Search results for: Avinash H. Anantharam

Authors: Snehal D. Ganjave, Hardik Dodia, Avinash V. Sunder, Swati Madhu, Pramod P. Wangikar

Abstract:

Batch cultivation of recombinant bacteria in shake flasks results in low cell density due to nutrient depletion. Previous protocols on high cell density cultivation in shake flasks have relied mainly on controlled release mechanisms and extended cultivation protocols. In the present work, we report an optimized fed-batch process for high cell density cultivation of recombinant E. coli BL21(DE3) for protein production. A cybernetic model-based, multi-objective optimization strategy was implemented to obtain the optimum operating variables to achieve maximum biomass and minimized substrate feed rate. A syringe pump was used to feed a mixture of glycerol and yeast extract into the shake flask. Preliminary experiments were conducted with online monitoring of dissolved oxygen (DO) and offline measurements of biomass and glycerol to estimate the model parameters. Multi-objective optimization was performed to obtain the pareto front surface. The selected optimized recipe was tested for a range of proteins that show different extent soluble expression in E. coli. These included eYFP and LkADH, which are largely expressed in soluble fractions, CbFDH and GcanADH , which are partially soluble, and human PDGF, which forms inclusion bodies. The biomass concentrations achieved in 24 h were in the range 19.9-21.5 g/L, while the model predicted value was 19.44 g/L. The process was successfully reproduced in a standard laboratory shake flask without online monitoring of DO and pH. The optimized fed-batch process showed significant improvement in both the biomass and protein production of the tested recombinant proteins compared to batch cultivation. The proposed process will have significant implications in the routine cultivation of E. coli for various applications.

Keywords: cybernetic model, E. coli, high cell density cultivation, multi-objective optimization

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Authors: Singh Deepeshwar, N. K. Manjunath, M. Avinash

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Meditation is a self-regulated conscious process associated with improved awareness, perception, attention and overall performance. Different traditional origin of meditation technique may have different effects on autonomic activity and brain functions. Based on this quest, the present study evaluated the effect of Tai-Chi Chuan (TCC, a Chines movement based meditation technique) and Cyclic Meditation (CM, an Indian traditional based stimulation and relaxation meditation technique) on the hemodynamic responses of the prefrontal cortex (PFC) and autonomic functions (such as R-R interval of heart rate variability and respiration). These two meditation practices were compared with simple walking. Employing 64 channel near infrared spectroscopy (NIRS), we measured hemoglobin concentration change (i.e., Oxyhemoglobin [ΔHbO], Deoxyhemoglobin [ΔHbR] and Total hemoglobin change [ΔTHC]) in the bilateral PFC before and after TCC, CM and Walking in young college students (n=25; average mean age ± SD; 23.4 ± 3.1 years). We observed the left PFC activity predominantly modulates sympathetic activity effects during the Tai-Chi whereas CM showed changes on right PFC with vagal dominance. However, the changes in oxyhemoglobin and total blood volume change after Tai-Chi was significant higher (p < 0.05, spam t-maps) on the left hemisphere, whereas after CM, there was a significant increase in oxyhemoglobin (p < 0.01) with a decrease in deoxyhemoglobin (p < 0.05) on right PFC. The normal walking showed decrease in Oxyhemoglobin with an increase in deoxyhemoglobin on left PFC. The autonomic functions result showed a significant increase in RR- interval (p < 0.05) along with significant reductions in HR (p < 0.05) in CM, whereas Tai-chi session showed significant increase in HR (p < 0.05) when compared to walking session. Within a group analysis showed a significant reduction in RR-I and significant increase in HR both in Tai-chi and walking sessions. The CM showed there were a significant improvement in the RR - interval of HRV (p < 0.01) with the reduction of heart rate and breath rate (p < 0.05). The result suggested that Tai-Chi and CM both have a positive effect on left and right prefrontal cortex and increase sympathovagal balance (alertful rest) in autonomic nervous system activity.

Keywords: brain, hemodynamic responses, yoga, meditation, Tai-Chi Chuan (TCC), walking, heart rate variability (HRV)

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Authors: Amlan Kumar Das, Avinash Marwal, Vikram Pareek

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Liposome plays an important role in medical and pharmaceutical science as e.g. nano scale drug carriers. Liposomes are vesicles of varying size consisting of a spherical lipid bilayer and an aqueous inner compartment. Magnet-driven liposome used for the targeted delivery of drugs to organs and tissues1. These liposome preparations contain encapsulated drug components and finely dispersed magnetic particles. Liposomes are vesicles of varying size consisting of a spherical lipid bilayer and an aqueous inner compartment that are generated in vitro. These are useful in terms of biocompatibility, biodegradability, and low toxicity, and can control biodistribution by changing the size, lipid composition, and physical characteristics2. Furthermore, liposomes can entrap both hydrophobic and hydrophilic drugs and are able to continuously release the entrapped substrate, thus being useful drug carriers. Magnetic liposomes (MLs) are phospholipid vesicles that encapsulate magneticor paramagnetic nanoparticles. They are applied as contrast agents for magnetic resonance imaging (MRI)3. The biological synthesis of nanoparticles using plant extracts plays an important role in the field of nanotechnology4. Green-synthesized magnetite nanoparticles-protein hybrid has been produced by treating Iron (III)/Iron(II) chloride with the leaf extract of Dhatura Inoxia. The phytochemicals present in the leaf extracts act as a reducing as well stabilizing agents preventing agglomeration, which include flavonoids, phenolic compounds, cardiac glycosides, proteins and sugars. The magnetite nanoparticles-protein hybrid has been trapped inside the aqueous core of the liposome prepared by reversed phase evaporation (REV) method using oleic and linoleic acid which has been shown to be driven under magnetic field confirming the formation magnetic liposome (ML). Chemical characterization of stealth magnetic liposome has been performed by breaking the liposome and release of magnetic nanoparticles. The presence iron has been confirmed by colour complex formation with KSCN and UV-Vis study using spectrophotometer Cary 60, Agilent. This magnet driven liposome using nanoparticles-protein hybrid can be a smart vesicles for the targeted drug delivery.

Keywords: nanoparticles-protein hybrid, magnetic liposome, medical, pharmaceutical science

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Authors: Sujata Sharma, Avinash Singh, Lovely Gautam, Pradeep Sharma, Mau Sinha, Asha Bhushan, Punit Kaur, Tej P. Singh

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Peptidyl-tRNA hydrolase (Pth) Pth is an essential bacterial enzyme that catalyzes the release of free tRNA and peptide moeities from peptidyl tRNAs during stalling of protein synthesis. In order to design inhibitors of Pth from Pseudomonas aeruginosa (PaPth), we have determined the structures of PaPth in its native state and in the bound states with two compounds, amino acylate-tRNA analogue (AAtA) and 5-azacytidine (AZAC). The peptidyl-tRNA hydrolase gene from Pseudomonas aeruginosa was amplified by Phusion High-Fidelity DNA Polymerase using forward and reverse primers, respectively. The E. coliBL21 (λDE3) strain was used for expression of the recombinant peptidyl-tRNA hydrolase from Pseudomonas aeruginosa. The protein was purified using a Ni-NTA superflow column. The crystallization experiments were carried out using hanging drop vapour diffusion method. The crystals diffracted to 1.50 Å resolution. The data were processed using HKL-2000. The polypeptide chain of PaPth consists of 194 amino acid residues from Met1 to Ala194. The centrally located β-structure is surrounded by α-helices from all sides except the side that has entrance to the substrate binding site. The structures of the complexes of PaPth with AAtA and AZAC showed the ligands bound to PaPth in the substrate binding cleft and interacted with protein atoms extensively. The residues that formed intermolecular hydrogen bonds with the atoms of AAtA included Asn12, His22, Asn70, Gly113, Asn116, Ser148, and Glu161 of the symmetry related molecule. The amino acids that were involved in hydrogen bonded interactions in case of AZAC included, His22, Gly113, Asn116, and Ser148. As indicated by fittings of two ligands and the number of interactions made by them with protein atoms, AAtA appears to be a more compatible with the structure of the substrate binding cleft. However, there is a further scope to achieve a better stacking than that of O-tyrosyl moiety because it is not still ideally stacked. These observations about the interactions between the protein and ligands have provided the information about the mode of binding of ligands, nature and number of interactions. This information may be useful for the design of tight inhibitors of Pth enzymes.

Keywords: peptidyl tRNA hydrolase, Acinetobacter baumannii, Pth enzymes, O-tyrosyl

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Authors: Rezaul K. Sk., Mohammad Ashiq, Avinash K. Srivastava

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The use of radiation shielding technology ranging from EMI to high energy gamma rays in various areas such as devices, medical science, defense, nuclear power plants, medical diagnostics etc. is increasing all over the world. However, exposure to different radiations such as X-ray, gamma ray, neutrons and EMI above the permissible limits is harmful to living beings, the environment and sensitive laboratory equipment. In order to solve this problem, there is a need to develop effective radiation shielding materials. Conventionally, lead and lead-based materials are used in making shielding materials, as lead is cheap, dense and provides very effective shielding to radiation. However, the problem associated with the use of lead is its toxic nature and carcinogenic. So, to overcome these drawbacks, there is a great need for lead-free radiation shielding materials and that should also be economically sustainable. Therefore, it is necessary to look for the synthesis of radiation-shielding glass by using other heavy metal oxides (HMO) instead of lead. The lead-free BaO-doped TeZnCaB glass systems have been synthesized by the traditional melt-quenching method. X-ray diffraction analysis confirmed the glassy nature of the synthesized samples. The densities of the developed glass samples were increased by doping the BaO concentration, ranging from 4.292 to 4.725 g/cm3. The vibrational and bending modes of the BaO-doped glass samples were analyzed by Raman spectroscopy, and FTIR (Fourier-transform infrared spectroscopy) was performed to study the functional group present in the samples. UV-visible characterization revealed the significance of optical parameters such as Urbach’s energy, refractive index and optical energy band gap. The indirect and direct energy band gaps were decreased with the BaO concentration whereas the refractive index was increased. X-ray attenuation measurements were performed to determine the radiation shielding parameters such as linear attenuation coefficient (LAC), mass attenuation coefficient (MAC), half value layer (HVL), tenth value layer (TVL), mean free path (MFP), attenuation factor (Att%) and lead equivalent thickness of the lead-free BaO-doped TeZnCaB glass system. It was observed that the radiation shielding characteristics were enhanced with the addition of BaO content in the TeZnCaB glass samples. The glass samples with higher contents of BaO have the best attenuation performance. So, it could be concluded that the addition of BaO into TeZnCaB glass samples is a significant technique to improve the radiation shielding performance of the glass samples. The best lead equivalent thickness was 2.626 mm, and these glasses could be good materials for medical diagnostics applications.

Keywords: heavy metal oxides, lead-free, melt-quenching method, x-ray attenuation

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Authors: Jonardan Koner, Avinash Purandare

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In today’s globalized world international business is a very key area for the country's growth. Some of the strategic areas for holding up a country’s international business to grow are in the areas of connecting Ports, Road Network, and Rail Network. India’s International Business is booming both in Exports as well as Imports. Ports play a very central part in the growth of international trade and ensuring competitive ports is of critical importance. India has a long coastline which is a big asset for the country as it has given the opportunity for development of a large number of major and minor ports which will contribute to the maritime trades’ development. The National Economic Development of India requires a well-functioning seaport system. To know the comparative strength of Indian ports over South-east Asian similar ports, the study is considering the objectives of (I) to identify the key parameters of an international mega container port, (II) to compare the five selected container ports (JNPT, Chennai, Singapore, Dubai, and Colombo Ports) according to user of the ports and iii) to measure the growth of selected five container ports’ throughput over time and their comparison. The study is based on both primary and secondary databases. The linear time trend analysis is done to show the trend in quantum of exports, imports and total goods/services handled by individual ports over the years. The comparative trend analysis is done for the selected five ports of cargo traffic handled in terms of Tonnage (weight) and number of containers (TEU’s). The comparative trend analysis is done between containerized and non-containerized cargo traffic in the five selected five ports. The primary data analysis is done comprising of comparative analysis of factor ratings through bar diagrams, statistical inference of factor ratings for the selected five ports, consolidated comparative line charts of factor rating for the selected five ports, consolidated comparative bar charts of factor ratings of the selected five ports and the distribution of ratings (frequency terms). The linear regression model is used to forecast the container capacities required for JNPT Port and Chennai Port by the year 2030. Multiple regression analysis is carried out to measure the impact of selected 34 explanatory variables on the ‘Overall Performance of the Port’ for each of the selected five ports. The research outcome is of high significance to the stakeholders of Indian container handling ports. Indian container port of JNPT and Chennai are benchmarked against international ports such as Singapore, Dubai, and Colombo Ports which are the competing ports in the neighbouring region. The study has analysed the feedback ratings for the selected 35 factors regarding physical infrastructure and services rendered to the port users. This feedback would provide valuable data for carrying out improvements in the facilities provided to the port users. These installations would help the ports’ users to carry out their work in more efficient manner.

Keywords: throughput, twenty equivalent units, TEUs, cargo traffic, shipping lines, freight forwarders

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