Chitin and chitosan from mushroom (Agaricus bisporus) using deep eutectic solvents.

In this study, chitin was isolated from a mushroom (Agaricus bisporus) using deep eutectic solvents, choline chloride: acetic acid (CCAA), choline chloride:lactic acid (CCLA) and choline chloride:glycerol (CCG). According to the results, three DES systems were also useful for the isolation of chitin from mushrooms. The deproteinization efficiency was 84.25 %. The degree of deacetylation of chitin isolated by microwave-assisted extraction using CCAA was 69 %. This result was promising to produce chitosan in a one-step, base-free process using deep eutectic solvents. FTIR, XRD, SEM and XPS were used to analyse the physicochemical properties of the chitin.

A colorimetric immunoassay for the detection of human vascular endothelial growth factor 165 (VEGF165) based on anti-VEGF-iron oxide nanoparticle conjugation.

Vascular endothelial growth factor (VEGF) is an indispensable element in many physiological processes, while alterations in its level in the circulating system are signs of pathology-associated diseases. Therefore, its precise and selective detection is critical for clinical applications to monitor the progression of the pathology. In this study, an optical immunoassay biosensor was developed as a model study for detecting recombinant VEGF165. The VEGF165 sample was purified from recombinant Kluyveromyces lactis GG799 yeast cells. Indirect ELISA was used during the detection, wherein iron oxide nanoparticles (FeNPs) were utilized to obtain optical signals. The FeNPs were synthesized in the presence of lactose p-amino benzoic acid (LpAB). VEGF165 antibody was conjugated to the LpAB-FeNPs through EDC/NHS chemistry to convert the iron oxide nanoparticles into VEGF165 specific probes. The specificity of the prepared system was tested in the presence of potential serum-based interferents (i.e., glucose, urea, insulin, C-reactive protein, and serum amyloid A), and validation studies were performed in a simulated serum sample. The proposed immunoassay showed a wide detection range (0.5 to 100 ng/mL) with a detection limit of 0.29 ng/mL. These results show that the developed assay could offer a sensitive, simple, specific, reliable, and high-throughput detection platform that can be used in the clinical diagnostics of VEGF.

Production of flavor compounds from rice bran by yeasts metabolisms of Kluyveromyces marxianus and Debaryomyces hansenii.

The aim of this study was to evaluate the biosynthesis of flavor compounds from rice bran by fermentation facilitated by Kluyveromyces marxianus and Debaryomyces hansenii. The growth of both yeasts was assessed by specific growth rates and doubling time. The biosynthesis of flavor compounds was evaluated by gas chromatography-olfactometry (GC-O), gas chromatography-mass spectrometry (GC-MS), and Spectrum™ sensory analysis. The specific growth rate (µ) and doubling time (td) of K. marxianus was calculated as 0.16/h and 4.21h, respectively, whereas that of D. hansenii was determined as 0.13/h and 5.33h, respectively. K. marxianus and D. hansenii produced significant levels of higher alcohols and acetate esters from rice bran. Results showed that K. marxianus can produce 827.27 µg/kg of isoamyl alcohol, 169.77 µg/kg of phenyl ethyl alcohol, and 216.08 µg/kg of phenyl ethyl acetate after 24-h batch fermentation. A significant amount of isovaleric acid was also synthesized by K. marxianus (4013 µg/kg) after the batch fermentation of 96 h. 415.64 µg/kg of isoamyl alcohol and 135.77 µg/kg of phenyl ethyl acetate was determined in rice bran fermented by D. hansenii after 24-h fermentation. Fermented cereals and rose were the characteristic flavor descriptors of the fermented rice bran samples. Rose flavor in fermented rice bran samples was found to be associated with phenyl ethyl alcohol, phenyl ethyl acetate, isoamyl acetate, and guaiacol. Thus, the findings of this study demonstrate that the valorization of rice bran can be achieved with the production of natural flavor compounds by yeast metabolism.

A review on the potential uses of deep eutectic solvents in chitin and chitosan related processes.

Ionic liquids (ILs) as well as closely related deep eutectic solvents (DESs) have been the most promising discoveries in the "Green Chemistry" world in recent years as types of solvents. Deep eutectic solvents are received as potential alternatives to the conventional organic solvents with other advantages such as renewability, reusability, biodegradability, nontoxicity, large-scale availability, having very low vapour pressure, low flammability and easy in preparation. Chitin is the second most abundant biopolymers in the nature that can be extracted from many aquatic and terrestrial organisms also some of the microorganisms. Chitin and its derivatives have applications in many fields such as medical, food-beverages, agriculture and cosmetics. This review reports the major contributions of DESs in chitin and chitosan related studies and summarizes recent advances in some applications such as extraction media, bio-film fabrication, nanomaterial preparation, chitosan methylation, chitin dissolution, composite material preparation.

The effect of medium and light wavelength towards Stichococcus bacillaris fatty acid production and composition.

Introduction of novel species will highlight technical feasibility of microalgae-based biofuels for commercial applications. This paper reports the effect of culture medium and light wavelength on biomass and fatty acid production of S. bacillaris which holds some advantages as short life cycle, easy cultivation, high lipid content, diversity of fatty acids and stability under harsh environmental conditions. The results displayed that, soil extract (SE) greatly enhance growth rate of cultures. Maximum biomass and lipid productivity were achieved in TAP medium as 81 mg/L·day, 19.44 mg/L·day; respectively. Light wavelength didn't significantly change growth kinetics but played a critical role on chlorophyll-a accumulation. C14:0, C16:0 and C18:0 fatty acids were abundant which are suitable for biodiesel conversion. Interestingly, blue and red light increased longer chain fatty acids content. These results indicated that; S. bacillaris holds potential for further development of biodiesel production and feasibility of algal biodiesel for fundamental and applied sciences.

Optimization of chitosanase production by Bacillus mojavensis EGE-B-5.2i.

Maximum production of industrially important enzymes such as chitosanases through media optimization still holds foremost interest. The present study was conducted to improve chitosanase activity of an indigenous strain identified as Bacillus mojavensis. Initially, carbon and nitrogen sources were optimized by one-variable-at-a-time approach. Further, fermentation medium was optimized using Plackett-Burman (PB) and central composite designs (CCD). PB verified soluble starch (SS), colloidal chitosan (CC) peptone, and NaCl as most significant variables affecting chitosanase production. CCD results predicted the optimum concentrations of SS, CC, peptone, and NaCl as 7.8, 7.0, 6.5, and 2.7 g L-1 , respectively to achieve maximum chitosanase activity (21.1 U ml-1 ). Discovery of the novel optimal medium has improved chitosanase production by B. mojavensis up-to 9.5 folds. Lastly, 18.6 U ml-1 chitosanase activity was achieved in stirred tank bioreactor using optimal medium, which is quite satisfactory to proclaim this strain as a potential candidate to provide commercial chitosanase.

Modification of chitosan-bead support materials with L-lysine and L-asparagine for α-amylase immobilization.

Maltose syrups have got wide-range utilizations in a variety of applications from bakery to drug-development. α-Amylases are among the most widely utilized industrial enzymes due to their high specificity in production of maltose syrup from starch. However, enzymes are not stable in ex vivo conditions towards alteration in pH, temperature, and such other parameters as high salt concentrations and impurities, where immobilization is required to advance the stability of the enzyme with which approach the requirement of isolation of the enzyme from media is eliminated as well. In this study, Termamyl® α-amylase was immobilized on the none-modified chitosan beads (NMCB), L-lysine-modified chitosan beads (LMCB), and L-asparagine-modified chitosan beads (AMCB) to assess effects of the support material on optimum conditions and kinetic parameters of the α-amylase activity in production of maltose from starch. Immobilization on NMCB, LMCB, and AMCB puts a strong influence on optimum pH, optimum temperature, stability, and kinetic parameters of α-amylase. Modification of chitosan beads with L-lysine and L-asparagine dramatically altered the overall immobilization yield, and enzyme's response to pH and temperature variations and the kinetic parameters. AMCB provided the best immobilization yield (49%), while LMCB only improved the yield by 2% from 22 to 24%.

Investigation of in vitro digestibility of dietary microalga Chlorella vulgaris and cyanobacterium Spirulina platensis as a nutritional supplement.

Microalgal proteins are promising sources for functional nutrition and a sustainable candidate for nutraceutical formulations. They also gain importance due to emerging focus on a healthy nutrition and increase in the number of chronic diseases. In this study, dried dietary species of microalga, Chlorella vulgaris, and cyanobacterium Spirulina platensis were hydrolyzed with pancreatin enzyme to obtain protein hydrolysates. The hydrolysis yield of biomass was 55.1 ± 0.1 and 64.8 ± 3.6% for C. vulgaris and S. platensis; respectively. Digestibility, as an indicator for dietary utilization, was also investigated. In vitro protein digestibility (IVPD) values depicted that cell wall structure due to the taxonomical differences affected both hydrolysis and digestibility yield of the crude biomass (p < 0.05). Epithelial cells (Vero) maintained their viability around 70%, even in relatively higher concentrations of hydrolysates in the culture. The protein hydrolysates showed no any antimicrobial activities. This study clearly shows that the conventional protein sources in nutraceutical formulations such as soy, whey, and fish proteins can be replaced by enzymatic hydrolysates of microalgae, which shows elevated digestibility values as a sustainable and reliable source.

Production of flavor compounds from olive mill waste by Rhizopus oryzae and Candida tropicalis

Abstract The purpose of this study was to investigate the production of flavor compounds from olive mill waste by microbial fermentation of Rhizopus oryzae and Candida tropicalis. Olive mill waste fermentations were performed in shake and bioreactor cultures. Production of flavor compounds from olive mill waste was followed by Gas Chromatography–Mass spectrometry, Gas chromatography- olfactometry and Spectrum Sensory Analysis ®. As a result, 1.73-log and 3.23-log cfu/mL increases were observed in the microbial populations of R. oryzae and C. tropicalis during shake cultures, respectively. C. tropicalis can produce a higher concentration of d-limonene from olive mill waste than R. oryzae in shake cultures. The concentration of d-limonene was determined as 185.56 and 249.54 µg/kg in the fermented olive mill waste by R. oryzae and C. tropicalis in shake cultures respectively. In contrast, R. oryzae can produce a higher concentration of d-limonene (87.73 µg/kg) d-limonene than C. tropicalis (11.95 µg/kg) in bioreactor cultures. Based on sensory analysis, unripe olive, wet towel, sweet aromatic, fermented aromas were determined at high intensity in olive mill waste fermented with R. oryzae meanwhile olive mill waste fermented with C. tropicalis had only a high intensity of unripe olive and oily aroma.

Production of flavor compounds from olive mill waste by Rhizopus oryzae and Candida tropicalis.

The purpose of this study was to investigate the production of flavor compounds from olive mill waste by microbial fermentation of Rhizopus oryzae and Candida tropicalis. Olive mill waste fermentations were performed in shake and bioreactor cultures. Production of flavor compounds from olive mill waste was followed by Gas Chromatography-Mass spectrometry, Gas chromatography- olfactometry and Spectrum Sensory Analysis®. As a result, 1.73-log and 3.23-log cfu/mL increases were observed in the microbial populations of R. oryzae and C. tropicalis during shake cultures, respectively. C. tropicalis can produce a higher concentration of d-limonene from olive mill waste than R. oryzae in shake cultures. The concentration of d-limonene was determined as 185.56 and 249.54µg/kg in the fermented olive mill waste by R. oryzae and C. tropicalis in shake cultures respectively. In contrast, R. oryzae can produce a higher concentration of d-limonene (87.73µg/kg) d-limonene than C. tropicalis (11.95µg/kg) in bioreactor cultures. Based on sensory analysis, unripe olive, wet towel, sweet aromatic, fermented aromas were determined at high intensity in olive mill waste fermented with R. oryzae meanwhile olive mill waste fermented with C. tropicalis had only a high intensity of unripe olive and oily aroma.

Characterization of products from hydrothermal carbonization of orange pomace including anaerobic digestibility of process liquor.

In this study, the effect of the temperature and reaction time on hydrothermal carbonization (HTC) of orange pomace was investigated. In addition, a set of anaerobic batch tests were performed to determine the resulting biogas and methane potential of the spent liquor. Hydrochar yields followed a decreasing trend with the increasing temperature, whereas reaction time had no considerably effect on the yield. The evolution of the H/C and O/C atomic ratios from the raw material to the hydrochars suggested that dehydration reactions prevail during HTC. The hydrochars tended to become enriched in Ca, Mg and P minerals by increasing HTC temperature. The heavy metal contents in hydrochars were found below limits and no PAH compound was detected. Anaerobic digestion tests showed that the aqueous phase from HTC can be used as feedstocks for biogas production.

Bioflavour production from tomato and pepper pomaces by Kluyveromyces marxianus and Debaryomyces hansenii.

Bioflavours are called natural flavour and/or fragrance compounds which are produced using metabolic pathway of the microorganism and/or plant cells or their enzyme systems with bioengineering approaches. The aim of this study was to investigate bioflavour production from tomato and red pepper pomaces by Kluyveromyces marxianus and Debaryomyces hansenii. Obtained specific growth rates of K. marxianus and D. hansenii in tomato pomace were 0.081/h and 0.177/h, respectively. The bioflavour profile differed between the yeasts. Both yeasts can produce esters and alcohols such as phenyl ethyl alcohol, isoamyl alcohol, isoamyl acetate, phenyl ethyl acetate and isovaleric acid. "Tarhana" and "rose" were descriptive flavour terms for tomato and pepper pomaces fermented by K. marxianus, respectively. Tomato pomace fermented by D. hansenii had the most intense "green bean" flavour while "fermented vegetable" and "storage/yeast" were defined as characteristic flavour terms for pepper pomaces fermented by D. hansenii.

Effect of operating conditions in production of diagnostic Salmonella Enteritidis O-antigen-specific monoclonal antibody in different bioreactor systems.

In this study, different cultivation systems such as roller bottles (RB), 5-L stirred-tank bioreactor (STR), and disposable bioreactors were used to cultivate hybridoma for lab-scale production of Salmonella Enteritidis O-antigen-specific monoclonal antibody (MAb). Hybridoma cell line was cultivated in either serum-containing or serum-free medium (SFM) culture conditions. In STR, MAb production scaled up to 4 L, and production capabilities of the cells were also evaluated in different featured production systems. Moreover, the growth parameters of the cells in all production systems such as glucose consumption, lactate and ammonia production, and also MAb productivities were determined. Collected supernatants from the reactors were concentrated by a cross-flow filtration system. In conclusion, cells were not adapted to SFM in RB and STR. Therefore, less MAb titer in both STR and RB systems with SFM was observed compared to the cultures containing fetal bovine serum-supplemented medium. A higher MAb titer was gained in the membrane-aerated system compared to those in STR and RB. Although the highest MAb titer was obtained in the static membrane bioreactor system, the highest productivity was obtained in STR operated in semicontinuous mode with overlay aeration.

Enhanced growth and lipid accumulation by a new Ettlia texensis isolate under optimized photoheterotrophic condition.

A green microalgae, named as Ettlia texensis was obtained from local freshwater in Turkey. The effects of autotrophic, photoheterotrophic and heterotrophic cultivations on biomass and lipid production were studied. Searching the preferences of the carbon and nitrogen source revealed that this strain could grow photoheterotrophically well with glucose and yeast extract. In the optimized medium, the highest biomass productivity and total lipid content achieved were 0.97 g/L d and 26% of dry weight basis, respectively. Moreover, the major fatty acid methyl esters were C16:0; C18:1; C18:2 and C18:3. In a scale-up attempt, productions were accomplished in a 3 L stirred tank bioreactor. The final biomass and lipid productivities obtained in bioreactor with 250 rpm agitation rate were 0.92 g/L d and 322 mg/L d, respectively. The biochemical compositions were monitored simultaneously by the FTIR spectroscopy during the production in bioreactor. E. texensis could be potent candidate for commercial production in the bioreactor photoheterotrophically.

Purification of alginate and feasible production of monoclonal antibodies by the alginate-immobilized hybridoma cells.

Alginate has an extensive usage in the immobilization of many cell types. Although they have high biocompatibility, commercial alginates contain various degrees of contaminants such as polyphenols, endotoxins and proteins. Thus, these alginates show cytotoxicity against sensitive cell types such as hybridoma cells. In the studies so far, owing to this fact, commercially purchased high-priced ultrapure alginates have been used in the immobilization of hybridoma cells for monoclonal antibody production. However in this study, as a novelty, low-priced commercial alginate was purified, and then the cultivation of alginate-immobilized hybridoma cells was performed for feasible monoclonal antibody production. Low-priced commercial alginate was purified with a profitability ratio of 40%. Then, an optimized immobilization procedure was conducted effectively by using the purified alginate. During more than 25 days of cultivation, serum concentration was kept low, and approximately 2 times greater monoclonal antibody production was achieved, in comparison with its free suspended counterpart. The results showed that the efficiency of monoclonal antibody production via alginate-immobilized hybridoma cultivation can be increased by performing a proved in-house purification method. By shedding light on the efficiency of the in-house purification method, the results also indicated a feasible way of monoclonal antibody production.

Alginate as an immobilization material for MAb production via encapsulated hybridoma cells.

Alginate has been widely used in various applications since its first extraction. What makes this biopolymer useful is its high biocompatibility and humid gelation conditions. Both of these features bring it into prominence as an ideal immobilization material. However, there are some complicated aspects of cell immobilization using alginate biopolymers. This review discusses and clarifies these crucial points, using as an example the bioprocessing of highly fragile cells (hybridoma cells). The review focuses on the cultivation and production of alginate encapsulated cells.

Cocultivation of Lactococcus lactis and Teredinobacter turnirae for biological chitin extraction from prawn waste.

In this study, mainly biological treatment of prawn waste for chitin production was investigated. Lactic acid and protease fermentations were applied to extract chitin from prawn waste in the presence of various glucose concentrations. The results obtained were also compared with those of chemical method which was consisted of first mineral removal and then protein removal sequence. Different strategies were applied using lactic acid producing bacterium, Lactococcus lactis, and a protease producer, marine bacterium Teredinobacter turnirae. Both bacteria were first cultivated individually and then cofermented. In their individual cultivation, L. lactis removed the inorganic materials efficiently, while T. turnirae performed better in deproteinization process. Cofermentation of both bacteria was also conducted using three different protocols. The highest process yield (95.5%) was obtained when T. turnirae was first inoculated. Although the extraction of chitin by biological treatment was incomplete compared to the chemical method, the biological treatment employed here could still be considered as an alternative method in a more environmentally benign approach.