Liquid–liquid extraction of fermentation inhibiting compounds in lignocellulose hydrolysate

Several compounds that are formed or released during hydrolysis of lignocellulosic biomass inhibit the fermentation of the hydrolysate. The use of a liquid extractive agent is suggested as a method for removal of these fermentation inhibitors. The method can be applied before or during the fermentation. For a series of alkanes and alcohols, partition coefficients were measured at low concentrations of the inhibiting compounds furfural, hydroxymethyl furfural, vanillin, syringaldehyde, coniferyl aldehyde, acetic acid, as well as for ethanol as the fermentation product. Carbon dioxide production was measured during fermentation in the presence of each organic solvent to indicate its biocompatibility. The feasibility of extractive fermentation of hydrolysate was investigated by ethanolic glucose fermentation in synthetic medium containing several concentrations of furfural and vanillin and in the presence of decanol, oleyl alcohol and oleic acid. Volumetric ethanol productivity with 6 g/L vanillin in the medium increased twofold with 30% volume oleyl alcohol. Decanol showed interesting extractive properties for most fermentation inhibiting compounds, but it is not suitable for in situ application due to its poor biocompatibility. Biotechnol. Bioeng. 2009;102: 1354–1360. © 2008 Wiley Periodicals, Inc.     

Use of a sequential strategy of experimental design to optimize the inulinase production in a batch bioreactor

A potential application of inulinase in the food industry is the production of fructoligosaccharides (FOS) by the transfructosilation of sucrose. The FOSs present many interesting functional properties besides their ability to increase the shelf-life and flavor of many products. The use of an industrial medium represents a good alternative to producing inulinase at low cost, since the activity may improve, or at least remain the same, as that obtained using a synthetic medium. This work was an optimization study of the inulinase production by Kluyveromyces marxianus NRRL Y-7571 using industrial pre-treated culture medium in a bioreactor employing a sequential strategy of experimental design. Initially, a Plackett–Burman (Screening Design) design was used, where the studied variables were molasses, corn steep liquor, yeast extract concentration, and agitation and aeration rates. After the analysis of the effects, a central composite rotational design (CCRD) was carried out. The optimized condition for the inulinase production was: 250 g/l of molasses, 80 g/l of corn steep liquor, 6 g/l of yeast extract, 300 rpm of agitation and 1.5 vvm aeration rate, which resulted in an enzymatic activity of 1,317 ± 65 U/ml.     

Technical viability of the production,partial purification and characterisation of inulinase using pretreated agroindustrial residues

The present work aimed to study the viability of the use of sugarcane molasses and corn steep liquor (CSL) in a sequential inulinase production performing an up-stream pretreatment of these agroindustrial residues. A sequential strategy was used applying three central composite rotatable designs (CCRDs) to optimise medium composition, followed by a down-stream step. The medium containing 150 g L⁻¹ molasses, 50 g L⁻¹ CSL and 6 g L⁻¹ yeast extract, yielded a maximum inulinase production of 1,294 ± 7 U mL⁻¹, after 72 h of fermentation. A down-stream evaluation was carried out using an expanded bed of Streamline DAE resin (Pharmacia), with and without the up-stream treatment. The results showed that the enzyme could not be recovered from the non-pretreated medium, whereas a yield of 91% was obtained in the adsorption stage from the medium prepared with the up-stream treatment, showing the viability of producing the enzyme inulinase from agroindustrial residues using the integrated process.     

Comparative studies on the biochemical properties of the malic enzymes from Trypanosoma cruzi and Trypanosoma brucei

The construction of engineered bacterial cells with a reduced genome allows the investigation of molecular mechanisms that may be cryptic in wild-type strains and derivatives. Previously, a large-scale combined deletion mutant of Escherichia coli that lacked 29.7% of the parental chromosome was constructed by combining large chromosome deletions. In this work, we improved the system for making markerless-chromosomal deletions and obtained mutants with a genome that lacked up to 38.9% of the parental chromosome. Although the large-scale deletion mutants possessed genes needed for resistance to oxidative stress, including superoxide dismutase, catalase, and RpoS, they were sensitive to menadione, which induces reactive oxygen species during stationary phase. Small genome size did not necessarily correlate with greater sensitivity to menadione as several mutants with large deletions were more resistant to menadione. The sensitivity to menadione depended on whether the mutants were grown aerobically or anaerobically, suggesting that the mechanism governing menadione resistance depended on the oxygen tension of the growth medium. Further analysis of the large-scale deletion mutants should help identify the regulatory networks that are important for cellular defense against oxidative stress.     

Humanised models of cancer in molecular medicine: the experimental control of disanalogy

This paper explores the epistemology of extrapolation from model organisms to humans in molecular medicine. We take into account two common views on the issue, the homology view and the disanalogy view. In response to both interpretations, we argue that the foundational basis of extrapolations cannot simply be provided by homology and that relevant disanalogies can, thanks to the techniques of molecular biology, be experimentally controlled and exploited to allow useful and reliable extrapolations. The case of "humanised mice" in the context of cancer stem cell research provides evidence of how animal models can be construed to approximate bona fide causal analogue models of human diseases. To supplement this view we show how the epistemology of model organisms needs to take into account the engineering side of molecular medicine. Model organisms are often manipulated to create analogies or remove disanalogies with the target system. We maintain that highlighting this feature is fundamental to explain what warrants extrapolation in the search for the molecular causes of disease.     

Identification of enzyme-producing thermophilic bacilli isolated from marine vents of Aeolian Islands (Italy)

Enzyme-producing thermophilic bacilli were isolated from different thermal sites located in and around Aeolian Islands (Italy) and characterised by both molecular and culture-based methods. Spore-forming bacteria with optimal growth from 45 to 70 °C were isolated from submarine vents and a geothermal well of Aeolian Islands (Italy). Eighteen selected strains were screened for extracellular enzyme presence by using nine substrates: Tween 20, Tween 80, tributyrin, soluble starch, xylan, dextran, carragenan, gelatine and casein. Sixteen isolates were able to grow at pH 9. The isolates were differentiated on the basis of restriction pattern of their amplified 16S rDNA (ARDRA) prior to 16S rRNA gene sequence analysis. On the basis of the most complete sequencing results strain V3 was identified asGeobacillus thermodenitrificans, most of isolates (10/14) was similar at high level (≥95%) to different reference strains of the speciesBacillus licheniformis. The remaining isolates, exhibiting sequence similarity below 95%, may represent novel species of the genusBacillus.     

Effects of glutamate on the serotonin-induced responses of vestibular neurons

The aim of this work was to verify whether and how spontaneous or glutamate(GLU)-induced enhancements of the neuronal firing rate modified the responsiveness of the vestibular neurons to microiontophoretic application of serotonin (5-HT). During experiments performed on anaesthetized Wistar rats the responses to 5-HT applications were studied in neurons of the lateral vestibular nucleus identified by the antidromic activation upon stimulation of the vestibulospinal tract. The magnitude (in percent) of the 5-HT induced excitatory responses decreased (hyperbolic correlation, r = 0.91) when the background mean firing rate was enhanced spontaneously or by long-lasting application of GLU. Even in high-discharging units, the response never changed its sign. The trend to a depression of the response to 5-HT in function of the background discharge was observed when either the enhancement of firing occurred spontaneously and it was induced by an application of GLU, no significant difference (F-test) being found between the two cases. It is concluded that serotoninergic afferents can exert a strong control upon the vestibular neurons when the background activity is depressed, and only a weak influence when the neuronal firing is enhanced by other excitatory afferents. It remains to verify whether the type of interference observed between GLU and 5-HT is specific or can be also detected between 5-HT and other excitatory neuromediators.