To compare quantitatively the concentrations of glucose and other reducing sugars in samples of fresh orange, lemon and grapefruit juice. The standard test for glucose (and other reducing sugars) is to use Benedict's reagent

Tessie Taylor 12 I

Glucose concentrations in orange, lemon and grapefruit juice

Introduction

In this experiment, I am going to have to compare quantitatively the concentrations of glucose and other reducing sugars in samples of fresh orange, lemon and grapefruit juice. The standard test for glucose (and other reducing sugars) is to use Benedict’s reagent. Benedict’s reagent is copper (II) sulphate in an alkaline solution, and so has a blue colour to it. If it is added to a reducing agent, its Cu2+ ions will be reduced to Cu+, resulting in the precipitant changing colour to the red of copper (I) sulphate. Reducing sugars have this effect on Benedict’s reagent because they have a –C=O group somewhere in their molecules which can contribute an electron to the copper, hence the name “reducing sugars” (1). If a reducing sugar is present when Benedict’s solution is added, the solution will change colour through green, yellow and orange, to brick red as the copper (I) sulphate forms a precipitant.

All fruit contains the sugars glucose and fructose. Fructose is an isomer of glucose, so both sugars have the molecular formula: C6H12O6. Both are monosaccharides, and are hexoses (they both have 6 carbon atoms in each molecule). Both glucose and fructose are reducing sugars, so I would really be finding the concentrations of both fructose and glucose in my fruit juices rather than just glucose, but for ease, I will only talk about glucose from now on.

Method

Before I test for the unknown concentrations in my three fruit juices, I need create a calibration curve to compare my unknown concentrations against.

Apparatus needed

14 x test tubes – two test tubes for each concentration; one for before filtering and one for the filtrate.
1 x colorimeter – this will allow very accurate readings of the concentration of unprecipitated blue Cu2+ ions in the filtrate, as is computerised. This removes any error obtained through human error.
7 x cuvettes – one for each concentration will prevent contamination.
2 x 1 cm3 syringe – these will ...

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Method

Before I test for the unknown concentrations in my three fruit juices, I need create a calibration curve to compare my unknown concentrations against.

Apparatus needed

14 x test tubes – two test tubes for each concentration; one for before filtering and one for the filtrate.
1 x colorimeter – this will allow very accurate readings of the concentration of unprecipitated blue Cu2+ ions in the filtrate, as is computerised. This removes any error obtained through human error.
7 x cuvettes – one for each concentration will prevent contamination.
2 x 1 cm3 syringe – these will allow a large degree of accuracy in measuring volumes of the different liquids. One for each liquid will prevent any contamination.
1 x 10 cm3 syringe – this will allow the measuring of the Benedict’s solution to be as accurate as possible.
1 x measuring cylinder – this will allow accurate measuring of the distilled water.
Filter paper – this will filter as much of the precipitant as possible.
Water bath – this will ensure that all of a solution will be heated equally and so fairly.
Test tube rack – for safe storage of test tubes.

Measure 2 cm3 of 4% (by mass) glucose solution in a 1 cm3 syringe and add it to a clean test tube.
Take 1 cm3 of the 4% glucose solution from test tube 1 and add it to test tube 2.
Measure 1 cm3 of distilled water in a measuring cylinder and add it to test tube 2. This will create a 2% glucose solution.
Wash the syringe in between each solution to avoid any contamination and to ensure that my concentrations are as accurate as possible.
Repeat this operation for tubes 3, 4, 5, 6 and 7. Once the solution has been mixed in test tube 7, I will remove 1 cm3 of the solution so that there is an equal volume of test solution in each test tube – this will make the test fair.
I will now have glucose concentrations of 4%, 2%, 1%, 0.5%, 0.25%, 0.125% and 0.0625%. These are known as serial concentrations. This is a good range of test solutions, as I would have a strong solution, a very weak solution and a lot of concentrations in the middle so that I can draw an accurate line of best fit, and so that my calibration curve is generally as accurate as possible. In previous work, I found out that Benedict’s solution stops detecting glucose between 0.1% and 0.01% glucose concentration, so it should still detect glucose at my lowest test concentration of 0.0625%.
Add 10 cm3 of Benedict’s solution to each test tube using a 10 cm3 syringe (I will use the same concentration of Benedict’s with each test solution in order to keep the test fair).

The reason for using excess Benedict’s solution is so that all of the sugar present will have definitely reacted (1).

Put all of the test tubes in a water bath at 75°C for 10 minutes. This will allow the Benedict’s to react as much as possible, resulting more accurate test concentration readings. heat all of the test solutions for an equal length of time in order to keep the test fair.
Filter each concentration using filter paper into new test tubes.
Then transfer equal volumes of each filtered solution into separate cuvettes so that the cuvettes are more than half full.
Place filter no 7 (orange) into the colorimeter and I will set the colorimeter to zero using the 4% solution cuvette.
Repeat each test solution 3 times in order to find the mean colorimeter readings for each of the glucose concentrations.

Recording results

I will record my results for the test solutions in a table like below.

In order to convert % concentration into mg cm-3 using 1 cm3 of solution, you must multiply the % by 10 e.g. there would be 40 mg cm-3 of glucose in 1 cm3 of 4% glucose solution.

Risk assessment

Wear a pair of goggles as Benedict’s solution is harmful.
Do not splash the test tubes when putting them in and out of the water bath, as the water is hot enough to burn.
Let the solutions cool sufficiently in a test tube rack before filtering them in order to prevent any burns if the solution splashes.

How to solve original question

Prepare the three fruit juices by filtering the freshly squeezed juice of each fruit so that all fruit bits are removed (these may affect any results I get).
Add 1 cm3 of the different juices to three different clean test tubes.
Add 10 cm3 of Benedict’s solution to each test tube.
Add all three solutions to a 75°C water bath for ten minutes.
Filter each fruit juice into clean test tubes.
Using a clean syringe, add equal volumes of each filtered juice into separate cuvettes so that the cuvettes are more than half full.
Check that filter no 7 (orange) is in place in the colorimeter, and take one colorimeter reading for each cuvette.
Repeat this part of the experiment three times as well in order to get a mean transmission of light for each fruit juice.
Estimate the concentration of each juice by reading off the concentration at each average transmission of light.

I will record my results on a similar table to the one on which I will record my test solutions on, but instead of having a column with known concentrations in, I will have the column with the name of the fruit juice.

Prediction

I predict that the lemon juice will have the smallest concentration of glucose, and the orange juice will have the largest. In previous research involving orange and lemon flavoured drinks, I have found out that the orange drink had a higher concentration of glucose than the lemon drink.

The only problem that may occur in using this method is the colour of the fruit juices may affect the colorimeter reading, but as there will be a very large volume of Benedict’s solution used compared to fruit juice volume, these effects should be kept to a minimum.

Bibliography