Antrodia cinnamomea May Interfere with the Interaction Between ACE2 and SARS-CoV-2 Spike Protein in vitro and Reduces Lung Inflammation in a Hamster Model of COVID-19.

Purpose: Coronavirus disease 2019 (COVID-19) poses a global health challenge with widespread transmission. Growing concerns about vaccine side effects, diminishing efficacy, and religious-based hesitancy highlight the need for alternative pharmacological approaches. Our study investigates the impact of the ethanol extract of Antrodia cinnamomea (AC), a native medicinal fungus from Taiwan, on COVID-19 in both in vitro and in vivo contexts. Methods: We measured the mRNA and protein levels of angiotensin-converting enzyme-2 (ACE2) in human lung cells using real-time reverse transcriptase-polymerase chain reaction and Western blotting, respectively. Additionally, we determined the enzymatic activity of ACE2 using the fluorogenic peptide substrate Mca-YVADAPK(Dnp)-OH. To assess the impact of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection, we used SARS-CoV-2 pseudovirus infections in human embryonic kidney 293T cells expressing ACE2 to measure infection rates. Furthermore, we evaluated the in vivo efficacy of AC in mitigating COVID-19 by conducting experiments on hamsters infected with the Delta variant of SARS-CoV-2. Results: AC effectively decreased ACE2 mRNA and protein levels, a critical host receptor for the SARS-CoV-2 spike protein, in human lung cells. It also prevented the spike protein from binding to human lung cells. Dehydrosulphurenic acid, an isolate from AC, directly inhibited ACE2 protease activity with an inhibitory constant of 1.53 µM. In vitro experiments showed that both AC and dehydrosulphurenic acid significantly reduced the infection rate of SARS-CoV-2 pseudovirus. In hamsters infected with the Delta variant of SARS-CoV-2, oral administration of AC reduced body weight loss and improved lung injury. Notably, AC also inhibited IL-1ß expression in both macrophages and the lung tissues of SARS-CoV-2-infected hamsters. Conclusion: AC shows potential as a nutraceutical for reducing the risk of SARS-CoV-2 infection by disrupting the interaction between ACE2 and the SARS-CoV-2 spike protein, and for preventing COVID-19-associated lung inflammation.

Correction: Systematic discovery of drug action mechanisms by an integrated chemical genomics approach: identification of functional disparities between azacytidine and decitabine.

[This corrects the article DOI: 10.18632/oncotarget.8455.].

Systematic discovery of drug action mechanisms by an integrated chemical genomics approach: identification of functional disparities between azacytidine and decitabine.

Polypharmacology (the ability of a drug to affect more than one molecular target) is considered a basic property of many therapeutic small molecules. Herein, we used a chemical genomics approach to systematically analyze polypharmacology by integrating several analytical tools, including the LINCS (Library of Integrated Cellular Signatures), STITCH (Search Tool for Interactions of Chemicals), and WebGestalt (WEB-based GEne SeT AnaLysis Toolkit). We applied this approach to identify functional disparities between two cytidine nucleoside analogs: azacytidine (AZA) and decitabine (DAC). AZA and DAC are structurally and mechanistically similar DNA-hypomethylating agents. However, their metabolism and destinations in cells are distinct. Due to their differential incorporation into RNA or DNA, functional disparities between AZA and DAC are expected. Indeed, different cytotoxicities of AZA and DAC toward human colorectal cancer cell lines were observed, in which cells were more sensitive to AZA. Based on a polypharmacological analysis, we found that AZA transiently blocked protein synthesis and induced an acute apoptotic response that was antagonized by concurrently induced cytoprotective autophagy. In contrast, DAC caused cell cycle arrest at the G2/M phase associated with p53 induction. Therefore, our study discriminated functional disparities between AZA and DAC, and also demonstrated the value of this chemical genomics approach that can be applied to discover novel drug action mechanisms.

Differential proteomics to explore the inhibitory effects of acidic, slightly acidic electrolysed water and sodium hypochlorite solution on Vibrio parahaemolyticus.

Slightly acidic electrolysed water (SlAEW) and acidic electrolysed water (AEW) have been demonstrated to effectively inactivate food-borne pathogens. However, the underlying mechanism of inactivation remains unknown. Therefore, in this study, a differential proteomic platform was used to investigate the bactericidal mechanism of SlAEW, AEW, and sodium hypochlorite (NaOCl) solutions against Vibrio parahaemolyticus. The upregulated proteins after SlAEW, AEW, and NaOCl treatments were identified as outer membrane proteins K and U. The downregulated proteins after the SlAEW, AEW, and NaOCl treatments were identified as adenylate kinase, phosphoglycerate kinase, glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and enolase, all of which are responsible for energy metabolism. Protein synthesis-associated proteins were downregulated and identified as elongation factor Tu and GAPDH. The inhibitory effects of SlAEW and AEW solutions against V. parahaemolyticus may be attributed to the changes in cell membrane permeability, protein synthesis activity, and adenosine triphosphate (ATP) biosynthesis pathways such as glycolysis and ATP replenishment.

A low toxicity synthetic cinnamaldehyde derivative ameliorates renal inflammation in mice by inhibiting NLRP3 inflammasome and its related signaling pathways.

Uncontrolled inflammation is a leading cause of various chronic diseases. Cinnamaldehyde (CA) is a major bioactive compound isolated from the essential oil of the leaves of Cinnamomum osmophloeum kaneh that exhibits anti-inflammatory activity; however, the use of CA is limited by its cytotoxicity. Here, we synthesized three CA derivatives and identified 4-hydroxycinnamaldehyde-galactosamine (HCAG) as a low toxicity anti-inflammatory compound in vitro (HCAG IC50 ≫ 1600 µM; CA IC50=40 µM) and in vivo. HCAG reduced pro-inflammatory mediator expression in LPS-activated macrophages by inhibiting MAPK and PKC-α/δ phosphorylation, decreasing ROS generation and reducing NF-κB activation. HCAG also reduced NLRP3 inflammasome-derived IL-1ß secretion by inhibiting the ATP-mediated phosphorylation of AKT and PKC-α/δ. In a mouse model of LPS-induced renal inflammation, we observed reduced albuminuria and a mild degree of glomerular proliferation, glomerular sclerosis and periglomerular inflammation in the HCAG-treated mice compared with the vehicle-treated mice. The underlying mechanisms for these renoprotective effects involved: (1) inhibited NLRP3 inflammasome activation; (2) decreased superoxide anion levels and apoptosis; and (3) suppressed activation of NF-κB and related downstream inflammatory mediators.

Glycosylation-dependent interaction between CD69 and S100A8/S100A9 complex is required for regulatory T-cell differentiation.

Cluster of differentiation (CD)69 is a leukocyte activation receptor involved in the maintenance of immune homeostasis and is positively selected in activated regulatory T (Treg) cells, implicating its role during Treg-cell differentiation. By RNA interference, we show that CD69 is not sufficient to support the conversion of CD4(+) naive T cells into Treg cells, whereas it does that of human peripheral blood mononuclear cells (hPBMCs) (P < 0.01), suggesting that a ligand-receptor interaction is required for CD69 function. Using immunoprecipitation and mass spectrometry, we identified the S100A8/S100A9 complex as the natural ligand of CD69 in hPBMCs. CD69 specifically associates with S100A8/S100A9 complex as confirmed by in vitro binding and competition assay, and the treatment of CD69 with peptide-N-glycosidase significantly abolishes such association. In agreement, the glycomics analysis determines the glycosylation site and the N-glycan composition of CD69, and terminal removal of sialic acid from that N-linked glycans reverses the generation of forkhead box P3-positive Treg cells (23.21%; P < 0.05). More specifically, we showed that CD69-S100A8/S100A9 association is required for the up-regulation of suppressor of cytokine signaling 3 resulting in inhibited signaling of signal transducer and activator of transcription 3 (36.54% increase upon CD69 silencing; P < 0.01). This might in turn support the secretion of key regulator TGF-ß (∼ 3.28-fold decrease upon CD69 silencing; P < 0.05), leading to reduced production of IL-4 in hPBMCs. Our results demonstrate the functional and mechanistic interplays between CD69 and S100A8/S100A9 in supporting Treg-cell differentiation.

Proteomic analysis of polysaccharide-milk protein interactions induced by chitosan.

The chitosan-induced coacervation of milk proteins was investigated using a proteomic approach. The addition of 0.8% chitosan to milk caused the milk proteins to coacervate after a 1 h incubation period. Approximately 86% of the milk proteins were present in the milk pellet fraction (MPF), and the protein concentration of the milk supernatant fraction (MSF) decreased from 29.4±0.2 to 4.2±0.6 mg/mL. SDS-PAGE analysis showed that the total intensities of serum albumin (BSA), αS-casein (αS-CN), ß-casein (ß-CN), κ-casein (κ-CN) and ß-lactoglobulin (ß-LG) in the MSF decreased to 8.5%±0.2%, 0.9%±0.3%, 0.7%±0.3%, 0.5%±0.2% and 15.0%±0.5%, respectively. Two-dimensional electrophoresis analysis indicated that αS1-, αS2-, ß- and κ-CN and a fraction of the ß-LG and BSA were found in the MSF following incubation with 0.8% chitosan. Isothermal titration calorimetry analysis indicated that binding of chitosan to milk proteins is an exothermic reaction based on binding titration curves of milk proteins dispersions with chitosan, and the enthalpy of binding (ΔH) and binding constant (Ka) were -7.85×10(4) cal/mol and 1.06×10(5)/mol, respectively. These results suggested that the addition of 0.8% chitosan causes milk proteins to coacervate due to polysaccharide-protein interactions.

Catalysis of rice straw hydrolysis by the combination of immobilized cellulase from Aspergillus niger on ß-cyclodextrin-Fe3O4 nanoparticles and ionic liquid.

Cellulase from Aspergillus niger was immobilized onto ß-cyclodextrin-conjugated magnetic particles by silanization and reductive amidation. The immobilized cellulase gained supermagnetism due to the magnetic nanoparticles. Ninety percent of cellulase was immobilized, but the activity of immobilized cellulase decreased by 10%. In this study, ionic liquid (1-butyl-3-methylimidazolium chloride) was introduced into the hydrolytic process because the original reaction was a solid-solid reaction. The activity of immobilized cellulase was improved from 54.87 to 59.11 U g immobilized cellulase(-1) at an ionic liquid concentration of 200 mM. Using immobilized cellulase and ionic liquid in the hydrolysis of rice straw, the initial reaction rate was increased from 1.629 to 2.739 g h(-1) L(-1). One of the advantages of immobilized cellulase is high reusability--it was usable for a total of 16 times in this study. Compared with free cellulase, magnetized cellulase can be recycled by magnetic field and the activity of immobilized cellulase was shown to remain at 85% of free cellulase without denaturation under a high concentration of glucose (15 g L(-1)). Therefore, immobilized cellulase can hydrolyze rice straw continuously compared with free cellulase. The amount of harvested glucose can be up to twentyfold higher than that from the hydrolysis by free cellulase.

Anti-diabetic effects of Ganoderma lucidum.

Ganoderma lucidum is a white rot fungus widely used as a tonic for the promotion of longevity and health. Extracts of G. lucidum have been recognized as an alternative adjuvant treatment for diabetes. Among the many biologically active constituents of G. lucidum, polysaccharides, proteoglycans, proteins and triterpenoids have been shown to have hypoglycemic effects. G. lucidum polysaccharides have been reported to have hypoglycemic activity by increasing plasma insulin levels and decreasing plasma sugar levels in mice. Protein tyrosine phosphatase 1B is a promising therapeutic target in diabetes, and G. lucidum proteoglycan can inhibit this enzyme in vitro. Moreover, G. lucidum triterpenoids were shown to have inhibitory activity on aldose reductase and α-glucosidase that can suppress postprandial hyperglycemia. In addition, a protein Ling Zhi-8 extracted from G. lucidum significantly decreased lymphocyte infiltration and increased the antibody detection of insulin in diabetic mice. This review summarizes most of the research about the hypoglycemic action effects of polysaccharides, proteoglycans, proteins and tritrerpenoids from G. lucidum as a guide for future research.

GM2-activator protein: a new biomarker for lung cancer.

INTRODUCTION: Effective biomarkers for early diagnosis of lung cancer are needed. A recent study demonstrated that urinary GM2-activator protein (GM2AP) level was increased in lung cancer patients. This study aims to validate the potential application of GM2AP as a biomarker for diagnosis of lung cancer. METHODS: Serum and urine samples were obtained from 189 participants (133 patients for treatment naive lung cancer, 26 healthy volunteers for urine, and 30 healthy volunteers for serum). GM2AP level was detected by Western blotting and quantified using enzyme-linked immunosorbent assay (ELISA). The GM2AP expression in tumors and nontumor parts of lung tissues from 143 nonsmall cell lung cancers was detected by immunohistochemical stains. RESULTS: There was an 8.11 ± 1.36 folds increase in urine and a 5.41 ± 0.73 folds increase in serum level of GM2AP in lung cancer patients compared with healthy volunteers (p < 0.0001), achieving a 0.89 AUC value in urine and 0.90 AUC value in serum for the receiver-operating characteristic curves. Both serum and urine levels of GM2AP correlated significantly with pathology stages (urine, p = 0.009; serum, p < 0.0001). Using immunohistochemical, positive expression of GM2AP was found at 83.9% of nonsmall cell lung cancers patients and none in normal tissue. The GM2AP expression was significantly correlated with pathology stage (p = 0.0001). Patients with higher GM2AP expression had shorter overall survival (p = 0.045) and disease-free survival (p = 0.049) than lower GM2AP expression. Moreover, the multivariate analysis suggested GM2AP as an independent predictors of disease-free survival and overall survival. CONCLUSIONS: Our study demonstrates that GM2AP might serve as potential diagnostic and prognostic biomarkers in patients with lung cancer.

Down-regulation of Slit-Robo pathway mediating neuronal cytoskeletal remodeling processes facilitates the antidepressive-like activity of Gastrodia elata Blume.

Nowadays, depression is a serious psychological disorder that causes extreme economic loss and social problems. Previously, we discovered that the water extract of Gastrodia elata Blume (WGE) improved depressive-like behavior by influencing neurotransmitters in rats subjected to the forced swimming test. To elucidate possible mechanisms, in the present study, we performed a proteomics and bioinformatics analysis to identify the related pathways. Western blot-validated results indicated that the core protein network modulated by WGE administration was closely associated with down-regulation of the Slit-Robo pathway, which modulates neuronal cytoskeletal remodeling processes. Although Slit-Robo signaling has been well investigated in neuronal development, its relationship with depression is not fully understood. We provide a potential hint on the mechanism responsible for the antidepressive-like activity of WGE. In conclusion, we suggest that the Slit-Robo pathway and neuronal cytoskeleton remodeling are possibly one of the pathways associated with the antidepressive-like effects of WGE.

MicroRNA-21-3p, a berberine-induced miRNA, directly down-regulates human methionine adenosyltransferases 2A and 2B and inhibits hepatoma cell growth.

Methionine adenosyltransferase (MAT) is the cellular enzyme that catalyzes the synthesis of S-adenosylmethionine (SAM), the principal biological methyl donor and a key regulator of hepatocyte proliferation, death and differentiation. Two genes, MAT1A and MAT2A, encode 2 distinct catalytic MAT isoforms. A third gene, MAT2B, encodes a MAT2A regulatory subunit. In hepatocellular carcinoma (HCC), MAT1A downregulation and MAT2A upregulation occur, known as the MAT1A:MAT2A switch. The switch is accompanied with an increasing expression of MAT2B, which results in decreased SAM levels and facilitates cancer cell growth. Berberine, an isoquinoline alkaloid isolated from many medicinal herbs such as Coptis chinensis, has a wide range of pharmacological effects including anti-cancer effects. Because drug-induced microRNAs have recently emerged as key regulators in guiding their pharmacological effects, we examined whether microRNA expression is differentially altered by berberine treatment in HCC. In this study, we used microRNA microarrays to find that the expression level of miR-21-3p (previously named miR-21*) increased after berberine treatment in the HepG2 human hepatoma cell line. To predict the putative targets of miR-21-3p, we integrated the gene expression profiles of HepG2 cells after berberine treatment by comparing with a gene list generated from sequence-based microRNA target prediction software. We then confirmed these predictions through transfection of microRNA mimics and a 3' UTR reporter assay. Our findings provide the first evidence that miR-21-3p directly reduces the expression of MAT2A and MAT2B by targeting their 3' UTRs. In addition, an overexpression of miR-21-3p increased intracellular SAM contents, which have been proven to be a growth disadvantage for hepatoma cells. The overexpression of miR-21-3p suppresses growth and induces apoptosis in HepG2 cells. Overall, our results demonstrate that miR-21-3p functions as a tumor suppressor by directly targeting both MAT2A and MAT2B, indicating its therapeutic potential in HCC.

The immunologically active oligosaccharides isolated from wheatgrass modulate monocytes via Toll-like receptor-2 signaling.

Wheatgrass is one of the most widely used health foods, but its functional components and mechanisms remain unexplored. Herein, wheatgrass-derived oligosaccharides (WG-PS3) were isolated and found to induce CD69 and Th1 cytokine expression in human peripheral blood mononuclear cells. In particular, WG-PS3 directly activated the purified monocytes by inducing the expression of CD69, CD80, CD86, IL-12, and TNF-α but affected NK and T cells only in the presence of monocytes. After further purification and structural analysis, maltoheptaose was identified from WG-PS3 as an immunomodulator. Maltoheptaose activated monocytes via Toll-like receptor 2 (TLR-2) signaling, as discovered by pretreatment of blocking antibodies against Toll-like receptors (TLRs) and also determined by click chemistry. This study is the first to reveal the immunostimulatory component of wheatgrass with well defined molecular structures and mechanisms.

Lengthening of 3'UTR increases with morphological complexity in animal evolution.

MOTIVATION: Evolutionary expansion of gene regulatory circuits seems to boost morphological complexity. However, the expansion patterns and the quantification relationships have not yet been identified. In this study, we focus on the regulatory circuits at the post-transcriptional level, investigating whether and how this principle may apply. RESULTS: By analysing the structure of mRNA transcripts in multiple metazoan species, we observed a striking exponential correlation between the length of 3' untranslated regions (3'UTR) and morphological complexity as measured by the number of cell types in each organism. Cellular diversity was similarly associated with the accumulation of microRNA genes and their putative targets. We propose that the lengthening of 3'UTRs together with a commensurate exponential expansion in post-transcriptional regulatory circuits can contribute to the emergence of new cell types during animal evolution.

Benzoxazinoids from Scoparia dulcis (sweet broomweed) with antiproliferative activity against the DU-145 human prostate cancer cell line.

Sweet broomweed (Scoparia dulcis) is an edible perennial medicinal herb widely distributed in tropical and subtropical regions of Asia, Africa, and the Americas. Four compounds, (2R)-7-methoxy-2H-1,4-benzoxazin-3(4H)-one 2-O-ß-galactopyranoside [(2R)-HMBOA-2-O-Gal], 3,6-dimethoxy-benzoxazolin-2(3H)-one (3,6-M2BOA), 3-hydroxy-6-methoxy-2-benzoxazolinone (3-OH-MBOA), and scutellarein 7-O-ß-glucuronamide, along with eight known compounds, including two 7-methoxy-1,4-benzoxazin-3(2H)-one 3-O-hexopyranosides [(2R)-HMBOA-2-O-Glc and (2R)-HDMBOA-2-O-Glc], 6-methoxy-benzoxazolin-2(3H)-one (MBOA), acteoside, sodium scutellarin, p-coumaric acid, and two monosaccharides (fructose and glucose), were isolated from the aqueous extract of S. dulcis. Antiproliferative activities of the six benzoxazinoid compounds against the DU-145 human prostate cancer cell line were assayed, and one of these displayed an IC50 of 65.8 µg/mL.

Chemical and biological evaluation of nephrocizin in protecting nerve growth factor-differentiated PC12 cells by 6-hydroxydopamine-induced neurotoxicity.

The neurotoxin 6-hydroxydopamine (6-OHDA) has been widely used to generate an experimental model of Parkinson's disease. This model is crucial in the search for compounds that diminish 6-OHDA-induced nerve growth factor (NGF)-differentiated PC12 cell death. Nephrocizin (luteolin-7-O-ß-D-glucopyranoside), a flavone glycoside, was isolated from widely distributed plants. The protective effects of pre-treatment with nephrocizin on the induced neurotoxicity in PC12 cells by 6-OHDA and its oxidative products, H2O2-, and p-quinone, were evaluated herein. Nephrocizin promoted cell viability, scavenged ROS-related products, increased cellular glutathione (GSH) levels, and reduced caspase-3 and -8 activities in 6-OHDA-, H2O2-, or p-quinone-treated PC12 cells. Furthermore, nephrocizin-conjugated metabolites in PC12 cells were identified with the boronate-affinity method and LC-MS technology, and preferential regioselectivity at the C2' and C5' positions by the nephrocizin-GSH (or NAC) adduct method was observed. These lines of evidence established that nephrocizin could form a dimer to diminish the intracellular ROS. These results demonstrate the first neuroprotective mechanism of nephrocizin against 6-OHDA-, H2O2- or p-quinone-induced cytotoxicity in PC12 cells via chemical and biological studies. These dietary antioxidants are potential candidates for use in intervention in neurodegenerative diseases.

Improved purity and immunostimulatory activity of ß-(1→3)(1→6)-glucan from Pleurotus sajor-caju using cell wall-degrading enzymes.

The objective of this work was to improve the purity of ß-(1→3)(1→6)-glucan in the native triple helical structure from the fruiting bodies of Pleurotus sajor-caju for effective biological function using cell wall-degrading enzymes. A crude carbohydrate was extracted with hot water, then treated with crude xylanase and cellulase from Paenibacillus curdlanolyticus B-6. ß-Glucan in the extract was purified to homogeneity with a single and symmetrical peak using 650M DEAE Toyopearl and Sepharose CL-6B column chromatography. The purity of ß-glucan was confirmed by high-performance size-exclusion chromatography. Purified ß-glucan was obtained at a purity of up to 90.2%. The Congo red reaction and atomic force microscopy indicated that the purified ß-glucan exhibited a triple helix conformation. Purified ß-glucan was able to effectively up-regulate the functions of macrophages such as nitric oxide (NO) and tumor necrosis factor (TNF-α) production.

Oligosaccharide and peptidoglycan of Ganoderma lucidum activate the immune response in human mononuclear cells.

The acid-hydrolyzed fragments of Ganoderma lucidum polysaccharides (GLPS) obtained by Smith degradation were separated by size-exclusion chromatography into two major water-soluble fractions: peptidoglycans (GLPS-SF1) and oligosaccharides (GLPS-SF2). Both fractions induced CD69 in human peripheral blood mononuclear cells (hPB-MNCs), and they displayed distinct immunomodulating properties. GLPS-SF1, with a molecular weight of around 20 kDa, were heterogeneous peptidoglycans composed of glucose/mannose (4:1) that exhibited biological activities with Th1 cytokines IL-12, IL-2, TNF-α, and IFN-γ in hPB-MNCs and stimulated macrophage cytokine expression via Toll-like receptor 4 (TLR4) signaling. For GLPS-SF2, with a molecular weight of around several kilodaltons, its sugar sequence was elucidated by mass spectrometry (MS) and nuclear magnetic resonance (NMR) spectroscopy as [-α-1,4-Glc-(ß-1,4-GlcA)(3)-](n). This oligosaccharide displayed specific immune property with low monocyte induction, greatly stimulated cell activation and proliferation of NK and T cells. This oligosaccharide isolated from G. lucidum polysaccharides with internal glucuronic acids/glucose repeat unit in a 3:1 ratio may be responsible for the active stimulation of NK and T cells.

Anti-inflammatory and free radial scavenging activities of the constituents isolated from Machilus zuihoensis.

A new biflavonol glycoside, quercetin-3-O-ß-D-glucopyranoside-(3'→O-3''')-quercetin-3-O-ß-D-galactopyranoside (9), together with eight known compounds was isolated for the first time from the leaves of Machilus zuihoensis Hayata (Lauraceae). The structure of compound 9 was elucidated by various types of spectroscopic data analysis. Analysis of the biological activity assay found that compound 9 showed significant superoxide anion scavenging activity (IC50 is 30.4 µM) and markedly suppressed LPS-induced high mobility group box 1 (HMGB-1) protein secretion in RAW264.7 cells. In addition, the HMGB-1 protein secretion was also inhibited by quercitrin (3), ethyl caffeate (6), and ethyl 3-O-caffeoylquinate (7) treatment. In the LPS-stimulated inducible nitric oxide synthase (iNOS) activation analysis, two known compounds, quercetin (1) and ethyl caffeate (6), were found to markedly suppress nitric oxide (NO) production (IC50 value, 27.6 and 42.9 µM, respectively) in RAW264.7 cells. Additionally, it was determined that ethyl caffeate (6) down-regulated mRNA expressions of iNOS, IL-1ß, and IL-10 in the LPS-treatment of RAW264.7 cells via a suppressed NF-kB pathway. These results suggested for the first time that the new compound 9 and other constituents isolated from M. zuihoensis have potential anti-inflammatory and superoxide anion scavenging effects. These constituents may be useful for treating various inflammatory diseases.