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Synthesis and Cytotoxicity Studies of Novel Triazolo- Benzoxazepine as New Anticancer Agents
No full textCancer continues to be one of the biggest threats to
the human civilization because there is no cure of it.
Small heterocyclic molecule with low molecular weight
and novel structural feature is therapeutically highly
demanding. These molecules have the capability to
disrupt signaling pathways leading to anticancer activities.
Therefore, the search for new anticancer agents
continues to draw attention to the research community.
In this study, a small triazolo-benzoxazepine scaffolds
was synthesized using a one-pot four-step synthetic methodology involving click reaction. Small libraries of 12 compounds were successfully synthesized and screened them against different cancer cell lines. Low micromolar anticancer activity was recorded using MTT assay, and further confirmation of cell death was obtained by phase contrast, fluorescent, and confocal image
Thermodynamic Investigations of Ligand–Protein Interactions: Binding of the Phenazinium Dyes Phenosafranin and Safranin O with Human Serum Albumin
No full textThe binding of the phenazinium dyes, phenosafranin (PSF) and safranin O (SO) with human serum albumin was investigated by calorimetry and spectroscopic techniques. Binding parameters revealed that PSF has a higher affinity (K = 1.60 � 105 M�1) compared to SO (K = 0.97 � 105 M�1). The binding of both dyeswas favoured by negative enthalpy and a stronger favourable entropy contribution. The heat capacity values were similar indicating the involvement of similar hydrophobic forces in the complexation. Enthalpyentropy compensation was also observed for both dyes. Both polyelectrolytic and non-polyelectrolytic forces contributed towards the binding but the later was dominant. The fluorescence data suggested a static quenching mechanism. Forster resonance energy transfer studies showed that the specific binding distances between Trp (donor) residue of the protein and the dye (acceptor) molecules were 3.95 and 4.07 nm, respectively, for PSF and SO. Both dyes bind to same site, viz. site I (subdomain II A) of HSA but SO having bulkier groups binds weakly due to steric hindranc
Binding of Plant Alkaloids Berberine and Palmatine to Serum Albumins: A Thermodynamic Investigation
No full textThe thermodynamics of the interaction of two
pharmaceutically important isoquinoline alkaloids berberine
and palmatine with bovine and human serum albumin was
investigated using calorimetric techniques, and the data was
supplemented with fluorescence and circular dichroism
studies. Thermodynamic results revealed that there was only
one class of binding sites for both alkaloids on BSA and
HSA. The equilibrium constant was of the order of 104 M-1
for both the alkaloids to serum albumins but the magnitude
was slightly higher with HSA. Berberine showed higher
affinity over palmatine to both proteins. The binding was
enthalpy dominated and entropy favoured for both the alkaloids
to BSA and HSA. Salt dependent studies suggested that
electrostatic interaction had a significant role in the binding
process, the binding affinity reduced as the salt concentration
increased. Temperature dependent calorimetric data yielded
heat capacity values that suggested the involvement of different
molecular forces in the complexation of the two alkaloids
with BSA and HSA. 3D fluorescence, synchronous
fluorescence and circular dichroism data suggested that the
binding of the alkaloids changed the conformation of proteins
by reducing their helicity. Destabilization of the protein
conformation was also revealed from differential scanning
calorimetry studies. Overall, the alkaloids bound strongly to
serum albumins, but berberine was a better binder to both
serum proteins compared to palmatin
Studies on Enzymes of Trehalose Metabolism from Candida Utilis
No full textTrehalose is a non-reducing disaccharide formed by the 1,1 linkage of two D-glucose molecules [Elbein et al., 2003]. The sugar is present in a wide range of organisms, including
bacteria, yeast, fungi, insects, invertebrates and plants where it may serve as a source of energy and carbon [Elbein et al., 2003]. In comparison with most sugars trehalose is far more stable to wide fluctuations of pH and heat and does not easily interact with proteinaceous molecules
[Richards et al., 2002]. The superior stability of trehalose is largely contributed by the glycosidic
bond joining the two glucose units which is not easily hydrolyzed by acid or alkali and is not cleaved by α-glucosidase [Richards et al., 2002]. The molecular formula and weight of trehalose are C12H22O11 and 342.31, respectively. When purified it is usually found in the dihydrate form, which is the typical commercial product. Physical properties that make trehalose unique are its
high degree of optical rotation ([a]D2 +178º) and its melting behavior [Richards et al., 2002].Trehalose first melts at 97ºC. Additional heat drives off the water of crystallization until the material resolidifies at 130ºC, and then the anhydrous trehalose melts at 203ºC. The combination of the molecular structure and physico - chemical properties of trehalose result in a very stable
disaccharide [Birch et al., 1963, Elbein et al., 1974]. Although α,β (neotrehalose) and β,β(isotrehalose) isomers of trehalose have been synthesized, they are rarely found in nature [Birch et al., 1963, Elbein et al., 1974]. The α,α form is the isomer commonly referred as trehalose
(α,α-trehalose, α-d-glucopyranosyl α-d-glucopyranoside, mushroom sugar or mycose) and is widespread throughout the plant and animal kingdoms [Birch et al.,, 1963, Elbein et al., 1974].
Trehalose is one of the most important storage carbohydrates and serves as a
carbohydrate reservoir in organisms like plants, algae, fungi, yeasts, bacteria, insects and other
invertebrates. Interestingly, for all of the species of insects where trehalose is present, it is the
principal sugar (approximately 80–90%) found in the hemolymph and constitute about 20% of
all carbohydrates during specific stages of insect development [Wyatt et al., 1957, Harding et al.,
1923]
Cell biology Of Antigen Processing And Presentation In Leishmania Infection And Its Implications In Antileishmanial Vaccine Development
No full textProtozoan parasites belonging to the genus Leishmania exhibit a pronounced tropism for macrophages. In this thesis, we have highlighted aspects of the surface
chemistry and intracellular antigen processing and presentation of Leishmania donovani infected macrophage.
We recall from our previous study that Leishmania donovani infection causes fluidity in the macrophage membrane. We have showed here the fluidity in infected macrophage membranes are due to decreased cholesterol content in the
membranes. Naturally the question arises, which physical parameters of a biological membrane are influenced by cholesterol? Membrane cholesterol regulates membrane protein function and dynamicity. We have seen a decrease in
antigen presenting ability of MHC class II in Leishmania infection. MHC class II molecule being a transmembrane protein may interact with plasma membrane
components which is responsible for its functional activity. Leishmania infection causes depletion of cholesterol from macrophage membrane which in turn affects
the antigen presenting ability of macrophage, although the precise nature of interaction is still unclear. Measurements of dynamics of membrane proteins
provide another window to show the effect of cholesterol depletion in Leishmania infected macrophage. Studying the local diffusion of membrane protein over time
provide information about the dynamic property of the membrane in Leishmania infection. Biochemically, the cholesterol forms rigid liquid ordered structures
within membranes called lipid rafts. The viscous drag of the lipid rafts restricts the mobility of the proteins. The novel technique allowed us to compare the
diffusion of proteins in normal and Leishmania infected macrophage membrane. Leishmania infection increases the mobility of PLC-δ1 (a lipid raft oriented membrane protein) in macrophages. The lateral diffusion of this protein is again restored back to normal with treatment with liposomal delivery of cholesterol
Phenazinium dyes methylene violet 3RAX and indoine blue bind to DNA by intercalation: Evidence from structural and thermodynamic studies
No full textThe structural and energetic aspects of the interaction of two important phenazinium dyes methylene
violet 3RAX and indoine blue with DNA were studied by spectroscopic and thermochemical techniques.
The binding affinity values of both the dyes were of the order of 105 M�1 but methylene violet 3RAX
bound to DNA more strongly. Both dyes bound DNA by intercalation but methylene violet 3RAX bound
stronger than indoine blue. The DNA binding of both the dyes was exothermic and favoured by a small
negative enthalpy and positive entropy contributions. The effect of ionic strength indicated that
electrostatic attraction is an important component of the dyeeDNA interaction although the major
contribution comes from hydrophobic forces. The temperature dependence of enthalpy changes yielded
negative heat capacity value which was higher for methylene violet 3RAX compared to indoine
blue suggesting higher contribution from hydrophobic forces in the interaction of the former.
Enthalpyeentropy compensation phenomenon was observed for the binding of both dyes to DNA
Synthesis and characterization of different types of nanoparticles and their applications
No full textNanoparticles are particles having size less than 100 nm. The properties of a conventional material changes when it forms nanoparticle. This is typically because nanoparticles have a greater surface area per unit weight than larger particles. This causes them to be more reactive to certain other molecules. As a result, nanoparticles are used or being evaluated for use in many fields. Our study has focused on three areas of application of nanoparticles, viz. (A) application of arsenite coated gold nanoparticles as anticancer agent, (B) role of gold nanoparticles in increasing polymerase chain reaction efficiency and (C) synthesis of carbon nanoparticles and its interaction with various biologically important proteins.(A) Application of arsenite coated gold nanoparticles as anticancer agent
Arsenic with its long history as a human poison has proven to be valuable as a potent anticancer agent, especially at low doses. In the present study, sodium arsenite conjugated gold nanoparticles were synthesized by reduction of
chloroauric acid with trisodium citrate followed by addition of sodium arsenite under boiling condition. Dynamic Light Scattering (DLS), Transmission Electron
Microscopy (TEM) and Atomic Force Microscopy (AFM) were used to confirm the formation and size distribution of arsenite coated gold nanoparticles. The arsenic-conjugated gold nanoparticles were characterized by a larger size (29 nm)as well as red shift in the absorption spectrum (532 nm) compared to gold nanoparticles (23 nm and 522 nm).
(B) Role of gold nanoparticles in increasing polymerase chain reaction efficiency The polymerase chain reaction (PCR) enhancing effect of gold nanoparticles (GNPs) might arise due to enhancement of heat transfer by high-conductivity, high-heat-capacity GNPs or PCR enhancing effect of GNP is mediated by surface interactions of GNPs rather than by heat-transfer enhancement. The present study
aimed at determining the most probable function of GNP during the PCR to regulate the yield of PCR. The influence of GNPs with different reacting components of PCR was monitored by UV-visible spectroscopy, fluorescence
quenching and circular dichroism spectroscopy. This study also involved the synthesis and characterization of GNPs by transmission electron microscopy, atomic force microscopy, dynamic light scattering and UV-visible spectroscopy.
An increase in the yield of PCR product was observed initially with addition of increasing concentration of GNPs, reached a maximum value and declined thereafter. Circular dichroism studies showed that Taq DNA polymerase had a greater affinity to interact with GNPs compared to primer or DNA template and CD melting study showed that pure Taq has an inversion temperature 73°C which increased to 81°C for Taq-GNP adduct. Melting temperature of the Taq-GNP adduct being 81°C, the available number of active Taq DNA molecules at the normal extension temperature (68-72°C) is much higher, which could be the reason for higher yield of PCR product in presence of GNP These findings would
shed light toward the trafficking power of nanoparticles on various enzymatic reactions to regulate the yield of the product.(C) Synthesis of carbon nanoparticle and its interaction with various biologically important proteins
Carbon nanoparticles (CNPs) with very small size and high purity were synthesized by dehydration of D-glucose using concentrated sulphuric acid as dehydrating agent. These were characterized by transmission electron microscopy,
atomic force microscopy, X-ray diffraction and UV-visible spectroscopy. CNPprotein interactions were studied by fluorescence spectroscopy, circular dichroism spectroscopy and isothermal titration calorimetry. The fluorescence quenching constants and thermodynamic parameters such as enthalpy change (ΔH0), entropy change (ΔS0) and free energy change (ΔG0) were calculated, which indicated a
strong static quenching and primary electrostatic interaction between the carbon nanoparticles and blood proteins. Circular dichroism spectra provided the
information about the secondary structure alteration of the proteins in presence of carbon nanoparticles. These findings have shed light toward an understanding of
the interactions between CNPs and serum proteins which may clarify the potential risks and undesirable health effects of carbon CNPs, as well as the related cellular
trafficking and systemic translocation
Studies on the Bioactive Constituents of Azadirachta indica and Shorea robusta
No full textThe work embodied in this thesis describes the isolation and structure elucidation of some phytochemical constituents like glycolipids, cerebrosides, flavone glycosides and triterpenoids from Indian medicinal plants Azadirachta indica A. Juss (Neem) and Shorea robusta Gaertner f. (Sal) that are receiving much attention in recent years for their useful biological activities.The chapter I of thesis contains a review on Shorea robusta. A list major classes of compounds triterpenoids, phenolic compounds, volatile oils, steroids and a steroid glycosides so far isolated from different parts of Shorea robusta and pharmacological activities of few extracts and compounds have been presented in this review.Chapter II describes the isolation, purification of phytochemical constituents from mature leaves of Azadirachta indica and characterization by modern spectroscopic technique like IR, ESI-MS, 1H NMR, 13C NMR with DEPT 90 and 135, 2D NMR– COSY, NOESY, HSQC, HMBC. It also describes anti-leukemic activity in human leukemic cell lines U937 and K562 with most active principle; sulfonoquinovosyldiacylglyceride (SQDG). SQDG also shows significant antibacterial as well as anti herpes virus activity. Chapter III of thesis describes the isolation, purification phytochemical constituents from Shorea robusta resin and characterization by modern spectroscopic technique like IR, ESI-MS, 1H NMR, 13C NMR with DEPT 90 and 135. Chapter also deals with spermicidal, microbicidal activity of Asiatic acid
Translating immune cell cross-talk into a treatment opportunity for visceral leishmaniasis
No full textResults from both murine and human studies suggest the emergence of immunotherapy, involving different agonists and antagonists to the cytokines and cells involved in the course of disease progression, as a promising treatment option against visceral leishmaniasi
Fluorescence Correlation Spectroscopy Study on the Effects of the Shape and Size of a Protein on Its Diffusion Inside a Crowded Environment
No full textFluorescence correlation spectroscopy (FCS)
has been commonly used to study the diffusional and
conformational fluctuations of labeled molecules at singlemolecule resolution. Here, we explored the applications of FCS inside a polyacrylamide gel to study the effects of molecular weight and molecular shape in a crowded environment. To understand the effect of molecular weight,we carried out FCS experiments with four model systems of different molecular weights in the presence of varying concentrations of acrylamide. The correlation curves were fit adequately using a model containing two diffusing components: one representing unhindered diffusion and one representing slow hindered diffusion in the gel phase. A large number of measurements carried out at different randomly chosen spots on a gel were used to determine the most probable diffusion time values using Gaussian distribution analysis. The variation of the
diffusivity with the molecular weight of the model systems could be represented well using the effective medium model. This model assumes a combination of hydrodynamic and steric effects on solute diffusivity. To study the effects of solute shape, FCS experiments were carried inside a urea gradient gel to probe the urea-induced unfolding transition of Alexa488Maleimide-labeled bovine serum albumin. We showed that the scaling behavior, relating the hydrodynamic radius and the number of amino acids, changes inside an acrylamide gel for both folded and unfolded proteins. We showed further that crowding induced by a polyacrylamide gel increases the resolution of measuring the difference in hydrodynamic radii between the unfolded and folded
states