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).

J Chromatogr A 1638, 461597 (2021). Samples were Isatis tinctoria (= I. indigotica) root extracts (Brassicaceae) and their fractions. Standards were oseltamivir acid (OA), a neuraminidase (NA) inhibitor; pinoresinol (PR, a lignan), β-sitosterol (SS, a sterol), and dihydro-neoascorbigen (DHNA, an alkaloid). HPTLC / TLC on silica gel with (1) petroleum ether – ethyl acetate – acetic acid 48:8:1 for petroleum ether extracts and SS, or 30:40:1 for ethyl acetate extracts, or 10:30:1 for PR; (2) with toluene – ethyl acetate – methanol – formic acid 16:3:1:2 or 10:4:1:2 also for ethyl acetate extracts and DHNA; (3) with n-butanol – acetic acid – water 25:4:3 for butanol extracts. OA was applied but not developed. RP-18, polyamide, cellulose, alumina layers were tested, but the resolution was lower. Derivatization by spraying with sulfuric acid (10 % in ethanol). Enzymatic assay by immersion of the plates into neuraminidase solution (6 U/mL), followed by 1 h incubation at 37 °C and by immersion into chromogenic substrate solution (1.75 mM 5-bromo-4-chloro-3-indolyl-α-D-N-acetylneuraminic acid). After 5 min, NA inhibitors were seen as white zones on blue background. The experiment was previously improved for the following parameters: incubation times, substrate and enzyme concentrations, followed by statistical evaluation and calculations using Box-Behnken design. Quantification by absorbance measurement (detection wavelength 605 nm, reference wavelength 420 nm). In optimal conditions, OA had LOD 300 ng/zone. Zones of interest on underivatized plates were directly submitted to MS, using EFISI (electrostatic-field-induced spray ionisation), as follows. Chromatograms were immersed 1–3 s into dimethicone – n-hexane 1:1 to form a hydrophobic film, and dried 30 min at room temperature; on the analyte spot, a hydrophilic droplet was formed with 5 µL methanol – water 1:1, extracting the analyte from the layer; the analyte was further attracted through a capillary tube (3–4 cm long, made of non-deactivated fused silica) under a strong electrostatic field, into the in-let orifice of the triple-quadrupole ­– linear ion-trap MS (induction voltage 4 kV; capillary voltage 40 V; tube lens voltage 100 V; capillary temperature 200 °C). Full-scan spectra were recorded in m/z range 50 – 1000, helium was used for collision-induced dissociation. 11 active compounds were identified in the extract: SS, 6 alkaloids (including cycloanthranilylproline, DHNA, hydroxy-indirubin, isatindigodiphindoside, isatindinoline A and), 3 lignans (including PR and isolariciresinol), 1 fatty acid (trihydroxy-octadecenoic acid).

Braz. J. Pharm. Sci. 58, e18691 (2022). HPTLC of paracetamol (1), diclofenac sodium (2), ibuprofen (3), and indomethacin (4) in wastewater effluents on silica gel with n-hexane - ethyl acetate - acetic acid 12:7:1. Quantitative determination by absorbance measurement at 254 nm. The hRF values for (1) to (4) were 18, 56, 69 and 44, respectively. Linearity was between 0.1 and 0.9 µg/zone for (1) to (4). Inter-day and intra-day precisions were below 1 % (n=3). Mean recovery was 99.7 % for (1), 99.8 % for (2), 99.7 % for (3) and 99.2 % for (4). 

J. Food. Biochem. 46, e13852 (2022). HPTLC of gallic acid (1), caffeic acid (2), quercetin (3) and ferulic acid (4) in the leaves of Aegle marmelos on silica gel with toluene - ethyl acetate - formic acid 9:10:1. Quantitative determination by absorbance measurement at 254 nm. The hRF values for (1) to (4) were 19, 37, 45 and 48, respectively. The LOD and LOQ were 9 and 24 ng/zone for (1), 6 and 17 ng/zone for (2), 8 and 23 ng/zone for (3) and 6 and 17 ng/zone for (4). Recovery was in the range of 97.9-101.3 % for (1), 100.6-104.0 % for (2), 99.2-110.4 % for (3) and 98.0-99.9 % for (4).

J. Sep. Sci. 45, 1305-1316 (2022). HPTLC of tamsulosin hydrochloride (1) and solifenacin succinate (2) along with their impurities on silica gel with ethyl acetate - butanol - glacial acetic acid 100:4:1. Quantitative determination by absorbance measurement at 225 nm. The hRF values for (1) and (2) were 61 and 26, respectively. Linearity was between 0.1 and 1.0 µg/zone for (1) and 1.0 and 15.0 µg/zone for (2). Inter-day and intra-day precisions were below 2 % (n=6). Mean recovery was 99.9 % for (1) and 99.8 % for (2). 

J. Sep. Sci. 45, 4187-4197 (2022). HPTLC of tadalafil (1) and canagliflozin (2) in spiked and real human plasma on silica gel with ethyl acetate - toluene - methanol 2:2:1. Quantitative determination by absorbance measurement at 291 nm. The hRF values for (1) and (2) were 73 and 55, respectively. Linearity was between 0.5 and 25 ng/zone for both (1) and (2). Inter-day and intra-day precisions were below 2 % (n=6). The LOD and LOQ were 0.14 and 0.43 ng/zone for (1) and 0.16 and 0.47 ng/zone for (2). Recovery was between 96.1 and 100.7 % for (1) and 97.6 and 102.0 % for (2).

J. Sep. Sci. 46, 2200695 (2023). HPTLC of phloroglucinol (1), trimethylphloroglucinol (2), and the impurity 3,5-dichloroaniline (3) on silica gel with ethyl acetate - butanol - ammonia 40:10:1. Quantitative determination by absorbance measurement at 210 nm. The hRF values for (1) to (3) were 23, 36 and 86. Linearity was between 2 and 12 µg/zone for (1), 1 and 10 µg/zone for (2) and 0.5 and 8 µg/zone for (3). Inter-day and intra-day precisions were below 1 % (n=3). The LOD and LOQ were 410 and 1250 ng/zone for (1), 310 and 940 ng/zone for (2) and 140 and 410 ng/zone for (3). Mean recovery was 98.9 % for (1), 100.7 % for (2) and 99.7 % for (3).

Chinese Medicine 15, 76 (2020). This review compared the 2020 editions of Chinese (ChP) and European Pharmacopoeas (EuP) in different aspects of quality control of traditional Chinese medicinal plants (73 of which drugs were common to both, but with differences in species or organs for 17 of them). Discussed points included history, identification, plant origin and processing, sample preparation, marker selection, tests and assays, as well as advanced analytical techniques for quality control and for the establishment of comprehensive quality standard. TLC was discussed in relation to its following aspects: purposes, markers/references, techniques and result description.
(A) The main uses of TLC and HPTLC were (1) chemical-based identification of the plant in a more accurate and precise method than by macroscopic and microscopic observation only, and in a more direct and easily interpretation than HPLC, and allowing the simultaneous analysis of multiple samples in parallel; (2) control of possible adulterants; (3) quantification of active compounds. Both uses (1) and (2) were combined in some EuP monographs: as example were given the roots of Angelica dahurica, A. pubescens, A. sinensis, using TLC for identification of the species and of adulterants from other species (Angelica, Levisticum and Ligusticum).
(B) In ChP, identification through TLC was in most cases achieved by fingerprint comparison to an official reference extract or herb (herbal reference substance). At the opposite, EuP often indicated analytical markers, irrespective of any pharmacological activity, but chosen only for analytical purposes in TCM identification and quantification. Examples were: aescin and arbutin as analytical markers for TLC identification of Anemarrhena asphodeloides rhizome and Panax notoginseng root.
For the TLC system suitability assessment tests, ChP used the same intensity markers or active markers that were chosen for the identification or assay; whereas EuP often used other specific references, e.g. isoeugenol and methyleugenol in the case of Ophiopogon japonicus roots.
(C) For the techniques, conventional separations and chemical derivatizations were used. Hyphenations of TLC to other analytical methods (e.g. MS) were absent. Only one monograph applied an effect-directed analysis directly on TLC chromatogram (free DPPH• radical scavenging assay for TLC identification of Rehmannia glutinosa root, in ChP).
Sometimes, the TLC methods were different between both reference books for the same species. Example was given for Belamcanda chinensis (=Iris domestica) rhizome: in EuP, development on silica gel with cyclohexane – ethyl acetate – acetic acid 20:80:1, detection under UV 254 nm, comparison to standards coumarin and irisflorentin; whereas in ChP, development on polyamide layer with chloroform – butanone – methanol (3:1:1), detection under UV 365nm after derivatization with aluminium chloride, comparison to a reference rhizome powder.
 (D) Finally, the results in ChP were described as a text stating the similarity of sample profile with the profile of the chosen reference, whereas the results in EuP were described with a schematic box indicating the positions of bands of interest.