Author Topic: TLC Time  (Read 19871 times)

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  • Guest
« Reply #20 on: July 15, 2002, 06:07:00 PM »
I'll dilute and check it,  and also run a plate so the rf's can be seen... this was the initial testing for the chromophores...

 extremely small amounts of material with this intensity...

Infinite Radiant Light - THKRA


  • Guest
second that
« Reply #21 on: July 15, 2002, 07:57:00 PM »
The LSD looks right and I'm guessing its 1 hit of blotter.

So try to scale the others accordingly
Good work by the way!  :)

Those who give up essential liberties for temporary safety deserve neither liberty nor safety


  • Guest
re: colors
« Reply #22 on: July 15, 2002, 08:37:00 PM »
the acid is 50 micrograms... I can't scale the others as such since the tests are so sensitive... I probably put 1 milligram of the dpt and 5meodmt on each dish and they are really intense... the tests work down to 25-50 nanograms, so I'll try different dilutions and see what gives a clearer color indication... probably should make up some standards and give the formulation... any suggestions on standards folks?

Infinite Radiant Light - THKRA


  • Guest
column and plate preparation
« Reply #23 on: August 22, 2002, 06:44:00 AM »
This web site details rolling your own tlc plates, by using the same materials you will use in your column. It gives % of binder and preparation methods.

Making TLC Plates from Bulk TLC Silica Gels
Many TLC users prefer to use pre-coated TLC plates, but others because of their special needs, or because they need special additives or a special thickness, coat their own TLC plates. This paper is a quick review of what is needed for the process.

Making TLC Plates from Bulk TLC Silica Gels

Many TLC users prefer to use pre-coated TLC plates, but others because of their special needs, or because they need special additives or a special thickness, coat their own TLC plates. This paper is a quick review of what is needed for the process.

Infinite Radiant Light - THKRA


  • Guest
Ghetto spray hood
« Reply #24 on: November 04, 2002, 04:36:00 AM »

  In the Journal of Chemical Education, oct 96, #73, pg 979
Josef G. Krause writes:

"A recent suggestion to visualize spots on thin layer plates by applying phosphomolybdic acid reagent with a cotton ball (1) may not be suitable for all visualization reagents especially concentrated sulfuric acid. If spraying a reagent with a power pac type device is preferred for this or some other reason, we suggest spraying the reagent inside of a twist tie-sealed, clear-plastic kitchen garbage bag. In this way, spray reagents are completely contained obviating the need for spraying the plates in a fume hood, and cleanup is simple. "

  So all you have to do is put your plates and your sprayer inside a clear trash bag, spray them, pull out your gear and toss the bag... this avoids the need for a fume hood and allow you to get a good mist on the plates.

Infinite Radiant Light - THKRA


  • Guest
« Reply #25 on: November 05, 2002, 01:08:00 AM »
In my lab we just have the development regent (DNPH,PMo,Sulfuric ect.) in a glass(mason actual) , and then just dip the plate into the glass. No nasty sprayin and fine result's.


  • Guest
Re: TLC for your reactions and synthesis
« Reply #26 on: November 11, 2002, 03:18:00 AM »
I thought that TLC might be more useful if applied in the synthesis process. One would get a known and spot against that ot verify one has the right stuff then as the procedur goes on one can check if the starting material is all used up and so on and always keep a sample of the  material from each step so next time one can spot agains a known  step material. By soing this one can check one's synthesis as one progresses through a reaction.  Depending on the solvent used to wet the plates, I've always used a small Iodine crystals to develop my slide type TLC plates to visualize my progress, and if a step is not pure on can see various spots on the run of the slide.  No spray   nothing fancy just two jars where your slides can fit standing. I also suggest  keeping notes and rf distance for future refrence because slides fade after a while .  I prefer the NH3OH-chloroform  -methanol solvent for my particular use in the synthesis of Methamphetamine  I look for the chlorephedrine and starting material marks in my finish product  spot.............this method is easy and although its not so colorful its effective  and keeps me on track.


  • Guest
« Reply #27 on: November 11, 2002, 09:39:00 AM »
Hmm... when you roll your own on sg gf, sometimes it tends to fall off on the dip...+ the visualization reagent shouldn't dissolve the target material...

Infinite Radiant Light - THKRA


  • Guest
Journal of Chromatography
« Reply #28 on: June 01, 2003, 07:45:00 PM »
Here is the Journal of Chromatography article mentioned earlier in the thread (Thanks ClearLight!):

The Van Urk-Salkowski Reagent - A Sensitive And Specific Chromogenic Reagent For Silica Gel Thin-Layer Chromatographic Detection And Identification Of Indole Derivatives

Axel Ehmann

Journal of Chromatography 132, 267-276 (1977)



  • Guest
TLC Visualization Reagents (TLC Stains)
« Reply #29 on: August 16, 2003, 09:51:00 PM »
Stains for Developing TLC Plates

Once a TLC has been developed, it is frequently necessary to aid in the visualization of the components of a reaction mixture. This is true primarily because most organic compounds are colorless. Frequently, the organic compounds of interest contain a chromophore which may be visualized by employing either a short or a long wave UV lamp. These lamps may be found as part of a standard organic chemistry research or teaching lab. Typical examples of functional groups which may be visualized through this method are: aromatic groups, a,b-unsat- urated carbonyl groups, and any organic compound containing other extensively p-conjugated systems. While exposing these TLC plates to UV light, you will notice that the silica gel will fluoresce while any organic molecule which absorbs UV light will appear as a dark blue spot. Circling these spots gently with a dull pencil will permit an initial method for visualization. Fortunately, there are a number of permanent or semi-permanent methods for visualization which will not only allow one to see these compounds but also provide a method for determining what functional groups are contained within the molecule. This method is referred to as staining the TLC plate, and experience will allow you to determine what functional groups will appear as what color upon visualization. Here are the most commonly employed stains, along with a typical preparation:


The staining of a TLC plate with iodine vapor is among the oldest methods for the visualization of organic compounds. It is based upon the observation that iodine has a high affinity for both unsaturated and aromatic compounds.

An chamber may be assembled as follows: To 100 mL wide mouth jar (with cap) is added a piece of filter paper and few crystals of iodine. Iodine has a high vapor pressure for a solid and the chamber will rapidly become saturated with iodine vapor. Insert your TLC plate and allow it to remain within the chamber until it develops a light brown color over the entire plate. Commonly, if your compound has an affinity for iodine, it will appear as a dark brown spot on a lighter brown background. Carefully remove the TLC plate at this point and gently circle the spots with a dull pencil. The iodine will not remain on the TLC plate for long periods of time so circling these spots is necessary if one wishes to refer to these TLC's at a later date.

Potassium Permanganate

This particular stain is excellent for functional groups which are sensitive to oxidation. Alkenes and alkynes will appear readily on a TLC plate following immersion into the stain and will appear as a bright yellow spot on a bright purple background. Alcohols, amines, sulfides, mercaptans and other oxidizable functional groups may also be visualized, however it will be necessary to gently heat the TLC plate following immersion into the stain. These spots will appear as either yellow or light brown on a light purple or pink background. Again it would be advantageous to circle such spots following visualization as eventually the TLC will take on a light brown color upon standing for prolonged periods of time.

Dissolve 1 g of KMnO4 and 2 g Na2CO3 into 100 mL of distilled water. Ideally, these stains may be stored in 100 mL wide mouth jars. A typical lifetime for this stain is approximately 3 months.

Bromocresol Green Stain

This particular stain is excellent for functional groups whose pKa is approximately 5.0 and lower. Thus, this stain provides an excellent means of selectively visualizing carboxylic acids. These will appear as bright yellow spots on either a dark or light blue background and typically, it is not necessary to heat the TLC plate following immersion. This TLC visualization method has a fairly long lifetime (usually weeks) thus, it is not often necessary to circle such spots following activation by staining.

To 100 ml of absolute ethanol is added 0.04 g of bromocresol green. Then a 0.1 M solution of aqueous NaOH is added dropwise until a blue color just appears in solution (the solution should be colorless prior to addition). Ideally, these stains may be stored in 100 mL wide mouth jars. The lifetime of such a solution typically depends upon solvent evaporation. Thus, it would be advantageous to tightly seal such jars in-between uses.

Cerium Molybdate Stain

This stain is a highly sensitive, multipurpose (multifunctional group stain). One word of caution, very minor constituents may appear as significant impurities by employing this stain. To ensure accurate results when employing this stain, it is necessary to heat the treated TLC plate vigorously. Thus, this may not be a stain to employ if your sample is somewhat volatile. The TLC plate itself will appear as either light blue or light green upon treatment, while the color of the spots may vary (although they usually appear as a dark blue spot). Typically, functional groups will not be distinguishable based upon the color of their spots; however, it would be worth while to make a list of potential colors of various functional groups as you experience variations in colors. This may permit future correlations which may prove beneficial when performing similar chemistry on related substrates.

To 235 mL of distilled water was added 12 g of ammonium Molybdate, 0.5 g of ceric ammonium molybdate, and 15 mL of concentrated sulfuric acid. Storage is possible in a 250 mL wide mouth jar. This stain has a long shelf-life so long as solvent evaporation is limited. It may also prove worth while to surround the jar with aluminum foil as the stain may be somewhat photo-sensitive and exposure to direct light may shorten the shelf-life of this reagent. It is worth while to also mention that it would be beneficial to circle the observed spots with a dull pencil following heating as this stain will eventually fade on the TLC plate after a few days.

p-Anisaldehyde Stain

This stain is an excellent multipurpose visualization method for examining TLC plates. It is sensitive to most functional groups, especially those which are strongly and weakly nucleophilic. It tends to be insensitive to alkenes, alkynes, and aromatic compounds unless other functional groups are present in the molecules which are being analyzed. It tends to stain the TLC plate itself, upon mild heating, to a light pink color, while other functional groups tend to vary with respect to coloration. It is recommended that a record is kept of which functional group stains which color for future reference, although these types of comparisons may be misleading when attempting to ascertain which functional groups are present in a molecule (especially in complex molecules). The shelf-life of this stain tends to be quite long except when exposed to direct light or solvent is allowed to evaporate. It is recommended that the stain be stored in a 100 mL wide mouth jar wrapped with aluminum foil to ensure a long life time.

To 135 mL of absolute ethanol was added 5 mL of concentrated sulfuric acid, 1.5 mL of glacial acetic acid and 3.7 mL of p-anisaldehyde. The solution is then stirred vigorously to ensure homogeneity. The resulting staining solution is ideally stored in a 100 mL wide mouth jar covered with aluminum foil.

Phosphomolybdic Acid (PMA) Stain

Phosphomolybdic acid stain is a good "universal" stain which is fairly sensitive to low concentrated solutions. It will stain most functional groups, however it does not distinguish between different functional groups based upon the coloration of the spots on the TLC plate. Most often, TLC's treated with this stain will appear as a light green color, while compounds of interest will appear as much darker green spots. It is necessary to heat TLC plates treated with this solution in order to activate the stain for visualization. The shelf life of these solutions are typically quite long, provided solvent evaporation is kept to a minimum.

Dissolve 10 g of Phosphomolybdic acid in 100 mL of absolute ethanol.

Occasionally, you may find it necessary to develop or investigate other staining techniques; the following references may be helpful:

* Handbook of Thin-Layer Chromatography J. Sherman and B. Fried, Eds., Marcel Dekker, New York, NY, 1991.
* Thin-Layer Chromatography 2nd ed. E. Stahl, Springer-Verlag, New York, NY, 1969.
* Thin-Layer Chromatography Reagents and Detection Methods, Vol. 1a: Physical and Chemical Detection Methods: Fundamentals, Reagents I H. H. Jork, W. Funk, W. Fischer, and H. Wimmer, VHC, Weinheim, Germany, 1990.
* Thin-Layer Chromatography: Techniques of Chemistry, Vol. XIV, 2nd ed. J. G. Kirchner and E. S. Perry, Eds., John Wiley and Sons, 1978.



  • Guest
What- no ninhydrin?
« Reply #30 on: August 17, 2003, 05:05:00 AM »
I'm surprised ninhydrin was not mentioned- it is a good stain for compounds containing amines or Boc-protecting groups, among others. Simply dissolve "a couple spatula-tip's worth" (what, maybe 10g? I always just eyeball it) in 95% ethanol. To visualize TLC plates, first circle UV-visible spots, then dip the plate in the ninhydrin solution. Lightly blot dry and heat with a heat-gun (hardcore hair dryer) until the plate takes on a brownish color/spots form.

Not sure about the lifetime of ninhydrin solutions- they always last long enough in lab to be used up, which may take a couple of months depending.



  • Guest
Carcinogenity or Cost?
« Reply #31 on: August 17, 2003, 05:19:00 AM »
It might be a cost issue, or because of the Carcinogenity of Ninhydrin.


  • Guest
Requested References
« Reply #32 on: August 20, 2003, 01:35:00 AM »
From Post 453943 (not existing)  ;)

Containment of Spray Reagents in TLC Spot Detection
Krause, Josef G.
J. Chem. Educ. 73, 979 (1996)

A recent suggestion to visualize spots on thin layer plates by applying phosphomolybdic acid reagent with a cotton ball may not be suitable for all visualization reagents especially concentrated sulfuric acid. If spray­ing a reagent with a power pac type device is preferred for this or some other reason, we suggest spraying the reagent inside of a twist tie-sealed, clear-plastic kitchen garbage bag. In this way, spray reagents are completely contained obviating the need for spraying the plates in a fume hood, and cleanup is simple.

Staining TLC Plate Using a Cotton Ball
Albert W. M. Lee and Zhong Zengpei
J. Chem. Educ. 73, 358 (1996)

Thin layer chromatography (TLC) is a simple but yet a very powerful analytical tool. It is widely used in unknown identification. Synthetic chemists routinely use TLC to monitor reactions and to check the collected fractions after column chromatography. After the TLC plate is developed, spot detection can be carried out under a ultraviolet lamp if the compounds are UV active, otherwise, some staining reagents have to be applied. Among many staining reagents, phosphomolybic acid (PMA) is widely used by organic chemists because most organic compounds can be visualized with PMA followed by heating. There are two common ways to apply the staining reagents, dipping and spraying. Dipping the TLC plate into the staining reagent may contaminate the reagent solution and may cause some diffusion of the spots. Spraying the TLC plate has to be carried in a fume hood and the mist generated can be very annoying.

We found that using a cotton ball to apply the staining reagent is a very convenient alternative. A cotton ball held by a pair of tweezers is dipped into the PMA ethanol solution. After the excess solution is squeezed out, the cotton ball is lightly wiped onto the TLC plate, followed by heating with a heat gun to visualize the spots.

Removing Silicone Greases from Round Bottom Flasks
Thomas J. Reilly
J. Chem. Educ. 73, 381 (1996)

If silicone greases are used on the standard taper joints of round bottom flasks they will eventually get inside the flask, contaminating the contents and introducing spurious peaks into the IR. and NM. spectra of samples. Removal of these deposits with solvents or conventional cleaning solutions is difficult, but the following method is fast and efficient: The flask to be cleaned is supported in a rubber ring and filled to the bottom of the joint surface with 5% aqueous hydrofluoric acid.

Caution: Hydrofluoric acid causes severe burns that may not become evident for several hours. This procedure should only be performed by an individual who is fully aware of the hazards involved and the safety precautions required. Hydrofluoric acid should never be handled by undergraduates.

After 20 minutes, the grease will separate from the glass surface in a thin film and can be removed from the hy­droflouric acid by gravity filtration. The procedure can be repeated if necessary and will not cause visible etching unless the contact time exceeds 200 minutes. The hydrofluoric acid can be reused as long as it remains strongly acidic to pH paper.



  • Guest
Dip Reagents for Visualization in TLC
« Reply #33 on: August 23, 2003, 12:39:00 AM »
Dip Reagents for Visualization in TLC

J. Chem. Educ. 73, A4-A6 (1996)


Thin-layer chromatography is used widely in the undergraduate organic laboratory (1-4). The technique often is illustrated with an early experiment involving the analysis of a mixture of colored substances such as those extracted from paprika or spinach. For colorless compounds, a step to make spots visible after development of the plate is needed.

Most Common Recommendations

* Commercially available plates can be used that incorporate a fluorescent dye in the adsorbent layer and make many substances visible when viewed with ultraviolet light.
* A TLC plate can be inserted into a jar containing iodine crystals to produce brown spots due to absorption of iodine from the vapor phase into the organic material.

Both methods, however, involve disadvantages. The UV method works only for UV active substances, and the iodine method is somewhat more general, but the spots produced tend to be fuzzy and indefinite, precluding fine resolution. We have found that these problems may be ameliorated largely by including "spray reagents" among the visualization methods available to students.


* 2,4-Dinitrophenylhydrazine (used to detect aldehydes and ketones)
* Potassium permanganate (detection of alcohols, alkenes, and other oxidizable compounds)
* Phosphomolybdic acid (non-specialized reagent)

The spots resulting from the use of these solutions usually are well-defined and often appear in characteristic colors. These solutions are applied to the TLC plate with any of several types of spraying devices available from major chemical companies. This step, however, has several objectionable features that have discouraged the use of these reagents in most student laboratories.

Objectionable Features

* The process is wasteful. Only a small fraction of the solution ever reaches the plate.
* Unless scrupulously maintained, sprayers tend to clog and splatter, resulting in an uneven coating.
* Most serious is the fact that most of the reagent is released into the atmosphere to remain as a noxious contaminant.

Employing a Dipping Technique Avoids Disadvantages

We have avoided these disadvantages by employing a dipping technique with the reagents. The solutions are stored in plastic centrifuge tubes of proper size (a few centimeters longer and a few millimeters wider than the TLC plate). After development, the plate is held at the solvent front end with forceps, submerged in reagent up to the solvent front, and immediately removed. Contact time with the solution should be kept to a minimum. Spots usually begin to emerge almost immediately but become more definite over a period of up to one hour. In some cases (e.g., after the phosphomolybdic acid dip) spot development requires the application of mild heat such as hot air from a hair dryer. A large number of general and specialized visualization reagents are listed in an easily available chemical-biochemical catalog (5). Use of such reagents with the dip technique should lead to optimal results with a minimum of cost, mess, wastage, and environmental contamination.

Materials and Procedures

The TLC plates are 2.5x7.5-cm plastic sheets with a silica gel coating containing a fluorescent dye (J. T. Baker catalog number IB2-F). The dip solution containers are 50 mL polypropylene centrifuge tubes, 2.8 x 11.5 cm, with screw cap (Fisher Scientific Company catalog number 05-538-55). The phosphomolybdic acid reagent is purchased most economically as a 20% solution in ethanol (Aldrich Chemical Company, catalog number 31,927-9). The permanganate dip was prepared as an aqueous solution, 1% in KMnO4, and 2% in K2CO3. Brady's reagent (6) was employed for the 2,4-dinitrophenyhydrazine solution.
Literature Cited

1. Fessenden, R. A.; A, Fessenden, J. S., Organic Laboratory Techniques, Brooks/Cole: Pacific Grove, CA, 1993, p 188.
2. Mayo, D.W,; Pike, R. M.; Trumper, P. K. Microscale Organic Laboratory; 3rd Ed. John Wiley New York, 1994, p 103.
3. Wilcox, C. F. Jr,; Wilcox, M. F., Experimental Organic Chemistry, Prentice-Hall: Englewood Cliffs, NY, 1995, p 131.
4. Zanger, M.; McKee, J. R. Small Scale Syntheses: A Laboratory Textbook of Organic Chemistry; William C. Brown: Boston, 1995, p 52.
5. Sigma Chemical Company Catalog, 1995; p 941.
6. Shriner, R. L.; Fuson, R. C., Curtin, D. Y., Morrill, T. C., The Systematic Identification of Organic Compounds, 6th. Ed.; John Wiley; New York, 1980, p 162.


  • Guest
TLC Variations
« Reply #34 on: August 23, 2003, 03:10:00 PM »
Thin-Layer Chromatography - Variations on a Theme
J. Anwar, S. A. Nagra, M. Nagi

J. Chem. Educ. 73(10), 977 (1996)

( (Retrieved by Lugh)


  • Guest
TLC Developing Chamber 4 Flash Columns
« Reply #35 on: November 27, 2003, 09:57:00 AM »
TLC Developing Chamber for Screening Column Chromatography Fractions
Samuel G. Levine
J. Chem. Ed. 73(1), 77 (1996)

Column chromatography, in all of its varied forms, is the mainstay technique for preparative separations in the organic chemical laboratory. The subject was possibly first introduced to undergraduates by Fieser's 1955 laboratory manual that includes complete instructions for preparing and running a column as well as two experiments that result in the separation of colorless substances. By contrast, most of the current texts include only less ambitious column chromatography experiments involving the much easier task of separating colored substances such as dye mixtures, plant pigments, or ferrocene derivatives. Barring the use of very expensive automatic monitoring equipment, the successful separation of colorless substances requires some means of screening the eluate fractions so that those of like composition may be combined. This process is customarily accomplished by thin layer chromatography (TLC). We describe here a novel TLC development chamber that greatly enhances the efficiency of this task.

The chamber (see figure), designed for use with 25x75 mm plastic TLC plates, is constructed from a 100-mm length of 25-mm OD Pyrex tubing of standard wall thickness. After both ends are sealed, a narrow 80-mm long slit is cut* along the length of the tube, equidistant from the ends. The chamber may be mounted conveniently, slit facing up, on a short length of 2x4 lumber with the help of two small lumps of "Hold-It" or other putty-like material. Ten fractions may be spotted 7 mm apart along one edge of the plate. The plate is then lowered through the slit into the development chamber that has been charged previously with 2 mL of eluant. (Care must be taken to maintain a horizontal orientation of the plate while lowering it into position.) Note that the upper edge of the plate extends a short distance outside the chamber. After elution is complete (45 sec using EtOAc/hexanes 1:4 as eluant) the plate is removed, dried, and subjected to spot visualization by any appropriate means. Viewing the plate will then disclose a pattern of fraction composition as a function of fraction number. If greater resolution of low Rf spots is desired, this often can be achieved by extending the elution time, e.g., to two minutes.

The methodology described here is advantageous for several reasons.
• It is very rapid.
• Highly efficient use is made of the least expensive form of commercial TLC plates.
• The perfectly straight base of the chamber results in very even flow of solvent.
• Very little solvent is required.

TLC developing chamber.

* Cutting was performed with a 3/32-in. mounted diamond-point cylinder from Starlite Industries, P. O. Box 990 Rosemont, PA 19010-0911; Cat. #117045, $15.60.


  • Guest
Determining TLC solvent systems
« Reply #36 on: November 29, 2003, 01:40:00 AM »
I did not find any info regarding the determination of a solvent system for a given substance. How is this determined??

Trial and error. You test different solvent mixtures until you find one which makes most of your spots end up between Rf 0.2 and 0.8 (very approximate - just see this as your highest goal, often spots either have a strong tendency to run away (non-polar) or stay beind (polar) on the TLC slide.

First try with a medium polarity solvent mixture (like petroleum ether/ethyl acetate in a 1:1 ratio). To make the spots run higher, use a more polar solvent mixture (like petroleum ether/ethyl acetate in a 1:5, or even 1:10 ratio). To make the spots run lower, use a less polar solvent mixture (like petroleum ether/ethyl acetate in a 5:1, or even 10:1 ratio). If 100% ethyl acetate isn't polar enough, you can begin mixing in some methanol or isopropanol (10% or so).

If you run TLC on amines, and if they tend to streak (form long oval spots) it usually helps to add 1% triethylamine to the solvent system. To find suitable solvents to use, see the polarity index here (and you'll see if they are miscible too, which naturally is a prerequisite):

Try to only mix solvents which have roughly the same evaporation rate or you will get inconsistent results, as one component will evaporate faster, changing the composition of the mixture over time.


  • Guest
Way better than triethylamine in the eluent is
« Reply #37 on: November 30, 2003, 06:30:00 PM »
Way better than triethylamine in the eluent is a vial with conc. aqueous ammonia in the chamber. This will not cause artifacts caused by the 'TEA-front' which might result in spot-doubling or smearing.


  • Guest
The latter.
« Reply #38 on: November 30, 2003, 09:56:00 PM »
The latter.


  • Guest
Phenethylamine TLC Visualization Reagent
« Reply #39 on: March 15, 2004, 05:17:00 PM »
Cactus Alkaloids XXVII.
Use of Fluorescamine as a Thin-Layer Chromatographic Visualization Reagent for Alkaloids

R.L. Ranieri & J.L. McLaughlin

Journal of Chromatography 111, 234-237 (1975)