Biotransformation-guided purification of a novel glycoside derived from the extracts of Chinese herb Baizhi.

Our pursuit of new compounds with enhanced bioavailability and bioactivity prompted us to employ the biotransformation-guided purification (BGP) approach which leverages proficient in vitro biotransformation techniques. Angelica dahurica roots, also called Baizhi in Chinese traditional medicine, are famous for their anti-inflammatory and analgesic properties. Herein, we applied the BGP methodology to Baizhi extracts, employing Deinococcus geothermalis amylosucrase (DgAS), an enzyme demonstrating catalytic competence across diverse substrates, for biotransformation. Initiating with a 70 % methanol extraction, we obtained the crude extract of commercial Baizhi powder, followed by an additional extraction using ethyl acetate. Notably, reactions performed on this extract yielded limited quantities of novel compounds. Subsequently, the extract underwent partitioning into four fractions based on HPLC profiling, leading to the successful isolation of a compound with significant yield from fraction 2 mixtures upon reaction with DgAS. Structural elucidation confirmed the compound as byakangelicin-7″-O-α-glucopyranoside (BG-G), a new alpha glycoside derivative of byakangelicin. Furthermore, validation experiments verified the capacity of DgAS to glycosylate pure byakangelicin, yielding BG-G. Remarkably, the aqueous solubility of BG-G exceeded that of byakangelicin by over 29,000-fold. In conclusion, BGP emerges as a potent strategy combining traditional medicinal insights with robust enzymatic tools for generating new compounds.

Novel Ganoderma triterpenoid saponins from the biotransformation-guided purification of a commercial Ganoderma extract.

Ganoderma sp. contains high amounts of diverse triterpenoids; however, few triterpenoid saponins could be isolated from the medicinal fungus. To produce novel Ganoderma triterpenoid saponins, biotransformation-guided purification (BGP) process was applied to a commercial Ganoderma extract. The commercial Ganoderma extract was partially separated into three fractions by preparative high-performance liquid chromatography, and the separated fractions were then directly biotransformed by a Bacillus glycosyltransferase (BsUGT489). One of the biotransformed products could be further purified and identified as a novel saponin: ganoderic acid C2 (GAC2)-3-O-ß-glucoside by nucleic magnetic resonance (NMR) and mass spectral analyses. Based on the structure of the saponin, the predicted precursor should be the GAC2, which was confirmed to be biotransformed into four saponins, GAC2-3-O-ß-glucoside, GAC2-3,15-O-ß-diglucoside and two unknown GAC2 monoglucosides, revealed by NMR and mass spectral analyses. GAC2-3-O-ß-glucoside and GAC2-3,15-O-ß-diglucoside possessed 17-fold and 200-fold higher aqueous solubility than that of GAC2, respectively. In addition, GAC2-3-O-ß-glucoside retained the most anti-α-glucosidase activity of GAC2 and was comparable with that of the anti-diabetes drug (acarbose). The present study showed that the BGP process is an efficient strategy to survey novel and bioactive molecules from crude extracts of natural products.

A Novel Soy Isoflavone Derivative, 3'-Hydroxyglycitin, with Potent Antioxidant and Anti-α-Glucosidase Activity.

This study demonstrated the enzymatic hydroxylation of glycitin to 3'-hydroxyglycitin, confirming the structure by mass and nucleic magnetic resonance spectral analyses. The bioactivity assays further revealed that the new compound possessed over 100-fold higher 1,1-diphenyl-2-picrylhydrazine free-radical scavenging activity than the original glycitin, although its half-time of stability was 22.3 min. Furthermore, the original glycitin lacked anti-α-glucosidase activity, whereas the low-toxic 3'-hydroxyglycitin displayed a 10-fold higher anti-α-glucosidase activity than acarbose, a standard clinical antidiabetic drug. The inhibition mode of 3'-hydroxyglycitin was noncompetitive, with a Ki value of 0.34 mM. These findings highlight the potential use of the new soy isoflavone 3'-hydroxyglycitin in biotechnology industries in the future.

A New Stilbene Glucoside from Biotransformation-Guided Purification of Chinese Herb Ha-Soo-Oh.

Ha-Soo-Oh is a traditional Chinese medicine prepared from the roots of Polygonum multiflorum Thunb. The herb extract has been widely used in Asian countries as a tonic agent and nutritional supplement for centuries. To identify new bioactive compounds in Chinese herbs, the biotransformation-guided purification (BGP) process was applied to Ha-Soo-Oh with Bacillus megaterium tyrosinase (BmTYR) as a biocatalyst. The result showed that a major biotransformed compound could be purified using the BGP process with preparative high-performance liquid chromatography (HPLC), and it was confirmed as a new compound, 2,3,5,3',4'-pentahydroxystilbene-2-O-ß-glucoside (PSG) following mass and nucleic magnetic resonance (NMR) spectral analyses. PSG was further confirmed as a biotransformation product from 2,3,5,4'-tetrahydroxystilbene-2-O-ß-glucoside (TSG) by BmTYR. The new PSG exhibited 4.7-fold higher 1,1-diphenyl-2-picrylhydrazine (DPPH) free radical scavenging activity than that of TSG. The present study highlights the potential usage of BGP in herbs to discover new bioactive compounds in the future.

Enzymatic Synthesis of Novel and Highly Soluble Puerarin Glucoside by Deinococcus geothermalis Amylosucrase.

Puerarin (daidzein-8-C-glucoside) is an isoflavone isolated from several leguminous plants of the genus Pueraria. Puerarin possesses several pharmacological properties; however, the poor solubility of puerarin limits its applications. To resolve this poor solubility, Deinococcus geothermalis amylosucrase (DgAS) was used to modify puerarin into more soluble derivatives. The results showed that DgAS could biotransform puerarin into a novel compound: puerarin-4'-O-α-glucoside. The biotransformation reaction was manipulated at different temperatures, pH values, sucrose concentrations, reaction times, and enzyme concentrations. The results showed that the optimal reaction condition was biotransformed by 200 µg/mL DgAS with 20% (w/v) sucrose at pH 6 and incubated at 40 °C for 48 h, and the optimal production yield was 35.1%. Puerarin-4'-O-α-glucoside showed 129-fold higher solubility than that of puerarin and, thus, could be further applied for pharmacological use in the future.

Novel Glycosylation by Amylosucrase to Produce Glycoside Anomers.

Glycosylation occurring at either lipids, proteins, or sugars plays important roles in many biological systems. In nature, enzymatic glycosylation is the formation of a glycosidic bond between the anomeric carbon of the donor sugar and the functional group of the sugar acceptor. This study found novel glycoside anomers without an anomeric carbon linkage of the sugar donor. A glycoside hydrolase (GH) enzyme, amylosucrase from Deinococcus geothermalis (DgAS), was evaluated to glycosylate ganoderic acid F (GAF), a lanostane triterpenoid from medicinal fungus Ganoderma lucidum, at different pH levels. The results showed that GAF was glycosylated by DgAS at acidic conditions pH 5 and pH 6, whereas the activity dramatically decreased to be undetectable at pH 7 or pH 8. The biotransformation product was purified by preparative high-performance liquid chromatography and identified as unusual α-glucosyl-(2→26)-GAF and ß-glucosyl-(2→26)-GAF anomers by mass and nucleic magnetic resonance (NMR) spectroscopy. We further used DgAS to catalyze another six triterpenoids. Under the acidic conditions, two of six compounds, ganoderic acid A (GAA) and ganoderic acid G (GAG), could be converted to α-glucosyl-(2→26)-GAA and ß-glucosyl-(2→26)-GAA anomers and α-glucosyl-(2→26)-GAG and ß-glucosyl-(2→26)-GAG anomers, respectively. The glycosylation of triterpenoid aglycones was first confirmed to be converted via a GH enzyme, DgAS. The novel enzymatic glycosylation-formed glycoside anomers opens a new bioreaction in the pharmaceutical industry and in the biotechnology sector.

Improving Aqueous Solubility of Natural Antioxidant Mangiferin through Glycosylation by Maltogenic Amylase from Parageobacillus galactosidasius DSM 18751.

Mangiferin is a natural antioxidant C-glucosidic xanthone originally isolated from the Mangifera indica (mango) plant. Mangiferin exhibits a wide range of pharmaceutical activities. However, mangiferin's poor solubility limits its applications. To resolve this limitation of mangiferin, enzymatic glycosylation of mangiferin to produce more soluble mangiferin glucosides was evaluated. Herein, the recombinant maltogenic amylase (MA; E.C. 3.2.1.133) from a thermophile Parageobacillus galactosidasius DSM 18751T (PgMA) was cloned into Escherichia coli BL21 (DE3) via the expression plasmid pET-Duet-1. The recombinant PgMA was purified via Ni2+ affinity chromatography. To evaluate its transglycosylation activity, 17 molecules, including mangiferin (as sugar acceptors), belonging to triterpenoids, saponins, flavonoids, and polyphenol glycosides, were assayed with ß-CD (as the sugar donor). The results showed that puerarin and mangiferin are suitable sugar acceptors in the transglycosylation reaction. The glycosylation products from mangiferin by PgMA were isolated using preparative high-performance liquid chromatography. Their chemical structures were glucosyl-α-(1→6)-mangiferin and maltosyl-α-(1→6)-mangiferin, determined by mass and nucleic magnetic resonance spectral analysis. The newly identified maltosyl-α-(1→6)-mangiferin showed 5500-fold higher aqueous solubility than that of mangiferin, and both mangiferin glucosides exhibited similar 1,1-diphenyl-2-picrylhydrazyl free radical scavenging activities compared to mangiferin. PgMA is the first MA with glycosylation activity toward mangiferin, meaning mangiferin glucosides have potential future applications.

Enzymatic Synthesis of Novel Vitexin Glucosides.

Vitexin is a C-glucoside flavone that exhibits a wide range of pharmaceutical activities. However, the poor solubility of vitexin limits its applications. To resolve this limitation, two glycoside hydrolases (GHs) and four glycosyltransferases (GTs) were assayed for glycosylation activity toward vitexin. The results showed that BtGT_16345 from the Bacillus thuringiensis GA A07 strain possessed the highest glycosylation activity, catalyzing the conversion of vitexin into new compounds, vitexin-4'-O-ß-glucoside (1) and vitexin-5-O-ß-glucoside (2), which showed greater aqueous solubility than vitexin. To our knowledge, this is the first report of vitexin glycosylation. Based on the multiple bioactivities of vitexin, the two highly soluble vitexin derivatives might have high potential for pharmacological usage in the future.

Glycosylation of Ganoderic Acid G by Bacillus Glycosyltransferases.

Ganoderma lucidum is a medicinal fungus abundant in triterpenoids, its primary bioactive components. Although numerous Ganoderma triterpenoids have already been identified, rare Ganoderma triterpenoid saponins were recently discovered. To create novel Ganoderma saponins, ganoderic acid G (GAG) was selected for biotransformation using four Bacillus glycosyltransferases (GTs) including BtGT_16345 from the Bacillus thuringiensis GA A07 strain and three GTs (BsGT110, BsUGT398, and BsUGT489) from the Bacillus subtilis ATCC 6633 strain. The results showed that BsUGT489 catalyzed the glycosylation of GAG to GAG-3-o-ß-glucoside, while BsGT110 catalyzed the glycosylation of GAG to GAG-26-o-ß-glucoside, which showed 54-fold and 97-fold greater aqueous solubility than that of GAG, respectively. To our knowledge, these two GAG saponins are new compounds. The glycosylation specificity of the four Bacillus GTs highlights the possibility of novel Ganoderma triterpenoid saponin production in the future.

A new macrocyclic diterpenoid from Anisomeles indica.

A new macrocyclic diterpenoid, 4ß,5ß-dihydroxyovatodiolide (1), together with twenty-two known compounds (2-23) were isolated from the MeOH extract of the dried aerial parts of Anisomeles indica (L.) O. Kuntze (Labiatae). The structure of 1 was established on the basis of spectral evidence. Phenylethanoids, acteoside (5) and isoacteoside (6) showed significant inhibitory to IL-2 secretion of with respect to phorbol myristate acetate and anti-CD28 monoclonal antibody co-stimulated activation of human peripheral blood T cells.

Tris(8-Hydroxyquinoline)iron induces apoptotic cell death via oxidative stress and by activating death receptor signaling pathway in human head and neck carcinoma cells.

BACKGROUND: 8-Hydroxyquinoline derivatives have highly sensitive fluorescent chemosensors for metal ions, which are associated with anti-oxidant, anti-tumor and anti-HIV-1 properties. Head and neck squamous cell carcinoma (HNSCC) is associated with a high rate of mortality and novel anti-HNSCC drugs must be developed. Therefore, effective chemotherapy agents are required to address this public health issue. HYPOTHESIS/PURPOSE: The aim of this study was to investigate the inhibitory effect of tris(8-hydroxyquinoline)iron (Feq3) on the HNSCC and the underlying mechanism. STUDY DESIGN/METHODS: A novel 8-hydroxyquinoline derivative, Feq3, was synthesized. The cell viabilities were analyzed using MTT reagent. Apoptosis and the cell cycle distributions were determined by flow cytometer. Reverse transcription-polymerase chain reaction (RT-PCR), immunofluorescence, western blot, MitoSOX and CellROX stain assay were used to study the mechanism of Feq3. Feq3 combined with antioxidants NAC (N-acetylcysteine) and BSO (buthionine sulfoximine) measured the cell viability and intracellular ROS. RESULTS: Feq3 induced the death of HNSCC cells and caused them to exhibit the morphological features of apoptosis. Feq3 also induced apoptosis of SCC9 cells by cell cycle arrest during the G2/M phase and the induced arrest of SCC25 cells in the G0/G1 and G2/M phases, which was associated with decreased cyclin B1/cdc2 and cyclin D/cdk4 expressions. Feq3 increases reactive oxygen species (ROS) and reduces glutathione (GSH) levels, and responds to increased p53 and p21 expressions. Feq3 induced apoptosis by mitochondria-mediated Bax and cytochrome c up-expression and down-expression Bcl-2. Feq3 also up-regulated tBid, which interacts with the mitochondrial pathway and tumor necrosis factor-α (TNF-α)/TNF-Rs, FasL/Fas, and TNF-related apoptosis inducing ligand receptors (TRAIL-Rs)/TRAIL-dependent caspases apoptotic signaling pathway in HNSCC cells. However, Feq3 activates Fas but not FasL in SCC25 cells. Feq3 arrests the growth of HNSCC cells and is involved in the mitochondria- and death receptor (DR)-mediated caspases apoptotic pathway. CONCLUSION: This study is the first to suggest that apoptosis mediates the anti-HNSCC of Feq3. Feq3 has potential as a cancer therapeutic agent against HNSCC.

Anti-inflammatory Activity of 8-Hydroxydaidzein in LPS-Stimulated BV2 Microglial Cells via Activation of Nrf2-Antioxidant and Attenuation of Akt/NF-κB-Inflammatory Signaling Pathways, as Well As Inhibition of COX-2 Activity.

It was demonstrated that isoflavones can cross the blood-brain barrier, making them desirable candidate agents for the prevention of neurological symptoms. 8-Hydroxydaidzein (8-OHD, 4',7,8-trihydoxyisoflavone) is an isoflavone found only in fermented soy food. Current results showed that 8-OHD inhibited LPS-stimulated production of nitric oxide (NO) and proinflammatory cytokines, such as tumor necrosis factor (TNF)-α and interleukin (IL)-6, by inhibiting gene expression in BV2 microglial cells. Moreover, 8-OHD markedly quenched reactive oxygen species (ROS) and activated NF-E2-related factor 2 (Nrf2) so as to upregulate expression of Phase II enzymes, including heme oxygenase (HO)-1, NAD(P)H quinone dehydrogenase 1 (NQO1), and the modifier subunit of glutamate cysteine ligase (GCLM). 8-OHD also suppressed LPS-stimulated phosphorylation of Akt and NF-κB-p65. The anti-inflammatory activity of 8-OHD was attenuated by the HO-1 inhibitor zinc protoporphyrin (Znpp) but augmented by the PI3K/Akt inhibitor LY294002. 8-OHD also diminished LPS-induced prostaglandin E2 (PGE2) production without affecting cyclooxygenase (COX)-2 expression. In vitro assay shows that 8-OHD displayed mixed-type inhibition of COX-2 with an IC50 of 8.9 ± 1.2 µM. These data suggest that the anti-inflammatory activity of 8-OHD may be associated with the activation of Nrf2/HO-1 and attenuation of Akt/NF-κB signaling pathways as well as inhibition of COX-2 enzyme activity. In conclusion, 8-OHD, a potent Nrf2 activator, Akt/NF-κB activation suppressor, and COX-2 enzyme inhibitor, may have health-promoting effects for mitigating microglia activation and preventing neuroinflammation.

Cleome rutidosperma and Euphorbia thymifolia Suppress Inflammatory Response via Upregulation of Phase II Enzymes and Modulation of NF-κB and JNK Activation in LPS-Stimulated BV2 Microglia.

Cleome rutidosperma DC. and Euphorbia thymifolia L. are herbal medicines used in traditional Indian and Chinese medicine to treat various illnesses. Reports document that they have antioxidant and anti-inflammatory activities; nonetheless, the molecular mechanisms involved in their anti-inflammatory actions have not yet been elucidated. The anti-neuroinflammatory activities and underlying mechanisms of ethanol extracts of Cleome rutidosperma (CR) and Euphorbia thymifolia (ET) were studied using lipopolysaccharide (LPS)-stimulated microglial cell line BV2. The morphology changes and production of pro-inflammatory mediators were assayed. Gene expression of inflammatory genes such as inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, interleukin (IL)-1ß, and CC chemokine ligand (CCL)-2, as well as phase II enzymes such as heme oxygenase (HO)-1, the modifier subunit of glutamate cysteine ligase (GCLM) and NAD(P)H quinone dehydrogenase 1 (NQO1), were further investigated using reverse transcription quantitative-PCR (RT-Q-PCR) and Western blotting. The effects of CR and ET on mitogen activated protein kinases (MAPKs) and nuclear factor (NF)-κB signaling pathways were examined using Western blotting and specific inhibitors. CR and ET suppressed BV2 activation, down-regulated iNOS and COX-2 expression and inhibited nitric oxide (NO) overproduction without affecting cell viability. They reduced LPS-mediated tumor necrosis factor (TNF) and IL-6 production, attenuated IL-1ß and CCL2 expression, but upregulated HO-1, GCLM and NQO1 expression. They also inhibited p65 NF-κB phosphorylation and modulated Jun-N terminal kinase (JNK) activation in BV2 cells. SP600125, the JNK inhibitor, significantly augmented the anti-IL-6 activity of ET. NF-κB inhibitor, Bay 11-7082, enhanced the anti-IL-6 effects of both CR and ET. Znpp, a competitive inhibitor of HO-1, attenuated the anti-NO effects of CR and ET. Our results show that CR and ET exhibit anti-neuroinflammatory activities by inhibiting pro-inflammatory mediator expression and production, upregulating HO-1, GCLM and NQO1, blocking NF-κB and modulating JNK signaling pathways. They may offer therapeutic potential for suppressing overactivated microglia and alleviating neurodegeneration.

The Inhibitory Mechanisms Study of 5,6,4'-Trihydroxy-7,3'-Dimethoxyflavone against the LPS-Induced Macrophage Inflammatory Responses through the Antioxidant Ability.

The whole plant of Anisomeles ovata has been widely used in Taiwan for treating inflammation-related skin and liver diseases, however, the detailed pharmacology mechanisms have yet to be elucidated. In the present study, one of the major components, 5,6,4'-trihydroxy-7,3'-dimethoxyflavone (5-TDMF), was purified from a methanol extract of Anisomeles ovata. A pharmacological study of this compound suggests that 5-TDMF possesses potent free radical scavenging activity both in vitro and ex vivo. Furthermore, 5-TDMF reduces nitric oxide and pro-inflammatory cytokine production in LPC-treated RAW 264.7 cells through the attenuation of nitric oxide synthase and cyclooxygenase-2. Additional experiments suggest that of 5-TDMF interferes with nuclear factor-κB translocation and mitogen-activated protein kinase pathways. These results identify 5-TDMF as an anti-oxidant and anti-inflammatory compound, explain the pharmacologic function of Anisomeles ovata and suggest its great potential as a new anti-inflammatory remedy.

Production of Two Novel Methoxy-Isoflavones from Biotransformation of 8-Hydroxydaidzein by Recombinant Escherichia coli Expressing O-Methyltransferase SpOMT2884 from Streptomyces peucetius.

Biotransformation of 8-hydroxydaidzein by recombinant Escherichia coli expressing O-methyltransferase (OMT) SpOMT2884 from Streptomyces peucetius was investigated. Two metabolites were isolated and identified as 7,4'-dihydroxy-8-methoxy-isoflavone (1) and 8,4'-dihydroxy-7-methoxy-isoflavone (2), based on mass, 1H-nuclear magnetic resonance (NMR) and 13C-NMR spectrophotometric analysis. The maximum production yields of compound (1) and (2) in a 5-L fermenter were 9.3 mg/L and 6.0 mg/L, respectively. The two methoxy-isoflavones showed dose-dependent inhibitory effects on melanogenesis in cultured B16 melanoma cells under non-toxic conditions. Among the effects, compound (1) decreased melanogenesis to 63.5% of the control at 25 µM. This is the first report on the 8-O-methylation activity of OMT toward isoflavones. In addition, the present study also first identified compound (1) with potent melanogenesis inhibitory activity.

Anti-inflammatory effect and mechanism of the green fruit extract of Solanum integrifolium Poir.

The green fruit of Solanum integrifolium Poir. has been used traditionally as an anti-inflammatory and analgesic remedy in Taiwanese aboriginal medicine. The goal of this study is to evaluate the anti-inflammatory activity and mechanism of the green fruit extract of S. integrifolium. A bioactivity-guided fractionation procedure was developed to identify the active partition fraction. The methanol fraction (ME), with the highest phenolic content, exhibited the strongest inhibitory effect against LPS-mediated nitric oxide (NO) release and cytotoxicity in RAW264.7 macrophages. ME also significantly downregulated the expression of LPS-induced proinflammatory genes, such as iNOS, COX-2, IL-1ß, IL-6, CCL2/MCP-1, and CCL3/MIP1α. Moreover, ME significantly upregulated HO-1 expression and stimulated the activation of extracellular-signal-regulated kinase 1/2 (ERK1/2). Pretreatment of cells with the HO-1 inhibitor zinc protoporphyrin and MEK/ERK inhibitor U0126 attenuated ME's inhibitory activity against LPS-induced NO production. Taken together, this is the first study to demonstrate the anti-inflammatory activity of green fruit extract of S. integrifolium and its activity may be mediated by the upregulation of HO-1 expression and activation of ERK1/2 pathway.

Antioxidant, anti-adipocyte differentiation, antitumor activity and anthelmintic activities against Anisakis simplex and Hymenolepis nana of yakuchinone A from Alpinia oxyphylla.

BACKGROUND: Alpinia oxyphylla is a common remedy in traditional Chinese medicine. Yakuchinone A is a major constituent of A. oxyphylla and exhibits anti-inflammatory, antitumor, antibacterial, and gastric protective activities. METHODS: Antioxidant and antitumor characteristics of yakuchinone A in skin cancer cells as well as novel mechanisms for the inhibition of adipocyte differentiation, cestocidal activities against Hymenolepis nana adults, and nematocidal activities against Anisakis simplex larvae are investigated. RESULTS: Yakuchinone A presents the ability of the removal of DPPH·and ABTS+ free radicals and inhibition of lipid peroxidation. Yakuchinone A suppresses intracellular lipid accumulation during adipocyte differentiation in 3 T3-L1 cells and the expressions of leptin and peroxisome proliferator-activated receptor γ (PPARγ). Yakuchinone A induces apoptosis and inhibits cell proliferation in skin cancer cells. The inhibition of cell growth by yakuchinone A is more significant for non-melanoma skin cancer (NMSC) cells than for melanoma (A375 and B16) and noncancerous (HaCaT and BNLCL2) cells. Treatment BCC cells with yakuchinone A shows down-regulation of Bcl-2, up-regulation of Bax, and an increase in cleavage poly (ADP-ribose) polymerase (PARP). This suggests that yakuchinone A induces BCC cells apoptosis through the Bcl-2-mediated signaling pathway. The anthelmintic activities of yakuchinone A for A. simplex are better than for H. nana. CONCLUSIONS: In this work, yakuchinone A exhibits antioxidative properties, anti-adipocyte differentiation, antitumor activity, and anthelmintic activities against A. simplex and H. nana.

Brazilein from Caesalpinia sappan L. Antioxidant Inhibits Adipocyte Differentiation and Induces Apoptosis through Caspase-3 Activity and Anthelmintic Activities against Hymenolepis nana and Anisakis simplex.

Brazilein, a natural, biologically active compound from Caesalpinia sappan L., has been shown to exhibit anti-inflammatory and antioxidant properties and to inhibit the growth of several cancer cells. This study verifies the antioxidant and antitumor characteristics of brazilein in skin cancer cells and is the first time to elucidate the inhibition mechanism of adipocyte differentiation, cestocidal activities against Hymenolepis nana, and reduction of spontaneous movement in Anisakis simplex. Brazilein exhibits an antioxidant capacity as well as the ability to scavenge DPPH(•) and ABTS(•+) free radicals and to inhibit lipid peroxidation. Brazilein inhibited intracellular lipid accumulation during adipocyte differentiation in 3T3-L1 cells and suppressed the induction of peroxisome proliferator-activated receptor γ (PPAR γ ), the master regulator of adipogenesis, suggesting that brazilein presents the antiobesity effects. The toxic effects of brazilein were evaluated in terms of cell viability, induction of apoptosis, and the activity of caspase-3 in BCC cells. The inhibition of the growth of skin cancer cells (A431, BCC, and SCC25) by brazilein is greater than that of human skin malignant melanoma (A375) cells, mouse leukemic monocyte macrophage (RAW 264.7 cells), and noncancerous cells (HaCaT and BNLCL2 cells). The anthelmintic activities of brazilein against Hymenolepis nana are better than those of Anisakis simplex.

trans-Caffeic acid stearyl ester from Paeonia suffruticosa inhibits melanin synthesis by cAMP-mediating down-regulation of α-melanocyte-stimulating hormone-stimulated melanogenesis signaling pathway in B16 cells.

trans-Caffeic acid stearyl ester (TCASE) from the root cortex of Paeonia suffruticosa ANDREWS is a traditional medicinal herb that has several beneficial properties. However, the inhibitory effect of TCASE on melanogenesis has not been explored. In the cell viability assay, TCASE did not show a cytotoxic effect at a dose of 65 µM for 48 h in B16, HaCaT and Hs68 cells. TCASE considerably inhibits melanin synthesis, and reduces intracellular cyclic adenosine monophosphate (cAMP) levels, tyrosinase activity and L-3-(3,4-dihydroxyphenyl)-alanine (DOPA) oxidase activity in a concentration-dependent manner in the presence of α-melanocyte-stimulating hormone (α-MSH) in B16 cells, and the inhibition efficiency of TCASE exceeds that of ascorbic acid and arbutin. TCASE reduces melanocortin-1 receptor (MC1R), microphthalmia transcription factor (MITF), tyrosinase, tyrosinase-related protein-2 (TRP-2) and TRP-1 mRNA and protein levels in B16 cells. Based on the findings, TCASE is posited to inhibit melanogenesis signaling while suppressing cAMP levels and, subsequently, MC1R, MITF, tyrosinase, TRP-2 and TRP-1 down-regulation, resulting in the suppression of tyrosinase activity, DOPA oxidase activity and melanin synthesis.

Melanogenesis inhibition by homoisoflavavone sappanone A from Caesalpinia sappan.

Homoisoflavanone, sappanone A, was isolated from Caesalpinia sappan and proven to dose-dependently inhibit both melanogenesis and cellular tyrosinase activity via repressing tyrosinase gene expression in mouse B16 melanoma cells. To our knowledge, sappanone A is the first homoisoflavanone to be discovered with melanogenesis inhibitory activity. Our results give a new impetus to the future search for other homoisoflavanone melanogenesis inhibitors.