Dear colleague,

when you need to test the ability of a certain microbial strain to synthetize lipases, it is necessary to include some triglyceride in the culture medium. One medium which is excellent for that purpose is: 8 g/L trypticase peptone, 4 g/L yeast extract, 3 g/L NaCl, 20 g/L agar and 10 g/L tributyrin. You have to emulsify with an ultraturrax prior to sterilize in an autoclave. The presence of lypolitic activity is detected by the appearance of a transparent halo around the colonies.

This assay is indicated for the detection of esterase activity. If you want to confirm the presence of a "true" lipase you have to use a long chain triglyceride. In this case, you have to use the following medium: 8 g/L trypticase peptone, 4 g/L yeast extract, 3 g/L NaCl, 20 g/L agar, 30 mL/L of olive oil and 2 mg/L rhodamine B. The presence of lipase activity is detected by the appearence of orangish fluorescent halos.

I hope that the protocol can be useful for you.

Best regards

Dear colleague,

when you need to test the ability of a certain microbial strain to synthetize lipases, it is necessary to include some triglyceride in the culture medium. One medium which is excellent for that purpose is: 8 g/L trypticase peptone, 4 g/L yeast extract, 3 g/L NaCl, 20 g/L agar and 10 g/L tributyrin. You have to emulsify with an ultraturrax prior to sterilize in an autoclave. The presence of lypolitic activity is detected by the appearance of a transparent halo around the colonies.

This assay is indicated for the detection of esterase activity. If you want to confirm the presence of a "true" lipase you have to use a long chain triglyceride. In this case, you have to use the following medium: 8 g/L trypticase peptone, 4 g/L yeast extract, 3 g/L NaCl, 20 g/L agar, 30 mL/L of olive oil and 2 mg/L rhodamine B. The presence of lipase activity is detected by the appearence of orangish fluorescent halos.

I hope that the protocol can be useful for you.

Best regards

Dear Ana Rodriguez,

Thank you very much.

You are welcome

Dear Ana Rodriguez,

Can we sterilize Rhodamine B containing medium by autoclave  ?

Ana Rodriguez mam is right incorporating a triglycerides in to a agar media and its detection by distinct halo zone is the best method to screen lipase on petri plate. Triglycerides along with dye like rhodamine B ,spirit blue  will also help to screen lipase on plates .Where in the released fatty acids are visualized by their distinct colour due to hydrolysis of lipidic substrates. 

Dear colleague

Which temperature will ı use for ıncubation and how many days will ı ıncubate ?

Dear college  can used methylene blue instead of  RhodamineB 

Dear Amany 

How wiil we observe the activity of lipase with methylene blue ? Wiil it give  fluorescent ? I will need  method without uv light..

The best thing using rhodamine b , which can give clear result than any other stain.

 Thank you for your attention but  how do you dissolve the oil in media 

I have a question regarding the detection of lypolitic activity. If you mixed olive oil in an agar culture medium, would you be able to observe halos arond colonies that have a lypolitic activity , or would you need another method ?

If microorganisms produce lipase, you will see orange colour under uv light because of interaction of rhodamine B with fatty acids released during the enzymatic hydrolysis of triglycerides.

Dear Colleague,

I have question .. To test lipase activity of microorganisms on a petri dish, can we sterilize Rhodamine-B containing medium (medium + Rhodamine-B) by autoclave .. ? or should we sterilize separately .. ?

Best regards

Use tributyrine (1%) in mineral based agar plate, lipase will hydrolyze this and a clear zone of hydrolysis will appeared in lipase isolate.