Enzyme loading dependence of cellulose hydrolysis of sugarcane bagasse

The enzymatic hydrolysis of steam-pretreated sugarcane bagasse, either delignified or non-delignified, was studied as a function of enzyme loading. Hydrolysis experiments were carried out using five enzyme loadings (2.5 to 20 FPU/g cellulose) and the concentration of solids was 2% for both materials. Alkaline delignification improved cellulose hydrolysis by increasing surface area. For both materials, glucose concentrations increased with enzyme loading. On the other hand, enzyme loadings higher than 15 FPU/g did not result in any increase in the initial rate, since the excess of enzyme adsorbed onto the substrate restricted the diffusion process through the structure.

Immunohistochemical detection of Clostridia species in paraffin-embedded tissues of experimentally inoculated guinea pigs

Blackleg is caused by Clostridium chauvoei, whereas malignant oedema is caused by C. chauvoei, C. septicum, C. sordellii, C. perfringens type A, and/or C. novyi type A. Anti-C. chauvoei, anti-C. septicum, anti-C. sordellii and anti-C. novyi type A polyclonal antibodies were produced in rabbits and purified in a column of DEAE-cellulose. Aliquots of the antisera were conjugated with fluorescein isothiocyanate and the remaining was used for the streptavidin biotin peroxidase technique (SBPT). SBPT was standardized to detect C. chauvoei, C. septicum, C. sordellii and C. novyi type A in formalin-fixed, paraffin-embedded tissues of guinea pigs. SBPT was compared to a fluorescent antibody technique (FAT). Sections and smears of muscle from inoculation area (MIA), heart, liver, spleen and kidney, were obtained for both SBPT and FAT. Cross-reactions between the different Clostridial species were not observed. C. chauvoei and C. septicum were detected in all specimens from the animals inoculated with these microorganisms, while only sections of muscle obtained from all the animals inoculated with C. sordellii and C. novyi type A were positive. The same results observed by the SBPT, were obtained on tissue smears of these microorganisms stained by the FAT. The results indicate that SBPT is suitable for detection of C. chauvoei, C. septicum, C. sordellii and C. novyi type A in formalin-fixed, paraffin-embedded tissues of guinea pigs.

Homocysteine induces glyceraldehyde-3-phosphate dehydrogenase acetylation and apoptosis in the neuroblastoma cell line Neuro2a

High plasma levels of homocysteine (Hcy) promote the progression of neurodegenerative diseases. However, the mechanism by which Hcy mediates neurotoxicity has not been elucidated. We observed that upon incubation with Hcy, the viability of a neuroblastoma cell line Neuro2a declined in a dose-dependent manner, and apoptosis was induced within 48 h. The median effective concentration (EC50) of Hcy was approximately 5 mM. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) nuclear translocation and acylation has been implicated in the regulation of apoptosis. We found that nuclear translocation and acetylation of GAPDH increased in the presence of 5 mM Hcy and that higher levels of acetyltransferase p300/CBP were detected in Neuro2a cells. These findings implicate the involvement of GAPDH in the mechanism whereby Hcy induces apoptosis in neurons. This study highlights a potentially important pathway in neurodegenerative disorders, and a novel target pathway for neuroprotective therapy.

Síntesis de espirobrasinoesteroides análogos de la 6-desoxocastasterona

The synthesis of two new spirostanic analogs of the natural occurring brassinosteroid 6-desoxocastasterone (1) is described. The scheme consists in the formation and elimination of tigogenin mesylate followed by catalytic dihydroxylation of the resulting D2-steroid (3) and acetylation of the 2a, 3a-diol introduced.Treatment diacetate (5) with NaNO2/BF3.Et2O and chromatography in alumina led to a 23-keto (6) which on reduction produced the 23S alcohol (8) as major product. Saponification of the 2a, 3a-diacetoxy-23-keto compound (6) and the 2a,3a-diacetoxy-23-hydroxy compound (8) led to the spirobrasinosteroids (7) and (9).13C NMR and ¹H RMN characteristics derived from substitution at C23 are briefly discussed.

Alginato bacteriano: aspectos tecnológicos, características e produção

Alginate is a biopolymer used for a variety of industrial applications, for example, in the textiles, cosmetics, foods, agricultural and biotechnological industries. This biopolymer is traditionally extracted from some brown seaweeds (Phaeophyceae) and can be produced by bacteria isolated from soil, as Azotobacter vinelandii, like capsular polysaccharide using glucose, sucrose, among others as carbon sources. The main difference between the alginate of seaweed and the bacterial ones, is the biggest degree of acetylation of this last one, with great influence in the gel force. These chemical characteristics and production of bacterial alginate are presented in this work.

Purificação e caracterização da quitinase de uva (Vitis vinífera L. cv Red Globe) para a produção de quitosana a partir de quitina de camarão

Chitinase is produced by a wide variety of plants as a defense against peste attacks. In this study, grape chitinases were purified 16 times by fractionation in 80% ammonium sulfate followed by dialysis and filtration. Purified chitinases exhibited enzymatic activity toward chitin azure. The yield of purified chitinase was 229 mg/L with chitinase activity of 563 U/g. Chitinases had molecular masses of 24 and 30 kDa, as evaluated by SDS-PAGE 12.5%. Two pH optima were determined 3.0 and 6.0. The optimal temperature was 42 °C. Pre hydrolysis of crystalline shrimp chitin by chitinases caused in an increase in the deacetylation ratio triggered by chitin deacetylase producing chitooligosaccharides with DA (degree acetylation) of 58.8%.

Derivados citotóxicos de vitanolidos isolados das folhas de Acnistus arborescens

In view of anticancer activity of 7 β-acetoxywithanolide D (2) and 7β-16α-diacetoxywithonide D (3), isolated from the leaves of Acnistus arborescens (Solanaceae), five withanolide derivatives were obtained and their structures were determined by NMR, MS and IV data analysis. The in vitro anticancer activity of these derivatives was evaluated in a panel of cancer cell lines: human breast (BC-1), human lung (Lu1), human colon (Col2) and human oral epidermoid carcinoma (KB). Compounds 2a (acetylation of 2), 3b (oxidation of 3) and 2c (hydrogenation of 2) exhibited the highest anticancer activity against human lung cancer cells, with ED50 values of 0.19, 0.25 and 0.63 μg/mL, respectively.

ESTUDO DA GLICOSILAÇÃO DO CICLO-HEXANOL COM DIFERENTES DERIVADOS DE D-GLICOSAMINA E PROMOTORES DE GLICOSILAÇÃO PELO MÉTODO DE KOENIGS-KNORR

We report herein a study on the glycosylation of cyclohexanol with four D-glucosamine-based peracetylated glycosyl chlorides bearing different substituents at C-2 and three glycosylation promoters, silver carbonate, silver triflate and mercury II chloride/mercury II oxide, by the Koenigs-Knorr method. Under the conditions studied, glycosylation was successful only when 3,4,6-tri-O -acetyl-2-deoxy-2-phthalimido-α-D-glucopyranosyl chloride was used as the glycosyl donor, with silver carbonate proving the best promoter. In order to investigate the influence of the nature of the halogen at C-1, we also carried out the glycosylation of cyclohexanol with 3,4,6-tri-O -acetyl-2-deoxy-2-phthalimido-α-D-glucopyranosyl bromide, a more reactive glycosyl donor. As expected, the yield with the bromide derivative was higher with the three promoters and, again, silver carbonate was the most efficient promoter. Finally, to illustrate the well-known efficient procedure for conversion of the phtalimido group at C-2 to the corresponding acetamido group, cyclohexyl 3,4,6-tri-O -acetyl-2-deoxy-2-phtalimido-β-D-glucopyranoside was converted into cyclohexyl 2-deoxy-2-acetamido-β-D-glucopyranoside in two steps, namely, hydrazinolysis of the phtalimido group followed by chemoselective acetylation of the free amino group by treatment with acetic anhydride in methanol, at 77% overall yield.

The relation of the concentration of macronutrients in the substrate and in the foliage to cell wall thickness and cellulose concentration in the xylem of slash pine (Pinus elliotti)

Sand culture experiments, using a sub-irrigation technique, were installed in order to find out the effects of the macronutrients N, P, K, Ca, Mg and S on growth, aspect, mineral composition, length of fibers, thickness of cell wall and cellulose concentration in slash pine. The aim was to obtain, under controlled conditions, basic information which could eventually lead to practical means designed to increase the rate of growth and to make of slash pine a richer source of cellulose. Nitrogen, Phosphorus, Potassium Experiment A 3 x 3 x 3 factorial design with two replicates was used. Nitrogen was supplied initially at the levels of 25, 50 and 100 ppm; phosphorus was given at the rates of 5, 10 and 20 ppm; potassium was supplied at the rates of 25, 50 and 100 ppm; six months after the experiment was started the first level for each element was dropped to zero. Others macro and all micronutrients were supplied at uniform rates. Fifteen hours of illumination per day were provided. The experimental technique for growing the slash pine seedlings proved quite satisfactory. Symptoms of deficiency of nitrogen, phosphorus and potassium were observed, described and recorded in photographs and water colors. These informations will help to identify abnormalities which may appear under field conditions. Chemical analysis of the several plant parts, on the other hand, give a valuable means to assess the nutritional status of slash pine, thus confirming when needed, the visual diagnosis. The correctness of manurial pratices, on the other hand, can be judged with the help of the analytical data tabulated. Under the experimental conditions nitrogen caused the highest increases on growth, as measured by increments in height and dry weights, whereas the effects of phosphorus and potassium were less marked. Cellulose concentration was not significantly affected by the treatments used. Higher levels of N seemed to decrease both length of fiber elements and the thickness of cell wall. The effects of P and K were not well defined. Calcium, Magnesium, Sulfur Experiment A 3 x 3 x 3 factorial design with two replicates was used. Calcium was supplied initially at the levels of 12.5, 25 and 50 ppm; magnesium and sulfur were given at the rates of 6, 12.5 and 25 ppm. Other macro and micronutrients were supplied at uniform rates, common to all treatments. Three months after starting the experiment the first level for each element was dropped to zero. Symptoms of deficiency of calcium, magnesium and sulfur were observed, described and recorded as in the case of the previous experiment. Chemical analysis were made, both for mineral content and cellulose concentration. Length of fibers and thickness of cell wall were measured. Both calcium and magnesium increase height, sulfur failing to give significant response. Dry weight was beneficially affected by calcium and sulfur. The levels of calcium, magnesium and sulfur in the needles associated with deficiency and maximum growth are comparable with those found in the literature. Cellulose concentration increased when the level of sulfur in the substrate was raised. The thickness of cell wall was negatively affected by the treatments; no effect was observed with regards to length of fibers.

A simple method to purify biologically and antigenically preserved bloodstream trypomastigotes of Trypanosoma cruzi using Deae-cellulose columns

A method to purify trypanosomastigotes of some strains of Trypanosoma cruzi (Y, CL, FL, F, "Berenice", "Colombiana" and "São Felipe") from mouse blood by using DEAE-cellulose columns was standardized. This procedure is a modification of the Lanham & Godfrey methods and differs in some aspects from others described to purify T. cruzi bloodstream trypomastigotes, mainly by avoidance of prior purifications of parasites. By this method, the broad trypomastigotes were mainly isolated, accounting for higher recoveries obtained with strains having higher percentages of these forms: processing of infected blood from irradiated mice could be advantageous by increasing the recovery of parasites (percentage and/or total number) and elution of more slender trypomastigotes. Trypomastigotes purified by this method presented normal morphology and motility, remained infective to triatomine bugs and mice, showing in the latter prepatent periods and courses parasitemia similar to those of control parasites, and also reproducing the polymorphism pattern of each strain. Their virulence and pathogenicity also remained considerably preserved, the latter property being evaluated by LD 50 tests, mortality rates and mean survival time of inoculated mice. Moreover, these parasites presented positive, clear and peripheral immunofluorescence reaction at titres similar to those of control organisms, thus suggesting important preservation of their surface antigens.

Use of glass beads and CF 11 cellulose for removal of leukocytes from malaria-infected human blood in field settings

Passage of malaria infected blood through a two-layered column composed of acid-washed glass beads and CF 11 cellulose removes white cells from parasitized blood. However, because use of glass beads and CF 11 cellulose requires filtration of infected blood separately through these two resins and the addition of ADP, the procedure is time-consuming and may be inapropriate for use in the field, especially when large numbers of blood samples are to be treated. Our modification of this process yields parasitized cells free of contaminating leukocytes, and because of its operational simplicity, large numbers of blood samples can be processed. Our procedure also compares well with those using expensive commercial Sepacell resins in its ability to separate leukocytes from whole blood. As a test of usefulness in molecular biologic investigations, the parasites obtained from the blood of malaria-infected patients using the modified procedure yield genomic DNA whose single copy gene, the circumsporozite gene, efficiently amplifies by polymerase chain reaction.

Molecular and morphological characterization of hydrochar produced by microwave-assisted hydrothermal carbonization of cellulose

The objective of this work was to characterize the morphology and molecular composition of the hydrochar produced by microwave-assisted hydrothermal carbonization of cellulose. The produced hydrochar consists mainly of aggregate microspheres with about 2.0 µm in diameter, with aliphatic and aromatic structures and the presence of carbonyl functional groups. The aromatic groups are formed mainly by benzofuran-like structures, being chemically different from common cellulose char. Microwave-assisted hydrothermal carbonization yields a functionalized carbon-rich material similar to that produced by the conventional hydrothermal process.

Optimization of wood flour acetylation by factorial design and partial least squares regression

Acetylation was performed to reduce the polarity of wood and increase its compatibility with polymer matrices for the production of composites. These reactions were performed first as a function of acetic acid and anhydride concentration in a mixture catalyzed by sulfuric acid. A concentration of 50%/50% (v/v) of acetic acid and anhydride was found to produced the highest conversion rate between the functional groups. After these reactions, the kinetics were investigated by varying times and temperatures using a 3² factorial design, and showed time was the most relevant parameter in determining the conversion of hydroxyl into carbonyl groups.

Hydrogels of cellulose acetate crosslinked with pyromellitic dianhydride: part I: synthesis and swelling kinetics

This work describes the synthesis of hydrogels of cellulose acetate (AC) crosslinked with 1,2,4,5-benzenotetracarboxylic dianhydride (PMDA). The crosslinking reaction was monitored by FTIR. Analysis of aromatic fragments from the alkaline hydrolysis of the gels by UV spectroscopy indicated that an increase in the stoichiometric ratio of dianhydride resulted in higher degrees of crosslinking. The non-porous nature of the gels was confirmed by analysis of nitrogen adsorption. Water absorption isotherms showed that as the temperature and degree of crosslinking increased, the percentage of water absorbed at equilibrium (%Seq) also increased. The hydrogels presented second order swelling kinetics.

ISOLATION AND CHARACTERIZATION OF NANOFIBRILLATED CELLULOSE FROM OAT HULLS

The objectives of this work were to investigate the microstructure, crystallinity and thermal stability of nanofibrillated cellulose obtained from oat hulls using bleaching and acid hydrolysis at a mild temperature (45 ºC) followed by ultrasonication. The oat hulls were bleached with peracetic acid, and after bleaching, the compact structure around the cellulosic fibers was removed, and the bundles became individualized. The extraction time (30 or 60 min) did not affect the properties of the nanofibrillated cellulose, which presented a higher crystallinity index and thermal stability than the raw material (oat hulls). The nanocellulose formed interconnected webs of tiny fibers with diameters of 70-100 nm and lengths of several micrometers, producing nanofibers with a relatively high aspect ratio, thus indicating that these materials are suitable for polymer reinforcement.