Optimisation of solid state fermentation of potato peel for the production of cellulolytic enzymes

This paper analyses the effects of water content, temperature and time on the kinetic activity of cellulolytic enzymes produced during the solid state fermentation of potato peel, using Aspergillus niger. Three main analytical steps – analysis of variance, regression analysis and plotting of response surface – were performed to obtain an optimum condition for enzymatic activity. The statistical results indicated that the best activity time for enzyme CMCase (carboxymethylcellulase) is 82.88 h, with water content of 51.48% and temperature of 29.46 °C; for FPase (filter paperase), the best activity time is 80.62 h, water content of 50.19% and temperature at 30.00 °C; for xylanase, time is 81.92 h, water content is 50.72% and temperature is 28.85 °C. Pareto charts have shown that all variables were significant in enzymatic activity for CMCase and xylanase. On the other hand, FPase shows that time and temperature have significant effect for this response variable.     

Improved biomass saccharification by Trichoderma reesei through heterologous expression of lacA gene from Trametes sp. AH28-2

The Trametes sp. AH28-2 laccase gene lacA fused to cellobiohydrolase I signal peptide coding sequence was heterologously expressed in T. reesei. The lacA cDNA was under the control of the Aspergillus nidulans glyceraldehyde-3-phosphate dehydrogenase promoter. Native PAGE analysis indicated that two transformants, L8 and L38, were able to secrete recombinant laccase A, and their laccase activities corresponding to ABTS oxidation reached 3.62 IUml(-1) and 1.50 IUml(-1) respectively. Most of the characteristics of the recombinant laccase were similar to those of the native enzyme. Reducing sugar yields of L8 and L38 obtained from saccharification of corn residue by crude enzyme increased by 31.3% and 71.6% respectively compared to the host strain. These results indicated that the engineering strains developed in this work could be potentially used for laccase production and tailoring cellulase properties with laccase proteins through genetic manipulation would be a feasible strategy to improve saccharification efficiency of biomass by cellulase preparation.     

Effects of food formulation and thermal processing on flavones in celery and chamomile

Flavones isolated from celery varied in their stability and susceptibility to deglycosylation during thermal processing at pH 3, 5, or 7. Apigenin 7-O-apiosylglucoside was converted to apigenin 7-O-glucoside when heated at pH 3 and 100 °C. Apigenin 7-O-glucoside showed little conversion or degradation at any pH after 5 h at 100 °C. Apigenin, luteolin, and chrysoeriol were most stable at pH 3 but progressively degraded at pH 5 or 7. Chamomile and celery were used to test the effects of glycosidase-rich foods and thermal processing on the stability of flavone glycosides. Apigenin 7-O-glucoside in chamomile extract was readily converted to apigenin aglycone after combination with almond, flax seed, or chickpea flour. Apigenin 7-O-apiosylglucoside in celery leaves was resistant to conversion by β-glucosidase-rich ingredients, but was converted to apigenin 7-O-glucoside at pH 2.7 when processed at 100 °C for 90 min and could then be further deglycosylated when mixed with almond or flax seed. Thus, combinations of acid hydrolysis and glycosidase enzymes in almond and flax seed were most effective for developing a flavone-rich, high aglycone food ingredient from celery.     

Lactobacillus pentosus dominates spontaneous fermentation of Italian table olives

Culture-dependent and -independent approaches were applied to identify the bacterial species involved in Italian table olive fermentation. Bacterial identification showed that Lactobacillus pentosus was the dominant species although the presence of Lactobacillus plantarum, Lactobacillus casei, Enterococcus durans, Lactobacillus fermentum and Lactobacillus helveticus was observed. Rep-PCR allowed to obtain strain-specific profiles and to establish a correlation with table olive environment. PCR-DGGE (Denaturing Gradient Gel Electrophoresis) confirmed the heterogeneity of bacterial community structure in fermented table olives as well as the prevalence of L. pentosus. The strains were characterized on the basis of technological properties (NaCl tolerance, β-glucosidase activity and the ability to grow in synthetic brine and in presence of 1 g/100 mL oleuropein). L. pentosus showed a high capacity of adaptation to the different conditions characterizing the olive ecosystem. This species showed the highest percentage of strains able to grow in presence of 10 g/100 mL NaCl, oleuropein and in the synthetic brine. Moreover, all the strains belonging to L. pentosus and L. plantarum species showed a β-glucosidase activity. This study allowed both to identify the main species and strains associated to Italian table olives and to obtain a lactic acid bacteria collection to apply as starter culture in the process of olive fermentation.     

Effect of lyoprotectants on β-glucosidase activity and viability of Bifidobacterium infantis after freeze-drying and storage in milk and low pH juices

The benefit of disaccharide protectants for maintaining viability and β-glucosidase activity of Bifidobacterium infantis UV16PR during freeze-drying and storage in different food matrices was investigated. Protectants used were cellobiose, lactose, sucrose and trehalose.     

Microbial transformation of tectoridin from Pueraria flos by Lactobacillus and bifidobacteria

Tectoridin could be hydrolyzed to tectorigenin by β-glucosidase-producing intestinal bacteria. In this study, nine strains of Lactobacillus and bifidobacteria were screened for high levels of β-glucosidase activity. We investigated their ability to transform tectoridin from Pueraria flos to tectorigenin. Lactobacillus reuteri DSM20016 showed the highest cell-envelope associated β-glucosidase activity, whereas the intracellular β-glucosidase activity from Bifidobacterium adolescentis ATCC15703 was higher than the other screened bacterial strains. L. reuteri DSM20016, Lactobacillus rhamnosus GGB41031 and B. adolescentis ATCC15703 showed high bioconversion rate of tectoridin. L. reuteri DSM20016 showed the highest bioconversion efficiency of tectoridin, 100% tectoridin was hydrolyzed and there was an approximate 185-fold increase in the concentration of tectorigenin after 24 h. The present study suggests that L. reuteri DSM20016, L. rhamnosus GGB41031 and B. adolescentis ATCC15703 have great potential for converting tectoridin from Pueraria flos to more bioactive tectorigenin.     

Production of trehalose by intramolecular transglucosylation of maltose catalysed by a new enzyme from Thermus thermophilus HB-8

Thermus thermophilus HB-8 is a source of trehalose synthase (GTase), which catalyses conversion of maltose into trehalose. Specific activity of maltose transglucosylation by cell-free extracts of the bacteria was about 0.1 U mg⁻¹ protein and precipitation at 28% saturation of ammonium sulphate caused 3.5-fold enzyme purification. The optimum temperature for conversion of maltose into trehalose was 65 °C with about 27% of maximum activity at 85 °C. The highest GTase productivity was achieved at cultivation temperature over 60 °C and at NaCl concentration range of 0.1–0.5% (w/v). However, larger concentrations of sodium chloride in the growth media caused a remarkable decrease of GTase synthesis. The results, of ammonium sulphate fractionation and activity towards maltotriose (0.028 U mg⁻¹), maltotetraose (0.16 U mg⁻¹) and GlcαpNp (0.27 U mg⁻¹), show that trehalose synthase and α-glucosidase activities reside in separate protein fractions of cell-free extracts from T. thermophilus cells.     

α-l-Arabinofuranosidase 3 from Penicillium purpurogenum (ABF3): Potential application in the enhancement of wine flavour and heterologous expression of the enzyme

An α-l-arabinofuranosidase (ABF3) from Penicillium purpurogenum was purified and its possible biotechnological application in the enhancement of wine flavour combined with P. purpurogenum β-glucosidase was studied. A must from Muscat of Alexandria was used to isolate the glycosides. The total monosaccharide (glucose, arabinose and xylose) levels in the glycosides were determined after acid hydrolysis, and were compared with the result of enzymatic hydrolysis. These results were analogous to those obtained in similar experiments using a commercial preparation, thus suggesting that the enzyme from P. purpurogenum may prove useful in this particular application. This prompted us to express the enzyme heterologously. The abf3 gene was thus expressed in Pichia pastoris. The recombinant enzyme was purified and it shows the same properties of the native ABF3 (substrate specificity, kinetic constants, pH and temperature optima and antibody cross-reactivity).     

Process development for mycelial growth and polysaccharide production in Tricholoma matsutake liquid culture

In this study, the effects of agitation and aeration on mycelial growth and exo-polysaccharide production were examined in batch cultures of Tricholoma matsutake. Agitation was varied from 100 to 300 rpm and aeration was varied from 0.5 to 1.5 vvm. Mycelial growth was 21.87 g/l at 150 rpm, and exo-polysaccharide production was 8.79 g/l at 1.5 vvm. When we analyzed the polysaccharide extractions from the cultured mycelium and the culture broth of T. matsutake, 1.4 g of crude polysaccharide was found per 100 g of dried weight in the cultured mycelium, and 1.47 g/l of polysaccharides was found in the culture broth. In addition, the amounts of β-Glucan in the soluble polysaccharide fractions of the cultured mycelium and culture broth were 75.4% and 83.6%, respectively. The cultured mycelium and the culture broth contained a higher amount of β-Glucan than that of the fruiting body.     

Ethanol production from non-pretreated napiergrass through a simultaneous saccharification and fermentation process followed by a pentose fermentation with Escherichia coli KO11

Efficient ethanol production from lignocellulosic napiergrass (Pennisetum purpureum Schumach) was examined by the combination of the simultaneous saccharification and fermentation (SSF) with commercial cellulase and Saccharomyces cerevisiae NBRC 2044 and subsequent pentose fermentation (PF) by Escherichia coli KO11. Under the optimized conditions, the combination of the SSF and PF processes resulted in the production of 144 mg g(-1) of ethanol from the non-pretreated napiergrass powder. The ethanol yield was 44.2% of the theoretical yield based on the hexose (37.5 g) and pentose (26.5 g) derived from 100 g of dry powdered napiergrass.     

Surface enhanced Raman scattering (SERS) with biopolymer encapsulated silver nanosubstrates for rapid detection of foodborne pathogens

A biopolymer encapsulated with silver nanoparticles was prepared using silver nitrate, polyvinyl alcohol (PVA) solution, and trisodium citrate. It was deposited on a mica sheet to use as SERS substrate. Fresh cultures of Salmonella Typhimurium, Escherichia coli, Staphylococcus aureus and Listeria innocua were washed from chicken rinse and suspended in 10 ml of sterile deionized water. Approximately 5 μl of the bacterial suspensions was placed on the substrate individually and exposed to 785 nm HeNe laser excitation. SERS spectral data were recorded over the Raman shift between 400 and 1800 cm^− 1 from 15 different spots on the substrate for each sample; and three replicates were done on each bacteria type. Principal component analysis (PCA) model was developed to classify foodborne bacteria types. PC1 identified 96% of the variation among the given bacteria specimen, and PC2 identified 3%, resulted in a total of 99% classification accuracy. Soft Independent Modeling of Class Analogies (SIMCA) of validation set gave an overall correct classification of 97%. Comparison of the SERS spectra of different types of gram-negative and gram-positive bacteria indicated that all of them have similar cell walls and cell membrane structures. Conversely, major differences were noted around the nucleic acid and amino acid structure information between 1200 cm^− 1 and 1700 cm^− 1 and at the finger print region between 400 cm^− 1 and 700 cm^− 1. Silver biopolymer nanoparticle substrate could be a promising SERS tool for pathogen detection. Also this study indicates that SERS technology could be used for reliable and rapid detection and classification of food borne pathogens.     

Antimicrobial and physical properties of edible chitosan films enhanced by lactoperoxidase system

Effects of lactoperoxidase system (LPOS) incorporated directly into chitosan films at different concentrations (0.5, 1 and 1.5%) were studied. Films obtained were tested on the inhibition of phytopathogenic strains such as Xanthomonas campestris pv. Mangifera indica, Colletotrichum gloeosporioides (C. 64, C. 4612 and C. 62) and Lasiodiplodia theobromae ngr 05A. Water vapor permeability and mechanical properties of films with LPOS and/or not iodine were also studied. Antibacterial effect obtained by disc diameter technique indicated that chitosan concentration at 1 and 1.5% (w/w) incorporated with LPOS and/or not iodine inhibited higher X. campestris pv. M. indica than chitosan film alone or at low concentrated of 0.5% incorporated by LPOS. The antimicrobial technique using puncture gave very good information on the antifungal effect and on the variability in susceptibility of strains of fungi. C. gloeosporioides C64 and L. theobromae were inhibited completely by 1 and 1.5% chitosan incorporated by LPOS contained or not iodine, while C. gloeosporioides C4612 was sensitive to the presence of iodine and C62 were resistant strains. The properties of chitosan films were not significantly changed by the incorporation of the enzyme system.