The thesis entitled “The Role of Liposomal Hybrids and Gold Nanoparticles in the Efficacious Transport of Nucleic Acids and Small Molecular Drugs for Cancer Nanomedicine” elucidates the preparation of various liposomal formulations of cationic monomeric and gemini lipids where hydrophobic domains were consisted of tocopherol, cholesterol and pseudoglyceryl backbone for the cellular transport of nucleic acids. The thesis continues while elucidating the role of various pH sensitive molecules and gold nanoparticles in liposomes to improve the delivery efficacy levels. This thesis also elucidates the role of gold nanoparticles stabilized with natural pH sensitive molecules for efficacious drug delivery applications. Additionally, the role of such pH sensitive gold nanoparticles in association with liposomes for the co-delivery of drug and gene has been discussed. The work has been divided into six chapters. Chapter 1A: Dimeric Lipids Derived from α-Tocopherol as Efficient Gene Transfection Agents. Mechanistic Insights into Lipoplex Internalization and Therapeutic Induction of Apoptotic Activity In this chapter, we present cationic dimeric (gemini) lipids for significant plasmid DNA (pDNA) delivery to different cell lines without any marked toxicity in the presence of serum. The six gemini lipids possess α-tocopherol as their hydrophobic backbone and differ from each other in terms of their spacer chain lengths. Each of these gemini lipids mixed with a helper lipid 1, 2-dioleoyl phosphatidyl ethanolamine (DOPE), was capable of forming stable aqueous suspensions. These co-liposomal systems were examined for their potential to transfect pEGFP-C3 plasmid DNA in to nine cell lines of different origins. The transfection efficacies noticed in terms of EGFP expression levels using flow cytometry were well corroborated using independent fluorescence microscopy studies. Significant EGFP expression levels were reported using the gemini co-liposomes which counted significantly better than one well known commercial formulation lipofectamine 2000 (L2K). Transfection efficacies were also analyzed in terms of the degree of intracellular delivery of labeled plasmid DNA (pDNA) using confocal microscopy which revealed an efficient internalization in the presence of serum. The cell viability assays performed using optimized formulations demonstrated no significant toxicity towards any of the cell lines used in the study. We also had a look at the lipoplex internalization pathway to profile the uptake characteristics. A caveolae/lipid raft route was attributed to their excellent gene transfection capabilities. The study was further advanced by using a therapeutic p53-EGFP-C3 plasmid and the apoptotic activity was observed using FACS and growth inhibition assay. Figure 1. The co-liposomes of tocopheryl gemini lipids and DOPE for efficient delivery of p53-EGFP-C3 plasmid DNA that induces significant apoptotic response. Chapter 1B: Efficacious Gene Silencing in Serum and Significant Apoptotic Activity Induction by Survivin Downregulation Mediated by Cationic Gemini Tocopheryl Lipids Non-viral gene delivery offers cationic liposomes as promising instruments for the delivery of double-stranded RNA (ds RNA) molecules for successful sequence-specific gene silencing (RNA interference). The efficient delivery of siRNA (small interfering RNA) to cells while avoiding the unexpected side effects is an important prerequisite for the exploitation of the power of this excellent tool. We discuss in this chapter about six tocopherol based cationic gemini lipids, which induce substantial gene knockdown without any obvious cytotoxicity. All the efficient co-liposomal formulations derived from each of these geminis and a helper lipid, dioleoyl phosphatidyl ethanolamine (DOPE) were well characterized using physical methods such as atomic force microscopy (AFM) and dynamic light scattering (DLS). Zeta potential measurements were conducted to estimate the surface charge of these formulations. Flow cytometric analysis showed that the optimized co-liposomal formulations could transfect anti-GFP siRNA efficiently in three different GFP expressing cell lines, viz. HEK 293T, HeLa and Caco-2 significantly better than a potent commercial standard Lipofectamine 2000 (L2K) both in the absence and presence of serum (FBS). Notably, the knockdown activity of co-liposomes of gemini lipids was not affected even in the presence of serum (10% and 50% FBS) while it dropped down for L2K significantly. Observations under a fluorescence microscope, RT-PCR and western blot analysis substantiated the flow cytometry results. The efficient cellular entry of labeled siRNA in GFP expressing cells as evidenced from confocal microscopy put forward these gemini lipids among the potent lipidic carriers for siRNA. The efficient transfection capabilities were also profiled in a more relevant fashion while performing siRNA transfections against survivin (an anti-apoptotic protein) which induced substantial apoptosis. Furthermore, the survivin downregulation improved the therapeutic efficacy levels of an anticancer drug, doxorubicin significantly. In short, the new tocopherol based gemini lipids appear to be highly promising for achieving siRNA mediated gene knockdown in various cell lines. Figure 2. The co-liposomes of tocopheryl gemini lipids and DOPE for efficient delivery of siRNA against survivin that induces significant apoptotic response. Chapter 2: Efficacious in Vitro EGFP Expression and Silencing in Serum by Cationic Pseudoglyceryl Gemini Lipids To elicit the desirable efficacy levels in cationic liposome mediated nucleic acid therapeutics has been part of extensive scientific efforts. This chapter describes three cationic gemini lipids and application of their co-liposomes with DOPE as potent pDNA (plasmid DNA) and siRNA (small interfering RNA) cytofectins for remarkably advanced efficacy levels in numerous cell lines in the presence of serum. The hydrophobic structural lineament of cationic gemini lipids is made up of pseudoglyceryl backbone linked to the hydrocarbon chains via oligo-oxyethylene units. The stable aqueous co-liposomal suspensions of gemini lipids showed an efficient binding to pDNA or siRNA and their significant intracellular delivery in various cell lines. The transfection capabilities of different co-liposomal formulations were profiled based on EGFP expression (pEGFP-C3 pDNA transfection) and EGFP knockdown (anti-GFP siRNA transfections) in EGFP expressing cell lines. The cellular EGFP expression levels and intracellular delivery of labeled nucleic acids were thoroughly studied using flow cytometry (FACS), fluorescence and confocal microscopy. The MTT based cell viability assay revealed no loss in cell viabilities for all of the transfection optimized lipoplexes of siRNA or pDNA. The transfection profile of gemini co-liposomes was noted to be significantly much better than a commercial lipofection reagent, Lipofectamine 2000 used for pDNA and siRNA applications in each of the cell lines studied. The co-liposomes and their transfection optimized lipoplexes were physiochemically characterized extensively by means of zeta potential, dynamic light scattering (DLS) and atomic force microscopy (AFM). In brief, these new gemini co-liposomal formulations seem to offer a great opportunity for successful nucleic acid (DNA and siRNA) delivery in a practical scenario. Figure 3. Efficacious EGFP expression (pDNA transfection) and EGFP silencing (anti GFP siRNA transfection) mediated by co-liposomes of pseudoglyceryl gemini lipids and DOPE. Chapter 3: Efficient Elicitation of Liposomal Nucleic acid delivery through the Eminence of Gold Nanoparticles Stabilized with pH Responsive Short Tripeptide Derived from Tyrosine Kinase NGF Receptors The prerequisite in the area of gene therapy today is to serve transfection efficient formulations nullifying the enduring key issues. To this end, we discuss in this chapter, the role of hybrid liposomal formulations derived from structurally distinct cationic lipids, a neutral lipid (DOPE) and pH responsive short tripeptide (KFG, Lys-Phe-Gly) capped gold nanoparticles (PAuNPs). The hybrid liposomes are presented to be efficient enough to transfect pDNA leading to remarkably high gene expression levels in various cell lines of different origins in the presence of serum (FBS). Hybrid liposomes could deliver pDNA more effectively than the native liposomes and commercial standard lipofectamine 2000 (L2K) across the entire range of N/P ratios studied under the influence of intracellular pH response and gold nanoparticles prominence. The gene transfection capabilities are profiled based on transfections performed using two different plasmids (pGL3, luciferase activity and p-EGFP-C3, green fluorescent protein expression). pDNA cellular internalization and subsequent gene expression levels are studied using flow cytometry, fluorescence microscopy and confocal microscopic studies. The extensive physiochemical characterization of hybrid liposomal formulation and their complexes with pDNA in comparison with respective native liposomes was performed using AFM, TEM, Zeta, DLS, gel retardation assay, U.V. and fluorescence emission measurements. The hybrid liposomes are shown to possess significantly higher fusion activity at lowered pH of intracellular compartments. These hybrid liposomes are fairly biocompatible across the concentration range used in transfection experiments. Precisely, introduction of these pH responsive tripeptide capped gold nanoparticles in to liposomal formulations straightforwardly must be more advantageous for a practical application in biomedical scenario to achieve therapeutic levels. Figure 4. The hybrid of liposomes and tri-peptide capped gold nanoparticles for significantly improved gene expression levels. Chapter 4: RNA Aptamer Decorated pH Sensitive Liposomes for Active Transport of Nucleic Acids in Specific Cancer Cells This chapter describes the target specific transport of pH sensitive liposomes loaded with a RNA aptamer for promising nucleic acid therapeutics. The pH sensitive liposomes are constructed from a cationic cholesteryl gemini lipid (CGL), neutral helper lipid (DOPE) and gemini analog of a pH sensitive lipid, palmitoyl homocysteine (GPHC). The liposomes are shown to be significantly fusogenic that deliver the cargoes upon lowerin the pH (6.0). The fusogenic behaviour of the liposomes was thoroughly studied by means of dynamic light scattering (DLS), zeta potential, lipid mixing, calcein dequenching and atomic force microscopy (AFM). The facile integration of cholesterol conjugated RNA aptamer in liposomes derived from cholesteryl gemini lipids was exploited for their delivery to specific cancer cells. The RNA aptamer specifically binds to epithelial cell adhesion molecule (EpCAM) with high affinity which is a cell surface marker in various solid cancers such as colorectal and breast carcinoma. These aptamer decorated pH sensitive liposomes could efficiently enter the EpCAM expressing COLO-205, Caco-2, MCF-7 and MDA-MB-231 cell lines while no such noticeable liposome transport was observed in EpCAM negative HEK 293T cells as evidenced by flow cytometry and confocal microscopy. Additionally, the liposomes are shown to be actively transported inside the cells, i.e., receptor mediated endocytosis. These liposomes could complex the nucleic acids (pDNA) in an efficient manner. The MTT based cell viability assay accounted no noticeable loss in cell viabilities for liposome treatments. Concisely, we have formulated RNA aptamer loaded pH sensitive liposomes that would certainly be promising tool in target based cancer nanomedicine. Figure 5. (A) Cellular internalization of DY-647 labeled aptamer loaded pH sensitive liposomes. (B) The liposomes were actively internalized through receptor mediated endocytosis. Each panel (A and B) represents (from left to right) bright field image, aptamer fluorescence, DAPI stained nuclei and merge of previous three impressions. Chapter 5: Natural Tri-peptide Capped Gold Nanoparticles for Efficacious Doxorubicin Delivery in Vitro and in Vivo Nanotechnology has gained ever increasing interest for the successful implementation of chemotherapy based treatment of cancer. This chapter describes the role of gold nanoparticles (AuNPs) capped with a natural pH responsive short tri-peptide (Lys-Phe–Gly or KFG) for significant intracellular delivery of an anti-cancer drug, doxorubicin (DOX). A significantly increased apoptotic response was noted for DOX treatments mediated by KFG-AuNPs in comparison with drug alone treatments in various cell lines (BT-474, HeLa, HEK 293T and U251) in vitro. Furthermore, KFG-AuNPs mediated DOX treatment significantly decreased cell proliferation and tumor growth in BT-474 cell xenograft model in nude mice. In addition, KFG-AuNPs showed efficacious drug delivery in DOX-resistant HeLa cells (HeLa-DOXR) in comparison with drug alone treatments. Figure 6. Representative images of excised tumors after doxorubicin treatment mediated by pH responsive tri-peptide capped gold nanoparticles (DOX-KFG-AuNPs) (C) in comparison with doxorubicin alone treatments (B) and untreated tumors (A). Extensive cell death as observed under Hematoxylin/eosin (H&E) (D) and TUNEL (E) staining of DOX-KFG-AuNPs treated tumor sections. Chapter 6: Significant Apoptotic Activity Induction by Efficacious Co-delivery of p53 Gene and Doxorubicin Mediated by the Combination of Co-liposomes of Cationic Gemini lipid and pH Responsive Tri-peptide Combining chemotherapy with gene therapy has appeared as an efficient tool to treat complex biological disorder like cancer. Herein, we show efficient co-delivery of DNA and an anti-cancer drug, doxorubicin (DOX) by means of gemini cationic liposome (GCL) based lipoplex nanoaggregates that are coated with DOX encapsulated pH responsive tripeptide nanovesicles. The lipoplex, tripeptide vesicles and their association was thoroughly studied using dynamic light scattering (DLS), zeta potential, atomic force microscopy (AFM). Flow cytometry, fluorescence and confocal microscopic analysis revealed that the GCL-tripeptide association could significantly co-deliver the p53 expression plasmid (p53-EGFP-C3) and DOX in HeLa and HEK 293T cells in the presence of serum. A synergistic increase in gene expression level and DOX internalization was observed in co-delivery which was even substantially higher than individual lipoplex transfection and DOX treatment. The apoptosis induced due to p53 expression and DOX was profiled with the help of annexin-V positivity analysis under flow cytometry and nuclear damage analysis by DAPI nuclei counterstaining under confocal microscopy which noted to be significantly higher in cells during co-delivery. The MTT based cell viability assay revealed a significantly increased loss in cell viability counts for co-delivery treatments. Such a system delivering synergistically increased significant efficacy levels in combinatorial drug and nucleic acid therapeutics would be certainly advantageous for practical biomedical applications. Figure 7. The co-delivery of pDNA and drug (doxorubicin) mediated by GCL-tripeptide association as observed under (A) confocal microscopy (pDNA; green and doxorubicin; red) and (B) flow cytometry.

The thesis entitled “Novel Cationic Gemini Lipids, Click Chemistry Based Adducts and Amphiphile-Capped Silver Nanostructures: Synthesis, Aggregation and Biological Properties” elucidates the design, synthesis, aggregation and gene transfection properties of novel gemini cationic lipids based on cholesterol and pseudoglyceryl backbone, and click chemistry based adducts. This thesis also elucidates the synthesis and aggregation properties of silver nanoparticles loaded cationic liposomes and silver nanorods stabilized by micellar solutions of gemini surfactants. The work has been divided into six chapters. Chapter 1: Introduction: Membrane Formation from Cholesterol-based Cationic Lipids and their use as Non-Viral Gene Delivery Agents This chapter describes the importance of cholesterol in biological membranes, the aggregation properties of cholesterol-based cationic lipids and their interactions with phospholipid membranes. This chapter also gives a comprehensive account of the research towards the development of novel cationic cholesterol-based monomeric and gemini lipids. It also reviews the utilization of cholesterol-based cationic monomeric and gemini lipids in gene transfection properties. Chapter 2A: Effect of Hydroxyl group on the Cationic Headgroups of Cholesterol-based Gemini Lipids on their Aggregation, DNA Binding Properties and Interaction with Phospholipid Membranes This chapter describes the syntheses and aggregation properties of two series of cholesterol-based monomeric and gemini cationic lipids with and without hydroxyl functionality (Figure 1). The gemini lipids of a given series differ in their spacer polymethylene -(CH2)n- chain lengths between the cationic headgroups. Figure 1. Molecular structures of non-hydroxylated and hydroxylated cationic cholesterol-based gemini lipids and their monomeric counterparts. All monomeric and gemini lipids were found to generate stable suspensions in aqueous media. Electron microscopic studies showed that all the lipids form vesicular aggregates in aqueous media. The structures seen under TEM for the non-hydroxylated series of monomeric (C-M) and gemini lipids are of variable sizes, they appeared like separated vesicular aggregates. For the hydroxylated series of lipids, however, both the monomeric lipid aggregates (CH-M) and aggregates of their gemini counterparts were found to be ‘connected’ with each other to form elongated chain of aggregates of different length scales. XRD studies with the cast films of lipids revealed that the monomeric lipids of either series have higher bilayer width than the corresponding gemini lipids. Incorporation of the -(CH2)n- spacer units at the head group level joining the two monomeric units lowered the bilayer thicknesses of both series of the lipid aggregates. Thus the monomeric lipids (C-M and CH-M) appear to form nearly regular bilayer type arrangements whereas gemini lipids form interdigited and tilted bilayer arrangements in their aggregates. Calorimetry studies of the coaggregates showed that ~10 mol-% of most of the cholesterol gemini lipids is enough to abolish the phase transition of DPPC membranes whereas more than 10 mol-% is required in case of their monomeric counterparts. Further these thermotropic properties depend upon the length of the spacer of the gemini lipid included in the mixture. We have observed greater quenching of the thermal phase transition of DPPC membranes with 10 mol-% of C-M as compared to CH-M doped liposomes. At 10 mol-% of all the cationic lipid doped DPPC covesicles, only CG-3 doped liposomes showed an observable transition temperature. Maximum broadening of the DPPC transition peak was observed in the case of the gemini lipids, CHG-6 and CHG-12. DNA binding and release studies show that the interactions between gemini lipids and DNA depend upon the nature of the head group as well as the length of the spacer between the cationic head groups. For the non-hydroxylated cholesterol-based cationic lipid series, the monomeric liposomes of C-M facilitates the dissociation of EB from the EB-DNA complex to an extent of 93% at a maximum lipid:DNA ratio of 3.0 whereas the liposomes of CG-4 and CG-12 showed the lowest extent of maximum EB exclusion (~74%) from the EB-DNA complex at lipid:DNA ratio of 3.0. For hydroxylated cholesterol-based cationic lipid series, the monomeric liposomes of CH-M facilitate the dissociation of EB from the intercalated EB-DNA complex to an extent of 81 % whereas the liposomes of CHG-3 showed the minimum binding to DNA. Thus the two monomeric liposomes C-M and CH-M were the more efficient formulations that allow dissociation of DNA from the corresponding lipoplexes. These findings have important being in their gene transfection activity compared their respective gemini lipid counterparts. Chapter 2B: Novel Cholesterol-based Cationic Gemini Lipids possessing Hydroxyethyl group on the Headgroup: Transfection Efficacy and Cell Toxicity Properties This chapter describes the transfection efficacy and cell toxicity properties of five cholesterol based gemini cationic lipids possessing hydroxyethyl functionality on each head group, which differ in the length of the polymethylene spacer [-(CH2)n-] chain (Figure 2). These gemini lipids are important to gene delivery processes as they possess pre-optimized molecular features, e.g., cholesterol backbone, ether linkage and a variable spacer chain between both the headgroups of the gemini lipids. Cationic liposomes were prepared from each of these lipids individually and as a mixture of individual cationic gemini lipid and 1,2-dioleoylphosphatidylethanolamine (DOPE). The gemini lipid with a hydroxyethylated headgroup and a -(CH2)5- spacer, CHG-5 showed the highest transfection activity at N/P (lipid/DNA) ratio of 0.5 and lipid:DOPE molar ratio of 2. Upon comparison of the relevant parameters, e.g., % transfected cells, the amount of DNA transfected to each cell and % cell viability all together against LipofectAMINE 2000, one of the most potent commercially available transfecting agents, the optimized lipid formulation based on CHG-5/DOPE was found to be comparable. In terms of its ability to induce gene-transfer in presence of serum and shelf-life CHG-5/DOPE liposome was found to be better than its commercial counterpart. Recording of confocal images confirmed that in presence of 10% serum using 1.2 µg DNA per well and lipid:DOPE ratio of 1:4 and N/P charge ratio of 0.75, CHG-5 is better than LipofectAMINE 2000. These properties render them to be reagents of practical value for various gene delivery applications. Figure 2. Molecular structures of cholesterol-based cationic monomeric lipid and gemini lipids possessing hydroxyethyl group on the headgroup synthesized. Chapter 3: Bilayer Membrane and Stable Monolayer Forming Properties of Cationic Pseudoglyceryl Gemini Lipids having Polymethylene Spacers and Oxyethylene Linkages This chapter describes the synthesis of five new cationic pseudoglyceryl gemini lipid versions of their monomeric counterpart (Figure 3). Each cationic lipid aggregate in aqueous media was found to form vesicular structures as evidenced from the negatively stained TEM experiments and DLS measurements. XRD experiments with their cast films of aqueous dispersions revealed that introduction of the polymethylene -(CH2)n-spacer chain joining the two monomers decreased the bilayer widths of the gemini lipid aggregates. The inter-lipidic packing and the hydration of the lipid vesicles were examined using fluorescence anisotropy and generalized polarization measurements using membrane-soluble probes, DPH and Paldan respectively. Fluorescence anisotropy measurements showed that the aggregates of lipid 2c with -(CH2)5- spacer chain were highly packed and ordered in the vesicular aggregates than that of the other cationic lipid aggregates in the series. Paldan hydration studies showed that incorporation of the polymethylene -(CH2)n- spacer chains joining two monomeric units lowered the hydration of the gemini lipid aggregates in the solid gel state. Each of these cationic lipid aggregates showed sharp transition temperatures (Tm) as observed from differential scanning calorimetric studies. DSC studies further revealed that the incorporation of oxyethylene group at the linker region of cationic pseudoglyceryl gemini lipid 2a with (CH2)3- spacer chain length lowered the thermotropic phase transition temperature (Tm) of the aggregates in aqueous media when compared with the corresponding gemini analogue without oxyethylene linkages. Langmuir film balance studies showed that each cationic gemini lipid and their monomeric counterpart were able to form stable monolayers at the air-water interphase. We observed that the mean molecular area (collapse area) of each of the cationic lipid obtained from the Langmuir monolayer studies increased with increase in the spacer chain lengths. Figure 3. Molecular structures of the cationic pseudoglyceryl gemini lipids and their monomeric counterpart. Chapter 4: Vesicle and Stable Monolayer Formation from Simple ‘Click’ Chemistry Adducts in Water This chapter describes successful use of Cu(I) catalyzed “Click Chemistry” for the syntheses of a series of hitherto unknown amphiphilic adducts (M1, M2, D1 and T1) which on dispersal in water afforded vesicular aggregates as evidenced from dye entrapment, TEM, SEM, AFM and DLS studies (Figure 4). Figure 4. Molecular structures of triazole based adducts. XRD experiments with their cast films of aqueous suspensions indicate the formation of a tilted bilayer arrangement for the aggregates of M1 whereas regular bilayer structures are predominant for the aggregates derived from M2, D1 and T1. Measurement of pKa values using UV-Vis spectroscopy showed that the aggregates of monomeric click adducts (M1 and M2) possess less pKa value than that of the aggregates of dimeric (D1) and tetrameric (T1) analogues and the values lie within the range of 2.8-3.2. The hydrodynamic diameter of the aggregates of each click adduct increased with decrease in the pH of the media. Thus, the protonation of the triazole groups in the aggregates of each click adduct increased the hydrodynamic diameter. Dye entrapment studies showed that each click chemistry based adduct formed closed vesicular aggregates with inner aqueous compartment in aqueous media. The temperature induced order-to-disorder transitions of the aggregates and the accompanying changes in hydration were examined using high sensitive DSC, fluorescence anisotropy and generalized polarization measurements using a membranesoluble probe, DPH and Paldan respectively. In the solid state, M1 remains as the most hydrated species whereas in the fluidized phase, D1 maintains as the most hydrated aggregate. Clearly simple variation in the adduct molecular architecture bring about significant changes in their packing in aggregates and also the hydration of the resulting vesicles. Langmuir monolayer studies confirmed that these click adducts do form stable monolayers as well on water subphase at the air-water interface. We also calculated the mean molecular areas from the Langmuir monolayer studies and as perhaps expected the adduct T1 has the highest head group area. Thus click chemistry based simple triazole adducts, which can be very easily prepared, are good candidates for further investigations involving syntheses of novel self-assembling structures. Chapter 5: Lipid Mediated Synthesis of Silver Nanoparticles, their Physical Characterizations and DNA Binding Abilities In this chapter, work on the Ag-NP (silver nanoparticle) loaded liposomes preparation using four cationic lipids (1-4) in which the Ag-NPs were entrapped within lipid bilayer has been described. A novel method was developed to synthesize the Ag-NPs where the lipid itself capped and stabilized the Ag-NPs. Consequently there was no need of inclusion of any other capping agents like citrate. Confocal microscopy confirmed that these Ag-nanoparticles are fluorescent in character. It was also demonstrated that silver nanoparticles are indeed entrapped in lipid bilayer with transmission electron microscopy (TEM). DLS experiments provided information about the hydrodynamic diameter of the lipid vesicles which increased with the increase in Ag concentrations. This could be due to the ‘loosening’ of the lipid packing in vesicles. Zeta potential measurements showed that the zeta potential value decreased with the increase in the concentration of Ag-NPs in the cationic lipid vesicles. XRD studies with the cast films of the lipid or Ag-NP loaded lipid suspensions revealed that when the Ag-NPs get entrapped into the bilayer of the multilamellar vesicles of the lipid in the aqueous media, the unit bilayer thickness of the aggregates increased. Paldan experiments showed that with the incorporation of Ag-NPs in the lipid vesicles, the hydration of the lipid vesicles increased to a significant extent but the phase transition temperatures remained practically unaltered for all the lipids. Fluorescence anisotropy experiments revealed that the hydrocarbon chain packing of the lipid vesicles ‘loosens’ with the incorporation of Ag-NPs. Ag-NP loaded liposomes showed enhanced DNA binding ability and also the presence of Ag-NPs in cationic liposomes induced the release of DNA from silver nanoparticle-loaded lipoplexes more effectively. Figure 5. Molecular structures of the cationic lipids mentioned in the present chapter. Chapter 6: Dependence of Spacer Chain Lengths in the Synthesis of Ag-Nanorods in Gemini Cationic Surfactant Micelles Figure 6. Chemical structures of cationic gemini surfactants. This chapter describes the synthesis of Ag-nanospecies by seed-mediated wet synthesis method using four gemini surfactants (16-2-16, 16-4-16, 16-5-16 and 16-1216) as the capping agents (Figure 6). For this, we first synthesized Ag-nanoseeds of diameter ~7 nm stabilized by trisodium citrate (as capping agent). Then the solution containing Ag-nanoseeds was used to synthesize Ag-nanorods of different aspect ratios. It was that with decreasing Ag-nanoseed concentration, the aspect ratios of Agnanorods stabilized by gemini surfactants (16-2-16 and 16-4-16) increased gradually as evidenced from TEM images. These Ag-nanoseeds and Ag-nanorods were further characterized using UV-Vis spectroscopy (to know the surface plasmon bands), transmission electron microscopy (TEM), energy-dispersive X-ray spectroscopy (EDAX) and X-ray diffraction (XRD). It was observed that when gemini surfactant 164-16 was used to stabilize Ag-nanorods, the λmax of the longitudinal band shifted more towards the red region (red-shift) as observed by UV-Vis spectroscopy when compared to that of gemini surfactant with shortest spacer, 16-2-16. Thus the gemini surfactants with shorter -(CH2)2- and -(CH2)4- spacer chains promoted the growth of Ag-nanorods in their micellar solutions whereas -(CH2)5- and -(CH2)12- spacer chains of gemini surfactants did not. So, the growth of Ag-nanorods in micellar solutions is found to be highly spacer-chain length specific. TEM micrographs revealed that the aspect ratios of Ag-nanorods stabilized by gemini surfactants 16-4-16 are larger than those compared to the Ag-nanorods stabilized by gemini surfactants 16-2-16 at a particular amount of Agnanoseed solution. TEM images of the samples containing micellar solutions of gemini surfactants 16-5-16 and 16-12-16 showed that the formation of only Ag-nanoparticles of larger sizes (compared to Ag-nanoseeds stabilized by trisodium citrate) and Agnanoprisms irrespective of the amount of Ag-nanoseed solution added. No Ag-nanorod formation in the micellar solutions of gemini surfactants 16-5-16 and 16-12-16 was observed. Gemini surfactants (16-2-16 and 16-4-16) formed bilayer arrangements to facilitate the growth and stabilization of Ag-nanorods in aqueous media where the inner layer is attached to the Ag-nanorod surface through the gemini surfactant ammonium headgroups. X-ray diffraction (XRD) studies showed that these Ag-nanorods stabilized by gemini surfactants 16-2-16 and 16-4-16 crystallized in the aqueous media via (111), (220) and (222) lattice faces. Thus this study demonstrated the way one can control structures and shapes of the silver nanoobjects using gemini surfactant micelles. (For structural formula pl refer the thesis)