An Overview of HPTLC: A Modern Analytical Technique with Excellent Potential for Automation, Optimization, Hyphenation, and Multidimensional Applications



High performance thin layer chromatography (HPTLC) is a sophisticated instrumental technique based on the full capabilities of thin layer chromatography. The advantages of automation, scanning, full optimization, selective detection principle, minimum sample preparation, hyphenation, etc. enable it to be a powerful analytical tool for chromatographic information of complex mixtures of inorganic, organic, and biomolecules. The chapter highlights related issues such as journey of thin-layer chromatography, basic principle, protocol, separation, resolution, validation process, recent developments, and modifications on TLC leading to the HPTLC, optimization, process control, automation, and hyphenation. It explains that HPTLC has strong potentials as a surrogate chromatographic model for estimating partitioning properties in support of combinatorial chemistry, environmental fate, and health effect studies.

Suggested Readings

  1. Adegbola AR, Dare AA and Lasisi EO. Spray reagents for the visualization and detection of sesame oil unsaponiiables on thin-layer chromatograms. Chromatographia, 2008, 68, (1–2), 151–155.CrossRefGoogle Scholar
  2. Ahmad S, Rizwan M, Abeaparveen R, Mujeeb M and Aquil M. A validated stability indicating TLC method for determination of forskolin in crude drug and pharmaceutical dosage form. Chromatographia, 2007, 67, (5–6), 441–447.Google Scholar
  3. Ahuja S (1992). Chromatography and Pharmaceutical Analysis. In Chromatography of Pharmaceuticals; published by American Chemical Society: Washington, DC.Google Scholar
  4. Bagócsi B, Végh Z and Ferenczi F. Optimization of the visualization of steroids separated by OPLC. J Planar Chromatogr, 2008, 21, 107–112.CrossRefGoogle Scholar
  5. Bard AJ, Faulkner LR (2000). Electrochemical Methods: Fundamentals and Applications, 2nd edition, published by John Wiley & Sons, New York.Google Scholar
  6. Bauer GK, Pfeifer AM, Hauck HE and Kovar K. A. Development of an optimized sorbent for direct HPTLC-FTIR on-line coupling. J Planar Chromatogr-Mod TLC, 1998, 11, 84–89.Google Scholar
  7. Bezuidenhout W and Brett MJ. Ultrathin layer chromatography on nanostructured thin films. J Chromatogr A, 2008, 118, (1–2), 179–185.CrossRefGoogle Scholar
  8. Burger K., Z. Anal. Chem., 1984, 318, 228.Google Scholar
  9. Burger K. Instrumental Thin-Layer Chromatography/Planar Chromatography, Proceedings of the International Symposium, Brighton, UK, 1989, 33–44.Google Scholar
  10. Cha S, Yeung ES. Colloidal graphite-assisted laser desorption/ionization mass spectrometry and MSn of small molecules. 1. Imaging of cerebrosides directly from rat brain tissue. Anal Chem, 2007, 79, 2373–2385.CrossRefGoogle Scholar
  11. Christensen JH, Mortensen J, Hansen AB and Andersen O. Chromatographic preprocessing of GC-MS data for analysis of complex chemical mixtures. J Chromatogr A, 2005, 1062, (1), 113–123.CrossRefGoogle Scholar
  12. Cserhati T, Forgacs E, Morais MH and Ramos AC. TLC-FTIR of color pigments of chestnut sawdust. J Liq Chromatogr Rel Technol, 2001, 24, 1435–1445.CrossRefGoogle Scholar
  13. Davld N, Rose WI Becker and Karen A. Sassic. Time optimization in thin-layer chromatography. Anal Chem. 1982, 54, 1955–1959.Google Scholar
  14. Ellis LA and Roberts DJ. Chromatographic and hyphenated methods for elemental speciation analysis in environmental media. J Chromatogr A, 1997, 774 (1–2), 3–19.CrossRefGoogle Scholar
  15. Fuchs B, Schiller J, Süß R, Zscharnack M, Bader A, Müller P, Schürenberg M and Becker M. Analysis of stem cell lipids by offline HPTLC-MALDI-TOF MS Detlev Suckau. Anal Bioanal Chem, 2008, 392, 849–860.CrossRefGoogle Scholar
  16. Gombosuren N, Nova Z, Kotschy A, Mincsovics E and Dibo G. A multidimensional overpressured layer chromatographic method for the characterization of tetrazine libraries. J Biochem Biophys Methods, 2007, 69, 239–249.CrossRefGoogle Scholar
  17. Gopu C, Aher S, Mehta H, Paradkar A and Mahadik K. Simultaneous determination of cinnamaldehyde, eugenol and piperine by HPTLC densitometric method Phytochem Anal, 2008, 19, 116–121.CrossRefGoogle Scholar
  18. Guetens G, and De Boeck G, Highley MS, Wood M, Maes RA, Eggermont AA, Hanauske A, de Bruijn EA, Tjaden UR. Hyphenated techniques in anticancer drug monitoring. II. Liquid chromatography-mass spectrometry and capillary electrophoresis-mass spectrometry. J Chromatogr A, 2002, 976, (1–2), 239–247.Google Scholar
  19. Guetens G, De Boeck G, Wood M, Maes RA, Eggermont AA, Highley MS, van Oosterom AT, de Bruijn EA and Tjaden UR. Hyphenated techniques in anticancer drug monitoring I. Capillary gas chromatography-mass spectrometry. J Chromatogr A, 2002, 976, (1–2), 229–238.Google Scholar
  20. Halpaap H. J. Chromatogr, 1973, 78, 77–78.Google Scholar
  21. Halpaap H, Krebs KF and Hauck HE. J. HRC and CC, 1980, 3, 215–240.Google Scholar
  22. Hauck HE and Jost W, ‘Instrumental High Performance, Thin-Layer Chromatography', Proceedings of 2nd International Symposium, Kaiser RE (ed.), Interlaken, Switzerland, 1982, 25–37.Google Scholar
  23. Hhodisan Y, Ccasoni D, Beldean-Ggalea MS and Cimpoiu C. Identification and quantification of tocopherols in vegetable oil by thin-layer chromatography. J Planar Chromatogr, 2008, 21, 213–215.CrossRefGoogle Scholar
  24. Saxena S, Jain DC, Gupta MM, Bhakuni RS, Mishra HO and Sharma RP. High-performance thin-layer chromatographic analysis of hepatoprote diterpenoids from Andrographis paniculata. Phytochem Anal, 2000, 11, 34–36.CrossRefGoogle Scholar
  25. Holt RM, Newman MJ, Pullen FS, Richards DS and Swanson AG. High-performance liquid chromatography/NMR spectrometry/mass spectrometry: further advances in hyphenated technology. J Mass spectrometry, 1997, 32, (1), 64–70.CrossRefGoogle Scholar
  26. Janecki DJ, Novotny AL, Woodward SD, Wiseman JM and Nurok D. A preliminary study of the coupling of desorption electrospray ionization-mass spectrometry with pressurized planar electrochromatography. J Planar Chromatogr, 2008, 21, 11–14.CrossRefGoogle Scholar
  27. Jäntschi L, Bolboacă DS, Ungureşan ML. Mobile Phase Optimization in Three Solvents High Performance Thin-Layer Chromatography: Methodology and Evaluation. 6th European Conference on Computational Chemistry, September 3–7, 2006, Slovakia.Google Scholar
  28. Kaiser RE. Instrumental Thin-Layer Chromatography/Planar Chromatography’, Proceedings of the International Symposium, Brighton, UK, 1989, 251–262.Google Scholar
  29. Kirchner JG, Miller JM and Keller GE. Anal. Chem., 1951, 23, 420.Google Scholar
  30. Kirchner JG. (1978) Thin-Layer Chromatography, 2nd edition, Techniques in Chemistry, vol. XIV, published by Wiley-Interscience, Chichester, UK.Google Scholar
  31. Kowalska T, Sherma J (2007). Thin-layer Chromatography in Chiral Separations and Analysis. Chromatographic Science Series, vol. 98, published by CRC Press, Boca Raton, New York.Google Scholar
  32. Kwiecien A, Krzek J and Biniek L. TLC-densitometric determination of azithromycin in pharmaceutical preparations. J Planar Chromatogr, 2008, 21, 177–181.CrossRefGoogle Scholar
  33. Leichert M, Eckhardt HS, Klein KF and Spangenberg F. A simple and reliable method for quantification of glucosamine in nutritional supplements. J Planar Chromatogr, 2008, 21, 55–59.CrossRefGoogle Scholar
  34. Mavle R, Katkar H, Daundkar B, Malve M and Krishnamurthy R. Thin layer chromatographic technique for detection and identification of endosulfan insecticide with m-dinitrobenzene reagent. J Planar Chromatogr, 2008, 21, 197–198.CrossRefGoogle Scholar
  35. Meinhard JE and Hall NF. Anal. Chem., 1949, 2l, 185.Google Scholar
  36. Mmirakor V, Vaidya V, Menon S and Champanerker P. HPTLC method for determination of colchicines in a pharmaceutical formulation. J Planar Chromatogr, 2008, 21, 187–189.Google Scholar
  37. Naik S, Desai SK, Nanda RK and Narayanan MS. Fermentation, isolation, purification, and biological activity of SJA-95, a heptaene polyene macrolide antibiotic produced by the Streptomyces sp. Strain S24. Arzneim-Forsch/Drug Res, 2007, 56, 171–179.Google Scholar
  38. Orinák A, Talian I, Efremov EV, Ariese F and Oriáakovác R. Diterpenoic acids analysis using a coupled TLC-surface-enhanced Raman spectroscopy system. Chromatographia, 2008, 67–74, 15–19.Google Scholar
  39. Patel BH, Suhagia BN, Patel MM and Patel JR. High-performance liquid chromatography and thin-layer chromatography for the simultaneous quantitation of rabeprazole and mosapride in pharmaceutical products. J Chromatogr Sci, 2008, 46, 10–14.Google Scholar
  40. Patel RK and Prajapati AM. Development and validation of a visible absorption densitometry method for quantitation of conessine in Holarrhena antidysenterica (Kurchi). J AOAC Int, 2008, 91.Google Scholar
  41. Peterson BL and Cummings BS. A review of chromatographic methods for the assessment of phospholipids in biological samples. Biomed Chromatogr, 2006, 20, 227–243.CrossRefGoogle Scholar
  42. Polak B, Halka A and Dzido T. Pressurized planar electrochromatographic separation of the enantiomers of tryptophan and valine. J Planar Chromatogr, 2008, 21.Google Scholar
  43. Poole CF. Thin layer chromatography: challenges and opportunities. J Chromatogr, 2003, 1000, 963–984.CrossRefGoogle Scholar
  44. Prichard E and Barwick V (2007). Quality Assurance in Analytical Chemistry. Analytical Techniques in the Sciences; published by Wiley-Interscience: Hoboken, NJ.Google Scholar
  45. Rahul G, Leena B, Asfak V and Mrinalini D. A validated high performance thin layer chromatographic method for simultaneous estimation of oloxacin and satranidazole in pharmaceutical dosage form. J Pharm Res, 2007, 6, (4).Google Scholar
  46. Randerath K. In Thin-Layer Chromatography, Academic Press, London, UK, 1962.Google Scholar
  47. Reich E, Schibli A and Debatt A. Validation of HPTLC methods for the identification of botanicals in a cGMP environment. J AOAC Int, 2008, 91, 10–20.Google Scholar
  48. Reitsema RH. Anal. Chem., 1954, 26, 960.Google Scholar
  49. Salo PK, Pertovaara AM, Salo VMA, Salomies HEM and Kostiainen RK. High-performance thin-layer chromatography method for assessment of the quality of combinatorial libraries, and comparison with liquid chromatography ultraviolet mass spectrometry. J Comb Chem, 2003, 5, 223–232.CrossRefGoogle Scholar
  50. Salo PK, Vilmunen S, Salomies H, Ketola RA and Kostiainen R. Two-dimensional ultra-thin-layer chromatography and atmospheric pressure matrix-assisted laser desorption/ionization mass spectrometry in bioanalysis. Anal Chem, 2007, 79, 2101–2108.CrossRefGoogle Scholar
  51. Sean X. Peng, Charles Henson, Michael J. Strojnowski, Adam Golebiowski, and Sean R. Klopfenstein. Automated high-throughput liquid extraction for initial purification of combinatorial libraries. Anal Chem, 2000, 72, 261–266.Google Scholar
  52. Somsen GW and Visser T. (2003) Liquid Chromatography/Infrared Spectroscopy. In Encyclopedia of Analytical Chemistry; Meyers RA, ed.; John Wiley & Sons Ltd: Chichester.Google Scholar
  53. Soponar F, Mot AC and Sârbu C. Quantitative determination of some food dyes using digital processing of images obtained by thin-layer chromatography. J Chromatogr A, 2008, 1188, (2), 295–300.CrossRefGoogle Scholar
  54. Stahl E. (1965) Thin-Layer Chromatography – A Laboratory Handbook, published by Springer-Verlag, Berlin, Germany.Google Scholar
  55. Stahlmann S and Kovar KA. Analysis of impurities by high-performance thin-layer chromatography with Fourier transform infrared spectroscopy and UV absorbance detection in situ measurement: chlordiazepoxide in bulk powder and in tablets. J Chromatogr A, 1998, 813, 145–152.CrossRefGoogle Scholar
  56. Tang T, Hong Wu. An image analysis system for thin-layer chromatography quantification and its validation. J Chromatogr Sci, 2008, 46, (6), 560–564.Google Scholar
  57. Tyihák E, Mincsovics E, Kátay G, Király-Véghely Z, Móricz AM and Ott PG. An unlimited possibility of biochemical interactions in the adsorbent layer after chromatographic separation. J Planar Chromatogr, 2008, 21, 15–20.CrossRefGoogle Scholar
  58. Urakova I, Pozharitskaya O, Shikov A, Kosman V and MakaRov V. Comparison of high performance TLC and HPLC for separation and quantification of chlorogenic acid in green coffee bean extracts. J Sep Sci, 2008, 1, 207–241.Google Scholar
  59. Waksmundzka-hajnos M and Józwiak GW. Special modes of development in preparative-layer chromatography of extracts of alkaloids from Fumaria oficinalis. J Planar Chromatogr, 2008, 21, 61–66.CrossRefGoogle Scholar
  60. Wall PE. (2005). Thin-layer Chromatography a Modern Practical Approach. RSC Chromatography Monographs Salesand Customer Care. Smith, RM, ed.; Royal Society of Chemistry, Thomas Graham House, Science Park, Milton Road, Cambridge.Google Scholar
  61. Wang PG (2008). High-Throughput Analysis in the Pharmaceutical Industry; Critical Reviews in Combinatorial Chemistry; CRC Press, Boca Raton, FL.Google Scholar
  62. Wilkins CL. Hyphenated techniques for analysis of complex organic mixtures, Science, 1983, 222, (4621), 291–296.CrossRefGoogle Scholar
  63. Workman J, Koch M, Lavine B and Chrisman R. Process analytical chemistry. Anal Chem, 2009, 81, 4623–4643.CrossRefGoogle Scholar
  64. Yuanling L, Mark EB, Todd Y and Bonner DM. quantitative analysis of aflatoxins by high-performance thin-layer chromatography utilizing a scientifically operated charge-coupled device detector. Anal Chem, 1996, 68, 3885–3891.CrossRefGoogle Scholar
  65. Zlatkis A and Kaiser RE. (1977) HPTLC High Performance Thin-Layer Chromatography, published by Elsevier, Amsterdam, Netherlands.Google Scholar

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© Springer-Verlag Berlin Heidelberg 2011

Authors and Affiliations

  1. 1.Department of Chemistry, Faculty of ScienceDayalbagh Educational InstituteAgraIndia

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