I am working on isolation, purification and characterization of biomolecules from leaf extracts. I have methanolic extract of leaf and I want to remove the chlorophyll from it first.
Extract your plant material with hexane to remove chlorophyll followed by polar solvents to get secondary metabolites.
Pour water into a medium-sized pot and set it on the stove. Bring the water to boil and set your leaf in it for two minutes. Remove the pot from the heat and take the leaf out. Pour 1 cup rubbing alcohol (Surgical spirit) into a tall, heat-safe glass and set it in the middle of the pot of hot water. Place the leaf in the glass with the rubbing alcohol. Make sure the leaf is entirely covered with the rubbing alcohol. Wait one hour and return to check on the leaf. The leaf should now show its true color. The rubbing alcohol will now be green. The green in the rubbing alcohol is the removed chlorophyll.
Thank Dr. Devang and Dr. Robin I will try your suggestions.
With regards
I have tried passing the extract through activated charcoal but other components may be lost in the process.
Carbon tetrachloride is the appropriate solvent for removal of chlorophyll from the extract without losing polar secondary metabolites. However, it may take away certain non-polar compounds.
Hello Dr. Devang J. Pandya Sir, Thanks for sharing ur knowledge here, Even i am also doing some work based on plant extract preparation.
Sir, Plz could you share any Link/ Research paper where related stuff will be given?
Here are some informative links on chlorophyll
https://www.researchgate.net/post/How_could_we_remove_chlorophyll_from_plant_extracts_during_purification_of_natural_compounds?tpr_view=cf963c49-6fdb-43d5-a01f-f1e47d31e8b6_1
https://www.researchgate.net/post/How_can_I_remove_chlorophyll_from_leaf_extract
https://www.researchgate.net/post/I_would_like_to_know_about_the_method_of_separating_chlorophyll_from_ethanolic_extracts_from_celtis_africana#557153915dbbbd835b8b4571
https://www.researchgate.net/post/How_can_I_measure_the_concentration_of_the_chlorophyll_in_the_eluent?tpr_view=8d0fb62a-4df7-4d4b-a492-edd46f43f3cb_2
https://www.researchgate.net/post/Can_I_use_oven-dried_plants_for_chlorophyll_Soxhlet_extraction?_trid=55409d7ad2fd64cb628b45b0_
https://www.researchgate.net/post/How_can_I_remove_chlorophyll_from_plant_extract?tpr_view=12502d85-0415-4a8a-9419-c5db2a876cb3_2
https://www.researchgate.net/post/What_is_the_best_way_to_isolate_leaves_with_a_lot_of_chlorophyll
https://www.researchgate.net/post/Before_extraction_from_leaf_material_how_can_I_get_rid_of_pigments_like_chlorophyll
Hello Punit R bhatt.
Thank you for your reply. Could you please elaborate me what you have written. I am working on extracting metabolites from barley plant. I extract the leaves , grind them and then add methanol and allow it to rotate for one hour in dark. Later, I evaporate it using rotatory evaporator. You mentioned to " add water", but this will not dissolve the grinded extracts which are present in the flask. Could you please throw some light on the procedure you follow? ALSO, please answer my following questions:
1. What do you meant by 80% water?
2.What is s.funnel?
Thanks,best
Timsy
This link is really helpful regarding leaf extract synthesis,http://www.instanano.com/uncategorized/leaf-extract-for-nanoparticles-synthesis/
first Extract with n-hexane then water. Both are used one by one for removing chlorophyll. Both solvents will not remove secondary metabolites from an extract which are methanolic in nature.
Hello Timsy,
Do you find a solution for it? If so, could you please elaborate.
you can remove chlorophyll by the following step:-
1. dry out the methanolic/ethanolic extract and then dissolved it in a sterile milliq or distilled water.
2. after dissolving with water some of the particles will not dissolve. So, to dissolve it you add hexane(analytical grade) double the amount of water and mix it and pour it in a separation funnel.
3. two layer will form the upper layer is hexane layer which will contain chlorophyll and it will turn dark green and the lower layer is aqueous layer. collect the aqueous layer in a separate beaker and hexane layer in a separate beaker.
4. to remove more traces of chlorophyll from aqueous layer than you have to add the equal amount of chloroform and follow the step 3
finally you will get the chlorophyll free aqueous fraction which will have 80% of secondary metabolite which are polar and semi polar.
hope this will help you. best of luck.
UV, but you will degrade the material you are after. However, if just Hemp Seed oil is your target to remove the green color UV light is great. Peroxide formation not likely but something to check. CBD unfortunately doesnt like UV light.
Timsy -- Excuse me to give some explanation, maybe Punit R Bhatt
describe about partition using separatory funnel. After extraction with methanol the next step is partition with N-Hexan or other non polar solvent (it depends on the kind of extract) and then add some water to clearly separation of polar and non polar phase- Badges
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- Biological Science
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