Quantitative tests for reducing sugars

What type of data is obtained from quantitative tests

Numerical data

How can numerical data be obtained by benedict’s test

As the concentration of reducing sugar in a solution increases, the mass of red precipitate formed will also increase. After performing Benedict’s test the solution can be filtered or centrifuged and the precipitate can be removed. Numerical data can then be obtained by either measuring the dry mass of the precipitate or measuring the absorbance of the filtrate (supernatant) using a colorimeter.

What does a colorimeter measure

% transmission (how much light gets through) or % absorbance (how much light is absorbed).

Why are filters used in colorimeters

To restrict other wavelengths of light so more accurate data can be obtained.

What is a suitable blank to calibrate a colorimeter and when should it be used

Distilled water, colorimeter must be calibrated before each reading.

First step of determining glucose concentration

Prepare a calibration curve by taking a range of solutions of known concentrations of glucose.

Second step of determining glucose concentration

Carry out Benedict’s test for reducing sugars on the glucose solutions. Use the same volume of test solution and Benedict’s reagent each time and heat in a water bath at 80°c for 10 minutes.

Third step of determining glucose concentration

Filter (or centrifuge) to remove the red precipitate. Transfer the filtrate (supernatant) to a cuvette using a pipette.

Fourth step of determining glucose concentration

Select the red filter and calibrate the colorimeter using a distilled water as a blank before each reading. Measure the absorbance of each solution.

Fifth step of determining glucose concentration

Plot the concentration of glucose (x axis) against the absorbance (y axis) to obtain a calibration curve.

Sixth step of determining glucose concentration

Repeat the Benedict’s test on the unknown sample in the same way, filter to remove the precipitate and measure the absorbance as before.

Seventh step of determining glucose concentration

Read a value for the concentration of reducing sugar off the calibration curve.

Why must excess Benedict’s reagent be used

To ensure all the reducing sugar reacts.

How could the method be modified to determine the concentration of protein

Use buiret’s reagent