High-Performance Thin
Layer Chromatography (HPTLC) is a modern and advanced form of traditional TLC
with higher resolution, sensitivity, and reproducibility. It is particularly
useful in the analysis of complex mixtures such as natural products,
including plant extracts, herbal
medicines, essential oils, and nutraceuticals.
Applications of HPTLC in Natural Product Studies
1. Qualitative Analysis:
o Identification of plant species
or raw materials.
o Fingerprinting of herbal
extracts to ensure authenticity and detect adulteration.
o Comparison with reference
standards or pharmacopeial standards.
2. Quantitative Analysis:
o Determination of marker
compounds (e.g., alkaloids, flavonoids, phenolics).
o Estimation of active
constituents in herbal formulations.
o Standardization of plant-based
products.
3. Quality Control:
o Routine testing in the
manufacturing of herbal drugs.
o Monitoring of raw material
consistency and batch-to-batch variation.
o Stability testing over time.
4. Phytochemical Screening:
o Detection of various classes of
natural products like saponins, terpenoids, alkaloids, glycosides, etc.
5. Chemotaxonomic Studies:
o Differentiation of plant species
based on chemical profile.
o Classification of plants based
on chemical constituents.
6. Bioautography:
o Integration with biological
assays (e.g., antimicrobial, antioxidant) on the chromatographic plate.
7. Metabolomics & Dereplication:
o Rapid profiling and comparison
of metabolite compositions in extracts.
o Early-stage dereplication in
natural product discovery.
Specifications Needed for HPTLC in Natural Product Studies
To effectively use HPTLC in natural product
research, certain technical
and methodological specifications are
needed:
1. Stationary
Phase:
Pre-coated
HPTLC plates (e.g., silica gel 60 F254).
Particle
size: ~5–7 µm for better resolution.
2. Sample
Application:
Automated
sample applicator (e.g., CAMAG Linomat or Automatic TLC Sampler).
Narrow,
consistent bands or spots for reproducibility.
3. Development
Chamber:
Automated
development chamber or twin-trough glass chamber.
Controlled
humidity and saturation time.
4. Mobile
Phase (Solvent System):
Optimized
based on polarity of compounds.
Should
allow good separation and resolution of targeted natural products.
5. Detection
System:
UV/Visible
light at 254 nm and/or 366 nm.
Derivatization
reagents for visualization (e.g., anisaldehyde, vanillin, Dragendorff’s).
6. Documentation
and Analysis:
High-resolution
digital documentation system.
Densitometric
scanning (e.g., CAMAG TLC Scanner) for quantification.
Software
(e.g., winCATS) for data analysis, peak integration, and comparison.
7. Reference
Standards:
Authentic
markers or standards for identification and quantification.
Use
of pharmacopoeial or published standards.
8. Validation
Parameters:
Precision,
accuracy, linearity, specificity, LOD/LOQ, robustness (according to ICH or AOAC
guidelines).
Minimum Specifications for HPTLC
Microprocessor
based fully automated Sample Applicator: for samples to be applied as “dot” or
“line” on HPTLC/TLC plates; Dosing and Filling Syringe; Fillport; memory for
multiple methods; Micro-preparative application; flexible delivery rate;
Resolution >=2000 steps/ul; Gas pressure : >2.5 bar and < 10 bar .
Integrated Windows based PC-Software (WIN) compatible with Applicator: to document, control and manage steps in HPLC analysis including application, development, scanning and photorecording; should be upgradable.
PC Controlled Densitometer with Scanning facility for automatic spectrum scanning, identification/purity check and quantitative measurement; Linear and meander scanning; Software for scanner; automatic quantitave measurement of absorbance & fluorescence; Deuterium, Halogen, Mercury as Light sources; High sensitivity and High resolution photomultiplier; Dual wavelength and Multiwavelength spectrum recording; Spot Optimisation.
3-D display of Chromatograms; Linear and Polynominal Calibration modes; Automatic and manual peak integration; Measurements enabled by user defined methods and user-friendly software; system should be upgradable.
Chambers for the chromatographic development of TLC/HPTLC plates: low solvent consumption; economical, reproducible.
UV-Box : for
UV detection in thin-layer chromatography.
PC & Printer specifications required for TLC System Operating System Windows 10 or 11 Professional Intel Dual Core Processor or Higher Version, 8 GB RAM, 500 GB HDD; Compatible Colour Printer; compatible UPS.
About HPTLC
HPTLC is a powerful tool in natural product
studies, particularly for quality
control, authentication,
and phytochemical profiling. When properly implemented with validated specifications,
it offers a reliable, cost-effective, and high-throughput platform for
analyzing complex herbal and plant-derived materials.
You may like to read more:
1. List of All SOPs and Documents for Laboratory Instruments and Equipment
2. List of All SOPs and Documents for Chemistry Laboratory
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