RESUMEN
Chitin is a structural polysaccharide abundant in the biosphere. Chitin possesses a highly ordered crystalline structure that makes its processing a challenge. In this study, chitin hydrogels and methanogels, prepared by dissolution in calcium chloride/methanol, were subjected to supercritical carbon dioxide (scCO2) to produce porous materials for use as scaffolds for osteoblasts. The control of the morphology, porosity, and physicochemical properties of the produced materials was performed according to the operational conditions, as well as the co-solvent addition. The dissolution of CO2 in methanol co-solvent improved the sorption of the compressed fluid into the hydrogel, rendering highly porous chitin scaffolds. The chitin crystallinity index significantly decreased after processing the hydrogel in supercritical conditions, with a significant effect on its swelling capacity. The use of scCO2 with methanol co-solvent resulted in chitin scaffolds with characteristics adequate to the adhesion and proliferation of osteoblasts.
RESUMEN
BACKGROUND/AIM: Enzyme-mediated grafting of poly (gallic acid) (PGAL) and L-arginine and a-L-lysine onto PGAL produces reactive oxygen species (ROS)-suppressor multiradical molecules with low cytotoxicity, high thermostability and water solubility with cancer treatment potential. This study examined the anticancer effects of these molecules in hepatic (HepG2, ATCC HB-8065), breast (MCF7, ATCC HTB-22), and prostate (PC-3, ATCC CRL-1435 and DU 145, ATCC HTB-81) cancer cell lines, as well as in fibroblasts from healthy human skin as control cells. MATERIALS AND METHODS: PGAL was synthesized by the oxidative polymerization of the naturally abundant GA using laccase from Trametes versicolor. Insertions of amino acids L-arginine and α-L-lysine on the PGAL chain were carried out by microwave. The cells of dermal fibroblast (Fb) were obtained from primary skin cultures and isolated from skin biopsies. The cancer cells lines of hepatic (HepG2), breast (MCF7), and prostate (PC-3, DU 145) were obtained from ATCC. The viability of the cancer cells and the primary culture was obtained by the MTT assay. Proliferation was demonstrated by crystal violet assay. Cell migration was determined by Wound healing assay. Finally, cell cycle analysis was carried out with cells. RESULTS: The results show that 200 µg/ml of PGAL cultured in vitro with prostate cancer cells decreased viability, proliferation, and migration, as well as arrested cells in the G1 and S phases of the cell cycle. In contrast, the dermal fibroblasts and the hepatic line remained unaffected. The random grafting of L-Arg and a-L-Lys onto the PGAL chain also decreased the viability of prostate cancer cells. CONCLUSION: PGAL and PGAL-grafted amino acids are potential adjuvants for prostate cancer treatment, with improved physicochemical characteristics compared to GA.
Asunto(s)
Ácido Gálico , Neoplasias de la Próstata , Salicilatos , Masculino , Humanos , Ácido Gálico/farmacología , Lisina , Trametes , Neoplasias de la Próstata/patología , Células MCF-7 , Arginina/farmacología , Proliferación CelularRESUMEN
Polygallic acid (PGAL) has been used in vitro to protect synoviocytes from monosodium urate (MSU) crystals due to its anti-inflammatory properties. However, MSU crystals can also activate other cells of the synovial fluid (SF). We studied the impact of PGAL on the phagocytosis of MSU crystals, inflammation, and oxidative stress using an in vitro model with SF leukocytes and THP-1 monocyte cells. SF leukocytes were stimulated with PGAL and MSU crystals, proinflammatory cytokines and phagocytosis were assessed. In THP-1 cells, the effect of PGAL on the phagocytosis of MSU crystals and the levels of IL-1ß, IL-6, TNF-α, and reactive oxygen species (ROS) was evaluated. PGAL was added to THP-1 cultures 24 h before MSU crystal addition as a pre-treatment, and IL-1ß was measured. One-way ANOVA with Tukey's post hoc test was performed, and a P value < 0.05 was considered statistically significant. PGAL (100 µg/mL) decreased phagocytosis in SF leukocytes by 14% compared to cells exposed to crystals without PGAL. In THP-1 cells, 100 and 200 µg/mL PGAL reduced phagocytosis by 17% and 15%, respectively. In SF cells, there was a tendency to decrease IL-1ß and IL-6. In THP-1 cells, decreases in IL-1ß and TNF-α, as well as a slight decrease in ROS, were identified. PGAL pre-treatment resulted in a reduction of IL-1ß. PGAL inhibits MSU phagocytosis by exerting an anti-inflammatory effect on cells exposed to crystals. The use of PGAL before an acute attack of gout suggests an important protective factor to control the inflammation.
Asunto(s)
Gota , Factor de Necrosis Tumoral alfa , Humanos , Especies Reactivas de Oxígeno , Interleucina-6 , Ácido Úrico/farmacología , Inflamación , AntiinflamatoriosRESUMEN
This study aimed to detect intracellular trehalose in boar sperm that were cryopreserved with liposomes and conduct an analysis of its effects on some characteristics of thawed sperm, including rheological properties. First, soybean lecithin cholesterol-based liposomes were produced and characterized in the presence of 300 mM trehalose. Next, semen samples were frozen in two freezing media: a control medium with 300 mM trehalose and an experimental medium supplemented with 300 mM trehalose and 10% liposomes, both of which were thawed and then studied to ascertain their integrity, motility, rheological response, and trehalose quantities by testing two methods of spermatic lysis via high-performance liquid chromatography with an evaporative light-scattering detector (HPLC-ELSD). The results found spherical liposomes measuring 357 nm that were relatively stable in an aqueous medium and had an entrapment efficiency of 73%. An analysis of the cryopreserved ejaculates showed that their viability and motility did not significantly differ between groups (P > 0.05). The viscous response of the samples was influenced by the extracellular medium rather than by the freezing-thawing process, which resulted in a loss of interaction between the cells and cryoprotectants. Finally, intracellular trehalose levels were determined using HPLC-ELSD, with no differences observed (P > 0.05) when comparing both sperm lysis methods. The use of liposomes with trehalose appears to be a promising option for boar semen cryopreservation, with a marked effect on rheological properties. The proposed HPLC-ELSD method was effective for measuring trehalose in cryopreserved cell samples.
Asunto(s)
Preservación de Semen , Semen , Masculino , Porcinos , Animales , Semen/fisiología , Trehalosa , Preservación de Semen/veterinaria , Preservación de Semen/métodos , Liposomas , Motilidad Espermática/fisiología , Disacáridos , Criopreservación/veterinaria , Criopreservación/métodos , Crioprotectores/farmacología , Espermatozoides/fisiologíaRESUMEN
The α-l-Lysine (LL) grafting onto the enzymatic poly(gallic acid) (PGAL) produces a helicoidal brush-like antimicrobial polymer containing outer positive-charged moieties. Best results are found with ca. 16 mol% α-LL-grafting for the inhibition of gram-positive Staphylococcus aureus and gram-negative Escherichia coli strains. Membrane permeability, confocal and scanning electron microscopy studies suggest a pore-formation and translocation mechanisms by initial electrostatic interaction of positive charged polymer at the negatively charged bacterial membranes. The attained polymer displays high concentration of hemolysis (Hc) in erythrocytes, and no lymphocyte mitochondrial activity. Interestingly, PGAL-LL is not cytotoxic on human dermal fibroblast. The antioxidant activity after the LL hybridization is also demonstrated by DPPH, ORAC, FRAP and hydroxyl radical scavenging, which enhances the preservation of human cells in addition to antimicrobial for this polymer.
Asunto(s)
Infecciones por Escherichia coli , Infecciones Estafilocócicas , Antibacterianos/farmacología , Escherichia coli , Ácido Gálico , Humanos , Lisina , Polímeros , Staphylococcus aureusRESUMEN
Microwave-mediated grafting of L-Arg onto naturally derived and stable multiradical poly(gallic acid) (PGAL) in aqueous media has been successfully achieved. This polymeric material has no adverse effect in human cells as there is no hemolytic activity upon MTT and Neutral Red assays. The analytical and computational characterization studies carried out in this study describe a helical molecular structure with random incorporation of L-Arginine pendant groups from PGAL's backbone. The antioxidant properties of the precursor polymer are preserved as proved by the elimination of stable DPPH and hydroxyl radical scavenging, as well as the FRAP and ORAC assays. Regarding the latter, the oxygen radical inhibition is enhanced compared to PGAL, which is attributed to the guanidyl moieties. PGAL-g-L-Arg displays antimicrobial activity against Gram (+) Listeria monocytogenes and Staphylococcus aureus strains with a MIC of 0.8â¯g/L and a bacteriostatic effect against Gram (-) Escherichia coli. Additionally, scanning electron and confocal fluorescence microscopies as well as crystal violet colorimetric assay demonstrate that the mechanism involved in the bacterial inhibition is related to the formation of porous channels on the membrane, which is discussed according to the helical secondary structure of the polymer and the amino acid guanidyl moieties interacting to bacterial membranes.
Asunto(s)
Antioxidantes , Ácido Gálico , Antibacterianos/farmacología , Antioxidantes/farmacología , Arginina , Ácido Gálico/farmacología , Humanos , Staphylococcus aureusRESUMEN
Enzymatic mediated poly (gallic acid) (PGAL), a stable multiradical polyanion with helicoidal secondary structure and high antioxidant capacity, was successfully grafted to poly(ε-caprolactone) (PCL) using UV-photo induction. PCL films were prepared with several levels of roughness and subsequently grafted with PGAL (PCL-g-PGAL). The results on the full characterization of the produced materials by mechanical tests, surface morphology, and topography, thermal and crystallographic analyses, as well as wettability and cell protection activity against oxidative stress, were adequate for tissue regeneration. The in vitro biocompatibility was then assessed with epithelial-like cells showing excellent adhesion and proliferation onto the PCL-g-PGAL films, most importantly, PCL-g-PGAL displayed a good ability to protect cell cultures on their surface against reactive oxygen species. These biomaterials can consequently be considered as novel biocompatible and antioxidant films with high-responsiveness for biomedical or tissue engineering applications.
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Células Epiteliales/citología , Ácido Gálico/farmacología , Estrés Oxidativo/efectos de los fármacos , Poliésteres/química , Animales , Adhesión Celular , Técnicas de Cultivo de Célula , Células Cultivadas , Chlorocebus aethiops , Perros , Células Epiteliales/efectos de los fármacos , Ácido Gálico/química , Células de Riñón Canino Madin Darby , Ensayo de Materiales , Células Vero , HumectabilidadRESUMEN
The bioconversion process of bioactive naringenin by whole-cells of Yarrowia lipolytica 2.2ab for the production of increased value-added compounds is successfully achieved in surface and liquid cultures. This approach is an alternative to the commercial production of these bioactive compounds from vegetable sources, which are limited due to their low concentrations and the complexity of the purification processes. The experimentation rendered seven value-added compounds in both surface and liquid bioconversion cultures. Some of the compounds produced have not been previously reported as products from the bioconversion processes, such as the case of ampelopsin. Biosynthetic pathways were suggested for the naringenin bioconversion using whole-cells of Y. lipolytica 2.2ab. Finally, the extracts obtained from the naringenin bioconversion in liquid cultures showed higher percentage of inhibition of DPPH· and ABTS· radicals up to 32.88 and 2.08 times, respectively, compared to commercial naringenin.
Asunto(s)
Flavanonas/metabolismo , Yarrowia/metabolismo , Antioxidantes/farmacología , Reactores Biológicos , Cromatografía Liquida/métodos , Medios de Cultivo , Fermentación , Flavanonas/farmacología , Flavonoides/biosíntesis , Hidroxilación , Espectrometría de Masas en Tándem/métodosRESUMEN
This work demonstrates that a biodegradable chitosan-based biocomposite packed in mini-reactors successfully removes copper ions from aqueous solutions. The chitosan is obtained by deacetylation of biological chitin, which is extracted from shrimp wastes by lactic acid fermentation. The polysaccharide is embedded in a biodegradable prepolymer matrix before extrusion to produce porous cylindrical pellets of 2 × 80 mm. The highest copper ion removal is 62.5 mg Cu2+ per g of the biodegradable adsorbent. Additionally, the adsorption capacity of the material, below its saturation, allows several cycles of reuse with a hydraulic retention time reduction of 1 h. This chitosan-based material is advantageous when compared with other approaches using non-biodegradable materials or costly commercial adsorbents for removing heavy metal ions in wastewater effluents as well as a filter component in water purification devices.
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Quitosano , Contaminantes Químicos del Agua , Purificación del Agua , Adsorción , Cobre , Concentración de Iones de Hidrógeno , Cinética , Aguas ResidualesRESUMEN
Yellowfin tuna by-products (Thunnus albacares) were processed to produce radical-scavenging peptides from hydrolysis by lactic acid fermentation (LAF) with Lactobacillus plantarum, papaya fruit (Carica papaya), and molasses as a carbon source for 72 h. A 15-kDa peptide was purified; after de novo sequencing, it was determined that fragments are rich in hydrophobic and neutral amino acids. The results suggest this effect is mainly to the hydrophobicity of the amino acids in their sequence. Further work is on progress to assess the ability of peptides to provide stability in lipids or in other types of samples sensitive to the action of free radicals.
Asunto(s)
Depuradores de Radicales Libres , Ácido Láctico/metabolismo , Péptidos , Análisis de Secuencia de Proteína , Atún , Animales , Carica/química , Depuradores de Radicales Libres/química , Depuradores de Radicales Libres/aislamiento & purificación , Frutas , Lactobacillus plantarum/crecimiento & desarrollo , Melaza , Péptidos/química , Péptidos/genética , Péptidos/aislamiento & purificaciónRESUMEN
In the present work, cell lines of different origin were exposed to BPA levels from food intake reported elsewhere. Specifically, we used an in vitro assay to determine cytotoxicity of BPA in three cell lines: MCF7 (breast cancer), PC3 (prostate cancer) and 3T3-L1 (mouse fibroblast). Cytotoxic effects were observed at concentrations higher than 50 µg/mL which is above the involuntary exposure level of BPA described before in fresh, canned and frozen foods and beverages. Furthermore, medial inhibitory concentrations (IC50) of 85.17 µg/mL and 88.48 µg/mL were observed for PC3 and 3T3-L1, respectively, and a slightly lower IC50 of 64.67 µg/mL for MCF7. These results highlight BPA's toxicity potential at current levels from food intake. The cell line-dependent divergent response to BPA reported herein is discussed.
Asunto(s)
Compuestos de Bencidrilo/efectos adversos , Compuestos de Bencidrilo/toxicidad , Línea Celular/efectos de los fármacos , Fenoles/efectos adversos , Fenoles/toxicidad , Células 3T3-L1/efectos de los fármacos , Animales , Contaminación de Alimentos , Humanos , Concentración 50 Inhibidora , Células MCF-7/efectos de los fármacos , Ratones , Células PC-3/efectos de los fármacosRESUMEN
Acinetobacter species are identified as producing surface-active and emulsifying molecules known as bioemulsifiers. Production, characterization and stability of bioemulsifiers produced by Acinetobacter bouvetii UAM25 were studied. A. bouvetii UAM25 grew in three different carbon and energy sources: ethanol, a glycerol-hexadecane mixture and waste cooking oil in an airlift bioreactor, showing that bioemulsifier production was growth associated. The three purified bioemulsifiers were lipo-heteropolysaccharides of high molecular weight (4866 ± 533 and 462 ± 101 kDa). The best carbon source and energy for bioemulsifier production was wasted cooking oil, with a highest emulsifying capacity (76.2 ± 3.5 EU mg-1) as compared with ethanol (46.6 ± 7.1 EU mg-1) and the glycerol-hexadecane mixture (49.5 ± 4.2 EU mg-1). The three bioemulsifiers in our study displayed similar macromolecular structures, regardless of the nature (hydrophobic or hydrophilic) of the carbon and energy source. Bioemulsifiers did not decrease surface tension, but the emulsifying capacity of all of them was retained under extreme variation in salinity (0-50 g NaCl L-1), pH (3-10) and temperature (25-121 °C), indicative of remarkable stability. These findings contribute to understanding of the relationship between: production, physical properties, chemical composition and stability of bioemulsifiers for their potential applications in biotechnology, such as bioremediation of hydrocarbon-contaminated soil and water.
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Acinetobacter/crecimiento & desarrollo , Alcanos/farmacología , Medios de Cultivo/farmacología , Emulsionantes/metabolismo , Etanol/farmacología , Glicerol/farmacología , Alcanos/química , Medios de Cultivo/química , Etanol/química , Glicerol/químicaRESUMEN
A nano-composite from biologically obtained chitin nanofillers homogenously dispersed in a poly(ε-caprolactone) matrix was successfully achieved by an ultrasonication-assisted non-toxic and non-aqueous methodology. For this purpose, biological chitin was obtained from lactic acid fermentation of shrimp wastes and converted into chitin whiskers by acidic hydrolysis in a novel process at low temperature (4°C) that enhanced the distribution and yield. Additionally, the polyester matrix was enzymatically produced in a non-toxic compressed fluid (1,1,1,2-tetrafluoroethane at 25bar and 65°C) medium. The homogeneous distribution of the nanofiller in the matrix was corroborated by confocal and atomic force microscopies. Films of the nanocomposite were physicochemically characterized to assess its adequate properties. Additionally, the qualitative viability of human fibroblasts and osteoblasts cells was studied on the produced nanocomposite films showing good biocompatibility.
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Quitina/química , Nanocompuestos/química , Nanopartículas/química , Poliésteres/química , Adulto , Animales , Candida/enzimología , Niño , Quitina/aislamiento & purificación , Fibroblastos , Tecnología Química Verde , Humanos , Hidrocarburos Fluorados/química , Hidrólisis , Lactobacillus plantarum/química , Lipasa/química , Osteoblastos , Tamaño de la Partícula , Penaeidae/químicaRESUMEN
The extraction of calcareous chitin from shrimp cephalothorax was successfully achieved using a subcritical water treatment to attain a deproteinization up to 96%. The treatments also increased the crystalline domain size in the α-chitin fibers. An experimental design of Taguchi allowed the optimization of experiments. The macroelements identified in all samples were Ca, P, S, K, Cl and Al, whereas Cr, Mn, Fe, Ni, Cu, Zn, Br and Sr were also detected as microelements. The assigned crystalline phases by XRD were α-chitin, calcite, HAP and traces of quartz. The presence of these phases was corroborated by ATR-FTIR and SEM-EDS analyses. The highest content of α-chitin (82.2wt%) was obtained for the 0.17 chitin:dH2O (wt/wt) ratio for 30min treatment at 260°C. Noteworthy, this treatment promotes the crystallization of both minerals as microcrystals of calcite and nanocrystals of hydroxyapatite with needle and flake shapes as well as intermediate morphologies.
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Quitina/aislamiento & purificación , Crustáceos , Durapatita/química , Animales , CristalizaciónRESUMEN
The poly(gallic acid), produced by laccase-mediated oxidation of gallic acid in aqueous media (pH5.5) to attain a novel material with well-defined molecular structure and high water solubility (500mg/mL at 25°C), has been investigated to understand its potential biological activities. In this regard, a biomedical approach based on cytoprotective effect on human fibroblast cells exposed to UV-irradiation in the presence of the polymer has been demonstrated. The results also shows that 200µg/mL of poly(gallic acid) inhibits the growth and migration of dermal fibroblasts and cancer cell lines without affecting cell viability. Poly(gallic acid) pretreatment with 10µg/mL protects dermal fibroblasts from UV induced cell death and additionally, the cytoprotective effect reduce ROS presence in the cells. This property can be correlated with the antioxidant power (IC50 of 23.5µg/mL) of this novel material, which was ascertained by electronic paramagnetic resonance spectroscopy and spectrophotometrically. Additionally, the antimicrobial activity of this material was corroborated with the inhibition of Staphylococcus aureus (ATCC 25923) and Enterococcus faecalis (ATCC 29212) strains (MIC=400mg/mL) common bacteria found in hospitals.
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Fibroblastos , Antioxidantes , Ácido Gálico , Humanos , Staphylococcus aureus , Rayos UltravioletaRESUMEN
A chitosan from biologically obtained chitin was successfully grafted with d,l-lactic acid (LA) in aqueous media using p-toluenesulfonic acid as catalyst to obtain a non-toxic, biodegradable packaging material that was characterized using scanning electron microscopy, water vapor permeability, and relative humidity (RH) losses. Additionally, the grafting in chitosan with LA produced films with improved mechanical properties. This material successfully extended the shelf life of fresh cheese and inhibited the growth of Listeria monocytogenes during 14 days at 4 °C and 22% RH, whereby inoculated samples with chitosan-g-LA packaging presented full bacterial inhibition. The results were compared to control samples and commercial low-density polyethylene packaging.
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Queso/microbiología , Quitosano/farmacología , Embalaje de Alimentos , Listeria monocytogenes/efectos de los fármacos , Quitina/química , Quitosano/química , Microbiología de Alimentos , Humanos , Ácido Láctico/química , Ácido Láctico/farmacología , Listeria monocytogenes/patogenicidadRESUMEN
The hydrolysis of chitin treated under supercritical conditions was successfully carried out using chitinases obtained by an optimized fermentation of the fungus Lecanicillium lecanii. The biopolymer was subjected to a pretreatment based on suspension in supercritical 1,1,1,2-tetrafluoroethane (scR134a), which possesses a critical temperature and pressure of 101°C and 40bar, respectively, followed by rapid depressurization to atmospheric pressure and further fibrillation. This methodology was compared to control untreated chitins and chitin subjected to steam explosion showing improved production of reducing sugars (0.18mg/mL), enzymatic hydrolysis and high acetylation (FA of 0.45) in products with degrees of polymerization between 2 and 5.
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Quitina/metabolismo , Quitinasas/química , Hidrocarburos Fluorados/química , Oligosacáridos/química , Acetilación , Quitinasas/aislamiento & purificación , Fermentación , Hidrólisis , Hypocreales/enzimología , Vapor , TemperaturaRESUMEN
The enzyme-mediated grafting of tert-butylhydroquinone (TBHQ) onto chitosan and further crosslinking to agave inulin (agavin) has been successfully achieved in a mild and non-toxic two-step route. The resulting products were characterized by nuclear magnetic resonance (NMR) and Infra-red spectroscopies to assess the molecular structure. The study of acute oral toxicity in mice revealed no adverse short-term effects of consumption in the synthesized materials with non-toxicity evidence until 2000 mg/kg through an oral acute administration. Importantly, this study proves that the compound maintains the radical scavenging capacity of the phenolic antioxidant upon ferric-reducing antioxidant power (FRAP) and oxygen radical absorbance capacity (ORAC) assays with a measured half-maximal inhibitory concentration (IC50) for the best case of 1.54 g/L based on inhibition of 2,2'-azino-bis(3-ethylbenzothiazoline)-6-sulfonic acid diammonium salt (ABTS). Additionally, the novel compound presented high prebiotic activities as ascertained in the presence of lactic acid bacteria (LAB).
Asunto(s)
Antioxidantes/química , Quitosano/química , Hidroquinonas/química , Inulina/química , Prebióticos/análisis , Agave/química , Animales , RatonesRESUMEN
The objective of this work was to evaluate the survival of a Brucella abortus aqpX mutant during the elaboration and conservation of fresh and ripened cheeses at 4 °C and 24 °C. The pH values and water activity were monitored for each type of cheese. The fresh cheese was elaborated with raw milk inoculated with 6×108 colony-forming units (CFU)/mL each of parental and mutant strain. Ripening cheeses were elaborated with both raw and pasteurized milk and inoculated with 12×108 CFU/mL each of parental and mutant strains. In fresh cheese, survival was observed during elaboration and conservation for 7 days at 4 °C in mutant and parental strains. The number of survivors of the mutant strain was 10 times lower compared with the parental strain at pH 5 and a(w) of 0.930. In the cheese elaborated with raw milk and ripened at 24 °C, both strains survived until day 17 at pH 4.0 and a(w) of 0.89. However, when the cheese was elaborated with pasteurized milk, the parental strain survived until day 31 of ripening, and the mutant strain survived 24 days at pH 4 and a(w) of 0.886. The survival of the mutant strain showed a diminution of one logarithm during elaboration and ripening of cheese as compared with the parental strain. When the cheese was elaborated with raw milk and ripened at 4 °C, survival of the parental strain was 24 days, whereas the mutant strain survived only 17 days (pH 5 and a(w) 0.90). Regarding the cheese elaborated with pasteurized milk and maturated at 4 °C, both strains survived 31 days (pH 5 and a(w) 0.90), with the same survival diminution during elaboration and ripening. Our results show that in both types of cheese, the mutated aqpX strain survived 10 times less than the parental strain, which shows that the aqpX gene can be related to the survival of Brucella abortus in this type of cheese.