Biochemical Engineering

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    Bio-Butanol Production from Napier Grass Hydrolysis Using Clostridium Acetobutylicum
    (Addis Ababa University, 2022-02) Biruk, Yohannes; Solomon, Kiros (PhD)
    Napier grass is a potential biomass for biofuel production due to its high adaptability and widespread availability in tropical areas such as Ethiopia. A compositional analysis of Napier grass collected from the Menagesha Mushroom farm in an Addis Ababa city suburb was performed, and microbes and enzymes were purchased from France. An alkaline pretreatment (2.5wt% NaOH, 1.5wt% urea, and 8wt% sample biomass) was mixed for 40 minutes at 121°C, followed by enzymatic hydrolysis (2ml Novozymes Cellic glucadase enzyme and 5gram pretreated biomass) shaken at 50rpm for 40 minutes at 50°C, and separated by centrifuge at 8500rpm to hydrolyze Napier grass. Carbohydrates make up 52.1 % of the Napier grass composition, making it an excellent carbon source. Acetone-Butanol-Ethanol fermentation (ABE) was carried out in a continuous fermentation process using co-cultured C. acetobutylicum bacteria. After optimizing the operating parameters pH, Temperature, substrate concentration, and fermentation time, the productivity of butanol was found to be 0.077 g lt -1 hr -1 with a 29.5 % efficiency. The yield of ABE was 0.0833 g lt -1 hr -1 when compared to the semi–Simultaneous Saccharification fermentation (sSSF) process, which combined alkaline pretreated biomass with enzymatic hydrolysis and cocultured bacteria after 24 hours of hydrolysis followed by 96hrs of fermentation. Butanol efficiency at the s-SSF process is 33.6 %. When compared to previous studies, ABE fermentation of Napier grass yields a competitive yield.
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    Investigating the Possibility of Producing Polyhydroxyalkanoates (PHAs) Using Hydrolysates of Napier Grass by Burkholderia Sacchari Strain
    (Addis Ababa University, 2022-02) Adane, Bazezew; Sintayehu, Nibret (PhD)
    Bacteria produces Polyhydroxyalkanoates (PHAs), which are macromolecules. They are inclusion bodies that build up as reserve resources while bacteria grow under various stress situations. PHAs have been chosen as alternatives for the development of biodegradable polymers due to their rapid degradability under natural environmental circumstances. The purpose of this study was to produce reducing sugars from lignocellulosic Napier grass and examine PHA production by the impact of temperature, pH, and culture time on growth yields by employing the produced reducing sugars as carbon sources using response surface methodology via Box- Behnken design BBD). Burkholderia Sacchari strain was chosen from more than 300 bacterial strains capable of PHA accumulating strains based on its ability to consume both hexose and pentose sugars and has a capacity of accumulating high mass PHA granules inside its cells. At 200 rpm, optimal pH, fermentation temperature, and incubation time for the isolate to produce the highest PHA were 7, 35 °C, and 48 hour, respectively. When hydrolysates of Napier grass was employed as carbon sources and ammonium sulphate as nitrogen sources, the strain was able to collect 62.1% PHA from its total biomass. The extracted PHAs' FTIR spectra show strong peaks at wavenumbers that are unique to PHAs as C–H, CH2, O-H, C=O, and C–O groups. The extract's resemblance appropriateness for bioplastic production were validated by UV–Vis spectrophotometric analysis.
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    Production of Antioxidant Rich Glucose Syrup from Ensete Ventricosum Starch through Enzymatic Hydrolysis and Preservation using Green Tea Extract
    (Addis Ababa University, 2021-10) Tigist, Abebe; Shimeles, Shumi (PhD)
    Ensete ventricosum is a plant native to Ethiopia with a high content of starch. This study aimed to produce antioxidant rich glucose syrup from enset starch by using alpha amylase and glucoamylase enzymes and preservatives using green tea polyphenols. The starch was isolated from the corm and pseudostem parts of Enset plant and characterized. The main process steps for enzymatic production of glucose syrup from enset starch were: gelatinization, liquefaction, saccharification, centrifugation followed by treatment with activated charcoal and evaporation to final solids ( >70%). The preservative effect of green tea extract on the shelf life of the syrup was also examined for 30days, by studying its effect on pH, moisture content and viscosity of the glucose syrup. Samples were treated with 0, 100, 200, and 300ppm of green tea polyphenols. The study showed that the moisture content, ash content, crude fat content, protein content, and amylose content of enset starch were: 8.98%, 0.15%, 0.38%, 0.1%, and 29.01%, respectively. Response Surface Methodology Design (RSMD), particularly, Box Behnken Design (BBD) was applied to investigate the effect of temperature (50, 60 and 70℃), time (12, 24 and 36hr), and glucoamylase concentration (0.1, 0.15 and 0.2wt.%), on the process of saccharification using Design expert® 12 software, which shows the significant influence on the value of dextrose equivalent (DE). The maximum concentration of glucose (95.64%) in terms of DE was found at the parameter interaction of 0.1wt.% (glucoamylase concentration), 60℃, and 36hr. The result showed that the optimum condition for the saccharification process was 62℃, 12hr and 0.2wt.% (glucoamylase dosage) which resulted in 91.79±0.5% of DE. The moisture content, total soluble solid, ash content, density, specific gravity, viscosity and pH of the product were: 24.01%, 75.32°Brix, 0.35%, 1380.8 kg/m3, 1.3833, 940 mPa.s, and 5.5, respectively. Generally, it was observed that green tea extracts with a concentration of 100ppm of TPC was found to be suitable as preservative to keep the quality and extend the shelf-life of glucose syrup at room temperature. Hence, Enset starch is a promised raw material to develop glucose syrup enriched with extracts of green tea to elevate its shelf life.
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    Production of Poly-hydroxybutyrate (PHB) from Glucose Derived from Sugar Cane Bagasse using Bacillus Subtilius
    (Addis Ababa University, 2021-03) Tinsae, Messay; Zebene, Kiflie (PhD)
    Most plastic materials are produced from petrochemicals. However, due to their non-degradable nature, they are causing serious environmental problems. The objective of this study was to obtain a bio-plastic material known as poly-hydroxybutyrate (PHB) based on glucose extracted from sugarcane bagasse with the help of bacillus subtilius. under stressing media condition. The extraction process stages were pretreatment, hydrolysis, fermentation, extraction, and recovery. The first stage was pretreating sugar cane bagasse and hydrolysis using dilute sulfuric acid in 1:8 solid/liquid ratios. The three factors analyzed were temperature, pH, and fermentation time for extraction of PHB. Synthesized PHB was characterized using Sudan staining, UV spectrophotometer and FTIR. The Response Surface Methodology with Box-Behnken experimental design was used to find the optimum extraction conditions. The effects of extraction parameters (temperature, pH, and fermentation time) on the response (poly-hydroxybutyrate yield) were analyzed. The optimum PHB yield of 6.82 g/l (0.17 g/g of bagasse) of was obtained at 37.3 0c, 7.2 pH and 48.6 hours. From the statistical analysis, all the factors had significant effect on the yield. Moreover, the interaction effects of temperature and pH, temperature and fermentation time, and pH and fermentation time had also significant effects on the yield.
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    Extraction and Characterization of Atropine from Jimson Seed (Datura Stramonium) as Biopesticide and Evaluate its Bioactivities Against Maize Weevil (Sitophilus Zeamais)
    (Addis Ababa University, 2021-03) Leta, Takele; Solomon, Kiros (PhD)
    The effect of synthetic pesticides on the environment and human health is persistent and very aggressive. A series health problem caused by synthetic pesticides every year in the world to a human being are major treat. The study investigates the extraction of atropine from Jimson seeds for biopesticide application using acid-water extraction method. The parameters used were particle size, extraction temperature, and extraction time. The extraction process stages performed in the experiments are pretreatment, extraction, separation and concentration. The first stage was drying and grinding the raw seed to a particulate size of 550, 750 and 1000μm. The raw seed have had 24.241 % (w/w) of moisture content, with ash and organic matter content of 6.607 %and 93.393% (w/w) respectively. Water was prepared in a ratio of 5:1(ml/gm) for extraction. Finally, the ground seed and the solvent were poured to an extraction vessel with overhead stirrer simultaneously and extraction was carried out until the extraction time becomes finished. The characteristics of atropine were then evaluated using different analytical methods. The result found; retention time of sample 5.5-minute, specific gravity of 1.4164 ± 0.07225, the flash point was 36.06 ± 0.9932oC, water solubility was 99.768 ± 0.0217% (w/v), acidity of atropine was 5.2 ± 0.19 and TVOC’s was 0.17426 ± 0.03%(w/w). Response surface methodology (RSM) of Box – Behnken method was used to justify the factors effect on the yield. The interaction effect of particle size and extraction temperature, particle size and extraction time, and extraction temperature and extraction time had also a significant effect on the yield. Depending on the model optimization design 713μm, 45.42oC, and 2.33h of the factors were found the optimum points with 10.811 yield of extract. Finally, the extracted atropine was characterized using UV Spectrophotometer, FTIR, and HPLC. The results obtained from the acid-water extraction method at different parameters confirmed that the yields were maximum and effect of prepared biopesticide is comparable with commercial synthetic pesticide. The result of assessment of the percent of death of maize weevil after 48hrs and 72hrs, the atropine based biopesticide was efficient in small quantity and less time when compared with chemical synthetic. The percentage of death of maize weevil was 41.6% for 48hours and 83.3% for 72hours for atropine based biopesticide.
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    Identification, Isolation and Characterization of Suitable Lactic Acid Bacterial Strain for Minimizing Fermentation Time of Kocho
    (Addis Ababa University, 2017-10) Redae, Nuguse; Adamu, Zegeye (Assoc.Prof.)
    Kocho is a traditional food prepared through fermentation of decorticated Enset in Southern Ethiopia. It has been produced for many years and is being consumed in the region. Various research findings were reported on Enset with major concerns such as food safety and security issues, microbial dynamics of enset fermentation, microbial spoilage and accompanying changes, biochemical changes during fermentation and the effect of altitude on microbial successions. However, gaps were observed during Kocho fermentation, involving lactic acid bacterial (LAB) strain for traditional Kocho fermentation, standardization and optimization of the process. Kocho fermentation has been done in underground pit for a minimum of four months. This long time of fermentation has become a problem of food insecurity, and its minimization is the main concern of this thesis research. Three Kocho samples (Month-1, Months-3 and Months-6) and three Enset trial varieties (Agade, Disho and Gimbo) were taken from Morsito Wereda of SNNP, Ethiopia. The temperature and pH of fermented mass samples (Month-1, Months-3 and Months-6) were measured and recorded as 32, 27 and 290C and 4.78, 4.47 and 4.39, respectively. From each of the three samples, 1mg was taken and serial dilution was prepared, thereafter 0.1ml from each serial dilution factor was inoculated to 54 petri plates and incubated at 25, 30 and 350C temperatures. Growth of mesophilic microbes was recorded from each plate. The results showed that colony forming units of microbes were the highest in Month-1 sample at the temperature of 300C. Based on morphological structure, color and size, LAB were identified, isolated, re-cultured and purified to single strain. White, small and round structure of the strain were stored at 40C. By using Enset trial variety samples as a source of carbon, broth media were prepared according to MRS composition. 30-35 CFU of the pure strain was inoculated to each broth medium, and lactic acid concentration was determined by pH meter for 36 days with 4 days interval at 35 and 300C incubation temperatures. Generally, pH dropped down linearly for both temperatures and for the three Enset trial variety samples up to 23rd day of fermentation and became constant (~pH=4.50) at 24-28th day and reached ~pH=5.30 at 36th day for all varieties. However, pH decreased faster at 350C for all Enset trial varieties, and Disho fermentation dropped faster for eight days than others, and long-delayed from 8-16th day. The pH and temperature of the experiment corresponded with pH and temperature that were recorded during sample collection of sample Month-6. The result showed that only 24-28 days are needed to get Kocho with good texture, fl
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    Characterization and Optimization of Bioinsecticide Extracted from Cigarette Waste to Control Cabbages Insect Pests
    (Addis Ababa University, 2020-11) Badhane, Gudeta; Solomon, Kiros (PhD)
    Chemical pesticides and fungicides have been in use since long period to protect the crops from insects and diseases. Plant extracts were likely the earliest agricultural bio-insecticides. Nicotine in the form of tobacco extracts was reported in 1690 as the first plant-derived insecticides. A huge mass of tobaccos is wasting in NTE during cigarette processing and within a cigarette butt disposal. Therefore, this study is aimed to characterize and optimize nicotine extract from these tobaccos wasted for insecticide application. The samples were collected from different parts of the countries. The tobacco fillers were removed from wastes, dried and resized for better extraction efficiency and quality. The processes involve the extraction, emulsification, and testing of efficiency on cabbage aphids. The extraction process was performed by the solvent extraction method using methanol to ethanol ratio of 1:4 determined by GCMS. The independent variables analyzed for extraction parameters were temperature (30-600C), extraction time (4-6hr), and molarity of sodium hydroxide (1-3M), and their optimum conditions analyzed by design expert, Response Surface methods, Central Composite design. Quality analysis was carried out by GCMS whereas quantity was analyzed by GCMS and UV/visible Spectrophotometer. The maximum yield and nicotine content obtained at the optimum condition of parameters were 17.749 and 3.258 % of the weight of tobacco filler. The emulsification process was carried out by mixing 10g nicotine-containing extracts (54. 4629 g) with 13.616 gram of the mixture of palm oil and surfactants prepared by 1:4 ratio of palm oil to surfactant at room temperature and atmospheric pressure. The physicochemical characteristic of the emulsified concentrated such as density, viscosity, pH, flash point, and surface tension were analyzed and their value was recorded as 1.0122 ± 0.0103 g/ml, 585.33 ± 2.52 mPas, 9.37 ± 0.03, 87.96 0C, and 34.10mN/m respectively. The best efficiency of the emulsified concentrated extract tested on the cabbage aphid was observed at the ratio of 1:100 (the emulsified concentrated to solvent).
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    Bioaccumulation of Cr(VI) from Tannery Wastewater Using Saccharomyces Cerevisiae
    (Addis Ababa University, 2018-06) Tessafa, Abrham; Shegaw, Ahmed (Assoc.Prof.)
    Chrome tanning is the most and widely used types of tanning process and the wastewater generated by these industries is a major source of chromium pollution in the form of Cr(III)&Cr(VI) which are released into the environment either after treated inefficiently or with no treatment and become a cause for many human health problem. Several researchers have published papers to find the solution for removal of Cr(VI) but at its low concentration and this research work investigates at high concentration of Cr(VI). Cell suspensions of S.cerevisiae at its exponential growth phase were added to 250ml plastic flasks of containing 50ml of YPD media at pH value of (3,5&8), initial Cr(VI) concentration of (5, 50&100mg/ml) and incubation time of(3,5&7 days) and shaken at 200rpm an from tannery wastewater using S.cerevisiae shows good result after Cr(VI)-contaminated wastewater has been treated at different treatment variables of pH, initial Cr(VI) concentration. Cr(VI) uptake efficiency of S.cerevisiae increases at low values of pH, Cr(VI) and incubation time and decreases at high values of these variables. And it can be conclude that Cr(VI)uptake efficiency is better at low values of the variables and the optimum conditions obtained by using the full factorial experimental design are pH value of 4.5, Cr(VI)concentration of 5mg/ml and incubation time of 5 days resulting 84.25 % theoretical Cr(VI) uptake efficiency . Cell viability test assay of this work shows the possibility of Cr (VI) removal at its high concentration using living cells that are tolerant to high concentration though the efficiency decreases as concentration increases due to inhibition. Bioaccumulation of Cr(VI) by the cell S.cerevisiae is analyzed by FTIR by comparing their IR spectrum of functional groups of the Cr(VI) free and Cr(VI) loaded S.cerevisiae cell wall that showed change of wavenumbers of biocomponents of cell wall of the two cells which confirms the binding of chromium ion with the functional groups of cell wall of S.cerevisiae and produces Cr complex compounds /precipitates. Freundlic equilibrium and pseudo second order kinetic model are used for expressing
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    io butanol Production from Brewery’s Spent Grain Hydrolysate by Clostridium/Eubacterium and Bacillus subtilis
    (Addis Ababa University, 2018-06) Meron, Asteraye; Solomon, Kiros (PhD)
    Butanol (butyl alcohol or n-butanol) is a four carbon straight chained alcohol which can be used as a liquid transportation biofuel. The objective of this study was production of bio butanol from brewery spent grain (BSG) which in effects to minimize energy cost and substituting nonrenewable energy by using renewable resources. The chemical and proximate compositions analysis was investigated. The moisture, ash, extractives, hemicellulose, lignin and cellulose contents were, 64.65, 4.2, 33.28, 17.79 and 17.87 wt. % respectively. The conversion of BSG to bio butanol can be achieved mainly by four process steps,pre-treatment, hydrolysis (dilute acid hydrolysis), fermentation and separation. Dilute acid hydrolysis was employed, because it is easy and productive process. Fermentation of the hydrolyzate was performed using 10 % v/v Clostridium/Eubacterium at 37 C temperature, pH 6.5 and 96 hr fermentation time for all samples. The experiment was designed by Box-Behnken Design (BBD) using Design expert 7.0.0 software with three factors to investigate the effect of (temperature, reaction time and acid concentration). Hydrolysis temperature was varied from 115°C to 135℃, reaction time was varied from 30 to 40 minutes and the acid concentration was varied from 1.5% to 2%. Significance of the process variables were analyzed using analysis of variance (ANOVA) and the quadratic model was fitted to the experimental results. Thus, the influence of all experimental variables, factors and interaction effects on the response was investigated. Among the investigated components hydrolysis temperature, reaction time, acid concentration, interaction between temperature and reaction time, interaction between temperature and acid concentration contributed a significant effect on the yield of bio butanol. As the result of RSM optimization, the best yield of total reducing sugar (TRS) and bio butanol were found at 127.42 0 C hydrolysis temperature, 34.32min reaction time and 1.65% w/w acid concentration. Under these conditions, 59.0 g/L total reducing sugar (TRS), 10.93 g/L biobutanol yield and 0.11 g/L.h of productivity were obtained. The optimized process variables were farther enhanced by co culturing Clostridium/Eubacterium and Bacillus subtilis. Different inoculum concentration ratios (10:1%, 10:3%, and 10:5% v/v) of Clostridium/Eubacterium to Bacillus subtilis) were used to ferment BSG hydrolysate. The results exhibited that the inoculum size of 10% Clostridium/Eubacterium with 1% Bacillus subtilis maximised the production of biobutanol to 13.76g/L and 0.14 g/L.h productivity.
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    Characterization and Treatment of Beseka Lake Water Quality Using Microbes to reduce Total Dissolved Solid (TDS) and Alkalinity
    (Addis Ababa University, 2018-07) Yasin, Ahmed; Solomon, Kiros (PhD)
    Water is one of the natural resource that used for irrigation, industrial, sanitation and other purpose that facilitate human activity. Lake Beseka is volcanically dammed, endorheic lake, which is located to the East direction, situated around 200 km from Addis Ababa, the capital city of Ethiopia. The Lake water is saline and alkaline with high concentration of fluoride so that the water could not be used for drinking and irrigation purpose. Different infrastructures such as a health center, high school, youth center, offices and others were affected by the lake flooding. So, this lake needs to be treatment. In present study, biological neutralization of saline- alkaline beseka lake water in the presence of glucose (carbohydrate) as carbon source was studied. To do so, determination of physiochemical characteristics of the lake water, which was pH, EC, TDS, HCO3 - , total alkalinity, chloride and fluoride were measured and also isolation of bacteria were carried out. In this study Bacillus alkalophilus bacteria was also used to compare its treatment performance with isolate (T1). Treatments of lake water sample at different pH, incubation time and inoculum size were monitored. Maximum removal of total alkalinity and TDS (Total Dissolved Solid) were observed at pH, incubation time and inoculum size of 9,72hr and 32ml respectively. Which is resulted in removal of total alkalinity; 774.669mg/l and TDS;1939.92mg/l for isolated (T1) and total alkalinity;779.631mg/l and TDS;1944.97mg/l for Bacillus alkalophilus from original value. Based on these findings, isolated (T1) bacteria were shown to be have good performance for biological treatment of beseka lake.
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    Production of Bioethanol from Brewery Spent Grain Using Cellulase Enzyme Hydrolysis
    (Addis Ababa University, 2018-07) Emru, Yidnekew; Eduardo, Ojito (Prof.)
    Bioethanol is a clear, colorless and mobile flammable liquid chemical compound with acceptable odour, and produced from biomass feed stocks through fermentation. The objective of this study was production of bioethanol from Barley spent grain by using cellulase enzyme hydrolysis. Decreasing of world petroleum reserves, fluctuation of petroleum price and environmental concerns, low yield cellulose acid hydrolysis and formation of inhibitory were the problem statement. Enzymatic hydrolysis method was used for cellulose hydrolysis, producing monomeric C6 sugars (glucose); enzymatic hydrolysis is more specific and high conversion rates can be achieved in long reaction times obtaining very high yields. Raw material characterization of BSG were, hemicelluloses content 28.4 %, cellulose content 27 %, and dry matter content 98.2 %. The aim of pretreatment is to disrupt recalcitrant structures of cellulosic biomass to make cellulose more accessible to the enzymes that convert carbohydrate polymers into fermentable sugars. The pretreated feedstock was mixed with 1.25 % (v/v) sulfuric acid solutions with a liquid-to-solid ratio of 20 %( w/w) and it was occurred on the temperature 121 ºC and time of 17 min, the severity factor coefficient was 0.198. Cellulase enzyme was produced from BSG using bacillus subtilus, under liquid state fermentation for 5 days at temperature of 37 ºC with pH of 5.0, in controlled incubator. For the enzyme hydrolysis, 26 full factorial central composite design (CCD) was applied to investigate the effect of temperature (40-50 ºC), pH (4.0-5.0), reaction time (24-72 hr.) and enzyme loading (1-2 %) using Design expert® 6 software. RSM was applied to investigate the interaction effect of hydrolysis process variables on the yield of ethanol from BSG. After hydrolysis process sugar content of the hydrolysate was quantified using spectrophotometer measuring its absorbance. Fermentation of the hydrolysate were performed 2.5 % Saccharomyces cerevisiae and 2.5 % fusarium oxysporum at 30 ºC temperature, pH 5.0 and 72 hr. fermentation time for all samples. Significance of the process variables were analyzed using analysis of variance. As the result of RSM optimization, the best yield of TRS was found at 40 ºC hydrolysis temperature, 4.5 PH, 48 hr. reaction time and 1.5 % v/v enzyme loading. Under these condition 96.55 %w/w and 53.68 % per 6.58 g dry barley spent grain of TRS and bio-ethanol respectively obtained.
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    Investigation on Mineral Characterization of Powder Injera
    (Addis Ababa University, 2019-10) Mulatu, Betsegaw; Eduardo, Ojito (Prof.)
    Injera is a fermented, sour bread consumed as a staple food in Ethiopia and Eritrea. The bread can be prepared from various cereals but tef [Eragrostis tef (Zucc.) Trotter] is the most preferred ingredient. A deep study on its inherent characteristics related to processing of injera is needed for its use in various food applications. This research attempt to use spray dryer technology to cook injera, as an alternative method of injera processing. And the product is forwarded for the use of as a food supplement. Also the researcher study the impact of type of tef white and red on nutritional composition of powder injera and impact of fermentation time at 24h, 48h, and 72h on moisture content, ash content, fiber content, iron content, zinc content, calcium content, potassium and sodium content of nutritional composition of powder injera was tested using statistic and regression analysis. Microsoft excel and two factorial three level design expert 11 is used to analyze the experimental result. It is observed that red tef has higher calcium, zinc, and iron value than the white tef but the white tef powder injera has sodium and potassium content than the red tef powder injera. And it is observed that at fermentation time of 48h the powder injera has higher mineral content than at fermentation time of 24h and 72h.
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    Determine the Potency of Caffeine Degradation Microbes and Process Optimization
    (Addis Ababa University, 2019-06) Zeynu, Shamil; Cespedes, Eduardo Ojito (Prof.)
    Bioremediation being a problem solver for many different environmental and other major hazards, they play an important and cost effective role in the decaffeination process. Form collected environmental sample coffee husk. From CH caffeine can be eliminated. With this experimental work take two spp of microorganism that were bacillus subtilis and rhizobium from Ethiopia bio diversity institute, that can be capability of improving soil fertility. Could grow on the medium supplemented with 2.5g/L caffeine and could effectively degrade 2.5 g/L of caffeine in the liquid media as a sole source of carbon and nitrogen. Morphological and biochemical characteristics were maintained for re-affirmation the organism as bacillus subtilis and rhizobium. The degradation of caffeine capability was authenticated by growth curve. The next task were that those authenticated caffeine degrading bacteria applicable for conserving soil fertility by means of N-fix and p- solubilized potential then propagation and growth of those selective caffeine degrading bacteria was maintained by selective medium with and without supplement of 2.5g/l caffeine then optimized the factors that affecting cell growth with prepared broth with and without supplement of caffeine and determined interaction of each factor from experiment design software 6.0.8) the result would be temp; 35 o c, pH 7.07 incubation time 36hr from those optimal condition obtained maximum cell growth in number was 3.41995E+008 and 5.0707E+008 with and without caffeine respectively for bacillus subtilis and temp 34 o c, pH 6.6 and incubation time 28hrs optimum condition maximum cell growth 3.51146E+008 and 4.09131E+008 was obtained for rhizobium sp. of organism. That viable cell grown microorganism ready for applicable of as liquid bio fertilizer those were easiest way of production, handle, cost effective and keep soil health care method of production.
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    ggshell and Membrane-Based Substrate for the Production, Optimization and Detergent Compatibility of Alkaline Protease Enzyme
    (Addis Ababa University, 2019-07) Tsegazeab, Kassa; Solomon, Kiros (PhD)
    In recent years the advancement of technology results not only development but also, environmental risks due to the use of chemicals, catalysts and environmentally hazardous materials moreover coasty. In spite of this, the use of biotechnological principles and products in the industrial sector is mandatory and expected to with stand the current problems. Though Microbial alkaline proteases are stimulating tremendous interests in the research and enzyme market. Hence, the aim of this thesis was the use of eggshell and membrane as a substrate for the production, optimization and detergent compatibility of alkaline protease using Bacillus subtilis mojavensis in submerged fermentation. Fermentation process for enzyme production was conducted using, four factors with three levels were used and 29 experiment had been studied. The optimum or maximum alkaline protease production was obtained at a temperature of 37 C, eggshell concentration of 20 %, pH of 9.0 and incubation time of 48 h with a maximum activity of 214 U/ml. Optimum parameters for the production of alkaline protease was found using design expert optimizer at 9.08 pH, 39.74 0 C, 19.78% eggshell and 48 h of incubation time were obtained. Alkaline protease enzyme was produced for further analysis such as, characterization and detergent compatibility of the enzyme. Enzyme activity was conducted in a pH range of 6-12, with maximum activity obtained at pH 10. It showed that more than 93.97% its original activity was retained using buffers pH ranging from 8-10, with the optimum activity at pH 10.0. Overall alkaline protease enzyme was stable between pH 8.0-10.0 attain 90%, 100% and 94% of residual activity. Its minimum residual activity was attained at pH value of 6.0 which is 39.6% of its original activity. Alkaline protease was found to be very active at all temperatures tested between 30 0 C and 80 0 0 C, alkaline protease was stable and retained more than 74% of its maximum activity. From 1M to 2.0M of NaCl concentration, exhibits > 97% of optimum activity was recorded at pH 8 and 10. Whereas; optimum stability was attained at these concentration range between 1.0 - 3.0 M NaCl. At pH 8.0 residual activity were raising as concentration increases and constant after 2.5 M. The detergent compatibility study was conducted. Results increasing the concentration of enzyme increases the degradation of blood stain. Therefore, alkaline protease enzyme has compatibility to industrial application including detergent as an alternative. C with maximum activity at 60 0 C. Within temperature range of 40 0 0 C - 70 0 0 C alkaline protease was stable and retained more than 74% of its maximum activity. From 1M to 2.0M of NaCl concentration, exhibits > 97% of optimum activity was recorded at pH 8 and 10. Whereas; optimum stability was attained at these concentration range between 1.0 - 3.0 M NaCl. At pH 8.0 residual activity were raising as concentration increases and constant after 2.5 M. The detergent compatibility study was conducted. Results increasing the concentration of enzyme increases the degradation of blood stain. Therefore, alkaline protease enzyme has compatibility to industrial application including detergent as an alternative.
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    Treatment of Waste Water Effluent from Coffee Industry using Anaerobic Mixed Culture Pseudomonas Florescence and Escherichia Coli Bacteria and with Gypsum: A Case in Dilla- Ethiopia
    (Addis Ababa University, 2019-06) Fitsum, Ashenafi; Cespedes, Eduardo Ojito (Prof.)
    Coffee processing industries generate huge amount of wastewater. The effluent is characterized by a lot of sludge, high amount of organic load and acidic nature due to fermentation process. Due to this, it contains high value of COD, BOD, TS and other contaminants. This effluent when disposed in natural water, it contaminates aquatic animals and plants environment. So, care has to be taken to ensure the quality of the releasing waters should be reduced below the prescribed standards. There are many physical and chemical treatment methods available for the removal of pollutants but all these methods have problems associated with secondary effluent, hazardous and harmful end products, high energy consuming, non-economic etc. These problems can be overcome by the use of biological treatment methods which are simple, eco-friendly and efficient where complete removal of the pollutants is possible. Cognizant of this, the researcher was initiated to study the treatment of wastewater effluent from coffee industry in the specific area by using anaerobic Pseudomonas florescence and Escherichia coli bacteria as mixed culture and with gypsum (CaSO4.2H2O). In this study, the wastewater value of BOD5, COD, TS and neutralization of the PH reduction was achieved. The results show that the treatment reduced BOD5, COD and TS from its initial concentration of 320.26 mg/l, 1261 mg/l, and 3545 mg/l to 35.66 mg/l, 97 mg/l and 68 mg/l respectively. The treatment in the study shows 87.74% reduced of BOD5 load, 92.02% reduced of COD and 98.01% reduced of TS from the initial load using optimization of the treatment method. Therefore, it can be concluded that mixed culture bacteria of Pseudomonas florescence and Escherichia coli in combination with gypsum as a new effective treatment having potential for BOD5, COD and TS reduce from the effluent treated. The mixed culture of microorganisms is capable to reduce the contaminants that made the approach cost effective, time saving compared with other results found by other authors without mixed culture. The microorganisms included in the study can be applied for the treatment of effluents containing the multiple contaminants.
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    Sugarcane Bagasse-Based Bioethanol Producing Biorefinery Plant Simulation and Technoeconomic Analysis
    (Addis Ababa University, 2019-10) Berhe, Tekle; Hundessa, Desalegn (PhD)
    Energy has been the driving force for economic development. However, the source of energy has become a big issue concerning the environmental pollution. Cellulosic bioethanol is one of the different types of alternative energy sources so far innovated. It is promised to replace the liquid fossil fuel and settle the fuel vs food competition. This area attracts researchers from around the world due to its abundant resource and environmentally friendly production process. Depending on that premises, this simulation-based bioethanol production research paper was dealt to investigate some of the scale up problems and to study the economic feasibility. The research paper carefully simulates the production process and analysis its techno-economic. The conceptual model used on this paper encompasses feedstock handling, pretreatment, saccharification, cofermentation, and production recovery. To validate the equation used by Aspen Plus to simulate enzyme kinetics, the mathematical equation for enzyme kinetics of hydrolysis and cofermentation were tested in Matlab. The techno-economic analysis for the overall production process was then studied depending on two models. The two models were bioethanol plant gate price assessment model (BPAM) and feedstock cost estimation model (FCEM). During the process sensitivity analysis is also performed using Aspen Plus built-in single objective function. The result of the sensitivity analysis showed that, the conversion rate of cellulose to glucose, feedstock cost, and enzyme loading have high factor in the minimum plant gate ethanol selling price determination. The relationship between scaling up and ethanol production capacity was uniformly corelated. The bioethanol for 5000 kg/h and 100000 kg/h of sugarcane bagasse was 296.518164 and 26443.563 respectively. This shows the positive correlation of the scale up and bioethanol production. In addition to that, the overall simulation result showed that the minimum plant gate ethanol selling price at breakeven point was $2.1174 per liter. This result indicates that, even in the most optimistic scenario, the price of cellulosic ethanol is much more expensive than the other similar fuels, gasoline. However, bioethanol can be competitive at some conditions. Some of the conditions that makes the biorefinery plant competitive enough on the local market are feed-in tariff and compulsory purchase of electricity, government direct subsidy, tax preference and R&D. In addition to the above condition, public awareness is also important. Finally, the researcher has recommended to government to revise the above condition to entertain such type of environmentally friendly biorefinery plants.
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    Utilization of Coffee Husk as A Substrate for Vinegar Production Using Acetobacter Aceti Bacteria
    (Addis Ababa University, 2019-06) Piniel, Belachew; Anuradha, Jabasingh (PhD)
    Advancement in industrial biotechnology offer potential opportunities for the economic utilization of agro-industrial residues. Coffee husk is a fibrous by-product obtained during the processing of coffee cherries by dry process. These wastes cause pollution to the environment, unless treated properly. This study employs coffee husk as a substrate to produce acetic acid using acetobacter aceti bacteria in a submerged fermentation process. Vinegar was formulated after the production of acetic acid from coffee husk. Three factors; temperature, ethanol concentration and time were optimized by conducting 17 experiments according to the Design Expert Software 11, using the response surface methodology. The optimum acetic acid concentration (16.59%) was found at 33.76 ℃, 7.84% (ethanol concentration), and 68h respectively and this was analyzed using High Performance Liquid Chromatography (HPLC) followed by the formulation of vinegar to 5% acetic acid.
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    Optimization of Drying Parameters for “Moisture Reduced Teff Injera Making” Using Response Surface Methodology
    (Addis Ababa University, 2019-06) Dessalegn, Abit; Eduardo Ojito, Cespedes (Prof.)
    Teff Injera is the staple Ethiopian fermented bread processed from fermented tef [Eragrostis tef (Zucc.) Trotter] grain flour.” Dirkosh” is the Amharic name given to the dried /moisture reduced injera. The current potential need of the super food, Teff and its products like injera and its dried form, Dirkosh needs a scientific research that will increase its quality, healthy and market demand. Therefore, the objective of the study was to prepare and optimize the drying parameters of Dirkosh using computer controlled tray dryer. The selected drying parameters were (drying air temperature, air velocity and sample loading density) in response of the most quality parameters of the final dried injera/Dirkosh. The response variables were the final moisture content, drying time, color, texture, overall acceptability, water activity and drying rate. The optimization process done by using Box-Behnken Design, response surface methodology (RSM). Drying process factors ( (drying air temperature(55-65 C), air velocity(0.5-1.5m/s) and sample loading density(12kg/m o 2 )) were considered and made 15 experimental runs of Dirkosh drying including 3 replicates using RSM. The investigated responses were final moisture content(5.43), color(6.02), texture(6.23), overall acceptability(6.05), water activity(0.274) but drying time and drying rate were not significant model terms but it was possible to estimate the drying kinetics using the better drying process parameters. RSM with Box-Behnken Design was used to develop and optimize drying process factors in response of the dried injera. Moreover, both numerical and graphical optimizations were carried out to determine the optimum values of Dirkosh in terms significant model terms. The optimum process factors of drying process parameters were drying air temperature of 55 with a desirability of 0.65. The study showed that all the drying process factors were important parameters influencing the physicochemical and sensory qualities of Dirkosh.
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    Extraction of Lectin from Banana Using Acid Solvent
    (Addis Ababa University, 2018-11) Fatuma, Abiyu; Shimeles, Admasu (PhD)
    Banana is a major tropical fruit crop distributed in more than 120 countries with an annual production of 102 million tones. Banana Lectin is a lectin from the jacalin-related lectin family isolated from the fruit of the bananas (Musa acuminate). It exists as a homodimer which are two identical kda (kilodalton) sub unit. The objective of the thesis was extraction of lectin from banana for glycoconjugates. A number of literatures were reviewed for extraction of lectin.Lectin was extracted from ripen and unripen banana. The protein was highly stable and binds to a certain carbohydrate structure. These heterogeneous class of carbohydrate-binding proteins or glycoproteins of non-immune origin are capable of specific recognition of, and reversible binding to, carbohydrates without altering their covalent structure. The maximum amount of lectin (3.6 mg) was found from ripen banana that socked in neutral pH, at 25 o C and 24 hours. And it was checked its glycoconjugates activity using KHB kit and agglutination was observed on the wall of the plate. As socking time increased from 12-36 hours the amount of lectin decreased. Large amount of lectin was observed in neutral pH than in acidic and basic solutions. Besides as temperature increased from 2530 o C, still amount of lectin decreased. This was due to the break-down of protein molecules in long socking time, acidic and basic solutions and high temperature.
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    Optimization of Parameters to Improve the Strength of Concrete Cube Using Bacteria Species
    (AAU, 2018-06) Eman, Negash; Solomon, Kiros (PhD)
    Concrete is the most commonly used building material, but the cracks in concrete create problem it was main reason for a decreased service life of concrete structures. Most research done used chemical method which not effective on cost and environmental. So, need of a new technology to overcome the bottle necking of concrete has led to the development of a very special concrete known as Bacterial Concrete. The objective of this thesis was optimizing of parameters to improve the strength of concrete using bacteria bacillus species (Bacillus subitilis, Sporosarcina pasteurii and Bacillus cibi) self-healing method. Proceed was the bacterial strains were obtained from Ethiopia Biodiversity Institute and for each bacterial growth in nutrient broth solution for 24-72 hour. Secondly, according to the mixing design twenty seven concrete mixes for each day and for each bacterial at C25 grades of concrete using water cement ratio 0.58, 0.53 and 0.45 and cement, fine aggregate and course aggregate contents of 360 kg / m 3, 800 kg / m 3 and 1087.5 kg / m 3 were prepared, respectively and tested in 7, 14 and 28 day. Significance of the process variables were analyzed using analysis of variance (ANOVA). Thus, the influence of all experimental variables, factors and interaction effects on the response was investigated. Dosage, time, water/cement ratio and in the case of bacteria Bacillus cibi use interaction between dosage and water cement ratio have significance effect on the compressive strength. As the result of RSM optimization, the best compressive strength were found on the 0.45 water cement ratio, 40 ml of dosage and the time after 14day. Also from the three bacterial Sporosarcina pasteurii selectable or give high compressive strength.