Qualitative observations were made on five powders using three different chemical indicators (Liquids #1, #2 & #3). Liquid #1 one was acetic acid (vinegar). Liquid #2 was Lugol's reagent (iodine and potassium iodide). Liquid #3 was Benedictls reagent (copper sulfate). Three of the five powders (A, B & C) were used as standards to identify two unknown powders (X₁& X₂). Powder A was the monosaccharide, glucose. Substance B was the polysaccharide amylose (plant starch) and C was sodium bicarbonate (baking soda). X₁ & X₂ were thought to contain only powders A, B or C, or mixtures of A, B and C.

METHODS AND MATERIALS:

1. Nine test tubes of two different types are needed to test powders A (glucose), B (amylose) & C (sodium bicarbonate) in this investigation. Six small (60 mm X 5 mm) glass tubes will be used to test all three powders with Liquids #1 (acetic acid) and #2 (Lugol's reagent). The remaining three tubes should be Pyrex, or Kimax as they will be used to test the powders with liquid #3 (Benedict's reagent). The Liquid #3 tests require a boiling water bath for three to five minutes.

The small glass tubes are disposable, so they may be thrown away after the tests have been performed and read. The Pyrex/Kimax test tubes are reusable and will need to been washed out after using.

2. Each of the jars of powder (A, B & C) has a spatula attached to it by a rubber band. To avoid contaminating other samples, and getting false data, be sure to use ony the spatula on each bottle to remove the powder from that bottle. Be sure you replace the spatula on the same bottle after use.

Using the spatula, put approximately 3 mm. of powders A, B and C in three separate tubes.

a. Two small glass tubes of glucose.

b. Two small glass tubes of amylose.

c. Two small glass tubes of sodium bicarbonate.

d. Three Pyrex/Kimax test tubes:

a. One containing powder glucose

b. One containing powder amylose

a. One containing powder sodium bicarbonate

3. Each of the powders in the small glass tubes will be tested with acetic acid and Lugol's reagent.

a. Put three drops of acetic acid in tubes A, B and C.

b. Put three drops of Lugol's reagent into tubes A, B and C.

Pay close attention to the reaction that you get with the liquids. These indicators produce instant results. Describe and record the results.

c. Put enough Benedict's reagent into the Pyrex/Kimax test tubes A, B and C to just cover the powder. Place these test tubes in the boiling water bath and wait 5 minutes. Remove the tubes with a test tube holder. Place them in your test tube rack. Describe and record the results.

4. IF THERE IS CONFIDENCE AT THIS POINT THAT THE GLUCOSE, AMYLOSE AND SODIUM BICARBONATE CAN BE IDENTIFIED WITH THE ACETIC ACID, LUGOL'S REAGENT AND BENEDICT'S REAGENT, THEN USE THE SAME TESTS TO DETERMINE THE CONTENTS OF BOTTLES X₁ AND X₂. IF A DEGREE OF UNCERTAINTY EXISTS ABOUT THE TEST RESULTS ON A, B AND C, THEN REDO THE TESTS.

 

	Powder A Glucose	Powder B Amylose	Powder C Sodium Bicarbonate	Powder X1	Powder X2
Liquid #1 Acetic Acid	-	-	+ Gas Release (CO2)	-	-
Liquid #2 Lugol's Reagent	-	+ Black color	-	+ Black color	-
Liquid #3 Benedictls Reagent	+ Orange color	+ Green/Orange color	-	+ Green/Orange color	-

The powders X₁ and X₂ were as expected in one case, but not the other. The indicators #1, #2 and #3 were sufficient to identify substances A, B and C.

Reagent #1, acetic acid, reacted with C, sodium bicarbonate to produce a distinctive CO₂ (carbon dioxide) release. Powder B, amylose, turned black when three drops of liquid #2, Lugol's reagent, was added to the tube. The data shows some conflict with A, glucose, and B, amylose, using indicator #3, Benedict's reagent. It appears that glucose and amylose share some chemical relationship as they have results that are similar. The Benedict's reagent reacted with glucose after being in the water bath, and appears to have chelated (bound to) the molecules of glucose producing a copper-colored (orange) substance. Amylose had a similar result, but a greenish (as in oxidized copper) residue was also present with an orange (copper-colored) portion of amylose. The fact that amylose turned black with Lugol's reagent will confirm its presence in a test, but the results of glucose and amylose with Benedict's reagent was thought to be conflicting and could make the identification of X₁ and X₂ confusing.

X₁ had a positive response to Lugol's in that it turned black when three drops of reagent were added. Benedict's produced a substance that appears to be relatively the same as standard B with Benedict's after heating. All other tests on X₁ were negative. Testing X₂ with acetic acid, Lugol's and Benedict's produced none of the results seen in powders A, glucose, B, amylose and C, sodium bicarbonate. All of the tests on X₂ were negative.

Indicator #2, Lugol's reagent, confirmed that substance X₁ contained standard B, amylose. Indicator #3, Benedict's, supports that B, amylose, is present in X₁, and it alludes to A, glucose, possibly being in what could be a mixture of other powders. Tests with A, B and Benedict's were repeated with similar results. The evidence points at X₁ containing B, amylose, but does not rule out other substances being present in the jar containing X₁. With only three standards available to identify the chemical content of containers X₁ and X₂, the results of this investigation are seen as tentative and indicates that other testing should be done.

X₂ does not contain any of standards A,glucose, B, amylose, or C, sodium bicarbonate. All tests here were negative. Again limited by only three standards the identification of X₂ remains to be determined. More testing is needed on X₂, with other different standards, if a positive identification is to be made.

 

 

Benedicts's Test for Reducing Sugars        

Alkaline solutions of copper are reduced by sugars having a free aldehyde or ketone group, with the formation of colored cuprous oxide.  Benedict's solution is composed of copper sulfate, sodium carbonate, and sodium citrate (pH 10.5).  The citrate will form soluble complex ions with Cu++, preventing the precipitation of CuCO3 in alkaline solutions.                  

Method:  Add 1 ml of the solution to be tested to 5 ml of Benedict's solution, and shake each tube.  Place the tube in a boiling water bath and heat for 3 minutes.  Remove the tubes from the heat and allow them to cool.  Formation of a green, red, or yellow precipitate is a positive test for reducing sugars.

 

 Iodine Test for Starch and Glycogen        

The use of Lugol's iodine reagent (IKI-iodine potassium iodide) is useful to distinguish starch and glycogen from other polysaccharides.  Lugol's iodine yields a blue-black color in the presence of starch.  Glycogen reacts with Lugol's reagent to give a brown-blue color.   Other polysaccharides and monosaccharides yield no color change; the test solution remains the characteristic brown-yellow of the reagent.  It is thought that starch and glycogen form helical coils.  Iodine atoms can then fit into the helices to form a starch-iodine or glycogen-iodine complex.  Starch in the form of amylose and amylopectin has less branches than glycogen.  This means that the helices of starch are longer than glycogen, therefore binding more iodine atoms.  The result is that the color produced by a starch-iodine complex is more intense than that obtained with a glycogen-iodine complex.   

Method:  Add 2-3 drops of Lugol's iodine solution to 5 ml of solution to be tested.  Starch gives a blue-black color.  A positive test for glycogen is a brown-blue color.  A negative test is the brown-yellow color of the test reagent.

 

Sodium Bicarbonate and Acetic Acid

Baking soda is sodium bicarbonate. Vinegar contains acetic acid dissolved in water. Sodium barcarbonate reacts with most acids. The products of the reaction with vinegar are carbon dioxide gas, sodium acetate, and water.

The reaction of sodium bicarbonate to form carbon dioxide gas is the basis of its use as a levening agent in baking. Cakes are solid foams. The foam is produced when bubbles of carbon dioxide from the reaction of sodium bicarbonate are trapped in the batter. As the cake bakes, the batter dries, and the trapped bubbles of carbon dioxide form the holes in the cake.