The objective of this experiment was to identify the reaction of carbohydrates in Benedict’s test, Barfoed’s test and Iodine test. Carbohydrates are essential to living organisms, and the principal role of carbohydrates is the production of energy. Carbohydrates are groups of sugars that contain carbon, oxygen, and hydrogen in a 1:2:1 ratio. Three main units of carbohydrates are monosaccharides, disaccharides, and polysaccharides. Monosaccharides are a single chain of CH2O while disaccharides are two units bonded together through dehydration reaction but they are both considered sugars.
Polysaccharides on the other hand are a combination of three or more units of monosaccharides put together. Carbohydrates are formed by the combination of carbon dioxide and water molecules. The carbohydrates contain two specific functional groups in which are the hydroxyl groups and carbonyl groups. In the carbohydrates, if the sugar contains aldehyde group it called reducing sugar and called non-reducing sugar if didn’t have the aldehyde group.
The Benedict’s test shows us which sugars are reducing or non-reducing. Benedict’s reagent, which contains Cu2+ ions in alkaline solution with sodium citrate added to keep the cupric ions in solution. The alkaline conditions of this test causes isomeric transformation of ketoses to aldoses, resulting in all monosaccharides and most disaccharides reducing the blue Cu2+ ion to cuprous oxide (Cu2O), a brick red-orange precipitate. Examples of reducing sugar are glucose. Examples of non-reducing sugar are sucrose.
Monosaccharide, which is smallest carbohydrates molecules, can be described by the number of carbons in the chain so that a monosaccharide with five carbons in a chain is a pentose and one with six carbons is a hexose. A monsaccharide contains one sugar unit, and has three to seven carbon atoms. Monosaccharides have a hydroxyl group bonded to each carbon atom; expect one, which is bonded to an oxygen atom thus forming a carbonyl group. If the carbonyl group is at the end of the chain then is it an aldehyde; therefore, if it is located in any other position it is a ketone.
Common monosaccharides are ribose, deoxyribose, glucose, fructose, and galatose. Barfoed’s test for monosaccharide relies on the fact that reducing monosaccharide produce Cu2O faster than reducing disaccharides. Barfoed’s solution contains cupric ions in an acidic medium. The milder condition allows oxidation of monosaccharides but does not oxidize disaccharides. If the time of heating is carefully controlled, disaccharides do not react while reducing monosaccharides give the positive result (red Cu2O precipitate). Ketoses do not isomerize with this reagent.
The Barfoed test showed whether the sample is a non-monosaccharide or a monosaccharide. Lastly the Iodine test identified whether the sample was a non-polysaccharide. If it wasn’t then it could identify if it was a branched or unbranched polysaccharide. Iodine test can easily determine a starch. Starches form deeply colored blue-black complex with iodine. Starches contain ?-amylose, a helix saccharide polymer, and amylopectin. The iodine molecule slips inside of the amylose coil. Iodine forms a large complex polysaccharide with the ?-amylose helix, producing the blue-black color.
Simpler oligosaccharides and monosaccharides do not form this complex with iodine. Methods/Procedures: For the Benedict test we had to reheat our water since we had difficulty with the burner and that was an extra procedure. Also what we found easier instead of using a cylinder to measure the appropriate solution we tried to put all the test tubes next to each other and match up the first measured test tube with the appropriate liquid. For the other tests we followed the same procedures as mentioned on pg 14 of “Experiments in Biology by Linda R.
Vanthiel”. Results: The results for the Benedict test showed that almost all samples were reducing except Sucrose, Glycogen, Starch, and Distilled water. Barfoed’s test proved that Fructose, Glucose, Galactose and Equal were all monosaccharides. Lastly the Iodine test showed that sweet and low, glycogen and starch are polysaccharides. To be more specific the results showed that distilled water is a non-reducing sugar and is neither monosaccharide nor polysaccharide. Glucose came out to be a reducing sugar that is a monosaccharide.
Fructose is also reducing sugar and monosaccharide just like Glucose and Galactose had the same results. Maltose on the other hand is a reducing sugar but it is not a monosaccharide or a polysaccharide. Lactose came out to have the same results as Maltose. Sucrose is a non-reducing sugar that is neither polysaccharide nor monosaccharide. Glycogen ss a non-reducing sugar that is a polysaccharide and further more it is also branched. Starch is a non-reducing polysaccharide that unbranched. Sweet and low, which was Unknown A, turned out to be reducing sugar and a branched polysaccharide.
While the second Unknown B that was Equal is also a reducing sugar but a monosaccharide. Discussion: In the Benedict test we started off with a blue solution and ended with a red color precipitate that indicated the sample was reducing. For the Barfoed test we started off the same but the end product in some samples had precipitate which showed that those samples were monosaccharide. Some of the samples that remained blue and indicated a positive test were starch and sucrose for example. While fructose, glucose and maltose formed the red precipitate which showed a negative test.
The blue solution remained after heated indicates the presence of disaccharide and the formation of brick red precipitate indicates the presence of monosaccharide. In the Iodine test the end results were mostly yellow which indicated non-polysaccharide but there were some rust or blue-black color that indicated the samples were either branched or unbranched polysaccharides. Glycogen and sweet and low were branched polysaccharides because they had a rusty color at the end while starch was an unbranched polysaccharide due to it blue-black color change.
With the Iodine and Barfoed test we were able to figure out which sample might be disaccharides. When the solution of carbohydrate and Benedict is heated, a formation of red brick precipitate is appeared. This red color indicates that the sample is reducing. The precipitate is the formation of CuO from the Benedict solution. This happen because monosaccharide will reduce Cu2SO4 in the Benedict solution to CuO. All type of monosaccharide that can do this reaction is known as reducing sugar.
Distilled water came out to be non-reducing because it had a blue color after the Benedicts test, it was also non-monosaccharides and non-polysaccharide because there was no precipitate after the Barfoed’s test was done and the color was yellow when it reacted with the Iodine. Therefore Distilled water must have been a disaccharide. Glucose was a reducing monosaccharide because during the Benedict test the color was red and it formed a precipitate after the Barfoed’s test therefore we can say that for the iodine test it will be non-polysaccharide.
Fructose was also reducing and monosaccharides just like Glucose and also had a red color after Benedict’s test. Galactose had the exact same reaction as Fructose and Glucose. Maltose was different because it showed to be a disaccharide since both Barfoed and Iodine test were negative and it is also a reducing sugar since the color was orange after the first test. Lactose has the exact same result as Malatose and I think that is because it is somewhat from the same grouping as Lactose. Sucrose came out to be non-reducing disaccharide sugar because the last two test were both negative and the Benedict test had blue solution at the end.
Glycogen seemed to differ from the rest because it was non-reducing and it came out to be branched polysaccharide which means that the Barfoed’s test was negative and the Iodine test had a rusty color in the end. Starch was also non-reducing and polysaccharide sugar but it was unbranched due to the results that showed the solution as blue-black. Sweet and Low was Unknown A and it turned out to be a reducing sugar which I had expected and it was also a branched polysaccharide due to the rusty color therefore I know that the second test was negative.
Unknown B was Equal and it was also a reducing sugar but it was a monosaccharide due to the precipitate. In conclusion I think that using all three test it was helpful to reinsure results from the other tests and find which of the samples were probably disaccharides. For example even though Glycogen came out non-reducing in the iodine test it did come out as a sugar. Therefore I think that being able to use many tests can prove a hypothesis right or wrong and give a more accurate answer rather than just assuming.