Assessment of recombinant glutathione-S-transferase (HaGST-8) silica nano-conjugates for effective removal of pesticides.

Diverse glutathione-S-transferases (GSTs) are produced by insect pests including Helicoverpa armigera (HaGSTs) for detoxification of insecticides or xenobiotic compounds that they encounter. In an earlier study, the HaGST-8 gene was isolated from H. armigera larvae exposed to pesticide mixtures and the recombinant protein was expressed in the yeast Pichia pastoris. In this investigation, HaGST-8 was successfully immobilized on glutaraldehyde-activated APTES functionalized silica nanoparticles to obtain SiAPT-HaGST-8 nano-conjugates. Although enzyme activity associated with these conjugates was comparable to that of free HaGST-8, the specific activity of the former was found to be 1.25 times higher than the latter. In comparison with the free enzyme (that demonstrated a pH optimum of 9.0), for the nano-conjugates, the pH range was extended between pH 8.0 to 9.0. The optimum temperature for activity of both forms of the enzyme was found to be 30 °C. Stability of the enzyme was improved from 20 d for free HaGST-8 to 30 d for SiAPT-HaGST-8 nano-conjugates. Some loss in GST activity was detected after every reuse cycle of nano-conjugates and in all, 63% reduction was observed after three cycles. When 3 kinds of pesticides (namely, chlorpyrifos, dichlorvos and cypermethrin) were reacted with SiAPT-HaGST-8, more than 80% reduction in levels were observed. On the basis of the results obtained, the use of such silica nanoparticle-based systems for stable enzyme conjugation followed by effective removal of pesticides from aqueous media is envisaged.

Layer-by-Layer Assembled Nanostructured Lipid Carriers for CD-44 Receptor-Based Targeting in HIV-Infected Macrophages for Efficient HIV-1 Inhibition.

Macrophages act as a cellular reservoir in HIV infection. Elimination of HIV from macrophages has been an unfulfilled dream due to the failure of drugs to reach them. To address this, we developed CD44 receptor-targeted, novel hyaluronic acid (HA)-coated nanostructured lipid carriers (NLCs) of efavirenz via washless layer-by-layer (LbL) assembly of HA and polyallylamine hydrochloride (PAH). NLCs were subjected to TEM analysis, size and zeta potential, in vitro release and encapsulation efficiency studies. The uptake of NLCs in THP-1 cells was studied using fluorescence microscopy and flow cytometry. The anti-HIV efficacy was evaluated using p24 antigen inhibition assay. NLCs were found to be spherical in shape with anionic zeta potential (-23.66 ± 0.87 mV) and 241.83 ± 5.38 nm particle size. NLCs exhibited prolonged release of efavirenz during in vitro drug release studies. Flow cytometry revealed 1.73-fold higher uptake of HA-coated NLCs in THP-1 cells. Cytotoxicity studies showed no significant change in cell viability in presence of NLCs as compared with the control. HA-coated NLCs distributed throughout the cell including cytoplasm, plasma membrane and nucleus, as observed during fluorescence microscopy. HA-coated NLCs demonstrated consistent and significantly higher inhibition (81.26 ± 1.70%) of p24 antigen which was 2.08-fold higher than plain NLCs. The obtained results suggested preferential uptake of HA-coated NLCs via CD44-mediated uptake. The present finding demonstrates that HA-based CD44 receptor targeting in HIV infection is an attractive strategy for maximising the drug delivery to macrophages and achieve effective viral inhibition.

Evaluation of silica nanoparticle mediated delivery of protease inhibitor in tomato plants and its effect on insect pest Helicoverpa armigera.

The inert and surface tunable nature of silica nanoparticles (SiNPs) makes them suitable for different applications. We have evaluated the potential of SiNPs for delivering proteins in tomato (Lycopersicon esculentum) plants. SiNPs of 20 and 100 nm (Si20 and Si100) were functionalized with (3-aminopropyl) triethoxysilane (APTES) to obtain Si20APT and Si100APT, respectively, that were non-toxic toward plants. The functionalized nanoparticles were taken up by plants through roots as well as leaf surfaces. They were seen to be localized near the vasculature, particularly around the xylem. Si20APT and Si100APT nanoparticles were conjugated with soybean trypsin inhibitor (STI) to yield Si20APT-STI and Si100APT-STI, respectively. Based on the trypsin inhibitory activity of loaded nanoparticles, optimum loading was obtained for 0.4 mg of STI per 0.8 mg of NPs. Si20APT nanoparticles retained higher contents of STI than Si100APT. Exposure of STI-conjugated nanoparticles to 25°C or pH 8.0 aided release of the inhibitor. The particle bound STI inhibited bovine trypsin by 80% and Helicoverpa armigera gut proteinase (HGP) activity by 50%. Second instar H. armigera larvae ingesting STI-loaded particles (incorporated in artificial diet or leaves) showed significant retardation in growth. In choice assays, Si20APT-STI applied leaf discs were strikingly avoided by insect larvae. On the basis of the results obtained in this investigation, we recommend the use of Si20 nanoparticles for developing plant delivery vehicles in the future.

Silica nanoparticle based techniques for extraction, detection, and degradation of pesticides.

Silica nanoparticles (SiNPs) find applications in the fields of drug delivery, catalysis, immobilization and sensing. Their synthesis can be mediated in a facile manner and they display broad range compatibility and stability. Their existence in the form of spheres, wires and sheets renders them suitable for varied purposes. This review summarizes the use of silica nanostructures in developing techniques for extraction, detection and degradation of pesticides. Silica nanostructures on account of their sorbent properties, porous nature and increased surface area allow effective extraction of pesticides. They can be modified (with ionic liquids, silanes or amines), coated with molecularly imprinted polymers or magnetized to improve the extraction of pesticides. Moreover, they can be altered to increase their sensitivity and stability. In addition to the analysis of pesticides by sophisticated techniques such as High Performance Liquid Chromatography or Gas chromatography, silica nanoparticles related simple detection methods are also proving to be effective. Electrochemical and optical detection based on enzymes (acetylcholinesterase and organophosphate hydrolase) or antibodies have been developed. Pesticide sensors dependent on fluorescence, chemiluminescence or Surface Enhanced Raman Spectroscopic responses are also SiNP based. Moreover, degradative enzymes (organophosphate hydrolases, carboxyesterases and laccases) and bacterial cells that produce recombinant enzymes have been immobilized on SiNPs for mediating pesticide degradation. After immobilization, these systems show increased stability and improved degradation. SiNP are significant in developing systems for effective extraction, detection and degradation of pesticides. SiNPs on account of their chemically inert nature and amenability to surface modifications makes them popular tools for fabricating devices for 'on-site' applications.