Sample Application

Samples should be placed on a TLC plate as a spot that is as small as possible. Sample volumes of 1 to 2 ml are common but when using the so called high performance TLC plates (HPTLC plates) coated with particles 5-7 mm in diameter a maximum loading of about 100-200 nl only can be tolerated. Ideally the sample area should be circular and not greater than 1 mm in diameter, on any HPTLC plate. As this requires the use of samples a few nanoliters in volume, most will need to be concentrated.

Manually, the sample can be applied with a micro-pipette and the solvent is then allowed to evaporate, This procedure is repeated until sufficient sample is placed on the plate. Micro-syringes can also be employed to place a sample on to the plate. With care, and a little local heating, sample concentration can also be accomplished. The contents of the syringe are slowly but continuously discharged onto the plate and at the same time the solvent is progressively evaporated. This procedure can be automated, using computer controlled syringes and, in this way, samples can be focused onto the plate and constrained to a very small area.

There are various devices that are commercially available that will apply samples to a TLC plate either individually or as multiple groups. An example of a device for automatically concentrating a sample and then placing it on a plate is shown in figure 33. The device consists of a plate with a small indentation, at the center of which is an aperture that can be connected either to a vacuum or to a source of gas pressure. A polymer film is placed over the plate surface and a vacuum applied to suck the film into the indentation.

Figure 33 A TLC Sampling Device Incorporating Sample Concentration