The Reactions Of Carbohydrates

Carbohydrates, or sachharides are sugars and starches that provide energy for us humans and animals. In this experiment, carbohydrates are tested with different reagents, and two unknown samples are examined to determine their identity. For the identification of unknown carbohydrate sample, seven different carbohydrates and two unknown samples with one ml volume was placed in separate test tubes. It was tested with Iodine test, wherein one ml of iodine reagent was added to each sample. For Benedict, Barfoed, Seliwanoff, and 2,4-DNP test, one ml of their corresponding reagent was supplemented on each carbohydrate sample and they were heated in a water bath afterwards. Results and reaction was taken and observed so that the identity of the two unknown samples can be determined. After carefully analyzing the data, the unknown samples were identified to be Fructose and Ribose. On the next part, hydrolysis of starch, fifty ml of five percent starch solution was placed in a one hundred ml beaker and five ml of concentrated HCl was added. It was heated while covered in foil. One ml of samples was placed in two distinct test tubes; the first one was supplemented with one ml of iodine reagent and the other one with Benedict’s reagent. The heating was continued and the process was repeated every five minutes until the formation of blue-black complex in iodine test, and formation of brick red color for the Benedict’s test. It took ten minutes until blue-black complex and brick red color was formed.

INTRODUCTION

Carbohydrates are carbon compounds that contain great amounts of hydroxyl groups. The simplest carbohydrates also comprise either an aldehyde moiety ( polyhydroxyaldehydes ) or a ketone moiety (polyhydroxyketones). All carbohydrates can be categorized as monosacchrides, oligosaccharides or polysaccharides. Somewhere from two to ten monosaccharide units, linked by glycosidic bonds, make up an oligosaccharide. Polysaccharides are much bigger, having hundreds of monosaccharide units. The existence of the hydroxyl groups permits carbohydrates to interrelate with the aqueous nature and to participate in hydrogen bonding, both inside and between chains. Derivatives of carbohydrates can contain nitrogen, phosphate and sulfur compounds. Carbohydrates also can unite with lipids to form glycolipids or with proteins to form glycoproteins. [1]

MATERIALS AND METHODS

Identification of Unknown Carbohydrate Sample

One ml of carbohydrate sample was added in two separate test tubes. One ml of Molisch reagent were added in the first test tube and 1ml of conc. H2SO4 where added to the next one. Results were observed and another 9, 1ml samples of different carbohydrates where prepared for and tested with the following: Iodine test, each test tube was added 1ml of iodine reagent. For the next tests, Benedict, Barfoed, Seliwanoff, and 2,4-DNP, 1ml of their respective reagent was added in each tube and were simultaneously heated afterwards in a water bath. Lastly, the unknown samples was compared with the other samples they were identified to be Fructose and Ribose.

Hydrolysis of Starch

A 50ml of 5% solution was placed in a 100ml beaker and 5ml of conc. HCl was added. It was covered with foil and heated. Then 1ml of sample was placed in two separate test tubes and 1 ml of iodine reagent was added with the first one and 1 ml of Benedict’s reagent was added on the second. It was continued and repeated until the formation of blue-black complex in iodine and development of brick red color in Benedict’s reagent.

RESULTS AND DISCUSSION

The table below summarizes the positive results for carbohydrates tested with different reagents.

Table 1. Positive Reactions for Carbohydrates Test

Test

Positive Color Change

Molisch

Deep purple

Iodine

Blue black

Benedict

Rust colored

Barfoed

Rust colored (monosaccharides)

Seliwanoff

Red color after heat

2,4-DNP

Yellow black

Carbohydrates can be identified through different tests. Molisch test which is a general test for all carbohydrates that gives a deep purple color for a positive reaction. Iodine test positively reacts on the presence of starch giving a blue-black color. Meanwhile, Benedict’s test is used to determine the occurrence of reducing sugars that produce rust-red color. Barfoed’s test is used for detecting monosaccharides that also have a rust-red color for a positive result. On the other hand, Seliwanoff is used to distinguish monosaccharide ketones that will give a red color after heating. And lastly, 2,4DNP is also a general test for all carbohydrates except for starch that has a yellow-black color for its positive reaction

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The results of the experiments where taken down, and the data were compared and thoroughly analyzed to recognize the identity of the 2 unknown samples that was distinguished to be Fructose for unknown 1 and Ribose for the unknown 2.

Table 2. Reaction of Carbohydrate Samples

Samples

Molisch

Iodine

Benedict

Barfoed

Seliwanoff

2,4-DNP

Glucose

+

–

+

++

–

+

Galactose

+

–

+

++

–

+

Ribose

+

–

++

+

–

+

Fructose

++

–

+

++

++

++

Lactose

+

–

+

–

–

+

Sucrose

++

–

–

–

False +

++

Starch

+

++

–

–

–

–

Unknown 1

++

–

+

++

++

++

Unknown 2

+

–

+

+

–

+

Identity of Unknown 1

Fructose

Identity of Unknown 2

Ribose

Legend: ++ fast reaction + slow reaction – no reaction

Molisch’s Test is a sensitive chemical test for all carbohydrates, and some compounds comprising carbohydrates in a shared form, based on the dehydration of carbohydrate by sulfuric to generate an aldehyde (furfural or a derivative) that is then compresses with the phenolic structure resulting in a deep-purple colored compound. [2]

Figure 1. Reaction of Carbohydrates with Molisch Test

Starch is a polysaccharide that can be simply recognized by the iodine test. The many glucose units in starch lock in the I2 molecules and appear as dark blue-black complex. [3]

Figure 2. Structure of Starch

Most disaccharides and polysaccharides can be simplified into their monosaccharide subunits by a method called hydrolysis. In live systems, molecules known as enzymes aids this breakdown. Hydrolysis of starch is essential for the organism to make use of the glucose monomers. A chemical hydrolysis can be done in the laboratory by heating the polysaccharides with acid in the presence of water. [4]

Table 3. Hydrolysis of Starch

Time (min)

Color w/ Benedict’s test

Time (min)

Color w/ Iodine test

5

Blue

5

Bluish black

10

Brick Red

10

Light Brown

The transformation in iodine color with the accumulation of starch and enzyme solution will signify starch hydrolysis. If the iodine turns blue-black, it indicates the presence of starch, therefore the denatured enzyme. If the iodine remains orange-yellow color, it shows the absence of starch, thus the proper function of amylase. In Benedict’s test, as the temperature increases, formation of brick red color begins; this is due to recognition of glucose. [5]

Figure 3. Reaction of Carbohydrates with Benedict’s Test

All monosaccharides and numerous disaccharides diminish feeble oxidizing agents like Cu2+ ion. These carbohydrates are called reducing sugars Benedict’s reagent changes color form blue to brick red in the presence of reducing sugars. [4]

Reducing sugar is a sugar that, in a mixture, has an aldehyde or a ketone group. The enolization of sugars under alkaline environments is a significant contemplation in reduction tests. The capability of a sugar to trim down alkaline test reagents depends on the promptness of an aldehyde or keto group for reduction reactions. [3]

Figure 4. Reaction of Carbohydrates with Seliwanoff Test

Seliwanoff’s test is used to differentiatealdohexoses from ketohexoses. A ketohexose just like fructose will create a deep red color with Seliwanoff’s reagent while an aldohexose will exhibit a light pink color and takes a longer time to produce the color. This test is based on the fact that, when settled on fire, ketoses are more precipitously dehydrated than aldoses. [4]

Figure 5. Reaction of Carbohydrates with Barfod’s Test

Barfoed’s reagent is composed of copper acetate in acetic acid and will not be reactive as Benedict’s reagent. It is also stable so that it can be only reduced by monosaccharids but not less authoritative reducing sugars. Dissacharides may also react with this reagent, but the response is not as fast when compared to monosaccharides. [3]

Figure 6. Reaction of Carbohydrates with 2,4-DNP Test

2,4-DNP test can react with both ketone and aldehyde. Therefore it’s also a general test for all carbohydrates except starch. A positive reaction results to a yellow-black color. Thus, absence of yellow-black color shows negative result.

When it comes to Diabetes test, urine taxing is n necessary part of complete physical test routine. Urine test helps in diagnosing the state of the body and give important diagnosis. [6]

Benedict’s test is used to sense the occurrence or absence of sugar in the urine is tremendously easy and ought to be leant by every diabetic. Even when sugar has been found to be in the urine, the analysis of diabetes should be completed by blood-sugar evaluation. [6]