Thin-layer chromatography (TLC)
Thin-layer chromatography (TLC) is a widely-used analytical technique that separates chemical compounds into their individual components. This method is particularly significant in forensic science, where it aids in identifying substances and linking samples from crime scenes to potential suspects. TLC can analyze various materials, including the dye composition of fibers, poisons in food, pigments in plants, and components of drugs or explosives.
In this technique, a thin layer of a solid stationary phase, such as silica gel or alumina, is applied to a flat surface, and a solvent serves as the mobile phase. The sample is spotted on the plate and placed in a solvent chamber, where capillary action causes the solvent to rise and carry the sample components with it. The distance each component travels depends on its solubility and affinity for the solvent.
For colorless compounds, visualization can be achieved through fluorescent compounds or iodine vapors. The separation efficiency can be measured using retention factor values, allowing for comparisons between known and unknown substances. Recent advancements have integrated TLC with mass spectrometry, enhancing its analytical capabilities by enabling detailed molecular identification. This makes TLC a valuable tool in both scientific research and forensic investigations.
On this Page
Subject Terms
Thin-layer chromatography (TLC)
DEFINITION: Technique used to separate chemical compounds into their individual components.
SIGNIFICANCE: By using thin-layer chromatography to determine the chemical components that make up particular substances, forensic scientists can help to identify the origins of those substances or link samples found at crime scenes to potential suspects.
Using thin-layer chromatography (TLC), forensic scientists can analyze the dye composition of fibers, poisons in food, pigments contained in plant specimens, or the ingredients in chemical weapons, explosives, or drugs. This method can also be used to detect the presence of a in urine or blood.
TLC involves a stationary phase (a solid) and a mobile phase (a liquid or gas). As the name suggests, the technique uses a thin layer of silica gel, alumina, or cellulose coated on a piece of flat and inert glass, acetate, metal, or plastic. The silica gel or alumina is the stationary phase. The mobile phase is the solvent used.
First, a solution containing the sample of interest is “spotted” or applied to the TLC plate alongside reference or control spots (of solutions containing known substances) near the bottom of the plate. The plate is dipped into a solvent, often ethanol or water, such that the plate is minimally submerged. The chamber containing the solvent and plate is covered. By capillary action, the solvent travels up the TLC plate. The spots are dissolved and moved up by the solvent. This is called chromatographic development. The rate and distance of movement depend on the molecular forces and solubility of the chemical compounds in the solvent. Solutes (the compounds contained within a spot) with a greater affinity for the solvent will tend to spend more time with the solvent than solutes with less affinity for the solvent.
Colorless substances can also be separated by TLC. One common method involves the addition of a fluorescent compound such as manganese-activated zinc silicate to the adsorbent and visualization under a black light. Another method is the use of iodine vapors as a general unspecific color reagent.
The movement of the solvents can be determined through the calculation of a retention factor value. Retention factor values of known and unknown compounds can be compared to provide an index of similarity. Compounds with similar retention factor values tend to share solubility characteristics.
In 2013, scientists used pre-coated plates to combine mass spectrometry with TLC. Mass spectrometry is a scientific practice that allows for the identification of a substance based on the size of its molecules. By combining the two practices, scientists were granted additional highly-detailed analytical tools.
Bibliography
Hahn-Deinstrop, Elke. Applied Thin-Layer Chromatography: Best Practice and Avoidance of Mistakes. Translated by R. G. Leach. 2d ed. Weinheim, Germany: Wiley-VCH, 2007.
"History of Thin-Layer Chromatography." Millipore Sigma, 2013, www.sigmaaldrich.com/US/en/technical-documents/technical-article/analytical-chemistry/thin-layer-chromatography/tlc-history?srsltid=AfmBOorA0SLr3MBUbP9tyGpuhQS0Og9ZKWFTkrFYB‗ceIkQ22Lmkvy-n. Accessed 18 Aug. 2024.
Houck, Max M., and Jay A. Siegel. Fundamentals of Forensic Science. 3d ed., Elsevier/Academic Press, 2015.
James, Stuart H., Jon J. Nordby, and Suzanne Bell, eds. Forensic Science: An Introduction to Scientific and Investigative Techniques. 4th ed. Boca Raton, Fla.: CRC Press, 2014.
Lewis, S. W., and C. E. Lenehan. "Liquid and Thin-Layer Chromatography." In Forensic Chemistry, edited by Max M. Houck. San Diego, Calif.: Academic Press, 2015.
Sherma, Joseph, and Bernard Fried, eds. Handbook of Thin-Layer Chromatography. 3d ed. New York: Marcel Dekker, 2003.
Zou, Xiaowei, et al. "Advances in Thin Layer Chromatography Coupled With Mass Spectrometry Technology." National Library of Medicine, doi: 10.3724/SP.J.1123.2022.03038. Accessed 18 Aug. 2024.