Simultaneous determination of five phenylethanoid glycosides in Cistanches Herba using quantitative analysis of multi-components by single marker

In the present study, a method for the quantitative analysis of multi-components by single marker(QAMS) has been developed and validated for the simultaneous determination of echinacoside(ECH), tubuloside A, acteoside, isoacteoside, and2’-acetylacteoside in Cistanches Herba. ECH was used as the internal standard(IS) to obtain the relative correction factors(RCFs) of the other four phenylethanoid glycosides(PhGs);meanwhile, various influencing factors on RCFs were investigated under different conditions. The content of each component was calculated with RCF. The results were compared with those obtained by the external standard method(ESM) to verify the feasibility and accuracy of the established QAMS method. No significant difference was found in the quantitative results of 10 batches of Cistanches Herba between QAMS and ESM. The proposed QAMS method for simultaneous determination of PhGs in Cistanches Herba is accurate and feasible, providing an efficient and economical approach for the quality control of Cistanches Herba.

Research progress of surface plasmon resonance technology in drug discovery

Surface plasmon resonance(SPR)technology is a powerful and sensitive tool for investigating molecular interactions.Technical improvements in recent years have brought new functionality to SPR apparatuses that can be widely applied at multiple stages of the drug discovery process.The technology allows users to conduct ligand fishing and offers rich and in-depth information regarding the affinity,specificity,kinetics,concentration and identification of covalent/allosteric/competitive binding behaviors.The present review highlights the principle,sample types,detection ranges,experimental methods,strengths,limitations,and the latest research progress on the application of SPR technology in the drug discovery process.

Cardioprotective effects of combination of notoginseng total saponins and safflower total flavonoids against myocardial infarction in rats

In this study, the cardioprotective mechanism of the combination of notoginseng total saponins and safflower total flavonoids（CNS） was investigated due to its excellently efficacy against myocardial infarction（MI） in rats. After the left anterior descending coronary artery（LADCA） ligation, rats were orally administered with CNS for 7 consecutive days. CNS prevented MI-induced pathophysiological changes and significantly decreased plasma levels of myocardial enzymes, including creatine kinase MB isoenzyme（CK-MB）, lactate dehydrogenase（LDH） and aspartate aminotransferase（AST）. Further investigation revealed that CNS attenuated the production of inflammatory factors in plasma, including tumor necrosis factor-alpha（TNF-α）, interleukin-6（IL-6） and interleukin-1β（IL-1β）. Moreover, CNS treatment decreased the expression of caspase-3 at the mR NA level in infarct tissue. Our findings demonstrated that the anti-inflammatory and anti-apoptotic properties of CNS might confer its cardioprotection against MI in rats.

Simultaneous quantification of evodiamine,rutaecarpine,and dehydroevodiamine in rat cerebrospinal fluid and cerebral nuclei after oral administration by UPLC-MS/MS

Evodiamine,rutaecarpine,and dehydroevodiamine have been demonstrated as the major alkaloids in the fruits of Euodia rutaecarpa,a well-known traditional Chinese medicine with central nervous system activities.To study their cerebrospinal fluid pharmacokinetics and cerebral nuclei distribution,the alkaloids were mixed at the weight ratio of 1:1:1 and orally administered via gavage to the rats at each dose of 15 mg/kg.A quick and reliable ultra-performance liquid chromatographic-tandem mass spectrometry method was developed and applied for the simultaneous analysis of the alkaloids in rat cerebrospinal fluid and cerebral nuclei collected at different time points.Non-compartmental pharmacokinetic profiles were calculated,and the distribution in cerebral nuclei was compared.All the tested compounds were absorbed into rat cerebrospinal fluid and distributed to the brain nuclei quickly.Their distribution in different nuclei varied,as evodiamine mainly in cerebellum and brainstem,rutaecarpine with its maximum in the brainstem,and dehydroevodiamine mostly in the cerebellum and hippocampus.They were eliminated from the brain rapidly without long-time accumulation.In summary,this study revealed the targeting discrepancy of evodiamine,rutaecarpine,and dehydroevodiamine in the brain,and highlighted the possibility for drug candidates in the encephalopathy treatment of the fruits of E.rutaecarpa.

Determination of inulin-type fructo-oligosaccharides in inulin by HPLC-ELSD

To develop an HPLC-ELSD method for the determination of nine inulin-type fructo-oligosaccharides in inulin,the HPLC-ELSD system consisted of Waters XBridge■ Amide column(4.6 mm×250 mm,5μm)with a gradient elution mobile phase consisting of acetonitrile and water at a flow rate of 1.2 mL/min at 30°C.The detector was an Agilent Technologies 380-ELSD.The drift tube temperature for the ELSD was set at 55°C with a nitrogen flow rate of 1.8 L/min.The injection volume was 15μL.The results showed that the detection range for the nine inulin-type fructo-oligosaccharides was 3.81–30.60μg R^(2)=0.99969 for kestose,3.73–29.97μg R^(2)=0.99981 for nystose,3.82–30.69μg R^(2)=0.99993 for fructosylnystose,3.80–30.48μg R^(2)=0.99995 for GF5,3.73–29.96μg R^(2)=0.99993 for GF6,3.78–30.30μg R^(2)=0.99983 for GF7,3.82–30μg R^(2)=0.99989 for GF8,3.71–29.80μg R^(2)=0.99974 for GF9,3.61–29.00μg R^(2)=0.99970 for GF10,respectively.The recovery of the nine oligosaccharides ranged between 96.48%–100.84%(n=6).The method was simple,accurate,and reproducible that it could be used as an analytical method for evaluating the quality of inulin effectively.

Applications of multiplexed immunohistochemistry/immunofluorescence and multispectral imaging technology in the field of tumor immunotherapy

Multiplexed immunohistochemistry/fluorescence(mIHC/IF)in combination with multispectral unmixing is a novel multitarget histopathological staining and imaging technique.By simultaneously revealing expression level and spatial information for up to eight biomarkers in situ,in addition to a nuclear stain within a single formalin-fixed paraffin-embedded(FFPE)tissue section,this technology can analyze the phenotype,abundance,morphology and intercellular relationship of cells while providing statistically significant results.In recent years,technical improvements have brought new insight into mIHC/IF and multispectral imaging approaches,which have been successfully applied in the field of cancer immunotherapy,specifically in regard to tumor microenvironment research,immunotherapy drug discovery,and prognostic and metastatic risk evaluation.This review highlights the principle,workflow,advantages and disadvantages of the technology,and discusses the latest applications of mIHC/IF-based imaging technology in the field of TME-related research and immunotherapy drug discovery.

Research on accurate quantitation technique using liquid chromatography coupled with triple quadrupole mass spectrometry and experiment optimization—take a novel diuretic as an example

Liquid chromatography coupled with the triple quadrupole mass spectrometry(LC-QQQ MS)technique is the most commonly used technique for quantitative analysis.It is widely used in pharmacology,targeted metabolomics,Chinese medicine quality control,and other research fields.The technique monitors only characteristic precursors and product ions through multiple reaction monitoring(MRM)mode and can detect targeted molecules in complex matrix with high specificity and sensitivity.In the present study,a diarylamide derivative diuretic was used as an example to introduce the method establishment and parameter optimization of this liquid chromatography-mass spectrometry technique.Diuretic and its internal standard could be completely separated within a 5-min gradient,and the quantitative linear range was 1–1000 ng/mL with R2=0.9996 in practical samples.This study showed that the key to the method development for LC-QQQ MS was the selection of the LC mobile phase,the elution gradient,the declustering potential,and the collision energy of compounds in MS.

Microscale thermophoresis in the investigation of biomolecular interactions

Accurate characterization of the interactions between biomolecules not only provides fundamental insights into cellular processes but also paves the way for drug discovery and development. With recent increases in throughput and sensitivity, biophysical technologies have become prominent tools for studying biomolecular interactions. Biophysical techniques that can reduce costs, shorten detection time, simplify the complexity of the system under analysis, and simultaneously provide high-quality data content are particularly favored. Here, we summarize the qualitative and quantitative analysis of biomolecular interactions using Micro Scale Thermophoresis(MST), as well as extend the application of MST functions to explore thermodynamics, enzyme kinetics and protein folding-unfolding processes. MST has emerged as a simple and powerful biophysical approach for identifying and quantifying binding events based on the movement of molecules along microscopic temperature gradients. The advantages of MST over other competitive biophysical techniques include freedom from immobilization, rapid analysis times, lower sample consumption, and the ability to analyze binding affinities in cell lysates. This article discusses the instrumental setups, principles, experimental workflows, and examples of MST application in practice.

High-throughput screening technologies for ion channel drug discovery

Ion channels are attractive targets for drug discovery as an increasing number of new ion channel targets have been uncovered in diseases, such as pain, cardiovascular disease, and neurological disorders. Despite their relevance in diseases and the variety of physiological functions they are involved in, ion channels still remain underexploited as drug targets. This, to a large extent, is attributed to the absence of screening technologies that ensure both the quality and the throughput of data. However, an increasing number of assays and technologies have evolved rapidly in the past decades. In this review, we summarized the currently available high-throughput screening technologies in ion channel drug discovery.

Integrated combination delivery of IDO inhibitor and paclitaxel for cancer treatment

In order to realize the combination of chemotherapy and immunotherapy,a reduction-responsive paclitaxel(PTX)prodrug PEG-SS-PTX was synthesized and used as a carrier to encapsulate IDO inhibitor CY-1-4 for preparing PEG-SS-PTX/CY-1-4 NPs.PEG-SS-PTX/CY-1-4 NPs were evaluated by cytotoxicity,immunogenic cell death(ICD)induction ability and anti-tumor efficacy.Dynamic light scattering(DLS)results showed that the size of PEG-SS-PTX/CY-1-4 NPs was about 149 nm.In vitro experiments indicated that its cytotoxicity was in a concentration-dependent manner,and it induced the ICD of B16-F10 cells.In vivo studies in melanoma mouse model indicated that PEG-SS-PTX/CY-1-4 NPs significantly inhibited the tumor growth and reduced the expression of IDO in tumor tissues.Moreover,it increased the rate of CD8+T cells in the spleen.In summary,PEG-SS-PTX/CY-1-4 NPs achieved good anti-tumor effects and reduced the dose of chemotherapy drugs,which was a safe and effective combined delivery system.

Proteomic analysis on cellular response induced by nanoparticles reveals the nano-trafficking pathway through epithelium

The application of nanomedicines in oral drug delivery effectively promotes the drug absorption and transportation through enterocytes.Nevertheless,the absence of mechanism studies on efficacy and safety limits their final translation in humans.Although the vesicular trafficking has been verified as the general character for transport of nanomedicines,the deeper mechanism in molecular mechanism is still unclear.Moreover,the cellular transport of nanomedicines is a dynamic process involved by different organelles and components.However,most of existing studies just pay attention to the static location of nanomedicines,but neglect the dynamic biological effects on cells caused by them.Here,we prepared gold nanoparticles(Au NPs)as the model and cultured epithelial cell monolayer to explore the nano-bio interactions at the molecular level.The traditional pharmacological inhibition strategy and subcellular imaging technology elucidated the macropinocytosis/endosome/MVB/lysosome pathway during the transportation of Au NPs.Proteomics strategy based on mass spectrometry(MS)was utilized to identify and quantify proteins involved in the cellular transport of nanomedicines.Multiple proteins related to subcellular structure,signal transduction,energy transformation and metabolism regulation were demonstrated to be regulated by nanoparticle transport.These alterations of protein expression clarified the effects of intracellular proteins and verified the conventional findings.More importantly,it revealed a feedback mechanism of cells to the nano-trafficking.We believed that these new regulatory mechanisms provided new insights into the efficient transport of nanomedicines through epithelial barriers.

Chemical constituents of the aerial parts of Glycyrrhiza uralensis and their inhibitory activities against PTP1B andα-glucosidase

In the present study,a total of 11 compounds were isolated from the aerial parts of Glycyrrhiza uralensis,including two new compounds,glycyuralin Q(1)and glycyuralin R(2),and nine known compounds,including licoriphenone(3),orobol(4),trifoliol(5),7,2′,4′-trihydroxy-5-methoxy-3-arylcoumarin(6),11-hydroxy-9(Z),12(Z)-octadecadienoic acid(7),11-hydroxy-9(E),12(E)-octadecadienoic acid(8),licoricone(9),glycyrin(10),and 2′-hydroxyformononetin(11).Structures of the new compounds were identified by 1 D,2 D NMR and HR-MS data analyses.Compounds 1,2 and 10 showed potent inhibitory activities against PTP1 B,with IC50 values of 1.43,4.71 and 3.79μM,respectively.Compounds 2,4 and 10 inhibitedα-glucosidase with IC50 values of 13.61,11.13 and 17.48μM,respectively.

Synthesis of two substrate mimics of thioesterase in the biosynthesis of cyclic depsipeptide WS9326A

WS9326 A is a tachykinin receptor antagonist and quorum sensing inhibitor discovered from several Streptomyces strains.The structure of WS9326 A features a(Z)-pentenylcinnamoyl moiety attached on a cyclic depsipeptide skeleton,which is biosynthesized by nonribosomal peptide synthetases(NRPS).The regioselective cyclization in the last step of NRPS catalysis,which is proposed to be catalyzed by a thioesterase(TE)domain in the last module,has not been experimentally characterized.We here report the synthesis of two substrate mimics(1 and 2)of the TE(WS9326 A-TE)in WS9326 A biosynthesis,by using Fmoc-based solid-phase peptide synthesis(SPPS)method.Compounds 1 and 2 are new compounds whose structures have been elucidated based on NMR and HRESIMS analyses.The N-terminal cinnamoyl moiety and C-terminal methylated L-Ser moiety in 2 were incorporated under the mild SPPS conditions.Given the isolation difficulties of substrate of WS9326 A-TE from the Streptomyces producers of WS9326 A,our synthesis of 1 and 2 set the stage for the reconstitution of WS9326 A-TE’s catalytic reaction in vitro in the future.

Synthesis of a series of novel homo-and hetero-glycoclusters and their binding activities to DC-SIGN

As a dendritic cell-specific C-type lectin receptor, DC-SIGN plays an important role in the early stages of many viral infections, including HIV and Ebola, making it an interesting therapeutic target. It has been found that DC-SIGN can recognize both highly mannosylated and branched fucosylated oligosaccharides. Herein, we synthesized a new series of homo-and Man-Fuc heteroglycoclusters with diverse structures. The binding properties of these compounds to tetrameric extracellular DC-SIGN were assessed by surface plasmon resonance(SPR). Heteroglycocluster 17 b showed high DC-SIGN-binding activity(K;= 2.6 μM). The structural determinants of this high affinity of 17 b were rationalized by docking and compared with its much less potent isomer 17 a. Therefore, 17 b might serve as a base for the development of potent inhibitors of DC-SIGN-dependent viral infection.

A concise total synthesis of secoisolariciresinol

A concise total synthesis of(R,R)-secoisolariciresinol was achieved in four steps, featuring a biomimetic β-β’ radical coupling of two dihydroferulic acid derivatives. The conversion of secoisolariciresinol to other related lignans was ongoing.

Surface-assisted laser desorption/ionization mass spectrometry for drug and metabolite analysis

Surface-assisted laser desorption/ionization mass spectrometry(SALDI-MS)uses inorganic nanomaterials as matrixes to facilitate desorption and ionization of analytes.Compared with the traditional matrix-assisted laser desorption/ionization mass spectrometry(MALDI-MS)technique,SALDI-MS has the advantages of less interference in the low mass range,better reproducibility and higher salt tolerance.It is highly suitable for the analysis of small molecule compounds.In recent years,researchers have developed a range of nanomaterials that are successfully applied to the field of small molecule drug and metabolite analysis including drug screening and quantification,drug delivery,metabolite profiling,biomarker discovery and so forth.This review summarizes the latest progress of SALDI-MS matrix materials such as metal-based,carbon-based,silica-based nanomaterials and organic framework nanomaterials and their applications.In addition,our perspective of SALDI-MS technology is also discussed for further advancement.

Microfluidic models in liver drug metabolism research

In pre-clinical phase of new drug development,it is particularly important to establish an in vitro model to mimic the metabolism situation of human body.The aim of the in vitro model is to reduce the usage of experimental animals and to make a more accurate prediction of the drug metabolism in vivo.Microfluidic chip is an emerging technology to establish predictive models.By integrating subcellular fractions,hepatocytes or liver tissue in the microfluidic chips,more predictive in vitro metabolism models can be established for drug development.The microfluidic platform offers dynamic and controlled fluids,as well as sophisticated liver tissue assembly to remodel the physiological and pathological microenvironment of liver in the human body.This review updates the microfluidic-based liver drug metabolism models since 2011,and summarizes the development of different models based on different chip vectors(subcellular components,primary hepatocytes,and tissue sections).It serves as a guide for newcomers to this dynamic field.

Discovery of tryptamine derivatives from Bacillus sp.PKU-TA00001

Tryptamine-derived natural products have been discovered from different sources including animals, plants and bacteria,and they show various biological activities. However, they are not discovered widely compared with the large amounts of tryptamine derivatives generated by chemical synthesis. We here report the discovery of five tryptamine-derived natural products(1-5) and one known polyketide 6 from Bacillus sp. PKU-TA00001. Compounds 1 and 2 are new compounds featuring methyl-hexanamide moieties, compound 4 is first discovered as a natural product, and 3 and 4’s NMR data are first provided. All compounds showed MIC(minimum inhibitory concentration) values higher than 50 μM against several Gram-positive and negative strains, and showed no cytotoxicity at the concentration of 100 μM against the human cancer cell lines A549, HCT-8 and MCF-7. The discovery of 1-4 expands the structural diversity of tryptamine-derived natural products, and sets the stage for revealing their biosynthetic mechanisms and biological activities in the future.

Synthetic studies toward resveratrol-based natural products polynapstilbenes A and B

Resveratrol-based natural products have received considerable attention as synthetic targets due to their versatile bioactivities and unique structures.Herein,we disclose our efforts toward the syntheses of polynapstilbenes A and B,which possess[C8-O-C-C-C7]-type dihydrobenzofuran skeleton that is distinctive compared with the other reported resveratrol-derived natural products.Our approach,featuring an acid-catalyzed conjugate addition followed by cyclization of para-quinone methides and phenols,affords two advanced intermediates that represents the dimethyl-protected aglycon of polynapstilbenes A and B.

Ela tablets improve sexual performance in mice with erectile dysfunction caused by repeated restraint stress

Ela tablets contain an optimized extract of a Chinese herbal prescription. The original prescription has been used to treat erectile dysfunction(ED), while the detailed indications and specific treatment mechanisms of Ela tablets remain unclear. In the present study, we aimed to investigate whether Ela tablets could protect mice from ED caused by repeated restraint stress and to explore the possible mechanisms. Mice were restrained daily in centrifuge tubes with venting holes for 14 consecutive days and administered Ela tablets at 1 h before restraining. On the 14 th night, mating experiments were conducted. Thereafter, blood, testicular, and penile samples were collected. Serum testosterone levels were measured using a kit. Testicular tissue morphology was observed by H&E staining. The PDE5 expression in the corpus cavernosum was measured by Western blotting analysis. The results indicated that Ela tablets significantly shortened the sexual arousal time, increased the number of sexual encounters, maintained the normal morphology of testicular tissue, increased the serum testosterone levels, and decreased the PDE5 expression in the corpus cavernosum. Our data showed that Ela tablets elicited protective effects on the sexual ability of mice exposed to repeated restraint stress. Moreover, the underlying mechanism was related to the preservation of testicular tissue morphology, maintenance of testosterone levels, and inhibition of penile PDE5 expression.