Heliyon 9(2), e13469 (2023). Samples were methanolic extracts of different organs (bark, leaves, fruit pericarps, roots, twigs, seed coats and seedlings) of Dysoxylon binectariferum (= D. gotadhora = D. ficiforme, Meliaceae), as well as rohitukine (chromone piperidine alkaloid) isolated from a bark Soxhlet extract through column chromatography. TLC was used to monitor the purity of rohitukine isolation and to compare the fingerprints of the organ extracts. TLC on silica gel in 2 steps, successively with ethyl acetate – hexane 2:1, and with methanol – chloroform – dichloromethane 4:4:1. Visualization under UV 254 nm and 366 nm. Rohitukine (hRF 16) was very concentrated in bark, but present also in pericarps, leaves, twigs, seed coats and seedlings. (Editors note: Mobile phases and distribution of rohitukine were explained directly by the author (successive 2-step development, not biphasic system). The TLC figures did not show unequivocally the presence in roots, but it was confirmed by the author (and already quantified by other methods in  doi.org/10.1371/journal.pone.0158099).

Heliyon 7(2), e06116 (2021). Samples were a methanolic extract of a semi-solid ayurvedic conserve (ashwagandhadi lehyam) prepared with Withania somnifera roots (Solanaceae) and five other plants, as well as standards: withaferin A and withanolide A (= withaniol), two ergostane triterpene steroids with lactone cycle and epoxide. HPTLC on silica gel with toluene – ethyl acetate – formic acid 6:4:1. Visualization and densitometric scanning at UV 254 nm and 366 nm (deuterium lamps). Derivatization by immersion into vanillin – sulfuric acid reagent, followed by oven heating at 105 °C until optimal coloration. Documentation under white light and densitometry scanning at 540 nm (tungsten lamp). Both analytes (hRF 35 and 45 respectively) were shown at 254 nm and 540 nm (but not at 366 nm), in the standards and in the extract.

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. Biol. 45, e13764 (2021). HPTLC of filamentous fungi (Endomelanconiopsis endophytica, Myxospora musae, Neopestalotiopsis cubana and Fusarium pseudocircinatum) isolated from the digestive tract of Phylloicus sp, on silica gel with acetone - chloroform 5:3 and acetone - petroleum ether 5:2. Detection with iodine vapor, ferric chloride solution, sulfuric vanillin solution, green bromocresol solution and chromic acid solution. Qualitative identification under UV light at 254 and 366 nm. 

Chinese Medicine 7, 12 (2012). TLC of a Soxhlet hydro-ethanolic extract of Trichosanthes lobata leaves (Cucurbitaceae) on silica gel with n-hexane – ethyl acetate 7:3. Derivatization with anisaldehyde – sulfuric acid reagent. The presence of flavonoids, saponins, and tannins was found.

J. Liq. Chromatogr. Relat. Technol. 44, 809-819 (2021). HPTLC of major cannabinoids in Cannabis sativa on silica gel with hexane - ethyl acetate - methanol 7:2:1. Detection by spraying with anisaldehyde  sulfuric acid reagent. Quantitative determination by absorbance measurement at 525 nm for lupeol. The hRF values for Δ9-tetrahydrocannabinol and tetrahydrocannabinolic acid were 55 and 36, respectively. Analytical Quality by Design (AQbD) tools, such as the Design of Experiments (DoE) was used for a better understanding of the analytes’ chromatographic behavior as a function of the mobile phase composition. 

J Chromatogr A, 1647, 462153 (2021). Samples were extracts of Pittosporum angustifolium leaves (Pittosporaceae), either pure or fermented 1-4 weeks in NaCl solution, as well as acarbose, gallic acid, β-sitosterol, caffeic and chlorogenic acids, as standards. HPTLC on silica gel (prewashed with methanol and dried 15 min at 105 °C) with n-hexane – ethyl acetate – acetic acid 15:9:1. Derivatization by immersion (speed 5 cm/s, time 1 s): (A) into DPPH• 0.2 % solution, to detect radical scavengers; (B) into neutralized ferric chloride (3 % in ethanol), followed by 5 min heating at 110 °C, for detection of phenolic compounds; (C) into anisaldehyde – sulphuric acid reagent, followed by 10 min heating at 110 °C, to detect terpenes and steroids. Effect-directed analysis (EDA) for α-amylase inhibition assay (D) by immersion into enzyme solution, incubation 15 min at 37 °C, immersion into substrate solution (starch 2 % in water), incubation 20 min at 37 °C and immersion into Gram’s iodine solution for detection (inhibition zones appear blue on white background). In all cases, visualization under white light. Quantification was performed on pictures using image processing software, and expressed as equivalents to the respective standards used for calibration curves: (A) and (B) gallic acid (LOQ 250 and 740 ng/band, respectively), (C) β-sitosterol (LOQ 1.5 µg/band), (D) acarbose (LOQ 8 µg/band). Zones of interest, scraped from untreated plates and washed with ethyl acetate, were submitted by ATR-FTIR analysis. An amylase inhibiting zone (hRF 85) present in all extracts was identified as fatty acid esters: ethyl palmitate in unfermented and methyl linoleate in fermented extracts. Moreover, fermented extracts contained antioxidant zones (hRF 15 – 20), identified as monomers and oligomers (including hydroxycinnamic, guaiacyl, syringyl derivatives) from decomposed lignin.

J Chromatogr A, 1625, 461230 (2020). Samples were extracts of Chinese plants: Acorus tatarinowii (= Acorus calamus var. angustatus) rhizomes (Araceae / Acoraceae) (1), Angelica sinensis roots (Apiaceae) (2), Gynura japonica rhizomes (Asteraceae) (3), Phellodendron chinense bark (Rutaceae) (4), Picrasma quassioides twigs and leaves (Simaroubaceae) (5), Rheum sp. roots and rhizomes (R. palmatum, R. tanguticum and/or R. officinale) (Polygonaceae) (6), Sophora flavescens roots (Fabaceae) (7), Dendrobium stems (D. aphyllum, D. aurantiacum var. dennaeanum, D. chrysanthum, D. chrysotoxum, D. gratiosissimum, D. hercoglossum, D. thyrsiflorum, D. trigonopus and D. williamsonii) (Orchidaceae) (8). Standards were: gigantol (from D. sonia); methoxycarbonyl-β-carboline (MCC from (5)); caffeic acid, emodin; senecionine and β-asarone; crategolic acid (= maslinic acid), corosolic acid, oleanic acid, ursolic acid; sesquiterpenoids (atractylenolides I – III) from Atractylodes macrocephala (Asteraceae); flavonoids (baicalein, baicalin, daidzin, hesperidin, wogonin) from Scutellaria baicalensis roots (Lamiaceae). HPTLC on silica gel with 10 mobile phases, depending on the samples. Detection under UV 254 nm and white light. For (3), derivatization with Dragendorff’s reagent (bismuth potassium iodide solution) for visualization of alkaloids. Zones of interest on underivatized plates were identified by a triple-quadrupole ­– linear ion-trap MS, the compounds being removed from the layer by a continuous-wave (445 nm) diode laser pointer through a DART interface (Direct Analysis in Real-Time, helium as gas for plasma-based ambient ionization, discharge needle voltage 1.5 kV, grid voltage 350 V, capillary temperature 300 °C and voltage 40 V, full scan in positive ionization mode in m/z range 150-800). Pigment standards were used for validation of this laser-assisted HPTLC-DART-MS method: malachite green, crystal violet, chrysoidin, auramine O, rhodamine B, Sudan red I – IV, Sudan red G, dimethyl yellow. Afterwards, the same HPTLC-MS method was applied to the origin / species determination of Dendrobium samples, based on the presence of four bibenzyl compounds erianin, gigantol, moscatilin, tristin. Erianin was present only in D. chrysotoxum, whereas none of these were detected in D. hercoglossum. Several components of the extracts were thus identified: asarone (a phenylpropanoid) in (1); phthalide lactones (butenylphthalide, ligustilide and chuanxiong lactone) in (2); co-eluting pyrrolizidine alkaloids (senecionine and seneciphylline) in (3); benzylisoquinoline alkaloid berberine in (4); alkaloids (canthinone alkaloids and MCC) in (5); anthraquinones (rhein, aloe-emodin, emodin, emodin methyl ether, chrysophanol) and (in negative mode) caffeic acid (a hydroxycinnamic acid) and corosolic, maslinic and oleanic acids (triterpenoids) in (6); quinolizidine alkaloids (matrine, oxymatrine, oxysophocarpine, sophoridine) in (7).