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Chitooligosaccharides (COS) have a great potential to be used by pharmaceutical industry due to their many biological activities. The use of enzymes to produce them is very advantageous, however it still faces many challenges, such as discovering new strains capable to produce enzymes that are able to generate bioactive oligosaccharides. In the present study a purification protein protocol was performed to purify chitosanases produced by Bacillus toyonensis CCT 7899 for further chitosan hydrolysis. The produced chitooligosaccharides were characterized by mass spectroscopy (MS) and their antiedematogenic effect was investigated through carrageenan-induced paw edema model. The animals were treated previously to inflammation by intragastric route with COS at 30, 300 and 600 mg/kg. The purification protocol showed a good performance for the chitosanases purification using 0.20 M NaCl solution to elute it, with a 9.54-fold purification factor. The treatment with COS promoted a decrease of paw edema at all evaluated times and the AUC0–4h, proving that COS produced showed activity in acute inflammation like commercial anti-inflammatory Dexamethasone (corticosteroid). Therefore, the strategy used to purification was successfully applied and it was possible to generate bioactive oligosaccharides with potential pharmacological use. [ABSTRACT FROM AUTHOR]

The search for promising biomolecules such as chitooligosaccharides (COS) has increased due to the need for healing products that act efficiently, avoiding complications resulting from exacerbated inflammation. Therefore, this study aimed to produce COS in two stages of hydrolysis using chitosanases derived from Bacillus toyonensis. Additionally, this study aimed to structurally characterize the COS via mass spectrometry, to analyze their biocompatibility in acute toxicity models in vivo, to evaluate their healing action in a cell migration model in vitro, to analyze the anti-inflammatory activity in in vivo models of xylol-induced ear edema and zymosan-induced air pouch, and to assess the wound repair action in vivo. The structural characterization process pointed out the presence of hexamers. The in vitro and in vivo biocompatibility of COS was reaffirmed. The COS stimulated the fibroblast migration. In the in vivo inflammatory assays, COS showed an antiedematogenic response and significant reductions in leukocyte migration, cytokine release, and protein exudate. The COS healing effect in vivo was confirmed by the significant wound reduction after seven days of the experiment. These results indicated that the presence of hexamers influences the COS biological properties, which have potential uses in the pharmaceutical field due to their healing and anti-inflammatory action. [ABSTRACT FROM AUTHOR]

Commiphora leptophloeos (Mart.) J.B. Gillett (Burseraceae) is a medicinal plant native from the brazilian northeast caatinga biome, known popularly as "imburana" or "imburana-de-cambão". The leaves of C. leptophloeos are widely used in folk medicine in the treatment of various inflammatory disorders. However, there is no scientific evidence to justify their popular use. This approach aimed to characterize the phytochemical profile of hydroethanolic leaf extract, as well as evaluate the anti-inflammatory and antioxidant potential activity and to investigate the acute toxicity with pre-clinical in vitro and in vivo methodologies. The phytochemical profile was characterized by UPLC-MS and FIA-ESI-IT-MS/MS. The in vitro anti-inflammatory potential the hydroethanolic extract of C. leptophloeos (1, 10, 100 and 200 μg/mL) was investigated by lipopolysaccharide (LPS) induced nitric oxide assay, in order to analyze the potential decrease of nitric oxide (NO) production. For carrageenan-induced paw edema and zymosan-induced air pouch models, the extract (100, 200 and 400 mg/kg) was administrated by intragastric gavage (i.g.) route and used for evaluating the anti-inflammatory effect in vivo. Related to the first animal model, the antiedematogenic activity and myeloperoxidase (MPO) levels could be investigated. In addition, the zymosan-induced air pouch model allowed the analyses of leukocytes migration, total MPO, malondialdehyde (MDA) and cytokines (TNF-α and IL-10) levels. The toxicity in vitro of the extract (1, 10, 100 and 200 μg/mL) was evaluated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and acute toxicity in vivo was tested using the extract at 2000 mg/kg by i. g. route. The phytochemical analyses of C. leptophloeos leaf extract pointed the presence of six glycosylated flavonoids, identified as orientin, isoorientin, vitexin and isovitexin, quercetrin and isoquercitrin. A decrease of NO in vitro was noticed by the use of the extract in the LPS-induced nitric oxide assay and an expressive reduction of the paw-edema followed by a decrease of myeloperoxidase activity at doses of 200 and 400 mg/kg. The zymosan-induced air pouch model indicated that the extract, in all doses, significantly reduced the leukocytes migration, total protein concentration, MPO and MDA levels. The levels of cytokines were verified by the administration of extract in this model, revealing a lower of TNF-α level and an increase of the IL-10 production. In the toxicity study, the MTT assay evidenced no cytotoxicity of the tested concentrations and acute toxicity in vivo test did not result in any sign of toxicity and mortality or significant changes on the biochemical parameters. Based on these results, is possible suggest that the anti-inflammatory activity revealed in this approach can be related to the modulating the level of cytokine, decrease of TNF-α, increase of IL-10 in vivo and also the inhibition of the production of nitric oxide RAW 264.7 activated by LPS. These results demonstrate the potential anti-inflammatory effect C. leptophloeos leaf extrat in inflammatory in vivo models, supporting its use in folk medicine for treatment of inflammatory diseases. Finally, glycosylated flavonoids can be responsible, at least in part, for this effect. Image 1 [ABSTRACT FROM AUTHOR]

Harpalyce brasiliana Benth (Leguminosae) is a shrub endemic to Brazil, popularly known as "snake's root." This species is used in folk medicine for the treatment of inflammation and snakebites. However, up to now there is no scientific research to justify its popular use. The study aimed to characterize the phytochemical profile of the hydroethanol extract from the roots of H. brasiliana (Hb), to evaluate its antioxidant and anti-inflammatory potential, as well as to investigate its cytotoxicity and acute toxicity. The extract was obtained by maceration method using a solution of ethanol:water (70: 30, v/v). The phytochemical profile was obtained by liquid chromatography coupled to mass spectrometry. The cytotoxicity of extract (31–2000 μg/mL) was evaluated in vitro , by the 3-methyl-[4-5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) method using murine macrophage and fibroblast cell lines (RAW 247.6 and 3T3, respectively) and by the hemolytic assay. For the in vivo acute toxicity, the extract (2000 mg/kg) was administered and after 14 days the weight (body and organs) and hematological and biochemical parameters were analyzed. Chemical free radical scavenging effect of the extract (125–2000 μg/mL) was investigated through diphenylpicryl hydrazine reduction, total antioxidant capacity, reducing power, hydroxyl radical scavenging, and iron and copper chelating assays. In vitro anti-inflammatory effect of the extract (125, 500, and 2000 μg/mL) was demonstrated through of nitric oxide (NO) analyzed in lipopolysaccharides stimulated RAW 264.7 cells. In vivo anti-inflammatory activities were evaluated in carrageenan-induced paw edema and zymosan-air-pouch models, with gavage administration (post-treatment) of extract at 100, 200, and 400 mg/kg. For the first animal model, the anti-edematogenic activity and myeloperoxidase (MPO) levels were investigated, while in the zymosan-air-pouch model the leukocyte number, MPO, total protein and pro-inflammatory cytokine (IL-1β, IL-6, and TNF-α) levels were quantified. In addition, the oxidative parameters such as malondialdehyde (MDA) and reduced glutathione (GSH) were determined. The phytochemical profile revealed the presence of 20 compounds, mainly prenylated and geranylated pterocarpans. The extract demonstrated no cytotoxicity in erythrocytes, macrophages and fibroblasts cells at the tested concentrations, as well as no sign of toxicity and mortality or significant alterations on the hematological and biochemical parameters in the acute toxicity model. The extract was also able to neutralize chemical free radicals, with copper and iron chelating effect. For the NO dosage, the extract evidenced the reduction of expression of NO after the administration of the extract (500 and 2000 μg/mL). The edematogenic model revealed a decrease in paw edema and MPO level, while the zymosan-air-pouch model evidenced a reduction of leukocyte number (especially of polymorphornuclears), MPO production, and total protein and cytokine levels, and demonstrated the antioxidant effect through a decrease in MDA and increase in GSH parameters. This approach demonstrates for the first time that Hb is not cytotoxic, has low acute toxicity, and possesses antioxidant and anti-inflammatory properties in preclinical analyses, corroborating its popular use. [Display omitted] • 20 compounds were identified of hydroethanol extract from the roots of Harpalyce brasiliana. • The extract is not cytotoxic and has low acute toxicity. • The extract demonstrated antioxidant and anti-inflammatory potential in vitro/vivo. [ABSTRACT FROM AUTHOR]