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Background: Biogenic amines are harmful to human health at a certain extent. As a kind of biogenic amine oxidase, multicopper oxidase can be used to degrade them. Currently, the literature about enzyme from Enterococcus spp. are limited, and recombinant multicopper oxidase might be an effective way to degrade biogenic amines. Objective: (i) Select and identify strains that can degrade biogenic amines, (ii) overexpress enzyme from Enterococcus spp., (iii) measure gene expression and probe amine-degradation differences among strains (native, E. coli DH5α, and L. delbruckii), and (iv) examine the biochemical properties of recombinant multicopper oxidase, (v) apply the recombinant enzyme into smoked horsemeat sausage. Methods: Reverse transcription PCR and high-performance liquid chromatography were performed to examine gene expression and amine degradation rate. Results: The results demonstrated that target enzymes were successfully overexpressed, accompanied by increased amine-degrading activity (P Conclusion: In this work, gene expression levels were higher in recombinant M5B than recombinant M2B, regardless of host. E. coli is more stable to express multicopper oxidase. Besides, the amine-degrading ability was markedly increased in the two recombinant strains. After prolonged incubation, the recombinant enzyme could degrade three amines, and it displayed high alkali resistance and thermostability.

Eight new flavonoids, including two β-hydroxy/methoxychalcones, velutones A and B (1 and 2), two 1,3-diarylpropan-1-ols, velutols C and D (3 and 4), a dihydroxychalcone, velutone E (5), a chalcone, velutone F (6), a furanoflavanone, velutone G (7), and a furanoflavonol, velutone H (8), and 14 known compounds were isolated from Millettia velutina. Their structures were determined by high-resolution electrospray ionisation mass spectrometry (HR-ESIMS) and spectroscopic data analyses and time-dependent density functional theory electronic circular dichroism (TD-DFT-ECD) calculations. Among the isolated constituents, compound 6 exhibited the most potent inhibitory effect (IC50: 1.3 μM) against nigericin-induced IL-1β release in THP-1 cells. The initial mechanism of action study revealed that compound 6 suppressed NLRP3 inflammasome activation via blocking ASC oligomerization without affecting the priming step, which subsequently inhibited caspase-1 activation and IL-1β secretion. Most importantly, compound 6 exerted potent protective effects in the LPS-induced septic shock mice model by improving the survival rate of mice and suppressing serum IL-1β release. These results demonstrated that compound 6 had the potential to be developed as a broad-spectrum NLRP3 inflammasome inhibitor for the treatment of NLRP3-related disease.

Black pepper (Piper nigrum L.) has been commonly utilized in food preparation and traditional medicine in several countries. Seven new amide alkaloids, pipernigramides A-G (3, 10, 38, and 41-44), a new piperic ester, pipernigrester A (48), along with 47 known compounds were isolated from the EtOH extract of P. nigrum. The inhibitory effects on nitric oxide (NO) of all compounds were then evaluated. Among the tested compounds, three of them (42-44) significantly inhibited inducible nitric oxide synthase (iNOS)-mediated NO (IC50 = 4.74 ± 0.18, 4.08 ± 0.19, and 3.71 ± 0.32 μM, respectively), and IL-1β, IL-6, TNF-α, and PGE2 release in RAW 264.7 cells stimulated by lipopolysaccharide. Moreover, 42-44 suppressed IκB degradation and further inhibited the cytosol-nucleus translocation of the p65 subunit by targeting IKK-β. In the carrageenan-induced paw edema test, 42-44 demonstrated anti-inflammatory effects as well. These results indicate that all three compounds from P.nigrum have the potential anti-inflammatory effects.

To develop highly efficient non-noble-metal electrocatalysts for oxygen evolution reaction is still a great challenge. Herein, cobalt and iron oxide with hollow structure has been fabricated using ZIF-67 as template, via reflux process followed by calcination. The simultaneous structural and electronic modulation are achieved due to the homogenous Fe incorporation. The Co sites in higher oxidation states and Fe sites with higher electron density can enhance the adsorption of H2O molecules and OOH species, and thus promote the water oxidation. The unique hollow structure also facilitates the mass transfer at the interface of electrolyte and active atoms. The optimal cobalt and iron oxide hollow nanocages exhibit superior OER performance, delivering an ultralow overpotential of 274 mV at 10 mA cm−2 and a very small Tafel slope of 31 mV dec−1 in alkaline media, which outperform the state-of-the-art RuO2. This work provides an efficient approach to design and explore non-precious electrocatalysts for electrochemical energy conversion and storage.

Magnolol and honokiol are the two major active ingredients with similar structure and anticancer activity from traditional Chinese medicine Magnolia officinalis, and honokiol is now in a phase I clinical trial (CTR20170822) for advanced non-small cell lung cancer (NSCLC). In search of potent lead compounds with better activity, our previous study has demonstrated that magnolol derivative C2, 3-(4-aminopiperidin-1-yl)methyl magnolol, has better activity than honokiol. Here, based on the core of 3-(4-aminopiperidin-1-yl)methyl magnolol, we synthesized fifty-one magnolol derivatives. Among them, compound 30 exhibited the most potent antiproliferative activities on H460, HCC827, H1975 cell lines with the IC50 values of 0.63–0.93 μM, which were approximately 10- and 100-fold more potent than those of C2 and magnolol, respectively. Besides, oral administration of 30 and C2 on an H460 xenograft model also demonstrated that 30 has better activity than C2. Mechanism study revealed that 30 induced G0/G1 phase cell cycle arrest, apoptosis and autophagy in cancer cells. Moreover, blocking autophagy by the autophagic inhibitor enhanced the anticancer activity of 30 in vitro and in vivo, suggesting autophagy played a cytoprotective role on 30-induced cancer cell death. Taken together, our study implied that compound 30 combined with autophagic inhibitor could be another choice for NSCLC treatment in further investigation.

Millettia pulchra is a renowned anti-inflammatory herbal medicine in southeast provinces of China. However, the underlying anti-inflammation mechanism remained incompletely understood. Herein, four new isoflavones, pulvones A-D and eleven reported constituents were isolated from the stems of Millettia pulchra with their structures being elucidated by HRMS and NMR analysis. The anti-inflammatory activities of pulvones A and C were further evaluated due to the better inhibitory activity on nitric oxide production in LPS-stimulated RAW264.7 cells and no obvious cytotoxicity to RAW264.7 cells. Western blot showed that pulvones A significantly decreased the levels of iNOS and COX-2 proteins and pulvones C only decreased the level of iNOS protein. ELISA analysis demonstrated that pulvones A inhibited the production of both interleukin-6 (IL-6) and IL-1β while pulvones C showed better suppression effect on IL-1β production in LPS-stimulated RAW264.7 cells. Then, their potential inhibitory effects on NF-κB pathway were tested in LPS-stimulated RAW264.7 cells. Immunofluorescence and western blot assay showed that pulvones A and C reduced the nuclear translocation of NF-κB(p65) and interrupted IκB phosphorylation. The ADP-Glo™ kinase assay showed pulvones A and C could directedly inhibit the IKKβ kinase activity with the inhibitory rate of 40%, which were also verified by docking study. Collectively, these results suggested that pulvones A and C's anti-inflammatory effects were relevant to the interruption of NF-κB activation by inhibiting IKKβ kinase.

Parkinson’s disease (PD) is a slowly progressive and complex neurodegenerative disorder. Up to date, there are no approved drugs that could slow or reverse the neurodegenerative process of PD. Here, we reported the synthesis of series of piperine analogues and the evaluation of their neuroprotective effects against hydrogen peroxide (H2O2) induced damage in the neuron-like PC12 cells. Among these analogues, 3b exhibited the most potent protection effect and its underlying mechanism was further investigated. Further results indicated that the ROS scavenging and cytoprotection effect of 3b might be related to the Nrf2 activation and upregulation of related phase II antioxidant enzymes, such as HO-1 and NQO1. In in vivo study, oral administration (100 mg/kg) of 3b significantly attenuated PD-associated behavioral deficits in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mouse model of PD and protected tyrosine hydroxylase-immunopositive dopaminergic neurons. Our results provided evidence that 3b might be a promising candidate for Parkinson’s disease treatment.

EGFR T790 M accounts for 50% to 60% of cases of non-small-cell lung carcinoma (NSCLC) resistance to the first-generation EGFR tyrosine kinase inhibitors (TKIs). Hence, identifying novel compounds with activity against TKIs resistant is of great value. In this study, twenty honokiol and magnolol derivatives were isolated from the EtOH extract of Magnolia officinalis and the antiproliferative activity was evaluated on HCC827 (19del EGFR mutation), H1975 (L858 R/T790 M EGFR mutation), and H460 (KRAS mutation) cell lines. Among the isolated compounds, piperitylmagnolol (a 3-substituted magnolol derivative) showed the best antiproliferative activity against those three cell lines with the IC50 values of 15.85, 15.60 and 18.60 μM, respectively, which provided a direction for the structural modification of magnolol. Further structural modification led to the synthesis of thirty-one magnolol derivatives, and compounds A13, C1, and C2 exhibited significant and broad-spectrum antiproliferative activity with the IC50 values ranging from 4.81 to 13.54 μM, which were approximately 4- and 8-fold more potent than those of honokiol and magnolol, respectively. Moreover, their aqueous solubility was remarkably improved with 12-, 400- and 105 fold greater than those of honokiol and magnolol. Anti-tumor mechanism research revealed that these three compounds were able to induce cell cycle arrest at G0/G1 phase, cause efficient apoptosis in H1975 cells, and also prevent the migration of HUVECs in a dose-dependent manner through Cdk2, Cdk4, Cyclin E, and Cyclin D1 inhibition as well as up-regulation of cleaved-PARP and cleaved-caspase 3 levels. In in vivo antitumor activity, C2 (10, 30 and 100 mg/kg, po) dose-dependently inhibited the tumor growth in H1975 xenograft model with the tumor inhibition rate of 46.3%, 59.3% and 61.2% respectively, suggesting that C2 is a potential oral anticancer agent deserving further investigation.

Inhibitors that bind to the paclitaxel- or vinblastine-binding sites of tubulin have been part of the pharmacopoeia of anticancer therapy for decades. However, tubulin inhibitors that bind to the colchicine-binding site are not used in clinical cancer therapy, because of their low therapeutic index. To address multidrug resistance to many conventional tubulin-binding agents, numerous efforts have attempted to clinically develop inhibitors that bind the colchicine-binding site. Previously, we have found that millepachine (MIL), a natural chalcone-type small molecule extracted from the plant Millettia pachycarpa, and its two derivatives (MDs) SKLB028 and SKLB050 have potential antitumor activities both in vitro and in vivo. However, their cellular targets and mechanisms are unclear. Here, biochemical and cellular experiments revealed that the MDs directly and irreversibly bind β-tubulin. X-ray crystallography of the tubulin–MD structures disclosed that the MDs bind at the tubulin intradimer interface and to the same site as colchicine and that their binding mode is similar to that of colchicine. Of note, MDs inhibited tubulin polymerization and caused G(2)/M cell-cycle arrest. Comprehensive analysis further revealed that free MIL exhibits an s-cis conformation, whereas MIL in the colchicine-binding site in tubulin adopts an s-trans conformation. Moreover, introducing an α-methyl to MDs to increase the proportion of s-trans conformations augmented MDs' tubulin inhibition activity. Our study uncovers a new class of chalcone-type tubulin inhibitors that bind the colchicine-binding site in β-tubulin and suggests that the s-trans conformation of these compounds may make them more active anticancer agents.

Two new lignan derivatives and nine known compounds were isolated from the EtOAc and n-BuOH extracts of Millettia dorwardi Coll. Et. Hemsl. The structures of the compounds were determined by 1D and 2D NMR, CD and HRMS. Then, cytotoxic effects of these compounds were evaluated against five cancer cell lines (HepG2, HCT116, MCF7, Raji and KG-1). Results showed the isolated compounds have no obvious cytotoxic activity, but millepurpan (5) showed moderate growth inhibitory activity against Raij and KG-1 cells, with IC50 values of 36.13 and 30.11 μM, respectively. Further activity evaluation revealed that millepurpan (5) could induce G1 arrest and apoptosis in KG-1 cells. The lignan skeleton was isolated from Millettia genus for the first time and all compounds were firstly reported from Millettia dorwardi Coll. Et.

Idiopathic pulmonary fibrosis (IPF) is a progressive, life-threatening and interstitial lung disease with the median survival of only 3-5 years. However, due to the unclear etiology and problems in accurate diagnosis, up to now only two drugs were approved by FDA for the treatment of IPF and their outcome responses are limited. Numerous studies have shown that TGF-β is the most important cytokine in the development of pulmonary fibrosis and plays a role through its downstream signaling molecule TGF-binding receptor Smads protein. In this paper, compounds bearing 2(1H)-quinolone scaffold were designed and their anti-fibrosis effects were evaluated. Of these compounds, 20f was identified as the most active one and could inhibit TGF-β-induced collagen deposition of NRK-49F cells and mouse fibroblasts migration with comparable activity and lower cytotoxicity than nintedanib in vitro. Further mechanism studies indicated that 20f reduced the expression of fibrogenic phenotypic protein α-SMA and collagen Ⅰ by inhibiting the TGF-β/Smad dependent pathways and ERK1/2 and p38 pathways. Moreover, compared with the nintedanib, 20f (100 mg/kg/day, p.o) more effectively alleviated collagen deposition in lung tissue and delayed the destruction of lung tissue structure both in bleomycin-induced prevention and treatment mice pulmonary fibrosis models. The immunohistochemical experiments further showed that 20f could block the expression level of phosphorylated Smad3 in the lung tissue cells, which resulted in its anti-fibrosis effects in vivo. In addition, 20f demonstrated good bioavailability (F = 41.55% vs 12%, compare with nintedanib) and an appropriate elimination half-life (T1/2 = 3.5 h), suggesting that 20f may be a potential drug candidate for the treatment of pulmonary fibrosis.

Idiopathic pulmonary fibrosis, characterized by excess accumulation of extracellular matrix, involved in many chronic diseases or injuries, threatens human health greatly. We have reported a series of compounds bearing coumarin scaffold which potently inhibited TGF-β-induced total collagen accumulation in NRK-49F cell line and migration of macrophages. Compound 9d also suppressed the TGF-β-induced protein expression of COL1A1, α-SMA, and p-Smad3 in vitro. Meanwhile, 9d at a dose of 100 mg/kg/day through oral administrations for 4 weeks effectively alleviated infiltration of inflammatory cells in lung tissue and fibrotic degree in bleomycin-induced pulmonary fibrosis model, which may related to its inhibition of TGF-β/Smad3 pathway and anti-inflammation efficacy. In addition, 9d demonstrated decent bioavailability (F = 39.88%) and suitable eliminated half-life time (T1/2 = 13.09 h), suggesting that 9d could be a potential drug candidate for the treatment of fibrotic diseases.

Natural products and their derivatives have historically been invaluable as a source of therapeutic agents. Honokiol, as a well-known natural product in Chinese herbal medicine Houpu, is finally being studied in a Phase I clinical trial (CTR20170822) in patients with Advanced Non-Small Cell Lung Cancer (NSCLS) in China this year. During the honokiol liposome formulation process, five major impurities were present in the range of 0.05–0.1% based on the HPLC analysis. These five major impurities were obtained from the forced degradation product of honokiol through countercurrent chromatography and prep-HPLC. The structure were elucidated with 1H NMR, 13C NMR, 2D NMR and MS spectral data. The proposed HPLC method was validated for specificity, linearity (concentration range 0.01–1.62, 0.003–0.96, 0.05–7.98, 0.04–6.52, 0.03–5.18 μg/ml for impurities I–V respectively, R2 > 0.9988), accuracy (99.11–100.67%), precision (CV

In this article, a CE with a new electrochemiluminescent (ECL) detection system was developed. A microfluidic ECL detection cell with less than 0.5 microL dead volumes was developed and used as detector for this system. A hydrofluoric acid-etched porous joint was made at 8 mm from the outlet of the separation capillary to isolate the CE high voltage from ECL detection. The proposed CE-ECL system was applied for separation and detection of some proteins labeled with tris(1,10-phenanthroline) ruthenium(II). High efficiency ECL-enhanced reagent, tripropylamine, was infused to the detection cell as coreactant by a micro-infusion system to obtain maximum and stable ECL signal. The performance of this setup was illustrated by the analysis of tris(1,10-phenanthroline) ruthenium(II)-labeled proteins. The background electrolyte for protein detection was 20 mM Tris-CH3COOH with 2.0% m/m PVP at pH 4.0. Under the optimal conditions, the corresponding LOD were 2.2 x 10(-10) M for HSA, 4.4 x 10(-10) M for casein (alpha-S1) and 5.1 x 10(-10) M for cytochrome c. The proposed method was also successfully used for the trace analysis of albumin in human urine without any pretreatment.