Benedicts Test - Overview, Preparation, Analysis, Limitations, FAQs

Reducing Sugar

A reducing sugar is any sugar (or carbohydrate) that has the ability to reduce other substances through the donation of electrons, and it contains a free aldehyde (-CHO) or ketone (-C=O) group. This means the sugar can donate electrons and act as a reducing agent in chemical reactions. The presence of a free aldehyde group (in glucose, fructose, etc.) or a ketone group (in fructose) allows these sugars to act as reducing agents.

Reducing Sugar Test

When Benedict’s reagent is subjected to reducing sugars, the reactions it undergoes result in the production of a brick-red precipitate, indicating a positive Benedict test principle. The changes in colour of Benedict’s reagent (from clear blue to brick-red) caused by exposure to reducing sugars are depicted in the figure below.

Benedict's sugar test is a straightforward method for determining the concentration of reducing monosaccharides in a solution. Benedict’s test can detect some disaccharides, however sucrose (table sugar) is an unreactive disaccharide. Benedict’s solution is a reagent, a substance that changes colour when exposed to another substance.

Benedict Test Reaction in the Benedict's Test for Urine Sample

Benedict’s test can also be used to determine whether or not there is glucose in a Benedict test for a urine sample. When Benedict’s test for glucose is present in the analyte, the test produces a positive result because it identifies any aldehydes and hydroxy ketones, and glucose is an aldose whose open-chain form an aldehyde group in reducing substances in urine. The presence of ascorbic acid, homogentisic acid, and other reducing chemicals in the Benedict test for urine, on the other hand, can cause a positive reaction. As a result, a positive Benedict reaction test does not always mean the person is diabetic.

Chemical Reaction:

$
\mathrm{Cu}^{2+}+\text { Reducing Sugar } \rightarrow \mathrm{Cu}^{+}+\text {Reduction Products }
$
For glucose ( $\mathrm{C}_6 \mathrm{H}_{12} \mathrm{O}_6$ ), the reaction can be represented as:

$
\mathrm{C}_6 \mathrm{H}_{12} \mathrm{O}_6 \text { (glucose) }+2 \mathrm{Cu}^{2+}+2 \mathrm{OH}^{-} \rightarrow \mathrm{C}_6 \mathrm{H}_{12} \mathrm{O}_6 \text { (oxidized) }+\mathrm{Cu}_2 \mathrm{O} \text { (copper(I) oxide) }+\mathrm{H}_2 \mathrm{O}
$

Principle of Benedict Test

A reducing sugar is transformed to an enediol formation when it is heated in the presence of an alkali (which is a relatively powerful reducing agent). The cupric ions (Cu²⁺) in Benedict’s reagent test are converted to cuprous ions (Cu⁺) when Benedict’s test for reducing sugars is present in the analyte. These cuprous ions combine with the reaction mixture to generate copper (I) oxide, which precipitates as a brick-red substance.

An Aldose + Benedict's reagent (blue) $\rightarrow$ Carboxylate ion + Brick Red precipitate

$
\mathrm{RCHO}+2 \mathrm{Cu}^{2+}(\text { citrate }) \rightarrow \mathrm{RCOO}^{-}+\mathrm{Cu}_2 \mathrm{~S}(\mathrm{~s})
$

Benedict reagent can be represented as:

$\mathrm{CuSO}_4($ Copper $($ II $)$ sulfate $)+\mathrm{Na}_2 \mathrm{CO}_3($ Sodium carbonate $)+\mathrm{Na}_3 \mathrm{C}_6 \mathrm{H}_5 \mathrm{O}_7($ Sodium citrate $)+$ Water $\left(\mathrm{H}_2 \mathrm{O}\right)$

Benedict's Test Procedure: Preparation of Benedict's Reagent

In distilled water, combine 17.3 g of copper sulphate pentahydrate (CuSO₄.5H₂O), 100 g of sodium carbonate (Na₂CO₃), and 173 g of sodium citrate to make one litre of Benedict’s reagent (required quantity). Copper (II) sulphate serves as a source of Cu²⁺ ions, sodium carbonate serves as an alkaline medium, and sodium citrate creates Cu²⁺ ion complexes. As a solvent, distilled water is employed.

• Alkalinity is imparted to the reaction media by anhydrous sodium carbonate.

• Sodium Citrate - creates a compound with cupric ions, preventing them from decomposing into cuprous ions during storage.

• The cupric ions are produced by copper (II) sulphate pentahydrate.

• As a solvent, distilled water is employed.

By heating Benedict’s reagent in a test tube, you can determine its purity. The fact that the blue benedict solution colour of the solution does not change when heated indicates that the reagent is pure.

Composition of Benedict reagent: $\mathrm{CuSO}_4 \cdot 5 \mathrm{H}_2 \mathrm{O}+\mathrm{Na}_2 \mathrm{CO}_3+\mathrm{Na}_3 \mathrm{C}_6 \mathrm{H}_5$

Also read :

Benedict's Test for Reducing Sugars

One millilitre of analyte sample must be combined with two millilitres of Benedict’s reagent and heated for 3 to 5 minutes in a bath of boiling water. The presence of reducing sugars in the analyte is confirmed by the formation of a brick-red coloured cuprous oxide precipitate.

Analysis of Benedict's Test Results

Depending on the amount of reducing sugar in the sample, the colour ranges from green to yellow to orange to brick-red; a sample containing 1% glucose produces a brick-red precipitate.

• Any mono or disaccharide having a hemiacetal or hemiketal group will change colour in Benedict's test.
• Because sucrose or table sugar lacks these groups, it will not result in a positive test.

Limitations of Benedict's Test

If some medicines are present, such as salicylates, isoniazid, streptomycin, penicillin, and p-amino salicylic acid, false-positive results in the test can occur. The compounds in the concentrated Benedict test for urine, such as urate, creatinine, and ascorbic acid, may diminish Benedict's reactivity (the reduction is slight).

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Some Solved Examples

Question 1: Fructose and glucose can be distinguished by:

1) Benedict's test

2) Fehling's test

3) Barfoed's test

4) (correct) Seliwanoff's test

Solution:

Fructose / fruit sugar -

Structure is similar to Glucose, except it contains a ketonic functional group at carbon number 2.

Fructose and glucose can be distinguished by -Seliwanoff's test

(1)Barfoed's test:- used for reducing sugar, both are reducing sugar. So both give this test

(2)Benedict's test:- used for reducing sugar

$\therefore$ Both give this test

(3)Fehling's test:- Given by reducing sugars. So Both gives this test

Hence, the answer is the option (4).

Question 2: A certain compound gives a negative test with ninhydrin and a positive test with Benedict's solution. The compound is:

1) Protein

2) (correct) Monosaccharide

3) Lipid

4) An amino acid

Solution:

A compound that gives a negative test with ninhydrin cannot be a protein or an amino acid. As it gives a positive test with Benedict's solution. So, it must be a monosaccharide but not a lipid.

Hence, the answer is option (2).

Question 3: Number of compounds from the following which will not produce orange red precipitate with Benedict solution
is _______________
Glucose, maltose, sucrose, ribose, 2-deoxyribose, amylose, lactose

Solution:

The test uses Benedict's reagent, a mixture of copper sulfate, sodium citrate, and sodium carbonate, to identify reducing sugars. When reducing sugars are present, the reagent's copper ions are reduced to cuprous ions, which form a brick-red precipitate

Neither Amylose nor Sucrose do not give the Benedict‘s test.

Ans = 2

Hence, the answer is (2).

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