Invertase

Safety

Results

Methods

Invertase is easy to obtain as a commercial preparation or from a culture of yeast and can be used for a variety of safe and simple yet interesting and rewarding investigations

Invertase, (β-fructofuranosidase, sucrase, EC 3.2.1.26), converts the disaccharide sucrose into the monosaccharides glucose and fructose by hydrolysing the bond between them.

Sucrose is a non-reducing sugar and gives negative results with Benedict’s and DNSA reagents. These reagents can therefore be used to monitor the enzyme by measuring the appearance of the reducing sugars glucose and fructose.

The reaction is particularly suitable for studying the immobilisation of enymes on alginate beads.

Invertase is found in the lining of the small intestine but the usual commercial source is yeast. Commercial preparations are available but the enzyme activity can easily be shown by culturing yeast and a certain amount of purification is relatively simple by precipitation with propanone, (acetone).

Sample results

The results shown here were obtained by incubating 1cm³ of invertase solution prepared from yeast with 9cm³ of 0.4M sucrose solution in citrate buffer pH6 at 35°C
0.3cm³ samples were removed and added to 0.3cm³ of DNSA reagent.
After about 10 minutes the sucrose has all been converted to the reducing sugars glucose and fructose.

Preparation of invertase

Invertase can be obtained from dried yeast sold for home baking.

We have obtained the best yield using the following recipe and growing for six hours at 35°C.

• Dried yeast 10g
• Sucrose 5g
• Yeast extract* 1g
• Peptone* 1g
• Water to 50cm³.
*Yeast extract and peptone can be replaced with 2g Marmite or Vegemite

Once growth has finished and the culture has separated out the clear liquid on top should contain a high concentration of invertase.

If a centrifuge is available a short spin will remove any remaining yeast cells.

Precipitation of invertase with propanone, (acetone)

Grow a culture of yeast and centrifuge it to separate the cells. Add the cell free supernatant to propanone in the ratio of 5 parts supernatant to 3 parts propanone. A cloudy precipitate will be observed. Centrifuge to separate the precipitate and resuspend it in water or buffer.

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