Optimization of enzymatic hydrolysis and fermentation conditions for improved bioethanol production from potato peel residues

The aim of this work was the optimization of the enzyme hydrolysis of potato peel residues (PPR) for bioethanol production. The process included a pretreatment step followed by an enzyme hydrolysis using crude enzyme system composed of cellulase, amylase and hemicellulase, produced by a mixed culture of Aspergillus niger and Trichoderma reesei. Hydrothermal, alkali and acid pretreatments were considered with regards to the enhancement of enzyme hydrolysis of potato peel residues. The obtained results showed that hydrothermal pretreatment lead to a higher enzyme hydrolysis yield compared to both acid and alkali pretreatments. Enzyme hydrolysis was also optimized for parameters such as temperature, pH, substrate loading and surfactant loading using a response surface methodology. Under optimized conditions, 77 g L^?1 of reducing sugars were obtained. Yeast fermentation of the released reducing sugars led to an ethanol titer of 30 g L^?1 after supplementation of the culture medium with ammonium sulfate. Moreover, a comparative study between acid and enzyme hydrolysis of potato peel residues was investigated. Results showed that enzyme hydrolysis offers higher yield of bioethanol production than acid hydrolysis. These results highlight the potential of second generation bioethanol production from potato peel residues treated with onsite produced hydrolytic enzymes. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:397–406, 2017     

A novel simple and rapid PCR-based site-directed mutagenesis method

Site-directed mutagenesis (SDM) is a powerful tool for exploring protein structure and function, and several procedures adjusted to specific purposes are still being developed. Herein we describe a straightforward and efficient method with versatile applications for introducing site-specific alterations in any deoxyribonucleic acid (DNA) sequence cloned in a plasmidic expression vector. In this polymerase chain reaction (PCR)-based SDM method, forward and reverse primers are used to amplify the plasmid containing the sequence of interest. The primers are designed so that the desired modifications are introduced at the 5′ end of one of the primers, whereas the other primer starts with the nucleotide at position (−1) of the one to be modified. The PCR is carried out using Pfu DNA polymerase. The blunt-ended PCR-generated DNA fragment is self-ligated and used to transform Escherichia coli. Mutant clones are screened by colony hybridization using the mutagenic primer as probe and the presence of the mutation is confirmed by direct DNA sequencing. This procedure was used efficiently to introduce substitutions, deletions, and insertions in the DNA sequences coding for a recombinant form (scFv) of antibody 107 specific of the human CR3 molecule, the rat α integrin CD11b A-domain and the human CD8β cloned in pPICZαB, pGEX-2T, and CDM8 expression vectors, respectively.     

Calcium-induced conformational changes in the amino-terminal half of gelsolin

Gelsolin is an actin-binding protein that is regulated by the occupancy of multiple calcium-binding sites. We have studied calcium induced conformational changes in the G1-2 and G1-3 sub-domains, and report the binding affinities for the three type II sites. A new probe for G3 has been produced and a K(d) of 5 microM has been measured for calcium in the context of G1-3. The two halves of gelsolin, G1-3 and G4-6 bind weakly with or without calcium, suggesting that once separated by apoptotic proteolysis, G1-3 and G4-6 remain apart allowing G1-3 to sever actin in a calcium free manner.     

Perception of cuticular hydrocarbons by the olfactory organs in Periplaneta americana (L.) (Insecta: Dictyoptera)

Despite the importance of cuticular hydrocarbons (CHs) in insect chemical communication, direct proof that they are detected and recognized by insects by contact or by olfactory receptors are rare. In Periplaneta americana, CHs induce aggregation. The aim of our study was to investigate how CHs are detected by P. americana antennae. Using solid phase microextraction and gas chromatography, the three main CHs of the species profile were identified in the volatiles emitted by these insects. Gas chromatography coupled to electroantennography recordings demonstrated that the antennae responded to these three CHs. Furthermore, CHs had an attraction effect in Y-olfactometer bioassays when presented at high concentrations. As CHs can be perceived by P. americana, at least from a short distance, they could play a role in attracting conspecifics during aggregation processes, in addition to inducing aggregation when direct contact is possible.     

Rare taxa and dark microbial matter: novel bioactive actinobacteria abound in Atacama Desert soils

An “in house” taxonomic approach to drug discovery led to the isolation of diverse actinobacteria from hyper-arid, extreme hyper-arid and very high altitude Atacama Desert soils. A high proportion of the isolates were assigned to novel taxa, with many showing activity in standard antimicrobial plug assays. The application of more advanced taxonomic and screening strategies showed that strains classified as novel species of Lentzea and Streptomyces synthesised new specialised metabolites thereby underpinning the premise that the extreme abiotic conditions in the Atacama Desert favour the development of a unique actinobacterial diversity which is the basis of novel chemistry. Complementary metagenomic analyses showed that the soils encompassed an astonishing degree of actinobacterial ‘dark matter’, while rank-abundance analyses showed them to be highly diverse habitats mainly composed of rare taxa that have not been recovered using culture-dependent methods. The implications of these pioneering studies on future bioprospecting campaigns are discussed.     

Effect of heated naringenin on immunomodulatory properties and cellular antioxidant activity

Naringenin is one of the most popular flavonoids derived from citrus. It has been reported to be an effective anti-inflammatory compound. Citrus fruit may be used raw, cooked, stewed, or boiled. The present study was conducted to investigate the effect of thermal processes on naringenin in its immunomodulatory and cellular antioxidant activities. The effects of flavonoids on B and T cell proliferation were assessed on splenocytes stimulated or not with mitogens. However, their effects on cytotoxic T lymphocyte (CTL) and natural killer (NK) activities were assessed in splenocytes co-incubated with target cells. The amount of nitric oxide production and the lysosomal enzyme activity were evaluated in vitro on mouse peritoneal macrophages. Cellular antioxidant activity in splenocytes and macrophages was determined by measuring the fluorescence of the dichlorofluorescin (DCF). Our findings revealed that naringenin induces B cell proliferation and enhances NK activity. The highest concentration of native naringenin exhibits a significant proliferation of T cells, induces CTL activity, and inhibits cellular oxidation in macrophages. Conversely, it was observed that when heat-processed, naringenin improves the cellular antioxidant activity in splenocytes, increases the cytotoxic activity of NK cells, and suppresses the cytotoxicity of T cells. However, heat treatment maintains the anti-inflammatory potency of naringenin.