Tetramethylpyrazine-loaded electroconductive hydrogels promote tissue repair after spinal cord injury by protecting the blood-spinal cord barrier and neurons.

Spinal cord injury (SCI) usually induces profound microvascular dysfunction. It disrupts the integrity of the blood-spinal cord barrier (BSCB), which could trigger a cascade of secondary pathological events that manifest as neuronal apoptosis and axonal demyelination. These events can further lead to irreversible neurological impairments. Thus, reducing the permeability of the BSCB and maintaining its substructural integrity are essential to promote neuronal survival following SCI. Tetramethylpyrazine (TMP) has emerged as a potential protective agent for treating the BSCB after SCI. However, its therapeutic potential is hindered by challenges in the administration route and suboptimal bioavailability, leading to attenuated clinical outcomes. To address this challenge, traditional Chinese medicine, TMP, was used in this study to construct a drug-loaded electroconductive hydrogel for synergistic treatment of SCI. A conductive hydrogel combined with TMP demonstrates good electrical and mechanical properties as well as superior biocompatibility. Furthermore, it also facilitates sustained local release of TMP at the implantation site. Furthermore, the TMP-loaded electroconductive hydrogel could suppress oxidative stress responses, thereby diminishing endothelial cell apoptosis and the breakdown of tight junction proteins. This concerted action repairs BSCB integrity. Concurrently, myelin-associated axons and neurons are protected against death, which meaningfully restore neurological functions post spinal cord injury. Hence, these findings indicate that combining the electroconductive hydrogel with TMP presents a promising avenue for potentiating drug efficacy and synergistic repair following SCI.

Folium Ginkgo extract and tetramethylpyrazine sodium chloride injection (Xingxiong injection) protects against focal cerebral ischaemia/reperfusion injury via activating the Akt/Nrf2 pathway and inhibiting NLRP3 inflammasome activation.

CONTEXT: Folium Ginkgo extract and tetramethylpyrazine sodium chloride injection (Xingxiong injection) is a compound preparation commonly used for treating cerebral ischaemia/reperfusion injury in ischaemic stroke in China. However, its potential mechanisms on ischaemic stroke remain unknown. OBJECTIVE: This study explores the potential mechanisms of Xingxiong injection in vivo or in vitro. MATERIALS AND METHODS: Sprague-Dawley (SD) rats were randomly assigned to five groups: the sham (normal saline), the model (normal saline) and the Xingxiong injection groups (12.5, 25 or 50 mL/kg). The rats were subjected to 2 h of middle cerebral artery occlusion (MCAO) followed by reperfusion for 14 d. Xingxiong injection was administered via intraperitoneal (i.p.) injection immediately after ischaemia induction for 14 d. Afterwards, rats were sacrificed at 14 d induced by administration of Xingxiong injection. RESULTS: Xingxiong injection significantly reduces infarct volume (23%) and neurological deficit scores (93%) compared with the MCAO/R group. Additionally, Xingxiong injection inhibits the loss in mitochondrial membrane potential (43%) and reduces caspase-3 level (44%), decreases NOX (41%), protein carbonyl (29%), 4-HNE (40%) and 8-OhdG (41%) levels, inhibits the expression of inflammatory factors, such as TNF-α (26%), IL-1ß (34%), IL-6 (39%), MCP-1 (36%), CD11a (41%) and ICAM-1 (43%). Moreover, Xingxiong injection can increase p-Akt/Akt (35%) and Nrf2 (47%) protein expression and inhibit NLRP3 (42%) protein expression. CONCLUSIONS: Xingxiong injection prevents cerebral ischaemia/reperfusion injury via activating the Akt/Nrf2 pathway and inhibiting NLRP3 inflammasome. These findings provide experimental evidence for clinical use of drugs in the treatment of ischaemic stroke.

Single Origin Coffee Aroma: From Optimized Flavor Protocols and Coffee Customization to Instrumental Volatile Characterization and Chemometrics.

In this study, the aroma profile of 10 single origin Arabica coffees originating from eight different growing locations, from Central America to Indonesia, was analyzed using Headspace SPME-GC-MS as the analytical method. Their roasting was performed under temperature-time conditions, customized for each sample to reach specific sensory brew characteristics in an attempt to underline the customization of roast profiles and implementation of separate roastings followed by subsequent blending as a means to tailor cup quality. A total of 138 volatile compounds were identified in all coffee samples, mainly furan (~24-41%) and pyrazine (~25-39%) derivatives, many of which are recognized as coffee key odorants, while the main formation mechanism was the Maillard reaction. Volatile compounds' composition data were also chemometrically processed using the HCA Heatmap, PCA and HCA aiming to explore if they meet the expected aroma quality attributes and if they can be an indicator of coffee origin. The desired brew characteristics of the samples were satisfactorily captured from the volatile compounds formed, contributing to the aroma potential of each sample. Furthermore, the volatile compounds presented a strong variation with the applied roasting conditions, meaning lighter roasted samples were efficiently differentiated from darker roasted samples, while roasting degree exceeded the geographical origin of the coffee. The coffee samples were distinguished into two groups, with the first two PCs accounting for 73.66% of the total variation, attributed mainly to the presence of higher quantities of furans and pyrazines, as well as to other chemical classes (e.g., dihydrofuranone and phenol derivatives), while HCA confirmed the above results rendering roasting conditions as the underlying criterion for differentiation.

Protective Effects of Astragalus Membranaceus and Ligustrazine on Rat Brain Microvascular Endothelial Cell Injury after Oxygen-Glucose Deprivation/ Reoxygenation by Suppressing the PKCδ/MARCKS Pathway.

AIM AND OBJECTIVE: Cell death is a main pathological change in brain ischemia. Astragalus membranaceus (Ast) and ligustrazine (Lig), as traditional Chinese herbs, have a protective effect against ischemia-reperfusion injury. We aim to find whether the underlying protective mechanism of Astragalus membranaceus and ligustrazine against Oxygen-glucose deprivation/reoxygenation (OGD/R) -induced injury in RBMECs is related to PKCδ/MARCKS pathway. MATERIALS AND METHODS: OGD/R preconditioning was instituted in rat brain microvascular endothelial cells (RBMECs). The survival and apoptosis of RBMECs were detected by a Cell Counting Kit-8 and TUNEL staining; PKCδ/MARCKS and MMP9 expression were examined by immunofluorescence, western blot and quantitative real-time PCR. RESULTS: OGD/R stimulation significantly increased RBMEC apoptosis, whereas Ast+Lig, Rottlerin or Ast+Lig+Rottlerin treatment evidently reduced cellular apoptosis and increased cell viability (P <0.05). Furthermore, Ast+Lig, Rottlerin or Ast+Lig+Rottlerin treatment significantly reduced mRNA expression levels of PKCδ/MARCKS and MMP9 (P <0.05), compared to OGD/R control group. Moreover, Ast+Lig, Rottlerin or Ast+Lig+Rottlerin treatment evidently reduced protein expression levels of PKCδ, MMP9, and MARCKS (P <0.05), compared to OGD/R control group, detected by western blotting or immunofluorescence. CONCLUSION: The administration of Astragalus membranaceus and ligustrazine protected RBMECs against OGD/R-induced apoptosis. PKCδ/MARCKS and MMP9 expression were significantly increased after OGD/R stimulation, while Astragalus membranaceus and ligustrazine treatment evidently suppressed. Collectively, Astragalus membranaceus and ligustrazine play protective effects against OGD/R-induced injury in RBMECs through regulating PKCδ/MARCKS pathway to inhibit MMP9 activation.

Design, synthesis, and biological evaluation of ligustrazine/resveratrol hybrids as potential anti-ischemic stroke agents.

To search for potent anti-ischemic stroke agents, a series of tetramethylpyrazine (TMP)/resveratrol (RES) hybrids 6a-t were designed and synthesized. These hybrids inhibited adenosine diphosphate (ADP)- or arachidonic acid (AA)-induced platelet aggregation, among them, 6d, 6g-i, 6o and 6q were more active than TMP. The most active compound 6h exhibited more potent anti-platelet aggregation activity than TMP, RES, as well as positive control ticlopidine (Ticlid) and aspirin (ASP). Furthermore, 6h exerted strong antioxidative activity in a dose-dependent manner in rat pheochromocytoma PC12 cells which were treated with hydrogen peroxide (H2O2) or hydroxyl radical (·OH). Importantly, 6h significantly protected primary neuronal cells suffered from oxygen-glucose deprivation/reoxygenation (OGD/R) injury, comparable to an anti-ischemic drug edaravone (Eda). Together, our findings suggest that 6h may be a promising candidate warranting further investigation for the intervention of ischemic stroke.

Aquatic interaction of uranium with two naturally ubiquitous pyrazine compounds: Speciation studies by experiment and theory.

The present studies interpret the speciation of uranyl (UO22+) with the most ubiquitous class of natural species named pyrazines in terms of stability, speciation and its identification, thermodynamics, spectral properties determined by a range of experimental techniques and further evidenced by theoretical insights. UO22+ forms ML and ML2 kind of species with a qualitative detection of ML3 species, while the ESI-MS identified the formation of all the complexes including ML3. Both the ligands act as bidentate chelators with a difference in ring size and coordinating atoms in the complex formed. The ML3 complexes involve the third ligand participation as monodentate via carboxylate only due to the restricted coordination number and space around the UO22+ ion to accommodate three ligand molecules in its primary coordination sphere. All the complexes are found to be endothermic and purely entropy driven formations. The complex formations showed redshift in the absorption spectra and the shift was further enhanced from ML to ML2 formation. The UO22+ ion redox properties are used to explore the redox potential and heterogeneous electron-transfer kinetic parameters as a function of pH and concentration of UO22+ in presence of pyrazine carboxylates. Interestingly, the cyclic voltammograms identified the ligands also as redox sensitive. The theoretical calculation gave inputs to understand the complex formation at the molecular level with major emphasis on geometry optimization, energetics, bonding parameters, molecular orbital diagrams and bond critical point analyses. The experimental observations in combination with theoretical addendum provided detailed knowledge on the interaction of UO22+ with pyrazine-2-carboxylate and pyrazine-2,3-dicarboxylates.

Discovery of a novel 2,3-dimethylimidazo[1,2-a]pyrazine-6-carboxamide M4 positive allosteric modulator (PAM) chemotype.

This Letter details our efforts to discover structurally unique M4 PAMs containing 5,6-heteroaryl ring systems. In an attempt to improve the DMPK profiles of the 2,3-dimethyl-2H-indazole-5-carboxamide and 1-methyl-1H-benzo[d][1,2,3]triazole-6-carboxamide cores, we investigated a plethora of core replacements. This exercise identified a novel 2,3-dimethylimidazo[1,2-a]pyrazine-6-carboxamide core that provided improved M4 PAM activity and CNS penetration.

Systems pharmacology-based approach for dissecting the mechanisms of pyrazine components in Maotai liquor.

Maotai liquor is a typical representative of sauce aroma-style flavor liquors and has been considered to be a precious cultural heritage of the oriental spirit culture. Aroma components are largely responsible for the characteristic aroma of liquor. Pyrazine compound is one of the most important categories of aroma components that affect the flavor of Maotai liquor. However, limited information is available regarding the systemic analysis of pyrazine compounds, especially the pharmacological effects of bioactive pyrazine components. Therefore, in the current study, a systemic analysis approach was provided by integrating absorption, distribution, metabolism, and excretion (ADME) screening, target identification, pharmacological evaluation and pathway analysis to explore the pharmacological mechanism of pyrazine compounds in Maotai liquor. As a result, 17 pyrazine components with adequate pharmacokinetic properties were filtered out using ADME models. Thirty eight potential targets of these active compounds were identified through target prediction. The pharmacological evaluation was proposed to uncover the pharmacological effect of pyrazine compounds in Maotai liquor from the holistic perspective. Finally, the pharmacological effects of the pathways perturbed by potential targets were interpreted based on the pathway analysis. Our study lays the foundation for formulating a comprehensive understanding of the pyrazine compounds in Maotai liquor, which would contribute to the development of Chinese liquor.

BA-12 Inhibits Angiogenesis via Glutathione Metabolism Activation.

There is a need for an efficient and low-cost leading compound discovery mode. However, drug development remains slow, expensive, and risky. Here, this manuscript proposes a leading compound discovery strategy based on a combination of traditional Chinese medicine (TCM) formulae and pharmacochemistry, using a ligustrazine-betulinic acid derivative (BA-12) in the treatment of angiogenesis as an example. Blocking angiogenesis to inhibit the growth and metastasis of solid tumors is currently one recognized therapy for cancer in the clinic. Firstly, based on a traditional Prunella vulgaris plaster, BA-12 was synthesized according to our previous study, as it exhibited better antitumor activities than other derivatives on human bladder carcinoma cells (T24); it was then uploaded for target prediction. Secondly, the efficacy and biotoxicity of BA-12 on angiogenesis were evaluated using human umbilical vein endothelial cells (HUVECs), a quail chick chorioallantoic membrane, and Caenorhabditis elegans. According to the prediction results, the main mechanisms of BA-12 were metabolic pathways. Thus, multiple metabolomics approaches were applied to reveal the mechanisms of BA-12. Finally, the predictive mechanisms of BA-12 on glutathione metabolism and glycerophospholipid metabolism activation were validated using targeted metabolomics and pharmacological assays. This strategy may provide a reference for highly efficient drug discovery, with the aim of sharing TCM wisdom for unmet clinical needs.

Discovery of the Oral Leukotriene C4 Synthase Inhibitor (1S,2S)-2-({5-[(5-Chloro-2,4-difluorophenyl)(2-fluoro-2-methylpropyl)amino]-3-methoxypyrazin-2-yl}carbonyl)cyclopropanecarboxylic Acid (AZD9898) as a New Treatment for Asthma.

While bronchodilators and inhaled corticosteroids are the mainstay of asthma treatment, up to 50% of asthmatics remain uncontrolled. Many studies show that the cysteinyl leukotriene cascade remains highly activated in some asthmatics, even those on high-dose inhaled or oral corticosteroids. Hence, inhibition of the leukotriene C4 synthase (LTC4S) enzyme could provide a new and differentiated core treatment for patients with a highly activated cysteinyl leukotriene cascade. Starting from a screening hit (3), a program to discover oral inhibitors of LTC4S led to (1S,2S)-2-({5-[(5-chloro-2,4-difluorophenyl)(2-fluoro-2-methylpropyl)amino]-3-methoxypyrazin-2-yl}carbonyl)cyclopropanecarboxylic acid (AZD9898) (36), a picomolar LTC4S inhibitor (IC50 = 0.28 nM) with high lipophilic ligand efficiency (LLE = 8.5), which displays nanomolar potency in cells (peripheral blood mononuclear cell, IC50,free = 6.2 nM) and good in vivo pharmacodynamics in a calcium ionophore-stimulated rat model after oral dosing (in vivo, IC50,free = 34 nM). Compound 36 mitigates the GABA binding, hepatic toxicity signal, and in vivo toxicology findings of an early lead compound 7 with a human dose predicted to be 30 mg once daily.

High-Throughput Screening Using Photoluminescence Probe to Measure Intracellular Calcium Levels.

Aequorin, a 22 kDa protein produced by the jellyfish Aequorea victoria, was the first probe used to measure Ca2+ concentrations ([Ca2+]) of specific intracellular organelles in intact cells. After the binding of Ca2+ to three high-affinity binding sites, an irreversible reaction occurs leading to the emission of photons that is proportional to [Ca2+]. While native aequorin is suitable for measuring cytosolic [Ca2+] after cell stimulation in a range from 0.5 to 10 µM, it cannot be used in organelles where [Ca2+] is much higher, such as in the lumen of endoplasmic/sarcoplasmic reticulum (ER/SR) and mitochondria. However, some modifications made on aequorin itself or on coelenterazine, its lipophilic prosthetic luminophore, and the addition of targeting sequences or the fusion with resident proteins allowed the specific organelle localization and the measurements of intra-organelle Ca2+ levels. In the last years, the development of multiwell plate readers has opened the possibility to perform aequorin-based high-throughput screenings and has overcome some limitation of the standard method. Here we present the procedure for expressing, targeting, and reconstituting aequorin in intact cells and for measuring Ca2+ in the bulk cytosol, mitochondria, and ER by a high-throughput screening system.

Ligustrazine promoted hypoxia-treated cell growth by upregulation of miR-135b in human umbilical vein endothelial cells.

BACKGROUND: Pressure ulcers are a kind of troublesome disease which caused by long-term pressure and subsequently lead to tissue festering necrosis because of sustained ischemia, hypoxia and malnutrition. In our study, we used hypoxia to stimulate human umbilical vein endothelial cells (HUVECs) to mimic pressure ulcers and investigated the effects of Ligustrazine (Lig) with multi-activities on HUVECs. METHODS: HUVECs were treated by hypoxia to induce cell injury. HUVECs were administrated with Lig and/or transfected with miR-135b inhibitor or negative control. Cell viability and cell apoptosis were detected by Cell Counting kit-8 assay and flow cytometry, respectively. The protein expression of Cyclin D1 and p53, the apoptosis-related proteins (Bcl-2, Bax, pro-/Cleaved-Caspas-3), and the JNK/SAPK and PI3K/AKT/mTOR pathways related proteins was examined by western blot. RESULTS: Hypoxia-induced injury presented by decreasing cell viability and increasing cell apoptosis. Then Lig administration enhanced cell viability and inhibited cell apoptosis. Importantly, miR-135b was upregulated by the treatment of Lig. Further studies revealed that transfection with miR-135b inhibitor led to the opposite result with decreasing cell viability and increasing cell apoptosis. In addition, Lig increased the phosphorylation of JNK, SAPK, PI3K, AKT and mTOR. CONCLUSION: Lig promoted hypoxia-treated HUVECs cell growth as evidenced by increasing cell viability and reducing cell apoptosis. This process might be modulated by upregulation of miR-135b and subsequent activation of JNK/SAPK and PI3K/AKT/mTOR pathways.

Ligustrazine inhibits platelet activation via suppression of the Akt pathway.

Aberrant activation of platelets has a critical role in thrombotic vascular events, including atherosclerosis, arterial thrombosis and myocardial infarction. The process of platelet activation is associated with multiple intracellular signaling pathways, including the phosphoinositide 3­kinase/AKT serine/threonine kinase (Akt) pathway. The well­known medicinal herb Rhizoma Ligusticum Wallichii (RLW) has long been used in China to clinically treat various cardiovascular disorders. As the most pharmacologically active component of RLW, ligustrazine has been demonstrated to possess a potent antiplatelet activity. However, the precise mechanisms mediating the bioactivities of ligustrazine have not been thoroughly elucidated. The present study evaluated the effects of ligustrazine hydrochloride (LH; the clinical­grade form of ligustrazine) on platelet activation and investigated the underlying molecular mechanisms. In vitro and ex vivo platelet activation models were used, established by stimulating rat platelet­rich plasma either with the platelet activator adenosine diphosphate (ADP) or with the specific Akt pathway activator insulin­like growth factor­1 (IGF­1). The results demonstrated that treatment with LH significantly and dose­dependently inhibited ADP­induced platelet aggregation, in addition to thromboxane A2 (TXA2) secretion and intracellular Ca2+ mobilization in platelets, in vitro and ex vivo. In addition, LH markedly suppressed ADP­induced Akt phosphorylation in vitro and ex vivo. Furthermore, LH markedly inhibited IGF­1­induced Akt phosphorylation, platelet aggregation, TXA2 formation and Ca2+ mobilization in vitro. Finally, LH was able to reverse adrenaline­induced shortening of bleeding time. Taken together, these results suggested that ligustrazine possesses a broad range of antiplatelet activities without apparent hemorrhagic side-effects, and suppression of Akt signaling may be one of the mechanisms by which ligustrazine exerts its antiplatelet activities.

Chemical and Sensory Evaluation of Magnetic Polymers as a Remedial Treatment for Elevated Concentrations of 3-Isobutyl-2-methoxypyrazine in Cabernet Sauvignon Grape Must and Wine.

3-Isobutyl-2-methoxypyrazine (IBMP) is a potent odorant present in grapes and wines that is reminiscent of green capsicum. Suprathreshold concentrations can lead to obvious vegetative characters and suppress desirable fruity aroma nuances in wines, but options to manage IBMP concentrations are limited. This work investigated pre- and postfermentation addition of a putative imprinted magnetic polymer (PIMP) as a remedial treatment for elevated concentrations of IBMP in Cabernet Sauvignon grape must in comparison to nonimprinted magnetic polymer (NIMP) and to a commercially available polylactic acid (PLA) based film added postfermentation. Chemical and sensory analyses of wines showed that PIMP treatments were more effective than PLA film for decreasing "fresh green" aroma nuances without negatively impacting overall aroma profiles and that postfermentation addition of a magnetic polymer removed up to 74% of the initial IBMP concentration compared to 18% for PLA. Prefermentation addition of magnetic polymers removed 20-30% less IBMP compared to that of postfermentation addition but also had less of an effect on other wine volatiles and color parameters.

Contribution of l-theanine to the formation of 2,5-dimethylpyrazine, a key roasted peanutty flavor in Oolong tea during manufacturing processes.

l-Theanine, the most abundant amino acid in tea, is widely believed to be associated with the tea taste, however, its contribution to the formation of tea aroma is still unknown. Volatiles were determined and nitrogen-containing compounds formed during manufacturing processes were quantified. Lower levels of total sugar and l-theanine were detected in the Oolong tea product undergoing full fire processing (FFOT) suggesting that l-theanine probably involved in the volatile formation during manufacturing processes. Methylpyrazine and 2,5-dimethylpyrazine, two newly formed compounds in FFOT, together with other volatiles were successfully detected in a model thermal reaction of d-glucose and l-theanine (GT-MTR) but not detectable in thermal reactions with single d-glucose (G-MTR) or l-theanine (T-MTR). The concentration of 2,5-dimethylpyrazine increased significantly by adding additional l-theanine to 2nd roasted tea. Our study demonstrated that l-theanine, at least partly, contributed to the formation of 2,5-dimethylpyrazine, a key roasted peanutty flavor in Oolong tea.

Screening and analysis of potentially active components in Shenxiong glucose injection using UHPLC coupled with photodiode array detection and MS/MS.

Shenxiong glucose injection, a pharmaceutical preparation containing a water extract of the roots of Salvia miltiorrhizae and ligustrazine hydrochloride, is widely used in clinical to treat cardiovascular diseases in China. The chemical components of the water extract have been reported and the cardioprotective effects of the injection have been evaluated. However, the chemical constituents of the injection and their correlations with its pharmacological effects have not been established. In this study, 13 chemical constituents of the injection have been identified or characterized by ultra-high performance liquid chromatography with diode array detection and electrospray ionization quadrupole time-of-flight tandem mass spectrometry. Besides, the potentially active compounds of this preparation that directly act on cardiac cells have been screened by cell extraction and ultra high performance liquid chromatography targeted multiple reaction monitoring. As a result, eight potentially active compounds, danshensu (1), ligustrazine hydrochloride (4), salvianolic acid I/H (7), lithospermic acid (8), salvianolic acid D (9), rosmarinic acid (10), salvianolic acid B (12), and salvianolic acid C (13), were obtained and structurally characterized from the 11 target compounds used for screening. The liquid chromatography with quadrupole time-of-flight mass spectrometry and liquid chromatography with multiple reaction monitoring tandem mass spectrometry combination method has demonstrated its potency for the screening, detection, and structural identification of bioactive compounds in a complex matrix.

Recent progress in the structural modification and pharmacological activities of ligustrazine derivatives.

Ligustrazine is a main active fraction of the traditional medicine known as Ligusticum chuanxiong hort, which has been used as clinical medication for cerebral thrombosis, coronary heart disease and stenocardia recently. The rapid metabolism and short half-life of ligustrazine seriously limits its application in clinical practice. Therefore, derivatives of ligustrazine are designed and synthesized in our and other labs, including piperazine, cinnamic acid, styrene, acylguanidine, amides, curcumin and triterpenes derivatives of ligustrazine. Most of these compounds present better pharmacodynamics activities and more favorable pharmacokinetic properties compared to the parent compound. Besides, some new biological activities of these compounds are discovered. Hence, this review continues the previous review of our group as well as aims to highlight recent prominent advances in this field in the past ten years.

Synthesis of New Isoxazole-, Pyridazine-, Pyrimidopyrazines and Their Anti-Inflammatory and Analgesic Activity.

BACKGROUND: Isoxazoles, pyridazines, and pyrimidopyrazines have recently attracted attention due to their potent pharmacological activities. They exhibited anticancer, neuroprotective, analgesic and anti-inflammatory effects. OBJECTIVE: The study aimed to synthesize novel isoxazoles, pyridazines, and pyrimidopyrazines through efficient high yield protocol for evaluating their analgesics and anti-inflammatory activities. METHOD: A series of novel isoxazole-, pyridazine-, pyrimidopyrazine derivatives was prepared from 5,8-alkyl-1,3-dimethyl-5,6-dihydropyrimido[5,6-e]pyrazine-2,4,7-trione (1a,b) as the starting material. RESULTS: The prepared derivatives were synthesized in moderate to good yields (60-75%) in a stepwise efficient protocol under mild condition. These new compounds have been proven by several spectroscopic techniques as IR, 1D and 2D NMR techniques and mass analysis. The in vivo anti-inflammatory was assessed for the synthesized compounds using carrageenan-induced rat hind paw edema model. Also, the in vivo analgesic activity for these products was examined utilizing hot-plate and acetic acid-induced writhing response assays. CONCLUSION: The isoxazole derivatives (3a-f) showed the most forceful anti-inflammatory and analgesic activities. Pyrimidopyrazines (4a-f) demonstrated weaker but comparable antiinflammatory and analgesic activities to the positive controls.

Isolation and identification of new chemical constituents from Chinese chive (Allium tuberosum) and toxicological evaluation of raw and cooked Chinese chive.

Chinese chive (jiu cai) is a popular vegetable in China and has a unique flavour and aroma. The molecular basis of the characteristic fragrance and nutritional properties of Chinese chive has not been previously identified. Sequential extractions in a series of solvents and high-performance liquid chromatography were used to isolate 40 compounds from Chinese chive. The compounds were identified based on high-resolution electrospray ionization mass spectra, 1D and 2D nuclear magnetic resonance techniques, and circular dichroism spectra. Eight novel compounds were identified-four new pyrazines, which have distinctive flavour; one new lignan; and three new flavonoids-together with 32 known compounds. Several of these compounds have potential applications as health-promoting dietary supplements, food additives, or seasonings. Additionally, the volatile organic compounds in fresh and steamed Chinese chive were compared, and the toxicological activity of extracts from fresh and steamed Chinese chive was tested in normal rat liver (IAR20) and kidney (NRK) cells. The results showed that Chinese chive is toxic to liver and kidney cells when fresh, but is safe after heating. This could explain why it is traditional to eat cooked Chinese chive. A possible metabolic rule regarding pyrazines is postulated based on this data, and a human metabolic pathway is suggested for two of the novel compounds which have the highest amount of Chinese chive extracts.

Ureidopyrazine Derivatives: Synthesis and Biological Evaluation as Anti-Infectives and Abiotic Elicitors.

Tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) has become a frequently deadly infection due to increasing antimicrobial resistance. This serious issue has driven efforts worldwide to discover new drugs effective against Mtb. One research area is the synthesis and evaluation of pyrazinamide derivatives as potential anti-TB drugs. In this paper we report the synthesis and biological evaluations of a series of ureidopyrazines. Compounds were synthesized by reacting alkyl/aryl isocyanates with aminopyrazine or with propyl 5-aminopyrazine-2-carboxylate. Reactions were performed in pressurized vials using a CEM Discover microwave reactor with a focused field. Purity and chemical structures of products were assessed, and the final compounds were tested in vitro for their antimycobacterial, antibacterial, and antifungal activities. Propyl 5-(3-phenylureido)pyrazine-2-carboxylate (compound 4, MICMtb = 1.56 µg/mL, 5.19 µM) and propyl 5-(3-(4-methoxyphenyl)ureido)pyrazine-2-carboxylate (compound 6, MICMtb = 6.25 µg/mL, 18.91 µM) had high antimycobacterial activity against Mtb H37Rv with no in vitro cytotoxicity on HepG2 cell line. Therefore 4 and 6 are suitable for further structural modifications that might improve their biological activity and physicochemical properties. Based on the structural similarity to 1-(2-chloropyridin-4-yl)-3-phenylurea, a known plant growth regulator, two selected compounds were evaluated for similar activity as abiotic elicitors.