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 methanolic extracts of commercial supplements containing Magnolia sp. bark (Magnoliaceae), as well as honokiol (1) and magnolol (2) (biphenyl neolignans) as separated or mixed standards. TLC and HPTLC on silica gel with n-hexane – ethyl acetate – ethanol 16:3:1. Visualization under UV 254 nm. Quantification of (1) and (2) by densitometric scanning in absorbance mode at 290 nm (hRF were 34 and 39, LOQ 200 ng and 280 ng/spot, respectively). Variability between samples from the same brand supplement was also determined, as well as extraction yields. Effect-directed analysis with 3 assays: A) to detect radical scavengers, immersion into DPPH• 0.02 % solution; B) to detect activity against Gram-negative bacteria, immersion into Aliivibrio fischeri suspension, followed by recording the bioluminescence; C) to detect activity against Gram-positive bacteria, immersion into Bacillus subtilis, followed by incubation 2 h at 28 °C and immersion into MTT 1 g/L. Compounds (1) and (2) were active in all assays. Identification of zones of interest by eluting with methanol from untreated TLC layer through the oval elution head of a TLC-MS interface directly to a single Quadrupole MS (electrospray ionization, interface temperature 350°C, heat block temperature 400°C, desolvation line temperature 250°C, detector voltage 4.5kV). Full mass scan spectra were recorded in the positive and negative ionization modes in m/z range 150–800. Other molecules (from other ingredients) were identified: piperine (alkaloid) and/or its geometrical isomers (active on A, hRF 29-30); and daidzein (active on A and B, hRF 18), isoflavone from Pueraria montana root (Fabaceae). Stability was assessed through 2D-HPTLC, by repeating the same development method in the orthogonal direction 4 h or 20 h after the first separation. Degradation products of (1) and (2) appeared after 20 h (but not at 4 h), including a honokiol dimer (formed in tracks of (1) and of (2)).

J Chromatogr A, 1628, 461461 (2020). Samples were hydro-methanolic extracts of 100 genuine saffron samples (Crocus sativus stigmata, Iridaceae) from South Khorasan (SK) and Razavi Khorasan (RK) provinces (Iran), pure or mixed in several proportions with common vegetal adulterants: C. sativus style, Calendula officinalis petals (Asteraceae, Asteroideae), Carthamus tinctorius petals (Asteraceae, Carduoideae), Rubia tinctorum rhizomes (Rubiaceae). Commercial saffron samples (containing artificial adulterants) were also tested. TLC on silica gel with ethyl acetate – methanol – water – acetic acid 66:23:11:1. Evaluation at 254 nm, 366 nm, and 440 nm. Crocin (carotenoid, hRF 38) was used for optimization of extraction (parameters being first calculated by chemometry), using multilinear regression and ANOVA. Image data (pixel intensities and colors of each sample under the three selected wavelengths) were unfolded into a data matrix and transformed into a vector, used for multivariate image analysis of the chromatogram fingerprints. This allowed: A) separation of genuine samples by principal component analysis (PCA) into 2 clusters according to origin (cold climate in Northern half of RK vs. warm climate in SK and Southern part of RK) with 92 % prediction accuracy; B) separation of samples according to purity / vegetal adulterant groups by partial least squares – discriminant analysis (PLS-DA) with 98 % accuracy (if 10 µL extract applied); C) separation with 100 % prediction accuracy by PCA between genuine, mixed, and commercial samples.

J Chromatogr A, 1620, 460970 (2020). Samples were ethyl acetate extracts of Lavandula angustifolia herb and flowers and of aerial parts of other Lamiaceae (Ocimum basilicum, Origanum vulgare, Thymus vulgaris, Rosmarinus officinalis, Salvia officinalis), as well as standards. HPTLC on silica gel (pre-washed with methanol and heated 30 min at 105 °C) with n-hexane – ethyl acetate – acetic acid 70:27:3. Documentation at UV 254 nm and 365 nm and white light before and after A) derivatization with anisaldehyde – sulfuric acid reagent, followed by 10 min heating at 110 °C; B) spraying with DPPH• (0.2 % in methanol), followed by 30 min incubation in the dark; C) α-amylase inhibition assay by immersion into enzyme solution, incubation 30 min at 37 °C, immersion into substrate solution (starch 1 % in water), incubation 20 min at 37 °C and immersion into Gram’s iodine solution for detection (inhibition zones appear blue on white background). Quantification was performed on pictures using image processing software, and expressed as equivalents to the respective standards used for calibration curves: A) β-sitosterol (LOQ 1.5 µg/band), B) gallic acid (LOQ 60 ng/band), C) acarbose (LOQ 8 µg/band). An amylase inhibiting zone (hRF 68) present in all samples (except L. angustifolia), scraped from untreated plates and washed with ethyl acetate, was tentatively identified by ATR-FTIR analysis as oleanolic acid (pentacyclic triterpene).

J. Planar Chromatogr. 35, 473-479 (2022). HPTLC of capsaicin and dihydrocapsaicin as capsaicinoids in Habanero pepper pods on silica gel with cyclohexane - chloroform - acetic acid 7:2:1. Quantitative determination by absorbance measurement at 254 nm. The hRF value for capsaicinoids was 29. Linearity was between 0.5 and 4.0 µg/zone. Interday and intra-day precisions were below 6 % (n=6). The LOD and LOQ were 251 and 750 ng/zone, respectively. 

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. Planar Chromatogr. 35, 313-330 (2022).  HPTLC of orange peel extract on silica gel with gradient multiple development using seven different polarity ranges: cyclohexane, cyclohexane - n-heptane 3:7, cyclohexane - methyl tert-butyl ether 43:7, cyclohexane - methyl tert-butyl ether 7:3, cyclohexane - methyl tert-butyl ether 3:7, methyl tert-butyl ether, methyl acetate - ethanol 9:1, ethyl acetate - ethanol - formic acid 44:5:1. Detection by spraying with vanillin reagent (100 mg vanillin dissolved in 9.8 mL ethanol and 0.2 mL sulfuric acid), followed by heating at 100 °C for 2 min. DPPH staining was performed with 2 mL of a DPPH solution (15 mg dissolved in 10 mL of methanol). Bioautography was performed by dipping into Aliivibrio fischeri bacteria suspension for 6 s, followed by measurement of bioluminescence within 15 min. In this sample, more than 50 compounds could be separated.

Chinese J. Food & Drug 23 (3), 229-232 (2021). Non-benzodiazepines are inhibitors directly acting in the central nervous system, with sedative, anxiolytic and anticonvulsant effects. They are used as third generation sedative-hypnotic drugs to treat insomnia. However, it has been found that they are illegally added to some health foods on the market, which may cause long-term abuse and cause harm to the human body. Therefore, it is necessary to strenghten a rapid screening to detect the illegal addition of non-benzodiazepines to health foods, to improve the efficiency of supervision, and to crack down illegal practices.  TLC of ethyl acetate extracts of health foods adulterated with non-benzodiazepines, on silica gel with dichloromethane - methanol - ethyl acetate 17:2:6. Detection in UV 254 nm, identification by fingerprint comparison with the standards zolpidem, zalepron and zoppilon. Using this method, 65 batches of health food samples obtained from the market were screened, and four of them were suspected of illegal addition of zopiclone. This result is consistent with the analysis using both HPLC-DAD and HPLC-MS, and proved the  TLC method to be simple, fast, accurate, easy to operate, and well suitable for quick screening.