Column chromatography

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msvnathan
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Column chromatography

Any reference book or protocol for separation of plant extract (not solvent extracted) using column chromatography.

Dr. Analytical
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Could you give a little more

Could you give a little more detail about what you are trying to do?

msvnathan
msvnathan's picture
Dr. Analytical wrote:Could

Dr. Analytical wrote:

Could you give a little more detail about what you are trying to do?

yes. Actually i am using plant extract for animal studies, ie taking plant extract and giving to animal to see the antifertility effect. i have used crude extract not soxhlet one. so the problem is when i am going for column i am getting first i am getting as usual chlorophyl and other compounds. i want to know what are all the other fraction can be collected and go for further purification.

Greg Pronger
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I guess what I am not

I guess what I am not following in the thread is how you are creating the extract itself.

Is this a aqueous extract of the plant material? A simple homogenization of the plant followed by filtering?

If so, have you by some means separated out the solid material?

I assume you've avoided solvents to forgo any concern that an observed antifertility effect is due to the plant material and not due to any residual solvents; correct?

Typically, I am most familiar with using the technique with compounds that are dissolved in an organic solvent.

Whenever I have been outside typical application use for most chromatography systems, I've turned around and approached the vendors with the products that to me seem most likely to fit my needs and see what their technical support people can do. I've been surprised by the amount of work some of the vendors are willing to do to assist a prospective customer.

Greg Pronger

Dr. Analytical
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A plant extract will probably

A plant extract will probably contain thousands of individual compounds, so identifying which one(s) are producing a desired biological effect will be a lengthy process.

I think what you want to know about is the order that compounds will elute from a column. There are some general rules that might help you get started.

If you have a "polar" stationary phase in the column (silica, alumina, Florisil, etc.) and a non-polar solvent, this is a "normal phase" system. Compounds will elute in order of increasing polarity (less polar to more polar). As you increase the polarity of the solvent, more polar compoiunds will elute.

If you have a polar solvent, and a less polar stationary phase (polymer, C18, C8 bonded phase, etc.), this is a "reversed phase" system. Molecules elute in order of decreasing polarity and increasing carbon number. That is, smaller more polar molecules come out first, followed by larger, less polar molecules.

If you can give us some more specifics, we may be able to provide a better explanation.

karthibiochem
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Better u try to contact prof

Better u try to contact prof Dr.Akbarsha from bharathidasan university trichy. They are doing these type of research, they identified anticancer drug from various plant extract so u contact that research lab they give better solution for u.

msvnathan
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Thankyou, i have some more

Thankyou, i have some more details, but i will explain detaily then i will try according to your suggestion.

Tirumal Rao
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Hellow,

Hellow,
High speed countercurrent chromatography (HSCCC) was used in a pre-fractionation pilot study to improve the quality of crude plant samples for primary screening in drug discovery efforts. The methanol extracts of sixty-four plant samples were (i) defatted, (ii) treated with poly-N-vinylpyrrolidone (PVP) for polyphenolic removal, and (iii) fractionated with a multilayer coil planet centrifuge.

The ternary solvent system CH2Cl2:MeOH:H2O (5:6:4, v/v/v) was used based upon elution of known plant natural product standards with ranging polarities. Elution was carried out until a partition coefficient (K) of 1, followed by column contents extrusion to exploit stationary phase separation and to increase the polarity range of compounds, fractionated.

Fractionation was found to be consistent for all separated extracts with respect to sample recovery, stationary phase fraction (Sf), and weight distribution by fraction number. Biological evaluation was conducted in 20 mechanism-based, in-vitro assays with an evaluation of biodata trends. Bioassay interfering agents such as polyphenolics and fatty acids were chromatographically localized and rapidly identified.
Plant membrane lipids were separated by multiple solid-phase extraction (SPE) in a single run. Elution was performed continuously through the modulated stationary phase employing only non-aqueous solvent systems. At the different stages of the glycerolipid separation the SPE manifold combined aminopropyl, aminopropyl/silica gel and silica gel/aminopropyl weak anion exchanger columns. The glycerolipid extract of pigment-containing plant tissues was cleared from the pigments onto the aminopropyl column. The aminopropyl column with the glycerolipid extract was then connected to a silica gel column from which monogalactosyldiacylglycerol, phosphatidylethanolamine, phosphatidylglycerol and digalactosyldiacylglycerol were eluted as individual fractions. The elution was performed under polarity, pH and temperature gradient conditions. To continue the separation, the aminopropyl column was discarded and the silica gel column containing the remaining glycerolipid extract was connected to an aminopropyl anion exchanger column. Individual fractions of sulfoquinovosyldiacylglycerol, phosphatidylcholine and phosphatidylinositol were now eluted. The separation process was supported by ammonium counter ions and by the polarity gradient of the elution systems used. The membrane lipids were isolated from pigment-containing (rice and maize leaves and rice leafy stems) and pigment-free (rice roots) tissues. The repeatability for a standard glycerolipid mixture was 2–6% (n=7), and for rice leaf lipid extracts, 3–7% (n=5). Glycerolipid recovery was 87–95%.
Hope this is help to u.
Thanks,
Tirumal
India

msvnathan
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Thankyou and it is usefull

Thankyou and it is usefull suggestion, i will try it.

ycjhz
ycjhz's picture
Preparative Chromatography

Preparative Chromatography Techniques by Kurt Hostettmann, Springer-Verlag,1998
I think this references book is the best effective for you and your research on pre-preparation of plant extrations