Nowadays it is possible to detect the

Nowadays it is possible to detect the AChE inhibitors by several methods such as gas chromatography-mass spectrometry [11], ion mobility spectrometry [12] and lab-on-chip devices [13,14]. However, almost all instrumental detection techniques that are able to conclusively determine the presence of the nerve agents, organophosphorus and carbamate insecticide are expensive, usually non-transferable and very time-consuming. For a quick personal and field detection by emergency and military services handheld chemical reactions based on biosensors are more desirable [15]. This is due to their high sensitivity and rapid determination but also a low cost value [16]. Some of the basic set-ups for these devices are so-called tube detectors, consisting of a sealed glass tube filled with a carrier containing immobilized chemical reagents. The analyzed air or liquid is fed to a working chamber inside the tube and the presence of the chemical warfare agent is subsequently indicated by coloration of the carrier. The most frequently used method for colorimetric detection of AChE inhibitors is the Ellman’s reaction. Acetylthiocholine or butyrylthiocholine (alternative substrates to physiological acetylcholine) is hydrolyzed by the immobilized acetylcholinesterase or butyrylcholinesterase (BuChE) to acetic MK-0752 or butyric acid, respectively, and thiocholine, which consequently reacts with the chromogenic reagent 5,5′-dithiobis(2-nitrobenzoic) acid forming 2-nitro-5-thiobenzoic acid, which manifests itself as a yellow color [17]. However, in the presence of the nerve agent or organophosphate/carbamate insecticides, the reaction is blocked with no color change being apparent. Besides the multiple advantages of this method, such as the speed of the reaction, sensitivity and simplicity, the main disadvantage is the indistinctive color transition from white to yellow. Reflecting the fact that the nerve agents (as AChE inhibitors) have already been misused in several terrorist acts or war conflicts [[18], [19], [20], [21]], and considering the current global security situation, it is of great importance to develop further effective and easy-to-maintain detection devices. Similarly there is a great risk of a contamination of water supplies in developing countries where organophosphorus insecticides are still widely used [22]. In our previous studies we have used a concept of enzyme impregnated matrix carriers based on microcrystalline cellulose, showing a promising potential [23,24]. This work was followed by a recently published study, which dealt with the addition of magnesium aluminometasilicate for improving the shape and structure of the carriers for a faster and more distinct transition from white to yellow, thus allowing an easier and more accurate detection of nerve agents by the naked eye [25]. However, in the impregnated matrix systems we experienced one major disadvantage – leaching of color substances to the testing solution, therefore making it harder to determine the test results. The presented study offers a novel pharmaceutical technological approach to this issue, utilizing double-coated pellets as a reagent carrier for detection of cholinesterase inhibitors in liquids. This carrier enables a more distinctive color transition during the detection of cholinesterase inhibitors. Our recently granted patent also confirms the uniqueness of this technology [26].
Experimental
Results and discussion In the presented study four samples of double-coated pellets each in three batches with different thicknesses of the second coat, composed of Eudragit® RL, Kollidon® 25 and PEG 400, were prepared. The semipermeable second coat was expected to prevent the enzyme leaching of carriers into the solution in which pellets were immersed during the detection. Moreover, due to this coating an improvement in color intensity change was expected together with a more distinctive difference between the color of the test solution and the created yellow color of the pellets during the detection in the case when the enzyme is not inhibited. All of these parameters are essential for ensuring fast, accurate and distinct detection of the cholinesterase inhibitors within liquids. Besides this, carriers intended as a filler for detection tubes should be an approximate size of 1 mm and spherical in shape for an optimal resolution of color change and easy filling into the detection tubes. Suitable carriers must also show a good mechanical resistance to enable filling the detection tubes.