Assessment of multispecies biofilm growth on root canal dentin under different radiation therapy regimens.

OBJECTIVES: To assess the growth of a multispecies biofilm on root canal dentin under different radiotherapy regimens. MATERIALS AND METHODS: Sixty-three human root dentin cylinders were distributed into six groups. In three groups, no biofilm was formed (n = 3): NoRT) non-irradiated dentin; RT55) 55 Gy; and RT70) 70 Gy. In the other three groups (n = 18), a 21-day multispecies biofilm (Enterococcus faecalis, Streptococcus mutans, and Candida albicans) was formed in the canal: NoRT + Bio) non-irradiated + biofilm; RT55 + Bio) 55 Gy + biofilm; and RT70 + Bio) 70 Gy + biofilm. The biofilm was quantified (CFUs/mL). Biofilm microstructure was assessed under SEM. Microbial penetration into dentinal tubules was assessed under CLSM. For the biofilm biomass and dentin microhardness pre- and after biofilm growth assessments, 45 bovine dentin specimens were distributed into three groups (n = 15): NoRT) non-irradiated + biofilm; RT55 + Bio) 55 Gy + biofilm; and RT70 + Bio) 70 Gy + biofilm. RESULTS: Irradiated specimens (70 Gy) had higher quantity of microorganisms than non-irradiated (p = .010). There was gradual increase in biofilm biomass from non-irradiated to 55 Gy and 70 Gy (p < .001). Irradiated specimens had greater reduction in microhardness after biofilm growth. Irradiated dentin led to the growth of a more complex and irregular biofilm. There was microbial penetration into the dentinal tubules, regardless of the radiation regimen. CONCLUSION: Radiotherapy increased the number of microorganisms and biofilm biomass and reduced dentin microhardness. Microbial penetration into dentinal tubules was noticeable. CLINICAL RELEVANCE: Cumulative and potentially irreversible side effects of radiotherapy affect biofilm growth on root dentin. These changes could compromise the success of endodontic treatment in oncological patients undergoing head and neck radiotherapy.

Smart labels based on polyvinyl alcohol incorporated with chitosan nanoparticles loaded with grape extract: Functionality, stability and food application.

Anthocyanins (ACNs) are natural compounds with potential applications due to their colorimetric response to pH. Due to their sensitivity to various environmental factors, nanoencapsulation with biopolymers is a successful strategy for stabilizing ACNs. In this work ACNs were extracted from grape skins and encapsulated into chitosan (CS) nanoparticles by ionic gelation using sodium tripolyphosphate (TPP) as a cross-linking agent. CS nanoparticles loaded with ACNs had particle sizes between 291 and 324 nm and polydispersity index around 0.3. The encapsulation efficiency of ACNs was approximately 60 %; and encapsulated anthocyanins (ACN-NPs) exhibited color change properties under different pH conditions. pH-sensitive labels based on polyvinyl alcohol (PVA) were prepared by the casting method. The effect of incorporating ACN-NPs on the physical, structural, and pH-sensitive properties of PVA labels was evaluated, and its application as shrimp freshness indicator was studied. The nanoencapsulation protected ACNs against heat and light treatments, preserving the original purple color. When applying the label, visible changes from red to blue until reaching yellow were observed with the change in the quality of the shrimp at the refrigeration temperature. The results suggest that PVA labels containing ACNs encapsulated in C-NPs can be used as smart packaging labels in the food industry.

The Potential of Crude and Partially Purified Black Rice Bran Extracts Obtained by Ultrasound-Assisted Extraction: Anti-Glycemic, Cytotoxicity, Cytoprotective, and Antitumoral Effects.

Recovering anthocyanins from black rice bran is a way of valuing this byproduct, by obtaining an extract with biological potential. The objective of this study was to recover anthocyanins using ultrasound-assisted extraction. Some of the extract was partially purified, and both (crude and partially purified) extracts were evaluated for their anthocyanin content, antioxidant activity, antidiabetic and antitumoral activities, cytotoxicity, and oxidative stress. An increase in the laboratory scale was also achieved, making possible to increase the extraction volume up to 20 times without significantly changing the content of anthocyanins (1.85 mg C3G/g DW). It was found that the purified sample presented a 4.2 times higher value of total anthocyanins compared to the crude sample. The best IC50 values for the purified sample were verified by DPPH and ABTS (0.76 and 0.33 mg/mL). The best results for antidiabetic activity were obtained for the partially purified sample: 0.82 µM C3G for α-glucosidase and 12.5 µM C3G for α-amylase. The extracts demonstrated protection (~70%) when subjected to the oxidative stress of L929 cells. An antitumoral effect of 25-30% for both extracts was found in A459 cells. The crude and partially purified extracts of black rice have antidiabetic and anticancer effects and more studies are needed to explore their potential.

Starch nanoparticles containing phenolic compounds from green propolis: Characterization and evaluation of antioxidant, antimicrobial and digestibility properties.

This study investigated the production of nanoparticles through nanoprecipitation using cassava and potato starches as carriers to stabilize phenolic compounds (PC) from green propolis extract (PE). Additionally, the antioxidant and antimicrobial activities of PC stabilized with starch nanoparticles (SNPs), as well as their release under gastrointestinal conditions were investigated. PE exhibited antioxidant and antibacterial properties, especially PE3 (PE produced using sonication by 20 min and stirring at 30 °C for 24 h) had the highest concentrations of p-coumaric acid, rutin, kaempferol and quercetin. SNPs displayed bimodal distribution with particle size lower than 340 nm. The stabilization of PC increased surface charge and hydrophobicity in SNPs. Moreover, SNPs containing PC from PE exhibited antibacterial activity against Listeria monocytogenes, at a concentration of 750 mg/mL. Low release of PC was observed from the nanoparticles when exposed under simulated gastrointestinal conditions. These nanomaterials could be used as natural ingredients with antioxidant and antimicrobial properties.

Bioactive compounds by UHPLC-MS/MS and molecular networking from Myrceugenia foveolata.

Myrceugenia foveolata, commonly known as 'guamirim', was analysed using UHPLC-MS/MS and molecular networking. Hydroalcoholic extracts of leaves and stem bark were obtained using ethanol:water (70:30) as solvent. Chemical composition of the extracts was identified using UHPLC-MS/MS and tested for antioxidant activity and growth inhibition against E. coli, S. aureus, L. monocytogenes and S. enterica. Ten compounds were tentatively identified in the extracts including myricitrin, quercitrin, and betulin (in leave extracts), and avicularin, kaemferol-3-O-arabinopyroniside, 2"-O-galloylquercitrin and the acid triterpenes ursolic, sumaresinolic, asiatic and maslinic (in both extracts). Both extracts showed similar antioxidant activities, phenolic composition, and growth inhibition. The most pronounced response was observed against L. monocytogenes, with a growth inhibition rate of 73% to leaves extract and 65% do stem bark extract.

Inhibitory impact of the anticancer drug doxorubicin on anaerobic microbial community.

The inhibitory effect of the anticancer drug doxorubicin (DOX) on biogas production was evaluated in short-term and long-term exposure assays. The short-term assays reached the DOX IC50 value on 648 ± 50 µg·L-1. In addition, it was found that inhibition caused by the exposure of 10×103 µg·L-1 was reversible after removing DOX from the feeding synthetic medium. Furthermore, DOX can be rapidly sorbed by the biomass (despite the low Kow), which might contribute to the inhibitory effect. The results of long-term exposure assays, when the DOX volumetric loading rate was increased from 100 µgDOX·L-1·day-1 to 200 µgDOX·L-1·day-1, showed that biogas production and COD removal decreased rapidly. However, the methanogenic Archaeas could recover from this exposure, corroborating the results on short-term exposure assays. In conclusion, DOX can play a key role in inhibiting biological wastewater treatment processes if its concentration in hospital wastewater treatment plants increases abruptly.

Membrane Emulsification as an Emerging Method for Lacticaseibacillus rhamnosus GG® Encapsulation.

Techniques capable of producing small-sized probiotic microcapsules with high encapsulation yields are of industrial and scientific interest. In this study, an innovative membrane emulsification system was investigated in the production of microcapsules containing Lacticaseibacillus rhamnosus GG® (Lr), sodium alginate (ALG), and whey protein (WPI), rice protein (RPC), or pea protein (PPC) as encapsulating agents. The microcapsules were characterized by particle size distribution, optical microscopy, encapsulation yield, morphology, water activity, hygroscopicity, thermal properties, Fourier-transform infrared spectroscopy (FTIR), and probiotic survival during in vitro simulation of gastrointestinal conditions. The innovative encapsulation technique resulted in microcapsules with diameters varying between 18 and 29 µm, and encapsulation yields > 93%. Combining alginate and whey, rice, or pea protein improved encapsulation efficiency and thermal properties. The encapsulation provided resistance to gastrointestinal fluids, resulting in high probiotic viability at the end of the intestinal phase (> 7.18 log CFU g-1). The proposed encapsulation technology represents an attractive alternative to developing probiotic microcapsules for future food applications. Supplementary Information: The online version contains supplementary material available at 10.1007/s11947-023-03099-w.

Black rice and its by-products: anthocyanin-rich extracts and their biological potential.

Recently, growing demand for products enriched with natural compounds that support human health has been observed. Black rice, its by-products, and residues are known to have in their composition a large amount of these compounds with biological potential, mainly anthocyanins. These compounds have reported effects on anti-obesity, antidiabetic, antimicrobial, anticancer, neuroprotective, and cardiovascular disease. Therefore, the extract from black rice or its by-products have great potential for application as ingredients in functional foods, supplements, or pharmacological formulations. This overview summarizes the methods employed for the extraction of anthocyanins from both black rice and its by-products. In addition, trends in applications of these extracts are also evaluated regarding their biological potential. Commonly, the extraction methods used to recover anthocyanins are conventional (maceration) and some emerging technologies (Ultrasound-Assisted Extraction - UAE, and Microwave-Assisted Extraction - MAE). Anthocyanin-rich extracts from black rice have presented a biological potential for human health. In vitro and in vivo assays (in mice) showed these compounds mainly with anti-cancer properties. However, more clinical trials are still needed to prove these potential biological effects. Extracts from black rice and its by-products have great potential in applying functional products with beneficial characteristics to humans and reducing agro-industrial residues.

Enhanced production of surfactin using cassava wastewater and hydrophobic inducers: a prospection on new homologues.

Biosurfactants are amphipathic molecules that can be applied in a wide range of areas. The cost of production limits the industrial application of biosurfactants. Nevertheless, the biosurfactant productivity can be easily enhanced by inducers. This work aimed to investigate the effect of hydrophobic inducers on surfactin production by B. subtilis ATCC 6633 using cassava wastewater as low-cost culture medium. The submerged cultivation was carried out at 30 °C, 150 rpm for 72 h. The fermentation parameters used were bacterial growth, consumption of sugars, and surfactin production, including surfactin homologues. The surface tension decreased by 40% after 12 h, when compared to control. Depletion of sugars was observed in all experiments. Palmitic acid led to the highest yield in terms of surfactin production (≈ 1.3 g·L- 1 of pure surfactin). The inducers triggered the production of new surfactin homologues, that represent, potentially, new biological activities.

Starch- and carboxymethyl cellulose-based films as active beauty masks with papain incorporation.

This work aimed to evaluate the effects of the incorporation of papain in biopolymeric-based beauty face masks with exfoliation activity of the skin. The masks were produced by casting with starch and carboxymethyl cellulose blend (50:50 weight percentages). The macro and microstructure, protein distribution, thickness, moisture content, water contact angle, solubility matter, and mechanical properties were evaluated. Moreover, the in vitro proteolytic and exfoliation activity and storage stability were also evaluated. The films with papain had a more concise matrix which provided higher mechanical properties and lower water solubility when compared to the control film (without papain). Films with 1, 2, and 5 % of papain had enzymatic activity for casein and porcine skin substrates. The micrographs of porcine skin treated with 2 and 5 % of papain showed more difference when compared to the control sample, indicating the enzymatic exfoliation. Differently from the solution of papain, the enzyme that was immobilized in the films maintained its activity for up 90 days during the storage stability assay. Based on the physicochemical properties and proteolytic activities, the films preserved the exfoliation activity of papain and have interesting characteristics to act as beauty masks in the cosmetic field.

A review on enzymatic acylation as a promising opportunity to stabilizing anthocyanins.

Anthocyanins are naturally occurring bioactive compounds found mainly in fruits, vegetables, and grains. They are usually extracted due to their biological properties and great potential for technological applications. These compounds have characteristic pH-dependent colorations that are natural dyes since they come in different colors. However, they are susceptible to processing conditions, remarkably light, temperature, and oxygen. The acylated anthocyanins showed better stability characteristics, and therefore, an acylation process of these compounds could improve their applications. The enzymatic acylation was effective and showed promising results. The current review provides an overview of the works that performed enzymatic acylation of anthocyanins and studies on the stability, antioxidant activity, and lipophilicity. In general, enzymatically acylated anthocyanins showed better stability to light and temperature than non-acylated compounds. In addition, they were liposoluble, a characteristic that allows their addition to products with lipid matrices. The results showed that these compounds formed by enzymatic acylation have perspectives of application mainly as natural colorants in food products. Therefore, the enzymatic acylation of anthocyanins appears viable to increase the industrial applicability of anthocyanins. There are still some gaps to be filled in process optimization, the reuse of enzymes, and toxicity analysis of the acylated compounds formed.

Mannosylerythritol lipids as green pesticides and plant biostimulants.

Biosurfactants can be applied in the formulation of personal care products, as food additives, and as biocontrol agents in the agricultural sector. Glycolipids and lipopeptides represent an important group of microbial-based biosurfactants with biostimulating properties. Among them, the mannosylerythritol lipids also presented antimicrobial activity, mostly against Gram-positive bacteria and phytopathogenic fungi. In this sense, mannosylerythritol lipids are a potential safer green alternative for partially replacing synthetic pesticides. This review aimed to critically discuss the current state of the art and future trends of mannosylerythritol lipids as green pesticides and biostimulants for seed germination and plant growth. Due to their chemical structure, mannosylerythritol lipids are likely related to energy pathways such as glycolysis and Krebs cycle, i.e. a direct cellular biostimulant potential. In this case, experimental evidence from other glycolipids indicated that structural and chemical changes as a potential drug vehicle due to morphological changes caused by biosurfactant-membrane interaction. In addition, like other biosurfactants, mannosylerythritol lipids can trigger self-defense mechanisms, leading to a lower frequency of phytopathogen infections. Therefore, mannosylerythritol lipids have the potential for biostimulation and antiphytopathogenic action, despite that to date no data are available on mannosylerythritol lipids as biostimulants and green pesticides simultaneously. Based on the current state of the art, mannosylerythritol lipids have great potential for a biotechnological advance toward more sustainable agriculture. © 2022 Society of Chemical Industry.

Colorimetric detection of Pseudomonas aeruginosa by aptamer-functionalized gold nanoparticles.

Novel rapid methodologies for the detection of bacteria have been recently investigated and applied. In hospital environments, infections by pathogens are very common and can cause serious health problems. Pseudomonas aeruginosa is one of the most common bacteria, which can grow in hospital equipment such as catheters and respirators. Even at low concentrations, it can cause severe infections as it is resistant to antibiotics and other treatments. Based on this subject's relevance, this work aimed to develop a colorimetric biosensor using aptamer-functionalized gold nanoparticles for identifying P. aeruginosa. The detection mechanism is based on the color change of gold nanoparticles (AuNPs) from red to blue-purple through NaCl induction after bacteria incubation and aptamer-target binding. First, AuNPs were synthesized and characterized. The influence of aptamer and sodium chloride concentration on the agglomeration of AuNPs was investigated. Optimization of aptamer concentration and salt addition were performed. The best condition for detection was 5 µM aptamers and 200 mM of NaCl. In this case, P. aeruginosa was detected after 5 h for concentrations from 108 to 105 CFU mL-1, being 105 and 104 CFU mL-1 the detection limit for color change by the naked eye and UV-Vis spectrometry, respectively. In addition, other bacteria such as E. coli, S. typhimurium, and Enterobacteriaceae bacterium were also detected with color changing from red to gray. Finally, it was confirmed that the salt incubation time can be 2 h, and that the ideal aptamer concentration is 5 µM. Thus, the colorimetric analysis can be a simple and fast detection method for P. aeruginosa in the range of 108 to 105 CFU mL-1 to the naked eye. KEY POINTS: • A new method for rapid detection of Pseudomonas aeruginosa • Aptamers conjugated with gold nanoparticles allow pathogen detection by colorimetry • No need for previous surface modification of nanoparticles.

Intelligent labels manufactured by thermo-compression using starch and natural biohybrid based.

This work aims to develop intelligent labels based on cassava starch and biohybrid pigments by thermo-compression. The biohybrid pigment (BH) was developed by the adsorption of anthocyanins (ACNs) extracted from the jambolan fruit (Syzygium cumini L.) into montmorillonite (Mt) in order to improve its stability. The effect of the addition of biohybrid on the physicochemical properties of the thermo-pressed starch labels was evaluated. ACNs from jambolan extract show a visible pH-dependent color-changing ability at pH 1 - 12, and the adsorption did not modify the color property. The intelligent labels presented a homogeneous surface, and the BH was well dispersed in the starch matrix. The presence of BH increased the solubility in the water of starch labels. Chemical structure characterization revealed that the BH interacted with starch matrices through hydrogen bonds. Furthermore, the thermal stability of starch labels increased with the presence of BH. Hence, the purple color of intelligent labels was preserved at high temperatures. Finally, labels containing BH show visible changes from purple to a blue color when exposed to ammonia vapor, which simulates the degradation of meat products. Thus, the label content jambolan pigments will be used to control meat deterioration.

In vitro effect on the proteolytic activity of papain with proteins of the skin as substrate.

OBJECTIVE: This work aims to evaluate the effects of enzyme concentration, pH, temperature and time course degree of hydrolysis (DH) of papain regarding further development of pharmaceutical and cosmetic formulations. METHODS: The hydrolysis of casein, collagen, keratin and porcine skin at pH and temperature ranges of the human skin was evaluated. Also, low contact times of enzyme-substrate were studied. The incorporation of 3mM of cysteine improved the caseinolytic (PU), collagenolytic (CU) and keratinolytic (KU) activities of papain. RESULTS: In general, the increase from 0.1 to 1.0 or 2.0 mgmL-1 of papain improved PU, CU and KU. When 2.0 mgmL-1 of papain was used, the highest DH of casein, collagen and keratin was obtained at 240min (14%, 35% and 6% respectively). The decrease in pH and temperature reduced all proteolytic activities, but papain maintained at least 50% and 40% of its activity at 26°C and pH  4.5 respectively. Scanning electron micrographs of the surface of the skin showed that papain application had exfoliating activity. CONCLUSION: This preformulation study demonstrated that papain concentration, time of application and pH of the product should be evaluated when developing a product to promote the hydrolysis of the proteins of the skin.

OBJECTIF: Ce travail vise à évaluer les effets de la concentration d'enzyme, du pH, de la température et du degré d'hydrolyse (DH) de la papaïne concernant le développement ultérieur de formulations pharmaceutiques et cosmétiques. MÉTHODES: L'hydrolyse de la caséine, du collagène, de la kératine et de la peau porcine à des plages de pH et de température de la peau humaine a été évaluée. De plus, des faibles temps de contact enzyme-substrat ont été étudiés. L'incorporation de 3mM de cystéine a amélioré les activités caséinolytiques (PU), collagénolytiques (CU) et kératinolytiques (KU) de la papaïne. RÉSULTATS: En général, l'augmentation de 0,1 à 1,0 ou 2,0 mg.mL−1 de papaïne a amélioré la PU, la CU et la KU. Lorsque 2,0 mg mL−1 de papaïne ont été utilisés, les DH les plus élevées de caséine, de collagène et de kératine ont été obtenues à 240 min (14, 35 et 6 %, respectivement). La diminution du pH et de la température a réduit toutes les activités protéolytiques, mais la papaïne a maintenu au moins 50 et 40 % de son activité à 26 °C et pH 4,5, respectivement. Des micrographies obtenues par microscopie électronique à balayage de la surface de la peau ont montré que l'application de la papaïne avait une activité exfoliante. CONCLUSION: Cette étude de pré-formulation a démontré que la concentration de papaïne, le temps d'application et le pH du produit doivent être évalués lors du développement d'un produit pour favoriser l'hydrolyse des protéines de la peau.

Melanoma growth in non-chemically modified translucid bacterial nanocellulose hollow and compartimentalized spheres.

BACKGROUND: Bacterial nanocellulose (BNC) has been used as cell support in numerous tissue engineering studies. Its use can be explained based on the fact its structure allows the creation of a required microenvironment for an ideal material, which supports 3D cell culture. Its structure and interconnected pores lead to animal cells adhesion and proliferation, also allowing oxygen and nutrients transportation. METHODS: We developed a new methodology to produce spherical platforms synthesized by Komagataebacter hansenii (ATCC 23769) under dynamic culture conditions in minimal medium. The chemical composition and physical properties of the platforms were evaluated. Then, human melanoma cells (SK-MEL-28) were encapsulated into the platforms and evaluated by metabolic activity, morphology and their ability on adhering to the Hollow Translucid BNC Spheres (BNC-TS-H) and Compartmentalized Translucid BNC Spheres (BNC-TS-C) up to 3 days. RESULTS: BNC-TS-H and BNC-TS-C platforms were produced as translucid spheroid platforms with distinct microenvironment under dynamic fermentation. The chemical and physical characterizations confirmed the platforms composition as BNC. The produced internal microenvironments in spherical platforms are relevant to determine tumor cell fate. In the first 12 h of culture, cells could adhere to nanocellulose microfibers assuming their typical tumorous phenotype in 72 h of culture. CONCLUSION: The dynamic fermentation in minimal medium produced distinct microstructured platforms of BNC-TS-H and BNC-TS-C. The platforms microstructure resulted in microenvironments that enabled distinct cell-cell and cell-matrix interactions. This behavior suggests several applications in tissue engineering. GENERAL SIGNIFICANCE: The method produced translucid BNC sphere platforms with distinct microenvironments for 3D cell culture.

Confocal Laser Microscopy for VM Analysis with DAPI and Phalloidin Staining.

Confocal laser scanning microscopy (CLSM) is one of the most prevalent fluorescence microscopy techniques for assessing the progression of cancer cells in three-dimensional structures, such as vasculogenic mimicry (VM). We show a basic approach for using DAPI and phalloidin dyes to detect the early stages of progression and VM of melanoma tumor cells grown in a 3D environment, as well as demonstrating how to acquire images and improve them by changing the software acquisition parameters.

New perspectives for banana peel polysaccharides and their conversion to oligosaccharides.

Banana peel is a source of polysaccharides: pectin, hemicellulose and cellulose. Recent studies have shown that these carbohydrate fractions can be converted into oligomers, which have applications in food, feed and pharmaceuticals, claiming important technical, functional and biological activities. Potential prebiotic activity of pectin and cellulose oligosaccharides obtained from banana peel was already reported. Based on technologies developed for fractionation and extraction of polysaccharides, such as pectin, hemicellulose and cellulose, banana peel can be explored to obtain functional oligosaccharides.

Laccase as an efficacious approach to remove anticancer drugs: A study of doxorubicin degradation, kinetic parameters, and toxicity assessment.

The degradation of an anticancer drug by laccase was investigated for the first time, bringing a new approach to treat these hazardous substances through the direct enzymatic application. Degradations of doxorubicin by laccase were performed in different enzymatic concentrations, pH values and temperatures through kinetic studies. The highest enzymatic degradation of doxorubicin was achieved at pH 7 and 30 ºC, which resembles effluent characteristics from wastewater treatment plants. Assays were carried out in different doxorubicin concentrations to comprehend the enzymatic kinetics of degradation. Michaelis-Menten kinetic parameters obtained were maximum velocity obtained (Vmax) of 702.8 µgDOX h-1 L-1 and Michaelis-Menten constant (KM) of 4.05 µM, which showed a good affinity for the substrate. The toxicity was evaluated against L-929 cell line, and the degraded doxorubicin solution did not show a reduction in cell viability in the concentration of 250 µg L-1. In contrast, the doxorubicin shows a reduction of 27% in cell viability. Furthermore, in the highest tested concentration (1000 µg L-1), enzymatic degradation reduced in up 41.4% the toxicity of doxorubicin, which indicates laccase degrades doxorubicin to non-toxic compounds. In conclusion, this study provides a new application to laccase since the results showed great potential to remove anticancer drugs from effluents.

In vitro evaluation of bilayer membranes of PLGA/hydroxyapatite/ß-tricalcium phosphate for guided bone regeneration.

Membranes for guided bone regeneration represent valuable resources, preventing fibroblast infiltration and aiding anatomical bone reconstruction. Nonetheless, available membranes lack bone regenerative capacity, suitable mechanical behavior, or adequate degradation profile. Therefore, to overcome these limitations, this study developed bilayer membranes with a dense layer (dry phase inversion) of PLGA (poly(lactic-co-glycolic acid)):HAp (hydroxyapatite) - 95:05 (wt%) - and an electrospun layer of PLGA and HAp:ß-TCP (ß-tricalcium phosphate) with ratios of 60:40, 70:30 and 85:15 (wt%), evaluating its mechanical, morphological and in vitro properties. The bilayer membranes displayed adequate interlayer adhesion, dense layer pore size of 4.20 µm and electrospun layer with porosity degree of 38.2%, thus capable of preventing fibroblast infiltration while allowing osteoblast migration and nutrient permeation. They also showed Tg of 82 °C and higher storage modulus, which was constant up to 54.6 °C, characteristics important for membrane implantation and use with no mechanical compromise. In vitro degradation mass loss was only 10% after 60 days, a profile suitable for the application requirement. Membranes with calcium phosphates had better osteoblast attachment, proliferation and migration. Taken together, results indicate the great potential of PLGA/HAp/ß-TCP bilayer membranes on bone reconstruction with proper degradation profile, morphology, mechanical behavior and bone regenerative capacity.