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Investigation of action of saliva and hydrochloric acid in two carbohydrate solution

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Procedures: 1. Prepared two boiling tubes with containing 1 ml solution A and 1 ml solution B respectively. 1 ml Benedict’s solution was added to each tube and heated both tubes together in the (~95°C) water bath for two minutes. Then, recorded the results in table 1.

2. Added a few drops of fresh solution A and B separately spaced on a white tile. On each solution, added 1-2 drops of iodine solution and mixed with pen cover. Recorded your observations in the table 1.

3. Pipetted 2 ml solution B into each of four boiling tubes. The tubes were labelled 1, 2, 3 and 4 respectively near mouth of tube. Labelled your group name.

4. Placed tubes 1 and 2 in a water bath of ~37°C.

5. Salivated into a small beaker until it reached about 5 ml.

6. At the same time, step (6) and (7) was to be done approximately. Measured out 4 ml of the saliva prepared in step (4) and pipetted 2 ml each into tubes 1 and 4. The contents of the tubes shook well to ensure through mixing.

7. Measured out 4 ml HCL and pipetted 2 ml each into tubes 2 and 3.

8. Let tubes 1, 2, 3, and 4 incubated at their respective temperatures (see Table 2) for 35 minutes from this moment.

9. Labelled 4 more new boiling tubes as follows: 1’, 2’, 3’ and 4’.

10. After 5 minutes of incubation of tubes 1 to 4, poured out about half of the contents from all these tubes into the respective newly labelled tubes (e. g., 1 into 1’, 2 into 2’ etc.). Straightaway, placed back the original tubes (labelled 1 to 4) back into the respective temperatures of incubation.

11. Benedict’s test performed on the contents of tubes 1’ to 4’ by pipetting 2 ml of Benedict’s solution into each tubes and heated them in 95°C water bath for one minute. The observations was recorded in Table 2.

12. Benedict’s test was carried out with an equal volume of Benedict’s solution (2 ml) for each tube. Remembered to heat the sample and recorded the observations.

All living organisms need nutrients to supply energy for their daily activities. A nutrient is a chemical that an organism needs to live and grow or a substance used in an organism’s metabolism which must be taken in from its environment. They are to build and repair tissues, regulate body process and are converted to and used as energy. Therefore, most nutrients are used to produce ATP such as carbohydrates, fat and the others. So, we need to break down foods into its most simple forms in order to absorb the nutrients through our digestive system.

In this experiment, we will test the action of saliva and hydrochloric acid in two carbohydrate solution with different of temperature. In this experiment, carbohydrate solution A acts a reducing sugar. Benedict’s test and iodine test were carried out at the same time. In the Benedict’s test, Copper(II) oxide is one of the components of Benedict’s reagent. When it is added in carbohydrate solution A(reducing sugar), the blue colour of copper(II) oxide is reduced to insoluble red-brown copper(I) oxide ions. That was caused by redox reaction because this reaction works as aldehydes nd oxidised the reducing sugar easily. Aldehydrates are oxidised to carboxylic acids. Benedict’s solution are weak oxidising agents. The copper(II) ion gains an electron to form copper(I) in the form of copper(I)oxide, Cu? O. A brick-red precipitate is formed. If the reducing sugar is present, the colour of solution would changed from blue to reddish-brown. In the iodine test, the yellowish of the mixture solution does not change colour after mixed with carbohydrate solution A(reducing sugar). That is because the absent of starch in the mixture solution.

At the same time, Benedict’s test and iodine test also carried out to test carbohydrate solution B, which is starch. In Benedict’s test, the benedict’s solution add into carbohydrate solution B but the reaction is not occur. It means the blue colour of the mixture solution remain the same after heating. That is because the benedict’s reagent only works on the reducing sugar, such as glucose, maltose, fructose, galactose, sucrose etc. Starch is not a reducing sugar but a polysaccharide. So, in order for starch be detected by benedict’s reagent if firstly has to be hydrolysed into its reducing sugar.

In the iodine test, iodine solution will react with carbohydrate solution B and change colour from yellowish to black-blue. That is because the complex of iodine stuck inside the amylose coil produces a characteristic black-blue colour when react with starch. Solution B represented starch. Starch also called complex carbohydrates and can be separated into two fractions–amylose and amylopectin. Natural starches are mixtures of amylose (10-20%) and amylopectin (80-90%). Starch is made up of groups of sugars that are bound tightly together. So, its structure is more complex and will break down into maltose during digestion.

Starch is composed of polymers of glucose and long linear chains are amylose. The higher the amount of amylose in a starch, the more slowly it is digested. This will cause blood glucose to rise less and over a longer period of time. Amylose coils into a helical secondary structure resembling a tube with a hollow core. Certain molecules including fatty acids and iodine can lodge inside the core as already mentioned. The complex of iodine stuck inside the amylose coil produces a characteristic blue-black colour. The starch itself is not altered. Starch-iodine complex becomes unstable at temperatures above 35°C.

This complex in presence of an oxidizing agent the solution turns blue, in the presence of reducing agent, the blue colour disappears because triiodide (I3- ) ions break up into three iodide ions, disassembling the complex. So starch turns into glucose molecules. Digestion begins in the mouth. The salivary glands secrete a liquid substance that contains various elements including water, mucin (a protein) and the enzyme amylase. This liquid called saliva. Amylase, an enzyme secreted by the salivary glands and the pancreas, rapidly hydrolyses amylopectin and amylose. Amylase works to begin the breakdown of polysaccharides to disaccharides.

The amylase in the mouth, salivary amylase, is called ptyalin. It like other enzymes, works as a catalyst. Amylase digests starch by catalysing hydrolysis, which is splitting by the addition of a water molecule. The body temperature (37°C) is the optimal heat for the best reaction of amylase. The process which can broken down the disaccharides and polysaccharides into their monosaccharide subunits called hydrolysis. Starch is polymer of glucose formed of glycosidic bonds. Thus, it can be broken either by enzymatic hydrolysis or acid hydrolysis only, and this hydrolysis would result into formation of monomers.

Glycosidic bond is ether linkage, having enough strength. This C-O-C bond, an ether linkage, can be broken through hydrolysis to release the constituent glucoses. In this experiment, hydrochloric acid (HCl) serves as an acid catalyst in water to hydrolyse starch and hydrochloric acid will puts H+ in the solution. The lone pairs of the oxygen in the C-O-C bond will attack this H+. Now oxygen is bound to three things; it is unstable because acid allows this destabilization to occur, it lets hydrolysis of the bond occur.? The temperature suitable for hydrochloric acid to hydrolysis the starch is among the body temperature.

That is because non-biological catalysts work at wide ranges of temperature and pH, whereas enzymes must work in mild conditions in cells, between 32°C and 40°C and at a pH between 6. 5 and 7. 5. Enzymes are highly specific both in the reaction catalysed and in their choice of reactants, which are called substrates. An enzyme usually catalyses a single chemical reaction or a set of closely related reactions. To catalyse a reaction, a substrate must have a matching shape into the active site of the enzyme. The products of the reaction then leave the active site, freeing it up for more similar reactions to take place.

The higher the concentration of enzyme, the faster the rate of reaction. In the Benedict’s test, there are 4 test tubes with the different contents and will heat in different temperature of water bath. At 37°C, the mix solution of test tube 1 change colour from blue to yellowish-brown. If the solution has a yellowish coloration, then a moderate amount of maltose is present (+++) . After 35 minutes, the colour changed from yellowish to greenish-brown because the amount of maltose increase. That was because the amylase enzyme in saliva speed up the process of break down the starch into maltose.

The mix solution of test tube 2 was remained the blue colour because the temperature (37°C) is too low and hydrochloric acid need more time to break down the starch. So, we can’t to observe the change in test tube 2 in a short time. At 95°C, the mix solution of test tube 3 change colour from blue to pale blue after 5 minutes of incubation because the pH of solution is acidic. Its mean if the more acid the solution, the less the colour formed. After 35 minutes, the colour change to reddish brown because hydrochloric acid speed up the process of breaking down the starch with the help of high temperature .

Then, the mix solution in test tube 4 also can change colour from blue to yellowish-brown after 5 minutes of incubation in 95°C water bath. That was because temperature rises slowly and give the time for amylase to break down the starch. After 35 minutes, the temperature is approximately achieve boiling point and amylase was denatured . So , the colour of test tube 4 became dark brown. There were a few of the precaution had to pay attention on it. Following the procedure when carry out the experiment in order to avoid getting the negative result.

Use the stopwatch to make sure the time taken of experiment to carried out. Make sure take the same volume of each test tube to get the accurate result. The temperature must be also measured correctly so the test tube is put into the water bath to desire the temperature was achieved. Be careful to the alkaline because it can affect the Benedict’s test. In conclusion, common test for reducing sugar is Benedict’s test. While test for break down of starch is added hydrochloric acid or saliva into Benedict’s solution with different temperature. Beside that, test of starch is Iodine test. Conclusion:

Benedict’s test was used to test reducing sugar , brick-red precipitate is formed. The colour of the solution changed from blue to reddish-brown. Then, use iodine test to test starch. The yellowish of the mixture solution was turned to black-blue in colour. In addition, Benedict’s test also used to test the starch which mix with saliva or hydrochloric acid in different temperature. The present of amylase in saliva effect the amount of maltose after break down the starch at the 37°C. The hydrochloric acid will also effect the amount of maltose after break down the starch but in high temperature.

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