J Chrom Sci, bmad042 (2023). Standards (separated and mixed) were antazoline (ANT) and tetryzoline (TET) hydrochlorides. Samples were one commercial ophthalmic solution containing both molecules (unspiked and spiked), and aqueous humour of untreated rabbits as biological fluid, spiked with various concentrations of ANT and TET. TLC on silica gel with ethyl acetate – ethanol 1:1. Visualization under UV 254 nm. Densitometric absorbance measurement at 220 nm (20mm/s scanning speed). The hRF was 47 for TET and 71 for ANT. System suitability was verified by resolution, selectivity, capacity and absence of tailing. The method was validated for linearity range (0.2 – 18 µg/band), for precision, for reproducibility, for robustness, and for accuracy expressed as average recovery values (100 % overall mean) at different concentrations. The method was also found statistically equivalent (Student’s t-test and F-test) to the official corresponding titrimetric methods of the European Pharmacopoeia. Finally, environmental and health impacts of the methods were qualitatively and quantitatively assessed better as the other described methods, using analytical greenness (AGREE), green analytical procedure index (GAPI), national environmental method index (NEMI), and analytical eco-scale (scores based on solvents/reagents, energy consumption, occupational hazard and waste generation).

J. Planar Chromatogr. 35, 609-616 (2022). HPTLC of chlorogenic acid (1), caffeic acid (2), β-sitosterol (3) and lupeol (4) in the flowers and roots of Inula grandiflora on silica gel with toluene - ethyl - acetate - formic acid - methanol 30:30:8:3 for (1) and (2), and toluene - ethyl acetate - glacial acetic acid 145:45:1 for (3) and (4). Detection by spraying with Natural product reagent for (1) and (2) and  p- anisaldehyde - sulfuric acid reagent for (3) and (4). Quantitative determination by absorbance measurement at 366 nm for (1) and (2) and 525 nm for (3) and (4). The hRF values for (1) to (4) were 18, 82, 71 and 88, respectively. Linearity was between 125 and 625 ng/zone for (1) to (4). Intermediate precisions were below 2 % (n=9). LOD and LOQ were 32 and 97 ng/zone for (1), 39 and 110 ng/zone for (2), 47 and 124 ng/zone for (3) and 29 and 89 ng/zone for (4), respectively. Average recovery was 98.9 % for (1), 96.7 % for (2), 97.8 % for (3) and 98.8 % for (4).

J Chromatogr A 1666, 462863 (2022). Theoretical discussion on the factors determining the RF value of a given substance in a chromatographic system: A) the stationary phase (SP); B) the mobile phase (MP), the composition of which can be different from the solvent mixture prepared because of evaporation, saturation and liquid or gas adsorption effects over migration time; C) the difference of the free energies for the analyte transfer from SP to MP; D) external parameters like temperature and humidity. The universal HPTLC mixture (UHM) is a mixture of reference compounds that can be used for the system suitability test (SST) for the full RF range in all HPTLC experiments. Its composition is: thioxanthen-9-one (0.001 %), guanosine (0.05 %), phthalimide (0.2 %), 9-hydroxyfluorene, octrizole, paracetamol, sulisobenzone and thymidine (each 0.1 %), in methanol. The purpose was to study the potential of UHM to replace SST (described with specific markers in European Pharmacopoeia monographs) and to assess the quality of HPTLC results. TLC and HPTLC silica gel on different support (aluminium, glass) or with different granulometries and binders (classic, Durasil, Adamant), of the UHM, an acetonitrile extract of Abelmoschus manihot flowers (Malvaceae), a methanol extract of Sambucus canadensis flowers (Adoxaceae), and essential oils of Lavandula angustifolia, of Mentha × piperita (Lamiaceae) and of Myristica fragrans (Myristicaceae), as well as the following specific markers (standards): borneol, bornyl acetate, linalool, linalyl acetate (terpenoids), isoeugenol, isoeugenol acetate, chlorogenic acid (phenylpropanoids), gossypin (flavone), gossypetin-glucuronide, hyperoside (flavonol heterosides). Development (after 20 min plate conditioning with a saturated MgCl2 solution) with one of the following mobile phases: (MP1) toluene – ethyl acetate 19:1, especially for essential oils; (MP2) ethyl acetate – butanone – formic acid – water 5:3:1:1, especially for S. canadensis; (MP3) ethyl acetate – acetic acid – formic acid – water 100:11:11:26, especially for A. manihot. Documentation in UV 254 nm and 350 nm, and with white light (reflection + transmission), before and after derivatization. RF values were determined by scanning densitometry at 254 nm in absorption mode (for octrizole, at 366 nm in fluorescence mode with mercury lamp and optical filter K400 nm). For each HPTLC condition, intra-laboratory precision assay of UHM separation was performed (at least 5 analyses) with average RF values and 95 % prediction intervals, and calculating RF differences between pairs of UHM constituents and 95 % confidence intervals, which were max. +/-0.012 of the RF values for all UHM and markers. The sensitivity of UHM, and thus its usefulness as generic SST was demonstrated by repeating the HPTLC experiments with modifying by 10 % the quantity of one of the solvent each time. There were always significant changes in RF values of UHM components and/or in RF differences between pairs of UHM bands; it was often but no always the case with the official specific markers. UHM underwent also significant changes (although less than A. manihot extract) when several silica gel phases were compared under the same HPTLC conditions. This property is crucial to verify the right stationary phase before doing any RF correlations, and could make UHM a universal tool to identify discrepancies between different analyses. Finally, the use of UHM for a computer-supported evaluation of HPTLC results was discussed, either for zone identification and RF corrections (within confidence intervals), or for correlations of entire fingerprints as first step to implement machine learning algorithms.

J Chromatogr A 1638, 461830 (2021). The purpose was to find the first universal HPTLC mixture (UHM), a mixture of reference compounds that could be used for the system suitability test (SST) for the full RF range in all HPTLC experiments.
(Part 1) UHM composition: First, 56 organic molecules, detectable without derivatization, were tested on HPTLC silica gel with 20 different mobile phases (MP) belonging to different Snyder’s selectivity groups and with several polarity indices. Visualization under UV 254 nm and 366 nm. Densitometry scanning at 254 nm in absorption mode, and at 366 nm in a fluorescence mode (mercury lamp 366 nm, with wavelength filter <400 nm). For selected bands, spectra were recorded in absorbance-reflectance mode (wavelength range 190 – 450 nm, deuterium and tungsten lamp). This procedure allowed 8 molecules to be selected for their better spot resolution and for their specific RF values (at least 3 different values distributed throughout the full RF range for each MP). The final composition of UHM was: thioxanthen-9-one (0.001 %), guanosine (0.05 %), phthalimide (0.2 %), 9-hydroxyfluorene, octrizole, paracetamol, sulisobenzone and thymidine (each 0.1 %), in methanol.
(Part 2) UHM validation: Afterwards, UHM was submitted again to a panel of HPTLC assays with always two MP: (A) toluene – methanol – diethylamine 8:1:1; (B) ethyl acetate – formic acid – water 15:1:1; and for each MP, the means, standard deviation and 95 % confidence intervals of the RF values were calculated. (a) UHM was validated for intermediate intra-laboratory precision, as well as for inter-laboratory reproducibility, with ΔRF 0.045. (b) The capacity of UHM to detect small variations was demonstrated by significant changes in at least some RF values, when separation was deliberately performed at different levels of relative humidity (0 %, 33 %, 75 %, 100 %), or with smaller humidity variations (7 % compared to 0–5 %, and 49 % compared to 33 %), or when performing vs. omitting the 10min chamber pre-saturation, or when modifying the MP (+/-10% of one solvent at each time). These response characteristics (the opposite of robustness) made UHM a powerful tool for SST. (c) Finally, UHM stability was studied with UHM aliquots under several storage conditions (-78 °C, -20 °C, 4 °C, room temperature, 45 °C; or 40 °C with 75 % relative humidity) and durations (2 weeks or 2 months). The densitometric peak profiles at 254 nm were compared to those of the fresh compounds, qualitatively (RF value, UV spectrum) and quantitatively (peak area). UHM was stable at room temperature or below, for 2 months (at higher temperature, guanosine, phthalimide and paracetamol degraded).

Plant Med 88(2), 163-178 (2022). TLC was used to verify the purity of acetoxychavicol acetate (a phenylpropanoid) isolated through column chromatography from a hexane extract of Alpinia galanga rhizomes (Zingiberaceae). TLC on silica gel with n-hexane – ethyl acetate 17:3, evaluation under UV 254 nm.

J. Ethnopharmacol. 289, 115035 (2022). HPTLC of stigmasterol and some polyphenolic compounds in the rhizome of Cyperus tegetum on silica gel with chloroform - ethyl acetate - formic acid 5:4:1 and toluene - methanol 9:1. Detection by spraying with p-anisaldehyde reagent in a cold solution of methanol - glacial acetic acid - phuric acid 17:2:1, followed by heating at 100 - 105 °C for 5-10 min.

Chinese Medicine 16, 98 (2021). Preparative TLC on silica gel for the isolation of bisbakuchiol N (a terpenophenolic) from a cyclohexane extract of Psoralea corylifolia (= Cullen corylifolia, Fabaceae) mature fruits, after fractionation on silica gel, cyclodextrane and reverse-phase columns. Mobile phase was petroleum ether – chloroform 10:1. Derivatization with sulfuric acid (10 % in ethanol – water, 19:1).

Food Chem. 133992 (2023). HPTLC of kiwi peel extracts on silica gel with ethyl acetate - formic acid - acetic acid - water 100:11:11:26. Detection of phenolics and tanning agents by heating the plate at 100 °C for 2 min, followed by dipping into fast blue B (140 mg fast blue salt B in a mixture of 140 mL methanol, 10 mL water, and 50 mL dichloromethane). Detection of flavanols, phenols and further natural compounds by dipping into a NPA solution (1 g 2-aminoethyl diphenylborinate in 200 mL of ethyl acetate), followed by drying, detection under UV light at 366 nm, followed by dipping into the anisaldehyde reagent (0.5 mL anisaldehyde, 10 mL acetic acid, 85 mL methanol, and 10 mL sulphuric acid), followed by heating at 100 °C for 5 min. Antioxidants were detected by dipping into a DPPH solution (0.4 g 2,2-diphenyl-1-picrylhydrazyl in 200 mL of methanol), followed by incubation in dark for 30 min.