High-performance Thin-layer Chromatography Fingerprinting Analysis Research Articles

Chemotaxonomy of Southeast Asian Peperomia (Piperaceae) Using High-Performance Thin-Layer Chromatography Colour Scale Fingerprint Imaging and Gas Chromatography–Mass Spectrometry

The morphological characters of Southeast Asia’s indigenous Peperomia species are very similar, especially in their flower structures. The flowers are simple, hermaphrodite and lack a perianth. Therefore, many species are hard to distinguish using morphological characters alone. Here, we apply chemometric data for species identification and classification, gathered using multiwavelength detection combined with the colour scale High-Performance Thin-Layer Chromatography (HPTLC) fingerprinting procedure and chemical compounds determined by Gas Chromatography–Mass Spectrometry (GC-MS). Fourteen taxa were investigated using hexane, ethyl acetate and ethanol solvent extractions. Principal component analysis (PCA) and hierarchical cluster analysis (HCA) were used with the colour scale fingerprints to classify the Peperomia species. The PCA and HCA using the chromatogram profile from hexane divided the taxa into six groups compared to the profile from ethyl acetate and ethanol, which each detected seven groups. The chromatogram from the combined dataset of all three solvents can differentiate all the species. The GC-MS data detected a total of 40 compounds from the hexane extract, and these differed among Peperomia species. This approach based on HPTLC fingerprinting and GC-MS analysis can therefore be used as a tool for authentication and identification studies of Peperomia species.

Pharmacognostic standardization and HPTLC fingerprinting analysis of Sahacharadi Kashayam: a classical ayurvedic polyherbal formulation

Multiherbal compositions are widely practiced in ayurvedic medical system, and ka??yam (decoction) is one such dose. Sahacharadi Kashayam is a traditional ayurvedic polyherbal medicine recommended by ayurvedic doctors for chronic and acute back pains, lower limb disorders, and conditions that mainly affect the nutrient level in the blood and the bone. The aim of this study was to develop pharmacognostic parameters for the standardization and quality evaluation of the Sahacharadi Kashayam. In-house polyherbal formulation was prepared as per the previously prescribed method, and physicochemical parameters were evaluated as per standard protocol and showed loss on drying at 99.31%, total ash content at 4.911 ± 0.120%, water-soluble ash at 3.022, and acid-insoluble ash at 0.489. Total phenolic content was estimated with FC reagent and expressed as gallic acid equivalent (9.95 + 0.004 mg/g). High-performance thin-layer chromatography (HPTLC) fingerprint profiling was obtained, and the quantity of total tannins was estimated as tannic acid equivalent using HPLC (0.47%). The results obtained from the study could be utilized as a reference for the quality control and quality assurance of Sahacharadi Kashayam.

Phytochemical, Physicochemical and HPTLC Analysis of Siddha Herbal Formulation Muppirandai Chooranam

Aim: The aim of the study was to evaluate the Phytochemical, Physicochemical, HPTLC analysis of Siddha herbal formulation Muppirandai Chooranam (MRC), indicated for various ailments including menorrhagia.
 Study design
 Place of study: The Phytochemical and Physicochemical analysis were conducted at the Tamilnadu Dr. MGR Medical University, located at no.69, Anna salai, Guindy, Chennai-32. The high-performance thin layer chromatography (HPTLC) was carried out at Noble research solutions, Kolathur, Chennai -99.
 Methodology: Siddha formulation Muppirandai Chooranam was prepared as per good manufacturing practices (GMP) guidelines and the phytochemical analysis, physicochemical analysis were carried out at the Tamilnadu Dr. MGR Medical University, located at no.69, Anna salai, Guindy, Chennai-32. As per PLIM guidelines (Pharmacopeia Laboratory Of Indian Medicine) in accordance with the guidelines established by AYUSH (Ayurveda, Yoga, Unani, Siddha, Homoeopathy), the governing body for traditional health systems in India. The HPTLC analysis was conducted at Noble Research Solutions, Kolathur, Chennai -99.
 Results: The Phytochemical screening of MRC shown the presence of Alkaloids, Carbohydrates, Saponins, Flavanoids, Diterpenes, Gum and Mucilage. The physicochemical analysis of MRC revealed that it had a loss on drying of 0.42% at 105°C, a total ash content of 23.3%, an acid-insoluble ash content of 3.02%, a water-soluble ash content of 2.30%, and a water-soluble extract content of 17.67%, and alcohol soluble extract content of 5.23%.HPTLC finger printing analysis of the sample reveals the presence of five prominent peaks corresponds to the presence of five versatile phytocomponents present within it. Rf value of the peaks ranges from 0.02 to 0.37.
 Conclusion: From the findings, it is concluded that it shown the compendious understanding of presence of Phytochemical components, Physicochemical characteristics, and HPTLC analysis of MRC and it is competent to assess the quality profile of Muppirandai Chooranam as a reference standard for the development of the standardised pharmaceutical product.

High-performance Thin-layer Chromatography Fingerprinting Analysis of Punarnavadi Kwatha

Introduction: Punarnavadi kwatha is one among the commonly used medicine in Ayurvedic practice, mentioned in Chakradatta in sopha chikitsa. Main indications of Punarnavadi kwatha are Sarvanga sopha (generalised oedema), Udara (ascites), Kasa (cough), Soolam (colicky pain), and Swasa (dyspnoea) associated with Pandu (anaemia). For the purpose of quality assurance and uniformity, scientific verification of Ayurvedic medications is important. Usage of modern analytical techniques helps in the proper authentication of medicines. In the present study, highperformance thin-layer chromatography (HPTLC) fingerprinting of Punarnavadi kwatha aimed to assess the different phytoconstituents. Materials and Methods: Methanolic extract of Punarnavadi kwatha was subjected to HPTLC fingerprinting using the solvent system toluene:ethyl acetate:formic acid (6:3:0.5). Toluene is a least polar solvent, ethyl acetate is mid polar solvent and formic acid is highly polar so in this solvent system, maximum separation of compounds took place. Results and Discussion: The number of peaks obtained at visualization 254 nm was 8 and total area of compounds in is 30309.5. Visualization at 366 nm, number of peaks was 12 and total area is 42473.2. Conclusion: Thus, the study revealed that HPTLC fingerprinting of formulations can be used in the quality control of ayurvedic medicaments. The maximum separation of compounds occurred at this solvent system.

Molecular docking analysis of estrogen receptor binding phytocomponents identified from the ethyl acetate extract of Salicornia herbacea (L).

It is of interest to evaluate the secondary metabolites using high performance thin layer chromatography (HPTLC) finger printing and Gas chromatography-Mass spectroscopy (GC-MS) in S. herbaceaextract. The powdered plant material extracted using different solvents were used for the qualitative analysis of alkaloids, flavonoids, terpenoids and saponins followed by HPTLC finger printing and GC-MS analysis. The components identified in the GC-MS were docked with estrogen receptor (ER) to identify the binding specificity of isolated compounds. The ethyl acetate extract of S. herbaceashowed the presence of high number of secondary metabolites when compared to other solvent system. The qualitative analysis of the plant material also showed the presence of carbohydrates, protein, amino acid, phenol, flavonoids, terpenoids, glycosides, saponins and steroids. The HPTLC finger printing analysis revealed the existence of alkaloid, flavonoid, terpenoid and saponin compounds and GC-MS. GC-MS was performed to identify the phytocomponents constituents in the extract. 8 phytocompounds were identified to analyse binding with ER. The binding affinity score (-6.8 kcal/mol) and interacting ER residues (28) the phyto compound di-n-octyl phthalate showed best docking score with ER α than the standard drugs lasofoxifene, and 4-hydroxytamoxifen. The binding affinity and number of interacting ER residues was -6.9 kcal/mol; 10 and -6.2; 11, respectively. The results identified the presence of ER antagonist in S. herbaceaand warrants further investigation to explore for treating ER regulated diseases.

Assessment of Biological Activities of Fungal Endophytes Derived Bioactive Compounds Isolated from Amoora rohituka.

Fungal endophytes have remarkable potential to produce bioactive compounds with numerous pharmacological significance that are used in various disease management and human welfare. In the current study, a total of eight fungal endophytes were isolated from the leaf tissue of Amoora rohituka, and out of which ethyl acetate (EA) extract of Penicillium oxalicum was found to exhibit potential antioxidant activity against DPPH, nitric oxide, superoxide anion and hydroxyl free radicals with EC50 values of 178.30 ± 1.446, 75.79 ± 0.692, 169.28 ± 0.402 and 126.12 ± 0.636 µg/mL, respectively. The significant antioxidant activity of EA extract of P. oxalicum is validated through highest phenolic and flavonoid content, and the presence of unique bioactive components observed through high-performance thin layer chromatography (HPTLC) fingerprinting. Moreover, EA extract of P. oxalicum also displayed substantial anti-proliferative activity with IC50 values of 56.81 ± 0.617, 37.24 ± 1.26 and 260.627 ± 5.415 µg/mL against three cancer cells HuT-78, MDA-MB-231 and MCF-7, respectively. Furthermore, comparative HPTLC fingerprint analysis and antioxidant activity of P. oxalicum revealed that fungal endophyte P. oxalicum produces bioactive compounds in a host-dependent manner. Therefore, the present study signifies that fungal endophyte P. oxalicum associated with the leaf of A. rohituka could be a potential source of bioactive compounds with antioxidant and anticancer activity.

Coriandrum sativum attenuates microglia mediated neuroinflammation and MPTP-induced behavioral and oxidative changes in Parkinson's disease mouse model.

Coriandrum sativum Linn. (family: Umbelliferae; C. sativum), is a potential herb widely used as a spice and traditional medicine. In the present work, the effects of C. sativum fruit extract (CSE), against lipopolysaccharide (LPS)-stimulated BV-2 microglia-mediated neuroinflammation in vitro and 1-methyl-4 phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) animal model in vivo was investigated. LPS-stimulated increase in nitric oxide (NO), inducible NO synthase, cyclooxygenase-2, interleukin-6 and tumor necrosis factor-alpha were significantly (p < 0.05 ~ p < 0.001) inhibited by CSE (25, 50 and 100 μg/mL) in BV-2 microglial cells. Further, CSE inhibited the LPS-induced nuclear factor of kappa-beta activation and IκB-α phosphorylation in BV-2 microglia. In vivo studies, CSE (100, 200 and 300 mg/kg) ameliorated the MPTP (25 mg/kg, i.p.)-induced changes in locomotor, cognitive and behavior functions evaluated by rotarod, passive avoidance and open field test significantly (p < 0.05 ~ p < 0.001). The MPTP-induced changes in brain oxidative enzyme levels such as superoxide dismutase, catalase, and lipid peroxidation were significantly (p < 0.01 and p < 0.001 at 200 and 300 mg/kg, respectively) restored with CSE treatment. High-performance thin-layer chromatography fingerprinting analysis of CSE exhibited several distinctive peaks with quercetin and kaempferol-3O-glucoside as identifiable compounds. In conclusion, our study indicated that CSE attenuated neuroinflammatory processes in LPS-stimulated microglia in vitro and restored the MPTP-induced behavioral deficits and brain oxidative enzyme status in vivo proving its therapeutic potential in the treatment of neuroinflammatory and oxidative stress-mediated neurodegeneration seen in PD.

HPTLC fingerprinting, photopigments, and nutrient analysis of Selinum tenuifolium along the altitudinal gradient

This study aimed to explore phenolic acids, nutrient status, and content of photopigments and their role in the production of secondary metabolites in Selinum tenuifolium collected from different altitudes (2150 m; 2593 m; 3178 m, asl) in Chopta region of Uttarakhand, India. High-performance thin-layer chromatography (HPTLC) fingerprinting, nutrient analysis, and chlorophyll (Chl) content were assessed in various plant parts (leaves, stem, and roots) of S. tenuifolium. HPTLC analysis revealed a number of bands in the methanolic extracts of S. tenuifolium of which two with retention factor (Rf) values, 0.36 pertaining to gallic acid (GA) and 0.58 relating to ferulic acid (FA) indicating their presence in all three plant parts of S. tenuifolium. The elemental analysis (Fe2+, Zn2+, Na + K+ and, Ca2+) of different plant parts reflected the nutrient status of the plant, the total amount of nutrients being highest in stems of plants growing at 3178 m and lowest in roots of plants growing at 2150 m. The photopigments analysis of leaves of S. tenuifolium showed variations in their contents, being maximum at the highest altitude (3178 m) and minimum at the lowest altitude (2150 m). The correlation matrix suggested a strong positive correlation between elements like Na+, K+and Ca+2 with that of carotenoids, and Fe+2 and Zn+2 with that of chlorophyll a + b (Chl a + b). On the other hand, a negative correlation existed between photopigments and phenolic acids probably because the plant enjoys a good nutrient status, and therefore secondary metabolites that are usually triggered during stress need not be produced in high quantities.

And validation of HPTLC method for the simultaneous estimation of ascorbic acid and gallic acid in amla juice preparation.

The aim of this study was to asses a simple, selective, precise, and reproducible high performance thin-layer chromatography (HPTLC) method for the simultaneous estimation analysis of ascorbic acid (AA) and gallic acid (GA) in amla juice preparation. The aluminium-based pre-coated TLC plates (Silica gel G 60F 254) were used for the HPTLC fingerprinting analysis. The chromatograms of samples were developed in twin trough glass chamber pre-saturated with mobile phase (toluene: ethyl acetate: methanol: formic acid; 3:3:2:1, v/v/v/v) at room temperature (25 ± 2°C). The densitometric analysis was carried out in absorbance mode at 254 nm. The optimized mobile phase showed compact spots of AA and GA at 0.59 and 0.86 Rf respectively. The linear regression analysis data for the calibration plots of AA and GA showed good linearity (r2= 0.992 and 0.996 respectively) with respect to peak area in the range of 200-1400 ng/spot. The method was validated as per International Conference on Harmonization (ICH) guidelines. The limits of detection and quantification (40 and 140 ng/spot, respectively) were also established. The proposed method has shown the excellent recovery (98.97–99.89%), which supports the suitability of the method for the analysis of AA and GA in the amla juice and other preparations containing these ingredients. Keywords: Amla juice, Ascorbic acid, Gallic acid, HPTLC, ICH guidelines, Validation.

ISOLATION AND HIGH-PERFORMANCE THIN-LAYER CHROMATOGRAPHY ANALYSIS OF CONDITIONAL AMINO ACIDS FROM THE FRESH LEAVES OF ALTERNANTHERA SESSILIS

Objective: The objective of the study was to isolate the conditional amino acids present in the fresh leaves extracts of Alternanthera sessilis and compare the isolated amino acids with amino acid standards.&#x0D; Method: The fresh plant material was used for extraction. For the extracted sample solution, paper chromatography and high-performance thin-layer chromatography (HPTLC) fingerprinting analysis were then carried out.&#x0D; Results: Since A. sessilis is rich in protein, various amino acids have been isolated in aggregates and paper chromatography has been performed. Amino acids were separated and identified using trial and error method. Cysteine, glycine, aspartic acid, and proline are the standard amino acids used for comparing with the isolated amino acids by HPTLC fingerprint analysis, which gives an idea for the authentication of the plant, its constituents and also provides a parameter for quality control.&#x0D; Conclusion: The results scientifically validate the use of A. sessilis in the traditional medicine and also as a food.

Erratum: Comparison of Granules for Prescription and Classical Decoctions by High-performance Thin-layer Chromatography-fingerprint Analysis

Erratum: Comparison of Granules for Prescription and Classical Decoctions by High-performance Thin-layer Chromatography-fingerprint Analysis

Preliminary Phytochemical Screening and HPTLC Fingerprinting of Extracts of Thuja occidentalis

Objective: To establish the fingerprint profile of Thuja occidentalis using high performance thin layer chromatography (HPTLC) technique. Methods: Preliminary phytochemical screening was done and HPTLC studies were carried out. CAMAG HPTLC system equipped with Linomat V applicator, TLC scanner 3 and visionCATS software were used. Results: Preliminary phytochemical screening of the extract showed the presence of alkaloids, triterpenes, tannins, saponins, glycosides, phenolic compounds and flavonoids. HPTLC finger printing of methanol extract of plant with Rf values at 0.5. Conclusions: It can be concluded that HPTLC fingerprint analysis of extract of Thuja occidentalis can be used as a diagnostic tool for the correct identification of the plant and it is useful as a phytochemical marker.

DETERMINATION OF PHYSICOCHEMICAL PROPERTIES, ANTIOXIDANT CONSTITUENTS BY HIGH-PERFORMANCE THIN-LAYER CHROMATOGRAPHY FINGERPRINTING, AND ANTIOXIDANT ACTIVITY OF CUCURBITA MAXIMA SEEDS

Objective: The present study is designed to evaluate preliminary phytochemical constituents, physiochemical properties evaluation, free radical scavenging activity, and high-performance thin-layer chromatography (HPTLC) fingerprinting analysis of Cucurbita maxima seed oil.Methods: Total phenol content, total flavonoid content, total tannin content, 1, 1-diphenyl, 2-picryl-hydrazyl (DPPH) radical scavenging method, and HPTLC fingerprinting analysis for compounds responsible for antioxidant activity were used.Result: Phytochemical screening shows the presence of constituents such as alkaloids, glycosides, carbohydrates, fixed oil and fats, phytosterols, saponin, phenolic compounds, tannins, proteins, amino acids, gums, and mucilage. Physicochemical studies show that the oil contains an acid value, saponification value, iodine value of 9.53 mg KOH/g, 183 mg KOH/g, and 116.51 mg I2/100 g, respectively. Total phenol content was 3.9498±3.2 mg gallic acid equivalent/g, total flavonoid content was 56.115±4.6 mg rutin equivalent/g, and total tannin content was 19.4±0.40 mg tannic acid equivalent/g. The radical scavenging activity by DPPH Model shows that the methanolic extract has maximum antioxidant activity with IC50 value of 238.31. HPTLC fingerprinting also proofs the presence of phenol and flavonoid compounds in the seed oil.Conclusion: The results in the paper show that the C. maxima seed oil is an natural antioxidant and can be used for the treatment of various disorders.

HPTLC Fingerprinting Analysis of Tannin Profile on Canthium coromandelicum and Flueggea leucopyrus willd.

The study of chemical composition of medicinal plants or drugs derived from plants is said to be Phytochemistry. Tannins are polyphenolic compounds; they are identified as better antiseptic due to the presence of phenolic group. The main objective of this study was to progress the High Performance Thin Layer Chromatography (HPTLC) fingerprinting of tannin profile on methanolic extracts of leaves and stem of Canthium coromandelicum and Flueggea leucopyrus willd aerial parts. The HPTLC fingerprinting analysis was carried for tannin using CAMAG LINOMAT 5 instrument which revealed that the methanolic extracts of Canthium coromandelicum leaves, stem and aerial parts Flueggea leucopyrus showed the presence of 10, 8 and 7 peaks with Rf values in the range of 0.04 to 0.97 respectively. Thus, the HPTLC methods developed in this study against the methanolic extract of selected plants provide useful and simple tools for the quality assessment and identification of phytochemical marker compounds. By isolating and identifying the biomarker compounds, new drugs can be synthesised to treat various ailments.

Comparison of granules for prescription and classical decoctions by high-performance thin-layer chromatography-fingerprint analysis

Objective: The so-called granules for prescription have been developed about 20 years ago as a new form of modernizing and simplification of the classical decoction common in Traditional Chinese Medicine (TCM) practice. Due to actual problems in Germany/Europe, which are caused by the lack of quality monographs and judicial classification of granules for prescription, the aim of the study was a comparison of the chemical composition of commercial granules versus decoctions. Taking an example, decoctions, commercial granules, and organic extracts of two well-established TCM herbal drugs, Scrophulariae Radix and Xanthii Fructus, were examined in their specific composition. Methods: Using high-performance thin-layer chromatography (HPTLC) for fingerprint analysis of different batches of herbal drugs and samples from various suppliers of Xanthii Fructus and Scrophulariae Radix were critically examined. The decoctions were prepared according to traditional rules, while the granules were dissolved in water in accordance with actual regulations. Furthermore, organic extracts of the plant material were examined and compared with aqueous extracts. Results: It could be demonstrated, that in some cases, there are remarkable differences in the specific composition between granules from different suppliers, the classical aqueous decoction and the organic extract used for the HPTLC fingerprinting. On the other hand, few examples exist for good comparability of decoctions and commercial granules. Conclusion: After critical evaluation of the above results, it can be questioned, if there is a so-called phytoequivalence between decoctions and commercial granules for prescription used in TCM practice.

High performance thin layer chromatography fingerprint analysis of guava (Psidium guajava) leaves

High-performance thin layer chromatography (HPTLC) fingerprint analysis is commonly used for quality control of medicinal plants in term of identification and authentication. In this study, we have been developed HPTLC fingerprint analysis for identification of guava (Psidium guajava) leaves raw material. A mixture of chloroform, acetone, and formic acid in the ratio 10:2:1 was used as the optimum mobile phase in HPTLC silica plate and with 13 bands were detected. As reference marker we chose gallic acid (Rf = 0.21) and catechin (Rf = 0.11). The two compound were detected as pale black bands at 366 nm after derivatization with sulfuric acid 10% v/v (in methanol) reagent. Validation of the method was met within validation criteria, so the developed method could be used for quality control of guava leaves.

An endangered medicinal plant, Plumbago zeylanica L. and its substitute, Dyerophytum indicum (Gibbs ex Wight) Kuntze: A comparative HPTLC, HPLC study, and screening of antioxidant activity

Plumbago zeylanica L. (PZ) is a significant medicinal plant in Ayurveda, so is used for the treatment of various disorders. The main active part of this plant is the root, and due to its inherent uses, it has been exploited hence becoming an endangered species. Dyerophytum indicum (Gibbs ex Wight) Kuntze (DI), mainly considered as a substitute of PZ, is also an important folklore medicine, used in many health problems. Both plants are much similar in their physical as well as chemical properties. However, an effective validation is required before declaration of substitution. In the present study, quantitative and qualitative estimations were performed on both plants with the help of modern analytical techniques. Simultaneous high-performance liquid chromatography (HPLC) and high-performance thin-layer chromatography (HPTLC) and quantitative determinations of β-sitosterol have been performed for comparing both plants, i.e., PZ and DI. HPTLC fingerprinting analysis was also performed comparatively in diffe...

HPTLC Fingerprinting of Swertia chirayita (Roxb. ex Fleming) Karsten from High Altitude Area of Western Himalaya

To establish the fingerprint profile of Swertia chirayita (Roxb. ex Fleming) Karsten from the high altitude area of the western Himalaya using high performance thin layer chromatography (HPTLC) technique. Dried and powdered plant material of S. chirayita was successively extracted with petroleum ether, ethyl acetate and methanol in the order of increasing polarity. The extracts so obtained were subjected to qualitative phytochemical screening and based on the presence and/or absence of phytochemical constituents, HPTLC fingerprinting analysis of ethyl acetate and methanol extract were carried out using CAMAG HPTLC system. Preliminary phytochemical screening of the extracts showed the presence of alkaloids, terpenoids, phenolics, flavonoids, carbohydrates, glycosides, tannins, saponins and lipids in various extracts. Methanol and ethyl acetate extracts contained the maximum number of phytochemicals. HPTLC phytochemical fingerprinting led to the detection of altogether 21 compounds in ethyl acetate and methanol extracts. However, greater numbers of compounds were detected in methanol extracts as compared to ethyl acetate extract. The chromatographic fingerprinting revealed the presence and variation in the phytochemical constituents with change in the polarity of the solvents. Further studies are required for the quantification of the compounds in various extracts and evaluation of their medicinal activities.

HPTLC Fingerprinting Method to Distinguish Total Extract of Caralluma fimbriata from the Modified Extracts of Caralluma fimbriata

To establish the fingerprint profile of total extract of Caralluma fimbriata using high performance thin layer chromatography (HPTLC) technique. HPTLC studies were carried out. CAMAG HPTLC system equipped with Linomat V applicator, TLC scanner 3, Reprostar 3 and WIN CATS-4 software were used. HPTLC finger printing of Caralluma fimbriata leaf extract revealed that the chromatogram obtained with the reference solution shows in the middle part a Dark blue zone [Slimalumaside A] at RF 0.33 and in the upper part a Blue zone [Slimalumaside B] at RF 0.94 was observed. It can be concluded that HPTLC fingerprint analysis of total extract of Caralluma fimbriata can be used as a diagnostic tool for the appropriate identification various extracts.

Pattern recognition methods and multivariate image analysis in HPTLC fingerprinting of propolis extracts

High‐performance thin‐layer chromatography (HPTLC) combined with image analysis and pattern recognition methods were used for fingerprinting and classification of 52 propolis samples collected from Serbia and one sample from Croatia. Modern thin‐layer chromatography equipment in combination with software for image processing and warping was applied for fingerprinting and data acquisition. The three mostly used chemometric techniques for classification, principal component analysis, cluster analysis and partial least square‐discriminant analysis, in combination with simple and fast HPTLC method for fingerprint analysis of propolis, were performed in order to favor and encourage their use in planar chromatography. HPTLC fingerprint analysis of propolis was for the first time performed on amino silica plates. All studied propolis samples have been classified in two major types, orange and blue, supporting the idea of existence of two types of European propolis. Signals at specific RF values responsible for classification of studied extracts have also been isolated and underlying compounds targeted for further investigation. Copyright © 2014 John Wiley &amp; Sons, Ltd.