Search results for: yeast Saccharomyces cerevisiae

Authors: Faranak Beigmohammadi, Seyed Hadi Peighambardoust, Seyed Jamaledin Peighambardoust

Abstract:

This study undertakes to investigate the ability of different kinds of nanocomposite films made of cloisite-30B with different percentages of silver and copper oxide nanoparticles incorporated into a low-density polyethylene (LDPE) polymeric matrix by a melt mixing method in order to inhibit the growth of microorganism in soy burger. The number of surviving cell of the total count was decreased by 3.61 log and mold and yeast diminished by 2.01 log after 8 weeks storage at 18 ± 0.5°C below zero, whilst pure LDPE did not has any antimicrobial effect. A composition of 1.3 % cloisite 30B-Ag and 2.7 % cloisite 30B-CuO for total count and 0 % cloisite 30B-Ag and 4 % cloisite 30B-CuO for yeast & mold gave optimum points in combined design test in Design Expert 7.1.5. Suitable microbial models were suggested for retarding above microorganisms growth in soy burger. To validation of optimum point, the difference between the optimum point of nanocomposite film and its repeat was not significant (p<0.05) by one-way ANOVA analysis using SPSS 17.0 software, while the difference was significant for pure film. Migration of metallic nanoparticles into a food stimulant was within the accepted safe level.

Keywords: modeling, nanocomposite film, packaging, soy burger

Procedia PDF Downloads 276

Authors: Lukas Stiburek, Jana Cesnekova, Josef Houstek, Jiri Zeman

Abstract:

Cellular function depends on mitochondrial function and integrity that is therefore maintained by several classes of proteins possessing chaperone and/or proteolytic activities. In this work, we focused on characterization of LACE1 (lactation elevated 1) function in mitochondrial protein homeostasis maintenance. LACE1 is the human homologue of yeast mitochondrial Afg1 ATPase, a member of SEC18-NSF, PAS1, CDC48-VCP, TBP family. Yeast Afg1 was shown to be involved in mitochondrial complex IV biogenesis, and based on its similarity with CDC48 (p97/VCP) it was suggested to facilitate extraction of polytopic membrane proteins. Here we show that LACE1, which is a mitochondrial integral membrane protein, exists as part of three complexes of approx. 140, 400 and 500 kDa and is essential for maintenance of fused mitochondrial reticulum and lamellar cristae morphology. Using affinity purification of LACE1-FLAG expressed in LACE1 knockdown background we show that the protein physically interacts with mitochondrial inner membrane protease YME1L. We further show that human LACE1 exhibits significant pro-apoptotic activity and that the protein is required for normal function of the mitochondrial respiratory chain. Thus, our work establishes LACE1 as a novel factor with the crucial role in mitochondrial homeostasis maintenance.

Keywords: LACE1, mitochondria, apoptosis, protease

Procedia PDF Downloads 290

Authors: Patarasuda Chaisupa, R. Clay Wright

Abstract:

The selection of appropriate biomolecular targets is a crucial aspect of biopharmaceutical development. The Auxin-Inducible Degron Degradation (AID) technology has demonstrated remarkable potential in efficiently and rapidly degrading target proteins, thereby enabling the identification and acquisition of drug targets. The AID system also offers a viable method to deplete specific proteins, particularly in cases where the degradation pathway has not been exploited or when the adaptation of proteins, including the cell environment, occurs to compensate for the mutation or gene knockout. In this study, we have engineered an improved AID system tailored to deplete proteins of interest. This AID construct combines the auxin-responsive E3 ubiquitin ligase binding domain, AFB2, and the substrate degron, IAA17, fused to the target genes. Essential genes of fungi with the lowest percent amino acid similarity to human and plant orthologs, according to the Basic Local Alignment Search Tool (BLAST), were cloned into the AID construct in S. cerevisiae (AID-tagged strains) using a modular yeast cloning toolkit for multipart assembly and direct genetic modification. Each E3 ubiquitin ligase and IAA17 degron was fused to a fluorescence protein, allowing for real-time monitoring of protein levels in response to different auxin doses via cytometry. Our AID system exhibited high sensitivity, with an EC50 value of 0.040 µM (SE = 0.016) for AFB2, enabling the specific promotion of IAA17::target protein degradation. Furthermore, we demonstrate how this improved AID system enhances quantitative functional studies of various proteins in fungi. The advancements made in auxin-inducible protein degradation in this study offer a powerful approach to investigating critical target protein viability in fungi, screening protein targets for drugs, and regulating intracellular protein abundance, thus revolutionizing the study of protein function underlying a diverse range of biological processes.

Keywords: synthetic biology, bioengineering, molecular biology, biotechnology

Procedia PDF Downloads 68

Authors: Vanessa De Carvalho, Chad MacPherson, Julien Tremblay, Julie Champagne, Stephanie-Anne Girard

Abstract:

Background: The interactions between bacteria and fungi in the human vaginal microbiome are fundamental to the concept of health and disease. The means by which the microbiota and mycobiota interact is still poorly understood and further studies are necessary to properly characterize this complex ecosystem. The aim of this study was to select a DNA extraction method capable of recovering high qualities of fungal and bacterial DNA from a single vaginal swab. Methods: 11 female volunteers ( ≥ 20 to < 55 years old) self-collected vaginal swabs in triplicates. Three commercial extraction kits: Masterpure Yeast Purification kit (Epicenter), PureLink™ Microbiome DNA Purification kit (Invitrogen), and Quick-DNA™ Fecal/Soil Microbe Miniprep kit (Zymo) were evaluated on the ability to recover fungal and bacterial DNA simultaneously. The extraction kits were compared on the basis of recovery, yield, purity, and the community richness of bacterial (16S rRNA - V3-V4 region) and fungal (ITS1) microbiota composition by Illumina MiSeq amplicon sequencing. Results: Recovery of bacterial DNA was achieved with all three kits while fungal DNA was only consistently recovered with Masterpure Yeast Purification kit (yield and purity). Overall, all kits displayed similar microbiota profiles for the top 20 OTUs; however, Quick-DNA™ Fecal/Soil Microbe Miniprep kit (Zymo) showed more species richness than the other two kits. Conclusion: In the present study, Masterpure Yeast purification kit proved to be a good candidate for purification of high quality fungal and bacterial DNA simultaneously. These findings have potential benefits that could be applied in future vaginal microbiome research. Whilst the use of a single extraction method would lessen the burden of multiple swab sampling, decrease laboratory workload and off-set costs associated with multiple DNA extractions, thoughtful consideration must be taken when selecting an extraction kit depending on the desired downstream application.

Keywords: bacterial vaginosis, DNA extraction, microbiota, mycobiota, vagina, vulvovaginal candidiasis, women’s health

Procedia PDF Downloads 180

Authors: Esa Abiso Godana, Qiya Yang, Kaili Wang, Zhang Hongyin, Xiaoyun Zhang, Lina Zhao

Abstract:

Blue mold decay caused by Penicillium expansum is among the recent identified diseases of grapes (Vitis vinifera). The increasing concern about use of chemical substance and pesticide in postharvest fruit push the trends of research toward biocontrol strategies which are more sustainable and ecofriendly. In this study, we determined the biocontrol efficacy of Pichia anomala alone and supplemented with 1% chitosan in the grapefruit against blue mold disease caused by P. expansum. The result showed that 1% chitosan better enhances the biocontrol efficacy P. anomala. Chitosan (1% w/v) also improved the number of population of P. anomala in grape wounds, surface and on nutrient yeast dextrose broth (NYDB). P. anomala supplemented with 1% w/v chitosan significantly reduced the disease incidence, lesion diameter and natural decay of grapefruits without affecting the fruit quality as compared to the control. The scanned electron microscope (SEM) concisely illustrates how the high number of yeast cells on the wounds reduced the growth of P. expansum. P. anomala alone or P. anomala supplemented with 1% w/v chitosan are presented as a potential biocontrol alternative against the postharvest blue mold of grapefruit.

Keywords: biocontrol, Pichia anomala, chitosan, Penicillium expansum, grape

Procedia PDF Downloads 88

Authors: Divya Choudhary, Hariprasad P., S. N. Naik

Abstract:

This study examines a low-alcohol nutra-beverage made with shatavari, a plant commonly used in traditional medicine. During fermentation, the addition of a specific strain of yeast affected the beverage's properties, including its pH level, yeast count, ethanol content, and antioxidant, phenolic, and flavonoid levels. We also analyzed the beverage's storage and shelf life. Despite its bitter taste, the low alcohol content of the beverage made it enjoyable to drink and visually appealing. Our analysis showed that the optimal time for fermentation was between the 14th and 21st day when the beverage had ideal levels of sugar, organic acids, and vitamins. The final product contained fructose and citric acid but not succinic, pyruvic, lactic, or acetic acids. It also contained vitamins B2, B1, B12, and B9. During the shelf life analysis, we observed changes in the beverage's pH, TSS, and cfu levels, as well as its antioxidant activity. We also identified volatile (GC-MS) and non-volatile compounds (LC-MS/MS) in the fermented product, some of which were already present in the Shatavari root. The highest yield of product contained the maximum concentration of antioxidant compounds, which depended on both the pH and the microorganisms' physiological status. Overall, our study provides insight into the properties and potential health benefits of this Nutra-beverage.

Keywords: antioxidants, fermentation, volatile compounds, acetonin, 1-butanol, non-volatile compounds, Shatavarin V, IX, kaempferol

Procedia PDF Downloads 45

Authors: Young-Ran Song, Byeong-Uk Lim, Sang-Ho Baik

Abstract:

This study was initiated in order to develop a method for soy sauce fermentation at low salt concentrations without decreasing quality. Soy sauce was fermented with the fermentation starter (meju) and different salt contents (8-14%, w/v) by inoculating two strains or not, in which Torulaspora delbrueckii and Pichia guilliermondii strains having different abilities to induce sterilizing effects or enhance flavor production were used. As the results, there were microbial and biochemical differences among prepared soy sauce. First, Staphylococcus and Enterococcus spp. in addition to Bacillus genus that is the most important bacteria in Korean fermented soy product were detected by salt reduction. However, application of yeast starters can inhibit the undesirable bacterial growth. Moreover, PCA bi-plots of major principal components on various biochemical parameters (final pH, total acidity, soluble sugar, reducing sugar, ethanol and 32 volatile flavor compounds) were drawn to demonstrate the physicochemical differences and similarities among the samples. It was confirmed that the soy sauce samples produced with different salt concentrations were clearly different since salt reduction induced low contents of acids, alcohols and esters with higher acidity. However despite low salt concentration, combining two different yeasts appeared to have similar characteristics to the high salt-fermented soy sauce with elevated concentrations of ethanol, some alcohols, and most ketones, hence resulted in a balance of more complex and richer flavors with a flavor profile pattern identical to that of high-salt.

Keywords: Soy sauce, low salt, fermentation, yeast.

Procedia PDF Downloads 367

Authors: Elizabeth Loza-Valerdi, Victor Pardiñas-Rios, Arnulfo Pluma-Pluma, Andres Breton-Toral, Julio Cercado-Jaramillo

Abstract:

The purification processes for mixtures of isomeric monosaccharides using industrial chromatographic methods poses a serious technical challenge. Mixtures of 2 or 3 monosaccharides are difficult to separate by strictly physical or chemical techniques. Differential fermentation by microbial cultures is an increasingly interesting way of selective enrichment in a particular kind of monosaccharides when a mixture of them is present in the solution, and only one has economical value. Osmotolerant yeast cultures provide an interesting source of biocatalysts for the selective catabolism of monosaccharides in media containing high concentrations of total soluble sugars. A collection of 398 yeast strains has been obtained using endemic and unique sources of fruit juices, industrial syrups, honey, and other high sugar content substrates, either natural or man made, products and by-products from Mexico. The osmotolerance of the strains was assessed by plate assay both in glucose (20-40-60%w/w). Strains were classified according to their osmotolerance in low, medium or highly tolerant to high glucose concentrations. The purified cultures were tested by their ability to growth in a solid plate media or liquid media of Yeas Nitrogen Base (YNB), added with specific monosaccharides as sole carbon source (glucose, galactose, lactose and fructose). Selected strains were subsequently tested in fermentation experiments with mixtures of two monosaccharides (galactose/glucose and glucose/fructose). Their ability to grow and selectively catabolize one monosaccharide was evaluated. Growth, fermentation activity and products of metabolism were determined by plate counts, CO2 production, turbidity and chromatographic analysis by HPLC. Selective catabolism of one monosaccharide in liquid media containing two monosaccharides was confirmed for 8 strains. Ion Exchange chromatographic processes were used in production of high fructose or galactose syrup. Laboratory scale processes for the production of fructose or galactose enriched syrups is now feasible, with important applications in food (like high fructose syrup as edulcorant) and fermentation technology (for GOS production).

Keywords: osmotolerant yeasts, selective metabolism, fructose syrup, GOS

Procedia PDF Downloads 434

Authors: Hossein Rassi, Marjan Moradi Fard, Samaneh Niko

Abstract:

Human papillomavirus type 16 and 18 are closely associated with the development of human cervical carcinoma, which is one of the most common causes of cancer death in women worldwide. At present, HPV type 18 accounts for about 34 % of all HPV infections in Iran and the most promising vaccine against HPV infection is based on the L1 major capsid protein. The L1 protein of HPV18 has the capacity to self-assemble into capsomers or virus-like particles (VLPs) that are non-infectious, highly immunogenic and allowing their use in vaccine production. The methylotrophic yeast Pichia pastoris is an efficient and inexpensive expression system used to produce high levels of heterologous proteins. In this study we expressed HPV18 L1 VLPs in P. pastoris. The gene encoding the major capsid protein L1 of the high-risk HPV type 18 was isolated from Iranian patient by PCR and inserted into pTG19-T vector to obtain the recombinant expression vector pTG19-HPV18-L1. Then, the pTG19-HPV18-L1 was transformed into E. coli strain DH5α and the recombinant protein HPV18 L1 was expressed under IPTG induction in soluble form. The HPV18 L1 gene was excised from recombinant plasmid with XhoI and EcoRI enzymes and ligated into the yeast expression vector pPICZα linearized with the same enzymes, and transformed into P. pastoris. Induction and expression of HPV18 L1 protein was demonstrated by BMGY/BMMY and RT PCR. The parameters for induced cultivation for strain in P. pastoris KM71 with HPV16L1 were investigated in shaking flask cultures. After induced cultivation BMMY (pH 7.0) medium supplemented with methanol to a final concentration of 1.0% every 24 h at 37 degrees C for 96 h, the recombinant produced 78.6 mg/L of L1 protein. This work offers the possibility for the production of prophylactic vaccine for cervical carcinoma by P. pastoris for HPV-18 L1 gene. The VLP-based HPV vaccines can prevent persistent HPV18 infections and cervical cancer in Iran. The HPV-18 L1 gene was expressed successfully in E.coli, which provides necessary basis for preparing HPV-18 L1 vaccine in human. Also, HPV type 6 L1 proteins expressed in Pichia pastoris will facilitate the HPV vaccine development and structure-function study.

Keywords: Pichia pastoris, L1 virus-like particles, human papillomavirus type 18, biotechnology

Procedia PDF Downloads 390

Authors: Zoran Herceg, Višnja Stulić, Tomislava Vukušić, Anet Režek Jambrak

Abstract:

High voltage electrical discharge is a new technology used for inactivation of pathogen microorganisms. Pathogen yeasts can cause diseases in humans if they are ingested. Nowadays new technologies have become the focus of researching all over the world. Rhodotorula is known as yeast that can cause diseases in humans. The aim of this study was to examine whether the high voltage electrical discharge treatment generated in gas phase has an influence on yeast reduction and recovery of Rhodotorula spp 74 in pure culture. Rhodotorula spp. 74 was treated in 200 mL of model solution. Treatment time (5 and 10 min), frequency (60 and 90 Hz) and injected gas (air or argon 99,99%) were changed. Titanium high voltage needle was used as high voltage electrode (positive polarity) through which air or argon was injected at the gas flow of 0.6 L/min. Experimental design and statistical analyses were obtained by Statgraphics Centurion software (StatPoint Technologies, Inc., VA, USA). The best inactivation rate 1.7 log10 reduction was observed after the 10 min of treatment, frequency of 90 Hz and injected air. Also with a longer treatment time inactivation rate was higher. After the 24 h recovery of treated samples was observed. Therefore the further optimization of method is needed to understand the mechanism of yeasts inactivation and cells recovery after the treatment. Acknowledgements: The authors would like to acknowledge the support by Croatian Science Foundation and research project ‘Application of electrical discharge plasma for preservation of liquid foods’.

Keywords: rhodotorula spp. 74, electrical discharge plasma, inactivation, stress response

Procedia PDF Downloads 215

Authors: Parmjit S. Panesar, Rupinder Kaur, Ram S. Singh

Abstract:

Enzymes are the biocatalysts which catalyze the biochemical processes and thus have a wide variety of applications in the industrial sector. β-Galactosidase (E.C. 3.2.1.23) also known as lactase, is one of the prime enzymes, which has significant potential in the dairy and food processing industries. It has the capability to catalyze both the hydrolytic reaction for the production of lactose hydrolyzed milk and transgalactosylation reaction for the synthesis of prebiotics such as lactulose and galactooligosaccharides. These prebiotics have various nutritional and technological benefits. Although, the enzyme is naturally present in almonds, peaches, apricots and other variety of fruits and animals, the extraction of enzyme from these sources increases the cost of enzyme. Therefore, focus has been shifted towards the production of low cost enzyme from the microorganisms such as bacteria, yeast and fungi. As compared to yeast and bacteria, fungal β-galactosidase is generally preferred as being extracellular and thermostable in nature. Keeping the above in view, the present study was carried out for the isolation of the β-galactosidase producing fungal strain from the food as well as the agricultural wastes. A total of more than 100 fungal cultures were examined for their potential in enzyme production. All the fungal strains were screened using X-gal and IPTG as inducers in the modified Czapek Dox Agar medium. Among the various isolated fungal strains, the strain exhibiting the highest enzyme activity was chosen for further phenotypic and genotypic characterization. The strain was identified as Rhizomucor pusillus on the basis of 5.8s RNA gene sequencing data.

Keywords: beta-galactosidase, enzyme, fungal, isolation

Procedia PDF Downloads 228

Authors: M. Javanmard

Abstract:

This study was aimed at analyzing the effects of packaging (MAP) and preservation conditions on the packaged fresh walnut kernel quality. The central composite plan was used for evaluating the effect of oxygen (0–10%), carbon dioxide (0-10%), and temperature (4-26 °C) on qualitative characteristics of walnut kernels. Also, the response level technique was used to find the optimal conditions for interactive effects of factors, as well as estimating the best conditions of process using least amount of testing. Measured qualitative parameters were: peroxide index, color, decreased weight, mould and yeast counting test, and sensory evaluation. The results showed that the defined model for peroxide index, color, weight loss, and sensory evaluation is significant (p < 0.001), so that increase of temperature causes the peroxide value, color variation, and weight loss to increase and it reduces the overall acceptability of walnut kernels. An increase in oxygen percentage caused the color variation level and peroxide value to increase and resulted in lower overall acceptability of the walnuts. An increase in CO₂ percentage caused the peroxide value to decrease, but did not significantly affect other indices (p ≥ 0.05). Mould and yeast were not found in any samples. Optimal packaging conditions to achieve maximum quality of walnuts include: 1.46% oxygen, 10% carbon dioxide, and temperature of 4 °C.

Keywords: shelled walnut, MAP, quality, storage temperature

Procedia PDF Downloads 365

Authors: Hue Dinh, Binh Nguyen, Vivian Mendez, Phillip W. Taylor, Fleur Ponton

Abstract:

Better understanding of how parental diet affects offspring traits is an important ecological and evolutionary question. In this study, we explored how maternal diet influences offspring physiology and resistance to infection using Bactrocera tryoni (Q-fly) as a system model. Female Q-flies were fed one of six single diets varying in their yeast-to-sugar ratio yielding six protein-to-carbohydrate ratios. As controls, we used females that were given a choice between yeast and sugar. Males were reared on a choice diet and allowed to mate with females 14 days post-emergence. Results showed that while maternal diet does not influence offspring developmental time, it has a strong effect on larval body weight. Mother fed either high-protein or high-sugar diet produced larger progeny. By challenging offspring with the bacterium Serratia marcescens, we found that female offspring from mothers fed high-sugar diet survived better the infection compared to those from mothers fed low-sugar diet. In contrast, male offspring produced by mother fed high-protein diet showed better resistance to the infection compared to those produced by mother fed low-protein diet. These results suggested sex-dependent transgenerational effects of maternal nutrition on offspring physiology and immunity.

Keywords: bacterial infection, Bactrocera tryoni, maternal diet, offspring, Serretia marcescens

Procedia PDF Downloads 121

Authors: Zeynep Yilmazer Hitit, Patrick C. Hallenbeck

Abstract:

Fuel reserve requirements due to depletion of fossil fuels have increased interest in biohydrogen since the 1990’s. In fermentative hydrogen production, pure, mixed, and co-cultures can be used to produce hydrogen. Several previous studies have evaluated hydrogen production by pure cultures of Clostridium butyricum or Enterobacter aerogenes. Evaluating hydrogen production by co-culture of these microorganisms is an interestıng approach since E. aerogenes is a facultative microorganism with resistance to oxygen in contrast to the strict anaerobe C. butyricum, and therefore has the ability to maintain anaerobic conditions. It was found that using co-cultures of facultative E. aerogenes (as a reducing agent and H2 producer) and the obligate anaerobe C. butyricum for producing hydrogen increases the yield of hydrogen by about 50% compared to C. butyricum by itself. Also, using different types of microorganisms for hydrogen production eliminates the need to use expensive reducing agents. C. butyricum strain pre-cultured anaerobically at 37 0C for 15h by inoculating 100 mL of GP medium (pH 6.8) consisting of 1% glucose, 2% polypeptone, 0.2% KH2PO4, 0.05% yeast extract, 0.05% MgSO4. 7H2O and E. aerogenes strain was pre-cultured aerobically at 30 0C, 150 rpm for 9 h by inoculating 100 mL of TGY medium (pH 6.8), consisting of 0.1% glucose, 0.5% tryptone, 0.1% K2HPO4, 0.5% yeast extract. All duplicate batch experiments were conducted in 100 mL bottles with different inoculum ratios of Clostridium butyricum and Enterobater aerogenes (C:E) using 5x diluted rich media (GP) consisting of 2 g/L glucose, 4g/L polypeptone, 0.4 g/L KH2PO4, 0.1 g/L yeast extract, 0.1 MgSO4.7H2O. The range of inoculum ratio of C. butyricum to E. aerogenes were 2:1,4:1,8:1, 1:2,1:4, 1:8, 1:0, 0:1. Using glucose as a carbon source aided in the observation of microbial behavior as well as making the effect of inoculum ratio more evident. Nearly all the glucose in the medium was used to produce hydrogen, except at a 1:0 ratio of inoculum (i.e. containing only C. butyricum). Low glucose consumption leads to a higher hydrogen yield due to cumulative hydrogen production and consumption of glucose, but not as much as C:E, 8:1. The lowest hydrogen yield was achieved in 1:8 inoculum ratio of C:E, 71.9 mL, 1.007±0.01 mol H2/mol glucose and the highest cumulative hydrogen, hydrogen yield and dry cell weight were achieved in 8:1 inoculum ratio of C:E, 117.4 mL, 2.035±0.082 mol H2/mol glucose, 0.4 g/L respectively. In this study effect of inoculum ratio on dark fermentative biohydrogen production using C. butyricum and E. aerogenes was investigated. The maximum hydrogen yield of 2.035mol H2/mol glucose was obtained using 2g/L glucose, an initial pH of 6 and an inoculum ratio of C. butyricum to E. aerogenes of 8:1. Results showed that inoculum ratio is an important parameter on hydrogen production due to competition between the two microorganisms in using substrate for growth and production of by-products. The results presented here could be of great significance for further waste management studies using co-culture hydrogen production.

Keywords: biohydrogen, Clostridium butyricum, dark fermentation, Enterobacter aerogenes, inoculum ratio in biohydrogen production

Procedia PDF Downloads 214

Authors: Aliaa M. El-Borai, Ehab A. Beltagy, Eman E. Gadallah, Samy A. ElAssar

Abstract:

Bioremediation technology is now used for treatment instead of traditional metal removal methods. A strain was isolated from Marsa Alam, Red sea, Egypt showed high resistance to high lead concentration and was identified by the 16S rRNA gene sequencing technique as Halomonas sp. ES015. Medium optimization was carried out using Plackett-Burman design, and the most significant factors were yeast extract, casamino acid and inoculums size. The optimized media obtained by the statistical design raised the removal efficiency from 84% to 99% from initial concentration 250 ppm of lead. Moreover, Box-Behnken experimental design was applied to study the relationship between yeast extract concentration, casamino acid concentration and inoculums size. The optimized medium increased removal efficiency to 97% from initial concentration 500 ppm of lead. Immobilized Halomonas sp. ES015 cells on sponge cubes, using optimized medium in loop bioremediation column, showed relatively constant lead removal efficiency when reused six successive cycles over the range of time interval. Also metal removal efficiency was not affected by flow rate changes. Finally, the results of this research refer to the possibility of lead bioremediation by free or immobilized cells of Halomonas sp. ES015. Also, bioremediation can be done in batch cultures and semicontinuous cultures using column technology.

Keywords: bioremediation, lead, Box–Behnken, Halomonas sp. ES015, loop bioremediation, Plackett-Burman

Procedia PDF Downloads 174

Authors: T. Hamaishi, Y. Morinaga, H. Morita

Abstract:

Introduction: Rice consumption in Japan has decreased, and Japanese government has recommended use of rice flour in order to expand the consumption of rice. There are two major protein components present in flour, called gliadin and glutenin. Gluten forms when water is added to flour and is mixed. As mixing continues, glutenin interacts with gliadin to form viscoelastic matrix of gluten. Rice flour bread does not expand as much as wheat flour bread. Because rice flour is not included gluten, it cannot construct gluten network in the dough. In recent years, some food additives have been used for dough-improving agent in bread making, especially surfactants has effect in order to improve dough extensibility. Therefore, we focused to fatty acid salt which is one of anionic surfactants. Fatty acid salt is a salt consist of fatty acid and alkali, it is main components of soap. According to JECFA(FAO/WHO Joint Expert Committee on Food Additives), salts of Myristic(C14), Palmitic(C16) and Stearic(C18) could be used as food additive. They have been evaluated ADI was not specified. In this study, we investigated to improving bread-making properties of rice flour dough adding fatty acid salt. Materials and methods: The sample of fatty acid salt is myristic (C14) dissolved in KOH solution to a concentration of 350 mM and pH 10.5. Rice dough was consisted of 100 g of flour using rice flour and wheat gluten, 5 g of sugar, 1.7 g of salt, 1.7g of dry yeast, 80 mL of water and fatty acid salt. Mixing was performed for 500 times by using hand. The concentration of C14K in the dough was 10 % relative to flour weight. Amount of gluten in the dough was 20 %, 30 % relative to flour weight. Dough expansion ability test was performed to measure physical property of bread dough according to the methods of Baker’s Yeast by Japan Yeast Industry Association. In this test, 150 g of dough was filled from bottom of the cylinder and fermented at 30 °C，85 % humidity for 120 min on an incubator. The height of the expansion in the dough was measured and determined its expansion ability. Results and Conclusion: Expansion ability of rice dough with gluten content of 20 %, 30% showed 316 mL, 341 mL for 120 min. When C14K adding to the rice dough, dough expansion abilities were 314 mL, 368 mL for 120 min, there was no significant difference. Conventionally it has been known that the rice flour dough contain gluten of 20 %. The considerable improvement of dough expansion ability was achieved when added C14K to wheat flour. The experimental result shows that c14k adding to the rice dough with gluten content more than 20 % was not improving bread-making properties. In conclusion, rice bread made with gluten content more than 20 % without C14K has been suggested to contribute to the formation of the sufficient gluten network.

Keywords: expansion ability, fatty acid salt, gluten, rice flour dough

Procedia PDF Downloads 226

Authors: Shahram Naghizadeh Raeisi, Ali Alghooneh

Abstract:

The loss of curd cohesiveness and syneresis are two common problems in the ultrafiltered cheese industry. In this study, by using membrane technology and protein modification, a modified cheese was developed and its properties were compared with a control sample. In order to decrease the lactose content and adjust the protein, acidity, dry matter and milk minerals, a combination of ultrafiltration, nanofiltration and reverse osmosis technologies was employed. For protein modification, a two-stage chemical and enzymatic reaction was employed before and after ultrafiltration. The physicochemical and microstructural properties of the modified ultrafiltered cheese were compared with the control one. Results showed that the modified protein enhanced the functional properties of the final cheese significantly (pvalue< 0.05), even if the protein content was 50% lower than the control one. The modified cheese showed 21 ± 0.70, 18 ± 1.10 & 25±1.65% higher hardness, cohesiveness and water-holding capacity values, respectively, than the control sample. This behavior could be explained by the developed microstructure of the gel network. Furthermore, chemical-enzymatic modification of milk protein induced a significant change in the network parameter of the final cheese. In this way, the indices of network linkage strength, network linkage density, and time scale of junctions were 10.34 ± 0.52, 68.50 ± 2.10 & 82.21 ± 3.85% higher than the control sample, whereas the distance between adjacent linkages was 16.77 ± 1.10% lower than the control sample. These results were supported by the results of the textural analysis. A non-linear viscoelastic study showed a triangle waveform stress of the modified protein contained cheese, while the control sample showed rectangular waveform stress, which suggested a better sliceability of the modified cheese. Moreover, to study the shelf life of the products, the acidity, as well as molds and yeast population, were determined in 120 days. It’s worth mentioning that the lactose content of modified cheese was adjusted at 2.5% before fermentation, while the lactose of the control one was at 4.5%. The control sample showed 8 weeks shelf life, while the shelf life of the modified cheese was 18 weeks in the refrigerator. During 18 weeks, the acidity of modified and control samples increased from 82 ± 1.50 to 94 ± 2.20 °D and 88 ± 1.64 to 194 ± 5.10 °D, respectively. The mold and yeast populations, with time, followed the semicircular shape model (R2 = 0.92, R2adj = 0.89, RMSE = 1.25). Furthermore, the mold and yeast counts and their growth rate in the modified cheese were lower than those for control one; Aforementioned result could be explained by the shortage of the source of energy for the microorganism in the modified cheese. The lactose content of the modified sample was less than 0.2 ± 0.05% at the end of fermentation, while this was 3.7 ± 0.68% in the control sample.

Keywords: non-linear viscoelastic, protein modification, semicircular shape model, ultrafiltered cheese

Procedia PDF Downloads 58

Authors: Andre Pastori D'Aurea, Lauriston Bertelli Feranades, Luis Eduardo Ferreira, Leandro Dias Pinto, Fabiana Ayumi Shiozaki

Abstract:

The aim of this study was to improve the production of beef cattle on tropical pasture without harming this environment. On tropical pastures, cattle's live weight gain is lower than feedlot, and forage production is seasonable, changing from season to season. Thus, concerned with sustainable livestock production, the Premix Company has developed strategies to improve the production of beef cattle on tropical pasture to ensure sustainability of welfare and production. There are two important principles in this productivity system: 1) increase individual gains with use of better supplementation and 2) increase the productivity units with better forage quality like corn silage or other forms of forage conservations, actually used only in winter, and adding natural additives in the diet. This production system was applied from June 2017 to May 2018 in the Research Center of Premix Company, Patrocínio Paulista, São Paulo State, Brazil. The area used had 9 hectares of pasture of Brachiaria brizantha. 36 steers Nellore were evaluated for one year. The initial weight was 253 kg. The parameters used were daily average gain and gain per area. This indicated the corrections to be made and helped design future fertilization. In this case, we fertilized the pasture with 30 kg of nitrogen per animal divided into two parts. The diet was pasture and protein-energy supplements (0.4% of live weight). The supplement used was added with natural additive Fator P® – Premix Company). Fator P® is an additive composed by amino acids (lysine, methionine and tyrosine, 16400, 2980 and 3000 mg.kg-1 respectively), minerals, probiotics (Saccharomyces cerevisiae, 7 x 10E8 CFU.kg-1) and essential fatty acids (linoleic and oleic acids, 108.9 and 99g.kg-1 respectively). Due to seasonal changes, in the winter we supplemented the diet by increasing the offer of forage, supplementing with maize silage. It was offered 1% of live weight in silage corn and 0.4% of the live weight in protein-energetic supplements with additive Fator P ®. At the end of the period, the productivity was calculated by summing the individual gains for the area used. The average daily gain of the animals were 693 grams per day and was produced 1.005 kg /hectare/year. This production is about 8 times higher than the average of Brazilian meat national production. To succeed in this project, it is necessary to increase the gains per area, so it is necessary to increase the capacity per area. Pasture management is very important to the project's success because the dietary decisions were taken from the quantity and quality of the forage. We, therefore, recommend the use of animals in the growth phase because the response to supplementation is greater in that phase and we can allocate more animals per area. This system's carbon footprint reduces emissions by 61.2 percent compared to the Brazilian average. This beef cattle production system can be efficient and environmentally friendly to the natural. Another point is that bovines will benefit from their natural environment without competing or having an impact on human food production.

Keywords: cattle production, environment, pasture, sustainability

Procedia PDF Downloads 122

Authors: Thalía Moreno-García Malo, Santiago Torres-Ríos, María G. González-Cruz, María M. Hernández-Arroyo, Sergio R. Trejo-Estrada

Abstract:

Agave atrovirens is the main source of agave sap, the raw material for the production of pulque, an artisanal fermented beverage, traditional since prehispanic times in the highlands of central Mexico. Agave sap is rich in glucose, sucrose and fructooligosaccharides, and strongly differs from agave syrup from A. tequilana, which is mostly a high molecular weight fructan. Agave sap is converted into pulque by a highly diverse microbial community which includes bacteria, yeast and even filamentous fungi. The bacterial diversity has been recently studied. But the composition of consortia derived from directed enrichments differs sharply from the whole fermentative consortium. Using classical microbiology methods, and selective liquid and solid media formulations, either bacterial or fungal consortia were developed and analyzed. Bacterial consortia able to catabolize specific prebiotic saccharides were selected and preserved for future developments. Different media formulations, selective for bacterial genera such as Bifidobacterium, Lactobacillus, Pediococcus, Lactococcus and Enterococcus were also used. For yeast, specific media, osmotic pressure and unique carbon sources were used as selective agents. Results show that most groups are represented in the enrichment cultures; although very few are recoverable from the whole consortium in artisanal pulque. Diversity and abundance vary among consortia. Potential bacterial probiotics obtained from agave sap and agave juices show tolerance to hydrochloric acid, as well as strong antimicrobial activity.

Keywords: Agave, pulque, microbial consortia, prebiotic activity

Procedia PDF Downloads 373

Authors: Jessica Pinheiro Silva, Brenda Rabello De Camargo, Daniel Gusmao De Morais, Eliane Ferreira Noronha

Abstract:

The utilization of lignocellulosic biomass as source of polysaccharides for industrial applications requires an arsenal of enzymes with different mode of action able to hydrolyze its complex and recalcitrant structure. Clostridium thermocellum is gram-positive, thermophilic bacterium producing lignocellulosic hydrolyzing enzymes in the form of multi-enzyme complex, termed celulossomes. This complex has several hydrolytic enzymes attached to a large and enzymically inactive protein known as Cellulosome-integrating protein (CipA), which serves as a scaffolding protein for the complex produced. This attachment occurs through specific interactions between cohesin modules of CipA and dockerin modules in enzymes. The present work aims to construct celulosomes in vitro with the structural protein CipA, a xylanase called Xyn10D and a cellulose called CelJ from C.thermocellum. A mini-scafoldin was constructed from modules derived from CipA containing two cohesion modules. This was cloned and expressed in Escherichia coli. The other two genes were cloned under the control of the alcohol oxidase 1 promoter (AOX1) in the vector pPIC9 and integrated into the genome of the methylotrophic yeast Pichia pastoris GS115. Purification of each protein is being carried out. Further studies regarding enzymatic activity of the cellulosome is going to be evaluated. The cellulosome built in vitro and composed of mini-CipA, CelJ and Xyn10D, can be very interesting for application in industrial processes involving the degradation of plant biomass.

Keywords: cellulosome, CipA, Clostridium thermocellum, cohesin, dockerin, yeast

Procedia PDF Downloads 214

Authors: Yahia Massinissa, Henhouda Affaf, Yahia Mouloud

Abstract:

The aim of this study was to investigate the toxicity; analgesic and anti-pyretic properties of standardized HA methanolic extract (HAMeOH) in vivo. The acute toxicity study was performed on rats while adopting the OECD-420 Guidelines (fixed dose procedure). Assessment of analgesic activity was performed in rats with two analgesic models. One was acetic acid induced writhing response and the other formalin-induced paw licking. The anti-pyretic effect was tested by brewer’s yeast induced fever in rats. For the acute toxicity test, the higher dose administration of 2000 mg/kg bw. of Hyoscyamus albus did not produce any toxic signs or deaths in rats. There were no significant differences (p>0.05) in the body and organ weights between control and treated groups. The (LD50) of Hyoscyamus albus was higher than 2000 g/kg bw. In subacute toxicity study, no mortality and toxic signs were observed with the doses of 100 and 200 mg/kg bw. of extracts of for 28 consecutive days. These analgesic experimental results indicated that HAMeOH (100 mg/kg and 200 mg/kg) decreased the acetic acid-induced writhing responses and HAMeOH (100 mg/kg and 200 mg/kg) decreased the licking time in the second phase of the formalin test. Moreover, in the model of yeast induced elevation of the body temperature HAMeOH showed dose-dependent lowering of the body temperature up to 3h at both the doses these results obtained, were comparable to that of paracetamol. The present findings indicate that the leaves of Hyoscyamus albus L. possess potent analgesic and antipyretic activity.

Keywords: Hyoscyamus albus, methanolic extract, toxicity, analgesic activity, antipyretic activity, formalin test

Procedia PDF Downloads 306

Authors: Yahia Massinissa, Afaf Benhouda, Mouloud Yahia

Abstract:

Objective: The aim of this study was to investigate the toxicity; analgesic and anti-pyretic properties of standardized HA methanolic extract (HAMeOH) in vivo. Methods: The acute toxicity study was performed on rats while adopting the OECD-420 Guidelines (fixed dose procedure). Assessment of analgesic activity was performed in rats with two analgesic models. One was acetic acid induced writhing response and the other formalin-induced paw licking. The anti-pyretic effect was tested by Brewer’s yeast induced fever in rats. Results: For the acute toxicity test, the higher dose administration of 2000 mg/kg bw. of H.albus did not produce any toxic signs or deaths in rats. There were no significant differences (p>0.05) in the body and organ weights between control and treated groups. The (LD50) of 'H. albus' was higher than 2000 g/kg bw. In subacute toxicity study, no mortality and toxic signs were observed with the doses of 100 and 200 mg/kg bw. of extracts of for 28 consecutive days. These analgesic experimental results indicated that HAMeOH (100 mg/kg and 200 mg/kg) decreased the acetic acid-induced writhing responses and HAMeOH (100 mg/kg and 200 mg/kg) decreased the licking time in the second phase of the formalin test. Moreover, in the model of yeast-induced elevation of the body temperature HAMeOH showed dose-dependent lowering of the body temperature up to 3h at both the doses these results obtained, were comparable to that of paracetamol. Conclusion: The present findings indicate that the leaves of Hyoscyamus albus L. possess potent analgesic and antipyretic activity.

Keywords: Hyoscyamus albus, Umbilicus rupestris, secondary metabolites, NMR with protons, pharmacobiologic activities, methanolic extract

Procedia PDF Downloads 399

Authors: András Fülöp, Lejla Heilmann, Zsolt Szabó, Ákos Koós

Abstract:

Microbial fuel cells (MFCs) present a sustainable biotechnological solution to future energy demands. The aim of this study was to construct soil based, single cell, membrane-less MFC systems, operated without treatment to continuously power on-site monitoring and control systems during the soil bioremediation processes. Our Pseudomonas aeruginosa 541 isolate is an ideal choice for MFCs, because it is able to produce pyocyanin which behaves as electron-shuttle molecule, furthermore, it also has a significant antimicrobial effect. We tested several materials and structural configurations to obtain long term high power output. Comparing different configurations, a proton exchange membrane-less, 0.6 m long with 0.05 m diameter MFC tubes offered the best long-term performances. The long-term electricity production were tested from starch, yeast extract (YE), carboxymethyl cellulose (CMC) with humic acid (HA) as a mediator. In all cases, 3 kΩ external load have been used. The two best-operated systems were the Pseudomonas aeruginosa 541 containing MFCs with 1 % carboxymethyl cellulose and the MFCs with 1% yeast extract in the anode area and 35% hydrogel in the cathode chamber. The first had 3.3 ± 0.033 mW/m2 and the second had 4.1 ± 0.065 mW/m2 power density values. These systems have operated for 230 days without any treatment. The addition of 0.2 % HA and 1 % YE referred to the volume of the anode area resulted in 1.4 ± 0.035 mW/m2 power densities. The mixture of 1% starch with 0.2 % HA gave 1.82 ± 0.031 mW/m2. Using CMC as retard carbon source takes effect in the long-term bacterial survivor, thus enable the expression of the long term power output. The application of hydrogels in the cathode chamber significantly increased the performance of the MFC units due to their good water retention capacity.

Keywords: microbial fuel cell, bioremediation, Pseudomonas aeruginosa, biotechnological solution

Procedia PDF Downloads 271

Authors: Chukwuma S. Ezeonu, Ikechukwu N. E. Onwurah, Uchechukwu U. Nwodo, Chibuike S. Ubani, Chigozie M. Ejikeme

Abstract:

Trichophyton soudanense and Trichophyton mentagrophyte were isolated from the rice mill environment, cultured and used singly and as di-culture in the treatment of measure quantities of preheated rice husk. Optimized conditions studied showed that carboxymethylcellulase (CMCellulase) activity of 57.61 µg/ml/min was optimum for Trichophyton mentagrophyte heat pretreated rice husk crude enzymes at 50oC and 80oC respectively. Duration of 120 hours (5 days) gave the highest CMcellulase activity of 75.84 µg/ml/min for crude enzyme of Trichophyton mentagrophyte heat pretreated rice husk. However, 96 hours (4 days) duration gave maximum activity of 58.21 µg/ml/min for crude enzyme of Trichophyton soudanense heat pretreated rice husk. Highest CMCellulase activities of 67.02 µg/ml/min and 69.02 µg/ml/min at pH of 5 were recorded for crude enzymes of monocultures of Trichophyton soudanense (TS) and Trichophyton mentagrophyte (TM) heat pretreated rice husk respectively. Biomass components showed that rice husk cooled after heating followed by treatment with Trichophyton mentagrophyte gave 44.50 ± 10.90 (% ± Standard Error of Mean) cellulose as the highest yield. Maximum total lignin value of 28.90 ± 1.80 (% ± SEM) was obtained from pre-heated rice husk treated with di-culture of Trichophyton soudanense and Trichophyton mentagrophyte (TS+TM). The hemicellulose content of 30.50 ± 2.12 (% ± SEM) from pre-heated rice husk treated with Trichophyton soudanense (TS); lignin value of 28.90 ± 1.80 from pre-heated rice husk treated with di-culture of Trichophyton soudanense and Trichophyton mentagrophyte (TS+TM); also carbohydrate content of 16.79 ± 9.14 (% ± SEM) , reducing and non-reducing sugar values of 2.66 ± 0.45 and 14.13 ± 8.69 (% ± SEM) were all obtained from for pre- heated rice husk treated with Trichophyton mentagrophyte (TM). All the values listed above were the highest values obtained from each rice husk treatment. The pre-heated rice husk treated with Trichophyton mentagrophyte (TM) fermented with palmwine yeast gave bio-ethanol value of 11.11 ± 0.21 (% ± Standard Deviation) as the highest yield.

Keywords: Trichophyton soudanense, Trichophyton mentagrophyte, biomass, bioethanol, rice husk

Procedia PDF Downloads 657

Authors: M. Mischopoulou, P. Naidis, S. Kalamaras, T. Kotsopoulos, P. Samaras

Abstract:

The treatment of high strength wastewater by an Upflow Anaerobic Sludge Blanket (UASB) reactor has several benefits, such as high organic removal efficiency, short hydraulic retention time along with low operating costs. In addition, high volumes of biogas are released in these reactors, which can be utilized in several industrial facilities for energy production. This study aims at the examination of the application potential of anaerobic treatment of wastewater, with high molasses content derived from yeast manufacturing, by a lab-scale UASB reactor. The molasses wastewater and the sludge used in the experiments were collected from the wastewater treatment plant of a baker’s yeast manufacturing company. The experimental set-up consisted of a 15 L thermostated UASB reactor at 37 ◦C. Before the reactor start-up, the reactor was filled with sludge and molasses wastewater at a ratio 1:1 v/v. Influent was fed to the reactor at a flowrate of 12 L/d, corresponding to a hydraulic residence time of about 30 h. Effluents were collected from the system outlet and were analyzed for the determination of the following parameters: COD, pH, total solids, volatile solids, ammonium, phosphates and total nitrogen according to the standard methods of analysis. In addition, volatile fatty acid (VFA) composition of the effluent was determined by a gas chromatograph equipped with a flame ionization detector (FID), as an indicator to evaluate the process efficiency. The volume of biogas generated in the reactor was daily measured by the water displacement method, while gas composition was analyzed by a gas chromatograph equipped with a thermal conductivity detector (TCD). The effluent quality was greatly enhanced due to the use of the UASB reactor and high rate of biogas production was observed. The anaerobic treatment of the molasses wastewater by the UASB reactor improved the biodegradation potential of the influent, resulting at high methane yields and an effluent with better quality than the raw wastewater.

Keywords: anaerobic digestion, biogas production, molasses wastewater, UASB reactor

Procedia PDF Downloads 255

Authors: Saranya Ganapathy, Megha N. Parajulee, Michael San Francisco, Hong Zhang

Abstract:

Insect pests are a serious threat to agricultural productivity. Use of chemical pesticides, the predominant control method thus far, has resulted in environmental damage, pest resurgence, and negative effects on non-target species. Genetically modified (GM) crops offer a promising alternative, and Bacillus thuringiensis endotoxin genes have played a major role in this respect. However, to overcome insect tolerance issues and to broaden the target range, it is critical to identify alternative-insecticidal toxins working through novel mechanisms. Our research group has identified a kinase from Chilo iridescent virus (CIV; Family Iridoviridae) that has insecticidal activity and designated it as ISTK (Iridovirus Serine/Threonine Kinase). A 35 kDa truncated form of ISTK, designated iridoptin, was obtained during expression and purification of ISTK in the yeast system. This yeast-expressed CIV toxin induced 50% mortality in cotton aphids and 100% mortality in green peach aphids (GPA). Optimized viral genes (o-ISTK and o-IRI) were stably transformed into the model plant, Arabidopsis. PCR analysis of genomic DNA confirmed the presence of the gene insert (oISTK/oIRI) in selected transgenic lines. The further screening was performed to identify the PCR positive lines that showed expression of respective toxins at the polypeptide level using Western blot analysis. The stable lines expressing either of these two toxins induced moderate to very high mortality in GPAs and significantly affected GPA development and fecundity. The aphicidal potential of these transgenic Arabidopsis lines will be presented.

Keywords: Chilo iridescent virus, insecticidal toxin, iridoviruses, plant-incorporated protectants, serine/threonine kinase

Procedia PDF Downloads 258

Authors: Genesis Julyus T. Agcaoili, Esperanza C. Cabrera

Abstract:

Thirty (30) isolates of lactic acid bacteria (LAB) from traditionally-prepared fermented products specifically fermented soy-bean paste, fermented mustard and fermented rice-fish mixture were studied for their in vitro antimicrobial and cytotoxic activities. Seventeen (17) isolates were identified as Lactobacillus plantarum, while 13 isolates were identified as Enterococcus spp using 16s rDNA sequences. Disc diffusion method was used to determine the antibacterial activity of LAB against Staphylococcus aureus (ATCC 25923) and Escherichia coli (ATCC 25922), while the modified agar overlay method was used to determine the antifungal activity of LAB isolates on the yeast Candida albicans, and the dermatophytes Microsporum gypseum, Trichophyton rubrum and Epidermophyton floccosum. The filter-sterilized LAB supernatants were evaluated for their cytotoxicity to mammalian colon cancer cell lines (HT-29 and HCT116) and normal human dermal fibrolasts (HDFn) using resazurin assay (PrestoBlueTM). Colchicine was the positive control. No antimicrobial activity was observed against the bacterial test organisms and the yeast Candida albicans. On the other hand, all of the tested LAB strains were fungicidal for all the test dermatophytes. Cytotoxicity index profiles of the supernatants of the 15 randomly picked LABs and negative control (brain heart infussion broth) suggest nontoxicity to the cells when compared to colchicine, whereas all LAB supernatants were found to be cytotoxic to HT-29 and HCT116 colon cancer cell lines. Results provide strong support for the role of the lactic acid bacteria studied in antimicrobial treatment and anticancer therapy.

Keywords: antimicrobial, fermented products, fungicidal activity, lactic acid bacteria, probiotics

Procedia PDF Downloads 218

Authors: Sara Franco Ortega, Alina Goldberg Cavalleri, Nawaporn Onkokesung, Richard Dale, Melissa Brazier-Hicks, Robert Edwards

Abstract:

Safeners are agrochemicals that enhance the selective chemical control of wild grasses by increasing the ability of the crop to metabolise the herbicide. Although these compounds are widely used, their mode of action is not well understood. It is known that safeners enhance the metabolism of herbicides, by up-regulating the associated detoxification system we have termed the xenome. The xenome proteins involved in herbicide metabolism have been previously divided into four different phases, with cytochrome P450s (CYPs) playing a key role in phase I metabolism by catalysing hydroxylation and dealkylation reactions. Subsequently, glutathione S-transferases (GSTs) and UDP-glucosyltransferases lead to the formation of Phase II conjugates prior to their transport into the vacuole by ABCs transporters (Phase III). Maize (Zea mays), was been treated with different safeners to explore the selective induction of xenome proteins, with a special interest in the regulation of the CYP superfamily. Transcriptome analysis enabled the identification of key safener-inducible CYPs that were then functionally assessed to determine their role in herbicide detoxification. In order to do that, CYP’s were codon optimised, synthesised and inserted into the yeast expression vector pYES3 using in-fusion cloning. CYP’s expressed as recombinant proteins in a strain of yeast engineered to contain the P450 co-enzyme (cytochrome P450 reductase) from Arabidopsis. Microsomes were extracted and treated with herbicides of different chemical classes in the presence of the cofactor NADPH. The reaction products were then analysed by LCMS to identify any herbicide metabolites. The results of these studies will be presented with the key CYPs identified in maize used as the starting point to find orthologs in other crops and weeds to better understand their roles in herbicide selectivity and safening.

Keywords: CYPs, herbicide detoxification, LCMS, RNA-Seq, safeners

Procedia PDF Downloads 115

Authors: Abu Yousuf, Maksudur Rahman Khan, Ahasanul Karim, Amirul Islam, Minhaj Uddin Monir, Sharmin Sultana, Domenico Pirozzi

Abstract:

Traditional biodiesel feedstock like edible oils or plant oils, animal fats and cooking waste oil have been replaced by microbial oil in recent research of biodiesel synthesis. The well-known community of microbial oil producers includes microalgae, oleaginous yeast and seaweeds. Conventional transesterification of microbial oil to produce biodiesel is lethargic, energy consuming, cost-ineffective and environmentally unhealthy. This process follows several steps such as microbial biomass drying, cell disruption, oil extraction, solvent recovery, oil separation and transesterification. Therefore, direct transesterification of biodiesel synthesis has been studying for last few years. It combines all the steps in a single reactor and it eliminates the steps of biomass drying, oil extraction and separation from solvent. Apparently, it seems to be cost-effective and faster process but number of difficulties need to be solved to make it large scale applicable. The main challenges are microbial cell disruption in bulk volume and make faster the esterification reaction, because water contents of the medium sluggish the reaction rate. Several methods have been proposed but none of them is up to the level to implement in large scale. It is still a great challenge to extract maximum lipid from microbial cells (yeast, fungi, algae) investing minimum energy. Electroporation technique results a significant increase in cell conductivity and permeability caused due to the application of an external electric field. Electroporation is required to alter the size and structure of the cells to increase their porosity as well as to disrupt the microbial cell walls within few seconds to leak out the intracellular lipid to the solution. Therefore, incorporation of electroporation techniques contributed in direct transesterification of microbial lipids by increasing the efficiency of biodiesel production rate.

Keywords: biodiesel, electroporation, microbial lipids, transesterification

Procedia PDF Downloads 258

Authors: Denis V. Axenov-Gribanov, Irina V. Voytsekhovskaya, Evgenii S. Protasov, Maxim A. Timofeyev

Abstract:

Isolated ecosystems existing under specific environmental conditions have been shown to be promising sources of new strains of actinobacteria. The taiga forest of Baikal Siberia has not been well studied, and its actinobacterial population remains uncharacterized. The proximity between the huge water mass of Lake Baikal and high mountain ranges influences the structure and diversity of the plant world in Siberia. Here, we report the isolation of eighteen actinobacterial strains from male cones of Pinus sylvestris trees growing on the shore of the ancient Lake Baikal in Siberia. The actinobacterial strains were isolated on solid nutrient MS media and Czapek agar supplemented with cycloheximide and phosphomycin. Identification of actinobacteria was carried out by 16S rRNA gene sequencing and further analysis of the evolutionary history. Four different liquid and solid media (NL19, DNPM, SG and ISP) were tested for metabolite production. The metabolite extracts produced by the isolated strains were tested for antibacterial and antifungal activities. Also, antiradical activity of crude extracts was carried out. Strain Streptomyces sp. IB 2014 I 74-3 that active against Gram-negative bacteria was selected for dereplication analysis with using the high-yield liquid chromatography with mass-spectrometry. Mass detection was performed in both positive and negative modes, with the detection range set to 160–2500 m/z. Data were collected and analyzed using Bruker Compass Data Analysis software, version 4.1. Dereplication was performed using the Dictionary of Natural Products (DNP) database version 6.1 with the following search parameters: accurate molecular mass, absorption spectra and source of compound isolation. Thus, in addition to more common representative strains of Streptomyces, several species belonging to the genera Rhodococcus, Amycolatopsis, and Micromonospora were isolated. Several of the selected strains were deposited in the Russian Collection of Agricultural Microorganisms (RCAM), St. Petersburg, Russia. All isolated strains exhibited antibacterial and antifungal activities. We identified several strains that inhibited the growth of the pathogen Candida albicans but did not hinder the growth of Saccharomyces cerevisiae. Several isolates were active against Gram-positive and Gram-negative bacteria. Moreover, extracts of several strains demonstrated high antioxidant activity. The high proportion of biologically active strains producing antibacterial and specific antifungal compounds may reflect their role in protecting pollen against phytopathogens. Dereplication of the secondary metabolites of the strain Streptomyces sp. IB 2014 I 74-3 was resulted in the fact that a total of 59 major compounds were detected in the culture liquid extract of strain cultivated in ISP medium. Eight compounds were preliminarily identified based on characteristics described in the Dictionary of Natural Products database, using the search parameters Streptomyces sp. IB 2014 I 74-3 was found to produce saframycin A, Y3 and S; 2-amino-3-oxo-3H-phenoxazine-1,8-dicarboxylic acid; galtamycinone; platencin A4-13R and A4-4S; ganefromycin d1; the antibiotic SS 8201B; and streptothricin D, 40-decarbamoyl, 60-carbamoyl. Moreover, forty-nine of the 59 compounds detected in the extract examined in the present study did not result in any positive hits when searching within the DNP database and could not be identified based on available mass-spec data. Thus, these compounds might represent new findings.

Keywords: actinobacteria, Baikal Lake, biodiversity, male cones, Pinus sylvestris

Procedia PDF Downloads 209