I am planning to extract the fungal metabolites which cultured from the potato dextrose broth (PDB) with solvent then run through TLC. I am just wondering, when we extracted the fungal compounds from PDB with solvent, is there any possibility that materials from PDB is extracted together too instead of only the fungal metabolites? Thanks.
is it solid culture media ?
if yes:
You will have for sure some components from PDB in the extract. to identify them you should do an blank extract of your culture medium alone (without fungi).
If you use a too polar solvent you will extract sugar from the media as well but you will also extract polar secondary metabolites.
There is many extraction solvant you can use. You will need to adapt your solvent system for TLC according to the solvent you use, or the other way.
you can look to those publication :
- https://www.researchgate.net/publication/235119345_Detection_of_metabolite_induction_in_fungal_co-cultures_on_solid_media_by_high-throughput_differential_ultra_high_pressure_liquid_chromatographytime-of-flight_mass_spectrometry_fingerprinting?ev=prf_pub
- https://www.researchgate.net/publication/45276107_Platotex_an_innovative_and_fully_automated_device_for_cell_growth_scale-up_of_agar-supported_solid-state_fermentation?ev=pub_cit
- https://www.researchgate.net/publication/14149059_Micro-scale_extraction_procedure_for_standardized_screening_of_fungal_metabolite_production_in_cultures?ev=pub_cit
Article Detection of metabolite induction in fungal co-cultures on s...
Article Platotex: An innovative and fully automated device for cell ...
Article Micro-scale extraction procedure for standardized screening ...
You should also run standards of the fungal metabolites and refer to tlc data bases of fungal metabolites like the one I published in J Chromat in the 1980s. You should use the same tlc systems of course. You should have told us which fungi you will be using. Remember you can use 2D tlc too.
if you are planning to extract the fungal metabolites, after filtration via Whatmann filter paper, pass it through 0.22um NCM membrane (make it cell free), then lyophilize it, reconstitute the residue in 10-15 ml of sterile distilled water and then start extracting sequentially with 5 volumes of the solvent based on its polarity (polar- slightly polar- slightly nonpolar-non polar), wash the residue with sodium sulphate and then evaporate the solvent at 37C. Reconstitute it either in DMSO/methanol or desired solvent. Repeat the above procedure for control (PDB) and then check their bio activity. All The best............
Thank you for the suggestion. For your information, I'm using Trichoderma
I run the blank on TLC already. Seems that the one that I thought is the major compound from trichoderma appears in the blank too. Which is PDB only.
you are working with liquide medium, you should let your culture enough incubation time (generally about 10 days) to make sure that the components(specially carbohydrate/ dextrose) of your medium were minimized and consumed by the fungus and that it produce more metabolites. then filtrate and concentrate your culture, you can directly strart extraction (liquide/lquide) with solvent that are not miscible to water or lyophilize your culture if you have to use miscible solvent. If the problem persist you can extract metabolites from scarped biomass on solide culture meduim , that works good with trichoderma.
Hello Lee Pei
Your question is not fairy specific, it is better if you mention the aim, about what you are going to do. Any way I attached some references I hope to be useful for your question
Good luck
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