Classification Tests for Carbonyl- and Hydroxyl- Containing Compounds

La Rosa, J. A., Lee, S. Q., Mangahas, N. K., Manota, G. E., Matuloy, L. S.

University of Santo Tomas
Faculty Of Pharmacy
2A-Biochemistry

Abstract

Classification of Hydroxyl and Carbonyl containing compounds was done through different tests. The
samples were n-butyl alcohol, sec-butyl alcohol, tert-butyl alcohol, acetaldehyde, n-butyraldehyde,
benzaldehyde, acetone, acetophenone, and isopropyl alcohol. Each of the samples were tested through
solubility test of alcohols in water, Lucas test, Chromic acid test or the Jones oxidation test, 2,4-
Dinitrophenylhydrazone (2,4-DNP) test, Fehling’s test, Tollens’ silver mirror test, and Iodoform test.
Solubility test of alcohols in water was used to determine the amount of water needed for it to produce a
homogenous dispersion and determine whether what kind of alcohol is soluble and insoluble in water.
Lucas test was used to differentiate primary, secondary, and tertiary alcohols. Chromic acid test was used
to determine if the sample is oxidized and can also be used to differentiate ketones from aldehydes. 2, 4-
Dinitrophenylhydrazone test was used to differentiate aldehydes and ketones. Fehling’s test and Tollens’
silver mirror test was used to determine whether the sample is an aldehyde. Lastly, Iodoform test was
used for classification of methyl ketones.

Introduction (Clayden& Bingham, 2008). Alcohols also have

the ability to hydrogen bond with water;
Alcohols are organic compounds that may be however, their solubility decreases as the size of
considered derivatives of water in which one of the R group and their hydrocarbon-like character
the hydrogen atoms of water molecule (H-O-H) increases (Clayden & Bingham, 2008).
has been replaced by an alkyl or substituted
alkyl group (Jaism& Mohammad, 2012).
Therefore, properties of alcohols may be related
to properties of both water and hydrocarbons
(Jaism& Mohammad, 2012). The alkyl group
could be primary, secondary, or tertiary, and
may be open chain or cyclic. Accordingly,
alcohols may be defined as organic compounds Figure 2. Classes of Alcohol
http://www.esu.edu/~scady/Experiments/Alcohols(summer).pdf
that contain hydroxyl groups attached to alkyl,
substituted alkyl, or cyclic alkyl group
Aldehydes and ketones are classes of organic
(Vollhardt, 2007).
compounds in which oxygen is covalently joined
to a carbonyl group (Tojo&Fernadez, 2006).
Ketones differ from aldehydes by a replacement
of hydrogen with an R group. Consequently,
aldehyde functional groups are always found at
the end of hydrocarbon chains (Tojo &
Figure 1. Structural formula of Alcohol Fernadez, 2006).
http://media-cache-ec0.pinimg.com/736x/39/23/bd/3923bde41273f511199d81a9f0431a6a.jpg

All the three classes of alcohol ( 1°, 2°, and 3°)

are capable of hydrogen-bonding in the pure
state or to other alcohol molecules because the –
OH functional group is always present
Figure 3. Structural formula of Aldehyde
http://media-cache-ec0.pinimg.com/736x/e3/ef/e7/e3efe7ea73142727b810ae434661a396.jpg

Figure 5. Lucas test reaction

Figure 4. Structural formula of Ketone http://www.esu.edu/~scady/Experiments/Alcohols(summer).pdf
http://media-cache-ak0.pinimg.com/736x/0f/dc/d4/0fdcd47ceb12b048fa82898a70099180.jpg

There are different types of differentiating tests The Jones oxidation (also known as Chromic
to determine whether its alcohol, aldehyde, and Acid Test) is an organic reaction for
ketones; the commonly used tests are Solubility the oxidation of primary and
test, Lucas test, Chromic Acid test (Jones secondary alcohols to carboxylic
Oxidation), 2,4-dinitophenylhydrazine (2,4- acids and ketones, respectively. Similar to
DNP) test, Fehling’s test, and Tollens’ Silver alcohols, aldehydes and ketones differ in their
Mirror test (Sinton, 2009). ability to be oxidized. As noted previously,
aldehydes can be oxidized to carboxylic acids.
Lucas test in alcohols is a test to differentiate Ketones, which lack hydrogen bonded to the
between primary, secondary, and tertiary carbonyl group, cannot be oxidized. The
alcohols. It is based on the difference in following reactions below show the expected
reactivity of the three classes of alcohols with products aldehydes, and ketones with the Jones
hydrogen halides (Lehman, 2004). The differing reagent (Pellegrini, 2000).
reactivity reflects the differing ease of formation
of the corresponding carbocations. Tertiary
carbocations are far more stable than secondary
carbocations, and primary carbocations are the
least stable (Lehman, 2004). The time taken for
turbidity to appear is a measure of the reactivity
of the class of alcohol, and this time difference
is used to differentiate between the three classes
of alcohols (Mariappan, 2009). No visible
reaction at room temperature and cloudy only on Figure 6. Jones oxidation reaction
heating means it is a primary alcohol, when the http://upload.wikimedia.org/wikipedia/commons/thumb/a/ab/Jones
_Oxidation_Scheme.png/640px-Jones_Oxidation_Scheme.png
solution turns cloudy in 3–5 minutes means it is
a secondary alcohol, and if solution turns cloudy
immediately, and/or phases separate it means Brady’s test or 2,4-dinitophenylhydrazine (2,4-
DNP) test is used to qualitatively detect the
that it is a tertiary alcohol (Mariappan, 2009).
carbonyl functionality of a ketone or aldehyde
functional group. A positive test is signaled by a
yellow, orange or red precipitate (known as a
dinitrophenylhydrazone.) If the carbonyl
compound is aromatic, then the precipitate will
be red; if aliphatic, then the precipitate will have
a more yellow color.Aldehydes and ketones
undergo a condensation reaction with 2,4-
dinitrophenylhydrazine to produce yellow to
orange precipitates as products. Alcohols do not
undergo this reaction (Simek & Wade, 2003).
Figure 7. 2,4-DNP test reaction
http://www.esu.edu/~scady/Experiments/Alcohols(summer).pdf
Fehling's test is a chemical test used to
differentiate between watersoluble carbohydrate
and ketone functional groups, and as a testfor
monosaccharides. Fehling's can be used to
determine whether a carbonyl-containing
compound is an aldehyde or a ketone. The
bistartratocuprate (II) complex in Fehling's
solution is an oxidizing agent and the active
reagent in the test. The compound to be tested is
added to the Fehling's solution and the mixture
is heated. Aldehydes are oxidized, giving a
positive result (precipitation of red copper (I)
oxide), but ketones do not react, unless they are
alpha-hydroxy-ketones (Jones & Gingrich,
2009).
Figure 8. Fehling’s test reaction
http://upload.wikimedia.org/wikipedia/commons/thumb/6/6
5/Fehling_orz.svg/1024px-Fehling_orz.svg.png
Tollens' test is used to determine whether a
known carbonyl-containing compound is an
aldehyde or alpha-hydroxy ketone. It is usually
ammoniacal silver nitrate, but can also be other
mixtures, as long as aqueous diammine silver(I)
complex is present. A positive test with Tollens'
reagent results in elemental silver precipitating
out of solution, occasionally onto the inner
surface of the reaction vessel, producing a
characteristic and memorable "silver mirror" on
the inner vessel surface. Aldehydes will be
positive in Tollens' test and a mirror-like
material will be formed (Jones, 2000).
Iodoform test or haloform reaction is a chemical
reaction where a haloform (CHX3, where X is a
halogen) is produced by the exhaustive
halogenation of a methyl ketone (a molecule
containing the R–CO–CH3 group) in the
presence of a base.[1] R may be H, alkyl or aryl.
The reaction can be used to produce chloroform
(CHCl3), bromoform (CHBr3), or
iodoform(CHI3). This reaction was traditionally
used to determine the presence of a secondary
alcohol oxidizable to a methyl ketone. When
iodine and sodium hydroxide are used as the
reagents, a positive reaction gives iodoform.
Iodoform (CHI3) is a pale-yellow substance.
Due to its high molar mass caused by the three
iodine atoms, it is solid at room temperature (cf.
chloroform and bromoform). It is insoluble in
water and has an antiseptic smell. A visible
precipitate of this compound will form from a
sample only when either a methyl ketone,
ethanal, ethanol, or a methyl secondary alcohol
is present (Martin & Gilbert, 2011).
Figure 9. Iodoform test reaction
http://upload.wikimedia.org/wikipedia/commons/3/3
1/Iodoform_test_alcohols.pn
Materials
The materials needed for this
experiment were Lucas reagent, chromic acid
reagent, 95% ethanol, Fehling’s A and B,
Tollens’ reagent, 5% NaOCl solution, iodoform
test reagent, 2-4 dinitrophenylhydrazine, Pasteur
pipette, test tubes, vials, beaker. The sample
compounds used were ethanol, n-butyl alcohol,
sec-butyl alcohol, tert-butyl alcohol, benzyl
alcohol, n-butyraldehyde, acetone,
acetophenone, isopropyl alcohol, and
acetaldehyde
Methods
A. Solubility of Alcohols in Water
Five test tubes were labeled A through E (A:
ethanol, B: n-butyl alcohol, C: sec-butyl alcohol,
D: tert-butyl alcohol, E: benzyl alcohol). With
the aid of a Pasteur pipette, 10 drops of each of
the alcohols was placed into the appropriate test
tube. One mL of water was added to the test tube
containing alcohol, the mixture was thoroughly
shakenfor each addition until a homogenous
dispersion resulted. The total volume of water
was noted. In cases where no cloudiness resulted
after the 2.0 mL water was added, the alcohol
was considered soluble in water. The results
were noted.
B. Lucas Test
The reagent was prepared by dissolving 16 g
of anhydrous zinc chloride in a 10 mL of
concentrated HCL. Fifty milligrams (2-3 drops)
of sample was added to 1 mL of the reagent in a
small vial. It was capped and was shaken
vigorously for a few seconds. This test was
performed on n-butyl alcohol, sec-butyl alcohol,
and tert-butyl alcohol as well. The rate of
formation of the cloudy suspension or the
formation of 2 layers was observed.
C. Chromic Acid Test (Jones Oxidation)
The reagent was prepared by dissolving 20 g
of chromium trioxide (CrO3) in 60 mL of cold
water in a beaker and adding of 20 mL of
concentrated sulfuric acid to the solution. One
drop of liquid or a small amount of the solid
sample was dissolved in 1 mL of acetone in a
small vial. Five drops of chromic acid reagent
was added. The test tubes were placed in a 60ºC
water bath for 5 minutes. The color of each
solution was noted.
D. 2,4-dinitophenylhydrazine (2,4-DNP)
Test
The reagent was prepared by adding 3 g if
2,4-dinitophenylhydrazine in 15 mL of
concentrated H2SO4 to a mixture of 20 mL of
water and 70 mL of 95% ethanol, and then it
was stirred and filtered. A drop of liquid sample
was placed into a small test tube. Five drops of
95% ethanol was added. Three drops of 2,4-
dinitrophenylhydrazine was added. In cases
where no yellow or orange-red precipitate
formed the solution was allowed to stand for at
least 15 minutes. This test was performed on
acetone, acetaldehyde, n-butyraldehyde,
benzaldehyde, and acetophenone. The results
were described.
E. Fehling’s Test
Fehling’s A was prepared by dissolving 7 g of
hydrated copper (II) sulfate in 100 mL of water,
while the Fehling’s B was prepared by
dissolving 35 g of potassium sodium tartrate and
10 g of NaOH in 100 mL of water. One milliliter
of freshly prepared Fehling’s reagent (made by
mixing equal amount of Fehling’s A and
Fehling’s B) was placed into each test tube.
Three drops was added to the sample that was
tested. The test tubes were placed in a beaker of
boiling water and the changes which occurred
within 10-15 minutes were observed. This test
was performed on acetaldehyde, n-
butyraldehyde, acetone, benzaldehyde, and
acetophenone as well.
F. Tollens’ Silver Mirror Test
The reagent was prepared by adding 2 drops
of 5% sodium hydroxide solution to 2 mL of 5%
silver nitrate solution. Then only enough 2%
ammonium hydroxide was added to dissolve the
precipitate. No excess ammonia was added
because it will jeopardize the results. Four test
tubes with 1 mL of freshly prepared Tollens’
reagent were prepared. Two drops each of the
sample was added separately: acetaldehyde,
benzaldehyde, acetone, n-butyraldehyde, and
acetophenone. It was allowed to stand for 10
minutes. In cases where there is no reaction
occurred, the test tubes were placed in a beaker
of warm water (30-50 ºC) for 5 minutes. The
observations were recorded.
G. Iodoform Test
Two drops of each sample (acetaldehyde,
acetone, acetophenone, benzaldehyde, and
isopropyl alcohol) was placed into its own vial.
Twenty drops of 10% KI solution was added.
Twenty drops of fresh chlorine bleach (5%
sodium hypochlorite) was added to each test
tube, and then mixed. Yellow precipitate
formation was noted.
Results and Discussion
For the test for solubility of alcohols in water, reaction was also observed. Table 2 presents the
turbidity of the solution was to be observed. results of the Lucas test.
Cloudiness of the solution indicated in solubility
of that specific alcohol to water. The amount of Table 2. Lucas test
water needed to produce homogenous dispersion
was also observed. Table 1 shows the data Sample Reaction Observed
gathered from the test. n-butyl alcohol colorless
sec-butyl alcohol slightly turbid
Table 1. Solubility of Alcohols in Water tert-butyl alcohol turbid
Alcohol Amount of water Solubility
(mL) needed to to water According to the table above, n-butyl alcohol
produce a was soluble in Lucas reagent while sec-butyl
homogenous
alcohol and tert-butyl alcohol were observed to
dispersion
ethanol 1 mL soluble have a formation of cloudy layer. Tert-butyl
n-butyl alcohol 1.50 mL soluble alcohol took the shortest time to form the layer
while sec-butyl alcohol took the longest time.
Sec-butyl alcohol 1 mL soluble The reaction mechanism involved in the Lucas
test is based on SN1 reaction, which depends on
tert –butyl alcohol 1 mL soluble the formation of stable carbocations. Reactivity
benzyl alcohol 2 mL insoluble of alcohols in SN1 reaction is 3˚ > 2˚> 1˚. 3˚
alcohols formed the second layer in less than a
As indicated in the table, only benzyl alcohol minute. Secondary alcohols required 5-10
was insoluble in water, while ethanol, n-butyl minutes while 1˚alcohols were usually
alcohol, sec-butyl alcohol and tert-butyl alcohol unreactive. The presence of ZnCl2, a good Lewis
were all soluble in water. This follows the acid, made the reaction mixture even more
principle “like dissolves like” and therefore, it acidic; thus, it enhanced the formation of
can be said that the alcohols that were soluble in carbocations.
water are polar compounds since water is polar.
The Chromic Acid test (JonesOxidation) tested
Of the alcohols that were soluble in water,
for oxidizable or any compounds that possess
ethanol, sec-butyl alcohol and tert-butyl alcohol
reducing property (has alpha acidic hydrogen).
all required only 1 mL of water to be added in
Table 3 shows the results gathered from the said
order to be considered soluble. This indicates
test.
that there are certain factors affecting solubility.
One of these is the presence of number of carbon Table 3. Chromic Acid test (Jones Oxidation)
atoms. The lower the number of carbon atoms
present, the more soluble or more miscible a Sample Reaction Observed
substance is. Branching of carbon chains also n-butyl alcohol blue-green solution
sec-butyl alcohol blue-green solution
affects solubility directly proportional. The more
tert -butyl alcohol orange solution
branching present, the more soluble a compound n-butyraldehyde blue-green solution
is. This is only true for organic compounds that benzaldehyde blue-green solution
have the same number of carbon atoms present. acetone orange solution
acetophenone orange solution
The Lucas test differentiated 1˚, 2˚ and3˚
alcohols. Alkyl chloride formation was observed
and caused turbidity or cloudiness. The rate of
According to Table 3, n-butyl alcohol, sec-butyl
alcohol, n-butyraldehyde and bezaldehyde gave
a positive result of blue-green solution while
tert-butyl alcohol, acetone and acetophenone
gave a result of orange solution. Chromic Acid
test orJones Oxidation involved reduction-
oxidation or redox reaction. 1˚ and 2˚ alcohols
and aldehydes underwent oxidation and
chromium underwent reduction from Cr6+ to
Cr3+. 1˚ and 2˚ alcohols and aldehydes reduced
the orange-red chromic acid/sulphuric
acidreagent to an opaque green or blue
suspension of Cr (III) salts in 2-5 seconds. 1˚
alcohols reacted with chromic acid to yield
aldehydes, which are further oxidized to
carboxylic acids. 2˚ alcohols reacted with
chromic acid to yield ketones, which do not
oxidize further. 3˚ alcohols were usually
unreactive and aldehydes were oxidized to
carboxylic acids. The 2,4
Dinitrophenylhydrazone (2,4-DNP) test detected
the presence of carbonyl groups and tests
positive for aldehydes and ketones. Table 4
shows the results from the test.
Table 4. 2,4-Ditrophenylhydrazone (2,4-DNP)
test
nucleophilic addition of NH2 to C=O and
elimination of H2O. Some high molecular
weight ketones may fail to react or may yield
oils. Most aromatic aldehydes and ketones
produce red dinitrophenylhydrazone while many
non aromatic aldehydes and ketones produced
yellow products.
Fehling’s test was another differentiating test for
aldehydes and ketones. In this test, aldehydes
gave a positive result of brick-red precipitate
while ketones did not produce any reaction.
Table 5 presents the results of the test.
Table 5. Fehling’s test
Sample Reaction observed
acetaldehyde brick-red ppt
n-butyraldehyde brick-red ppt
benzaldehyde brick-red ppt
acetone blue solution
acetophenone blue solution
As shown in the given table, only acetone and
acetophenone did not react to forma precipitate
while the rest gave a positive result of brick-red
precipitate. Fehling’s test involved reduction-
oxidation or redox reaction. Aldehydes were
oxidized to carboxylic acidswhile ketones did
not undergo oxidation. Inhere, copper was
reduced from Cu2+ to Cu1+.

Sample Reaction observed Tollens’ Silver Mirror test differentiated

acetaldehyde yellow ppt aldehydes from ketones wherein aldehydes were
expected to be oxidized while ketones did not
n-butyraldehyde yellow ppt
undergo any oxidation. Table 6 shows the results
benzaldehyde yellow ppt
from the said test.
acetone yellow ppt
acetophenone red-orange ppt Table 6. Tollens’ Silver Mirror test

Sample Result observed

As indicated in the table, only acetophenone
acetaldehyde silver mirror
gave a result of red-orange precipitate while the
n-butyraldehyde flesh solution
rest of the samples gave a result of yellow
precipitate. A result of red-orange precipitate benzaldehyde light yellow solution
with globules
indicated the presence of conjugated carbonyl
compounds while a result of yellow precipitate acetone dark-gray solution
indicated the presence of unconjugated carbonyl acetophenone turbid gray solution
compounds. The reaction of 2,4-DNPH with
aldehydes and ketones in an acidic solution is a
According to Table 6, only acetaldehyde formed
dependable and sensitive test. Its reaction
a silver mirror. The samples n-butyraldehyde
mechanism involved condensation or
and benzaldehyde, although they are aldehydes,
did not form any silver mirror. The ketones
acetone and acetophenone formed dark-gray
solution and turbid gray solution, respectively.
The preparation of Tollens’ reagent was based
on the formation of a silver diamine complex
that is water soluble in basic solution. The
Tollens’ Silver Mirror test involved reduction-
oxidation or redox reaction. Aldehydes were
oxidized to carboxylic acids while ketones did
not undergo oxidation except alpha-
hydroxyketone. Silver was reduced from Ag1+ to
Ag0
Iodoform test was used to detect the presence of
methyl carbinol (2˚ alcohol with adjacent methyl
group) and methyl carbonylgroups. Table 7
shows the results from the said test.
Table 7. Iodoform test
Sample Reaction observed
acetaldehyde yellow ppt
n-butyraldehyde no reaction
benzaldehyde red ppt with globules
acetone yellow ppt
acetophenone yellow ppt
isopropyl alcohol yellow crystal ppt
According to the give table, acetaldehyde,
acetone and acetophenone gave a result of
yellow precipitate. Benzaldehyde gave a result
of red precipitate with globules while isopropyl
alcohol gave a result of yellow crystal
precipitates. No reaction was observed from n-
butyraldehyde. In this test, yellow crystals or
precipitate gave a positive result. An alkaline
solution of sodium hypoiodite, formed from
sodium hydroxide and iodine, converted
acetaldehyde and aliphatic methyl ketones into
iodoform (haloform reaction). Since the reagent
was also an oxidizing agent, alcohols which are
readily oxidized to acetaldehydes or methyl
ketones also gave a positive reaction. The
mechanism of iodoform synthesis occurred
through a series of enolate anions, which are
iodinated; hydroxide displaced the Cl3- anion
through an addition/elimination pathway.
References
Clayden, J., & Bingham, M. J. (2008). Alcohols.
Stuttgart: Thieme.
Lehman, J. W. (2004). The student's lab
companion: laboratory techniques for organic
chemistry. Upper Saddle River, N.J.: Prentice-
Hall.
Jaism, A. M., & Mohammad, D. H.,
(2012). Practical Organic Chemistry. 1st ed.
Baghdad: Baghdad University Publishing.
Jones, M. (2000). Organic chemistry (2nd ed.).
New York: W.W. Norton.
Jones, M., & Gingrich, H. L. (2005). Study
guide/solutions manual for Jones's Organic
Chemistry (3rd ed.). New York: W.W. Norton.
Mariappan, M. (2009). Organic Chemistry Lab
Manual. Dubuque, IA: Kendall Hunt.
Martin, S. F., & Gilbert, J. C. (2011). Organic
Chemistry Lab Experiments: Miniscale and
Microscale (5th ed., Int'l ed.). Boston, MA.:
Brooks/Cole, Cengage Learning.
Pellegrini, F. (2000). Organic chemistry II.
Foster City, CA: IDG Books Worldwide.
Simek, J. W., & Wade, L. G. (2003). Organic
Chemistry: Solutions Manual (5th ed.). New
York: Prentice Hall.
Sinton, M. (2009). Organic chemistry. Dubuque,
Iowa: Kendall Hunt Pub. Co..
Tojo, G., & Fernandez, M. (2006). Oxidation of
alcohols to aldehydes and ketones: a guide to
current common practice. New York, NY:
Springer.
Vollhardt, K. C., & Neil E. S., (2007). Organic
Chemistry. 5th ed. New York: W.H. Freeman.
Appendix
http://www.esu.edu/~scady/Experiments/Alcoho
ls(summer).pdf
Schematic diagram of the differentiating tests
http://upload.wikimedia.org/wikipedia/commo
ns/thumb/5/53/Butan-1-ol_Skelett.svg/1280px-
Butan-1-ol_Skelett.svg.png
n-butyl alcohol
http://upload.wikimedia.org/wikipedia/commo
ns/thumb/d/d4/Ethanol2.svg/1280px-
Ethanol2.svg.png
Ethanol
http://upload.wikimedia.org/wikipedia/commo
ns/thumb/e/e6/2-
Butanol_Structural_Formula_V.1.svg/1280px-2-
Butanol_Structural_Formula_V.1.svg.png
Sec-butyl alcohol
http://upload.wikimedia.org/wikipedia/commo
ns/thumb/4/4a/Tert-
Butanol_Structural_Formula_V.1.svg/785px-
Tert-Butanol_Structural_Formula_V.1.svg.png
Tert-butyl alcohol
http://upload.wikimedia.org/wikipedia/commo
ns/5/5a/Benzyl-alcohol-2D-skeletal.png
Benzyl alcohol
http://upload.wikimedia.org/wikipedia/commo
ns/thumb/1/1c/Butyraldehyde_200.svg/1280px
-Butyraldehyde_200.svg.png
n-butyraldehyde
http://upload.wikimedia.org/wikipedia/commo
ns/thumb/2/21/Aceton.svg/1280px-
Aceton.svg.png
Acetone

http://upload.wikimedia.org/wikipedia/commo
ns/thumb/8/88/Acetophenone_200.svg/1280px
-Acetophenone_200.svg.png
Acetophenone

http://upload.wikimedia.org/wikipedia/commo
ns/thumb/5/57/2-Propanol2.svg/1280px-2-
Propanol2.svg.png
Isopropyl alcohol

http://upload.wikimedia.org/wikipedia/commo
ns/thumb/4/47/Acetaldehyde_200.svg/1280px-
Acetaldehyde_200.svg.png
Acetaldehyde