I would like to know the best protocols for:
quantitative and qualitative phytochemicals from powder or extracts of seeds from medicinal plant
and for extraction of secondary metabolites by soxhlet , microwave, ultrasonic and supercritical fluid and their analyses by chromatographic and spectrometry methods
Thank you for your helpful
Amel Nait Marzoug There is no fixed protocol for phytochemical analyzes. You can follow either WHO guidelines (readily available online) for the same or refer "Phytochemical analysis" by Harbone. In short it is an experience based approach.
1- Try to collect the plant by yourself
2- siscribe the season, time and the place,
because these three points effect the
quantity of the extracts.
3- Classify the plant potanicaly.
4- Dry at shade and open place.
5- Start extraction with non-polar solvent
(n-hexane or petroluim ether) then increase the polarity of the solvent,ethyl acetate, methanol and 70:30 methanol:water.
6- Take TLC for each extract to know the
number of the components.
7- Sepatate using different types of chromatography, besr one is flash (mplc).
8- Separate single compounds and identify
by H nmr ,C nmr, ms...etc
9- Test their biological activities....
Be specific in your plants ( which are all the medicinal plants you have to study belongs to family, genus and species etc.). Study their chemotypic classification whether belongs to leaf, stem or roots with specific related species. Then the analysis part comes. Links below will take you to the standard classic book for phyto-extraction.
https://www.springer.com/in/book/9780412572609
https://link.springer.com/book/10.1385/1592599559
Regards
Surendra Kumar Swarnkar thanks but what about the protocols of extraction by soxhlet , microwave, supercritical fluid ? and their analyzes by GC-MS or other methods , i mean the evaluation of phytochemical constituents by GC-MS for example and i want please your opinion about Peganum harmala
H.N. Nagendra Prasad Thanks, yes i am specific in my choice but the plant which i have chosen has been used extensively what about the originality ?
Faiq H S Hussain Thanks ,what do you mean about time?and the collection of the plant by myself
my problem is : the medicinal plant which i have chosen has been used extensively .I'm forced to do methods that have not been used before and the big problem is that i have to to these abroad we don't have this methods in our laboratory
Can i talk to you privately please ?
Dear Amel
I am sorry ,I am in holiday now till 24th March. abuot time ( morning, afternon, night...) ,the best time for terpenoids( volatile compounds) is morning before sunrise, high B.P. compounds in any time...etc
About collection I mean the plant should be fresh , if buy at market it is not scientific route, may be very old and most of volatile compounds are evaporate and some compds. their structure are changed by the effect of sun light and moisture or may be the plant micture of some species from same family.
Dear Amel,
The medicinal plants you are working with belong to which family? Depending on the family of the plant, you can find specific secondary metabolites. One class of seconday metabolite you can find more easily is the flavonoids and they are commonly analyzed by liquid cromatography. Do you have acces to this equipment? I do not know whether you have already worked with secondary metabolites in GC. Does your lab have a library to identify seconday metabolites? I am asking you because in my lab, we only have NIST and it was not good for secondary metabolites identifcation. However, the identification of primary metabolites was better.
Alice Nagai
Dear Alice
we don't have a library to identify secondary metabolites by GC but you have achance to do it in other laboratory
for the plant i want to find all the metabolites that are found for example in the seeds or leaves of the plant specially alcaloides
Faiq H S Hussain
Dear professor
Good holiday and i apologize for the inconvenience
what is the best time for polyphenols , flavonoids and alcaloids
and i will collect the plant on ground , i will not buy it from super market
Regarding quantitative and qualitative phytochemical analysis. For quantifying secondary metabolite such as total phenolic contents you can use Follin- Ciocalteu methods and for Flavonoid you can use aluminium chloride methods.
while for qualitative phytochemical analysis you can simply use the standard method of Treas and Evans. I hope this helps.
Abdullahi Hassan Ndanusa
Thanks but what about alcaloids and the extraction with soxhlet, microwave..
If you want to extract the secondary metabolites, the best method is extraction, fractionation and separation by column method, as well as preparative HPLC, and just want to identify then use GC-MS
Bakhodir S. Okhundedaev
Thanks
what are the best protocols to do the extraction of polyphenols ,flavonoids and alcaloids with different methods please?
to identify the secondary metabolites i can use just GC-MS what about GC-FID
is there other methods to identify the molecules please
thanks for your helpful
Dear Amel, First, you can dissolve the chosen plant parts (Powder) with ethanol or ethanol:water (80:20) for getting crude extract. Then crude extract subsequently fractionated with different solvents (Petroleum ether, Ethyl acetate, n-butanol). If you will follow bioassay-guided fractionation that would better for you. Thereafter, each fraction could be separated by preparative TLC, HPLC or column chromatography to isolate the compounds. Depending on yielded fraction you can choose whether you will identify the compounds by analysing GC-MS or isolate and identify the compounds by NMR and Mass spectrometry.
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