High performance thin-layer chromatography HPTLC

7 HIGH PERFORMANCE THIN-LAYER CHROMATOGRAPHY (HPTLC) 

Recent developments in the practice of thin-layer chromatography have resulted in a breakthrough in performance which has led to the expression high performance thin-layer chromatography . These developments have not been the result of any specific advance in instrumentation (as with HPLC), but rather the culmination of improvements in the various operations involved in TLC. The three chief features of HPTLC are summarised below, but for a comprehensive account of the subject the reader is recommended to consult a more specialised text.59 

Quality of the adsorbent layer. Layers for HPTLC are prepared using specially purified silica gel with average particle diameter of 5-15 /mi and a narrow particle size distribution. The silica gel may be modified if necessary, e.g. chemically bonded layers are available commercially as reverse-phase plates. Layers prepared using these improved adsorbents give up to about 5000 theoretical plates and so provide a much improved performance over conventional TLC this enables more difficult separations to be effected using HPTLC, and also enables separations to be achieved in much shorter times. 

Methods of sample application. Due to the lower sample capacity of the H PTLC layer, the amount of sample applied to the layer is reduced. Typical sample volumes are 100-200 nL which give starting spots of only 1.0-1.5 mm diameter after developing the plate for a distance of 3-6 cm, compact separated spots are obtained giving detection limits about ten times better than in conventional TLC. A further advantage is that the compact starting spots allow an increase in the number of samples which may be applied to the HPTLC plate. 

The introduction of the sample into the adsorbent layer is a critical process in HPTLC. For most quantitative work a platinum-iridium capillary of fixed volume (100 or 200 nL), sealed into a glass support capillary of larger bore, provides a convenient spotting device. The capillary tip is polished to provide a smooth, planar surface of small area (ca 0.05 mm2), which when used with a mechanical applicator minimises damage to the surface of the plate spotting by manual procedures invariably damages the surface. 

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Commercially available pre-coated plates with a variety of adsorbents are generally very good for quantitative work because they are of a standard quality. Plates of a standardised silica gel 60 (as medium porosity silica gel with a mean porosity of 6mm) released by Merck have a specific surface of 500 m /g and a specific pore volume of 0.75 mL/g. They are so efficient that they have been called high performance thin layer chromatography (HPTLC) plates (Ropphahn and Halpap J Chromatogr 112 81 1975). In another variant of thin layer chromatography the... 

The reason for this lies not least in the increasing instrumentalization and delibei automation of all those processes which were earlier particularly subject to eri (Fig. 2). Modem high performance thin-layer chromatography (HPTLC) is no Ion inferior to other liquid chromatographic techniques with respect to precision and s sitivity (Fig. 3) [6]. 

Morita et al. [69] optimized the mobile phase composition using the PRISMA model for rapid and economic determination of synthetic red pigments in cosmetics and medicines. The PRISMA model has been effective in combination with a super modihed simplex method for fadhtating optimization of the mobile phase in high performance thin layer chromatography (HPTLC). 

Principles and Characteristics High-performance thin-layer chromatography (HPTLC), also known as planar chromatography, is an analytical technique with separation power and reproducibility superior to conventional TLC, which was first used in 1938 [7] and modified in 1958 [8]. HPTLC is based on the use of precoated TLC plates with small particle sizes (3-5 xm) and precise instruments for each step of the chromatographic process. 

High-performance thin-layer chromatography (HPTLC) has become widely used and while it follows the same principles as TLC, it makes use of modern technology including automatic application devices and smaller plates, which allow for better sensitivity. 

A. Jamshidi, M. Adjvadi and S.W. Husain, Determination of kaempferol and quercetin in the extract of Ginkgo biloba leaves by high-performance thin-layer chromatography (HPTLC). J. Plan. Chromatogr.—Mod. TLC. 13 (2000) 57-59. 

M. Lambri, M. Jourdes, Y. Glories and C. Saucier, High performance thin layer chromatography (HPTLC) analysis of red wine pigments. J. Planar Chromatogr.—Mod. TLC 16 (2003) 88-94. 

Muscarine is detected by high-performance thin-layer chromatography (HPTLC), with a limit of detection of 50 ng (Stijve, 1981). Using Dragendorff reagent, muscarine appears as an orange spot. 

A number of compounds have been quantified by tic or high performance thin-layer chromatography (hptlc) using absorption or fluorescence scanning densitometry. 

Benzimidazole carbamates have generally been determined by HPLC (20,124-126), although other methods have been reported based on enzyme immunoassay (147) or in gas chromatography (124). The commercial availability of high-performance thin-layer chromatography (HPTLC) plates with layers of very different polarities has improved the sensitivity, selectivity, and analytical precision of TLC, which has led to a reconsideration of the use of this technique for qualitative and quantitative analysis (54). 

High performance thin-layer chromatography (HPTLC) on silica gel plates was used for the separation >3-adrenoceptor blocking drugs. The detection limit for atenolol was 25.o ng, and the absorption wavelength selected for its determination was 2o5 nm.(8)... 

Note that Jenke " also cites references to the use of other related analytical techniques, including Head-space GC, high performance thin layer chromatography (HPTLC), supercritical fluid chromatography (SFC) /MS, and supercritical fluid chromatography/ Fourier transform infrared spectroscopy (SFC/FTIR). 

In this article, we will review some examples of these three areas of plant chemistry and see how thin-layer chromatography (TLC) or high-performance thin-layer chromatography (HPTLC) can fulfill the various requirements. 

The benefit of using thin-layer chromatography (TLC) for the identification of unknown vitamins and related compounds by comparing Rf values of the unknown compounds with authentic vitamins is beyond doubt. The quantification of the separated vitamins can be performed by the use of modem densitometry. TLC [or high-performance thin-layer chromatography (HPTLC)], as a powerful separation and analytic tool, is used particularly with pharmaceutical preparations and food products. Because amounts of most hydrophilic vitamins are low, or very low, in tissues or body fluids, bioautography or de-rivatization is used before densitometry. 

Since the introduction of commercial precoated plates in the mid-1960s, continual developments with regard to the increase of selectivity and improvement of separation efficiency were pursued [i.e., ready-to-use layers suitable for high-performance thin-layer chromatography (HPTLC), polar and hydrophobic bonded phases, plates with concentrating zones],... 

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