Your solvent system is Chloroform:methanol:water (10:3:0.2). A ten minute saturation is not sufficient. Instead of a beaker, use a small TLC jar, if possible and use a filter paper wrapped on inner sides of the chamber to see the solvent vapors rising and the chamber being saturated properly. Let's see.

Are these spots of same compound? what was solvent the solvent system?

Due to thickness of TLC plate which may not be uniform.

@Santosh Yele these spots for different fractions' tubes from the same column with same mobile phase.

I am not sure if the same compound or not !

Mr Santosh's Question is Right!!! Because without complet information wee can not give Judgment for the same.

but i had some logical solution which was applicable if your all spot from same compound

1) your Compound was very sensitive for polarity,

2) in your solvent system may be you have used heighly voletile solvents and from the surface of the plates its got evaporate and both side (left and rifht ) 17 no and 23 no got same environment for separation so they give same RT value .

3) may be you had used open TLC chamber.

Please excuse me for sending suggestion

Regards

Dr vipul

@Dr. Vipul Davariya thanks for your valued answer and comments

in facts yes, for TLC mobile phase I am using chloroform/methanol/water 10/3/0.2

the chamber is a beaker covered in good way with aluminum foil.

I would like to share more information about this TLC, in fact this TLC is a result of fractions that on TLC gave only Two fragments (compounds) for the same spots so one of them was UV active and the other appeared with vanillin. I applied to another column then I got this result which is the vanillin active !

I would suggest you to prepare a fresh TLC chamber and saturate it with the solvent system properly. Make sure the TLC chamber is properly washed and dried before adding the solvent system in it. Use a filter paper to see if the chamber is saturated properly.

Is your solvent system kept in the TLC jar for a long time? Means is there a possibility that you have over-saturated the chamber. Because this is not a problem of solvent, this is a problem of saturation because vapors are rising from both the sides in an uneven manner. Every-time u perform a TLC with this solvent system, prepare a fresh chamber and activate the plate before spotting by heating properly for 30 mins at 110 C. Then compare the results.

As of now, I will advice you to do this only. Afterwards, we can think of other solutions once we are clear that this is not the problem. Otherwise this is over here only :)

Gud Luck and best regards

@Dr. Alok Nahata,

regarding to the solvent, it is fresh I have prepared it a minutes before running the TLC ! and put in the chamber not more than 10 minutes before putting the TLC plate.

I did not wash the beaker before using it, but I was sure that it is dry!

I will repeat the experiment again then share the results

many thanks :)

thank you Helmi for sharing information, actually separation method development is my hoby and Natural products compd. Separation is you know better!!!!!!!

we never sure for that separeted single spot is a single compd!!! Right?

feel free to interract me for separation techniques i will try to searve at my level

regards

Dr vipul

Your solvent system is Chloroform:methanol:water (10:3:0.2). A ten minute saturation is not sufficient. Instead of a beaker, use a small TLC jar, if possible and use a filter paper wrapped on inner sides of the chamber to see the solvent vapors rising and the chamber being saturated properly. Let's see.

Some close observations:

1. Spots No. 17-19 are moving slightly towards right i.e. towards 20 whereas

2. Spot No. 21-23 are moving slightly towards left i.e. towards 20.

Concluding migration of spots towards center of the plate and this is simply some errors while TLC Development. Solvent system is appropriate but this might be due to improper mixing of solvents while solvent system preparation OR over saturation of the chamber.

As the spots near extremities 17 and 23 are tallest as compared to adjacent spots 18 and 22, it shows HORIZONTAL UPWARD EFFECTS of the solvent adsorbed on the filter paper for tank saturation indication of OVER SATURATION.

I advice you to perform the experiment again under standard conditions. Please consider the Laboratory temp. while developing the plate.

As stated above (Dr. Nahata)....10min of chamber saturation won´t be enough due to the fact that your solvent systems includes water. Extend your chamber saturation time. Good luck

The problem is your solvent system (main culprit is water ). Repeat the expt. with other solvent mixture. Secondly you may try Two dimensional TLC chromatography for more clarity/ separation of spots

Dr. Eichmann is right- 10 minutes isn't enough saturation time with an aqueous solvent system. Like mentioned above, use some filter paper to improve and enhance the saturation in the chamber. Water is OK as a part of a solvent system for your plates as the binder is still in place.

I'm curious about the "over saturation" a couple have posted- unless the lab cools and there is condensation on the sides of the TLC chamber, the atmosphere in the TLC chamber should be in equilibrium with liquid solvent.

When you use CHCl3:H2O:MeOH mixture the maximum saturation of a TLC chamber expected is CHCl3 ( saturation has a role but not to that extent to change Rf values by 50%).

TLC plates (silicagel -G ) when used resolution of spots and Rf values depend basically on both adsorption as well partition chromatography. and also whether used plates are self coated or pre-coated ones and also the edges of plate looks uneven (which effect solvent movement )

It is clearly seen on the plates that spots developed as U shaped curve and definitely water is playing a role (the tag question has not specified the ratio of solvents used ) as depicted by solvent front line also.

Better try some other suitable system

Thanks all for your valued comments and suggestion

@Dr. Alok I have followed your suggestions and they worked with me very nice and gave me better results :) all the bands were almost a line !

thanks again

How is your solvent front moving ? Straight horizontal (plane) ? Obviously yes considering the straight grey line at the top. Try to spot a bit higher or reduce solvent volume, increase saturation time. Some people suggested 2D migration, but with that result it could be a mess finally, I would suggest do to two migrations by reversing the plate after the solvent went first at the top. (slightly dried between)

@ Hemli Alfarra. Pleased to know that my suggestions worked for you and your problem is solved. In case you encounter any problem in future, feel free to ask.

Gud Luck and best regards :)

Dear Bro. Zarei, was alikom assalam thanks for your suggestions and comments its kind of u to share with me ur experience

@Dr.Alok thanks again for ur reply again and ur help

@Zarei, Absolutely, my email is [email protected], waiting for you email :)

your TLC results looks nice, what were your TLC conditions ?

regards

Yes, Zarei

Have you try again and have same result? use the same way.. avoiding the central arch of the plate