Is that true that crude extract should be subjected to column and fraction of that and be used for HPLC?
An full answer to your question is complicated.
It really depend on what polarity has your extract. Was it obtained by MeOH, CH2Cl2 ?
If you do C18 RP HPLC what you can do to be sure, is SPE on C18 cartridge prior to any sample HPLC analysis (your can look to that example among many other : 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).
By doing that you will remove any apolar compounds which will be stuck into your column. If you have very long sequences it may be very important to avoid retention time drift. Do not forget that if you column is too dirty (too much compounds stuck inside) you may have to clean it.
And it may take some time if you have injected very apolar samples.
To clean you can pass trough isopropanol for 30-60min. If it does not work well pas this sequence of solvent isopropanol, EtOAc, Hexane, EtOAc, isopropanol; with 30-60min for each steps.
If you just want to pass some samples (not large sequence) then there should nor be any problems as long as you do some king of cleaning regularly. Do not forget that pre-column will take most of your apolar compounds, anyway to some extend.
The main question to ask yourself before, is still which solvent did I used for the extraction? And why do I need to do HPLC analysis.
Article Detection of metabolite induction in fungal co-cultures on s...
Sure ... the problem is the durability of the column. It is advisable to make a open column fractionation in different media (different polarities) and then you, make fractionation. in analytical HPLC
Yes, you can use any type of samples depends on column for the separation of chemical constituents. Once you complete the analysis, make sure to wash the columns according to recommended solvents to extend the column life.
Sure, You can inject any type of samples in HPLC but make sure that according to sample you may choose different type of column. After completion of analysis make sure to wash the column properly with recommended solvent and one more thing don't forget to connect guard column along with analytical column. It is always extend the column life.
So far as the crude product is well filtered using appropriate filter, you can use it.
An full answer to your question is complicated.
It really depend on what polarity has your extract. Was it obtained by MeOH, CH2Cl2 ?
If you do C18 RP HPLC what you can do to be sure, is SPE on C18 cartridge prior to any sample HPLC analysis (your can look to that example among many other : 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).
By doing that you will remove any apolar compounds which will be stuck into your column. If you have very long sequences it may be very important to avoid retention time drift. Do not forget that if you column is too dirty (too much compounds stuck inside) you may have to clean it.
And it may take some time if you have injected very apolar samples.
To clean you can pass trough isopropanol for 30-60min. If it does not work well pas this sequence of solvent isopropanol, EtOAc, Hexane, EtOAc, isopropanol; with 30-60min for each steps.
If you just want to pass some samples (not large sequence) then there should nor be any problems as long as you do some king of cleaning regularly. Do not forget that pre-column will take most of your apolar compounds, anyway to some extend.
The main question to ask yourself before, is still which solvent did I used for the extraction? And why do I need to do HPLC analysis.
Article Detection of metabolite induction in fungal co-cultures on s...
If it is a routine analyses (means you inject these samples every day), I advice you to use a guard column. It can help to protect the column from the impurities. Wash it carefully after each run.
Yes if u want to analyze specific compounds in crude extract.
But you should also go for solvent partition technique using immisible solvent combinations (Hexane, chloroform, ethyl acetate and water etc.). The compounds of interest were than analyzed in different solvents fractions after removing the solvents. In this way u can analyze compounds of your interest without using column chromatograpgy.
Yes, for analytical purpose you can use. But for preparative HPLC, you need to fractionate the extract.
Crude plant extracts can be used directly in HPLC, yet selection of appropriate column of proper size and thorough pre-filtration of the extract sample is very important. However, by using crude extract, you may not achieve proper separation of the constituent compounds. Total crude extract of a plant contain a number of constituents of different phytochemical groups like steroids, alkaloids, terpenoids, phenolics, carbohydrates, etc of varying physicochemical traits and polarities.There are dedicated HPLC columns of plant different types of compounds. Hence, it is advisable to first separate different class of compounds through fractionation and/or gross separation by column chromatography and subject there fraction to HPLC separation that will give much better recovery of pure individual constituents.
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