Use of confocal scanning laser microscopy to measure the concentrations of aerial and penetrative hyphae during growth of Rhizopus oligosporus on a solid surface

In order to develop a method for use in investigations of spatial biomass distribution in solid-state fermentation systems, confocal scanning laser microscopy was used to determine the concentrations of aerial and penetrative biomass against height and depth above and below the substrate surface, during growth of Rhizopus oligosporus on potato dextrose agar. Penetrative hyphae had penetrated to a depth of 0.445 cm by 64 h and showed rhizoid morphology, in which the maximum biomass concentration, of 4.45 mg dry wt cm(-3), occurred at a depth of 0.075 cm. For aerial biomass the maximum density of 39.54 mg dry wt(-3) occurred at the substrate surface. For both aerial and penetrative biomass, there were two distinct regions in which the biomass concentration decayed exponentially with distance from the surface. For aerial biomass, the first exponential decay region was up to 0.1 cm height. The second region above the height of 0.1 cm corresponded to that in which sporangiophores dominated. This work lays the foundation for deeper studies into what controls the growth of fungal hyphae above and below the surfaces of solid substrates. (C) Wiley Periodicals, Inc.

Use of confocal microscopy to follow the development of penetrative hyphae during growth of Rhizopus oligosporus in an artificial solid-state fermentation system

Two methods were compared for determining the concentration of penetrative biomass during growth of Rhizopus oligosporus on an artificial solid substrate consisting of an inert gel and starch as the sole source of carbon and energy. The first method was based on the use of a hand microtome to make sections of approximately 0.2- to 0.4-mm thickness parallel to the substrate surface and the determination of the glucosamine content in each slice. Use of glucosamine measurements to estimate biomass concentrations was shown to be problematic due to the large variations in glucosamine content with mycelial age. The second method was a novel method based on the use of confocal scanning laser microscopy to estimate the fractional volume occupied by the biomass. Although it is not simple to translate fractional volumes into dry weights of hyphae due to the lack of experimentally determined conversion factors, measurement of the fractional volumes in themselves is useful for characterizing fungal penetration into the substrate. Growth of penetrative biomass in the artificial model substrate showed two forms of growth with an indistinct mass in the region close to the substrate surface and a few hyphae penetrating perpendicularly to the surface in regions further away from the substrate surface. The biomass profiles against depth obtained from the confocal microscopy showed two linear regions on log-linear plots, which are possibly related to different oxygen availability at different depths within the substrate. Confocal microscopy has the potential to be a powerful tool in the investigation of fungal growth mechanisms in solid-state fermentation. (C) 2003 Wiley Periodicals, Inc.

Fermentación en estado sólido del mijo perla (Pennisetum americahum (L.) Leeke) por Rhizopus oligosporus para la obtención de un producto rico en proteína

Tesis (Maestría en Ciencias con Especialidad en Alimentos) UANL

Control of lipase production by Rhizopus oligosporus under various growth conditions

Some factors influencing the growth and production of extracellular lipase by Rhizopus oligosporus were studied. Highest yields of enzyme were obtained when Tweens were the carbon source. Soybean meal extract supported good growth and enzyme production. Carbohydrates, vegetable oils, proteins or amino acids did not stimulate lipase production. The fungus grew well with carbohydrate- or protein-supplemented media but not with oils, unless emulsified with a non-metabolizable gum. The production of biomass in static cultures was maximum at 35-40°C after 4 d at pH 5.5. The yield of lipase was maximum at 25°C after 3 d at pH 6.5. Shaking cultures enhanced growth but decreased lipase production.

Bioconversão energética da folha e bagaço de mandioca pelo fungo Rhizopus oligosporus para obtenção de alimento funcional

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Influência da temperatura na atividade de amilase e protease de Rhizopus oligosporus cultivado por fermentação em estado sólido

Amilases e proteases constituem um dos principais grupos de enzimas industriais pelo seu amplo espectro de aplicações biotecnológicas. Elas podem ser obtidas a partir de fontes microbianas e com altos rendimentos por processos de fermentação em estado sólido (FES). Conhecer as características bioquímicas das enzimas é fundamental para adequação aos processos industriais. O objetivo do trabalho foi determinar a melhor temperatura para atividade das enzimas amilase e protease de Rhizopus oligosporus obtidas por fermentação em estado sólido utilizando farelo de trigo como substrato. Os melhores valores para atividade amilolítica e proteolítica foram obtidos nas temperaturas de 55 - 65 °C e de 50 - 60 °C, respectivamente. Estes resultados sugerem que as enzimas estudadas podem ser utilizadas em processos que empregam elevadas temperaturas.

Desenvolvimento de um bioprocesso utilizando-se resíduos para produção de amilases por Rhizopus oligosporus e etanol por Saccharomyces cerevisiae

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Viabilidade de obtenção de alimento funcional a base de farinha de mesocarpo de babaçu (Orbignya sp.) e folhas de mandioca (Manihot esculenta) mediante fermentação por Rhizopus microsporus var. oligosporus

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Biotechnological production of lactic acid integrated with potato wastewater treatment by Rhizopus arrhizus

This paper describes a feasibility study of a for lactic acid production integrated with are treatment of wastewater from an industrial starch plant. Rhizopus oryzae two strains, Rhizopus arrhizus and Rhizopus oligosporus were tested with respect to their capability to carry out simultaneous saccharification and fermentation to lactic acid using potato wastewater. Rhizopus arrhizus DAR 36017 was identified as a suitable strain that demonstrated a high capacity for starch saccharification and lactic acid synthesis. The optimal conditions, in terms of pH, temperature and starch concentration, for lactic acid production were determined. The selected fungal strain grew well in a pH range from 3.0 to 7.0. The addition of CaCO(3)10 g dm(-3) maintained the pH at 5.0-6.0 and significantly enhanced lactic acid production. Kinetic study revealed that almost complete starch saccharification and a lactic acid yield of 450g kg(-1) could be achieved in 20 h and 28 h cultivation, respectively. The maximum lactic acid production 21 g dm(-3) and mycelial biomass (1.7 g dm(-3)) were obtained at 30degreesC. Besides the multiple bioproducts, total removal of suspended solids and 90% reduction of COD were achieved in a single no-aseptic operation. (C) 2003 Society of Chemical Industry.

O-2 uptake during solid-state fermentation in a rotating drum bioreactor

The maximum O-2 uptake by Rhizopus oligosporus grown in a 200 litre rotating drum bioreactor at 0.5 rpm ranged from 6.7 to 7.6 mmol per min per kg initial dry substrate (IDS), for runs done with 4 baffles each 17 cm wide, and 12 baffles each 5 cm wide. Without baffles, the maximum O-2 uptake rate at 5 rpm was 6.9 mmol/(min.kg IDS), compared to 5.1 mmol/(min.kg IDS) obtained at 0.5 rpm. Therefore O-2 supply is adequate in rotating drum bioreactors as long as slumping flow regimes of the substrate bed are avoided.

Tempeh flour as a substitute for soybean flour in coconut cookies

The objective of this study was to prepare roasted and lyophilized tempeh flour with soybean cultivar BRS 267 to apply them in the formulation of coconut biscuits. The cookies produced with whole soy flour and mixed flour of soybean and tempeh were evaluated for proximate composition, fatty acid profile, and isoflavone aglycones in order to verify the effects of inoculation with the fungus Rhizopus oligosporus and those of the drying processes of roasting and lyophilization on the chemical characteristics of the final product. Sensory acceptance and purchase intention of the formulated products were also evaluated. The results indicate the maintenance of linolenic acid, which is important in the prevention of coronary diseases, and an increase in the aglycones levels when the tempeh flour was used. Lipids and proteins showed differences, and the sensory analyses demonstrated similarity between the cookies with satisfactory scores for aroma, flavor, texture, and overall acceptability for both samples. when compared to the control. Purchase intent was also positive for the lyophilized and toasted tempeh flours, thus enabling the use of the roasting process as a simple drying method, for processing tempeh and obtaining a flour rich in proteins and aglycones that can be used as a partial substitute for soy flour in cookies and other bakery products.

Cloning, sequencing and characterization of a chitin synthase gene fragment from the fungus Phascolomyces articulosus /

Phascolomyces articulosus genomic DNA was isolated from 48 h old hyphae and was used for amplification of a chitin synthase fragment by the polymerase chain reaction method. The primers used in the amplification corresponded to two widely conserved amino acid regions found in chitin synthases of many fimgi. Amphfication resulted in four bands (820, 900, 1000 and 1500 bp, approximately) as visualized in a 1.2% agarose gel. The lowest band (820 bp) was selected as a candidate for chitin synthase because most amplified regions from other fimgi so far exhibited similar sizes (600-750 bp). The selected fragment was extracted from the gel and cloned in the Hinc n site of pUC19. The derived plasmid and insert were designated ^\5C\9'PaCHS and PaCHS respectively. The plasmid pUC19-PaC/fS was digested by several restriction enzymes and was found to contain BamHl and HincU sites. Sequencing of PaCHS revealed two intron sequences and a total open reading frame of 200 amino acids. The derived polypeptide was compared with other related sequences from the EMBL database (Heidelberg, Germany) and was matched to 36 other fiilly or partially sequenced fimgal chitin synthase genes. The closest resemblance was with two genes (74.5% and 73.1% identity) from Rhizopus oligosporus. Southern hybridization with the cloned fragment as a probe to the PCR reaction showed a strong signal at the fragment selected for cloning and weaker signals at the other two fragments. Southern hybridization with partially digested Phascolomyces articulosus genomic DNA showed a single band. The amino acid sequence was compared with sequences from other chitin synthase gene classes using the CLUSTALW program. The chitin synthase fragment from Phascolomyces articulosus was initially grouped in class n along with chitin synthase fragments from Rhizopus oligosporus and Phycomyces blakesleeanus which also belong to the same class, Zygomycetes. Bootstrap analysis using the neighbor-joining method available by CLUSTALW verified such classification. Comparison of PaCHS revealed conservation of intron positions that are characteristic of chitin synthase gene fragments of zygomycetous fungi.

Funcionalidade dos extratos fenólicos obtidos pelo cultivo semi-sólido de resíduos de abacaxi (Ananas comusus L.) e goiaba (Psidium guajava L.)

Solid substrate cultivation (SSC) has become an efficient alternative towards rational use of agro industrial wastes and production of value-added products, mainly in developing countries. This work presents the production and functional application results of phenolic extracts obtained by solid substrate cultivation of pineapple (Ananas comosus L.) and guava (Psidium guajava L.) residues associated to soy flour and bioprocessed by Rhizopus oligosporus fungus. Two experimental groups were tested: (1) 9g of fruit residue and 1g of soy flour (A9 or G9); (2) 5g of fruit residue and 5g of soy flour (A5 or G5). After SSC, 100ml of distilled water was added to each Erlenmeyer flask containing 10g of bioprocessed material in order to obtain the phenolic extracts. Samples were taken every two days for total phenolic concentration (TPC) and antioxidant capacity evaluation by DPPH test during 12-day cultivation. The 2-day and 10-d ay extracts were selected and concentrated by ebullition until 1/10 of original volume was reached. After that, both non-concentrated and concentrated extracts were evaluated for their antimicrobial activity against Staphylococcus aureus and Salmonella enterica and a-amylase inhibitory capacity. It was observed an inverse relationship between total phenolic concentration (TPC) and antioxidant capacity during the cultivation. Besides that, the concentrated pineapple samples after two days were able to inhibit both pathogens tested, especially S. aureus. Guava concentrated extracts after 2 days showed expressive inhibition against S. enterica, but negative results against S. aureus growth. When it comes to a-amylase inhibition, A9 extracts after 2 days, both concentrated or not, completely inhibited enzyme activity. Similar behavior was observed for G9 samples, but only for concentrated samples. It was shown that concentration by ebullition positively affected the enzymatic inhibition of G9 and A9 samples, but on the other side, decreased antiamylase activity of A5 and G5 samples

Estudo de biorremediação dos azo corantes têxteis Acid Blue 161 e Procion Red MX-5B por fungos filamentosos em solução simples e solução binária associado a testes de toxicidade com Lactuca sativa e Artemia salina

Pós-graduação em Ciências Biológicas (Microbiologia Aplicada) - IBRC

Proceedings of the Nineteenth Annual Biochemical Engineering Symposium

The nineteenth symposium was held at the University of Missouri–Columbia on April 22, 1989. A total of eighteen papers were scheduled for presentation, of which nine were in poster session. Finally, fifteen papers were presented and sixteen were submitted for this proceedings. It was attended by 53 participants from five institutions. A sixth group (from Colorado State University) was kept from attending the symposium due to mechanical problems on the road and we missed them. Since they worked hard at their presentations, I requested CSU-group to submit their papers for the proceedings and I am happy that they did. ContentsMathematical modelling of a flour milling system. K. Takahashi, Y. Chen, J. Hosokoschi, and L. T. Fan. Kansas State University A novel solution to the problem of plasmid segregation in continuous bacterial fermentations. K.L. Henry, R. H. Davis, and A. L. Taylor. University of Colorado Modelling of embryonic growth in avian and reptile Eggs. C.L. Krause, R. C. Seagrave, and R. A. Ackerman. Iowa State University Mathematical modeling of in situ biodegradation processes. J.C. Wu, L. T. Fan, and L. E. Erickson. Kansas State University Effect of molecular changes on starch viscosity. C.H. Rosane and V. G. Murphy. Colorado State University Analysis of two stage recombinant bacterial fermentations using a structured kinetic model. F. Miao and D. S. Kampala. University of Colorado Lactic acid fermentation from enzyme-thinned starch by Lactobacillus amylovorus. P.S. Cheng, E. L. Iannotti, R. K. Bajpai, R. Mueller, and s. Yaeger. University of Missouri–Columbia Solubilization of preoxidized Texas lignite by cell-free broths of Penicillium strains. R. Moolick, M. N. Karim, J. C. Linden, and B. L. Burback. Colorado State University Separation of proteins from polyelectrolytes by ultrafiltration. A.G. Bazzano and C. E. Glatz. Iowa State University Growth estimation and modelling of Rhizopus oligosporus in solid state fermentations. D.-H. Ryoo, V. G. Murphy, M. N. Karim, and R. P. Tengerdy. Colorado State University Simulation of ethanol fermentations from sugars in cheese whey. C.J. Wang and R. K. Bajpai. University of Missouri–Columbia Studies on protoplast fusion of B. licheniformis. B. Shi, Kansas State University Cell separations of non-dividing and dividing yeasts using an inclined settler. C.-Y. Lee, R. H. Davis, and R. A. Sclafani. University of Colorado Effect of·serum upon local hydrodynamics within an airlift column. G.T. Jones, L. E. Erickson, and L. A. Glasgow. Kansas State University Optimization of heterologous protein secretion in continuous culture. A. Chatterjee, W. F. Remirez, and R. H. Davis. University of Colorado An improved model for lactic acid fermentation. P. Yeh, R. K. Bajpai, and E. L. Iannotti. University of Missouri–Columbia