Potassium carbonate-catalyzed (3 + 2) cycloaddition reaction between N-2,2,2-trifluoroethylisatin ketimines and azodicarboxylates has been developed, constructing a series of novel N-heterocycle infused spirooxindoles in good to excellent yields (up to 98%) under milder conditions. The presence of both biologically active oxindole and trifluoromethyl-1,2,4-triazoline moieties in these novel spirocyclic compounds would provide new lead structures in the discovery of heterocyclic compounds with potential pharmaceutical activities.

Designing photoelectrochemical sensors for detecting ultra-trace amounts of Aflatoxin B1 (AFB1) in complex samples poses a significant challenge. In pursuit of this objective, we engineered hollow-structured ZnCdS (denoted as h-Zn1Cd5S) material and applied an electrodeposition method to introduce Au nanoparticles (AuNPs) on the surface. This approach resulted in a significant enhancement of the photocurrent response of the photoelectrode, attributed to the synergistic effects of the hollow structure and surface plasmon resonance, thereby underscoring its potential for advanced applications. Utilizing density functional theory, we computed the transfer route and energy band formation of photogenerated electrons. Subsequently, we developed an AFB1 photoelectrochemical (PEC) aptasensor based on the competition mechanism, coupled with aptamer probe technology. This aptasensor allows for the ultra-trace determination of AFB1, exhibiting a remarkable detection limit as low as 1.32 fg/mL and a linear range from 1.0×10-2 to 100.0pg/mL. The PEC aptasensor for AFB1 demonstrates excellent sensitivity and selectivity. When applied to the detection of AFB1 in Tsaoko, wheat, and corn samples, its performance aligns closely with high-performance liquid chromatography (HPLC) results, validating its applicability for analyzing real-world samples.

Endometrial cancer (EC) is one of the most common malignant tumors in women, and the increasing incidence and mortality pose a serious threat to the public health. Early diagnosis of EC could prolong the survival period and optimize the survivorship, greatly alleviating patients’ suffering and social medical pressure. In this study, we collected urine and serum samples from the recruited patients, analyzed the samples using LC-MS approach, and identified the differential metabolites through metabolomic analysis. Then, the differentially expressed genes were identified through the systematic transcriptomic analysis of EC-related dataset from Gene Expression Omnibus (GEO), followed by network profiling of metabolic-reaction-enzyme-gene. In this experiment, a total of 83 differential metabolites and 19 hub genes were discovered, of which 10 different metabolites and 3 hub genes were further evaluated as more potential biomarkers based on network analysis. According to the KEGG enrichment analysis, the potential biomarkers and gene-encoded proteins were found to be involved in the arginine and proline metabolism, histidine metabolism, and pyrimidine metabolism, which was of significance for the early diagnosis of EC. In particular, the combination of metabolites (histamine, 1-methylhistamine, and methylimidazole acetaldehyde) as well as the combination of RRM2, TYMS and TK1 exerted more accurate discrimination abilities between EC and healthy groups, providing more criteria for the early diagnosis of EC.

Red Yeast Rice (RYR) is a functional product prepared by fermenting rice with the fungus Monascus purpureus which has long been used as a traditional food and medicine. However, little is known about the effects of RYR on the body's overall metabolism. The untargeted and pseudotargeted metabolomics with HPLC-MS integrating network analysis were used to reveal the interventional effects of RYR administrated to rats. Serum untargeted metabolomics screened 187 multiple reaction monitoring (MRM) ion transitions. Five hundred and forty-six MRM ion transitions were detected by pseudotargeted metabolomics. Significant differentiation between the RYR and control groups was observed through principal component analysis (PCA) and systematic cluster analysis. With the assistance of orthogonal partial least squares discriminant analysis (OPLS-DA) and analysis of variance, 24 metabolites were identified as differentially expressed. Three key differential metabolites were screened including LPC (P-18:1), PI (34:2), and LPC (17:2). Pathway analysis revealed that RYR could influence lipid metabolism in rats by regulating glycerophospholipid metabolism and D-glutamine and D-glutamate metabolic pathways. Nineteen hub genes, including GOT1, GOT2, PLD2, and GGT1, were ultimately screened through integrative network analysis.

Since the overexpression of folate receptors (FRs) in certain types of cancers, a variety of FR-targeted fluorescent probes for tumor detection have been developed. However, the reported probes almost all have the same targeting ligand of folic acid with various fluorophores and/or linkers. In the present study, a series of novel tumor-targeted near-infrared (NIR) molecular fluorescent probes were designed and synthesized based on previously reported 6-substituted pyrrolo[2,3-d]pyrimidine antifolates. All newly synthesized probes showed specific FR binding in vitro, whereas GT-NIR-4 and GT-NIR-5 with a benzene and a thiophene ring, respectively, on the side chain of pyrrolo[2,3-d]pyrimidine exhibited better FR binding affinity than that of GT-NIR-6 with folic acid as targeting ligand. GT-NIR-4 also showed high tumor uptake in KB tumor-bearing mice with good pharmacokinetic properties and biological safety. This work demonstrates the first attempt to replace folic acid with antifolates as targeting ligands for tumor-targeted NIR probes.

Tripterygium glycosides tablets (TGT) are the commonly used preparation for rheumatoid arthritis (RA). However, the changes in TGT on RA are still unclear at the metabolic level. This study aimed to reveal the biological processes of TGT in collagen-induced arthritis (CIA) rats through integrated metabolomics and network analysis. First, the CIA model in rats was established, and the CIA rats were given three doses of TGT. Then, the endogenous metabolites in the serum from normal rats, CIA rats, and CIA rats treated with varying doses of TGT were detected by UHPLC-QTOF-MS/MS. Next, univariate and multivariate statistical analyses were performed to find the differential metabolites. Finally, differential metabolites, metabolic pathways, and hub genes were analyzed integrally to reveal the biological processes of TGT in CIA rats. The paw diameter, arthritis score, immunoglobulin G (IgG) concentration, CT image, and histological assay showed that TGT had evident therapeutic effects on CIA rats. Untargeted metabolomics revealed that TGT could ameliorate the down-regulation of lipid levels in CIA rats. Four key differential metabolites were found including LysoP(18:0), LysoPA(20:4), LysoPA(18:2), and PS(O-20:0/17:1). The glycerophospholipid metabolic pathway was perturbed in treating CIA with TGT. A total of 24 genes, including PLD1, LPCAT4, AGPAT1, and PLA2G4A, were found to be the hub genes of TGT in CIA rats. In conclusion, the integrated analysis provided a novel and holistic perspective on the biological processes of TGT in CIA rats, which could give helpful guidance for further TGT on RA. Future studies based on human samples are necessary.

Background: Tripterygium glycoside tablets (TGT) is the most common preparation from Tripterygium wilfordii Hook F, which is widely used in clinical for treating rheumatoid arthritis (RA) and other autoimmune diseases. However, its serious reproductive toxicity limits its application. Purpose: This study aimed to elucidate the toxic effects of TGT on the reproductive system of male RA rats and its potential toxic components and mechanism. Methods: Collagen-induced arthritis (CIA) rat model was established, and TGT suspension was given at low, medium, and high doses. Gonadal index, pathological changes, and the number of spermatogenic cells were used to evaluate the toxic effects of TGT on the reproductive system. Non-targeted metabolomics of testicular tissue was conducted by UHPLC-QTOF/MS. Combined with network toxicology, the key targets of TGT-induced reproductive toxicity were screened and RT-qPCR was used to validation. In vitro toxicity of 19 components of TGT was evaluated using TM3 and TM4 cell lines. Molecular docking was used to predict the interaction between toxic components and key targets. Results: TGT reduced testicular and epididymis weight. Pathology analysis showed a lot of deformed and atrophic spermatogenic tubules. The number of spermatogenic cells decreased significantly (P<0.0001). A total of 58 different metabolites including platelet-activating factor (PAF), lysophosphatidylcholine (Lyso PC), phosphatidylinositol (PI), glutathione (GSH), and adenosine monophosphate (AMP) were identified by testicular metabolomics. Glycerophospholipid metabolism, ether lipid metabolism, and glutathione metabolism were key pathways responsible for the reproductive toxicity of TGT. Ten key reproductive toxicity targets were screened by network toxicology. The cytotoxicity test showed that triptolide, triptonide, celastrol, and demethylzeylasteral could significantly reduce the viability of TM3 and TM4 cells. Alkaloids had no apparent toxic effects. Molecular docking showed that the four toxic components had a good affinity with 10 key targets. All binding energies were less than -7 kcal/mol. The RT-qPCR results showed the Cyp19a1 level was significantly up-regulated. Pik3ca and Pik3cg levels were significantly down-regulated. Conclusion: Through testicular metabolomics, we found that TGT may cause reproductive toxicity through CYP19A1, PIK3CA, and PIK3CG three target, which was preliminarily revealed. This study laid the foundation for elucidating the toxicity mechanism of TGT and evaluating its safety and quality.

Herein, we report an unprecedented intramolecular cross-nucleophile coupling strategy of indole tethered β-amino acrylates using a catalyst system combining λ3-iodanes and Lewis acids to achieve the chemodivergent synthesis of three unique alkaloid skeletons. It was worth noting that the acquisition of spiroindolenines and azepino[4,5-b]indoles derivatives was switchable with choice of the Lewis acids. Moreover, the polycyclic spiroindolines containing a lactone fragment could also be accessed for the first time via cross-nucleophile coupling cascade intramolecular condensation sequence.

A novel benzylbenzimidazolium based dicationic ionic liquids modified magnetic nanoparticles (MNP@BDILs) was successfully synthesized by a simple one-step reaction at mild condition and employed as adsorbent for determination of polycyclic aromatic hydrocarbons (PAHs) by magnetic solid-phase extraction (MSPE) coupled with high performance liquid chromatography (HPLC). Main parameters affecting the MSPE efficiency, including extraction time, extraction solvent, desorption solvent, desorption time and the amount of sorbent were studied. Under the optimized conditions, the proposed method provided good linearity with determination coefficients over 0.9991, and excellent reproducibility. Finally, for the determination of six PAHs in soil, milk and black wolfberry, satisfactory spiked recoveries were between 91.2% and 104.1% with the relative standard deviations below 6.5%. The results demonstrated that MNP@BDILs synthesized by the simple one-step reaction was a promising strategy for the application in sample preparation.

Twelve new compounds, aspergorakhins A-L (1-12) coupled with one known xanthone leptosphaerin D (13), were isolated from the extract of soil-derived fungus Aspergillus gorakhpurensis F07ZB1707. Their structures were elucidated by spectroscopic data analysis including UV, IR, NMR, and HRESIMS. The absolute configurations of 5 and 8-11 were identified using ECD and OR calculations. All compounds were tested by enzyme inhibitory activity assay in vitro. Aspergorakhin A (1) showed selective activities against PTP1B and SHP1 over TCPTP with IC50 values 0.57, 1.19, and 22.97 μM, respectively. Compounds 1 and 2 exhibited modest cytotoxicity against tumor cell lines A549, HeLa, Bel-7402, and SMMC-7721 with IC50 values in the range of 6.75-83.4 μM.

A major challenge in drug development is that the majority of drugs are water insoluble, and a powerful method to conquer this obstacle is to transfer a crystalline drug into its amorphous phase (AP) or coamorphous phase (CAP) with a coformer. In the present study, the physical and chemical stabilities of an AP and a CAP based on the dihydropyridine calcium ion antagonist azelnidipine (AZE) were investigated using thermal analysis and a solution chemistry method. The identification of two APs (named α-AP and β-AP, from crystalline α-AZE and β-AZE, respectively) and one AZE-piperazine CAP was attempted using powder X-ray diffraction, temperature modulated differential scanning calorimetry and Fourier-transform infrared spectroscopy. The transition thermodynamics from the two APs and the CAP to stable crystalline β-AZE (β-Cry) were investigated using a solubility method. The solubility of the two APs, the CAP and β-Cry in 0.01 M HCl medium at 298, 304, 310, 316 and 322 K was investigated; the values obtained were used to calculate the thermodynamic parameters of the transition reaction. The transition temperatures of α-AP, β-AP and the CAP to form β-Cry in 0.01 M HCl were 237.7, 400.3, and 231.4 K, respectively. The glass transition temperature (T g) values of α-AP, β-AP and the CAP were 365.5, 358.9 and 347.6 K, respectively, indicating a high physical stability for α-AP. However, β-AP proved to be the most thermodynamically stable form at room temperature compared with α-AP and CAP in the 0.01 M HCl medium. As evidenced by those observations, no general relationship occurred between the solid physical stability and the solution chemical stability for AP and CAP. The kinetics of the solid-state decomposition, studied using DSC analysis, showed that the activation energies for decomposition of α-AP, β-AP and CAP at high temperatures were 133.0, 114.2 and 131.6 kJ mol-1, respectively.

Reverse micelles have good thermodynamic stability, small particle size, good self-assembly performance, and controlled drug release versus other microparticle transdermal drug delivery systems. Here, a reverse micelles solution and a gel system formed by self-assembly of insulin aqueous solution/lecithin/isopropyl myristate (IPM) were used for transdermal delivery of insulin. The size, size distribution, morphology and rheology of the reverse micelles were characterized with dynamic light scattering (DLS), small angle x-ray scattering (SAXS), differential scanning calorimetry (DSC), viscosity measurement, and freeze fracture transmission electron microscopy (FF-TEM). The findings suggested that lecithin/isopropyl myristate self-assembled reverse micelles were spherical with uniform particle sizes (< 100nm). The water in the reverse micelle was in the form of bound water, and the diameter of the water pool was about 4nm. The reverse micelle solution and gel appeared as a pseudo-plastic fluid. The gel was thixotropic, which benefits transdermal delivery.

The marine mangrove Xylocarpus granatum is used as a folk medicine and is rich in bioactive limonoids. The quantitative determination of the chemical composition and distribution of limonoids in different parts of X. granatum fruit (fruit peel, seed coat, seed kernels, seed, and fruit) is significant for authentication and quality control purposes. However, the quantitative determination of limonoids in X. granatum has not yet been reported. In this study, a chemometric-assisted liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed and validated for the simultaneous determination of 17 limonoids to reveal the chemical composition and distribution in different parts of X. granatum fruit. Ultrasonic-assisted extraction, optimized by response surface methodology (RSM), was more accurate than the general one-variable-at-a-time method. The overall distribution of 17 limonoids in different parts of X. granatum fruit had the following order: seed kernels > seed > fruit, and 13 limonoids showed a rank order of seed kernels > seed > fruit > fruit peel > seed coat. Furthermore, principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) were used to analyze the LC–MS/MS data and provide a chemometric model for easy visualization and interpretation to classify the different parts of X. granatum fruit. In addition, the study indicated that the chemometric-assisted strategy, consisting of RSM, PCA, and OPLS-DA for the development, optimization, and data analysis of multicomponent quantitation by LC–MS/MS, is effective and feasible. This study provided the chemical composition and distribution evidence for the authentication and quality control of X. granatum fruit.

OBJECTIVE:A RP-HPLC method was developed for simultaneous determination of bigelovin, ergolide and tomentosin in Inula hupehensis.METHOD:An Agilent C18 column (4.6 mm x 250 mm, 5 microm) was used for separation at 40 degrees C. The mobile phase was acetonitrile-water, and the flow rate was 1.2 mL x min(-1). The detection wavelength was set at 210 nm.RESULT:The method has good linearity in the ranges of 0.01792-0.1792 g x L(-1) (r =0.9999) for bigelovin, 0.0424-0.4240 g x L(-1) (r =0.9996) for ergolide, and 0.044 8-0.4480 g x L(-1) (r = 0.9996) for tomentosin. The average recoveries of bigelovin, ergolide, and tomentosin were 98.5%, 98.2%, 98.4%, with the RSD of 1. 3%, 1.3%, 1.7%, respectively. The results demonstrated that there was a significant difference in the contents of three sequterpene lactones among the tested Inulae Flos.CONCLUSION:The results indicated that the present RP-HPLC method is simple, quick and accurate, and can be used for the quality control of I. hupehensis, especially for the authentication of Inulae Flos.

Voltage-gated Kv7/KCNQ/M-potassium channels play a pivotal role in controlling neuronal excitability. Genetic reduction of KCNQ channel activity as a result of mutations causes various human diseases such as epilepsy and arrhythmia. Therefore, discovery of small molecules that activate KCNQ channels is an important strategy for clinical intervention of membrane excitability related disorders. In this study, a series of pyrazolo[1,5-a]pyrimidin-7(4H)-ones (PPOs) have been found to be novel activators (openers) of KCNQ2/3 potassium channels through high-throughput screening by using atomic absorption rubidium efflux assay. Based on structure-activity relationship (SAR), the substituted PPOs have been optimized. The 5-(2,6-dichloro-5-fluoropyridin-3-yl)-3-phenyl-2-(trifluoromethyl) pyrazolo[1,5-a]pyrimidin-7(4H)-one (17) was identified as a novel, potent, and selective KCNQ2/3 potassium channel opener by patch-clamp recording assay.

BACKGROUND AND PURPOSECelecoxib is a selective cyclooxygenase‐2 (COX‐2) inhibitor used for the treatment of pain and inflammation. Emerging and accumulating evidence suggests that celecoxib can affect cellular targets other than COX, such as ion channels. In this study, we characterized the effects of celecoxib on Kv7 K+ channels and compared its effects with the well‐established Kv7 channel opener retigabine.EXPERIMENTAL APPROACHA perforated whole‐cell patch technique was used to record Kv7currents expressed in HEK 293 cells and M‐type currents from rat superior cervical ganglion neurons.KEY RESULTSCelecoxib enhanced Kv7.2–7.4, Kv7.2/7.3 and Kv7.3/7.5 currents but inhibited Kv7.1 and Kv7.1/KCNE1 currents and these effects were concentration dependent. The IC50 value for inhibition of Kv7.1 channels was approximately 4 µM and the EC50 values for activation of Kv7.2–7.4, Kv7.2/Kv7.3 and Kv7.3/Kv7.5 channels were approximately 2–5 µM. The effects of celecoxib were manifested by increasing current amplitudes, shifting the voltage‐dependent activation curve in a more negative direction and slowing the deactivation of Kv7 currents. 2,5‐Dimethyl‐celecoxib, a celecoxib analogue devoid of COX inhibition activity, has similar but greater effects on Kv7currents. Kv7.2(A235T) and Kv7.2(W236L) mutant channels, which have greatly attenuated responses to retigabine, showed a reversed response to celecoxib, from activation to inhibition.CONCLUSIONS AND IMPLICATIONSThese results suggest that Kv7 channels are targets of celecoxib action and provide new mechanistic evidence for understanding the effects of celecoxib. They also provide a new approach to developing Kv7 modulators and for studying the structure–function relationship of Kv7 channels.

A simple, sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous determination of m-nisoldipine and its three metabolites in rat plasma has been developed using nitrendipine as an internal standard (IS). Following liquid-liquid extraction, the analytes were separated using an isocratic mobile phase on a reverse phase C(18) column and analyzed by MS in the multiple reaction monitoring (MRM) mode. To avoid contamination by residual sample in the injection syringe, a special injection protocol was developed. We found that m-nisoldipine, metabolite M1 and IS could be ionized under positive or negative electrospray ionization conditions, whereas metabolite M and M2 could only be ionized in the positive mode. The mass spectrometry fragmentation pathways for these analytes are analyzed and discussed herein. The total analysis time required less than 5 min per sample. We employed this method successfully to study the metabolism of m-nisoldipine when it was orally administered to rats at a dose of 9 mg/kg. Three metabolites of m-nisoldipine and an unknown compound of molecular weight 386 were found for the first time in rat plasma. The concentration of the potentially active metabolite was approximately equal to its parent compound concentration.

A reversed-phase high-performance liquid chromatographic method was developed for the first time to simultaneously determine six major sesquiterpene lactones in Inula britannica L, that is, britannilactone, 1-O-acetylbritannilactone, hydroxylantolactone, 8-epi-ivangustin, 6-isobutylbritannilactone and 1,6-O,O-diacetylbritannilactone. The chromatographic separation was achieved with a linear gradient of acetonitrile in water with a flow rate at 1.0 mL min−1 using an Inertsil ODS-3 column (250 × 4.6 mm, 5.0 μm). Detection was carried out using a photodiode array detector at 210 nm. The calibration curves for the determination of all analytes showed good linearity over the investigated ranges (R 2 > 0.999). Repeatability was evaluated by intra-day and inter-day assays and RSD values were less than 3.0%. The recoveries were between 93.9 and 104.6%. This developed method can be used for quality control purposes and allows for the differentiation between Inula britannica L., Inula hupehensis Ling., and Inula japonica Thunb.

A new sesquiterpene lactone, 6 beta O-isobutyrylbritannilactone (5) and five known sesquiterpene lactones; britannilactone (1), 1-O-acetylbritannilactone (2), Hydroxy alantolactone (3), 8-epi-ivangustin (4) and 1,6-O,O-diacetylbritannilactone (6), were isolated from the flowers of Inula britannica. In the known sesquiterpene lactones, (3) and (4) were isolated for the first time from Inula britannica. The structures of these compounds were established unambiguously by UV, MS and a series of 1D and 2D NMR analyses. Compounds 3, 4, 5 and 6 exhibited significant cytotoxicity for tumor cells in vitro in bioassay.

To prepare reference substance for quality control of Inula Britannica L., 1-O-acetylbritannilactone was extracted and separated from chloroform extraction of Inula Britannica L. Chemical structure of the 1-O-acetylbritannilactone product was elucidated by ultraviolet spectrometry (UV), infrared spectroscopy (IR), nuclear magnetic resonance (NMR) and mass spectrometry (MS), and the results were in agreement with the reference. The purity of the 1-O-acetylbritannilactone product was 99.5%, which satisfies the need of reference substances of traditional Chinese medicines. A method of high performance liquid chromatography-evaporative light scattering detection (HPLC-ELSD) is described for the determination of 1-O-acetylbritannilactone in Inula Britannica L. The chromatographic conditions include Hypersil ODS-2 column, a mixture of methanol-water (52: 48, v/v) with the flow rate of 1.0 mL/min used as mobile phase. The temperature of drift tube of the ELSD was 90 degrees C. Flow rate of carrier gas was 2.5 L/min. Linear range of 1-O-acetylbritannilactone was 1.37 - 8.21 microLg (r = 0.999 8). The average recovery of 1-O-acetylbritannilactone was 100.2% with the relative standard deviation (RSD) of 1.3% (n = 6). The method is simple, accurate, time saving and reproducible.