Mechanistic Understanding of Tyrosinase Inhibition by Polymeric Proanthocyanidins from Acacia confusa Stem Bark and Their Effect on the Browning Resistance of Fresh-Cut Asparagus Lettuce.

Tyrosinase inhibitors are capable of preventing unfavorable enzymatic browning of fruits and vegetables. In this study, the capacity of Acacia confusa stem bark proanthocyanidins (ASBPs) to inhibit tyrosinase activity was evaluated. ASBPs were shown to be a high-potential inhibitor of tyrosinase with IC50 values of 92.49 ± 4.70 and 61.74 ± 8.93 µg/mL when using L-tyrosine and L-DOPA as the substrate, respectively. The structural elucidation performed with UV-vis, FT-IR spectroscopy, ESI-MS and thiolysis coupled to HPLC-ESI-MS suggested that ASBPs had structural heterogeneity in monomer units and interflavan linkages and consisted mainly of procyanidins dominant with B-type linkages. To gain insights into the inhibitory mechanisms of ASBPs against tyrosinase, different spectroscopic and molecular docking methods were further conducted. Results validated that ASBPs possessed the ability to chelate copper ions and could prevent the oxidation process of substrates by tyrosinase. The hydrogen bond formed with Lys-376 residue played a key role in the binding force of ASBPs with tyrosinase that induced a certain alteration in the microenvironment and secondary structure of tyrosinase, resulting in the enzymatic activity being ultimately restricted. It was also observed that ASBPs treatment effectively inhibited the activities of PPO and POD to retard the surface browning of fresh-cut asparagus lettuce and thus extended their shelf-life. The results provided preliminary evidence supporting the exploitation of ASBPs into potential antibrowning agents for the fresh-cut food industry.

Condensed tannins from Ulmus pumila L. leaves induce G2/M phase arrest and apoptosis via caspase-cascade activation in TFK-1 cholangiocarcinoma cells.

Condensed tannins the polyphenolic compounds that are widespread in plants have been proved to have antitumor potential. Here, we purified the bioactive condensed tannins from leaves of Ulmus pumila L. and explored their structural characteristics, antitumor effect on TFK-1 cholangiocarcinoma cells as well as the related potential mechanism. The UV-Vis, FT-IR spectroscopy, ESI-Full-MS, and thiolysis-HPLC-ESI-MS demonstrated that U. pumila condensed tannins (UCTs) consisted essentially of procyanidins with epicatechin as the main flavan-3-ol extension unit. The UCTs could significantly reduce the survival rate of human cholangiocarcinoma TFK-1, SK-CHA-1, and MZ-CHA-1 cells with the better inhibitory effect on TFK-1 cell proliferation. Flow cytometric assay showed that UCTs affected TFK-1 survival by G2/M phase arrest and inducing apoptosis in a dose-dependent manner. In addition, a total of 6592 differentially expressed genes (DEGs), consisting of 94 upregulated and 6498 downregulated DEGs, were identified between untreated and UCTs-treated TFK-1 cells using RNA-seq technology. Enrichment analysis based on the KEGG database revealed that these DEGs were closely associated with cell cycle and p53 apoptotic signaling pathways. Furthermore, qRT-PCR confirmed that treatment of UCTs to TFK-1 cells caused significant changes in the expression of cyclin E, cdc25 A, cytochrome c, caspase-3, and caspase-8. These results indicated that UCTs exhibited the growth inhibition effect on TFK-1 cells possibly via G2/M cell cycle arrest and activation of caspase-cascade to induce apoptosis, and had potential as an anti-cholangiocarcinoma drug for further development. PRACTICAL APPLICATIONS: Ulmus pumila L. as a valuable tree species has been widely used in fields of medicine and food. Condensed tannins, the polyphenolic compounds widespread in plants, have been proved to have antitumor potential and be safe to normal cells. In this study, the condensed tannins from leaves of U. pumila (UCTs) remarkably suppressed cholangiocarcinoma (CCA) cell viability possibly via G2/M cell cycle arrest and activation of caspase-cascade to induce apoptosis. The results provided evidence for the application of UCTs as a potential therapeutic drug for CCA tumor.

Conformational changes of tyrosinase caused by pentagalloylglucose binding: Implications for inhibitory effect and underlying mechanism.

Tyrosinase is a critical enzyme related to various pigmentation disorders and browning of fruits and vegetables. In this study, a novel inhibitor pentagalloylglucose (PGG) against tyrosinase was prepared from tannic acid with the chemical structure elucidated using HPLC, ESI-MS, 1H- and 13C NMR. Its inhibitory effect and the underlying mechanism on tyrosinase were explored by enzyme kinetics, UV-scanning, copper-ion chelation, fluorescence, circular dichroism, fourier transform infrared spectroscopy and molecular docking simulation. Results revealed that the yield of PGG reached 18.0% and the purity was up to 99.09%. PGG was a high-potential inhibitor of tyrosinase with IC50 values of (15.54 ± 0.56) × 10-6 and (50.89 ± 3.34) × 10-6 mol/L for monophenolase and diphenolase, respectively. PGG could disturb the formation of dopachrome and had strong capacity to chelate copper ions. The fluorescence of tyrosinase was efficiently quenched by PGG through a static mechanism. The binding of PGG to tyrosinase was a spontaneous exothermic process that induced unfolding of the tyrosinase structure to expose more buried hydrophobic residues. Docking results implied that PGG interacted with tyrosinase by forming hydrogen bonds with amino acid residues Glu-173, Glu-208, Lys-158, Lys-180, Gln-44 and Gln-159. This study would enhance our understanding of the inhibitory mechanism of PGG on tyrosinase at the molecular level and provide scientific guidance for the application of PGG in food and pharmaceutical industries.