Qualitative tests of Carbohydrate, Lecture notes of Organic Chemistry

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Qualitative tests of Carbohydrate

• Carbohydrates are the key source of energy used by living things.
• Also serve as extracellular structural elements as in cell wall of bacteria and plant.
• Carbohydrates are defined as the polyhydroxy aldehydes or polyhydroxy ketones.
• Most , but not all carbohydrate have a formula (CH 2 O)n (hence the name hydrate of carbon)
• In human body, the D-glucose is used.
• Simple sugars ends with – ose.

Carbohydrate :

Classification :

1 - Simple sugar (one unit) : Monosaccharides contain one monosaccharide unit.

2-Complex sugar (more than one) :

• Disaccharides contain two monosaccharide units.
• Oligosaccharides contain 3-9 monosaccharide units.
• Polysaccharides can contain more than 9 monosaccharide units.

-Complex carbohydrates can be broken down into smaller sugar units through a process known as hydrolysis.

Reducing and non reducing sugars

Reducing and non reducing sugar :If the oxygen on the anomeric carbon of a sugar is not attached to any other structure, that sugar can act as a reducing agent and is termed a reducing sugar.

reducing Non-reducing

Chemical Properties of Carbohydrates:

1 - Molisch Test: specific for carbohydrates.

2- Benedict's Test: presence of reducing sugars.

3- Barfoed's Test: test used for detecting the presence of monosaccharides.

4- Bial's Test: used to detect pentose [5C] monosacharides.

5- Seliwanoff's Test: distinguish between aldoses and ketoses.

1 .Molisch test:

This test is specific for all carbohydrates Monosaccharide gives a rapid positive test, Disaccharides and polysaccharides react slower.

Objective: To identify the carbohydrate from other macromolecules, lipids and proteins.

Principle:

• The test reagent(H2SO4) dehydrates pentose to form furfural and dehydrates hexoses to form 5- hydroxymethyl furfural.
• The furfural and 5- hydroxymethyl furfural further react with α-naphthol present in the test reagent to produce a purple ring.

Method :

1-Two ml of a sample solution is placed in a test tube. 2-Two drops of the Molisch reagent (which α-napthol in 95 % ethanol) is added. 3 - The solution is then poured slowly into a tube containing two ml of concentrated sulfuric acid so that two layers form, producing violet ring appear as liaison between the surface separations.

Tube Observation Glucose Lactose Starch

2 .Benedict's test:

Objective: To detect the presence of reducing sugars.

• All monosaccharides are reducing sugars; they all have a free reactive carbonyl group.
• Some disaccharides have exposed carbonyl groups and are also reducing sugars. Other disaccharides such as sucrose are non-reducing sugars and will not react with Benedict's solution

-Large polymers of glucose, such as starch, are not reducing sugars, since the concentration of hemiacetal groups is very low.

• One ml of a sample solution is placed in a test tube.
• Two ml of Benedict's reagent is added.
• The solution is then heated in a boiling water bath for five minutes.
• A positive test is indicated by: The formation of a reddish precipitate.

Method :

Tube observation 1 -glucose 2-lactose 3-starch

3 .Barfoed’s Test:

Objective : To distinguish between mono- , di- and poly saccharides. Principle : Barfoed’s test used copper (II) ions in a slightly acidic medium. Reducing monosaccharides are oxidized by the copper ion in solution to form a carboxylic acid and a reddish precipitate of copper (I) oxide within three minutes. Reducing disaccharides undergo the same reaction, but do so at a slower rate.

-The nonreducing sugars give negative result.

Objective: To distinguish between pentose monosaccharide and hexose

monosaccharide

4. Bial’s Test:

Principle: Bial’s test uses concentrated HCl as a dehydrating acid and orcinol +

traces of ferric chloride [FeCl 3 ] as condensation reagent. The test reagent dehydrates pentoses to form furfural. Furfural further reacts with orcinol and the iron ion present in the test reagent to produce a bluish or green product, while hexoses yield muddy-brown to grey condensation product.

• Put 2 ml of a sample solution in a test tube.
• Add 3 ml of Bial's reagent (a solution of orcinol, HCl and ferric chloride) to each tube.
• Heat the tubes gently in hot water bath.
• If the color is not obvious, more water can be added to the tube.

Method :

Tube observation 1 -glucose 2 -ribose

• One half ml of a sample solution is placed in a test tube.
• Two ml of Seliwanoff's reagent (a solution of resorcinol and HCl) is added.
• The solution is then heated in a boiling water bath for two minutes.

Method :

Tube observation 1 -glucose 2-fructose

Questions:

1- Name the complex formed by the addition of concentrated sulfuric acid to sugar solution and explain the reaction?

2- Why sucrose gives negative Benedict test?

3- Explain, although starch has free hemiacetal bond it gives negative Benedict test?

4- Why glucose (monosaccharide) and maltose (disaccharide) give positive Benedict test?

5- What is the difference between Benedict and Barfoed's reaction?

6- What are the carbohydrates’ that give positive result with Seliwanoff? why?