Effect of Blueberry Supplementation on a Diet-Induced Rat Model of Prediabetes-Focus on Hepatic Lipid Deposition, Endoplasmic Stress Response and Autophagy.

Blueberries, red fruits enriched in polyphenols and fibers, are envisaged as a promising nutraceutical intervention in a plethora of metabolic diseases. Prediabetes, an intermediate state between normal glucose tolerance and type 2 diabetes, fuels the development of complications, including hepatic steatosis. In previous work, we have demonstrated that blueberry juice (BJ) supplementation benefits glycemic control and lipid profile, which was accompanied by an amelioration of hepatic mitochondrial bioenergetics. The purpose of this study is to clarify the impact of long-term BJ nutraceutical intervention on cellular mechanisms that govern hepatic lipid homeostasis, namely autophagy and endoplasmic reticulum (ER) stress, in a rat model of prediabetes. Two groups of male Wistar rats, 8-weeks old, were fed a prediabetes-inducing high-fat diet (HFD) and one group was fed a control diet (CD). From the timepoint where the prediabetic phenotype was achieved (week 16) until the end of the study (week 24), one of the HFD-fed groups was daily orally supplemented with 25 g/kg body weight (BW) of BJ (HFD + BJ). BW, caloric intake, glucose tolerance and insulin sensitivity were monitored throughout the study. The serum and hepatic lipid contents were quantified. Liver and interscapular brown and epidydimal white adipose tissue depots (iBAT and eWAT) were collected for histological analysis and to assess thermogenesis, ER stress and autophagy markers. The gut microbiota composition and the short-chain fatty acids (SCFAs) content were determined in colon fecal samples. BJ supplementation positively impacted glycemic control but was unable to prevent obesity and adiposity. BJ-treated animals presented a reduction in fecal SCFAs, increased markers of arrested iBAT thermogenesis and energy expenditure, together with an aggravation of HFD-induced lipotoxicity and hepatic steatosis, which were accompanied by the inhibition of autophagy and ER stress responses in the liver. In conclusion, despite the improvement of glucose tolerance, BJ supplementation promoted a major impact on lipid management mechanisms at liver and AT levels in prediabetic animals, which might affect disease course.

The Chemical Profile, and Antidermatophytic, Anti-Inflammatory, Antioxidant and Antitumor Activities of Withania chevalieri A.E. Gonç. Ethanolic Extract.

Withania chevalieri, endogenous from Cape Verde, is a medicinal plant used in ethnomedicine with a large spectrum of applications, such as treating skin fungal infections caused by dermatophytes. The aim of this work was to chemically characterize the W. chevalieri crude ethanolic extract (WcCEE), and evaluate its bioactivities as antidermatophytic, antioxidant, anti-inflammatory and anticancer, as well as its cytotoxicity. WcCEE was chemically characterized via HPLC-MS. The minimal inhibitory concentration, minimal fungicidal concentration, time-kill and checkerboard assays were used to study the antidermatophytic activity of WcCEE. As an approach to the mechanism of action, the cell wall components, ß-1,3-glucan and chitin, and cell membrane ergosterol were quantified. Transmission electron microscopy (TEM) allowed for the study of the fungal ultrastructure. WcCEE contained phenolic acids, flavonoids and terpenes. It had a concentration-dependent fungicidal activity, not inducing relevant resistance, and was endowed with synergistic effects, especially terbinafine. TEM showed severely damaged fungi; the cell membrane and cell wall components levels had slight modifications. The extract had antioxidant, anti-inflammatory and anti-cancer activities, with low toxicity to non-tumoral cell lines. The results demonstrated the potential of WcCEE as an antidermatophytic agent, with antioxidant, anti-inflammatory and anticancer activity, to be safely used in pharmaceutical and dermocosmetic applications.

Effect of Thymbra capitata (L.) Cav. on Inflammation, Senescence and Cell Migration.

Aromatic plants are reported to display pharmacological properties, including anti-aging. This work aims to disclose the anti-aging effect of the essential oil (EO) of Thymbra capitata (L.) Cav., an aromatic and medicinal plant widely used as a spice, as well as of the hydrodistillation residual water (HRW), a discarded by-product of EO hydrodistillation. The phytochemical characterization of EO and HRW was assessed by GC-MS and HPLC-PDA-ESI-MSn, respectively. The DPPH, ABTS, and FRAP assays were used to disclose the antioxidant properties. The anti-inflammatory potential was evaluated using lipopolysaccharide-stimulated macrophages by assessing NO production, iNOS, and pro-IL-1ß protein levels. Cell migration was evaluated using the scratch wound assay, and the etoposide-induced senescence was used to assess the modulation of senescence. The EO is mainly characterized by carvacrol, while the HRW is predominantly characterized by rosmarinic acid. The HRW exerts a stronger antioxidant effect in the DPPH and FRAP assays, whereas the EO was the most active sample in the ABTS assay. Both extracts reduce NO, iNOS, and pro-IL-1ß. The EO has no effect on cell migration and presents anti-senescence effects. In opposition, HRW reduces cell migration and induces cellular senescence. Overall, our study highlights interesting pharmacological properties for both extracts, EO being of interest as an anti-aging ingredient and HRW relevant in cancer therapy.

Exploiting the Crithmum maritimum L. Aqueous Extracts and Essential Oil as Potential Preservatives in Food, Feed, Pharmaceutical and Cosmetic Industries.

Chritmum maritimum, sea fennel, is a facultative halophyte used in salads, soups, and sauces, as well as used to prepare medicinal juices and aqueous extracts (AE) to treat several ailments. Its essential oil (EO) is used as a spice and aromatizing. In this work, the nutritional (crude protein, fiber, lipids, and ashes content) and HPLC-PDA phenolic profiles were determined. Furthermore, the antioxidant potential of the infusion and of the decoction, as well as the antibacterial activity of both, the AE and EO, were assessed against food-contaminating bacteria. The composition of the EO was also established. Sea fennel exhibited considerable fiber (34.3 ± 1.92%) and mineral content (23.6 ± 4.8%). AE contains chlorogenic acid as the major phenolic compound, 49.7 ± 0.8 mg/g in the infusion dry extract and (26.8 ± 0.9 mg/g in the decoction dry extract). EO contains high amounts of monoterpene hydrocarbons, namely γ-terpinene and sabinene. In regards to the antioxidant activity, IC50 values for the infusion and decoction were, respectively: 36.5 ± 1.4 µg/mL and 44.7 ± 4.4 µg/mL in the DPPH assay; 37.3 ± 2.6 µg/mL and 38.4 ± 1.8 µg/mL, in the ABTS assay. EO is particularly active against Bacillus cereus and Lactobacillus plantarum. The results support the use of sea fennel AE and EO as a potential alternative preservative ingredient for feeds, foods, pharmaceutical, and cosmetic industries, due to the antioxidant activity of infusion and decoction, and antibacterial properties of essential oil.

Acanthus mollis Formulations for Transdermal Delivery: From Hydrogels to Emulsions.

Topical formulations of Acanthus mollis L. leaf and the optimization of the release of their active compounds and their topical bioavailability were investigated for the first time. In vitro, the release of active compounds from three formulations-an oil-in-water cream and two hydrogels (Carbopol 940 and Pluronic F-127)-was determined using Franz diffusion cells. Detection and quantification of the compounds was performed via high-performance liquid chromatography with a photodiode array (HPLC-PDA). DIBOA, a bioactive compound of this medicinal plant, exhibited release kinetics of the Weibull model for the Carbopol and Pluronic F-127 formulation, identifying it as a potential active agent to optimize the topical distribution of the formulations. The implications extend to applications in inflammation treatment and tyrosinase inhibition, suggesting that it can make a significant contribution to addressing skin conditions, including melanoma and various inflammatory diseases.

Effect of Phenolic Compounds from Cymbopogon citratus (DC) Stapf. Leaves on Micellar Solubility of Cholesterol.

Dyslipidemias are one of the risk factors for cardiovascular diseases, the leading cause of death and hospitalization worldwide. One way to control cholesterol levels is to control the exogenous cholesterol intake in the body. Natural polyphenolic compounds, namely theaflavins from plant extracts such as black tea, showed the ability to inhibit the formation of the micellar structure, essential for the absorption of cholesterol in the intestine. There are several methodologies to determine this effect, many of which are expensive and time-consuming. Due to these facts, the main purposes of this work were to optimize an inexpensive colorimetric method to study, in vitro, the micellar solubility of cholesterol and applied it to plant extracts. In this work, Cymbopogon citratus leaf extracts, its phenolic fractions, and flavonoids were evaluated. The non-delipidified infusion (CcI) obtained a maximum percentage of micelle destruction of 59.22% for a concentration of 50 µg/mL and the delipidified infusion (CcdI) obtained a maximum percentage of micelle destruction of 58.01% for a concentration of 200 µg/mL. In the case of the fraction of phenolic acids (CcPAs), 23.85% of maximum micellar destruction was recorded for the concentration of 100 µg/mL, while for the fraction of flavonoids (CcF), the micellar destruction was 92.74% at 1 µg/mL, and for the tannin fraction (CcT) of 99.45% at 25 µg/mL. Luteolin presented a percentage of micelle destruction of 94.83% in the concentration of 1 ng/mL, followed by luteolin-7-O-glucoside with 93.71% and luteo-lin-6-C-glucoside with 91.26% at the concentrations of 25 ng/mL and 50 ng/mL, respectively. These results suggest the capability of polyphenols from Cymbopogon citratus to prevent the cholesterol absorption in the gut by micellar destruction, and its contribution for cholesterol-lowering activity.

Forest Biomass as a Promising Source of Bioactive Essential Oil and Phenolic Compounds for Alzheimer's Disease Therapy.

Alzheimer's disease (AD) is the most common neurodegenerative disorder affecting elderly people worldwide. Currently, there are no effective treatments for AD able to prevent disease progression, highlighting the urgency of finding new therapeutic strategies to stop or delay this pathology. Several plants exhibit potential as source of safe and multi-target new therapeutic molecules for AD treatment. Meanwhile, Eucalyptus globulus extracts revealed important pharmacological activities, namely antioxidant and anti-inflammatory properties, which can contribute to the reported neuroprotective effects. This review summarizes the chemical composition of essential oil (EO) and phenolic extracts obtained from Eucalyptus globulus leaves, disclosing major compounds and their effects on AD-relevant pathological features, including deposition of amyloid-ß (Aß) in senile plaques and hyperphosphorylated tau in neurofibrillary tangles (NFTs), abnormalities in GABAergic, cholinergic and glutamatergic neurotransmission, inflammation, and oxidative stress. In general, 1,8-cineole is the major compound identified in EO, and ellagic acid, quercetin, and rutin were described as main compounds in phenolic extracts from Eucalyptus globulus leaves. EO and phenolic extracts, and especially their major compounds, were found to prevent several pathological cellular processes and to improve cognitive function in AD animal models. Therefore, Eucalyptus globulus leaves are a relevant source of biological active and safe molecules that could be used as raw material for nutraceuticals and plant-based medicinal products useful for AD prevention and treatment.

Vascular Effects of Polyphenols from Agrimonia eupatoria L. and Role of Isoquercitrin in Its Vasorelaxant Potential in Human Arteries.

Agrimonia eupatoria L. has been traditionally used for the treatment of inflammatory diseases but also as a hypotensive. To our knowledge, only one study has previously suggested an improvement in vascular endothelial function in diabetic conditions, as the underlying mechanisms and responsible compounds are unknown. In this study, we aimed to assess the direct vascular effects of Agrimonia eupatoria L. in human arteries. The infusion elicited a mild increase in basal vascular tone and a significant potentiation of the adrenergic contraction of 49.18% at 0.02 mg/mL, suggesting the presence of compounds with mild vasoconstrictor activity. In contrast, the ethyl acetate fraction inhibited adrenergic contraction by 80.65% at 2 mg/mL and elicited no effect on basal vascular tone. A potent concentration-dependent vasorelaxation was observed for both the infusion and the ethyl acetate fraction (maximal relaxation above 76% and 47%, respectively). Inhibition of nitric oxide synthase and cyclooxygenase elicited significant decreases in the vasorelaxation to the infusion, as, for the ethyl acetate fraction, only the cyclooxygenase pathway appeared to be involved. Isoquercitrin elicited a vasoactivity consistent with the ethyl acetate fraction, suggesting this is a major component responsible for the vasorelaxant properties of A. eupatoria. Further research is warranted to fully evaluate its vasoprotective properties with therapeutic potential in several conditions, e.g., atherosclerosis.

A review of the ethnomedicinal uses, chemistry, and pharmacological properties of the genus Acanthus (Acanthaceae).

ETHNOPHARMACOLOGICAL RELEVANCE: The Acanthus genus belongs to the Acanthaceae family, and its species are distributed in all continents, mainly in tropical and subtropical regions. Several traditional applications are referred to, but few scientific studies validate them. Despite this, studies in animal models corroborate some of its uses in folk medicine, such as anticancer, antidiabetic, anti-inflammatory, and antinociceptive, which encourages the research on plants of this genus. AIM OF THE REVIEW: To our knowledge, this document is the first comprehensive review study that provides information on the geographic distribution, botanical characteristics, ethnomedicinal uses, phytochemicals, and pharmacological activities of some Acanthus species to understand the correlation between traditional uses, phytochemical, and pharmacological activities, providing perspectives for future studies. RESULTS: In traditional medicine, Acanthus species are mainly used for diseases of respiratory, nervous and reproductive system, gastrointestinal and urinary tract, and skin illness. The most used species are A. montanus, A. ilicifolius, and A. ebracteatus. Chemical compounds (125) from different chemical classes were isolated and identified in seven species, mainly from A. ilicifolius, about 80, followed by A. ebracteatus and A. montanus, appearing with a slightly lower number with fewer phytochemical profile studies. Isolated phytoconstituents have been mainly alkaloids, phenylpropanoid glycosides, and phenylethanoids. In addition, aliphatic glycosides, flavonoids, lignan glycosides, megastigmane derivatives, triterpenoids, steroids, fatty acids, alcohols, hydroxybenzoic acids, simple phenols were also cited. Scientific studies from Acanthus species extracts and their phytoconstituents support their ethnomedical uses. Antimicrobial activity that is the most studied, followed by the antioxidant, anti-inflammatory, and anticancer properties, underlie many Acanthus species activities. A. dioscoridis, A. ebracteatus, A. hirsutus, A. ilicifolius, A. mollis, A. montanus, and A. polystachyus have studies on these activities, A. ilicifolius being the one with the most publications. Most studies were essentially performed in vitro. However, the anticancer, antidiabetic, anti-inflammatory and antinociceptive properties have been studied in vivo. CONCLUSION: Acanthus species have remarkable phytoconstituents with different biological activities, such as antioxidant, antimicrobial, anti-inflammatory, antinociceptive, hepatoprotective, and leishmanicidal, supporting traditional uses of some species. However, many others remain unexplored. Future studies should focus on these species, especially pharmacological properties, toxicity, and action mechanisms. This review provides a comprehensive report on Acanthus genus plants, evidencing their therapeutic potential and prospects for discovering new safe and effective drugs from Acanthus species.

Formulation Effects in the Antioxidant Activity of Extract from the Leaves of Cymbopogon citratus (DC) Stapf.

Cymbopogon citratus DC (Stapf.) is a perennial grass and it is distributed around the world. It is used as a condiment for food and beverage flavouring in the form of infusions and decoctions of its dried leaves. Our previous studies have shown antioxidant, anti-inflammatory and gastroprotective activities for the infusion and its phenolic fractions. The aim of the present work was to develop oral dosage forms from a Cymbopogon citratus extract to be used as a functional food with antioxidant properties. Initially, an essential oil-free infusion was prepared, lyophilized and characterized by HPLC-PDA. Total phenols were quantified with the Folin-Ciocalteu method and the antioxidant activity was assessed by DPPH assay. Gelatine capsules containing the extract with different excipients, selected after DSC and IR trials, were prepared. A formulation exhibiting better antioxidant behaviour in a gastric environment was attained. These results suggest that the proposed formulation for this extract could be a valuable antioxidant product and, consequently, make an important contribution to "preventing" and minimizing diseases related to oxidative stress conditions.

Chemical characterization and bioactive potential of Artemisia campestris L. subsp. maritima (DC) Arcang. essential oil and hydrodistillation residual water.

ETHNOPHARMACOLOGICAL RELEVANCE: Artemisia campestris L. is widely used in traditional medicine for their anti-inflammatory, antirheumatic, antimicrobial and antioxidant properties. A. campestris subsp. maritima Arcang., a halophyte plant ("madorneira" or "erva-lombrigueira" in Portugal), is traditionally used for gastric disorders, rheumatism and hypertension. AIM OF THE STUDY: The present study aims to characterize the essential oil (EO) and the hydrodistillation residual water (HRW), a by-product of the EO production, of Artemisia campestris subsp. maritima from Portugal and evaluate the antioxidant, antifungal, anti-inflammatory and wound healing activities of both extracts at concentrations without toxicity. MATERIALS AND METHODS: The phenolic profile of HRW was determined by HPLC-PDA-ESI/MSn and the EO was analyzed by gas chromatography (GC-FID and GC-MS). The antioxidant activity of both extracts were determined by several assays (ABTS, NO FRAP, ß-carotene and DPPH). The antifungal activity (MIC and MLC) was evaluated against yeasts, dermatophytes and Aspergillus strains using broth macrodilution methods. The anti-inflammatory potential was determined measuring the nitric oxide (NO) production by stimulated mouse leukemic macrophages (RAW 264.7). Cell viability was evaluated for RAW 264.7 and mouse fibroblasts (NIH/3T3). The wound healing activity was evaluated in mouse fibroblasts (NIH/3T3) by the scratch assay. RESULTS: The HRW is mainly characterized by hydroxycinnamic acids and the essential oil is characterized by high amounts of ß-pinene (54.5%), cadin-4-en-7-ol (9.5%), Z-ß-ocimene (6.0%) and Ƴ-terpinene (4.6%). Both extracts showed high antioxidant potential in different assays. Additionally, only the hydrodistillation residual water showed anti-inflammatory activity (IC50 of 330 µg/mL). On the other hand, only the EO showed antifungal activity, particularly against Epidermophyton floccosum (MIC and MLC values of 0.16 µL/mL), and wound healing activity. Bothe extracts were not cytotoxic to macrophages CONCLUSIONS: The by-product HRW contains safe bioactive compounds with antioxidant and anti-inflammatory effect and the EO shows antioxidant properties, antifungal activity against dermatophytes and wound healing effect in skin cells. Overall, our results support the interest and economic value of two extracts obtained from a Portuguese native species and provide scientific validation to some of its traditional uses.

Seaweed Phenolics: From Extraction to Applications.

Seaweeds have attracted high interest in recent years due to their chemical and bioactive properties to find new molecules with valuable applications for humankind. Phenolic compounds are the group of metabolites with the most structural variation and the highest content in seaweeds. The most researched seaweed polyphenol class is the phlorotannins, which are specifically synthesized by brown seaweeds, but there are other polyphenolic compounds, such as bromophenols, flavonoids, phenolic terpenoids, and mycosporine-like amino acids. The compounds already discovered and characterized demonstrate a full range of bioactivities and potential future applications in various industrial sectors. This review focuses on the extraction, purification, and future applications of seaweed phenolic compounds based on the bioactive properties described in the literature. It also intends to provide a comprehensive insight into the phenolic compounds in seaweed.

Urtica spp.: Phenolic composition, safety, antioxidant and anti-inflammatory activities.

Urtica dioica and other less studied Urtica species (Urticaceae) are often used as a food ingredient. Fifteen hydroxycinnamic acid derivatives and sixteen flavonoids, flavone and flavonol-type glycosides were identified in hydroalcoholic extracts from aerial parts of Urtica dioica L., Urtica urens L. and Urtica membranacea using HPLC-PDA-ESI/MSn. Among them, the 4-caffeoyl-5-p-coumaroylquinic acid and three statin-like 3-hydroxy-3-methylglutaroyl flavone derivatives were identified for the first time in Urtica urens and U. membranacea respectively. Urtica membranacea showed the higher content of flavonoids, mainly luteolin and apigenin C-glycosides, which are almost absent in the other species studied. In vitro, Urtica dioica exhibited greater antioxidant activity but Urtica urens exhibited stronger anti-inflammatory potential. Interestingly, statin-like compounds detected in Urtica membranacea have been associated with hypocholesterolemic activity making this plant interesting for future investigations. None of the extracts were cytotoxic to macrophages and hepatocytes in bioactive concentrations (200 and 350µg/mL), suggesting their safety use in food applications.

The Flavone Luteolin Inhibits Liver X Receptor Activation.

Luteolin is a dietary flavonoid with medicinal properties including antioxidant, antimicrobial, anticancer, antiallergic, and anti-inflammatory. However, the effect of luteolin on liver X receptors (LXRs), oxysterol sensors that regulate cholesterol homeostasis, lipogenesis, and inflammation, has yet to be studied. To unveil the potential of luteolin as an LXRα/ß modulator, we investigated by real-time RT-PCR the expression of LXR-target genes, namely, sterol regulatory element binding protein 1c (SREBP-1c) in hepatocytes and ATP-binding cassette transporter (ABC)A1 in macrophages. The lipid content of hepatocytes was evaluated by Oil Red staining. The results demonstrated, for the first time, that luteolin abrogated the LXRα/ß agonist-induced LXRα/ß transcriptional activity and, consequently, inhibited SREBP-1c expression, lipid accumulation, and ABCA1 expression. Therefore, luteolin could abrogate hypertriglyceridemia associated with LXR activation, thus presenting putative therapeutic effects in diseases associated with deregulated lipid metabolism, such as hepatic steatosis, cardiovascular diseases, and diabetes.

Cymbopogon citratus industrial waste as a potential source of bioactive compounds.

BACKGROUND: Cymbopogon citratus (Cc), commonly known as lemongrass, is a very important crop worldwide, being grown in tropical countries. It is widely used in the food, pharmaceutical, cosmetic and perfumery industries for its essential oil. Cc aqueous extracts are also used in traditional medicine. They contain high levels of polyphenols, which are known for their antioxidant and anti-inflammatory properties. Hydrodistillation of lemongrass essential oil produces an aqueous waste (CcHD) which is discarded. Therefore a comparative study between CcHD and Cc infusion (CcI) was performed to characterize its phytochemical profile and to research its antioxidant and anti-inflammatory potential. RESULTS: HPLC-PDA/ESI-MS(n) analysis showed that CcI and CcHD have similar phenolic profiles, with CcHD presenting a higher amount of polyphenols. Additionally, both CcI and CcHD showed antioxidant activity against DPPH (EC50 of 41.72 ± 0.05 and 42.29 ± 0.05 µg mL(-1) respectively) and strong anti-inflammatory properties, by reducing NO production and iNOS expression in macrophages and through their NO-scavenging activity, in a dose-dependent manner. Furthermore, no cytotoxicity was observed. CONCLUSION: The data of this study encourage considering the aqueous solution from Cc leaf hydrodistillation as a source of bioactive compounds, which may add great industrial value to this crop.

Bioactivity of Fragaria vesca leaves through inflammation, proteasome and autophagy modulation.

ETHNOPHARMACOLOGICAL RELEVANCE: Fragaria vesca leaves have been used in folk medicine for the treatment of several diseases, namely gastrointestinal, cardiovascular and urinary disorders, which could be related with the potential anti-inflammatory properties of the extract. This work aims to disclose the bioactivity and the underlying action mechanism of an extract from Fragaria vesca leaves in order to support its traditional uses. MATERIALS AND METHODS: A hydroalcoholic extract was prepared from Fragaria vesca leaves and its anti-inflammatory potential was evaluated through inhibition of nitric oxide production and expression of several pro-inflammatory proteins in lipopolysaccharide-triggered macrophages. Nitric oxide scavenger activity was also assessed using a standard nitric oxide donor. Since numerous inflammatory proteins are tightly regulated by ubiquitination and proteasomal degradation, the putative effect of the extract on these cellular proteolytic pathways was also disclosed. The phytochemical characterization was performed by HPLC-PDA-ESI/MSn and compared with an infusion prepared according to the traditional method. RESULTS: For non-cytotoxic concentrations (80 and 160µg/mL) the extract inhibited nitrite production, probably due to a direct nitric oxide scavenging. Furthermore, inhibition of proteasome activity was verified, leading to accumulation of ubiquitinated proteins. The extract also increased the conversion of the microtubule-associated protein light chain LC3-I to LC3-II, a marker of autophagy. Polyphenols, namely ellagitannins, proanthocyanidins, and quercetin and kaempferol glucuronide derivatives were identified in Fragaria vesca leaves extract. Most of the identified phenolic compounds matched with those found in traditional preparation, the infusion. CONCLUSIONS: The extract has a direct nitric oxide scavenging activity giving support to the traditional use of this plant for the treatment of inflammatory disorders. Furthermore, the extract affects the proteolytic systems but its role in cancer treatment requires further studies.

Anti-inflammatory activity of Cymbopogon citratus leaves infusion via proteasome and nuclear factor-κB pathway inhibition: contribution of chlorogenic acid.

ETHNOPHARMACOLOGICAL RELEVANCE: Cymbopogon citratus (DC.) Stapf leaves infusion is used in traditional medicine for the treatment of inflammatory conditions, however little is known about their bioactive compounds. AIM OF THE STUDY: Investigate the compounds responsible for anti-inflammatory potential of Cymbopogon citratus (Cy) on cytokines production induced by lipopolysaccharide (LPS) in human and mouse macrophages, and the action mechanisms involved. MATERIALS AND METHODS: An essential oil-free infusion of Cy was prepared and polyphenol-rich fractions (PFs) were obtained from it by column chromatography. Chlorogenic acid (CGA) was identified, by HPLC/PDA/ESI-MS(n). The expression of cytokines, namely TNF-α and CCL5, was analyzed by real-time RT-PCR, on LPS-stimulated human macrophages. Activation of nuclear factor (NF)-κB, a master regulator of inflammation, was investigated by western blot and gene reporter assay. Proteasome activity was assessed using a fluorogenic peptide. RESULTS: Cymbopogon citratus extract and its polyphenols inhibited the cytokine production on human macrophages. This supports the anti-inflammatory activity of Cy polyphenols in physiologically relevant cells. Concerning the effect on the activation of NF-κB pathway, the results pointed to an inhibition of LPS-induced NF-κB activation by Cy and PFs. CGA was identified, by HPLC/PDA/ESI-MS(n), as the main phenolic acid of the Cy infusion, and it demonstrated to be, at least in part, responsible by that effect. Additionally, it was verified for the first time that Cy and PFs inhibited the proteasome activity, a complex that controls NF-κB activation, having CGA a strong contribution. CONCLUSIONS: The results evidenced, for the first time, the anti-inflammatory properties of Cymbopogon citratus through proteasome inhibition and, consequently NF-κB pathway and cytokine expression. Additionally, Cy polyphenols, in particular chlorogenic acid, were highlighted as bioactive compounds.

Cymbopogon citratus as source of new and safe anti-inflammatory drugs: bio-guided assay using lipopolysaccharide-stimulated macrophages.

ETHNOPHARMACOLOGICAL RELEVANCE: Aqueous extracts of Cymbopogon citratus (Cy) leaves are used in traditional medicine for the treatment of inflammatory conditions, however, little is known about their mechanism of action. AIM OF THE STUDY: The aim of this study is to explore the anti-inflammatory properties of Cymbopogon citratus leaves and their polyphenol-rich fractions (PFs), as well its mechanism of action in murine macrophages. MATERIALS AND METHODS: A lipid- and essential oil-free infusion of Cy leaves was prepared (Cy extract) and fractionated by column chromatography. Anti-inflammatory properties of Cy extract (1.115 mg/ml) and its PFs, namely phenolic acids (530 µg/ml), flavonoids (97.5 µg/ml) and tannins (78 µg/ml), were investigated using lipopolysaccharide (LPS)-stimulated Raw 264.7 macrophages as in vitro model. As inflammatory parameters, nitric oxide (NO) production was evaluated by Griess reaction, as well as effects on cyclooxygenase-2 (COX-2), inducible NO synthase (iNOS) expression and on intracellular signaling pathways activation, which were analyzed by Western blot using specific antibodies. RESULTS: Cy extract inhibited iNOS expression, NO production and various LPS-induced pathways like p38 mitogen-activated protein kinase (MAPK), c-jun NH(2)-terminal kinase (JNK) 1/2 and the transcription nuclear factor (NF)-κB. The extracellular signal-regulated kinase (ERK) 1/2 and the phosphatidylinositol-3-kinase (PI3K)/Akt activation were not affected by Cy extract. Both phenolic acid- and tannin-rich fractions significantly inhibited NF-κB activation, iNOS expression and NO production but none of the PFs modulated MAPKs or PI3K/Akt activation. Neither Cy extract nor PFs affected LPS-induced COX-2 expression but LPS-induced PGE(2) production is inhibited by Cy extract and by phenolic acid-rich fraction. CONCLUSIONS: Our data provide evidence that support the usage of Cymbopogon citratus leaves extract in traditional medicine, and suggest that Cy, in particular its polyphenolic compounds, could constitute a natural source of a new and safe anti-inflammatory drug.

Anti-inflammatory activity of Cymbopogon citratus leaf infusion in lipopolysaccharide-stimulated dendritic cells: contribution of the polyphenols.

Cymbopogon citratus, an herb known worldwide as lemongrass, is widely consumed as an aromatic drink, and its fresh and dried leaves are currently used in traditional cuisine. However, little is known about the mechanism of action of C. citratus, namely, the anti-inflammatory effects of its dietary components. Because nitric oxide (NO), produced in large quantities by activated inflammatory cells, has been demonstrated to be involved in the pathogenesis of acute and chronic inflammation, we evaluated the effects of the infusion of dried leaves from C. citratus, as well as its polyphenolic fractions--flavonoid-, tannin-, and phenolic acid-rich fractions (FF, TF, and PAF, respectively)--on the NO production induced by lipopolysaccharide (LPS) in a skin-derived dendritic cell line (FSDC). C. citratus infusion significantly inhibited the LPS-induced NO production and inducible NO synthase (iNOS) protein expression. All the polyphenolic fractions tested also reduced the iNOS protein levels and NO production stimulated by LPS in FSDC cells, without affecting cell viability, with the strongest effects being observed for the fractions with mono- and polymeric flavonoids (FF and TF, respectively). Our results also indicated that the anti-inflammatory properties of FF are mainly due to luteolin glycosides. In conclusion, C. citratus has NO scavenging activity and inhibits iNOS expression and should be explored for the treatment of inflammatory diseases, in particular of the gastrointestinal tract.