Quaternity method for integrated screening, separation, extraction optimization, and bioactivity evaluation of acetylcholinesterase inhibitors from Sophora flavescens Aiton.

INTRODUCTION: Sophora flavescens Aiton (Fabaceae), a ubiquitous plant species in Asia, contains a wide range of pharmacologically active compounds, such as flavonoids, with potential anti-Alzheimer's disease (anti-AD) effects. OBJECTIVES: The objective of the study is to develop a quaternity method for the screening, isolation, extraction optimization, and activity evaluation of acetylcholinesterase (AChE)-inhibiting compounds from S. flavescens to realize high-throughput screening of active substances in traditional Chinese medicine and to provide experimental data for the development of anti-AD drugs. METHODS: With AChE as the target molecule, affinity ultrafiltration and liquid chromatography-mass spectrometry were applied to screen for potential inhibitors of the enzyme in S. flavescens. Orthogonal array experiments combined with the multi-objective Non-Dominated Sorting Genetic Algorithm III was used for the first time to optimize the process for extracting the active substances. Enzyme inhibition kinetics and molecular docking studies were performed to verify the potential anti-AD effects of the active compounds. RESULTS: Five AChE-inhibiting compounds were identified: kushenol I, kurarinone, sophoraflavanone G, isokurarinone, and kushenol E. These were successfully separated at purities of 72.88%, 98.55%, 96.86%, 96.74%, and 95.84%, respectively, using the n-hexane/ethyl acetate/methanol/water (4.0/5.0/4.0/5.0, v/v/v/v), n-hexane/ethyl acetate/methanol/water (5.0/5.0/6.0/4.0, v/v/v/v), and n-hexane/ethyl acetate/methanol/water (4.9/5.1/5.7/4.3, v/v/v/v) mobile phase systems. Enzyme inhibition kinetics revealed that kushenol E had the best inhibitory effect. CONCLUSION: This study elucidates the mechanism of action of five active AChE inhibitors in S. flavescens and provides a theoretical basis for the screening and development of anti-AD and other therapeutic drugs.

Catalytic conversion of carbohydrates into 5-ethoxymethylfurfural using γ-AlOOH and CeO2@B2O3 catalyst synergistic effect.

Selective catalytic conversion of carbohydrates to 5-ethoxymethylfurfural (EMF) is a critical approach to the biorefinery. In this work, solid acid catalysts of γ-AlOOH and CeO2@B2O3 were used to convert carbohydrates to EMF in a one-pot process, performed in an ethanol/DMSO solvent system. The synergistic effect of γ-AlOOH and CeO2@B2O3 was studied. Furthermore, the morpho-structural properties of the catalysts were characterized, and the effects of reaction time, reaction temperature, catalyst load, and the amount of cosolvent on the conversion of glucose to EMF were examined and optimized. Under the reaction conditions of 170 °C for 20 h, glucose, sucrose, cellobiose, inulin and starch were used as raw materials, and the EMF yield range was 9.2-27.7%. The results showed that the synergistic effect of γ-AlOOH and CeO2@B2O3 further causes the combination of multiple acid sites with different types and strength distributions. Particularly, the collaboration between weak, medium-strong, and strong acid, as well as between Lewis and Brønsted acidity, is of great significance for EMF generation. The reusability experiments showed that the combined catalytic system was easily separated and maintained catalytic activity for five successive reactions without further intermediate regeneration steps. This work provides a promising route for the catalytic conversion of biomass-derived carbohydrates into EMF.

Sulfonic acid-functionalized PCP(Cr) catalysts with Cr3+ and -SO3H sites for 5-ethoxymethylfurfural production from glucose.

5-Ethoxymethylfurfural (EMF) has been identified as a potential biofuel and fuel additive, for which the production from glucose (the most abundant and inexpensive monosaccharide) in a one-step process would be highly desirable. Here, the synthesis of sulfonic acid-functionalized porous coordination polymers (PCPs) and their application as catalysts for EMF synthesis are reported. PCP(Cr)-BA (PCP material with Cr3+ ions and H2BDC-SO3H linkers) and PCP(Cr)-NA (PCP material with Cr3+ ions and H2NDC(SO3H)2 linkers) materials containing both Cr3+ sites and Brønsted-acidic -SO3H sites were prepared. The morphology, pore structure, acidity, chemical composition, and thermal stability of the two functionalized PCP(Cr) catalysts were analyzed by systematic characterization. The catalysts featured a porous morphology and dual Cr3+ and -SO3H sites, which enabled the cascade conversion of glucose to EMF. PCP(Cr)-BA exhibited higher performance than PCP(Cr)-NA with an EMF yield of 23.1% in the conversion of glucose at 140 °C after 22 h in an ethanol/water system. In addition, the as-prepared catalyst exhibited a high stability in the current catalytic system for EMF production from glucose with a constant catalytic activity in a four-run recycling test without an intermediate regeneration step.

Adsorption of copper(II) and lead(II) from seawater using hydrothermal biochar derived from Enteromorpha.

The objective of this research was to evaluate the capacity of Enteromorpha derived biochar to adsorb heavy metals from seawater. The biochar characteristics were determined, and isothermal and kinetic data were obtained using batch experiments. Copper [Cu(II)] and lead [Pb(II)] adsorption by the biochar was favored by high pH conditions, while elevated salinity had a relatively weak negative effect on adsorption. The Langmuir isotherm and adsorption kinetics pattern enabled interpretation of the equilibrium and kinetics of Cu(II) and Pb(II) removal by the biochar. The maximum removal rates of Cu(II) and Pb(II) by the biochar in 60 min were estimated to be 91% and 54%, respectively. A model describing the adsorption processes was developed to predict the efficiency of heavy metal removal by the biochar. The outcomes of the present study indicate that Enteromorpha derived biochar could be an effective and environmentally friendly adsorbent for removing heavy metals from marine environments.

Transformation of corncob into furfural by a bifunctional solid acid catalyst.

A transformation route was developed for the conversion of raw corncob into furfural by a Clbearing solid acid catalyst (HSCSO3H) prepared by the hydrothermal carbonization and sulfonation of sucralose. The catalytic performances of HSCSO3H in selected solvents were demonstrated and optimized, where a furfural yield of 90.8 mol% (20.9 wt%) was achieved at 448 K in 30 min in γ-valerolactone/water system. Interestingly, significant furfural yields were also obtained from cellulose. The effect of elevated temperature on furfural yield from high initial feedstock loading was also investigated. HSCSO3H with COOH, phenolicOH, and Cl as binding sites and SO3H as the catalytic site on its surface presents a bifunctional catalyst, and synergic effects of these functional groups, reaction solvent property and temperature are made responsible for the good catalytic performances. The catalytic strategy proposed in this study demonstrated an effective transformation of corncob into furfural with a high yield.

Distribution, bioaccumulation and trophic transfer of chlorinated polyfluoroalkyl ether sulfonic acids in the marine food web of Bohai, China.

Chlorinated polyfluoroalkyl ether sulfonic acids (Cl-PFESAs) caused great concerns recently as novel fluorinated alternatives. However, information on their bioconcentration, bioaccumulation and biomagnification in marine ecosystems is limited. In this study, 152 biological samples including invertebrates, fishes, seabirds and mammals collected from Bohai Sea of China were analyzed to investigate the residual level, spatial distribution, bioaccumulation and biomagnification of Cl-PFESAs. 6:2 Cl-PFESA was found in concentrations ranging from

Occurrence and seasonal variations of per- and polyfluoroalkyl substances (PFASs) including fluorinated alternatives in rivers, drain outlets and the receiving Bohai Sea of China.

A simultaneous sampling campaign was undertaken to study the pollution by 21 per- and polyfluoroalkyl substances (PFASs) in rivers, drain outlets and their receiving Bohai Sea of China. Chlorinated polyfluoroalkyl ether sulfonic acids (Cl-PFESAs) are being used as fluorinated alternatives and they were included in this study. In comparison with other regions and countries, high concentrations of ∑21PFASs in seawater samples from the Bohai Sea, ranging from 5.03 to 41 700 ng/L (median: 64.8 ng/L), were observed. The spatial distribution of PFAS levels in this sea area was in the ranking of Laizhou Bay > Liaodong Bay > Bohai Bay > other sea areas. By comparing the levels and composition profiles of PFASs in the seawater and their sources (rivers and drain outlets), it was concluded that rivers and drain outlets are the primary sources of PFAS contamination to the Bohai Sea. These PFAS levels varied seasonally among the rivers and drain outlets, but statistically significant changes were not observed. Levels of 6:2 and 8:2 Cl-PFESAs in rivers, drain outlets and receiving sea were firstly reported in the present study. Relatively high concentrations of 6:2 Cl-PFESA were found in drain outlets, ranging from below method limits of quantification (MLQ) to 7600 ng/L, but 8:2 Cl-PFAES detection was infrequent and all median concentration below MLQ. Mass discharges to the sea of 6:2 Cl-PFESA from rivers and drain outlets to the sea were estimated to be 37 and 17 kg/y, respectively.

Occurrence and inputs of perfluoroalkyl substances (PFASs) from rivers and drain outlets to the Bohai Sea, China.

Concentrations of 19 PFASs in riverwater, coastal wastewater and effluents from WWTPs which were directly discharged into the Bohai Sea of China were measured and their inputs to this sea area were calculated accordingly. For riverwater samples, the total PFAS concentrations ranged from 13.1 to 69 238 ng/L. PFAS levels in riverwater collected from Liaoning Province were comparable to those from Shandong Province, while they were two orders of magnitude greater than those from Hebei Province and the city of Tianjin. The dominant PFAS patterns were spatially different. PFBS and PFOA were the predominant PFASs in riverwater samples at sites where fluorochemical industry parks are located in Liaoning Province and Shandong Province, respectively. For other sites, PFOA and PFOS were the most abundant PFASs. In contrast, the total PFAS concentrations in coastal wastewater and effluent samples ranged from 16.7 to 7 522 ng/L and from 13.1 to 319 ng/L, respectively. PFOA was dominant in these samples. Inputs of PFASs to the Bohai Sea via riverine flow, discharge of coastal wastewater and effluents were estimated to be 87.3 tons per year. As compared with coastal wastewater and effluent discharge, riverine input was a major source for the PFAS pollution in the Bohai Sea except for PFBS.

Occurrence, spatial and temporal distributions of perfluoroalkyl substances in wastewater, seawater and sediment from Bohai Sea, China.

In this study, surface seawater and sediment samples were collected from Bohai Sea, China, to investigate the temporal and spatial distributions of perfluoroalkyl substances (PFASs) in that area. The total concentrations of PFASs in seawater and sediment samples ranged from below method limit of quantification (

Levels and spatial distribution of perfluoroalkyl substances in China Liaodong Bay basin with concentrated fluorine industry parks.

Eighteen different perfluoroalkyl substances (PFASs) were investigated in 35 river water samples and 34 sediment samples collected from rivers in the Liaodong Bay basin containing two fluorine industry parks. Perfluorooctanoate (PFOA) and perfluorooctane sulfonate (PFOS) were the predominant PFASs in freshwater, with median concentrations of 26.5 ng/L and 1.87 ng/L, respectively. However, perfluorobutane sulfonate (PFBS) had the highest maximum concentration (up to 124.1 ng/L, approximately two orders of magnitude higher) in water at a site which is the nearest to the industrial source of PFASs. Total PFASs in water at this site were also the highest. In contrast, PFOA and perfluorooctadecanoate (PFOcDA) were the most abundant PFASs in sediment, with median concentrations of 1.19 ng/g and 0.35 ng/g, respectively. Total PFAS concentrations in sediment from the site near to the industrial park were significantly higher than the other rivers. Mass loading of total PFASs from the rivers flowing into Liaodong Bay was estimated to be 506 kg/year.

[Screen and validation of differentially expressing genes related to silicotic pulmonary fibrosis in rats].

OBJECTIVE: To screen the differentially expressing genes between silicotic lung tissue and normal lung tissue, to identify the differentially expressing genes of matrix metalloproteinase-12 (MMP-12) and Cathepsin E and to explore the roles of those genes in silicosis development. METHODS: Thirty male SD rats were divided randomly into two groups: control group (6 rats) and exposure group (24 rats) which was exposed to SiO2 by intra-tracheal perfusion. On the 30 th, 60 th and 90 th days after exposure, 8 rats in model group and 2 rats in control group were executed and the lung tissues were obtained. The morphologic changes of lung tissues were observed with HE staining and VG staining under a light microscope. The gene microarrays were used to identify differentially expressing genes of lung tissues in rats exposed to SiO2 for 60 days. Two significantly up-regulated genes, MMP-12 and Cathepsin E, were validated using RT-PCR, immunohistochemistry and Western Blot assay. RESULTS: A total of 338 differentially expressing genes were identified from the 26 962 genes between silicotic rats and normal rats, including 267 up-regulated genes and 71 down-regulated genes. The results of RT-PCR showed that in the lung tissues of exposure group on the 30 th, 60 th and 90 th days, the mRNA expression levels of MMP-12 were 4.306, 5.338, 6.713 times higher than those in the control group, the mRNA expression levels of Cathepsin E were 1.434, 2.974, 3.889 times higher than those in the control group, respectively. The results of immunohistochemical showed that in the lung tissues of exposure group on the 30th, 60th and 90th days, the mRNA expression levels of MMP-12 were 1.435, 1.746, 2.069 times higher than those in the control group, the mRNA expression levels of Cathepsin E were 1.372, 1.663, 2.103 times higher than those in the control group, respectively. The results of immunohistochemical showed that in the lung tissues of exposure group on the 30th, 60th and 90th days, the expression levels of MMP-12 protein were 1.214, 1.531, 1.959 times higher than those in the control group, the expression levels of Cathepsin E protein were 1.262, 1.828, 1.907 times higher than those in the control group, respectively. Compared with the control group, the mRNA and protein expression levels of MMP-12 and Cathepsin E in lung tissues of exposure group were significantly up-regulated (P < 0.05). CONCLUSION: The differentially expressing genes in rat lung tissues screened by gene chip were validated, which suggested that a complex gene regulatory network may be contributed to occurrence of silicosis. MMP-12 and Cathepsin E genes may be involved in the development of silicotic pulmonary fibrosis by degrading the basement membrane of alveolar wall and participating in the immune response.

[Expression of vascular endothelial growth factor and microvessel density in rabbit radius defects repaired with allogeneic and autogenic bone].

OBJECTIVE: To investigate the expression levels and significance of vascular endothelial growth factor (VEGF) and microvessel density (MVD) in rabbit radius defects repaired with allogeneic and autogenic bone. METHODS: Forty adult New Zealand rabbits were chosen, and 10 mm bone defect model was created in the bilateral radii of 28 experimental rabbits. The other 12 rabbits provided allogeneic bone under the standard of American Association of Tissue Bank. In the left side, allogeneic bone were used to repair bone defect (experimental group), equal capacity autogenous iliac bone was used in the right side (control group). Animals were sacrificed at 2, 4, 8, and 12 weeks postoperatively. Immunohistochemical method was used to determine the expression of VEGF, CD34 protein and MVD counting. Bone histomorphometric parameters, including percent trabecular area (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th), and trabecular separation (Tb.Sp) were measured by von Kossa staining undecalcified slices. The relation was analyzed between VEGF and MVD, histomorphometric parameters. RESULTS: The positive signals of VEGF protein were detected in cytoplasm of vascular endothelial cells, chondrocytes, osteoblasts, fibroblasts and osteoclasts. At 2 weeks, there was no significant difference in VEGF protein expression between experimental group and control group (P > 0.05); at 4 and 8 weeks, the expression of VEGF in control group was significantly higher than that in experimental group (P < 0.05); and at 12 weeks, there was no significant difference between two groups (P > 0.05). There was a positive correlation (P < 0.01) between VEGF expression and MVD value in two groups at 2, 4, 8, and 12 weeks postoperatively. There was no significant difference in bone histomorphometric parameters (BV/TV, Tb.Th, Tb.N, Tb.Sp) between two groups at 12 weeks postoperatively (P > 0.05), but there was a positive correlation between VEGF expression and parameters of BV/TV, Tb.Th, and Tb.N (P < 0.01); and a negative correlation between VEGF and Tb.Sp (P < 0.01). CONCLUSION: VEGF can express diversity at different time and positions, and the different expressions indicated various biology significances in the process of the bone healing. It can coordinate growth of cartilage and bone and profit vascular reconstruction of allogeneic bone. VEGF may participate in promoting osteogenesis in the course of allogeneic bone transplantation.

[Experimental study on adhesiveness of osteoblasts and vascular endothelial cells from rat BMSCs co-cultured on allogeneic freeze-dried partially bone in vitro].

OBJECTIVE: To investigate the adhesiveness of osteoblasts and vascular endothelial cells from rat BMSCs. METHODS: The BMSCs were isolated from 4-week-old SD co-cultured on allogeneic freeze-dried partially bone in vitro. rats (weighing 100-110 g) and cultured in vitro. The third generation of BMSCs were induced into osteoblasts and vascular endothelial cells. The osteoblasts and vascular endothelial cells after being induced for 7 days in a ratio of 1 to 1 were directly co-cultured (experimental group), while the second generation of uninduced BMSCs was used as a control (control group). The growth and proliferation ability were analyzed by MTT examination and the growth curve was drawn at 1-8 days. The osteoblasts and vascular endothelial cells after being induced for 14 days were implanted in the allogeneic freeze-dried partially bone coated by 20% Col I or not at different densities (0.25 x 10(6)/mL, 0.50 x 10(6)/mL, 1.00 x 10(6)/mL, 2.00 x 10(6)/mL, 4.00 x 10(6)/mL), as modified group and unmodified group, the cell adherence rate was calculated after 24 hours. These two kinds of cells were implanted in the pre-disposal treated allogeneic freeze-dried partially bone and observed by scanning electron microscope. RESULTS: ALP staining of osteoblasts showed that there were blue grains in cytoplasm at 7 days. CD31 and CD34 immunocytochemical staining of vascular endothelial cell showed that there were positive signals in the cytoplasm at 14 days. The MTT test showed that the proliferation level of the experimental group was lower than those of the control group. There were significant differences in absorbance value between two group from 3 days to 8 days (P < 0.05). The cell adherence rate increased with increasing seeding density when the seeding density was (0.25-1.00) x 10(6)/mL. The cell adherence rate reached the peak when the seeding density was 1.00 x 10(6)/mL. The cell adherence rate decreased when the seeding density was more than 2.00 x 106/mL. There were significant differences in cell adherence rate between modified group and unmodified group at different seeding densities (P < 0.05). The proliferation of the osteoblasts and endothelial cells presented better growth and histocompatibility under scanning electron microscope. CONCLUSION: The growing behavior of two kinds of cells is good in the allogeneic freeze-dried partially bone coated by 20% Col I, which can be used in reconstruction of vascularized tissue engineered bone.