CARBOHYDRATES I. OBJECTIVES 1. To be able to perform tests that will classify carbohydrates as reducing sugar or non-reducing sugar, etc. 2. To be able to define the condition necessary in the hydrolysis of carbohydrates. II. INTRODUCTION Carbohydrates are one of the three main classes of foods and a source of energy. Carbohydrates are mainly sugar and starches that the body breaks down into glucose (a simple sugar that the body can use to feed its cells). Although a number of classification schemes have been devised for carbohydrates, the division into four major groups monosaccharides, disaccharides, oligosaccharides, and polysaccharides. Most monosaccharides, or simple sugars, are found in grapes, other fruits, and honey. Although they can contain from three to nine carbon atoms, the most common representatives consist of five or six joined together to form a chainlike molecule Oligosaccharides, which consist of three to six monosaccharide units, are rather infrequently found in natural sources, although a few plant derivatives have been identified. Two molecules of a simple sugar that are linked to each other form a disaccharide, or double sugar. The disaccharide sucrose, or table sugar, consists of one molecule of glucose and one molecule of fructose; the most familiar sources of sucrose are sugar beets and cane sugar. Polysaccharide represent most of the structural and energy-reserve carbohydrates found in nature. Large molecules that may consist of as many as 10,000 monosaccharide units linked together, polysaccharides vary considerably in size, in structural complexity, and in sugar content; several hundred distinct types have thus far been identified Reducing sugars are sugars where the anomeric carbon has an OH group attached that can reduce other compounds. Non-reducing sugars do not have an OH group attached to the anomeric carbon so they cannot reduce other compounds. The Benedict's test heats a mixture of Benedict's reagent (a deep-blue alkaline solution) and sugar. If a reducing sugar is present, the reagent changes color: from green to dark red or rusty-brown, depending on the quantity and type of sugar. If you add a non-reducing sugar, like sucrose, the reagent remains blue. III. RESULTS AND DISCUSSIONS A. Test for Carbohydrates Samples Glucose Fructose Maltose General Test for Carbohydrates Molisch Test Observation Result (+/-) Purple ring + at the junction of two liquid Purple ring + at the junction of two liquid Purple ring + at the junction of two liquid Tests for Reducing Action of Carbohydrates Fehhling’s Test Observation Result (+/-) Brick red + precipitate Benedict’s Test Observation Result (+/-) Brick red + precipitate Brick red precipitate + Brick red precipitate + Brick red precipitate + Brick red precipitate + Sucrose Lactose Starch Cellulose Purple ring at the junction of two liquid Purple ring at the junction of two liquid Purple ring at the junction of two liquid Purple ring at the junction of two liquid + No precipitate - No precipitate + Brick red precipitate + Brick red precipitate + No precipitate - No precipitate - No precipitate - No precipitate - + + Discussions: 1. Which samples are positive for Molisch test? Which samples are negative? What do your results indicate about your samples? - They are all positive for Molisch test. 2. What is your result for fructose on Fehling’s and Benedict’s test? Is fructose a reducing sugar? Why? - Fructose on Fehling’s and Benedict’s test the same results which is brick red precipitate. A reducing sugar is any sugar that is capable of acting as a reducing agent because it has a free aldehyde group or a free ketone group. Ketoses must first tautomerize to aldoses before they can act as reducing sugars. The common dietary monosaccharides galactose, glucose and fructose are all reducing sugars. 3. What is your result for sucrose on Fehling’s and Benedict’s test? Is sucrose a reducing sugar? Why? -In Fehling’s and Benedict’s test, sucrose does not change in color. Sucrose is a non-reducing sugar because the two monosaccharide units are held together by a glycosidic linkage between C1 of α-glucose and C2 of β-fructose. Since the reducing groups of glucose and fructose are involved in glycosidic bond formation sucrose is a non-reducing sugar. 4. Maltose, sucrose and lactose are all disaccharides. Do they give the same results on Fehling’s and Benedict’s test? Why or why not? - Maltose and lactose gave the same results on Fehling’s and Benedict’s test but sucrose did not change in color. The acid breaks down the sucrose, the base does not. Sucrose does not react with Benedict's so when the base test tube stays blue we know there is still sucrose present and it did not break down. 5. Starch and cotton are polysaccharides. What do your results on Fehling’s and Benedict’s test reveal about polysaccharides? -The result of starch on Fehling’s and Benedicts test they did not change in color which has no precipitate. B. HYDROLYSIS OF CARBOHYDRATES Hydrolysis of Sucrose Observation to Benedict’s test: Brick red precipitate Hydrolyzed sucrose: A positive Benedict's test is evidenced by the formation of a brownish-red cuprous oxide precipitate. Unhydrolyzed sucrose: Sucrose contains two sugars (fructose and glucose) joined by their glycosidic bond in such a way as to prevent the glucose undergoing isomerization to an aldehyde, or fructose to alpha-hydroxy-ketone form. Sucrose is thus a non-reducing sugar which does not react with Benedict's reagent. Do both hydrolysed and unhydrolyzed sucrose have the same results to Benedict’s test? Explain why. They are the same result on Benedict’s test because hydrolyzed sucrose give positive result while unhydrolyzed is negative. Hydrolysis of Starch Observation to Benedict’s test: Brick red precipitate Hydrolyzed starch: starch will be hydrolyzed to shorter polysaccharides, dextrins, maltose, and glucose. When a blue solution of Benedict's reagent is added to a glucose solution, the color will change to green (at low glucose concentrations) or reddish-orange (at higher glucose concentrations). Unhydrolyzed starch: Iodine proved that there was no longer any starch in the solution by staying a red/brown color in the solution Do both hydrolysed and unhydrolyzed starch have the same results to Benedict’s test? Explain why. No, the test tube will be more dark red/brown, indicating that the hydrochloric acid hydrolyzed the starch in the solution and produced greater quantities of single sugars. C. IODINE TEST Sample Starch Agar-agar Gum Arabic Observation A deep blue/blue black Yellowish brown yellow Discussions: 1. What is the significance of the iodine test? What is being tested when you perform the test? Iodine staining distinguishes starch (a polysaccharide) from monosaccharides, disaccharides, and other polysaccharides. The basis for this test is that starch is a coiled polymer of glucose. Iodine interacts with these coiled molecules and becomes bluish black. Other non-coiled carbohydrates do not react with iodine. Therefore, a bluish black color is a positive test for starch, and a yellowish brown color (no color change) is a negative test for starch. Conclusion: Not all carbohydrates are reducing sugar some of it are non- reducing sugar. The common dietary monosaccharides galactose, glucose and fructose are all reducing sugars. Disaccharides are formed from two monosaccharides and can be classified as either reducing or nonreducing. Carbohydrates are also known as sugars or saccharides. Hydrolysis is a reaction with water. Acid hydrolysis of disaccharides and polysaccharides produces monosaccharides by breaking the glycosidic links (ether bonds) between monomer units in the structure of the molecule. References: https://www.flinnsci.com/api/library/Download/962c97b1f85d4627b535f24cdde91c5f https://laney.edu/cheli-fossum/wp-content/uploads/sites/210/2012/01/10-Enzymes.pdf https://msu.edu/course/lbs/145/luckie/Lab1.html https://www.ausetute.com.au/hydrolysiscarbs.html
