Effect of Dioscorea villosa extract and the phytoestrogen diosgenin on ovariectomized mice with zymosan-induced arthritis

Abstract Humans are exposed to natural compounds such as phytoestrogens primarily through diet and supplements. These compounds promote health by alleviating the symptoms and illnesses associated with menopause and arthritis. Diosgenin (DSG) occurs naturally in plants such as Dioscorea villosa (DV) and binds to estrogen receptors, so it may have similar effects to this hormone, including against arthritis. Thus, we investigated the effect of chronic treatment with dry extract of DV and its phytoestrogen DSG on ovariectomized mice with arthritis. We found that dry extract of Dioscorea villosa (DV) contains the phytoestrogen diosgenin (DSG) in its composition. Furthermore, arthritic mice treated with DV and DSG showed reduced neutrophil accumulation in the articular cartilage. Also, the dry extract of DV administered orally (v.o) did not alter the leukocyte count in the joints or promote changes in the reproductive tract. However, DSG altered these parameters, with possible beneficial effects by reducing symptoms related to reproductive aging. Thus, oral treatment with dry extract of DV and subcutaneous (s.c) treatment with DSG showed promise by acting against inflammation caused by arthritis and reducing symptoms in the reproductive tract due to menopause.

Antinociceptive effect of Nephelium lappaceum L. fruit peel and the participation of nitric oxide, opioid receptors, and ATP-sensitive potassium channels.

Introduction: Nephelium lappaceum L. (Sapindaceae) is a plant known as rambutan. It is used for various purposes in traditional medicine. Objective: We aimed to evaluate the antinociceptive effects of the ethanol extract of the fruit peel of N. lappaceum (EENL), the mechanisms involved in these effects, and the acute toxicity in zebrafish. Methods: We performed chromatography coupled to mass spectrometry, acute toxicity assay in zebrafish, and evaluation in mice submitted to models of nociception and locomotor activity. Results: We identified (epi)-catechin, procyanidin B, and ellagic acid and its derivatives in EENL. We did not find any toxicity in zebrafish embryos incubated with EENL. The locomotor activity of mice submitted to oral pretreatment with EENL was not changed, but it reduced the abdominal constrictions induced by acetic acid, the licking/biting time in both the first and second phase of formalin testing and capsaicin testing, and carrageenan-induced paw mechanical allodynia. Oral pretreatment with EENL increased latency time in the hot plate test. This antinociceptive effect was significantly reversed by naloxone, L-arginine, and glibenclamide respectively showing the participation of opioid receptors, nitric oxide, and KATP channels as mediators of EENL-induced antinociception. Conclusion: EENL causes antinociception with the participation of opioid receptors, nitric oxide, and KATP channels, and is not toxic to zebrafish.

Prevalence of SARS-CoV-2 infection among urban cleaning and solid waste management workers during transmission of the Omicron variant in Brazil.

This study estimated the prevalence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in urban cleaning and solid waste management workers during the transmission of the Omicron variant in one of the poorest regions of Brazil (the state of Sergipe). Nasopharyngeal swabs were collected from 494 workers, and the presence of SARS-CoV-2 RNA was tested by quantitative reverse-transcriptase polymerase chain reaction. Data on socio-demographic characteristics, comorbidities, vaccination status, mask use, and use of public transport to commute to the workplace were collected. The prevalence with a 95% confidence interval (CI) was calculated from the proportion of SARS-CoV-2 positive cases among the total number of individuals tested. The prevalence ratio (PR) with a 95% CI was the measure of association used to evaluate the relationship between SARS-CoV-2 infection and the exposure variables. The prevalence of SARS-CoV-2 infection was 22.5% (95% CI, 19.0 to 26.4). Individuals under the age of 40 had a higher prevalence of infection (PR, 1.53; 95% CI, 1.03 to 2.30) as well as those who did not believe in the protective effect of vaccines (PR, 1.78; 95% CI, 1.05 to 2.89). Our results indicate the need for better guidance on preventive measures against coronavirus disease 2019 among urban cleaning and solid waste management workers.

Dynamics of SARS-CoV-2 seroprevalence during the first year of the COVID-19 pandemic in the Northeast region of Brazil.

In this household-based seroepidemiological survey, we analyzed the dynamics of SARS-CoV-2 seroprevalence during the first year of the COVID-19 pandemic in Sergipe State, Northeast Brazil, the poorest region of the country. A total of 16,547 individuals were tested using a rapid IgM-IgG antibody test and fluorescence immunoassay (FIA). Seroprevalence rates were presented according to age, sex, and geographic region. A comparative analysis was performed between the results obtained in July 2020 (peak of the first wave), August - November 2020 (end of the first wave), and February - March 2021 (beginning of the second wave). Seroprevalence rates in the three phases were estimated at 9.3% (95% CI 8.5-10.1), 12.0% (95% CI 11.2-12.9) and 15.4% (95% CI 14.5-16.4). At the end of the first wave, there was a rise in seroprevalence in the countryside (p < 0.001). At the beginning of the second wave, we found an increase in seroprevalence among women (p < 0.001), adults aged 20 to 59 years (p < 0.001), and the elderly (p < 0.001). In this phase, we found an increase in estimates both in metropolitan areas and in the countryside (p < 0.001). This study showed an increase in SARS-CoV-2 seroprevalence over the first year of the pandemic, with approximately one in six people having anti-SARS-CoV-2 antibodies at the beginning of the second wave of COVID-19. Furthermore, our results suggest a rapid spread of COVID-19 from metropolitan areas to the countryside during the first months of the pandemic.

Biochanin A as a modulator of the inflammatory response: An updated overview and therapeutic potential.

Uncontrolled inflammation and failure to resolve the inflammatory response are crucial factors involved in the progress of inflammatory diseases. Current therapeutic strategies aimed at controlling excessive inflammation are effective in some cases, though they may be accompanied by severe side effects, such as immunosuppression. Phytochemicals as a therapeutic alternative can have a fundamental impact on the different stages of inflammation and its resolution. Biochanin A (BCA) is an isoflavone known for its wide range of pharmacological properties, especially its marked anti-inflammatory effects. Recent studies have provided evidence of BCA's abilities to activate events essential for resolving inflammation. In this review, we summarize the most recent findings from pre-clinical studies of the pharmacological effects of BCA on the complex signaling network associated with the onset and resolution of inflammation and BCA's potential protective functionality in several models of inflammatory diseases, such as arthritis, pulmonary disease, neuroinflammation, and metabolic disease.

Gastroprotective action of the ethanol extract of Leonurus sibiricus L. (Lamiaceae) in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Leonurus sibiricus L. (Lamiaceae) is a medicinal plant known in Brazil as "rubim" or "erva de macaé". It is used for various purposes, including stomach disorders. AIM OF THE STUDY: To evaluate the effect of the ethanol extract of the aerial parts of L. sibiricus (EELs) in models of gastric damage in mice. MATERIAL AND METHODS: The effect of EELs (50, 100 and 300 mg/kg, p.o., 1 h before induction) was tested on acidified ethanol (ACEt)-induced gastric ulcers. Additionally, we tested the effect of EELs (by intraduodenal administration) in the pylorus ligation (PL) model. RESULTS: Pretreatment with EELs, at 300 mg/kg, but not 50 and 100 mg/kg, reduced the relative area of gastric ulcers induced by ACEt (p < 0.01) and lipoperoxidation (p < 0.001), and increased the sulfhydryl content (p < 0.01) in the stomach in comparison with the vehicle group. Pretreatment with N-ethylmaleimide (a blocker of non-protein sulfhydryl groups, 10 mg/kg, i.p.) or glibenclamide (a KATP channel blocker, 10 mg/kg, i.p.) inhibited the gastroprotective response caused by EELs (300 mg/kg; p < 0.001), but there were no alterations due to pretreatments with inhibitors of the synthesis of prostaglandins (indomethacin, 10 mg/kg), nitric oxide (L-NAME, 70 mg/kg) or hydrogen sulfide (DL-propargylglycine, 10 mg/kg). Treatment with EELs (300 mg/kg) reduced mucus production (p < 0.001) and the volume of gastric secretion (p < 0.001) after PL without affecting gastric acidity or pH. CONCLUSIONS: These results provide evidence that EELs exerts gastroprotective action in mice, with the participation of oxidative stress and mediation of NP-SH, KATP channels and mucus production.

Biochanin A Regulates Key Steps of Inflammation Resolution in a Model of Antigen-Induced Arthritis via GPR30/PKA-Dependent Mechanism.

Biochanin A (BCA) is a natural organic compound of the class of phytochemicals known as flavonoids and isoflavone subclass predominantly found in red clover (Trifolium pratense). It has anti-inflammatory activity and some pro-resolving actions, such as neutrophil apoptosis. However, the effect of BCA in the resolution of inflammation is still poorly understood. In this study, we investigated the effects of BCA on the neutrophilic inflammatory response and its resolution in a model of antigen-induced arthritis. Male wild-type BALB/c mice were treated with BCA at the peak of the inflammatory process (12 h). BCA decreased the accumulation of migrated neutrophils, and this effect was associated with reduction of myeloperoxidase activity, IL-1ß and CXCL1 levels, and the histological score in periarticular tissues. Joint dysfunction, as seen by mechanical hypernociception, was improved by treatment with BCA. The resolution interval (Ri) was also quantified, defining profiles of acute inflammatory parameters that include the amplitude and duration of the inflammatory response monitored by the neutrophil infiltration. BCA treatment shortened Ri from ∼23 h observed in vehicle-treated mice to ∼5.5 h, associated with an increase in apoptotic events and efferocytosis, both key steps for the resolution of inflammation. These effects of BCA were prevented by H89, an inhibitor of protein kinase A (PKA) and G15, a selective G protein-coupled receptor 30 (GPR30) antagonist. In line with the in vivo data, BCA also increased the efferocytic ability of murine bone marrow-derived macrophages. Collectively, these data indicate for the first time that BCA resolves neutrophilic inflammation acting in key steps of the resolution of inflammation, requiring activation of GPR30 and via stimulation of cAMP-dependent signaling.

Seroprevalence of SARS-CoV-2 antibodies in the poorest region of Brazil: results from a population-based study.

Population-based seroprevalence studies on coronavirus disease 2019 (COVID-19) in low- and middle-income countries are lacking. We investigated the seroprevalence of severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) antibodies in Sergipe state, Northeast Brazil, using rapid IgM-IgG antibody test and fluorescence immunoassay. The seroprevalence was 9.3% (95% CI 8.5-10.1), 10.2% (95% CI 9.2-11.3) for women and 7.9% (IC 95% 6.8-9.1) for men (P = 0.004). We found a decline in the prevalence of SARS-CoV-2 antibodies according to age, but the differences were not statistically significant: 0-19 years (9.9%; 95% CI 7.8-12.5), 20-59 years (9.3%; 95% CI 8.4-10.3) and ≥60 years (9.0%; 95% CI 7.5-10.8) (P = 0.517). The metropolitan area had a higher seroprevalence (11.7%, 95% CI 10.3-13.2) than outside municipalities (8.0%, 95% CI 7.2-8.9) (P < 0.001). These findings highlight the importance of serosurveillance to estimate the real impact of the COVID-19 outbreak and thereby provide data to better understand the spread of the virus, as well as providing information to guide stay-at-home measures and other policies. In addition, these results may be useful as basic data to follow the progress of COVID-19 outbreak as social restriction initiatives start to be relaxed in Brazil.

Reduced cutaneous inflammation associated with antioxidant action after topical application of the aqueous extract of Annona muricata leaves.

Annona muricata L. is used in folk medicine for treatment of diseases related to inflammatory and oxidative processes. This study investigated the effect of the aqueous extract of A. muricata leaves (AEAM) on TPA-induced ear inflammation and antioxidant capacity, both in vitro and in vivo. The in vitro antioxidant capacity of AEAM was measured by the 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing/antioxidant power (FRAP) and lipoperoxidation assays. Cytotoxicity and reactive oxygen species (ROS) release were evaluated in the L929 fibroblasts. Swiss mice were submitted to TPA application and were topically treated with AEAM (0.3, 1 or 3 mg/ear). After 6 h, inflammatory and oxidative parameters were evaluated. Quercetin 3-glucoside, rutin, chlorogenic acid, catechin and gallic acid were identified in AEAM. It also presented antioxidant activity in all in vitro assays used. Incubation with AEAM did not cause cell cytotoxicity but reduced ROS release from fibroblasts. Compared with the control group, treatment with AEAM significantly reduced ear oedema and mieloperoxidase activity in inflamed ears, as well as histological parameters of inflammation. These results were associated with the reduction of total hydroperoxides and modulation of catalase, but not superoxide dismutase activity. These findings show the anti-inflammatory effect of AEAM is associated with antioxidant capacity.

Biochanin A attenuates zymosan-induced arthritis in mice similarly to 17-ß estradiol: an alternative to hormone replacement therapy?

OBJECTIVE AND DESIGN: Biochanin A (BCA), a phytoestrogen, has various pharmacological properties. This study was conducted to compare BCA's therapeutic property against 17-ß estradiol replacement therapy in zymosan-induced arthritis (ZIA) in mice. Additionally, we further investigated in vitro the anti-inflammatory action on neutrophils. TREATMENT: Ovariectomized (OVX) and non-OVX mice were pretreated with BCA (1, 3 and 9 mg/kg) or estrogen (50 µg/kg) for 14 days prior to ZIA. Neutrophils were pretreated with BCA (1, 10 and 100 µM) for 1 h prior to phorbol 12-myristate 13-acetate. METHODS: Anti-inflammatory effects of BCA were evaluated by cellular infiltrate, paw edema and cytokine measurement. In vitro, apoptosis was assessed by morphology and flow cytometry. Neutrophil extracellular traps (NET) were determined by fluorescent microscopy and DNA release. Statistical differences were determined by one- or two-way ANOVA. RESULTS: BCA inhibited neutrophil accumulation, paw edema and proinflammatory cytokine (TNF-α and IFN-γ) and increased anti-inflammatory cytokines (IL-4 and IL-10) in OVX and non-OVX mice, similar to 17-ß estradiol replacement therapy. In vitro, BCA increased apoptosis and consequently reduced NETs. CONCLUSION: BCA has a notable anti-inflammatory effect, similar to 17-ß estradiol, and is especially effective for treatment of ZIA. These results suggest that BCA may be promising for the treatment of postmenopausal arthritis.

Anti-inflammatory effect of nano-encapsulated nerolidol on zymosan-induced arthritis in mice.

Nerolidol is naturally occurring sesquiterpene has wide range of biological properties including anti-inflammatory activity. However, it has high volatility with low solubility in nature. The present study aimed to develop and characterized nano-encapsulated nerolidol and evaluated its activity on zymosan-induced arthritis model. Nano-capsules were produced by interfacial deposition of preformed polymer method and characterized by particle size, pH, polydispersity index (PDI), zeta potential, drug content and transmission electron microscopy (TEM). In vitro cytotoxicity of formulations was evaluated by alamar blue and MTT assays. In vivo neutrophils migration assay was performed on intra-articular zymosan-induced arthritis model in mice. Nano-encapsulated nerolidol suspensions presented adequate properties: mean diameter of particles 219.5 ±â€¯8.4 nm, pH: 6.84 ±â€¯0.5, PDI≤0.2, the zeta potential was -20.3 ±â€¯3.6 mV and drug content 71,2 ±â€¯1.3%. The formulations did not demonstrated cytotoxicity under the conditions assessed. Nerolidol 300 mg/kg inhibited neutrophils migration into joint cavity by 18.8% remains compared with control group, and nano-encapsulated nerolidol 3 mg/kg inhibited (26.7% remains) similar to free nerolidol 10 mg/kg (27.4% remains). Histological, quantification of pro-inflammatory and anti-inflammatory cytokines proves the same results. In conclusion the data suggests that nanoencapsulation of nerolidol improved its anti-inflammatory effect on arthritis in mice.

The inclusion complex of carvacrol and ß-cyclodextrin reduces acute skeletal muscle inflammation and nociception in rats.

BACKGROUND: Skeletal muscle inflammation is strongly associated with pain and may impair regeneration and functional recovery after injury. Since anti-inflammatory and antinociceptive effects have been described for the inclusion complex of carvacrol and ß-cyclodextrin (ßCD-carvacrol), this study investigated the effects of ßCD-carvacrol in a model of acute skeletal muscle inflammation. METHODS: Muscle injury was induced in male Wistar rats by injection of 3% carrageenan in the gastrocnemius muscle. Rats were orally pretreated with saline (vehicle) or ßCD-carvacrol (20, 40, 80 and 180 mg/kg) one hour before administration of carrageenan. RESULTS: The injection of carrageenan in the gastrocnemius muscle increased tissue myeloperoxidase (MPO) activity (p < 0.001), edema (p < 0.001) and the levels of tumoral necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6, macrophage inflammatory protein (MIP-2), but not IL-10 levels. Also, it increased mechanical hyperalgesia and decreased the grip force of animals. Pretreatment with ßCD-carvacrol (80 or 160 mg/kg) significantly decreased muscle MPO activity and edema 24 h after injury in comparison to vehicle-pretreated group. Animals pretreated with ßCD-carvacrol (160 mg/kg) presented significantly lower levels of IL-1ß, IL-6 and MIP-2 and higher levels of IL-10 six hours after induction and lower levels of TNF-α and MIP-2 after 24 h when compared to the vehicle group. Pretreatment with ßCD-carvacrol also reduced mechanical hyperalgesia and limited the decrease of grip force (80 or 160 mg/kg; p < 0.001) 6 and 24 h after injury. CONCLUSION: These results show that ßCD-carvacrol reduces inflammation and nociception in a model of acute injury to skeletal muscles.

Vaccinium macrocarpon Aiton Extract Ameliorates Inflammation and Hyperalgesia through Oxidative Stress Inhibition in Experimental Acute Pancreatitis.

We evaluated the effect of the hydroethanolic extract of fruits of Vaccinium macrocarpon (HEVm) in a model of acute pancreatitis (AP) in mice. AP was induced by two injections of L-arginine and animals were treated with HEVm (50, 100, and 200 mg/kg, p.o.) or vehicle (saline) every 24 h, starting 1 h after the induction of AP. Phytochemical analysis of the extract and measurement of inflammatory and oxidative stress parameters, as well as abdominal hyperalgesia, were performed. Catechin, epicatechin, rutin, and anthocyanins were identified in HEVm. Treatment with HEVm decreased L-arginine-induced abdominal hyperalgesia (from 48 to 72 h). Also, treatment with HEVm decreased L-arginine-induced pancreatic edema, pancreatic and pulmonary neutrophil infiltration, and levels of tumor necrosis factor-α, interleukin-1ß, and interleukin-6, after 72 h of induction. L-arginine-induced hyperamylasemia and hyperlipasemia were also reduced by the treatment with HEVm in comparison to vehicle-treated group. Moreover, lipoperoxidation, carbonyl radicals, nonprotein sulfhydryl groups, and activity of catalase and superoxide dismutase, but not glutathione peroxidase, were restored by the treatment with HEVm. These results show that treatment with HEVm decreased hyperalgesia and pancreatic/extrapancreatic inflammation and oxidative damage in L-arginine-induced AP, making this extract attractive for future approaches designed to treat this condition.

A single administration of fish oil inhibits the acute inflammatory response in rats.

OBJECTIVE: To investigate the antiinflammatory effects of a single administration of fish oil (FO) on the acute inflammatory response. METHODS: The paw edema and pleurisy models were used to evaluate the effects of FO dissolved in olive oil (FOP) orally administered in a single dose in rats. Nitric oxide (NO) concentrations in the pleural exudate were performed according to the Griess method and the cytokine concentrations were determined by Luminex bead-based multiplex assay. RESULTS: FOP treatment (30 and 300 mg/kg) significantly reduced paw edema. FOP treatment at 18.75, 37.5, 75.0, 150.0, and 300 mg/kg decreased both the volume of pleural exudate and cellular migration into the pleural cavity and each of these doses presented the same effectiveness. Treatment with FOP (300 mg/kg) reduced NO, TNF-α, IL-1ß, and IL-6 concentrations in the pleural exudate. CONCLUSIONS: The present data provide evidence that FO has inhibitory effects on the acute inflammatory response when administered in a single dose in rats. This effect might be attributable to a direct inhibitory effect of FO on the production or release of inflammatory mediators that are involved in the pathological processes evaluated herein.

The aggravation of arthritis by periodontitis is dependent of IL-17 receptor A activation.

AIM: To evaluate whether Porphyromonas gingivalis-induced periodontitis aggravates the antigen-induced arthritis (AIA) model, and whether this effect is dependent on the Th17/IL-17 signalling pathway. MATERIALS AND METHODS: Antigen-induced arthritis was triggered by local injection of methylated bovine serum albumin into the knee joint of previously immunized C57BL/6 wild-type (WT) and IL-17 receptor A (IL-17RA)-knockout mice. Periodontal disease in naïve or arthritic mice was induced by oral infection with P. gingivalis. Animals were sacrificed 7, 15 and 30 days after infection. Alveolar bone loss, joint histopathology, articular hyperalgesia and joint cytokine production were assessed, in addition to the proportion of Th17 and Treg cells isolated from the inguinal lymph nodes. RESULTS: No influence of experimentally-induced arthritis was found on the alveolar bone resorption induced by P. gingivalis. However, mice with experimentally-induced arthritis that were exposed to P. gingivalis presented higher joint damage and Th17 frequencies when compared to non-infected mice. The aggravation of arthritis by periodontitis was accompanied by increased TNF and IL-17 production and articular neutrophil infiltration, whereas arthritis aggravation and changes in neutrophil infiltration were absent in IL-17RA-deficient mice. CONCLUSION: The effects of P. gingivalis-induced periodontitis on arthritis are dependent on Th17 expansion and IL-17RA signalling, which lead to increased neutrophil infiltration into the joints.

Therapeutic Treatment of Arthritic Mice with 15-Deoxy Δ12,14-Prostaglandin J2 (15d-PGJ2) Ameliorates Disease through the Suppression of Th17 Cells and the Induction of CD4+CD25-FOXP3+ Cells.

The prostaglandin, 15-deoxy Δ12,14-prostaglandin J2 (15d-PGJ2), is a lipid mediator that plays an important role in the control of chronic inflammatory disease. However, the role of prostanoid in rheumatoid arthritis (RA) is not well determined. We demonstrated the therapeutic effect of 15d-PGJ2 in an experimental model of arthritis. Daily administration of 15d-PGJ2 attenuated the severity of CIA, reducing the clinical score, pain, and edema. 15d-PGJ2 treatment was associated with a marked reduction in joint levels of proinflammatory cytokines. Although the mRNA expression of ROR-γt was profoundly reduced, FOXP3 was enhanced in draining lymph node cells from 15d-PGJ2-treated arthritic mice. The specific and polyclonal CD4+ Th17 cell responses were limited during the addition of prostaglandin to cell culture. Moreover, in vitro 15d-PGJ2 increased the expression of FOXP3, GITR, and CTLA-4 in the CD4+CD25- population, suggesting the induction of Tregs on conventional T cells. Prostanoid addition to CD4+CD25- cells selectively suppressed Th17 differentiation and promoted the enhancement of FOXP3 under polarization conditions. Thus, 15d-PGJ2 ameliorated symptoms of collagen-induced arthritis by regulating Th17 differentiation, concomitant with the induction of Tregs, and, consequently, protected mice from diseases aggravation. Altogether, these results indicate that 15d-PGJ2 may represent a potential therapeutic strategy in RA.

Hepatoprotective Effect of Silymarin (Silybum marianum) on Hepatotoxicity Induced by Acetaminophen in Spontaneously Hypertensive Rats.

This study was aimed to investigate the effect of Silymarin (SLM) on the hypertension state and the liver function changes induced by acetaminophen (APAP) in spontaneously hypertensive rat (SHR). Animals normotensive (N) or hypertensive (SHR) were treated or not with APAP (3 g/kg, oral) or previously treated with SLM. Twelve hours after APAP administration, plasmatic levels of liver function markers: alanine aminotransferase (ALT), aspartate aminotransferase (AST), glucose (GLU), gamma glutamyl transferase (γ-GT), and alkaline phosphatase (ALP) of all groups, were determined. Liver injury was assessed using histological studies. Samples of their livers were then used to determine the myeloperoxidase (MPO) activity and nitric oxide (NO) production and were also sectioned for histological analysis. No differences were observed for ALT, γ-GT, and GLU levels between SHR and normotensive rats groups. However, AST and ALP levels were increased in hypertensive animals. APAP treatment promoted an increase in ALT and AST in both SHR and N. However, only for SHR, γ-GT levels were increased. The inflammatory response evaluated by MPO activity and NO production showed that SHR was more susceptible to APAP effect, by increasing leucocyte infiltration. Silymarin treatment (Legalon) restored the hepatocyte functional and histopathological alterations induced by APAP in normotensive and hypertensive animals.

Antifungal activity and inhibition of fumonisin production by Rosmarinus officinalis L. essential oil in Fusarium verticillioides (Sacc.) Nirenberg.

The chemical composition of Rosmarinus officinalis L. essential oil (REO) was analysed by gas chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy. The main compounds of the REO were 1.8 cineole (52.2%), camphor (15.2%) and α-pinene (12.4%). The mycelial growth of Fusarium verticillioides (Sacc.) Nirenberg was reduced significantly by 150 µg/mL of REO. Significant microscopic morphological changes were visualised, such as the rupture of the cell wall and the leakage of cytoplasm at 300 µg/mL of REO. At lower concentrations of REO, the effects on the production of ergosterol and the biomass of mycelium varied, as did the effects on the production of fumonisins, but at ≥300 µg/mL of REO, these processes were significantly inhibited, showing the effectiveness of the REO as an antifungal agent. The results suggested that the REO acts against F. verticillioides by disrupting the cell wall and causing the loss of cellular components, subsequently inhibiting the production of fumonisins and ergosterol.

Antifungal properties and inhibitory effects upon aflatoxin production of Thymus vulgaris L. by Aspergillus flavus Link.

The antifungal and antiaflatoxigenic properties of Thymus vulgaris essential oil (TEO) were evaluated upon Aspergillus flavus "in vitro". Suspension containing 10(6) of A. flavus were cultivated with TEO in concentrations ranging from 50 to 500 µg/mL. TEO reached minimum inhibitory concentration (MIC) and minimum fungicidal concentration (MFC) at 250 µg/mL. Inhibition of ergosterol biosynthesis was detected at a concentration of 100 µg/mL of TEO. Morphological evaluation performed by both light microscopy and scanning electron microscopy showed that antifungal activity of TEO could be detected starting at a concentration of 50 µg/mL and the fungicide effect at a concentration of 250 µg/mL. TEO completely inhibited production of both B1 and B2 aflatoxins (AFB1 and AFB2) at a concentration of 150 µg/mL. This way, fungal biomass development and aflatoxin production were dependent on TEO concentration. Therefore, TEO was capable of controlling the growth of A. flavus and its production of aflatoxins.