Open Access Repository of Indian Theses

Mitra, Puja

Study of size-based isolation and detection of circulating tumour cells (cancer cells) using micro-
fluidics and image processing 
 
In recent years, micro-fluidics has offered an immense contribution in the area of circulating tumour 
cells (cancer cells) isolation from whole blood. The conventional micro-fluidic approaches utilize 
expensive and sophisticated instruments to fabricate micro-channels with precise geometry. At this 
juncture, a cost-effective and user-friendly fabrication method is required. In this context, a spiral 
micro-fluidic device has been developed, in this research, via laser-based machining. As an initial 
attempt, two different sizes (25μm and 10μm) of polystyrene micro-particles (by mimicking the size 
or dimensions of cancer cells and healthy blood cells) have been used for size-based isolation purpose. 
Inside the fabricated spiral micro-channel, the development of Dean vortices or counter-rotating 
vortices due to centrifugal forces not only has helped in isolation but also has reduced the effect of 
clogging. A 3D simulation study has been performed to understand the flow behaviour inside the 
developed spiral microchannel.  
 
Post isolation analysis of circulating tumour cells (cancer cells) is also necessary as cancer cells are rare 
in peripheral blood. Image processing, in this regard, has become popular to analyse and identify 
cancer cells with the advancement of computing hardware and the machine vision algorithms, 
recently. Generally, the CTCs have been identified from stained blood sample images, where staining 
is costly and operator dependent technique. However, classification and detection of cancer cells from 
unstained blood from its microscopic images is still challenging. Therefore, in this study, an attempt 
has been made to classify and identify CTCs (cancer cells) using image processing. At first, classification 
between unstained healthy blood samples and blood samples spiked with cancer cells has been 
accomplished from their microscopic images. Image pre-processing based on unsharp masking and 
contrast limited adaptive histogram equalization (CLAHE) to enhance the cell boundaries has been 
performed. Then statistical texture analyses based on gray level co-occurrence matrix (GLCM) and run 
length statistics (RLS) have been used on pre-processed images for features extraction. Operating 
parameters of the texture analyses are adaptively selected for making this more operator 
independent. Fisher Discriminant Ratio (FDR) based feature selection has been used for overcoming 
rigorous manual feature selection, where eight features have been selected out of twenty extracted 
features. Then the eight features are fed to train k-nearest neighbour (k-NN), support vector machine 
(SVM), Naïve-Bayes classification (NBC) and LogitBoost classification models where 60% data has been 
utilized for training and remaining for testing purpose. These four machine learning methods have 
been compared on the testing data based on performance metrics i.e., accuracy, precision, recall and 
F1-score. SVM and LogitBoost classifier achieved the best classification accuracy. In the next step, 
detection of cancer cells has been performed on images classified as spiked blood sample or blood 
samples spiked with cancer cells, where edge detection based on ant colony optimization (ACO) has 
been used to identify the cell boundaries. Finally, circular hough transform (CHT) has been applied to 
detect cancer cells. 


English

Study of size-based isolation and detection of circulating tumour cells (cancer cells) using microfluidics and image processing

Srivastava, Shikha

Investigating the Role of Signal Transducer and Activator of Transcription 3 (STAT3) in Dengue 
Virus Propagation  
 
 
Dengue fever is an arthropod-borne viral infection affecting millions of people annually during the 
breeding season of its vector mosquito. To date, no specific antiviral drug or vaccination is invented 
to fight against dengue virus. Since viruses depend on host factors for their propagation, recent 
approaches to developing antivirals include targeting host factors for inhibiting viral propagation. 
DENV has developed several co-existing strategies to avoid eradication by the host antiviral 
mechanism. DENV modulates host interferon response by attenuating its components, inhibiting 
their activation or promoting their degradation. This dissertation summarizes the studies we carried 
out to identify the role of a host transcription factor; STAT3 in dengue virus propagation to identify 
its potential as a suitable antiviral target. We first wanted to understand how DENV manipulates 
STAT3 induced interferon response to its benefit. By utilizing various cell culture techniques, we 
found that STAT3 is increased and activated in DV-2 infected A549 cells and knocking it out 
resulted in a substantial decrease in viral protein production and viral replication. Our results also 
demonstrated that DV-2 purposefully manipulates STAT3, which negatively regulates type I IFN 
signaling, to avoid host interferon response. These results thus established the role of STAT3 as a 
proviral factor. Moving ahead, we explored the potential of STAT3 inhibitor stattic as an antiviral 
agent. For this we performed a whole proteome analysis of infected cells by high throughput liquid 
chromatography mass spectrometry (LC-MS), wherein we observed that treatment with stattic 
induces multiple host defense mechanisms to counteract dengue infection. We also observed that 
treatment with stattic downregulates several pathways involved in viral transcription. Taken 
together these results suggest that inhibiting STAT3 by stattic or its derivatives may prove to be an 
effective strategy for the development of antiviral interventions.  
Furthermore, to identify residues of STAT3 for the development of novel therapeutics we explored 
its interaction with some of the viral proteins translocating into the nucleus e.g., capsid and NS5 
we identified some critical residues which mediate the interaction of STAT3 with viral NS5 protein. 
Since both STAT3 and NS5 are important antagonists of human innate immune response we 
propose that targeting the residues which mediate their interaction may aid in the screening of 
alternative antivirals against dengue. Overall, this study strongly indicates the regulatory role of a 
host transcription factor STAT3 in modulating viral propagation that may be targeted for the 
development of effective antiviral therapeutics.  
 


Investigating The Role of Signal Transducer and Activator of Transcription 3 (STAT3) in Dengue Virus Propagation

Roychowdhury, Prasun

Study of Poincaré-Hardy type inequalities and eigenvalue problems for 
second-order elliptic PDEs 
 
The major text of this thesis is studying Poincaré-Hardy and Hardy-Rellich 
type inequalities on one of the most discussed Cartan-Hadamard manifold 
namely hyperbolic space and studying eigenvalue problems for second-order 
elliptic PDEs. The thesis is divided into two parts. In the first part we have 
centralized our attention on the following three problems: 
 
• On some strong Poincaré inequalities on Riemannian models and their 
improvements. 
• On higher order Poincaré inequalities with radial derivatives and Hardy 
improvements on the hyperbolic space. 
• Hardy-Rellich and second order Poincaré identities on the hyperbolic 
space via Bessel pairs. 
In the second part we have focused our essence on the following two 
problems: 
• Generalized principal eigenvalues of convex nonlinear elliptic operators 
in RN. 
• On ergodic control problem for viscous Hamilton-Jacobi equations for 
weakly coupled elliptic systems. 


Study of Poincaré-Hardy type inequalities and eigenvalue problems for second-order elliptic PDEs

Kamei, K. Rana

	
	
98 
 
 
CHAPTER 6 
 
SUMMARY 
 
6. SUMMARY: 
Lectins are proteins or glycoproteins which can reversibly bind to specific sugar 
moiety present in glycoconjugates. They are ubiquitous in nature and found in 
all organisms ranging from viruses, bacteria, plants, and animals. They play 
significant roles at the cellular and molecular level for many biological 
recognition processes including carbohydrates, and proteins. Lectins are also 
responsible for mediating the attachment and binding of bacteria, viruses, and 
fungi to their intended targets or host cells. Accumulating literature reports 
suggest that Legumes are the most studied and rich source of lectins. A large 
number of legume lectins have been isolated from various sources and are well 
characterized for their specificity and biological activities. Because of their 
diverse roles and applications, there is an expanding emphasis on the isolation, 
purification, and characterization of lectins from different sources.  
In the present study, a lectin was isolated and purified from the seeds of 
a leguminous plant Meizotropis buteiformis collected from in and around Chini-
inkhol Village, Kangpokpi District, Manipur, India. The main aim of this study 
is to purify and characterize the lectin for its biochemical and physical properties. 
The lectin was assayed by the hemagglutination assay method of Devi et al., 
2009 involving a standard serial dilution procedure using 2% suspension (v/v) 
	
	
99 
of rabbit RBC. Lectin activity was taken as hemagglutination titre, the reciprocal 
of the highest dilution of lectin sample at which complete hemagglutination 
occurred and it was expressed in terms of hemagglutination unit (HAU), that is 
the minimum amount of lectin required for complete agglutination under the 
standard assay condition. Protein was estimated following the method of Lowry 
et al., 1951 using crystalline Bovine serum albumin as standard. Carbohydrate 
specificity of the lectin activity was determined by hemagglutination inhibition 
assay taking various carbohydrates and glycoproteins following a slight 
modification of the method of Sawhney et al., 1996. 
The lectin was purified from the crude lectin preparation by a procedure 
involving successive steps of ammonium sulphate fractionation, and affinity 
chromatography. The crude lectin preparation was found to have a typical 
specific activity of 729.6HAU/mg protein. To undergo further purification steps, 
the crude lectin preparation was subjected to 50-80% ammonium sulphate 
fractionation, and its corresponding specific activity was found to be 2402.8 
HAU/mg with 3.29 purification fold and 70.59% recovery. Among the sugars 
and sugar derivatives tested for preliminary sugar specificity of lectin, 
disaccharide Lactose was found to be the most potent inhibitory sugar followed 
by a monosaccharide D-Galactose. So, the partially purified ammonium sulphate 
fraction was subjected after dialysis to affinity chromatography of lactose-
sepharose 4B column. Then the bound lectin was eluted using 10mM Lactose in 
PBS solution. The affinity eluted lection fractions were pooled and found to have 
a specific activity of 360658.8HAU/mg and a percentage recovery of 35.22% 
with 494.32 fold purification when compared to the crude preparation. The 
electrophoretic homogeneity of the affinity eluted lectin was determined by 
acidic native PAGE (pH 4.3) which showed a single band under native 
conditions. 
 Following the successful purification, the lectin was characterized with 
respect to certain physicochemical properties. The native molecular weight of 
the purified lectin was determined by gel filtration using Sephadex G-75 column 
	
	
100 
and found to be 75kDa. This native molecular weight of the lectin compares well 
with many of the leguminous plant lectins reported in the literature. The subunits 
molecular weights of the lectin, determined by SDS-PAGE both in reduced and 
non-reduced condition coupled with silver staining, were found to be 36kDa and 
38kDa which are designated as subunit A and B respectively. Combining the 
result of native molecular weight determination with that of the subunit 
molecular weight determination by SDS-PAGE, it is inferred that the purified 
lectin is a heterodimeric protein made up of two non-identical subunits (subunit 
A and B) linked non-covalently without a disulfide bond. These subunit 
structures are also in agreement with many of the earlier reports on the subunit 
structure of leguminous lectins. Like other leguminous lectins, the purified lectin 
in the present investigation is strongly inhibited by disaccharide Lactose 
followed by monosaccharide D-Galactose. Though many other leguminous 
lectins required divalent metal ions for their activity, the activity of the lectin 
purified from M. buteiformis seeds does not show any effect either in the 
presence or absence of divalent metal ions (Ca2+, Mg2+, Zn2+, Mn2+, and Sn2) and 
also does not require the given divalent metal ions. Similar results of non-
requirement of the metal ions by other lectins are also reported in previous works 
and compared well with the purified lectin.  
The pH optima of the lectin activity were found to be relatively broad 
ranging from 6.0 to 8.0 which compares well with the pH optima of other 
leguminous lectins. The lectin was completely stable from pH 6.5 to 7.5 which 
is in conformity with those lectins from the leguminous family. Full activity of 
the lectin was observed at the assay temperature from 300C to 500C. A decrease 
in lectin activity was observed below 300C and above 500C as well. No activity 
was observed at 800C and above. With respect to thermal stability, the purified 
lectin subjected to 10mins incubation in PBS exhibited complete stability from 
00C to 500C with a t1/2 value of 610C. These observed effects of assay 
temperature and thermal stability are comparable to or in conformity with 
literature reports on other leguminous lectins. Moreover, the M. buteiformis 
	
	
101 
lectin does not show blood group specificity as it agglutinates well with all RBC 
belonging to human blood group A, B, AB, and O, with the O group being 16 
times more specific than either A or B group. However, the observed 
hemagglutination activity of the lectin towards human blood groups A, B, or AB, 
and O was 32 and 2 times lesser respectively than that of observed lectin activity 
towards rabbit RBC. These results are comparable with those of many other 
lectins reported in the literature. The purified M. buteiformis lectin is a 
glycoprotein in nature and the total neutral carbohydrate content of the lectin 
was found to be 11.4% (w/w) of the total weight of the lectin. These results 
compare well with other reported leguminous lectins in the literature. The 
purified lectin does not show any antibacterial activity against the tested three 
bacteria namely E-coli, Bacillus subtillis, Micrococcus luteus. In contrast, some 
other leguminous lectins reported were found to show anti-microbial properties 
against many bacteria. 
Based on the results of peptide mass fingerprinting using Liquid 
chromatography tandem mass spectrometry (LC-MS/MS) and subsequent 
analysis by Mascot Sequence matching software (Matrix Science), no match was 
found with any of the known proteins in the database. However, from the protein 
view report for the matched protein, it was found that M. buteiformis lectin 
showed only upto 50% protein sequence coverage against β-chain of Butea 
monosperma lectin (ANNEXURE I). From this result, it was concluded that the 
lectin, purified from the seeds of Meizotropis buteiformis is a novel lectin which 
is not reported and characterized earlier. For the future work plan, more 
fundamental and functional characteristics of this lectin including three-
dimensional structure, binding site structure, carbohydrate interaction kinetics, 
mitogenicity, cytotoxicity, etc. are yet to be determined. Research works may 
also be directed towards a better understanding of this newly purified lectin with 
respect to its physicochemical and functional characteristics. 
 


Purification and characterization of lectin from the seeds of Meizotropic Buteiformis

-, Raman

 
 Bioefficacy of Vermicompost on Meloidogyne incognita (Kofoid & White) 
Chitwood Infecting Tomato Plants 
 
Use of vermicompost is an alternative way to manage plant parasitic nematodes as well as to 
increase crop productivity. Therefore, the aim of the present study was to use an eco-friendly and 
inexpensive approach of applying vermicompost as a replacement for pesticides for enhancing the 
biomass and bioactive components along with mitigation of M incognita induced stress in tomato. 
Tomato seeds were pre-soaked in various dilutions of vermicompost extract before being planted 
in clay pots. Plants at the true leaf stage were transferred to separate pots having grading mlixtures 
of vermicompost and soil. The transplanted plants were infected with freshly hatched J2s (@ 2J2/g 
soil) and water was sprayed on a regular basis to keep the potting mixture wet. After 10 days of 
nematode infection, the plants were harvested. Several morphological characteristics were 
evaluated, including root and shoot length, fresh weights of roots and shoots, number of leaves, 
and number of galls on roots. Further, Biochemical parameters like Protein content, flavonoid 
content, phenol content, Anthocyanin, Total Glutathione content, proline content, MDA, H202 
content, photosynthetic pigments (Chlorophyll and Carotenoid), Antioxidant enzymes like CAT, 
APOX, GST were studied using standard protocols. The data were subjected to statistical analysis 
by using SPSS software version 16.0. Analysis of Variance (ANOVA) and Tukey's Test were 
employed to observations. 
 
Experimental data shows that all the morphological and biochemical parameters like Protein 
content, flavonoid content, phenol content, Anthocyanin, Total Glutathione content, proline 
content, photosynthetic pigments (Chlorophyll and Carotenoid), Antioxidant enzymes like CAT, 
APOX of tomato seedlings treated with vermicompost were significantly higher as compared to 
control. This increase was in concentration dependent manner. There was non-significant increase 
in antioxidant enzymes like DHAR, GST. Further, stress indices like MDA and H202 content were 
significantly reduced as compared to control. The current investigation demonstrates the possible 
impact of vermicompost in boosting tomato plant development. Protein content and anti-oxidative 
enzyme levels in tomato seedlings increased considerably after the administration of 
vermicompost. In addition to enzymes, non-enzymatic antioxidants such as ascorbic acid and 
glutathione concentration were increased in tomato plants fed with vermicompost. Vermicompost 
also aided in the reduction of MDA and H202 levels, both of which are considered plant stress 
signals. As a result of the foregoing observations, vermicompost appears to have a high potential 
for improving plant development under nematode stress. 
 


Bioefficacy of Vermicompost on Meloidogyne incognita (Kofoid & White) Chitwood Infecting Tomato Plants

Verma, Satish Kumar

i  
 
Abstract 
 
With the increasing concern on several global issue likes’ shortage in fossil fuel, 
climate change, environmental pollution, and sustainability has become one of the 
essential principles to meet the objectives of human being’s development. To ensure 
sustainable development, we have to rely on clean, green, and renewable energy 
resources such as solar, wind, tidal, geothermal, hydrogen, etc. Among these 
resources, hydrogen is the most abundant element in the universe and a potent energy 
carrier due to its clean, renewable, indigenous, and higher energy density (33.33 
kWh/kg) nature. However, despite all such merits, a flawless economic system based 
upon H2 fuel (hydrogen economy) could not be readily framed due to technical 
challenges in hydrogen production, hydrogen storage, nd its onboard applications. 
Out of these challenges, hydrogen storage presents a significant challenge due to its 
very low density (0.08988 g/L). To increase hydrogen density, work must either be 
applied to compress the gas, the temperature decreased below the critical temperature 
or the repulsion reduced by the interaction of hydrogen with another material. All 
these processes are cost-intensive and risky; thus,e main concern regarding the 
onboard application of hydrogen energy is the development of safe and efficient solid-
state hydrogen storage materials. In order to have saf and efficient solid-state 
hydrogen storage materials, the United States Department of Energy (US-DOE) has 
set some targets for onboard hydrogen storage particular for vehicular purpose. These 
targets include the system gravimetric density needs to be 5.5 wt% (1.8 kWh/kg), 
system volumetric density 1.3 kWh/L (0.04 kg/ L), system fill time (5 kg) 3.3 min, 
operational cycle life more than 1500 cycles, and the operating ambient temperature 
below 60 ºC. Unfortunately, we could not develop such materials that meet these 
requirements so far. 
 MgH2 is one of the most attractive options as a solid-state hydrogen storage 
material owing to its high hydrogen storage capacity ~ 7.60 wt% (volumetric capacity 
of 110 g/L). Other factors which make MgH2 as one of the promising materials for 
hydrogen storage are high abundance of Mg in earth crust (6th abundant), light weight 
Abstract 
 
ii   
 
etc.,. However, the pristine MgH2 has a high operating temperature ≥ 400 °C and slow 
hydrogen ab/de-sorption kinetics (< 1 wt%/min). Besid  MgH2, the complex hydride 
NaAlH4 is also one of the solid-state hydrogen storage materials with high hydrogen 
density and manageable thermodynamics and fulfills the proposed US-DOE target for 
an onboard hydrogen storage system. However, the rev rsibility (hydrogen ab/de-
sorption during cycling) and dehydrogenation kinetics of NaAlH4 is quite sluggish 
below its melting point i.e., 183 oC. In this thesis, the main emphasis is given on 
solving the various issues related with MgH2 and NaAlH4 as hydrogen storage 
materials i.e., improving thermodynamics and kinetics of MgH2 and NaAlH4.  Finally, 
the studies on hydrogen storage has been extended to the system which can be a 
viable hydrogen storage option especially for space vehicle application i.e. Metal-
organic frameworks (MOF) i.e., M2(m-dobdc) MOFs (M= Fe and Mn) (m-dobdc
4- = 
4,6-dioxido-1,3-benzene-di-carboxylate). We have also studied these MOF materials 
as electrode materials for supercapacitor application. The thesis embodies six chapters 
which are briefly described and discussed in the following: 
 The first chapter discusses the background of hydrogen energy and the 
principle of hydrogen storage in various storage modes with special emphasis on 
metal hydrides. The principle and mechanism of hydrogen storage in metal hydrides 
has been discussed. The limitations of conventional alloys/intermetallics for reversible 
hydrogen storage are pointed out. It also discusses the hydrogen storage behavior of 
light weight hydrides, especially, MgH2, NaAlH4, porous nanomaterials like MOF and 
some new materials with improved hydrogen storage cpa ity. 
 The second chapter describes the experimental and theoretical 
techniques/methods employed in the thesis. These techniques include ball-milling, 
glove box etc., X-ray diffraction (XRD) technique has been employed for gross 
structural characterization. In addition, Scanning electron microscopy (SEM) using 
secondary electron imaging mode, Transmission electron microscopy (TEM), high 
resolution electron microscopy (HR-TEM), X-ray photoelectron spectroscopy (XPS) 
and EDAX have been used for microstructural and compositional characterization. 
The temperature based Raman, Fourier transforms infrared spectroscopy (FT-IR), and 
Nuclear magnetic resonance (NMR) spectroscopy has been used for spectroscopic 
Abstract 
 
iii   
 
analysis. Hydrogen storage measurements were carried out using an automated one 
and two channel Gas Reaction Controller (GRC) operated by a Lab-View based 
program. 
 The third chapter investigated the comparative study of graphene, Vanadium 
disulfide (VS2), Tungsten disulfide (WS2), and Molybdenum disulfide (MoS2) as a 
catalyst for improving the hydrogen sorption properties of MgH2. Based on the de/re-
hydrogenation study; it is found that VS2 works as an optimum catalyst over 
graphene, MoS2, and WS2 for MgH2. The MgH2-VS2 has the onset dehydrogenation 
temperature of ~ 242 
o
C, which is ~ 134 
o
C than the desorption temperature of pristine 
MgH2 and ~88 
o
C, ~58 
o
C, ~35 
o
C than MgH2-MoS2, MgH2-Gr, MgH2-WS2, 
respectively. There is a slight degradation in the hydrogen storage capacity (0.2 wt%) 
even after 25th cycles. However, the catalysts VS2 graphene, MoS2, and WS2 do not 
play a significant role to lower the thermodynamics of the MgH2. The hydrogen 
desorption activation energy barrier to converting M H2 into Mg in the presence of 
VS2 catalyst was found to be 98.09 kJ/mol, which is 36.86 kJ/mol, 19 kJ/mol, 5.91 
kJ/mol and ~ 42 kJ/mol less than from activation energies of MgH2-Gr, MgH2-MoS2, 
MgH2-WS2 and pristine MgH2 respectively. The studies in the present investigation 
indicate that VS2 is a better catalyst that replaces graphene and possibly other 2D 
materials (like MoS2, WS2, etc.,) as a catalyst for MgH2. 
 
 The fourth chapter reports the results of investigations of de/re-
hydrogenation in the Mg/MgH2-graphene templated Ti-based catalysts (Ti, TiO2, and 
TiH2) system. Based on de/re-hydrogenation study graphene t mplated TiH2 
(TiH2@Gr) was found to be the optimum catalyst as TiH2@Gr catalyzed MgH2 has 
the onset dehydrogenation temperature of ~204 
o
C which is 31 
o
C, 36 
o
C, and 66 
o
C 
lower than the dehydrogenation temperature of MgH2-Ti@Gr, MgH2-TiO2@Gr, and 
BM MgH2. Most importantly, the effect on the thermodynamics of MgH2 which has 
not been observed in case of VS2 catalyst has been seen for MgH2-TiH2@Gr, the 
change in enthalpy of MgH2-TiH2@Gr is ~6.12 kJ/mol lower than ball-BM MgH2. 
There is slight decrease (0.1 wt%) in hydrogen storage capacity of MgH2-TiH2@Gr 
up to 5 cycles and then capacity remains unchanged up to 25 cycles. The improved 
Abstract 
 
iv  
 
hydrogen sorption in MgH2-TiH2@Gr can be attributed to three effects namely 
catalytic effect of TiH2, TiH1.924 which forms due to interaction of TiH2 and graphene, 
co-catalytic effect of graphene and absence of agglomeration of TiH2 and TiH1.924 due 
to their anchoring on graphene. Graphene besides ceasing the agglomeration of TiH2 
also helps in electron transfer to and fro from TiHx to MgH2. The studies carried out 
in the present investigation indicate that further r search on graphene and possibly 
other two-dimensional (2D) material as a template for the existing catalyst may open 
a new avenue for catalyzing MgH2, so as to make it a viable hydrogen storage system 
for practical applications. 
 
 The fifth chapter investigates the effect of quasicrystal (QC) (neither 
amorphous nor crystalline) as catalyst on yet another highly efficient solid-state 
hydrogen storage material i.e., the complex metal hydride NaAlH4 system. The 
present study found leached ball-milled Al65Cu20Fe15 QC as a better catalyst for the 
NaAlH4. The leached ball-milledAl65Cu20Fe15 (LBMACF) catalyzed NaAlH4 sample 
has the lowest hydrogen desorption temperature of 140 °C as compared to NaAlH4 
catalyzed by other versions of QC as well as with uncatalyzed NaAlH4. NaAlH4-
LBMACF shows faster rehydrogenation kinetics by absor ing ~ 4.68 wt% in 10 
minutes at 130 ºC under 100 atm hydrogen pressure and promising cyclic stability up 
to 25 cycles. Based on XRD, electron microscopy, temp rature based Raman 
spectroscopy, temperature based NMR spectroscopy, and XPS analysis, it has been 
established that the weakening of Al-H in NaAlH4 is one of the main reasons for 
destabilizing NaAlH4. The destabilization of the Al-H bond is found greater in 
LBMACF due to a more multi-electronic state environment around NaAlH4 than 
other catalysts. Therefore, synthesizing multivalent QC-based catalysts seems to be a 
promising approach for destabilizing NaAlH4 and promotes the use of complex 
hydrides for onboard applications. 
 
 In chapters 3-5, we have discussed about the catalyzed and uncatalyzed 
version of metal hydrides like MgH2, NaAlH4 for hydrogen storage applications 
keeping the US-DOE target for hydrogen fuel cell vehicle in view. In chapter six we 
Abstract 
 
v  
 
have investigated porous nanomaterial Metal-organic frameworks (MOF) for 
hydrogen storage application in space vehicle. Here the hydrogen uptake and release 
temperatures and pressures are drastically different f om the need of US-DOE target. 
The hydrogen storage properties are usually evaluated  -196 °C (liquid (LQ) N2 
temperature) for space vehicle application. The hydrogen storage capacity of Mn2(m-
dobdc) MOF is determined to be the highest of 7.72 wt% at -196 °C (LQN2 
temperature) under 5-100 atm H2 pressure, which is 3.81 wt% higher than Fe2(m-
dobdc) MOF under identical temperature and pressure conditions. As a result, Mn2(m-
dobdc) out performs Fe2(m-dobdc) when it comes to gravimetric hydrogen storage 
capacity. The isosteric heats of adsorption and desorption for Mn2(m-dobdc) were 
also computed and found to be -4.83 kJ/mol and -4.68 kJ/mol, respectively. The 
isosteric heats of adsorption and desorption for Fe2(m-dobdc) are -15.89 kJ/mol and -
16.74 kJ/mol, respectively. On the other hand, the supercapacitance application for 
the Mn2(m-dobdc) and Fe2(m-dobdc) based electrode materials were evaluated. In the 
comparative study present in the current work indicates that Mn2(m-dobdc) based 
supercapacitor exhibited higher capacity (~25 F/g at 0.41A/g current density) as 
compared to Fe2(m-dobdc) based supercapacitor.  
 
 In conclusion, the thesis work has beensummarized, and their potential 
outcomes with future aspectshave beendiscussed. 
 
 
 
 
 
 
 


Studies of nanomaterials for hydrogen storage and other energy applications

Sharma, Rahul

Isolation, Purification, Structural Characterization and Assessment of Larvicidal Activity of Major N-alkylamides from Acmella ciliata HBK
In the present study, an attempt was made to develop methods for isolation and estimation of major N-alkylamides from A. ciliata that exhibit larvicidal activity (LA) against late III – early IV instar larvae of Aedes aegypti and Culex quinquefasciatus. The isolated compounds were 2E,4E-N-isobutyl undeca-2,4-diene-8,10-diynamide (NUD) and 2E,4E,8Z,10E-N-isobutyl dodeca-2,4,8,10-tetraenamide (NDT). LC50 of NUD and NDT against Ae. aegypti was 44.19 ppm and 18.28 ppm, and for Cx. quinquefasciatus these were 30.89 ppm and 11.75 ppm, respectively. NDT was found more toxic than NUD with both having a higher potency than the commercial larvicide methoprene (MET). HPLC of different parts of the plant showed that NAAs accumulate maximally in flowers as compared to other parts of the plant



Isolation, Purification, Structural Characterization and Assessment of Larvicidal Activity of Major N-alkylamides from Acmella ciliata HBK

CHAUDHARY, NIDHI

 
Deciphering the Mechanism of a Host Cellular Factor High Mobility Group Box1 Protein in 
Dengue Virus Pathogenesis  
 
Dengue disease is a highly prevalent mosquito-borne infection-causing millions of infections 
annually. The global rapid emergence and distribution of dengue in tropical and sub-tropical 
regions are exposing almost half of the world’s population to the threat. Dengue virus (DENV) 
causes the overexpression and secretion of many proinflammatory cytokines, including high-
mobility group box-1 protein (HMGB1). HMGB1 is the most abundant, extremely conserved, 
ubiquitously expressed nuclear protein that executes the localization-dependent function. 
HMGB1 translocation and secretion have been implicated in many chronic infections, 
metabolic disorders, cancer, and viral and inflammatory diseases. In this study, we explored 
the contribution of HMGB1 in dengue virus replication in A549 cells. Results showed that 
HMGB1 is translocated and secreted out during dengue infection. Moreover, blockage of 
HMGB1 release using ethyl pyruvate resulted in enhanced dengue replication and the 
knockdown of HMGB1 abolished viral replication. In-silico, in-vitro assays, and co-
immunoprecipitation revealed the binding of HMGB1 to both untranslated regions of viral 
RNA. This interaction further induces the expression of proinflammatory cytokines like TNF-
α, IL-6, and IL-1β which contributes to the severity of DENV disease. Therefore, our study 
suggests that the DENV tweaks HMGB1 translocation and its interaction with the DENV 
genome to stimulate proinflammatory cytokines expression in A549 cells.  
DENV exploits various cellular pathways including autophagy to assure enhanced virus 
propagation. The mechanisms of DENV-mediated control of the autophagy pathway are 
largely unknown. Our investigations have revealed a novel role for HMGB1 protein in the 
regulation of the cellular autophagy process in the DENV-2 infected A549 cell line. While 
induction of autophagy by rapamycin treatment resulted in enhanced DENV-2 propagation, 
the blockade of autophagy flux with bafilomycin A1 suppressed viral replication. Furthermore, 
siRNA-mediated silencing of HMGB1 significantly abrogated DENV-induced autophagy, while 
LPS-induced HMGB1 expression counteracted these effects. Interestingly, silencing of HMGB1 
showed a reduction of BECN1 and stabilization of BCL-2 protein. On the contrary, LPS 
induction of HMGB1 resulted in enhanced BECN1 and a reduction in BCL-2 levels. This study 
shows that the modulation of autophagy by DENV-2 in A549 cells is HMGB1/ BECN1 
dependent. In addition, glycyrrhizic acid (GA), a potent HMGB1 inhibitor suppressed 
autophagy as well as DENV-2 replication. Altogether, our data suggest that HMGB1 induces 
BECN1-dependent autophagy to promote DENV-2 replication. Therefore, we postulate 
HMGB1 as a crucial host element promoting viral propagation and must be considered as an 
alternative approach for targeting DENV infection. The complex pathogenesis of DENV 
infection can be better resolved by understanding the complicated correlation between the 
DENV virus and its host factors.  
Growing pieces of evidence demonstrate the importance of metabolic control on 
proinflammatory response, however, the regulatory mechanism is not known. Here, we 
suggest a novel mechanism for pyruvate kinase M2 (PKM2) in mediating HMGB1 release 
during DENV infection. Results showed that the PKM2 expression and nuclear localization 
were increased in the DENV-2 infection group. Inhibition of PKM2 nuclear translocation by 
DASA-58 and ML-265 results in decreased HMGB1 release in the supernatant. Furthermore, 
we observed that DASA-58 and ML-265 resulted in reduced viral protein expression and 
autophagy. Altogether, our results shed light on an important process for metabolic 
regulation of proinflammatory response by controlling HMGB1 release and emphasize the 
role of a metabolic enzyme in understanding the DENV disease pathogenesis.  
Collectively, we have deciphered the proviral mechanism of HMGB1 in DENV virus replication 
by its interaction with the viral genome to induce a proinflammatory response and inducing 
autophagy. Furthermore, we unraveled that release of HMGB1 during DENV infection is 
controlled metabolically by a glycolytic enzyme PKM2 expression and its nuclear 
translocation. Therefore, the molecular mechanisms associated with host factors that 
regulate inflammation must be further explored to understand the pathogenesis of severe 
DENV disease.  


DECIPHERING THE MECHANISM OF A HOST CELLULAR FACTOR HIGH MOBILITY GROUP BOX1 PROTEIN IN DENGUE VIRUS PATHOGENESIS

Chowdhury, Sayantan Nag

ABSTRACT
A complex network of coupled dynamical units has often been the object of active investi-
gation in a wide range of interdisciplinary researches ranging from physical, biological to
socio-technical and engineering systems. Diverse collective behaviors emanate in complex
systems of interacting dynamical entities not only because of the local systems’ unique at-
tributes but mainly because of the variability in the fashion of interactions among the units.
Both coupling topology and coupling function play quite decisive roles in the emergence
of the collective asymptotic state. However, most of the existing investigations on coupled
oscillators focus only on attractive pairwise interactions. Nevertheless, there exist vari-
ous realistic, relevant scenarios in several scientific, technological, and natural instances,
where competing types of couplings affect the collective behavior of coupled oscillator
ensembles. One of the primary goals of this thesis is to study the diverse collective phe-
nomena of an ensemble of coupled nonlinear oscillators under the coexistence of attractive
and repulsive interactions in the static as well as in time-varying networks.
While complete synchronization is the least expected outcome in a static network with
mixed positive-negative couplings, such an emergent state is still possible under the time-
varying network formalisms. We encounter diverse collective behaviors ranging from com-
plete and cluster synchronization to inhomogeneous small oscillation due to the interplay
between the attractive and repulsive coupling strengths. In the absence of repulsive cou-
pling, we successfully calculate the critical positive coupling strength required for acquir-
ing complete synchronization using the time-average Laplacian matrix. We also recognize
that suitable repulsive strength can make the error dynamics intermittent and leaves the
signature of largely deviated extreme events. The formal practice to differentiate extreme
events from other events is choosing a threshold T = m+dσ, where m is the sample mean
of the observable and σ is the corresponding standard deviation. d is generally chosen ran-
v
domly from the interval [4, 8]. We provide an analytical formulation to derive a threshold
HS = m+ 8σ that distinguishes extreme events from other intermittent states. We further
calculate an upper bound for the probability of occurrence of extreme events depending
on the choice of HS. Besides, we approximate the mean return interval of these extreme
events using time series analogs of Kac’s lemma.
In the case of static networks of coupled oscillators, we propose a unique algorithm
by identifying a suitable repulsive path in a connected graph for establishing zero frustra-
tion (antiphase states). Moreover, we provide an analytical understanding with the help
of graph theory to prove that antiphase synchronization is only possible in a connected
network if it is bipartite in nature. In addition, we formulate a universal 0 − π algorithm
for predicting the non-zero frustration value of arbitrary undirected non-bipartite graphs
of attractive-repulsively coupled limit cycle oscillators. Using this algorithm, one can eas-
ily construct a sparse non-bipartite network with desired frustration from a highly diluted
graph. We validate our findings in the single layer as well as multiplex networks. We
identify a mixed state of interlayer antisynchronization and intralayer synchronization in
a multiplex network when the interlinks’ strength between layers is solely negative. We
successfully calculate the amplitude of each Stuart-Landau oscillator, placed at each node
of a multiplex network, exhibiting this mixed state. This derived attractor size remains
independent of network size and agrees excellently with our numerical simulations for
negative interlayer coupling strength. Apart from deriving the necessary condition for
the existence of interlayer antisynchronization together with intralayer synchronization,
we analytically derive the invariance of the intralayer synchronization manifold. We also
derive the local stability condition of the interlayer antisynchronization state using the
master stability function approach.
vi


Emerging collective phenomena in static and temporal dynamical networks

Inapurapu, Santhi Priya

vii
Abstract
Structural and mechanical reliability of brittle solids for both conventional and 
advanced applications is determined by the rate at which it can deform and sustain 
externally applied static or dynamic strain at the microstructural length scale. Hence, the 
effect of loading rate on nanomechanical properties of brittle solids like soda-lime silica 
glass (SLS), MgAl2O4 Spinel glass ceramics and polycrystalline alumina ceramics have 
been investigated at different microstructural as well as measurement length scales. The 
nanohardness of brittle solids was found to increase with the loading rate. Further, 
signatures of serrations were observed in the P-h plots. These serrations were linked to 
the formation of shear bands inside and around the indentation cavities. A new concept of 
inelastic deformation (IED) parameter was found out to describe well the observed trend 
of loading rate dependencies of the nanohardness data. Further, it is predicted from the 
present wok that the shear stresses developed at various loading rates just beneath the tip 
of the nanoindenter were sufficient enough to cause shear induced plastic flow and / or 
deformation band formation inside the nanoindentation cavities. Time-dependent 
deformation behavior was also investigated for commercially available SLS glass with 
the help of a sharp Vicker’s diamond pyramidal indenter at a given peak load of 10 N by 
varying contact duration from 1- 90 sec. The results showed that the deformation 
behavior may be represented in terms of a simple mechanical model analogous to a 
viscoelastic system. 
Further, an attempt was also made to understand how the high strain rate flyer 
plate impact affects the nanohardness of alumina ceramics. The load controlled 
nanoindentation experiments were conducted with a Berkovich indenter on an as received 
coarse grain (~10 m) alumina and shock recovered tiny fragments of the same alumina 
obtained from gas gun experiments conducted at 6.5 GPa and 12 GPa shock pressure. 
The nanohardness of the as received alumina (ARA) was much higher than that of the 6.5 
GPa and 12 GPa shock recovered (SA) alumina. The indentation size effect (ISE) was 
strongest in 12 GPa SA sample, relatively stronger in 6.5 GPa SA sample but mild in the 
ARA sample. These results were rationalized by analysis of the experimental load depth 
data and evidences obtained from field emission scanning electron microscopy.


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