i would like to to extract the secondary metabolites trapped in the fungus mycelium without homogenizing them ?
Any ideas
A suitable procedure would consist of taking the dry-pressed mycelium and directly put it in ethylacetate (or your solvent of choice) and let it stir over night. Of course, efficiency of extraction would be much higher, if you are able to homogenise the mycelium directly in your solvent of choice. After incubation you may continue with a second cycle of extraction. Combine both fractions, dry by your method of choice and solve in a suitable solvent (generally methanol for RP-HPLC). Filter the solution prior to applying it to a HPLC column!!!
It depends on the nature of the metabolites you want to extract and wheter they are partially excrted into the surrounding merdium.
An easy way to start with is to incubate the fungus in solid medium. When the fungus has grown just add the dissolvent to the plate and incubate with gentle shaking for 10-15 min. Recover the dissolvent and analyze.
We work with different Penicillia and use PDA. After 7 days of incubation we wash the palte with methanol.
Hope this helps as starting point.
Dear Sir Friedrich thank you for your answer, actually I'm working on small scale there for there is no enough amount of mycelium to stir over night.
Dear Sir Luis, thank you for your answer, it's a very good idea to add solvent and then incubat it for 15 minutes shaking, but how about the secondary metabolites which are impeded in between the mycelium layers ?? How to get them out..
In the case you want to analyze intracellular secondary metabolites you have to disrupt the cells. But you can do so even with a small amount of tissue by using a cell disruptor with glass-steal beads using 1.5-2.0 ml tubes.
Just overlay the agar culture of the fungus with solvent (Ethylacetate + Methanol 100:1). Leave the solvent for 24 h, collect it and re-extract after 24 h. Pay attention: your culture container must be glass or plastic resistant to the solvent you use. Use anhydrous Na2SO4 to adsorb water, evaporate the solvent and purify the extract.
Dear Rasha,
Attached is an example, for both mycelial exopolysaccharide (mostly water-soluble, if its water-insoluble - sulfation is needed) and intracellular polysaccharide (simply autoclave the mycelial biomass and proceed just like exopolysaccharide procedure).
Article Optimisation of biomass, exopolysaccharide and intracellular...
Hope it helps
Best wishes
Wan
- Badges
- Science topic
- Similar topics
- Phytochemistry
- Extracts