Potent Anti-inflammatory Compound Research Articles

Experimental dapsone administration induces infertility in male Wistar rats: Mechanisms and clinical implications

Dapsone (4, 4′-diaminodiphenylsulfone, DDS) is a potent anti-inflammatory and antibacterial compound which has been used in the treatment of leprosy, vasculitis and dermatitis herpetiformis, lupus erythematosus profundus and even as an antimalarial in combination with proguanil. This study investigated the effect of the administration of dapsone on the reproductive activities of male rats using in vivo and in vitro techniques. In the in vivo study, dapsone was administered orally to male Wistar rats for 5 days or 6 weeks after which their body weight, relative reproductive organ weights, sperm parameters and reproductive hormones were determined while testicular and epididymal histology were also assessed. Data were compared using analysis of variance and Students-Newman-Keuls multiple comparison test. For the in vitro study, Sertoli cells were cultured and treated with varying doses of dapsone at different durations, thereafter Sertoli cell viability and nuclei integrity were determined. Also, the genetic expressions of Glial cell line-derived neurotrophic factor (GDNF) and transferrin were assessed. The results obtained from the in vivo study showed a duration-dependent significant decrease in body and reproductive organ weights, sperm parameters and serum testosterone concentration. Testicular and epididymal histology also showed duration-dependent degenerative changes. However, all these changes were restored towards control values in the recovery experiment. The viability and deoxyribonucleic acid (DNA) integrity of the treated Sertoli cells showed dose and duration-dependent adverse effects while GDNF and transferrin showed normal genetic expressions. These results suggest that dapsone could induce male reproductive stress by affecting testicular and epididymal structure and function.

Methyl jasmonate abrogates rotenone-induced parkinsonian-like symptoms through inhibition of oxidative stress, release of pro-inflammatory cytokines, and down-regulation of immnopositive cells of NF-κB and α-synuclein expressions in mice

Oxidative stress and neuroinflammation play key roles in the initiation and progression of Parkinson's disease (PD), a neurodegenerative disorder, associated with the loss of nigrostriatal dopaminergic pathway. Thus, compounds that can mitigate oxidative stress and neuroinflammation are being investigated as promising agents for the treatment of PD. This study was designed to evaluate the effects of methyl jasmonate (MJ), a potent antioxidant and anti-inflammatory compound on parkinsonian-like symptoms and the underlying biochemical changes induced by rotenone (Rot) in mice. To this end, the effects of graded doses of MJ (25, 50 and100 mg/kg, i.p.) on motor dysfunctions, cognitive and depressive-like disorders induced by Rot (2.5 mg/kg, i.p.) were evaluated. The specific brain regions (striatum, prefrontal cortex and hippocampus) of the animals were processed for various biochemical studies. Rot-treated mice showed reduced motor activity, postural instability, cognitive and depressive-like disorders. Rot also increased brain levels of malondialdehyde (MDA), nitrite, tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and acetyl-cholinesterase (AChE) activity. Moreover, Rot reduced the concentration of glutathione (GSH) and increased immnopositive cells of NF-κB and α-synuclein expressions in these brain regions. However, pretreatment with MJ, attenuated the parkinsonian-like symptoms and reduced the brain levels of MDA/nitrite, TNF-α and IL-6 induced by Rot. MJ also reduced AChE activity and down-regulate the expressions of NF-κB and α-synuclein in the brain of Rot-treated mice. These findings suggest that MJ has anti-parkinsonian-like activity, which may be related to the inhibition of oxidative stress, release of pro-inflammatory cytokines, and down regulation of NF-κB and α-synuclein expressions.

Ursolic acid exhibits anti-inflammatory effects through blocking TLR4-MyD88 pathway mediated by autophagy

There is an urgent need for effective treatments to reduce the large and growing burden of acute kidney injury (AKI) and its consequences. Inflammation is believed to play a vital role in the pathophysiology of AKI. Macrophage autophagy is considered protective against inflammation. Previous study discovered that ursolic acid (UA), a natural pentacyclic triterpene carboxylic acid found in many plants as apples, bilberries, cranberries and so on, promoted cancer cell autophagy. In the present study, we aimed to explore the effect of UA on ameliorating AKI and the role of macrophage autophagy in the context of inflammation. The data from in vivo experiments showed that pretreatment of mice with UA significantly suppressed xylene-induced ear oedema as well as protected against LPS-induced AKI. Related mechanisms were further studied through in vitro experiment. As expected, UA decreased inflammatory factors TNF-α, IL-6 and IL-1β secretion in macrophages in response to lipopolysaccharide (LPS) stimulation. Furthermore, UA blocked LPS-induced TLR4/MyD88 pathway. More importantly, enhanced autophagy of macrophages by UA through increasing the expression of both LC3B and Beclin-1 led to alter macrophage function. What is more, similar to UA, autophagy inhibitor 3-MA obviously decreased inflammation factors releases hinting the vital role of autophagy in regulating inflammation. In all, above study suggested that UA is a potential anti-inflammatory natural compound for treating AKI by inducing autophagy.

LC-MSn and HR-MS characterization of secondary metabolites from Hypericum japonicum Thunb. ex Murray from Nepalese Himalayan region and assessment of cytotoxic effect and inhibition of NF-κB and AP-1 transcription factors in vitro

Hypericum japonicum Thunb. ex Murray is traditionally used in Nepal to treat several diseases, among whom inflammation and acute pain. Although several secondary metabolites from the same Hypericum species have been already characterized and considered for their pharmacological use, an exhaustive phytochemical characterization of H. japonicum from Nepal is lacking, as well as the assessment of its potential pharmacological properties. Hence, the aims of this study were the characterization of a methanolic extract of H. japonicum (HJME) collected from the Northern region of Nepal by LC–MSn and UPLC-QTOF. The assessment of in vitro inhibition of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and activator protein 1 (AP-1) transcription factors and HJME’s cytotoxic effect on human cell lines was performed to evaluate the potential use of this herb as a source of anti-inflammatory and cytotoxic lead compounds. Fifty-seven phytoconstituents were identified, being mainly flavonoids, phloroglucinols, phenolic acids and xanthones. Although compounds characteristic of H. japonicum were detected (quercetin, quercetin-7-O-α-l-rhamnoside, quercitrin and hyperoside), several others are here reported for the first time in this species. The results from bioassays indicated that HJME could significantly reduce the viability of human THP-1 cells (IC50 = 5.4 ± 1.1 μg mL−1), showing the promising potential of HJME as anti-tumor agent. Furthermore, HJME significantly decreased the activation of both NF-κB and AP-1 at the concentration of 2 μg mL−1. Overall, these data suggest that H. japonicum from Nepal could be used as a source of potential natural anti-inflammatory and anti-tumor lead compounds.

Simultaneous HPLC Analysis of Acemetacin and Indometacin in Plasma and Its Application to Bioavailability Study

Acemetacin is a new potent non-steroidal anti-inflammatory compound which is used for the treatment of arthritis and rheumatic diseases. To establish an assay condition for bioavailability study of acemetacin, high-performance liquid chromatographic (HPLC) method using UV detection for the determination of acemetacin and its metabolite indometacin in blood has been developed. Samples were analysed with Luna C18 column (250×4.6 mm, 5 μm, Phenomenex) which maintained at 40oC. The mobile phase was composed of 0.02M phosphate buffer (pH 4.5):methanol=45:55. The flow rate was 1.4 mL/min. UV absorbance was monitored at 254 nm. Typical retention times were 23, 31 and 34 min for flubiprofen (internal standard), acemetacin and indometacin, respectively. Calibration curves for the determination of acemetacin and indometacin in plasma showed a good linearity at a concentration range from 100 to 3000 ng/mL(r2=0.999). Blood samples of eight human volunteers who had received 3 tablets of 60 mg of acemetacin orally were analysed with HPLC and pharmacokinetic parameters were evaluated. The developed analytical method afforded a simple, rapid and sensitive assay for the measurements of acemetacin and its metabolite, indometacin in plasma and can be employed for bioavailability study for acemetacin formulations.

The parmodulin NRD-21 is an allosteric inhibitor of PAR1 Gq signaling with improved anti-inflammatory activity and stability

Novel analogs of the allosteric, biased PAR1 ligand ML161 (parmodulin 2, PM2) were prepared in order to identify potential anti-thrombotic and anti-inflammatory compounds of the parmodulin class with improved properties. Investigations of structure-activity relationships of the western portion of the 1,3-diaminobenzene scaffold were performed using an intracellular calcium mobilization assay with endothelial cells, and several heterocycles were identified that inhibited PAR1 at sub-micromolar concentrations. The oxazole NRD-21 was profiled in additional detail, and it was confirmed to act as a selective, reversible, negative allosteric modulator of PAR1. In addition to inhibiting human platelet aggregation, it showed superior anti-inflammatory activity to ML161 in a qPCR assay measuring the expression of tissue factor in response to the cytokine TNF-alpha in endothelial cells. Additionally, NRD-21 is much more plasma stable than ML161, and is a promising lead compound for the parmodulin class for anti-thrombotic and anti-inflammatory indications.

Attenuation of Palmitic Acid-Induced Lipotoxicity by Chlorogenic Acid through Activation of SIRT1 in Hepatocytes.

Saturated free fatty acids (FFAs) induce hepatocyte lipotoxicity, wherein oxidative stress-associated mitochondrial dysfunction is mechanistically involved. Chlorogenic acid (CGA), a potent antioxidant and anti-inflammatory compound, protects against high-fat-diet-induced oxidative stress and mitochondrial dysfunction in liver. This study investigates whether CGA protects against FFA-induced hepatocyte lipotoxicity via the regulation of mitochondrial fission/fusion and elucidates its underlying mechanisms. AML12 cell, a non-transformed hepatocyte cell line, is treated with palmitate. Here, it is shown that CGA prevents palmitate-induced lipotoxicity by activation of SIRT1 regulated mitochondrial morphology. CGA treatment mitigates oxidative stress and mitochondrial dysfunction, as evidenced by a decrease in reactive oxygen species (ROS) production, and an increase in mitochondrial mass and mitochondrial membrane potential. CGA also significantly decreases Bax expression and thereby reduces mitochondria-mediated caspase-dependent apoptosis. Mechanistically, CGA attenuates ROS-induced mitochondrial fragmentation by inhibiting dynamin-related protein 1 (Drp1) and enhancing Mfn2 expression. In contrast, the inhibitory effects of CGA on the generation of mitochondrial ROS and Drp1 are blocked by siRNA knockdown of SIRT1. Collectively, these findings show that supplementation with CGA protects hepatocytes from FFA-induced lipotoxicity through activation of SIRT1, which reverses the oxidative stress and dysfunction of mitochondrial biogenesis directly.

Computational Evaluation and Anti-inflammatory and Analgesic Activities of Nebrodenside A Isolated From Dodonaea viscosa

Nebrodenside A was isolated from the plant Dodonaea viscosa and its chemical structure was elucidated by spectroscopic methods. Molecular docking simulation predicted its strong ability to be tested as a potential anti-inflammatory compound. Carrageen-induced paw model and formalin test were utilized for the assessment of anti-inflammatory and analgesic potential. Nebrodenside A was found to be significantly active in reducing inflammation, when compared with diclofenac as a standard drug. Similarly, the compound also possesses substantial analgesic activity in the formalin-induced writhing test. Thus, nebrodenside A revealed a robust potential to be developed as a possible leading medicinal agent for better management of pain and inflammation.

Evaluation of the protective and therapeutic effects of Pistacia atlantica gum aqueous extract on cellular and pathological aspects of experimental asthma in Balb/c mice.

The purpose of this study was to investigate the protective and therapeutic effects of aqueous extract of P. atlantica gum on an experimental asthma in BALB/c mice. Aqueous extract of dried and milled P. atlantica gum was assemble and evaluate by GC-MS. In order to investigate the effect of P. atlantica gum extract on cellular and pathological aspects of asthma, 60 BALB/c mice were divided into six groups as: negative control, asthmatic group, asthmatic group receiving dexamethasone (1mg/kg; intraperitoneal (IP)) and three asthmatic groups receiving different concentrations of the extract (100, 200 and 400 mg/kg, orally) from the beginning of the study and continued for 84 days. The examined parameters included cell population, IgE antibody production, levels of IL-4, IL-5, TGF-β, INF-γ, IL-10, and IL-17 cytokines, and lung tissue damage. Regardless of the dose, aqueous extract of P. atlantica gum, caused significant decrease in the number of BALF eosinophilic cells and levels of anti-ovalbumin IgE, IL-4, IL-5 and IL-17 cytokine levels, as well as pathologic damage of the lung tissue. In addition, the amount of anti-inflammatory IL-10, TGF-β, and INF-γ Th1 cytokines significantly increased in the extract-treated groups compared to the asthmatic and dexamethasone-treated groups. Moreover, IFN-γ/IL-4 ratio significantly increased in a dose-dependent manner compared to the un-treated asthma group. The aqueous extract of P. atlantica gum can be considered as a potent anti-inflammatory and immunomodulatory compound and may be used as a natural compound for treatment of immune system disorders.

Metabolomic Screening of Anti-Inflammatory Compounds from the Leaves of Actinidia arguta (Hardy Kiwi).

The metabolomic screening of potential anti-inflammatory compounds in the leaves of Actinidia arguta was performed by using LC-MS/MS. Ethanol extracts were prepared, and the anti-inflammatory effects were investigated based on nitric oxide (NO) synthesis and inducible nitric oxide synthase expression in lipopolysaccharide-induced RAW 264.7 macrophages. The 75% ethanol extract showed the highest inhibitory effect on nitric oxide (NO) production, and it was further separated by in vitro bioassay-guided fractionation using preparative LC with reversed-phase column separation. Through multiple steps of fractionation, sub-fraction 1-3 was finally purified, and caffeic acid derivatives, such as caffeoylthreonic acid and danshensu (salvianic acid A), were successfully identified as key anti-inflammatory compounds by LC-MS/MS and metabolomics analyses. This is the first study identifying anti-inflammatory compounds in A. arguta (Actinidia arguta) leaves through bioassay-guided fractionation and metabolomics screening. Results of this study would be useful for the application of A. arguta leaves as a nutraceutical.

Identification of anti-inflammatory components in Sinomenii Caulis based on spectrum-effect relationship and chemometric methods

The present study aimed to identify the anti-inflammatory components in Sinomenii Caulis (SC) based on spectrum-effect relationship and chemometric methods. A phytochemical investigation of SC extract was performed firstly and afforded eleven potential bioactive compounds. The HPLC fingerprints of 19 batches of SC samples were evaluated by the chemometric methods such as similarity analysis (SA) and hierarchical clustering analysis (HCA). The anti-inflammatory effects of these samples were determined by inhibition of Nitric Oxide (NO) production. Partial least squares regression (PLSR) and artificial neural network (ANN) were used to explore the spectrum-effect relationship of SC. The results indicated that there was a close correlation between chemical fingerprint and anti-inflammatory activity of SC, and peaks 8, 9, 12, 13, 14, 16, 19 and 22 might be potential anti-inflammatory compounds in SC. The verification experiments by testing individual compounds and a combination of them indicated that sinomenine (P8), magnoflorine (P13), menisperine (P16) and stepharanine (P19) were the major anti-inflammatory compounds in SC. Collectively, the present study established the spectrum-effect relationship mode of SC and discovered the anti-inflammatory compounds in SC, which could be used for exploration of bioactive components and quality control of herbal medicines.

Oleoylethanolamide, Neuroinflammation, and Alcohol Abuse.

Neuroinflammation is a complex process involved in the physiopathology of many central nervous system diseases, including addiction. Alcohol abuse is characterized by induction of peripheral inflammation and neuroinflammation, which hallmark is the activation of innate immunity toll-like receptors 4 (TLR4). In the last years, lipid transmitters have generated attention as modulators of parts of the addictive process. Specifically, the bioactive lipid oleoylethanolamide (OEA), which is an endogenous acylethanolamide, has shown a beneficial profile for alcohol abuse. Preclinical studies have shown that OEA is a potent anti-inflammatory and antioxidant compound that exerts neuroprotective effects in alcohol abuse. Exogenous administration of OEA blocks the alcohol-induced TLR4-mediated pro-inflammatory cascade, reducing the release of proinflammatory cytokines and chemokines, oxidative and nitrosative stress, and ultimately, preventing the neural damage in frontal cortex of rodents. The mechanisms of action of OEA are discussed in this review, including a protective action in the intestinal barrier. Additionally, OEA blocks cue-induced reinstatement of alcohol-seeking behavior and reduces the severity of withdrawal symptoms in animals, together with the modulation of alcohol-induced depression-like behavior and other negative motivational states associated with the abstinence, such as the anhedonia. Finally, exposure to alcohol induces OEA release in blood and brain of rodents. Clinical evidences will be highlighted, including the OEA release and the correlation of plasma OEA levels with TLR4-dependent peripheral inflammatory markers in alcohol abusers. In base of these evidences we hypothesize that the endogenous release of OEA could be a homeostatic signal to counteract the toxic action of alcohol and we propose the exploration of OEA-based pharmacotherapies to treat alcohol-use disorders.

Anti-inflammatory, Anti-thrombotic and Molecular Docking Studies of 1,3,4 Oxadiazole Derivatives in Rats

Background: 1,3,4-Oxadiazole have been reported for the diverse use in medicine because of its versatile pharmacological properties. In continuation of our on-going research the present study is established to investigate the in-vitro and in-vivo anti-inflammatory activity of 5-{[2-(4-chlorophenoxy)propan-2-yl]-1,3,4-oxadiazole-2-thiol (1) and its derivatives N-substituted 5-{[2-(4-chlorophenoxy)propan-2-yl]-1,3,4-oxadiazol-2ylthio} acetamides (3a-3i). Methods: In-vitro and in-vivo anti-inflammatory activity was evaluated by albumin protein denaturation method and carrageenan-induced edema respectively. Ligands 1, standard, and 3a-3h were docked with COX-2 (3LN1) by Patch Dock to find the docking transformations. All the values are expressed in mean ± SEM. Statistical significance was calculated by one way ANOVA with Dennett’s test. Results: Experimental results for in-vitro and in-vivo anti-inflammatory activity suggested that 3a and 3c as potent anti-inflammatory compounds. Moreover, in the in-vivo rat model, the compound 3h and 3a showed a significant role in enhancing clotting time. In silico study of tested compounds showed that all these compounds except 1 and 3e show higher docking score than standard drug diclofenac sodium. In addition, impressive inhibitory potential against COX-2 was observed with higher docking scores (5172-5938) with Atomic Contact Energy (ACE) values (-143.21 to -380.56 kcal/mol) than the standard diclofenac sodium (Docking score 4612; ACE -242.58 kcal/mol). Conclusion: All the results suggest that these compounds may be used in the development of anti-inflammatory pharmaceutical products.

A rare case report on dapsone induced haemolytic anaemia in a hansen’s disease patient with type-1 leprae reactions

Dapsone also known as diaminodiphenyl sulfone is a principal drug in a multidrug regimen recommended by the WHO used for the treatment of leprosy. It is a potent anti-inflammatory and anti-parasitic compound mainly used in the treatment of wide variety of dermatological inflammatory diseases. A 58 years old male patient was admitted in the dermatology ward with chief complaints of body pains, scaly red raised lesions all over the body since 10 days. His past medical history includes h/o BB (Borderline Borderline) leprosy & started MB–MDT (is a combination of Tab. Dapsone, Tab. Rifampicin, Tab. Clofazimine) since 2 months. Now he developed ­ erythema & scaling of lesions in body & body pains, burning sensation of eyes since 10 days. Dapsone is associated with seven fold increased risk of hemolytic anemia so better vigilance is necessary for implementation of safe and effective treatment for each individual.

Curcumin Elevates TFH Cells and Germinal Center B Cell Response for Antibody Production in Mice.

Curcumin is a natural product extracted from Curcuma longa. It has been reported as a potent antioxidant and anti-inflammatory compound. Previous studies have demonstrated that curcumin suppresses pro-inflammatory cytokine production via inhibition of NF-κB in macrophages. However, its role in adaptive immune cells such as T cells, in vivo, has not clearly been elucidated. Here, we examined the effects of curcumin in T follicular helper (TFH) cells and on Ab production during NP-ovalbumin immunization in mice. The results revealed that curcumin administered daily significantly increased CXCR5+B-cell lymphoma 6+ TFH cells and CD95+GL-7+ germinal center (GC) B cells in draining lymph nodes. In addition, curcumin treatment in mice induced total Ab production as well as high affinity IgG1 and IgG2b Ab production. Collectively, these results suggest that curcumin has positive regulatory roles in TFH cell functions and GC responses. Thus, this could be an advantageous supplement to enhance humoral immunity against infectious diseases and cancer.

Old age-associated phenotypic screening for Alzheimer's disease drug candidates identifies sterubin as a potent neuroprotective compound from Yerba santa

Alzheimer's disease (AD) is the most frequent age-associated dementia with no treatments that can prevent or slow its progression. Since age is by far the major risk factor for AD, there is a strong rationale for an alternative approach to drug discovery based upon the biology of aging. Phenotypic screening assays that reflect multiple, age-associated neurotoxicity pathways rather than single molecular targets can identify compounds that have therapeutic efficacy by targeting aspects of aging that contribute to AD pathology. And, while the suitability of any single assay can be questioned, a combination of assays can make reliable predictions about the neuroprotective effects of compounds in vivo. Therefore, our lab has developed a combination of phenotypic screening assays that are ideally suited not only to identify novel neuroprotective compounds for the treatment of AD but also their target pathways, thereby potentially providing new therapeutic targets for disease treatment. Using these assays, we screened a large library of extracts from plants with identified pharmacological uses. Analysis of one of these extracts from the plant Yerba santa (Eriodictyon californicum) identified the flavanone sterubin as the active component and further studies showed it to be a potent neuroprotective and anti-inflammatory compound.

Synthesis and Anti-Inflammatory Activities of Phloroglucinol-Based Derivatives.

The natural product phloroglucinol-based derivatives representing monoacyl-, diacyl-, dimeric acyl-, alkylated monoacyl-, and the nitrogen-containing alkylated monoacylphloro- glucinols were synthesized and evaluated for inhibitory activities against the inflammatory mediators such as inducible nitric oxide synthase (iNOS) and nuclear factor kappaB (NF-κB). The diacylphloroglucinol compound 2 and the alkylated acylphloroglucinol compound 4 inhibited iNOS with IC50 values of 19.0 and 19.5 µM, respectively, and NF-κB with IC50 values of 34.0 and 37.5 µM, respectively. These compounds may serve as leads for the synthesis of more potent anti-inflammatory compounds for future drug discovery.

Restoration of skeletal muscle homeostasis by hydrogen sulfide during hyperhomocysteinemia-mediated oxidative/ER stress condition 1.

Elevated homocysteine (Hcy), i.e., hyperhomocysteinemia (HHcy), causes skeletal muscle myopathy. Among many cellular and metabolic alterations caused by HHcy, oxidative and endoplasmic reticulum (ER) stress are considered the major ones; however, the precise molecular mechanism(s) in this process is unclear. Nevertheless, there is no treatment option available to treat HHcy-mediated muscle injury. Hydrogen sulfide (H2S) is increasingly recognized as a potent anti-oxidant, anti-apoptotic/necrotic/pyroptotic, and anti-inflammatory compound and also has been shown to improve angiogenesis during ischemic injury. Patients with CBS mutation produce less H2S, making them vulnerable to Hcy-mediated cellular damage. Many studies have reported bidirectional regulation of ER stress in apoptosis through JNK activation and concomitant attenuation of cell proliferation and protein synthesis via PI3K/AKT axis. Whether H2S mitigates these detrimental effects of HHcy on muscle remains unexplored. In this review, we discuss molecular mechanisms of HHcy-mediated oxidative/ER stress responses, apoptosis, angiogenesis, and atrophic changes in skeletal muscle and how H2S can restore skeletal muscle homeostasis during HHcy condition. This review also highlights the molecular mechanisms on how H2S could be developed as a clinically relevant therapeutic option for chronic conditions that are aggravated by HHcy.

Anti-inflammatory bibenzyls from the stems of Dendrobium huoshanense via bioassay guided isolation

The stems of Dendrobium huoshanense have long been used to prevent various diseases, including inflammatory diseases. This study was aimed to explain the anti-inflammatory effect of D. huoshanense stems in LPS-induced RAW 264.7 macrophages and to discover potential anti-inflammatory compounds. Results exhibited that D. huoshanense stems ethanol extract could significantly inhibit LPS-induced production of NO, TNF-α and IL-1β. Based on bioassay guided strategy, four bibenzyls (1–4) were isolated from D. huoshanense stems for the first time. Anti-inflammatory assay showed 1–4 could remarkably inhibit the production of NO in LPS-induced macrophages. Moreover, quantitative RT-PCR analysis displayed that the mRNA levels of iNOs, TNF-α and IL-1β could also be significantly reduced by 1–4. These results suggested that D. huoshanense stems ethanol extract and bibenzyls 1–4 might be well developed as therapeutic agent to prevent inflammatory diseases.

Bioassay-guided isolation and UHPLC-DAD-ESI-MS/MS quantification of potential anti-inflammatory phenolic compounds from flowers of Inula montana L

Ethnopharmacological relevanceFlowers of Inula montana L. (Asteraceae), commonly known as “Arnica de Provence”, are used in the traditional medicine of Provence in France with the same indication as Arnica montana, for the relief of bruises, as an anti-inflammatory agent. Aims of the studyThe aim of our study is to evaluate its anti-inflammatory properties and to justify its traditional uses. Its potential valorization is evaluated in order to propose Inula montana as an alternative to Arnica montana. Materials and methodsBio-guided fractionation of ethanolic extract allowed the isolation of compounds responsible of the inhibition of NO production. The fractionation was realized using chromatographic techniques and structure elucidation was conducted by ESI-MS and NMR spectral data. Anti-inflammatory effect of ethanolic extract, different fractions and isolated pure compounds was studied in vitro on immortalized mouse macrophages RAW 264.7. An analytical UHPLC-DAD-ESI-MS/MS method was developed for the identification of these compounds in the herbal drug. This UHPLC-DAD method was validated and was used to compare the phenolic profile and content in plant material from the two collection sites: Bonnieux and Merindol. ResultsEleven compounds were identified by UHPLC-MS. Chlorogenic acid (1), Luteolin (2), Nepetin (3), 3,5-O-Dicaffeoylquinic acid (4), 1,5-O-Dicaffeoylquinic acid (5), Nepitrin (6), Hispiduloside (7) and Jaceosid (8) were isolated and identified by NMR. Compounds 9, 10 and 11 were confirmed to be 6-Hydroxykaempferol 3,7-dimethyl ether, Hispidulin and Chrysosplenol C, respectively by comparing retention times and MS/MS data with those of the authentic substances. Six compounds: 1 and 4–8 are reported for the first time in Inula montana L. Compounds 2–8 showed promising anti-inflammatory activity with the release of NO with IC50 value < 7 µM. The UHPLC-DAD method of quantification of three major bioactive compounds (1, 3 and 5) was validated. ConclusionFlowers extracts and isolated compounds present promising anti-inflammatory activity which provides a scientific basis for the traditional use of Inula montana and may be proposed in the same indications as Arnica montana. The developed and validated simple, accurate and rapid UHPLC method can be used for the quality control of the herbal drug.