Anchrom HPTLC

TLC photodocumentation system

Discover the TLC Photodocumentation System by Anchrom!

Capture and document your Thin Layer Chromatography (TLC) results with ease using Anchrom's TLC Photodocumentation System. Our innovative system is specifically designed for efficient and accurate imaging of TLC plates, allowing you to capture high-resolution images of your chromatograms.

Hptlc in food Analysis

HPTLC for Drug Analysis 

Method Of Separation of Coloured Dyes 

Chromatography is the science of the separation of components from a mixture. These mixtures could be anything from food, pharma, herbal, forensic or textile samples. 

Of these e.g., textile dyes, food dyes, and ink are examples of different dyes. These dyes are separated by chromatography. 

TLC/HPTLC (High-Performance Thin Layer Chromatography) is one of the best chromatographic techniques for the effective separation of dye mixtures. 

There are many examples of dyes i.e., Solvent dyes (e.g., Sudan Red R, Sudan orange G, Sudan Blue G etc.), Disperse Dyes (e.g., Celliton Fast Blue B, Celliton Fast Pink B), Azo pigments Permanent Red FR extra, Amaplast orange ORC, basic dyes (e.g., Rhodamine B, Malachite Green etc.), Acid dyes (Metanil yellow, Ponceau R) etc. 

These dyes have applications in various industries like paper, leather, textile, polyamide fibres, inks etc. Only certain acid dyes are permitted for colouring foodstuffs and such food colourants are treated under different categories) of permissible food colours. 

High-Performance Thin Layer Chromatography (HPTLC) is a powerful analytical methodology suitable for qualitative and quantitative analysis. 

HPTLC practice is performed using highly sophisticated instruments/modules and is GLP/USP/EP-compliant chromatography. 

The unique features of HPTLC are that it is the fastest, simplest, most economical and flexible, “visible” technique which can analyse more than 100 samples in parallel. It is risk-free and multiple detections can be made without repeating chromatograms. 

After doing a single analysis, the analyst gets many data points. Data is obtained at 254 nm, 366 nm, in white light and after derivatisation in white light or at 366 nm. 

For quantitative analysis, densitometric scanning is done by means of a scanner. Spectral evaluation is also possible, as it has a wide scanning range starting from 190-900nm. 

Nowadays HPTLC can also be coupled with mass spectrometry by means of a TLC-MS interface for the identification of compounds by mass-to-charge ratio and it can also be subjected to other techniques and like IR, and  NMR for further characterization studies.

Let us take one example to understand the application of HPTLC for the separation of coloured dyes simultaneous identification and quantification of synthetic food colours in Kala khatta syrup and Halwa barfi (Indian sweet) samples.

 Synthetic colours like Carmoisine, Sunset Yellow FCF and Brilliant blue FCF are FSSAI permitted colours to be used in food products.  These colours are added to food to make it visually appealing. 

The FSSAI has set a limit of 100 ppm for the usage of these food colours. Identification and quantification of these food colours in the food products can be efficiently performed by HPTLC. 

HPTLC identification and quantification of Carmoisine, Sunset Yellow FCF and Brilliant Blue FCF was performed in Halwa Barfi (sweet) and Kala khatta (syrup) samples.

After plate development, Brilliant blue FCF, Sunset Yellow FCF, and Carmoisine are separated at Rf 0.34, 0.39, 0.45 respectively in the standard mixture (combination standard) and respective food samples. 

Carmoisine and Sunset Yellow FCF are detected in Halwa Barfi and Brilliant blue FCF is detected in Kala Khatta sample. Quantification was done at λ max values of (Carmoisine) 516 nm, (Sunset Yellow FCF ) 427 nm and (Brilliant Blue FCF) 630 nm. 

The amount of Carmoisine and Sunset yellow FCF added in Halwa Barfi was detected at 8.28ppm and 3.91ppm respectively. In Kala Khatta samples, the concentration of brilliant blue FCF detected is 5.25ppm. 

HPTLC Analytical Instruments Dealers

Anchrom is the sole HPTLC analytical instruments dealer of CAMAG across India. CAMAG instruments are Swiss-made and are known for their quality products. Anchrom is known for its excellent service post-sales.

Thin Layer Chromatography (TLC) is a separation technique used to separate compounds in a mixture. It is based on the principle of separation through adsorption type. HPTLC is a most versatile analytical technique which offers a great separation power using precise sample application, software-controlled chromatographic steps, chromatogram development, scanning and photo documentation. TLC plates have big absorbent particles, whereas HPTLC plates have very minute absorbent particles. HPTLC is more  sophisticated technology than TLC.

This video explains in detail the Narcotics application note on Screening Method for Identification of Forensic Drugs using CAMAG HPTLC and further confirmation by CAMAG TLC-MS Interface 2. 

High Performance Thin Layer Chromatography (HPTLC) offers identification as well as qualitative and quantitative analysis for toxic substances, rapid identification of such toxins for antidote administration. 

Forensic applications of HPTLC include detection of forged documents by analyzing the ink used, identification of narcotic substances, viscera samples, etc.

In case of any questions, please write to us at [email protected] 

This video explains in detail the Narcotics application note on Screening Method for Identification of Forensic Drugs using CAMAG HPTLC and further confirmation by CAMAG TLC-MS Interface 2. 

High Performance Thin Layer Chromatography (HPTLC) offers identification as well as qualitative and quantitative analysis for toxic substances, rapid identification of such toxins for antidote administration. 

Forensic applications of HPTLC include detection of forged documents by analyzing the ink used, identification of narcotic substances, viscera samples, etc. 

How to identify whether a reaction has been completed using TLC?

Reaction monitoring is a process that can be carried out during the research and manufacturing processes. During R & D, reaction monitoring can be used to optimize process conditions.

In the manufacturing process, reaction monitoring can help to make sure that the reaction is progressing as expected.

Thin Layer Chromatography for Reaction Monitoring:

There is a number of instrumental techniques used to monitor reactions. Thin Layer Chromatography is one of the most popular techniques used in pharma industries for reaction monitoring. 

As stated above, it can be used to monitor the progress of a reaction, identify compounds present in a given reaction mixture and determine the purity of a substance.

Three spots/bands are made on the TLC, one with the Starting material (SM), another one with the reaction mixture (RM), and another one in the middle (co-spot or cross-spot) in which you put both a solution of the SM and the reaction mixture. 

This way you can clearly visualize, after elution, that your SM actually reacted to form a new product. This is particularly important if both SM and product have very similar Rf, and it is difficult to see if you actually have a new product or just SM in the reaction mixture.

TLC and HPTLC difference:

Although, TLC is useful for reaction monitoring it has many drawbacks or limitations it is only a qualitative technique, non-GLP compliant and prone to manual errors. These limitations can be overcome by subjecting them to HPTLC analysis. 

HPTLC is a modern version of TLC wherein the principles are the same but the practice is fully automated and GLP/USP/EP compliant. HPTLC has the latest generation equipment with USP-compliant software. It is 21CFR Rule 11 compliant and designed for rapid analysis, with more output at a reduced cost.

CAMAG HPTLC in pharmaceutical analysis:

CAMAG HPTLC system has many applications in pharmaceutical analysis like identification, quantification and validation studies of API, identification of known/unknown impurities, content uniformity test (CUT studies), cleaning validation and reaction monitoring etc.

Advantages of HPTLC for drug analysis -In reaction monitoring:

A number of enhancements can be made to the original method to automate the different steps, to increase the resolution achieved with TLC and allow more accurate quantitative analysis. This can be achieved by performing High-performance Thin Layer Chromatography (HPTLC).

Unique features of HPTLC are that it is the fastest, simplest, most economical and flexible, “visible” technique which can analyse in parallel more than 100 samples. It is risk-free and multiple detections can be made without repeating chromatograms.

While doing reaction monitoring, samples from many batches can be applied simultaneously on the same plate, solvent requirement and analysis time is significantly less as a 20x20cm plate require only 20ml mobile phase and plate development time is roughly 20-30 minutes.

HPTLC Chromatography instrumentation:

CAMAG, the world’s pioneer and leader in HPTLC instrumentation has introduced a range of instruments/modules for each step in HPTLC i.e., application(CAMAG Linomat 5/ATS4), development(CAMAG ADC2), photo documentation(CAMAG Visualizer 1 / 2) and derivatization (CAMAG Derivatizer).

 One can get a very clear picture of reaction progress as known/unknown intermediates and products can be visualized and identified by Rf values, spectral evaluation and if required by using a suitable derivatizing reagent. 

Even we can go for quantitative analysis of the respective components and if unknown intermediates are observed, that can be further subjected to MS analysis by means of HPTLC-MS interface or FTIR for further identification and characterisation of an unknown compound.

Thus, HPTLC has many advantages in the reaction monitoring process whether it is a research or manufacturing process.

Reference:

Application of thin layer chromatography for qualitative analysis of gunpowder in the purpose of life prediction of ammunition, Namir H. et al., International Journal of Biosensors and Bioelectronics, 5(1):4-12, 2019.

https://chemistryhall.com/thin-layer-chromatography/