicts solutions?
Ans. Benedicts soln. is made of only one solution while
Fehlings is made of two. Fehlings is not specific to
aldehydes.
b. Between Benedicts and Fehlings solution,
which is more sensitive? Why?
Ans. Benedicts. Because it uses only one solution and
it undergoes a series of change in color when exposed to
heat.
4. What is the name of the colored precipitate
formed in Nylanders test?
- Metallic bismuth

5.

a. In phloroglucinol-HCl test, is there a development

of red color in all sugar solutions?

- No
b. Phloroglucinol-HCl test is a specific test for what
sugar?
- Galactose and Pentoses (like Ribose)

DISACCHARIDES
-COMPOSED OF TWO MONOSACCHARIDE SUBUNITS HOOKED
TOGETHER BY AN ACETAL LINKAGE

1.

MALTOSE
-D-glucose linked to -D-glucose by an
(1-4) glycosidic bond
- A reducing sugar
- repeating disaccharide unit of starch

DISACCHARIDES

Maltose
Present in malt, the juice from sprouted barley and
other cereal grains.
Maltose is a reducing sugar.

HOCH2 O
HO

CH2 OH

OH

OH
HO

OH

HO
O OH HO

-1,4-glycosidic
bond

CH2 OH
O
1

OH 4 CH2 OH
O
O
OH
HO
OH

2. LACTOSE

-D-galactopyranosyl-(1-4) -D-glucopyranoside
Made up of -D-galactose linked to -D-glucose
by a (1-4) glycosidic bond (galactose: brain sugar)
A reducing sugar

Lactose is the principal sugar present in milk; it

makes up about 5 to 8 percent of human milk and 4
to 6 percent of cow's milk

DISACCHARIDES
Lactose

CH2 OH
OH
O
OH

CH2 OH
O
OH

O
1

OH

OH

OH

OH
CH2 OH
HO

-1,4-glycosidic bond
O

4
1

OH

O
HO

CH2 OH

O
OH

OH

3. SUCROSE
Made

up of -D-fructose linked to an -Dglucose by a (2-1) glycosidic bond

non-reducing sugar (Why?)

Table

sugar

 Sucrose

(table sugar)

Sucrose is the most abundant disaccharide in the

biological world; it is obtained principally from the juice of
sugar cane and sugar beets.

Sucrose is a nonreducing sugar.

CH2 OH
O
OH

HO
HO

OH

HO
OH O
O
HO 2
CH2 OH
1
OH

HOCH2

a unit of -Dglucopyranose

CH2 OH
O

HOCH2

O
HO

O
2

-1,2-glycosidic bond
a unit of -Dfructofuranose

CH2 OH

OH

THE DISACCHARIDES

EXPT. 7 THE DISACCHARIDES

Data and Results
A. Moores test
Color
change
Odor
produced

Sucrose
colorless

Maltose
brown

Lactose
brown

Odorless

Caramel
like

Caramel
like

PHENYLHYDRAZINE REACTION
Sucrose

No crystals

Maltosazone

Lactosazone

C. Reduction Test
Sucrose

Maltose

Lactose

Trommers Test

colorless

Brick red ppt.

Brick red ppt.

Fehlings Test

Blue soln.

Brick red ppt.

Brick red ppt

Benedicts Test

Blue soln.

Brick red ppt.

Brick red ppt.

Nylanders Test

colorless

Black ppt.

Black ppt.

Barfoeds Test

Blue soln.

Blue soln.

Blue soln.

Picric Acid Test

Yellow solution

Mahogany red

Mahogany red

D. Phloroglucinol-HCl Test

Color produced

Sucrose

Maltose

Lactose

black

black

red

E. Mucic Acid Test

-only lactose can produce white sandy crystals
F.

Seliwanoffs Test

Sucrose

Maltose

Lactose

red

No red color

No red color

Time required for

the development
of red color
Color of the
dissolved
precipitate in
alcohol

G. Inversion of Sucrose
Test

Visible Result (with the

filtrate)

a. Reduction Test

Fehlings test

Brick red ppt

Benedicts test

Brick red ppt

Barfoeds test

Brick red ppt

Picric Acid test

Mahogany red

Phenylhydrazine Test
Ans. Formation of yellow
osazone crystals

EXPERIMENT 8-A & B

The Polysaccharides

POLYSACCHARIDES
Homopolysaccharides

Yield one kind of monosaccharide upon

hydrolysis

Heteropolysaccharides

Yields mixtures of monosaccharides and

derived products

HOMOPOLYSACCHARIDES
Starch

(C6H10O5)x

Found abundantly in plant kingdom

Most important constituent of human diet
A granule is made up of 98% amylose (300 glucose
units in unbranched chain)

Starch (amylose)
Acid

Glucose

Enzyme
(Amylase)

maltose

HOMOPOLYSACCHARIDES
Starch

(C6H10O5)x
The ratios of the two molecules amylose
and amylopectin, are different
depending on the plant which made
them High-amylose corn starch is 85%
amylose, while waxy corn is more than
99% amylopectin

STARCH COMPONENTS

Amylose. The simple glucose molecules have a

bond between the 1 carbon in the ring and the 4
carbon in the ring. They are joined by an oxygen.

Amylopectin is another component of starch and it

also consists entirely of repeating chains of glucose. It
is different from amylose because it has two different
types of linkages between the glucose subunits:
the 1C-4C and the 1C-6C. The 1:6 linkage makes a
branched structure instead of a linear structure. The
branches occur every 24 to 30 glucose units. The
molecules of Amylopectin are composed of between
2000 and 200,000 units of glucose.

STAGES IN STARCH HYDROLYSIS

Reaction with iodine

Benedicts test

Starch

Blue

Soluble starch

Blue

Amylodextrin

Purple

Erythrodextrin

Red

Achrodextrin

Colorless

++

Maltose

Colorless

+++

2 Glucose

Colorless

++++

HOMOPOLYSACCHARIDES
Inulin

(C6H10O5)x

Found in bulb of onions and garlic

White, odorless, tasteless powder; soluble in hot
water, slightly soluble in cold water and insoluble
in 60% alcohol
Optically active
Yields fructose upon hydrolysis

Glycogen

(C6H10O5)x

Animal starch, found mostly in liver as storage

material; in muscles as source of energy
Enzymatic (amylase) hydrolysis yields maltose
Acid hydrolysis yields glucose

CARBOHYDRATE AND COLOR IT YIELDS

WITH IODINE

Carbohydrate
Starch
Amylodextrin
Erythrodextrin
Archrodextrin
Glycogen/
Amylopectin

Color
Blue
Purple
Red
Colorless
Faint red

EXPT. 8-A THE POLYSACCHARIDES

(STARCH, INULIN AND GLYCOGEN)
Data and Results
Starch
A. Preparation of Potato Starch:
B. Microscopic Study

Potato starch

Cassava starch

Corn starch

Arrow root starch

EXPT. 8-A THE POLYSACCHARIDES

(STARCH, INULIN AND GLYCOGEN)
Data and Results
Macroscopic Appearance
Potato
Corn
starch
starch

Color
Odor
Form

Arrow
root
starch

Cassava
starch

White

White

White

White

Odorless

Odorless

Odorless

Odorless

Amorphous

Amorphous

Amorphous

Amorphous

D. Solubility
Solvent

Solubility

Color reaction with

iodine solution

Water

Insoluble

Yellow

Hydrochloric acid

Soluble

Blue

Sodium hydroxide

Slightly soluble

Colorless

Alcohol

Insoluble

Yellow

E. Iodine Test on Starch Powder

- Blue black or dark blue
- the blue black color is due to starch-iodine
complex

F. Iodine test on Starch Paste

- Blue black or dark blue

G. Hydrolysis of Starch
1. Reduction Tests
Test Performed

Visible Result with

Hydrolyzed starch

Fehlings Test

Brick red ppt

Benedicts Test

Brick red ppt

2. Phenylhydrazine Test
- Formation of glucosazone crystals

H. Influence of Tannic acid

Yellow to brown solution with white jelly-like ppt.

INULIN
A. Solubility
Solvents

Solubility of Inulin

Hot Water

Soluble

Cold water

Slightly soluble

Alcohol

Insoluble

B.

Iodine Test on Inulin Powder- yellow

C.

Iodine test on Inulin Solution - yellow

D.

Benedicts Test- blue color

E.

Hydrolysis of Inulin
Test Performed

Visible Result with

Hydrolyzed Inulin

Benedicts Test

Brick red ppt

Seliwanoffs Test

Brick red ppt

HYDROLYSIS OF INULIN
Inulin
acid/ inulase

Fructose

Glycogen
A.

B.

Preparation and Test of Glycogen

Test Performed

Visible Result with

Glycogen Solution

Benedicts Test

Blue

Iodine Test

Red

Hydrolysis of Glycogen
a. Benedicts Test brick red ppt.
b. Hydrolyzed glycogen contains reducing sugar
c. End product of glycogen hydrolysis is GLUCOSE

HYDROLYSIS OF GLYCOGEN
Glycogen
amylase

Maltose
acid

2 Glucose

EXPT. 8-B: THE POLYSACCHARIDES

( CELLULOSE AND HEMICELLULOSE)
Data and Results
Cellulose
A.
Solubility
Solvents

Solubility of Cotton

H2O

Insoluble

Dilute H2 SO4

Insoluble

Concentrated H2 SO4

Soluble

Dilute HCl

Insoluble

Concentrated HCl

Insoluble

95% alcohol

Insoluble

Dilute NaOH

Insoluble

Concentrated NaOH

Insoluble

Iodine Test -reddish brown color

Schweitzers Test - white flocculent precipitate
(cellulose acetate)
Cross and Bevans Test- white gelatinous ppt
(ethyl cellulose)

Formation of Amyloid blue-black or blue

Hydrolysis of Amyloid
Test Performed

Visible Result with

Hydrolyzed Amyloid Solution

Fehlings Test

Brick red ppt

Benedicts Test

Brick red ppt

Picric Acid Test

Mahogany Red

Hemicellulose
Pentosans: Gum Arabic
A. Macroscopic Appearance
Gum Arabic
Color

Flesh (dirty white)

Odor

Odorless

Form

Amorphous

Taste

Tasteless

B. Solubility

Solvents

Solubility of Gum
Arabic

Hot water

Freely soluble

Cold water
95% alcohol

Soluble
insoluble

C. Iodine Test
Ans. Yellowish color
D. Hydrolysis of Gum Arabic
ans. Liberation of arabinose

D-arabinose

Test Performed

Visible Result with

Hydrolyzed Gum Arabic

BenedictsTest

Brick red

Fehlings Test

Brick red

E. Phenylhydrazine Test
ans. Yellow crystals of
arabinosazone

Hexosan: Agar-Agar
A. Solubility
Solvents

Solubility of Agar-Agar

Hot water
Cold water
95% alcohol

Soluble forming gel

Insoluble
Insoluble

B. Iodine Test on Agar-Agar powder

- yellow

C. Iodine Test on Agar-agar solution

- yellow
D. Hydrolysis of Agar-Agar
Test Performed

Visible Result with

Hydrolyzed Agar-Agar

BenedictsTest
Fehlings Test

Brick red
Brick red

E. Mucic Acid Test

ans. White sandy crystals

1,4
linkage

1,4
linkage

CELLULOSE VS. STARCH

IODINE TEST FOR POLYSACCHARIDES

Glucose: Product upon hydrolysis

REMINDERS

February 23 - Monday
Submit Summary of Scores reflected in the Logbook
(1/4 piece of paper)
Submit BLUE answer sheet

February 24

Quizzes
Midterm Exam