This document summarizes research on the production and characterization of amylase enzyme from a strain of Aspergillus niger using solid-state fermentation. Key findings include:
- Amylase was produced using optimized fermentation conditions and showed highest activity at pH 4-8 and temperatures of 30-60°C.
- Partial purification using ammonium sulfate increased the specific activity and concentration of amylase.
- The purified enzyme was able to hydrolyze starches from various sources like maize, yam and cassava, producing reducing sugars.
- Different starch sources showed varying susceptibility to hydrolysis, with maize starch being most easily broken down.
Production of alcohol by yeast isolated from apple, orange and bananaPremier Publishers
The purpose of this study was to isolate wild yeast strains present in different fruits (apple, orange and banana) and to determine the yeast growth and the amount of alcohol production at various glucose concentrations. Three fruits namely apple, banana and orange were selected as natural sources for yeast isolation. Medium used for isolation of yeast from fruits was consisting of 50 glucose, 3 malt extract, 3 yeast extract, 5 peptone and 15g/L agar. For fermentation MGYP medium used with different glucose concentrations of 5, 10, 30, 50 and 70g/L. Inocula were prepared by loop transfers from stock slants to 50 mL of the 20 percent medium in 500-mL Erlenmeyer flasks. The results showed that with higher concentration of glucose (30g/L) higher amount of alcohol (0.22g/100mL) was produced. Similarly, the yeast isolated from apple showed maximum yeast biomass (0.38g/100mL) at 70g/L glucose concentration and minimum on 50g/L (0.03g/100mL) glucose concentration. In conclusion, the wild yeast produce higher ethanol amount in case of banana fruits.
IJRET : International Journal of Research in Engineering and Technology is an international peer reviewed, online journal published by eSAT Publishing House for the enhancement of research in various disciplines of Engineering and Technology. The aim and scope of the journal is to provide an academic medium and an important reference for the advancement and dissemination of research results that support high-level learning, teaching and research in the fields of Engineering and Technology. We bring together Scientists, Academician, Field Engineers, Scholars and Students of related fields of Engineering and Technology
Statistical optimization of process parameters for gelatinization of potato (...eSAT Journals
Abstract Potato, a cost effective source of starch, a root vegetable is grown in more than 100 countries in the world. India holds 2nd position in potato production in the world and West Bengal 1st position in India. Potato is processed into a variety of products ranging from potato powder, potato starch, frozen potato flakes, baby food and alcohol, potato chips, French fries, potato flakes/powder etc. In this study, the main objective was statistical optimization of the condition for potato gelatinization and effect of potassium metabisulphite (KMS) concentration on potato starch in gelatinization process. The range of the factors employed were gelatinization pressure, time and different concentration of KMS. The optimized gelatinization condition was 10psig for 15 minutes and 200ppm KMS. The percentage of gelatinized starch at optimized gelatinized condition was 35.67 (dry weight basis). Optimization of effective concentration of potassium metabisulphite (KMS) to retain the colour of potato was also studied and the optimized effective concentration of KMS to retain the colour was 200ppm. RSM modeling, simple regression was used as a tool to study different interactive and linear effect of different factors and significance of the factors. Keywords: potato starch, gelatinization, KMS concentration, colour retention, RSM modeling, regression equation.
Effect of Different Physico-Chemical Parameters on Production ofAmylase by Ba...IOSR Journals
The present study is concerned with the production of amylase by Bacillus species strain. In this
study 12 bacterial strains were isolated and screened for their α-amylase activity. These strains were
maintained on nutrient agar medium. Fermentation for the production of amylase was carried out in the enzyme
production medium (EPM). All the 12 strains were tested for amylase production. On the basis of maximum
amylase activity strain no.1 was selected for further studies. Different starch concentrations, 0.75,1.00,1.25%,
pH labels 6.5,7.0,7.5,8.0, aeration (RPM), 100,120,140, temperatures 250C,280C,370C, and 400C and inoculums
level 0.5%,1.0%, 1.5% and 2.0% were studied
Study on the Use of Agricultural Wastes for Cellulase Production by Using Asp...IOSR Journals
It was the goal to investigate the Cellulase enzyme production ability of fungal strains such as
Aspergillus niger against the lignocellulosic bio-waste like Rice husks, Millet husks, Maize cobs, Wheat straws
and leaves at varying environmental parameters of pH, substrate concentration and dry weight and wet weight.
The enzyme production was analyzed individually by Miller‘s modified method of Dinitrosalicylic acid ( DNSA
). In this study the high level of enzyme production was achieved at pH 3 by using Rice husks, Millet husks,
Maize cobs, Wheat straws and leaves and pH 4 by using Maize cobs at varying substrate concentrations. Out of
all the five used substrates rice husks gave the higher production of the cellulose enzyme. The importance of
cellulase enzyme in industries cannot be over emphasized.
Production of alcohol by yeast isolated from apple, orange and bananaPremier Publishers
The purpose of this study was to isolate wild yeast strains present in different fruits (apple, orange and banana) and to determine the yeast growth and the amount of alcohol production at various glucose concentrations. Three fruits namely apple, banana and orange were selected as natural sources for yeast isolation. Medium used for isolation of yeast from fruits was consisting of 50 glucose, 3 malt extract, 3 yeast extract, 5 peptone and 15g/L agar. For fermentation MGYP medium used with different glucose concentrations of 5, 10, 30, 50 and 70g/L. Inocula were prepared by loop transfers from stock slants to 50 mL of the 20 percent medium in 500-mL Erlenmeyer flasks. The results showed that with higher concentration of glucose (30g/L) higher amount of alcohol (0.22g/100mL) was produced. Similarly, the yeast isolated from apple showed maximum yeast biomass (0.38g/100mL) at 70g/L glucose concentration and minimum on 50g/L (0.03g/100mL) glucose concentration. In conclusion, the wild yeast produce higher ethanol amount in case of banana fruits.
IJRET : International Journal of Research in Engineering and Technology is an international peer reviewed, online journal published by eSAT Publishing House for the enhancement of research in various disciplines of Engineering and Technology. The aim and scope of the journal is to provide an academic medium and an important reference for the advancement and dissemination of research results that support high-level learning, teaching and research in the fields of Engineering and Technology. We bring together Scientists, Academician, Field Engineers, Scholars and Students of related fields of Engineering and Technology
Statistical optimization of process parameters for gelatinization of potato (...eSAT Journals
Abstract Potato, a cost effective source of starch, a root vegetable is grown in more than 100 countries in the world. India holds 2nd position in potato production in the world and West Bengal 1st position in India. Potato is processed into a variety of products ranging from potato powder, potato starch, frozen potato flakes, baby food and alcohol, potato chips, French fries, potato flakes/powder etc. In this study, the main objective was statistical optimization of the condition for potato gelatinization and effect of potassium metabisulphite (KMS) concentration on potato starch in gelatinization process. The range of the factors employed were gelatinization pressure, time and different concentration of KMS. The optimized gelatinization condition was 10psig for 15 minutes and 200ppm KMS. The percentage of gelatinized starch at optimized gelatinized condition was 35.67 (dry weight basis). Optimization of effective concentration of potassium metabisulphite (KMS) to retain the colour of potato was also studied and the optimized effective concentration of KMS to retain the colour was 200ppm. RSM modeling, simple regression was used as a tool to study different interactive and linear effect of different factors and significance of the factors. Keywords: potato starch, gelatinization, KMS concentration, colour retention, RSM modeling, regression equation.
Effect of Different Physico-Chemical Parameters on Production ofAmylase by Ba...IOSR Journals
The present study is concerned with the production of amylase by Bacillus species strain. In this
study 12 bacterial strains were isolated and screened for their α-amylase activity. These strains were
maintained on nutrient agar medium. Fermentation for the production of amylase was carried out in the enzyme
production medium (EPM). All the 12 strains were tested for amylase production. On the basis of maximum
amylase activity strain no.1 was selected for further studies. Different starch concentrations, 0.75,1.00,1.25%,
pH labels 6.5,7.0,7.5,8.0, aeration (RPM), 100,120,140, temperatures 250C,280C,370C, and 400C and inoculums
level 0.5%,1.0%, 1.5% and 2.0% were studied
Study on the Use of Agricultural Wastes for Cellulase Production by Using Asp...IOSR Journals
It was the goal to investigate the Cellulase enzyme production ability of fungal strains such as
Aspergillus niger against the lignocellulosic bio-waste like Rice husks, Millet husks, Maize cobs, Wheat straws
and leaves at varying environmental parameters of pH, substrate concentration and dry weight and wet weight.
The enzyme production was analyzed individually by Miller‘s modified method of Dinitrosalicylic acid ( DNSA
). In this study the high level of enzyme production was achieved at pH 3 by using Rice husks, Millet husks,
Maize cobs, Wheat straws and leaves and pH 4 by using Maize cobs at varying substrate concentrations. Out of
all the five used substrates rice husks gave the higher production of the cellulose enzyme. The importance of
cellulase enzyme in industries cannot be over emphasized.
Proximate and Microbial Profile of Couscous Yoghurt Produced from Soya MilkIJEAB
This study investigated the effect of milk type and mixture ratio on the proximate composition and microbial profile counts of couscous yoghurt. Yoghurts were first made from cow milk (CM), soya milk (SM) and equal mixture of both types of milk at ratio 50:50. Couscous was then mixed with yoghurts from cow milk (CMCY); soya milk (SMCY) and cow-soya milk (CSCY) at ratios of 90:10, 80:20 and 70:30 (yoghurt: couscous), w/w for the three respectively. The experiment was designed based on 2 factors (milk type and mixing ratio) at 3 levels, each resulting in a total of 9 treatments. Cow milk yoghurt without couscous was used as the control. Proximate compositions were determined using standard methods. Total viable microbial counts of samples were also determined. There were significant differences (p<0.05) in the proximate composition and CSCY at ratio 70:30 had the highest crude protein. In addition, CMCY at ratio 90:10 recorded the highest mean value for fat, while SMCY at ratio 80:20 and 70:30 recorded the least mean value for fat. All the couscous yoghurt samples had total viable cell counts of (<9 log CFU) that are within the acceptable range according to Codex Standards. In conclusion, the study has shown that CSCY at 70:30 had the highest nutrient content. Moreover, the products were also found to have low levels of microbial profile.
Optimization of Cultural Parameters for Cellulase Enzyme Production from Fung...IOSR Journals
Cellulalytic fungi synthesize cellulose enzyme for biodegradation of cellulose. This depends on various condition which include the source f isolation. This study was designed to determine the optimum condition necessary for cellulose production by fungi. Cellulose activities at different temperatures, pH and nitrogen sources by Rhizopus oryzae Aspergillus niger; A. flams, P. expansum and A. oryzae in liquid medium was studied and cellulose enzyme assay carried out by dinitrosalicylic acid method. All the fungal isolates have their highest cellulose activity at 400c except Penicillium expansum whose highest value of 1.28mg/ml was obtained at 320c. Cellulase produced 6m was found to be highest in all the isolate at pH 4.0 exception P expansum which occur at pH 5.5 (1.21mg/ml). The highest value e1.45mg/ml was obtained in A niger. Highest cellulose activity for A. niger, A. oryzae & P. expansum occurred in peptone. The study shows the need to determine the best physiological condition that allow for the optimal cellulose activity of fungal isolate. This will enhance their enzyme production.
Effect of Different Drying Methods on Chemical Composition of Unripe Plantain...YogeshIJTSRD
Food processing is often thought to bring about changes in nutrients content, thus decreasing its patronage. To investigate this in a Nigerian staple, unripe plantain Musa paradisiaca flours were prepared following sun drying and oven drying methods. These were compared against fresh plantain for their nutritional composition. Proximate composition and minerals contents were determined using standard AOAC methods. The results showed that the unripe plantains pulp contained 59.77 , 1.42 , 1.51 , 1.40 , 7.65 , 28.23 , 40.22 and 38.80 of moisture, ash, fat oils, crude fibre, crude protein, carbohydrates, dry matter and organic matter respectively. Calcium, sodium, potassium, iron, and nitrogen were determined to be 0.1534 ppm, 0.2613 ppm, 0.3034 ppm, 0.7808 ppm and 0.2240 ppm respectively. The processing methods produced flour with similar nutritional composition. However, oven drying gave the lowest moisture content in the flour, suggesting a higher capacity to prevent microbial growth and decay in the dried sample, hence prolonging storage life. Segilola, V. O | Amodu, S. O | Olatunji, C. A "Effect of Different Drying Methods on Chemical Composition of Unripe Plantain Flour" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-5 | Issue-3 , April 2021, URL: https://www.ijtsrd.com/papers/ijtsrd38725.pdf Paper URL: https://www.ijtsrd.com/biological-science/allied-sciences/38725/effect-of-different-drying-methods-on-chemical-composition-of-unripe-plantain-flour/segilola-v-o
Fungal cellulase xylanase production and corresponding hydrolysis using pretr...zhenhua82
Three pretreated corn stover (ammonia fiber expansion, dilute acid, and dilute alkali) were used as carbon source to culture Trichoderma reesei Rut C-30 for cellulase and xylanase production. The results indicated that the cultures on ammonia fiber expansion and alkali pretreated corn stover had better enzyme production than the acid pretreated ones. The consequent enzymatic hydrolysis was performed applying fungal enzymes on pretreated corn stover samples. Tukey’s statistical comparisons exhibited that there were significant differences on enzymatic hydrolysis among different combination of fungal enzymes and pretreated corn stover. The higher sugar yields were achieved by the enzymatic hydrolysis of dilute alkali pretreated corn stover.
Utilization of Banana Peel Powder in ConcreteYogeshIJTSRD
Analysis of properties of concrete using banana peel as admixture is studied and verified the strength of concrete and temperature emitted due to chemical reaction to the normal Portland cement. The percentage of transmission temperature and reduction time of temperature has decreased hence it is clear that the exothermal reaction in concrete has been reduced by using dried banana peel powder as admixture. The percentage of transmission temperature and reduction time of temperature has decreased hence it is clear that the exothermal reaction in concrete has been reduced by using dried banana peel powder as admixture. Ingredients other than cement, water and aggregates that import a specific quality to either plastic fresh mix or the hardened concrete ASTMC 496 is called concrete admixture. The flexural strength of concrete by using banana peel powder as admixture has increased, but considerable lesser compressive strength has increased. Rahul Mohabe | G. D. Dhawade | R. K. Kakpure "Utilization of Banana Peel Powder in Concrete" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-5 | Issue-4 , June 2021, URL: https://www.ijtsrd.compapers/ijtsrd41186.pdf Paper URL: https://www.ijtsrd.comengineering/civil-engineering/41186/utilization-of-banana-peel-powder-in-concrete/rahul-mohabe
protease activity of extracellular enzyme produced by b. subtilis isolated fr...IJEAB
Background: Proteases produced by enzymatic method are more environments friendly than chemical process, and they have tremendous potential in the leather industry and in other several industries. In this study extracellular protease producing non pathogenic Bacillus subtilis was isolated from soil sample and relationship between sporulation and extracellular protease synthesis in large scale cultivation was studied. The enzyme was further characterized, purified, and tested for potential application. Result: The molecular weight of the protease was found to be ~30 KDa. Enzyme activity was checked on the presence of different metal ions and effectors. The enzyme was slightly modulated by MG++ ion, and significantly by Hg++ ion, while Zn++ ion slightly decrease the proteolytic activity. Sulfahydryl reagents, DTT slightly and β-ME significantly inhibit the enzyme. EDTA showed no effect on the enzyme suggesting that the enzyme might not be metalloprotease. PMSF, a known serine protease inhibitor was seen to totally inhibit the enzyme which indicates that the enzyme is a serine protease. The optimum enzyme activity was observed after 22 hours of incubation of B. subtilis at 37o C. Conclusions: Crude enzyme contains 285 units of enzyme which have direct dehairing activity. The enzyme was also seen to be able to remove blood and curry stain from clothes; making it a very promising candidate to be used in a leather and detergent industry. Apart from protease the bacterium was also seen to have lipase and collagenase activity. So, the bacteria are potentially good candidate for industrial application.
Utilization of Banana Peel Powder in Concrete A Resultijtsrd
Analysis of properties of concrete using banana peel as admixture is studied and verified the strength of concrete and temperature emitted due to chemical reaction to the normal Portland cement. The percentage of transmission temperature and reduction time of temperature has decreased hence it is clear that the exothermal reaction in concrete has been reduced by using dried banana peel powder as admixture. The percentage of transmission temperature and reduction time of temperature has decreased hence it is clear that the exothermal reaction in concrete has been reduced by using dried banana peel powder as admixture. Ingredients other than cement, water and aggregates that import a specific quality to either plastic fresh mix or the hardened concrete ASTMC 496 is called concrete admixture. The flexural strength of concrete by using banana peel powder as admixture has increased, but considerable lesser compressive strength has increased. Rahul M Mohabe | Prof. G. D. Dhavale | R. K. Kakpure "Utilization of Banana Peel Powder in Concrete: A Result" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-5 | Issue-4 , June 2021, URL: https://www.ijtsrd.compapers/ijtsrd42560.pdf Paper URL: https://www.ijtsrd.comengineering/civil-engineering/42560/utilization-of-banana-peel-powder-in-concrete-a-result/rahul-m-mohabe
IJRET : International Journal of Research in Engineering and Technology is an international peer reviewed, online journal published by eSAT Publishing House for the enhancement of research in various disciplines of Engineering and Technology. The aim and scope of the journal is to provide an academic medium and an important reference for the advancement and dissemination of research results that support high-level learning, teaching and research in the fields of Engineering and Technology. We bring together Scientists, Academician, Field Engineers, Scholars and Students of related fields of Engineering and Technology.
Isolation and characterization of plant growth promoting bacteria (PGPB) from...Agriculture Journal IJOEAR
— The use of anaerobic digestate as fertilizer is considered beneficial since it provides plant nutrients and organic matter to soils. However, there is limited information about plant growth promoting bacteria (PGPB) in digestate. In this study, we isolated Bacillus and Pseudomonas from two types of anaerobic digestates, and selected three different plant growth promoting traits and antifungal activity to screen 200 bacteria isolated from each digestate. Then 6 isolates based on plant growth promoting traits were selected and inoculated with common wheat seeds to evaluate their plant growth promoting activity. Cultivable population of Bacillus and Pseudomonas were 2.20 × 10 6 and 6.98 × 10 4 CFU g-1 dry matter in mesophilic digestate, while were 6.86 × 10 5 and 5.65 × 10 4 CFU g-1 dry matter in thermophilic digestate. Twenty-five bacterial isolates from mesophilic digestate and 12 bacterial isolates from thermophilic digestate showed positive plant growth promoting traits or antifungal activity. In plant growth promoting assay, all isolates significantly promoted growth of wheat seedlings (p < 0.05). Seedlings stem length was increased from 28.5% to 38.6% by bacteria inoculation. In addition, bacteria inoculation increased seedlings stem weight from 113.3% to 214.2% and root weight from 108.6% to 207.2% as compared to un-inoculated control. The results showed that anaerobic digestate was a potential source for isolation of PGPB, and PGPB in digestate would be beneficial for plant growth with fertilizer application.
Production and Purification of Amylase from Bacillus subtilis Isolated from SoilDr. Amarjeet Singh
In spite of progress in biotechnology and
enzymology, the enzymes have been industrialized in recent
years for the mounting up the product development in
various arena. The ultimate goal of this study comprises the
production and purification the amylase enzyme from the
bacterial strain. A powerful amylase producer, Bacillus
subtilis ISOLATE-4 was isolated, screened and identified
from the soil sample. In order to produce extracellular
amylase, various physico-chemical parameters were
optimized. During optimization, the maximal production of
amylase by the isolate at 48 hrs of incubation in 100 rpm was
found to be 6.93U/ml, 5.94U/ml, 6.0U/ml at 45ºC, pH 6 with
1% substrate concentration respectively. Ammonium
sulphate fractionation was done for rapid precipitation of the
amylase at a concentration of 60% and exposed to dialysis
showed the 25% purification fold of an enzyme. The dialyzed
product was further subjected to DEAE-Cellulose column
chromatography resulted in an increase up to 75%
purification fold than crude enzyme. The amylase enzyme
might be suitable for the liquefaction of starch, detergent,
textile and several additional industrial applications.
Proximate and Microbial Profile of Couscous Yoghurt Produced from Soya MilkIJEAB
This study investigated the effect of milk type and mixture ratio on the proximate composition and microbial profile counts of couscous yoghurt. Yoghurts were first made from cow milk (CM), soya milk (SM) and equal mixture of both types of milk at ratio 50:50. Couscous was then mixed with yoghurts from cow milk (CMCY); soya milk (SMCY) and cow-soya milk (CSCY) at ratios of 90:10, 80:20 and 70:30 (yoghurt: couscous), w/w for the three respectively. The experiment was designed based on 2 factors (milk type and mixing ratio) at 3 levels, each resulting in a total of 9 treatments. Cow milk yoghurt without couscous was used as the control. Proximate compositions were determined using standard methods. Total viable microbial counts of samples were also determined. There were significant differences (p<0.05) in the proximate composition and CSCY at ratio 70:30 had the highest crude protein. In addition, CMCY at ratio 90:10 recorded the highest mean value for fat, while SMCY at ratio 80:20 and 70:30 recorded the least mean value for fat. All the couscous yoghurt samples had total viable cell counts of (<9 log CFU) that are within the acceptable range according to Codex Standards. In conclusion, the study has shown that CSCY at 70:30 had the highest nutrient content. Moreover, the products were also found to have low levels of microbial profile.
Optimization of Cultural Parameters for Cellulase Enzyme Production from Fung...IOSR Journals
Cellulalytic fungi synthesize cellulose enzyme for biodegradation of cellulose. This depends on various condition which include the source f isolation. This study was designed to determine the optimum condition necessary for cellulose production by fungi. Cellulose activities at different temperatures, pH and nitrogen sources by Rhizopus oryzae Aspergillus niger; A. flams, P. expansum and A. oryzae in liquid medium was studied and cellulose enzyme assay carried out by dinitrosalicylic acid method. All the fungal isolates have their highest cellulose activity at 400c except Penicillium expansum whose highest value of 1.28mg/ml was obtained at 320c. Cellulase produced 6m was found to be highest in all the isolate at pH 4.0 exception P expansum which occur at pH 5.5 (1.21mg/ml). The highest value e1.45mg/ml was obtained in A niger. Highest cellulose activity for A. niger, A. oryzae & P. expansum occurred in peptone. The study shows the need to determine the best physiological condition that allow for the optimal cellulose activity of fungal isolate. This will enhance their enzyme production.
Effect of Different Drying Methods on Chemical Composition of Unripe Plantain...YogeshIJTSRD
Food processing is often thought to bring about changes in nutrients content, thus decreasing its patronage. To investigate this in a Nigerian staple, unripe plantain Musa paradisiaca flours were prepared following sun drying and oven drying methods. These were compared against fresh plantain for their nutritional composition. Proximate composition and minerals contents were determined using standard AOAC methods. The results showed that the unripe plantains pulp contained 59.77 , 1.42 , 1.51 , 1.40 , 7.65 , 28.23 , 40.22 and 38.80 of moisture, ash, fat oils, crude fibre, crude protein, carbohydrates, dry matter and organic matter respectively. Calcium, sodium, potassium, iron, and nitrogen were determined to be 0.1534 ppm, 0.2613 ppm, 0.3034 ppm, 0.7808 ppm and 0.2240 ppm respectively. The processing methods produced flour with similar nutritional composition. However, oven drying gave the lowest moisture content in the flour, suggesting a higher capacity to prevent microbial growth and decay in the dried sample, hence prolonging storage life. Segilola, V. O | Amodu, S. O | Olatunji, C. A "Effect of Different Drying Methods on Chemical Composition of Unripe Plantain Flour" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-5 | Issue-3 , April 2021, URL: https://www.ijtsrd.com/papers/ijtsrd38725.pdf Paper URL: https://www.ijtsrd.com/biological-science/allied-sciences/38725/effect-of-different-drying-methods-on-chemical-composition-of-unripe-plantain-flour/segilola-v-o
Fungal cellulase xylanase production and corresponding hydrolysis using pretr...zhenhua82
Three pretreated corn stover (ammonia fiber expansion, dilute acid, and dilute alkali) were used as carbon source to culture Trichoderma reesei Rut C-30 for cellulase and xylanase production. The results indicated that the cultures on ammonia fiber expansion and alkali pretreated corn stover had better enzyme production than the acid pretreated ones. The consequent enzymatic hydrolysis was performed applying fungal enzymes on pretreated corn stover samples. Tukey’s statistical comparisons exhibited that there were significant differences on enzymatic hydrolysis among different combination of fungal enzymes and pretreated corn stover. The higher sugar yields were achieved by the enzymatic hydrolysis of dilute alkali pretreated corn stover.
Utilization of Banana Peel Powder in ConcreteYogeshIJTSRD
Analysis of properties of concrete using banana peel as admixture is studied and verified the strength of concrete and temperature emitted due to chemical reaction to the normal Portland cement. The percentage of transmission temperature and reduction time of temperature has decreased hence it is clear that the exothermal reaction in concrete has been reduced by using dried banana peel powder as admixture. The percentage of transmission temperature and reduction time of temperature has decreased hence it is clear that the exothermal reaction in concrete has been reduced by using dried banana peel powder as admixture. Ingredients other than cement, water and aggregates that import a specific quality to either plastic fresh mix or the hardened concrete ASTMC 496 is called concrete admixture. The flexural strength of concrete by using banana peel powder as admixture has increased, but considerable lesser compressive strength has increased. Rahul Mohabe | G. D. Dhawade | R. K. Kakpure "Utilization of Banana Peel Powder in Concrete" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-5 | Issue-4 , June 2021, URL: https://www.ijtsrd.compapers/ijtsrd41186.pdf Paper URL: https://www.ijtsrd.comengineering/civil-engineering/41186/utilization-of-banana-peel-powder-in-concrete/rahul-mohabe
protease activity of extracellular enzyme produced by b. subtilis isolated fr...IJEAB
Background: Proteases produced by enzymatic method are more environments friendly than chemical process, and they have tremendous potential in the leather industry and in other several industries. In this study extracellular protease producing non pathogenic Bacillus subtilis was isolated from soil sample and relationship between sporulation and extracellular protease synthesis in large scale cultivation was studied. The enzyme was further characterized, purified, and tested for potential application. Result: The molecular weight of the protease was found to be ~30 KDa. Enzyme activity was checked on the presence of different metal ions and effectors. The enzyme was slightly modulated by MG++ ion, and significantly by Hg++ ion, while Zn++ ion slightly decrease the proteolytic activity. Sulfahydryl reagents, DTT slightly and β-ME significantly inhibit the enzyme. EDTA showed no effect on the enzyme suggesting that the enzyme might not be metalloprotease. PMSF, a known serine protease inhibitor was seen to totally inhibit the enzyme which indicates that the enzyme is a serine protease. The optimum enzyme activity was observed after 22 hours of incubation of B. subtilis at 37o C. Conclusions: Crude enzyme contains 285 units of enzyme which have direct dehairing activity. The enzyme was also seen to be able to remove blood and curry stain from clothes; making it a very promising candidate to be used in a leather and detergent industry. Apart from protease the bacterium was also seen to have lipase and collagenase activity. So, the bacteria are potentially good candidate for industrial application.
Utilization of Banana Peel Powder in Concrete A Resultijtsrd
Analysis of properties of concrete using banana peel as admixture is studied and verified the strength of concrete and temperature emitted due to chemical reaction to the normal Portland cement. The percentage of transmission temperature and reduction time of temperature has decreased hence it is clear that the exothermal reaction in concrete has been reduced by using dried banana peel powder as admixture. The percentage of transmission temperature and reduction time of temperature has decreased hence it is clear that the exothermal reaction in concrete has been reduced by using dried banana peel powder as admixture. Ingredients other than cement, water and aggregates that import a specific quality to either plastic fresh mix or the hardened concrete ASTMC 496 is called concrete admixture. The flexural strength of concrete by using banana peel powder as admixture has increased, but considerable lesser compressive strength has increased. Rahul M Mohabe | Prof. G. D. Dhavale | R. K. Kakpure "Utilization of Banana Peel Powder in Concrete: A Result" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-5 | Issue-4 , June 2021, URL: https://www.ijtsrd.compapers/ijtsrd42560.pdf Paper URL: https://www.ijtsrd.comengineering/civil-engineering/42560/utilization-of-banana-peel-powder-in-concrete-a-result/rahul-m-mohabe
IJRET : International Journal of Research in Engineering and Technology is an international peer reviewed, online journal published by eSAT Publishing House for the enhancement of research in various disciplines of Engineering and Technology. The aim and scope of the journal is to provide an academic medium and an important reference for the advancement and dissemination of research results that support high-level learning, teaching and research in the fields of Engineering and Technology. We bring together Scientists, Academician, Field Engineers, Scholars and Students of related fields of Engineering and Technology.
Isolation and characterization of plant growth promoting bacteria (PGPB) from...Agriculture Journal IJOEAR
— The use of anaerobic digestate as fertilizer is considered beneficial since it provides plant nutrients and organic matter to soils. However, there is limited information about plant growth promoting bacteria (PGPB) in digestate. In this study, we isolated Bacillus and Pseudomonas from two types of anaerobic digestates, and selected three different plant growth promoting traits and antifungal activity to screen 200 bacteria isolated from each digestate. Then 6 isolates based on plant growth promoting traits were selected and inoculated with common wheat seeds to evaluate their plant growth promoting activity. Cultivable population of Bacillus and Pseudomonas were 2.20 × 10 6 and 6.98 × 10 4 CFU g-1 dry matter in mesophilic digestate, while were 6.86 × 10 5 and 5.65 × 10 4 CFU g-1 dry matter in thermophilic digestate. Twenty-five bacterial isolates from mesophilic digestate and 12 bacterial isolates from thermophilic digestate showed positive plant growth promoting traits or antifungal activity. In plant growth promoting assay, all isolates significantly promoted growth of wheat seedlings (p < 0.05). Seedlings stem length was increased from 28.5% to 38.6% by bacteria inoculation. In addition, bacteria inoculation increased seedlings stem weight from 113.3% to 214.2% and root weight from 108.6% to 207.2% as compared to un-inoculated control. The results showed that anaerobic digestate was a potential source for isolation of PGPB, and PGPB in digestate would be beneficial for plant growth with fertilizer application.
Production and Purification of Amylase from Bacillus subtilis Isolated from SoilDr. Amarjeet Singh
In spite of progress in biotechnology and
enzymology, the enzymes have been industrialized in recent
years for the mounting up the product development in
various arena. The ultimate goal of this study comprises the
production and purification the amylase enzyme from the
bacterial strain. A powerful amylase producer, Bacillus
subtilis ISOLATE-4 was isolated, screened and identified
from the soil sample. In order to produce extracellular
amylase, various physico-chemical parameters were
optimized. During optimization, the maximal production of
amylase by the isolate at 48 hrs of incubation in 100 rpm was
found to be 6.93U/ml, 5.94U/ml, 6.0U/ml at 45ºC, pH 6 with
1% substrate concentration respectively. Ammonium
sulphate fractionation was done for rapid precipitation of the
amylase at a concentration of 60% and exposed to dialysis
showed the 25% purification fold of an enzyme. The dialyzed
product was further subjected to DEAE-Cellulose column
chromatography resulted in an increase up to 75%
purification fold than crude enzyme. The amylase enzyme
might be suitable for the liquefaction of starch, detergent,
textile and several additional industrial applications.
Enhancing the Nutritive Values of Agrowastes for Animal Feed Production Using...iosrjce
IOSR Journal of Environmental Science, Toxicology and Food Technology (IOSR-JESTFT) multidisciplinary peer-reviewed Journal with reputable academics and experts as board member. IOSR-JESTFT is designed for the prompt publication of peer-reviewed articles in all areas of subject. The journal articles will be accessed freely online.
Cassava Retting Water An Alternative Source for Industrial Cellulase EnzymeYogeshIJTSRD
Cassava fermentation is one of the teaming businesses in almost all the tribes of Nigeria. Among all the methods of cassava processing to food, fermentation is the most used. This produces foul smelling waste water that causes environmental pollution to man and animals. The retted cassava water was checked for cellulase enzyme activity to see if it can be a source of cellulase for used in food, paper, textile and other industries. The aim of this work is to seek a way of utilizing the waste water as source of enzymes as this will reduce the importation of these enzymes and make them available always. Cassava tubers were peeled, cut into cylindrical portions of about 3 5 cm and washed. Two hundred grams of the washed tubers were soaked in 5 liters of water and allowed to ret. The retting water was analyzed daily for titratable acidity, cyanide content, pH, cellulase activity and the microbial flora were isolated and identified. Results showed that titratable acidity rose from 0.20 to 2.76 mg g and cyanide content increased from 0.28 to 4.69 mg ml while pH fall from 7.2 – 6.0 tending acidic. Retting started on the 2nd day and complete retting was achieved on the 4th day. ß glucosidase activity rose from 0.05 to 8.0 µ mol, Filter paper activity increased from 0.06 to 7.5 µ mol and carboxyl methyl cellulase CMC activity increased from 0.05 to 7.7 µ mol. Ten organisms Aspergillus fumigatus, Rhizopus stolonifer, Bacillus subtilis, Candida utilis, Citrobacter sp, Enterobacter aerogenes, Lactobacillus coryneformis, Saccharomyces cerevisiae, Staphylococcus aureus, Streptococcus feacalis were isolated from the retting water. Daily increase in the enzyme activities showed that cassava retting when done in a large scale can yield large quantity of enzymes. This will reduce the importation of industrial enzymes and reduce the environmental pollution caused by the waste water. Umeh, S. O. | Nwiyi, I. U. | Dimejesi, S. A. | Ikele, M. O. | Ugwu, C. H. "Cassava Retting Water: An Alternative Source for Industrial Cellulase (Enzyme)" Published in International Journal of Trend in Scientific Research and Development (ijtsrd), ISSN: 2456-6470, Volume-5 | Issue-5 , August 2021, URL: https://www.ijtsrd.com/papers/ijtsrd43889.pdf Paper URL: https://www.ijtsrd.com/biological-science/microbiology/43889/cassava-retting-water-an-alternative-source-for-industrial-cellulase-enzyme/umeh-s-o
ABSTRACT- Microbial source of amylase is preferred to other sources because of its plasticity, vast availability, higher yield and
thermostability even at elevated temperatures.Various physical and chemical factors have been known to affect the production of α-
amylase such as temperature, pH, period of incubation, carbon sources acting as inducers, surfactants, nitrogen sources, phosphate,
different metal ions, moisture. Interactions of these parameters are reported to have a significant influence on the production of
the enzyme.Study was mainly aimed to isolate a bacterium capable of hydrolyzing a starch source and to check effect of different physiological
parameters on amylase enzyme activity. To conduct this research, study was mainly focused on three objectives i.e. 1st Screening
and morphological characterization of the isolated bacteria. 2nd Characterization of amylase production by selected isolates. 3rd
Time course of Enzyme production and Partial purification with Ammonium Sulphate saturation.Amylases of isolate-6 and isolate-9
were concentrated by ammonium sulfate precipitation which can be used as partially purified enzyme for further study. Isolate-6 and
Isolate-9 showed the activity 0.34 and 0.28 units/ml/min respectively.Enzyme derived from isolate-6 and isolate-9 was stable at different
physiological conditions. So, it is useful in fermentation industry and in pharmaceuticals.
Key words- Amylase, Starch hydrolyzing bacteria, fermentation and pharmaceutical industries
IOSR Journal of Pharmacy and Biological Sciences(IOSR-JPBS) is an open access international journal that provides rapid publication (within a month) of articles in all areas of Pharmacy and Biological Science. The journal welcomes publications of high quality papers on theoretical developments and practical applications in Pharmacy and Biological Science. Original research papers, state-of-the-art reviews, and high quality technical notes are invited for publications.
Production of α-amylase using new strain of Bacillus polymyxa isolated from s...IOSR Journals
In this study, a new amylase producer strain was isolated from sweet potato tuber. This strain was able to grow at 37 °C and produce α-amylase in high quantity compared to other standard strain cultures. In the first part, cultivation in shake flask in standard medium was carried out to give complete information about the growth and production kinetics of this strain. The results clearly demonstrate that the isolated strain is able to production α-amylase in submerged culture with concentration up to 2050 kat/L after 20 h cultivation. Furthermore, medium optimization was carried out by changing the starch concentration and cell cultivation in medium of mixed carbon source (composed of starch and glucose of ratio 15:5 g/g) to enhance the production process and to increase the growth rate. The volumetric and specific α-amylase production in this optimized medium were 4550 kat/L and 1060 kat/g, respectively. Further improvement in enzyme production process was achieved by scaling up the process from shake flask to 3-L stirred tank bioreactor under non-oxygen limiting condition. The maximal volumetric and specific α-amylase productions in bioreactor batch culture were 5210 kat/L and 1095kat/g, respectively, after only 14 h cultivation
Detection of Alpha-Amylase Activity from Soil Bacteriaiosrjce
Alpha-amylase is one of the industrial enzymes that hydrolyze starch molecules into polymers
composed of glucose units. The enzyme has potential application in a wide number of industrial processes such
as food, textile, paper, detergent, fermentation and pharmaceutical industries. Alpha-amylase can be produced
by microorganisms, plants or animals.
Aim: The aim of this study is to detect the activity of alpha-amylase from bacteria isolated from soil
environment.
Method: Soil samples were inoculated onto the media that are rich in nutrient that favour the growth of the
bacteria and incubated for 24 hours at 37oC after which the bacterial growth was detected in form of colonies.
In this study, bacterial species belonging to the genus Bacillus were identified through phylogenetic analysis
using 16s-ribosomal RNA sequencing for detection of the enzymatic activity. Effects of pH and temperature on
the enzymatic activity were observed using DNS activity assay method.
Results: Positive response to alpha-amylase activity by the soil bacteria was observed by the formation of clear
zone of inhibition shown by the colonies on the petri plates.
Conclusions: The optimal pH and temperature activities showed that the bacteria exhibit enzymatic activity at
mesophilic temperature and acidophilic or alkalophilic pH.
IJERA (International journal of Engineering Research and Applications) is International online, ... peer reviewed journal. For more detail or submit your article, please visit www.ijera.com
This pdf is about the Schizophrenia.
For more details visit on YouTube; @SELF-EXPLANATORY;
https://www.youtube.com/channel/UCAiarMZDNhe1A3Rnpr_WkzA/videos
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Cancer cell metabolism: special Reference to Lactate PathwayAADYARAJPANDEY1
Normal Cell Metabolism:
Cellular respiration describes the series of steps that cells use to break down sugar and other chemicals to get the energy we need to function.
Energy is stored in the bonds of glucose and when glucose is broken down, much of that energy is released.
Cell utilize energy in the form of ATP.
The first step of respiration is called glycolysis. In a series of steps, glycolysis breaks glucose into two smaller molecules - a chemical called pyruvate. A small amount of ATP is formed during this process.
Most healthy cells continue the breakdown in a second process, called the Kreb's cycle. The Kreb's cycle allows cells to “burn” the pyruvates made in glycolysis to get more ATP.
The last step in the breakdown of glucose is called oxidative phosphorylation (Ox-Phos).
It takes place in specialized cell structures called mitochondria. This process produces a large amount of ATP. Importantly, cells need oxygen to complete oxidative phosphorylation.
If a cell completes only glycolysis, only 2 molecules of ATP are made per glucose. However, if the cell completes the entire respiration process (glycolysis - Kreb's - oxidative phosphorylation), about 36 molecules of ATP are created, giving it much more energy to use.
IN CANCER CELL:
Unlike healthy cells that "burn" the entire molecule of sugar to capture a large amount of energy as ATP, cancer cells are wasteful.
Cancer cells only partially break down sugar molecules. They overuse the first step of respiration, glycolysis. They frequently do not complete the second step, oxidative phosphorylation.
This results in only 2 molecules of ATP per each glucose molecule instead of the 36 or so ATPs healthy cells gain. As a result, cancer cells need to use a lot more sugar molecules to get enough energy to survive.
Unlike healthy cells that "burn" the entire molecule of sugar to capture a large amount of energy as ATP, cancer cells are wasteful.
Cancer cells only partially break down sugar molecules. They overuse the first step of respiration, glycolysis. They frequently do not complete the second step, oxidative phosphorylation.
This results in only 2 molecules of ATP per each glucose molecule instead of the 36 or so ATPs healthy cells gain. As a result, cancer cells need to use a lot more sugar molecules to get enough energy to survive.
introduction to WARBERG PHENOMENA:
WARBURG EFFECT Usually, cancer cells are highly glycolytic (glucose addiction) and take up more glucose than do normal cells from outside.
Otto Heinrich Warburg (; 8 October 1883 – 1 August 1970) In 1931 was awarded the Nobel Prize in Physiology for his "discovery of the nature and mode of action of the respiratory enzyme.
WARNBURG EFFECT : cancer cells under aerobic (well-oxygenated) conditions to metabolize glucose to lactate (aerobic glycolysis) is known as the Warburg effect. Warburg made the observation that tumor slices consume glucose and secrete lactate at a higher rate than normal tissues.
Earliest Galaxies in the JADES Origins Field: Luminosity Function and Cosmic ...Sérgio Sacani
We characterize the earliest galaxy population in the JADES Origins Field (JOF), the deepest
imaging field observed with JWST. We make use of the ancillary Hubble optical images (5 filters
spanning 0.4−0.9µm) and novel JWST images with 14 filters spanning 0.8−5µm, including 7 mediumband filters, and reaching total exposure times of up to 46 hours per filter. We combine all our data
at > 2.3µm to construct an ultradeep image, reaching as deep as ≈ 31.4 AB mag in the stack and
30.3-31.0 AB mag (5σ, r = 0.1” circular aperture) in individual filters. We measure photometric
redshifts and use robust selection criteria to identify a sample of eight galaxy candidates at redshifts
z = 11.5 − 15. These objects show compact half-light radii of R1/2 ∼ 50 − 200pc, stellar masses of
M⋆ ∼ 107−108M⊙, and star-formation rates of SFR ∼ 0.1−1 M⊙ yr−1
. Our search finds no candidates
at 15 < z < 20, placing upper limits at these redshifts. We develop a forward modeling approach to
infer the properties of the evolving luminosity function without binning in redshift or luminosity that
marginalizes over the photometric redshift uncertainty of our candidate galaxies and incorporates the
impact of non-detections. We find a z = 12 luminosity function in good agreement with prior results,
and that the luminosity function normalization and UV luminosity density decline by a factor of ∼ 2.5
from z = 12 to z = 14. We discuss the possible implications of our results in the context of theoretical
models for evolution of the dark matter halo mass function.
What is greenhouse gasses and how many gasses are there to affect the Earth.moosaasad1975
What are greenhouse gasses how they affect the earth and its environment what is the future of the environment and earth how the weather and the climate effects.
Deep Behavioral Phenotyping in Systems Neuroscience for Functional Atlasing a...Ana Luísa Pinho
Functional Magnetic Resonance Imaging (fMRI) provides means to characterize brain activations in response to behavior. However, cognitive neuroscience has been limited to group-level effects referring to the performance of specific tasks. To obtain the functional profile of elementary cognitive mechanisms, the combination of brain responses to many tasks is required. Yet, to date, both structural atlases and parcellation-based activations do not fully account for cognitive function and still present several limitations. Further, they do not adapt overall to individual characteristics. In this talk, I will give an account of deep-behavioral phenotyping strategies, namely data-driven methods in large task-fMRI datasets, to optimize functional brain-data collection and improve inference of effects-of-interest related to mental processes. Key to this approach is the employment of fast multi-functional paradigms rich on features that can be well parametrized and, consequently, facilitate the creation of psycho-physiological constructs to be modelled with imaging data. Particular emphasis will be given to music stimuli when studying high-order cognitive mechanisms, due to their ecological nature and quality to enable complex behavior compounded by discrete entities. I will also discuss how deep-behavioral phenotyping and individualized models applied to neuroimaging data can better account for the subject-specific organization of domain-general cognitive systems in the human brain. Finally, the accumulation of functional brain signatures brings the possibility to clarify relationships among tasks and create a univocal link between brain systems and mental functions through: (1) the development of ontologies proposing an organization of cognitive processes; and (2) brain-network taxonomies describing functional specialization. To this end, tools to improve commensurability in cognitive science are necessary, such as public repositories, ontology-based platforms and automated meta-analysis tools. I will thus discuss some brain-atlasing resources currently under development, and their applicability in cognitive as well as clinical neuroscience.
A brief information about the SCOP protein database used in bioinformatics.
The Structural Classification of Proteins (SCOP) database is a comprehensive and authoritative resource for the structural and evolutionary relationships of proteins. It provides a detailed and curated classification of protein structures, grouping them into families, superfamilies, and folds based on their structural and sequence similarities.
Comparing Evolved Extractive Text Summary Scores of Bidirectional Encoder Rep...University of Maribor
Slides from:
11th International Conference on Electrical, Electronics and Computer Engineering (IcETRAN), Niš, 3-6 June 2024
Track: Artificial Intelligence
https://www.etran.rs/2024/en/home-english/
Seminar of U.V. Spectroscopy by SAMIR PANDASAMIR PANDA
Spectroscopy is a branch of science dealing the study of interaction of electromagnetic radiation with matter.
Ultraviolet-visible spectroscopy refers to absorption spectroscopy or reflect spectroscopy in the UV-VIS spectral region.
Ultraviolet-visible spectroscopy is an analytical method that can measure the amount of light received by the analyte.
Hydrolytic activity of amylase produced in solid state fermentation by a local isolate of aspergillus niger
1. International Journal of Scientific and Research Publications, Volume 8, Issue 7, July 2018 125
ISSN 2250-3153
http://dx.doi.org/10.29322/IJSRP.8.6.2018.p7821 www.ijsrp.org
Hydrolytic activity of amylase produced in solid - state
fermentation by a local isolate of Aspergillus niger
Samuel Kwatia*
, Victoria P. Dzogbefia*
Department of Biochemistry and Biotechnology, Kwame Nkrumah University of Science and Technology, Kumasi, Ghana
DOI: 10.29322/IJSRP.8.7.2018.p7921
http://dx.doi.org/10.29322/IJSRP.8.7.2018.p7921
Abstract- The increase in demand for local enzymes for industrial activities has stimulated the exploration of the extracellular
enzymatic activities of several microorganisms. Using validated values from a central composite designed experiment, amylase from a
locally isolated strain of Aspergillus niger was produced in a larger quantity by solid-state fermentation for characterization and its
subsequent application in the hydrolysis of raw native starches. The optimum activity of the enzyme was 30.96 U/ml-min. Partial
purification using ammonium sulphate saturation was attained at 50% (NH4)2SO4. The enzyme was stable over a wide range of pH (4 -
8) and temperature (30o
C – 60o
C). Other optimum values determined were; Ca2+
concentration – 5 mM, starch (substrate)
concentration – 3%, enzyme - substrate reaction time – 6 min and enzyme dosage – 4 mg total protein / 3% starch solution. The
enzyme was able to hydrolyse the raw starchy substrates studied, producing glucose concentrations of 31.90 mg/ml, 24.78 mg/ml,
19.31 mg/ml and 8.85 mg/ml from maize, sweet potato, yam, and cassava respectively. The optimal substrate concentration of these
substrates investigated also showed that different starchy substrates behaved differently towards hydrolysis by amylase. The study has
shown the possible production of a local enzyme that was capable of hydrolyzing raw native starches with good activity. This result
indicates the possibility of less dependence on imported enzymes.
Index Terms- Amylase, Aspergillus niger, Hydrolysis, Native starch, Solid-state fermentation
I. INTRODUCTION
the application of biotechnology to produce industrially competent enzymes has stimulated the quest for the extracellular
enzymatic activities of several microorganisms (Saranraj and Stella, 2013). Amylase is a very common enzyme found in most
starch processing industries due to its hydrolytic action on starches to produce simple sugars (Reddy et al., 2003). The potential
technological importance and economic profits of amylases have made it receive a great deal of attention in developing countries
(Suganthi et al., 2011). Amylase sourced from molds, bacteria, and yeasts have been reported and their biochemical characteristics
documented (Buzzini and Martini, 2002; Liaquat et al., 2015; Suganthi et al., 2011). Molds are identified to produce high amounts of
amylase (Suganthi et al., 2011) and amongst these, Aspergillus niger is seen as the most important source of industrial enzymes
(Sakthi et al., 2012). Commercially, fungal amylases have been reported to be more stable than bacterial amylases (Suganthi et al.,
2011); this has therefore called for several researchers to optimize the culture conditions of suitable fungal strains (Abu et al., 2005).
Solid - state fermentation (SSF) holds unlimited potential for the production of enzymes (Pandey et al., 1999), particularly when fungi
are involved (Bhargav et al., 2008). SSF present benefits such as; simplified operations, high volumetric production, and the
requirement for low initial capital (Omemu et al., 2005; Pandey et al., 1999).
Starchy tubers (such as yam, cassava, sweet potato, and cocoyam) and cereals occur abundantly in most developing countries in the
tropics, with the former being the most cultivated; since these remain the important staple foods in the diet of most people living in
these developing countries (Okolo et al., 1995). Annual losses of large proportions of these tubers to spoilage agents are encountered
due to inadequate storage facilities and improper post-harvest practices (Okolo et al., 1995; Omemu et al., 2005; Owusu - Darko et al.,
2016). These tubers, rich sources of starch, could be put to an alternative use; enzymatic conversion into reducing sugars by
saccharification (Omemu et al., 2005). Enzymatic hydrolysis has been shown to be economically superior in terms of process
simplicity and energy utilization as compared to the conventional method that uses pregelatinized starch as substrates (Okolo et al.,
1995; Omemu et al., 2005). Resistance to amylase enzymatic action has been observed in starches from tubers as compared to the
susceptibility in cereal starches (Omemu et al., 2005).
This paper reports on some properties of amylase produced by a locally isolated strain of Aspergillus niger as well as its hydrolytic
characteristics on some raw native starches.
T
2. International Journal of Scientific and Research Publications, Volume 8, Issue 7, July 2018 126
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II. MATERIALS AND METHODS
Preparation of starches from tubers
Cassava, sweet potato and yam starches were prepared according to the procedure outlined by (Rasper, 1969). Maize grains were
steeped in water for 3 days with intermittent changes of the water. It was milled using a double disc attrition milling machine and the
resulting dough was mixed with adequate water. The suspension was filtered through a 250 μm sized mesh gauze. The filtrate was
allowed to stand for 3h and the water was decanted leaving the settled starch slurry at the bottom. The starch was carefully dried in
circulating air temperatures below 45o
C. Corn-derived soluble starch (Sigma – Aldrich) was used as the standard. All other chemicals
were of analytical grade and obtained from Sigma – Aldrich St. Louis, MO, USA.
Source of fungal culture
Five (5) fungal colonies were isolated from a serial dilution of a mouldy bread sample and they were identified as Aspergillus niger by
employing the lactophenol cotton blue staining technique and studying morphology using standard identification manuals (Barnett and
Hunter, 1972). These fungal colonies were screened for their ability to synthesize amylase on potato dextrose agar (PDA) amended
with 2% (W/V) soluble starch medium (Uguru et al., 1997). The ability of individual colonies to produce amylase was indicated by
the zone of clearance shown on the medium. The isolate, Aspergillus niger KV5B showed the highest clearance zone (29 mm) and
thus was selected for further studies. Aspergillus niger KV5B was maintained on PDA slants and stored in the refrigerator at 4o
C.
Culture conditions and solid state fermentation
A spore suspension of Aspergillus niger KV5B was obtained from a 5-day old culture grown on PDA plates at room temperature by
adding 10 ml of sterile distilled water to the spores within a 1 cm cork borer and making the suspension to a 60 ml mark (Bentil et al.,
2015). Two (2 ml) of the spore suspension, containing about 3.96 x 106
cells/ml, was used as the inoculum. Yam peels, ground into
about 0.3 mm particle sizes, was used as the substrate for the amylase production. The enzyme was produced by SSF using the central
composite designed optimized conditions (pH - 5.95, Temperature - 49.53o
C and incubation time – 104 h) as previously described
(Kwatia et al., 2017). The fungi were grown in a 100 ml Erlenmeyer flask containing 5 g of the ground yam peel and moistened with 5
ml of a mineral solution (comprising KH2PO4 – 0.35 g, NH4NO3 – 2.5 g, KCl – 1.25 g, MgSO4.7H20 – 0.025 g, FeSO4.7H2O – 0.0025
g, soluble starch – 5 g in 250 ml of distilled water) (Sethi and Gupta, 2015).
Extraction of the enzyme from the fermentation medium
The enzyme was extracted using 50 ml of 0.1 M phosphate buffer (pH 6). The buffer on the substrate bed was shaken on an orbital
shaker for 30 min at 250 rpm, the resulting mixture was filtered through cheesecloth and the filtrate centrifuged at 3600 g for 15 mins
(Abu et al., 2005). The supernatant was decanted and used as the crude enzyme.
Amylase assay
The activity of amylase was assayed as described by (Uguru et al., 1997). The reaction mixture comprised of 0.5 ml of the crude
enzyme, 0.5 ml of 1% (W/V) starch solution in 0.02 M phosphate buffer with 0.006 M NaCl (pH 6.9). The reaction was incubated for
3 min at 37o
C and was terminated using 1 ml of 3, 5 dinitrosalicylic acid (Miller, 1959), followed by boiling for 5 min, cooled and
absorbance were taken at 540 nm. The amount of reducing sugars liberated (Miller, 1959) was estimated using glucose as standard. A
unit of amylase activity was expressed as the amount of enzyme that released 1 μmol of reducing sugars (maltose/glucose) per minute
under the assay conditions (Sakthi et al., 2012).
The protein content (Lowry et al., 1951) of the extract was also determined using bovine serum albumin as the standard at 540 nm.
Partial Purification of the crude amylase
Precipitation of the crude enzyme was carried out using ammonium sulphate ((NH4)2SO4). The extracted crude amylase was saturated
with ammonium sulphate up to 80%. Precipitates were obtained by gently stirring the mixture and leaving on ice for 30 min after
which the mixture was centrifuged at 3600 g for 15 min. The resultant precipitates were solubilized in 0.1M phosphate buffer (pH 6)
and their activities and protein concentrations determined.
Characterization studies
3. International Journal of Scientific and Research Publications, Volume 8, Issue 7, July 2018 127
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Effect of pH on enzyme activity and pH stability of the enzyme
The effect of pH on the enzyme activity was investigated by dissolving 1% (W/V) soluble starch in 0.1 M
sodium citrate buffer (pH 4 – 6), 0.1 M phosphate buffer ( pH 7 – 8) and glycine NaOH buffer (pH 9 – 10)
(Shah et al., 2014). A fraction of the enzyme was incubated at different pH values (4 – 10) without the substrate
in order to determine the pH stability of the enzyme. The DNS method was used to estimate the enzyme activity
using 0.5 ml of the crude enzyme.
Effect of temperature and temperature stability of the enzyme
An enzyme substrate reaction mixture was incubated at different temperatures of 30o
C – 90o
C for 3 min to determine the optimum
temperature of the enzyme. Thermostability of the enzyme was also determined by incubating the enzyme fraction without a substrate
at temperatures of 30o
C – 80o
C. The DNS method was used to estimate the enzyme activity using 0.5 ml of the crude enzyme.
Effect of calcium ion concentration on the stability of the enzyme
Amylase stability at 60o
C was investigated by incubating the enzyme-substrate reaction mixture with different concentrations of CaCl2
(2 mM, 5 mM, 7 mM and 10 mM). An untreated sample was used as the control.
Effect of substrate (starch) concentration on amylase activity
Starch concentrations of 1%, 2%, 3%, 4%, and 5% suspended in 0.1 M phosphate buffer (pH 6. 9) with 0.05 M NaCl were used to
investigate the effect of starch concentration on amylase activity. The DNS method was used for the amylase activity determination.
Enzyme substrate reaction time
The effect of reaction time on amylase activity was investigated by incubating the enzyme-substrate reaction mixture at different time
intervals of 0, 3, 6, 9, 12 and 15 min and the enzyme activity assayed using the DNS method.
Effect of enzyme concentration
The enzyme saturated at 50% of (NH4)2SO4 with a protein concentration of 1.90 mg/ml (approximated to 2 mg/ml). Thus, multiples of
2 mg/ml i.e. 2 ml (4 mg of protein), 3 ml (6 mg of protein), 4 ml (8 mg of protein) and 5 ml (10 mg of protein) were used to determine
the effect of enzyme concentration on amylase activity. The enzyme assay was by the DNS method.
Hydrolysis of starchy substrates
The ability of the enzyme to hydrolyze starch was investigated by using starches from cassava, sweet potato, yam, and maize.
Commercial corn – derived soluble starch (Sigma – Aldrich, St. Louis MO, USA) was used as the standard. The starch hydrolysis was
investigated using the optimized conditions that were obtained after the characterization. The rate of hydrolysis of each starch was
evaluated based on the quantity of reducing sugars (mg /ml) produced.
In order to extend the substrate range of the substrates industrially, the optimal substrate concentrations of the native starches were
also investigated with starch concentrations of 1 %, 2 %, 3 %, 4% and 5 %. The amylase activity was determined using the DNS
method after incubating the enzyme – substrate mixture for 30 min.
Statistical analysis and experimental design
All determinations were of 3 independent experiments. Mean values of each experiment were reported and the least significant
difference (LSD) test was used to identify means that differed significantly. Sigma plot (Ver. 10.1) software was used for the analysis
and graphs.
III. RESULTS AND DISCUSSION
Partial purification of amylase
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The stages of purifying amylase produced by Aspergillus niger KV5B using ammonium sulphate (NH4)2SO4 is shown in table 1.
Table 1 indicates that the specific activity of amylase increased from one purification step to another until the highest activity (18.04
U/mg) was reached at 50% (NH4)2SO4 saturation. The increase of specific activity from 0% to 50% (NH4)2SO4 saturation was an
indication that the purification steps used were effective in eliminating contaminant proteins. The ability of the (NH4)2SO4 to
concentrate enzymes makes it employable even in later stages of purification to concentrate enzymes from dilute solutions.
Table 1: Purification table for the crude extracted amylase using ammonium sulphate
Purification step Total activity
(U/ml-min)
Total protein
(mg/ml)
Specific activity
(U/mg)
Purification
fold (X)
Crude extract 16.06±2.02 8.71±2.00 1.93±0.66 1.00
20% (NH4)2SO4 30.83±0.49 6.99±1.29 4.52±0.87 2.34
50% (NH4)2SO4 35.16±1.44 1.96±2.20 18.04±1.46 9.33
80% (NH4)2SO4 39.05±2.22 4.89±1.60 8.67±3.15 4.48
Effect of pH and pH stability of the enzyme
Figure 1 shows the effect of pH as well as pH stability on the activity of the enzyme. The highest activity (31.88 U/ml-min) was
observed at pH 5. A pH of 28 also recorded a high activity of 29.91 U/ml-min. This pH range shows the versatility of the enzyme for
several process conditions. These optima observed indicated the enzyme was at its satisfactory conformation (Sachdev et al., 2016).
Kumar and Duhan (2011) and Alva et al. (2007) reported similar pH ranges of (4.2 and 8.4) and (5.8 and 9) for Aspergillus niger
MTCC – 104 and Aspergillus sp JG112 amylases respectively. They attributed such observations to the existence of at least two
amylolytic activities, i.e. glucoamylase and α-amylase since both enzymes synergistically degrade starch molecules.
The enzyme retained more than 80% of its activity across a pH range of 4 – 8. This implied the enzyme was stable over the pH ranges
4 – 8. The drop in activity at pH 9 and 10 (Figure 1) could be attributed to severe denaturation of the structure of the enzyme due to
the changes in pH thus leading to the reduction of activity at such pH levels (Kumar and Duhan, 2011).
pH
3 4 5 6 7 8 9 10
Amylaseactivity(U/ml-min)
10
15
20
25
30
35
pH
pH stability
Figure 1: Effect of pH and pH stability of A. niger KV5B enzyme activity
Effect of temperature and temperature activity of the enzyme
5. International Journal of Scientific and Research Publications, Volume 8, Issue 7, July 2018 129
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The effect of temperature on the activity of amylase showed that the enzyme activity increased progressively with an increase in
temperature from 30 o
C (16.73 U/ml-min) to a maximum of 28.41 U/ml-min at 60 o
C (Figure 2). Reduction in amylase activities was
observed at temperatures below 60o
C. The decrease in amylase activity may be attributed to the disruption of the secondary, tertiary
and quaternary bonds of the amylase enzyme (Sachdev et al., 2016; Schokker et al., 1998). An optimum temperature of 70 o
C was
recorded for an amylase produced by Aspergillus niger on yam peels (Uguru et al., 1997). The enzyme showed thermal stability
across a temperature range of 30 o
C – 60 o
C, beyond which the activity decreased (Figure 2), this may probably be the result of amino
acids destruction, peptide chain hydrolysis and aggregation thus causing incorrect conformation of the enzyme (Alva et al., 2007).
Temperature (o
C)
20 30 40 50 60 70 80
Amylaseactivity(U/ml-min)
5
10
15
20
25
30
35 Temperature
Temperature stability
Figure 2: Effect of temperature and temperature stability of A. niger KV5B amylase activity
Effect of calcium concentration on the stability of the amylase
The influence of Ca2+
varied with concentrations of calcium that the enzyme was exposed to at 60o
C (Figure 3). A concentration of 5
mM of calcium ions recorded the highest activity (39.51 U/ml-min) and the least was found with 2 mM calcium concentration.
Amylase is a metalloenzyme and is stabilized in the presence of calcium ions (Konsoula and Liakopoulou-Kyriakides, 2007; Prajapati
et al., 2015). The results of this study indicated that amylase from Aspergillus niger KV5B was stable in the presence of Ca2+
.
The stability of an enzyme from Aspergillus sp cmst 04 at 5 mM concentration has been reported (Ajikumar et al., 2014). An
inhibitory action of Ca2+
at 10 mM was also reported by (Babu and Satyanarayana, 1993). Other reports also suggest that Ca2+
does
not affect the enzyme activity (Asgher et al., 2007).
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CaCl2 concentration (mM)
0 2 5 7 10
Amylaseactivity(U/ml-min)
0
10
20
30
40
50
Figure 3: Effect of calcium concentration on A. niger KV5B amylase activity
Effect of substrate (starch) concentration on amylase activity
Amylase activity increased with increase in starch concentrations from 1 % (21.88 U/ml-min) to 3% (37.92 U/ml-min), but the activity
decreased beyond 3% (Figure 4). The active sites of the enzyme were not saturated at low substrate concentrations; therefore, increase
in activity was expected when the substrate concentration increases (Kumar and Duhan, 2011). Low activities recorded beyond 3%
might have been a result of blockage of the active site of the enzyme due to competition for these sites by the substrates, thus
preventing other molecules from binding (Nyamful, 2013).
Starch concentration ( %)
1 2 3 4 5
Amylaseactivity(U/ml-min)
0
10
20
30
40
50
Figure 4: Effect of starch concentration on A. niger KV5B amylase activity
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Effect of enzyme-substrate reaction time
The time allowed for starch hydrolysis increased the enzyme activity from 3 min (25.35 U/ml-min) to a maximum of 6 min (32.41
U/ml-min) (Figure 5). Incubation beyond 6 min, however, decreased the activity (Figure 5). The enzyme showed efficiency within a
relatively shorter time, thus saving time and energy and also a possible prevention of the formation of undesirable products (Dzogbefia
et al., 2008).
Time (min)
0 2 4 6 8 10 12 14 16
Amylaseactivity(U/ml-min)
0
10
20
30
40
Figure 5: Effect of reaction time on A. niger KV5B amylase activity
Enzyme dosage
Figure 6 depicts the effect of enzyme dosage on the hydrolysis of starch. Two (2) ml of the enzyme extract (i.e. 4 mg of total protein)
per 3% starch solution gave the highest activity of 58.10 U/ml-min and the least activity was observed with 1 ml of the enzyme extract
(i.e. 2 mg of total protein). A probable inhibitory action of the enzyme beyond 4 mg total protein / 3% starch solution as a result of
increased enzyme dosage thus introducing competition for binding could have resulted in the decrease in amylase activity. The
enzyme application could be cost-effective since lower concentrations will be required if it should be used by industries for starch
hydrolysis.
Enzyme dosage (mg total protein / 3% starch solution
0 2 4 6 8 10
Amylaseactivity(U/ml-min)
0
10
20
30
40
50
60
70
Figure 6: Effect of enzyme dosage on A. niger KV5B amylase activity
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Hydrolysis of native starchy substrates
The ability of the amylase to hydrolyse starches from tubers (cassava, yam, and sweet potato) and cereal (maize) was studied using
corn-derived soluble starch as a standard. The results that portrayed the amount of reducing sugars (mg/ml) produced are shown in
Figure 7. The corn-derived soluble starch was also used as a standard (100%) to evaluate the susceptibility of each starchy source to
amylase hydrolysis. By keeping all the previously determined parameters at their optimized levels, rapid hydrolysis was observed on
the corn-derived soluble starch and maize to give reducing sugar concentrations of 30.68 mg/ml and 31.90 mg/ml respectively. In
general, the order of starch hydrolysis was maize > soluble starch > sweet potato > yam > cassava (Figure 7). Statistically, reducing
sugars produced by maize, sweet potato and yam were significantly higher (p<0.05) than that produced by cassava starch within the
six minutes of hydrolysis. However, no statistical difference (p>0.05) was observed between hydrolysis of sweet potato and yam
starches. The conversion efficiency of the substrates as shown in Table 2 indicated that starch from maize was rapidly hydrolyzed
(103. 98%) followed by sweet potato, yam, and cassava with conversion efficiencies of 80.76%, 62.56%, and 28.83% respectively.
The optimal substrate concentration of each substrate after 30 min incubation is summarized in figure 8. Maize starch recorded the
fastest optimal level and thus highest hydrolysis and activity at 2% concentration (65.29 U/ml-min) followed by soluble starch (60.55
U/ml-min), cassava (56.54 U/ml-min) and sweet potato (48.45 U/ml-min) starches, all at 3% concentration. Yam recorded the least
hydrolysis and thus the least activity (34.24 U/ml-min) at 4% concentration (Figure 8). Results in figure 8 also showed that the ability
of amylase of Aspergillus niger KV5B to digest raw starches is directly proportional to incubation time
Table 2: Conversion efficiencies of the native starchy substrates at 6 min of incubation
Starches Amount of reducing sugars (mg/ml) Relative conversion efficiency (%)
Soluble starch 30.69 ± 0.45 100.00
Maize 31.91 ± 0.48 103.98
Yam 19.31 ± 0.43 62.56
Sweet potato 24.78 ± 0.11 80.76
Cassava 8.85 ± 0.18 28.83
Source of starch
Soluble starch Cassava Maize Sweet potato Yam
Amylaseactivity(U/ml-min)
0
10
20
30
40
Figure 7: Hydrolysis of raw native starches by A. niger KV5B amylase
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Starch concentration (%)
1 2 3 4 5
Amylaseactivity(U/ml-min)
0
20
40
60
80 Soluble starch
Maize
Sweet potato
Yam
Cassaava
Figure 8: Optimum substrate concentration of the raw native starches upon hydrolysis by amylase of A. niger KV5B
Many factors have been attributed to the differences in enzymatic susceptibilities of starches; viz, starch source, extension of
association between starch components, granule size, crystallinity, rate of amylose and amylopectin, polymeric type (A,B,C),
amylose-lipid complex, type of enzyme and hydrolysis condition ( pH, temperature, concentration)(Oates, 1997; Rocha et al., 2010).
Maize starch granules have been identified to exhibit A –type crystal polymorphism with characteristics such as having a porous
surface, faster chemical penetration and derivatization reactions, weak points and more susceptible to the enzyme-catalyzed reactions
(Jane, 2009). Starches with such A polymorphism are more susceptible to hydrolysis by enzymes because the crystalline structure of
such starches contains the A and B1 chains which are unstable and more vulnerable to rearrangement; therefore making them
susceptible to hydrolysis (Rocha et al., 2010).
Cassava and sweet potato starches also exhibited more susceptibility to the enzyme hydrolysis as compared to yam starches (Figure 8).
The high susceptibility of cassava and sweet potato to the enzymatic hydrolysis may also be attributed to its A – type polymorphism as
reported by (Rocha et al., 2010). Yam starches, however, displays the B – type polymorphism which normally displays a nonporous
internal granule structure thus, making them less susceptible to amylase hydrolysis (Jane, 2009). B – type starches present higher
proportions of long B – chains, which extend for two or more clusters and stabilize the internal structure of granules thus becoming
more resistant to enzymatic action (Jane, 2009; Rocha et al., 2010).
Amylose content of maize, sweet potato, cassava and yam starches have been stated as 70 %w/w (Lim et al., 1994; Morrison et al.,
1984), 28.9 ±0.35 %w/w (Tan et al., 2006), 23.7±0.1 %w/w 8 -25 %w/w (Paes et al., 2008; Yuan et al., 2007) and 26.3±0.2 %w/w
(Yuan et al., 2007) respectively. A report by Tester et al. (2006) suggested that the amount of native starch hydrolysis by amylases is
inversely proportional to the amylose content, indicating that high amylose starches are more resistant to amylase hydrolysis. This is
supported by (Jane, 2009) who also reported that such amyloses are strongly associated with the periphery of the granules thus making
them less susceptible to enzyme attack. These might be some contributing factors to the results recorded in this investigation.
Results from the study agree with reports made by Omemu et al., (2005) and Okolo et al., (1995) that starch susceptibility to amylase
hydrolysis is dependent on the botanical source and the duration of the amylase treatment. The ability of the partially purified amylase
of Aspergillus niger KV5B to hydrolyse roots starches especially cassava starch presents an incredible property since these roots are
the most abundant in the tropics.
In conclusion, the amylase of Aspergillus niger KV5B selected for this study could be effectively used in hydrolysis of both native
tuber and cereal starches to produce sugars which will find applications in several industries such as the brewing industry.
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First Author – Samuel Kwatia, MSc, Department of Biochemistry and Biotechnology, KNUST, Ghana
samuelkwatias@gmail.com
Second Author - Prof. (Mrs) Victoria P. Dzogbefia, Ph.D., Department of Biochemistry and Biotechnology, KNUST,
victoriadzogbefia@gmail.com
Correspondence Author – Samuel Kwatia, samuelkwatias@gmail.com, skwatia@gsa.gov.gh, +233240140026, +233261431206.