In vitro fermentation of Pennisetum clandestinum Hochst. Ex Chiov increased methane production with ruminal fluid adapted to crude glycerol.

The aim of this trial was to determine the effect of ruminal fluid adapted to crude glycerol supplementation on in vitro methane and fermentation parameters of the forage Pennisetum clandestinum Hochst. Ex Chiov (Kikuyu forage) of 35 (D35) and 45 days (D45) of regrowth. Four non-lactating cannulated Holstein cows with 600 ± 33 kg LW, under a randomized Latin square design (4 animals × 4 periods), were used to obtain ruminal fluid after adaptation for 13 days within each period to supplementation in the rumen with crude glycerol (GL): 0 (GL0), 500 (GL1), 1000 (GL2), and 1500 g/cow/day (GL3). All the variables were analyzed under a 4 × 2 split-plot arrangement, where the ruminal fluid adaptation to supplementation with GL and its interaction with the incubated diets (GL× D) were analyzed. Ruminal pH values and in vitro apparent degradability of the dry matter (DMd) and organic matter (OMd) were not affected, with a decrease in the concentrations of NH3-N and acetate by the ruminal fluid adapted to crude glycerol. The concentrations of propionate and butyrate increased, without affecting the total concentration of VFA in the ruminal fluid. The in vitro methane production increased with forage D35 and GL1, GL2, GL3. The in vitro methane production with the D45 forage increased with the ruminal fluid adapted to GL2 and GL3. The ruminal fluid adapted to crude glycerol supplementation increased in vitro methane production when incubating the forage Kikuyu.

Production and partial characterization of serine and metallo peptidases secreted by Aspergillus fumigatus Fresenius in submerged and solid state fermentation.

Enzyme production varies in different fermentation systems. Enzyme expression in different fermentation systems yields important information for improving our understanding of enzymatic production induction. Comparative studies between solid-state fermentation (SSF) using agro-industrial waste wheat bran and submerged fermentation (SmF) using synthetic media were carried out to determinate the best parameters for peptidase production by the fungus Aspergillus fumigatus Fresen. Variables tested include: the concentration of carbon and protein nitrogen sources, the size of the inoculum, the pH of the media, temperature, and the length of the fermentation process. The best peptidase production during SSF was obtained after 96 hours using wheat bran at 30 °C with an inoculum of 1 × 10(6) spores and yielded 1500 active units (U/mL). The best peptidase production using SmF was obtained after periods of 72 and 96 hours of fermentation in media containing 0.5% and 0.25% of casein, respectively, at a pH of 6.0 and at 30 °C and yielded 40 U/mL. We also found examples of catabolite repression of peptidase production under SmF conditions. Biochemical characterization of the peptidases produced by both fermentative processes showed optimum activity at pH 8.0 and 50 °C, and also showed that their proteolytic activity is modulated by surfactants. The enzymatic inhibition profile using phenylmethylsulfonyl fluoride (PMSF) in SmF and SSF indicated that both fermentative processes produced a serine peptidase. Additionally, the inhibitory effect of the ethylene-diaminetetraacetic acid (EDTA) chelating agent on the peptidase produced by SmF indicated that this fermentative process also produced a metallopeptidase.

Fermentation conditions for yeast selection and effect of yeast-bacterial interaction in developing a starter culture for tequila fermentation.

Similar to other beverages, tequila fermentation can be improved using selected native strains. During fermentation, yeast and bacteria frequently act together and can improve product quality. However, their influence during tequila fermentation is not fully understood. Three Saccharomyces cerevisiae strains isolated in a previous study were used to determine their fermentation parameters. Fermentation of agave juice by a selected yeast strain in the presence of seven native bacterial species (individually and in consortium) was also evaluated. Optimal temperature, initial fructose, and pH were determined using a rotatory central composite design, and concomitant fermentative parameters were used to select the best tequila yeast strain. The yeast strain Teq-199 presented the best fructose consumption (91.0%), ethanol production (33.5 g/L), and yield (72%), as well as produced low concentrations of acetic acid (0.2 g/L). The optimal fermentation temperature was 32°C, similar to that used at distilleries. During fermentation of agave juice by strain Teq-199 in the presence of individual native bacteria, the yeast dominated fermentation, while bacterial species impaired its quality due to lactic acid production (>3.3 g/L). In the presence of a bacterial consortium, strain Teq-199 also dominated fermentation; however, a high concentration of acetic acid (6.5 g/L) was generated, reducing the quality of the fermentation parameters. A comparison with a commercial wine strain K1-V1116, revealed that the strain Teq-199 exhibited the best fermentation parameters, indicating its potential use as an inoculant in industrial processing. Since bacteria were detrimental, distilleries should exert efforts to reduce their impact. PRACTICAL APPLICATION: The optimal fermentation conditions for pre-selected yeast strains isolated during tequila production were determined, which led to the selection of the best-adapted yeast strain to fulfill the actual requirements and conditions in the tequila industry. This study allows the selection of yeast strains that could be used by local producers. In addition, this yeast was able to overcome a well-adapted bacterial consortium, showing dominance throughout the fermentation process. This study is unique since it explored bacterial interactions during fermentation process and proposes a method to assess the effect of these concomitant microorganisms to properly select and design a starter culture.

Effect of concentrate feeding sequence on equine hindgut fermentation parameters.

Feeding a diet with a small amount of hay and a high proportion of concentrate given in large meals entails a risk of colic to horses as this can impact the hindgut microbial ecosystem. To counteract this potential negative effect, one feeding strategy is to modify the distribution sequence of concentrate and hay. The purpose of the present study was to assess the impact of feeding the concentrate meal before or with the hay meal on fermentative parameters postprandial variations in caecum and right ventral colon contents and faeces. We focused on the comparison of pH value and volatile fatty acid (VFA) concentrations between the three compartments. Four horses fitted with two cannulas in the caecum and right ventral colon were fed a 100% hay diet (H) for a 3-week period, and then a 40% concentrate-60% hay diet for two consecutive periods of 3 weeks each. Concentrate and hay were offered simultaneously in two equal meals at 0800 and 1700 h (HC1) during one period, or separately in two equal meals at 0800 and 1600 h for the concentrate and at 1000 and 1730 h for hay (HC2) during the second one. During the 2nd week of each diet period (H, HC1, HC2), caecal, right ventral colonic and faecal samples were collected before (0) and at 2, 4, 6 and 8 h after the morning meal. pH values and VFA concentrations were measured. Adding concentrate to the hay was associated with a decrease of pH value in the caecum, right ventral colon and faeces and an increase of faecal VFA concentrations (P<0.01). An impact of the concentrate feeding sequence was observed on caecal pH and faecal propionate concentration (P<0.001) but none on the other parameters investigated. In the caecum and right ventral colon, pH values decreased and VFA concentrations increased within the range of 4 to 8 h after the morning meal. Our data suggested that the variation of faecal pH could be an indicator of those of the caecal and right ventral colonic pH with a delay of 2 h. The present study showed that hay and commercial pelleted could be safely distributed simultaneously, thus making easier every day horse feeding.

Production and partial characterization of serine and metallo peptidases secreted by Aspergillus fumigatus Fresenius in submerged and solid state fermentation

Enzyme production varies in different fermentation systems. Enzyme expression in different fermentation systems yields important information for improving our understanding of enzymatic production induction. Comparative studies between solid-state fermentation (SSF) using agro-industrial waste wheat bran and submerged fermentation (SmF) using synthetic media were carried out to determinate the best parameters for peptidase production by the fungus Aspergillus fumigatus Fresen. Variables tested include: the concentration of carbon and protein nitrogen sources, the size of the inoculum, the pH of the media, temperature, and the length of the fermentation process. The best peptidase production during SSF was obtained after 96 hours using wheat bran at 30 ºC with an inoculum of 1 x 10(6) spores and yielded 1500 active units (UµmL). The best peptidase production using SmF was obtained after periods of 72 and 96 hours of fermentation in media containing 0.5% and 0.25% of casein, respectively, at a pH of 6.0 and at 30 ºC and yielded 40 UµmL. We also found examples of catabolite repression of peptidase production under SmF conditions. Biochemical characterization of the peptidases produced by both fermentative processes showed optimum activity at pH 8.0 and 50 ºC, and also showed that their proteolytic activity is modulated by surfactants. The enzymatic inhibition profile using phenylmethylsulfonyl fluoride (PMSF) in SmF and SSF indicated that both fermentative processes produced a serine peptidase. Additionally, the inhibitory effect of the ethylene-diaminetetraacetic acid (EDTA) chelating agent on the peptidase produced by SmF indicated that this fermentative process also produced a metallopeptidase.