Question on Hard Layer TLC Binders

[ClearLight]

TLC plates are normally prepared w/ a polyvinyl alcohol (although centipoise density is not mentioned). For rolling your own hard layer tlc plates (I'm looking for otc.. $132/100 5x7cm's is a bit rich), does anyone know the method or have a link to it?

  Really would like something that indicated % of PV Alch. and heating parameters etc.. enough to duplicate in OTC application.

[Lilienthal]

Use plaster from the hardware store (gypsum) as a binder for preparative layers. I think it doesn't make much sense to prepare your own analytical sheets.

[hest]

Anyeone who know's wath the fluricent material in TLC plates is ??

[lugh]

From Experimental Organic Chemistry

Thin-layer chromatography (TLC) is another form of solid-liquid adsorption chromatography and involves the same fundamental principles as column chro­matography. The properties of the adsorbents and the relative eluting abilities of the solvents given for column chromatography  also apply to TLC. However, unlike column chromatography where the mobile phase moves down the column, the mobile phase in TLC ascends the thin layer of adsorbent. Another significant difference is that a TLC experiment may be performed more rapidly and requires much less sample, sometimes as little as 10-9 g. Thin-layer chromatography is an excellent analytical tool and is frequently used to deter­mine the optimal combinations of solvent and adsorbent for preparative column chromatographic separations as well as to monitor the progress of a separation by column chromatography. Note that TLC is limited to relatively nonvolatile compounds such as solids and liquids whose boiling points are above 150 °C at atmospheric pressure.

For TLC, the adsorbent, usually alumina or silica gel, is mixed with a small quantity of a binder, such as starch or calcium sulfate, and spread as a layer approximately 250 µ thick on either a glass or plastic plate. The binder is neces­sary for proper adhesion of the thin layer of adsorbent to the plate. Thin-layer chromatography plates should be dried in an oven for an hour or more at 110 °C prior to use to remove any adsorbed moisture. The presence of water molecules on the surface of the adsorbent decreases its activity and effectiveness in binding and separating the components of the mixture.

In a typical TLC analysis, a small drop of a solution of the mixture to be analyzed is carefully applied or spotted from a capillary to a small rectangular chromatographic plate near the edge of the narrow end. The plate is placed with spotted end down in a closed jar, called a devel­oping chamber, containing a suitable eluting solvent, which may be either a pure solvent or a mixture of two or more solvents; the level of the eluent should be just below that of the spot. As the solvent moves up the plate, the components of the mixture are carried along at different rates to pro­duce a series of spots on the plate. When the solvent front has advanced nearly to the top of the TLC plate, the development of the chromatogram is complete, and the plate is withdrawn from the developing chamber.

The components of the mixture may be detected in a variety of ways. If the compounds being separated are colored, visual detection of the spots is easy.

Many organic compounds are colorless, however, and a variety of methods have been developed to detect their presence on the plate:

1. Compounds that fluoresce may be located by placing the plate under an ultraviolet light. Since the spots disappear when the light is removed, it is necessary to circle the spots with a pencil in order to have a permanent record of the chromatogram. There are also commercially available plates that contain a fluorescent material as part of their coating; compounds that do not fluoresce but do absorb ultraviolet light then appear as dark spots under ultraviolet light.

2. The chromatographic plate may be sprayed with a variety of reagents such as sulfuric acid, potassium permanganate, phosphomolybdic acid, and nin­hydrin; these reagents will react with the individual components to produce colored or dark spots.

3. The chromatographic plate may be exposed to iodine vapor by placing it in a closed chamber containing several crystals of iodine. As the iodine forms complexes with the various organic compounds, the spots become brown. Since the process is reversible and the spots fade, it is wise to circle the spots with a pencil in order to have a permanent record of the chro­matogram.

Once the separation of the components of the mixture is complete and the individual spots have been detected, the retention factor (Rf) of each com­pound may be calculated. The Rf value for a compound is a physical constant for a given set of chromato­paphic conditions, and consequently the adsorbent and the eluting solvent should be recorded together with the experimentally determined Rf values.

There are many important applications of TLC in modern organic chem­istry. For example, TLC is commonly used to identify components of an unknown mixture by running chromatograms of the unknown sample side by side with known standards. Multiple aliquots of samples collected from chro­matographic columns may be analyzed by TLC to follow the chro­matographic separation. Alternatively, it is possible to follow the course and progress of a reaction by TLC by monitoring the disappearance of starting mate­rial or the appearance of product. Samples are simply withdrawn from a reaction mixture and subjected to TLC analysis. Thin-layer chromatography may also be used for preparative purposes. Specially prepared TLC plates with thicker coats of adsorbent can accommodate samples as large as 5-10 mg. In such cases. recovery of the components can be achieved by scraping the individual spots from the plate and extracting them from the adsorbent with an appropriate solvent. It may be necessary to repeat the process  number of times in order to get an amount of compound sufficient for spectral analysis.

[lugh]

From Organic Laboratory Techniques

The two adsorbent materials most often used for tlc are alumina G (alumi­num oxide) and silica gel G (silicic acid). The G designation stands for gypsum (calcium sulfate). Calcined gypsum, CaSO4 •2H2O, is better known as plaster of Paris. When exposed to water or moisture, gypsum sets in a rigid mass, CaSO4.2H20, which binds the adsorbent together and to the glass plates used as a backing support. In the adsorbents used for tic, about 10-13% by weight of gypsum is added as a binder. The adsorbent materials are otherwise like those used in column chromatography; the adsorbents used in column chromatogra­phy have a larger particle size, however. The material for thin-layer work is a fine powder. The small particle size, along with the added gypsum, makes it impossible to use silica gel G or alumina G for column work. In a column, these adsorbents generally set so rigidly that solvent virtually stops flowing through the column.

For qualitative work such as identifying the number of components in a mixture or trying to establish that two compounds are identical, small tlc plates made from microscope slides are especially convenient. Coated microscope slides are easily made by dipping the slides into a container holding a slurry of the adsorbent material. Although numerous solvents can be used to prepare a slurry, methylene chloride is probably the most convenient solvent. It has the two advantages of low boiling point (40 °C) and inability to cause the adsorbent to set or form lumps. The low boiling point means that it is not necessary to dry the coated slides in an oven. Its inability to cause the gypsum binder to set means that slurries made with it are stable for several days. It has the disadvan­tage that the layer of adsorbent formed is fragile and must be treated carefully. For this reason, some persons prefer to add a small amount of methanol to the methylene chloride to enable the gypsum to set more firmly. The methanol solvates the calcium sulfate much as water does. More durable plates can be made by dipping plates into a slurry prepared from water. These plates must be oven-dried before use. Also, a slurry prepared from water must be used soon after its preparation. If it is not, it will begin to set and to form lumps. Thus, an aqueous slurry must be prepared immediately before use; it cannot be used after it has stood for any length of time. For microscope slides, a slurry of silica gel G in methylene chloride is not only convenient but also adequate for most purposes.
The slurry is most conveniently prepared in a 4-oz wide-mouthed screw­cap jar. About 3 mL of methylene chloride is required for each gram of silica gel G. For a smooth slurry without lumps, the silica gel should be added to the solvent while the mixture is being either stirred or swirled. Adding solvent to the adsorbent usually causes lumps to form in the mixture. When the addition is complete, the cap should be placed on the jar tightly and the jar shaken vigorously to ensure thorough mixing. The slurry may be stored, in the tightly
capped jar, until it is to be used. More methylene chloride may have to be added to replace evaporation losses.
If new microscope slides are available, they can be used without any spe­cial treatment. However, it is more economical to reuse or recycle used micro­scope slides. The slides should be washed with soap and water, rinsed with water, and then rinsed with 50% aqueous methanol. The plates should be al­lowed to dry thoroughly on paper towels. They should be handled by the edge because fingerprints on the plate surface will make it difficult for the adsorbent to bind to the glass.
The slides are coated with adsorbent by dipping them into the container of slurry. Two slides can be coated simultaneously by sandwiching them together before dipping them in the slurry. The slurry should be shaken vigorously just before dipping the slides. Since the slurry settles on standing, it should be mixed in this way before each set of slides is dipped. The depth of the slurry in the jar
should be about 3 in., and the plates should be dipped into the slurry until only about 0.25 in. at the top remains uncoated. The dipping operation should be done smoothly. The plates may be held at the top, where they will not be coated. They are dipped into the slurry and withdrawn with a slow and steady motion. The dipping operation takes about 2 seconds. Some practice may be required to get the correct timing. After dipping, the cap should be replaced on the jar, and the plates should be held for a minute until most of the solvent has evaporated. The plates may then be separated and placed on paper towels to complete the drying.
The plates should have an even coating; there should be no streaks and no thin spots where glass shows through the adsorbent. The plates should not have a thick and lumpy coating. Two conditions cause thin and streaked plates. First, the slurry may not have been thoroughly mixed before the dipping operation; the adsorbent might then have settled to the bottom of the jar, and the thin slurry at the top would not have coated the slides properly. Second, the slurry simply may not have been thick enough; more silica gel G must then be added to the slurry until the consistency is proper. If the slurry is too thick, the coating on the plates will be thick, uneven, and lumpy. To correct this, the slurry should be diluted with enough solvent to achieve the proper consistency.
Plates with an unsatisfactory coating may be wiped clean with a paper towel and redipped. Care must be taken to handle the plates only from the top or by the sides.
For separations involving large amounts of material, or for difficult separa­tions, it may be necessary to use larger thin-layer plates. Plates with dimensions up to 20-25 sq cm are common. With larger plates, it is desirable to have a more durable coating, and a water slurry of the adsorbent should be used to prepare them. If silica gel is used, the slurry should be made up in the ratio about 1 g of silica gel G to each 2 mL of water. The glass plate used for the thin-layer plate should be washed, dried, and placed on a sheet of newspaper. Along two edges of the plate are placed two strips of masking tape. More than one layer of masking tape is used if a thicker coating is desired on the plate. A slurry is prepared, shaken well, and poured along one of the untaped edges of the plate. A heavy piece of glass rod, long enough to span the taped edges, is used to level and spread the slurry over the plate. While the rod is resting on the tape, it is pushed along the plate from the end at which the slurry was poured toward the opposite end of the plate. After the slurry is spread, the masking-tape strips are removed, and the plates are dried in a 110 °C oven for about 1 hour. Plates of 20-25 sq cm are easily prepared by this method. Larger plates present more difficulties.

[lugh]

If the compounds separated by tlc are colored, it is a fortunate result, because the separation can be followed visually. More often than not, however, the compounds are colorless. Then the separated materials must be made visi­ble by some reagent or some method that makes the separated compounds visible. Reagents that give rise to colored spots are called visualization reagents.
Methods of viewing that make the spots apparent are visualization methods.
The visualization reagent used most often is iodine. Iodine reacts with many organic materials to form complexes that are either brown or yellow. In this visualization method, the developed and dried tlc plate is placed in a 4-oz wide-mouthed screw-cap jar along with a few crystals of iodine. The jar is capped and gently warmed on a steam bath at low heat. The jar fills with iodine vapors, and the spots begin to appear. When the spots are sufficiently intense, the plate is removed from the jar, and the spots are outlined with a pencil. The spots are not permanent. Their appearance results from the formation of com­plexes the iodine makes with the organic substances. As the iodine sublimes off the plate, the spots fade. Hence, they should be marked immediately. Nearly all compounds except saturated hydrocarbons and alkyl halides form complexes with iodine. The intensities of the spots do not accurately indicate the amount of material present, except in the crudest way.
The second most common method of visualization is by an ultraviolet lamp. Under uv light, compounds often look like bright spots on the plate. This often suggests the structure of the compounds, because certain types of com­pounds shine very brightly under uv light since they fluoresce.
Another method with good results involves adding a fluorescent indicator to the adsorbent used to coat the plates. A mixture of zinc and cadmium sulfides is often used. When treated in this way and held under uv light, the entire plate fluoresces. However, dark spots appear on the plate where the separated com­pounds are seen to quench this fluorescence.
In addition to the above methods, several chemical methods are available that either destroy or permanently alter the separated compounds through reac­tion. Many of these methods are specific only for particular functional groups.
Alkyl halides can be visualized if a dilute solution of silver nitrate is sprayed on the plates. Silver halides are formed. These halides decompose if exposed to light, giving rise to dark spots (free silver) on the tlc plate.
Most organic functional groups can be made visible if they are charred with sulfuric acid. Concentrated sulfuric acid is sprayed on the plate, which is then heated in an oven at 110 °C to complete the charring. Permanent spots are thus created.
Colored compounds can be prepared from colorless compounds by mak­ing derivatives before spotting them on the plate. An example of this is the preparation of 2,4-dinitrophenylhydrazones from aldehydes and ketones, to produce yellow and orange compounds. One may also spray the 2,4-dinitro­phenyhydrazine reagent on the plate after the ketones or aldehydes have sepa­rated. Red and yellow spots form where the compounds are located. Other examples of this method are using ferric chloride for visualizing phenols and using bromocresol green for detecting carboxylic acids. Chromium trioxide, potassium dichromate, and potassium permanganate can be used for visualiz­ing compounds that are easily oxidized. p-Dimethylaminobenzaldehyde easily detects amines. Ninhydrin reacts with amino acids to make them visible. Nu­merous other methods and reagents available from various supply outlets are specific for certain types of functional groups. These visualize only the class of compounds of interest.

[lugh]

If large plates are used, materials can be separated and the separated components individually recovered from the plates. Plates used in this way are said to be preparative plates. For preparative plates, a thick layer of adsorbent is generally used. Instead of being applied as a spot or a series of spots, the mixture to be separated is applied as a line of material about 1 cm from the bottom of the plate. As the plate is developed, the separated materials form bands. After development, the separated bands are observed, usually by uv light, and the zones are outlined in pencil, If the method of visualization is destructive, most of the plate is covered with paper to protect it, and the reagent is applied only at the extreme edge of the plate.
Once the zones have been identified, the adsorbent in those bands is scraped from the plate and extracted with solvent to remove the adsorbed material. Filtration removes the adsorbent, and evaporation of the solvent gives the recovered component from the mixture.

THE Rf VALUE Thin-layer-chromatography conditions include:
1. Solvent system
2. Adsorbent
3. Thickness of the adsorbent layer
4. Relative amount of material spotted
Under an established set of such conditions, a given compound always travels a fixed distance relative to the distance the solvent front travels. This ratio of the distance the compound travels to the distance the solvent front travels is called the Rf value. The symbol Rf stands for "ratio to front," and it is expressed as a decimal fraction.
When the conditions of measurement are completely specified, the Rf value is constant for any given compound, and it corresponds to a physical property of that compound.
The Rf value can be used to identify an unknown compound; but like any other identification based on a single piece of data, the Rf value is best con­firmed with some additional data. Many compounds can have the same Rf value, just as many different compounds have the same melting point.
It is not always possible, in measuring an Rf value, to duplicate exactly the conditions of measurement another worker has used. Therefore, Rf values tend to be of more use to a single worker in one laboratory than they are to workers in different laboratories. The only exception to this is when two workers use tlc plates from the same source, as in commercial plates, or know the exact details of how the plates were prepared. Nevertheless, the Rf value can be a useful guide. If exact values cannot be relied on, the relative values can provide another worker with useful information about what to expect. Anyone using pub­lished Rf values will find it a good idea to check them by comparing them with standard substances whose identity and Rf values are known.
To calculate the Rf value for a given compound, one measures the distance that the compound has traveled from the point at which it was originally spot­ted. For spots that are not too large, one measures to the center of the migrated spot. For large spots, the measurement should be repeated on a new plate, using less material. For spots that show tailing, the measurement is made to the "center of gravity" of the spot; this first distance measurement is then divided by the distance the solvent front has traveled from the same original spot.

[lugh]

Thin-layer chromatography has several important uses in organic chemis­try. It can be used in the following applications:
1. To establish that two compounds are identical.
2. To determine the number of components in a mixture.
3. To determine the appropriate solvent or adsorbent for a column chromatographic separation.
4. To monitor a column chromatographic separation.
5. To check the effectiveness of a separation achieved on a column, by crystallization, or by extraction.
6. To monitor the progress of a reaction.
In all these applications, thin-layer chromatography has the advantage that only small amounts of material are necessary. Material is not wasted. With many of the visualization methods, less than a tenth of a microgram (10-7 g) of material can be detected. On the other hand, samples as large as a milligram may also be used. With preparative plates that are large (about 9 in. on a side) and have a relatively thick (>500 µm) coating of adsorbent, it is often possible to separate from 0.2 to 0.5 g of material at one time. The main disadvantage of tic is that volatile materials cannot be used, since they evaporate from the plates.
Thin-layer chromatography can establish that two compounds suspected to be identical are in fact identical. One simply spots both compounds side by side on a single plate and develops the plate. If both compounds travel the same distance on the plate (have the same Rf value), they are probably identical. If the spot positions are not the same, the compounds are definitely not identical. It is important to spot both compounds on the same plate. This is especially important with hand-dipped microscope slides, since they vary widely from plate to plate, no two plates having exactly the same thickness of adsorbent. If commercial plates are used, this precaution is not necessary, although it is nevertheless a good idea.
Thin-layer chromatography can establish whether a compound is a single substance or a mixture. A single substance gives a single spot no matter what solvent is used to develop the plate. On the other hand, the number of compo­nents in a mixture can be established by trying various solvents on a mixture. A word of caution should be given. It may be difficult, in dealing with compounds of very similar properties, isomers for example, to find a solvent that will sepa­rate the mixture. Inability to achieve a separation is not absolute proof that a compound is a single pure substance. Many compounds can be separated only by multiple developments of the tic slide, with a fairly nonpolar solvent. In this method, the plate is removed after the first development and allowed to dry. After being dried, it is placed in the chamber again and developed once more. This effectively doubles the length of the slide. At times, several developments may be necessary.
When a mixture is to be separated, tic can be used to choose the best solvent to separate it if column chromatography is contemplated. Various sol­vents can be tried on a plate coated with the same adsorbent as will be used in the column. The solvent that resolves the components best will probably work well on the column. These small-scale experiments are quick, use very little material, and save time that would be wasted by attempting to separate the entire mixture on the column. Similarly, tic plates can monitor a column. A solvent was found that would separate the mixture into four components (A-D). A column was run using this solvent and eleven fractions of 15 mL each were collected. Thin-layer analysis of the various fractions showed that fractions 1-3 contained component A; fractions 4-7, component B; fractions 8 and 9, component C; and fractions 10 and 11, component D. A small amount of cross-contamination was observed in fractions 3, 4, 7, and 9.
In another tlc example, a worker found a product from a reaction to be a mixture. It gave two spots, A and B, on a tlc slide. After the product was crystallized, the crystals were found by tlc to be pure A, whereas the mother liquor was found to have a mixture of A and B. The crystallization was judged to have purified A satisfactorily.
Finally, it is often possible to monitor the progress of a reaction by tlc. At various points during a reaction, samples of the reaction mixture are taken and subjected to tlc analysis. In this case, the desired reaction was the conversion of A to B. At the beginning of the reaction (0 hr), a tic slide was prepared that was spotted with pure A, pure B, and the reaction mixture. Similar slides were prepared at 0.5, 1, 2, and 3 hours after the start of the reaction. The slides showed that the reaction was complete in 2 hours. When the reaction was run longer than 2 hours, a new compound, side product C, began to appear. Thus, the optimum reaction time was judged to be 2 hours.

Anyeone who know's wath the fluricent material in TLC plates is ??

A mixture of zinc and cadmium sulfides is often used.
 

I think it doesn't make much sense to prepare your own analytical sheets.

While most of time this is undoubtedly the case, if one is attempting to determine the proper adsorbent mixture for column chromatography, as mentioned above, it's often necessary to roll your own, as not all mixtures are available  

[ClearLight]

Thx! to lugh for posting all that info!  I have made my own plates using tlc Silica gel 60 F254 G and also have the hard layer plates as well. Unfortunately the G (gypsum/plaster of paris) plates are waaay to soft to mark up.
  The fluorescent indicators are available @ 100gms for $48.00 in either 254nm or 338 nm wavelengths.  I'm going to build a roll your own concentration plate today using Cellite and gypsum for the Concentration/spotting zone and the regular SG 60 F254 for the rest of the layer and see how that works on a separation that has a lot of tailing on it. (i'm not overloading the plate).

  Another problem i've had is that some of the roll your own plates tend to slough off in the solvent, i'm not sure if I haven't activated them long enough or my water ratios are too high.. 2:1 makes a really thick slurry that is not easy to get a thin coating w/ even using the two plate dip and separate method.  The disadvantage of diluting further is that the UV indicator is not uniform.

  So I wanted the Hard Layer organic binder plates so that the wouldn't fall off and I can mark them up since some of the visualization reagents ( marquis for example) destroy the Fluorescent indicators.

[hest]

Thank'sa luge, guess the zinksulfide is the choise, don't like Cd

[Lilienthal]

lugh: why not write a digest  
ClearLight: Tailing often happens with amines in the absence of a base in the solvent. Simply put an open vial with concentrated ammonia into the chamber.