Thin-layer chromatography (TLC) is often used in laboratories dealing with chemical analysis thanks to many methodological possibilities for conducting the separation process and modern equipment. Recently, the combination of high-performance thin-layer chromatography (HPTLC) with instrumental analysis techniques, especially mass spectrometry, has been developed [1], [2], [3], [4], [5], [6], [7], [8], [9], [10], [11]. The combination of thin-layer chromatography and mass spectrometry (MS) can be divided into two groups of techniques. The first involves solvent extraction, and the obtained extract is sent to a mass spectrometer for analysis using either an online or offline approach. The second group comprises techniques that combine desorption in the liquid or gas phase with the ionization of substances, which are then directed straight to the mass spectrometer. The first group of techniques includes extraction of substances from the scraped adsorbent layer, direct elution of substances from a spot in the adsorbent layer on a chromatographic plate, and liquid microjunction surface sampling probe. In the second group, some examples include techniques such as desorption electrospray ionization [9], laser desorption ionization [12] matrix-assisted laser desorption/ionization [13], direct analysis in real-time [14], atmospheric pressure chemical ionization [15]. Among the techniques of the first group, the technique of eluting substances from spots on a chromatographic plate has gained great commercial importance [16], [17], [18]. It involves pressing a special piston/stamp against the adsorbent layer, which tightly covers a spot of the substance or part of it with its pressing area against the chromatographic plate. There are two channels in the piston/stamp, one for supplying (inlet channel) the elution solvent and the other for removing (output channel) the solvent. The liquid pump pumps the solvent for elution through the inlet channel, across the adsorbent layer pressed by the piston/stamp, and out through the output channel. From the output channel, the solution of the substance(s) eluted from the adsorbent layer can be collected into vials or directly supplied into analytical instruments for analysis using techniques like high-performance liquid chromatography (HPLC), liquid chromatography-mass spectrometry (LC-MS), or MS. A certain disadvantage of this approach is the need to use a liquid pump, which significantly increases the price of the entire set for eluting substances from the adsorbent layer. Moreover, there is a need to wash the piston and the output channel before the process of eluting the substance from the next spot on/in the adsorbent layer. Therefore, there is a risk of cross-contamination between samples. Furthermore, there are rare problems with clogging of the inlet and outlet channels, as well as leakage of the eluting solvent at the connection of the pressure piston/stamp with the carrier plate of the adsorbent layer. In the liquid microjunction surface sampling probe technique [19], the extracting liquid is continuously supplied to a very small surface of the adsorbent layer and discharged continuously using a special coaxial tube system. Due to this, substances are extracted from the place where the liquid contacts the adsorbent layer. The condition for using this technique is that the extraction liquid cannot wet the adsorbent layer, which significantly limits the use of this technique for chromatographic plates.

In our paper, we present a new approach to extracting substances from a spot in an adsorbent layer of a chromatographic plate to use the obtained solution for analysis with instrumental techniques, especially LC-MS. A prototype device was used for the research, based on the example of extraction of six selected coccidiostats, which are substances used to prevent coccidiosis in farm animals, especially poultry. Coccidiosis is caused by protozoa of Eimeria. This approach is characterized by the elimination of some of the unfavorable features associated with the use of extraction techniques belonging to the above-mentioned first group of techniques. The use of this new approach is characterized by performing several simple operations using the device described in our paper.