testing for biological molecules (q)

Question 1

Describe Benedict’s test for reducing sugars

Answer 1

1. Place sample in boiling tube (if not liquid, then grind up and blend with water)
2. Add an equal volume of Benedict’s reagent
3. Heat the mixture gently in boiling water bath for 5 minutes

Question 2

What is Benedict’s solution?

Answer 2

An alkaline solution of copper (II) sulfate

Question 3

What are reducing sugars (examples)?

Answer 3

All monosaccharides + some disaccharides (e.g. maltose and lactose) are reducing sugars - they can donate electrons or reduce another molecule or chemical

Question 4

What order is the colour change if reducing sugars are present?

Answer 4

Blue, green, yellow, orange, brick red

traffic lights

Question 5

Why does the solution change colour? (Benedict’s)

Answer 5

Reducing sugars react with the copper ions. This results in the addition of electrons to the blue Cu2+ ions, reducing them to brick red copper ions, indicating a positive result.

Question 6

Relationship between amount of reducing sugar and colour of solution

Answer 6

The more the reducing sugar present, the more precipitate formed and the less Cu2+ (blue) ions are left in the solution.

Question 7

Qualitative data for Benedict’s

Answer 7

The colour seen is a mixture of red and blue, depending on the concentration of the reducing sugar.

Question 8

Quantitative data for Benedict’s

Answer 8

Solutions can be filtered and the precipitate can be weighed. Or colorimeter can be used to give a quantitative value for the colour.

Question 9

Describe Benedict’s for non-reducing sugars

Answer 9

1. Get a new sample of test solution and add HCl
2. Carefully heat it in the water bath that’s been brought to boil
3. Neutralise it with sodium hydrogen carbonate
4. Carry out the Benedict’s test

Question 10

Why does the Benedict’s test not work on non-reducing sugars?

Answer 10

Because non-reducing sugars must be broken down into monosaccharides. They don’t react with Benedict’s, so remain blue at first. The non-reducing sugar must be hydrolysed by the acid to a reducing sugar.

Question 11

Describe reagent strip test for reducing sugars

Answer 11

• strips are dipped in a test solution and change colour is glucose is present
• colour change can be compared to a chart to give an indication of the concentration of glucose present.

Question 12

When are reagent strips used?

Answer 12

Useful for testing urine for glucose, for the diagnosis of diabetes.

Question 13

Describe a starch test

Answer 13

• add iodine dissolved in potassium iodide solution to the test sample

Question 14

Results of a starch test

Answer 14

• if present, sample changes from brown/orange to blue/black colour - positive result
• if no starch is present the sample stays brown/orange - negative result

Question 15

Describe emulsion test for lipids

Answer 15

1. Add the test substance to ethanol
2. Shake well
3. Add the solution to water

Question 16

Results for emulsion test

Answer 16

• if lipid is present, solution will turn milky. the more lipid there is, then more noticeable the milky colour
• if no lipid is present, the solution will stay clear

Question 17

Describe Biuret test for proteins

Answer 17

1. The test solution must be alkaline, so add a few drops of sodium hydroxide solution
2. Then add copper (II) sulphate solution
   (mixture of an alkali and copper sulphate solution is called Biuret reagent)

Question 18

Results of Biuret test

Answer 18

• If protein is present, solution turns purple

- if no protein is present, solution remains blue

Question 19

What is colorimetry?

Answer 19

• equipment used to quantitatively measure the absorbance or transmission of light by a coloured solution
• more concentrated solution = more light absorbed = less transmitted
• can be used to calculate concentration of reducing sugar present

Question 20

How to use colorimetry

Answer 20

1. Place a filter in the colorimeter
2. Calibrate with distilled water
3. Perform the test, e.g. Benedict’s for glucose concentrations
4. Filter solutions to remove precipitates
5. Measure % transmission for each solution
6. Plot a calibration curve

Question 21

What are biosensors used for?

Answer 21

They use biological components to determine the presence and concentration of molecules such as glucose.

Question 22

Components of biosensors

Answer 22

• molecular recognition: a protein or single strand of DNA is immobilised to a surface, which will interact with the molecule under investigation
• transduction: this interaction causes a change in a transducer, it detects the change and produces a response
• display: produces a visible, qualitative or quantitative signal

Question 23

Describe thin-layer chromatography to separate biological molecules

Answer 23

• stationary phase = thing layer of silica gel applied to a sheet of glass of metal
• mobile phase = organic solvent
• the molecules, e.g. amino acids, are added to one end of the gel; this end is submerged in the solvent
• they separate based on their interactions with the stationary phase + solubility in the mobile phase
• can be used to separate proteins, carbs, vitamins, nucleic acid

Question 24

Rf values

Answer 24

Rf = distance travelled by component / distance travelled by solvent

Question 25

Describe the procedure for TLC

Answer 25

1. Draw pencil line, handling the plate only by the edges
2. Spot the solution onto the pencil line, allow to dry and respot, label in pencil
3. Place the place in a jar of solvent lower than the pencil line and close the jar
4. Leave plate for a while, then draw the solvent front and allow the plate to dry
5. Spray with ninhydrin in a fume cupboard - amino acids react with ninhydrin forming a purple/brown colour