Pragya Sharma

Piper betle-mediated green synthesis of biocompatible gold nanoparticles

Here, we report the novel use of the ethonolic leaf extract of Piper betle for gold nanoparticle (AuNP) synthesis. The successful formation of AuNPs was confirmed by UV-visible spectroscopy, and different parameters such as leaf extract concentration (2%), gold salt concentration (0.5 mM), and time (18 s) were optimized. The synthesized AuNPs were characterized with different biophysical techniques such as transmission electron microscopy (TEM), Fourier transform infrared (FT-IR) spectroscopy, X-ray diffraction (XRD), and energy-dispersive X-ray spectroscopy (EDX). TEM experiments showed that nanoparticles were of various shapes and sizes ranging from 10 to 35 nm. FT-IR spectroscopy revealed that AuNPs were functionalized with biomolecules that have primary amine group –NH2, carbonyl group, –OH groups, and other stabilizing functional groups. EDX showed the presence of the elements on the surface of the AuNPs. FT-IR and EDX together confirmed the presence of biomolecules bounded on the AuNPs. Cytotoxicity of the AuNPs was tested on HeLa and MCF-7 cancer cell lines, and they were found to be nontoxic, indicating their biocompatibility. Thus, synthesized AuNPs have potential for use in various biomedical applications.

Photochemical activation of a polycarbonate surface for covalent immobilization of a protein ligand

Polycarbonate-a thermostable polymer is activated by a simple and rapid method using a photolinker, 1-fluoro-2-nitro-4-azidobenzene (FNAB) for covalent immobilization of a biomolecule. Horseradish peroxidase (HRP) is used as a model enzyme to check the efficacy of the activated surface. HRP is immobilized on the activated polycarbonate surface without addition of any reagent or catalyst and is found to give 2-2.5-fold increase in absorbance with the substrate as compared to the directly adsorbed enzyme. Photochemical attachment of FNAB to the PC surface is confirmed by X-ray photoelectron spectroscopy (XPS), which shows the presence of nitrogen and fluorine in the ratio of 2:1 in the activated polycarbonate. Disappearance of fluorine peak in the XP spectra of PC bound enzyme further confirms the covalent binding of HRP, through displacement of fluorine moiety of the activated PC by the amino group of the protein. Optimized concentration of the photolinker is found as 6mumol of FNAB per well and time of photo irradiation is 8min for activation of a PCR polycarbonate plate. PC bound HRP has shown enhanced thermal and storage stability. Kinetic studies of the immobilized HRP shows improved catalytic activity. The potential application of activated polycarbonate surface includes immobilization of biomolecules for biosensors, immunoassays, and protein and DNA micro-arrays. Due to the stability of the polycarbonate at high temperature, the activated polycarbonate has an advantage for immobilization of thermostable biomolecule such as thermostable enzyme for reaction at elevated temperature.

Green synthesis of gold nanoparticles using aqueous extract of Dillenia indica

In this study, we report a novel method of gold nanoparticle (AuNP) synthesis using aqueous fruit extract of Dillenia indica. The phytochemicals present in the fruit extract act as an effective reducing and capping agent to synthesize AuNPs. The synthesized AuNPs were characterized by spectrophotometry, transmission electron microscopy (TEM), x-ray diffraction (XRD), and Fourier transform infrared (FTIR) spectroscopy. TEM studies revealed the particles of various sizes and mainly spherical in shape. Selected-area electron diffraction (SAED) patterns and high-resolution transmission electron microscopy (HRTEM) images confirmed the crystallinity of the particles. The XRD patterns showed peaks at (111), (200), (220) which exhibited preferential orientation of the AuNPs as face-centered cubic crystal. FTIR measurements confirmed the coating of phenolic compounds on the AuNPs indicating a possible role of biomolecules for the capping and efficient stabilization of the AuNPs. The synthesized AuNPs did not show any form of cytotoxicity in the normal fibroblast cell line L929.

Ultrasound wave-mediated enzyme-linked immunosorbent assay technique

With the increasing burden of infectious diseases, it has become important to develop a rapid ELISA that could facilitate early diagnosis. Herein, we have shown that ultrasound waves can dramatically reduce the ELISA timing without losing its specificity or sensitivity. Ultrasound-mediated ELISA was best achieved on an activated microtiter plate which was able to covalently bind antigen or antibody in 10 min when subjected to ultrasound waves in a sonicator bath having a temperature of 37 degrees C, operating at a frequency of 40 KHz and an output power of 120 W. Blocking, antibody binding and secondary antibody-enzyme conjugate binding were also accomplished in 10 min each in the sonicator bath under similar conditions. The validation of SELISA method was demonstrated by detecting IgE in allergic patients sera. Total IgE detection by 40 min-SELISA method gives similar absorbance value to that obtained by 20 h conventional or 3 h-HELISA procedure. As SELISA method can detect IgE even at the serum dilution of 1/50 (v/v) on photoactivated surface it could be significantly useful to confirm false negative cases. The inhibition assay ruled out the possibility of any cross reactivity or non-specific binding. As SELISA procedure is sensitive, specific and reproducible (intra- and inter-assay CVs were 9.65% and 8.47%) it could be an excellent alternative to conventional ELISA or HELISA procedures.

Microwave-Mediated Rapid Synthesis of Gold Nanoparticles Using Calotropis procera Latex and Study of Optical Properties

We report a microwave-mediated simple and rapid method of gold nanoparticles (GNPs) synthesis using latex of Calotropis procera. UV-Vis spectroscopic studies indicated the formation of GNPs. The GNPs were stable at room temperature (25°C) for six months. Transmission electron microscope (TEM) micrographs of the synthesized GNPs showed the formation of spherical nanoparticles with an average size of 13 ± 5 nm. SAED and XRD confirmed the crystalline nature of GNPs. Fourier transform infrared (FTIR) analysis indicated the presence of organic coating on the nanoparticles. Cytotoxicity of the GNPs was tested on HeLa and A549 and found to be nontoxic which was indicating that latex of Calotropis procera provided the nontoxic coating on GNPs, thus can be used as biomedical and pharmacological applications.

Photoimmobilization of unmodified carbohydrates on activated surface.

Herein, we demonstrate a simple, versatile and efficient method for immobilization of unmodified carbohydrates onto a solid surface. The method employs a photoreactive-cellulose membrane which is prepared by the introduction of a photoreactive group to a cellulose membrane through 1-fluoro-2-nitro-4-azidobenzene (FNAB). Upon exposure to UV light photo-reactive azido group of the activated cellulose membrane transforms itself into highly reactive nitrene which covalently binds to underivatized carbohydrates through an insertion reaction. Maximum immobilization of carbohydrate is achieved at the UV exposure time of 60min and carbohydrate concentration of 100microg/disk. The immobilized carbohydrate is assayed by affinity binding of biotin-labeled lectins. The amount of Con A required for detecting immobilized carbohydrate ranges from 4-10microg/ml. The binding affinities of the lectin to the immobilized carbohydrates are analyzed by calculating their IC(50) values. Overall, the present work demonstrates an efficient immobilization of underivatized carbohydrate onto cellulose surface and has the potential to be applied to other surfaces.

Gold nanoparticles: sonocatalytic synthesis using ethanolic extract of Andrographis paniculata and functionalization with polycaprolactone-gelatin composites

Gold nanoparticles (AuNPs) were synthesized by sonication using ethanolic leaf extract of Andrographis paniculata. We investigated the optimum parameters for AuNP synthesis and functionalization with polycaprolactone-gelatin (PCL-GL) composites. The AuNPs were characterized with various biophysical techniques such as TEM, XRD, FT-IR and EDX spectroscopy. TEM images showed that nanoparticles were spherical in shape with a size range from 5 to 75 nm. EDX analysis revealed the presence of molecular oxygen and carbon on the surface of AuNPs. The synthesized AuNPs were tested for their effect on HeLa (human cervical cancer) and MCF-7 (human breast cancer) cell lines and found to be nontoxic and biocompatible, which are potential carriers for hydrophobic drugs.

Rapid Immobilization of Enzymes onto Solid Supports by Ultrasound Waves

Abstract: Herein, we describe a non-conventional method for immobilization of enzymes onto different solid surfaces using ultrasound as a source of energy. When horseradish peroxidase (HRP) was taken on the surface of an activated support and allowed to float on a sonicator bath operating at a frequency of 40 KHz, it readily started binding itself to the surface. Maximum binding was observed in 10 min whereas a control experiment carried out similarly without ultrasound waves showed insignificant immobilization. Ultrasound wave-mediated immobilization is rapid and reproducible and is better suitable for versatile applications in different fields, including fabrication of enzyme-based biosensors or bioreactors.

Green Synthesis and Characterization of Biocompatible Gold Nanoparticles Using Solanum indicum Fruits

This paper reports on the eco-friendly synthesis of gold nanoparticles (AuNPs) using Solanum indicum fruit extract (SFE). We have evaluated various parameters for synthesis of AuNPs such as SFE (0.03%), HAuCl4 (0.5 mM) and reaction time (20 seconds). The synthesized AuNPs were characterized with different physical techniques such as transmission electron microscopy (TEM), Fourier transform infrared (FT-IR) spectroscopy, X-ray diffraction (XRD) and energy dispersive X-ray spectroscopy (EDX). TEM experiments showed that AuNPs presented an anisotropic shape and size ranging from 5-50nm. FT-IR spectroscopy revealed that biomolecules containing an amine group (-NH2), a carbonyl group, -OH groups and other stabilizing functional groups were adsorbed on the surface of the synthesized AuNPs. EDX showed the presence of the elements on the surface of the AuNPs. The cytotoxicity of the synthesized AuNPs were tested on two different human cancer cell lines, HeLa and MCF-7 and were found to be nontoxic, thus providing an opportunity to be used in biomedical applications.

The mitochondrial genome of Muga silkworm (Antheraea assamensis) and its comparative analysis with other lepidopteran insects

Muga (Antheraea assamensis) is an economically important silkmoth endemic to the states of Assam and Meghalaya in India and is the producer of the strongest known commercial silk. However, there is a scarcity of genomic and proteomic data for understanding the organism at a molecular level. Our present study is on decoding the complete mitochondrial genome (mitogenome) of A. assamensis using next generation sequencing technology and comparing it with other available lepidopteran mitogenomes. Mitogenome of A. assamensis is an AT rich circular molecule of 15,272 bp (A+T content ~80.2%). It contains 37 genes comprising of 13 protein coding genes (PCGs), 22 tRNA and 2 rRNA genes along with a 328 bp long control region. Its typical tRNAMet-tRNAIle-tRNAGln arrangement differed from ancestral insects (tRNAIle-tRNAGln-tRNAMet). Two PCGs cox1 and cox2 were found to have CGA and GTG as start codons, respectively as reported in some lepidopterans. Interestingly, nad4l gene showed higher transversion mutations at intra-species than inter-species level. All PCGs evolved under strong purifying selection with highest evolutionary rates observed for atp8 gene while lowest for cox1 gene. We observed the typical clover-leaf shaped secondary structures of tRNAs with a few exceptions in case of tRNASer1 and tRNATyr where stable DHU and TΨC loop were absent. A significant number of mismatches (35) were found to spread over 19 tRNA structures. The control region of mitogenome contained a six bp (CTTAGA/G) deletion atypical of other Antheraea species and lacked tandem repeats. Phylogenetic position of A. assamensis was consistent with the traditional taxonomic classification of Saturniidae. The complete annotated mitogenome is available in GenBank (Accession No. KU379695). To the best of our knowledge, this is the first report on complete mitogenome of A. assamensis.