Shells of compacted DNA as nanocontainers transporting proteins in multiplexed delivery.

Polyethyleneimine (PEI) polymers are known to compact DNA strands into spheroid, toroid, or rod structures. A formulation with mannose-grafted PEI (PEIm), however, was reported to compact DNA into ~100 nm spheroids that indented like thin-walled pressurized shells. The goal of the study is to understand why mannose bristles divert the traditional pathway of PEI-DNA compaction to produce shell-like structures, and to manipulate the process so that proteins can be packed into the core of the assembling shells for co-delivering DNA and proteins into cells. DLS, AFM, and TEM imaging provide a consistent picture that BSA proteins can be packed into the shells without altering the shell architecture, as long as the proteins were added during the time course of shell assembly. Force spectroscopy studies reveal that DNA shells that buckle also have a rich surface-coating of mannose, indicating that a micelle-like partitioning of hydrophobic and hydrophilic layers governs shell assembly. When HEK293T cells are spiked with BSA-laden DNA shells, co-transfection of DNA and BSA is observed at higher levels than control formulations. Distinct micron-sized features appear having both green fluorescence from BSA-FITC and blue fluorescence from NucBlue DNA stain, suggesting BSA release in nucleus and secretory granules. With DNA nanocontainers, proteins can take advantage of the efficiency of PEI-based DNA transfection for hitchhiking into cells while being shielded from the challenges of the intracellular route. DNA nanocontainers are rapid to assemble, not dependent on the DNA sequence, and can be adapted for different protein types; thereby having potential to serve as a high-throughput platform in scenarios where DNA and protein have to be released at the same site and time within cells (e.g., theranostics, multiplexed co-delivery, gene editing).

Unusual Salt and pH Induced Changes in Polyethylenimine Solutions.

Linear PEI is a cationic polymer commonly used for complexing DNA into nanoparticles for cell-transfection and gene-therapy applications. The polymer has closely-spaced amines with weak-base protonation capacity, and a hydrophobic backbone that is kept unaggregated by intra-chain repulsion. As a result, in solution PEI exhibits multiple buffering mechanisms, and polyelectrolyte states that shift between aggregated and free forms. We studied the interplay between the aggregation and protonation behavior of 2.5 kDa linear PEI by pH probing, vapor pressure osmometry, dynamic light scattering, and ninhydrin assay. Our results indicate that: At neutral pH, the PEI chains are associated and the addition of NaCl initially reduces and then increases the extent of association.The aggregate form is uncollapsed and co-exists with the free chains.PEI buffering occurs due to continuous or discontinuous charging between stalled states.Ninhydrin assay tracks the number of unprotonated amines in PEI.The size of PEI-DNA complexes is not significantly affected by the free vs. aggregated state of the PEI polymer. Despite its simple chemical structure, linear PEI displays intricate solution dynamics, which can be harnessed for environment-sensitive biomaterials and for overcoming current challenges with DNA delivery.

Biogenic synthesis of Ag, Au and bimetallic Au/Ag alloy nanoparticles using aqueous extract of mahogany (Swietenia mahogani JACQ.) leaves.

In this paper, we have demonstrated for the first time, the superb efficiency of aqueous extract of dried leaves of mahogany (Swietenia mahogani JACQ.) in the rapid synthesis of stable monometallic Au and Ag nanoparticles and also Au/Ag bimetallic alloy nanoparticles having spectacular morphologies. Our method was clean, nontoxic and environment friendly. When exposed to aqueous mahogany leaf extract, competitive reduction of Au(III) and Ag(I) ions present simultaneously in same solution leads to the production of bimetallic Au/Ag alloy nanoparticles. UV-visible spectroscopy was used to monitor the kinetics of nanoparticles formation. UV-visible spectroscopic data and TEM images revealed the formation of bimetallic Au/Ag alloy nanoparticles. Mahogany leaf extract contains various polyhydroxy limonoids which are responsible for the reduction of Au(III) and Ag(I) ions leading to the formation and stabilization of Au and Ag nanopaticles.

Proton transfer reactions in nanoscopic polar domains: 3-hydroxyflavone in AOT reverse micelles.

A dramatic reduction in the excited-state intramolecular proton transfer (ESIPT) rate is observed for 3-hydroxyflavone (3-HF) within the nanoscopic polar domains of Aerosol-OT (AOT)/n-heptane reverse micelle solutions. It is attributed to the formation of intermolecularly hydrogen-bonded 3-HF:AOT complexes, which cause a significant disruption of intramolecular hydrogen bonding within the complex-bound 3-HF molecules, thereby limiting the overall rate of the ESIPT process. Introduction of strong hydrogen-bonding polar solvents like water or methanol into the reverse micelles causes extensive solvation of the AOT head groups, leading to the collapse of the 3-HF:AOT complex and eventual release of intramolecularly hydrogen-bonded 3-HF molecules which are then able to undergo ultrafast ESIPT. With increasing W-value (W=[polar solvent]:[AOT]), a larger number of 3-HF:AOT complexes are decimated, thus accelerating the overall ESIPT process. In contrast, in presence of solvents like acetonitrile, whose hydrogen-bonding power is inherently weak, the AOT head groups are poorly solvated, so that the 3-HF:AOT complexes are hardly affected at any W-value. Consequently the ESIPT dynamics of 3-HF in acetonitrile-containing AOT reverse micelles is nearly independent of the W-value, and always slower compared to that in water- or methanol-containing AOT reverse micelles. The results highlight the importance of hydrogen-bonding property of the polar solvent on the ESIPT of 3-HF within a nanoscopic domain.

Biogenic synthesis of Au and Ag nanoparticles by Indian propolis and its constituents.

In an attempt to find natural, environmentally benign, green-chemical agents for the synthesis of metal nanoparticles, we have demonstrated for the first time the excellent efficiency of ethanol and water extracts of a natural, non-toxic material, Indian propolis and two of its chemical constituents, pinocembrin and galangin in the rapid synthesis of stable Ag and Au nanoparticles having wide spectrum of fascinating morphologies. Both of these two extracts were found to be extremely efficient in the synthesis of Ag and Au nanoparticles under alkaline condition. For a given metal ion precursor, the kinetics of particle synthesis were remarkably similar in all the cases, as it is evident from the absorption spectra monitored over time. Moreover they exhibited similar redox behavior under alkaline condition (pH approximately 10.62). The efficiency of the ethanol and water extracts of Indian propolis towards Ag and Au nanoparticles synthesis was compared with that of naturally occurring hydroxyflavonoids, pinocembrin and galangin isolated from Indian propolis; which are equally efficient in the rapid synthesis of Ag and Au nanoparticles and stabilization of the resultant particles.

Biogenic synthesis of Au and Ag nanoparticles using aqueous solutions of Black Tea leaf extracts.

We explored the application of three different aqueous solutions derived from Black Tea leaf extracts in the synthesis of Au and Ag nanoparticles. The plain tea leaf broth, as well as that containing the ethyl acetate extract of tea leaves, were found to be extremely efficient, leading to rapid formation of stable nanoparticles of various shapes: spheres, trapezoids, prisms and rods. For a given metal ion precursor, the kinetics of particle synthesis were remarkably similar in these two solutions, as evidenced from their absorption spectroscopy monitored over time. Moreover, they exhibited similar redox behavior. In contrast, with the other solution, containing the dichloromethane (CH(2)Cl(2)) extract of tea leaves, we failed to detect any nanoparticle generation under similar reaction conditions. Our results suggest that the reduction of metal ions and stabilization of the resultant particles in the first two solutions involved the same class of biomolecules. We identified these biomolecules as the tea polyphenols, including flavonoids, which were present in comparable amounts in both the tea leaf broth and ethyl acetate extract, but are absent in the CH(2)Cl(2) extract of tea leaves. The efficiency of the tea leaf extracts towards Au and Ag nanoparticle synthesis were compared with that of a naturally occurring hydroxyflavonoid, quercetin.