Fumaric acid production by Rhizopus species from acid hydrolysate of oil palm empty fruit bunches.

The hemicellulosic fraction of lignocellulosic biomass is a very important material, due to the significant concentration of pentoses present in its composition and that can be used sustainably in biotechnological processes such as the production of fumaric acid. Research efforts are currently being promoted for the proper disposal and valorization of empty fruit bunches (EFB) from oil palm. In this work, seventeen Rhizopus species were evaluated in a fermentation medium with EFB hydrolyzate, without detoxification, as a carbon source for fumaric acid production. Rhizopus circicans 1475 and Rhizopus 3271 achieved productions of 5.65 g.L-1 and 5.25 g.L-1 of fumaric acid at 30 °C, 120 rpm for 96 h, respectively. The percentage of consumed sugars, mainly pentoses, was 24.88% and 34.02% for R. circicans 1475 and R 3271, respectively. Soy peptone and ammonium sulfate were evaluated as nitrogen sources, where soy peptone stimulated the formation of biomass pellets while ammonium sulfate produced mycelia and clamps.

A non-waste strategy for enzymatic hydrolysis of cellulose recovered from domestic wastewater.

Cellulose is a potential resource to be recovered from wastewater treatment plants (WWTP). Enzyme formulations can be employed to hydrolyze cellulose into fermentable sugars, to be further used as biochemical building blocks or reducing its recalcitrance to further treatment processes. This study proposed the production, recovery and formulation of cellulase using domestic wastewater as culture medium and its application for the hydrolysis of cellulosic residues recovered from WWTPs. Cellulose was recovered from raw sanitary wastewater using a fine-mesh sieve (0.35 mm) and quantified through enzymatic hydrolysis and thermogravimetric analysis. The production, concentration and formulation of cellulase enzyme resulted in an enzymatic blend of endoglucanases (7.3 UFP/mL), cellobiohydrolases (7.4 UCMC/mL) and beta-glucosidases (4.4 UBGL/mL). The content of the recovered cellulosic material was 21.3% according to enzymatic hydrolysis and 27.7 for thermogravimetric results. The enzymatic hydrolysis of the WWTP residue using the produced cellulase (107.6 ± 10.2 mgreduc/gresidue) showed better results than using the commercial cellulase complex (66.4 ± 2.5 mgreduc/gresidue). This fact showed the potential of application of the produced enzyme for the hydrolysis of cellulosic residues recovered from WWTP processes. In a non-waste biorefinery approach, the generated hydrolysate can be further used for producing added-value biomolecules including biofuels and biochemicals.

In vitro cytotoxic effect of a chitin-like polysaccharide produced by Mortierella alpina on adrenocortical carcinoma cells H295R, and its use as mitotane adjuvant.

This study presents an in vitro evaluation of the antitumor potential of a chitin-like exopolysaccharide (EPS, produced by Mortierella alpina) on Adrenocortical carcinoma cells (ACC) compared to mitotane, a commercial drug commonly used in ACC treatment, and known for its side effects. Techniques of cellular viability determination such as MTT and fluorescence were used to measure the cytotoxic effects of the EPS and mitotane in tumoral cells (H295R) and non-tumoral cells (VERO), observing high cytotoxicity of mitotane and a 10% superior pro-apoptotic effect of the EPS compared to mitotane (p < 0.05). The cytotoxic effect of the EPS was similar to the effect of 50 µM mitotane on tumoral cells (p < 0.05). A decrement of the lysosomal volume was also noted in tumoral cells treated with the EPS. To enhance the antitumor effect, a combination of mitotane at a lower dosage and the EPS (as adjuvant) was also tested, showing a slight improvement of the cytotoxicity effect on tumoral cells. Therefore, the results indicate a cytotoxic effect of the EPS produced by Mortierella alpina on adrenocortical carcinoma, and a possible application in biomedical formulations or additional treatments.

Lignin from oil palm empty fruit bunches: Characterization, biological activities and application in green synthesis of silver nanoparticles.

Lignin was extracted from oil palm empty fruit bunches under four different conditions. The lignin samples were characterized and employed in the green synthesis of silver nanoparticles. Two-dimensional HSQC NMR analysis showed that lignins extracted under more aggressive conditions (3.5% acid, 60 min) exhibited less signals and thus, presented a more degraded chemical structure. Additionally, those lignins obtained under harsh conditions (3.5% acid, 60 min) exhibited higher antioxidant capacity than those obtained under mild conditions (1.5% acid, 20 min). Formation of lignin-mediated silver nanoparticles was confirmed by color change during their synthesis. The surface plasmon resonance peaks (423-427 nm) in UV-visible spectra also confirmed the synthesis of AgNPs. AgNPs showed spherical shape, polycrystalline nature and average size between 18 and 20 nm. AgNPs, in suspension, presented a negative Zeta potential profile. Lignin was assumed to contribute in the antioxidant capacity exhibited by AgNPs. All AgNPs presented no significant differences on the disk diffusion antimicrobial susceptibility test against E. coli. The minimum inhibitory concentration of HAL3-L AgNPs (62.5 µg·mL-1) was better than other physicochemically produced AgNPs (100 µg·mL-1).

Influence of organic solvents in the extraction and purification of torularhodin from Sporobolomyces ruberrimus.

OBJECTIVE: This work aimed at evaluating the influence of organic solvents and stationary phases in the extraction with glass beads and chromatographic purification of carotenoids, especially torularhodin, from Sporobolomyces ruberrimus. RESULTS: The combinations of acetone:hexane (1:1 v/v) and acetone:ethyl ether (1:1 v/v) yielded 171.74 and 172.19 µg of total carotenoids.g of cells-1, respectively. The first blend resulted in the highest percent of cell lysis of 57.4%. Among different proportions of acetone:hexane, the 9:1 v/v mixture showed a significant difference (p < 0.05), resulting in a recovery of total carotenoids of 221.88 µg.g of cells-1. The purification of carotenoids was made by preparative chromatography and the yield of the silica-containing stationary phase was higher (24 µg torularhodin.g cells-1). The analyses of the purified fractions in thin layer chromatography and high performance liquid chromatography indicated that the purification of carotenoids, especially of torularhodin, was successfully performed. CONCLUSIONS: The combination of polar (acetone) and non-polar solvents (hexane) and the use of silica as stationary phase was efficient to recover and purify torularhodin from the intracellular pigments of Sporobolomyces ruberrimus.

Effect of sequential acid-alkaline treatment on physical and chemical characteristics of lignin and cellulose from pine (Pinus spp.) residual sawdust.

Timber industry generates large amounts of residues such as sawdust. Softwoods have a significant economic value for timber production and the Pinus genus is widely utilized. Thus, the aim of this work was to study the hemicellulose extraction and lignin recovery from pine (Pinus spp.) residual sawdust (PRS) by sequential acid-alkaline treatment, generating a cellulose-rich solid fraction. The hemicellulose removed was 87.11% (wt·wt-1) after dilute acid treatment at 130 °C, 4.5% (wt·wt-1) of H2SO4 for 20 min at 120 rpm. Three temperatures were evaluated for recovering the lignin and the highest yield, 93.97% (wt·wt-1), was achieved at 170 °C, 10% (wt·wt-1) of NaOH for 90 min at 120 rpm. Lignin was characterized by Fourier-transform infrared spectroscopy, nuclear magnetic resonance and thermogravimetry. The resulting cellulose-rich fraction exhibited polymorphic transformation. The results demonstrated that PRS is a promising lignocellulosic residue whose lignin and carbohydrates can be readily obtained.

Alternative methods for gibberellic acid production, recovery and formulation: A case study for product cost reduction.

The aim of this work was to develop a new production, recovery and formulation process of gibberellic acid (GA3). Low-cost byproducts - citrus pulp (CP) and soybean hulls (SH) - were employed as substrate for GA3 production by Gibberella fujikuroi in semisolid fermentation. A CP/SH mixture (70%/30%) promoted high productivities both in bubble column reactor (1.66 mg L/h), and in stirred tank reactor (2.13 mg L/h). GA3 production medium cost (US$ 6.70/m3) was reduced by 85% when compared to previously reported synthetic media (US$ 44.96/m3). It was described that GA3 fermented extract has low stability, and that liquid and powder formulation of the fermented extract maintained the biomolecule activity over 6 months. Alginate and alginate/kefiran beads containing GA3 showed encapsulation efficiency of 70% and 60%, respectively. This work supports good perspectives for GA3 production using cheap substrates and simple formulation of clarified extract to favour its use in agricultural countries.

Phytochemical analysis and biological activities of in vitro cultured Nidularium procerum, a bromeliad vulnerable to extinction.

This study reports the first phytochemical and biological characterization in treatment of adrenocortical carcinoma cells (H295R) of extracts from Nidularium procerum, an endemic bromeliad of Atlantic Forest vulnerable to extinction. Extracts of dry leaves obtained from in vitro-grown plants were recovered by different extraction methods, viz., hexanoic, ethanolic, and hot and cold aqueous. Chromatography-based metabolite profiling and chemical reaction methods revealed the presence of flavonoids, steroids, lipids, vitamins, among other antioxidant and antitumor biomolecules. Eicosanoic and tricosanoic acids, α-Tocopherol (vitamin E) and scutellarein were, for the first time, described in the Nidularium group. Ethanolic and aqueous extracts contained the highest phenolic content (107.3 mg of GAE.100 g-1) and 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) radical scavenging activity, respectively. The immunomodulatory and antitumoral activities of aqueous extracts were assessed using specific tests of murine macrophages modulation (RAW 264.7) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay against adrenocortical carcinoma cell line, respectively. The aqueous extract improved cell adhesion and phagocytic activities and phagolysossomal formation of murine macrophages. This constitutes new data on the Bromeliaceae family, which should be better exploited to the production of new phytomedicines for pharmacological uses.

A comparative study of extraction techniques for maximum recovery of bioactive compounds from Ganoderma lucidum spores / Estudio comparativo de técnicas de extracción para la máxima recuperación de compuestos bioactivos a partir esporas de Ganoderma lucidum

SUMMARY This study aimed at evaluating effective methods for breaking the hard and insoluble spores of Ganoderma lucidum to recover functional biomolecules. Rupture techniques were evaluated such as manual maceration (RM), maceration with spheres of various materials (BR), and microwave exposure plus maceration with steel/ chrome spheres (MBR1). Spore rupture was evaluated using UV-Vis spectroscopy, which showed vibrations of 2955, 1642, 1240, 1080 and 1746 cm-1 corresponding to changes in spore walls. The MBR1 extract contained the largest amounts of carbohydrates (19.80 mg.g-1 spores) and polyphenols (2.21 mg.g-1 spores), whereas the BR extract had higher antioxidant activity (57.22%Inb DPPH). The MBR1 and BR extracts contained 62.2 and 73.5% glucose, respectively. Both methods also involved significant extraction of carbohydrates and proteins. The best way to extract biomolecules from spore walls is to perform a microwave heat treatment and break the walls with steel/chrome spheres; this produces large quantities of carbohydrates with antioxidant properties.

RESUMEN El objetivo de este estudio fue evaluar varios métodos de ruptura de las esporas de Ganoderma lucidum y extraer sus propiedades bioactivas. Para este propósito se evaluaron diferentes técnicas de rompimiento como: la maceración manual (RM), la maceración con esferas de diversos materiales (BR) y la exposición a microondas junto la maceración de las esporas con esferas de acero/cromo (MBR1). La ruptura de las esporas fue evaluada por espectroscopia UV-Vis, la cual mostró que las vibraciones 2955, 1642, 1240, 1080 y 1746 cm-1 correspondieron a cambios estructurales en las paredes de las esporas. El extracto MBR1 presento el mayor contenido de carbohidratos (19,80 mg.g-1) y polifenoles (2,21 mg.g-1), mientras que el extracto BR tuvo una mayor actividad antioxidante (57,22% Inb DPPH). Los extractos MBR1 y BR también presentaron en el análisis de monosacáridos un 62,2 y 73,5% de contenido glucosa. Como conclusión la mejor metodología para extraer biomoléculas de las paredes de las esporas de G. lucidum fueron el tratamiento térmico con microondas y la ruptura de las paredes con esferas de acero/cromo, porque este proceso permitió la extracción de una mayor cantidad de carbohidratos con posibles propiedades antioxidantes.

Feasibility of ramie fibers as raw material for the isolation of nanofibrillated cellulose.

In this study, a strategy was adopted to enhance the use of ramie fibers as raw material for isolation of cellulose nanofibers (CNFs). Ramie pulp was produced by alkaline organosolv followed by bleaching. CNFs were produced by mechanical defibrillation, and films were fabricated via casting. Effects of number of passes in the mechanical grinding on physical and mechanical properties of CNF films were comprehensively studied. Potential of ramie fibers was proved by fabricating homogeneous nanofibers with average thickness of 8.72 nm, which led to CNF films with dense and non-porous networks, and crystallinity index of 76-78%. Tensile strength (42-82 MPa) and dynamic mechanical (9-11 GPa) performance were good only for less severe mechanical defibrillation. Lower solubility (1.85-2.43%). and activity (0.69) in water, and outstanding barrier properties against water vapor and oxygen make ramie suitable for more sustainable extraction of cellulose nanofibers and production of CNF films for diverse applications.

Definition of Liquid and Powder Cellulase Formulations Using Domestic Wastewater in Bubble Column Reactor.

Raw domestic wastewater was used as a culture medium for cellulase production in a bubble column reactor (6.2 UFP/mL, 64.6 U/L h) using the strain Trichoderma harzianum TRIC03-LPBII. Cellulases presented optimum pH and temperature between 4 and 5 and 50 and 70 °C, respectively. Enzymatic extract was concentrated through ultrafiltration and then a cellulolytic formulation was prepared with the addition of sorbitol (50% w/v) and benzoic acid (0.05% w/v). High cellulase stability of around 100% was reached after 30 days at 4 °C. The concentrated extract was also dried in a spray-dryer with the addition of maltodextrin at 20% (w/v), resulting in powder enzymatic formulation with 85% stability after 60 days. With these characteristics, the liquid and powder cellulase products have potential to be used in different industrial applications.