J Chromatogr A, 1608, 460415 (2019). Samples were acetonitrile extracts of Annona cherimola fruit peel, pulp and seeds (Annonaceae), as well as caffeic acid as standards. HPTLC on silica gel with chloroform – ethyl acetate – propanol 21:2:2 for peel extracts, with chloroform – methanol 9:1 for seed extracts. Derivatization by spraying Dragendorff’s reagent for alkaloids, secondary amines and non-nitrogenous oxygenated compounds.  Effect-directed assay was performed for inhibitors of α-glucosidase. Before sample application, plates were developed with enzyme substrate (2-naphthyl-α-D-glucopyranoside 0.1 % in methanol) and dried 20 min at 60 °C. Then, samples were applied and separated, and mobile phase was removed by heating 10 min at 60 °C. The chromatogram was sprayed with 4 mL enzyme solution (5 unit/mL in 100 mM phosphate buffer,  pH 7.4), liquid excess was removed under lukewarm air stream, the plate was incubated 10 min at 37 °C in a moisture box, followed by spraying chromogenic reagent Fast Blue salt B 0.1 % in water, giving after 2 min white inhibition bands visible on purple background under white light. Plate image was documented under illumination (reflectance mode) with white light. The bands of 3 inhibiting compounds were analyzed in a triple quadrupole mass spectrometer. 1) Full scan mass spectra (m/z 50−1000) in the positive ionization mode were recorded using electrospray ionization (ESI, spray voltage 3 kV, desolvation line temperature 250 °C, block temperature 400 °C) for compounds directly eluted with methanol – acetonitrile through the oval elution head of a TLC-MS interface pump. 2) Compounds were also isolated (either eluted directly from the plate into a vial through the same interface, or scraped from the plate and extracted with methanol – chloroform into a vial), dried, and submitted to HPLC-DAD-MS/MS; MS-MS spectra were recorded in the same conditions, using argon as collision gas and collision cell voltages from -20 and -40 V. Inhibitors were identified as phenolamides (phenylethyl cinnamides): moupinamide (hRF 66 in peels, 56 in seeds), N-trans-feruloyl phenethylamine (hRF 76 in peels), N-trans-p-coumaroyl tyramine (hRF 44 in seeds).

J Chromatogr A, 1598, 209-215 (2019). Samples were methanolic extracts of honeys from Robinia pseudoacacia (Fabaceae) or from Tilia spp. (Tiliaceae / Malvaceae), as well as standards: abscisic acid (sesquiterpenoid), caffeic acid, chlorogenic acid, cinnamic acid, ferulic acid (phenolic acids), chrysin (flavone), myricetin, quercetin (flavonols), naringenin (flavanone). HPTLC on silica gel with chloroform – ethyl acetate – formic acid 5:4:1. Visualization under UV 254 nm and 366 nm, before and after derivatization by spraying with aluminium chloride (1 % in methanol), which rendered flavone bands bright yellow. Quantitative absorbance measuremet by densitometry at 366 nm. Linearity was in the range of 12,5–200 µg/mL for most standards (25–400 µg/mL for chrysin). Main differences observed in samples: 1) abscisic acid (hRF 56) and chrysin (hRF 82) were present only in Tilia honey samples, quercetin (hRF 55) only in Robinia honey; 2) ferulic acid (hRF 60) was the most prominent blue band in Tilia honey samples (1.35–18.73 g/kg of honey), and less intense in Robinia honey (0–1.24 g/kg of honey). Multivariate analysis was performed in two different ways with principal component analysis.

Molecules 25 (17), E3928 (2020). Samples were curcumin (as standard) and methanolic extracts of Curcuma xanthorrhiza and C. aeruginosa (Zingiberaceae) rhizomes, both separately and in mixtures. Separation on TLC silica gel with chloroform – methanol – formic acid 94:3:3. Densitometry of curcumin (hRF 50) in absorption mode at UV 427 nm. This method was validated with curcumin standard for selectivity (vs. demethoxycurcumin hRF 32), linearity range (250 - 450 ng), LOD (21 ng) and LOQ (69 ng), accuracy and precision. Curcumin contents were between 0.74 and 1.23 % in pure C. xanthorrhiza extracts, but decreased when adulterated with C. aeruginosa.

Molecules, 26 (5), 1468 (2021). Summary: Samples were fortified extracts produced with iPowder technology (involving spray-drying of a rich first extract on a new batch of the same plant) from following plants: Camellia sinensis final bud and two leaves (Theaceae), Cynara scolumus leaves and Echinacea purpurea roots (Asteraceae), Eleutherococcus senticosus roots (Araliaceae), Equisetum arvense aerial part (Equisetaceae), Eschscholzia californica aerial parts (Papaveraceae), Humulus lupulus cones (Cannabaceae), Ilex paraguariensis leaves (Aquifoliaceae), Melissa officinalis aerial parts and Rosmarinus officinalis leaves (Lamiaceae), Passiflora incarnata aerial part (Passifloraceae), Raphanus sativus var. niger roots (Brassicaceae), Ribes nigrum leaves (Grossulariaceae), Spiraea ulmaria floral tops (Rosaceae), Valeriana officinalis roots (Caprifoliaceae), Vitis vinifera leaves or pomace (Vitaceae). HPTLC on silica gel with 1) ethyl acetate – toluene – formic acid – water 16:4:3:2,  or 2) cyclohexane – ethyl acetate – formic acid 30:19:1. Detection under white light, UV 254 nm and 366 nm. Extract stability after 2 years was also checked through HPTLC. Neutralization by spraying phosphate-citrate buffer, and drying in cold air stream. Effect-directed analysis using automated piezoelectrical spraying: A) for enzymatic inhibition (acetyl-cholinesterase, glucosidase, glucuronidase, tyrosinase); B) for activity against Gram-negative bacteria (Aliivibrio fischeri bioluminescence assay). Active bands of multipotent compounds were eluted from HPTLC layers with methanol through the oval elution head of a TLC-MS interface pump, into a quadrupole-Orbitrap mass spectrometer. Full scan mass spectra (m/z 100−1000) in the positive and negative ionization modes were recorded using heated electrospray ionization (HESI, spray voltage 3.5 kV, capillary temperature 270 °C). By comparison to literature, the following compounds were assigned: caffeine, catechins, carnosol, chlorogenic acid, cynaratriol, dicaffeoylquinic acid, feruloyl quinic acid, gallic acid, linoleic and linolenic acids, oleanic or ursolic acid, rosmarinic acid.

J Chromatogr A, 1609, 460438 (2020). HPTLC of defatted hydro-methanolic extract of Anarrhinum pubescens (= A. duriminium) aerial parts (Plantaginaceae) on silica gel with chloroform – methanol 9:2. When intended for MS experiments, layers were previously washed twice with methanol – water 4:1 and heated 20 min at 110 °C. Derivatization by automatic piezoelectric spraying of anisaldehyde sulfuric acid reagent, followed by heating 4 min at 105 °C. Effect-directed analysis for acetyl-cholinesterase (AChE) inhibitors was performed by successive piezoelectric sprayings with TRIS buffer, with AChE solution and (after 30 min incubation at 37 °C) with naphthyl acetate and Fast Blue salt B solution. White inhibiting zones on purple background were documented under white light, and densitometry was measured by scanning in fluorescence mode at 500 nm. One of the active bands was eluted from untreated layer with methanol through the oval elution head of a TLC-MS interface pump, into a quadrupole-Orbitrap mass spectrometer using heated electrospray ionization (HESI); a full scan mass spectrum (m/z 50−750) in the positive ionization mode was recorded, as well as HRMS/MS data across a range of collision energies (10–50 V). The compound was identified as foliamenthoyl-cinnamoyl-antirrhinoside. It was applied with two other active antirrhinosides (iridoids), all isolated from the extract through column chromatography, on an HPTLC layer without migration and submitted to AChE assay; their activity was expressed as equivalency towards rivastigmine tartrate as positive control. 

J Chromatogr A, 1629, 461511 (2020). HPTLC of methanolic extracts of female inflorescences from ten hemp varieties (Cannabis sativa, Cannabaceae) on silica gel with toluene – ethyl acetate 1:1 or (for yeast assays) on RP-18W with toluene – ethyl acetate 7:3. When intended for MS experiments, layers were previously washed twice with methanol – formic acid 10:1, once with acetonitrile – methanol 2:1 and air-dried. Chromatograms were documented under white light, UV 254 nm and for fluorescence detection (FLD) at 366 nm. Afterwards, 6 derivatization assays were performed with the following reagents, either without heating: primuline; or requiring heating 5 min at 120 °C: p-aminobenzoic acid; anisaldehyde sulfuric acid; diphenylamine aniline phosphoric acid; ninhydrin; vanillin sulfuric acid. Besides, 8 effect-directed assays (EDA) were performed for free radical (DPPH•) scavengers, for antimicrobial compounds (Gram-positive Bacillus subtilis assay, Gram-negative Aliivibrio fischeri bioluminescence assay), for phytoestrogens (planar yeast estrogen assay), for inhibitors of the following enzymes: acetyl-cholinesterase (AChE), α- and β-glucosidase, tyrosinase. AChE assay was performed by immersion (speed 3.5 cm/s, time 5 s) into AChE solution (666 units in TRIS buffer 0.05 M, with bovine serum albumin 0.1 %, pH 7.8), incubation 25 min at 37 °C, spraying with substrate solution, and heating 2 min at 50 °C. Two AChE substrate solutions were used: A) α-naphthyl acetate 0.1 % and chromogenic reagent Fast Blue salt B 0.18 % in ethanol – water 1:2, giving white inhibition bands visible on purple background under white light; B) with 3-indoxyl-3-acetate, giving black inhibition bands on blue background under UV 254 nm, which was useful to prevent false negatives when Fast Blue Salt B formed colored bands with analytes. Two bands of multipotent compounds were eluted from normal-phase layer with methanol through the oval elution head of a TLC-MS interface pump, into a quadrupole-Orbitrap mass spectrometer. Full scan mass spectra (m/z 50−750) in the positive and negative ionization modes were recorded using heated electrospray ionization (HESI, spray voltage 3.5 kV, capillary temperature 270 °C). By comparison to literature and standards, they were identified as cannabidivarinic acid (hRF 55) and cannabidiolic acid (hRF 60-70).

J Chromatogr A, 1616, 461434 (2020). Samples were acetonic extracts of Malus domestica fruit peels (Rosaceae) and of Salvia officinalis, Thymus vulgaris and Origanum vulgare spice powders (Lamiaceae), as well as standards of maleic acid (dicarboxylic acid), carvacrol, thymol (phenolic monoterpenes), rosmanol (phenolic diterpene), betulinic acid, corosolic acid (CA), maslinic acid (MA), oleanolic acid (OA) and its isomer ursolic acid (UA) (triterpenes). HPTLC on silica gel, when intended for MS and NMR experiments, layers were prewashed twice with methanol – water 3:1, followed by 30 min drying at 120 °C. When intended for quantitative densitometry, start zones were submitted to prechromatographic derivatization with iodine solution (10 g/L in chloroform) allowed to migrate up to 12 mm, incubated 10 min at 27 °C and dried under cold air stream; this allowed separation of isomeric triterpenes. Separation with toluene – methanol – ethyl acetate 17:2:1 after 5 min chamber saturation at 50 % relative humidity. CA coeluted with MA, and OA with UA. Four hyphenations: A) Quantitative HPTLC densitometry for active analytes was performed by measuring absorption at 665 nm with a tungsten lamp after immersion of the chromatograms in anisaldehyde sulfuric acid reagent and heating 5 min at 110 °C. Linear range was obtained at 25 - 200 ng/band for OA and 100 - 400 ng/band for UA. B) Effect-directed analysis by immersing the chromatograms into Gram-positive Bacillus subtilis suspension for antibacterial activity and into acetyl-cholinesterase and tyrosinase solutions for enzymatic inhibition. C) Active bands were eluted with methanol through the oval elution head and in-line filter frit of a TLC-MS interface pump, into a quadrupole-Orbitrap mass spectrometer. Full scan mass spectra (m/z 100−1000) in the positive and negative ionization modes were recorded using heated electrospray ionization (HESI, spray voltage 3.5 kV, capillary temperature 270 °C, probe heater temperature 200 °C). D) With higher amounts applied, preparative HPTLC, by scraping the multipotent band corresponding to OA and UA, and dissolving these analytes in methanol, for NMR analyses (1H raw or deconvoluted, and 2D 1H–13C Heteronuclear Single Quantum Coherence). Both isomers were distinguished by their allylic H-18 protons and separately quantified by applying PULCON method (PUlse Length-based CONcentration). LOQ was 267 μM for OA and 173 μM for UA; optimal range was 300 – 4600 mM, corresponding to 126 - 2090 μg of triterpenes.

J Chromatogr A, 1616, 460774 (2020). Methanolic extracts of leaves of Musa acuminata, M. balbisiana and M. sapientum (Musaceae), either from fields or from in vitro cultures or from the plantlets derived from in vitro culture and acclimatized in isolated warm room, were separated on HPTLC silica gel layers with toluene – ethyl acetate – methanol 6:3:1 or ethyl acetate – toluene – formic acid – water 34:5:7:5. When intended for MS experiments, layers were previously washed twice with methanol – formic acid 10:1, once with acetonitrile – methanol 2:1 and air-dried. Evaluation under white light, UV 254 nm and 366 nm. Derivatization by immersion (2s, 2cm/s) into natural product reagent preceded by heating at 110 °C for 5 min, or into anisaldehyde sulfuric acid reagent, diphenylamine aniline reagent, ninhydrin reagent, followed by the same heating procedure. Besides, plates were neutralized by cold air stream followed with phosphate buffer (8 %, pH 7.5) piezoelectrically sprayed on the plates and automated plate drying. Thereafter, 9 effect-directed assays (EDA) were performed for free radical (DPPH•) scavengers, for enzymatic inhibitors (α-amylase, acetyl- and butyryl-cholinesterase, α- and β-glucosidase), for antimicrobial compounds (Gram-positive Bacillus subtilis assay, Gram-negative Aliivibrio fischeri bioluminescence assay), and for mutagenic compounds (SOS response – UMU-C test using Salmonella typhimurium suspension and 4-nitroquinoline 1-oxide as positive control). The bands of 4 active compounds were eluted with methanol through a TLC-MS interface pump into a quadrupole-Orbitrap mass spectrometer. Full scan mass spectra (m/z 50−800) in the positive and negative ionization modes were recorded using electrospray ionization (ESI, spray voltage 3.3kV, capillary temperature 320°C, collision energy 35 eV). By comparison to a standard, one band present in all samples was identified as linolenic acid. For the other bands, only present in in vitro grown accessions, only raw molecular formulas and phytochemical classes were assigned (a pyrrolidine alkaloid, an amino-acid, a phenolic derivative).