Molecules

1621 KiB

Open AccessArticle

Time-dependent Inhibition of CYP2C8 and CYP2C19 by Hedera helix Extracts, A Traditional Respiratory Herbal Medicine

by Shaheed Ur Rehman, In Sook Kim, Min Sun Choi, Seung Hyun Kim, Yonghui Zhang and Hye Hyun Yoo

Molecules 2017, 22(7), 1241; https://doi.org/10.3390/molecules22071241 - 24 Jul 2017

Cited by 8 | Viewed by 7033

The extract of Hedera helix L. (Araliaceae), a well-known folk medicine, has been popularly used to treat respiratory problems, worldwide. It is very likely that this herbal extract is taken in combination with conventional drugs. The present study aimed to evaluate the effects [...] Read more.

The extract of Hedera helix L. (Araliaceae), a well-known folk medicine, has been popularly used to treat respiratory problems, worldwide. It is very likely that this herbal extract is taken in combination with conventional drugs. The present study aimed to evaluate the effects of H. helix extract on cytochrome P450 (CYP) enzyme-mediated metabolism to predict the potential for herb–drug interactions. A cocktail probe assay was used to measure the inhibitory effect of CYP. H. helix extracts were incubated with pooled human liver microsomes or CYP isozymes with CYP-specific substrates, and the formation of specific metabolites was investigated to measure the inhibitory effects. H. helix showed significant inhibitory effects on CYP2C8, CYP2C19 and CYP2D6 in a concentration-dependent manner. In recombinant CYP2C8, CYP2C19 and CYP2D6 isozymes, the IC₅₀ values of the extract were 0.08 ± 0.01, 0.58 ± 0.03 and 6.72 ± 0.22 mg/mL, respectively. Further investigation showed that H. helix extract has a positive time-dependent inhibition property on both CYP2C8 and CYP2C19 with IC₅₀ shift value of 2.77 ± 0.12 and 6.31 ± 0.25, respectively. Based on this in vitro investigation, consumption of herbal medicines or dietary supplements containing H. helix extracts requires careful attention to avoid any CYP-based interactions. Full article

(This article belongs to the Special Issue Herbal Remedies Meet Modern Day Medicine: Challenges and Opportunities)

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Figure 1
Representative multiple reaction monitoring (MRM) chromatograms of human liver microsome samples of, (A) control; (B) H. helix extract-treated, and (C) Hederacoside C (HDC)-treated. A fraction of human liver microsomal was incubated with the substrate mixture, in an NADPH-generating system, and H. helix extract (2.5 mg/mL) or HDC (100 µM) for 30 min and the cytochromes P450 (CYP)-specific metabolite formation was determined by LC-MS/MS. (a) Acetaminophen; (b) 7-OH coumarin; (c) 6-OH-paclitaxel (d) 4-OH-diclofenac; (e) 4-OH-mephenytoin; (f) dextrorphan; (g) 1-OH-madazolam; and (h) terfenadine (IS). Full article ">Figure 2
Effects of H. helix extract and hederacoside C on the CYP-specific metabolite formation in human liver microsomes. (A) H. helix extract (5 mg/mL); (B) Hederacoside C (500 μM); (C) Effects of H. helix extract on the metabolic activities of CYP2C8, CYP2C19, and CYP2D6; and (D) Effects of hederacoside C on the metabolic activities of CYP2C8, CYP2C19, and CYP2D6. Data was presented as the mean ± S.D. of the data obtained from two independent experiments. Full article ">Figure 2 Cont.
Effects of H. helix extract and hederacoside C on the CYP-specific metabolite formation in human liver microsomes. (A) H. helix extract (5 mg/mL); (B) Hederacoside C (500 μM); (C) Effects of H. helix extract on the metabolic activities of CYP2C8, CYP2C19, and CYP2D6; and (D) Effects of hederacoside C on the metabolic activities of CYP2C8, CYP2C19, and CYP2D6. Data was presented as the mean ± S.D. of the data obtained from two independent experiments. Full article ">Figure 3
CYP-specific metabolite formation as the percent of control after co-incubation and pre-incubation of H. helix extracts with and without NADPH in c-DNA expressed CYP isozymes; (A) CYP2C8; (B) CYP2C19, and (C) CYP2D6. Data was presented as the mean ± S.D. of the data obtained from two independent experiments. Full article ">Figure 3 Cont.
CYP-specific metabolite formation as the percent of control after co-incubation and pre-incubation of H. helix extracts with and without NADPH in c-DNA expressed CYP isozymes; (A) CYP2C8; (B) CYP2C19, and (C) CYP2D6. Data was presented as the mean ± S.D. of the data obtained from two independent experiments. Full article ">Figure 4
Hederacoside C, (A) product ion mass spectra of the [M − H]− ions of HDC, and (B) MRM chromatograms of HDC in H. helix extracts. Full article ">

1006 KiB

Open AccessFeature PaperReview

Role of G Protein-Coupled Receptors in the Regulation of Structural Plasticity and Cognitive Function

by Crystal C. Y. Leung and Yung H. Wong

Molecules 2017, 22(7), 1239; https://doi.org/10.3390/molecules22071239 - 24 Jul 2017

Cited by 35 | Viewed by 15607

Abstract

Cognition and other higher brain functions are known to be intricately associated with the capacity of neural circuits to undergo structural reorganization. Structural remodelling of neural circuits, or structural plasticity, in the hippocampus plays a major role in learning and memory. Dynamic modifications [...] Read more.

Cognition and other higher brain functions are known to be intricately associated with the capacity of neural circuits to undergo structural reorganization. Structural remodelling of neural circuits, or structural plasticity, in the hippocampus plays a major role in learning and memory. Dynamic modifications of neuronal connectivity in the form of dendritic spine morphology alteration, as well as synapse formation and elimination, often result in the strengthening or weakening of specific neural circuits that determine synaptic plasticity. Changes in dendritic complexity and synapse number are mediated by cellular processes that are regulated by extracellular signals such as neurotransmitters and neurotrophic factors. As many neurotransmitters act on G protein-coupled receptors (GPCRs), it has become increasingly apparent that GPCRs can regulate structural plasticity through a myriad of G protein-dependent pathways and non-canonical signals. A thorough understanding of how GPCRs exert their regulatory influence on dendritic spine morphogenesis may provide new insights for treating cognitive impairment and decline in various age-related diseases. In this article, we review the evidence of GPCR-mediated regulation of structural plasticity, with a special emphasis on the involvement of common as well as distinct signalling pathways that are regulated by major neurotransmitters. Full article

(This article belongs to the Special Issue G-protein Coupled Receptor Structure and Function)

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1931 KiB

Open AccessArticle

Quality Assessment of Gentiana rigescens from Different Geographical Origins Using FT-IR Spectroscopy Combined with HPLC

by Zhe Wu, Yanli Zhao, Ji Zhang and Yuanzhong Wang

Molecules 2017, 22(7), 1238; https://doi.org/10.3390/molecules22071238 - 24 Jul 2017

Cited by 30 | Viewed by 5528

Abstract

Gentiana rigescens is a precious herbal medicine in China because of its liver-protective and choleretic effects. A method for the qualitative identification and quantitative evaluation of G. rigescens from Yunnan Province, China, has been developed employing Fourier transform infrared (FT-IR) spectroscopy and high [...] Read more.

Gentiana rigescens is a precious herbal medicine in China because of its liver-protective and choleretic effects. A method for the qualitative identification and quantitative evaluation of G. rigescens from Yunnan Province, China, has been developed employing Fourier transform infrared (FT-IR) spectroscopy and high performance liquid chromatography (HPLC) with the aid of chemometrics such as partial least squares discriminant analysis (PLS-DA) and support vector machines (SVM) regression. Our results indicated that PLS-DA model could efficiently discriminate G. rigescens from different geographical origins. It was found that the samples which could not be determined accurately were in the margin or outside of the 95% confidence ellipses. Moreover, the result implied that geographical origins variation of root samples were more obvious than that of stems and leaves. The quantitative analysis was based on gentiopicroside content which was the main active constituent in G. rigescens. For the prediction of gentiopicroside, the performances of model based on the parameters selected through grid search algorithm (GS) with seven-fold cross validation were better than those based on genetic algorithm (GA) and particle swarm optimization algorithm (PSO). For the SVM-GS model, the result was satisfactory. FT-IR spectroscopy coupled with PLS-DA and SVM-GS can be an alternative strategy for qualitative identification and quantitative evaluation of G. rigescens. Full article

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4033 KiB

Open AccessArticle

Synthesis and Biological Evaluation of Ginsenoside Compound K Derivatives as a Novel Class of LXRα Activator

by Yan Huang, Hongmei Liu, Yingxian Zhang, Jin Li, Chenping Wang, Li Zhou, Yi Jia and Xiaohui Li

Molecules 2017, 22(7), 1232; https://doi.org/10.3390/molecules22071232 - 24 Jul 2017

Cited by 24 | Viewed by 4855

Abstract

Compound K is one of the active metabolites of Panaxnotoginseng saponins, which could attenuate the formation of atherosclerosis in mice modelsvia activating LXRα. We synthesized and evaluated a series of ginsenoside compound K derivatives modified with short chain fatty acids. All of the [...] Read more.

Compound K is one of the active metabolites of Panaxnotoginseng saponins, which could attenuate the formation of atherosclerosis in mice modelsvia activating LXRα. We synthesized and evaluated a series of ginsenoside compound K derivatives modified with short chain fatty acids. All of the structures of this class of ginsenoside compound K derivative exhibited comparable or better biological activity than ginsenoside compound K. Especially structure 1 exhibited the best potency (cholesteryl ester content: 41.51%; expression of ABCA1 mRNA: 319%) and low cytotoxicity. Full article

(This article belongs to the Section Medicinal Chemistry)

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2191 KiB

Open AccessArticle

Study on Chemical Profile and Neuroprotective Activity of Myrica rubra Leaf Extract

by Pinghong Chen, Xianzong Lin, Ching-Hsu Yang, Xu Tang, Yu-Wei Chang, Weibing Zheng, Lianzhong Luo, Changan Xu and Yung-Husan Chen

Molecules 2017, 22(7), 1226; https://doi.org/10.3390/molecules22071226 - 24 Jul 2017

Cited by 11 | Viewed by 5480

Abstract

The chemical profile of Myrica rubra (a native species in China) leaf extract was investigated by UPLC-PDA-HRMS, and the neuroprotective activity of two characteristic constituents, myricanol and myricetrin, was evaluated with N2a cells using H₂O₂-inducedoxidative challenge through a series [...] Read more.

The chemical profile of Myrica rubra (a native species in China) leaf extract was investigated by UPLC-PDA-HRMS, and the neuroprotective activity of two characteristic constituents, myricanol and myricetrin, was evaluated with N2a cells using H₂O₂-inducedoxidative challenge through a series of methods, e.g., MTT assay, ROS assay and [Ca²⁺]i assay. Among the 188 constituents detected in the extract of Myrica rubra leaf, 116 were identified definitely or tentatively by the comprehensive utilization of precise molecular weight and abundant multistage fragmentation information obtained by quadrupole orbitrap mass spectrometry. In addition, 14 potential new compounds were reported for the first time. This work established an example for the research of microconstituents in a complex analyte and revealed that suppression of H₂O₂-induced cytotoxicity in N2a cells was achieved by the pretreatment with myricanol. The evidence suggested myricanol may potentially serve as a remedy for prevention and therapy of neurodegenerative diseases induced by oxidative stress. Full article

(This article belongs to the Section Natural Products Chemistry)

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The schematic diagram of the proposed approach. Full article ">Figure 2
Total ion chromatograms (TICs) of the extract of Myrica rubra by UPLC-PDA-HRMS. (A) Negative ion mode; (B) positive ion mode. Full article ">Figure 3
Chemical structures of some constituents identified in the extract of Myrica rubra. Full article ">Figure 4
Identification of flavonoids isolated from Myrica rubra leaf extract according to their structure units. Full article ">Figure 5
Effect of the whole extract (20 μg/mL), myricetrin (0.65 mM) and myricanol at various concentrations on H2O2-induced oxidative stress in N2a cells by MTT assay. Full article ">Figure 6
Effect of myricanol on H2O2-induced cell morphological changes. Full article ">Figure 7
Effect of myricanol and myricetrin on intracellular ROS in N2a cells. Myricanol and myricetrin at the concentration of 0.84 mM and 0.65 mM, respectively. Full article ">

5444 KiB

Open AccessArticle

Neighbor Affinity-Based Core-Attachment Method to Detect Protein Complexes in Dynamic PPI Networks

by Xiujuan Lei and Jing Liang

Molecules 2017, 22(7), 1223; https://doi.org/10.3390/molecules22071223 - 24 Jul 2017

Cited by 7 | Viewed by 5876

Abstract

Protein complexes play significant roles in cellular processes. Identifying protein complexes from protein-protein interaction (PPI) networks is an effective strategy to understand biological processes and cellular functions. A number of methods have recently been proposed to detect protein complexes. However, most of methods [...] Read more.

Protein complexes play significant roles in cellular processes. Identifying protein complexes from protein-protein interaction (PPI) networks is an effective strategy to understand biological processes and cellular functions. A number of methods have recently been proposed to detect protein complexes. However, most of methods predict protein complexes from static PPI networks, and usually overlook the inherent dynamics and topological properties of protein complexes. In this paper, we proposed a novel method, called NABCAM (Neighbor Affinity-Based Core-Attachment Method), to identify protein complexes from dynamic PPI networks. Firstly, the centrality score of every protein is calculated. The proteins with the highest centrality scores are regarded as the seed proteins. Secondly, the seed proteins are expanded to complex cores by calculating the similarity values between the seed proteins and their neighboring proteins. Thirdly, the attachments are appended to their corresponding protein complex cores by comparing the affinity among neighbors inside the core, against that outside the core. Finally, filtering processes are carried out to obtain the final clustering result. The result in the DIP database shows that the NABCAM algorithm can predict protein complexes effectively in comparison with other state-of-the-art methods. Moreover, many protein complexes predicted by our method are biologically significant. Full article

(This article belongs to the Special Issue Computational Analysis for Protein Structure and Interaction)

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2951 KiB

Open AccessArticle

A Novel and Practical Chromatographic “Fingerprint-ROC-SVM” Strategy Applied to Quality Analysis of Traditional Chinese Medicine Injections: Using KuDieZi Injection as a Case Study

by Bin Yang, Yuan Wang, Lanlan Shan, Jingtao Zou, Yuanyuan Wu, Feifan Yang, Yani Zhang, Yubo Li and Yanjun Zhang

Molecules 2017, 22(7), 1237; https://doi.org/10.3390/molecules22071237 - 23 Jul 2017

Cited by 27 | Viewed by 6135

Abstract

Fingerprinting is widely and commonly used in the quality control of traditional Chinese medicine (TCM) injections. However, current studies informed that the fingerprint similarity evaluation was less sensitive and easily generated false positive results. For this reason, a novel and practical chromatographic “Fingerprint-ROC-SVM” [...] Read more.

Fingerprinting is widely and commonly used in the quality control of traditional Chinese medicine (TCM) injections. However, current studies informed that the fingerprint similarity evaluation was less sensitive and easily generated false positive results. For this reason, a novel and practical chromatographic “Fingerprint-ROC-SVM” strategy was established by using KuDieZi (KDZ) injection as a case study in the present article. Firstly, the chromatographic fingerprints of KDZ injection were obtained by UPLC and the common characteristic peaks were identified with UPLC/Q-TOF-MS under the same chromatographic conditions. Then, the receiver operating characteristic (ROC) curve was used to optimize common characteristic peaks by the AUCs value greater than 0.7. Finally, a support vector machine (SVM) model, with the accuracy of 97.06%, was established by the optimized characteristic peaks and applied to monitor the quality of KDZ injection. As a result, the established model could sensitively and accurately distinguish the qualified products (QPs) with the unqualified products (UPs), high-temperature processed samples (HTPs) and high-illumination processed samples (HIPs) of KDZ injection, and the prediction accuracy was 100.00%, 93.75% and 100.00%, respectively. Furthermore, through the comparison with other chemometrics methods, the superiority of the novel analytical strategy was more prominent. It indicated that the novel and practical chromatographic “Fingerprint-ROC-SVM” strategy could be further applied to facilitate the development of the quality analysis of TCM injections. Full article

(This article belongs to the Collection Herbal Medicine Research)

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Open AccessArticle

A Novel Brominated Alkaloid Securidine A, Isolated from the Marine Bryozoan Securiflustra securifrons

by Priyanka Michael, Kine Ø. Hansen, Johan Isaksson, Jeanette H. Andersen and Espen Hansen

Molecules 2017, 22(7), 1236; https://doi.org/10.3390/molecules22071236 - 23 Jul 2017

Cited by 12 | Viewed by 5596

Abstract

A novel brominated alkaloid, Securidine A, was isolated from the cold water marine bryozoan Securiflustra securifrons. Securidine A was isolated using semi-preparative HPLC, and the structure was elucidated by spectroscopic methods. The isolated Securidine A was tested for cytotoxic, antibacterial, and anti-diabetic [...] Read more.

A novel brominated alkaloid, Securidine A, was isolated from the cold water marine bryozoan Securiflustra securifrons. Securidine A was isolated using semi-preparative HPLC, and the structure was elucidated by spectroscopic methods. The isolated Securidine A was tested for cytotoxic, antibacterial, and anti-diabetic activities as well as for its potential for inhibition of biofilm formation. No significant biological activity was observed in the applied bioassays, thus expanded bioactivity profiling is required, in order to reveal any potential applications for Securidine A. Full article

(This article belongs to the Section Natural Products Chemistry)

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4254 KiB

Open AccessArticle

Molecular Dynamics Simulations of the Host Defense Peptide Temporin L and Its Q3K Derivative: An Atomic Level View from Aggregation in Water to Bilayer Perturbation

by Andrea Farrotti, Paolo Conflitti, Saurabh Srivastava, Jimut Kanti Ghosh, Antonio Palleschi, Lorenzo Stella and Gianfranco Bocchinfuso

Molecules 2017, 22(7), 1235; https://doi.org/10.3390/molecules22071235 - 22 Jul 2017

Cited by 13 | Viewed by 6946

Abstract

Temporin L (TempL) is a 13 residue Host Defense Peptide (HDP) isolated from the skin of frogs. It has a strong affinity for lipopolysaccharides (LPS), which is related to its high activity against Gram-negative bacteria and also to its strong tendency to neutralize [...] Read more.

Temporin L (TempL) is a 13 residue Host Defense Peptide (HDP) isolated from the skin of frogs. It has a strong affinity for lipopolysaccharides (LPS), which is related to its high activity against Gram-negative bacteria and also to its strong tendency to neutralize the pro-inflammatory response caused by LPS release from inactivated bacteria. A designed analog with the Q3K substitution shows an enhancement in both these activities. In the present paper, Molecular Dynamics (MD) simulations have been used to investigate the origin of these improved properties. To this end, we have studied the behavior of the peptides both in water solution and in the presence of LPS lipid-A bilayers, demonstrating that the main effect through which the Q3K substitution improves the peptide activities is the destabilization of peptide aggregates in water. Full article

(This article belongs to the Special Issue Biomolecular Simulations)

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Graphical abstract

Graphical abstract
Full article ">Figure 1
Final structures of the MD simulations in water. For each peptide (TempL, F5,8L-TempL, F5,8A-TempL and Q3K-TempL from the top) the structures from two independent simulations are reported. Peptides are represented as cyan ribbon; and residues 3, 5 and 8 are reported in stick representation (blue for Q/K residues at position 3 and red for the F/L/A at positions 5 and 8). Full article ">Figure 2
PMF profiles of the insertion of TempL (black) and Q3K-TempL (red) into a bilayer formed by Lipid-A. As reference, the density profiles of different parts of an unperturbed lipid-A bilayer are also reported. In particular, the water density profile is reported in blue and that of lipid tails and phosphate groups are reported in gray and orange, respectively. Roughly, the two peaks of the phosphates represent the boundaries of the membrane environment. Full article ">Figure 3
Representative structures for each one of the three clusters that characterize the configurations sampled in the global minimum of the PMF profile, as obtained using the g_cluster tool in the GROMACS software package. The structures reported above refer to the TempL peptide, those reported below to Q3K-TempL. The phosphorus atoms of the lipid-A, marking the position of the lipid polar heads, are indicated as gold spheres. The peptide backbone is reported as a cyan ribbon, the water molecules and the third residue of the peptides (Q and K in TempL and Q3K-TempL, respectively) are reported as sticks, with O, N and H atoms colored in red, blue and white, respectively. For the sake of clarity, the lipid tails are not reported. Full article ">Figure 4
Density profile along the axis perpendicular to the bilayer plane (Z axis) calculated on the structures in the absolute minimum of the PMF profile. The reported profiles refer to water (broken blue, solid blue and solid cyan for unperturbed bilayer, with TempL and with Q3K-TempL, respectively), lipid-A (broken gray, solid gray and solid light gray for unperturbed bilayer, with TempL and with Q3K-TempL, respectively), the saccharide moiety of lipid-A (broken dark green, solid dark green and solid light green for unperturbed bilayer, with TempL and with Q3K-TempL, respectively). The black and red lines refer respectively to the whole TempL and Q3K-TempL peptides (below) and to the Q3 and K3 side chain residues (above). To highlight the effects of the mutated residue, the graph above reports a magnification of the central region of the density profile. Full article ">Figure 5
Deuterium order parameter for the six lipid chains of the lipid-A calculated starting from the structures in the global minimum of the PMF profile. Black, red and dashed-gray lines refer to TempL, Q3K-TempL, and unperturbed bilayer, respectively. Full article ">Scheme 1
(Left) Helical wheel representation of TempL: In the scheme, the hydrophilic, hydrophobic and potentially positively charged residues are reported as circles, diamonds and pentagons, respectively. The third residue is marked with a red bar. (Right) View along the helix-axis of the TempL 3D helical structure: The side chains of the leucine, isoleucine, valine, phenylalanine and tryptophan residues are reported as green sticks; those of arginine and lysine are reported in blue; and those of glutamine and serine in violet. Full article ">

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Open AccessArticle

New Cinchona Oximes Evaluated as Reactivators of Acetylcholinesterase and Butyrylcholinesterase Inhibited by Organophosphorus Compounds

by Maja Katalinić, Antonio Zandona, Alma Ramić, Tamara Zorbaz, Ines Primožič and Zrinka Kovarik

Molecules 2017, 22(7), 1234; https://doi.org/10.3390/molecules22071234 - 22 Jul 2017

Cited by 29 | Viewed by 6625

Abstract

For the last six decades, researchers have been focused on finding efficient reactivators of organophosphorus compound (OP)-inhibited acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). In this study, we have focused our research on a new oxime scaffold based on the Cinchona structure since it was [...] Read more.

For the last six decades, researchers have been focused on finding efficient reactivators of organophosphorus compound (OP)-inhibited acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). In this study, we have focused our research on a new oxime scaffold based on the Cinchona structure since it was proven to fit the cholinesterases active site and reversibly inhibit their activity. Three Cinchona oximes (C1, C2, and C3), derivatives of the 9-oxocinchonidine, were synthesized and investigated in reactivation of various OP-inhibited AChE and BChE. As the results showed, the tested oximes were more efficient in the reactivation of BChE and they reactivated enzyme activity to up to 70% with reactivation rates similar to known pyridinium oximes used as antidotes in medical practice today. Furthermore, the oximes showed selectivity towards binding to the BChE active site and the determined enzyme-oxime dissociation constants supported work on the future development of inhibitors in other targeted studies (e.g., in treatment of neurodegenerative disease). Also, we monitored the cytotoxic effect of Cinchona oximes on two cell lines Hep G2 and SH-SY5Y to determine the possible limits for in vivo application. The cytotoxicity results support future studies of these compounds as long as their biological activity is targeted in the lower micromolar range. Full article

(This article belongs to the Special Issue The Cholinesterases—Structure, Mechanism, Function and Drug Design: The 25th Anniversary of the Solution of the Crystal Structure of Acetylcholinesterase by Joel L. Sussman and Israel Silman)

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Synthesis of Cinchona derivatives starting from cinchonidine. Full article ">Figure 2
Screening of organophosphorus compound (OP)-inhibited human acetylcholinesterase (hAChE) reactivation by 0.1 mM oximes C1, C2 and C3 at 25 °C. Results in terms of the maximal obtained reactivation percentage within 23 h are presented as a mean of two to three experiments (experimental deviation was less than 10%). Full article ">Figure 3
[H+]-dependence of oximolysis rate for C1, C2 and C3 oximes (0.1 mM) and their in vitro and in silico determined pKa values. Acid dissociation constant (Ka) of the oxime group was determined by measuring the degradation of substrate acetylthiocholine (1 mM) by 100 µM oxime at 412 nm and in pH range 4.4–11.3 and was calculated using the Equation (1) (see <a href="#sec3dot4-molecules-22-01234" class="html-sec">Section 3.4</a>). Full article ">Figure 4
Screening of OP-inhibited human butyrylcholinesterase (hBChE) reactivation by 0.1 mM oximes C1, C2 and 0.05 mM C3 at 25 °C. Results in terms of maximal obtained reactivation percentage within 22–24 h are presented as a mean of two to three experiments (experimental deviation was less than 10%). Full article ">Figure 5
Reversible inhibition of (A) hAChE and (B) hBChE by the tested oximes. The graphs present Hunter-Downs plots with the average of the experimentally obtained values from at least three experiments. Ki presenting the Y-intercept was studied in 0.3–1.0 mM substrate acetylthiocholine (ATCh) concentration range. Full article ">Figure 6
Cytotoxicity of oximes C1, C2, and C3 on two cell lines (A) HepG2, human Caucasian hepatocyte carcinoma, epithelial; (B) SH-SY5Y human neuroblastoma cell line. Full article ">

6857 KiB

Open AccessArticle

Synthesis, Physico-chemical Characterization, Crystal Structure and Influence on Microbial and Tumor Cells of Some Co(II) Complexes with 5,7-Dimethyl-1,2,4-triazolo[1,5-a]pyrimidine

by Luminiţa Măruţescu, Larisa Calu, Mariana Carmen Chifiriuc, Coralia Bleotu, Constantin-Gabriel Daniliuc, Denisa Fălcescu, Crina Maria Kamerzan, Mihaela Badea and Rodica Olar

Molecules 2017, 22(7), 1233; https://doi.org/10.3390/molecules22071233 - 22 Jul 2017

Cited by 9 | Viewed by 5591

Abstract

Three complexes, namely [Co(dmtp)₂(OH₂)₄][CoCl₄] (1), [Co(dmtp)₂Cl₂] (2) and [Co(dmtp)₂(OH₂)₄]Cl₂∙2H₂O (3) (dmtp: 5,7-dimethyl-1,2,4-triazolo[1,5-a]pyrimidine), were [...] Read more.

Three complexes, namely [Co(dmtp)₂(OH₂)₄][CoCl₄] (1), [Co(dmtp)₂Cl₂] (2) and [Co(dmtp)₂(OH₂)₄]Cl₂∙2H₂O (3) (dmtp: 5,7-dimethyl-1,2,4-triazolo[1,5-a]pyrimidine), were synthesized and characterized by spectral (IR, UV-Vis-NIR), and magnetic measurements at room temperature, as well as single crystal X-ray diffraction. Complex (1) crystallizes in monoclinic system (space group C2/c), complex (2) adopts an orthorhombic system (space group Pbca), and complex (3) crystallizes in triclinic system (space group P1). Various types of extended hydrogen bonds and π–π interactions provide a supramolecular architecture for all complexes. All species were evaluated for antimicrobial activity towards planktonic and biofilm-embedded microbial cells and influence on HEp-2 cell viability, cellular cycle and gene expression. Full article

(This article belongs to the Section Organometallic Chemistry)

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3739 KiB

Open AccessArticle

Spectrum Effect Relationship and Component Knock-Out in Angelica Dahurica Radix by High Performance Liquid Chromatography-Q Exactive Hybrid Quadrupole-Orbitrap Mass Spectrometer

by Jinmei Wang, Linna Peng, Mengjun Shi, Changqin Li, Yan Zhang and Wenyi Kang

Molecules 2017, 22(7), 1231; https://doi.org/10.3390/molecules22071231 - 21 Jul 2017

Cited by 29 | Viewed by 7031

Abstract

Different extracts of Angelica dahuricae were available for whitening or treating vitiligo clinically. They showed inhibitory or activating effects on tyrosinase, a rate-limiting enzyme of melanogenesis. This study aimed to identify active compounds on tyrosinase in water extract of Angelica dahurica Radix. We [...] Read more.

Different extracts of Angelica dahuricae were available for whitening or treating vitiligo clinically. They showed inhibitory or activating effects on tyrosinase, a rate-limiting enzyme of melanogenesis. This study aimed to identify active compounds on tyrosinase in water extract of Angelica dahurica Radix. We applied spectrum-effect relationship and component knock-out methods to make it clear. HPLC was used to obtain the specific chromatograms. The effects on tyrosinase activity were examined by measuring the oxidation rate of levodopa in vitro. Partial least squares method was used to examine the spectrum-effect relationships. The knocked-out samples were prepared by HPLC method, and the identification of knocked-out compounds was conducted by the high performance liquid chromatography-four stage rod-electrostatic field orbit trap high resolution mass spectrometry. Results showed that S6, S14, S18, S21, S35, S36, S37, S40, and S41 were positively correlated to inhibitory activity of Angelica dahuricae on tyrosinase whereas S9, S11, S8, S12, S22, and S30 were negatively correlated. When the concentration of each sample was 1 g·mL⁻¹, equal to the amount of raw medicinal herbs, oxypeucedanin hydrate, imperatorin, cnidilin, and isoimperatorin had inhibitory effects on tyrosinase activity whereas byakangelicin and bergapten had activating effects. Full article

(This article belongs to the Collection Herbal Medicine Research)

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1944 KiB

Open AccessArticle

The Influence of Glycosylation of Natural and Synthetic Prenylated Flavonoids on Binding to Human Serum Albumin and Inhibition of Cyclooxygenases COX-1 and COX-2

by Tomasz Tronina, Paulina Strugała, Jarosław Popłoński, Aleksandra Włoch, Sandra Sordon, Agnieszka Bartmańska and Ewa Huszcza

Molecules 2017, 22(7), 1230; https://doi.org/10.3390/molecules22071230 - 21 Jul 2017

Cited by 38 | Viewed by 7025

Abstract

The synthesis of different classes of prenylated aglycones (α,β-dihydroxanthohumol (2) and (Z)-6,4’-dihydroxy-4-methoxy-7-prenylaurone (3)) was performed in one step reactions from xanthohumol (1)—major prenylated chalcone naturally occurring in hops. Obtained flavonoids (2–3) [...] Read more.

The synthesis of different classes of prenylated aglycones (α,β-dihydroxanthohumol (2) and (Z)-6,4’-dihydroxy-4-methoxy-7-prenylaurone (3)) was performed in one step reactions from xanthohumol (1)—major prenylated chalcone naturally occurring in hops. Obtained flavonoids (2–3) and xanthohumol (1) were used as substrates for regioselective fungal glycosylation catalyzed by two Absidia species and Beauveria bassiana. As a result six glycosides (4–9) were formed, of which four glycosides (6–9) have not been published so far. The influence of flavonoid skeleton and the presence of glucopyranose and 4-O-methylglucopyranose moiety in flavonoid molecule on binding to main protein in plasma, human serum albumin (HSA), and inhibition of cyclooxygenases COX-1 and COX-2 were investigated. Results showed that chalcone (1) had the highest binding affinity to HSA (8.624 × 10⁴ M⁻¹) of all tested compounds. It has also exhibited the highest inhibition of cyclooxygenases activity, and it was a two-fold stronger inhibitor than α,β-dihydrochalcone (2) and aurone (3). The presence of sugar moiety in flavonoid molecule caused the loss of HSA binding activity as well as the decrease in inhibition of cyclooxygenases activity. Full article

(This article belongs to the Special Issue Synthesis and Biological Applications of Glycoconjugates)

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Full article ">Figure 1
Synthesis of α,β-dihydroxanthohumol (2) from xanthohumol (1) [<a href="#B34-molecules-22-01230" class="html-bibr">34</a>]. Full article ">Figure 2
Synthesis of (Z)-6,4’-dihydroxy-4-methoxy-7-prenylaurone (3) from xanthohumol (1). Full article ">Figure 3
Fungal transformations of prenylated flavonoids 1-3 catalyzed by: (A) Absidia coeruela AM93; (B) Absidia glauca AM177; (C) Rhizopus nigricans UPF701; (D) Beauveria bassiana AM278; and (E) B. bassiana AM446. Full article ">Figure 3 Cont.
Fungal transformations of prenylated flavonoids 1-3 catalyzed by: (A) Absidia coeruela AM93; (B) Absidia glauca AM177; (C) Rhizopus nigricans UPF701; (D) Beauveria bassiana AM278; and (E) B. bassiana AM446. Full article ">Figure 4
Emission spectra of HSA in the presence of various concentrations of tested compounds and Stern-Volmer plots of F0/F against the concentration of tested compounds (HSA = 1.5 × 10−5 M, λex = 280 nm, T = 310 K). Control is marked red and consecutive spectra of the studied compounds (marked grey) are in the following concentrations 1, 3, 5…, 15 µM. Top graphs present xanthohumol (1) and its glycosides (4–5); middle graphs present α,β-xanthohumol (2) and its glycosides (6–7); and bottom graphs present aurone 3 and its glycosides (8–9) ((A) aglycones (1–3); (B) glycosides (4, 6, and 8); and (C) 4-O-methylglycosides (5, 7, and 9)). Full article ">

4798 KiB

Open AccessArticle

Effects of K11R and G31P Mutations on the Structure and Biological Activities of CXCL8: Solution Structure of Human CXCL8_(3-72)K11R/G31P

by Hsi-Tsung Cheng, Hui-Yuan Yu, John R. Gordon, Fang Li and Jya-Wei Cheng

Molecules 2017, 22(7), 1229; https://doi.org/10.3390/molecules22071229 - 21 Jul 2017

Cited by 5 | Viewed by 5377

Abstract

The ELR-CXC chemokines are important to neutrophil inflammation in many acute and chronic diseases. Among them, CXCL8 (interleukin-8, IL-8), the expression levels of which are elevated in many inflammatory diseases, binds to both the CXCR1 and CXCR2 receptors with high affinity. Recently, an [...] Read more.

The ELR-CXC chemokines are important to neutrophil inflammation in many acute and chronic diseases. Among them, CXCL8 (interleukin-8, IL-8), the expression levels of which are elevated in many inflammatory diseases, binds to both the CXCR1 and CXCR2 receptors with high affinity. Recently, an analogue of human CXCL8, CXCL8_(3–72)K11R/G31P (hG31P) has been developed. It has been demonstrated that hG31P is a high affinity antagonist for both the CXCR1 and CXCR2. Herein, we have determined the solution structure and the CXCR1 N-terminal peptide binding sites of hG31P by NMR spectroscopy. We have found that the displacement within the tertiary structure of the 30 s loop and the N-terminal region and more specifically change of the loop conformation (especially H33), of hG31P may affect its binding to the CXCR1 receptor and thereby inhibit human neutrophil chemotactic responses induced by ELR-CXC chemokines. Our results provide a structural basis for future clinical investigations of this CXCR1/CXCR2 receptor antagonist and for the further development of CXCL8 based antagonists. Full article

(This article belongs to the Special Issue Recent Advances in Biomolecular NMR Spectroscopy)

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Full article ">Figure 1
1H-15N HSQC of hG31P. Cross-peaks are labeled according to the residue types and numbers. Full article ">Figure 2
(A) A multiple sequence alignment of hG31P and CXCL1 to CXCL10. Secondary structural elements are shown above the alignment; (B) A stereoview of the backbone superimposition of the final 20 structures; (C) Ribbon diagram of the final 20 structures of hG31P. Full article ">Figure 3
(A) Stereo view of the superposition of the ribbon structures of hG31P (cyan) and CXCL8 (PDB code 1ILP; brown) showing the displacement of the ELR N-terminal region and the E29–T37 loop between hG31P and CXCL8. The structures were superimposed by residues F21–E29, G46–L51, and N56–S72; (B) Stereo view of the schematic representations of the residues involved in inter-residue NOEs of hG31P. None of such NOEs were found in CXCL8. Full article ">Figure 4
Superposition of the ribbon structures of the 30 s loop between (A) CXCL8 (PDB: 1IL8, H33 in red) and hG31P (H33 in blue); (B) CXCL8 (PDB: 1IL8, H33 in red) and CXCL4 (PDB: 1F9Q, H35 in green); (C) CXCL8 (PDB: 1IL8, H33 in red) and CXCL10 (PDB: 1LV9, H35 in pink). Full article ">Figure 5
(A) Overlay of the 1H-15N HSQC spectrum of hG31P (red) and the hG31P/CXCR1 N-terminal peptide complex (black). Cross-peaks with significant shifts are labeled according to the residue types and numbers; (B) Chemical shift change versus residue number of hG31P upon CXCR1 N-terminal peptide binding. The chemical shift changes are calculated according to the equation: Δδ = [Δδ(1H)2] + [[0.2xΔδ(15N)]2]1/2; (C) Surface rendering of hG31P bound to CXCR1 N-terminal peptide. The blue shading corresponds to the residues that give rise to a change in Δδ chemical shift great than 0.04 ppm. The peptide is shown as a stick model in red; (D) Equilibrium binding of hG31P to the N-terminal peptide. Solid squares are the hG31P dependent fluorescence signal intensity at 516 nm (Kd = 48 ± 5 μM). Solid line represents curve fitting of the single binding site model. Full article ">Figure 5 Cont.
(A) Overlay of the 1H-15N HSQC spectrum of hG31P (red) and the hG31P/CXCR1 N-terminal peptide complex (black). Cross-peaks with significant shifts are labeled according to the residue types and numbers; (B) Chemical shift change versus residue number of hG31P upon CXCR1 N-terminal peptide binding. The chemical shift changes are calculated according to the equation: Δδ = [Δδ(1H)2] + [[0.2xΔδ(15N)]2]1/2; (C) Surface rendering of hG31P bound to CXCR1 N-terminal peptide. The blue shading corresponds to the residues that give rise to a change in Δδ chemical shift great than 0.04 ppm. The peptide is shown as a stick model in red; (D) Equilibrium binding of hG31P to the N-terminal peptide. Solid squares are the hG31P dependent fluorescence signal intensity at 516 nm (Kd = 48 ± 5 μM). Solid line represents curve fitting of the single binding site model. Full article ">Figure 6
Model for the binding of hG31P (ribbon structure) to the CXCR1 receptor (blue) on a neutrophil. The side chains of the N-terminal ELR, R11, P31 residues of hG31P are shown in sticks (blue). Site I indicates the interactions between the hG31P N-loop residues and receptor N-domain, and site II indicates the hG31P N-terminal and 30 s loop residues and receptor exoloop residues. The displacement of the N-terminal ELR region and the 30 s loop and the change of the 30 s loop conformation may affect its binding and activation to the CXCR1 receptor thus inhibit the neutrophil chemotaxis signaling pathway. Full article ">Figure 7
(A) Amino acid sequences of the designed CXCL8-IP10 molecule. Residues derived from CXCL8 are shown in black, residues derived from CXCL10 (IP10) are shown in red, K11R mutation site is shown in blue; (B) CXCL8-IP10 effectively antagonizes human neutrophil responses to ELR-CXC chemokine CXCL8. Full article ">

8438 KiB

Open AccessArticle

Effects of Chlorhexidine-Encapsulated Mesoporous Silica Nanoparticles on the Anti-Biofilm and Mechanical Properties of Glass Ionomer Cement

by Huiyi Yan, Hongye Yang, Kang Li, Jian Yu and Cui Huang

Molecules 2017, 22(7), 1225; https://doi.org/10.3390/molecules22071225 - 21 Jul 2017

Cited by 42 | Viewed by 7593

Abstract

One of the primary causes for the failure of glass ionomer cement (GIC) is secondary caries. To enhance the anti-microbial performance of GIC without affecting its mechanical properties, chlorhexidine (CHX) was encapsulated in expanded-pore mesoporous silica nanoparticles (pMSN) to synthesize CHX@pMSN. CHX@pMSN was [...] Read more.

One of the primary causes for the failure of glass ionomer cement (GIC) is secondary caries. To enhance the anti-microbial performance of GIC without affecting its mechanical properties, chlorhexidine (CHX) was encapsulated in expanded-pore mesoporous silica nanoparticles (pMSN) to synthesize CHX@pMSN. CHX@pMSN was added at three mass fractions (1%, 5%, and 10% (w/w)) to GIC powder as the experimental groups. Pure GIC was set as the control group. The mechanical and anti-biofilm properties of GIC from each group were tested. The results demonstrated that CHX was successfully encapsulated on/into pMSN, and the encapsulating efficiency of CHX was 44.62% in CHX@pMSN. The anti-biofilm ability was significantly enhanced in all experimental groups (p < 0.001) compared with that in the control group. CHX was continuously released, and anti-biofilm ability was maintained up to 30 days. In addition, the mechanical properties (compressive strength, surface hardness, elastic modulus, water sorption, and solubility) of 1% (w/w) group were maintained compared with those in the control group (p > 0.05). In conclusion, adding 1% (w/w) CHX@pMSN to GIC led to conspicuous anti-biofilm ability and had no adverse effect on the mechanical properties of this restorative material. This study proposes a new strategy for preventing secondary caries by using CHX@pMSN-modified GIC. Full article

(This article belongs to the Special Issue Mesoporous Silica in Biomedical Applications)

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2308 KiB

Open AccessArticle

Application of Ammonium Persulfate for Selective Oxidation of Guanines for Nucleic Acid Sequencing

by Yafen Wang, Chaoxing Liu, Tingting Hong, Fan Wu, Shuyi Yu, Zhiyong He, Wuxiang Mao and Xiang Zhou

Molecules 2017, 22(7), 1222; https://doi.org/10.3390/molecules22071222 - 21 Jul 2017

Cited by 2 | Viewed by 6656

Abstract

Nucleic acids can be sequenced by a chemical procedure that partially damages the nucleotide positions at their base repetition. Many methods have been reported for the selective recognition of guanine. The accurate identification of guanine in both single and double regions of DNA [...] Read more.

Nucleic acids can be sequenced by a chemical procedure that partially damages the nucleotide positions at their base repetition. Many methods have been reported for the selective recognition of guanine. The accurate identification of guanine in both single and double regions of DNA and RNA remains a challenging task. Herein, we present a new, non-toxic and simple method for the selective recognition of guanine in both DNA and RNA sequences via ammonium persulfate modification. This strategy can be further successfully applied to the detection of 5-methylcytosine by using PCR. Full article

(This article belongs to the Special Issue Nucleic Acid Chemistry and Nucleic Acid Drugs: A Themed Issue in Honor of Professor Lihe Zhang on the Occasion of His 80th Birthday)

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Figure 1
(A) Schematic illustration of nucleic acid sequences oxidized by AP and then cleaved by hot piperidine or aniline acetate; (B) Polyacrylamide gel electrophoresis experiments showing cleavage products of HEX-labeled DNA of ODN-mismatch (20 pmol) incubated with AP in Tris-HCl buffer (pH = 7.4). Lane 1: DNA alone; lane 2: ODN with AP; lane 3: ODN with piperidine; lane 4: ODN with AP and piperidine; lane 5: G-ladder of ODN (treated with DMS). Full article ">Figure 2
Polyacrylamide gel electrophoresis analysis of DNA. The oxidized DNA treated with piperidine at 90 °C for 40 min. (A) Cleavage of the hairpin-loop structure; (B) Cleavage of the double strand structure; (C) Cleavage of the terminal structure; (D) Cleavage of the mismatch structure; (E) Cleavage of the loop and bulge structure; (F) Cleavage of the bulge structure; (G) Cleavage of the G-quadruplex structure. Full article ">Figure 2 Cont.
Polyacrylamide gel electrophoresis analysis of DNA. The oxidized DNA treated with piperidine at 90 °C for 40 min. (A) Cleavage of the hairpin-loop structure; (B) Cleavage of the double strand structure; (C) Cleavage of the terminal structure; (D) Cleavage of the mismatch structure; (E) Cleavage of the loop and bulge structure; (F) Cleavage of the bulge structure; (G) Cleavage of the G-quadruplex structure. Full article ">Figure 3
Polyacrylamide gel electrophoresis analysis of 76-mer DNA. The ODN (20 pmol) was oxidized by AP and then treated with piperidine. Lane 1: DNA alone; lane 2: DNA treated with AP; lane 3: G-ladder of ODN (treated with DMS). Full article ">Figure 4
Polyacrylamide gel electrophoresis analysis of RNA sequences treated with AP and aniline acetate. (A) Lane 1: RNA alone; lane 2: RNA was treated with RNase T1; lane 3: RNA was treated with AP and aniline acetate; (B) Lane 1: RNA alone; lane 2: RNA was treated with AP and aniline acetate; lane 3: RNA was treated with RNase T1. Full article ">Figure 5
Polyacrylamide gel electrophoresis analysis of 20 mer-DNA. Lane 1: DNA alone; Lane 2: DNA was treated with AP; Lane 3: DNA was treated with methylene blue in the presence of light and oxygen for 15 min. Full article ">Figure 6
(A) Schematic illustration of detection of the loci of 5-methylcytosine in DNA by PCR, the product of PCR was treated with AP and hot piperidine; (B,C) Poly-acrylamide gel electrophoresis analysis of 76-mer template containing one or two 5-methylcytosine sites. Lane 1: marker; lane 2: PCR product without treatment; lane 3: PCR product treated with AP and hot piperidine. Full article ">

4073 KiB

Open AccessArticle

High-Resolution Inhibition Profiling Combined with HPLC-HRMS-SPE-NMR for Identification of PTP1B Inhibitors from Vietnamese Plants

by Binh Thi Dieu Trinh, Anna K. Jäger and Dan Staerk

Molecules 2017, 22(7), 1228; https://doi.org/10.3390/molecules22071228 - 20 Jul 2017

Cited by 12 | Viewed by 6355

Abstract

Protein tyrosine phosphatase 1B (PTP1B) plays a key role as a negative regulator in insulin signal transduction by deactivating the insulin receptor. Thus, PTP1B inhibition has emerged as a potential therapeutic strategy for curing insulin resistance. In this study, 40 extracts from 18 [...] Read more.

Protein tyrosine phosphatase 1B (PTP1B) plays a key role as a negative regulator in insulin signal transduction by deactivating the insulin receptor. Thus, PTP1B inhibition has emerged as a potential therapeutic strategy for curing insulin resistance. In this study, 40 extracts from 18 different plant species were investigated for PTP1B inhibitory activity in vitro. The most promising one, the EtOAc extract of Ficus racemosa, was investigated by high-resolution PTP1B inhibition profiling combined with HPLC-HRMS-SPE-NMR analysis. This led to the identification of isoderrone (1), derrone (2), alpinumisoflavone (3) and mucusisoflavone B (4) as PTP1B inhibitors. IC₅₀ of these compounds were 22.7 ± 1.7, 12.6 ± 1.6, 21.2 ± 3.8 and 2.5 ± 0.2 µM, respectively. Kinetics analysis revealed that these compounds inhibited PTP1B non-competitively with K_i values of 21.3 ± 2.8, 7.9 ± 1.9, 14.3 ± 2.0, and 3.0 ± 0.5 µM, respectively. These findings support the important role of F. racemosa as a novel source of new drugs and/or as a herbal remedy for treatment of type 2 diabetes. Full article

(This article belongs to the Special Issue Bioactive Compounds for Metabolic Syndrome and Type 2 Diabetes)

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Protein tyrosine phosphatase 1B (PTP1B) inhibitory activity of EtOAc and n-BuOH extracts of 18 different plant species, at a concentration of 100 µg/mL, represented as mean ± standard deviation, n = 3. Full article ">Figure 2
HPLC trace at 254 nm and high-resolution PTP1B inhibition profile of EtOAc extract of Ficus racemosa. Full article ">Figure 3
Trapping peaks 1–7 of Fr.1 of Ficus racemosa EtOAc extract, analyzed by HPLC-PDA-HRMS-SPE-NMR using an analytical-scale HPLC. Blue line: UV chromatogram at 254 nm, pink line: base peak chromatogram. Full article ">Figure 4
Structures of compounds 1–4 isolated from EtOAc extract of Ficus racemosa. Full article ">Figure 5
Dose–response curves of compounds 1–4. Each point represents the average of triplicate measurements. Full article ">Figure 6
Lineweaver–Burk plots of inhibition kinetics of PTP1B inhibitory effects by compounds 1–4. Each point represents the average of triplicate measurements. Full article ">

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Open AccessArticle

Characterization and Purification of Bergamottin from Citrus grandis (L.) Osbeck cv. Yongjiazaoxiangyou and Its Antiproliferative Activity and Effect on Glucose Consumption in HepG2 cells

by Yilong Liu, Chuanhong Ren, Yunlin Cao, Yue Wang, Wenyi Duan, Linfeng Xie, Chongde Sun and Xian Li

Molecules 2017, 22(7), 1227; https://doi.org/10.3390/molecules22071227 - 20 Jul 2017

Cited by 33 | Viewed by 5951

Abstract

Bergamottin is a natural furanocoumarin compound with weak polarity. Characterization and quantification of bergamottin were carried out in different fruit tissues of various citrus cultivars. Among the four citrus tissues tested, i.e., flavedo, albedo, segment membrane (SM), and juice sacs (JS) in eight [...] Read more.

Bergamottin is a natural furanocoumarin compound with weak polarity. Characterization and quantification of bergamottin were carried out in different fruit tissues of various citrus cultivars. Among the four citrus tissues tested, i.e., flavedo, albedo, segment membrane (SM), and juice sacs (JS) in eight citrus cultivars, the highest bergamottin content was found in the flavedo of Citrus grandis (L.) Osbeck cv. Yongjiazaoxiangyou (YJZXY, 666.54 μg·g⁻¹ DW). A combination of silica gel column chromatography and high-speed counter-current chromatography (HSCCC) was established to efficiently purify bergamottin from the flavedo of YJZXY. Bergamottin showed significant antiproliferative activity on three cancer cell lines, i.e., human liver cancer HepG2, promyelocytic leukemia HL-60, and gastric cancer BGC-823 cells, which showed a marked inhibition effect on these cell lines in a dose-dependent manner. In addition, bergamottin significantly increased glucose consumption in HepG2 cells also in a dose-dependent manner, which is the first report of its potential in anti-diabetes applications. Full article

(This article belongs to the Section Natural Products Chemistry)

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Structure of bergamottin. Full article ">Figure 2
High-speed counter-current chromatography (HSCCC) chromatogram of the purification of bergamottin extracted from the silica gel-refined sample. Two-phase solvent system: hexane–ethyl acetate–methanol–water (1:1:2:0.625, v/v/v/v); stationary phase: upper phase; mobile phase: lower phase; flow rate: 2.0 mL·min−1; revolution speed: 900 rpm; detection wavelength: 250 nm. Full article ">Figure 3
HPLC chromatogram of crude extract before (A) and after (B) treatment with silica gel column; the HSCCC purified product (C); LC-MS2 chromatogram of the final purified bergamottin (D) (λ = 250 nm). Full article ">Figure 4
Effect of purified bergamottin on the growth of three cancer cell lines. Human liver cancer HepG2 (A); promyelocytic leukemia HL-60 (B); and gastric cancer BGC-823 (C) were used in the experiment. Taxol was used as a positive control. Full article ">Figure 5
Effect of purified bergamottin (BGM) on glucose consumption after 24 h treatment in HepG2 cells. * p < 0.05, ** p < 0.01, *** p < 0.001, compared to the DMSO blank control. MET, metformin as a positive control. Full article ">

949 KiB

Open AccessArticle

Characterization of Essential Oil Composition in Different Basil Species and Pot Cultures by a GC-MS Method

by Andrea Muráriková, Anton Ťažký, Jarmila Neugebauerová, Alexandra Planková, Josef Jampílek, Pavel Mučaji and Peter Mikuš

Molecules 2017, 22(7), 1221; https://doi.org/10.3390/molecules22071221 - 20 Jul 2017

Cited by 61 | Viewed by 9152

Abstract

Basil (Ocimum L.) species are used as medicinal plants due to their essential oils exhibiting specific biological activity. The present work demonstrated that both the variety and season/conditions of cultivation had a significant effect on (i) the produced amount (extraction yield), (ii) [...] Read more.

Basil (Ocimum L.) species are used as medicinal plants due to their essential oils exhibiting specific biological activity. The present work demonstrated that both the variety and season/conditions of cultivation had a significant effect on (i) the produced amount (extraction yield), (ii) qualitative, as well as (iii) quantitative profile of basil essential oil. Among studied basil varieties, a new variety, ‘Mánes’, was characterized for the first time. Based on our quantitative evaluation of GC-MS profiles, the following chemotypes and average concentrations of a main component were detected in the studied basil varieties: ‘Ohře’, ‘Lettuce Leaf’, ‘Purple Opaal’, ‘Dark Green’ (linalool, 5.99, 2.49, 2.34, 2.01 mg/mL, respectively), and ‘Mammolo Genovese’, ‘Mánes’, ‘Red Rubin’ (eucalyptol, 1.34, 0.96, 0.76 mg/mL, respectively). At the same time, when considering other compounds identified in GC-MS profiles, all the studied varieties, except from ‘Lettuce Leaf’, were methyl eugenol-rich with a strong dependence of the eugenol:methyl eugenol ratio on the seasonal changes (mainly solar irradiation, but also temperature and relative humidity). More complex and/or variable (depending on the season and cultivation) chemotypes were observed with ‘Lettuce Leaf’ (plus estragole, 2.27 mg/mL), ‘Dark Green’ (plus eucalyptol, 1.36 mg/mL), ‘Mammolo Genovese’ (plus eugenol, 1.19 mg/mL), ‘Red Rubin’ (plus linalool and eugenol, 0.46 and 0.56 mg/mL, respectively), and ‘Mánes’ (plus linalool and eugenol, 0.58 and 0.40 mg/mL, respectively). When considering superior extraction yield (ca. 17 mL·kg⁻¹, i.e., two to five times higher than other examined varieties) and consistent amounts (yields) of essential oil when comparing inter-seasonal or inter-year data (RSD and inter-year difference in mean yield values ˂2.5%), this new basil variety is very promising for use in the pharmaceutical, food, and cosmetic industries. Full article

(This article belongs to the Section Natural Products Chemistry)

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165 KiB

Open AccessEditorial

Special Issue: Adenosine Receptors

by Francisco Ciruela and Eddy Sotelo

Molecules 2017, 22(7), 1220; https://doi.org/10.3390/molecules22071220 - 20 Jul 2017

Cited by 4 | Viewed by 3616

Abstract

Nearly 90 years ago, Drury and Szent-Györgyi revealed that adenosine produced profound hypotension and bradycardia, and it affected kidney function in mammals [1]. [...]
Full article

(This article belongs to the Special Issue Adenosine Receptors)

2852 KiB

Open AccessArticle

Fluorination of Naturally Occurring N⁶-Benzyladenosine Remarkably Increased Its Antiviral Activity and Selectivity

by Vladimir E. Oslovsky, Mikhail S. Drenichev, Liang Sun, Nikolay N. Kurochkin, Vladislav E. Kunetsky, Carmen Mirabelli, Johan Neyts, Pieter Leyssen and Sergey N. Mikhailov

Molecules 2017, 22(7), 1219; https://doi.org/10.3390/molecules22071219 - 20 Jul 2017

Cited by 17 | Viewed by 5233

Abstract

Recently, we demonstrated that the natural cytokinin nucleosides N⁶-isopentenyladenosine (iPR) and N⁶-benzyladenosine (BAPR) exert a potent and selective antiviral effect on the replication of human enterovirus 71. In order to further characterize the antiviral profile [...] Read more.

Recently, we demonstrated that the natural cytokinin nucleosides N⁶-isopentenyladenosine (iPR) and N⁶-benzyladenosine (BAPR) exert a potent and selective antiviral effect on the replication of human enterovirus 71. In order to further characterize the antiviral profile of this class of compounds, we generated a series of fluorinated derivatives of BAPR and evaluated their activity on the replication of human enterovirus 71 in a cytopathic effect (CPE) reduction assay. The monofluorination of the BAPR-phenyl group changed the selectivity index (SI) slightly because of the concomitant high cell toxicity. Interestingly, the incorporation of a second fluorine atom resulted in a dramatic improvement of selectivity. Moreover, N⁶-trifluoromethylbenzyladenosine derivatives (9–11) exhibited also a very interesting profile, with low cytotoxicity observed. In particular, the analogue N⁶-(3-trifluoromethylbenzyl)-adenosine (10) with a four-fold gain in potency as compared to BAPR and the best SI in the class represents a promising candidate for further development. Full article

(This article belongs to the Special Issue Nucleoside and Nucleotide Analogues)

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5020 KiB

Open AccessArticle

Magnolol, a Natural Polyphenol, Attenuates Dextran Sulfate Sodium-Induced Colitis in Mice

by Ling Zhao, Hai-tao Xiao, Huai-xue Mu, Tao Huang, Ze-si Lin, Linda L. D. Zhong, Guang-zhi Zeng, Bao-min Fan, Cheng-yuan Lin and Zhao-xiang Bian

Molecules 2017, 22(7), 1218; https://doi.org/10.3390/molecules22071218 - 20 Jul 2017

Cited by 54 | Viewed by 9116

Abstract

Magnolol is a lignan with anti-inflammatory activity identified in Magnolia officinalis. Ulcerative colitis (UC), one of the types of inflammatory bowel disease (IBD), is a disease that causes inflammation and ulcers in the colon. To investigate the effect of magnolol in dextran [...] Read more.

Magnolol is a lignan with anti-inflammatory activity identified in Magnolia officinalis. Ulcerative colitis (UC), one of the types of inflammatory bowel disease (IBD), is a disease that causes inflammation and ulcers in the colon. To investigate the effect of magnolol in dextran sulfate sodium (DSS)-induced experimental UC model, male C57 mice were treated with 2% DSS drinking water for 5 consecutive days followed by intragastric administration with magnolol (5, 10 and 15 mg/kg) daily for 7 days. The results showed that magnolol significantly attenuated disease activity index, inhibited colonic shortening, reduced colonic lesions and suppressed myeloperoxidase (MPO) activity. Moreover, colonic pro-inflammatory cytokines (TNF-α, IL-6, and IL-1β) induced by colitis were dramatically decreased by magnolol. To further unveil the metabolic signatures upon magnolol treatment, mass spectrometry-based metabolomic analysis of the small molecular metabolites in mice serum were performed. Compared with controls, abnormality of serum metabolic phenotypes in DSS-treated mice were effectively reversed by different doses of magnolol. In particular, magnolol treatment effectively elevated the serum levels of tryptophan metabolites including kynurenic acid (KA), 5-hydroxyindoleacetic acid, indoleacetic acid (IAA), indolelactic acid and indoxylsulfuric acid, which are potential aryl hydrocarbon receptor (AHR) ligands to impact colitis. These findings suggest that magnolol exerts anti-inflammatory effect on DSS-induced colitis and its underlying mechanisms are associated with the restoring of tryptophan metabolites that inhibit the colonic inflammation. Full article

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Figure 1
The phenotypic severity of dextran sulfate sodium (DSS)-induced colitis in mice can be effectively attenuated by one-week treatment of magnolol. (A) The body weight loss was significantly improved in all treatment groups versus DSS group; (B) The enhanced disease activity index of DSS mice was significantly reduced in all treatment groups; (C) DSS-induced shortened colon was significantly improved by high dose of magnolol and infliximab. The value in the plot was expressed as means ± SEM, and statistically significant was marked by asterisk (* p < 0.05; ** p < 0.01, vs. DSS group). Full article ">Figure 1 Cont.
The phenotypic severity of dextran sulfate sodium (DSS)-induced colitis in mice can be effectively attenuated by one-week treatment of magnolol. (A) The body weight loss was significantly improved in all treatment groups versus DSS group; (B) The enhanced disease activity index of DSS mice was significantly reduced in all treatment groups; (C) DSS-induced shortened colon was significantly improved by high dose of magnolol and infliximab. The value in the plot was expressed as means ± SEM, and statistically significant was marked by asterisk (* p < 0.05; ** p < 0.01, vs. DSS group). Full article ">Figure 2
Medium and high dosages of magnolol effectively attenuated histopathological changes and myeloperoxidase activity in the colon of DSS-treated mice. (A) Representative images of hematoxylin/eosin (H&E) staining (magnification, 10×): (a) Control group; (b) DSS group; (c) Infliximab group; (d) Magnolol 5 mg/kg; (e) Magnolol 10 mg/kg; (f) Magnolol 15 mg/kg; (B) Histological scores; (C) MPO activity. The value in the plot was expressed as means ± SEM, and statistically significant was marked by asterisk (* p < 0.05; ** p < 0.01). Full article ">Figure 3
Medium and/or high dosages of magnolol significantly attenuated DSS-induced high levels of proinflammatory cytokines TNF-α (A), IL-1β (B) and IL-6 (C) in the colonic tissues. The value in the plot was expressed as means ± SEM, and statistically significant was marked by asterisk (* p < 0.05). Full article ">Figure 4
Magnolol majorly reversed abnormality of serum metabolome in colitis mice. (A) Two dimensional partial least squares discriminant analysis (PLS-DA) scatter plots displayed distinct metabolic profiles among model mice with and without drug treatment through UPLC/MS-based serum metabolomic analysis in both ESI modes. The variables explained 23% (t1) and 13.6% (t2) in ESI positive mode, while the variables explained 20.9% (t1) and 10.9% (t2) in ESI negative mode. (B) The log2 fold changes of all identified metabolic features between groups of DSS and control, groups of medium dose of magnolol and DSS as well as groups of infliximab and DSS. Full article ">Figure 5
Reduction of serum metabolites kynurenic acid (A), 5-hydroxyindoleacetic acid (B), indoleacetic acid (C), indolelactic acid (D) and indoxylsulfuric acid (E) involved in tryptophan metabolism was significantly raised by magnolol treatment. The value in the plot was expressed as means ± SEM, and statistically significant was marked by asterisk (* p < 0.05; ** p < 0.01). Full article ">Figure 6
The schematic in the anti-inflammatory effect of magnolol on DSS-induced colitis. Full article ">

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Open AccessReview

Cyclic Peptides as Novel Therapeutic Microbicides: Engineering of Human Defensin Mimetics

by Annarita Falanga, Ersilia Nigro, Margherita Gabriella De Biasi, Aurora Daniele, Giancarlo Morelli, Stefania Galdiero and Olga Scudiero

Molecules 2017, 22(7), 1217; https://doi.org/10.3390/molecules22071217 - 20 Jul 2017

Cited by 79 | Viewed by 8939

Abstract

Cyclic peptides are receiving significant attention thanks to their antimicrobial activity and high serum stability, which is useful to develop and design novel antimicrobial agents. Antimicrobial peptides appear to be key components of innate defences against bacteria, viruses, and fungi. Among the others, [...] Read more.

Cyclic peptides are receiving significant attention thanks to their antimicrobial activity and high serum stability, which is useful to develop and design novel antimicrobial agents. Antimicrobial peptides appear to be key components of innate defences against bacteria, viruses, and fungi. Among the others, defensins possess a strong microbicidial activity. Defensins are cationic and amphipathic peptides with six cysteine residues connected by three disulfide bonds found in plants, insects, and mammals; they are divided in three families: α-, β-, and θ-defensins. α-Defensins are contained in the primary granules of human neutrophils; β-defensins are expressed in human epithelia; and θ-defensins are pseudo-cyclic defensins not found in humans, but in rhesus macaques. The structural diversities among the three families are reflected in a different antimicrobial action as well as in serum stability. The engineering of these peptides is an exciting opportunity to obtain more functional antimicrobial molecules highlighting their potential as therapeutic agents. The present review reports the most recent advances in the field of cyclic peptides with a specific regard to defensin analogs. Full article

(This article belongs to the Special Issue Peptide-Based Drugs and Drug Delivery Systems)

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Open AccessArticle

Synthesis, Antitumor Evaluation and Molecular Docking of New Morpholine Based Heterocycles

by Zeinab A. Muhammad, Mastoura M. Edrees, Rasha A. M. Faty, Sobhi M. Gomha, Seham S. Alterary and Yahia N. Mabkhot

Molecules 2017, 22(7), 1211; https://doi.org/10.3390/molecules22071211 - 20 Jul 2017

Cited by 15 | Viewed by 6047

Abstract

A series of new morpholinylchalcones was prepared and then used as building blocks for constructing a series of 7-morpholino-2-thioxo-2,3-dihydropyrido[2,3-d]pyrimidin-4(1H)-ones via their reaction with 6-aminothiouracil. The latter thiones reacted with the appropriate hydrazonoyl chloride to give the corresponding pyrido[2,3-d [...] Read more.

A series of new morpholinylchalcones was prepared and then used as building blocks for constructing a series of 7-morpholino-2-thioxo-2,3-dihydropyrido[2,3-d]pyrimidin-4(1H)-ones via their reaction with 6-aminothiouracil. The latter thiones reacted with the appropriate hydrazonoyl chloride to give the corresponding pyrido[2,3-d][1,2,4]triazolo[4,3-a]pyrimidin-5(1H)-ones. The assigned structures for all the newly synthesized compounds were confirmed on the basis of elemental analyses and spectral data and the mechanisms of their formation were also discussed. Most of the synthesized compounds were tested for in vitro activity against human lung cancer (A-549) and human hepatocellular carcinoma (HepG-2) cell lines compared with the employed standard antitumor drug (cisplatin) and the results revealed that compounds 8, 4e and 7b have promising activities against the A-549 cell line (IC₅₀ values of 2.78 ± 0.86 μg/mL, 5.37 ± 0.95 μg/mL and 5.70 ± 0.91 μg/mL, respectively) while compound 7b has promising activity against the HepG-2 cell lines (IC₅₀ = 3.54 ± 1.11 μg/mL). Moreover, computational studies using MOE 2014.09 software supported the biological activity results. Full article

(This article belongs to the Collection Heterocyclic Compounds)

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Open AccessComment

The Hydrogen Sulfide-Vitamin B12-Folic Acid Axis: An Intriguing Issue in Chronic Kidney Disease. A Comment on Toohey JI: “Possible Involvement of Hydrosulfide in B12-Dependent Methyl Group Transfer”. Molecules 2017, 22, 582, pii: E582

by Giuseppe Cianciolo, Maria Cappuccilli and Gaetano La Manna

Molecules 2017, 22(7), 1216; https://doi.org/10.3390/molecules22071216 - 19 Jul 2017

Cited by 2 | Viewed by 4238

Abstract

Dear Editor, We read with great interest the recent article by John I. Toohey entitled “Possible Involvement of Hydrosulfide in B12-Dependent Methyl Group Transfer”, recently published in Molecules 2017, and we wish to discuss some additional insights raised by this important issue into [...] Read more.

Dear Editor, We read with great interest the recent article by John I. Toohey entitled “Possible Involvement of Hydrosulfide in B12-Dependent Methyl Group Transfer”, recently published in Molecules 2017, and we wish to discuss some additional insights raised by this important issue into the nephrological area [1].[...] Full article

(This article belongs to the Special Issue Sulfur Atom: Element for Adaptation to an Oxidative Environment 2016)

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Open AccessArticle

Identification of a Novel Vasodilatory Octapeptide from the Skin Secretion of the African Hyperoliid Frog, Kassina senegalensis

by Qiang Du, Hui Wang, Chengbang Ma, Yue Wu, Xinping Xi, Mei Zhou, Tianbao Chen, Chris Shaw and Lei Wang

Molecules 2017, 22(7), 1215; https://doi.org/10.3390/molecules22071215 - 19 Jul 2017

Cited by 4 | Viewed by 4464

Abstract

The defensive skin secretions of amphibians continue to be an excellent source of novel biologically-active peptides. Here we report the identification and pharmacological activity of a novel C-terminally amided myotropic octapeptide from the skin secretion of the African hyperoliid frog, Kassina senegalensis. [...] Read more.

The defensive skin secretions of amphibians continue to be an excellent source of novel biologically-active peptides. Here we report the identification and pharmacological activity of a novel C-terminally amided myotropic octapeptide from the skin secretion of the African hyperoliid frog, Kassina senegalensis. The 8-amino acid peptide has the following primary structure: WMSLGWSL-amide and has a molecular mass of 978 Da. The primary structure and organisation of the biosynthetic precursor of WL-8 amide was successfully deduced from cloned skin secretion-derived cDNA. The open-reading frame encoded a single copy of WL-8, located at the C-terminus. Synthetic WL-8 amide was found to cause relaxation of rat tail artery smooth muscle with an EC₅₀ of 25.98 nM. This peptide is unique in terms of its primary structure and is unlike any other peptide previously isolated from an amphibian source which has been archived in the NCBI database. WL-8 amide thus represents the prototype of a novel family of myotropic peptide from amphibian defensive skin secretions. Full article

(This article belongs to the Special Issue Bioactive Natural Peptides As A Pipeline For Therapeutics)

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Open AccessArticle

Molecular Weights of Bovine and Porcine Heparin Samples: Comparison of Chromatographic Methods and Results of a Collaborative Survey

by Sabrina Bertini, Giulia Risi, Marco Guerrini, Kevin Carrick, Anita Y. Szajek and Barbara Mulloy

Molecules 2017, 22(7), 1214; https://doi.org/10.3390/molecules22071214 - 19 Jul 2017

Cited by 11 | Viewed by 5984

Abstract

In a collaborative study involving six laboratories in the USA, Europe, and India the molecular weight distributions of a panel of heparin sodium samples were determined, in order to compare heparin sodium of bovine intestinal origin with that of bovine lung and porcine [...] Read more.

In a collaborative study involving six laboratories in the USA, Europe, and India the molecular weight distributions of a panel of heparin sodium samples were determined, in order to compare heparin sodium of bovine intestinal origin with that of bovine lung and porcine intestinal origin. Porcine samples met the current criteria as laid out in the USP Heparin Sodium monograph. Bovine lung heparin samples had consistently lower average molecular weights. Bovine intestinal heparin was variable in molecular weight; some samples fell below the USP limits, some fell within these limits and others fell above the upper limits. These data will inform the establishment of pharmacopeial acceptance criteria for heparin sodium derived from bovine intestinal mucosa. The method for MW determination as described in the USP monograph uses a single, broad standard calibrant to characterize the chromatographic profile of heparin sodium on high-resolution silica-based GPC columns. These columns may be short-lived in some laboratories. Using the panel of samples described above, methods based on the use of robust polymer-based columns have been developed. In addition to the use of the USP’s broad standard calibrant for heparin sodium with these columns, a set of conditions have been devised that allow light-scattering detected molecular weight characterization of heparin sodium, giving results that agree well with the monograph method. These findings may facilitate the validation of variant chromatographic methods with some practical advantages over the USP monograph method. Full article

(This article belongs to the Special Issue Looking Forward to the Future of Heparin: New Sources, Developments and Applications)

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Figure 1
Histogram plots of Phase 1 summary results for 24 heparin samples (see <a href="#app1-molecules-22-01214" class="html-app">Table S1</a>) (A) Mw, (B) M24,000 and (C) M8000–16,000/M16,000–24,000 for bovine lung heparin (green), porcine mucosal heparin (grey), and bovine mucosal heparin from sample donors 1 (orange), 3 (blue), 4 (pink), and 6 (yellow). Values recorded are the mean values from five or six laboratories (see <a href="#app1-molecules-22-01214" class="html-app">Table S2A–C</a>) The vertical lines indicate upper and lower limit acceptance criteria in the USP monograph for heparin sodium. Full article ">Figure 2
Phase 1: Molecular weight distributions for the samples of bovine mucosal heparin with highest (R-2) and lowest (E-2) molecular weights as measured by the USP Heparin Sodium monograph method. Chromatographic profiles refer to Method 1. Full article ">Figure 3
Phase 2: Overlay view of USP Heparin Sodium Identification RS chromatographic profiles in 12 distinct chromatographic systems. Panels: (A) Methods 1 and 2; (B) Methods 3–7; (C) Methods 8–12. Full article ">Figure 4
Phase 2: Column chart indicating similarity in molecular weight results for 24 heparin samples, between the USP Heparin Sodium monograph method (Method 1) and 11 other distinct chromatographic methods (<a href="#app1-molecules-22-01214" class="html-app">Table S5</a>). Blue columns plot Mw data and orange columns plot M24,000. Data are taken from <a href="#app1-molecules-22-01214" class="html-app">Table S3A,B</a>; similarity criteria are for Mw, values differ from Method 1 by less than 500 Da; for M24,000 values differ from Method 1 by less than 10%. Full article ">

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Open AccessArticle

The Anti-Inflammatory Properties of Citrus wilsonii Tanaka Extract in LPS-Induced RAW 264.7 and Primary Mouse Bone Marrow-Derived Dendritic Cells

by Liping Cheng, Yujie Ren, Dingbo Lin, Shu’ang Peng, Bo Zhong and Zhaocheng Ma

Molecules 2017, 22(7), 1213; https://doi.org/10.3390/molecules22071213 - 19 Jul 2017

Cited by 45 | Viewed by 8402

Abstract

‘Zhique’ (Citrus wilsonii Tanaka) is a traditional Chinese medicine. Its fruits have been used to treat inflammation-related symptoms, such as cough and sputum, though the underlying mechanism remains poorly understood. The aim of this study was to investigate the anti-inflammatory properties of ‘Zhique’ [...] Read more.

‘Zhique’ (Citrus wilsonii Tanaka) is a traditional Chinese medicine. Its fruits have been used to treat inflammation-related symptoms, such as cough and sputum, though the underlying mechanism remains poorly understood. The aim of this study was to investigate the anti-inflammatory properties of ‘Zhique’ pulp extract (ZQE) in lipopolysaccharide (LPS)-induced RAW 264.7 macrophages and primary mouse bone marrow-derived dendritic cells (BMDCs). The flavonoid profiles of the ZQE were determined by high performance liquid chromatography. The anti-inflammatory activity was evaluated in LPS-induced inflammatory RAW 264.7 macrophages and BMDCs through enzyme-linked immunosorbent assay, quantitative real-time polymerase chain reaction, and Western blot assays. Naringin was a predominant flavonoid occurring in ZQE, followed by eriocitrin, hesperidin, neohesperidin, rhoifolin, naringenin, and poncirin. ZQE exhibited a very low cytotoxicity in LPS-stimulated RAW 264.7 macrophages. Meanwhile, ZQE significantly inhibited the production of prostaglandins E2 and secretion of cyclooxygenase-2 protein in LPS-stimulated RAW 264.7 macrophages, and markedly suppressed the mRNA expression of inflammatory mediators, such as cyclooxygenase-2, tumor necrosis factor alpha, interleukin-1 beta (IL-1β), and IL-6 in LPS-induced RAW 264.7 macrophages and/or primary BMDCs. The ZQE inhibited the inflammatory responses in RAW 264.7 macrophages and BMDCs triggered by LPS. The results suggested that ‘Zhique’ has a high potential as a novel therapeutic agent to treat chronic inflammatory diseases. Full article

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The extraction methods used to isolate ‘Zhique’ pulp extract (ZQE) from ‘Zhique’ fruits. Full article ">Figure 2
High performance liquid chromatography (HPLC) analysis with detection at 283 nm (a) and 330 nm (b) of the ZQE. Samples were analyzed using 15–75% methanol in 0.3% formic acid for 70 min. 1: Eriocitrin, 2: Naringin, 3: Hespiridin, 4: Neohesperidin, 5: Rhoifolin, 6: Naringenin, 7: Poncirin. Full article ">Figure 3
Effects of ZQE on cell viability in RAW 264.7 macrophages. The cells were treated with ZQE (0, 62.5, 125, 250, 500, and 1000 μg/mL) in the presence of LPS (1 μg/mL) for 18 h. Cell viability was measured by a MTT assay. The data shown are from three independent experiments expressed as mean ± SD. * p < 0.05 compared to the control (without ZQE and LPS). LPS: lipopolysaccharide; MTT: Methylthiazolyldiphenyl-tetrazolium bromide. Full article ">Figure 4
Effects of ZQE on the PGE2 production in LPS-induced RAW 264.7 macrophages. The cells were pretreated with ZQE (250 μg/mL) and aspirin (250 μg/mL) in the presence of LPS (1 μg/mL) for 12 h, and aspirin was used as a positive control. The production of PGE2 in the supernatant were assayed using ELISA kits. The data shown are from three independent experiments and expressed as mean ± SD. * p < 0.05 and # p < 0.05 compared the control (with no LPS, ZQE, or aspirin) and LPS alone, respectively. LPS: lipopolysaccharide; PGE2: Prostaglandins E2; ELISA: enzyme-linked immunosorbent assay. Full article ">Figure 5
Effects of ZQE on LPS-induced COX-2, IL-1β and IL-6 expression in RAW 264.7 macrophages. The concentration of LPS is 1 μg/mL, and aspirin was used as a positive control, and the concentrations of ZQE and aspirin are both 250 μg/mL. (a) COX-2 mRNA expression; (b) COX-2 protein expression, β-actin as a loading control; (c) IL-1β mRNA expression; (d) IL-6 mRNA expression. The data shown are from three independent experiments and expressed as mean ± SD. * p < 0.05 and # p < 0.05 compared the control (with no LPS, ZQE, or aspirin) and LPS alone, respectively. COX-2, cyclooxygenase-2; LPS, lipopolysaccharide; PGE2, Prostaglandins E2; IL-1β, interleukin 1β; IL-6, interleukin 6. Full article ">Figure 6
Effects of ZQE on LPS-induced mRNA expression of COX-2, TNF-α, IL-1β, and IL-6 (a–d) in primary BMDCs. The concentrations of LPS and ZQE is 100 ng/mL and 1 μg/mL, respectively. The data shown are from three independent experiments and expressed as mean ± SD. * p < 0.05 and # p < 0.05 compared the control (with no LPS or ZQE) and LPS alone, respectively. LPS: lipopolysaccharide; COX-2: cyclooxygenase-2; TNF-α: tumor necrosis factor-alpha; IL-1β: interleukin 1β; IL-6: interleukin 6; BMDCs: bone marrow-derived dendritic cells. Full article ">

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Open AccessArticle

Isoegomaketone Alleviates the Development of Collagen Antibody-Induced Arthritis in Male Balb/c Mice

by Chang Hyun Jin, Yangkang So, Bomi Nam, Sung Nim Han and Jin-Baek Kim

Molecules 2017, 22(7), 1209; https://doi.org/10.3390/molecules22071209 - 19 Jul 2017

Cited by 9 | Viewed by 5734

Abstract

In this study, we attempted to identify and assess effects of isoegomaketone (IK) isolated from Perilla frutescens var. crispa on the development of rheumatoid arthritis (RA). RA was induced in male Balb/c mice by collagen antibody injection. Experimental animals were randomly divided into [...] Read more.

In this study, we attempted to identify and assess effects of isoegomaketone (IK) isolated from Perilla frutescens var. crispa on the development of rheumatoid arthritis (RA). RA was induced in male Balb/c mice by collagen antibody injection. Experimental animals were randomly divided into five groups: normal, collagen antibody-induced arthritis (CAIA), CAIA + IK (5 mg/kg/day), CAIA + IK (10 mg/kg/day), and CAIA + apigenin (16 mg/kg/day) and respective treatments were administered via oral gavage once per day for four days. Mice treated with IK (10 mg/kg/day) developed less severe arthritis than the control CAIA mice. Arthritic score, paw volume, and paw thickness were less significant compared to the control CAIA mice at day seven (73%, 15%, and 14% lower, respectively). Furthermore, histopathological examination of ankle for inflammation showed that infiltration of inflammatory cells and edema formation were reduced by IK treatment. Similarly, neutrophil to lymphocyte ratio (NLR) in whole blood was lower in mice treated with IK (10 mg/kg/day) by 85% when compared to CAIA mice. Taken together, treatment with IK delays the onset of the arthritis and alleviates the manifestations of arthritis in CAIA mice. Full article

(This article belongs to the Section Natural Products Chemistry)

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Image of representative microscopic features of knee joint (A) and mice joint (B). Isoegomaketone (IK) and apigenin (API) were administered via oral gavage once per day for 4 days. Arrow indicates infiltration of neutrophils and arrowhead indicates the necrosis. Full article ">Figure 2
Effect of IK and API on mean histopathological arthritis scores in CAIA mice. Results were expressed as a score (means ± SD) of six mice. * p < 0.05 vs. CAIA-group and ** p < 0.05 vs. IK(10 mg/kg)-group. Full article ">Figure 3
Effect of IK and API on paw volume in CAIA mice. Paw volume were measured using a Digital Plethysmometer every day after lipopolysaccharide (LPS) injection and oral administration of treatments. The average volume of both hind legs was used. Data are presented as means ± SD (n = 6). # p < 0.05 vs. PBS-group and * p < 0.05 vs. CAIA-group. Full article ">Figure 4
Effect of IK and API on paw thickness in CAIA mice. Paw thickness was measured using a digital caliper every day after LPS injection and oral administration of treatments. The average thickness of both hind legs was used. Data are presented as means ± SD (n = 6). # p < 0.05 vs. PBS-group and * p < 0.05 vs. CAIA-group. Full article ">Figure 5
Effect of IK and API on arthritic score in CAIA mice. Arthritic score was done blindly by using a system based on the number of inflamed joints in front and hind paws, inflammation being defined by swelling and redness at the scale from 0 (no redness and swelling) to 3 (severe swelling with joint rigidity or deformity; maximal score for four paws, 12). Data are presented as means ± SD (n = 6). # p < 0.05 vs. PBS-group, * p < 0.05 vs. CAIA-group, and ** p < 0.05 vs. IK(10 mg/kg)-group. Full article ">Figure 6
Effect of IK and API on neutrophil-to-lymphocyte ratio in CAIA mice. Whole blood samples were collected by cardiac puncture. Data are presented as means ± SD (n = 6). # p < 0.05 vs. normal-group and * p < 0.05 vs. CAIA-group. Full article ">

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Open AccessArticle

Structural Insight into the Recognition of r(UAG) by Musashi-1 RBD2, and Construction of a Model of Musashi-1 RBD1-2 Bound to the Minimum Target RNA

by Ryo Iwaoka, Takashi Nagata, Kengo Tsuda, Takao Imai, Hideyuki Okano, Naohiro Kobayashi and Masato Katahira

Molecules 2017, 22(7), 1207; https://doi.org/10.3390/molecules22071207 - 19 Jul 2017

Cited by 23 | Viewed by 7307

Abstract

Musashi-1 (Msi1) controls the maintenance of stem cells and tumorigenesis through binding to its target mRNAs and subsequent translational regulation. Msi1 has two RNA-binding domains (RBDs), RBD1 and RBD2, which recognize r(GUAG) and r(UAG), respectively. These minimal recognition sequences are connected by variable [...] Read more.

Musashi-1 (Msi1) controls the maintenance of stem cells and tumorigenesis through binding to its target mRNAs and subsequent translational regulation. Msi1 has two RNA-binding domains (RBDs), RBD1 and RBD2, which recognize r(GUAG) and r(UAG), respectively. These minimal recognition sequences are connected by variable linkers in the Msi1 target mRNAs, however, the molecular mechanism by which Msi1 recognizes its targets is not yet understood. We previously determined the solution structure of the Msi1 RBD1:r(GUAGU) complex. Here, we determined the first structure of the RBD2:r(GUAGU) complex. The structure revealed that the central trinucleotide, r(UAG), is specifically recognized by the intermolecular hydrogen-bonding and aromatic stacking interactions. Importantly, the C-terminal region, which is disordered in the free form, took a certain conformation, resembling a helix. The observation of chemical shift perturbation and intermolecular NOEs, together with increases in the heteronuclear steady-state {¹H}-¹⁵N NOE values on complex formation, indicated the involvement of the C-terminal region in RNA binding. On the basis of the two complex structures, we built a structural model of consecutive RBDs with r(UAGGUAG) containing both minimal recognition sequences, which resulted in no steric hindrance. The model suggests recognition of variable lengths (n) of the linker up to n = 50 may be possible. Full article

(This article belongs to the Special Issue Recent Advances in Biomolecular NMR Spectroscopy)

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Graphical abstract

Graphical abstract
Full article ">Figure 1
Interaction of r(GUAGU) with Msi1(109–200). (a) Domain organization of Msi1 (top), the protein construct used in this study (middle), and 5-mer RNA (bottom) originating from numb mRNA are shown; (b) Overlay of the 15N-1H HSQC spectra of Msi1(109–200) (80 μM) in the absence (black) and presence of r(GUAGU) with Msi1 RBD2:r(GUAGU) ratios of 1:0.25 (green), 1:0.50 (blue), 1:1.00 (yellow), and 1:1.30 (red); (c) The chemical shift perturbation, Δδ, was calculated using Δδ = [(ΔδH)2 + (ΔδN/6.5)2]1/2, where ΔδH and ΔδN are the chemical shift differences between free and complex forms for HN and 15N, respectively. The dashed line indicates the mean value (0.167 ppm). The secondary structure elements of Msi1(109–200) are shown at the top as blue arrows (strands) and yellow cylinders (helices). The amino acid residues that were not assigned (open circles) or whose 1H-15N resonance vanished upon addition of r(GUAGU) (closed circles) are indicated beneath the amino acid sequence at the bottom. Full article ">Figure 2
Solution structure of the Msi1 RBD2(109–191):r(GUAGU) complex. (a) Backbone traces of the 20 conformers of the complex are superimposed. Only the r(UAG) portion of r(GUAGU) is shown for RNA. The protein backbone (residues 109–190) is colored white. RNA is shown as a stick model: carbon (yellow), nitrogen (blue), oxygen (red), and phosphorus (orange); (b) Ribbon representation of the lowest energy conformer of the complex. RNA is shown as a ball-and-stick model: color-coded as in (a). Full article ">Figure 2 Cont.
Solution structure of the Msi1 RBD2(109–191):r(GUAGU) complex. (a) Backbone traces of the 20 conformers of the complex are superimposed. Only the r(UAG) portion of r(GUAGU) is shown for RNA. The protein backbone (residues 109–190) is colored white. RNA is shown as a stick model: carbon (yellow), nitrogen (blue), oxygen (red), and phosphorus (orange); (b) Ribbon representation of the lowest energy conformer of the complex. RNA is shown as a ball-and-stick model: color-coded as in (a). Full article ">Figure 3
r(UAG) recognition by Msi1 RBD2(109–191). (a) The Ura2 base fits in a shallow basin. Msi1 RBD2(109–190) is depicted as a ribbon model with a semi-transparent molecular surface. F112 (orange), E180 (green), and K182 (blue), whose side chains are shown (oxygen and nitrogen atoms in red and blue, respectively), form a rim, while G114 (magenta) and G115 (cyan) form the bottom of the basin. RNA is shown as a ball-and-stick model: carbon is color-coded yellow (Ura2) and white (Ade3), while nitrogen, oxygen, and phosphorus are color-coded blue, red, and yellow, respectively; (b) Recognition of Ura2. Intermolecular hydrogen bonds, Ura2 N3-E180 Oɛ and Ura2 O2-K182 Hζ, are formed; (c) Recognition of Ade3. The Ade3 base stacks onto the F112 aromatic ring, and the M190 side chain likely makes a van der Waals contact with Ade3, such that the adenine is sandwiched between F112 and M190. Hydrogen bonds, Ade3 H6-K183 O and Ade3 N1-Q185 HN, are also formed. Sδ of M190 is colored yellow; (d) Recognition of Gua4. The Gua4 base stacks onto the F154 aromatic ring, and hydrogen bonds, Gua4 N1-Q185 O, Gua4 H2-Q185 O, and Gua4 O6-K110 Hζ, are also formed. The M141 side chain makes a van der Waals contact with the ribose of Gua4. A salt bridge between K187 Hζ and the 5′ phosphate group of Gua4 may also be formed. Sδ of M141 is colored yellow. RNA is shown as a ball-and-stick model: carbon (yellow), nitrogen (blue), oxygen (red), and phosphorus (yellow). Hydrogen bonds are indicated by cyan dotted lines. Full article ">Figure 3 Cont.
r(UAG) recognition by Msi1 RBD2(109–191). (a) The Ura2 base fits in a shallow basin. Msi1 RBD2(109–190) is depicted as a ribbon model with a semi-transparent molecular surface. F112 (orange), E180 (green), and K182 (blue), whose side chains are shown (oxygen and nitrogen atoms in red and blue, respectively), form a rim, while G114 (magenta) and G115 (cyan) form the bottom of the basin. RNA is shown as a ball-and-stick model: carbon is color-coded yellow (Ura2) and white (Ade3), while nitrogen, oxygen, and phosphorus are color-coded blue, red, and yellow, respectively; (b) Recognition of Ura2. Intermolecular hydrogen bonds, Ura2 N3-E180 Oɛ and Ura2 O2-K182 Hζ, are formed; (c) Recognition of Ade3. The Ade3 base stacks onto the F112 aromatic ring, and the M190 side chain likely makes a van der Waals contact with Ade3, such that the adenine is sandwiched between F112 and M190. Hydrogen bonds, Ade3 H6-K183 O and Ade3 N1-Q185 HN, are also formed. Sδ of M190 is colored yellow; (d) Recognition of Gua4. The Gua4 base stacks onto the F154 aromatic ring, and hydrogen bonds, Gua4 N1-Q185 O, Gua4 H2-Q185 O, and Gua4 O6-K110 Hζ, are also formed. The M141 side chain makes a van der Waals contact with the ribose of Gua4. A salt bridge between K187 Hζ and the 5′ phosphate group of Gua4 may also be formed. Sδ of M141 is colored yellow. RNA is shown as a ball-and-stick model: carbon (yellow), nitrogen (blue), oxygen (red), and phosphorus (yellow). Hydrogen bonds are indicated by cyan dotted lines. Full article ">Figure 4
Comparison of the heteronuclear steady-state {1H}-15N nuclear Overhauser effect (NOE) values for Msi1(109–200) in its free and bound forms. The heteronuclear steady-state {1H}-15N NOE values are shown for Msi1(109–200) in the free form (black squares) and in the complex form with r(GUAGU) (red circles). The secondary structure elements (helices: yellow cylinders; β-strands: blue arrows) of Msi1(109–200) are shown at the top. The amino acid residues whose 1H-15N resonances were not assigned are indicated by black circles (free form) and a red square (complex form) beneath the amino acid sequence at the bottom. Daggers indicate the residues with overlapped 1H-15N resonances, while double-daggers indicate the residues whose 1H-15N resonance was missing in the free form. Full article ">Figure 5
A model structure of Msi1 RBD1-2 bound to the minimal target sequence, r(UAGGUAG). (a) Consecutive RBDs, RBD1-2, of Hrp1 (Lys156-His322), which are connected by a short linker (Ile234–Gly243), reportedly binds to r(UAUAUAUA). The structures of the Msi1 RBD1:r(GUAG) complex (green and yellow) and Hrp1 RBD1:r(AUAU) complex (pink and red) (left), and those of the Msi1 RBD2:r(UAG) complex (cyan and yellow) and Hrp1 RBD2:r(UAU) complex (pink and red) (right) are superimposed. Note that in both pairs, the binding regions are superimposed very well. One of the characteristic interactions between Msi1 RBD1 and r(GUAG), stacking of the aromatic rings of W29 and Gua1, was similarly found in the Hrp1 RBD1:RNA complex (W168 and Ade4) (labeled in the Figure); (b) A schematic illustration of the model building steps; (c) On the basis of the similarities found in (a), we used the solution structure of the Hrp1:r(UAUAUAUA) complex (PDB ID: 2CJK) (left) as a template and built a model structure containing the both RBDs of both Msi1 and r(UAGGUAG) (right). The W29 in Msi1 and W168 in Hrp1 are shown as green and magenta sticks, and labelled, respectively; (d) Alignment of the 3′-UTR regions of the Msi family target mRNAs, containing the r(UAGGUAG) sequence (highlighted in grey): H. sapiens APC (accession code NM_001127511.2), H. sapiens doublecortin (NM_000555.3), H. sapiens Jagged 1 (NM_000214.2), and H. sapiens Smad3 (NM_005902.3). Full article ">

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Molecules, Volume 22, Issue 7 (July 2017) – 209 articles

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