Lab 2 - Identification Of Functional Groups

Question 1

How do you test alcohols?

Answer 1

Lucas’s test (hydrochloric acid/ zink chloride).
- The tertiary alcohol will show a positive result fast in the presence of anhydrous zink. Secondary alcohols after 5-10min. Primary alcohols do not react with ZnCl2

Question 2

How do you test unsaturated compunds (eg. cyclohexene and cholesterol)?

Answer 2

1) Bromine in carbon tetrachloride (Br2/CCl4).

2) Potassium permaganate test (KMnO4)-[Baeyer’s test]

Question 3

How do you test phenols?

Answer 3

Ferric chloride(FeCl3)

• phenols = OH group in aromatic ring
• The phenol will form a colord solution depending on the substituents of the aromatic ring. (Purple, blue, red, or green)

Question 4

Bromine in carbon tetrachloride (Br2/CCl4).

Answer 4

This test is useful for indicating the presence of many olefinic or acetylic functional groups. Unsaturated hydrocarbons undergo addition reactions. Bromine solution added to unsaturated hydrocarbons solution CC4 is discolored.

Question 5

Aqueus Potassium permaganate test (KMnO4)-[Baeyer’s test]

Answer 5

This test is useful for indicating the presence of unsaturated hydrocarbons. Potassium permaganate solution is discolored.

Question 6

How do we test aceton (carbonyl group in ketone)?

Answer 6

2,4-dinitrophenylhydrazine

Question 7

How do we test glucose (carbonyl group in aldehyde)?

Answer 7

• 2,4-dinitrophenylhydrazine

- Tollens test

Question 8

2,4-dinitrophenylhydrazine

Answer 8

this test is useful for the identification of aldehydes and ketones. Most aldehydes and ketones give a solid 2,4-dinitrophenylhydrazone. This derivatives is yellow, orange, or red colored.

Question 9

Silver-ammonia complex ion - Tollens test:

Answer 9

• This test is useful for distinguishing aldehydes (and “reducing sugars”) from ketones and other carbonyl compounds.
• Aldehydes and other substances can be oxidized by the silver-ammonia complex ion, will reduce the silver ion to metallic silver, which will precipitate as a “mirror” on the test tube, or as a black colloidal suspension.

Question 10

How do we test the carbohydrates?

Answer 10

Molisch´s test

Question 11

How do we test the amino acids?

Answer 11

Ninhydrin test

Question 12

Ninhydrin test color change:

Answer 12

• Ninhydrin reacts with compounds containing the PRIMARY amine group causes the blue coloration of solution.
• Ninhydrin reacts with proline and yellow product is formed. Proline has SECONDARY amine.

Question 13

Ninhydrin reaction:

Answer 13

• Free α-amino group of amino acid while reacting with ninhydrin produces a deep blue or purple color known as Ruhemann’s purple.
• During this reaction, the amino acid is oxidized, decarboxylated and is subjected to oxidative deamination.
• Released ammonia reacts with ninhydrin derivative (obtained as a result of heating) to form Ruhemann’s
  purple.

Question 14

What will get a positive reaction with Ninhydrin reaction?

Answer 14

• Positive result of this reaction can be visible for compounds including: proteins, peptides, ammonia, amines, and amino sugars.

Question 15

What does the colors in a positive Ninhydrin reaction tell us:

Answer 15

• The intensity of the color depends on the concentration of amino acid in analyzed solution.
• This property is used for the quantitative determination of amino acids by the colorimetric method.
• Proline forms with ninhydrin compound of different structure that is brick-red color.

Question 16

Reaction with nitrous acid - HNO2 (Van Slyke’s reaction):

Answer 16

• Free amino acids react with nitric acid (III) releasing a stoichiometric amount of nitrogen (deamination).
• This reaction is used for quantitative determination of amino acid nitrogen by Van Slyke method.
• The unstable nitric acid (III) is prepared in situ by the reaction of nitrate (III) sodium salt with acetic acid.

Question 17

Xanthoproteic reaction:

Answer 17

• Amino acids containing aromatic ring can undergo the nitration reaction with concentrated nitric acid to form a yellow nitro derivatives.
• The addition of NaOH deeps orange color due to the formation of salt.
• Tyrosine and tryptophan can be easily subjected to the xantoproteic reaction but the positive effect of it for phenylalanine is difficult to observe.

Question 18

Detection of tryptophan (Voisent’s reaction):

Answer 18

• In acidic environment, compounds containing an indole ring (tryptophan) react with aldehydes to give a violet condensation products.
• The most often used aldehydes are glyoxal and formaldehyde.

Question 19

Tyrosine detection (Millon’s reaction):

Answer 19

• Detection of tyrosine with Millon’s reagent (a mixture of ionic mercury (I) and (II) and nitrate ions (III) and (V)), is based on the formation, in a concentrated nitric acid (V), at a temperature of 1000 C, nitro derivatives of tyrosine which form red products with a mercury ion (II) and mercury (I).
• Positive effect gives other compounds containing a phenolic ring.

Question 20

Detection of sulfur amino acids (cystine reaction):

Answer 20

• Compounds containing sulfhydryl group while heating under strongly alkaline, (e.g.. free or protein-bound cysteine or cystine) release sulfur in the form of sulphide ions, which react with lead (II) salts to form black lead sulphide precipitate.
• Other sulfuric amino acid, methionine, which has a thioether sulfur, does not give a positive result of the
  reaction.

Question 21

Methionine detection (McCarthy-Sullivan’s reaction):

Answer 21

• The sulfur present in the methionine is released in an alkaline medium.
• Methionine can be detected using sodium nitroprusside in the presence of glycine (the structure of a formed compound is unknown).
• Appearing pinkish-brown color indicates the presence of methionine.

Question 22

Histidine detection (Pauly’s reaction):

Answer 22

• Imidazole-containing compounds (e.g. histidine), and aromatic compounds having hydroxy substituent within the ring, or an amine (e.g. tyrosine) are coupled under alkaline conditions with diazonium salts to give colored products (azo dyes).
• Histidine forms red, and tyrosine - yellow-orange (tea) color products.
• Pauly’s reagent consists of three solutions: sulfanilic acid, nitrate (III) and sodium carbonate.
• Mixing of all aforementioned reagents lead to diazotization reaction in which diazobenzensulfonic acid is formed and coupled with histidine or tyrosine.

Question 23

Arginine detection (Sakaguchi’s reaction):

Answer 23

Arginine has a guanidino group, which reacts with α naphthol in the presence of an oxidant sodium bromate (I) to form orange-red dye solution. Excess of bromate oxidize ammonia to free nitrogen to form bubbles in the reaction mixture.