Abstract The present study aimed to investigate the effect of different culture conditions on biomass content and flavonoid production of the elite C. asiatica accession UPM03. When 0.1 g inoculum 25 mL-1 of cell was grown in Murashige & Skoog (MS) medium supplemented with 2 mg 2,4-D l-1 and 1 mg kinetin l-1, flavonoids were not significantly produced in cells or released into the culture medium. Production of flavonoid was correlated with the aggregation size and inoculum density. At aggregate size of 250-500 μm with 0.3 g inoculum density 25 mL-1, it gave the highest luteolin content with 35.45 μg g-1 DW. After investigating the effect of culture conditions, i.e. pH, inoculum density, light irradiation and plant growth regulator, we found that with the optimized condition (i.e. 250-500 μm aggregate size, 0.5 g wet cell 25 mL-1 supplemented with 3 mg L-1 2,4-D and 1 mg L-1 kinetin at pH 5.7 under 16h photoperiod) the luteolin content was 11-fold higher than the cell suspension cultured at an inoculum size of 0.3 g wet cell 25 mL-1 with 250-500 μm aggregate size. Index Terms: Centella asiatica, flavonoid, light irradiation, pH regime, plant growth regulators .
In Vitro Cell Tests for Functional FoodInstitut Kurz
The relationship between the food we eat and our health is
clear. In the constant search for healthier foods rich in
bioactive compounds that promote health and healthy
ageing, a wide variety of functional foods have appeared on
the market.
To know the real function of these functional foods
in our body, it is necessary to carry out different types of in
vitro cell tests.
Institut Kurz specializes in conducting in vitro cell tests for functional foods.
Contact us for more information:
info@institut-kurz.com
https://www.institut-kurz.com/
International Journal of Pharmaceutical Science Invention (IJPSI)inventionjournals
is an international journal intended for professionals and researchers in all fields of Pahrmaceutical Science. IJPSI publishes research articles and reviews within the whole field Pharmacy and Pharmaceutical Science, new teaching methods, assessment, validation and the impact of new technologies and it will continue to provide information on the latest trends and developments in this ever-expanding subject. The publications of papers are selected through double peer reviewed to ensure originality, relevance, and readability. The articles published in our journal can be accessed online
International Journal of Engineering Research and Applications (IJERA) is an open access online peer reviewed international journal that publishes research and review articles in the fields of Computer Science, Neural Networks, Electrical Engineering, Software Engineering, Information Technology, Mechanical Engineering, Chemical Engineering, Plastic Engineering, Food Technology, Textile Engineering, Nano Technology & science, Power Electronics, Electronics & Communication Engineering, Computational mathematics, Image processing, Civil Engineering, Structural Engineering, Environmental Engineering, VLSI Testing & Low Power VLSI Design etc.
This study aimed to determine optimal yeast growth conditions for undergraduate student schedules. Yeast were cultured under varying conditions of inoculation mass, media composition, and flask type. An inoculation of 0.01g yeast grown in rich media at 25°C and 100rpm reached an optimal OD600 of 0.5-0.7 within 14-24 hours, suitable for protoplasting. Preliminary results showed yeast with OD600 over 0.7 resisted protoplasting. The initial inoculation mass was critical to prevent overgrowth. Overall, an inoculation of 0.01g yeast in rich media at 25°C and 100rpm provided optimal growth conditions for student timelines and downstream protoplasting.
Solid state fermentation of Lathyrus sativus and sugarcane bagasse by Pleurot...Innspub Net
This research paper investigates the solid state fermentation of Lathyrus sativus (khesari plant) and sugarcane bagasse by Pleurotus sajor-caju. The researchers pretreated the substrates with lime and alkali to increase their digestibility. They found that lime treatment and presoaking increased the accumulation of reducing sugars and soluble proteins during the 8-week fermentation. The highest levels of reducing sugars (22.15 mg/g) and soluble proteins (22.80 mg/g) were observed in the lime treated and presoaked mixture of khesari plant and sugarcane bagasse. Pretreatment also enhanced the cellulolytic enzyme activity of the fungal
DEVELOPING PROTOCOLS FOR EFFICIENT GENETIC TRANSFORMATION IN EURYCOMA LONGIFOLIAVijendren Krishnan
This research proposal aims to develop genetic transformation protocols for Eurycoma longifolia using particle bombardment. The objectives are to transform E. longifolia with therapeutic protein genes, analyze transformants for gene insertion and expression, and facilitate production of therapeutic proteins in medicinal plants. The methodology involves optimizing callus induction, regeneration, and genetic transformation parameters. Preliminary results show E. longifolia and L. pumila have the best callus growth. Upon completing the optimization and verification steps, transgenic E. longifolia expressing the therapeutic protein gene will be produced.
52.Screeing of industrial production of CellulaseAnnadurai B
1) The study screened 73 microorganism species for their ability to produce cellulase enzymes. Alternaria, Bacillus, Aspergillus, Trichoderma, Fusarium and Penicillium showed good cellulase production.
2) Trichoderma viride and Aspergillus terreus produced the highest amounts of cellulase. Maximum enzyme production of 1.76 U/ml was achieved by A. terreus at 30°C, pH 5.0 over 6 days of incubation.
3) Supplementing the culture medium with plant waste materials like banana leaves, rice husk, and wheat bran further improved cellulase production, with 6% supplementation
Optimization of Bacillus Subtilis Natto Immobilization Process on Alginate – ...inventionjournals
Nattokinase is a potent fibrinolytic enzyme with the potential for fighting cardiovascular diseases. In this study, Bacillus subtilis natto were immobilized in the alginate – chitosan complex for fermentation of nattokinase enzyme. Six factors affecting the efficiency of immobilization cells were screened by Plackett – Burman design including: concentration of alginate, concentration of chitosan, pH of chitosan, concentration of CaCl2, added cells density, shaking time after supplementing chitosan. Results of optimization have identified two factors affecting the efficiency of cell immobilization. They are concentration of alginate (2.5%) and added cells density (approximately 5.86 million colonies per milliliter). With these two factors optimized and others kept at the normal level, immobilization efficiency reached 90.73%. After Bacillus subtilis natto had been immobilized by optimization of parameters, we conducted application for fermenting nattokinase. For 24 hours of fermentation, nattokinase enzyme activity reached 71.80 ± 0.19 FU/ml. Immobilized Bacillus subtilis natto cells were reused 6 times and on the 6 th time of reuse, nattokinase enzyme activity only decreased 2.7% in compared with the 1st reuse.
In Vitro Cell Tests for Functional FoodInstitut Kurz
The relationship between the food we eat and our health is
clear. In the constant search for healthier foods rich in
bioactive compounds that promote health and healthy
ageing, a wide variety of functional foods have appeared on
the market.
To know the real function of these functional foods
in our body, it is necessary to carry out different types of in
vitro cell tests.
Institut Kurz specializes in conducting in vitro cell tests for functional foods.
Contact us for more information:
info@institut-kurz.com
https://www.institut-kurz.com/
International Journal of Pharmaceutical Science Invention (IJPSI)inventionjournals
is an international journal intended for professionals and researchers in all fields of Pahrmaceutical Science. IJPSI publishes research articles and reviews within the whole field Pharmacy and Pharmaceutical Science, new teaching methods, assessment, validation and the impact of new technologies and it will continue to provide information on the latest trends and developments in this ever-expanding subject. The publications of papers are selected through double peer reviewed to ensure originality, relevance, and readability. The articles published in our journal can be accessed online
International Journal of Engineering Research and Applications (IJERA) is an open access online peer reviewed international journal that publishes research and review articles in the fields of Computer Science, Neural Networks, Electrical Engineering, Software Engineering, Information Technology, Mechanical Engineering, Chemical Engineering, Plastic Engineering, Food Technology, Textile Engineering, Nano Technology & science, Power Electronics, Electronics & Communication Engineering, Computational mathematics, Image processing, Civil Engineering, Structural Engineering, Environmental Engineering, VLSI Testing & Low Power VLSI Design etc.
This study aimed to determine optimal yeast growth conditions for undergraduate student schedules. Yeast were cultured under varying conditions of inoculation mass, media composition, and flask type. An inoculation of 0.01g yeast grown in rich media at 25°C and 100rpm reached an optimal OD600 of 0.5-0.7 within 14-24 hours, suitable for protoplasting. Preliminary results showed yeast with OD600 over 0.7 resisted protoplasting. The initial inoculation mass was critical to prevent overgrowth. Overall, an inoculation of 0.01g yeast in rich media at 25°C and 100rpm provided optimal growth conditions for student timelines and downstream protoplasting.
Solid state fermentation of Lathyrus sativus and sugarcane bagasse by Pleurot...Innspub Net
This research paper investigates the solid state fermentation of Lathyrus sativus (khesari plant) and sugarcane bagasse by Pleurotus sajor-caju. The researchers pretreated the substrates with lime and alkali to increase their digestibility. They found that lime treatment and presoaking increased the accumulation of reducing sugars and soluble proteins during the 8-week fermentation. The highest levels of reducing sugars (22.15 mg/g) and soluble proteins (22.80 mg/g) were observed in the lime treated and presoaked mixture of khesari plant and sugarcane bagasse. Pretreatment also enhanced the cellulolytic enzyme activity of the fungal
DEVELOPING PROTOCOLS FOR EFFICIENT GENETIC TRANSFORMATION IN EURYCOMA LONGIFOLIAVijendren Krishnan
This research proposal aims to develop genetic transformation protocols for Eurycoma longifolia using particle bombardment. The objectives are to transform E. longifolia with therapeutic protein genes, analyze transformants for gene insertion and expression, and facilitate production of therapeutic proteins in medicinal plants. The methodology involves optimizing callus induction, regeneration, and genetic transformation parameters. Preliminary results show E. longifolia and L. pumila have the best callus growth. Upon completing the optimization and verification steps, transgenic E. longifolia expressing the therapeutic protein gene will be produced.
52.Screeing of industrial production of CellulaseAnnadurai B
1) The study screened 73 microorganism species for their ability to produce cellulase enzymes. Alternaria, Bacillus, Aspergillus, Trichoderma, Fusarium and Penicillium showed good cellulase production.
2) Trichoderma viride and Aspergillus terreus produced the highest amounts of cellulase. Maximum enzyme production of 1.76 U/ml was achieved by A. terreus at 30°C, pH 5.0 over 6 days of incubation.
3) Supplementing the culture medium with plant waste materials like banana leaves, rice husk, and wheat bran further improved cellulase production, with 6% supplementation
Optimization of Bacillus Subtilis Natto Immobilization Process on Alginate – ...inventionjournals
Nattokinase is a potent fibrinolytic enzyme with the potential for fighting cardiovascular diseases. In this study, Bacillus subtilis natto were immobilized in the alginate – chitosan complex for fermentation of nattokinase enzyme. Six factors affecting the efficiency of immobilization cells were screened by Plackett – Burman design including: concentration of alginate, concentration of chitosan, pH of chitosan, concentration of CaCl2, added cells density, shaking time after supplementing chitosan. Results of optimization have identified two factors affecting the efficiency of cell immobilization. They are concentration of alginate (2.5%) and added cells density (approximately 5.86 million colonies per milliliter). With these two factors optimized and others kept at the normal level, immobilization efficiency reached 90.73%. After Bacillus subtilis natto had been immobilized by optimization of parameters, we conducted application for fermenting nattokinase. For 24 hours of fermentation, nattokinase enzyme activity reached 71.80 ± 0.19 FU/ml. Immobilized Bacillus subtilis natto cells were reused 6 times and on the 6 th time of reuse, nattokinase enzyme activity only decreased 2.7% in compared with the 1st reuse.
Isolation, Screening and Selection of Fungal Strains for Potential Cellulase ...inventionjournals
The present study was aimed to isolate, screen and identify the potential cellulase and xylanase producing fungi from the soil samples collected from different areas of Haryana. Total one hundred fifty one fungal isolates were isolated from these soil samples were then screened by using selective media (i.e. CMC and Xylan agar) in order to determine the potency of microbes in producing cellulase and xylanase which were indicated by clear zones formation around the cultures. This qualitative screening which showing greater cellulase and xylanase indexes were subjected to enzyme activity tests by Dinitrosalicylic acid (DNS) method. Maximum enzyme production was achieved at 30°C, pH of 6.0 by Trichoderma atroviride on 5th day of incubation.
Extraction and purification of product from fermentation is known as Downstream Processing ( DSP) or Product Recovery
It is an essential step in the manufacture of pharmaceuticals product
Cost of the product is determined by the DSP involved
The results of the study show a promising role of Acacia Honey, a natural product with proven therapeutic effects on skin wound healing. It accelerated the initial stage of corneal wound healing without the side effects found when using conventional treatments which contain preservatives. Corneal keratocytes cultured in media supplemented with 0.025% Acacia Honey showed an increase in proliferative capacity while retaining their morphology, gene and protein expressions with normal cell cycle.
The current study investigated the immunomodulatory
potential of ethyl acetate soluble supernatant of
Lactobacillus casei (LC-EAS) in vitro. The effect of
LC-EAS on nitric oxide release was analyzed in RAW
264.7 cells, wherein, an inhibition in nitric oxide production
through suppression of inducible nitric oxide synthase
mRNA expression was observed. Evaluation of LC-EAS
on LPS-induced peripheral blood mononuclear cells
showed a down-regulation in TNF-a and IL-6 genes and an
upregulation of IL-10. An inhibition in the protein
expression of NF-kB, ERK1/2 and STAT3 phosphorylation
confirms the immunomodulatory potential of LC-EAS. The
effect of LC-EAS on in vitro intestinal epithelial cells was
investigated using HT-29 human colon adenocarcinoma
cancer cells. LC-EAS exhibited an inhibition of NF-jB and
ERK1/2 phosphorylation, whereas STAT3 phosphorylation
was unregulated. To evaluate the downstream target of
STAT3 upregulation, expression of the intestinal trefoil
factor TFF3 which is a NF-jB regulator and STAT3
downstream target was studied. LC-EAS was observed to
elevate TFF3 mRNA expression. Overall the study shows
that the anti-inflammatory potential of LC-EAS is through
inhibition of NF-kB in different cell types.
ABSTRACT- Background: Declining trend in semen quality is receiving attention worldwide. The aim of the study to investigate the role of lifestyle factors with respect to semen quality.
Methods: Semen samples were collected from 351 male partners attending OPD of Obstetrics and Gynecology at civil hospital and IKD hospital, Ahmedabad, India. They were subjected to assess the quality of semen according to WHO criteria and semen quality were analyzed with respect to self reported history of tobacco smoking and/or chewing and alcohol consumption as lifestyle factors.
Results: The result revealed that sperm count was lower in subjects with habit of tobacco smoking, chewing and alcohol consumption as compared to subjects without such habits and decline was found statistically significant among smokers. Total progressive motility and normal morphology percentage was significantly decreased in tobacco chewers compared to non-chewers. Further analysis of data with respect to alcohol consumption indicated non-significantly lower total progressive motility and normal sperm morphology percentage as compared to subjects with no such habit.
Conclusion: The data obtained suggested, the role of lifestyle factors especially tobacco smoking and chewing in declining semen quality.
Key-words- Semen quality, Tobacco, Alcohol, Lifestyle, Sperm motility, Smoking
1) A Penicillium sp. fungus produced high levels of the softwood-specific enzyme beta-mannanase when grown in solid and liquid cultures containing locust bean gum.
2) Adding a crude enzyme extract from this Penicillium sp. to a mixture of commercial enzymes improved saccharification of steam-exploded radiata pine by 7.8%.
3) Producing crude enzyme extracts on-site from native fungi is a potential strategy to lower the cost of enzymes for breaking down softwood biomass.
This study developed a novel biodegradable porous scaffold for skin regeneration composed of collagen, hyaluronic acid, and gelatin. The scaffold was tested in vitro by culturing human keratinocytes, melanocytes, and fibroblasts, which all proliferated normally. In vivo tests on rats found the scaffold treated wounds healed faster than untreated wounds, with over 50% closure within 7 days versus only 60% for untreated wounds. Histological analysis further supported the scaffold's ability to facilitate more rapid wound healing and regeneration of skin tissue.
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.
This study evaluated the antioxidant and antimicrobial properties of Syzygium jambos. Methanolic extracts and aqueous soluble fractions of S. jambos stem bark exhibited significant antioxidant activity. The chloroform and carbon tetrachloride soluble fractions showed mild antimicrobial activity against several bacteria. The chloroform soluble fraction also demonstrated moderate free radical scavenging activity. Overall, S. jambos extracts displayed antioxidant and limited antimicrobial effects, suggesting potential for further investigation into their bioactive properties.
Study on Culture Conditions for A Cellulase Production From As Pergillus UnguisIJMERJOURNAL
1) The document studies the optimal culture conditions for cellulase production by Aspergillus unguis through solid state fermentation.
2) Experiments were conducted to determine the effect of various carbon sources, nitrogen sources, moisture content, duration and citrate buffer content on cellulase production.
3) The highest cellulase activity of 110.92 U/ml was achieved under the following optimal conditions: rice husk as the carbon source, yeast extract as the nitrogen source, 64% moisture content, 7 days of incubation, and a citrate buffer to byproduct ratio of 7:1.
- Digested black tea induces the secretion of pro-inflammatory cytokines TNF-α and IL-6 in macrophages after 8 hours of exposure, but does not induce secretion of the anti-inflammatory cytokine IL-10. Simulated digestive fluid alone also induces some inflammatory response, but to a lesser extent than digested black tea.
- The study investigated the effects of digested black tea on cytokine secretion in macrophages to better understand how consumed black tea impacts the immune system, as previous studies have not considered the effects of digestion.
This document summarizes a study that transformed the plant Artemisia dubia with the rolA gene using Agrobacterium tumefaciens to increase production of artemisinin and its derivatives. The rolA gene was inserted into A. dubia and transgenic plants were regenerated. Thin layer chromatography analysis found that both rolA and rol ABC transgenic A. dubia produced comparable amounts of artemisinin and derivatives, demonstrating that rolA transformation is effective for enhancing secondary metabolite production. Optimization of the transformation protocol improved regeneration of transgenic A. dubia plants.
Ảnh hưởng của màu sắc đèn led trong kĩ thuật nấm đông trùng hạ thảoHo Chi Minh
Ảnh hưởng của màu sắc đèn led trong kĩ thuật nấm đông trùng hạ thảo
GIA BAO CO.
www.dongtrunghathaogiabao.com
Add: 119A Tran Binh Trong, District 5, Ho Chi Minh City, Viet Nam
Hotline: 0776 950 960
DOI: 10.21276/ijlssr.2016.2.4.7
ABSTRACT- Most of the Euphorbian plants secrete fluid which contain a proteolytic enzyme for defensive role against
insects, pests and hence eco physiological inheritance to sustain vegetation eventually in adverse environmental
conditions. Evaluation has been carried out on twenty five Euphorbian garden plants for their proteolytic activities using
casein as a substrate. Out of these, Euphorbia nerifolia, Euphorbia milli, Euphorbia tirucalli, Euphorbia lactea,
Synadenium granti, Jatropha curcas, Euphorbia nivulia, Euphorbia antiquoram, Pedilanthus tithymaloides, Euphorbia
viguieri, E. heterophylla and E. leucocephala are the good enzyme source. Moderate activity found in Jatropha
integerrima, Jatropha multifida, Jatropha podagrica, Euphorbia pulcherrima, and Dalechampia scandens. While
different tissues of Acalypha hispida, Acalypha wilkesiana, Breynia nivosa, Cicca acida, Codiaeum variegatum, Drypetes
roxburghii are devoid of proteolytic activity. This paper describes in detail about name of plants, habitat and presence of
proteolytic enzyme in them. Results show that the out of twenty five plants 50% plant tissue synthesize protease in
appreciable amount, while 10% are not able to produce it. However 40% plants demonstrate only detectable amount of
protease. A comparative account of proteolytic activity reveals some promising plants good source of enzyme. Some
plants produce combination of cysteine and serine proteases. A single plant i.e. Euphorbia nerifolia latex contains
cysteine, serine, metallo-protease and aspartic proteases. In turn, these proteases may be used in various industrial uses in
general and cheese production Key-words- Garden Euphorb iiann a p plaarnttisc,u Clayr.s t e ine and serine protease, E. leucocephala, Euphorbia viguieri
Effect of Different Concentrations of Glycerol on survival of Azotobacter chr...IIJSRJournal
This document discusses a study that evaluated the effect of different concentrations of glycerol on the survival of the nitrogen-fixing bacterium Azotobacter chroococcum in liquid formulations. Azotobacter was inoculated into Jensen's broth containing various concentrations of glycerol from 5 mM to 25 mM. The number of surviving Azotobacter colonies was counted over 360 days. Results showed that survival was highest in the 25 mM glycerol formulation, with 22 colonies surviving after 360 days. Survival was lowest in the 5 mM glycerol formulation, with only 5 colonies surviving after 360 days. Additionally, three Azotobacter strains showed different survival rates in the 25 mM glycerol formulation over 360 days, demonstrating an
IRJET- An Overview on Fungi as Self Healing Agent in Biomineralization of...IRJET Journal
This document discusses the use of fungi as self-healing agents in biomineralization of calcite in cement concrete. While bacteria have been used for their ability to precipitate calcite via urea hydrolysis, fungi are also efficient in this process. The document reviews studies demonstrating fungal calcite precipitation and describes the methodology for culturing fungi, producing bioconcrete incorporating fungi, and testing the physicochemical and bioengineering properties of the resulting bioconcretes. Key properties examined include compressive strength, crack healing ability, and calcite precipitation as observed using SEM and EDX analysis.
The Studies of Effect of Bio Fertilizers Rhizobium, Phosphobacteria, and Root...IIJSRJournal
A pot culture experiment was carried out and to the find out the effect of Rhizobium, Phosphobacteria and Root Nodule extract on the vegetative growth and biochemical changes in Black gram and Maize. The Black gram and Maize is vegetative parameters such as seed germination, shoot and root length, leaf fresh weight, leafs dry weight, shoot and root dry weight had analyzed and biochemical studies of both Black gram and Maize was carried such as total chlorophyll content, leave soluble protein and in vivo nitrate reductase activity. Among microbial inoculants, the Rhizobium + PB mixer was found most effective in terms of seed germination, shoot and root length, leaf fresh weight, leafs dry weight, shoot and root dry weight and also showed increase total chlorophyll content, soluble protein and nitrate activities in both Black gram and Maize.
Deals with various methods adapted for the Improvement in Microbial Cell Culture alongwith the procedure and instruments used to carry out the operation through illustrative diagrams
SunModo is a solar racking company founded in 2008 that provides a complete line of roof and ground mounting solutions. Their mission is to accelerate adoption of affordable clean energy technologies by providing high quality, low cost, and easily installable racking systems. They offer innovative racking products for roof, flat roof, pitched roof, ground mount, and self-grounding applications. Their focus is on professional installers through responsive support, engineering services, and quick to install systems.
Este documento presenta las normas de comportamiento para usar Internet de manera respetuosa. Súper compu le pide a una persona que use un lenguaje más respetuoso al comunicarse en Internet. Luego, Súper NETI enumera principios como el respeto, la libertad y la intimidad que deben guiar las interacciones en línea. Finalmente, la persona se disculpa por su comportamiento anterior.
O documento analisa as tendências de compra de eletroeletrônicos e eletrodomésticos para o Natal de 2010 no Brasil, com base em dados de mídias sociais e notícias online. Televisões, celulares e computadores aparecem como os produtos eletrônicos mais desejados no Twitter e blogs, enquanto eletrodomésticos recebem menos menções. As vendas online devem crescer 41% em relação ao ano anterior.
Isolation, Screening and Selection of Fungal Strains for Potential Cellulase ...inventionjournals
The present study was aimed to isolate, screen and identify the potential cellulase and xylanase producing fungi from the soil samples collected from different areas of Haryana. Total one hundred fifty one fungal isolates were isolated from these soil samples were then screened by using selective media (i.e. CMC and Xylan agar) in order to determine the potency of microbes in producing cellulase and xylanase which were indicated by clear zones formation around the cultures. This qualitative screening which showing greater cellulase and xylanase indexes were subjected to enzyme activity tests by Dinitrosalicylic acid (DNS) method. Maximum enzyme production was achieved at 30°C, pH of 6.0 by Trichoderma atroviride on 5th day of incubation.
Extraction and purification of product from fermentation is known as Downstream Processing ( DSP) or Product Recovery
It is an essential step in the manufacture of pharmaceuticals product
Cost of the product is determined by the DSP involved
The results of the study show a promising role of Acacia Honey, a natural product with proven therapeutic effects on skin wound healing. It accelerated the initial stage of corneal wound healing without the side effects found when using conventional treatments which contain preservatives. Corneal keratocytes cultured in media supplemented with 0.025% Acacia Honey showed an increase in proliferative capacity while retaining their morphology, gene and protein expressions with normal cell cycle.
The current study investigated the immunomodulatory
potential of ethyl acetate soluble supernatant of
Lactobacillus casei (LC-EAS) in vitro. The effect of
LC-EAS on nitric oxide release was analyzed in RAW
264.7 cells, wherein, an inhibition in nitric oxide production
through suppression of inducible nitric oxide synthase
mRNA expression was observed. Evaluation of LC-EAS
on LPS-induced peripheral blood mononuclear cells
showed a down-regulation in TNF-a and IL-6 genes and an
upregulation of IL-10. An inhibition in the protein
expression of NF-kB, ERK1/2 and STAT3 phosphorylation
confirms the immunomodulatory potential of LC-EAS. The
effect of LC-EAS on in vitro intestinal epithelial cells was
investigated using HT-29 human colon adenocarcinoma
cancer cells. LC-EAS exhibited an inhibition of NF-jB and
ERK1/2 phosphorylation, whereas STAT3 phosphorylation
was unregulated. To evaluate the downstream target of
STAT3 upregulation, expression of the intestinal trefoil
factor TFF3 which is a NF-jB regulator and STAT3
downstream target was studied. LC-EAS was observed to
elevate TFF3 mRNA expression. Overall the study shows
that the anti-inflammatory potential of LC-EAS is through
inhibition of NF-kB in different cell types.
ABSTRACT- Background: Declining trend in semen quality is receiving attention worldwide. The aim of the study to investigate the role of lifestyle factors with respect to semen quality.
Methods: Semen samples were collected from 351 male partners attending OPD of Obstetrics and Gynecology at civil hospital and IKD hospital, Ahmedabad, India. They were subjected to assess the quality of semen according to WHO criteria and semen quality were analyzed with respect to self reported history of tobacco smoking and/or chewing and alcohol consumption as lifestyle factors.
Results: The result revealed that sperm count was lower in subjects with habit of tobacco smoking, chewing and alcohol consumption as compared to subjects without such habits and decline was found statistically significant among smokers. Total progressive motility and normal morphology percentage was significantly decreased in tobacco chewers compared to non-chewers. Further analysis of data with respect to alcohol consumption indicated non-significantly lower total progressive motility and normal sperm morphology percentage as compared to subjects with no such habit.
Conclusion: The data obtained suggested, the role of lifestyle factors especially tobacco smoking and chewing in declining semen quality.
Key-words- Semen quality, Tobacco, Alcohol, Lifestyle, Sperm motility, Smoking
1) A Penicillium sp. fungus produced high levels of the softwood-specific enzyme beta-mannanase when grown in solid and liquid cultures containing locust bean gum.
2) Adding a crude enzyme extract from this Penicillium sp. to a mixture of commercial enzymes improved saccharification of steam-exploded radiata pine by 7.8%.
3) Producing crude enzyme extracts on-site from native fungi is a potential strategy to lower the cost of enzymes for breaking down softwood biomass.
This study developed a novel biodegradable porous scaffold for skin regeneration composed of collagen, hyaluronic acid, and gelatin. The scaffold was tested in vitro by culturing human keratinocytes, melanocytes, and fibroblasts, which all proliferated normally. In vivo tests on rats found the scaffold treated wounds healed faster than untreated wounds, with over 50% closure within 7 days versus only 60% for untreated wounds. Histological analysis further supported the scaffold's ability to facilitate more rapid wound healing and regeneration of skin tissue.
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.
This study evaluated the antioxidant and antimicrobial properties of Syzygium jambos. Methanolic extracts and aqueous soluble fractions of S. jambos stem bark exhibited significant antioxidant activity. The chloroform and carbon tetrachloride soluble fractions showed mild antimicrobial activity against several bacteria. The chloroform soluble fraction also demonstrated moderate free radical scavenging activity. Overall, S. jambos extracts displayed antioxidant and limited antimicrobial effects, suggesting potential for further investigation into their bioactive properties.
Study on Culture Conditions for A Cellulase Production From As Pergillus UnguisIJMERJOURNAL
1) The document studies the optimal culture conditions for cellulase production by Aspergillus unguis through solid state fermentation.
2) Experiments were conducted to determine the effect of various carbon sources, nitrogen sources, moisture content, duration and citrate buffer content on cellulase production.
3) The highest cellulase activity of 110.92 U/ml was achieved under the following optimal conditions: rice husk as the carbon source, yeast extract as the nitrogen source, 64% moisture content, 7 days of incubation, and a citrate buffer to byproduct ratio of 7:1.
- Digested black tea induces the secretion of pro-inflammatory cytokines TNF-α and IL-6 in macrophages after 8 hours of exposure, but does not induce secretion of the anti-inflammatory cytokine IL-10. Simulated digestive fluid alone also induces some inflammatory response, but to a lesser extent than digested black tea.
- The study investigated the effects of digested black tea on cytokine secretion in macrophages to better understand how consumed black tea impacts the immune system, as previous studies have not considered the effects of digestion.
This document summarizes a study that transformed the plant Artemisia dubia with the rolA gene using Agrobacterium tumefaciens to increase production of artemisinin and its derivatives. The rolA gene was inserted into A. dubia and transgenic plants were regenerated. Thin layer chromatography analysis found that both rolA and rol ABC transgenic A. dubia produced comparable amounts of artemisinin and derivatives, demonstrating that rolA transformation is effective for enhancing secondary metabolite production. Optimization of the transformation protocol improved regeneration of transgenic A. dubia plants.
Ảnh hưởng của màu sắc đèn led trong kĩ thuật nấm đông trùng hạ thảoHo Chi Minh
Ảnh hưởng của màu sắc đèn led trong kĩ thuật nấm đông trùng hạ thảo
GIA BAO CO.
www.dongtrunghathaogiabao.com
Add: 119A Tran Binh Trong, District 5, Ho Chi Minh City, Viet Nam
Hotline: 0776 950 960
DOI: 10.21276/ijlssr.2016.2.4.7
ABSTRACT- Most of the Euphorbian plants secrete fluid which contain a proteolytic enzyme for defensive role against
insects, pests and hence eco physiological inheritance to sustain vegetation eventually in adverse environmental
conditions. Evaluation has been carried out on twenty five Euphorbian garden plants for their proteolytic activities using
casein as a substrate. Out of these, Euphorbia nerifolia, Euphorbia milli, Euphorbia tirucalli, Euphorbia lactea,
Synadenium granti, Jatropha curcas, Euphorbia nivulia, Euphorbia antiquoram, Pedilanthus tithymaloides, Euphorbia
viguieri, E. heterophylla and E. leucocephala are the good enzyme source. Moderate activity found in Jatropha
integerrima, Jatropha multifida, Jatropha podagrica, Euphorbia pulcherrima, and Dalechampia scandens. While
different tissues of Acalypha hispida, Acalypha wilkesiana, Breynia nivosa, Cicca acida, Codiaeum variegatum, Drypetes
roxburghii are devoid of proteolytic activity. This paper describes in detail about name of plants, habitat and presence of
proteolytic enzyme in them. Results show that the out of twenty five plants 50% plant tissue synthesize protease in
appreciable amount, while 10% are not able to produce it. However 40% plants demonstrate only detectable amount of
protease. A comparative account of proteolytic activity reveals some promising plants good source of enzyme. Some
plants produce combination of cysteine and serine proteases. A single plant i.e. Euphorbia nerifolia latex contains
cysteine, serine, metallo-protease and aspartic proteases. In turn, these proteases may be used in various industrial uses in
general and cheese production Key-words- Garden Euphorb iiann a p plaarnttisc,u Clayr.s t e ine and serine protease, E. leucocephala, Euphorbia viguieri
Effect of Different Concentrations of Glycerol on survival of Azotobacter chr...IIJSRJournal
This document discusses a study that evaluated the effect of different concentrations of glycerol on the survival of the nitrogen-fixing bacterium Azotobacter chroococcum in liquid formulations. Azotobacter was inoculated into Jensen's broth containing various concentrations of glycerol from 5 mM to 25 mM. The number of surviving Azotobacter colonies was counted over 360 days. Results showed that survival was highest in the 25 mM glycerol formulation, with 22 colonies surviving after 360 days. Survival was lowest in the 5 mM glycerol formulation, with only 5 colonies surviving after 360 days. Additionally, three Azotobacter strains showed different survival rates in the 25 mM glycerol formulation over 360 days, demonstrating an
IRJET- An Overview on Fungi as Self Healing Agent in Biomineralization of...IRJET Journal
This document discusses the use of fungi as self-healing agents in biomineralization of calcite in cement concrete. While bacteria have been used for their ability to precipitate calcite via urea hydrolysis, fungi are also efficient in this process. The document reviews studies demonstrating fungal calcite precipitation and describes the methodology for culturing fungi, producing bioconcrete incorporating fungi, and testing the physicochemical and bioengineering properties of the resulting bioconcretes. Key properties examined include compressive strength, crack healing ability, and calcite precipitation as observed using SEM and EDX analysis.
The Studies of Effect of Bio Fertilizers Rhizobium, Phosphobacteria, and Root...IIJSRJournal
A pot culture experiment was carried out and to the find out the effect of Rhizobium, Phosphobacteria and Root Nodule extract on the vegetative growth and biochemical changes in Black gram and Maize. The Black gram and Maize is vegetative parameters such as seed germination, shoot and root length, leaf fresh weight, leafs dry weight, shoot and root dry weight had analyzed and biochemical studies of both Black gram and Maize was carried such as total chlorophyll content, leave soluble protein and in vivo nitrate reductase activity. Among microbial inoculants, the Rhizobium + PB mixer was found most effective in terms of seed germination, shoot and root length, leaf fresh weight, leafs dry weight, shoot and root dry weight and also showed increase total chlorophyll content, soluble protein and nitrate activities in both Black gram and Maize.
Deals with various methods adapted for the Improvement in Microbial Cell Culture alongwith the procedure and instruments used to carry out the operation through illustrative diagrams
SunModo is a solar racking company founded in 2008 that provides a complete line of roof and ground mounting solutions. Their mission is to accelerate adoption of affordable clean energy technologies by providing high quality, low cost, and easily installable racking systems. They offer innovative racking products for roof, flat roof, pitched roof, ground mount, and self-grounding applications. Their focus is on professional installers through responsive support, engineering services, and quick to install systems.
Este documento presenta las normas de comportamiento para usar Internet de manera respetuosa. Súper compu le pide a una persona que use un lenguaje más respetuoso al comunicarse en Internet. Luego, Súper NETI enumera principios como el respeto, la libertad y la intimidad que deben guiar las interacciones en línea. Finalmente, la persona se disculpa por su comportamiento anterior.
O documento analisa as tendências de compra de eletroeletrônicos e eletrodomésticos para o Natal de 2010 no Brasil, com base em dados de mídias sociais e notícias online. Televisões, celulares e computadores aparecem como os produtos eletrônicos mais desejados no Twitter e blogs, enquanto eletrodomésticos recebem menos menções. As vendas online devem crescer 41% em relação ao ano anterior.
Systematic screening was implemented in rural health clinics to improve preventative care delivery. Staff were trained to screen patients for various health issues using a checklist and provide additional services if needed. Shadowing patients found that screening often identified additional needed services but these were not always provided due to time constraints. Interviews found that screening facilitated difficult conversations but implementation challenges included maintaining staff motivation over time. Further analysis will assess the effectiveness, feasibility and value of systematic screening to determine if a revised approach should be tested and scaled up to help reduce health disparities in underserved populations in the US.
Este documento describe los números enteros, los cuales forman un conjunto que incluye números positivos y negativos. Los números enteros se leen por las cifras que están a la izquierda de la coma decimal y representan la cantidad de unidades enteras.
O documento é muito curto e não fornece informações claras ou contexto. Consiste de poucas palavras soltas em inglês e espanhol sem uma estrutura ou significado aparente.
Este documento presenta lineamientos sobre el buen uso y comportamiento en las redes sociales, conocido como netiqueta. En menos de 3 oraciones, resume que se debe mostrar respeto a la privacidad de otros al no publicar información o fotos sin permiso, no agregar amigos desconocidos que podrían dañar la información personal, y usar las redes de forma responsable mostrando la misma conducta que en la vida real.
2016-06 Italy Car Sales Volkswagen June 2016Uli Kaiser
Volkswagen sold 70,667 vehicles domestically in Italy in June 2016. The top 5 selling models were the Golf, Polo, Up!, Passat, and Tiguan. The next most popular models, ranking from 6 to 11, included the Touran, Golf Sportsvan, Caddy, Maggiolino, and Sharan. Of these top 11 models, the Golf, Polo, Up!, Tiguan sold the highest that month at 10,482 units combined.
Um homem encontra a Morte que veio buscá-lo, levando uma maleta vazia. A Morte explica que nada realmente pertencia ao homem - suas posses eram da terra, suas recordações do tempo, seus talentos das circunstâncias, seus amigos e família do caminho, seu corpo da terra e sua alma do Universo. No final, a Morte conforta o homem dizendo que cada momento vivido foi dele.
Paul Rowe has over 20 years of experience in transport, fleet, and operations management. He has held various roles such as Depot Manager, Area Manager, and Area Sales Representative. In his most recent role as a self-employed taxi driver, he was responsible for safely transporting customers to their destinations. Rowe is now looking for a new supervisory or managerial position in transport, fleet, or operations management.
The Staples managers use an administrative management style that incorporates elements of Henri Fayol and Mary Parker Follett's principles. They follow a democratic leadership style, encouraging employee participation and initiative. The managers could improve by ensuring each employee reports to one manager to avoid confusion, and continuing to motivate employees with promotions in addition to compensation.
This document summarizes patient characteristics and antibiotic resistance testing results for 16 cases of Shigella Flexneri infection. The majority of patients were male (100%), white (50%), and between the ages of 23-53 years old (median age 29.5). Most cases were typed as serotype 1B (81%) and displayed resistance to ampicillin (19%) and sulfamethoxazole/trimethoprim (25%). Resistance testing was performed on about a third of cases, with varying levels of susceptibility and resistance seen for the antibiotics tested.
ORDEN CYT/49/2017, de 25 de enero, por la que se modifica la Orden CYT/343/2016, de 14 de abril, por la que se establecen las bases reguladoras para la concesión de subvenciones destinadas a Entidades Locales con una población inferior a 20.000 habitantes para financiar actuaciones en bienes inmuebles integrantes del Patrimonio Cultural de Castilla y León.
The document contains testimonials from several students who have completed Lesson 8 of the Bible correspondence course from the Worldwide Church of God. The students praise the course for opening their eyes to biblical truths they had not understood before. One student says they feel they have learned more real biblical truth from the course than in all previous years. Another says their mind can now accept teachings they previously heard from the pulpit. A third writes that Lessons 7 and 8 alone taught them so much about God's divine plan. All express eagerness to continue learning from the course.
Similar to Synergism effect between inoculum size and aggregate size on flavonoid production in centella asiatica (l.) urban (pegaga) cell suspension cultures
This document provides a summer training report on mammalian cell culture and western blotting techniques conducted at Jawaharlal Nehru University. It describes experiments on reviving cryopreserved cells, subculturing cells, cryopreserving cells, and treating cells with different drug concentrations. It then details the process of western blotting, including extracting whole cell lysates, estimating protein concentration using the Bradford assay, running SDS-PAGE gel electrophoresis, electrotransferring proteins to a membrane, and immunoblotting. The document provides a concise overview of key cell culture and western blotting procedures.
International Journal of Pharmaceutical Science Invention (IJPSI) is an international journal intended for professionals and researchers in all fields of Pahrmaceutical Science. IJPSI publishes research articles and reviews within the whole field Pharmacy and Pharmaceutical Science, new teaching methods, assessment, validation and the impact of new technologies and it will continue to provide information on the latest trends and developments in this ever-expanding subject. The publications of papers are selected through double peer reviewed to ensure originality, relevance, and readability. The articles published in our journal can be accessed online.
CELL SUSPENSION CULTURE AND SECONDARY METABOLITES.pptxBulBulsTutorial
1) Cell suspension cultures involve dispersing plant cell aggregates or single cells in liquid media through agitation, allowing for mass cultivation. This allows independent study of secondary metabolite biosynthesis.
2) Production of secondary metabolites is highest during stationary phase when cell division stops and primary metabolites are converted to secondary metabolites. Medium composition, growth regulators, and culture system (batch vs continuous) impact metabolite yield.
3) Suspension cultures provide advantages over whole plants for industrial production of valuable compounds, including independence from climate and ability to propagate cells clonally.
Abstract — The most common polymer in plastics is
polyethylene (PE), which is made from ethylene monomers
(CH2=CH2). In natural form it is not biodegradable. Low density
polyethylene is a vital cause of environmental pollution. It occurs
by choking sewer line through mishandling thus posing an
everlasting ecological threat, the making of the genetically
engineered microbes for bioremediation, the latter being a
strategy to develop an accelerated evolution of pathways by DNA
restructuring
Abstract — The most common polymer in plastics is
polyethylene (PE), which is made from ethylene monomers
(CH2=CH2). In natural form it is not biodegradable. Low density polyethylene is a vital cause of environmental pollution. It occurs by choking sewer line through mishandling thus posing an everlasting ecological threat, the making of the genetically engineered microbes for bioremediation, the latter being a strategy to develop an accelerated evolution of pathways by DNA restructuring. To enhance the biodegradation of polyethylene, pretreatment strategies were followed. Three different pretreatment strategies were employed for the present study, three duration of Pseudomanas putida treatment to PE were
analyzed on 7, 14, and 28th day. In the first, PE films were
subjected for Bleach with Alkali treatment and in the second they were subjected to UV light (UV-C,>300nm wavelength). Third with the EMS induction of bacterial strains and assessed for polymer biodegradation by Biomass weight loss, estimation of total carbohydrates and total protein in the culture supernatant, followed by DNA isolation for Gel electrophoresis, and Mutated DNA Stability analysis by Capillary Gel electrophoresis were carried out.
This document discusses plant tissue culture, including its definition, history, advantages, disadvantages, and procedures. Some key points:
- Plant tissue culture involves growing plant cells, tissues, or organs in a sterile nutrient medium under controlled conditions. It allows for the production of metabolites and regeneration of whole plants.
- The history of plant tissue culture dates back to the 19th century with early work on cell theory. Significant developments occurred throughout the 20th century, including the discovery of plant growth hormones and establishment of callus culture.
- Advantages include availability of raw materials, disease-free propagation, biosynthetic pathway studies, while disadvantages include requiring expertise, instability, and slow growth.
- Proced
Cells were thawed and plated, then trypsinized and passaged to detach and transfer cells to new plates. Cells were quantified using a hemocytometer after staining with trypan blue. Around 58% viability was observed. Cells were then cryopreserved in DMSO for storage in liquid nitrogen. Proper techniques like quick thawing, plating in fresh media, and passaging help keep cells alive through multiple procedures in cell culture work.
Molecular farming uses plants or plant cells as bioreactors to produce valuable pharmaceutical proteins through recombinant DNA techniques. This document discusses the history, hosts, strategies, applications and case study of molecular farming. It summarizes a study that produced the antibody M12 in tobacco hairy root cultures. The study found optimized conditions that increased secreted antibody yields by 30-fold. Microscopy showed morphological changes and increased protein content in root tissues after induction, supporting more efficient protein secretion.
www.ijera.com 1|P a g e Production of microbial enzymes by new method of cult...IJERA Editor
1. The document describes a new method for cultivating microorganisms using immobilization on a substrate to produce enzymes. Traditional cultivation methods using free-floating cells have limitations like short production cycles.
2. The new immobilization method forms a loose, filamentous-spongy mycelium structure with good nutrient and oxygen access. This extends the active enzyme production phase from 3 to 11-14 days compared to traditional methods.
3. Other advantages of immobilization include increased enzyme productivities up to 10-12 times higher than traditional methods, longer overall cultivation periods of up to 60 days, and the ability to repeatedly harvest enzymes every 1-2 days from the same initial culture.
Suspension Culture and Single Cell Cultures, Culturing methods, maintenance a...Ananya Sinha
Suspension Culture and Single Cell Cultures, Culturing methods, maintenance and application
Generally, suspension culture is a one stop technology to produce secondary metabolites on a large scale in-vitro, irrespective of the climatic condition or nutrient availability (as required in field plants).
In this presentation, we will see the importance of suspension culture, culturing methods and it's application (mostly with respect to plants) and also focus on what exactly is a single cell culture.
I'm delighted to share the PDFs of lab courses in Microbial Physiology and Microbial Genetics. These comprehensive resources cover essential topics in understanding the intricate workings of microbes at a physiological and genetic level. these PDFs provide a detailed roadmap for our laboratory explorations.
Bioprocess development for enhanced spore production in shake flask and pilot...iosrjce
1) The document describes a study that optimized the production medium and cultivation conditions for enhanced spore production of Bacillus thuringiensis var. israelensis in shake flasks and a 16-L bioreactor.
2) In shake flask experiments, the maximal cell dry mass was 4.26 g/L at 36 hours and maximal spore production was 3.29×106 spores/mL.
3) In the 16-L bioreactor under uncontrolled pH, the maximal cell dry mass was 4.14 g/L at 36 hours and maximal spore production was 3.7×106 spores/mL, representing increases of 23% and 47% respectively over the controlled pH
The Role of Cell Wall-Degrading Enzymes in the Development of Anthracnose Dis...Agriculture Journal IJOEAR
This document summarizes a study that evaluated the production of cell wall-degrading enzymes by Colletotrichum truncatum CP2, a fungal pathogen that causes anthracnose disease in chili peppers. The study found that polygalacturonase (PG) was the first cell wall-degrading enzyme detected, with higher activity levels than other enzymes. After PG degraded the cell wall, further degradation was carried out by pectin methylesterases, pectin lyase, and pectate lyase. C. truncatum CP2 produced higher levels of these enzymes compared to the reference fungus C. gloeosporiodes. The timing of peak enzymatic activity suggests each enzyme plays a specific
In vitro experiments of prokaryotic and eukaryotic antimicrobial peptide cyto...AI Publications
These proteinaceous molecules, called antimicrobial peptides (AMPs), are a varied collection of antimicrobial peptides. The ability of AMPs to combat gut infections necessitates further study of the AMP-GI tract interaction. These peptides need to be tested in vitro for cytotoxicity before they may be considered for use in clinical infections. Using the MTT conversion assay, neutral red dye absorption assay, and a comparison to vancomycin, researchers examined the cytotoxicity of gallidermin, nisin A, natural magainin peptides, and melittin in two gastrointestinal cell types (HT29 and Caco-2). Sheep erythrocyte hemolytic activity was also studied, and the influence of AMPs on paracellular permeability was assessed using transepithelial resistance (TEER) and TEM. Gallidermin, nisin A, magainin I, magainin II, and melittin were the least cytotoxic AMPs. To our knowledge, only Melittin and NIS caused considerable hemolysis. There are two distinct ways that melittin and nisin differ in their ability to kill bacteria. It was the only AMP that had an effect on the permeability of the paracellular space. Intestinal tight junctions and cell–cell adhesion were destroyed by long-term melittin therapy, as were microvilli, cell debris, and cell–cell adhesion. Antimicrobial activity and low cytotoxicity make Gallidermin a promising therapeutic drug. The antibacterial properties of Melittin are limited, but its ability to transport poorly bioavailable medicines may be useful.
Isolation and characterization of coprophilous cellulolytic fungi from asian ...Alexander Decker
This document summarizes a study that isolated and characterized coprophilous (dung-loving) cellulolytic fungi from Asian elephant dung in Malaysia. Eight new fungi were isolated from elephant dung samples collected from a forest reserve, identified morphologically and through molecular analysis, and had their DNA sequences deposited in GenBank. Two isolates, Trichoderma aureoviride and Fusarium equiseti, showed potential for cellulase production when tested on carboxymethyl cellulose. The study suggests that natural environments like elephant dung harbor cellulolytic fungi that could provide cheaper cellulase enzymes for applications like biofuel production.
This document describes research aimed at improving the efficiency of somatic embryogenesis in coconut palms. The researchers evaluated secondary somatic embryogenesis and embryogenic callus multiplication, starting with plumule explants. Through a stepwise process involving primary and secondary somatic embryogenesis and callus multiplication over multiple cycles, the yield of somatic embryos produced from one original plumule explant was calculated to be 98,000 embryos, representing as much as a 50,000-fold increase compared to primary somatic embryogenesis alone. Histological analysis provided insights into the cellular development at different stages. The improved protocol allows for significantly higher somatic embryo production efficiency in coconut regeneration.
This document discusses several in vitro methods for assessing the cytotoxicity of chemotherapeutic drugs, including assays using brine shrimp, MTT, sulforhodamine B, trypan blue dye, and acridine orange/ethidium bromide staining. It describes maintaining cell lines from cervical carcinoma and breast adenocarcinoma in culture, assessing cell viability, and preserving cells in liquid nitrogen. Specific methods are provided for assays including brine shrimp lethality, MTT, sulforhodamine B, trypan blue dye exclusion, acridine orange/ethidium bromide staining, and DNA fragmentation to evaluate the cytotoxic effects of test compounds on cultured malignant cell lines.
Optimization of Media and Feeding Strategy for Improved Capsular Polysacchari...theijes
Meningococcal serogroup W-135 (MenW-135) has turned out to be the reason for recent outbreak of meningitis in the sub-Saharan regions of Africa including Cameroon, Chad, Gambia, Burkina Faso, and other countries. The capsular polysaccharide (PS) of Neisseria meningitidis serogroups A, C, X, Y, and W-135 are excellent vaccine targets against meningitis caused by this bacterium. This study promotes a novel feed solution including amino acids, vitamins, and glucose along with an optimal fermentation medium; and feeding strategy to cultivate high yield polysaccharide at harvest level. The study is based on the hypothesis that glucose is one of the major growth determining factors with the combinations of nitrogen sources. In this study, fed-batch cultivation was performed in a 2L fermenter, maintaining the following conditions: (i) Temperature = 37ºC (ii) pH = 7.0 (iii) Agitation frequency in the range 150 rotations per minute (rpm) to 500 rpm, and (iv) Dissolved oxygen (DO) 20%-25%. The bacterial growth and polysaccharide production were found to be enhanced when the feed flow was increased periodically from late lag phase to early decline phase. The polysaccharide production was found to be maximum in the early decline phase of the bacterial growth. The said method produced polysaccharide of about 1000mg/L, which is twice the reported method. There is limited research information available on fed-batch cultivation and feeding strategy for growing Neisseria meningitidis W-135. The major findings of this study can provide significant contribution in the field of capsular polysaccharide production from which MenW polysaccharide vaccines are manufactured.
This document describes research into the microbial synthesis of platinum nanoparticles using Saccharomyces boulardii and evaluation of the anticancer activity of the synthesized platinum nanoparticles. Key findings include:
1) Platinum nanoparticles were successfully synthesized using the cell free extract of S. boulardii when reacted with chloroplatinic acid.
2) Various parameters like metal salt concentration, temperature, cellmass concentration, pH, and reaction time were optimized to control the yield and properties of the synthesized nanoparticles.
3) The synthesized platinum nanoparticles showed anticancer activity against A431 and MCF-7 cell lines with IC50 values between 57-100 μg/ml, indicating potential for use as an antic
This document summarizes an experiment optimizing expression of G-protein coupled receptors (GPCRs) in HEK293 cells using a bioreactor. The researchers found that using a bioreactor allowed for higher cell densities and expression levels of over 1 mg of recombinant GPCR per liter of culture, compared to 0.1-0.2 mg/L using spinner flasks. A large-scale run in the bioreactor produced 10 mg of purified, bioactive recombinant GPCR. The bioreactor setup improved expression levels for structural characterization of this important class of membrane proteins.
Similar to Synergism effect between inoculum size and aggregate size on flavonoid production in centella asiatica (l.) urban (pegaga) cell suspension cultures (20)
Mechanical properties of hybrid fiber reinforced concrete for pavementseSAT Journals
Abstract
The effect of addition of mono fibers and hybrid fibers on the mechanical properties of concrete mixture is studied in the present
investigation. Steel fibers of 1% and polypropylene fibers 0.036% were added individually to the concrete mixture as mono fibers and
then they were added together to form a hybrid fiber reinforced concrete. Mechanical properties such as compressive, split tensile and
flexural strength were determined. The results show that hybrid fibers improve the compressive strength marginally as compared to
mono fibers. Whereas, hybridization improves split tensile strength and flexural strength noticeably.
Keywords:-Hybridization, mono fibers, steel fiber, polypropylene fiber, Improvement in mechanical properties.
Material management in construction – a case studyeSAT Journals
Abstract
The objective of the present study is to understand about all the problems occurring in the company because of improper application
of material management. In construction project operation, often there is a project cost variance in terms of the material, equipments,
manpower, subcontractor, overhead cost, and general condition. Material is the main component in construction projects. Therefore,
if the material management is not properly managed it will create a project cost variance. Project cost can be controlled by taking
corrective actions towards the cost variance. Therefore a methodology is used to diagnose and evaluate the procurement process
involved in material management and launch a continuous improvement was developed and applied. A thorough study was carried
out along with study of cases, surveys and interviews to professionals involved in this area. As a result, a methodology for diagnosis
and improvement was proposed and tested in selected projects. The results obtained show that the main problem of procurement is
related to schedule delays and lack of specified quality for the project. To prevent this situation it is often necessary to dedicate
important resources like money, personnel, time, etc. To monitor and control the process. A great potential for improvement was
detected if state of the art technologies such as, electronic mail, electronic data interchange (EDI), and analysis were applied to the
procurement process. These helped to eliminate the root causes for many types of problems that were detected.
Managing drought short term strategies in semi arid regions a case studyeSAT Journals
Abstract
Drought management needs multidisciplinary action. Interdisciplinary efforts among the experts in various fields of the droughts
prone areas are helpful to achieve tangible and permanent solution for this recurring problem. The Gulbarga district having the total
area around 16, 240 sq.km, and accounts 8.45 per cent of the Karnataka state area. The district has been situated with latitude 17º 19'
60" North and longitude of 76 º 49' 60" east. The district is situated entirely on the Deccan plateau positioned at a height of 300 to
750 m above MSL. Sub-tropical, semi-arid type is one among the drought prone districts of Karnataka State. The drought
management is very important for a district like Gulbarga. In this paper various short term strategies are discussed to mitigate the
drought condition in the district.
Keywords: Drought, South-West monsoon, Semi-Arid, Rainfall, Strategies etc.
Life cycle cost analysis of overlay for an urban road in bangaloreeSAT Journals
Abstract
Pavements are subjected to severe condition of stresses and weathering effects from the day they are constructed and opened to traffic
mainly due to its fatigue behavior and environmental effects. Therefore, pavement rehabilitation is one of the most important
components of entire road systems. This paper highlights the design of concrete pavement with added mono fibers like polypropylene,
steel and hybrid fibres for a widened portion of existing concrete pavement and various overlay alternatives for an existing
bituminous pavement in an urban road in Bangalore. Along with this, Life cycle cost analyses at these sections are done by Net
Present Value (NPV) method to identify the most feasible option. The results show that though the initial cost of construction of
concrete overlay is high, over a period of time it prove to be better than the bituminous overlay considering the whole life cycle cost.
The economic analysis also indicates that, out of the three fibre options, hybrid reinforced concrete would be economical without
compromising the performance of the pavement.
Keywords: - Fatigue, Life cycle cost analysis, Net Present Value method, Overlay, Rehabilitation
Laboratory studies of dense bituminous mixes ii with reclaimed asphalt materialseSAT Journals
Abstract
The issue of growing demand on our nation’s roadways over that past couple of decades, decreasing budgetary funds, and the need to
provide a safe, efficient, and cost effective roadway system has led to a dramatic increase in the need to rehabilitate our existing
pavements and the issue of building sustainable road infrastructure in India. With these emergency of the mentioned needs and this
are today’s burning issue and has become the purpose of the study.
In the present study, the samples of existing bituminous layer materials were collected from NH-48(Devahalli to Hassan) site.The
mixtures were designed by Marshall Method as per Asphalt institute (MS-II) at 20% and 30% Reclaimed Asphalt Pavement (RAP).
RAP material was blended with virgin aggregate such that all specimens tested for the, Dense Bituminous Macadam-II (DBM-II)
gradation as per Ministry of Roads, Transport, and Highways (MoRT&H) and cost analysis were carried out to know the economics.
Laboratory results and analysis showed the use of recycled materials showed significant variability in Marshall Stability, and the
variability increased with the increase in RAP content. The saving can be realized from utilization of recycled materials as per the
methodology, the reduction in the total cost is 19%, 30%, comparing with the virgin mixes.
Keywords: Reclaimed Asphalt Pavement, Marshall Stability, MS-II, Dense Bituminous Macadam-II
Laboratory investigation of expansive soil stabilized with natural inorganic ...eSAT Journals
This document summarizes a study on stabilizing expansive black cotton soil with the natural inorganic stabilizer RBI-81. Laboratory tests were conducted to evaluate the effect of RBI-81 on the soil's engineering properties. The tests showed that with 2% RBI-81 and 28 days of curing, the unconfined compressive strength increased by around 250% and the CBR value improved by approximately 400% compared to the untreated soil. Overall, the study found that RBI-81 effectively improved the strength properties of the black cotton soil and its suitability as a soil stabilizer was supported.
Influence of reinforcement on the behavior of hollow concrete block masonry p...eSAT Journals
Abstract
Reinforced masonry was developed to exploit the strength potential of masonry and to solve its lack of tensile strength. Experimental
and analytical studies have been carried out to investigate the effect of reinforcement on the behavior of hollow concrete block
masonry prisms under compression and to predict ultimate failure compressive strength. In the numerical program, three dimensional
non-linear finite elements (FE) model based on the micro-modeling approach is developed for both unreinforced and reinforced
masonry prisms using ANSYS (14.5). The proposed FE model uses multi-linear stress-strain relationships to model the non-linear
behavior of hollow concrete block, mortar, and grout. Willam-Warnke’s five parameter failure theory has been adopted to model the
failure of masonry materials. The comparison of the numerical and experimental results indicates that the FE models can successfully
capture the highly nonlinear behavior of the physical specimens and accurately predict their strength and failure mechanisms.
Keywords: Structural masonry, Hollow concrete block prism, grout, Compression failure, Finite element method,
Numerical modeling.
Influence of compaction energy on soil stabilized with chemical stabilizereSAT Journals
This document summarizes a study on the influence of compaction energy on soil stabilized with a chemical stabilizer. Laboratory tests were conducted on locally available loamy soil treated with a patented polymer liquid stabilizer and compacted at four different energy levels. The study found that increasing the compaction effort increased the density of both untreated and treated soil, but the rate of increase was lower for stabilized soil. Treating the soil with the stabilizer improved its unconfined compressive strength and resilient modulus, and reduced accumulated plastic strain, with these properties further improved by higher compaction efforts. The stabilized soil exhibited strength and performance benefits compared to the untreated soil.
Geographical information system (gis) for water resources managementeSAT Journals
This document describes a hydrological framework developed in the form of a Hydrologic Information System (HIS) to meet the information needs of various government departments related to water management in a state. The HIS consists of a hydrological database coupled with tools for collecting and analyzing spatial and non-spatial water resources data. It also incorporates a hydrological model to indirectly assess water balance components over space and time. A web-based GIS portal was created to allow users to access and visualize the hydrological data, as well as outputs from the SWAT hydrological model. The framework is intended to facilitate integrated water resources planning and management across different administrative levels.
Forest type mapping of bidar forest division, karnataka using geoinformatics ...eSAT Journals
Abstract
The study demonstrate the potentiality of satellite remote sensing technique for the generation of baseline information on forest types
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crop land, 1.778% of the area is covered by dry deciduous forest, 1.38 % of mixed plantation, which is very threatening to the
environmental stability of the forest, future plantation site has been mapped. With the use of latest Geo-informatics technology proper
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to shorter edge); (c) edge column (bending parallel to shorter edge); (d) corner column. These design sheets are prepared as per
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GIS techniques are being increasingly used for planning, management and development of natural resources. The study area, Nallur
Amanikere watershed geographically lies between 110 38’ and 110 52’ N latitude and 760 30’ and 760 50’ E longitude with an area of
415.68 Sq. km. The thematic layers such as land use/land cover and soil maps were derived from remotely sensed data and overlayed
through ArcGIS software to assign the curve number on polygon wise. The daily rainfall data of six rain gauge stations in and around
the watershed (2001-2011) was used to estimate the daily runoff from the watershed using Soil Conservation Service - Curve Number
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Tools & Techniques for Commissioning and Maintaining PV Systems W-Animations ...Transcat
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Supermarket Management System Project Report.pdfKamal Acharya
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Synergism effect between inoculum size and aggregate size on flavonoid production in centella asiatica (l.) urban (pegaga) cell suspension cultures
1. IJRET: International Journal of Research in Engineering and Technology eISSN: 2319-1163 | pISSN: 2321-7308
__________________________________________________________________________________________
Volume: 02 Issue: 08 | Sep-2013, Available @ http://www.ijret.org 244
SYNERGISM EFFECT BETWEEN INOCULUM SIZE AND AGGREGATE
SIZE ON FLAVONOID PRODUCTION IN CENTELLA ASIATICA (L.)
URBAN (PEGAGA) CELL SUSPENSION CULTURES
Suat-HianTan1
, Maziah Mahmood2
, Arbakariya Ariff 3
1
Doctor, Faculty of Industrial Science and Technology, University Malaysia Pahang, Lebuhraya Tun Razak, 26300
Gambang, Kuantan, Pahang Darul Makmur, tshian@ump.edu.my
2
Professor Doctor, Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra
Malaysia, 43400 UPM Serdang, Selangor Darul Ehsan, Malaysia, maziahm@biotech.upm.edu.my
3
Professor Doctor, Department of Bioprocess Technology, Faculty of Biotechnology and Biomolecular Sciences,
Universiti Putra Malaysia, 43400 UPM Serdang, Selangor Darul Ehsan, Malaysia, arbarif@upm.edu.my
Abstract
The present study aimed to investigate the effect of different culture conditions on biomass content and flavonoid production of the
elite C. asiatica accession UPM03. When 0.1 g inoculum 25 mL-1
of cell was grown in Murashige & Skoog (MS) medium
supplemented with 2 mg 2,4-D l-1
and 1 mg kinetin l-1
, flavonoids were not significantly produced in cells or released into the culture
medium. Production of flavonoid was correlated with the aggregation size and inoculum density. At aggregate size of 250-500 µm
with 0.3 g inoculum density 25 mL-1
, it gave the highest luteolin content with 35.45 µg g-1
DW. After investigating the effect of culture
conditions, i.e. pH, inoculum density, light irradiation and plant growth regulator, we found that with the optimized condition (i.e.
250-500 µm aggregate size, 0.5 g wet cell 25 mL-1
supplemented with 3 mg L-1
2,4-D and 1 mg L-1
kinetin at pH 5.7 under 16h
photoperiod) the luteolin content was 11-fold higher than the cell suspension cultured at an inoculum size of 0.3 g wet cell 25 mL-1
with 250-500 µm aggregate size.
Index Terms: Centella asiatica, flavonoid, light irradiation, pH regime, plant growth regulators .
-----------------------------------------------------------------------***-----------------------------------------------------------------------
1. INTRODUCTION
Centella asiatica is an important medicinal herb and
commercially grown in Malaysia. This herbaceous plant has
widely been used for wound healing, memory improvement,
treatment for mental fatigue, bronchitis, urethritis, anti-allergic
and anti-cancer purposes [1]. The different uses claimed for
this plant are reported to be mainly due to its high content of
secondary metabolites especially flavonoids and triterpenoids
[2]. The interesting biological activities of flavonoids and their
low content in intact plants impose the development of
alternative ways for higher production of flavonoids. Cultured
plant cells are considered as known producers of secondary
metabolites. Cell cultures not only have a higher rate of
metabolism than differentiated plants, but also have a shorter
period of biosynthetic cycles [3]. Since plant cell culture
systems are relatively easy to manipulate by experimental
processes, they could be developed into a large-scale culture
which the secondary metabolites could be extracted. This
method can supply the continuous and reliable source of
natural products. Discoveries of cell culture capability in
producing specific medicinal compounds at a rate similar or
superior to the intact plants have accelerated in the last few
years [4].
Plant cells have the natural ability to aggregate into
macroscopic clumps. Xu and co-researchers [5] reported that
uncontrolled clumping often causes operating problems during
large-scale suspension cultures, but the tendency to aggregate
can in some cases eliminate the need for artificial
immobilization supports and may be exploited for large-scale
phytochemical synthesis. When a smaller size of cell
aggregates is preferred from the stand point of process
engineering, a certain degree of cell-cell contact and cell
differentiation is required for the synthesis of secondary
metabolites [6].
Although for many species [7, 8] for commercial production
of secondary metabolites, the low yield is one of the major
problems that challenges the profitable commercialization.
The morphological differentiation and biochemical processes
are often linked in tissue cultures. For many plant species, the
synthetic capacity of dedifferentiated cells is lower than that of
the fully differentiated tissues, both quantitatively and
qualitatively [9]. Chemical gradients in differentiated tissue
2. IJRET: International Journal of Research in Engineering and Technology eISSN: 2319-1163 | pISSN: 2321-7308
__________________________________________________________________________________________
Volume: 02 Issue: 08 | Sep-2013, Available @ http://www.ijret.org 245
complex or cell aggregates encourage the synthesis of
secondary metabolites. The morphological differentiation of
the cell is manipulated by changing the biological, chemical
and physical factors [10]. Hence, this study focused on each or
different combination of the following factors, namely; the
cellular/tissue differentiation status, external phytohormones,
culture media and culture conditions to increase the yields of
flavonoids in the cultured C. asiatica cell suspensions.
2. MATERIALS AND METHODS
2.1 Induction and Maintenance of Callus Cultures
In vitro C. asiatica leaf explants of the UPM03 accession were
cultured on Murashige and Skoog (MS) [11] medium with B5
vitamins [12], 30 g L-1
sucrose, 2.75 g L-1
gelrite, 2 mg L-1
2,4-dichlorophenoxy acetic acid (2,4-D) and 1mg L-1
kinetin
to initiate callus formation and maintained at 25±2°C under
16/8h photoperiods.
2.2 Cell Suspension Cultures
The cell suspension cultures were established according to
Tan et al. [13] and were maintained in the MS basal medium
supplemented with 2 mg L-1
2,4-D and 1 mg L-1
kinetin on a
rotary shaker at 110 rpm. In order to obtain suitable and
homogenous growing cells, sub-culturing and sieving with
750 µm pore size were carried out every 12 days. Sub-
culturing was performed by transferring 5 mL of the
suspension cultures into 20 mL of fresh liquid medium in 100
mL Erlenmeyer flasks. This stock culture was used as inocula.
2.3 Culture Condition Studied
The 12-day-old cell suspension was partitioned into three
fractions by sieving through a stainless-steel mesh with 750
µm pore size and followed by filtration through a stainless-
steel mesh with a smaller pore size of 500 µm. Finally, the
filtrates were sieved through another smaller pore size of 250
µm. The first two fractions were quickly transferred back into
the autoclaved media after flushing with the rinsing medium
(same composition as maintenance medium) to avoid any
damage of the cells.
The effects of cell inoculum size were tested, ranging from 0.1
g to 1.5 g of wet cells in 25 mL (4-60 g fresh weight per litre).
For the preparation of 0.1 g wet cells 25 mL-1 inoculum size,
the stock cultures were sieved through a stainless steel mesh
750 µm and 5 mL (approximately 0.10 ± 0.02 g) of the cells
were then transferred into 20 mL of fresh liquid medium. To
determine the accurate density, 5 mL inoculum of sieved stock
cultures was centrifuged at 10,000 rpm for 15 min at 4ºC
(Sigma K-20, USA). The supernatant was discarded and the
fresh cell weight was determined. In the case where the fresh
weight was less than 0.10 ± 0.02 g, more inoculums were
added to achieve the control density. Conversely, and if the
fresh cell weight was more than 0.10 ± 0.02 g, some media
were added into the inoculum. A total of 5 mL inoculum
containing 0.10 ± 0.02 g fresh cells was then inoculated into
20 mL of medium in a 100 mL Erlenmeyer flask. The same
procedures were repeated for other inoculum sizes.
The effects of initial pH values were also tested, ranging from
3 to 9 with ph 5.7 as the control value. The pH of the media
was adjusted to pH 3, 5, 5.7 and 9 with 0.01 M HCl or 0.01 M
NaOH using the pH meter (Mettler Toledo FE20,
Schwarzenbach, Switzerland) prior to autoclaving.
For light irradiation investigation, the cultures were incubated
under either darkness or continuous exposure to light intensity
of 433.33 µMol m-2
s-2
. To create a dark condition for the
cultures, all the culture flasks were covered with a layer of
black sugar paper.
The two PGRs chosen were 2,4-D and kinetin because they
had shown potential influences both in cell growth and
flavonoid production during the previous establishment of cell
suspension cultures using picloram, 2,4-D, 1-
Naphthaleneacetic acid (NAA), thidiazuron (TDZ), kinetin
and 6-Benzylaminopurine (BAP) [13]. The concentrations of
2,4-D used were 1-3 mg/L while for kinetin was ranged from
1-2mg/L. All twelve combinations were tested in a Latin
Square arrangement in order see which combination produces
the optimum level of flavonoid. The control was MS medium
without any hormone.
2.4 Sampling, Determination of Biomass and
Residual Sugar
For sampling, three flasks were taken each time. In the cell
suspension establishment, the cells were sieved with opening
750 µm after 10 days. The cells were collected on filter paper
by vacuum filtration. The fresh weight of cells was determined
before being dried in the oven at 50ºC until a constant weight
was reached and recorded as dry weight (DW). After
sampling, the residual sugar level in the supernatant was
assayed by Anthrone method [14].
2.5 Measurement of Flavonoid in Cell and in Media
The media (40 mL) was concentrated by using a rotor
evaporator at 40ºC. Flavonoid from the dried cells and
concentrated media were extracted as described by Crozier et
al. [15] and was measured according to Marinova et al. [16].
The main compound, luteolin, was quantified using HPLC
method as explained by Crozier et al. [15]. For analysis of the
flavonoid content, the extraction and hydrolysis conditions
were based on the method described by Crozier et al. [15]. The
oven-dried cells were extracted with 10 mL of 60% aqueous
methanol containing 20 mM sodium diethyldithiocarbamate
(NaEDTC) as an antioxidant. They were hormogenized with
mortar and pestle before 2.5 mL 6 M HCl was added to each
extract to give a 12.5 ml solution of 1.2 M HCl in 50%
3. IJRET: International Journal of Research in Engineering and Technology eISSN: 2319-1163 | pISSN: 2321-7308
__________________________________________________________________________________________
Volume: 02 Issue: 08 | Sep-2013, Available @ http://www.ijret.org 246
aqueous methanol. The extracts were refluxed at 90°C for two
h and known as hydrolyzed extracts. The total flavonoid
content was determined by using the aluminium chloride
colorimetric assay method according to Marinova et al. [16].
Luteolin was used as the standard flavonoid.
For HPLC analysis, the hydrolysed extracts were filtered
through a 0.45 μm filter (Minisart RC 15, Sartorius, Germany)
and analysed using the Waters (Milford, MA, USA) liquid
chromatograph comprising an empowered chromatography
manager, a 717 plus autoinjector, 501 HPLC pump and a 486-
tunable absorbance detector. Reverse-phase separations were
carried out at room temperature using a 150 x 3.9 mm I.D.,
4μm C18 Nova-Pak column (Waters). The column was eluted
using gradient of 22%-35% acetonitrile in water, adjusted to
pH 2.5 with Trifluoroacetic Acid (TFA) at a flow rate of 0.6
mL min−1
. The flavonoid compound was detected at 365 nm.
Identification of the compound was achieved by comparison
with the commercial standard, luteolin. The quantity of
flavonoid compounds in each sample was determined from the
standard curve.
2.6 Statistical Analysis
All the experiment was run in triplicate and repeated thrice.
The data were analyzed using the one-way ANOVA. The
mean values were compared utilizing Duncan’s multiple range
test at 5% (p = 0.05) significance level, using the SPSS
software version 11.5 (SPSS Inc. USA).
3.0 RESULTS
3.1 Effect of Aggregate Sizes and Inoculum Densities
A preliminary investigation had shown that 0.1 g wet cells 25
mL-1
inoculum was sufficient to support the cell growth and
provided a sigmoidal growth curve. However, there was no
significant flavonoid production during the cultivation period
until the cells went through the necrosis process where the
cultures turned brown in colour (data not shown). Since it had
been reported that a relative bigger size of aggregation helped
to increase the secondary compound formation remarkably [5,
17, 18], thus, an investigation was carried out by using
different aggregation sizes. The effects of aggregate sizes on
the cell growth in the cell suspension of Malaysian C. asiatica
accession of UPM03 are shown in Fig. 1a. For the aggregation
sizes within the range of 250-750 µm, a lag phase of four days
was observed. Apparently, there was no growth of the cells
with the aggregate being less 250 µm. This suggested that 0.1
g wet cells 25 mL-1
could not support the cell growth of all the
fractions during the cultivation of C. asiatica cell suspensions.
For the aggregation sizes of 250-500 µm and 500-750 µm, the
maximum cell concentrations were 0.18 g DW and 0.21 g
DW, respectively. The final cell concentration was very low
with a small aggregate fraction. In addition, the cell growth
was greatly enhanced with the increment of aggregate
fractions under 0.1 g wet cells 25 mL-1
. It appeared that the
maximal biomass of 0.21g DW/culture obtained at the biggest
aggregate fraction of 500-750 µm was about 35-fold higher
compared to the minimal biomass of 0.006 g DW/culture
obtained at the smallest aggregate fraction (< 250 µm).
a
b
c
Fig -1: The effects of different cell aggregation sizes on the
biomass accumulation (a) flavonoid production in the cell (b)
and in the medium (c) of C. asiatica accession of UPM03. An
inoculums size of 0.1 g wet cells were cultured on a 25 mL
MS medium for 26 days. The bar indicates the standard
deviation of mean (n=3).
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Fig. 1b and 1c show the time profiles of the flavonoid
production under various aggregate fractions in the cells and
the media, respectively. An increase of aggregate
fractionsfrom <250 µm to 750 µm enhanced the flavonoid
contents in the media. However, an opposite trend was
observed in the flavonoid production on the dry weight basis.
It appeared that, there were no significant differences between
the flavonoid production and the cultivation period of 26 days
for the various aggregate fractions either in the cells or in the
media. The three aggregate fractions were found not only
unable to sustain the growth of the cells but also unable to
promote the flavonoid production either in the cells or the
media. Thus, a further increment of inoculum size to 0.3 g wet
cells 25 L-1
was used with the three aggregate fractions to
investigate the cell growth and flavonoid production.
a
b
c
Fig -2: The effect of different cell aggregation sizes on the
biomass accumulation (a), flavonoid production in the
medium (c) of C. asiatica accession of UPM03. The inoculum
size of 0.3 g wet cell were cultured in a 25 ml MS medium for
26 days.
Fig. 2a shows that 500-750 µm fractions reached the
stationary growth phase on the 22nd
day in the suspension
culture under the described conditions, whereas 250-500 µm
and <250 µm fractions have a longer growth phase up to the
24th
day. Similarly, this observation was the same with the C.
asiatica cultures using 0.1 g wet cells 25 mL-1
MS medium,
whereby 500-750 µm was the highest biomass accumulation
on the 16th
day and 250-500 µm on the 20th
day. During the
middle of exponential stage in the suspension culture, the
flavonoid content in the cells increased rapidly to high levels
of 7.38 ± 0.09, 7.05 ± 0.07 and 6.02 ± 0.05 mg g-1
DW for the
samples collected on day 18 (<250 µm), day 12 (250-500 µm)
and day 16 (500-750 µm), respectively. The content reduced
when the cells entered the end of the exponential phase and
reached the low levels of 2.38 ± 0.07 mg g-1
DW (<250 µm),
3.85 ± 0.55 mg g-1
DW (250-500 µm) and 3.99 ± 0.26 mg g-1
DW (500-750 µm) on around the 22nd
day (Fig. 2b). The main
compound, luteolin was quantified for the samples, which
have the highest flavonoid content. Luteolin differed with the
aggregate size. The highest luteolin level was detected in the
250-500 µm aggregate size with 35.45 ± 2.2 µg g-1
DW (Table
1).
For flavonoid accumulation in the media as illustrated in Fig.
2c, the results showed that 250-500 µm of fractions attained
the highest flavonoid content, i.e. 3.02 ± 0.05 mg g-1
, on day
10, followed by 500-750 µm (2.89 ± 0.43 mg g-1
) on day 22
and <250 µm (1.74 ± 0.11 mg g-1
) on day 16. The flavonoid
accumulation patterns for the three aggregate sizes were
different among themselves (Fig. 2c).
Fig. 3 shows the dynamic profiles of sucrose consumption
under various aggregation sizes. The sucrose consumption
corresponded to the growth of C. asiatica. While medium
sucrose was almost consumed on day 18, the cell density
reached the peak on day 22 (Fig. 2a). Both the cell growth and
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sucrose consumption rates were relatively lower at an
aggregate size of <250 µm, while they were found almost
constant at bigger aggregate sizes. The average glucose
consumption rates were 0.58, 0.59 and 0.69 g glucose l-1
per
day, and the corresponding cell yields were 0.025, 0.018,
0.019 g cell/g glucose for <250 µm, 250-500 µm and 500-750
µm of aggregate size, respectively. No significant effect was
observed on the cell yields against sucrose during the
cultivation of C. asiatica for the bigger aggregate sizes.
Fig -3: Sucrose consumption profiles of different aggregation
sizes of C. asiatica accession of UPM03 cells in the
suspension cultures. The inoculums size of 0.3 g wet cells
were cultured in a 25 ml MS medium for 26 days.
3.2 Effect of Inoculum Densities
The effects of inoculum size within the range of 0.1 to 1.5 g
wet cells 25 mL-1
on the C. asiatica cell cultures were
investigated. The differences in biomass accumulation by the
use of different inoculum sizes are shown in Fig. 4a for fresh
and dry weight. The results revealed that the biomass cells
were very low at low inoculum size and the cell growth was
greatly enhanced with the increment of inoculum size from 0.1
to 1.5 g wet cells 25 mL-1
. The maximum biomass yields of
3.05 g FW and 0.14 g DW were obtained at inoculum size of
1.5 g wet cells 25 mL-1
. It appeared that the flavonoid content
of cell suspensions on the dry weight basis differed
remarkably with the inoculums sizes.
a
b
Fig -4: The effect of cell inoculum size on the biomass
accumulation (a) and flavonoid production in the cell and
media (b) of C. asiatica accession of UPM03. The different
letters indicate that the values are significantly different
(p≤0.05).
Fig. 4b. In the inoculums with the sizes less than 1 g wet cells
25 mL-1
, the flavonoid content increased with the inoculum
sizes. However, when the inoculum sizes exceeded 1.0 g wet
cells 25 mL-1
, the value remained the same. Likewise, a
similar pattern was observed in the flavonoid production in the
media. These findings indicated that large sizes of inoculum
resulted in low flavonoid contents.
3.3 Effect of Initial pH Values
The cell suspension was exposed to four pH regimes. The
profiles for biomass production in fresh and dry weights are
shown in Fig. 5a. Both pH 5 and pH 5.7 obtained a very
similar biomass concentration of 0.158 g/culture and 0.157
g/culture on the dry weight basis, respectively. Meanwhile,
Fig. 5b shows the flavonoid production in different pH values
intracellularlly and extracellularly, respectively. In the cells,
the accumulation of flavonoids was maximum at pH 5.7, i.e.
3.658 mg g-1
DW. However, the extracellular flavonoid
content in the media was increased gradually from pH 3 to pH
9. In the high pH value of 9, the media changed colour from
clear to reddish on day 6.
3.4 Effect of Light Irradiation
Fig. 6a illustrates that continuous high level light irradiation
was found to be able to increase the cell growth significantly
in both fresh and dry cell weights. Additionally, flavonoid
biosynthesis was stimulated considerably by high level light
irradiation in C. asiatica cell suspension either in the cells or
in the media (Fig. 6b). The flavonoid content was stimulated
almost 2.5-fold for the light-treated culture with 3.66 mg g-1
as
compared to the dark-treated cell cultures with only 1.41 mg g-
1
. Although the extracellular flavonoid content was
comparably lower than the intracellular flavonoid content, the
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Volume: 02 Issue: 08 | Sep-2013, Available @ http://www.ijret.org 249
extracellular flavonoid content in the light-treated cultures was
0.79 mg g-1
, which showed 3.6-fold increase compared to the
dark-treated cultures (0.23 mg g-1
).
3.5 Effect of Plant Growth Regulators
In the preliminary study on the establishment of cell
suspension cultures using different auxins and cytokinins, 2,4-
D and kinetin were found to be the most potential PGRs in
promoting biosynthesis of flavonoids [7]. Thus, in this study,
both plant growth regulators were tested at different
concentrations and various combinations. Fig. 7a shows the
biomass accumulation of fresh and dry weight of the cultured
cells obtained under different combinations of 2,4-D and
kinetin. When the concentration of 2,4-D was increased in the
treatment, the cell growth was decreased. On the contrary,
with the increased concentration of the kinetin, the cell growth
was also increased. The increase in cell growth could be due
to the expansion of cells which increased the cell size to form
bigger clumps. The results showed that the flavonoid
production in the cells was significantly increased with the
increasing concentrations of 2,4-D (Fig. 7b). Flavonoid
content was significantly declined with the increasing
concentration of Kinetin. Evidently, the flavonoid content
increased up to 11.87 mg g-1
DW in the cells as well as 6.62
mg g-1
in the media (Fig. 7b), four-fold and 1.3-fold
increments compared to the control medium when the medium
was supplemented with 3 mg L-1
2,4-D and 1 mg L-1
kinetin,
respectively. Furthermore, minimal growths of 0.07 mg g-1
FW and 0.063 mg g-1
DW were obtained. The trend
demonstrated that when the cell growth decreased, the
flavonoid content in the cells increased. However, there were
no significant differences among the flavonoid content in
media within all the concentrations except for the
supplementation of 2,4-D and kinetin with the ratio 3:1 and
3:2. A high concentration of 2,4-D was more effective in
combination with a low concentration of kinetin for enhancing
the flavonoid content in in vitro cultures. Luteolin content was
greatly enhanced to 390±30 µg/g DW in the cell when treated
with 3 mg L-1
2,4-D and 1 mg L-1
kinetin (Table -1).
a
b
Fig -5: The effect of initial pH values on the biomass
accumulation (a) and flavonoid production in the cell and
media (b) of C. asiatica accession of UPM03. They were
cultured in a 100 ml conical flask at an inoculums size of 0.5 g
wet cells in 25ml for 12 days. The size of aggregate used was
250-500 µm. The different letters indicate that the values are
significantly different (p≤0.05).
Table -1: Luteolin production in each size aggregation with it
harvested day in the C. asiatica cell suspension cultures at pH
5.7 with a photoperiod of 16h under the fluorescent light.
Cell culture condition
Inoculum
size
Luteolin
production
(µg/g DW)
2mg L-1
2,4-D
+ 1mg L-1
kinetin
+ 0.3 g wet cell 25 mL-1
500-750 µm
(day18)
30.32 ± 2.1
250-500 µm
(day 12)
35.45 ± 2.2
<250 µm
(day 16)
20.55 ± 1.9
2 mg L-1
2,4-D
+ 1mg L-1
kinetin
+ 0.5 g wet cell 25 mL-1
250-500 µm
(day 12)
110 ± 13
3mg L-1
2,4-D
+ 1mg L-1
kinetin
+ 0.5 g wet cell 25 mL-1
250-500 µm
(day 12)
390±33
Each value represents the mean ± SE, n=3
b
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a
b
Fig -6: The effect of different light sources on the biomass
accumulation (a) and flavonoid production in the cell and
media (b) of C. asiatica accession of UPM03. They were
cultured in a 100 ml conical flask at an inoculum size of 0.5 g
wet cells in 25 ml after cultured for 12 days. The size of
aggregate used was 250-500 µm. The different letters indicate
that the values are significantly different (p≤0.05).
a
b
Fig -7: The effect of 2,4-D (D) and kinetin (K) in combination
on the biomass accumulation (a) flavonoid production in the
cell and media (b) of C. asiatica accession of UPM03. They
were cultured at an inoculum size of 0.5 g wet cells in 25 ml
after cultured for 12 days. The size of aggregate used was 250-
500 µm. The different letters indicate that the values are
significantly different (p≤0.05).
4. DISCUSSION
Primary metabolism is associated with the exponential phase
of a culture where the sole products of metabolism are either
essential for growth or the by-products of energy-yielding
metabolism [19]. In most batches of cell suspension cultures,
secondary metabolite accumulation tends to increase at the
end of the rapid cell division in the growth cycle [20].
However, in some cases, the production of secondary products
did not show a positive correlation with the maximal growth
rate of the culture [19].
The early production of flavonoid during the cell cultivation
was consistent with the previous studies and suggested that the
secondary metabolites could be accumulated maximally
during the exponential phase. Wu et al. [9] reported that the
maximum salidroside yield of Rhodiola sachalenensis
suspension cultures was 19.69 ± 1.84 mg L-1
salidroside on
day 10 (early exponential phase) after inoculation.
Additionally, Jacinda et al. [19] found that the highest
concentrations of the targeted triterpenoids in the cell
suspension of C. asiatica were observed on day 16 and day 20
for both investigated accessions at the end of the exponential
phase.
It appeared that high cell density favored mass tissue
formation and played an important role in the flavonoid
production either in the cells or in the media as well as the
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Volume: 02 Issue: 08 | Sep-2013, Available @ http://www.ijret.org 251
time of flavonoid accumulation. These observations might
reflect a competition for metabolites utilized in the primary
metabolism with those pathways leading to the formation of
the secondary products [19]. A similar phenomenon also
occurred in the cucurbit cell cultures for cucurbitacin
production [21]. Rama et al. [22] also found that the cell
aggregates at 500 µm diameter favoured the biomass
accumulation, while the accumulation of berberine increased
with the increase in cell aggregate size.
There was a minimum size of explants or quantity of separated
cells per unit culture volume for successful culture initiation
[23]. Large explants generally survive more frequently and
grow more rapidly than relatively small explants at the initial
stage of culture [24]. Inoculation density was found to affect
the activities of individual enzymes in suspended plant cell
cultures for the synthesis of secondary metabolites [25]. The
higher final cell mass could be achieved in the cell culture
with higher inoculum sizes. A similar phenomenon was also
observed in the cell growth of Panax ginseng cultures [26].
Wu et al. [27] also reported that the final weight of root
biomass was low when less dense inoculum sources were
used, but the development was greatly enhanced with
inoculum sizes of 10 to 20 g/l. Previous research studies stated
that the cell growth kinetics and secondary metabolite
formation in plant cell cultures were greatly affected by the
inoculum size. For example, Sakurai and co-researchers [28]
reported that the anthocyanin productivity in strawberry
suspension culture increased in proportion to the inoculum
size up to 0.25 g 25 mL-1
, whereas the maximum cell
concentration was achieved at 0.5 g 25 mL-1
. The stimulatory
effects of inoculum size on the secondary metabolite
production were also observed in ginsenoside accumulation in
ginseng suspension culture [29] as well as ajmalicine and
catharanthine production in immobilized Catharanthus roseus
cell cultures [30].
Notably, C. asiatica cell suspension can be maintained in a
good condition when the medium pH is between 5.0 and 6.0,
but the concentration of hydrogen ions in the medium changes
during the development period, decreases during ammonia
assimilation and increases during nitrate uptake [27]. Initial
medium pH range of 5.0 to 6.0 was best in promoting the
growth of Echinacea roots and their accumulation of phenols
and flavonoids. However, the growth was inhibited at the
initial pH lower than 5.0 or higher than 6.0 [27]. In addition,
Shu and Lung [31] also stated that a higher culture pH might
facilitate the transport of methanol-soluble metabolites from
mycelia of Antrodia camphorate into the medium. The
formation of reddish culture during high pH regime might be
due to the presence of high content of anthocyanin, which was
the end product in the flavonoid pathway. The intermediate
flavonoid might have been secreted out into the medium for
anthocyanin formation which caused lower flavonoid content
in the cells. This showed that flavonoid in this culture has the
tendency to be secreted out into the medium instead of retain
in the cell. Hence, the metabolites presented in cells were
relatively lower as compared to the media.
Light is thought to have effects on enzyme induction and
activation [32]. The increment of flavonoid production in
response to light in this study is similar with previous
researchers who also reported that the secondary metabolites
were induced by light in cell or tissue cultures of several
species, such as Rudbeckia hirta L. [33] and Vitis vinifera
[34]. It was reported that light enhanced the production
involving the protection function of the plants against UV
damage.
Notably, cytokinins are known to enhance the production of
secondary metabolites and play an important role in
cotydifferentiation [35]. However, externally added kinetin
inhibited both salidroside accumulation and growth of CCA
suspension culture [21]. In addition, Pasqua and co-workors
[36] also reported that high concentrations of cytokinins
reduced the anthocyanin production. On the other hand, with
regard to auxins, the highest anthocyanin production was
obtained in the presence of low auxin concentration, which
contradicted with the finding of this present study.
Nevertheless, the current result of high auxin concentration
that enhanced the secondary metabolites production was found
to be consistent with the studies carried out on Tinospora
cordifolia Miers [22].
CONCLUSIONS
Cultivation of cell suspension is an efficient method for
producing useful phytomolecules. We found that there is a
positive relationship between the flavonoid production and the
cell aggregate size. A bigger size of cell aggregation with a
high inoculum density is attributed as a useful finding
flavonoid production either intercellularly or extracellularly.
In this study, in vitro conditions strongly affected the cell
growth and accumulation of flavonoid content in C. asiatica
tissues. The best performance was obtained in the MS medium
supplemented with 3.0 mg L-1 2,4-D and 1 mg L-1 kinetin ,
3% (w/v) sucrose, pH and the inoculum size of 0.5g wet cells
25 mL-1 at 250-500 µm aggregate size in the presence of light
or without light. This particular culture protocol will prove
beneficial for large-scale biomass production and secondary
metabolites of C. asiatica.
ACKNOWLEDGEMENTS
One of the author, SH Tan, would like to thank Universiti
Malaysia Pahang RDU120332 for financial support.
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