Selective utilization of fructose to glucose by Candida magnoliae , an erythritol producer

Candida magnoliae isolated from honeycomb is an industrially important yeast with high erythritol-producing ability. Erythritol has been used as functional sugar substitute for various foods. In order to analyze the physiological properties of C. magnoliae, a study on sugar utilization pattern was carried out. The fermentation kinetics of glucose and fructose revealed that C. magnoliae has the discrepancy in glucose and fructose utilization when it produces erythritol. In contrast to most yeasts, C. magnoliae showed preference for fructose to glucose as a carbon source, deserving the designation of fructophilic yeast. Such a peculiar pattern of sugar utilization in C. magnoliae seems to be related to the evolutionary environment.     

Effect of media composition and growth conditions on production of β-glucosidase by Aspergillus niger C-6

The hydrolytic activity of fungal originated β-glucosidase is exploited in several biotechnological processes to increase the rate and extent of saccharification of several cellulosic materials by hydrolyzing the cellobiose which inhibits cellulases. In a previous presentation, we reported the screening and liquid fermentation with Aspergillus niger, strain C-6 for β-glucosidase production at shake flask cultures in a basal culture medium with mineral salts, corn syrup liquor, and different waste lignocellulosic materials as the sole carbon source obtaining the maximum enzymatic activity after 5–6 d of 8.5 IU/mL using native sugar cane bagasse. In this work we describe the evaluation of fermentation conditions: growth temperature, medium composition, and pH, also the agitation and aeration effects for β-glucosidase production under submerged culture using a culture media with corn syrup liquor (CSL) and native sugar cane bagasse pith as the sole carbon source in a laboratory fermenter. The maximum enzyme titer of 7.2 IU/mL was obtained within 3 d of fermentation. This indicates that β-glucosidase productivity by Aspergillus niger C-6 is function of culture conditions, principally temperature, pH, culture medium conditions, and the oxygen supply given in the bioreactor. Results obtained suggest that this strain is a potential microorganism that can reach a major level of enzyme production and also for enzyme characterization.     

Analysis of Antioxidant Capacity and Antimicrobial Properties of Selected Polish Grape Vinegars Obtained by Spontaneous Fermentation

Nowadays, products of natural origin with health-promoting properties are increasingly more common. Research shows that fruit vinegars can be a source of compounds with antioxidant activity. Research on the total antioxidant capacity, total phenolic content, and antimicrobial properties against Staphylococcus aureus, Escherichia coli, and Candida albicans of grape vinegars were conducted. Moreover, gas chromatography was used to measure acetic acid content in the vinegars. The research material consisted of vinegars produced from five different grape varieties. For each variety, two variants were prepared: with and without the addition of sugar in the fermentation process. The highest antimicrobial activity against all micro-organisms was observed in vinegar produced from Solaris grapes with added sugar. The highest polyphenol content was observed in vinegar produced from the Prior grape variety with added sugar and the highest total antioxidant capacity is the Johanniter grape variety with added sugar. The vinegars examined in this study differed, depending on grape variety, in terms of antimicrobial properties, antioxidant capacity, total phenolic content, as well as acetic acid content. Sugar addition caused significant differences in the antioxidant capacity of vinegar samples.     

Black Liquor Decolorization by Selected White-Rot Fungi

Five different strains of white-rot fungi have been tested for their ability to decolorize black liquor on plates and on solid-state fermentation using vermiculite as the solid inert support. Since the high salt concentration inhibited the growth of all fungi, the black liquor was dialyzed against distilled water prior to use. A preliminary step on plates was carried out to qualitatively determine the capacity of the fungal strains for black liquor decolorization. Out of the five fungi studied, Phanerochaete sordida, Pycnoporus sanguineus, and Trametes elegans exhibited the more conspicuous decolorization halos in malt extract medium, while the decolorization by all the strains was not evident when a defined culture medium was used. Cultures on solid-state fermentation using vermiculite as solid support were also tested, the liquid phase was malt extract or glucose-based medium and supplemented with different black liquor concentrations. Decolorization of black liquor was largely affected by the fungal strain, the concentration of black liquor, and the carbon source. The percentage of color removal ranged from 6.14% to 91.86% depending on the fungal strain and culture conditions. Maximal decolorization was observed in malt extract cultures after 60 cultivation days. Interestingly, decolorization in malt extract medium increased with increasing black liquor concentration. The highest decolorization value was achieved by Steccherinum sp. which reduced up to 91.86% the color of the black liquor in malt extract medium; this percentage is equivalent to 5.2 g L⁻¹ of decolorized black liquor, the highest value reported to date. Traditional technologies used for the treatment of black liquor are not always effective and may not to be an environmentally friendly process. Vermiculite–white-rot fungi systems are presented in this work as a promising efficient alternative for the treatment of black liquor.     

Succinic acid production from acid hydrolysate of corn fiber by Actinobacillus succinogenes

Dilute acid hydrolysate of corn fiber was used as carbon source for the production of succinic acid by Actinobacillus succinogenes NJ113. The optimized hydrolysis conditions were obtained by orthogonal experiments. When corn fiber particles were of 20 mesh in size and treated with 1.0% sulfuric acid at 121 °C for 2 h, the total sugar yield could reach 63.3%. It was found that CaCO₃ neutralization combined with activated carbon adsorption was an effective method to remove fermentation inhibitors especially furfural that presented in the acid hydrolysate of corn fiber. Only 5.2% of the total sugar was lost, while 91.9% of furfural was removed. The yield of succinic acid was higher than 72.0% with the detoxified corn fiber hydrolysate as the carbon source in anaerobic bottles or 7.5 L fermentor cultures. It was proved that the corn fiber hydrolysate could be an alternative to glucose for the production of succinic acid by A. succinogenes NJ113.     

Screening of potential antibiotic action of cellulolytic fungi

Twenty different strains of filamentous fungi were initially selected for evaluation of cellulolytic activity using a single test in a simple mineral salts culture medium with filter paper as the only carbon source. Those fungi strains that were capable of completely breaking the filter paper strip within 4–8 d were assayed also for antimicrobial action, using Staphyloccocus a ureus ATCC 6538P according to the so-called agar piece method. We screened three different strains with both capacities: the production of cellulolytic activity and antibiotic action. The experimental results suggest that the fungi Pinicillium sp. FOPCO1, Aspergillus sp. F0Q001, and Cephalosporium sp. F03800 have both capabilities because they grew rapidly on cellulose as the only carbon source and were able to produce an area of growth inhibition in S. aureus of approx 2.04, 1.57, and 2.39 cm, respectively, on agar plates using the agar piece method. Subsequently, the antibiotic production obtained with those cellulolytic strains was evaluated by submerged fermentation at the flask level, in a simple culture medium containing lactose without biosynthesis precursor, obtaining 3670, 2830, and 4060 antibiotic units/mL, referred to as penicillin G, whereas for cellulolytic activity, the results were 1.34, 1.81 and 0.57 FPU/mL, respectively.     

Production and characterization of pullulan from beet molasses using a nonpigmented strain of Aureobasidium pullulans in batch culture

The production of pullulan from beet molasses by a pigment-free strain of Aureobasidium pullulans on shake-flask culture was investigated. Combined pretreatment of molasses with sulfuric acid and activated carbon to remove potential fermentation inhibitors present in molasses resulted in a maximum pullulan concentration of 24 g/L, a biomass dry wt of 14 g/L, a pullulan yield of 52.5%, and a sugar utilization of 92% with optimum fermentation conditions (initial sugar concentration of 50 g/L and initial pH of 7.0). The addition of other nutrients as carbon and nitrogen supplements (olive oil, ammonium sulfate, yeast extract) did not further improve the production of the exopolysaccharides. Structural characterization of the isolated polysaccharides from the fermentation broths by ¹³C-nuclear magnetic resonance spectroscopy and pullulanase digestion combined with size-exclusion chromatography confirmed the identity of pullulan and the homogeneity (>93% dry basis) of the elaborated polysaccharides by the microorganism. Using multiangle laser light scattering and refractive index detectors in conjunction with high-performance size-exclusion chromatography molecular size distributions and estimates of the molecular weight (M _w =2.1−4.1×10⁵), root mean square of the radius of gyration (R _g =30−38 nm), and polydispersity index (M _w /M _n =1.4−2.4) were obtained. The fermentation products of molasses pretreated with sulfuric acid and/or activated carbon were more homogeneous and free of contaminating proteins. In the concentration range of 2.8−10.0 (w/v), the solution’s rheologic behavior of the isolated pullulans was almost Newtonian (within 1 and 1200 s⁻¹ at 20°C); a slight shear thinning was observed at 10.0 (w/v) for the high molecular weight samples. Overall, beet molasses pretreated with sulfuric acid and activated carbon appears as an attractive fermentation medium for the production of pullulan by A. pullulans.     

Synthesis and Antimicrobial Activity of New Substituted 5‐(Pyridine‐3‐yl)‐1,3,4‐Thiadiazoles and Their Sugar Derivatives

New substituted 5‐(pyridine‐3‐yl)‐1,3,4‐thiadiazoles, their sugar hydrazones and acyclic C‐nucleoside analogs as well as the corresponding thioglycoside derivatives were synthesized. The synthesized compounds were tested for their antimicrobial activity against Escherichia coli, Bacillus subtilis, Staph aureus, Aspergillus niger, and Candida albicans The obtained results indicated that most of tested compounds exhibited moderate to high antimicrobial activity while few compounds were found to exhibit little or no activity against the tested microorganisms.     

Biotechnological Production of Phenyllactic Acid and Biosurfactants from Trimming Vine Shoot Hydrolyzates by Microbial Coculture Fermentation

Coculture fermentations show advantages for producing food additives from agroindustrial wastes, considering that different specified microbial strains are combined to improve the consumption of mixed sugars obtained by hydrolysis. This technology dovetails with both the growing interest of consumers towards the use of natural food additives and with stricter legislations and concern in developed countries towards the management of wastes. The use of this technology allows valorization of both cellulosic and hemicellulosic fractions of trimming vine shoots for the production of lactic acid (LA), phenyllactic acid (PLA), and biosurfactants (BS). This work compares the study of the potential of hemicellulosic and cellulosic fractions of trimming vine shoots as cheaper and renewable carbon sources for PLA and BS production by independent or coculture fermentations. The highest LA and PLA concentrations, 43.0 g/L and 1.58 mM, respectively, were obtained after 144 h during the fermentation of hemicellulosic sugars and simultaneous saccharification and fermentation (SSF) carried out by cocultures of Lactobacillus plantarum and Lactobacillus pentosus. Additionally, cell-bond BS decreased the surface tension (ST) in 17.2 U; meanwhile, cell-free supernatants (CFS) showed antimicrobial activity against Salmonella enterica and Listeria monocytogenes with inhibition halos of 12.1?±?0.6 mm and 11.5?±?0.9 mm, respectively.     

Effect of surfactants on carbon monoxide fermentations byButyribacterium methylotrophicum

Butyribacterium methylotrophicum has been grown on carbon monoxide as its carbon and energy source in the presence of various surfactants that are capable of forming microbubble dispersions for the screening of surfactants for use in microbubble-sparged synthesis gas fermentations. In the range of 0–3 times the critical micelle concentration, the presence of Tween surfactants was not significantly inhibitory to growth, final cell density, and fermentation stoichiometry, although some of the Brij surfactants caused significant inhibition. As the batch fermentations entered the stationary phase both the pH and the ratio of acetate to butyrate decreased.     

Inulin-containing biomass for ethanol production

The use of stalks instead of tubers as a source of carbohydrates for ethanol production has been investigated. The inulin present in the stalks of Jerusalem artichoke was extracted with water and the effect of solid-liquid ratio, temperature, and acid addition was studied and optimized in order to attain a high-fructose fermentable extract. The maximum extraction efficiency (corresponding to 35 g/L) of soluble sugars was obtained at 1/6 solidliquid ratio. Fermentations of hydrolyzed extracts by baker's yeast and direct fermentation by an inulinease activity yeast were also performed and the potential to use this feedstock for bioethanol production assessed. The results show that the carbohydrates derived from Jerusalem artichoke stalks can be converted efficiently to ethanol by acidic hydrolysis followed by fermentation with Saccharomyces cerevisiae or by direct fermentation of inulin using Kluyveromyces marxianus strains. In this last case about 30 h to complete fermentation was required in comparison with 8–9 h obtained in experiments with S. cerevisiae growth on acid extracted juices.     

Rheological Analysis of Biopolymer Produced by Aureobasidium Pullulans in Different Sources of Nitrogen

Summary: In this study, the rheological behaviors of media fermented by two Aureobasidium pullulans strains (IOC 3467 and IOC 3011) were evaluated in different nutritional conditions. The media consisted of crystal sugar (sucrose), as the main carbon source, and different nitrogen sources (ammonium sulfate, sodium nitrate, ammonium nitrate, urea or residual brewery yeast - RBY). Viscosity measurements were performed on cell-free supernatants, from 48-hour fermentation assays, at 25 ^°C. Shear rates ranged between 0.1000 and 500 s⁻¹. All samples showed pseudoplastic behavior. Nevertheless, the viscosimetric profile of each one varied according to the nitrogen source, its concentration and the strain used. The maximum viscosity of 0.06 Pa.s. was achieved at 15.6 s⁻¹ for IOC 3011 strain grown on RBY as nitrogen source.     

Enhanced production of xylanase from locally isolated fungal strain using agro-industrial residues under solid-state fermentation

This study is related to the isolation of fungal strain for xylanase production using agro-industrial residues. Forty fungal strains with xylanolytic potential were isolated by using xylan agar plates and quantitatively screened in solid-state fermentation. Of all the tested isolates, the strain showing highest ability to produce xylanase was assigned the code Aspergillus niger LCBT-14. For the enhanced production of the enzyme, five different fermentation media were evaluated. Out of all media, M4 containing wheat bran gave maximum enzyme production. Effect of different variables including incubation time, temperature, pH, carbon and nitrogen sources has been investigated. The optimum enzyme production was obtained after 72 h at 30°C and pH 4. Glucose as a carbon source while ammonium sulphate and yeast extract as nitrogen sources gave maximum xylanase production (946 U/mL/min). This study was successful in producing xylanase by A. niger LCBT-14 economically by utilising cheap indigenous substrate.     

Fermentations of pectin-rich biomass with recombinant bacteria to produce fuel ethanol

Pectin-rich residues from sugar beet processing contain significant carbohydrates and insignificant amounts of lignin. Beet pulp was evaluated for conversion toethanol using recombinant bacteria as biocatalysts. Hydrolysis of pectin-rich residues followed by ethanolic fermentations by yeasts has not been productive because galacturonic acid and arabinose are not ferm entable toethanol by these organisms. The three recombinant bacteria evaluated in this study, Escherichia coli strain KO11, Klebsiella oxytoca strain P2, and Erwinia chrysanthemi EC 16 pLOI 555, ferment carbohydrates in beet pulp with varying efficiencies. E. coli KO11 is able to convert pure galactu ronic acid to ethanol with minimal acetate production. Using an enzyme loading of 10.5 filter paper un its of cellulase, 120.4 polygalactu ronase units of pectinase, and 6.4 g of cellobiase (per gram of dry wt sugar beet pulp), with substrate addition after 24 h of fermentation, 40 g of ethanol/L was produced. Other recombinants exhibited lower ethanol yields with increases in acetate and succinate production.     

Preparation of some novel thiazolidinones,imidazolinones, and azetidinone bearing pyridine and pyrimidine moieties with antimicrobial activity

By aiming to design new antimicrobial agents, we prepared new series of thiazolidin-4-ones (12a–d), imidazolin-4-ones (13a–d), and azetidin-2-ones (14a–d), having pyridine and pyrimidine moieties. Chemical structures of these derivatives were elucidated by the use of spectral and elemental analyses. All the new substituted pyridopyrimidines were subjected to in vitro antimicrobial testing by estimating the zone of inhibition toward Bacillus subtilis and Staphylococcus aureus, as examples of bacterial species, in addition to Aspergillus flavus and Candida albicans, as examples of fungal species. The results of antimicrobial testing detected that all the screened derivatives displayed antibacterial effect; especially azetidin-2-one derivative, ( 14c ), was the most active one. Regarding the antifungal potential, only thiazolidinone derivatives, 12a and 12c, and the imidazolinone, 13c, displayed inhibitory activity toward Aspergillus flavus, while all the tested compounds, 12a–d , 13a–d, and 14a–d, except 14a, produced inhibitory potential toward Candida albicans. Docking studies of the most active antimicrobial agents, 12c, 13c, and 14c, within GLN-6-P, recorded good scores with several binding interactions with the active site.     

Cell-Free Supernatants Obtained from Fermentation of Cheese Whey Hydrolyzates and Phenylpyruvic Acid by Lactobacillus plantarum as a Source of Antimicrobial Compounds, Bacteriocins, and Natural Aromas

Cheese whey hydrolyzates supplemented with phenylpyruvic acid (PPA) and commercial nutrients can be efficiently metabolized by Lactobacillus plantarum CECT-221 to biosynthesize some compounds with attractive applications in the food market. The main metabolites of cell-free extracts were antimicrobial compounds such as phenyllactic acid (PLA) and lactic acid (LA). The production of PLA by L. plantarum CECT-221 was evaluated in the Man–Rogosa–Sharpe broth supplemented with two biosynthetic precursors: phenylalanine or PPA. Using 30.5 mM PPA, the microorganism increased sevenfold the concentration of PLA producing 16.4 mM PLA in 46 h. A concentration of 40 mM PPA was a threshold to avoid substrate inhibition. The biosynthesis of whey hydrolyzates as a carbon source was enhanced by fed-batch fermentation of PPA; the average productivity of PLA increased up to 45.4?±?3.02 mM after 120 h with a product yield of 0.244 mM mM^?1; meanwhile, LA reached 26.1?±?1.3 g L^?1 with a product yield of 0.72 g g^?1. Cell-free fed-batch extracts charged in wells showed bacteriocin activity with halos of 7.49?±?1.44 mm in plates inoculated with Carnobacterium piscicola and antimicrobial activity against Staphylococcus aureus (11.54?±?1.14 mm), Pseudomonas aeruginosa (10.17?±?2.46 mm), Listeria monocytogenes (7.75?±?1.31 mm), and Salmonella enterica (3.60?±?1.52 mm). Additionally, the analysis of the volatile composition of the headspace of this cell-free extract revealed that L. plantarum is a potential producer for natural aromas, such as acetophenone, with high price in the market. This is the first report of PLA production from cheese whey and PPA. The extracts showed bacteriocin activity and potential to be applied as an antimicrobial in the elaboration of safer foods.     

Use of steam explosion liquor from sugar cane bagasse for lignin peroxidase production by Phanerochaete chrysosporium

The possibility of using two by-products of the sugar cane industry, molasses and bagasse steam explosion liquor (SEL), for lignin peroxidase (LiP) production by Phanerochaete chrysosporium was investigated. For comparison, the fungus was initially cultivated in synthetic media containing either glucose, sucrose, xylose, or xylan as sole carbon sources. The effect of veratryl alcohol (VA) was also investigated in relation to the enzyme activity levels. Results showed that sucrose was not metabolized by this fungus, which precluded the use of molasses as a carbon source. Glucose, xylose, and xylan promoted equivalent cell growth. Enzyme levels in the absence of VA were lower than 28 UI/L and in the presence of VA reached 109 IU/L with glucose and 85 IU/L with xylose or xylan. SEL was adequate for P. chrysosporium LiP production as LiP activity reached 90 IU/L. When VA was added to this medium, enzyme concentration increased to 155 IU/L.     

Cellulase Production by <Emphasis Type="Italic">Streptomyces viridobrunneus</Emphasis> SCPE-09 Using Lignocellulosic Biomass as Inducer Substrate

An actinomycete strain, isolated from a soil sample under a sugar cane plantation in Brazil and identified as Streptomyces viridobrunneus SCPE-09, was selected as a promising cellulolytic strain, and tested for its ability to produce cellulases from agro-industrial residues. Sugar cane bagasse or wheat bran was tested as carbon source, and corn steep liquor tested as nitrogen source. Different concentrations of carbon and nitrogen were tested using factorial design to identify optimal cellulose production. The results showed that media containing wheat bran 2.0% (w/v) and corn steep liquid 0.19% (w/v) lead to the highest production, 2.0 U mL⁻¹ of CMCase, obtained on the fifth day of fermentation. The pH and temperature profile showed optimal activity at pH 4.9 and 50°C. As for thermostability, endoglucanases were most tolerant at 50°C, retaining more than 80% of maximal activity even after 2 h of incubation. Zymogram analyses using supernatant from growth under optimized conditions revealed the presence of two CMCase bands with apparent molecular masses of 37 and 119 kDa. The combination of pH tolerance and CMCase production from agro-industrial residues by S. viridobrunneus SCPE-09 offers promise for future bioethanol biotechnologies.     

Cellulases and Hemicellulases from Endophytic <Emphasis Type="Italic">Acremonium</Emphasis> Species and Its Application on Sugarcane Bagasse Hydrolysis

The aim of this work was to have cellulase activity and hemicellulase activity screenings of endophyte Acremonium species (Acremonium zeae EA0802 and Acremonium sp. EA0810). Both fungi were cultivated in submerged culture (SC) containing l-arabinose, d-xylose, oat spelt xylan, sugarcane bagasse, or corn straw as carbon source. In solid-state fermentation, it was tested as carbon source sugarcane bagasse or corn straw. The highest FPase, endoglucanase, and xylanase activities were produced by Acremonium sp. EA0810 cultivated in SC containing sugarcane bagasse as a carbon source. The highest β-glucosidase activity was produced by Acremonium sp. EA0810 cultivated in SC using d-xylose as carbon source. A. zeae EA0802 has highest α-arabinofuranosidase and α-galactosidase activities in SC using xylan as a carbon source. FPase, endoglucanase, β-glucosidase, and xylanase from Acremonium sp. EA0810 has optimum pH and temperatures of 6.0, 55 °C; 5.0, 70 °C; 4.5, 60 °C; and 6.5, 50 °C, respectively. α-Arabinofuranosidase and α-galactosidase from A. zeae EA0802 has optimum pH and temperatures of 5.0, 60 °C and 4.5, 45 °C, respectively. It was analyzed the application of Acremonium sp. EA0810 to hydrolyze sugarcane bagasse, and it was achieved 63% of conversion into reducing sugar and 42% of conversion into glucose.     

Exploration of quinoxaline derivatives as antimicrobial and anticancer agents

Novel 2 and 3‐substituted quinoxaline derivatives were synthesized through various synthetic pathways, among which cyanoacetamide and cyanoacetohydrazide quinoxaline derivatives 4a‐c and 11a‐c , respectively, were synthesized. Furthermore, methoxy quinoxaline derivatives 3c and quinoxaline derivatives bearing substituted pyridines 6a,b , 12a,b , and 13a,b were designed to be synthesized. However, we have synthesized acrylohydrazide 5a,b and 7 /acrylamide derivatives, Schiff base analogues 14a‐f , pyrazole derivatives 15a‐e, amide derivatives 16a‐f , guanidine derivatives 16 g,h as well as, quinoxalin‐2‐methylallyl propionate derivative 14g . All the synthesized compounds were confirmed via spectral data and elemental analyses. Moreover, the newly synthesized compounds were evaluated for their antimicrobial activity (Gm +ve, Gm ?ve in comparison to Gentamycin a standard) and fungi (in comparison to Ketoconazole as a standard). Thus, compound 16b showed promising antimicrobial activity against B. subtilis, P. vulgaris, and S. mutants with values ranging from 20 to 27‐mm zone of inhibition. While compounds 5a , 14e,f, and 16a,c,d,g,h showed potent antimicrobial activity. Moreover, the National Cancer Institute (NCI) selected 20 compounds that were submitted for anticancer screening against 60 types of cancer cell lines. The most active compounds are 5b and 12a where compound 5b containing 2,4‐dichlorophenyl moiety at cyanoacetamide linkage of hydrazine quinoxaline backbone exerted significant growth inhibition activity against Leukemia MOLT‐4, Renal cancer UO‐31, and Breast cancer MCF‐7. In addition, compound 12a having 4,6‐diaminopyridinone side chain at position‐3 of quinoxaline nucleus exhibited remarkable anticancer activity against renal cancer UO‐31.