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).

J. Food. Biochem. 45, e13764 (2021). HPTLC of diosgenin in Trigonella foenum-graecum on silica gel with toluene - ethyl acetate - formic acid 5:4:1. Detection by spraying with anisaldehyde-sulfuric acid, followed by heating at 110 °C for 10 min. Quantitative determination by absorbance measurement at 620 nm. The hRF value for diosgenin was 83. 

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.

Pharmacogn. Mag. 18, 836-843 (2022). HPTLC of shatavarin IV in the roots of Asparagus racemosus on silica gel with ethyl acetate - methanol - water 15:3:2. Detection by dipping into anisaldehyde-sulfuric acid reagent, followed by heating at 110 °C for 5 min. Quantitative determination by absorbance measurement at 425 nm. The hRF value for hatavarin IV was 40. Linearity was between 72 and 432 ng/zone. Intermediate precisions were below 2 % (n=3). The LOD and LOQ were 24 and 72 ng/zone, respectively. Mean recovery was 97.5 %.

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.

J. Sep. Sci. 45, 1616-1635 (2022). HPTLC of gallic acid (1), protocatechuic acid (2), vanillic acid (3), cinnamic acid (4), piperine (5), guggulsterone-E (6), and guggulsterone-Z (7) in Mahayograj Guggul on silica gel with toluene - ethyl acetate - formic acid 10:9:2 for (1) to (3) and toluene - acetone 9:1 for (4) to (7). Quantitative determination by absorbance measurement at 250 nm for (6) and (7), 280 nm for (1), (2), (3) and (4) and 343 nm for (5). The hRF values for (1) to (7) were 30, 41, 47, 15, 33, 41 and 45, respectively. Linearity was between 100 and 1000 µg/mL for (1), 5 and 60 µg/mL for (2), 10 and 80 µg/mL for (3), (4) and (7), 20 and 100 µg/mL for (5) and 40 and 120 µg/mL for (6). Inter-day and intra-day precisions were below 4 % (n=18). The LOD and LOQ were 4 and 14 µg/g for (1), 7 and 21 µg/g for (2), 24 and 72 µg/g for (3), 0.8 and 2.4 µg/g for (4), 12 and 35 µg/g for (5), 2 and 6 µg/g for (6) and 4 and 14 µg/g for (7), respectively. Recovery was between 86.6 and 102.0 % for (1) to (7).

J. Ethnopharmacol. 289, 115035 (2022). HPTLC of Unani pharmacopoeial preparations Itrifal Hakim Ali (1) and Habb-e-Kalaf (2) on silica gel with toluene - ethyl acetate 9:1 for (1) and toluene - ethyl acetate 4:1. Detection under UV light at 254 and 366 nm. 

Chinese Medicine 10, 13 (2015). Samples were root and rhizome extracts of Panax notoginseng (Araliaceae), either raw or in the form of commercial granules. Standards were ginsenosides Rg1, Rb1, Rd, Re and Rg2, notoginsenoside NR1. TLC on silica gel with chloroform – ethyl acetate – methanol – water 15:40:22:9, followed by 10 min air drying. Derivatization for ginsenosides by immersion into sulfuric acid (10 % in ice cold methanol), followed by 10 min air drying and 5 min heating at 100 °C. Quantification by densitometric fluorescence measurement (deuterium and tungstene lamp, 366 nm). For each standard the linear range was 0.05-1 mg/mL (LOQ comprised between 38 and 431 µg/µL). As NR1 and Re (ratio ca. 2:1) had almost the same hRF, they were quantified together as one substance. Multivariate analysis through hierarchical (HCA) and principal component analyses (PCA) was used to order the samples into two clusters, according to the analyte concentrations, the raw plant extracts being richer than most of the commercial products. This TLC method was compared to quantification through UPLC-PDA (Ultra-performance liquid chromatography with photo diode array), which was more sensitive (LOQ between 10 and 49 µg/µL) but did not allow the separation between Rg1 and Re (ratio ca. 6:1).