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Papers by amulya panda

Additional file 1: Figure S1. Analysis of NATA fluorescence and acrylamide quenching of NATA. a) ... more Additional file 1: Figure S1. Analysis of NATA fluorescence and acrylamide quenching of NATA. a) Fluorescence spectra of NATA incubated in different buffers. b) Sternâ Volmer plots for acrylamide quenching of NATA in presence of different buffers.

Journal of basic and clinical physiology and pharmacology, Jan 10, 2017

Curcumin and nisin have been widely reported for their antibacterial and anticancer potency. Howe... more Curcumin and nisin have been widely reported for their antibacterial and anticancer potency. However, their therapeutic applications are hampered by several factors, which necessitate their development into nanosize ranges for improved delivery and activities. Their incorporation into a single nanosynthesized form may suggest desirable efficacy on parasites. The aim of the study was to assess the ovicidal activity of the curcumin-nisin polylactic acid (PLA) entrapped nanoparticle on the Fasciola eggs and its reproductive toxicity. The nanoparticle was formulated by double emulsion method. The eggs of the adult Fasciola spp. were exposed to different concentrations (0.3125-5 mg/mL) of the nanoparticle to monitor hatchability. Mice were exposed to 0.5 mL of the formulated drug at varying concentrations (10-20 mg/kg) and then sacrificed for sperm morphology assay. The mean particle size, polydispersity index, and drug entrapment efficiency of the formulated drug were 288.4±24.3 nm, 0.2...

European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft für Pharmazeutische Verfahrenstechnik e.V, 2016

Rapidly increasing malignant neoplastic disease demands immediate attention. Several dietary comp... more Rapidly increasing malignant neoplastic disease demands immediate attention. Several dietary compounds have recently emerged as strong anti-cancerous agents. Among, Bromelain (BL), a protease from pineapple plant, was used to enhance its anti-cancerous efficacy using nanotechnology. In lieu of this, hyaluronic acid (HA) grafted PLGA copolymer, having tumor targeting ability, was developed. BL was encapsulated in copolymer to obtain BL-copolymer nanoparticles (NPs) that ranged between 140 to 281nm in size. NPs exhibited higher cellular uptake and cytotoxicity in cells with high CD44 expression as compared with non-targeted NPs. In vivo results on tumor bearing mice showed that NPs were efficient in suppressing the tumor growth. Hence, the formulation could be used as a self-targeting drug delivery cargo for the remission of cancer.

Journal of Nanopharmaceutics and Drug Delivery, 2013

Polylacticcoglycolic acid (PLGA) 50:50 and Eudragit RS 100 nanoparticles entrapping glipizide alo... more Polylacticcoglycolic acid (PLGA) 50:50 and Eudragit RS 100 nanoparticles entrapping glipizide along with excipients were prepared using single emulsion solvent evaporation method. The objective was to develop single oral dose glipizide nano particles for reducing blood sugar level in diabetes induced experimental animals. Incorporation of Polyethylene glycol (PEG) (0.5%), Hydroxypropyl methylcellulose (HPMC) (0.5%) and Tween 20 (0.5%) in the organic phase during particle formulation improved release profile of glipizide from the polymer particles. Entrapment efficiency of glipizide in all the polymeric formulations was around ~70 %. Around 80 % of glipizide was released from both PLGA and Eudragit RS 100 nanoparticles when 0.5% of PEG and Tween 20 were added during preparation. Incorporation of amphiphilic polymer during particle formulation not only improved entrapment efficiency of glipizide but also resulted in uniform stabilized nanoparticles having desired control release characteristics. Both PLGA and Eudragit nanoparticles were biocompatible to SW 480 adenocarcinoma human cell line at concentration ranges from 12.5 to 500 µg/ml. The efficacy of glipizide loaded particle formulations were evaluated in female out breed Wistar rats. Significant reduction of blood glucose level was observed (p ≤ 0.05) for 24 hours from a single oral dose using stabilized nanoparticles formulations.

Protein Expression and Purification, 2015

A large number of cancers express human chorionic gonadotropin (hCG) or its subunits ectopically.... more A large number of cancers express human chorionic gonadotropin (hCG) or its subunits ectopically. Patients harboring such cancers have poor prognosis and adverse survival. PiPP is a monoclonal antibody of high affinity and specificity for hCGb/hCG. Work was carried out to develop a PiPP based recombinant immunotoxin for the immunotherapy of hCG expressing cancers. Recombinant PiPP antibody was constructed in scFv format in which gene encoding the VH and VL domains were joined through a linker. This scFv gene was fused to the gene expressing Pseudomonas exotoxin (PE38), and cloned in a Escherichia coli based expression vector under the control of strong bacteriophage T7 promoter. Immunotoxin conjugating scFv(PiPP) and PE38, was expressed in E. coli as recombinant protein. Recombinant PiPP immunotoxin was purified from the bacterial cell lysate and tested for binding and killing of hCGb expressing lymphoma, T-lymphoblastic leukemia and lung carcinoma cells in vitro. Immunotoxin showed nearly 90% killing on the cells. This is the first ever report on recombinant immunotoxin for binding and cytotoxicity to hCG expressing cancer cells, and thus can be a potential candidate for the immunotherapy of hCG expressing cells.

International Journal of Nanomedicine, 2011

Background: Particulate systems have received increasing attention for oral delivery of biomolecu... more Background: Particulate systems have received increasing attention for oral delivery of biomolecules. The objective of the present study was to prepare submicron particulate formulations of papain for pH-dependent site-specific release using pH-sensitive polymers. Methods: Enteric submicron particle formulations of papain were prepared by w/o/w emulsion solvent evaporation using hydroxypropyl methylcellulose phthalate (HPMCP), Eudragit L100, and Eudragit S100, to avoid gastric inactivation of papain. Results: Smaller internal and external aqueous phase volumes provided maximum encapsulation efficiency (75.58%-82.35%), the smallest particle size (665.6-692.4 nm), and 25%-30% loss of enzyme activity. Release studies in 0.1 N HCl confirmed the gastroresistance of the formulations. The anionic submicron particles aggregated in 0.1 N HCl (ie, gastric pH 1.2) due to protonation of carboxylic groups in the enteric polymer. Aggregates , 500 µm size would not impede gastric emptying. However, at pH. 5.0 (duodenal pH), the submicron particles showed deaggregation due to restoration of surface charge. HPMCP submicron particles facilitated almost complete release of papain within 30 minutes at pH 6.0, while Eudragit L100 and Eudragit S100 particles released 88.82% and 53.00% of papain at pH 6.8 and pH 7.4, respectively, according to the Korsmeyer-Peppas equation. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and fluorescence spectroscopy confirmed that the structural integrity of the enzyme was maintained during encapsulation. Fourier transform infrared spectroscopy revealed entrapment of the enzyme, with powder x-ray diffraction and differential scanning calorimetry indicating an amorphous character, and scanning electron microscopy showing that the submicron particles had a spherical shape. Conclusion: In simulated gastrointestinal pH conditions, the HPMCP, Eudragit L100, and Eudragit S100 submicron particles showed good digestion of paneer and milk protein, and could serve as potential carriers for oral enzyme delivery. Stability studies indicated that formulations with approximately 6% overage would ensure a two-year shelf-life at room temperature.

Protein expression and purification, Jan 12, 2014

Pectate lyase (EC 4.2.2.2) gene from Bacillus subtilis RCK was cloned and expressed in Escherichi... more Pectate lyase (EC 4.2.2.2) gene from Bacillus subtilis RCK was cloned and expressed in Escherichia coli to maximize its production. In addition to soluble fraction, bioactive pectate lyase was also obtained from inclusion body aggregates by urea solubilization and refolding under in vitro conditions. Enzyme with specific activity ∼3194IU/mg and ∼1493IU/mg were obtained from soluble and inclusion bodies (IBs) fraction with recovery of 56% and 74% in terms of activity, respectively. The recombinant enzyme was moderately thermostable (t1/2 60min at 50°C) and optimally active in wider alkaline pH range (7.0-10.5). Interaction of protein with its cofactor CaCl2 was found to stimulate the change in tertiary structure as revealed by near UV CD spectra. Intrinsic tryptophan fluorescence spectra indicated that tryptophan is involved in substrate binding and there might be independent binding of Ca(2+) and polygalacturonic acid to the active site. The recombinant enzyme was found to be capabl...

Vaccine, 2006

Low adjuvanticity of microparticles based vaccine formulation necessitates the use of alum along ... more Low adjuvanticity of microparticles based vaccine formulation necessitates the use of alum along with particles to elicit improved antibody titers from single point immunization. It was observed that antibody response from immunization with admixture of alum and polymer entrapped antigen was dependent on particle size, amount of antigen released during burst phase and dose of microencapsulated antigen. In the animals immunized with polymer entrapped tetanus toxoid (TT) very large particles (50-150 m) did not elicited high antibody titers where as microparticles in the range 2-8 m exhibited remarkable improvement in the antibody response. Very small size particles (<2 m) were also not as effective as 2-8 m size particles for generation of antibody response. Presence of alum improved the immune response by adsorbing the burst released antigen from the particle surfaces. Role of alum in potentiation of immune response from polymer entrapped TT was highly significant at lower dose regimes. Polymer entrapped TT as little as 0.1 Lf when immunized along with alum generated antibody responses superior to those elicited by 10 Lf soluble antigen. Immunization with admixture of particles and alum generated two to three times higher antibody titer than that observed from immunization with similar single doses of alum adsorbed TT. Single point immunization of admixture of particles entrapped TT and alum generated sustained long lasting antibody responses comparable to two divided doses of alum-adsorbed antigen. Superiority of single dose polymeric formulation in comparison to two divided doses of alum adsorbed TT was more evident at lower doses of TT immunization. This reflected the profound synergistic effect of both the adjuvants at lower doses. Affinity of antibodies generated from single point immunization was comparable to that achieved with two doses of alum adsorbed TT immunization. Particle alone elicited more of IgG2a type antibody where as immunization with admixture of alum and particles improved the overall antibody response and more of Th2 type response.

Protein Expression and Purification, 2000

Recombinant human growth hormone (r-hGH) was expressed in Escherichia coli as inclusion bodies. I... more Recombinant human growth hormone (r-hGH) was expressed in Escherichia coli as inclusion bodies. In 10 h of fed-batch fermentation, 1.6 g/L of r-hGH was produced at a cell concentration of 25 g dry cell weight/L. Inclusion bodies from the cells were isolated and purified to homogeneity. Various buffers with and without reducing agents were used to solubilize r-hGH from the inclusion bodies and the extent of solubility was compared with that of 8 M urea as well as 6 M Gdn-HCl. Hydrophobic interactions as well as ionic interactions were found to be the dominant forces responsible for the formation of r-hGH inclusion bodies during its high-level expression in E. coli. Complete solubilization of r-hGH inclusion bodies was observed in 100 mM Tris buffer at pH 12.5 containing 2 M urea. Solubilization of r-hGH inclusion bodies in the presence of low concentrations of urea helped in retaining the existing native-like secondary structures of r-hGH, thus improving the yield of bioactive protein during refolding. Solubilized r-hGH in Tris buffer containing 2 M urea was found to be less susceptible to aggregation during buffer exchange and thus was refolded by simple dilution. The r-hGH was purified by use of DEAE-Sepharose ion-exchange chromatography and the pure monomeric r-hGH was finally obtained by using size-exclusion chromatography. The overall yield of the purified monomeric r-hGH was ϳ50% of the initial inclusion body proteins and was found to be biologically active in promoting growth of rat Nb2 lymphoma cell lines.

YAKUGAKU ZASSHI, 2011

Enteric microspheres formulations of papain were prepared by w/o/w emulsion solvent evaporation u... more Enteric microspheres formulations of papain were prepared by w/o/w emulsion solvent evaporation using hydroxypropyl methylcellulose phthalate (HPMCP), Eudragit L 100 and Eudragit S 100, to avoid gastric inactivation of papain. Smaller internal and external aqueous phase volume provided maximum encapsulation e‹ciency (74.49 79.76 ％), least particle size (52.4 60.2 mm) and 21 26％ loss of enzyme activity. Release studies in 0.1 N HCl conˆrmed the gastro-resistance of formulations. The anionic microspheres, zeta potential between-18.21 and-20.06 mV, aggregated in 0.1 N HCl (i.e., gastric pH 1.2), due to protonation of carboxylic groups of enteric polymer and loss of surface charge with subsequent change in zeta potential. The aggregates being ＜500 mm size would not impede gastric emptying. However, at pH＞5.0 (duodenal pH) the microspheres showed de-aggregation due to restoration of surface charge. HPMCP and Eudragit L 100 microspheres facilitated almost complete release of papain within an hour at pH 6.0 and 6.8, respectively while Eudragit S 100 microspheres released 84.56％ papain at pH 7.4, following Higuchi kinetics. FTIR spectroscopy revealed entrapment of enzyme; PXRD & DSC indicated amorphous character and SEM showed spherical shape of microspheres. In simulated gastro-intestinal pH condition, HPMCP, Eudragit L 100 and Eudragit S 100 microspheres showed good digestion of paneer and milk protein. Thus, enteric microspheres formulations could serve as potential carrier for oral enzyme delivery. Stability studies indicated the formulations with around 5％ overage would ensure 2 years shelf life at room temperature.

Journal of Microencapsulation, 2005

Low encapsulation efficiency, incomplete and erratic release profiles are the most common feature... more Low encapsulation efficiency, incomplete and erratic release profiles are the most common features of controlled released protein delivery systems employing biodegradable polymers. In the present study, lysozyme as a model protein was encapsulated in biodegradable microspheres using solvent evaporation method and the effect of amphiphilic stabilizer, a basic salt and a lyoprotectant on microparticle formulation was evaluated. Incorporation rat serum albumin (RSA) in the internal aqueous phase during emulsion increased the encapsulation efficiency of lysozyme and maintained the bioactivity. Use of NaHCO3 improved the encapsulation efficiency of lysozyme from 15-94%, but at the cost of reduced in vitro release characteristics. Incorporation of both RSA and NaHCO3 improved the bioactivity of lysozyme and decreased burst release of the protein from the polymer particle, but reduced the encapsulation efficiency from 90-70%. Addition of sucrose in the internal aqueous phase lowered the encapsulation efficiency which was restored by its addition in the external aqueous phase. Maintenance of internal aqueous phase pH close to the iso-electric point of the protein and osmotic balance between the internal aqueous phase and the external aqueous phase during solvent evaporation method helped in better encapsulation of the protein drug. In vitro release of the lysozyme correlated with the effect of different excipients on entrapment in polymer matrix. Entrapment efficiency as high as 76%, low burst effect and high bioactivity of the entrapped lysozyme was observed from the polymer particles. Use of RSA, sucrose and NaHCO3 helped in a co-operative way towards the formulation of particles entrapping bioactive lysozyme.

Journal of Indian Society of Periodontology, 2011

The preservation or reduction of alveolar ridge resorption following tooth extraction is importan... more The preservation or reduction of alveolar ridge resorption following tooth extraction is important in patients especially for those intended for implants at a later stage. One way to achieve this is by using membranes, graft materials, and biodegradable space fillers to prevent alveolar bone resorption and promote regeneration. A major attraction for using biodegradable and biocompatible polymers as space fillers for ridge preservation is their safety profile in comparison to xenograft materials like lyophilized bone and collagen. Biocompatible polylactide space fillers were fabricated by fusing porous polylactide particles. The sponges were loaded with drugs by placing them in the respective solutions. Pseudomonas aeruginosa was isolated from a chronic periodontitis patient and in vitro anti-microbial evaluation was done with the drug loaded sponges. Chlorhexidine loaded space filler showed significant anti microbial effect against multiple drug resistant Pseudomonas aeruginosa isolated from a patient with chronic periodontitis. The results of this study indicate that biodegradable drug releasing polylactide space fillers has the potential to be used for ridge preservation following tooth extraction. Release of drugs in the socket may prove useful in preventing development of alveolar osteitis post extraction which can interfere with normal healing of the socket. Synthetic biodegradable polymers also exhibit a controlled degradation rate to achieve complete resorption within the intended time.

Journal of Biotechnology, 1999

A process for maximizing the volumetric productivity of recombinant ovine growth hormone (r-oGH) ... more A process for maximizing the volumetric productivity of recombinant ovine growth hormone (r-oGH) expressed in Escherichia coli during high cell density fermentation process has been devised. Kinetics of r-oGH expression as inclusion bodies and its effect on specific growth rates of E. coli cells were monitored during batch fermentation process. It was observed that during r-oGH expression in E. coli, the specific growth rate of the culture became an intrinsic property of the cells which reduced in a programmed manner upon induction. Nutrient feeding during protein expression phase of the fed-batch process was designed according to the reduction in specific growth rate of the culture. By feeding yeast extract along with glucose during fed-batch operation, high cell growth with very little accumulation of acetic acid was observed. Use of yeast extract helped in maintaining high specific cellular protein yield which resulted in high volumetric productivity of r-oGH. In 16 h of fed-batch fermentation, 3.2 g l − 1 of r-oGH were produced at a cell OD of 124. This is the highest concentration of r-oGH reported to date using E. coli expression system. The volumetric productivity of r-oGH was 0.2 g l − 1 h − 1 , which is also the highest value reported for any therapeutic protein using IPTG inducible expression system in a single stage fed-batch process.

Journal of Biotechnology, 1995

Plasmid content, its stability and the expression of B-subunit of Escherichiu coli heat-labile en... more Plasmid content, its stability and the expression of B-subunit of Escherichiu coli heat-labile enterotoxin (LTB) in Vibrio cholerue/r-pMMB68 system have been studied in batch as well as in chemostat cultures. Upon induction with isopropyl-Po-thiogalactopyranoside (IPTG), cultures secreted LTB into the extracellular milieu. Highest specific LTB production rate of 7.3 mg mg-' h-' was achieved in batch culture induced at the late exponential growth phase. The plasmid pMMB68 was fairly stable up to 20 generations, even in the absence of selection pressure. Instability of the plasmid was accelerated in the presence of IPTG and. at higher dilution rates. Maximum productivity of 2.1 mg 1-l h-' was achieved in continuous culture, which remained constant at a range of dilution rates from 0.20 to 0.35 h-l.

Journal of Biomaterials Applications, 2006

Eudragit L100 microspheres were prepared using water-in-oil-in water (w/o/w) emulsion-solvent eva... more Eudragit L100 microspheres were prepared using water-in-oil-in water (w/o/w) emulsion-solvent evaporation with polysorbate 20 as dispersing agent in the internal aqueous phase, and PVA/PVP as stabilizer in the external aqueous phase. Smaller internal and external aqueous phases provided higher drug encapsulation. The PVA-stabilized microspheres having maximum drug encapsulation (84.5 2.8%) released 7% insulin at pH 1.0 in 2 h. In phosphate buffer (pH 7.4), microspheres showed an initial burst release of 21% in 1 h with additional 35% release in the next 5 h. The smaller the volumes of internal and external aqueous phases, the lower the initial burst release. The release of drug from microspheres followed Higuchi kinetics. Scanning electron microscopy of PVA stabilized microspheres demonstrated spherical particles with smooth surface and laser diffractometry revealed a mean particle size (Vm) of 59.11 30 m.

Gene, 2013

Purification and characterization of a novel high molecular weight alkaline protease produced by ... more Purification and characterization of a novel high molecular weight alkaline protease produced by an endophytic Bacillus halotolerans strain CT2

Expert Review of Vaccines, 2011

Induction of activated T-cell responses is a prerequisite for the development of vaccine against ... more Induction of activated T-cell responses is a prerequisite for the development of vaccine against intracellular infection and for the control of cancer. Particulate nanoscale delivery systems deliver antigens intracellularly and help in inducing T-cell responses. T-cell responses can be further augmented by targeting these particles to dendritic cells, which have the ability to induce both innate and adaptive immune responses. Flagellin, which acts as a TLR-5 ligand, has been extensively explored for its adjuvant activity. The paper under evaluation reports a novel vaccine delivery platform technology for induction of a T-cell response using a nanoscale liposome containing antigen and a small synthetic peptide representing TLR-5-binding motifs of flagellin. Vaccination using this nanodelivery system activated dendritic cells through TLR-5 activation and induced both innate and adaptive immune responses. Such novel delivery systems can improve modern vaccine formulation, particularly for the generation of activated T-cell responses and anti-tumor immunity.

European Journal of Pharmaceutical Sciences, 2012

Polylactide (PLA) polymer particles entrapping diphtheria toxoid (DT) or tetanus toxoid (TT) were... more Polylactide (PLA) polymer particles entrapping diphtheria toxoid (DT) or tetanus toxoid (TT) were formulated with surface coatings of soybean agglutinin to have dendritic cells (DCs) targeting ability through ctype lectin receptors (CLR). It was observed that soybean agglutinin coating resulted in more association of polymer particles with DCs. Immunization with soybean agglutinin coated polymer particles entrapping DT or TT elicited antibody response better than the plain particles entrapping antigens. Both for TT and DT, single point immunization of soybean agglutinin coated polymer particles along with alum resulted in very high antibody titers; much higher than that observed while immunizing with alum adsorbed antigens or admixture of particle entrapped antigens and alum. More interestingly, single point immunization with soybean agglutinin coated polymer particles also elicited very high secondary antibody response which sustained for more than six weeks in mice. Interactions of different polymeric microparticles formulations with DCs correlated with antibody response. Improved primary and sustained secondary antibody response from single point immunization of antigen entrapped soybean agglutinin coated particles was attributed to the N-linked glycan mediated targeting of polymer particles to DCs.

Drug Delivery, 2003

Poly(lactide) (PLA) polymer particles entrapping immunoreactive tetanus toxoid (TT) were used for... more Poly(lactide) (PLA) polymer particles entrapping immunoreactive tetanus toxoid (TT) were used for generation of immune response using single point immunization. Immunization with different sizes of polymer particles encapsulating immunoreactive TT elicited anti-TT antibody titers that persisted for more than 5 months. However, antibody response generated by single point immunization of either nanoparticles or microparticles were lower than the conventional two doses of alum adsorbed TT. To overcome this limitation, alum was used with particles that improved anti-TT antibody response. Immunization with nanoparticles along with alum resulted in very high and early immune response: high anti-TT antibody titers were detected as early as 15 days postimmunization. However anti-TT antibody titers declined rapidly with time. Immunization with admixture of microparticles and alum elicited higher antibody titers than the particles alone and the antibody titers were high particularly during the later part of the postimmunization period. Single point immunization with admixture of PLA microparticles and alum resulted in an antibody response very close to that achieved by two injection of alum-adsorbed TT. Physical mixture of both a nano-and microparticles along with alum resulted in sustained anti-TT antibody response from very early days of postimmunization until 150 days. The antibody titers were maintained around 50 µg/ml for more than 5 months. These results indicated that immune response from polymer particles can be further improved by use of additional adjuvant. Furthermore, using various size particles or physical mixture of different size particles along with alum, it is possible to modulate the kinetics of immune response using polymer particles based immunization.