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Antioxidants derived from food by-products are known for their bioactive properties and impact on human health. However, the gastrointestinal behavior is often poor due to their degradation during digestion. The development of Ca(II)-alginate beads supplemented with biopolymers and enriched with cowpea ( Vigna unguiculata ) extract could represent a novel environmentally friendly technological solution to produce functional ingredients in the food industry. The present study evaluates the impact of in vitro digestion/fermentation by analyzing global antioxidant response (GAR), production of short-chain fatty acids (SCFAs) as a modulation of gut microbiota, and behavior of proton transverse relaxation times by low-field nuclear magnetic resonance (as an indicator of gelation state and characterization of microstructure). Results revealed that guar gum and cowpea protein preserved a high GAR of total phenolic compounds and antioxidant capacity by ABTS and FRAP methods after digestion/fermentation, promoting an adequate protection of the bioactives for their absorption. Alginate-based beads have great potential as prebiotics, with the guar gum-containing system contributing the most to SCFAs production. Finally, the overall higher mobility of protons observed in the intestinal phase agrees with structural changes that promote the release of phenolic compounds during this stage. Beads are excellent carriers of bioactive compounds (cowpea phenolic compounds and peptides) with potential capacities.

Carvacrol has demonstrated antioxidant activity; however, its high volatility and low water solubility limit its direct application in food matrices. Then, an effective encapsulation system is required to protect it. This study aimed to design and characterize a carvacrol-based additive encapsulated in a spray-dried multilayer emulsion based on chitosan/sodium alginate/maltodextrin. Spray-drying temperature of 120 °C and 3 %(w/w) maltodextrin content maximized both encapsulation efficiency (~97 %) and loading capacity (~53 %). The powder's antioxidant properties were evaluated in two food simulant media: water (SiW) and water-ethanol (SiD). The highest antioxidant activity was observed in SiW for both ABTS •+ (8.2 ± 0.3mg EAG /g) and FRAP (4.1 ± 0.2mg EAG /g) methods because of the reduced release of carvacrol in SiD vs. SiW, as supported by micro- and macrostructural observations by SAXS and microscopy, respectively. An increase from 143 to 157 °C attributable to carvacrol protection and T g  = 44.4 °C (> ambient) were obtained by TGA and DSC, respectively. FT-IR confirmed intermolecular interactions (e.g. -COO - and -NH 3 + ) as well as H-bonding formation. High water solubility (81 ± 3 %), low hygroscopicity (8.8 ± 0.2 %(w/w), poor flowability (CI:45 ± 4), and high cohesiveness (HR:1.8 ± 0.1) between particles were achieved, leading to a powdered antioxidant additive with high potential for applications which required avoiding/reducing oxidation on hydrophilic and hydrophobic food products., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Bioaccessibility analysis and antioxidant activity along in vitro digestion and a consumer-oriented sensory analysis were conducted in three potential functional foods based on Ca(II)-alginate beads containing bioactive compounds extracted from beet stems. Ca(II)-alginate beads per se , and two selected products (cookies and turkish delights supplemented with the beads) were prepared. Regarding the beads, among the attributes rated by consumers, visual appreciation predominates, being color in the just-as-right (JAR) category and in the like preference. Instead, both flavor and sweet taste were attributes highly penalized and should be improved in beads to be accepted as food per se . A higher percentage of customers preferred cookies and turkish delights instead of only beads, considering global satisfaction. Regarding in vitro digestion, there was a significant content of phenolic compounds in the products with beads, showing a bioaccessibility greater than 80% (for cookies) and 26% (for turkish delights). Also, the antioxidant capacity measured by ABTS ranged between 50 and 109% for cookies and turkish delights, being lower when measured by FRAP (between 20 and 30%, respectively). Thus, including the beads with beet stem extract in both products leads to a significant increase in the content of phenolic compounds and in the antioxidant capacity compared to their counterparts, protecting the compound during oral and gastric phases. These results allow the generation of improved Ca(II)-alginate systems with promising functional properties for the development of ingredients and functional foods., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2022 The Authors.)

The aim of the present paper was to unravel the effect of a standardized in vitro European protocol of digestion-fermentation over Ca(II)-alginate beads synthesized with sugars and biopolymers. Special emphasis on the antioxidant capacity using methods that simulate physiological conditions, short-chain fatty acids (SCFAs) production, and a detailed study of the microstructure of the gel network by SAXS at several scales (1-100 nm) were considered. Beads released high antioxidant capacity during digestion assessed by several methods, comparable to some common foods; antioxidant capacity was improved with sucrose and arabic gum inclusion in the formulation. After fermentation by gut microbiota, a ten-fold increase in the antioxidant values and an important SCFAs production were obtained, revealing the enhanced ability to produce these functional biomolecules. The microstructural analysis of Ca(II)-alginate showed an advantageous behavior: they slightly changed in oral and gastric fluids and partially dissolved their structure in intestinal fluid, where absorption occurs., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The aim of the present work was to analyze the effect of in vitro gastrointestinal digestion-fermentation on antioxidant capacity, total phenols and production of short chain fatty acids (SCFAs) from biocompounds derived from beet waste (leaf and stem) encapsulated in different formulations of Ca(ii)-alginate beads. The encapsulated systems presented higher antioxidant capacity in different phases (digested and fermented) than the extracts without encapsulation, making Ca(ii)-alginate beads a suitable delivery vehicle. Levels of total phenolic compounds and antioxidant capacity of the fermented fraction were up to ten times higher than those of the digested fraction, boosted by the contribution of bioactive compounds from the by-product of beet as well as by sugars and biopolymers. Among the formulations used, those that had excipients (sugars and/or biopolymers) presented a better overall antioxidant response than the beads with just alginate. Guar gum and sucrose lead to a promising enhancement of Ca(ii)-alginate beads not only for preservation and protection but also in terms of stability under in vitro digestion-fermentation and production of SCFAs.

Currently and according to the growing worldwide interest in the revaluation of agricultural by-products, the use of legumes waste presents great potential to obtain bioactive compounds. In this context, an extract rich in phenolic compounds was obtained from Vigna unguiculata (cowpea) pods by optimizing the high-intensity ultrasound conditions (10 min and 36% of amplitude) using response surface methodology. Then, the extract was encapsulated in Ca(II)-alginate beads with the addition of arabic or guar gums or cowpea isolated proteins. A complete morphological study by image analysis and microstructural evaluation by SAXS has been carried out. Results showed that beads containing alginate and alginate-guar gum have the highest loading efficiency of total phenolic compounds (47 ± 5%) and antioxidant activity (44 ± 3%). However, the coupled effect of the cowpea extract and the isolated proteins (at it higher concentration) increased the antioxidant capacity of the beads due to the contribution of the phenolic compounds and the amino acids with anti-radical activity, reaching a value of 67 ± 3 % of inhibition of ABTS .+ . Finally, the microstructural analyses revealed that cowpea pod extract increased the interconnectivity of the rods due to the presence of trivalent cations, conferring versatility, and larger coordination to the network. Also, it was observed that the addition of cowpea proteins produced more interconnected bigger and fewer compacts rods than beads containing only alginate, increasing 12 and 49 % the interconnection and the size, respectively, and decreasing 10 % their compactness. This research demonstrated the use of cowpea sub-products as a source of bioactive compounds that further modulate the microstructure of the hydrogel network, and the outstanding potential for being incorporated in techno-functional foods by using Ca(II)-alginate as a carrier., (© 2020 Published by Elsevier Ltd.)

Lycopene extracted from pink grapefruit was encapsulated on Ca(II)-alginate beads with the addition of trehalose and galactomannans to improve its stability against freezing and drying. Three galactomannans of different physicochemical properties were studied since their inclusion affects both loading efficiency and release of lycopene in wet beads; however, there is no information about their performance during freezing and dehydration operations. The remaining lycopene and its stability towards isomerization were analyzed in beads subjected to continuous freezing, freezing/thawing cycles and vacuum- and freeze-drying. Isothermal crystallization studies were conducted by LF-NMR and related to beads formulation and lycopene stability. In the absence of excipients, lycopene was severely affected by all the treatments, retaining less than 20% of the original content. Alginate beads containing trehalose with guar gum protected more than 80% of the lycopene regardless of the employed freezing or drying methods. These beads concomitantly showed higher solid fraction than the other two galactomannans-containing systems, displaying guar gum ability to associate water. On the other hand, the addition of vinal gum affected lycopene stability (between 40 and 60% were recovered after treatments), even compromising the positive effect of a well-established cryoprotectant as trehalose. Thus, the addition of secondary excipients should be carefully conducted. The differences among galactomannans could be related to the substitution degree of the polymer chains, affecting the overall systems interactions. These results can contribute to excipients selection for the encapsulation of labile biomolecules in Ca(II)-alginate beads subjected to freezing and drying., Competing Interests: Conflict of interestThe author declares that there is no conflict of interests regarding the publication of this paper.

The Ca(II)-alginate beads were formulated changing some synthesis variables: pH (3.8-6.8), extrusion tip size (0.25-0.50 mm) and washing/storage protocol, to evaluate possible implications in the structural properties of beads, which are critical to scale and standardize their production at industrial level. Regardless of the macro- (diameter, area, perimeter and roundness, studied by image analysis) and micro-structural (size, density and interconnectivity of rods assessed by SAXS) parameters analyzed, there are no effects related to the washing and storage protocol employed for any synthesis conditions. Structural parameters are only influenced by the synthesis pH. Both washing protocol and extrusion tip size effects on the consolidation of the alginate network are negligible at each pH value. Besides, none of the synthesis variables affected the availability of water within the beads as assessed by diffusion coefficient and water activity measurements. A model, relating chain-chain interactions and polymer chain packing, is proposed., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Context: Indigofera suffruticosa Miller (Fabaceae) and I. truxillensis Kunth produce compounds, such as isatin (ISA) and indirubin (IRN), which possess antitumour properties. Their effects in mammalian cells are still not very well understood., Objective: We evaluated the activities of ISA and/or IRN on cell viability and apoptosis in vitro, their genotoxic potentials in vitro and in vivo, and the IRN- and ISA-induced expression of ERCC1 or BAX genes., Materials and Methods: HeLa and/or CHO-K1 cell lines were tested (3 or 24 h) in the MTT, Trypan blue exclusion, acridine orange/ethidium bromide, cytokinesis-blocked micronucleus (CBMN) and comet (36, 24 and 72 h) tests after treatment with IRN (0.1 to 200 μM) or ISA (0.5 to 50 μM). Gene expression was measured by RT-qPCR in HeLa cells. Swiss albino mice received IRN (3, 4 or 24 h) by gavage (50, 100 and 150 mg/kg determined from the LD 50 - 1 g/kg b.w.) and submitted to comet assay in vivo., Results: IRN reduced the viability of CHO-K1 (24 h; 5 to 200 μM) and HeLa cells (10 to 200 μM), and was antiproliferative in the CBMN test (CHO-K1: 0.5 to 10 μM; HeLa: 5 and 10 μM). The drug did not induce apoptosis, micronucleus neither altered gene expression. IRN and ISA were genotoxic for HeLa cells (3 and 24 h) at all doses tested. IRN (100 and 150 mg/kg) also induced genotoxicity in vivo (4 h)., Conclusion: IRN and ISA have properties that make them candidates as chemotherapeutics for further pharmacological investigations.

β-Galactosidase (lactase) is a widely used enzyme in the food industry; however, it has low stability against thermal and mechanical treatments. Due to this, the purpose of the present research was to analyze the encapsulation of lactase in alginate-Ca(II) beads in order to maintain its enzymatic activity toward freezing, freezing/thawing, and storage. Also, the effect of the addition of trehalose, and arabic and guar gums and their influence on the microstructure as well as on thermal properties and molecular mobility were studied. Lactase was successfully encapsulated in alginate-Ca(II) beads, and the inclusion of trehalose was critical for activity preservation toward treatments, being improved in guar gum-containing systems. The gums increased the T m ' values, which represents a valuable technological improvement. Finally, the presence of secondary excipients affected the microstructure, showing rods with smaller outer diameter and with lower compactness than alginate-Ca(II) beads. Also, bead composition greatly affects the size, shape, and relaxation times.

Background: Syngonanthus arthrotrichus and Syngonanthus bisulcatus, currently known for Comanthera aciphylla (Bong.) L.R.Parra & Giul. and Comanthera bisulcata (Koern.) L.R. Parra & Giul, popularly known in Brazil as "sempre-vivas," are plants from the family Eriocaulaceae. They are found in the states of Minas Gerais and Bahia. The species are known to be rich in flavonoids to which their gastroprotective activity has been attributed. In this research, experimental protocols were performed to elucidate the associated mechanisms of action., Methods: The activity was evaluated using induced gastric ulcer models (acetic acid and ethanol-induced gastric lesions in NEM or L-NAME pre-treated mice, and by ischemia/reperfusion). Antioxidant enzymes, serum somatostatin, and gastrin were also evaluated., Results: In chronic gastric ulcers, a single daily oral dose of Sa-FRF or Sb-FRF (100 mg/kg body wt.) for 14 consecutive days accelerated ulcer healing to an extent similar to that seen with an equal dose of cimetidine. The pre-treatment of mice with NEM (N-ethylmaleimide) or L-NAME (N-nitro-L-arginine) abolished the protective activity of Sa-FRF, Sa-FDF, Sb-FDF and Sb-FRF or Sa-FRF and Sb-FRF, respectively, which indicates that antioxidant compounds and nitric oxide synthase activity are involved in the gastroprotective. Sa-FRF and Sb-FRF (100 mg/kg p.o) protected the gastric mucosa against ulceration that was induced by ischemia/reperfusion (72 and 76 %, respectively). It also decreased lipid peroxidation and restored total thiols in the gastric wall of mice that had been treated with ethanol. When administered to rats submitted to ethanol-induced gastric lesions, Sa-FRF and Sb-FRF (100 mg/kg, p.o.) increased the somatostatin serum levels, while the gastrin serum levels were proportionally decreased., Conclusions: The results indicate significant healing effects and gastroprotective activity for the Sa-FRF and Sb-FRF, which probably involves the participation of SH groups, nitric oxide (NO), the antioxidant system, somatostatin, and gastrin. All are integral parts of the gastrointestinal mucosa's cytoprotective mechanisms against aggressive factors.

Isatin (1H-indole-2,3-dione) is a chemical found in various medicinal plant species and responsible for a broad spectrum of pharmacological and biological properties that may be beneficial to human health, as an anticonvulsant, antibacterial, antifungal, antiviral, and anticancer agent. The aim of the present study was to determine in vitro the cytotoxic, mutagenic, and apoptotic effects of isatin on CHO-K1 and HeLa cells using the MTT viability assay (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide), micronucleus (MN) test, apoptosis index, and nuclear division index (NDI). The 5 isatin concentrations evaluated in the mutagenicity and apoptosis tests were 0.5, 1, 5, 10, and 50 μM, selected through a preliminary MTT assay. Positive (doxorubicin, DXR) and negative (phosphate buffered saline, PBS) control groups were also included in the analysis. Isatin did not exert a mutagenic effect on CHO-K1 after 3 and 24 h of treatment or on HeLa cells after 24 h. However, 10 and 50 μM concentrations inhibited cell proliferation and promoted apoptosis in both CHO-K1 and HeLa cells. Data indicate that the cytotoxic, apoptotic, and antiproliferative effects of isatin were concentration independent and cell line independent.

A recent type 1 diabetes (T1D) clinical trial of rituximab (a B cell-depleting anti-CD20 antibody) achieved some therapeutic benefit in preserving C-peptide for a period of approximately nine months in patients with recently diagnosed diabetes. Our previous data in the NOD mouse demonstrated that co-administration of antigen (insulin) with anti-CD3 antibody (a T cell-directed immunomodulator) offers better protection than either entity alone, indicating that novel combination therapies that include a T1D-related autoantigen are possible. To accelerate the identification and development of novel combination therapies that can be advanced into the clinic, we have evaluated the combination of a mouse anti-CD20 antibody with either oral insulin or a proinsulin-expressing DNA vaccine. Anti-CD20 alone, given once or on 4 consecutive days, produced transient B cell depletion but did not prevent or reverse T1D in the NOD mouse. Oral insulin alone (twice weekly for 6 weeks) was also ineffective, while proinsulin DNA (weekly for up to 12 weeks) showed a trend toward modest efficacy. Combination of anti-CD20 with oral insulin was ineffective in reversing diabetes in NOD mice whose glycemia was controlled with SC insulin pellets; these experiments were performed in three independent labs. Combination of anti-CD20 with proinsulin DNA was also ineffective in diabetes reversal, but did show modest efficacy in diabetes prevention (p = 0.04). In the prevention studies, anti-CD20 plus proinsulin resulted in modest increases in Tregs in pancreatic lymph nodes and elevated levels of proinsulin-specific CD4+ T-cells that produced IL-4. Thus, combination therapy with anti-CD20 and either oral insulin or proinsulin does not protect hyperglycemic NOD mice, but the combination with proinsulin offers limited efficacy in T1D prevention, potentially by augmentation of proinsulin-specific IL-4 production.

The present study evaluated the antiulcerogenic activity and mechanisms of the aqueous (AqF 100 mg/kg) and ethyl acetate (AcF 50 mg/kg) fractions from Indigofera truxillensis leaves. This dose was selected to assess its activity on ulcer healing and its action on gastric acid and mucus secretion, prostaglandin production and antioxidant enzyme activity (superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and glutathione reductase (GSH-Rd)). Gastric ulcer was induced by absolute ethanol. Antisecretory action, mucus and prostaglandin production, healing and antioxidant enzyme activities were evaluated for both fractions. AqF and AcF significantly inhibited the gastric mucosal damage caused by ethanol. This effect was statistically significant at 100 and 50 mg/kg compared with the vehicle. Neither fraction interfered with gastric secretion. AcF increased the PGE(2) production, and both fractions increased mucus production. l-NAME did not alter the gastroprotection exerted by the fractions, but N-ethylmaleimide attenuated only AcF. In the ischemia/reperfusion model both fractions inhibited the mucosal damage. AcF increased SOD, GSH-Px and GSH-Rd activity, but AqF increased only SOD and GSH-Px. In the acetic acid-induced ulcer model AcF only accelerated ulcer healing. These results showed that Indigofera truxillensis acted as a gastroprotective agent, stimulating protective factors and antioxidants enzymes.

One of the major disadvantages of the current cancer therapy is the suppression of the immune system. Brazilian flora is considered one of the most diverse in the world and many plants were found to contain active constituents that can be valuable sources of new drugs. The plant Indigofera suffruticosa was studied to determine its potential to stimulate the immune system and also to be effective against tumour cells. We investigated the effects of the alkaloidal fraction and the pure alkaloid indigo obtained from I. suffruticosa on macrophage activation by measuring nitric oxide (NO) and tumour necrosis factor-α (TNF-α) production. Cytotoxic activity was also evaluated against the two tumour murine cells lines, LM2 (breast adenocarcinoma) and LP07 (lung adenocarcinoma). The alkaloidal fraction induced a high NO production and a moderated TNF-α release. The pure indigo demonstrated an elevated NO and TNF-α production. The fraction and the pure compound also exhibited cytotoxic activity against both adenocarcinoma cell lines and indigo showed the strongest cytotoxic activity with IC₅₀ value of 0.89 µg mL⁻¹ against LM2 and 1.44 µg mL⁻¹ against LP07. Our results presented the immunostimulatory and cytotoxic activity of I. suffruticosa, enhancing macrophage function and therefore contributing to the host defence against tumours.

Alchornea triplinervia (Spreng.) Muell. Arg (Euphorbiaceae) is a medicinal plant commonly used by people living in the Cerrado region of Brazil to treat gastrointestinal ulcers. We previously described the gastroprotective action of methanolic extract (ME) of Alchornea triplinervia and the ethyl acetate fraction (EAF) in increasing of prostaglandin E₂ (PGE₂) gastric levels in the mucosa. In this work we evaluated the effect of EAF in promoting the healing process in rats with acetic acid-induced gastric ulcers. In addition, toxicity was investigated during treatment with EAF. After 14 days of treatment with EAF, the potent stimulator of gastric cell proliferation contributed to the acceleration of gastric ulcer healing. Upon immunohistochemical analysis, we observed a pronounced expression of COX-2, mainly in the submucosal layer. The 14-day EAF treatment also significantly increased the number of neutrophils in the gastric mucosa regeneration area. The EAF induced angiogenesis on gastric mucosa, observed as an increase of the number of blood vessels supplying the stomach in rats treated with EAF. Oral administration for 14 days of the ethyl acetate fraction from Alchornea triplinervia accelerated the healing of gastric ulcers in rats by promoting epithelial cell proliferation, increasing the number of neutrophils and stimulation of mucus production. This fraction, which contained mainly phenolic compounds, contributed to gastric mucosa healing.

Ethnopharmacological Relevance: Indigofera suffruticosa is specie typical of the "Cerrado" or Brazilian savannah; it is a member of the Fabaceae family - in folkmedicine is used for gastric disorders, infection and inflammation., Aim of the Study: Ethyl acetate fraction (AcF) and aqueous fraction (AqF) of the methanolic extract of I. suffruticosa leaves were evaluated against acute gastric ulcer. The AcF fraction was selected to assess its activity in ulcer healing and its gastroprotective effects via mucus and gastric secretion., Materials and Methods: The gastroprotective action of AcF and AqF fractions were evaluated in a rodent experimental model. The action mechanisms, involvements of the antisecretory action, mucus and prostaglandin production, toxicological and healing activity of the AcF (100mg/kg, p.o.) were evaluated. We also used histological analysis (HE and PAS) and immunohistochemical (PCNA and HSP-70) assays to evaluate the effects of I. suffruticosa., Results: AcF significantly inhibited the gastric mucosal damage caused by ethanol. This effect was statistically significant in 100mg/kg group compared vehicle. AcF did not interfered with gastric secretion, significantly increased the PGE(2) and mucus production (validated in PAS technique). The gastroprotection was attenuated by pretreatment with N-ethylmaleimide, but not L-NAME. In acid-acetic-induced ulcer model AcF accelerated ulcer healing. Immunohistochemistry analysis showed induction of proliferating cell (PCNA) and heat shock protein (HSP 70)., Conclusions: These results showed that AcF acted as gastroprotective agent stimulating prostaglandin, mucus and HSP70., (Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.)

Isatin (1H-indole-2,3-dione) is a synthetically versatile substrate used for the synthesis of heterocyclic compounds and as a raw material for drug synthesis. Isatin and its derivatives demonstrate anticonvulsant, antibacterial, antifungal, antiviral, and anticancer properties. We evaluated the genotoxic and mutagenic effects of acute (24h) and repeated (14d) exposure to isatin in vivo, using the comet assay and the micronucleus test. Three doses (50, 100, and 150mg/kgb.w.) were administered to mice via gavage. Doses were selected according to the LD(50) of isatin, estimated in a preliminary test to be 1g/kgb.w. To evaluate the results, parametric (ANOVA/Tukey) and non-parametric (Kruskal-Wallis/Dunn's post hoc test) tests were used, according to the nature of the data distribution. At all doses (50, 100 and 150mg/kgb.w.), after acute treatment with isatin, alterations in DNA migration (comet assay) were not observed and mutagenic effects were not seen (micronucleus test on peripheral blood cells). After repeated doses, only the highest dose of isatin (150mg/kgb.w.) induced alterations in the DNA that gave rise to micronuclei in the bone marrow and peripheral blood cells of the mice. Our results show that the mutagenic and genotoxic effects of isatin depend on dose and on period of exposure., (Copyright © 2010 Elsevier B.V. All rights reserved.)

Indigofera truxillensis and I. suffruticosa, are used as a source of indigo dye and to treat several diseases. The mutagenic activity of the methanolic extracts from aerial parts, glycerolipid, flavonoid and alkaloid fractions of the extract were evaluated by means of Salmonella/microsome assays using TA100, TA98, TA102 and TA97a strains. The methanolic extract of I. truxillensis showed mutagenic activity in the TA98 strain without S9 while glycerolipid fraction was devoid of activity. The flavonoid and alkaloid fractions of both plants showed mutagenicity. Chemical analysis of flavonoid fractions of I. truxillensis and I. suffruticosa resulted in the identification of kaempferol, quercetin and their derivatives. The alkaloid fraction of both the species contained indigo and indirubin and indigo was found mainly responsible for the mutagenic activity.

Context: Alchornea triplinervia (Spreng.) Müll. Arg. (Euphorbiaceae) is a tree widespread in many Brazilian states. This plant naturally occurs in different ecosystems including tropical Atlantic forest, Amazon rain forest, moist tropical mixed forest, savanna forest, among others. Local populations traditionally use it in tea form to treat gastric disturbances., Objective: The objective of this research was to evaluate the plant A. triplinervia as a potential inhibitor of some macrophage functions involved in the inflammatory process., Materials and Methods: The effects of Alchornea triplinervia ethyl acetate fraction (AtF) on hydrogen peroxide (H₂O₂), nitric oxide (NO) and tumor necrosis factor-α (TNF-α) production in peritoneal macrophages were investigated using phenol red, Griess reagent and a sandwich immunoassay, respectively., Results: AtF chromatographic analyses indicate the presence of flavonoids as majority compounds. The fraction also showed an intense inhibition of H₂O₂ and NO production. The inhibitory effects of the fraction in H₂O₂ and NO production ranged from 72.25 ± 4.68 to 69.64 ± 4.21 and from 47.8 ± 8.96 to 76.77 ± 8.11%, respectively in the two tested concentrations, 15.62 and 62.5 μg/mL. TNF-α production was partially inhibited in the tested concentrations and the inhibitory rate was around 18%., Discussion and Conclusion: It is supposed that the elevated biological potential of A. triplinervia is related to the presence of phenolic compounds in the plant leaves. According to the results observed in this study, it is suggested that AtF presents anti-inflammatory activity, supporting the traditional use of A. triplinervia in Brazilian folk medicine.

Mycobacterium tuberculosis is responsible for over 8 million cases of tuberculosis (TB) annually. Natural products may play important roles in the chemotherapy of TB. The antimycobacterial activity and the innate immune response of methanol (METH) and dichloromethane (DCM) extracts of Indigofera suffruticosa Miller (Fabaceae) were evaluated. We observed that the minimum inhibitory concentrations (MICs) for METH and DCM extracts were 125 and 1000 microg/mL, respectively. However, they were able to induce the innate immune response through the production of high levels of NO and TNF-alpha (p < 0.001) by peritoneal exudate cells (PECs). These results suggest that I. suffruticosa extracts may have an important immunological role in the control of TB once macrophage activity is induced by them.

Phenolic compounds are produced by secretory idioblasts and hypodermis, and by specialized cells of the epidermis and chlorenchyma of leaves of Alchornea triplinervia. Phytochemical investigation of these leaves led to the isolation of the known substances quercetin, quercetin-7-O-beta-D-glucopyranoside, quercetin-3-O-beta-D-glucopyranoside, quercetin-3-O-beta-D-galactopyranoside, quercetin-3-O-alpha-L-arabinopyranoside, amentoflavone, brevifolin carboxylic acid, gallic acid, and methyl gallate from the methanolic extract, and stigmasterol, campesterol, sitosterol, lupeol, friedelan-3-ol, and friedelan-3-one from the chloroform extract. In studies of antibacterial activity and mutagenicity, the methanolic extract showed promising activity against Staphylococcus aureus (MIC = 62.5 microg/mL) and was slightly mutagenic in vitro and in vivo at the highest concentrations tested (1335 mg/kg b.w.).

Among the current treatment strategies for the peptic ulcer patient with Helicobacter pylori infection, the method of choice is triple therapy based on the concurrent use of proton inhibitors and two antibiotics. Alchornea triplinervia is a medicinal plant commonly used by people living in the Cerrado region of Brazil to treat gastrointestinal ulcers. In the present work we proposed therapy based on this medicinal plant that presents effective gastroprotective action with antibiotic effects. Oral pretreatment with methanolic extract (ME) of A. triplinervia in rats and mice decreased the gastric injuries induced by ethanol and HCl/ethanol. Increasing the dose reduced the gastroprotective effects of ME on the gastric lesions induced by nonsteroidal anti-inflammatory drug. After pylorus ligature of mice, oral administration of ME induced a decrease not only in total acid but also in the ulcer index. We also observed that ME displayed antibacterial activity against H. pylori. Liquid-liquid separation of ME indicated that active constituents responsible for the gastroprotective action are concentrated in the ethyl acetate fraction (EAF) (50% protection) rather than in the aqueous fraction, which did not induce significant gastroprotection at the same dose (100 mg/kg). EAF induced an increase of gastric mucosa prostaglandin (PG) E(2) levels, which remained high even after previous administration of indomethacin. The phytochemical profile of ME revealed that EAF contains mainly flavonoids. In conclusion, all these results suggest that ME did not show acute toxicity, but exhibited an antisecretory property, anti-H. pylori effect, and gastroprotective action. The observed effect did not involve the participation of nitric oxide or endogenous sulfhydryl groups. However, EAF showed a more efficient gastroprotective effect than ME at a lower dose and protected the gastric mucosa by increasing PGE(2).

Ethanol-induced oxidative damage is commonly associated with the generation of reactive oxygen molecules, leading to oxidative stress. Considering that antioxidant activity is an important mechanism of action involved in cytoprotection, the aim of this work was to evaluate the antioxidant properties of the alkaloid indigo (1) (2 mg/kg, P. O.), obtained from the leaves of Indigofera truxillensis Kunth (Fabaceae), on rat gastric mucosa submitted to ethanol-induced (100%, 1 mL, P. O.) gastric ulcer. Enzymatic assays and DNA fragmentation analysis were performed. When ethanol was administered to the control group, the sulfhydryl content (SH) and the glutathione peroxidase (GPx) activity decreased by 41% and 50%, respectively; in contrast, superoxide dismutase (SOD) and glutathione reductase (GR) activities increased by 56% and 67%, respectively. Additionally, myeloperoxidase (MPO) activity, a marker for free radical generation caused by polymorphonuclear neutrophil (PMN) tissue infiltration, also increased 4.5-fold after ethanol treatment. Rat gastric mucosa exposed to ethanol showed DNA fragmentation. Indigo alkaloid pretreatment protected rats from ethanol-induced gastric lesions. This effect was determined by the ulcerative lesion area (ULA), indicating an inhibition of around 80% at 2 mg/kg. This alkaloid also diminished GPx activity, which was higher than that observed with ethanol alone. However, this effect was counterbalanced by increased GR activity. Indigo was unable to restore alterations in SOD activity promoted by ethanol. After indigo pretreatment, SH levels and MPO activity remained normal and gastric mucosa DNA damage caused by ethanol was also partially prevented by indigo. These results suggest that the gastroprotective mechanisms of indigo include non-enzymatic antioxidant effects and the inhibition of PMN infiltration which, in combination, partially protect the gastric mucosa against ethanol-induced DNA damage.

Alchornea glandulosa (Euphorbiaceae) is a plant used in folk medicine as an antiulcer agent. Rats pretreated with methanolic extract obtained from the leaves of A. glandulosa (AG) showed a dose-dependent effect and significant reduction of gastric ulcers induced by absolute ethanol at the doses of 500 (57%) and 1000 mg/kg (85%) in relation to the control group. Pretreatment of mice with AG (500, 1000 mg/kg, p.o.) showed dose-dependent activity and significantly decreased the severity of lesions caused by HCl/ethanol and by non steroidal anti inflammatory drug-induced gastric lesions. Pretreatment with AG also induced antisecretory action via local and systemic routes and a significant decrease in the total gastric acid content. The gastroprotective effects of AG involved the participation of nitric oxide and increased levels of endogenous sulfhydryl compounds, which are defensive mechanisms of the gastrointestinal mucosa against aggressive factors. The ability of AG to heal gastric ulcers was evaluated after 14 consecutive days of treatment. The results showed that single oral administrations of AG (250 mg/kg/once daily) potently stimulates gastric epithelial cell proliferation that contributes to the accelerated healing of gastric ulcers induced by acetic acid. In addition, no subacute toxicity (body weight gain, vital organs, and serum biochemical parameters) was observed during treatment with AG. Phytochemical investigation of AG led to the isolation of myricetin-3-O-alpha-L-rhamnopyranoside, quercetin-3-O-alpha-L-arabinopyranoside, quercetin-3-O-beta-D-galactopyranoside, quercetin, amentoflavone, methyl gallate, gallic acid, and pterogynidine. We also established the phytochemical profile of AG with the quantification of total phenolic compounds. These compounds may contribute to the observed antiulcerogenic effects of AG.

The hydroethanolic extract of the leaves (HEL) and bark (HEB) obtained from Alchornea castaneaefolia (Euphorbiaceae) were investigated for their ability to prevent ulceration of the gastric mucosa in animal models. HEL (500 and 1000 mg/kg) and HEB (1000 mg/kg) significantly reduced the gastric injuries induced by the combination of HCl/ethanol and lowered the severity of gastric damage formation induced by indomethacin/bethanechol in mice. Further investigation showed that HEL also inhibited formation of ulcers in mice submitted to stress and pylorus ligature, but HEL did not modify gastric juice parameters in Shay mice. HEL was also effective in promoting the healing process in chronic gastric ulcer induced by acetic acid in rats. An enriched flavonoidic fraction (EFF at dose of 100mg/kg) obtained from HEL reduced gastric lesions induced by HCl/ethanol and indomethacin/bethanechol in mice. Although EFF did not modify the amount of free mucus production by gastric mucosa, it was able to increase prostaglandin production. When administered to rats submitted to ethanol-induced gastric lesions, EFF increased the somatostatin serum levels, while the gastrin serum levels were proportionally decreased. Phytochemical investigation on HEL and EFF led to the isolation of flavonoids glycosides as the main compounds, thus suggesting that these substances may be involved in the observed antiulcer activity.

The effects of Alchornea glandulosa ethyl acetate fraction (AGF) on hydrogen peroxide (H2O2), nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) production in peritoneal macrophages activated with lipopolysaccharide (LPS) or phorbol myristate acetate (PMA) were investigated. Analysis by thin layer chromatography (TLC) of AGF showed several constituents, including flavonoids, which may have anti-inflammatory activity. Inhibitory effects of the fraction in H2O2 and NO production ranged from 8.59+/-7.84% to 70.56+/-4.16% and from 16.06+/-3.65% to 38.73+/-3.90%, respectively. The TNF-alpha production was only partially inhibited in the tested concentrations (12.21+/-6.23% - 15.16+/-0.96%). According to these results, it is suggested that AGF has anti-inflammatory activity. This medicinal plant may have therapeutic potential in the control of inflammatory disorders.

Syngonanthus arthrotrichus SILVEIRA, popularly known as "sempre-vivas mini-saia," is found in mountains of the Espinhaço range in the Brazilian states of Bahia and Minas Gerais. Extracts of this species contain several constituents, including flavonoids which may have antiulcerogenic activity. An ethanolic extract (EEOH), and flavonoid-rich (FRF) and flavonoid-deficient (FDF) fractions obtained from the scapes of S. arthrotrichus were investigated for their ability to prevent ulceration of the gastric mucosa in mice and rats. In the ethanol/HCl-induced ulcer model, lansoprazole (30 mg/kg), EEOH (50, 100, 250 mg/kg) given orally protected the gastric mucosal against injury in mice by 79%, 78%, 73%, and 64% respectively. In the ethanol-induced gastric ulcer model in rats, the lansoprazole (30 mg/kg), FRF and FDF (100 mg/kg) significantly protected the gastric mucosal of rats by 65%, 38% and 25% respectively when compared with the negative control group. In indomethacin/bethanechol-induced gastric ulcers, cimetidine (100 mg/kg) and the EEOH (100, 250 mg/kg) inhibited gastric ulcer formation by 73%, 55% and 32% respectively. In this exactly model other treatments as cimetidine, FRF and FDF (100 mg/kg) each caused 54%, 36% and 45% inhibition, respectively. In the stress-induced gastric ulcer model, cimetidine (100 mg/kg) and the EEOH (50, 100, 250 mg/kg), inhibited gastric ulcer formation by 63%, 73%, 68% and 69% respectively. In the same model, cimetidine, FRF and FDF (100 mg/kg) significantly protected the gastric mucosal of the mice by 60%, 51% and 47% when compared to the control group. In pylorus-ligated mice, cimetidine (positive control) and FRF significantly decreased gastric acid secretion, increased gastric pH and reduced the acid output when compared to the negative control. FDF had no significant effect on these parameters. The protection provided by FRF probably involved an antisecretory mechanism mediated by flavonoids which were absent in FDF. The amount of adherent mucous in the stomach contents was also evaluated with the treatments carbenoxolone (200 mg/kg), FRF and FDF (100 mg/kg) treatment. Each treatments significantly increased the amount of adherent mucous in the gastric juice (8.67+/-1.73, 3.35+/-1.59, 2.1+/-0.41 mg/g of wet tissue, respectively) compared to the control group, indicating a cytoprotective action on the gastric mucosa. Treatment with FRF plus indomethacin and FDF plus indomethacin reduced the prostaglandin biosyntesis (13.6+/-6.5, 27+/-5.5 pg/well) by the mucosa, indicating that the cytoprotective action on the gastric mucosa was not related to the level of prostaglandins. Only FDF (38+/-17 pg/well) maintained the level of prostaglandins and guaranteed the integrity of the mucosa. The results indicate that the EEOH, FRF and FDF have antisecretory and cytoprotective actions, that may be related to the presence of luteoline in the extract and active fractions.