Thin Layer Chromatography is by far the most popular chromatography technique for qualitative and semi-quantitative analysis, since the 1960s!
Over the years, TLC started to be performed instrumentally, overcoming some of the limitations, such as sample application as bands instead of spots, quantitative evaluation by densitometry etc. However no SOP for HPTLC was ever established. This led to confusion between TLC and HPTLC, which led to instrumental TLC being wrongly called as HPTLC.
TLC is a manual method that uses devices designed in the 1960s and its methods cannot meet cGLP and other modern regulatory requirements. Instrumental TLC requires TLC software and instrumentation for sample application, chromatogram development, evaluation by scanner, chromatogram “image” visualisation and documentation all controlled by one software. However, the SOP is still missing. HPTLC requires the above instrumentation plus control of all parameters that affect chromatography, which is where the USP SOP comes into play.
The moment of truth came on August 1, 2015 when USP Chapter 203 became effective. PhEur has adopted exactly the same SOP albeit in different words, in 2017.
Both TLC and HPTLC are pharmacopoeial procedures; however, the following table shows the parameters that are required to be controlled in the two.
Column chromatography is to HPLC what TLC is to HPTLC.
No procedure can become a regulatory method in trillion dollar trades unless numerous experts and authorities are satisfied about the quality of results.
The main drawback of instrumental TLC i.e. chromatographic development done in non-standardised manner in an open system has been completely eliminated by the development of an automated development chamber, in HPTLC
HPTLC must not be looked through LC eyes. They are two separate techniques, each with its own strengths and limitations. Generally speaking LC scores in a technical sense while HPTLC scores in practical sense.
Limitations of HPTLC include short separation bed (62 mm effective length), limited number of samples per plate, presence of silica gel during detection (unless MS is the detector). However the advantages are too numerous, which are evident to an open and scientific mind. HPTLC is the fastest chromatography method; result is visible both to detector and human eye (!), analyses 50-60 samples of 5-6 different types in parallel, has very low cost of analysis, negligible maintenance and a log machine life. HPTLC is shareable since it is not dedicated to any sample. Multiple detection modes like UV, Visible, Fluorescence, Bio-assay, MS, post-chromatographic derivatisation etc. can be used without repeating chromatography. Sample cannot contaminate chromatograph. Cross contamination is impossible. Samples and standards are chromtographed in identical conditions. HPTLC instrument can be share. No set-up time. Hyphenation with MS can produce several dozen MS per day. These are but a few advantages.
HPTLC is the ideal method for first opinion on any organic, non-volatile sample.
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