Skin unhairing proteases of Bacillus subtilis: production andpartial characterization

Proteases able to unhair sheep skins were produced from Bacillus subtilis. Protease activity was increased from 640 to 990 U/mL by using a fed-batch culture with glucose added sequentially up to 10 g/L. The crude enzyme prep-aration was mainly a mixture of metallo- and serine-proteases with optimal pH and temperature for protease activity at 7 and 50-55°C, respectively. At pH 8 and 30°C, protease activity of the crude enzyme was 33% of the maximal value and 97% of the original activity (900 U/mL) was retained after incubation for 4 hours.     

Isolation and selection of native microorganisms for the aerobic treatment of simulated dairy wastewaters

Milk fat/protein degrading microorganisms were isolated from different locations of a dairy wastewater treatment system with the goal of developing an inoculum for bioaugmentation strategies. Eight isolates, identified by 16S rRNA gene sequence analysis as belonging to the genera Bacillus, Pseudomonas, and Acinetobacter, were tested for their ability to remove COD and protein from a milk-based medium (3000 mg/L COD) and compared to a commercial bioaugmentation inoculum. The Acinetobacter isolate exhibited a pellet-type growth in liquid culture, a property that could potentially aid in the separation of microbes and liquid phase following treatment. Based on the individual degradation capacity and growth behavior of the isolates, three microorganisms were further selected and tested together. This consortium exhibited a COD removal similar to the commercial inoculum (57% and 63%, respectively), but higher protein (consortium: 93%; commercial inoculum: 54%), and fat removals (consortium: 75%; commercial inoculum: 38%).     

Baker's yeast production from molasses/cheese whey mixtures

Partial substitution of sugarcane molasses by cheese whey in the fed-batch production of baker's yeast was evaluated. A sugar feeding profile based in a commercial process and different modes of addition of -galactosidase were used. Molasses substitution of 46%, in terms of sugar fed to the bioreactor, was reached and no significant differences in biomass volumetric productivity, by-products yields, and baking quality were observed. However the biomass yield was 6% lower.     

Antarctic microorganisms as source of the omega-3 polyunsaturated fatty acids

Docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) are long-chain polyunsaturated fatty acids (PUFAs) that belong to the omega-3 group. They are essential fatty acids found in phospholipid of cell membranes. There is strong evidence that these nutrients may also favorably modulate many diseases. Primary sources of omega-3 PUFAs in the human diet are fish and fish-derived products. The fishing industry worldwide, however, is becoming unable to satisfy the growing demand for these PUFAs. A promising cost-effective alternative source of PUFAs is bacterial production. We identified 40 Antarctic marine bacterial isolates by 16S rRNA gene sequence analysis. Fifteen genera in three phyla were represented in the collection. Isolates were tested for ability to produce EPA using a method in which their ability to reduce 2,3,5-triphenyltetrazolium chloride (TTC) is determined and by gas chromatography coupled to mass spectrometry (GC–MS). All isolates could reduce TTC, and GC–MS analysis showed that four produced EPA and that six produced DHA. We show for the first time that isolates identified as Cellulophaga, Pibocella and Polaribacter can produce EPA and DHA, only DHA or only EPA, respectively. One isolate, Shewanella sp. (strain 8-5), is indicated to be a good candidate for further study to optimize growth and EPA production. In conclusion, a rapid method was tested for identification of new EPA producing strains from marine environments. New EPA and DHA producing strains were found as well as a potentially useful PUFA production strain.     

Kinetic properties of a commercial and a native inoculum for aerobic milk fat degradation

The aerobic fat biodegradation potential and growth characteristics of a commercial and a native inoculum (activated sludge from a dairy wastewater treatment pond), were evaluated. Batch tests were conducted with a medium based on butter oil, as the sole source of carbon, and mineral salts. Residual fat, biomass and CO(2) production were measured. Overall fat removal values were above 78% for both inocula. The growth kinetics of the commercial and native inocula followed Haldane and Monod models respectively. Both inocula showed a similar behaviour when butter oil concentration was under 360 mg/l; at higher values, the difference between the growth rates increased as a consequence of the inhibition exhibited by the commercial inoculum. The selection of an inoculum for bioaugmentation of bioreactors in the wastewater treatment requires a comprehensive knowledge of their degradation ability and tolerance to fluctuating compounds and of the operational conditions that will be utilized.     

Performance of a commercial inoculum for the aerobic biodegradation of a high fat content dairy wastewater

The effectiveness of a commercial inoculum for degrading a dairy wastewater with high fat content was evaluated, and compared with an activated sludge inoculum from a dairy wastewater treatment pond. Both inocula reached similar chemical oxygen demand removal in batch experiments. The population dynamics was also studied by determining heterotrophic counts. Predominant microorganisms were differentiated by colony morphology and genomic fingerprinting (BOX-PCR) analysis. The higher population diversity and the wider range of CO2 production rate observed in batch reactors inoculated with activated-sludge, indicated that microorganisms from this inoculum were well adapted and may have had synergic activity for the degradation of the dairy effluent. When the bioreactor was operated with the commercial inoculum in continuous mode, according to its microbial growth kinetics, other microorganisms became predominant. These results showed that inoculated microorganisms did not persist in the open system and periodic addition of microorganisms may be needed to achieve a high performance treatment.     

Ethanol production from concentrated whey permeate using a fed-batch culture ofKluyveromyces fragilis

Summary Fed-batch fermentation of non-supplemented concentrated whey permeate resulted in high ethanol productivity for feeds of lactose for which batch fermentation had a poor performance. At an initial lactose concentration of 100 g/L and a constant lactose feeding rate of 18 g/h we have obtained: ethanol concentration 64 g/L, ethanol productivity 3.3 g/Lh, lactose consumption 100%, ethanol yield 0.47 g/g, and biomass yield 0.058 g/g.Nomenclature St total lactose fed per medium volume in the bioreactor, g/L - Si initial lactose concentration, g/L - F lactpse feeding rate, g/h - P final ethanol concentration, g/L - Y_p/s ethanol yield, g ethanol/g lactose - Y_x/s biomass yield, g biomass/g lactose - X_S lactose consumption, % - Qp overall ethanol volumetric productivity, g/Lh - m maximum specific growth rate, h - qsm maximum specific lactose consumption rate, g/gh - qpm maximum specific ethanol production rate, g/gh     

Extracellular enzymes produced by microorganisms isolated from maritime Antarctica

Antarctic environments can sustain a great diversity of well-adapted microorganisms known as psychrophiles or psychrotrophs. The potential of these microorganisms as a resource of enzymes able to maintain their activity and stability at low temperature for technological applications has stimulated interest in exploration and isolation of microbes from this extreme environment. Enzymes produced by these organisms have a considerable potential for technological applications because they are known to have higher enzymatic activities at lower temperatures than their mesophilic and thermophilic counterparts. A total of 518 Antarctic microorganisms, were isolated during Antarctic expeditions organized by the Instituto Antártico Uruguayo. Samples of particules suspended in air, ice, sea and freshwater, soil, sediment, bird and marine animal faeces, dead animals, algae, plants, rocks and microbial mats were collected from different sites in maritime Antarctica. We report enzymatic activities present in 161 microorganisms (120 bacteria, 31 yeasts and 10 filamentous fungi) isolated from these locations. Enzymatic performance was evaluated at 4 and 20°C. Most of yeasts and bacteria grew better at 20°C than at 4°C, however the opposite was observed with the fungi. Amylase, lipase and protease activities were frequently found in bacterial strains. Yeasts and fungal isolates typically exhibited lipase, celullase and gelatinase activities. Bacterial isolates with highest enzymatic activities were identified by 16S rDNA sequence analysis as Pseudomonas spp., Psychrobacter sp., Arthrobacter spp., Bacillus sp. and Carnobacterium sp. Yeasts and fungal strains, with multiple enzymatic activities, belonged to Cryptococcus victoriae, Trichosporon pullulans and Geomyces pannorum.     

Short Communication: Effect of medium composition on the production by a new Bacillus subtilis isolate of protease with promising unhairing activity

In a search for microorganisms producing extracellular protease with unhairing activity, Bacillus subtilis IIQDB32 was isolated. Protease formation was significantly stimulated by glucose, tryptone, yeast extract, Ca²⁺ and Mn²⁺, but was repressed by ammonia and Fe²⁺.