Jazan University

Faculty of Science
Chemistry Department

PRACTICAL
ORGANIC CHEMISTRY
3rd Year Students
Part I

Name:………………..…………………………
Group:……………....…………………………
Ac. Year:……….…..………………………….
  

INVESTIGATION
OF SIMPLE ORGANIC COMPOUNDS

A – PHYSICAL PROPERTIES:
A knowledge as to whether a given compound is liquid or solid (if
solid, then crystalline or amorphous) is of great help in its identification
(this can by consulting tables of known compounds).
I-1- Colour:
The colour of the original sample is noted. Some compound show
colour due to impurities, which is frequently produced as a results of their
slow oxidation by moist air (freshly distilled aniline is nearly colourless but
goes reddish brown when kept for a longer time), while many others show
colour due to the presence of chromophoric groups in them. A brief
summery of conclusions that can be drawn by the observation of colour is
given here.

Observation Inference
Nitoanilines, azo compound, β-
Orange-red
naphthoquinone, alizarin
Brown Higher aromatic amines, diamines
Pink Naphthols
Greenish-yellow p- nitroso compounds, quinhydrone
Iodoform, nitrocompounds, quinines, α-
Yellow
diketones
Colourless, turning reddish
Phenol, aniline, aminophenols, tannic
brown or pinkish (due to air
acid, α- and β- aphthylamines
oxidation)
Colourless, turning yellowish or Anthracene, cinnamic acid, cresols,
yellow-brown …etc
Carbohydrates, aldehydes, ketones,
Colourless acids, esters, alcohols, ethers, many
hydrocarbons

 
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I-2- Odour :
Many types of organic compounds have characteristic odour and so
the ability to detect and remember an odour is helpful in qualitative
analysis. Some of the odour characteristics of compounds are given here.

Observation Inference
Benzaldehyde, benzonitrile,
Odour of better almond
nitrobenzene
Mouse-like Acetamide, acetonitrile
Cinnamon-like Cinnamic acid
Pleasant-fruity Ester
Pleasant, sweet Chloroform, diphenylamine, alcohols
Garlic Thiophenols, thioalcohols
Phenolic (carbolic) Many phenols
Lower acids, lower aldehydes, acid
Pungent, irritating
halides, thioacids
Camphor-like Pinacol, hexachloroethane
Intensely disagreeable Isocyanides
Vingar-like Acetic acid, aspirin
Cucumber-like Chloral
Pyridine-like Heterocyclic bases
Lower aliphatic amines, side-chain
Fish-like
aromatic amines
Carbohydrates, aromatic acid, glycerol,
odourless
solid aliphatic acid, …etc.

B- CHEMICAL PROPERTIES:

II-1- Elements Test (Lassaigne,s Test)

a) Required chemicals:
Soda limes, sodium metal (Na), distilled water, ammonium oxalate,
ammonium tartarate, acetanilide, benzamide, acetamide, antharnilic acid,

 
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glucose, fructose, sulfanilic acid, aniline HC1, aniline H2SO4, lead acetate,
ferrous sulfate, sulfuric acid, sodium nitroprusside and silver nitrate.
b) Apparatus:
Beakers, conical flasks, glass rod, ignition tubes, Bunsen flame, filter
papers, funnels, test tubes, rack for test tubes and litmus papers.
c) Experimental tests:
This test is aimed to identify the nitrogen, sulphur and halogen in
organic compounds and it can be carried out by fusion of the organic
substance with sodium metal in the presence of excess soda lime
(NaOH/CaO). The following equations express the reaction of element
after reacted with sodium:-
Na + C + N →NaCN (nitrogen compounds)
2 Na + S → Na2S (sulfur compounds)
Na + X → NaX (halogen compounds, X = Cl, Br, ..)

Method:
1- A small portion of the finely powdered substance is mixed with a
small amount of powdered soda lime and the mixture is then rolled
within a small flattered piece of clean sodium metal.
2- The roll is introduced into a dry micro ignition tube and covered
with an amount of soda lime.
3- Heat the tube gently, near the outer post of the flame. When any
reaction sets in, remove the tube from the flame and wait till the
reaction subsides.
4- Heating of the ignition tube is continued gently until no more
reaction occurs.
5- Heating is then continued more strongly until the lower part of the
tube becomes red, then plunge the hot tube at once, into a beaker
containing about 10 ml of distilled water.
6- Crush the tube well, boil the contents of the beaker and break any
lumps present, then filter.
 
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7- The N, S and halogen should be detected in the alkaline filtrate

according to the following tests.
II-1-1- Test for Nitrogen:-

To carry out this experiment in a clean test tubes take about 1 ml of

the filtrate and 1 ml of FeSO4 solution followed by addition of one-two
drops only of concentrated H2SO4 the color of the solution turns to green or
blue indicted of the presence of nitrogen. If no nitrogen in the compound
the color will not change.

Equations:

Na + C + N → NaCN
FeSO4 + 2 NaOH→ Fe(OH)2 + Na2SO4
Fe(OH)2 + 6 NaCN → Na4[Fe(CN)6] + 2 NaOH
Sodium ferrocyanide
Na4 [Fe(CN)6] + Fe2 (SO4)3→ Fe4 [Fe(CN)6]3
Ferric-ferrocyanide (Prussian blue)

II-1- 2- Test for Sulfur:-

i) Sodium nitroprusside test.

In a clean test tube take about 1ml of filtrate then add about 1ml of
sodium nitroprusside the color changes to purple in case of the presence of
sulphur.

2 Na + S → Na2S
Na2S + Na2 [Fe(CN)5NO] → Na4 [Fe(CN)5] NOS

ii) Lead sulfide test.

In a clean test tube take about 1ml of the filtrate then add few drops
of acetic acid followed by 3 drops lead acetate, then shake the tube, the
black precipitate of lead sulfide indicates the presence of sulfur.
CH COOH
Na2S + (CH3COO)2Pb  
3
 PbS + 2 CH3COONa

 
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II-1-3- Test for Halogen:-

i) In the presence of (Nitrogen or Sulfur).

In a test tube take about 1ml of the filtrate then add about 1 ml of
conc. HNO3 then boil the mixture for few minutes. Cool the solution and
then add about 0.5 ml of silver nitrate (AgNO3) solution:-
 White precipitate formed in case of chlorine (Cl).
 Pale yellow precipitate formed in case of bromine (Br).
 Yellow precipitate formed in case of iodine (I).
ii) In the absence of (Nitrogen and Sulfur)
In a test tube, take about 1 ml of the filtrate then added about 1-2 ml
of dilute HNO3 followed by the addition of about 1 ml of AgNO3 solution
and observe the results as described above.
Equations:-
Na + X → NaX
NaX + HNO3 + AgNO3 → AgX + NaNO3
Where: X = Cl, Br, I

The results should be written as:

No N S X

II-2- Action of Heat (Ignition Test):

Place a little of the substance (0.1g) on a piece of porcelain and then

heat it over a low flame, raising the temperature gradually. Observe
whether the compound is volatile; inflammable (if so, nature of the flame);
or melts, decomposes or a residue is left. If a residue is left, it is cooled and
drop of distilled H2O is added to the residue and tested with litmus paper

 
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(or ph.ph. indicator). Then a little of dil. HCl is added to see whether any
gas is evolved or not.

Observation Inference
Aromatic compounds, or
a) Burn with a smoky flame
chorolhydrates
b) Burn with a non-smoky flame Aliphatic compounds
c) Melts, darkness, swells and then
Sugar
chars with odour of burnt sugar.
d) Chars without melting with the Starch, inulin, tartaric, citric, or their
odour of burnt sugar salts, lactate
Organo metallic compounds or
e) Residual ash
metallic salts of acids

II-3- Heating with Soda-Lime:

Mix well the compound (0.g) with powdered soda-lime (1g) in a dry
test tube, cover with a layer of soda-lime and heat first gently and then
strongly, notice any odour or change in colour.

Observation Inference
Ammonium salts of amides, imides,
a) Ammonia or ammonical vapours
nitriles, aliphatic amino acids or poly
evolved
nitro-compounds.
b) Chloroform produced (gives a
Chloral hydrates
characteristic odour while cold)
Carbohydrates or some aliphatic
c) Odour of burnt sugar
hydroxyl acid and their salts
Phenolic acids or phenyl esters of
d) Phenolic odour
carboxylic acids
e) Benzene produced Benzene carboxylic acid or their salts
Amine salts, anilinides or aromatic
f) Odour of aromatic amine
amino-carboxylic acids
g) Colour changes yellow to brown
Many aromatic nitro compounds
and then black

 
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Ammonia odour

RCOONH4 + NaOH / CaO RCOONa + NH3 + H2O

RCOONH2 + NaOH / CaO RCOONa + NH3

RCO
NH + NaOH / CaO RCOONa + NH3
RCO

Benzene odour

COOH

+ NaOH / CaO + Na2CO3

Aniline odour

NH3Cl

+ NaOH / CaO + NaCl + H2O

NH2 NH2

+ NaOH / CaO + Na2CO3

COOH
NHCOR

+ NaOH / CaO + RCOONa

Phenolic odour

OH OH
COONa
+ NaOH / CaO + Na2CO3

Chloroform odour

CCl3CHO.H2O + NaOH / CaO CHCCl3 + HCOONa

 
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II-4- Treatment with 20% NaOH solutions:

Add about 5 ml of 20% NaOH solution to the compound (0.1g).

Note the reaction while cold and then warm or heat if necessary.

Observation Inference

a) Dissolves on cold (warm) but Some aromatic acid or water

not soluble in water insoluble phenols.

Some polyhydric phenols,

b) Colour deepens on standing
aminophenone or benzoquinone

c) Ammonia evolves on cold Mostly ammonium salts of acids

d) Chloroform produced as oil Chloral or its hydrates

e) Oil or ppt. formed Salts of aliphatic or aromatic amines

Carbohydrates (sucrose and

f) Yellow to brown resin on starchgive only a faintly yellow
heating coloured product) or aliphatic
aldehydes (except HCHO)

g) Ammonia evolves on heating Amides, imides, nitriles or urea.

h) Oil or ppt. is formed on heating Anilides.

 
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II-5- Treatment with conc. H2SO4 acid:

To a small quantity (0.1g) of the substances contained in a dry test

tube, add about 1ml of conc. H2SO4 acid and note the reaction first while
the mixture is cold and then after it has been gently warmed.

Observation Inference

a) Evolution of CO and CO2

without charring but the mixture Citric acid and its salts.
turns yellow.

b) rapid charring with

effervescence ( evolution of CO Carbohydrates, tartarates, lactates.
and/or CO2 and SO2)

c) evolution of CO and CO2 but no

Oxalates
blackening

d) evolution of pungent vapour but

Many simple carboxylic acids and
no blackening and no marked
their salts
effervescence

e) blackening without
Phenols.
effervescence

CH2COOH CH2COOH
H2SO4
HO C COOH C O + CO + CO2 + H2O
CH2COOH CH2COOH
Acetone dicarboxylic acid

HO CHCOOH H2SO4
CH3COCOOH + CO2 + H2O
HO CHCOOH
Pyruvic acid

COOH H2SO4
CO2 + CO2 + H2O
COOH

 
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II-6- Nitration test:

Boil a portion of the solid with a mixture of equal volumes of conc.
HNO3 and H2SO4 acid in a dry test tube. When the brown fumes cease to
evolved. Pour the contents into a beaker containing about 30ml of water.
Nitration of liquid
Great care must be taken during nitration of liquids. Alcohol under
any conditions are not to be nitrated. Liquid in general, react more
vigorously and nitrated more easily than solids.
Procedures:
To one drop of the liquid add continuously one drop of conc. Nitric
acid and if necessary heat gently, then add another drop of acid, warm and
finally add 1 ml of conc. sulphuric acid, then proceed as usual.
Notes on nitration:
1. some aromatic compound are not nitrated under the above
experimental conditions (e.g. polyhydroxy compound: resorcinol,
gallic acid).
2. Other aromatic compound give rise to almost colourless nitro-
compounds which are not precipitated in the nitration medium (e.g.
some substituted and polycarboxylic benzoic acid, phthalic acid.
3. Some aliphatic compound (specislly some salt of aliphatic acids),
when subject to the nitration test give an indication of an unture
aromatic nature, e.g. iron and cobalt salts, when nitrated give
coloured solution (due to their cations) and lead and barium salts
give, on nitration white ppt. of the insoluble sulphates. In this latter
case reference is made to reaction π-2 (metallic residue).

Observation Inference

a) a yellow , orange or red coloured

Aromatic compound
solution, oil or ppt.

b) –ve Aliphatic compound

 
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II-7- Acidity test:

A cold solution of NaHCO3 is added to a cold solution of the

substance in H2O or alcohol or even to a suspension in H2O.

Observation Inference

A carboxylic acid, aniline salts, urea

a) Effervescence with evolution of
salts sulphonic acid or phenolic
CO2
compounds activated by nitro groups.

b) –ve Non acidic compound

RCOOH + NaHCO3 RCOONa + CO2 + H2O

SO3H SO3Na

+ NaHCO3 + CO2 + H2O

NH2 NH2

NH3Cl NH2

+ NaHCO3 + CO2 + H2O + NaCl

OH OH
NO2 NO2
NO2 NO2
+ NaHCO3 + CO2 + H2O

NO2 NO2

NH2CONH3NO3 + NaHCO3 NH2CONH2 + CO2 + H2O + NaNO3

 
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II-8- Solubility and Reverse precipitation :

Observation Inference

a) Soluble in 5% Na2CO3 soln. and reprecipitated by Aromatic

dil. HCl acid and not soluble in H2O. carboxylic acid

b) Soluble in 5% NaOH soln. (but insoluble in Na2CO3 Water-insoluble

soln. and H2O) and reprecipitated by dil. HCl acid. phenol

c) Soluble in dil. HCl ( not soluble in H2O) and

Organic base
reprecipitated by NaOH solution.

COOH COONa COOH

HCl
+ Na2CO3 + NaCl

Insoluble soluble Insoluble

OH ONa OH

HCl
+ NaOH

Insoluble soluble Insoluble

NH2 NH3Cl NH2

NaOH
+ HCl

R R R
Insoluble soluble Insoluble

 
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II-9- Acidity test:

Procedure:
To obtain a neutral FeCl3 soln., add a few drops of NH4OH soln. to
FeCl3 soln. till a permanent ppt. is formed and filter. Add a few drops of
this FeCl3 soln. to dilute soln. of the substance in water or in alcohol or
(neutral soln. in case of acids).

Observation Inference

Phenol, phenolic acids, esters

a) Intense purple, blue or red colour.
and amides

b) Buff ppt. (cleared by dil. HCl but

Aromatic acid and succinate.
replaced by a white ppt. of free acid.

Catechol, qunol or β-
c) Greenish colour of ppt.
naphthol.

d) Red colour (discharges on addition of Amino acids, formic or acetic

dil. HCl ) acids.

e) Greenish colour ( the substance in dil.

Some amines
HCl )

 
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REACTION OF CHARACTERISTIC GROUPS

III-Characteristic group having no additional elements:
Characteristic groups considered in this section contain C and H
(with or without O) elements.
III-1-Carbohydrates:
All the compounds of this class given for identification are solids and
are covered under the classes given below:
I-1-a-Monosacharides (glucose, galactose and fructose).
I-1-b-Disacharides (maltose, lactose and sucrose).
I-1-c-Polysaccharides (starch and inulin).

Glucose Galactose Fructose

Sucrose

Maltose Lactose

 
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Physical properties:
Colourless solids, which decomposes on heating and therefore have
no definite melting points. All except starch and inulin are soluble in water.
III-1-1-Molish's tes (characteristic test for carbohydrates):
Place 0.025g of the substance in a test-tube containing 2.5ml of H2O
and mix it with 2 drops of a 10% solution of 1-naphthol in ethanol. Allow 1
ml of conc. H2SO4 acid to follow down the side of the test tube (the acid
forms a heavy layer at the bottom) and allow to solution to stand for a
minute. If a carbohydrate is present a red-violet ring appears at the interface
of the two liquids : the colour quickly changes on standing or shaking, a
violet solution being formed. Shake and allow the mixture to stand for 2
minutes, then dilute with 5ml of H2O. In the presence of a carbohydrate, a
violet precipitate will appear immediately.

Glucose Hydroxyl methyl furfural

Identification of sugars:
III-1-2-Reduction:
a) Barfoed's reagent:(Cu acetate in acetic acid)
Heat a test-tube containing 1ml of the Barfoed's reagent and 1ml of a
dilute solution of the carbohydrate in a boiling water bath.

Observation Inference

Red Cu2O ppt. within 2 minutes Monosaccharides

Red ppt. after 5 minutes boiling Disaccharides
No red ppt. polysaccharide

Glucose Gluconic acid

 
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b) Fehling's solution:
(Fehling 1 : CuSO4. H2O and Fehling 2 : sod. Tartarate in NaOH soln.)
Place 3 ml of Fehling's solution(prepared by mixing equal volumes
of Fehling's solution of 0.1g of the carbohydrate in 2 ml of H2O and
continue the boiling for 1-2 minutes.

Fehling reagent

Observation Inference

A yellow-red Cu2O ppt. Reducing sugar

No red ppt. Non reducing sugar (sucrose)

Ketose Enediol Aldose

c) Bendict's solution:
This is a modification of Fehling's solution. To 3 ml of Bendict's
solution, add 2 ml of dil. soln. of carbohydrate, boil for 2 minutes and
allow to cool.

Observation Inference

A red Cu2O ppt. Reducing sugar

Clear solution Non reducing sugar

 
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Bendict reagent
d) Tollen's reagent ( Ammonical silver nitrate)
Add 1 ml of Tollen reagent to 1ml of carbohydrate soln. and heat
gently.

Observation Inference

A silver mirror or black ppt. on cold or Monosaccharide.

after gentle heating

A silver mirror on heating. Lactose or maltose.

-ve. Sucrose.

Glucose + (Ag(NH3)2)OH → Gluconic acid + Ag↓

III-1-3-Osazoneformation:
Carbohydrates react with phenylhydrazine to give crystalline
compounds known as osazone. Such compounds are used to identify the
different types of carbohydrates by observing the crystallineform of the
osazone under the microscope.
Dissolve 0.2 g of the carbohydrate, 0.4g of phenylhydrazine
hydrochloride and 0.6g of sod. acetate in 4 ml of water and dip the tube in a
beaker containing boiling water and leave it as such for 20 minutes then
place it a side to cool slowly. The osazone separates as a yellow crystalline
precipitate.

 
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HC NNHPh
Glucose or Fructose + PhNHNH2
C NNHPh

Osazone derivatives
Glucosazone or Fructosazone

 
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III-1-4-Rapid furfural test:

Add 1 ml of alc. 1-naphthol solution to 1 ml of the carbohydrate
solution and then add 6-8 ml conc. HCl acid and heat to boiling.

Observation Inference

Violet colour (immediately). Fructose or Sucrose

Violet colour (after a while). Glucose, Maltose or Lactose
-ve. Galactose.

III-1-5-Test for Fructose:

Upon heating a little fructose with dilute CoCl2 solution, cooling and
treat with a little ammonia solution.

Observation Inference

A violet purple colour, gradually fading. Fructose.

Green Co(OH)2 ppt. All other carbohydrates.

Identification of polysaccharide:
Starch
Starch occurs in plant tissues in the form of small white granules. It
is insoluble in water but it absorbs about 35% of a weight when subjected
to moist air.
III-1-6-Action of Iodine:
Starch may be identified by the blue colour with iodine ions. The
formed compound is easily decomposed and its colour is discharged on
heating and is restored on cooling.
III-1-7-Hydrolysis:
When starch is heated with dil. Acids, the so called soluble starch is
formed which gives on further heating gum-like materials known as
dextrin. The final hydrolysis products are maltose and glucose.

 
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Prepare a paste from starch and little water. Add to part of this 1/2 ml
of dil. Sulphuric acid and heat gently for 5 minutes. Test this solution every
now and then by taking one drop with a glass rod and adding it to iodine
solution. When no colour is produced, it shows that hydrolysis of starch to
glucose is complete.

III-2-Carboxylic acids:

All acids give a positive acidity test as they contain a carboxylic

group. They react with NaHCO3 or Na2CO3 to give the sodium salt of acid
with evolution of CO2 gas. The occurrence of effervescence denotes the
presence of a free carboxylic group.

III-2-1-Acidity test( characteristic test for acid):

Add 1 ml of NaHCO3 solution to 1 ml of an aqueous or alc. Solution

of compound (unknown). Effervescence take place due to the evolution of
CO2 gas.

RCOOH + NaHCO3 → RCOONa + CO2↑ + H2O

III-2-2-Ester formation test:

To 0.5 g of the compound, add abs. ethanol (1ml), a few drops of
conc H2SO4 and heat the content, in a dry test tube, gently for 1 minute.
Note the fruity smell of the resulted ester.

RCOOH + C2H5OH → RCOOC2H5 + H2O

III-2-3-Iodate-iodine test:
Take 4-5 drops of an aqueous or alc. soln. of the compound in a test
tube, add 2 ml of 2% KI solution and 2 ml of 4% KIO3 soln. into it and
keep the test tube in a boiling water bath for 1-2 minutes, then cool and add
3-4 drops of starch solution. A blue or violet colour indicates the presence
of COOH group.

5KI + KIO3 + 6 RCOOH → 3I2 + 3H2O + 6 RCOOK

 
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Identification of acids
Acids may be aliphatic or aromatic.

A) For aliphatic acids:

Malonic acid Fumaric acid Maleic acid Succinic acid

Oxalic acid Tartaric acid Citric acid

III-2-4- Effect of heat on solid:
Odour of acetic acid. (Solution of solid in acetic anhydride
→yellowish red solution with green fluorescence)
Malonic acid

III-2-5- Neutral solution + neutral FeCl3:

Observation Inference

Red colour turn brown Maleic acid ( unsaturation test +ve , fluorescein
ppt. by boiling. test, red colour).

Red brown ppt. Fumaric ( unsaturation test +ve , fluorescein

test, red colour).

Buff ppt. Succinic acid (fluorescein test green)

Yellow colour Oxalic , tartaric or citric acid

 
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Notes:
1-Neutral solution of an acid:
Add ammonia solution in excess to a small amount of the solid acid
or its solution. Heat until no more ammonia gas evolved. The residual is the
neutral solution.

2- fluorescein test:
In a clean dry test tube, fuse carefully together, few crystals of
resorcinol and an equal quantity of the solid acid, or its anhydride
moistened with 2 drops of conc. H2SO4. Pour into excess NaOH.

HO OH HO OH HO O OH

H H conc. H2SO4 C
O
O
C
C
O
O
C

O OH H

O O O

COO

Green f luorrescence

 
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III-2-6- Neutral solution + neutral CaCl2:

Observation Inference

White ppt. immediately on cold insoluble in Oxalic acid

acetic acid.

White ppt. after scratching and warming, Tartaric acid (Fenton's test)
soluble in acetic acid.

White ppt. after boiling, insoluble in acetic Citric acid (Deng's test)
acid

Deng's test:
Add 1 ml of Deng's No. 1 to 3 ml of neutral citric acid and heat to
boiling. Add 2 drops of Deng's No. 2 and note that the colour of pot.
permanganate is discharged. (redox reaction).
Fenton's test:
Add 1 drop of FeSO4 soln. to 2 ml of tartaric acid soln. then add
dropwise H2O2 soln. till the solution acquires a green colouration. Add
excess of NaOH soln. a violet colour is produced. (redox reaction).
B) For Aromatic Acids:

Salicylic acid Phthalic acid Cinnamic acid

Phenylacetic acid Benzoic acid

 
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III-2-7-Neutral solution + neutral FeCl3:

Observation Inference

Violet colour. Salicylic acid

Buff ppt. Phthalic , Cinnamic, Phenylacetic or Benzoic acid

Amm. Benzoate Ferric benzoate Basic Ferric benzoate

To differentiate :
III-2-8-Phthalein test:
In a clean dry test tube, fuse carefully together, few crystals of the
acid and 2 drops of phenol, moistened with 2 drops of conc. H2SO4. Pour
into excess NaOH, if red colour, then phthalic acid.
HO OH HO OH

H H conc. H2SO4 C
O
O
C
C
O
O
C

O OH H

O O

COO

Red Colour

 
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III-2-9-Fluorescein test: +ve phthalic acid

III-2-10-Unsaturation test:
If the solution of the substance is sod. carbonate discharge the colour
of bromine water and also a few drops of dilute KMnO4. If +ve, then
cinnammic acid

III-2-11-Boiling with test:

If oily drops are formed, then phenyl acetic acid. If all the above tests
(phthalein, fluorescein, unsaturation and boiling with water) are negative,
then Benzoic acid.

III-3-Hydroxyl compounds (phenol and alcohols):

The test for –OH group may be accomplished in any one of the
following methods, provided that –COOH group is absent.
III-3-1-Acetyl chloride test:
A portion of the substance (solid or liquid) is added to 1 ml of
CH3COCl, in a dry test tube, a vigorous evolution of HCl gas indicates the
presence of –OH group.

III-3-2-Benzoyl chloride test:

Benzoyl chloride may be used instead of CH3COCl, but warming is
necessary in this case.

 
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III-3-3-FeCl3 Test:
Hydroxyl group of phenols and enolic compound gives colour
(usually violet) with FeCl3. This test is valid in presence of carboxylic
group.
Add few drops of neutral FeCl3 solution to a solution of the
substance in water or in alcohol (note the colour produced).

HO OH + FeCl3 O O

Benzoquinone
OH HO

OH

2 + FeCl3

2-Dinaphthol
OH
HO OH

2 + FeCl3

III-3-4-Sodium metal Test:

Clean pieces of sodium are added to small amount of dry ether or
benzene until evolution of H2 ceases if present. Then add a little of the
substance, if bubbles of H2 gas evolve, -OH group is present.

Identification of Phenols:

 
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Water soluble phenols

Catechol Resorcinol Quinol Pyrogallol

Aq. or alc. soln. of phenol + Neutral FeCl3

Observation Inference

Violet discharged by HCl. phenol

Violet discharged by AcONa. Resorcinol

Green turns red by NaOH. Catechol

Green crystals Hydroquinone

Red-turns violet by NaOH. Pyrogallol.

Greenish turns to violet α-Naphthol.

Faint green β-Naphthol.

III-3-5-Liebermann-nitroso reaction:

Phenol react with nitrous acid to give p-nitrosophenols which

condense with excess phenol to give indophenols (usually red). When
solution of the latter is rendered alkaline, a blue or green anion is liberated.
To a small amount of NaNO2 in a clean dry test tube, add 0.5g of
phenol and heat gently for 1 min., cool, add 1 ml of conc. H2SO4 with
shaking. A deep green or blue colouration develops (sometimes only after
1-2 min.). Dilute cautiously, the solution, turns red. Now add an excess of
NaOH soln., the green or blue colouration reappears.

 
‐ 28 - 
  

Phenol H2SO4 Dilution NaOH

phenol Green or blue Red Green or blue
α-Naphthol. Green Ill-defined Ill-defined
β-Naphthol. Brownish back Ill-defined Ill-defined
Catechol Deep green Dirty brown Red
Resorcinol Deep blue red brown

HO + HONO HO NO

C6H5OH

NaOH
NaO N O HO N O

III-3-6-Chloroform and NaOH (Reimer-Timann reaction):

Dissolve the phenol in conc. NaOH (20%), add 1-2 ml of chloroform
and heat gently. Notice the colouration produced.

Phenolic comp. colour Phenolic comp. colour

phenol No-colour Catechol Green

α-Naphthol. Deep blue fading Resorcinol Red with
to green fluorecence
β-Naphthol Deep blue fading quinol -ve
to green

Phenol Salicylaldehyde

 
‐ 29 - 
  

III-4-Metallic salts of acids:

III-4-1-Test for metallic residue (Action of heat):
Heat a small portion of the substance on a piece of porcelain, leaves
a residue, when dissolved in dil. HCl, CO2 evolves with effervescence. In
this case a metallic salt of acid is present.

a) White residue; Alkali metal in the form of carbonate or alkaline

earth metal left as oxide.
b) Red or yellow residue, lead oxide.
c) Yellow residue when hot and white when cold, zinc oxide.
d) Black residue, CuO, Co2O3, Fe2O3.
The metallic residue can be further identified by dissolving it in dil.
HCl acid and searching for the basic radical in the usual way.
III-4-2-Nitration test:
-ve salt of aliphatic acid
+ve salt of aromatic acid
Identification of salt of an aliphatic acid:
III-4-3- Treatment with FeCl3:
i) Red colour turns to brown by boiling.
Formate : Reduces amm. AgNO3.
Acetate : Esterify to ethylacetate.
HCOONH4 + FeCl3 HCOO Fe HCOOFe(OH)2
3
Basic ferric formate
ii) Carry out iodoform test, if +ve then lactate.
iii) Follow then the same steps as in the case of free aliphatic acids.
Identification of salt of an aromatic acid:
Follow the same steps as in the case of free aromatic acids

 
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III-5-Aldehydes:

III-5-1-Schieff's reagent:
Add 2-3 drops (if the substance is a liquid) or 0.1 gm of the solid to 2
ml of Schiff's reagent. A pink colour developes within 2 minutes. (Schiff's
reagent must not be warmed, nor alcohol must be used as a solvent).
Note :
1. Schiff's reagent (rosaniline hydrochloride (magenta) is dissolved
in water (deep-red solution) and SO2 is passed in until the
magenta colour is discharged.
2. The reaction of aldehydes with Schiff's reagent is not very
characteristic due to the reducing properties of the aldehydes.

Schiff reagent
III-5-2-Tollen's reagent:
Aldehydes reduce Tollen's reagent (ammonical AgNO3 soln.) to
metallic Ag.
Place 1 ml of Tollen's reagent in a clean test tube, then add a small
portion of the aldehyde. Warm the reaction mixture and notice the
formation of a silver mirror (or ppt.) on the inner wall of the tube.
III-5-3-Fehling's solution:
Aldehydes reduce Fehling's soln. to a red cuprous oxide.
R-CHO + Cu2 RCOOH + Cu

 
‐ 31 - 
  

Mix together two equal volumes of Fehling 1 and Fehling 2. Heat the
mixture then add 1 ml of the aldehyde and reheat gently. Note the
formation of a yellow-red ppt. of Cu2O.
N.B. Reaction with Tollen's reagent and Fehling's solution is characteristic
only for aldehydes.

III-6-Ketones:

In the absence of aldehydes, ketones may be detected by formation

of derivatives such as oxime, phenylhydrazone or 2,4-
dinitrophenylhydrazone.
III-6-1-Formation of 2,4-dinitrophenylhydrazone:
For detection, add 0.05-0.1 g of the substance to be tested to 3 ml of
the 2.4-dinitrophenylhydrazine reagent, and shake. If no precipitate forms
immediately allow to stand for 5-10 minutes. A crystalline precipitate
indicates the presence of carbonyl compound. Occasionally the precipitate
is oily at first, but this becomes crystalline upon standing.

 
‐ 32 - 
  

Identification of ketones:

Acetophenone Benzophenone
O

O O

p-Benzoquinone Anthraquinone
III-6-2-Boiling with water:

Observation Inference

Oily drops benzphenone

No oily drops p-benzoquinone or anthraquinone

III-6-3-Tollen's reagent:
Add 1 ml of Tollen's reagent to a few drops of cold aqueous solution
of ketone, give only, black ppt. or silver mirror with p-Benzoquinone.
III-6-4-Confirmatory tests:
For p-benzoquinone
a) Dissolve 0.2 g of benzoquinone in 1 ml of alcohol, add 2 drops of
aniline and warm gently, a reddish ppt. of quinine-dianiline is
formed.

Quinonedianiline

 
‐ 33 - 
  

b) Reduction (to quinol):

Add aq. benzoquinone solution to KI solution acidified with few
drops of H2SO4, I2 is librates (test by starch).
For anthaquinone:
III-6-5-Reduction with Zn dust:
Heat 0.2 g of anthraquinone with about 5 ml of NaOH solution add a
small amount of Zn dust, a red colouration is produced. When all Zn dust
has dissolved, the solution is slowly decolouration on shaking (due to
atmospheric oxidation).

Oxanthranol

III-7-Hydrocarbons

Naphthalene Anthracene
Many hydrocarbon form molecular compound with picric acid, for
example, naphthalene picrate C10H8-C6H2(NO2)3OH.
III-7-1-Picrate test:
Dissolve 0.1 g of the hydrocarbon and 0.2 g of picric acid separately
in the minimum volume of benzene (about 2 ml), mix the two solutions and
allow to cool. Filter and wash with 2 ml of ethanol.

 
‐ 34 - 
  

Observation Inference

Yellow crystals Naphthalene (150 ºC)

Red crystals Anthracene (138 ºC)

III-7-2-Nitation test:
Nitric acid reacts with naphthalene, in the presence of glacial acetic
acid to give a nitronaphthalene while anthracene is not readily nitrated with
nitric acid.
Dissolve by heating, 0.5 g of naphthalene in 3 ml of glacial acetic
acid, then cool and add 1ml of conc. HNO3 and heat the mixture gently for
1 minute. Cool and pour the solution into a beaker which contains about 25
ml of cold H2O and note the separation of nitronaphthalene as a yellow
solid (1-,1,8-and 1,5-nitronaphthalene).

 
‐ 35 - 
  

IV- Compounds containing C, H,O and N

This class includes ammonium salts of acids, amides, imides, amines,
anilides, nitro compounds and amino acids.
IV-1- ammonium salts of acids:
IV-1-1- Grind in amorter, aportion of the substance with solid Na2CO3,
moisten wiyh afew drops of H2O, ammonia evolves.
IV-1-2- Add NaOH solution to the solid substance in a test tube, ammonia
is given on cold. The above two tests indicates ammonium salts of acids
now detect and identify the acid radical of the amm. salts as under metallic
salts of acids (III-7).
IV-2- Amides, imides and nitriles:
IV-2-1- Boiling with 30% NaOH solution:
Boil the compound, in a test tube with 30-40% NaOH solution (3-4 ml)
evolution of ammonia indicates the presence of an amide, imide or nitrile
group.
RCONH2 + NaOH RCOONa + NH3

R(CO)2NH + 2NaOH R(COONa)2 + NH3

RCN + NaOH + H2O RCOONa + NH3
Amides liberate ammonia comparatively rapidly, whereas nitriles require
longer heating.

Identification of Amides, imides and nitriles:

 
‐ 36 - 
  
CONH2

CONH2
CH3CONH2
CONH2

Oxamide Benzamide
Accetamide

H2N C NH2
NH2CONH2
S
Urea Thiourea
O
O

C
H2C
NH
NH
H2C
C

O
O

Succinimide Phthlimide
IV-2-2- hydrolysis of amides, imides and nitriles:
Place about 2g of the substance in a round bottom flask and 20 ml of
20% NaOH fit the flask with a reflux condenser and boil for about 0.5 h or
more. cool the flask and add excess dil. H2SO4 while cooling:
a) A white ppt. indicates an aromatic acid, filter, wash with cold water
and identify the acid as usual (amide of benzoic or phthalic acid).
b) No ppt. is formed. Then the acid is aliphatic, it is identified in the
usual way. (Acetic acid, Oxalic or succinic acid).
c) A brisk evolution of CO2 occurs but no ppt. indicates urea.

CO(NH2)2 + 2NaOH Na2CO3 + NH3

 
‐ 37 - 
  

Confirmatory test:
IV-2-3- Biuret test (for urea and its salts):
Place 0.5 g urea in dry test tube, heat gently for 1-2 minutes and cool the
residue.
2CO(NH2)2 H2NCONHCONH2 (biuret) + NH 3
Dissolve the formed biuret in a few ml. of warm 10%NaOH.by cooling and
adding 1 drop of very dilute CuSO4 solution, a purple or violet colouration
is obtained.
HN
H2N HN
O
O OH
HN Cu
CuSO4
HN HN
O
O OH
HN
H2N HN Purple colour
For differentiation
Acidity test
observation Inference
-ve Urea
+ve Urea salts

- Aq. Soln + CaCl2

Urea oxalate give white ppt. insol.in ACOH acid

IV-2-4- For Oxamide:
Biuret test: shake 0.1 g of oxamide with 1 ml of 10% NaOH solution, add
1-2 drops of very dilute CuSO4 soln. and mix well. A pink colouration is
produced
IV-2-5- For succinimide:
It gives positive fluorescin reaction.
IV-2-6- For phthalimide:
It gives positive phthalein and flurescin reactions.
IV-3-primary aromatic amines:
IV-3-1- diazotization and coupling with β-naphtho:
Add dil. HCl to the substance in the test tube (the substance does not need
to dissolve completely in HCl). Cool in ice, then add cold alkaline solution
of β-naphtho in excess, a brilliant red dye indicates primary aromatic
amines.

 
‐ 38 - 
  
HO

NH2 N NCl N N

NaNO 2 + HCl B Naphthol

NaOH

Benzenediazonium chloride Azo dye

IV-3-2- test for nitro group in the presence of amino group
Diazotize the amine as before. Stir the solution from time to time, wait for
10 minutes, then take a portion of the diazotised solution in a test tube and
boil it till all N2 gas is evolved. Cool in ice and couple (small amount)by
adding alkaline solution of b-naphthol, if a colour or ppt. is formed, this
indicates that the amino group is not removed completely. Continue
diazotizing step for the original solution by adding HCl and NaNO2 again
boil , then test a portion by coupling with β-naphthol soln. of no dye ia
ginen, then diazotization is complete. Next, boil the original soln. with Zn
and HCl for 10 minutes at least. This step destroys the diazonium
compound and reduces the nitro group to amino group. Cool again and
diazotize as before and couple, a red dye indicates the presence of a nitro
group.
NH2
N N OH

-H2O -N
O N OH

NO2
NO2
o-Nitrobenzenediazonium hydroxide

OH OH

Zn/HCl
(test for NH2)

NO2 NH2
Differentiation between nitro anilines
All nitro anilines are coloured yellow or orange compounds and are
soluble in hot water.

 
‐ 39 - 
  

NH2 NH2
NH2

NO2

NO2

NO2
o-Nitroaniline m-Nitroaniline p-Nitroaniline
IV-3-3- formyl derivatives as in IV-3-8.
o-nitri aniline m.p. 122oC
m--nitri aniline m.p. 134oC
p--nitri aniline m.p. 194oC
IV-3-4- test for –COOH group in the presence of amino group (acidity
test)
COOH COOH
COOH

NH2

NH2

NH2
o-Aminobenzoic acid m-Aminobenzoic acid p-Aminobenzoic acid
Anthranilic acid
The above acids are sparingly sol. In cold H2O, sol. In minerl acids and
caustic alkalies, dissolve readily in alcohol.
IV-3-5- Confirmatory test:
a) Give aniline on heating with soda-lime
b) Give diazo-derivatives which couple with alkaline β-naphthol to give
an azo-dye.
c) Decompose on heating to give the corresponding phenolic
derivatives e.g. o-aminobenzoic acid yields salicylic acid.
d) Amide derivatives

Dissolve or suspend the requisite acid chloride of amino benzoic acid

in 5-10 ml. of dry ether or benzene and add concentrated ammonia
solution and warm for a few minutes in a water-bath. If no solid
separates, evaporate the solvent. Recrystallize the amide from water
or dil. Ethanol.

 
‐ 40 - 
  

If IV-3-2- and IV-3-4- are negative then detect for unsubstituted

aromatic amino compounds.
Differentiation between primary amines.
NH2
CH3

NH2

NH2
p-Toluidine Naphthylamine Naphthylamine
IV-3-6- boiling the solid substance with water:
observation Inference
No oil drops β-naphthylamine
Oily drops P-toluidine or α-naphthylamine

IV-3-7- amine in dil. HCl+FeCl3

observation Inference
Blue colour α-naphthylamine
-ve P-toluidine

Confirmatory test:
IV-3-8- formyl derivatives:
Formic acid condenses with primary and secondary aminesto yield
formyl derivatives:
ArNHR + HCOOH ArN(CHO)R + H2O
Reflux 0.5 g. of the amino with 5 ml. of 90% formic acid (caution in
handling) for 10 minutes, and dilute the hot solution with 10 ml. of cold
H2O. cool in ice and, in some cases, saturate with salt if the derivative
does not separate immediately. Filter, wash with cold H2O and
recrystallize from water, ethanol or light petroleuim.
P-toluidine (formyl) m.p. 53oC
α-naphthylamine m.p. 139oC
β-naphthylamine m.p. 129oC
IV-3-9- Action of bleaching powder (oxidation)
Shake 1-2 drops of amine with 10 ml. of H2O and add a few drops of
bleaching powder soln.(NaOCl), characteristic colour appears.
observation Inference
Yellowish colour P-toluidine
Pale brown α-naphthylamine

 
‐ 41 - 
  

-ve β-naphthylamine

IV-4- secondary aromatic amines:

IV-4-1- reaction with nitrous acid:
Nitrosamines are formed, these are usually yellow oils solids of low m.p.
and give the Liebermann's nitroso reaction.
IV-4-2- Liebermann's nitroso reaction:
Dissolve 0.1 g.of the secondary amine (unknown) in 3-5 ml. of dil. HCl or
ethanol (in the latter case, add 1 ml. of conc. HCl acid). Cool to about 5oC
and add 4-5 ml. of 10% NaNO2 soln. and allow to stand for 5 minutes.
Add10 ml. of water, transfer to a small separator funnel and extract the oil
with about 20 ml. ether. Wash the ethereal extract successively with water,
dil. NaOH solution and water. Remove the ether on a wormed water bath.
Place 1 drop or 0.01- 0.02 g. of the extracted nitroso compound in a dry test
tube, add 0.05 g. of phenol and warm together for 20 seconds, cool, and
add 1ml. of conc. H2SO4 acid. And an intense green (or greenish-blue)
colouration will be developed, which changes to pale red upon-pouring in
to 30-50 ml. of cold H2O the colour becomes deep blue or green upon
adding excess of NaOH solution.
R2NH + HONO R2N-NO + H2O

H2SO4 C6H5OH
R2N-NO HONO HO NO

C6H5OH
HO N O

Indophenol

NaOH NaO N O

Diphenylamine (C6H5)2NH, m.p. 54oC

IV-4-3- Reactions:
a) Dissolve a few crystals of diphenylamine in 1 ml. of conc. H2SO4 or
H3PO4. Add 2 drops of conc. HNO3 to about 10 ml. of H2O. shake
and add one drop of this latter solution an intense blue colouration is
produced.
b) With FeCl3,diphenylamine give green colour

 
‐ 42 - 
  

c) N-Nitrosodiphenylamine Dissolve 0.5 g of diphenylamine in about 5

ml. of warm alcohol, then add 0.5 ml. HCl, cool in ice and add, drop
by drop, a cold soln. of 3 g of NaNO2 in 10 ml. H2O. leave the
reaction in ice for 4 minutes and then filter off the yellow crystals
which have separated, wash with water and carry out liebermann's
reaction as described before.
d) Simon's test:
To a suspension of diphenylamine (0.001 g.) in H2O (3 ml.),
add acetaldehyde (1 ml) and then 1% aq. Soln. of nitroprusside (2-3
drops). Adeep blue colour is developed with in5 min. which changes
gradually through greenish blue to pale yellow.
e) Carbon disulphide reagent test:

Prepare a soln. of diphenylamine (0.015 g) in dil. HCl (5.ml.) in a

test tube. Place the reagent (1 ml.) in an anther test tube, add conc. NH3
soln. (0.5-1 ml) and the amine soln. (0.5-1ml). a white (brown) colour
produced.
R2NH + CS2 + 2NH4OH + NiCl NH4Cl + H2O + R2N C S
2Ni

N.B. test for thee nitro group

IV-5- Tertiary Amines:

N(CH3)2 N(C2H5)2

N,N-Dimethylaniline N,N-Diethylaniline Triphenylamine

IV-5-2- Reaction with nitrous acid:
The dialkylanilines yield green solid P-nitroso compounds.

(H3C)2N
HONO (H3C)2N NO + H2O

IV-5-2- Nessler's reagent test:

 
‐ 43 - 
  

In a test tube dissolve the compound (0.1 g) in dil. HCl and add into it
Nessler's solution. the appearance of a white ppt. indicates
tertiaryamines.
IV-5-3- action of nitric acid:
Dissolve a few crystals of amine in 1 ml. of conc. H2SO4. Add 2
drops of conc. HNO3 to about 10 ml. of H2O. shake, and add one
drop of this solution to the former one,
Deep yellow colour Dimethylaniline
Intense greenish-blue Triphenylamine
IV-6- Anilides R-CONHC6H5:
O

NHCOCH3 HN C

Acetanilide Benzanilide
IV-6-1- Hydrolysis with 70%H2SO4 test

Hydrolyse about 2 g. of the anilide with 20 ml. of 70% H2so4 (4

volumes of conc. H2so4 to 3 volumes of water) in a small flask fitted.
with a reflux condenser and boil for 20-30 minutes. In case of
acetanilide. distill the acetic acid, and test for aniline in the distilling
flask, while in case of benzanilide filter the ppt. (benzoic acid), purify
from any unchanged anilide by dissolving in Na2Co3 , filter then acidify,
and refilter the ppted .acid (then identify both amine and acid as usual).
IV-6-2- Anilides Give the Liebermann's nitroso reaction for
secondary amines (see IV-4-2)
IV-6-3- Dichromate test:
Place the compound (0.001 g) in a dry test tube, add conc. H2SO4, shak
the contents vigorously and then add finely powdered K2Cr2O7 (0.05-
0.07 g). the immediate appearance of a bluish pink or deep blue colour
indicates anilide.
This is a positive test for those anilides which do not have
substituents in the ring.
IV-7- Test for Nitro Aromatic Compounds:
Nitro compounds are usually coloured, yellow to red. They react
vigorously with metallic sodium during the element test.

 
‐ 44 - 
  

IV-7-1- Reduction to the hydroxylamine with neutral reducing

agent:
Dissolve 0.5 g. of the substance in 10 ml. of 50% ethanol, add 0.5 g. of
solid NH4Cl and about 0.5 g. of Zn powder. Heat the mixture to boiling
for 5 minutes. Filter from axcess of Zn powder and test the filtrate with
Tollen's reagent. An immediate black or grey ppt. or a silver mirror
indicates the presence of –NO2 group (Fehling's soln. may be used).
Make sure that the original compound does not affect the reagent used.
RNO2 + 4H RNHOH + H2O

RNHOH + 2[Ag(NH3)2]OH RNO + 2Ag + 4NH3 + 2H2O

IV-7-2- Reduction to amino group:
NO2-group on reduction, is transformed to the primary amino group
which can be detected by diazotiasation and coupling.
Place about 0.5 ml. or 0.5 g. of the nitro compoundin small beaker, add
5 ml. conc. HCl and few pieces of granulated tin or Zinc. Warm gently
with stirring till all the compound has disappeared. Filter and pour into
about 20 ml. of water. Cool, carry out diazotiasation test and couple
with alkaline β-naphthol.
In order to separate the base, cool the solution after dilution, then add
NaOH soln. till alkaline and extract with ether. Separate the ethereal
layer and evaporate it, the amine is left behind. Identify as usual.
RNO2 RNH2 + H2O

IV-7-3- compound containing one or more NO2 group will oxidize

Fe(OH)2, the colour change is from green to brown.
RNO2 + 6Fe(OH)2 +4H2O RNH2 + 6Fe(OH)3
For differentiation
IV-7-4-compound soln. + FeCl3
observation Inference
Aviolet – red colour Nitro-phenols
-ve Mono-or dinitro hydrocarbons

IV-7-5-Nitro-Phenols:

 
‐ 45 - 
  
OH

OH

OH

NO2 NO2 NO2

o-Nitrophenol m-Nitrophenol p-Nitrophenol

The presence of the nitro-group renders the phenol acidic enough to

dissolve in NaCO3 soln. with the evolution of CO2.
o-Nitrophenol:
Bright yellow needles, m.p. 46oC, its oddour resembles both that of phenol
and of nitrobenzene, volatile in steam,sparingly soluble in cold water, sol.
In hot water.
m-Nitrophenol:
m.p. 95oC, sol. In hot water, non volatile in steam.
p-Nitrophenol:
yellow prisms, gradually turns red in light, m.p. 114oC, very sol. In hot
water.
General reaction:
a-NaOH:
the dissolve in sodium hydroxid solution giving on orange soln. the
colour is discharged an adding an acid, but is restored on making the soln.
alkaline
b- Na2CO3:
they dissolve in sodium carbonate solution with the evolution of
CO2
c- FeCl3:
when a few drops of FeCl3 are added to the nitrophenol solution, a
violt-red colour is produced. O-Nitrophenol does not give a colouration.
d- They do not give the liebermann's nitrose reaction.
e- Reduction:
they are reduced (as befor) to amino phenols (diazotize and couple).
The solution after reduction does not give a colouration when rendered
alkaline with NaOH.
IV-7-6- Nitro Hydrocarbons:

 
‐ 46 - 
  
NO2

NO2 NO2

Nitrobenzene m-diNitrobenzene
Nitrobenzene:
A pale yellow liquid, insol. In water and heavier than water, with a
characteristic odour of bitter almonds. On reduction as before it gives
aniline which can be identified as usual.
m-Dinitrobenzene:
A pale yellow solid, m.p. 90oC, insol. In water
General reaction:
a- Acetone alkali colouration: dissolve a few crystals in 1-2 ml. of
acetone, then add a few drops of aqueous NaOH solution, adeep
colouration is produced, and it turns red by acetic acid, but destroyed
by mineral acids.
b- It is reduced in acid solution to m-phenylenediamine NH2C6H4NH2
which:
1) On diazotization and coupling, a deep-brown soln. or a brown
ppt. of the dye Bismark brown appears.
2) When a few drops of FeCl3 is added to a very dil. Soln. of the
base in water, a reddish colouration appears.

IV-8- Test for Amino acid:

H
H2N C COOH

R
IV-8-1- Ninhydria test:
Place 2-3 drops of 0.2%aq.soln. of ninhydrin on a piece of filter paper
followed by drying it in air (or oven).Drop aq. Soln. of amino acid (1
drop) on this paper and redry it for afew minutes, Ablue (sometimes violet)
spot or ring appears on the paper.

 
‐ 47 - 
  
O

CH(OH) H -H2O
H2N C COOH
-CO2
C
R
O
Ninhydrin
O O

C C

H2O
C NCH2R C N CHR

C C

O OH
O

Ninhydrin
C NH2 RCHO -2H2O
C

OH O O

C C

C N C

C C

OH O

Blue-violet

 
‐ 48 - 
  

V- COMBINATION OF SULPHUR
This class includes: alkaline bisulphate addition compounds of
aldehydes and ketones, alkyl sulphates, addition compounds of amines with
sulphuric acid, sulphonic acid, sulphonic acids and their salts
V-1- Alkaline bisulphate addition compounds of aldehydes and ketones
:
Heat the solid substance in a crucible, note the odour of the evolved
aldehyde or ketone . the residue left in the crucible is tested for sulphite and
sulphate. (add dil. HCL, SO2 gas is evolved known by its odour of burnt
sulphur and turns acidified dichromate paper green).
R-CH (OH)SO3Na + HCL RCHO + NaCL + SO2 + H2O
V-2-Ionisable sulphate:
Dissolve the substance in water (or alcohol) and add barium nitrate soln. a
white ppt. of barium sulphate, indicates ionisable sulphate, e.g. aniline
sulphate (it must be taken into consideration that some salts of acids which
are sol. In water may pptate the barium salt of the acid).
C6H5NH2H2SO4 + Ba(NO3)2 C6H5NH2 + BaSO4 + 2HNO3
V-3- hydrolysable sulphate:
Boil the substance with NaOH soln. then acidify the soln. strongly with
nitric acid ( if a ppt. appears, filter and use the filtrate) and add barium
nitrate soln., awhite ppt. indicates hydrolysable sulphate.
S=C(NH2)2 + 2H2O CO2 + H2S + 2HN3
V-4- sulphonic acids:
If the above tests are negative, the sulphur present is directly linked to the
aromatic nucleus, most probably sulphonic acid group or fuedthe substance
with solid sodium hydroxide. Sulphonic acid group is decomposed into
sodium sulphite and sodium sulphate. Dissolve in water, acidify with dil.
Acid, add barium nitrate solution, a white ppt. is produced.
OH
SO3H

NaOH NaHSO3 Na2SO4

BaCl2
NH2
NH2
NaCl + BaSO White ppt.

HCl
NaCl + H2O + SO2
V-5-sulphonamide group (-SO2.NH2):

 
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Fuse the compound (0.5 g.) with KOH (2 g.) in a tube for 10 minutes and
bring a filter paper moistened with saturated ethanolic 2,4-
dinitrochlorobenzene solution. The appearance of a strong yellow colour on
the paper indicates sulphonamide.
Fusion with 80% H2SO4 at 160-170 o results in rapid hydrolysis.
ArSO2NRR' + H2O ArSO2OH + RR'NH.H2SO4
The reaction product may then be examined for asulphonic acid and an
amine.
For differentiation:
SO3H

NH2

H2SO4 (NH2)2C=S

NH2
Aniline Sulphate thiourea Sulphanilic acid
a) Acidity test:

observation inference
Aniline sulphate or Sulphanilic
+ve
acid
-ve thiourea

b) Compound soln. +BaCl2:

observation inference
A white ppt. insol. In dil. acids Aniline sulphate
-ve Sulphanilic acid
 
c) Azo-dye test

observation inference
Deep yellow orange Sulphanilic acid
Scarler red Aniline sulphate
d) Mercury oxide removes a molecule of H2S from
thiourea at room temperature to form cyanamide.

(NH2)2CS + HgO H2NCN + HgS + H2O

Black

 
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e) Oxidation with alkaline

KMNO4 (removes its violet colour) converts thiourea into urea.
(NH2)2C=S + [O] (NH2)2C=O + S yellow

VI- COMBINATION OF SULPHUR

This class includes: halogenated aldehydes and ketones, e.g. chloral
hydrate, acid halides, halogenated hydrocarbons, acids or phenols.
VI-1- ionisable halogen:
Dissolve the substance in water (or alcohol), add AgNO3 soln., a ppt.
indicates ionisable halogen e.g. amine salts, acid chlorides.
N.B.: acid chlorides specially aliphatic, are very easily hydrolysable in
contact with water, that they give ppts. When treated with AgNO3 and
appear as if they contain ionisable halogen.
C6H5NH2.HCl + AgNO3 C6H5NH2 + AgCl + HNO3
VI-2- Hydrolysable halogen (aliphatic combination):
Hydrolyse by boiling the substance with 30% NaOH soln. for some time,
cool, acidify with dil. Nitric acid (filter from any ppt. formed). And add
AgNO3 soln. a ppt. indicates a hydrolysable halogen (halogen attached to
an aliphatic linkage or to an aromatic side chain). e.g. chloralhydrate.
VI-3-Non-hydrolysable halogen (aromatic combination):
IF the above tests are negative then the halogen is in aromatic linkage
(directly linked to an aromatic nucleus).
For Differentiation:
NH2

CCl3CHO.H2O CHI3

Aniline hydrochloride Chloralhydrate Iodoform

a) Acidity test:

observation inference
+ve Aniline hydrochloride
-ve Iodoform or chloral hydrate

b) Action of heat: violet fumes Iodoform

c) Treatment with NaOH: CHCl3 evolves. Chloralhydrate
d) Reduction (for chloral hydrate):

 
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CCl3CHO CCl3-COOH
With Tollen's reagent a silver mirror
With Fehling 1 and 2 Red ppt. of Cu2O
e) Azo-dye test: Scarlet red dye aniline hydrochloride

Lastly
1) The melting point of the original compound is recorded, (or the
boiling point)
2) After the group in the compound has been ascertained, asuitable
solid derivative (which depends upon the group present) is prepared
and its m.p. is determined.
3) The compound is then identified if possible.
 

 
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