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Sample records for 1-ethyl-3-3-dimethylaminopropyl carbodiimide edc

  1. Comparison of 1-Ethyl-3-(3-Dimethylaminopropyl) Carbodiimide Based Strategies to Crosslink Antibodies on Amine-Functionalized Platforms for Immunodiagnostic Applications.

    PubMed

    Vashist, Sandeep Kumar

    2012-01-01

    1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) alone, and in combination with N-hydroxysuccinimide (NHS) or sulfoNHS were employed for crosslinking anti-human fetuin A (HFA) antibodies on 3-aminopropyltriethoxysilane (APTES)-functionalized surface plasmon resonance (SPR) gold chip and 96-well microtiter plate. The SPR immunoassay and sandwich enzyme linked immunosorbent immunoassay (ELISA) for HFA clearly demonstrated that EDC crosslinks anti-HFA antibodies to APTES-functionalized bioanalytical platforms more efficiently than EDC/NHS and EDC/sulfoNHS at a normal pH of 7.4. Similar results were obtained by sandwich ELISAs for human Lipocalin-2 and human albumin, and direct ELISA for horseradish peroxidase. The more efficient crosslinking of antibodies by EDC to the APTES-functionalized platforms increased the cost-effectiveness and analytical performance of our immunoassays. This study will be of wide interest to researchers developing immunoassays on APTES-functionalized platforms that are being widely used in biomedical diagnostics, biosensors, lab-on-a-chip and point-of-care-devices. It stresses a critical need of an intensive investigation into the mechanisms of EDC-based amine-carboxyl coupling under various experimental conditions. PMID:26859395

  2. Effect of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide concentrations on the mechanical and biological characteristics of cross-linked collagen fibres for tendon repair

    PubMed Central

    Ahmad, Zafar; Shepherd, Jennifer H.; Shepherd, David V.; Ghose, Siddhartha; Kew, Simon J.; Cameron, Ruth E.; Best, Serena M.; Brooks, Roger A.; Wardale, John; Rushton, Neil

    2015-01-01

    Reconstituted type I collagen fibres have received considerable interest as tendon implant materials due to their chemical and structural similarity to the native tissue. Fibres produced through a semi-continuous extrusion process were cross-linked with different concentrations of the zero-length cross-linker 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) in combination with N-hydroxysuccinimide (NHS). Tensile properties of the fibres were considered, along with imaging of both surface structure and fibrillar alignment. Resistance of the fibres to bacterial collagenase was investigated and fibre sections seeded with human tendon cells for biological characterization, including cell adhesion and proliferation. The work clearly demonstrated that whilst the concentration of EDC and NHS had no significant effect on the mechanics, a higher concentration was associated with higher collagenase resistance, but also provided a less attractive surface for cell adhesion and proliferation. A lower cross-linking concentration offered a more biocompatible material without reduction in mechanics and with a potentially more optimal degradability. PMID:26816633

  3. Characterization of porous collagen/hyaluronic acid scaffold modified by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide cross-linking.

    PubMed

    Park, Si-Nae; Park, Jong-Chul; Kim, Hea Ok; Song, Min Jung; Suh, Hwal

    2002-02-01

    In order to develop a scaffolding material for tissue regeneration, porous matrices containing collagen and hyaluronic acid were fabricated by freeze drying at -20 degrees C, -70 degrees C or -196 degrees C. The fabricated porous membranes were cross-linked using 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) in a range of 1-100 mM concentrations for enhancing mechanical stability of the composite matrix. Scanning electron microscope (SEM) views of the matrices demonstrated that the matrices obtained before cross-linking process had interconnected pores with mean diameters of 40, 90 or 230 microm and porosity of 58-66% according to the freezing temperature, and also the porous structures after cross-linking process were retained. The swelling test and IR spectroscopic measurement of different cross-linked membranes were carried out as a measure of the extent of cross-linking. The swelling behavior of cross-linked membranes showed no significant differences as cross-linking degree increased. FT-IR spectra showed the increase of the intensity of the absorbencies at amide bonds (1655, 1546, 1458 cm(-1)) compared to that of CH bond (2930 cm(-1)). In enzymatic degradation test, EDC treated membranes showed significant enhancement of the resistance to collagenase activity in comparison with 0.625% glutaraldehyde treated membranes. In cytotoxicity test using L929 fibroblastic cells, the EDC-cross-linked membranes demonstrated no significant toxicity. PMID:11791924

  4. Structural changes in photosystem II after treatment with the zero-length bifunctional cross-linker 1-ethyl-3-(3-dimethylaminopropyl)carbodi-imide: an electron microscopic study.

    PubMed Central

    Collins, R F; Flint, T D; Holzenburg, A; Ford, R C

    1996-01-01

    Two-dimensional (2D) crystals of photosystem II (PS II) treated with various concentrations of the zero-length crosslinker 1-ethyl-3-(3-dimethylaminopropyl))carbodi-imide (EDC) were analysed by electron microscopy in conjunction with crystallographic image processing. The preparations were characterized by SDS/PAGE and oxygen-evolution measurements, and the effectiveness of cross-linking was monitored by measuring the level of protection afforded against high concentrations of NaCl and CaCl2, which normally remove extrinsic proteins from PS II. We found that low concentrations of EDC (0.25%) increase the order of 2D crystals of PS II. Treatments with EDC concentrations higher than 0.5% did not improve the order of 2D crystals but induced gross structural changes, which were correlated with a decrease in oxygen evolution activity. Structural changes due to cross-linking did not affect packing or symmetry of the 2D crystals, further supporting the conclusion that PS II has a monomeric nature in vivo. PMID:8912698

  5. Quantum dot-antibody conjugates via carbodiimide-mediated coupling for cellular imaging.

    PubMed

    East, Daniel Alistair; Todd, Michael; Bruce, Ian James

    2014-01-01

    This chapter describes the processes of antibody (Ab) production, purification, conjugation to quantum dots (QDs), and the use of the conjugates produced in intracellular imaging of cell components and structures. Specifically, information is provided on the conjugation of carboxyl surface-terminated QDs to Abs via a one-step reaction using the water-soluble carbodiimide, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC). The chapter details the process of conjugate optimization in terms of its final fluorescence and biological activity. The method described should guarantee the production of QD-Ab conjugates, which outperform classic organic fluorophore-Ab conjugates in terms of both image definition produced and the longevity of the imaging agent. PMID:25103800

  6. Stability of water-soluble carbodiimides in aqueous solution.

    PubMed

    Gilles, M A; Hudson, A Q; Borders, C L

    1990-02-01

    A dimethylbarbituric acid reagent has been used to follow the kinetics of loss of two water-soluble carbodiimides, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and the structurally related 1-ethyl-3-(4-azonia-4,4-dimethylpentyl) carbodiimide (EAC), in aqueous solution as a function of pH and added chemical reagents. In 50 mM 2-(N-morpholino)ethanesulfonic acid at 25 degrees C, EDC has t1/2 values of 37, 20, and 3.9 h at pH 7.0, 6.0, and 5.0, respectively, while the corresponding values for EAC are 12, 2.9, and 0.32 h. Iodide, bromide, or chloride, at 0.1 M, has very little or no effect on carbodiimide stability. However, 0.1 M glycine methyl ester or 0.1 M ethylenediamine causes a significant increase in the rate of loss of EAC and EDC, while the presence of 0.1 M phosphate, 0.1 M hydroxylamine, or 0.01 M ATP decreases the half-lives to less than or equal to 0.4 h at all pH values. PMID:2158246

  7. Preparation of thin film nanofibrous composite NF membrane based on EDC/NHS modified PAN-AA nanofibrous substrate

    NASA Astrophysics Data System (ADS)

    Yang, Y.; Wang, X.; Hsiao, B. S.

    2016-07-01

    A novel kind of thin-film nanofibrous composite (TFNC) nanofiltration (NF) membranes consisting of a polyamide (PA) barrier layer were successfully fabricated by interfacial polymerization (IFP) based on electrospun double-layer nanofibrous substrates, which have an ultrathin poly (acrylonitrile-co-acrylic acid) (PAN-AA) nanofibrous layer as top layer and a thicker polyacrylonitrile (PAN) nanofiber layer as bottom porous support layer. Immersing PAN/PAN-AA nanofibrous substrates into 1-ethyl-(3-3-dimethylaminopropyl) carbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) aqueous solution and piperazine (PIP) aqueous solution (0.20 wt%) sequentially for a period of time, the carboxyl groups on PAN-AA nanofibers were activated by carbodiimide and then reacted with the amide groups. The as prepared composite membrane has an integrated structure with high rejection rate (98.0%); high permeate flux (40.4 L/m2h) for MgSO4 aqueous solution (2 g/L).

  8. Carbodiimide Inactivation of MMPs and Effect on Dentin Bonding

    PubMed Central

    Mazzoni, A.; Apolonio, F.M.; Saboia, V.P.A.; Santi, S.; Angeloni, V.; Checchi, V.; Curci, R.; Di Lenarda, R.; Tay, F.R.; Pashley, D.H.; Breschi, L.

    2014-01-01

    The use of protein cross-linking agents during bonding procedures has been recently proposed to improve bond durability. This study aimed to use zymography and in situ zymography techniques to evaluate the ability of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) cross-linker to inhibit matrix metalloproteinase (MMP) activity. The hypotheses tested were that: (1) bonding procedures increase dentin gelatinolytic activity and (2) EDC pre-treatment prevents this enzymatic activity. The zymographic assay was performed on protein extracts obtained from dentin powder treated with Optibond FL or Scotchbond 1XT with or without 0.3M EDC pre-treatment. For in situ zymography, adhesive/dentin interfaces were created with the same adhesives applied to acid-etched dentin slabs pre-treated or not with EDC conditioner. Zymograms revealed increased expression of dentin endogenous MMP-2 and -9 after adhesive application, while the use of EDC as a primer inactivated dentin gelatinases. Results of in situ zymograpy showed that hybrid layers of tested adhesives exhibited intense collagenolytic activity, while almost no fluorescence signal was detected when specimens were pre-treated with EDC. The correlative analysis used in this study demonstrated that EDC could contribute to inactivate endogenous dentin MMPs within the hybrid layer created by etch-and-rinse adhesives. PMID:24334409

  9. Carbodiimide inactivation of MMPs and effect on dentin bonding.

    PubMed

    Mazzoni, A; Apolonio, F M; Saboia, V P A; Santi, S; Angeloni, V; Checchi, V; Curci, R; Di Lenarda, R; Tay, F R; Pashley, D H; Breschi, L

    2014-03-01

    The use of protein cross-linking agents during bonding procedures has been recently proposed to improve bond durability. This study aimed to use zymography and in situ zymography techniques to evaluate the ability of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) cross-linker to inhibit matrix metalloproteinase (MMP) activity. The hypotheses tested were that: (1) bonding procedures increase dentin gelatinolytic activity and (2) EDC pre-treatment prevents this enzymatic activity. The zymographic assay was performed on protein extracts obtained from dentin powder treated with Optibond FL or Scotchbond 1XT with or without 0.3M EDC pre-treatment. For in situ zymography, adhesive/dentin interfaces were created with the same adhesives applied to acid-etched dentin slabs pre-treated or not with EDC conditioner. Zymograms revealed increased expression of dentin endogenous MMP-2 and -9 after adhesive application, while the use of EDC as a primer inactivated dentin gelatinases. Results of in situ zymograpy showed that hybrid layers of tested adhesives exhibited intense collagenolytic activity, while almost no fluorescence signal was detected when specimens were pre-treated with EDC. The correlative analysis used in this study demonstrated that EDC could contribute to inactivate endogenous dentin MMPs within the hybrid layer created by etch-and-rinse adhesives. PMID:24334409

  10. The process of EDC-NHS cross-linking of reconstituted collagen fibres increases collagen fibrillar order and alignment

    SciTech Connect

    Shepherd, D. V. Shepherd, J. H.; Cameron, R. E.; Best, S. M.; Ghose, S.; Kew, S. J.

    2015-01-01

    We describe the production of collagen fibre bundles through a multi-strand, semi-continuous extrusion process. Cross-linking using an EDC (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide), NHS (N-hydroxysuccinimide) combination was considered. Atomic Force Microscopy and Raman spectroscopy focused on how cross-linking affected the collagen fibrillar structure. In the cross-linked fibres, a clear fibrillar structure comparable to native collagen was observed which was not observed in the non-cross-linked fibre. The amide III doublet in the Raman spectra provided additional evidence of alignment in the cross-linked fibres. Raman spectroscopy also indicated no residual polyethylene glycol (from the fibre forming buffer) or water in any of the fibres.

  11. Long-term effect of carbodiimide on dentin matrix and resin-dentin bonds

    PubMed Central

    Bedran-Russo, Ana Karina B.; Vidal, Cristina M.P.; Dos Santos, Paulo H.; Castellan, Carina S.

    2010-01-01

    Objectives To characterize the interaction of 1-Ethyl-3-[3-dimethylaminopropyl] carbodiimide Hydrochloride (EDC) with dentin matrix and its effect on the resin-dentin bond. Methods Changes to the stiffness of demineralized dentin fragments treated with EDC/N-hydroxysuccinimide (NHS) in different solutions were evaluated at different time points. The resistance against enzymatic degradation was indirectly evaluated by ultimate tensile strength (UTS) test of demineralized dentin treated or not with EDC/NHS and subjected to collagenase digestion. Short- and long-term evaluations of the strength of resin-dentin interfaces treated with EDC/NHS for 1 hour were performed using microtensile bond strength (µTBS) test. All data (MPa) were individually analyzed using ANOVA and Tukey HSD tests (α=0.05). Results The different exposure times significantly increased the stiffness of dentin (p<0.0001, control - 5.15 and EDC/NHS - 29.50), while no differences were observed among the different solutions of EDC/NHS (p=0.063). Collagenase challenge did not affect the UTS values of EDC/NHS group (6.08) (p>0.05), while complete degradation was observed for the control group (p=0.0008, control - 20.84 and EDC/NHS - 43.15). EDC/NHS treatment did not significantly increase resin-dentin µTBS, but the values remained stable after 12 months water storage (p<0.05). Conclusions Biomimetic use of EDC/NHS to induce exogenous collagen cross-links resulted in increased mechanical properties and stability of dentin matrix and dentin-resin interfaces. PMID:20524201

  12. Effect of Carbodiimide on Bonding Durability of Adhesive-cemented Fiber Posts in Root Canals.

    PubMed

    Shafiei, F; Yousefipour, B; Mohammadi-Bassir, M

    2016-01-01

    This study was undertaken to investigate whether using a protein cross-linker, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), improves bonding stability of fiber posts to root dentin using three resin cements. Sixty human maxillary central incisor roots were randomly divided into six groups after endodontic treatment, according to the cements used with and without EDC pretreatment. In the etch-and-rinse group, 0.3 M EDC aqueous solution was applied on acid-etched root dentin prior to Excite DSC/Variolink II for post cementation. In the self-etch and self-adhesive groups, EDC was used on EDTA-conditioned root space prior to application of ED Primer II/Panavia F2.0 and Clearfil SA, respectively. After microslicing the root dentin, a push-out bond strength (BS) test was performed immediately or after one-year of water storage for each group. Data were analyzed using three-way analysis of variance and Tukey tests (α=0.05). A significant effect of cement type, time, EDC, and Time × Cement and Time × EDC interactions were observed (p≤0.001). EDC pretreatment did not affect immediate bonding of the three cements (p>0.05). Aging significantly reduced the BS in all the groups (p≤0.001), but EDC groups exhibited a higher BS compared with the respective control groups (p<0.001). Despite the significant effect of aging on decreasing the BS of fiber post to radicular dentin, EDC could diminish this effect for the three tested cements. PMID:26794191

  13. Carbodiimide Cross-linking Inactivates Soluble and Matrix-bound MMPs, in vitro

    PubMed Central

    Tezvergil-Mutluay, A.; Mutluay, M.M.; Agee, K.A.; Seseogullari-Dirihan, R.; Hoshika, T.; Cadenaro, M.; Breschi, L.; Vallittu, P.; Tay, F.R.; Pashley, D.H.

    2012-01-01

    Matrix metalloproteinases (MMPs) cause collagen degradation in hybrid layers created by dentin adhesives. This in vitro study evaluated the feasibility of using a cross-linking agent, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), to inactivate soluble rhMMP-9, as an example of dentin MMPs, and matrix-bound dentin proteases. The inhibitory effects of 5 EDC concentrations (0.01-0.3 M) and 5 incubation times (1-30 min) on soluble rhMMP-9 were screened with an MMP assay kit. The same EDC concentrations were used to evaluate their inhibitory effects on endogenous proteinases from completely demineralized dentin beams that were incubated in simulated body fluid for 30 days. Decreases in modulus of elasticity (E) and dry mass of the beams, and increases in hydroxyproline content of hydrolysates derived from the incubation medium were used as indirect measures of matrix collagen hydrolysis. All EDC concentrations and pre-treatment times inactivated MMP-9 by 98% to 100% (p < 0.05) compared with non-cross-linked controls. Dentin beams incubated in 0.3 M EDC showed only a 9% decrease in E (45% decrease in control), a 3.6% to 5% loss of dry mass (18% loss in control), and significantly less solubilized hydroxyproline when compared with the control without EDC cross-linking (p < 0.05). It is concluded that EDC application for 1 min may be a clinically relevant and effective means for inactivating soluble rhMMP-9 and matrix-bound dentin proteinases if further studies demonstrate that EDC is not toxic to pulpal tissues. PMID:22058118

  14. The effect of solvent composition on grafting gallic acid onto chitosan via carbodiimide.

    PubMed

    Guo, Ping; Anderson, John D; Bozell, Joseph J; Zivanovic, Svetlana

    2016-04-20

    The primary antioxidant (AOX) activity of chitosan can be introduced by grafting of phenolic compound - gallic acid (GA) to its amino and/or hydroxyl groups. The objective of this study was to investigate the effect of ethanol (EtOH) concentration (0%, 25%, 50%, and 75% in water) on efficiency of grafting GA onto chitosan in the presence of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC)/N-hydroxysuccinimide (NHS). The grafting was confirmed by FTIR and the efficiency was quantified as Folin's total phenolics. When pure deionized water was used as a sole solvent (0% EtOH), GA was grafted to chitosan at the largest extent (285.9mg GA/g chitosan). As the concentration of EtOH increased, the grafting efficiency proportionally decreased. NMR studies showed that EtOH inhibited grafting of GA by prohibiting the production of the intermediate - NHS ester. The results confirm that the concentration of EtOH in grafting solution significantly affects grafting efficiency of GA on chitosan. PMID:26876841

  15. Effect of carbodiimide cross-linking of decellularized porcine pulmonary artery valvular leaflets.

    PubMed

    Xu, Xiu-Fang; Guo, Hai-Ping; Ren, Xue-Jun; Gong, Da; Ma, Jin-Hui; Wang, Ai-Ping; Shi, Hai-Feng; Xin, Yi; Wu, Ying; Li, Wen-Bin

    2014-01-01

    Decellularization provides low immunogenicity and is only slightly subject to calcification in tissue engineering. However, the mechanical properties of the tissues are weakened after decellularization. We adopted cross-linking agent 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) to treat decellularized porcine pulmonary artery valvular leaflets to improve their mechanical properties. Twenty porcine pulmonary artery valvular leaflets were divided into three groups: the fresh control group A, group B treated with trypsin and Triton X-100 to remove cells, and group C cross-linked with EDC after decellularization. All samples were evaluated the physical and mechanical properties and were then subcutaneously embedded in rabbits. These valvular leaflets were removed after 1, 2, or 4 weeks and checked for pathological changes. The cells of the valvular leaflets were completely removed. The thickness of the valvular leaflets was thinner in group B than in group A (P<0.01). In the subcutaneous embedding of the group B samples, there was mild immunological response after 1-2 weeks, and parts of the scaffolds were degraded. After 4 weeks, fibroblasts had grown into the scaffolds. In group C, there was an increase in the tensile strength and thermal shrinkage temperature in group C compared with group B (P<0.01). In subcutaneous embedding of the group C samples, there was a mild immunological response after 1-2 weeks. The fibroblasts had grown into the samples. The EDC-based cross-linking procedure can enhance the tensile strength of decellularized pulmonary artery valvular leaflets and both decrease the valvular leaflets' rejection and promote tissue regeneration in vivo. PMID:24753759

  16. The Anti-calcification Effect of Dithiobispropionimidate, Carbodiimide and Ultraviolet Irradiation Cross-linking Compared to Glutaraldehyde in Rabbit Implantation Models

    PubMed Central

    Park, Samina; Kim, Soo Hwan; Lim, Hong-Gook; Lim, Cheong

    2013-01-01

    Background Glutaraldehyde (GA) is a widely used cross-linking agent for improving mechanical properties and resistance to enzymatic degradation of collagenous tissue, but it has several drawbacks such as calcification and cytotoxicity. The aim of this study was to find the alternative effective cross-linking methods to GA. Materials and Methods Bovine pericardium was processed with GA with ethanol+octanol and glycine detoxification, and polyethylene glycol (PG) space filler, dimethyl 3,3'-dithiobispropionimidate (DTBP), 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) treatment, and the physical fixation of ultraviolet irradiation were done. The biologic material properties of variously treated pericardial tissues were assessed by biochemical, mechanical and histological tests. Treated pericardial tissues were also implanted subcutaneously or intramuscularly into the rabbit for 10 weeks to assess the xenoreactive antibody response of immunoglobulin G and M, their anti-calcification effect. Results The biochemical and mechanical properties of EDC fixed pericardial tissues were comparable to the GA fixed tissue. The cytotoxicity was lowest in space filler treated GA fixed group. In rabbit subcutaneous or intramuscular implantation models, decellularization, space filler, EDC treatment group showed significantly lower calcium content than GA only and DTBP treatment group (p<0.05, analysis of variance). The titer of anti Galα1-3Galβ1-4GlcNAc-R antibodies did not change in the postimplantation serial enzyme-linked immunosorbent assay. Hematoxylin and eosin and von Kossa staining showed that decellularization, space filler, EDC, and ultraviolet treatment had less inflammatory cell infiltration and calcium deposits. Conclusion The decellularization process, PG filler, and EDC treatments are good alternative cross-linking methods compared to GA only fixation and primary amine of DTBP treatment for cardiovascular xenograft preservation in terms of the collagen cross

  17. Comparison of EDC and DMTMM efficiency in glycoconjugate preparation.

    PubMed

    Farkaš, Pavol; Čížová, Alžbeta; Bekešová, Slávka; Bystrický, Slavomír

    2013-09-01

    A series of mono- or disaccharide-protein conjugates were prepared by amide bond formation. Incorporation efficiency of the aminosaccharides on bovine serum albumin was determined by colorimetry and matrix assisted laser desorption/ionization mass spectrometry. Study compares two amide bond coupling agents, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride. Paper demonstrates a practical usefulness of natural bovine serum albumin succinylation. Large number of its carboxyl groups results in its superior reactivity towards simple aminosaccharides. PMID:23791662

  18. New method and characterization of self-assembled gelatin-oleic nanoparticles using a desolvation method via carbodiimide/N-hydroxysuccinimide (EDC/NHS) reaction.

    PubMed

    Park, Chulhun; Vo, Chau Le-Ngoc; Kang, Taehee; Oh, Euichaul; Lee, Beom-Jin

    2015-01-01

    In this study, we investigated a new method for the preparation of gelatin-oleic conjugate (GOC) as an amphiphilic biomaterial to load model anti-cancer drugs into self-assembled nanoparticles (NPs). Oleic acid (OA) was covalently bound to gelatin via carbodiimide/N-hydroxysuccinimide (EDC/NHS) reaction in water-ethanol cosolvent to form a GOC. Fourier transform infrared (FT-IR) spectroscopy and proton nuclear magnetic resonance ((1)H NMR) clearly indicated the successful synthesis of GOC. The percentage of gelatin amino groups reacted with OA was up to 50% as determined using the 2,4,6-trinitrobenzene sulfonic acid (TNBS) method. Subsequently, gelatin-oleic nanoparticles (GONs) were prepared using a desolvation method with glutaraldehyde or genipin used as a crosslinker for comparison. Irinotecan hydrochloride (IRT) was used as a model drug to load into GONs using incubation or an in-process adding method for comparison. Dynamic light scattering (DLS) and transmission electron microscopy (TEM) data showed that the sizes of GONs and IRT-loaded GONs (IRT-GONs) were below 250 nm. The zeta potentials of the GONs and irinotecan-loaded IRT-GONs were below -20 mV, which was found to be stable in suspension against the aggregation process. The incubation method was more suitable for drug loading because it did not affect the process of GON formation and thus did not increase their size much compared to the change in size with the in-process adding method. The lipophilic property of the oleic moiety in the GOC increased the affinity between GOC molecules, thus reducing the amount of crosslinking agents needed to stabilize GONs compared to gelatin nanoparticles (GNs). As novel approaches for the synthesis of protein-fatty acid complexes, chemical reaction has been suggested for the synthesis of GOC. The above results show that GOC synthesized via new method is a promising biomaterial based upon preparation of nanoparticles. PMID:25536111

  19. Amperometric determination of hydrogen peroxide by functionalized carbon nanotubes through EDC/NHS coupling chemistry.

    PubMed

    Jeykumari, D R Shobha; Narayanan, S Sriman

    2007-06-01

    The electrochemistry of the redox mediator Toluidine blue (TB) which was covalently linked to the carboxyl group of the multiwalled carbon nanotubes (MWNTs) by coupling reactions, in which N-hydroxysuccinimide was used to assist 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride catalyzed amidation reaction is described. The results from cyclic voltammetry (CV) and amperometry suggested that the redox mediator is linked to the surface of the MWNTs and the nanotubes showed an obvious promotion for the direct electron-transfer between the redox mediator and the electrode. A couple of well-defined redox peak of TB was observed in a phosphate buffer solution (pH 7.0). The redox mediator immobilized to MWNTs exhibits remarkable electrocatalytic activity for the reduction of hydrogen peroxide (H2O2). The analytical applicability of the modified electrode for the determination of hydrogen peroxide was examined. A linear response in the concentration range of 6.8 x 10(-7)-3.4 x 10(-2) M (r = 0.9958) was obtained with detection limit of 3.4 x 10(-7) M for the determination of hydrogen peroxide. The modified electrode has advantages of being highly stable, sensitive, ease of construction and use. PMID:17654948

  20. The carboxyl modifier 1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide (EDC) inhibits half of the high-affinity Mn-binding site in photosystem II membrane fragments

    SciTech Connect

    Preston, C.; Seibert, M. )

    1991-10-08

    The diphenylcarbazide (DPC)/Mn{sup 2+} assay was used to assess the amount of the high-affinity Mn-binding site in manganese-depleted photosystem II (PS II) membrane fragments from spinach and Scenedesmus obliquus. The assay mechanism at high DPC concentration was shown to involve noncompetitive inhibition of only half of the control level of DPC donation to PS II by micromolar concentrations of Mn at pH 6.5. At low DPC concentration both DPC and Mn{sup 2+} donate to PS II additively. Treatment with the carboxyl amino acid modifier 1-ethyl-3-(3(dimethylamino) propyl) carbodiimide (EDC) inhibited half of the high affinity Mn-binding site in spinach and Scenedesmus WT PS II membranes and all of the available site in Scenedesmus LF-1 mutant PS II membranes. A similar EDC concentration dependence was observed in all cases. This protection was specific for Mn{sup 2+}; six other divalent cations were ineffective. The authors conclude that EDC modifies that half of the high-affinity Mn-binding site that is insensitive to the histidine modifier diethyl pyrocarbonate (DEPC) and directly affects ligands that bind Mn. The effects of EDC and DEPC that influence the high-affinity site are mutually exclusive and are specific to the lumenal side of the PS II membrane. They suggest that carboxyl residues on reaction center proteins are associated with half of the high-affinity Mn-binding site in PS II and are involved along with histidine residues in binding Mn functional in the O{sub 2}-evolving process.

  1. Production and Characterization of Chemically Inactivated Genetically Engineered Clostridium difficile Toxoids.

    PubMed

    Vidunas, Eugene; Mathews, Antony; Weaver, Michele; Cai, Ping; Koh, Eun Hee; Patel-Brown, Sujata; Yuan, Hailey; Zheng, Zi-Rong; Carriere, Marjolaine; Johnson, J Erik; Lotvin, Jason; Moran, Justin

    2016-07-01

    A recombinant Clostridium difficile expression system was used to produce genetically engineered toxoids A and B as immunogens for a prophylactic vaccine against C. difficile-associated disease. Although all known enzymatic activities responsible for cytotoxicity were genetically abrogated, the toxoids exhibited residual cytotoxic activity as measured in an in vitro cell-based cytotoxicity assay. The residual cytotoxicity was eliminated by treating the toxoids with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide. Mass spectrometry and amino acid analysis of the EDC-inactivated toxoids identified crosslinks, glycine adducts, and β-alanine adducts. Surface plasmon resonance analysis demonstrated that modifications resulting from the chemical treatment did not appreciably affect recognition of epitopes by both toxin A- and B-specific neutralizing monoclonal antibodies. Compared to formaldehyde-inactivated toxoids, the EDC/N-hydroxysuccinimide-inactivated toxoids exhibited superior stability in solution with respect to reversion of cytotoxic activity. PMID:27233688

  2. Oxidized SWCNT chemically attached to a modified copper substrate

    NASA Astrophysics Data System (ADS)

    Hernandez, Dionne; Cabrera, Carlos R.; Mendez, Laura; Diaz-Serrano, Madeline; Vega, Omar; Weiner, Brad R.; Rosa, Luis G.

    2015-08-01

    Oxidized single-wall carbon nanotubes were chemically attached to a modified copper substrate by means of a 4-aminothiophenol (4-ATP) self-assembly monolayer (SAM). This bottom up approach offers a low cost route to attach carbon nanotubes by using a condensation reaction. Here, the exposed amine from the 4-ATP SAM reacts with the carboxylic group (COOH) on the CNT. The reaction was followed using ATR-FTIR and the surface was characterized using SEM, XPS, and Raman. The analytical techniques used on this study suggest that the condensation reaction employed using a carbodiimide such as 1-Ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride (EDC) produced SWCNT attached to the copper substrate.

  3. Bioactive and metal uptake studies of carboxymethyl chitosan-graft-D-glucuronic acid membranes for tissue engineering and environmental applications.

    PubMed

    Jayakumar, R; Rajkumar, M; Freitas, H; Sudheesh Kumar, P T; Nair, S V; Furuike, T; Tamura, H

    2009-08-01

    Carboxymethyl chitosan-graft-D-glucuronic acid (CMCS-g-D-GA) was prepared by grafting D-GA onto CMCS in the presence of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and then the membranes were made from it. In this work, the bioactivity studies of CMCS-g-D-GA membranes were carried out and then characterized by SEM, CLSM, XRD and FT-IR. The CMCS-g-D-GA membranes were found to be bioactive. The adsorption of Ni2+, Zn2+ and Cu2+ ions onto CMCS-g-D-GA membranes has also been investigated. The maximum adsorption capacity of CMCS-g-D-GA for Ni2+, Zn2+ and Cu2+ was found to be 57, 56.4 and 70.2 mg/g, respectively. Hence, these membranes were useful for tissue engineering, environmental and water purification applications. PMID:19409415

  4. Growth factors-loaded stents modified with hyaluronic acid and heparin for induction of rapid and tight re-endothelialization.

    PubMed

    Choi, Dong Hoon; Kang, Sung Nam; Kim, Seong Min; Gobaa, Samy; Park, Bang Ju; Kim, Ik Hwan; Joung, Yoon Ki; Han, Dong Keun

    2016-05-01

    Rapid re-endothelialization of damaged vessel lining efficiently prevents restenosis and thrombosis and restores original vascular functions. In this study, we designed a novel metallic stent with a heparin-modified surface and used different methods, including 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and divinyl sulfone (DVS), to load growth factors. First we loaded heparin into a dopamine-conjugated hyaluronic acid (HA) coating to serve as a growth factor reservoir. In a second step, we took advantage of the heparin-binding domain of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) to gain advanced re-endothelialization capabilities. We demonstrated that DVS technique offered higher amount of growth factor loading. In vitro assessment also showed better capillary-like structure formation and localized gap junctions when DVS coating was employed. This study suggested that growth factor loaded stent modified by HA and heparin provided the advantage to rapid and tight restoration of endothelium. PMID:26928466

  5. Antifouling ultrafiltration membranes via post-fabrication grafting of biocidal nanomaterials.

    PubMed

    Mauter, Meagan S; Wang, Yue; Okemgbo, Kaetochi C; Osuji, Chinedum O; Giannelis, Emmanuel P; Elimelech, Menachem

    2011-08-01

    Ultrafiltration (UF) membranes perform critical pre-treatment functions in advanced water treatment processes. In operational systems, however, biofouling decreases membrane performance and increases the frequency and cost of chemical cleaning. The present work demonstrates a novel technique for covalently or ionically tethering antimicrobial nanoparticles to the surface of UF membranes. Silver nanoparticles (AgNPs) encapsulated in positively charged polyethyleneimine (PEI) were reacted with an oxygen plasma modified polysulfone UF membrane with and without 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) present. The nucleophilic primary amines of the PEI react with the electrophilic carboxyl groups on the UF membrane surface to form electrostatic and covalent bonds. The irreversible modification process imparts significant antimicrobial activity to the membrane surface. Post-synthesis functionalization methods, such as the one presented here, maximize the density of nanomaterials at the membrane surface and may provide a more efficient route for fabricating diverse array of reactive nanocomposite membranes. PMID:21736330

  6. The Dimerization State of the Mammalian High Mobility Group Protein AT-Hook 2 (HMGA2)

    PubMed Central

    Frost, Lorraine; Baez, Maria A. M.; Harrilal, Christopher; Garabedian, Alyssa; Fernandez-Lima, Francisco; Leng, Fenfei

    2015-01-01

    The mammalian high mobility group protein AT-hook 2 (HMGA2) is a chromosomal architectural transcription factor involved in cell transformation and oncogenesis. It consists of three positively charged “AT-hooks” and a negatively charged C-terminus. Sequence analyses, circular dichroism experiments, and gel-filtration studies showed that HMGA2, in the native state, does not have a defined secondary or tertiary structure. Surprisingly, using combined approaches of 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) chemical cross-linking, analytical ultracentrifugation, fluorescence resonance energy transfer (FRET), and mass spectrometry, we discovered that HMGA2 is capable of self-associating into homodimers in aqueous buffer solution. Our results showed that electrostatic interactions between the positively charged “AT-hooks” and the negatively charged C-terminus greatly contribute to the homodimer formation. PMID:26114780

  7. Effect of condensation agents and minerals for oligopeptide formation under mild and hydrothermal conditions in related to chemical evolution of proteins

    NASA Astrophysics Data System (ADS)

    Kawamura, Kunio; Takeya, Hitoshi; Kushibe, Takao

    2009-07-01

    The role of condensation agents and minerals for oligopeptide formation was inspected to see whether minerals possess catalytic activity under mild and hydrothermal conditions. Under mild conditions, oligopeptide formation from negatively charged amino acids (Asp and Glu) using different minerals and the elongation of alanine oligopeptides ((Ala) 2-(Ala) 5) were attempted using apatite minerals. Oligo(Asp) up to 10 amino acid units from Asp were observed in the presence of 1-ethyl-3-(3-dimethylaminopropyl carbodiimide (EDC). Notable influence of minerals was not detected for the oligo(Asp) formation. Oligo(Asp) was gradually degraded by the further incubation in the presence of EDC in both the absence and presence of minerals. The formation of oligo(Glu) was less efficient in the presence of carbonyldiimidazole. The elongation from (Ala) 3, (Ala) 4, and (Ala) 5 and the formation of diketopiperazine from (Ala) 2 proceeded immediately in the presence of EDC in the meantime of the sample preparations. In addition, it was unexpected that the disappearance of the products and the reformation of the reactants occurred by the further incubation for 24 h; for instance, (Ala) 5 decreased but (Ala) 4 increased with increasing the reaction time in the reaction of (Ala) 4 with EDC. These facts suggest that the activation of the reactant amino acids or peptides immediately occurs. Under the simulated hydrothermal conditions, EDC did not enhance the formation of oligopeptides from Asp, Glu or Ala nor the spontaneous formation of (Ala) 5 from (Ala) 4.

  8. Controlling Stem Cell-mediated Bone Regeneration through Tailored Mechanical Properties of Collagen Scaffolds

    PubMed Central

    Sun, Hongli; Zhu, Feng; Hu, Qingang; Krebsbach, Paul H.

    2014-01-01

    Mechanical properties of the extracellular matrix (ECM) play an essential role in cell fate determination. To study the role of mechanical properties of ECM in stem cell-mediated bone regeneration, we used a 3D in vivo ossicle model that recapitulates endochondral bone formation. Three-dimensional gelatin scaffolds with distinct stiffness were developed using 1-Ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride (EDC) mediated zero-length crosslinking. The mechanical strength of the scaffolds was significantly increased by EDC treatment, while the microstructure of the scaffold was preserved. Cell behavior on the scaffolds with different mechanical properties was evaluated in vitro and in vivo. EDC-treated scaffolds promoted early chondrogenic differentiation, while it promoted both chondrogenic and osteogenic differentiation at later time points. Both micro-computed tomography and histologic data demonstrated that EDC-treatment significantly increased trabecular bone formation by transplanted cells transduced with AdBMP. Moreover, significantly increased chondrogenesis was observed in the EDC-treated scaffolds. Based on both in vitro and in vivo data, we conclude that the high mechanical strength of 3D scaffolds promoted stem cell mediated bone regeneration by promoting endochondral ossification. These data suggest a new method for harnessing stem cells for bone regeneration in vivo by tailoring the mechanical properties of 3D scaffolds. PMID:24211076

  9. Cytotoxic evaluation of biomechanically improved crosslinked ovine collagen on human dermal fibroblasts.

    PubMed

    Awang, M A; Firdaus, M A B; Busra, M B; Chowdhury, S R; Fadilah, N R; Wan Hamirul, W K; Reusmaazran, M Y; Aminuddin, M Y; Ruszymah, B H I

    2014-01-01

    Earlier studies in our laboratory demonstrated that collagen extracted from ovine tendon is biocompatible towards human dermal fibroblast. To be able to use this collagen as a scaffold in skin tissue engineering, a mechanically stronger scaffold is required that can withstand manipulation before transplantation. This study was conducted to improve the mechanical strength of this collagen sponge using chemical crosslinkers, and evaluate their effect on physical, chemical and biocompatible properties. Collagen sponge was crosslinked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and glutaraldehyde (GA). Tensile test, FTIR study and mercury porosimetry were used to evaluate mechanical properties, chemical property and porosity, respectively. MTT assay was performed to evaluate the cytotoxic effect of crosslinked collagen sponge on human dermal fibroblasts. The FTIR study confirmed the successful crosslinking of collagen sponge. Crosslinking with EDC and GA significantly increased the mechanical strength of collagen sponge, with GA being more superior. Crosslinking of collagen sponge significantly reduced the porosity and the effect was predominant in GA-crosslinked collagen sponge. The GA-crosslinked collagen showed significantly lower, 60% cell viability towards human dermal fibroblasts compared to that of EDC-crosslinked collagen, 80% and non-crosslinked collagen, 100%. Although the mechanical strength was better when using GA but the more toxic effect on dermal fibroblast makes EDC a more suitable crosslinker for future skin tissue engineering. PMID:24948455

  10. Structural analysis of photosystem I polypeptides using chemical crosslinking

    NASA Technical Reports Server (NTRS)

    Armbrust, T. S.; Odom, W. R.; Guikema, J. A.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    Thylakoid membranes, obtained from leaves of 14 d soybean (Glycine max L. cv. Williams) plants, were treated with the chemical crosslinkers glutaraldehyde or 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) to investigate the structural organization of photosystem I. Polypeptides were resolved using lithium dodecyl sulfate polyacrylamide gel electrophoresis, and were identified by western blot analysis using a library of polyclonal antibodies specific for photosystem I subunits. An electrophoretic examination of crosslinked thylakoids revealed numerous crosslinked products, using either glutaraldehyde or EDC. However, only a few of these could be identified by western blot analysis using subunit-specific polyclonal antibodies. Several glutaraldehyde dependent crosslinked species were identified. A single band was identified minimally composed of PsaC and PsaD, documenting the close interaction between these two subunits. The most interesting aspect of these studies was a crosslinked species composed of the PsaB subunit observed following EDC treatment of thylakoids. This is either an internally crosslinked species, which will provide structural information concerning the topology of the complex PsaB protein, a linkage with a polypeptide for which we do not yet have an immunological probe, or a masking of epitopes by the EDC linkage at critical locations in the peptide which is linked to PsaB.

  11. Conjugation behaviours of CdTe quantum dots and antibody by a novel immunochromatographic method.

    PubMed

    Wang, Y; Bai, Y; Wei, X

    2011-03-01

    Three water-soluble CdTe quantum dots (QDs) (green-emitting, yellow-emitting and red-emitting) were synthesised for different refluxing time with 3-mercaptopropionic acid (MPA) as stabiliser. Then the red-emitting CdTe QDs and mouse immunoglobulin G (IgG) were taken as the representative to study the conjugation behaviour of QDs and antibody by a novel immunochromatographic method. After comparing with several methods, that is, direct conjugation, 1-ethyl-3(3-dimethylaminopropyl) carbodiimides hydrochloride (EDC)-mediated conjugation, N-hydroxysuccinimide (NHS)-mediated conjugation, EDC/NHS-mediated conjugation by immunochromatographic strips, EDC and NHS were selected together as coupling agents to conjugate QDs with antibody efficiently. Finally, the K562 leukaemia cells were incubated with the EDC/NHS-mediated conjugates to evaluate the performance in practical application, and the result from fluorescence images showed that it was successfully applied to label cells. The immunochromatographic strip was a superior method to study the conjugation of the fluorophore and antibody. PMID:21241157

  12. Collagen based film with well epithelial and stromal regeneration as corneal repair materials: Improving mechanical property by crosslinking with citric acid.

    PubMed

    Zhao, Xuan; Liu, Yang; Li, Weichang; Long, Kai; Wang, Lin; Liu, Sa; Wang, Yingjun; Ren, Li

    2015-10-01

    Corneal disease can lead to vision loss. It has become the second greatest cause of blindness in the world, and keratoplasty is considered as an effective treatment method. This paper presents the crosslinked collagen (Col)-citric acid (CA) films developed by making use of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS). The results showed that the Col-CA films had necessary optical performance, water content. The collagenase resistance of CA crosslinked films was superior to that of EDC crosslinked films. And CA5 film (Col:CA:EDC:NHS=60:3:10:10) had the best mechanical properties. Cell experiments showed that CA5 film was non-cytotoxic and human corneal epithelial cells could proliferate well on the films. Lamellar keratoplasty showed that the CA5 film could be sutured in the rabbit eyes and was epithelialized completely in about 10 days, and the transparency was restored quickly in 30±5 days. No inflammation and corneal neovascularization were observed at 6 months. Corneal stroma had been repaired; stromal cells and neo-stroma could be seen in the area of operation from the hematoxylin-eosin stained histologic sections and anterior segment optical coherence tomography images. These results indicated that Col-CA films were highly promising biomaterials that could be used in corneal tissue engineering and a variety of other tissue engineering applications. PMID:26117756

  13. ncd and kinesin motor domains interact with both alpha- and beta-tubulin.

    PubMed Central

    Walker, R A

    1995-01-01

    Motor domains of the Drosophila minus-end-directed microtubule (MT) motor protein ncd, were found to saturate microtubule binding sites at a stoichiometry of approximately one motor domain per tubulin dimer. To determine the tubulin subunit(s) involved in binding to ncd, mixtures of ncd motor domain and MTs were treated with the zero-length cross-linker 1-ethyl-3-(3-dimethylaminopropyl-carbodiimide) (EDC). EDC treatment generated covalently cross-linked products of ncd and alpha-tubulin and of ncd and beta-tubulin, indicating that the ncd motor domain interacts with both alpha- and beta-tubulin. When the Drosophila kinesin motor domain protein was substituted for the ncd motor domain, cross-linked products of kinesin and alpha-tubulin and of kinesin and beta-tubulin were produced. EDC treatment of mixtures of ncd motor domain and unassembled tubulin dimers or of kinesin motor domain and unassembled tubulin dimers produced the same motor-tubulin products generated in the presence of MTs. These results indicate that kinesin family motors of opposite polarity interact with both tubulin monomers and support a model in which some portion of each protein's motor domain overlaps adjacent alpha- and beta-tubulin subunits. Images Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:7597061

  14. Evaluation of synthetic schemes to prepare immunogenic conjugates of Vibrio cholerae O139 capsular polysaccharide with chicken serum albumin.

    PubMed

    Kossaczka, Z; Szu, S C

    2000-06-01

    Vibrio cholerae serotype O139 is a new etiologic agent of epidemic cholera. There is no vaccine available against cholera caused by this serotype. V. cholerae O139 is an encapsulated bacterium, and its polysaccharide capsule is an essential virulent factor and likely protective antigen. This study evaluated several synthetic schemes for preparation of conjugates of V. cholerae O139 capsular polysaccharide (CPS) with chicken serum albumin as the carrier protein (CSA) using 1-ethyl-3(3-dimethylaminopropyl)carbodiimide (EDC) or 1-cyano-4-dimethylaminopyridinium tetrafluoroborate (CDAP) as activating agents. Four conjugates described here as representative of many experiments were synthesized in 2 steps: 1) preparation of adipic acid hydrazide derivative of CPS (CPS(AH)) or of CSA (CSA(AH)), and 2) binding of CPS(AH) to CSA or of CPS to CSA(AH). Although all conjugates induced CPS antibodies, the conjugate prepared by EDC-mediated binding of CPS and CSA(AH) (EDC:CPS-CSA(AH)) was statistically significantly less immunogenic than the other three conjugates. Representative sera from mice injected with these three conjugates contained antibodies that mediated the lysis of V. cholerae O139 inoculum. Evaluation of the different synthetic schemes and reaction conditions in relation to the immunogenicity of the resultant conjugates provided the basis for the preparation of a V. cholerae O139 conjugate vaccine with a medically useful carrier protein such as diphtheria toxin mutant. PMID:11294508

  15. Effect of curing time and concentration for a chemical treatment that improves surface gliding for extrasynovial tendon grafts in vitro

    PubMed Central

    Tanaka, Toshikazu; Sun, Yu-Long; Zhao, Chunfeng; Zobitz, Mark E.; An, Kai-Nan; Amadio, Peter C.

    2013-01-01

    The purpose of this study was to evaluate whether treatment time and concentration of these reagents have an effect on the resulting gliding resistance. Forty peroneus longus (PL) tendons were used, from 20 adult mongrel dogs, along with the A2 pulley obtained from the ipsilateral hind paw. After the baseline gliding resistance was measured, the PL tendons were treated with one of three concentrations of hyaluronic acid (HA) and 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride (EDC) or N-hydroxysuccinimide (NHS) mixed with 10% gelatin for various times (5, 30, and 60 min). Tendon friction was measured over 1000 cycles of simulated flexion/extension motion. Gliding resistance of the untreated PL tendons had no significant difference among the groups. After surface treatment with low concentration of HA and EDC/NHS for 5-min cure, the gliding resistance was similar to that of the untreated PL tendon and significantly higher than its 30- and 60-min treatment. For the rest of high concentration of HA and EDC/NHS groups, the gliding resistance was lower than that of untreated PL tendon. However, there was no significant difference among the timing points. It is possible to optimize the effect of surface treatment on friction and durability by regulating cure time and concentration of reagents in a canine extrasynovial tendon in vitro. PMID:16752399

  16. Detection of vibrio cholerae O1 by using cerium oxide nanowires - based immunosensor with different antibody immobilization methods

    NASA Astrophysics Data System (ADS)

    Tam, Phuong Dinh; Hoang, Nguyen Luong; Lan, Hoang; Vuong, Pham Hung; Anh, Ta Thi Nhat; Huy, Tran Quang; Thuy, Nguyen Thanh

    2016-05-01

    In this work, we evaluated the effects of different antibody immobilization strategies on the response of a CeO2-nanowires (NWs)-based immunosensor for Vibrio cholerae O1 detection. Accordingly, the changes in the electron-transfer resistance ( R et ) from before to after cells bind to an antibody-modified electrode prepared by using three different methods of antibody immobilization were determined. The values were 16.2%, 8.3%, and 6.65% for the method that utilized protein A, antibodies activated by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxysuccinimide (NHS), and absorption, respectively. Cyclic voltammetry confirmed that the change in the current was highest for the immunosensors prepared using protein A (11%), followed by those prepared with EDC/NHS-activated antibodies (9%), and finally, those prepared through absorption (7.5%). The order of the antibody immobilization strategies in terms of resulting immunosensor detection limit and sensitivity was as follows order: absorption (3.2 × 103 CFU/mL; 45.1 Ω/CFU·mL-1) < EDC/NHS-activated antibody (1.0 × 103 CFU/mL; 50.6 Ω/CFU·mL-1) < protein A (1.0 × 102 CFU/mL; 65.8 Ω/CFU·mL-1). Thus, we confirmed that the protein A - mediated method showed significantly high cell binding efficiencies compared to the random immobilization method.

  17. Development of an ELISA and Immunochromatographic Strip for Highly Sensitive Detection of Microcystin-LR

    PubMed Central

    Liu, Liqiang; Xing, Changrui; Yan, Huijuan; Kuang, Hua; Xu, Chuanlai

    2014-01-01

    A monoclonal antibody for microcystin–leucine–arginine (MC-LR) was produced by cell fusion. The immunogen was synthesized in two steps. First, ovalbumin/ bovine serum albumin was conjugated with 6-acetylthiohexanoic acid using a carbodiimide EDC (1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride)/ NHS (N-hydroxysulfosuccinimide) reaction. After dialysis, the protein was reacted with MC-LR based on a free radical reaction under basic solution conditions. The protein conjugate was used for immunization based on low volume. The antibodies were identified by indirect competitive (ic)ELISA and were subjected to tap water and lake water analysis. The concentration causing 50% inhibition of binding of MC-LR (IC50) by the competitive indirect ELISA was 0.27 ng/mL. Cross-reactivity to the MC-RR, MC-YR and MC-WR was good. The tap water and lake water matrices had no effect on the detection limit. The analytical recovery of MC-LR in the water samples in the icELISA was 94%–110%. Based on this antibody, an immunochromatographic biosensor was developed with a cut-off value of 1 ng/mL, which could satisfy the requirement of the World Health Organization for MC-LR detection in drinking water. This biosensor could be therefore be used as a fast screening tool in the field detection of MC-LR. PMID:25120158

  18. DNA attachment to nanocrystalline diamond films

    NASA Astrophysics Data System (ADS)

    Wenmackers, S.; Christiaens, P.; Daenen, M.; Haenen, K.; Nesládek, M.; van Deven, M.; Vermeeren, V.; Michiels, L.; Ameloot, M.; Wagner, P.

    2005-09-01

    A biochemical method to immobilize DNA on synthetic diamond for biosensor applications is developed. Nanocrystalline diamond is grown using microwave plasma-enhanced chemical vapour deposition. On the hydrogen-terminated surface 10-undecenoic acid is tethered photochemically under 254 nm illumination, followed by 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide crosslinker-mediated attachment of amino modified DNA. The attachment is functionally confirmed by comparison of supernatant fluorescence and gel electrophoresis. The linking procedure allowed for 35 denaturation and rehybridisation steps.

  19. EDC EXPOSURE METHODS

    EPA Science Inventory

    Endocrine disrupter compounds (EDCs) are exogenous agents that interfere with the production, release, transport, metabolism, binding action, or elimination of the natural hormones in the body responsible for the maintenance of homeostasis and regulation of developmental processe...

  20. Endocrine Disrupting Chemicals (EDCs)

    MedlinePlus

    ... Pesticides (use to kill insect pests) —methoxychlor, chlorpyrifos, DDT Fungicides (used to kill fungus) —vinclozolin Herbicides (used ... similar effects in humans. Some EDCs such as DDT, BPA, phthalates, and PCBs can mimic or block ...

  1. Binding of mouse immunoglobulin G to polylysine-coated glass substrate for immunodiagnosis

    NASA Astrophysics Data System (ADS)

    Vashist, Sandeep Kumar; Tewari, Rupinder; Bajpai, Ram Prakash; Bharadwaj, Lalit Mohan; Raiteri, Roberto

    2006-12-01

    We report a method for immobilizing mouse immunoglobulin G (IgG) on polylysine-coated glass substrate for immunodiagnostic applications. Mouse IgG molecules were immobilized on polylysine-coated glass substrate employing 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and protein A. The amino groups of the polylysine-coated glass slide were cross linked to the carboxyl groups of protein A employing EDC crosslinker. Protein A was employed as it binds to the constant Fc region of antibodies keeping their antigen binding sites on the variable F ab region free to bind to antigens. The qualitative analysis of surface immobilized mouse IgG was done by fluorescent microscopy employing fluorescein isothiocyanate (FITC) labeled mouse IgG molecules. The immobilization densities of protein A and mouse IgG were determined by 3, 3', 4, 4'-tetramethyl benzidine (TMB) substrate assay employing horse radish peroxidise labelled molecules and were found to be 130 +/- 17 ng/cm2 and 596 +/- 31 ng/cm2 respectively. The biomolecular coatings analyzed by atomic force microscopy (AFM) were found to be uniform.

  2. Preparation of cross-linked carboxymethyl chitosan for repairing sciatic nerve injury in rats.

    PubMed

    Wang, Gan; Lu, Guangyuan; Ao, Qiang; Gong, Yandao; Zhang, Xiufang

    2010-01-01

    A successful nerve regeneration process was achieved with nerve repair tubes made up of 1-ethyl-3(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) cross-linked carboxymethyl chitosan (CM-chitosan) with improved biodegradability. Chitosan has a very slow degradation rate, while the EDC cross-linked CM-chitosan tubes degraded to 30% of original weight during 8 weeks of incubation in lysozyme solution. In vitro cell culture indicated that the CM-chitosan films presented no cytotoxicity to Schwann cells. From in vivo studies using a 10 mm rat sciatic nerve defect model investigated by histomorphometry analysis, the average diameter of the fibers and the average thickness of myelin sheath in the CM-chitosan tubes were 3.7 +/- 0.33 and 0.33 +/- 0.04 mum, respectively, which demonstrated equivalence to nerve autografts (the current "gold" standard); furthermore, the average fiber density in the CM-chitosan tubes was 20.5 x 10(3)/mm(2), which was similar to that of autografts (21 x 10(3)/mm(2)) and significantly higher than that of common chitosan tubes (15.3 x 10(3)/mm(2)). PMID:19760120

  3. Increasing Mechanical Strength of Gelatin Hydrogels by Divalent Metal Ion Removal

    PubMed Central

    Xing, Qi; Yates, Keegan; Vogt, Caleb; Qian, Zichen; Frost, Megan C.; Zhao, Feng

    2014-01-01

    The usage of gelatin hydrogel is limited due to its instability and poor mechanical properties, especially under physiological conditions. Divalent metal ions present in gelatin such as Ca2+ and Fe2+ play important roles in the gelatin molecule interactions. The objective of this study was to determine the impact of divalent ion removal on the stability and mechanical properties of gelatin gels with and without chemical crosslinking. The gelatin solution was purified by Chelex resin to replace divalent metal ions with sodium ions. The gel was then chemically crosslinked by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC). Results showed that the removal of divalent metal ions significantly impacted the formation of the gelatin network. The purified gelatin hydrogels had less interactions between gelatin molecules and form larger-pore network which enabled EDC to penetrate and crosslink the gel more efficiently. The crosslinked purified gels showed small swelling ratio, higher crosslinking density and dramatically increased storage and loss moduli. The removal of divalent ions is a simple yet effective method that can significantly improve the stability and strength of gelatin hydrogels. The in vitro cell culture demonstrated that the purified gelatin maintained its ability to support cell attachment and spreading. PMID:24736500

  4. Increasing Mechanical Strength of Gelatin Hydrogels by Divalent Metal Ion Removal

    NASA Astrophysics Data System (ADS)

    Xing, Qi; Yates, Keegan; Vogt, Caleb; Qian, Zichen; Frost, Megan C.; Zhao, Feng

    2014-04-01

    The usage of gelatin hydrogel is limited due to its instability and poor mechanical properties, especially under physiological conditions. Divalent metal ions present in gelatin such as Ca2+ and Fe2+ play important roles in the gelatin molecule interactions. The objective of this study was to determine the impact of divalent ion removal on the stability and mechanical properties of gelatin gels with and without chemical crosslinking. The gelatin solution was purified by Chelex resin to replace divalent metal ions with sodium ions. The gel was then chemically crosslinked by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC). Results showed that the removal of divalent metal ions significantly impacted the formation of the gelatin network. The purified gelatin hydrogels had less interactions between gelatin molecules and form larger-pore network which enabled EDC to penetrate and crosslink the gel more efficiently. The crosslinked purified gels showed small swelling ratio, higher crosslinking density and dramatically increased storage and loss moduli. The removal of divalent ions is a simple yet effective method that can significantly improve the stability and strength of gelatin hydrogels. The in vitro cell culture demonstrated that the purified gelatin maintained its ability to support cell attachment and spreading.

  5. Radio frequency glow discharge-induced acidification of fluoropolymers.

    PubMed

    Krawczyk, Benjamin M; Baltrusaitis, Jonas; Yoder, Colin M; Vargo, Terrence G; Bowden, Ned B; Kader, Khalid N

    2011-12-01

    Fluoropolymer surfaces are unique in view of the fact that they are quite inert, have low surface energies, and possess high thermal stabilities. Attempts to modify fluoropolymer surfaces have met with difficulties in that it is difficult to control the modification to maintain bulk characteristics of the polymer. In a previously described method, the replacement of a small fraction of surface fluorine by acid groups through radio frequency glow discharge created a surface with unexpected reactivity allowing for attachment of proteins in their active states. The present study demonstrates that 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride (EDC) reacts with the acid groups on fluoropolymer surfaces in a novel reaction not previously described. This reaction yields an excellent leaving group in which a primary amine on proteins can substitute to form a covalent bond between a protein and these surfaces. In an earlier study, we demonstrated that collagen IV could be deposited on a modified PTFE surface using EDC as a linker. Once collagen IV is attached to the surface, it assembles to form a functional stratum resembling collagen IV in native basement membrane. In this study, we show data suggesting that the fluorine to carbon ratio determines the acidity of the fluoropolymer surfaces and how well collagen IV attaches to and assembles on four different fluoropolymer surfaces. PMID:21887736

  6. Label Free QCM Immunobiosensor for AFB1 Detection Using Monoclonal IgA Antibody as Recognition Element.

    PubMed

    Ertekin, Özlem; Öztürk, Selma; Öztürk, Zafer Ziya

    2016-01-01

    This study introduces the use of an IgA isotype aflatoxin (AF) specific monoclonal antibody for the development of a highly sensitive Quartz Crystal Microbalance (QCM) immunobiosensor for the detection of AF in inhibitory immunoassay format. The higher molecular weight of IgA antibodies proved an advantage over commonly used IgG antibodies in label free immunobiosensor measurements. IgA and IgG antibodies with similar affinity for AF were used in the comparative studies. Sensor surface was prepared by covalent immobilization of AFB1, using self assembled monolayer (SAM) formed on gold coated Quartz Crystal, with 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxy succinimide (EDC/NHS) method using a diamine linker. Nonspecific binding to the surface was decreased by minimizing the duration of EDC/NHS activation. Sensor surface was chemically blocked after AF immobilization without any need for protein blocking. This protein free sensor chip endured harsh solutions with strong ionic detergent at high pH, which is required for the regeneration of the high affinity antibody-antigen interaction. According to the obtained results, the detection range with IgA antibodies was higher than IgG antibodies in QCM immunosensor developed for AFB1. PMID:27529243

  7. Development of indirect competitive ELISA for quantification of mitragynine in Kratom (Mitragyna speciosa (Roxb.) Korth.).

    PubMed

    Limsuwanchote, Supattra; Wungsintaweekul, Juraithip; Keawpradub, Niwat; Putalun, Waraporn; Morimoto, Satoshi; Tanaka, Hiroyuki

    2014-11-01

    Monoclonal antibody (MAb) against mitragynine (MG), an analgesic alkaloid from Kratom leaves (Mitragyna speciosa), was produced. MG was coupled to carrier proteins employing either 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS), a zero-length cross linker or a 5-carbon length glutaraldehyde cross linker. To confirm the immunogenicity, the hapten numbers were determined using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Preparation of the MAb was accomplished by the electrofusion method. Hybridoma 1A6 that was constructed from the fusion between splenocytes of EDC/NHS conjugate immunized mice and SP2/0-Ag14 myeloma cells was selected, cloned twice and expanded. The cross-reactivities (CRs) of this MAb 1A6 with a series of indole alkaloids were 30.54%, 24.83% and 8.63% for speciogynine, paynantheine and mitraciliatine, respectively. Using this MAb, an indirect competitive enzyme-linked immunosorbent assay (icELISA) was developed with a measurement range of 32.92-250 μg/mL. Quantitative analysis of the MG contents in plant samples by icELISA correlated well with the standard high performance liquid chromatography method (R(2)=0.994). The MAb against mitragynine provided a tool for detection of MG in Kratom preparations. PMID:25216455

  8. Enzymatic cross-linking of carboxymethylpullulan grafted with ferulic acid.

    PubMed

    Dulong, Virginie; Hadrich, Ahdi; Picton, Luc; Le Cerf, Didier

    2016-10-20

    Carboxymethylpullulan (CMP) has been modified in a two-step grafting reaction of ferulic acid (FA). Acid adipic dihydrazyde (ADH) was first reacted with FA activated with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC). Then the product of this first reaction was reacted with CMP (activated with EDC). Grafted polysaccharides structure was confirmed by FTIR and (1)H NMR spectroscopy. Analyses by size-exclusion chromatography (SEC) coupling on-line with a multi-angle light scattering detector (MALS), a viscometer and a differential refractive index detector (DRI) (SEC/MALS/DRI/Visco) showed that associations between FA moieties occurred due to hydrophobic interactions. The grafting rates of FA were determined by the Folin-Ciocalteu method and were found between 1.0% and 11.2% (mol/mol anhydroglucose unit). The CMP-FA were then enzymatically cross-linked with laccase from Pleurotus ostreatus. The crosslinking reactions were followed by rheological measurements, demonstrating the influence of laccase concentration on kinetics. Elastic modulus and swelling rates of hydrogels depends on FA content only for low values. PMID:27474545

  9. Ocular biocompatibility of carbodiimide cross-linked hyaluronic acid hydrogels for cell sheet delivery carriers.

    PubMed

    Lai, Jui-Yang; Ma, David Hui-Kang; Cheng, Hsiao-Yun; Sun, Chi-Chin; Huang, Shu-Jung; Li, Ya-Ting; Hsiue, Ging-Ho

    2010-01-01

    Due to its innocuous nature, hyaluronic acid (HA) is one of the most commonly used biopolymers for ophthalmic applications. We recently developed a cell sheet delivery system using carbodiimide cross-linked HA carriers. Chemical cross-linking provides an improvement in stability of polymer gels, but probably causes toxic side-effects. The aim of this study was to investigate the ocular biocompatibility of HA hydrogels cross-linked by 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC). HA discs without cross-linking and glutaraldehyde (GTA) cross-linked HA samples were used for comparison. The disc implants were inserted in the anterior chamber of rabbit eyes for 24 weeks and characterized by slit-lamp biomicroscopy, histology and scanning electron microscopy. The ophthalmic parameters obtained from biomicroscopic examinations were also scored to provide a quantitative grading system. Results of this study showed that the HA discs cross-linked with EDC had better ocular biocompatibility than those with GTA. The continued residence of GTA cross-linked HA implants in the intraocular cavity elicited severe tissue responses and significant foreign body reactions, whereas no adverse inflammatory reaction was observed after contact with non-cross-linked HA or EDC cross-linked HA samples. It is concluded that the cross-linking agent type gives influence on ocular biocompatibility of cell carriers and the EDC-HA hydrogel is an ideal candidate for use as an implantable material in cell sheet delivery applications. PMID:20178691

  10. In-Depth Electrochemical Investigation of Surface Attachment Chemistry via Carbodiimide Coupling.

    PubMed

    Booth, Marsilea Adela; Kannappan, Karthik; Hosseini, Ali; Partridge, Ashton

    2015-07-28

    Aminoferrocene is used as an electroactive indicator to investigate carbodiimide coupling reactions on a carboxylic acid-functionalized self-assembled monolayer. The commonly used attachment chemistry with 1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) is used for surface activation. A number of conditions are investigated, including EDC and NHS concentration, buffer solutions, incubation timing, and aminoferrocene concentration. Ferrocene is a well-documented electroactive species, and the number of surface-bound ferrocene species can be calculated using electrochemical methods. This capability allows determination of optimal conditions, as well as providing a method for comparing and investigating novel carboxylated surfaces. An EDC-mediated procedure with ∼5 mM EDC and NHS (1:1) made in water, with a full acid monolayer, with 250 μM aminoferrocene for 40 min was found to give the highest ferrocene attachment. An application of this is demonstrated for preparing a probe-DNA-coated surface for DNA sensing. By backfilling with aminoferrocene, a differential quantification of the amount of probe DNA available for sensing can be obtained. This provides an elegant method to monitor an important aspect, namely, probe surface characterization, which will be highly useful for biosensing purposes. PMID:26107592

  11. Electrodeposition of chitosan/gelatin/nanosilver: A new method for constructing biopolymer/nanoparticle composite films with conductivity and antibacterial activity.

    PubMed

    Wang, Yifeng; Guo, Xuecheng; Pan, Ruihao; Han, Di; Chen, Tao; Geng, Zenghua; Xiong, Yanfei; Chen, Yanjun

    2015-08-01

    Electrodeposition of chitosan provides a controllable means to simultaneously assemble biological materials and nanoparticles for various applications. Here, we present a new method to construct biopolymer/nanoparticle composite films with conductivity and antibacterial activity by electrodeposition of chitosan/gelatin/nanosilver. Besides, this method can be employed to build biopolymer/nanoparticle composite hydrogels or coatings on various electrodes or conductive substrates. We initially use a simple approach to prepare the aqueous nanosilver that can be well-dispersed in water. Then, the codeposition mixture containing chitosan, gelatin and nanosilver is prepared, and it can be electrodeposited onto different electrodes or conductive substrates in response to imposed electrical signals. After electrodeposition, it is found that the deposited hydrogels and their dried films are smooth and homogeneous due to the elimination of H2 bubbles by addition of H2O2 in electrodeposition process. Importantly, the composite films are strong enough to completely and readily peel from the electrodes after they reacted with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC), which can build a type of biopolymer/nanoparticle film for further applications. Furthermore, the electrodeposition technique is able to offer controllable and convenient method to construct the composite films with diverse shapes. The composite films display improved conductivity and in vitro antibacterial activity against Escherichia coli and Staphylococcus aureus, which may provide attractive applications in biomedical fields such as artificial muscles, skin biomaterials and neuroprosthetic implants. PMID:26042710

  12. Direct competitive chemiluminescence immunoassays based on gold-coated magnetic particles for detection of chloramphenicol.

    PubMed

    Liang, Xiaohui; Fang, Xiangyi; Yao, Manwen; Yang, Yucong; Li, Junfeng; Liu, Hongjun; Wang, Linyu

    2016-02-01

    Direct competitive chemiluminescence immunoassays (CLIA) based on gold-coated magnetic nanospheres (Au-MNPs) were developed for rapid analysis of chloramphenicol (CAP). The Au-MNPs were modified with carboxyl groups and amino groups by 11-mercaptoundecanoic acid (MUA) and cysteamine respectively, and then were respectively conjugated with CAP base and CAP succinate via an activating reaction using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). NSP-DMAE-NHS, a new and effective luminescence reagent, was employed to label anti-CAP antibody (mAb) as a tracer in direct CLIA for CAP detection using a 'homemade' luminescent measurement system that was set up with a photomultiplier tube (PMT) and a photon counting unit linked to a computer. The sensitivities and limits of detection (LODs) of the two methods were obtained and compared according to the inhibition curves. The 50% inhibition concentration (IC50 ) values of the two methods were about 0.044 ng/mL and 0.072 ng/mL respectively and LODs were approximately 0.001 ng/mL and 0.006 ng/mL respectively. To our knowledge, they were much more sensitive than any traditional enzyme-linked immunosorbent assay (ELISA) ever reported. Moreover, the new luminescence reagent NSP-DMAE-NHS is much more sensitive and stable than luminol and its derivatives, contributing to the sensitivity enhancement. PMID:26031849

  13. Three-Dimensional Supermacroporous Carrageenan-Gelatin Cryogel Matrix for Tissue Engineering Applications

    PubMed Central

    Sharma, Archana; Bhat, Sumrita; Vishnoi, Tanushree; Nayak, Vijayashree; Kumar, Ashok

    2013-01-01

    A tissue-engineered polymeric scaffold should provide suitable macroporous structure similar to that of extracellular matrix which can induce cellular activities and guide tissue regeneration. Cryogelation is a technique in which appropriate monomers or polymeric precursors frozen at sub-zero temperature leads to the formation of supermacroporous cryogel matrices. In this study carrageenan-gelatin (natural polymers) cryogels were synthesized by using glutaraldehyde and 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride and N-hydroxysuccinimide (EDC-NHS) as crosslinking agent at optimum concentrations. Matrices showed large and interconnected pores which were in the range of 60–100 μm diameter. Unconfined compression analysis showed elasticity and physical integrity of all cryogels, as these matrices regained their original length after 90% compressing from the original size. Moreover Young's modulus was found to be in the range of 4–11 kPa for the dry cryogel sections. These cryogels also exhibited good in vitro degradation capacity at 37 °C within 4 weeks of incubation. Supermacroporous carrageenan-gelatin cryogels showed efficient cell adherence and proliferation of Cos-7 cells which was examined by SEM. PI nuclear stain was used to observe cell-matrix interaction. Cytotoxicity of the scaffolds was checked by MTT assay which showed that cryogels are biocompatible and act as a potential material for tissue engineering and regenerative medicine. PMID:23936806

  14. The Effect of Sterilization Methods on the Structural and Chemical Properties of Fibrin Microthread Scaffolds.

    PubMed

    Grasman, Jonathan M; O'Brien, Megan P; Ackerman, Kevin; Gagnon, Keith A; Wong, Gregory M; Pins, George D

    2016-06-01

    A challenge for the design of scaffolds in tissue engineering is to determine a terminal sterilization method that will retain the structural and biochemical properties of the materials. Since commonly used heat and ionizing energy-based sterilization methods have been shown to alter the material properties of protein-based scaffolds, the effects of ethanol and ethylene oxide (EtO) sterilization on the cellular compatibility and the structural, chemical, and mechanical properties of uncrosslinked, UV crosslinked, or 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) crosslinked fibrin microthreads in neutral (EDCn) or acidic (EDCa) buffers are evaluated. EtO sterilization significantly reduces the tensile strength of uncrosslinked microthreads. Surface chemistry analyses show that EtO sterilization induces alkylation of EDCa microthreads leading to a significant reduction in myoblast attachment. The material properties of EDCn microthreads do not appear to be affected by the sterilization method. These results significantly enhance the understanding of how sterilization or crosslinking techniques affect the material properties of protein scaffolds. PMID:26847494

  15. Chitosan grafted monomethyl fumaric acid as a potential food preservative.

    PubMed

    Khan, Imran; Ullah, Shafi; Oh, Deog-Hwan

    2016-11-01

    The present study aims at in vitro antibacterial and antioxidant activity evaluation of chitosan modified with monomethyl fumaric acid (MFA) using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) as mediator. Three different kinds of chitosan derivatives Ch-Ds-1,Ch-Ds-2 and Ch-Ds-3 were synthesized by feeding different concentration of MFA. The chemical structures of resulting materials were characterized by (1)H NMR, (13)C NMR, HR-XRD, FT-IR and TNBS assay. The results showed that Ch-Ds-1, Ch-Ds-2 and Ch-Ds-3 were successfully synthesized. The % amino groups of chitosan modified by MFA were evaluated by TNBS assay and ranging from 1.82±0.05% to 7.88±0.04%. All the chitosan derivatives are readily soluble in water and swelled by dimethyl sulfoxide (DMSO), toluene and dimethyl formamide (DMF). The antioxidant activity for all the chitosan derivatives have been significantly improved (P<0.05) compared to the chitosan. Upon antibacterial activity at pH 4.0, all the chitosan derivatives showed significant (P<0.05) antibacterial activity against Gram positive Staphylococcus aureus, Listeria monocytogenes strains and Gram negative Escherichia coli and Salmonella enteritidis strains compared to chitosan. In conclusion, MFA modified chitosan has shown enhanced activities along with solubility, and could be used as a novel food preservative and packaging material for long time food safety and security. PMID:27516253

  16. An Optically-Transparent Aptamer-Based Detection System for Colon Cancer Applications Using Gold Nanoparticles Electrodeposited on Indium Tin Oxide

    PubMed Central

    Ahmadzadeh-Raji, Mojgan; Ghafar-Zadeh, Ebrahim; Amoabediny, Ghasem

    2016-01-01

    In this paper, a label-free aptamer based detection system (apta-DS) was investigated for detecting colon cancer cells. For this purpose, we employed an aptamer specific to colon cancer cells like HCT116 expressing carcinoembryonic antigen (CEA) on their surfaces. Capture aptamers were covalently immobilized on the surface of gold nanoparticles (GNPs) through self-assembly monolayer of 11-mercaptoundecanoic acid (11-MUA) activated with EDC (1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide)/N-hydroxysuccinimide (NHS). The cyclic voltammetry (CV) and chronopotentiometry (CP) methods were used for electrodeposition of GNPs on the surface of indium tin oxide (ITO). In this work, the CV method was also used to demonstrate the conjugation of GNPs and aptamers and identify the cancer cell capturing events. Additionally, Field Emission Scanning Electron Microscopy (FE-SEM) confirmed the deposition of GNPs on ITO and the immobilization of aptamer on the apta-DS. The electrodeposited GNPs played the role of nanoprobes for cancer cell targeting without losing the optical transparency of the ITO substrate. A conventional optical microscope also verified the detection of captured cancer cells. Based on this study’s results relying on electrochemical and optical microscopic methods, the proposed apta-DS is reliable and high sensitive with a LOD equal to 6 cell/mL for colon cancer detection. PMID:27420059

  17. Characterization of bioactive RGD peptide immobilized onto poly(acrylic acid) thin films by plasma polymerization

    NASA Astrophysics Data System (ADS)

    Seo, Hyun Suk; Ko, Yeong Mu; Shim, Jae Won; Lim, Yun Kyong; Kook, Joong-Ki; Cho, Dong-Lyun; Kim, Byung Hoon

    2010-11-01

    Plasma surface modification can be used to improve the surface properties of commercial pure Ti by creating functional groups to produce bioactive materials with different surface topography. In this study, a titanium surface was modified with acrylic acid (AA) using a plasma treatment and immobilized with bioactive arginine-glycine-aspartic acid (RGD) peptide, which may accelerate the tissue integration of bone implants. Both terminals containing the -NH2 of RGD peptide sequence and -COOH of poly(acrylic acid) (PAA) thin film were combined with a covalent bond in the presence of 1-ethyl-3-3-dimethylaminopropyl carbodiimide (EDC). The chemical structure and morphology of AA film and RGD immobilized surface were investigated by X-ray photoelectron spectroscopy (XPS), Fourier transform infrared (FT-IR), atomic force microscopy (AFM), and scanning electron microscopy (SEM). All chemical analysis showed full coverage of the Ti substrate with the PAA thin film containing COOH groups and the RGD peptide. The MC3T3-E1 cells were cultured on each specimen, and the cell alkaline phosphatase (ALP) activity were examined. The surface-immobilized RGD peptide has a significantly increased the ALP activity of MC3T3-E1 cells. These results suggest that the RGD peptide immobilization on the titanium surface has an effect on osteoblastic differentiation of MC3T3-E1 cells and potential use in osteo-conductive bone implants.

  18. Surface plasmon resonance-based immunoassay for procalcitonin.

    PubMed

    Vashist, Sandeep Kumar; Schneider, E Marion; Barth, Eberhard; Luong, John H T

    2016-09-28

    A surface plasmon resonance (SPR) biosensor has been developed for rapid immunoassay of procalcitonin (PCT) with high detection sensitivity and reproducibility. The 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC)-activated protein A (PrA), diluted in 1% (v/v) 3-aminopropyltriethoxysilane (APTES) was dispensed on a KOH-treated Au-coated SPR chip, resulting in the covalent binding of PrA in 30 min. This "single-step" PrA immobilization strategy led to the oriented binding of the anti-PCT antibody (Ab) on a PrA-functionalized gold (Au) chip. The leach-proof immobilization procedure is five-fold faster than conventional counterparts, enabling high detection specificity and reproducibility. The IA detects 4-324 ng mL(-1) of PCT with a limit of detection (LOD) and a limit of quantification (LOQ) of 4.2 ng mL(-1) and 9.2 ng mL(-1), respectively. It was capable of detecting PCT in real sample matrices and patient samples with high precision. The Ab-bound SPR chips were stable for more than five weeks. PMID:27619095

  19. In vitro characterization of MG-63 osteoblast-like cells cultured on organic-inorganic lyophilized gelatin sponges for early bone healing.

    PubMed

    Rodriguez, Isaac A; Saxena, Gunjan; Hixon, Katherine R; Sell, Scott A; Bowlin, Gary L

    2016-08-01

    The development of three-dimensional porous scaffolds with enhanced osteogenic and angiogenic potential would be beneficial for inducing early-stage bone regeneration. Previous studies have demonstrated the advantages of mineralized and nonmineralized acellular 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC) cross-linked gelatin sponges enhanced with preparations rich in growth factors, hydroxyapatite, and chitin whiskers. In this study, those same scaffolds were mineralized and dynamically seeded with MG-63 cells. Cell proliferation, protein/cytokine secretion, and compressive mechanical properties of scaffolds were evaluated. It was found that mineralization and the addition of growth factors increased cell proliferation compared to gelatin controls. Cells on all scaffolds responded in an appropriate bone regenerative fashion as shown through osteocalcin secretion and little to no secretion of bone resorbing markers. However, compressive mechanical properties of cellularized scaffolds were not significantly different from acellular scaffolds. The combined results of increased cellular attachment, infiltration, and bone regenerative protein/cytokine secretion on scaffolds support the need for the addition of a bone-like mineral surface. Cellularized scaffolds containing growth factors reported similar advantages and mechanical values in the range of native tissues present in the early stages of bone healing. These results suggest that the developed composite sponges exhibited cellular responses and mechanical properties appropriate for promoting early bone healing in various applications. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2011-2019, 2016. PMID:27038217

  20. An Optically-Transparent Aptamer-Based Detection System for Colon Cancer Applications Using Gold Nanoparticles Electrodeposited on Indium Tin Oxide.

    PubMed

    Ahmadzadeh-Raji, Mojgan; Ghafar-Zadeh, Ebrahim; Amoabediny, Ghasem

    2016-01-01

    In this paper, a label-free aptamer based detection system (apta-DS) was investigated for detecting colon cancer cells. For this purpose, we employed an aptamer specific to colon cancer cells like HCT116 expressing carcinoembryonic antigen (CEA) on their surfaces. Capture aptamers were covalently immobilized on the surface of gold nanoparticles (GNPs) through self-assembly monolayer of 11-mercaptoundecanoic acid (11-MUA) activated with EDC (1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide)/N-hydroxysuccinimide (NHS). The cyclic voltammetry (CV) and chronopotentiometry (CP) methods were used for electrodeposition of GNPs on the surface of indium tin oxide (ITO). In this work, the CV method was also used to demonstrate the conjugation of GNPs and aptamers and identify the cancer cell capturing events. Additionally, Field Emission Scanning Electron Microscopy (FE-SEM) confirmed the deposition of GNPs on ITO and the immobilization of aptamer on the apta-DS. The electrodeposited GNPs played the role of nanoprobes for cancer cell targeting without losing the optical transparency of the ITO substrate. A conventional optical microscope also verified the detection of captured cancer cells. Based on this study's results relying on electrochemical and optical microscopic methods, the proposed apta-DS is reliable and high sensitive with a LOD equal to 6 cell/mL for colon cancer detection. PMID:27420059

  1. Effectiveness of tobramycin conjugated to iron oxide nanoparticles in treating infection in cystic fibrosis

    NASA Astrophysics Data System (ADS)

    Brandt, Yekaterina I.; Armijo, Leisha M.; Rivera, Antonio C.; Plumley, John B.; Cook, Nathaniel C.; Smolyakov, Gennady A.; Smyth, Hugh D. C.; Osiński, Marek

    2013-02-01

    Cystic fibrosis (CF) is an inherited childhood-onset life-shortening disease. It is characterized by increased respiratory production, leading to airway obstruction, chronic lung infection and inflammatory reactions. The most common bacteria causing persisting infections in people with CF is Pseudomonas aeruginosa. Superparamagnetic Fe3O4 iron oxide nanoparticles (NPs) conjugated to the antibiotic (tobramycin), guided by a gradient of the magnetic field or subjected to an oscillating magnetic field, show promise in improving the drug delivery across the mucus and P. aeruginosa biofilm to the bacteria. The question remains whether tobramycin needs to be released from the NPs after the penetration of the mucus barrier in order to act upon the pathogenic bacteria. We used a zero-length 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride (EDC) crosslinking agent to couple tobramycin, via its amine groups, to the carboxyl groups on Fe3O4 NPs capped with citric acid. The therapeutic efficiency of Fe3O4 NPs attached to the drug versus that of the free drug was investigated in P. aeruginosa culture.

  2. Injectable pullulan hydrogel for the prevention of postoperative tissue adhesion.

    PubMed

    Bang, Sumi; Lee, Eungjae; Ko, Young-Gwang; Kim, Won Il; Kwon, Oh Hyeong

    2016-06-01

    Methods for reducing and preventing postoperative abdominal adhesions have been researched for decades; however, despite these efforts, the formation of postoperative peritoneal adhesions is continuously reported. Adhesions cause serious complications such as postoperative pain, intestinal obstruction, and infertility. Tissue adhesion barriers have been developed as films, membranes, knits, sprays, and hydrogels. Hydrogels have several advantages when used as adhesion barriers, including flexibility, low tissue adhesiveness, biodegradability, and non-toxic degraded products. Furthermore, compared with preformed hydrogels, injectable hydrogels can fill and cover spaces of any shape and do not require a surgical procedure for implantation. In this study, pullulan was modified through reaction with 2,2,6,6-tetramethyl-1-piperidinyloxy (TEMPO) and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) to introduce carboxyl and phenyl groups as crosslinking sites. The grafting of tyramine on pullulan allows crosslinking branches on pullulan backbone. We successfully fabricated pullulan hydrogel with an enzymatic reaction using horseradish peroxidase (HRP) and hydrogen peroxide (H2O2). The chemical structure of modified pullulan was analyzed with ATR-FTIR and (1)H NMR spectroscopies. Rheological properties were tested by measuring storage modulus with varying H2O2, HRP, polymer solution concentrations and tyramine substitution rates. Cell viability and animal tests were performed. The modified pullulan hydrogel is an invaluable advance in anti-adhesion agents. PMID:26879910

  3. Bioconjugate functionalization of thermally carbonized porous silicon using a radical coupling reaction†

    PubMed Central

    Sciacca, Beniamino; Alvarez, Sara D.; Geobaldo, Francesco; Sailor, Michael J.

    2011-01-01

    The high stability of Salonen’s thermally carbonized porous silicon (TCPSi) has attracted attention for environmental and biochemical sensing applications, where corrosion-induced zero point drift of porous silicon-based sensor elements has historically been a significant problem. Prepared by the high temperature reaction of porous silicon with acetylene gas, the stability of this silicon carbide-like material also poses a challenge—many sensor applications require a functionalized surface, and the low reactivity of TCPSi has limited the ability to chemically modify its surface. This work presents a simple reaction to modify the surface of TCPSi with an alkyl carboxylate. The method involves radical coupling of a dicarboxylic acid (sebacic acid) to the TCPSi surface using a benzoyl peroxide initiator. The grafted carboxylic acid species provides a route for bioconjugate chemical modification, demonstrated in this work by coupling propylamine to the surface carboxylic acid group through the intermediacy of pentafluorophenol and 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC). The stability of the carbonized porous Si surface, both before and after chemical modification, is tested in phosphate buffered saline solution and found to be superior to either hydrosilylated (with undecylenic acid) or thermally oxidized porous Si surfaces. PMID:20967329

  4. Biodistribution and pharmacokinetics of uniform magnetite nanoparticles chemically modified with polyethylene glycol

    NASA Astrophysics Data System (ADS)

    Ruiz, A.; Hernández, Y.; Cabal, C.; González, E.; Veintemillas-Verdaguer, S.; Martínez, E.; Morales, M. P.

    2013-11-01

    The influence of polyethylene glycol (PEG) grafting on the pharmacokinetics, biodistribution and elimination of iron oxide nanoparticles is studied in this work. Magnetite nanoparticles (12 nm) were obtained via thermal decomposition of an iron coordination complex as a precursor. Particles were coated with meso-2,3-dimercaptosuccinic acid (DMSA) and conjugated to PEG-derived molecules by 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide (EDC) chemistry. Using a rat model, we explored the nanoparticle biodistribution pattern in blood and in different organs (liver, spleen and lungs) after intravenous administration of the product. The time of residence in blood was measured from the evolution of water proton relaxivities with time and Fe analysis in blood samples. The results showed that the residence time was doubled for PEG coated nanoparticles and consequently particle accumulation in liver and spleen was reduced. Post-mortem histological analyses showed no alterations in the liver and confirm heterogeneous distribution of NPs in the organ, in agreement with magnetic measurements and iron analysis. Finally, by successive magnetic resonance images we studied the evolution of contrast in the liver and measured the absorption, time of residence and excretion of nanoparticles in the liver during a one month period. On the basis of these results we propose different metabolic routes that determine the fate of magnetic nanoparticles.

  5. The synthesis and coupling of photoreactive collagen-based peptides to restore integrin reactivity to an inert substrate, chemically-crosslinked collagen.

    PubMed

    Malcor, Jean-Daniel; Bax, Daniel; Hamaia, Samir W; Davidenko, Natalia; Best, Serena M; Cameron, Ruth E; Farndale, Richard W; Bihan, Dominique

    2016-04-01

    Collagen is frequently advocated as a scaffold for use in regenerative medicine. Increasing the mechanical stability of a collagen scaffold is widely achieved by cross-linking using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS). However, this treatment consumes the carboxylate-containing amino acid sidechains that are crucial for recognition by the cell-surface integrins, abolishing cell adhesion. Here, we restore cell reactivity to a cross-linked type I collagen film by covalently linking synthetic triple-helical peptides (THPs), mimicking the structure of collagen. These THPs are ligands containing an active cell-recognition motif, GFOGER, a high-affinity binding site for the collagen-binding integrins. We end-stapled peptide strands containing GFOGER by coupling a short diglutamate-containing peptide to their N-terminus, improving the thermal stability of the resulting THP. A photoreactive Diazirine group was grafted onto the end-stapled THP to allow covalent linkage to the collagen film upon UV activation. Such GFOGER-derivatized collagen films showed restored affinity for the ligand-binding I domain of integrin α2β1, and increased integrin-dependent cell attachment and spreading of HT1080 and Rugli cell lines, expressing integrins α2β1 and α1β1, respectively. The method we describe has wide application, beyond collagen films or scaffolds, since the photoreactive diazirine will react with many organic carbon skeletons. PMID:26854392

  6. The synthesis and coupling of photoreactive collagen-based peptides to restore integrin reactivity to an inert substrate, chemically-crosslinked collagen

    PubMed Central

    Malcor, Jean-Daniel; Bax, Daniel; Hamaia, Samir W.; Davidenko, Natalia; Best, Serena M.; Cameron, Ruth E.; Farndale, Richard W.; Bihan, Dominique

    2016-01-01

    Collagen is frequently advocated as a scaffold for use in regenerative medicine. Increasing the mechanical stability of a collagen scaffold is widely achieved by cross-linking using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS). However, this treatment consumes the carboxylate-containing amino acid sidechains that are crucial for recognition by the cell-surface integrins, abolishing cell adhesion. Here, we restore cell reactivity to a cross-linked type I collagen film by covalently linking synthetic triple-helical peptides (THPs), mimicking the structure of collagen. These THPs are ligands containing an active cell-recognition motif, GFOGER, a high-affinity binding site for the collagen-binding integrins. We end-stapled peptide strands containing GFOGER by coupling a short diglutamate-containing peptide to their N-terminus, improving the thermal stability of the resulting THP. A photoreactive Diazirine group was grafted onto the end-stapled THP to allow covalent linkage to the collagen film upon UV activation. Such GFOGER-derivatized collagen films showed restored affinity for the ligand-binding I domain of integrin α2β1, and increased integrin-dependent cell attachment and spreading of HT1080 and Rugli cell lines, expressing integrins α2β1 and α1β1, respectively. The method we describe has wide application, beyond collagen films or scaffolds, since the photoreactive diazirine will react with many organic carbon skeletons. PMID:26854392

  7. In Situ Imidazole Activation of Ribonucleotides for Abiotic RNA Oligomerization Reactions

    NASA Astrophysics Data System (ADS)

    Burcar, Bradley T.; Jawed, Mohsin; Shah, Hari; McGown, Linda B.

    2015-06-01

    The hypothesis that RNA played a significant role in the origin of life requires effective and efficient abiotic pathways to produce RNA oligomers. The most successful abiotic oligomerization reactions to date have utilized high-energy, modified, or pre-activated ribonucleotides to generate strands of RNA up to 50-mers in length. In spite of their success, these modifications and pre-activation reactions significantly alter the ribonucleotides in ways that are highly unlikely to have occurred on a prebiotic Earth. This research seeks to address this problem by exploring an aqueous based method for activating the canonical ribonucleotides in situ using 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and imidazole. The reactions were run with and without a montmorillonite clay catalyst and compared to reactions that used ribonucleotides that were pre-activated with imidazole. The effects of pH and ribonucleotide concentration were also investigated. The results demonstrate the ability of in situ activation of ribonucleotides to generate linear RNA oligomers in solution, providing an alternative route to produce RNA for use in prebiotic Earth scenarios.

  8. Quartz crystal microbalance (QCM) as biosensor for the detecting of Escherichia coli O157:H7

    NASA Astrophysics Data System (ADS)

    Thanh Ngo, Vo Ke; Giang Nguyen, Dang; Phuong Uyen Nguyen, Hoang; Tran, Van Man; Nguyen, Thi Khoa My; Phat Huynh, Trong; Lam, Quang Vinh; Dat Huynh, Thanh; Truong, Thi Ngoc Lien

    2014-12-01

    Although Escherichia coli (E. coli) is a commensalism organism in the intestine of humans and warm-blooded animals, it can be toxic at higher density and causes diseases, especially the highly toxic E. coli O157:H7. In this paper a quartz crystal microbalance (QCM) biosensor was developed for the detection of E. coli O157:H7 bacteria. The anti-E. coli O157:H7 antibodies were immobilized on a self-assembly monolayer (SAM) modified 5 MHz AT-cut quartz crystal resonator. The SAMs were activated with 16-mercaptopropanoic acid, in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and ester N-hydroxysuccinimide (NHS). The result of changing frequency due to the adsorption of E. coli O157:H7 was measured by the QCM biosensor system designed and fabricated by ICDREC-VNUHCM. This system gave good results in the range of 102-107 CFU mL-1 E. coli O157:H7. The time of bacteria E. coli O157:H7 detection in the sample was about 50 m. Besides, QCM biosensor from SAM method was comparable to protein A method-based piezoelectric immunosensor in terms of the amount of immobilized antibodies and detection sensitivity.

  9. Preparation and characterization of nanoparticles based on histidine-hyaluronic acid conjugates as doxorubicin carriers.

    PubMed

    Wu, Jing-liang; Liu, Chen-guang; Wang, Xiao-lei; Huang, Zhen-hua

    2012-08-01

    Histidine-hyaluronic acid (His-HA) conjugates were synthesized using hyaluronic acid (HA) as a hydrophilic segment and histidine (His) as hydrophobic segment by 1-ethyl-3(3-dimethylaminopropyl)carbodiimide (EDC) mediated coupling reactions. The structural characteristics of the His-HA conjugates were investigated using (1)H NMR. His-HA nanoparticles (HH-NPs) were prepared based on His-HA conjugates, and the characteristics of HH-NPs were investigated using dynamic light scattering, transmission electron microscopy (TEM), scanning electron microscopy (SEM), and fluorescence spectroscopy. The particles were between 342 and 732 nm in size, depending on the degree of substitution (DS) of the His. TEM and SEM images indicated that the morphology of HH-NPs was spherical in shape. The critical aggregation concentrations of HH-NPs ranged from 0.034 to 0.125 mg/ml, which decreased with an increase in the DS of the His. Images of fluorescence microscopy indicate that HH-NPs were taken up by the cancer cell line (MCF-7), and significantly decreased by competition inhibition of free HA. From the cytotoxicity test, it was found that DOX-loaded HH-NPs exhibited similar dose and time-dependent cytotoxicity against MCF-7 cells with free DOX. PMID:22580754

  10. Effect of hyaluronic acid molecular weight on the morphology of quantum dot-hyaluronic acid conjugates.

    PubMed

    Kim, Jiseok; Park, Kitae; Hahn, Sei Kwang

    2008-01-01

    The morphological analysis of novel quantum dot-hyaluronic acid (QDot-HA) conjugates was carried out with a transmission electron microscope (TEM). Adipic acid dihydrazide-modified HA (HA-ADH) was synthesized and conjugated to quantum dots (QDots) having carboxyl terminal ligands which were activated with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysulfosuccinimide (sulfo-NHS). HA molecules with a molecular weight (MW) of 20K, 234 K and 3000 K were used to investigate the effect of MW on the morphology of QDot-HA conjugates. The TEM micrographs of QDot-HA conjugates showed branched and multi-layered chain type morphology formed by inter- and intra-molecular conjugation of QDots to HA molecules. The size of QDot-HA conjugate increased with the MW of HA. QDot-HA conjugate could be successfully used for real-time bio-imaging of HA derivatives in nude mice. The novel QDot-HA conjugate will be further used to investigate the biological roles of HA with a different MW in the body. PMID:17936350

  11. Hypromellose succinate-crosslinked chitosan hydrogel films for potential wound dressing.

    PubMed

    Jiang, Qiong; Zhou, Wei; Wang, Jun; Tang, Rupei; Zhang, Di; Wang, Xin

    2016-10-01

    The objective of this study was to develop novel hydrogel films based on carboxyl-modified hypromellose-crosslinked chitosan for potential wound dressing. Hypromellose (HPMC) was grafted with succinic acid to yield hypromellose succinate (HPMCS), and then the reinforced hydrogel films of HPMCS-crosslinked chitosan (HPMCS-CS) were prepared through amide bond formation using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N- hydroxysuccinimide (NHS) as a catalyst. Compared to that of blend film, mechanical properties of HPMCS-CS hydrogel films were significantly enhanced both in dry and swollen state. To assess the applicability of HPMCS-CS hydrogel films as wound dressing, the swelling behavior, water vapor transmission rate (WVTR), oxygen permeability, biocompatibility (cytotoxicity and hemolysis), in vitro drug release and bactericidal properties were analyzed. The results indicated that HPMCS-CS hydrogel films with good biocompatibility possess high swelling ratio, proper WVTR, and oxygen permeability, which might accelerate tissue regeneration. Meanwhile, gentamycin sulfate release from drug-loaded HPMCS-CS hydrogel films were sustained, which would help to protect wound from infection. PMID:27222285

  12. Crosslinked chitosan-dextran sulfate nanoparticle for improved topical ocular drug delivery

    PubMed Central

    Chaiyasan, Wanachat; Srinivas, Sangly P.

    2015-01-01

    Purpose To examine the benefits of chitosan-dextran sulfate nanoparticles (CDNs) as a topical ocular delivery system with lutein as a model drug. Methods CDNs were developed by polyelectrolyte complexation of positively charged chitosan (CS) and negatively charged dextran sulfate (DS). 1-Ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) and polyethylene glycol 400 (PEG400) were used as co-crosslinking and stabilizing agents, respectively. The influence of these on the properties of CDNs, including drug release and mucoadhesiveness, was examined. The chemical stability of lutein in CDNs (LCDNs) was also examined. Results Typically, LCDNs showed a spherical shape, possessing a mean size of ~400 nm with a narrow size distribution. The entrapment efficiency of lutein was in the range of 60%–76%. LCDNs possessing a positive surface charge (+46 mV) were found to be mucoadhesive. The release profile of LCDNs followed Higuchi’s square root model, suggesting drug release by diffusion from the polymer matrix. Lutein in LCDNs showed increased chemical stability during storage compared to its solution form. Conclusions These characteristics of CDNs make them suitable for drug delivery to the ocular surface. PMID:26604662

  13. Fast self-assembly kinetics of alkanethiols on gold nanoparticles: simulation and characterization by localized surface plasmon resonance spectroscopy

    NASA Astrophysics Data System (ADS)

    Asiaei, Sasan; Denomme, Ryan C.; Marr, Chelsea; Nieva, Patricia M.; Vijayan, Mathilakath M.

    2012-03-01

    This study demonstrates improved kinetics for the formation of self-assembled monolayers (SAMs) of alkanethiols on gold nanoparticle substrates. A computational model was developed to predict SAM growth kinetics. Based on the predictions from the model, SAMs of 11-mercaptoundecanoic acid (11-MUA) and 1-octanethiol (1-OT) were formed by incubation of gold nanoparticle chips in an ethanolic 10 mM solution within 20 min. The performance of this novel rapid SAM formation protocol was compared with a conventional 24 hour incubation protocol. Binding capacity of the alkanethiol SAM was investigated for a 20 min incubation protocol using biotin-streptavidin. For this purpose, the SAM loaded gold nanoparticle chips were modified with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) to allow attachment of EZ-Link amine PEG3 biotin to the 11-MUA molecules. Binding reactions were monitored in real time using localized surface plasmon resonance (LSPR) spectroscopy. The resulting LSPR absorbance peak shift was comparable to the experimental results for biotin-streptavidin reported in literature. Results of this study suggest that formation of a high quality alkanethiol SAM within 20 min on gold nanoparticles surfaces is possible and could greatly reduce the time and cost compared to conventional 24 h incubation protocols.

  14. VEGF-conjugated alginate hydrogel prompt angiogenesis and improve pancreatic islet engraftment and function in type 1 diabetes.

    PubMed

    Yin, Nina; Han, Yongming; Xu, Hanlin; Gao, Yisen; Yi, Tao; Yao, Jiale; Dong, Li; Cheng, Dejun; Chen, Zebin

    2016-02-01

    Type 1 diabetes was a life-long disease that affected numerous people around the world. Insulin therapy has its limitations that may involve hyperglycemia and heavy burden of patient by repeated dose. Islet transplantation emerged as a promising approach to reach periodical reverse of diabetes, however, transplanted islets suffer from foreign body reaction and lack of nutrition and oxygen supply, especially in the blood-vessel-shortage subcutaneous site which was preferred by patient and surgeon. In this study, we designed and synthesized a vascular endothelial growth factor (VEGF) conjugated alginate material to encapsulate the transplanted islets via 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) reaction, and successful conjugation was confirmed by Nuclear Magnetic Resonance H1 spectrum. The best VEGF concentration (100ng/ml) was determined by the combined studies of the mechanical property and endothelial cell growth assay. In vivo study, conjugated VEGF on alginate exhibited sustained promoting angiogenesis property after subcutaneous transplantation by histology study and islets encapsulated in this material achieved long term therapeutic effect (up to 50days) in the diabetic mice model. In conclusion, this study establishes a simple biomaterial strategy for islet transplantation to enhance islet survival and function, which could be a feasible therapeutic alternative for type 1 diabetes. PMID:26652453

  15. Cellular behaviour of hepatocyte-like cells from nude mouse bone marrow-derived mesenchymal stem cells on galactosylated poly(D,L-lactic-co-glycolic acid).

    PubMed

    Roh, Hyun; Yang, Dae Hyeok; Chun, Heung Jae; Khang, Gilson

    2015-07-01

    Previously, the galactosylation of poly(d,l-lactic-co-glycolic acid) (PLGA) surface was accomplished by grafting allylamine (AA), using inductively coupled plasma-assisted chemical vapour deposition (ICP-CVD) and conjugating lactobionic acid (LA) with AA via 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide (EDC/NHS) activation for hepatic tissue-engineering purposes. As a continuation study, the cellular behaviour of hepatocyte-like cells (HLCs) on the surface of the galactosylated PLGA were investigated. Nude mouse bone marrow-derived mesenchymal stem cells (MSCs) were cultured under hepatogenic conditions and the differentiated cells were characterized by reverse-transcription polymerase chain reaction (RT-PCR), immunofluorescence and periodic acid-Schiff (PAS) staining. Galactosylated PLGA enhanced the proliferation rate of HLCs compared to the control; HLCs on the surface of the sample became aggregated and formed spheroids after 3 days of culture. A large number of cells on the surface of the sample exhibited increased liver-specific functional activities, such as albumin and urea secretions. In addition, multicellular spheroids in the sample strongly expressed phospholyated focal adhesion kinase (pFAK) (cell-matrix interactions), E-cadherin (cell-cell interactions) and connexin 32 (Cox32; gap junction). PMID:23784953

  16. Enhanced stability of catalase covalently immobilized on functionalized titania submicrospheres.

    PubMed

    Wu, Hong; Liang, Yanpeng; Shi, Jiafu; Wang, Xiaoli; Yang, Dong; Jiang, Zhongyi

    2013-04-01

    In this study, a novel approach combing the chelation and covalent binding was explored for facile and efficient enzyme immobilization. The unique capability of titania to chelate with catecholic derivatives at ambient conditions was utilized for titania surface functionalization. The functionalized titania was then used for enzyme immobilization. Titania submicrospheres (500-600 nm) were synthesized by a modified sol-gel method and functionalized with carboxylic acid groups through a facile chelation method by using 3-(3,4-dihydroxyphenyl) propionic acid as the chelating agent. Then, catalase (CAT) was covalently immobilized on these functionalized titania submicrospheres through 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) coupling reaction. The immobilized CAT retained 65% of its free form activity with a loading capacity of 100-150 mg/g titania. The pH stability, thermostability, recycling stability and storage stability of the immobilized CAT were evaluated. A remarkable enhancement in enzyme stability was achieved. The immobilized CAT retained 90% and 76% of its initial activity after 10 and 16 successive cycles of decomposition of hydrogen peroxide, respectively. Both the Km and the Vmax values of the immobilized CAT (27.4 mM, 13.36 mM/min) were close to those of the free CAT (25.7 mM, 13.46 mM/min). PMID:23827593

  17. Transition mode long period grating biosensor with functional multilayer coatings.

    PubMed

    Pilla, Pierluigi; Malachovská, Viera; Borriello, Anna; Buosciolo, Antonietta; Giordano, Michele; Ambrosio, Luigi; Cutolo, Antonello; Cusano, Andrea

    2011-01-17

    We report our latest research results concerning the development of a platform for label-free biosensing based on overlayered Long Period Gratings (LPGs) working in transition mode. The main novelty of this work lies in a multilayer design that allows to decouple the problem of an efficient surface functionalization from that of the tuning in transition region of the cladding modes. An innovative solvent/nonsolvent strategy for the dip-coating technique was developed in order to deposit on the LPG multiple layers of transparent polymers. In particular, a primary coating of atactic polystyrene was used as high refractive index layer to tune the working point of the device in the so-called transition region. In this way, state-of-the-art-competitive sensitivity to surrounding medium refractive index changes was achieved. An extremely thin secondary functional layer of poly(methyl methacrylate-co-methacrylic acid) was deposited onto the primary coating by means of an original identification of selective solvents. This approach allowed to obtain desired functional groups (carboxyls) on the surface of the device for a stable covalent attachment of bioreceptors and minimal perturbation of the optical design. Standard 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide / N-hydrosuccinimide (EDC / NHS) coupling chemistry was used to link streptavidin on the surface of the coated LPG. Highly sensitive real-time monitoring of multiple affinity assays between streptavidin and biotinylated bovine serum albumin was performed by following the shift of the LPGs attenuation bands. PMID:21263591

  18. Protein-functionalized hairy diamond nanoparticles.

    PubMed

    Dahoumane, Si Amar; Nguyen, Minh Ngoc; Thorel, Alain; Boudou, Jean-Paul; Chehimi, Mohamed M; Mangeney, Claire

    2009-09-01

    Diazonium salt chemistry and atom transfer radical polymerization (ATRP) were combined in view of preparing new bioactive hairy diamond nanoparticles containing, or potentially containing, nitrogen-vacancy (NV) fluorescent centers (fluorescent nanodiamonds, or fNDs). fNDs were modified by ATRP initiators using the electroless reduction of the diazonium salt BF(4)(-),(+)N(2)-C(6)H(4)-CH(CH(3))-Br. The strongly bound aryl groups -C(6)H(4)-CH(CH(3))-Br efficiently initiated the ATRP of tert-butyl methacrylate (tBMA) at the surface of the nanodiamonds, which resulted in obtaining ND-PtBMA hybrids. The grafted chain thickness, estimated from X-ray photoelectron spectroscopy (XPS), was found to increase linearly with respect to time before reaching a plateau value of ca. 2 nm. These nanoobjects were further hydrolyzed into ND-PMAA (where PMAA is the poly(methacrylic acid) graft) and further decorated by bovine serum albumin through the 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) coupling procedure. PMID:19634873

  19. Preparation, blood coagulation and cell compatibility evaluation of chitosan-graft-polylactide copolymers.

    PubMed

    Wang, Qi; Liu, Pei; Liu, Peifeng; Gong, Tao; Li, Suming; Duan, Yourong; Zhang, Zhirong

    2014-02-01

    Biodegradable chitosan-graft-polylactide (PLA-CS) copolymers were prepared by the grafting of a poly(L-lactide) (PLLA) or poly(D-lactide) (PDLA) precursor to the backbone of chitosan using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC ⋅ HCl) and N-hydroxysuccinimide (NHS) as a coupling agent. The blood and cell compatibility of the graft copolymers were investigated in comparison to PLLA and PDLA homopolymers. The coagulation properties of PLA-CS were evaluated by hemolysis, plasma recalcification, dynamic blood clotting and protein absorption assays. PLA-CS copolymers present similar hemolysis ratio and plasma recalcification time as PLA, but slower dynamic blood clotting and lower protein absorption. The cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), agar diffusion and lactate dehydrogenase (LDH) experiments. All the samples presented no effect on the viability to cells. Inflammatory cytokine analysis using sandwich ELISAs revealed that PLA-CS would not stimulate inflammatory activity. PMID:24448591

  20. Real-time monitoring of peptide grafting onto chitosan films using capillary electrophoresis.

    PubMed

    Taylor, Danielle L; Thevarajah, Joel J; Narayan, Diksha K; Murphy, Patricia; Mangala, Melissa M; Lim, Seakcheng; Wuhrer, Richard; Lefay, Catherine; O'Connor, Michael D; Gaborieau, Marianne; Castignolles, Patrice

    2015-03-01

    Chitosan, being antimicrobial and biocompatible, is attractive as a cell growth substrate. To improve cell attachment, arginine-glycine-aspartic acid-serine (RGDS) peptides were covalently grafted to chitosan films, through the widely used coupling agents 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC-HCl) and N-hydroxysuccinimide (NHS), via the carboxylic acid function of the RGDS molecule. The grafting reaction was monitored, for the first time, in real time using free-solution capillary electrophoresis (CE). This enabled fast separation and determination of the peptide and all other reactants in one separation with no sample preparation. Covalent RGDS peptide grafting onto the chitosan film surface was demonstrated using solid-state NMR of swollen films. CE indicated that oligomers of RGDS, not simply RGDS, were grafted on the film, with a likely hyperbranched structure. To assess the functional properties of the grafted films, cell growth was compared on control and peptide-grafted chitosan films. Light microscopy and polymerase chain reaction (PCR) analysis demonstrated greatly improved cell attachment to RGDS-grafted chitosan films. PMID:25680633

  1. Conjugation of ampicillin and enrofloxacin residues with bovine serum albumin and raising of polyclonal antibodies against them

    PubMed Central

    Kumar, B. Sampath; Ashok, Vasili; Kalyani, P.; Nair, G. Remya

    2016-01-01

    Aim: The aim of this study is to test the potency of bovine serum albumin (BSA) conjugated ampicillin (AMP) and enrofloxacin (ENR) antigens in eliciting an immune response in rats using indirect competitive enzyme-linked immunosorbent assay (icELISA). Materials and Methods: AMP and ENR antibiotics were conjugated with BSA by carbodiimide reaction using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) as a cross-linker. The successful conjugation was confirmed by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Sprague-Dawley rats were immunized with the conjugates and blood samples were collected serially at 15 days time interval after first immunization plus first booster, second booster, third booster, and the fourth sampling was done 1½ month after the third booster. The antibody titres in the antisera of each antibiotic in all the four immunization cycles (ICs) were determined by an icELISA at various serum dilutions ranging from 1/100 to 1/6400. Results: Analysis of antibiotic-BSA conjugates by sodium dodecyl sulfate polyacrylamide gel electrophoresis and coomassie blue staining revealed high molecular weight bands of 85 kDa and 74 kDa for AMP-BSA and ENR-BSA respectively when compared to 68 kDa band of BSA. Both the antibiotic conjugates elicited a good immune response in rats but comparatively the response was more with AMP-BSA conjugate than ENR-BSA conjugate. Maximum optical density 450 value of 2.577 was recorded for AMP-BSA antisera, and 1.723 was recorded for ENR-BSA antisera at 1/100th antiserum dilution in third IC. Conclusion: AMP and ENR antibiotics proved to be good immunogens when conjugated to BSA by carbodiimide reaction with EDC as crosslinker. The polyclonal antibodies produced can be employed for detecting AMP and ENR residues in milk and urine samples. PMID:27182138

  2. Synthesis and immunological properties of Vi and di-O-acetyl pectin protein conjugates with adipic acid dihydrazide as the linker.

    PubMed Central

    Kossaczka, Z; Bystricky, S; Bryla, D A; Shiloach, J; Robbins, J B; Szu, S C

    1997-01-01

    The Vi capsular polysaccharide of Salmonella typhi, a licensed vaccine for typhoid fever in individuals > or = 5 years old, induces low and short-lived antibodies in children, and reinjection does not elicit booster responses at any age. Its immunogenicity was improved by binding Vi to proteins by using N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP) as a linker. Similar findings were observed with the structurally related, di-O-acetyl derivative of pectin [poly-alpha(1-->4)-D-GalpA] designated OAcP. Protein conjugates of Vi and OAcP were synthesized by carbodiimide-mediated synthesis with adipic acid dihydrazide (ADH) as the linker. Hydrazide groups were introduced into proteins (bovine serum albumin or recombinant Pseudomonas aeruginosa exoprotein A) by treatment with ADH and 1-ethyl-3(3-dimethylaminopropyl carbodiimide (EDC). The resultant adipic acid hydrazide derivatives (AH-proteins), containing 2.3 to 3.4% AH, had antigenic and physicochemical properties similar to those of the native proteins. The AH-proteins were bound to Vi and OAcP by treatment with EDC. The immunogenicity of Vi or OAcP, alone or as protein conjugates, was evaluated in young outbred mice and guinea pigs by subcutaneous injection of 2.5 and 5.0 microg, respectively, of polysaccharide, and antibodies were measured by enzyme-linked immunosorbent assay. All conjugates were significantly more immunogenic than Vi or OAcP alone and induced booster responses with 5- to 25-fold increases of antibodies. Vi conjugates were significantly more immunogenic than their OAcP analogs. A carboxymethyl derivative of yeast beta-glucan enhanced the anti-Vi response elicited by an OAcP conjugate but had no effect on the immunogenicity of Vi or of OAcP alone. Vi and OAcP conjugates synthesized by this scheme will be evaluated clinically. PMID:9169736

  3. Three-dimensional printing of soy protein scaffolds for tissue regeneration.

    PubMed

    Chien, Karen B; Makridakis, Emmanuella; Shah, Ramille N

    2013-06-01

    Fabricating three-dimensional (3D) porous scaffolds with controlled structure and geometry is crucial for tissue regeneration. To date, exploration in printing 3D natural protein scaffolds is limited. In this study, soy protein slurry was successfully printed using the 3D Bioplotter to form scaffolds. A method to verify the structural integrity of resulting scaffolds during printing was developed. This process involved measuring the mass extrusion flow rate of the slurry from the instrument, which was directly affected by the extrusion pressure and the soy protein slurry properties. The optimal mass flow rate for printing soy slurry at 27°C was 0.0072±0.0002 g/s. The addition of dithiothreitol to soy slurries demonstrated the importance of disulfide bonds in forming solid structures upon printing. Resulting Bioplotted soy protein scaffolds were cured using 95% ethanol and post-treated using dehydrothermal treatment (DHT), a combination of freeze-drying and DHT, and chemical crosslinking using 1-ethyl-3-(3 dimethylaminopropyl)carbodiimide (EDC) chemistry. Surface morphologies of the different treatment groups were characterized using scanning electron microscopy. Scaffold properties, including relative crosslink density, mass loss upon rinsing, and compressive modulus revealed that EDC crosslinked scaffolds were the most robust with moduli of approximately 4 kPa. Scaffold geometry (45° and 90° layer rotations) affected the mechanical properties for DHT and EDC crosslinked scaffolds. Seeding efficiency of human mesenchymal stem cells (hMSC) was highest for nontreated and thermally treated scaffolds, and all scaffolds supported hMSC viability over time. PMID:23102234

  4. Development of a simple bioelectrode for the electrochemical detection of hydrogen peroxide using Pichia pastoris catalase immobilized on gold nanoparticle nanotubes and polythiophene hybrid.

    PubMed

    Nandini, Seetharamaiah; Nalini, Seetharamaiah; Sanetuntikul, Jakkid; Shanmugam, Sangaraju; Niranjana, Pathappa; Melo, Jose Savio; Suresh, Gurukar Shivappa

    2014-11-21

    In this paper, a simple and innovative electrochemical hydrogen peroxide biosensor has been proposed using catalase (CATpp) derived from Pichia pastoris as bioelectrocatalyst. The model biocomponent was immobilized on gold nanoparticle nanotubes (AuNPNTs) and polythiophene composite using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide and N-hydroxysuccinimide (EDC-NHS) coupling reagent. In this present work, we have successfully synthesized gold nanoparticles (AuNPs) by ultrasonic irradiation. The tubular gold nanostructures containing coalesced AuNPs were obtained by sacrificial template synthesis. The assembly of AuNPNTs onto the graphite (Gr) electrode was achieved via S-Au chemisorption. The latter was pre-coated with electropolymerized thiophene (PTh) to enable S groups to bind AuNPNTs. The combination of AuNPNTs-PTh, i.e., an inorganic-organic hybrid, provides a stable enzyme immobilization platform. The physical morphology of the fabricated biosensor Gr/PTh/AuNPNTs/EDC-NHS/CATpp was investigated using scanning electron microscopy and energy-dispersive microscopy. The analytical performance of the bioelectrode was examined using cyclic voltammetry, differential pulse voltammetry and chronoamperometry. Operational parameters such as working potential, pH, and thermal stability of the modified electrode were examined. The beneficial analytical characteristics of the proposed electrode were demonstrated. Our results indicate that the Gr/PTh/AuNPNTs/EDC-NHS/CATpp bioelectrode exhibits a wide linear range from 0.05 mM to 18.5 mM of H2O2, fast response time of 7 s, excellent sensitivity of 26.2 mA mM(-1) cm(-2), good detection limit of 0.12 μM and good Michaelis-Menten constant of 1.4 mM. In addition, the bioelectrode displayed good repeatability, high stability and acceptable reproducibility, which can be attributed to the AuNPNTs-PTh composite that provides a biocompatible micro-environment. PMID:25208248

  5. Corneal Stromal Cell Growth on Gelatin/Chondroitin Sulfate Scaffolds Modified at Different NHS/EDC Molar Ratios

    PubMed Central

    Lai, Jui-Yang

    2013-01-01

    A nanoscale modification strategy that can incorporate chondroitin sulfate (CS) into the cross-linked porous gelatin materials has previously been proposed to give superior performance for designed corneal keratocyte scaffolds. The purpose of this work was to further investigate the influence of carbodiimide chemistry on the characteristics and biofunctionalities of gelatin/CS scaffolds treated with varying N-hydroxysuccinimide (NHS)/1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC) molar ratios (0–1) at a constant EDC concentration of 10 mM. Results of Fourier transform infrared spectroscopy and dimethylmethylene blue assays consistently indicated that when the NHS to EDC molar ratio exceeds a critical level (i.e., 0.5), the efficiency of carbodiimide-mediated biomaterial modification is significantly reduced. With the optimum NHS/EDC molar ratio of 0.5, chemical treatment could achieve relatively high CS content in the gelatin scaffolds, thereby enhancing the water content, glucose permeation, and fibronectin adsorption. Live/Dead assays and interleukin-6 mRNA expression analyses demonstrated that all the test samples have good cytocompatibility without causing toxicity and inflammation. In the molar ratio range of NHS to EDC from 0 to 0.5, the cell adhesion ratio and proliferation activity on the chemically modified samples significantly increased, which is attributed to the increasing CS content. Additionally, the materials with highest CS content (0.143 ± 0.007 nmol/10 mg scaffold) showed the greatest stimulatory effect on the biosynthetic activity of cultivated keratocytes. These findings suggest that a positive correlation is noticed between the NHS to EDC molar ratio and the CS content in the biopolymer matrices, thereby greatly affecting the corneal stromal cell growth. PMID:23337203

  6. Magnesium Catalysis for the Hydroboration of Carbodiimides.

    PubMed

    Weetman, Catherine; Hill, Michael S; Mahon, Mary F

    2016-05-17

    A β-diketiminato magnesium alkyl complex, [CH{C(Me)NDipp}2 }MgnBu] (Dipp=2,6-iPr2 C6 H3 ), was shown to be an effective pre-catalyst for the first reported catalytic hydroboration of alkyl- and aryl-substituted carbodiimides with pinacol borane (HBpin). The catalytic reactions proceed under mild conditions to afford the corresponding N-borylated formamidine compounds in good yields. The reactions were observed to proceed through the intermediacy of magnesium amidinate and formamidinatoborate intermediates and an example of one of these latter species has been structurally characterised by an X-ray diffraction analysis. Crucially, no formation of the N-boryl formamidine products was observed in the absence of additional equivalents of the carbodiimide and HBpin substrates. This observation, supported by the evolution of a sigmoidal kinetic profile for the hydroboration of dicyclohexylcarbodiimide, has been rationalised as the consequence of an allosteric effect of the pinacol borane and carbodiimide on the magnesium formamidinatoborate intermediates. PMID:27072429

  7. Effect of EDTA Conditioning and Carbodiimide Pretreatment on the Bonding Performance of All-in-One Self-Etch Adhesives

    PubMed Central

    Singh, Shipra; Nagpal, Rajni; Tyagi, Shashi Prabha; Manuja, Naveen

    2015-01-01

    Objective. This study evaluated the effect of ethylenediaminetetraacetic acid (EDTA) conditioning and carbodiimide (EDC) pretreatment on the shear bond strength of two all-in-one self-etch adhesives to dentin. Methods. Flat coronal dentin surfaces were prepared on one hundred and sixty extracted human molars. Teeth were randomly divided into eight groups according to two different self-etch adhesives used [G-Bond and OptiBond-All-In-One] and four different surface pretreatments: (a) adhesive applied following manufacturer's instructions; (b) dentin conditioning with 24% EDTA gel prior to application of adhesive; (c) EDC pretreatment followed by application of adhesive; (d) application of EDC on EDTA conditioned dentin surface followed by application of adhesive. Composite restorations were placed in all the samples. Ten samples from each group were subjected to immediate and delayed (6-month storage in artificial saliva) shear bond strength evaluation. Data collected was subjected to statistical analysis using three-way ANOVA and post hoc Tukey's test at a significance level of p < 0.05.  Results and Conclusion. EDTA preconditioning as well as EDC pretreatment alone had no significant effect on the immediate and delayed bond strengths of either of the adhesives. However, EDC pretreatment on EDTA conditioned dentin surface resulted in preservation of resin-dentin bond strength of both adhesives with no significant fall over six months. PMID:26557850

  8. Potential role of surface wettability on the long-term stability of dentin bonds after surface biomodification

    PubMed Central

    Leme, Ariene A.; Vidal, Cristina M. P.; Hassan, Lina Saleh; Bedran-Russo, Ana K.

    2015-01-01

    Degradation of the adhesive interface contributes to the failure of resin composite restorations. The hydrophilicity of the dentin matrix during and after bonding procedures may result in an adhesive interface that is more prone to degradation over time. This study assessed the effect of chemical modification of dentin matrix on the wettability and the long-term reduced modulus of elasticity (Er) of the adhesive interfaces. Human molars were divided into groups according to the priming solutions: distilled water (control), 6.5% Proanthocyanidin-rich grape seed extract (PACs), 5.75% 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride/1.4% n-hydroxysuccinimide (EDC/NHS) and 5% Glutaraldehyde (GA). The water-surface contact angle was verified before and after chemical modification of the dentin matrix. The demineralized dentin surface was treated with the priming solutions and restored with One Step Plus (OS) and Single Bond Plus (SB) and resin composite. The Er of the adhesive, hybrid layer and underlying dentin was evaluated after 24 h and 30 months in artificial saliva. The dentin hydrophilicity significantly decreased after application of the priming solutions. Aging significantly decreased the Er in the hybrid layer and underlying dentin of control groups. The Er of GA groups remained stable over time at the hybrid layer and underlying dentin. Significant higher Er was observed for PACs and EDC/NHS groups at the hybrid layer after 24 h. The decreased hydrophilicity of the modified dentin matrix likely influence the immediate mechanical properties of the hybrid layer. Dentin biomodification prevented substantial aging at the hybrid layer and underlying dentin after 30 months storage. PMID:25869721

  9. Growth Factors Cross-Linked to Collagen Microcarriers Promote Expansion and Chondrogenic Differentiation of Human Mesenchymal Stem Cells.

    PubMed

    Bertolo, Alessandro; Arcolino, Fanny; Capossela, Simona; Taddei, Anna Rita; Baur, Martin; Pötzel, Tobias; Stoyanov, Jivko

    2015-10-01

    Tissue engineering is a field in progressive expansion and requires constant updates in methods and devices. One of the central fields is the development of biocompatible, biodegradable, and injectable scaffolds, such as collagen microcarriers. To enhance cell attachment and produce a cost-effective cell culture solution with local stimulation of cells, basic fibroblast growth factor (bFGF) or transforming growth factor-β1 (TGF-β1) was covalently immobilized on microcarriers either by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) or riboflavin/UV (RB/UV) light-mediated cross-linking. Collagen microcarriers cross-linked with bFGF or TGF-β1 were used for expansion and chondrogenic differentiation of human mesenchymal stem cells (MSCs). Evaluation methods included cell viability test, chondrogenic marker expression (aggrecan and collagen type I and type II), histological detection of proteoglycans, and immunohistochemical analysis. Cross-linking strengthened the collagen structure of the microcarriers and reduced collagenase-mediated degradation. MSCs effectively proliferated on microcarriers cross-linked with bFGF, especially by EDC/NHS cross-linking. Chondrogenic differentiation of MSCs was induced by TGF-β1 cross-linked on microcarriers, promoting gene expression and protein accumulation of aggrecan and collagen type I and type II, as well as proteoglycans. Cross-linking by RB/UV enhanced chondrogenesis more than any other group. In addition, cross-linking reduced scaffold shrinkage exerted by MSCs during chondrogenesis, a desirable feature for microcarriers if used as tissue defect filler. In conclusion, cross-linking of bFGF or TGF-β1 to collagen microcarriers supported in vitro proliferation and chondrogenesis, respectively. If translated in vivo and in clinical practice, such approach might lead a step closer to development of a cost-effective and locally acting device for cell-based therapy. PMID:26222829

  10. Preparation and characterization of group A meningococcal capsular polysaccharide conjugates and evaluation of their immunogenicity in mice.

    PubMed

    Jin, Zhigang; Chu, Chiayung; Robbins, John B; Schneerson, Rachel

    2003-09-01

    Epidemic and endemic meningitis caused by group A Neisseria meningitidis remains a problem in sub-Saharan Africa. Although group A meningococcal capsular polysaccharide (GAMP) vaccine confers immunity at all ages, the improved immunogenicity of a conjugate and its compatibility with the World Health Organization's Extended Program on Immunization offers advantages over GAMP alone. Conjugates of GAMP bound to bovine serum albumin (BSA) were synthesized, characterized, and evaluated for their immunogenicities in mice. Two methods, involving adipic acid dihydrazide (ADH) as a linker, were used. First, ADH was bound to GAMP activated with cyanogen bromide (CNBr) or with 1-cyano-4(dimethylamino)-pyridinium tetrafluoroborate (CDAP) to form GAMP(CNBr)AH and GAMP(CDAP)AH. These derivatives were bound to BSA by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) to form GAMP(CNBr)AH-BSA and GAMP(CDAP)AH-BSA. Second, ADH was bound to BSA with EDC to form AHBSA. AHBSA was bound to activated GAMP to form GAMP(CNBr)-AHBSA and GAMP(CDAP)-AHBSA. The yield of GAMP(CDAP)-AHBSA (35 to 40%) was higher than those of the other conjugates (5 to 20%). GAMP conjugates elicited immunoglobulin G (IgG) anti-GAMP in all mice after three injections of 2.5 or 5.0 microg of GAMP: the geometric mean (GM) was highest in recipients of GAMP(CDAP)-AHBSA (11.40 enzyme-linked immunosorbent assay units). Although the difference was not statistically significant, the 5.0- microg dose elicited a higher GM IgG anti-GAMP than the 2.5- microg dose. Low levels of anti-GAMP were elicited by GAMP alone. GAMP(CDAP)-AHBSA elicited bactericidal activity roughly proportional to the level of IgG anti-GAMP. PMID:12933854

  11. Stiffness-controlled three-dimensional collagen scaffolds for differentiation of human Wharton's jelly mesenchymal stem cells into cardiac progenitor cells.

    PubMed

    Lin, Yun-Li; Chen, Chie-Pein; Lo, Chun-Min; Wang, Hwai-Shi

    2016-09-01

    Stem cell-based regenerative therapy has emerged as a promising treatment for myocardial infarction. The aim of this study is to develop stiffness-controlled collagen scaffolds to allow proliferation and differentiation of mesenchymal stem cell (MSCs) into cardiac progenitor cells. In this study transforming growth factor β2 (TGF-β2), was used to induce stem cell differentiation into cardiac lineage cells. Collagen scaffolds were cross-linked with cross-linkers, 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC), and N-Hydroxysuccinimide (NHS). The results showed that collagen scaffolds cross-linked with 25/50 and 50/50 of EDC mM/NHS mM cross-linkers exhibited little difference in shape and size, the scaffold cross-linked with 50/50 of cross-linkers demonstrated better interconnectivity and higher Young's modulus (31.8 kPa) than the other (15.4 kPa). SEM observation showed that MSCs could grow inside the scaffolds and interact with collagen scaffolds. Furthermore, greater viability and cardiac lineage differentiation were achieved in MSCs cultured on stiffer scaffolds. The results suggest that three-dimensional type I collagen scaffolds with suitable cross-linking to adjust for stiffness can affect MSC fate and direct the differentiation of MSCs into cardiac progenitor cells with/without TGF-β2. These stiffness-controlled collagen scaffolds hold great potential as carriers for delivering MSCs differentiated cardiac progenitor cells into infracted hearts. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2234-2242, 2016. PMID:27120780

  12. Coupling Strategies for the Synthesis of Peptide-Oligonucleotide Conjugates for Patterned Synthetic Biomineralization

    PubMed Central

    Carter, Joshua D.; LaBean, Thomas H.

    2011-01-01

    This work describes preparation strategies for peptide-oligonucleotide conjugates that combine the self-assembling behavior of DNA oligonucleotides with the molecular recognition capabilities of peptides. The syntheses include a solution-phase fragment coupling reaction and a solid-phase fragment coupling strategy where the oligonucleotide has been immobilized on DEAE Sepharose. The yield of four coupling reagents is evaluated, two reagents in water, EDC (1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride) and DMTMM (4-(4,6-dimethoxy[1,3,5]triazin-2-yl)-4-methyl-morpholinium chloride), and two in dimethylformamide (DMF), PyBOP ((Benzotriazol-1-yloxy) tripyrrolidinophosphonium hexafluorophosphate) and HBTU (O-benzotriazole-N,N,N′,N′-tetramethyluronium hexafluorophosphate), while the oligonucleotide fragment is either in solution or immobilized on DEAE. These coupling strategies rely on an unprotected 5′ amino linker on the oligonucleotide reacting with the peptide C-terminus. The peptide, selected from a combinatorial library for its gold-binding behavior, was 12 amino acids long with an N-terminus acetyl cap. Formation of the conjugates was confirmed by gel electrophoresis and mass spectrometry while molecular recognition functionality of the peptide portion was verified using atomic force microscopy. Solution-phase yields were superior to their solid-phase counterparts. EDC resulted in the highest yield for both solution-phase (95%) and solid-phase strategies (24%), while the DMF-based reagents, PyBOP and HBTU, resulted in low yields with reduced recovery. All recoverable conjugates demonstrated gold nanoparticle templating capability. PMID:22007290

  13. Coupling strategies for the synthesis of Peptide-oligonucleotide conjugates for patterned synthetic biomineralization.

    PubMed

    Carter, Joshua D; Labean, Thomas H

    2011-01-01

    This work describes preparation strategies for peptide-oligonucleotide conjugates that combine the self-assembling behavior of DNA oligonucleotides with the molecular recognition capabilities of peptides. The syntheses include a solution-phase fragment coupling reaction and a solid-phase fragment coupling strategy where the oligonucleotide has been immobilized on DEAE Sepharose. The yield of four coupling reagents is evaluated, two reagents in water, EDC (1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride) and DMTMM (4-(4,6-dimethoxy[1,3,5]triazin-2-yl)-4-methyl-morpholinium chloride), and two in dimethylformamide (DMF), PyBOP ((Benzotriazol-1-yloxy) tripyrrolidinophosphonium hexafluorophosphate) and HBTU (O-benzotriazole-N,N,N',N'-tetramethyluronium hexafluorophosphate), while the oligonucleotide fragment is either in solution or immobilized on DEAE. These coupling strategies rely on an unprotected 5' amino linker on the oligonucleotide reacting with the peptide C-terminus. The peptide, selected from a combinatorial library for its gold-binding behavior, was 12 amino acids long with an N-terminus acetyl cap. Formation of the conjugates was confirmed by gel electrophoresis and mass spectrometry while molecular recognition functionality of the peptide portion was verified using atomic force microscopy. Solution-phase yields were superior to their solid-phase counterparts. EDC resulted in the highest yield for both solution-phase (95%) and solid-phase strategies (24%), while the DMF-based reagents, PyBOP and HBTU, resulted in low yields with reduced recovery. All recoverable conjugates demonstrated gold nanoparticle templating capability. PMID:22007290

  14. Integrated reactive ion etching to pattern cross-linked hydrophilic polymer structures for protein immobilization

    NASA Astrophysics Data System (ADS)

    Bhatnagar, Parijat; Strickland, Aaron D.; Kim, Il; Malliaras, George G.; Batt, Carl A.

    2007-04-01

    Patterning of cross-linked hydrophilic polymer features using reactive ion etching (RIE) capable of covalently immobilizing proteins has been achieved. Projection photolithography was used to pattern photoresist to create micromolds. Vapor phase molecular self-assembly of polymerizable monolayer in molds allowed covalent binding of hydrogel on surface during free-radical polymerization. Excess hydrogel blanket film was consumed with oxygen RIE resulting into hydrogel pattern of 1μm size aligned to prefabricated silicon oxide structures. Proteins were finally coupled through their primary amine groups selectively to acid functionalized hydrogel features through stable amide linkages using 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride and N-hydroxysulfosuccinimide.

  15. Polymerization of beta-amino acids in aqueous solution

    NASA Technical Reports Server (NTRS)

    Liu, R.; Orgel, L. E.; Bada, J. L. (Principal Investigator)

    1998-01-01

    We have compared carbonyl diimidazole (CDI) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDAC) as activating agents for the oligomerization of negatively-charged alpha- and beta-amino acids in homogeneous aqueous solution. alpha-Amino acids can be oligomerized efficiently using CDI, but not by EDAC. beta-Amino acids can be oligomerized efficiently using EDAC, but not by CDI. Aspartic acid, an alpha- and beta-dicarboxylic acid is oligomerized efficiently by both reagents. These results are explained in terms of the mechanisms of the reactions, and their relevance to prebiotic chemistry is discussed.

  16. Impact of carbondiimide crosslinker used for magnetic carbon nanotube mediated GFP plasmid delivery

    NASA Astrophysics Data System (ADS)

    Hao, Yuzhi; Xu, Peng; He, Chuan; Yang, Xiaoyan; Huang, Min; Xing, James; Chen, Jie

    2011-07-01

    1-ethyl-3-(3-dimethylaminopropyl) carbondiimide hydrochloride (EDC) is commonly used as a crosslinker to help bind biomolecules, such as DNA plasmids, with nanostructures. However, EDC often remains, after a crosslink reaction, in the micro-aperture of the nanostructure, e.g., carbon nanotube. The remaining EDC shows positive green fluorescent signals and makes a nanostructure with a strong cytotoxicity which induces cell death. The toxicity of EDC was confirmed on a breast cancer cell line (MCF-7) and two leukemic cell lines (THP-1 and KG-1). The MCF-7 cells mainly underwent necrosis after treatment with EDC, which was verified by fluorescein isothiocyanate (FITC) annexin V staining, video microscopy and scanning electronic microscopy (SEM). If the EDC was not removed completely, the nanostructures with remaining EDC produced a green fluorescent background that could interfere with flow cytometry (FACS) measurement and result in false information about GFP plasmid delivery. Effective methods to remove residual EDC on macromolecules were also developed.

  17. Systematic Investigation of EDC/sNHS-Mediated Bioconjugation Reactions for Carboxylated Peptide Substrates.

    PubMed

    Totaro, Kyle A; Liao, Xiaoli; Bhattacharya, Keshab; Finneman, Jari I; Sperry, Justin B; Massa, Mark A; Thorn, Jennifer; Ho, Sa V; Pentelute, Bradley L

    2016-04-20

    1-Ethyl-3-(3-(dimethylamino)propyl)carbodiimide (EDC) bioconjugations have been utilized in preparing variants for medical research. While there have been advances in optimizing the reaction for aqueous applications, there has been limited focus toward identifying conditions and side reactions that interfere with product formation. We present a systematic investigation of EDC/N-hydroxysulfosuccinimide (sNHS)-mediated bioconjugations on carboxylated peptides and small proteins. We identified yet-to-be-reported side products arising from both the reagents and substrates. Model peptides used in this study illustrate particular substrates are more susceptible to side reactions than others. From our studies, we found that bioconjugations are more efficient with high concentrations of amine nucleophile but not sNHS. Performing bioconjugations on a model affibody protein show that the trends established with model peptides hold for more complex systems. PMID:26974183

  18. Microfluidic Generation of Haptotactic Gradients through 3D Collagen Gels for Enhanced Neurite Growth

    PubMed Central

    Sundararaghavan, Harini G.; Masand, Shirley N.

    2011-01-01

    Abstract We adapted a microfluidic system used previously to generate durotactic gradients of stiffness in a 3D collagen gel, to produce haptotactic gradients of adhesive ligands through the collagen gel. Oligopeptide sequences that included bioactive peptide sequences from laminin, YIGSR, or IKVAV, were grafted separately onto type I collagen using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC). Solutions of peptide-grafted collagen and untreated collagen were then used as source and sink input solutions, respectively, in an H-shaped microfluidic network fabricated using traditional soft lithography. One-dimensional gradients of the peptide-grafted collagen solution were generated in the channel that connected the source and sink channels, and these gradients became immobilized upon self-assembly of the collagen into a 3D fibrillar gel. The slope and average concentration of the gradients were adjusted by changing the concentration of the source solutions and by changing the length of the cross-channel. A separate, underlying channel in the microfluidic construct allowed the introduction of a chick embryo dorsal root ganglion into the network. Neurites from these explants grew significantly longer up steep gradients of YIGSR, but shallow gradients of IKVAV in comparison to untreated collagen controls. When these two gradients were presented in combination, the bias in growth acceleration was the largest and most consistent. No differences were observed in the number of neurites choosing to grow up or down the gradients in any condition. These results suggest that the incorporation of distinct gradients of multiple bioactive ligands can improve directional acceleration of regenerating axons. PMID:21473683

  19. Synthesis and characterization of gibberellin-chitosan conjugate for controlled-release applications.

    PubMed

    Liu, Yao; Sun, Yan; He, Shun; Zhu, Yuncong; Ao, Mingming; Li, Jianqiang; Cao, Yongsong

    2013-06-01

    Controlled release formulations (CRFs) are promising in improving the efficiency of pesticide and minimizing the spreading of hazardous residues in environment. By coupling with the pesticide covalently, chitosan can be used as a carrier material for the vulnerable ingredient. For the first time, gibberellic acid (GA3), one of plant growth regulators, was attached to chitosan (CS) to form a GA3-CS conjugate via the formation of an amide bound using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and N-hydroxysuccinimide. The novel conjugate was structurally characterized by fourier transform infrared spectroscopy, ultraviolet spectrophotometer, and thermal gravimetric analysis. Effects of pH, temperature, and UV irradiation on the release of this conjugate were investigated. The results showed that the new conjugate had a remarkable modification degree for CS (more than 60%, w/w) and the optimal coupling conditions were defined as: the molar ratio of GA3:EDC/NHS:CS was 1:1.2:1.2, at pH 6.0 for 24 h. The release data showed the novel conjugate protected GA3 against photo- and thermal-degradation effectively and the concentration of GA3 in GA3-CS kept unchangeable about 60 d in different pH conditions. Compared with GA3 technical, the conjugate had better water solubility and stability and have potential applications. The present study also provides a novel preparation method of CRFs comprising a pesticide with long duration, sustained-release performance and good environmental compatibility. This method may be extended to other pesticides that possess a carboxyl group. PMID:23511059

  20. Fluorescent identification and detection of Staphylococcus aureus with carboxymethyl chitosan/CdS quantum dots bioconjugates.

    PubMed

    Wang, Xiaohui; Du, Yumin; Li, Yan; Li, Dong; Sun, Runcang

    2011-01-01

    A fast and sensitive method based on fluorescent carboxymethyl chitosan/CdS quantum dots (CMCS-CdS QDs) composites was developed for specific detection of Staphylococcus aureus in food and the environment. Fluorescent CMCS-CdS QDs were prepared in aqueous solution through a green method. A human immunoglobulin (IgG) antibody was then bioconjugated to the QDs in the presence of 1-ethyl-3-(3)-dimethylaminopropyl carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) to make a novel type of mono-dispersed water-soluble fluorescent bioprobes. The fluorescent bioprobes were employed to identify S. aureus by incubating them with the bacteria for a certain time and observing the marked cells under fluorescence microscopy after removing free fluorescent QDs. Fluorescence microscopy images showed the S. aureus cells were successfully recognized by the bioprobes. Several other bacteria commonly found in environment such as Escherichia coli and Bacillus subtilis were also incubated with the bioprobes to test their specificity. It was found that the novel QDs-CMCS-IgG bioprobes had specific identification to S. aureus cells. Fluorescence measurement using a luminescence spectrometer could be applied to quantify S. aureus cells. The fluorescence intensity of the samples at 600 nm was proportional to the cell concentration in the range of 10(3)-10(7) cfu/ml, and the detection limit was as low as 900 cfu/ml. Considering the simplicity and cost-efficiency of this method, its application in the identification and quantification of bacteria in clinical, food and environmental samples is anticipated. PMID:20961493

  1. Localized surface plasmon resonance immunoassay and verification using surface-enhanced Raman spectroscopy

    NASA Astrophysics Data System (ADS)

    Yonzon, Chanda R.; Zhang, Xiaoyu; Van Duyne, Richard P.

    2003-11-01

    This work exploits the localized surface plasmon resonance (LSPR) spectroscopy of noble metal nanoparticles to achieve sensitive and selective detection of biological analytes. Noble metal nanoparticles exhibit an LSPR that is strongly dependent on their size, shape, material, and the local dielectric environment. The LSPR is also responsible for the intense signals observed in surface-enhanced Raman scattering (SERS). Ag nanoparticles fabricated using the nanosphere lithography (NSL) technique exploits this LSPR sensitivity as a signal transduction method in biosensing applications. The current work implements LSPR biosensing for the anti dinitrophenyl (antiDNP) immunoassay system. Upon forming the 2,4 dinitrobenzoic acid/antiDNP complex, this system shows a large LSPR shift of 44 nm when exposed to antiDNP concentration of 1.5 x 10-6 M. In addition, due to the unique molecular characteristics of the functional groups on the biosensor, it can also be characterized using SERS. First, the nanoparticles are functionalized with a mixed self-assembled monolayer (SAM) comprised of 2:1 octanethiol and 11-amino undecanethiol. The SAM is exposed to 2,4-dinitrobenzoic acid with the 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC) coupling reagent. Finally, the 2,4-dinitrophenyl terminated SAM is exposed to various concentration of antiDNP. LSPR shifts indicate the occurrence of a binding event. SER spectra confirm binding of 2,4 dinitrobenzoic acid with amine-terminated SAM. This LSPR/SERS biosensing method can be generalized to a myriad of biologically relevant systems.

  2. Generation and characterization of phage-GnRH chemical conjugates for potential use in cat and dog immunocontraception.

    PubMed

    Samoylov, A; Cox, N; Cochran, A; Wolfe, K; Donovan, C; Kutzler, M; Petrenko, V; Baker, H; Samoylova, T

    2012-12-01

    Overpopulation of cats and dogs is a serious worldwide problem that demands novel, safe and cost-effective solutions. The objective of this study was to generate and characterize phage-peptide conjugates with gonadotropin-releasing hormone (GnRH) for potential use as an immunocontraceptive. A filamentous phage vector f5-8 with wild-type phage coat proteins was used as a carrier for construction of chemical conjugates with GnRH, a peptide that acts as a master reproductive hormone. In such conjugates, the phage body plays the role of a carrier protein, while multiple copies of GnRH peptide stimulate production of neutralizing anti-GnRH antibodies potentially leading to contraceptive effects. To generate the constructs, four different GnRH-based peptides were synthesized and conjugated to phage particles in a two-step procedure: (i) peptides were reacted with phage to form a conjugate using 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride chemistry (EDC) and (ii) the conjugates were separated from remaining free peptides by dialysis. Formation and specificity of phage-GnRH conjugates were confirmed by three independent methods: spectrophotometry, electron microscopy and ELISA. When the conjugates were tested for interaction with sera collected from cats and dogs immunized with GnRH-based vaccines in independent studies, strong specific ELISA signals were obtained, suggesting the potential use of the conjugates for cat and dog immunosterilization. The ability of the conjugates to stimulate production of anti-GnRH antibodies in vivo was evaluated in mice. While optimization of dose, immunization route and adjuvant still requires investigation, our preliminary results demonstrated the presence of anti-GnRH antibodies in sera of mice immunized with such conjugates. Fertility trials in cats and dogs will be needed to evaluate contraceptive potentials of the phage-GnRH peptide chemical conjugates. PMID:23279551

  3. A comparative study of different protein immobilization methods for the construction of an efficient nano-structured lactate oxidase-SWCNT-biosensor

    PubMed Central

    Pagán, Miraida; Suazo, Dámaris; del Toro, Nicole; Griebenow, Kai

    2014-01-01

    We constructed lactate biosensors by immobilization of lactate oxidase (LOx) onto a single-walled carbon nanotube (SWCNT) electrode. The first step of the sensor construction was the immobilization of oxidized SWCNT onto a platinum electrode modified with 4-aminothiophenol (4-ATP). Two enzyme immobilization methods were used to construct the biosensors, i.e., covalent immobilization using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and physical adsorption. Atomic force microscopy (AFM) experiments confirmed the immobilization of SWCNT during the biosensor construction and X-ray photoelectron spectroscopy (XPS) experiments confirmed covalent immobilization of LOx onto the SWCNT in the first method. The biosensor based on covalent enzyme immobilization showed a sensitivity of 5.8 μA/mM, a linearity up to 0.12 mM of L-lactate, and a detection limit of 4.0 μM. The biosensor based on protein adsorption displayed a sensitivity of 9.4 μA/mM, retaining linearity up to 0.18 mM of L-lactate with a detection limit of 3.0 μM. The difference in the biosensor response can be attributed to protein conformational or dynamical changes during covalent immobilization. The stability of the biosensors was tested at different temperatures and after different storage periods. The thermostability of the biosensors after incubation at 60°C demonstrated that the biosensor with covalently immobilized LOx retained a higher response compare with the adsorbed protein. Long-term stability experiments show a better residual activity of 40% for the covalently immobilized protein compared to 20% of residual activity for the adsorbed protein after 25 d storage. Covalent protein immobilization was superior compared to adsorption in preserving biosensor functionality over extended time period. PMID:25216450

  4. Investigation and characterization of oxidized cellulose and cellulose nanofiber films

    NASA Astrophysics Data System (ADS)

    Yang, Han

    Over the last two decades, a large amount of research has focused on natural cellulose fibers, since they are "green" and renewable raw materials. Recently, nanomaterials science has attracted wide attention due to the large surface area and unique properties of nanoparticles. Cellulose certainly is becoming an important material in nanomaterials science, with the increasing demand of environmentally friendly materials. In this work, a novel method of preparing cellulose nanofibers (CNF) is being presented. This method contains up to three oxidation steps: periodate, chlorite and TEMPO (2,2,6,6-tetramethylpiperidinyl-1-oxyl) oxidation. The first two oxidation steps are investigated in the first part of this work. Cellulose pulp was oxidized to various extents by a two step-oxidation with sodium periodate, followed by sodium chlorite. The oxidized products can be separated into three different fractions. The mass ratio and charge content of each fraction were determined. The morphology, size distribution and crystallinity index of each fraction were measured by AFM, DLS and XRD, respectively. In the second part of this work, CNF were prepared and modified under various conditions, including (1) the introduction of various amounts of aldehyde groups onto CNF by periodate oxidation; (2) the carboxyl groups in sodium form on CNF were converted to acid form by treated with an acid type ion-exchange resin; (3) CNF were cross-linked in two different ways by employing adipic dihydrazide (ADH) as cross-linker and water-soluble 1-ethyl-3-[3-(dimethylaminopropyl)] carbodiimide (EDC) as carboxyl-activating agent. Films were fabricated with these modified CNF suspensions by vacuum filtration. The optical, mechanical and thermo-stability properties of these films were investigated by UV-visible spectrometry, tensile test and thermogravimetric analysis (TGA). Water vapor transmission rates (WVTR) and water contact angle (WCA) of these films were also studied.

  5. Vibrio cholerae O139 conjugate vaccines: synthesis and immunogenicity of V. cholerae O139 capsular polysaccharide conjugates with recombinant diphtheria toxin mutant in mice.

    PubMed

    Kossaczka, Z; Shiloach, J; Johnson, V; Taylor, D N; Finkelstein, R A; Robbins, J B; Szu, S C

    2000-09-01

    Epidemiologic and experimental data provide evidence that a critical level of serum immunoglobulin G (IgG) antibodies to the surface polysaccharide of Vibrio cholerae O1 (lipopolysaccharide) and of Vibrio cholerae O139 (capsular polysaccharide [CPS]) is associated with immunity to the homologous pathogen. The immunogenicity of polysaccharides, especially in infants, may be enhanced by their covalent attachment to proteins (conjugates). Two synthetic schemes, involving 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and 1-cyano-4-dimethylaminopyridinium tetrafluoroborate (CDAP) as activating agents, were adapted to prepare four conjugates of V. cholerae O139 CPS with the recombinant diphtheria toxin mutant, CRMH21G. Adipic acid dihydrazide was used as a linker. When injected subcutaneously into young outbred mice by a clinically relevant dose and schedule, these conjugates elicited serum CPS antibodies of the IgG and IgM classes with vibriocidal activity to strains of capsulated V. cholerae O139. Treatment of these sera with 2-mercaptoethanol (2-ME) reduced, but did not eliminate, their vibriocidal activity. These results indicate that the conjugates elicited IgG with vibriocidal activity. Conjugates also elicited high levels of serum diphtheria toxin IgG. Convalescent sera from 20 cholera patients infected with V. cholerae O139 had vibriocidal titers ranging from 100 to 3,200: absorption with the CPS reduced the vibriocidal titer of all sera to < or =50. Treatment with 2-ME reduced the titers of 17 of 20 patients to < or =50. These data show that, like infection with V. cholerae O1, infection with V. cholerae O139 induces vibriocidal antibodies specific to the surface polysaccharide of this bacterium (CPS) that are mostly of IgM class. Based on these data, clinical trials with the V. cholerae O139 CPS conjugates with recombinant diphtheria toxin are planned. PMID:10948122

  6. A novel electrospinning approach to fabricate high strength aqueous silk fibroin nanofibers.

    PubMed

    Singh, B N; Panda, N N; Pramanik, K

    2016-06-01

    The present paper describes a rapid method of producing concentrated aqueous regenerated Bombyx mori silk fibroin (RSF) solution by applying mild shearing under forced dehumidified air and generation of electrospun SF nanofibers from concentrated solution with high mechanical strength using free liquid surface electrospinning machine. The shear induced concentrating mechanism favoured the electrospinning process by enhancing the viscosity (>2.43Pas as onset for electrospinning) and decreasing the surface tension of the solution (40.1-37.7mN/m). Shearing reduced the β-turns and random coil molecular conformation and thereby, intensified the β-sheet content from 16.9% to 34% which is the minimum content needed to commence RSF nanofibers formation. Subsequently, electrospun nanofibrous mats were produced from different batches of concentrated SF solutions (15-21wt%). Among the concentrated RSF, 17wt% RSF solution was the most favourable concentration producing electrospun nanofibrous mat having lowest average fiber diameters of 183±55nm and good tensile strength. The mechanical strength of the nanofibrous sheet was further improved by cross-linking with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and N-hydroxysuccinimide (EDC+NHS) which might be due to enhancement of β-sheet content. These nanofibers exhibited 17.57±1.13MPa ultimate tensile strength, 12.48±1.46% tensile strain at break and 37.7% increase in root mean square surface roughness which is favourable feature for cell adhesion and neo-tissue formation. PMID:26905467

  7. Preparation of aminated chitosan/alginate scaffold containing halloysite nanotubes with improved cell attachment.

    PubMed

    Amir Afshar, Hamideh; Ghaee, Azadeh

    2016-10-20

    The chemical nature of biomaterials play important role in cell attachment, proliferation and migration in tissue engineering. Chitosan and alginate are biodegradable and biocompatible polymers used as scaffolds for various medical and clinical applications. Amine groups of chitosan scaffolds play an important role in cell attachment and water adsorption but also associate with alginate carboxyl groups via electrostatic interactions and hydrogen bonding, consequently the activity of amine groups in the scaffold decreases. In this study, chitosan/alginate/halloysite nanotube (HNTs) composite scaffolds were prepared using a freeze-drying method. Amine treatment on the scaffold occurred through chemical methods, which in turn caused the hydroxyl groups to be replaced with carboxyl groups in chitosan and alginate, after which a reaction between ethylenediamine, 1-ethyl-3,(3-dimethylaminopropyl) carbodiimide (EDC) and scaffold triggered the amine groups to connect to the carboxyl groups of chitosan and alginate. The chemical structure, morphology and mechanical properties of the composite scaffolds were investigated by FTIR, CHNS, SEM/EDS and compression tests. The electrostatic attraction and hydrogen bonding between chitosan, alginate and halloysite was confirmed by FTIR spectroscopy. Chitosan/alginate/halloysite scaffolds exhibit significant enhancement in compressive strength compared with chitosan/alginate scaffolds. CHNS and EDS perfectly illustrate that amine groups were effectively introduced in the aminated scaffold. The growth and cell attachment of L929 cells as well as the cytotoxicity of the scaffolds were investigated by SEM and Alamar Blue (AB). The results indicated that the aminated chitosan/alginate/halloysite scaffold has better cell growth and cell adherence in comparison to that of chitosan/alginate/halloysite samples. Aminated chitosan/alginate/halloysite composite scaffolds exhibit great potential for applications in tissue engineering, ideally in

  8. Covalent immobilization of porcine pancreatic lipase on carboxyl-activated magnetic nanoparticles: characterization and application for enzymatic inhibition assays.

    PubMed

    Zhu, Yuan-Ting; Ren, Xiao-Yun; Liu, Yi-Ming; Wei, Ying; Qing, Lin-Sen; Liao, Xun

    2014-05-01

    Using carboxyl functionalized silica-coated magnetic nanoparticles (MNPs) as carrier, a novel immobilized porcine pancreatic lipase (PPL) was prepared through the 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride/N-hydroxysuccinimide (EDC/NHS) coupling reaction. Transmission electron microscopic images showed that the synthesized nanoparticles (Fe3O4-SiO2) possessed three dimensional core-shell structures with an average diameter of ~20 nm. The effective enzyme immobilization onto the nanocomposite was confirmed by atomic force microscopic (AFM) analysis. Results from Fourier-transform infrared spectroscopy (FT-IR), Bradford protein assay, and thermo-gravimetric analysis (TGA) indicated that PPL was covalently attached to the surface of magnetic nanoparticles with a PPL immobilization yield of 50mg enzyme/g MNPs. Vibrating sample magnetometer (VSM) analysis revealed that the MNPs-PPL nanocomposite had a high saturation magnetization of 42.25 emu·g(-1). The properties of the immobilized PPL were investigated in comparison with the free enzyme counterpart. Enzymatic activity, reusability, thermo-stability, and storage stability of the immobilized PPL were found significantly superior to those of the free one. The Km and the Vmax values (0.02 mM, 6.40 U·mg(-1) enzyme) indicated the enhanced activity of the immobilized PPL compared to those of the free enzyme (0.29 mM, 3.16 U·mg(-1) enzyme). Furthermore, at an elevated temperature of 70 °C, immobilized PPL retained 60% of its initial activity. The PPL-MNPs nanocomposite was applied in the enzyme inhibition assays using orlistat, and two natural products isolated from oolong tea (i.e., EGCG and EGC) as the test compounds. PMID:24656379

  9. A comparative study of different protein immobilization methods for the construction of an efficient nano-structured lactate oxidase-SWCNT-biosensor.

    PubMed

    Pagán, Miraida; Suazo, Dámaris; Del Toro, Nicole; Griebenow, Kai

    2015-02-15

    We constructed lactate biosensors by immobilization of lactate oxidase (LOx) onto a single-walled carbon nanotube (SWCNT) electrode. The first step of the sensor construction was the immobilization of oxidized SWCNT onto a platinum electrode modified with 4-aminothiophenol (4-ATP). Two enzyme immobilization methods were used to construct the biosensors, i.e., covalent immobilization using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and physical adsorption. Atomic force microscopy (AFM) experiments confirmed the immobilization of SWCNT during the biosensor construction and X-ray photoelectron spectroscopy (XPS) experiments confirmed covalent immobilization of LOx onto the SWCNT in the first method. The biosensor based on covalent enzyme immobilization showed a sensitivity of 5.8 μA/mM, a linearity up to 0.12 mM of L-lactate, and a detection limit of 4.0 μM. The biosensor based on protein adsorption displayed a sensitivity of 9.4 μA/mM, retaining linearity up to 0.18 mM of L-lactate with a detection limit of 3.0 μM. The difference in the biosensor response can be attributed to protein conformational or dynamical changes during covalent immobilization. The stability of the biosensors was tested at different temperatures and after different storage periods. The thermostability of the biosensors after incubation at 60 °C demonstrated that the biosensor with covalently immobilized LOx retained a higher response compared with the adsorbed protein. Long-term stability experiments show a better residual activity of 40% for the covalently immobilized protein compared to 20% of residual activity for the adsorbed protein after 25 d storage. Covalent protein immobilization was superior compared to adsorption in preserving biosensor functionality over extended time period. PMID:25216450

  10. Protein immobilization on the surface of polydimethylsiloxane and polymethyl methacrylate microfluidic devices.

    PubMed

    Khnouf, Ruba; Karasneh, Dina; Albiss, Borhan Aldeen

    2016-02-01

    PDMS and PMMA are two of the most used polymers in the fabrication of lab-on-chip or microfluidic devices. In order to use these polymers in biological applications, it is sometimes essential to be able to bind biomolecules such as proteins and DNA to the surface of these materials. In this work, we have evaluated a number of processes that have been developed to bind protein to PDMS surfaces which include passive adsorption, passive adsorption with glutaraldehyde cross-linking, (3-aminopropyl) triethoxysilane functionalization followed by glutaraldehyde or 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride cross-linkers. It has been shown that the latter technique--using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride--results in more than twice the bonding of protein to the surface of PDMS microchannels than proteins binding passively. We have also evaluated a few techniques that have been tested for the functionalization of PMMA microchannels where we have found that the use of polyethyleneimine (PEI) has led to the strongest protein-PMMA microchannel bond. We finally demonstrated the effect of PDMS curing methodology on protein adsorption to its surface, and showed that increased curing time is the factor that reduces passive adsorption the most. PMID:26534833

  11. Artificial intelligence techniques to optimize the EDC/NHS-mediated immobilization of cellulase on Eudragit L-100.

    PubMed

    Zhang, Yu; Xu, Jing-Liang; Yuan, Zhen-Hong; Qi, Wei; Liu, Yun-Yun; He, Min-Chao

    2012-01-01

    Two artificial intelligence techniques, namely artificial neural network (ANN) and genetic algorithm (GA) were combined to be used as a tool for optimizing the covalent immobilization of cellulase on a smart polymer, Eudragit L-100. 1-Ethyl-3-(3-dimethyllaminopropyl) carbodiimide (EDC) concentration, N-hydroxysuccinimide (NHS) concentration and coupling time were taken as independent variables, and immobilization efficiency was taken as the response. The data of the central composite design were used to train ANN by back-propagation algorithm, and the result showed that the trained ANN fitted the data accurately (correlation coefficient R(2) = 0.99). Then a maximum immobilization efficiency of 88.76% was searched by genetic algorithm at a EDC concentration of 0.44%, NHS concentration of 0.37% and a coupling time of 2.22 h, where the experimental value was 87.97 ± 6.45%. The application of ANN based optimization by GA is quite successful. PMID:22942683

  12. Investigating thiol-modification on hyaluronan via carbodiimide chemistry using response surface methodology.

    PubMed

    Santhanam, Sruthi; Liang, Jue; Baid, Rinku; Ravi, Nathan

    2015-07-01

    Hyaluronan (HA) is a naturally occurring glycosaminoglycan widely researched for its use as a biomaterial in tissue engineering, drug delivery, angiogenesis, and ophthalmic surgeries. The mechanical properties of this biomaterial can be altered to a required extent by chemically modifying the pendant reactive groups. However, derivatizing these polymers to a predetermined extent has been the Achilles heel for this process. In this study, we have investigated the factors controlling the derivatization of the carboxyl moieties of HA with amine containing thiol, cystamine dihydrochloride (Cys), via carbodiimide crosslinking chemistry. We used fractional factorial design to screen and identify the significant factor(s) affecting the reaction, and response surface methodology (RSM) to develop a model equation for predicting the degree of thiolation of HA. Also, we analyzed the reaction mechanism for potential side reactions. We observed that N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) (mole ratio with repeat unit of HA) is the significant factor controlling the degree of amidation. The quadratic equations developed from RSM predict the formulation for a desired degree of amidation of HA and percentage of potential side product. Hence, derivatizing HA to a predetermined extent with minimal side product can be achieved using the statistical design of experiments. PMID:25369214

  13. A novel benign solution for collagen processing

    NASA Astrophysics Data System (ADS)

    Arnoult, Olivier

    Collagen is the main protein constituting the extracellular matrix (ECM) of tissues in the body (skin, cartilage, blood vessels...). It exists many types of collagen, this work studies only fibrillar collagen (e.g. collagen type I contained in the skin) that exhibits a triple helical structure composed of 3 alpha-helical collagen chains. This particular and defined hierarchical structure is essential to the biological and mechanical properties of the collagen. Processing collagen into scaffolds to mimic the ECM is crucial for successful tissue engineering. Recently collagen was processed into fibrous and porous scaffold using electrospinning process. However the solvent (HFIP) used for electrospinning is extremely toxic for the user and expensive. This work shows that HFIP can be replaced by a benign mixture composed of water, salt and alcohol. Yet only three alcohols (methanol, ethanol and iso-propanol) enable the dissolution of large quantity of collagen in the benign mixture, with a wide range of alcohol to buffer ratio, and conserve the collagen hierarchical structure at least as well as the HFIP. Collagen can be electrospun from the benign mixture into sub-micron fibers with concentrations as low as 6 wt-% for a wide range of alcohol to buffer ratio, with at least 10wt-% of salt, and any of the three alcohols. Specific conditions yield nano size fibers. After processing from HFIP or a benign mixture, collagen is water soluble and needs to be chemically crosslink for tissue engineering application. Post-crosslinking with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) results in the loss of the scaffold fibrous aspect and porosity, hence it is useless for tissue engineering. Such issue could be prevented by incorporating the crosslinker into the mixture prior to electrospinning. When EDC is used alone, collagen forms a gel in the mixture within minutes, preventing electrospinning. The addition of N-hydroxysuccinimide (NHS) in excess to EDC

  14. EDC BIOASSAYS FOR RISK MANAGEMENT PROJECTS

    EPA Science Inventory

    Overall goal for this research is to develop 3 bioassays for use in EDC projects across NRMRL (estrogenic, androgenic and thyroid assays). Currently, research is focused on estrogenic assays. A literature search was conducted to identify potential assays. The Yeast Estrogen Sc...

  15. EDC RESEARCH AT EPA ATLANTIC ECOLOGY DIVISION: DO ENVIRONMENTAL EDCS IMPACT FISH POPULATIONS

    EPA Science Inventory

    The Atlantic Ecology Division, Office of Research and Development, EP A is a marine laboratory situated on Narragansett Bay, Rhode Island. Researchers at AED are investigating the effects endocrine disrupting chemicals (EDCs) in the aquatic environment might have on reproductive ...

  16. RESEARCH ON RISK MANAGEMENT OF ENDOCRINE DISRUPTING CHEMICALS (EDCS)

    EPA Science Inventory

    The activity is a research program that will develop technical tools to manage sources of EDCs to the environment and to manage EDC accumulated in the environment. Several sources are under investigation including wastewater treatment plants, concentrated animal feeding operatio...

  17. Intra-molecular cross-linking of acidic residues for protein structure studies.

    PubMed

    Novak, Petr; Kruppa, Gary H

    2008-01-01

    Intra-molecular cross-linking has been suggested as a method of obtaining distance constraints that would help to develop structural models of proteins. Recent work published on intra-molecular cross-linking for protein structural studies has employed commercially available primary amine (lysine, the amino terminus) selective reagents. Previous work using these cross-linkers has shown that for several proteins of known structure, the number of cross-links that can be obtained experimentally may be small compared to what would be expected from the known structure, due to the relative reactivity, distribution and solvent accessibility of the lysines in the protein sequence. To overcome these limitations, we have investigated the use of cross-linking reagents that can react with other reactive side chains in proteins. We used 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) to activate the carboxylic acid containing residues, aspartic acid (D), glutamic acid (E) and the carboxy terminus (O), for cross-linking reactions. Once activated, the DEO side chains can react to form "zero-length" cross-links with nearby primary amine containing residues, lysines (K) and the amino terminus (X), via the formation of a new amide bond. We also show that the EDC-activated DEO side chains can be cross-linked to each other using dihydrazides, two hydrazide moieties connected by an alkyl cross-linker arm of variable length. Using these reagents, we have found three new "zero-length" cross-links in ubiquitin consistent with its known structure (M1-E16, M1-E18 and K63-E64). Using the dihydrazide cross-linkers, we have identified two new cross-links (D21-D32 and E24-D32) unambiguously. Using a library of dihydrazide cross-linkers with varying arm length, we have shown that there is a minimum arm length required for the DEO-DEO cross-links of 5.8 A. These results show that additional structural information can be obtained by exploiting new cross-linker chemistry

  18. Intra-molecular cross-linking of acidic residues for protein structure studies.

    SciTech Connect

    Kruppa, Gary Hermann; Young, Malin M.; Novak, Petr; Schoeniger, Joseph S.

    2005-03-01

    Intra-molecular cross-linking has been suggested as a method of obtaining distance constraints that would be useful in developing structural models of proteins. Recent work published on intra-molecular cross-linking for protein structural studies has employed commercially available primary amine selective reagents that can cross-link lysine residues to other lysine residues or the amino terminus. Previous work using these cross-linkers has shown that for several proteins of known structure, the number of cross-links that can be obtained experimentally may be small compared to what would be expected from the known structure, due to the relative reactivity, distribution, and solvent accessibility of the lysines in the protein sequence. To overcome these limitations we have investigated the use of cross-linking reagents that can react with other reactive sidechains in proteins. We used 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) to activate the carboxylic acid containing residues, aspartic acid (D), glutamic acid (E), and the carboxy terminus (O), for cross-linking reactions. Once activated, the DEO sidechains can react to form 'zero-length' cross-links with nearby primary amine containing resides, lysines (K) and the amino terminus (X), via the formation of a new amide bond. We also show that the EDC-activated DEO sidechains can be cross-linked to each other using dihydrazides, two hydrazide moieties connected by an alkyl cross-linker ann of variable length. Using these reagents, we have found three new 'zero-length' cross-links in ubiquitin consistent with its known structure (M1-E16, M1-E18, and K63-E64). Using the dihydrazide cross-linkers, we have identified 2 new cross-links (D21-D32 and E24-D32) unambiguously. Using a library of dihydrazide cross-linkers with varying arm length, we have shown that there is a minimum arm length required for the DEO-DEO cross-links of 5.8 angstroms. These results show that additional structural information

  19. An Immunoassay for Dibutyl Phthalate Based on Direct Hapten Linkage to the Polystyrene Surface of Microtiter Plates

    PubMed Central

    Wei, Chenxi; Ding, Shumao; You, Huihui; Zhang, Yaran; Wang, Yao; Yang, Xu; Yuan, Junlin

    2011-01-01

    Background Dibutyl phthalate (DBP) is predominantly used as a plasticizer inplastics to make them flexible. Extensive use of phthalates in both industrial processes and other consumer products has resulted in the ubiquitous presence of phthalates in the environment. In order to better determine the level of pollution in the environment and evaluate the potential adverse effects of exposure to DBP, immunoassay for DBP was developed. Methodology/Principal Findings A monoclonal antibody specific to DBP was produced from a stable hybridoma cell line generated by lymphocyte hybridoma technique. An indirect competitive enzyme-linked immunosorbent assay (icELISA) employing direct coating of hapten on polystyrene microtiter plates was established for the detection of DBP. Polystyrene surface was first oxidized by permanganate in dilute sulfuric acid to generate carboxyl groups. Then dibutyl 4-aminophthalate, which is an analogue of DBP, was covalently linked to the carboxyl groups of polystyrene surface with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). Compared with conjugate coated format (IC50 = 106 ng/mL), the direct hapten coated format (IC50 = 14.6 ng/mL) improved assay sensitivity after careful optimization of assay conditions. The average recovery of DBP from spiked water sample was 104.4% and the average coefficient of variation was 9.95%. Good agreement of the results obtained by the hapten coated icELISA and gas chromatography-mass spectrometry further confirmed the reliability and accuracy of the icELISA for the detection of DBP in certain plastic and cosmetic samples. Conclusions/Significance The stable and efficient hybridoma cell line obtained is an unlimited source of sensitive and specific antibody to DBP. The hapten coated format is proposed as generally applicable because the carboxyl groups on modified microtiter plate surface enables stable immobilization of aminated or hydroxylated hapten with EDC. The developed hapten

  20. Psb27, a transiently associated protein, binds to the chlorophyll binding protein CP43 in photosystem II assembly intermediates

    PubMed Central

    Liu, Haijun; Huang, Richard Y.-C.; Chen, Jiawei; Gross, Michael L.; Pakrasi, Himadri B.

    2011-01-01

    Photosystem II (PSII), a large multisubunit pigment–protein complex localized in the thylakoid membrane of cyanobacteria and chloroplasts, mediates light-driven evolution of oxygen from water. Recently, a high-resolution X-ray structure of the mature PSII complex has become available. Two PSII polypeptides, D1 and CP43, provide many of the ligands to an inorganic Mn4Ca center that is essential for water oxidation. Because of its unusual redox chemistry, PSII often undergoes degradation followed by stepwise assembly. Psb27, a small luminal polypeptide, functions as an important accessory factor in this elaborate assembly pathway. However, the structural location of Psb27 within PSII assembly intermediates has remained elusive. Here we report that Psb27 binds to CP43 in such assembly intermediates. We treated purified genetically tagged PSII assembly intermediate complexes from the cyanobacterium Synechocystis 6803 with chemical cross-linkers to examine intermolecular interactions between Psb27 and various PSII proteins. First, the water-soluble 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) was used to cross-link proteins with complementary charged groups in close association to one another. In the His27△ctpAPSII preparation, a 58-kDa cross-linked species containing Psb27 and CP43 was identified. This species was not formed in the HT3△ctpA△psb27PSII complex in which Psb27 was absent. Second, the homobifunctional thiol-cleavable cross-linker 3,3′-dithiobis(sulfosuccinimidylpropionate) (DTSSP) was used to reversibly cross-link Psb27 to CP43 in His27△ctpAPSII preparations, which allowed the use of liquid chromatography/tandem MS to map the cross-linking sites as Psb27K63↔CP43D321 (trypsin) and CP43K215↔Psb27D58AGGLK63↔CP43D321 (chymotrypsin), respectively. Our data suggest that Psb27 acts as an important regulatory protein during PSII assembly through specific interactions with the luminal domain of CP43. PMID:22031695

  1. Psb27, a transiently associated protein, binds to the chlorophyll binding protein CP43 in photosystem II assembly intermediates.

    PubMed

    Liu, Haijun; Huang, Richard Y-C; Chen, Jiawei; Gross, Michael L; Pakrasi, Himadri B

    2011-11-01

    Photosystem II (PSII), a large multisubunit pigment-protein complex localized in the thylakoid membrane of cyanobacteria and chloroplasts, mediates light-driven evolution of oxygen from water. Recently, a high-resolution X-ray structure of the mature PSII complex has become available. Two PSII polypeptides, D1 and CP43, provide many of the ligands to an inorganic Mn(4)Ca center that is essential for water oxidation. Because of its unusual redox chemistry, PSII often undergoes degradation followed by stepwise assembly. Psb27, a small luminal polypeptide, functions as an important accessory factor in this elaborate assembly pathway. However, the structural location of Psb27 within PSII assembly intermediates has remained elusive. Here we report that Psb27 binds to CP43 in such assembly intermediates. We treated purified genetically tagged PSII assembly intermediate complexes from the cyanobacterium Synechocystis 6803 with chemical cross-linkers to examine intermolecular interactions between Psb27 and various PSII proteins. First, the water-soluble 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) was used to cross-link proteins with complementary charged groups in close association to one another. In the His27△ctpAPSII preparation, a 58-kDa cross-linked species containing Psb27 and CP43 was identified. This species was not formed in the HT3△ctpA△psb27PSII complex in which Psb27 was absent. Second, the homobifunctional thiol-cleavable cross-linker 3,3'-dithiobis(sulfosuccinimidylpropionate) (DTSSP) was used to reversibly cross-link Psb27 to CP43 in His27△ctpAPSII preparations, which allowed the use of liquid chromatography/tandem MS to map the cross-linking sites as Psb27K(63)↔CP43D(321) (trypsin) and CP43K(215)↔Psb27D(58)AGGLK(63)↔CP43D(321) (chymotrypsin), respectively. Our data suggest that Psb27 acts as an important regulatory protein during PSII assembly through specific interactions with the luminal domain of CP43. PMID:22031695

  2. Antimicrobial activity of four cationic peptides immobilised to poly-hydroxyethylmethacrylate.

    PubMed

    Dutta, Debarun; Kumar, Naresh; D P Willcox, Mark

    2016-04-01

    The objective of this study was to immobilise and characterise a variety of antimicrobial peptides (AMPs) onto poly-hydroxyethylmethacrylate (pHEMA) surfaces to achieve an antibacterial effect. Four AMPs, viz. LL-37, melimine, lactoferricin and Mel-4 were immobilised on pHEMA by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) which assisted covalent attachment. Increasing concentrations of AMPs were immobilised to determine the effect on the adhesion of Pseudomonas aeruginosa and Staphylococcus aureus. The AMP immobilised pHEMAs were characterised by X-ray photoelectron spectroscopy (XPS) to determine the surface elemental composition and by amino acid analysis to determine the total amount of AMP attached. In vitro cytotoxicity of the immobilised pHEMA samples to mouse L929 cells was investigated. Melimine and Mel-4 when immobilised at the highest concentrations showed 3.1 ± 0.6 log and 1.3 ± 0.2 log inhibition against P. aeruginosa, and 3.9 ± 0.6 log and 2.4 ± 0.5 log inhibition against S. aureus, respectively. Immobilisation of LL-37 resulted in up to 2.6 ± 1.0 log inhibition against only P. aeruginosa, but no activity against S. aureus. LFc attachment showed no antibacterial activity. Upon XPS analysis, immobilised melimine, LL-37, LFc and Mel-4 had 1.57 ± 0.38%, 1.13 ± 1.36%, 0.66 ± 0.47% and 0.73 ± 0.32% amide nitrogen attached to pHEMA compared to 0.12 ± 0.14% in the untreated controls. Amino acid analysis determined that the total amount of AMP attachment to pHEMA was 44.3 ± 7.4 nmol, 3.8 ± 0.2 nmol, 6.5 ± 0.6 nmol and 48.9 ± 2.3 nmol for the same peptides respectively. None of the AMP immobilised pHEMA surfaces showed any toxicity towards mouse L929 cells. The immobilisation of certain AMPs at nanomolar concentration to pHEMA is an effective option to develop a stable antimicrobial surface. PMID:26934297

  3. New insights into the effectiveness of alpha-amylase enzyme presentation on the Bacillus subtilis spore surface by adsorption and covalent immobilization.

    PubMed

    Gashtasbi, Fatemeh; Ahmadian, Gholamreza; Noghabi, Kambiz Akbari

    2014-10-01

    Most of the studies in the field of enzyme immobilization have sought to develop a simple, efficient and cost-effective immobilization system. In this study, probiotic Bacillus spores were used as a matrix for enzyme immobilization, because of their inherent resistance to extreme temperatures, UV irradiation, solvents and drying. Above all, their preparation is cost-effective. The alpha-amylase enzyme was immobilized on the spore surface by the covalent and adsorption methods. For the covalent method, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N hydroxysulfosuccinimide (NHS) were used. The maximum concentration of the alpha-amylase immobilized by the two methods onto the spore surface was 360 μg/1.2×10(11) spore. However, maximum activity was achieved at an enzyme concentration of approximately 60 μg/.4×10(10), corresponding to an estimated activity of 8×10(3) IU mg(-1)/1.2×10(11) spore for covalent immobilization and 8.53×10(3) for the adsorption method. After washing the enzyme with 1M NaCl and 0.5% Triton X-100, the enzyme immobilization yield was estimated to be 77% and 20.07% for the covalent and adsorption methods, respectively. The alpha-amylase immobilized by both methods, displayed improved activity in the basic pH range. The optimum pH for the free enzyme was 5 while it shifted to 8 for the immobilized enzyme. The optimum temperatures for the free and immobilized enzymes were 60 °C and 80 °C, respectively. The covalently-immobilized alpha-amylase retained 65% of its initial activity, even after 10 times of recycling. The Km and Vmax values were determined by the GraphPad Prism software, which showed that the Vmax value decreased moderately after immobilization. This is the first study which reports the covalent immobilization of an enzyme on the spore surface. PMID:25152412

  4. Chemical and physical characterization of a novel poly(carbonate urea) urethane surface with protein crosslinker sites.

    PubMed

    Phaneuf, M D; Quist, W C; LoGerfo, F W; Szycher, M; Dempsey, D J; Bide, M J

    1997-10-01

    evaluation slightly increased water vapor permeation through the films. Covalent linkage of the radiolabelled blood protein albumin (125I-BSA) to the cPU segments incubated with the heterobifunctional crosslinker 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) was greatest in the higher percent of ethanol as compared to controls. These results serve as foundation for developing a novel poly(carbonate urea) urethane with physical characteristics comparable to other medical-grade polyurethanes while having protein binding capabilities. PMID:9399137

  5. Response of human macrophages to wound matrices in vitro.

    PubMed

    Witherel, Claire E; Graney, Pamela L; Freytes, Donald O; Weingarten, Michael S; Spiller, Kara L

    2016-05-01

    Chronic wounds remain a major burden to the global healthcare system. Myriad wound matrices are commercially available but their mechanisms of action are poorly understood. Recent studies have shown that macrophages are highly influenced by their microenvironment, but it is not known how different biomaterials affect this interaction. Here, it was hypothesized that human macrophages respond differently to changes in biomaterial properties in vitro with respect to phenotype, including pro-inflammatory M1, anti-inflammatory M2a, known for facilitating extracellular matrix deposition and proliferation, and M2c, which has recently been associated with tissue remodeling. Using multiple donors, it was found that collagen scaffolds cross-linked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide (EDC/NHS) promoted the least inflammatory phenotype in primary human macrophages compared with scaffolds cross-linked with formaldehyde or glutaraldehyde. Importantly, gene expression analysis trends were largely conserved between donors, especially TNFa (M1), CCL22 (M2a), and MRC1 (M2a). Then the response of primary and THP1 monocyte-derived macrophages to four commercially available wound matrices were compared-Integra Dermal Regeneration Template (Integra), PriMatrix Dermal Repair Scaffold (PriMatrix), AlloMend Acellular Dermal Matrix (AlloMend), and Oasis Wound Matrix (Oasis). Gene expression trends were different between primary and THP1 monocyte-derived macrophages for all six genes analyzed in this study. Finally, the behavior of primary macrophages cultured onto the wound matrices over time was analyzed. Integra and Oasis caused down-regulation of M2a markers CCL22 and TIMP3. PriMatrix caused up-regulation of TNFa (M1) and CD163 (M2c) and down-regulation of CCL22 and TIMP3 (both M2a). AlloMend caused up-regulation in CD163 (M2c). Lastly, Oasis promoted the largest increase in the combinatorial M1/M2 score, defined as the sum of M1 genes divided by

  6. Folate-conjugated polymer micelles for active targeting to cancer cells: preparation, in vitro evaluation of targeting ability and cytotoxicity

    NASA Astrophysics Data System (ADS)

    You, Jian; Li, Xin; de Cui, Fu; Du, Yong-Zhong; Yuan, Hong; Hu, Fu qiang

    2008-01-01

    To obtain an active-targeting carrier to cancer cells, folate-conjugated stearic acid grafted chitosan oligosaccharide (Fa-CSOSA) was synthesized by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)-mediated coupling reaction. The substitution degree is 22.1%. The critical micelle concentrations (CMCs) of Fa-CSOSA were 0.017 and 0.0074 mg ml-1 in distilled water and PBS (pH 7.4), respectively. The average volume size range of Fa-CSOSA micelles was 60-120 nm. The targeting ability of Fa-CSOSA micelles was investigated against two kinds of cell lines (A549 and Hela), which have different amounts of folate receptors in their surface. The results indicated that Fa-CSOSA micelles presented a targeting ability to the cells (Hela) with a higher expression of folate receptor during a short-time incubation (<6 h). As incubation proceeded, the special spatial structure of the micelles gradually plays a main role in cellular internalization of the micelles. Good internalization of the micelles into both Hela and A549 cells was shown. Then, paclitaxel (PTX) was encapsulated into the micelles, and the content of PTX in the micelles was about 4.8% (w/w). The average volume size range of PTX-loaded micelles was 150-340 nm. Furthermore, the anti-tumor efficacy in vitro was investigated by a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) method. The IC50 of Taxol (a clinical formulation containing PTX) on A549 and Hela cells was 7.0 and 11.0 µg ml-1, respectively. The cytotoxicity of PTX-loaded micelles was improved sharply (IC50 on A549: 0.32 µg ml-1 IC50 on Hela: 0.268 µg ml-1). This is attributed to the increased intracellular delivery of the drug. The Fa-CSOSA micelles that are presented may be a promising active-targeting carrier candidate via folate mediation.

  7. Bacteriorhodopsin protein hybrids for chemical and biological sensing

    NASA Astrophysics Data System (ADS)

    Winder, Eric Michael

    Bacteriorhodopsin (bR), an optoelectric protein found in Halobacterium salinarum, has the potential for use in protein hybrid sensing systems. Bacteriorhodopsin has no intrinsic sensing properties, however molecular and chemical tools permit production of bR protein hybrids with transducing and sensing properties. As a proof of concept, a maltose binding protein-bacteriorhodopsin ([MBP]-bR) hybrid was developed. It was proposed that the energy associated with target molecule binding, maltose, to the hybrid sensor protein would provide a means to directly modulate the electrical output from the MBP-bR bio-nanosensor platform. The bR protein hybrid is produced by linkage between bR (principal component of purified purple membrane [PM]) and MBP, which was produced by use of a plasmid expression vector system in Escherichia coli and purified utilizing an amylose affinity column. These proteins were chemically linked using 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS), which facilitates formation of an amide bond between a primary carboxylic acid and a primary amine. The presence of novel protein hybrids after chemical linkage was analyzed by SDS-PAGE. Soluble proteins (MBP-only derivatives and unlinked MBP) were separated from insoluble proteins (PM derivatives and unlinked PM) using size exclusion chromatography. The putatively identified MBP-bR protein hybrid, in addition to unlinked bR, was collected. This sample was normalized for bR concentration to native PM and both were deposited onto indium tin oxide (ITO) coated glass slides by electrophoretic sedimentation. The photoresponse of both samples, activated using 100 Watt tungsten lamp at 10 cm distance, were equal at 175 mV. Testing of deposited PM with 1 mM sucrose or 1 mM maltose showed no change in the photoresponse of the material, however addition of 1 mM maltose to the deposited MBP-bR linked hybrid material elicited a 57% decrease in photoresponse

  8. Development of a new antibacterial biomaterial by tetracycline immobilization on calcium-alginate beads.

    PubMed

    Ozseker, Emine Erdogan; Akkaya, Alper

    2016-10-20

    In recent years, increasing risk of infection, caused by resistant microorganism to antibiotics, has become the limelight discovery of new and natural antibacterial materials. Heavy metals, such as silver, copper, mercury and titanium, have antibacterial activity. Products, which improved these metals, do not have stable antibacterial property. Therefore, use of these products is restricted. The aim of this study was to immobilize tetracycline to alginate and improve an antibacterial biomaterial. For this purpose, calcium-alginate beads were formed by dropping to calcium-chloride solution and tetracycline was immobilized to beads using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide at optimum conditions. After immobilization, actualization of immobilization was investigated by analyzing ATR-FTIR spectrum and SEM images. Also, antibacterial property of obtained product was tested. Improved product demonstrated antibacterial property. It has potential for open wound, surgical drapes, bed and pillow sheath in hospitals and it may also be used for increasing human comfort in daily life. PMID:27474587

  9. Polymerization on the rocks: beta-amino acids and arginine

    NASA Technical Reports Server (NTRS)

    Liu, R.; Orgel, L. E.; Bada, J. L. (Principal Investigator)

    1998-01-01

    We have studied the accumulation of long oligomers of beta-amino acids on the surface of minerals using the 'polymerization on the rocks' protocol. We find that long oligopeptides of beta-glutamic acid which cannot be formed in homogeneous aqueous solution are accumulated efficiently on the surface of hydroxylapatite using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDAC) as condensing agent. The EDAC-induced oligomerization of aspartic acid on hydroxylapatite proceeds even more efficiently. Hydroxylapatite can also facilitate the ligation of the tripeptide (glu)3. The 'polymerization on the rocks' scenario is not restricted to negatively-charged amino acids. Oligoarginines are accumulated on the surface of illite using carbonyldiimidizole (CDI) as condensing agent. We find that FeS2 catalyzes the CDI-induced oligomerization of arginine, although it does not adsorb oligoarginines. These results are relevant to the formation of polypeptides on the primitive earth.

  10. Crosslinked hydrogels based on biological macromolecules with potential use in skin tissue engineering.

    PubMed

    Vulpe, Raluca; Popa, Marcel; Picton, Luc; Balan, Vera; Dulong, Virginie; Butnaru, Maria; Verestiuc, Liliana

    2016-03-01

    Zero-length crosslinked hydrogels have been synthesized by covalent linking of three natural polymers (collagen, hyaluronic acid and sericin), in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide. The hydrogels have been investigated by FT-IR spectroscopy, microcalorimetry, in vitro swelling, enzymatic degradation, and in vitro cell viability studies. The obtained crosslinked hydrogels showed a macroporous structure, high swelling degree and in vitro enzymatic resistance compared to uncrosslinked collagen. The in vitro cell viability studies performed on normal human dermal fibroblasts assessed the sericin proliferation properties indicating a potential use of the hydrogels based on collagen, hyaluronic acid and sericin in skin tissue engineering. PMID:26704998

  11. Actinide-Catalyzed Intermolecular Addition of Alcohols to Carbodiimides.

    PubMed

    Batrice, Rami J; Kefalidis, Christos E; Maron, Laurent; Eisen, Moris S

    2016-02-24

    The unprecedented actinide-catalyzed addition of alcohols to carbodiimides is presented. This represents a rare example of thorium-catalyzed transformations of an alcoholic substrate and the first example of uranium complexes showing catalytic reactivity with alcohols. Using the uranium and thorium amides U[N(SiMe3)2]3 and [(Me3Si)2N]2An[κ(2)-(N,C)-CH2Si(CH3)2N(SiMe3)] (An = Th or U), alcohol additions to unsaturated carbon-nitrogen bonds are achieved in short reaction times with excellent selectivities and high to excellent yields. Computational studies, supported by experimental thermodynamic data, suggest plausible models of the profile of the reaction which allow the system to overcome the high barrier of scission of the actinide-oxygen bond. Accompanied by experimentally determined kinetic parameters, a plausible mechanism is proposed for the catalytic cycle. PMID:26844823

  12. EDC-37 Deflagration Rates at Elevated Pressures

    SciTech Connect

    Maienschein, J L; Koerner, J G

    2008-01-31

    We report deflagration rates on EDC-37 at high pressures. Experiments are conducted using the Lawrence Livermore National Laboratory High Pressure Strand Burner (HPSB) apparatus. The HPSB contains a deflagrating sample in a small volume, high pressure chamber. The sample consists of nine, 6.35 mm diameter, 6.35 mm length cylinders stacked on end, with burn wires placed between cylinders. Sample deflagration is limited to the cross-sectional surface of the cylinder by coating the cylindrical surface of the tower with Halthane 88-2 epoxy. Sample deflagration is initiated on one end of the tower by a B/KNO{sub 3} and HNS igniter train. Simultaneous temporal pressure history and burn front time of arrival measurements yield the laminar deflagration rate for a range of pressures and provide insight into deflagration uniformity. These measurements are one indicator of overall thermal explosion violence. Specific details of the experiment and the apparatus can be found in the literature.

  13. The effects of hyaluronic acid incorporated as a wetting agent on lysozyme denaturation in model contact lens materials.

    PubMed

    Weeks, Andrea; Boone, Adrienne; Luensmann, Doerte; Jones, Lyndon; Sheardown, Heather

    2013-09-01

    Conventional and silicone hydrogels as models for contact lenses were prepared to determine the effect of the presence of hyaluronic acid on lysozyme sorption and denaturation. Hyaluronic acid was loaded into poly(2-hydroxyethyl methacrylate) and poly(2-hydroxyethyl methacrylate)/TRIS--methacryloxypropyltris (trimethylsiloxy silane) hydrogels, which served as models for conventional and silicone hydrogel contact lens materials. The hyaluronic acid was cross-linked using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide in the presence of dendrimers. Active lysozyme was quantified using a Micrococcus lysodeikticus assay while total lysozyme was determined using 125-I radiolabeled protein. To examine the location of hyaluronic acid in the gels, 6-aminofluorescein labeled hyaluronic acid was incorporated into the gels using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide chemistry and the gels were examined using confocal laser scanning microscopy. Hyaluronic acid incorporation significantly reduced lysozyme sorption in poly(2-hydroxyethyl methacrylate) (p < 0.00001) and poly(2-hydroxyethyl methacrylate)/TRIS--methacryloxypropyltris (trimethylsiloxy silane) (p < 0.001) hydrogels, with the modified materials sorbing only 20% and 16% that of the control, respectively. More importantly, hyaluronic acid also decreased lysozyme denaturation in poly(2-hydroxyethyl methacrylate) (p < 0.005) and poly(2-hydroxyethyl methacrylate)/TRIS--methacryloxypropyltris (trimethylsiloxy silane) (p < 0.02) hydrogels. The confocal laser scanning microscopy results showed that the hyaluronic acid distribution was dependent on both the material type and the molecular weight of hyaluronic acid. This study demonstrates that hyaluronic acid incorporated as a wetting agent has the potential to reduce lysozyme sorption and denaturation in contact lens applications. The distribution of hyaluronic acid within hydrogels appears to affect denaturation, with more surface mobile, lower

  14. Evaluation of surfactant flushing for remediating EDC-tar contamination.

    PubMed

    Liang, Chenju; Hsieh, Cheng-Lin

    2015-01-01

    Ethylene dichloride tar (EDC-tar) is a dense non-aqueous phase liquid (DNAPL) waste originated from the process of vinyl chloride production, with major constituents including chlorinated aliphatic and aromatic hydrocarbons. This study investigated the feasibility of Surfactant Enhanced Aquifer Remediation (SEAR) for treating EDC-tar contaminated aquifers. Initial experiments explored the potential to enhance the apparent solubility of EDC-tar using single or mixed surfactants. The results showed that an aqueous solution mixed anionic and non-ionic surfactants (i.e., SDS/Tween 80) exhibited higher EDC-tar apparent solubility and lower surface tension than other surfactant systems tested. Additionally, alkaline pH aids in increasing the EDC-tar apparent solubility. In column flushing experiments, it was seen that the alkaline mixed SDS/Tween 80 solution showed better removal of pure EDC-tar from silica sand porous media. Furthermore, separation of EDC-tar in the surfactant solution was conducted employing a salting-out effect. Significant separation of DNAPL was observed when 13 wt.% or more NaCl was added to the solution. Overall, this study evaluates the feasibility of using SEAR for remediating EDC-tar contaminated subsurface soil and groundwater. PMID:25941757

  15. Evaluation of surfactant flushing for remediating EDC-tar contamination

    NASA Astrophysics Data System (ADS)

    Liang, Chenju; Hsieh, Cheng-Lin

    2015-06-01

    Ethylene dichloride tar (EDC-tar) is a dense non-aqueous phase liquid (DNAPL) waste originated from the process of vinyl chloride production, with major constituents including chlorinated aliphatic and aromatic hydrocarbons. This study investigated the feasibility of Surfactant Enhanced Aquifer Remediation (SEAR) for treating EDC-tar contaminated aquifers. Initial experiments explored the potential to enhance the apparent solubility of EDC-tar using single or mixed surfactants. The results showed that an aqueous solution mixed anionic and non-ionic surfactants (i.e., SDS/Tween 80) exhibited higher EDC-tar apparent solubility and lower surface tension than other surfactant systems tested. Additionally, alkaline pH aids in increasing the EDC-tar apparent solubility. In column flushing experiments, it was seen that the alkaline mixed SDS/Tween 80 solution showed better removal of pure EDC-tar from silica sand porous media. Furthermore, separation of EDC-tar in the surfactant solution was conducted employing a salting-out effect. Significant separation of DNAPL was observed when 13 wt.% or more NaCl was added to the solution. Overall, this study evaluates the feasibility of using SEAR for remediating EDC-tar contaminated subsurface soil and groundwater.

  16. SMALL-SCALE IMPACT SENSITIVITY TESTING ON EDC37

    SciTech Connect

    HSU, P C; HUST, G; MAIENSCHEIN, J L

    2008-04-28

    EDC37 was tested at LLNL to determine its impact sensitivity in the LLNL's drop hammer system. The results showed that impact sensitivities of the samples were between 86 cm and 156 cm, depending on test methods. EDC37 is a plastic bonded explosive consisting of 90% HMX, 1% nitrocellulose and binder. We recently conducted impact sensitivity testing in our drop hammer system and the results are presented in this report.

  17. Impact of carbondiimide crosslinker used for magnetic carbon nanotube mediated GFP plasmid delivery.

    PubMed

    Hao, Yuzhi; Xu, Peng; He, Chuan; Yang, Xiaoyan; Huang, Min; Xing, James; Chen, Jie

    2011-07-15

    1-Ethyl-3-(3-dimethylaminopropyl) carbondiimide hydrochloride (EDC) is commonly used as a crosslinker to help bind biomolecules, such as DNA plasmids, with nanostructures. However, EDC often remains, after a crosslink reaction, in the micro-aperture of the nanostructure, e.g., carbon nanotube. The remaining EDC shows positive green fluorescent signals and makes a nanostructure with a strong cytotoxicity which induces cell death. The toxicity of EDC was confirmed on a breast cancer cell line (MCF-7) and two leukemic cell lines (THP-1 and KG-1). The MCF-7 cells mainly underwent necrosis after treatment with EDC, which was verified by fluorescein isothiocyanate (FITC) annexin V staining, video microscopy and scanning electronic microscopy (SEM). If the EDC was not removed completely, the nanostructures with remaining EDC produced a green fluorescent background that could interfere with flow cytometry (FACS) measurement and result in false information about GFP plasmid delivery. Effective methods to remove residual EDC on macromolecules were also developed. PMID:21654030

  18. EDC/NHS cross-linked collagen foams as scaffolds for artificial corneal stroma.

    PubMed

    Vrana, N E; Builles, N; Kocak, H; Gulay, P; Justin, V; Malbouyres, M; Ruggiero, F; Damour, O; Hasirci, V

    2007-01-01

    In this study, a highly porous collagen-based biodegradable scaffold was developed as an alternative to synthetic, non-degradable corneal implants. The developed method involved lyophilization and subsequent stabilization through N-ethyl-N'-[3-dimethylaminopropyl] carbodiimide/N-hydroxy succinimide (EDC/NHS) cross-linking to yield longer lasting, porous scaffolds with a thickness similar to that of native cornea (500 microm). For collagen-based scaffolds, cross-linking is essential; however, it has direct effects on physical characteristics crucial for optimum cell behavior. Hence, the effect of cross-linking was studied by examining the influence of cross-linking on pore size distribution, bulk porosity and average pore size. After seeding the foam with human corneal keratocytes, cell proliferation, cell penetration into the scaffold and ECM production within the scaffold were studied. After a month of culture microscopical and immunohistochemical examinations showed that the foam structure did not undergo any significant loss of integrity, and the human corneal keratocytes populated the scaffold with cells migrating both longitudinally and laterally, and secreted some of the main constituents of the corneal ECM, namely collagen types I, V and VI. The foams had a layer of lower porosity (skin layer) both at the top and the bottom. Foams had an optimal porosity (93.6%), average pore size (67.7 microm), and chemistry for cell attachment and proliferation. They also had a sufficiently rapid degradation rate (73.6+/-1.1% in 4 weeks) and could be produced at a thickness close to that of the natural corneal stroma. Cells were seeded at the top surface of the foams and their numbers there was higher than the rest, basically due to the presence of the skin layer. This is considered to be an advantage when epithelial cells need to be seeded for the construction of hemi or full thickness cornea. PMID:17988518

  19. Assessing EDCs in the Field: Challenges and New Approaches

    EPA Science Inventory

    Assessing the occurrence and effects of EDCs in the environment can be challenging from a number of perspectives. For example, conventional analytical approaches and/or toxicity tests may not be appropriate to detecting very potent chemicals that impact specific pathways, and oft...

  20. Minireview: Epigenomic Plasticity and Vulnerability to EDC Exposures.

    PubMed

    Walker, Cheryl Lyn

    2016-08-01

    The epigenome undergoes significant remodeling during tissue and organ development, which coincides with a period of exquisite sensitivity to environmental exposures. In the case of endocrine-disrupting compounds (EDCs), exposures can reprogram the epigenome of developing tissues to increase susceptibility to diseases later in life, a process termed "developmental reprogramming." Both DNA methylation and histone modifications have been shown to be vulnerable to disruption by EDC exposures, and several mechanisms have been identified by which EDCs can reprogram the epigenome. These include altered methyl donor availability, loss of imprinting control, changes in dioxygenase activity, altered expression of noncoding RNAs, and activation of cell signaling pathways that can phosphorylate, and alter the activity of, histone methyltransferases. This altered epigenomic programming can persist across the life course, and in some instances generations, to alter gene expression in ways that correlate with increased disease susceptibility. Together, these studies on developmental reprogramming of the epigenome by EDCs are providing new insights into epigenomic plasticity that is vulnerable to disruption by environmental exposures. PMID:27355193

  1. EVALUATION OF DRINKING WATER TREATMENT TECHNIQUES FOR EDC REMOVAL

    EPA Science Inventory

    Many of the chemicals identified as potential endocrine disrupting chemicals (EDCs) may be present in surface or ground waters used as drinking water sources, due to their disposal via domestic and industrial sewage treatment systems and wet-weather runoff. In order to decrease t...

  2. Carbodiimide or periodate method to prepare peroxidase hydrazide for its use in immunoassay.

    PubMed

    Shrivastav, Tulsidas G

    2004-01-01

    Peroxidase hydrazides were prepared by conjugating horseradish peroxidase (HRP) to adipic acid dihydrazide (ADH) by carbodiimide or periodate oxidation method. The resulting HRP hydrazides (ADH-HRP) were conjugated to cortisol-21-hemisuccinate (cortisol-21-HS) by forming diimide bonds using the N-hydroxysuccinimide (NHS) carbodiimide mediated reaction. The prepared cortisol-21-HS-ADH-HRP enzyme conjugates were utilized for the development of an enzyme linked immunosorbent assay (ELISA) for direct estimation of cortisol. To the cortisol antibody coated microtiter wells, standard or serum sample (50 microL), along with 100 microL of cortisol-21-HS-ADH-HRP enzyme conjugate (ADH-HRP used is prepared by either carbodiimide or periodate oxidation method), was incubated for 1 hr at 37 degrees C. Bound enzyme activity was measured by using tetramethyl benzidine/hydrogen peroxide (TMB/H202) as substrate. The sensitivity, specificity, and recovery of the assays were found to be identical when ELISAs were employed with cortisol enzyme conjugates prepared by conjugating cortisol-21-HS to HRP hydrazide, made either by the carbodiimide method or periodate oxidation method. PMID:15461389

  3. ASSESSEMNT OF GONAD SIZE IN TAUTOGOLABRUS ADSPERSUS AS AN INDICATOR OF REPRODUCTION AND EDC EXPOSURE

    EPA Science Inventory

    Cunner habitat includes estuarine and marine areas where sewage treatment and other discharges containing estrogenic (EDCs) are likely.

    Endocrine-disrupting chemicals (EDCs) in the environment may disturb the population dynamics of wildlife by affecting their reproductive...

  4. A review of the genotoxicity of 1,2-dichloroethane (EDC).

    PubMed

    Gwinn, Maureen R; Johns, Douglas O; Bateson, Thomas F; Guyton, Kathryn Z

    2011-01-01

    1,2-Dichloroethane (EDC, CAS#107-06-2) is a high production volume halogenated aliphatic hydrocarbon that is used mainly in the manufacture of vinyl chloride. EDC has been found in ambient and residential air samples, as well as in groundwater, surface water and drinking water. EDC has been well-studied in a variety of genotoxicity assays, and appears to involve the metabolic activation of the parent compound. We critically evaluated the genotoxicity data of EDC and its metabolites as part of an evaluation of carcinogenic mechanisms of action of EDC. EDC is genotoxic in multiple test systems via multiple routes of exposure. EDC has been shown to induce DNA adduct formation, gene mutations and chromosomal aberrations in the presence of key activation enzymes (including CYP450s and/or GSTs) in laboratory animal and in vitro studies. EDC was negative for clastogenesis as measured by the micronucleus assay in mice. In general, an increased level of DNA damage is observed related to the GSH-dependent bioactivation of EDC. Increased chromosomal aberrations with increased CYP450 expression were suggestive of a role for the oxidative metabolites of EDC in inducing chromosomal damage. Taken together, these studies demonstrate that EDC exposure, in the presence of key enzymes (including CYP450s and/or GSTs), leads to DNA adduct formation, gene mutations and chromosomal aberrations. PMID:21255676

  5. Carbodiimide cross-linking of amniotic membranes in the presence of amino acid bridges.

    PubMed

    Lai, Jui-Yang

    2015-06-01

    The purpose of this study was to investigate the carbodiimide cross-linking of amniotic membrane (AM) in the presence of amino acid bridges. The biological tissues were treated with glycine, lysine, or glutamic acid and chemically cross-linked to examine the role of amino acid types in collagenous biomaterial processing. Results of zeta potential measurements showed that the use of uncharged, positively and negatively charged amino acids dictates the charge state of membrane surface. Tensile strength and water content measurements demonstrated that the addition of lysine molecules to the cross-linking system can increase the cross-linking efficiency and dehydration degree while the introduction of glutamic acid in the AM samples decreases the number of cross-links per unit mass of chemically modified tissue collagen. The differences in the cross-linking density further determined the thermal and biological stability by differential scanning calorimetry and in vitro degradation tests. As demonstrated in matrix permeability studies, the improved formation of covalent cross-linkages imposed by lysine facilitated construction of stronger cross-linking structures. In contrast, the added glycine molecules were insufficient to enhance the resistances of the proteinaceous matrices to thermal denaturation and enzymatic degradation. The cytocompatibility of these biological tissue membranes was evaluated by using human corneal epithelial cell cultures. Results of cell viability, metabolic activity, and pro-inflammatory gene expression level showed that the AM materials cross-linked with carbodiimide in the presence of different types of amino acids are well tolerated without evidence of detrimental effect on cell growth. In addition, the amino acid treated and carbodiimide cross-linked AM implants had good biocompatibility in the anterior chamber of the rabbit eye model. Our findings suggest that amino acid type is a very important engineering parameter to mediate

  6. Annulation of thioimidates and vinyl carbodiimides to prepare 2-aminopyrimidines, competent nucleophiles for intramolecular alkyne hydroamination. Synthesis of (-)-crambidine.

    PubMed

    Perl, Nicholas R; Ide, Nathan D; Prajapati, Sudeep; Perfect, Hahdi H; Durón, Sergio G; Gin, David Y

    2010-02-17

    A convergent synthesis of (-)-crambidine is reported. The sequence capitalizes on two novel key transformations, including a [4+2] annulation of thioimidates with vinyl carbodiimides and an alkyne hydroamination employing 2-aminopyrimidine nucleophiles. PMID:20095555

  7. Preparation and properties of EDC/NHS mediated crosslinking poly (gamma-glutamic acid)/epsilon-polylysine hydrogels.

    PubMed

    Hua, Jiachuan; Li, Zheng; Xia, Wen; Yang, Ning; Gong, Jixian; Zhang, Jianfei; Qiao, Changsheng

    2016-04-01

    In this paper, a novel pH-sensitive poly (amino acid) hydrogel based on poly γ-glutamic acid (γ-PGA) and ε-polylysine (ε-PL) was prepared by carbodiimide (EDC) and N-hydroxysuccinimide (NHS) mediated polymerization. The influence of PGA/PL molar ratio and EDC/NHS concentration on the structure and properties was studied. Fourier transform infrared spectroscopy (FT-IR) and X-ray photoelectron spectroscopy (XPS) proved that hydrogels were crosslinked through amide bond linkage, and the conversion rate of a carboxyl group could reach 96%. Scanning electron microscopy (SEM) results showed a regularly porous structure with 20 μm pore size in average. The gelation time in the crosslink process of PGA/PL hydrogels was within less than 5 min. PGA/PL hydrogels had excellent optical performance that was evaluated by a novel optotype method. Furthermore, PGA/PL hydrogels were found to be pH-sensitive, which could be adjusted to the pH of swelling media intelligently. The terminal pH of swelling medium could be controlled at 5 ± 1 after equilibrium when the initial pH was within 3-11. The swelling kinetics was found to follow a Voigt model in deionized water but a pseudo-second-order model in normal saline and phosphate buffer solution, respectively. The differential swelling degrees were attributed to the swelling theory based on the different ratio of -COOH/-NH2 and pore size in hydrogels. The results of mechanical property indicated that PGA/PL hydrogels were soft and elastic. Moreover, PGA/PL hydrogels exhibited excellent biocompatibility by cell proliferation experiment. PGA/PL hydrogels could be degraded in PBS solution and the degradation rate was decreased with the increase of the molar ratio of PL. Considering the simple preparation process and pH-sensitive property, these PGA/PL hydrogels might have high potential for use in medical and clinical fields. PMID:26838920

  8. Performance of Electrostatic Dust Collectors (EDCs) for Endotoxin Assessment in Homes: Effect of Mailing, Placement, Heating and Electrostatic Charge

    PubMed Central

    Kilburg-Basnyat, Brita; Metwali, Nervana; Thorne, Peter S.

    2016-01-01

    Electrostatic Dust Collectors (EDCs) are in use for passive sampling of bioaerosols, but particular aspects of their performance have not yet been evaluated. This study investigated the effect of mailing EDCs on endotoxin loading and the effect of EDC deployment in front of and away from heated ventilation on endotoxin sampling. Endotoxin sampling efficiency of heated and unheated EDC cloths was also evaluated. Cross-country express mailing of dust-spiked EDCs yielded no significant changes in endotoxin concentrations compared to dust-only samples for both high spiked-EDCs (p=0.30) and low spiked-EDCs (p=0.36). EDCs were also deployed in 20 identical apartments with one EDC placed in front of the univent heater in each apartment and contemporaneous EDC placed on the built-in bookshelf in each apartment. The endotoxin concentrations were significantly different (p=0.049) indicating that the placement of EDC does impact endotoxin sampling. Heated and unheated EDCs were deployed for 7 days in pairs in farm homes. There was a significant difference between endotoxin concentrations (p=0.027) indicating that heating EDCs may diminish their electrostatic capabilities and impact endotoxin sampling. The last study investigated the electrostatic charge of 12 heated and 12 unheated EDC cloths. There was a significant difference in charge (p=0.009) which suggests that heating EDC cloths may make them less effective for sampling. In conclusion, EDCs can be mailed to and from deployment sites, EDC placement in relationship to ventilation is crucial, and heating EDCs reduces their electrostatic charge which may diminish their endotoxin sampling capabilities. PMID:26325020

  9. Performance of electrostatic dust collectors (EDCs) for endotoxin assessment in homes: Effect of mailing, placement, heating, and electrostatic charge.

    PubMed

    Kilburg-Basnyat, Brita; Metwali, Nervana; Thorne, Peter S

    2016-01-01

    Electrostatic Dust Collectors (EDCs) are in use for passive sampling of bioaerosols, but particular aspects of their performance have not yet been evaluated. This study investigated the effect of mailing EDCs on endotoxin loading and the effect of EDC deployment in front of, and away from, heated ventilation on endotoxin sampling. Endotoxin sampling efficiency of heated and unheated EDC cloths was also evaluated. Cross-country express mailing of dust-spiked EDCs yielded no significant changes in endotoxin concentrations compared to dust-only samples for both high-spiked EDCs (p = 0.30) and low-spiked EDCs (p = 0.36). EDCs were also deployed in 20 identical apartments with one EDC placed in front of the univent heater in each apartment and contemporaneous EDC placed on the built-in bookshelf in each apartment. The endotoxin concentrations were significantly different (p = 0.049) indicating that the placement of EDC does impact endotoxin sampling. Heated and unheated EDCs were deployed for 7 days in pairs in farm homes. There was a significant difference between endotoxin concentrations (p = 0.027) indicating that heating EDCs may diminish their electrostatic capabilities and impact endotoxin sampling. The last study investigated the electrostatic charge of 12 heated and 12 unheated EDC cloths. There was a significant difference in charge (p = 0.009) which suggests that heating EDC cloths may make them less effective for sampling. In conclusion, EDCs can be mailed to and from deployment sites, EDC placement in relationship to ventilation is crucial, and heating EDCs reduces their electrostatic charge which may diminish their endotoxin sampling capabilities. PMID:26325020

  10. Modification of poly(L-lactic acid) electrospun fibers and films with poly(propylene imine) dendrimer

    NASA Astrophysics Data System (ADS)

    Khaliliazar, Sh.; Akbari, S.; Kish, M. H.

    2016-02-01

    Poly(L-lactic acid) (PLLA) electrospun fibers and films were modified with the second generation of poly(propylene imine) dendrimer (PPI-G2) by three different approaches, namely, sodium hydroxide hydrolysis, plasma treatment and direct application of PPI-G2. For the first and the second approaches, PLLA was modified by sodium hydroxide hydrolysis or plasma treatment to produce carboxylic acid groups. Then, the carboxylic acid groups were activated by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDAC) and N,N‧-dicyclohexyl carbodiimide (DCC) as a hetero bi-functional cross-linker. The cross-linkers promoted the grafting of carboxylic acid groups on the modified PLLA with NH2 groups of PPI-G2. In the third approach, the PPI-G2 dendrimer was directly used as an aminolysis agent for the functionalization of PLLA in a one step process. FTIR analysis confirmed the presence of sbnd NH2 groups of PPI-G2 on the modified PLLA samples, resulting from each one of the three modification methods. Studies by SEM shows bead free electrospun fibers. Also, FE-SEM shows nano-cracks on the surface of films after modification. Contact angle, drug release tests, antibacterial effects and the dying results confirmed that these functionalization methods increased hydrophilicity and reactive side-chains of PLLA in the wet chemical process resulted in providing host-guest properties on the PLLA surface for adsorbing various kinds of guest molecules.

  11. EUROLAS Data Center (EDC) - A new website for tracking the SLR data flow

    NASA Astrophysics Data System (ADS)

    Schwatke, C.

    2012-04-01

    Since the foundation of the International Laser Ranging Service (ILRS) in 1998, the EDC acts as one of the two global ILRS data centers, the Crustal Dynamics Data Information System (CDDIS) and the EUROLAS data center (EDC) at the DGFI. In 2009, the EDC became also an ILRS Operation Center (OS), which has the responsibility to verify the format of all incoming SLR observations. The SLR observations are submitted in different formats (Normal Points (CSTG), Normal Point (CRD), Full Rate (MERIT-II) and Full Rate (CRD)). In addition to the SLR observations, the EDC treats predictions (CPF), orbits and station coordinates. All data and products are handled in the internal data flow at the EDC. After processing data and products successful without errors they will be stored on the FTP at EDC (ftp://edc.dgfi.badw.de) or send via mail to the users. This web site (http://edc.dgfi.badw.de) contains an interface for station managers for uploading and quality control of tracking data. The interface gives feedback in case the upload was sucessful or a detailed description of errors is displayed. In addition summary information is provided on the latest data acquired by satellite and station.

  12. Organization versus activation: the role of endocrine-disrupting contaminants (EDCs) during embryonic development in wildlife.

    PubMed Central

    Guillette, L J; Crain, D A; Rooney, A A; Pickford, D B

    1995-01-01

    Many environmental contaminants disrupt the vertebrate endocrine system. Although they may be no more sensitive to endocrine-disrupting contaminants (EDCs) than other vertebrates, reptiles are good sentinels of exposure to EDCs due to the lability in their sex determination. This is exemplified by a study of alligators at Lake Apopka, Florida, showing that EDCs have altered the balance of reproductive hormones resulting in reproductive dysfunction. Such alterations may be activationally or organizationally induced. Much research emphasizes the former, but a complete understanding of the influence of EDCs in nature can be generated only after consideration of both activational and organizational alterations. The organizational model suggests that a small quantity of an EDC, administered during a specific period of embryonic development, can permanently modify the organization of the reproductive, immune, and nervous systems. Additionally, this model helps explain evolutionary adaptations to naturally occurring estrogenic compounds, such as phytoestrogens. PMID:8593864

  13. Heat-induced reshaping and coarsening of metal nanoparticle-graphene oxide hybrids

    NASA Astrophysics Data System (ADS)

    Pan, Hanqing

    coalesce or undergo reshaping at a lower temperature. Nanoparticle- and nanorod-graphene oxide hybrid materials were also used to study the effect of covalent and non-covalent interactions between gold nanoparticles or nanorods and graphene oxide during coarsening or reshaping, respectively. Non-covalent interactions were studied by directly adding graphene oxide to aqueous solutions containing water-soluble metal nanoparticles or nanorods, and covalent interactions were achieved by the self-assembly of the same nanoparticles onto thiolated graphene oxide that was prepared by coupling L-cystine using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS). When nanoparticles and nanorods are attached to graphene oxide through additional -covalent bonds, they are more strongly immobilized and therefore would undergo less coalescence and slower reshaping upon heating.

  14. The methods of identification, analysis, and removal of endocrine disrupting compounds (EDCs) in water.

    PubMed

    Chang, Hyun-Shik; Choo, Kwang-Ho; Lee, Byungwhan; Choi, Sang-June

    2009-12-15

    The information regarding endocrine disrupting compounds (EDCs) was reviewed, including the definition and characteristics, the recent research trends concerning identification and analytical methods, and the applicable removal processes. EDCs include various types of natural and synthetic chemical compounds presenting the mimicking or inhibition of the reproductive action of the endocrine system in animals and humans. The ubiquitous presence with trace level concentrations and the wide diversity are the reported characteristics of EDCs. Biologically based assays seem to be a promising method for the identification of EDCs. On the other hand, mass-based analytical methods show excellent sensitivity and precision for their quantification. Several extraction techniques for the instrumental analysis have been developed since they are crucial in determining overall analytical performances. Conventional treatment techniques, including coagulation, precipitation, and activated sludge processes, may not be highly effective in removing EDCs, while the advanced treatment options, such as granular activated carbon (GAC), membrane, and advanced oxidation processes (AOPs), have shown satisfactory results. The oxidative degradation of some EDCs was associated with aromatic moieties in their structure. Further studies on EDCs need to be conducted, such as source reduction, limiting exposure to vulnerable populations, treatment or remediation of contaminated sites, and the detailed understanding of transport mechanisms in the environment. PMID:19632774

  15. EDCs DataBank: 3D-Structure database of endocrine disrupting chemicals.

    PubMed

    Montes-Grajales, Diana; Olivero-Verbel, Jesus

    2015-01-01

    Endocrine disrupting chemicals (EDCs) are a group of compounds that affect the endocrine system, frequently found in everyday products and epidemiologically associated with several diseases. The purpose of this work was to develop EDCs DataBank, the only database of EDCs with three-dimensional structures. This database was built on MySQL using the EU list of potential endocrine disruptors and TEDX list. It contains the three-dimensional structures available on PubChem, as well as a wide variety of information from different databases and text mining tools, useful for almost any kind of research regarding EDCs. The web platform was developed employing HTML, CSS and PHP languages, with dynamic contents in a graphic environment, facilitating information analysis. Currently EDCs DataBank has 615 molecules, including pesticides, natural and industrial products, cosmetics, drugs and food additives, among other low molecular weight xenobiotics. Therefore, this database can be used to study the toxicological effects of these molecules, or to develop pharmaceuticals targeting hormone receptors, through docking studies, high-throughput virtual screening and ligand-protein interaction analysis. EDCs DataBank is totally user-friendly and the 3D-structures of the molecules can be downloaded in several formats. This database is freely available at http://edcs.unicartagena.edu.co. PMID:25451822

  16. Roles of Edc3 in the oxidative stress response and CaMCA1-encoded metacaspase expression in Candida albicans.

    PubMed

    Jung, Jong-Hwan; Kim, Jinmi

    2014-11-01

    The Edc3 protein is an enhancer of mRNA decapping, and acts as a scaffold protein for the mRNA granules that are known as processing bodies in yeast. In the pathogenic yeast Candida albicans, various stresses, such as glucose depletion, oxidative stress, and filamentation defects, induce the accumulation of processing bodies. Here, we report that the edc3/edc3 deletion strain showed increased resistance to various stresses, including hydrogen peroxide, acetic acid, and high temperature. Oxidative stress is known to induce the intracellular accumulation of reactive oxygen species (ROS) and apoptotic cell death in C. albicans. We found that the ROS level was lower in edc3/edc3 cells than in wild-type cells following oxidative stress. We also observed that expression of the metacaspase gene CaMCA1 was decreased in edc3/edc3 cells. Overexpression of CaMCA1 suppressed the decreased accumulation of ROS and the increased resistance to hydrogen peroxide in edc3/edc3 cells. The catalase Cat1 and the superoxide dismutase Sod1 were upregulated in edc3/edc3 cells as compared with wild-type cells. On the basis of these findings, we suggest that EDC3 plays a critical role in the expression of CaMCA1 and the oxidative stress response in C. albicans. PMID:25158786

  17. The Measured Temperature and Pressure of EDC37 detonation products

    NASA Astrophysics Data System (ADS)

    Ferguson, James; Richley, James; Ota, Tom; Sutton, Ben; Price, Ed

    2015-06-01

    We present the experimentally determined temperature and pressure of the detonation products of EDC37; a HMX based conventional high explosive. These measurements were performed on a series of cylinder tests. The temperature measurements were performed at the end of the cylinder with optical fibres observing the bare explosive through a LiF window. The temperature of the products was measured for 2 microseconds using single colour pyrometry, multicolour pyrometry and spectroscopy with the results from all three methods being consistent. The peak temperature was found to be ~ 3600 K dropping to ~ 2400 K at the end of the measurement window. The spectroscopy was time integrated and showed that the emission spectra can be approximated using a grey body curve with no other emission or absorption lines being present. The pressure was obtained using an analytical method which used the velocity of the expanding cylinder wall, measured using heterodyne velocimetry (HetV), and the velocity of detonation, measured with chirped fibre Bragg gratings. The pressure drops from an initial CJ value of ~38 GPa to ~4 GPa at the end of the 2 microsecond temperature measurement window.

  18. In Vitro Selection of Optimal DNA Substrates for Ligation by a Water-Soluble Carbodiimide

    NASA Technical Reports Server (NTRS)

    Harada, Kazuo; Orgel, Leslie E.

    1994-01-01

    We have used in vitro selection to investigate the sequence requirements for efficient template-directed ligation of oligonucleotides at 0 deg C using a water-soluble carbodiimide as condensing agent. We find that only 2 bp at each side of the ligation junction are needed. We also studied chemical ligation of substrate ensembles that we have previously selected as optimal by RNA ligase or by DNA ligase. As anticipated, we find that substrates selected with DNA ligase ligate efficiently with a chemical ligating agent, and vice versa. Substrates selected using RNA ligase are not ligated by the chemical condensing agent and vice versa. The implications of these results for prebiotic chemistry are discussed.

  19. Nitrile imines: matrix isolation, IR spectra, structures, and rearrangement to carbodiimides.

    PubMed

    Bégué, Didier; Qiao, Greg GuangHua; Wentrup, Curt

    2012-03-21

    The structures and reactivities of nitrile imines are subjects of continuing debate. Several nitrile imines were generated photochemically or thermally and investigated by IR spectroscopy in Ar matrices at cryogenic temperatures (Ph-CNN-H 6, Ph-CNN-CH(3)17, Ph-CNN-SiMe(3)23, Ph-CNN-Ph 29, Ph(3)C-CNN-CPh(3)34, and the boryl-CNN-boryl derivative 39). The effect of substituents on the structures and IR absorptions of nitrile imines was investigated computationally at the B3LYP/6-31G* level. IR spectra were analyzed in terms of calculated anharmonic vibrational spectra and were generally in very good agreement with the calculated spectra. Infrared spectra were found to reflect the structures of nitrile imines accurately. Nitrile imines with IR absorptions above 2200 cm(-1) have essentially propargylic structures, possessing a CN triple bond (typically PhCNNSiMe(3)23, PhCNNPh 29, and boryl-CNN-boryl 39). Nitrile imines with IR absorptions below ca. 2200 cm(-1) are more likely to be allenic (e.g., HCNNH 1, PhCNNH 6, HCNNPh 43, PhCNNCH(3)17, and Ph(3)C-CNN-CPh(3)34). All nitrile imines isomerize to the corresponding carbodiimides both thermally and photochemically. Monosubstituted carbodiimides isomerize thermally to the corresponding cyanamides (e.g., Ph-N═C═N-H 5 → Ph-NH-CN 8), which are therefore the thermal end products for nitrile imines of the types RCNNH and HCNNR. This tautomerization is reversible under flash vacuum thermolysis conditions. PMID:22364289

  20. Transition Metal Carbodiimides as Molecular Negative Electrode Materials for Lithium and Sodium Ion Batteries with Excellent Cycling Properties

    SciTech Connect

    Sougrati, M. T.; Darwiche, Ali; Liu, Xiaohiu; Mahmoud, A.; Hermann, Raphael P; Jouen, Samuel; Monconduit, Laure; Dronskowski, Richard; Stievano, L.

    2016-01-01

    We report evidence for the electrochemical activity of transition-metal carbodiimides versus lithium and sodium. In particular, iron carbodiimide, FeNCN, can be efficiently used as a negative electrode material for alkali-metal-ion batteries, similar to its oxide analogue FeO. Based on 57Fe M ssbauer and infrared spectroscopy (IR) data, the electrochemical reaction mechanism can be explained by the reversible transformation of the Fe NCN into Li/Na NCN bonds during discharge and charge. These new electrode materials exhibit higher capacity compared to well-established negative electrode references such as graphite or hard carbon. Contrary to its oxide analogue, iron carbodiimide does not require heavy treatments (nanoscale tailoring, sophisticated textures, coating etc.) to obtain long cycle life with density current as high as 9 A/g for hundreds of charge/discharge cycles. Similar to the iron compound, several other transition-metal carbodiimides Mx(NCN)y with M = Mn, Cr, Zn can cycle successfully versus lithium and sodium. Their electrochemical activity and performances open the way to the design of a novel family of anode materials.

  1. Transition-Metal Carbodiimides as Molecular Negative Electrode Materials for Lithium- and Sodium-Ion Batteries with Excellent Cycling Properties

    DOE PAGESBeta

    Sougrati, Moulay T.; Darwiche, Ali; Liu, Xiaohiu; Mahmoud, Abdelfattah; Hermann, Raphael P.; Jouen, Samuel; Monconduit, Laure; Dronskowski, Richard; Stievano, Lorenzo

    2016-03-16

    Here we report evidence for the electrochemical activity of transition-metal carbodiimides versus lithium and sodium. In particular, iron carbodiimide, FeNCN, can be efficiently used as a negative electrode material for alkali-metal-ion batteries, similar to its oxide analogue FeO. Based on 57Fe M ssbauer and infrared spectroscopy (IR) data, the electrochemical reaction mechanism can be explained by the reversible transformation of the Fe NCN into Li/Na NCN bonds during discharge and charge. These new electrode materials exhibit higher capacity compared to well-established negative electrode references such as graphite or hard carbon. Contrary to its oxide analogue, iron carbodiimide does not requiremore » heavy treatments (nanoscale tailoring, sophisticated textures, coating etc.) to obtain long cycle life with density current as high as 9 A/g-1 for hundreds of charge/discharge cycles. Similar to the iron compound, several other transition-metal carbodiimides Mx(NCN)y with M = Mn, Cr, Zn can cycle successfully versus lithium and sodium. Ultimately, their electrochemical activity and performances open the way to the design of a novel family of anode materials.« less

  2. Transition-Metal Carbodiimides as Molecular Negative Electrode Materials for Lithium- and Sodium-Ion Batteries with Excellent Cycling Properties.

    PubMed

    Sougrati, Moulay T; Darwiche, Ali; Liu, Xiaohiu; Mahmoud, Abdelfattah; Hermann, Raphael P; Jouen, Samuel; Monconduit, Laure; Dronskowski, Richard; Stievano, Lorenzo

    2016-04-11

    We report evidence for the electrochemical activity of transition-metal carbodiimides versus lithium and sodium. In particular, iron carbodiimide, FeNCN, can be efficiently used as negative electrode material for alkali-metal-ion batteries, similar to its oxide analogue FeO. Based on (57)Fe Mössbauer and infrared spectroscopy (IR) data, the electrochemical reaction mechanism can be explained by the reversible transformation of the Fe-NCN into Li/Na-NCN bonds during discharge and charge. These new electrode materials exhibit higher capacity compared to well-established negative electrode references such as graphite or hard carbon. Contrary to its oxide analogue, iron carbodiimide does not require heavy treatments (such as nanoscale tailoring, sophisticated textures, or coating) to obtain long cycle life with current density as high as 9 A g(-1) for hundreds of charge-discharge cycles. Similar to the iron compound, several other transition-metal carbodiimides M(x)(NCN)y with M=Mn, Cr, Zn can cycle successfully versus lithium and sodium. Their electrochemical activity and performance open the way to the design of a novel family of anode materials. PMID:26989882

  3. Effects of carbon-based nanoparticles (CNPs) on the fate of endocrine disrupting chemicals (EDCs) in different agricultural soils.

    NASA Astrophysics Data System (ADS)

    Stumpe, Britta; Wolski, Sabrina; Marschner, Bernd

    2013-04-01

    Nanotechnology is a major innovative scientific and economic growth area. To date there is a lack about possible adverse effects that may be associated with manufactured nanomaterial in terrestrial environments. Since it is known that on the one hand carbon-based nanoparticles (CNPs) and endocrine disrupting chemicals (EDCs) strongly interact in wastewater and that on the other hand CNPs and EDCs are released together via wastewater irrigation to agricultural soils, knowledge of CNP effects on the EDC fate in the soil environment is needed for further risk assessments. The overall goal of this project is to gain a better understanding of interaction of CNPs with EDCs within the soil system. Three different soil samples were applied with different CNPs, EDCs and CNP-EDC complexes and incubated over a period of 6 weeks. The EDC mineralization as well as their uptake by soil microorganisms was monitored to describe impacts of the nanomaterial on the EDC fate. As quality control for the biological soil activity soil respiration, enzyme activities and the soil microbial biomass were monitored in all incubated soil samples. Clearly, EDCs bound in CNP complexes showed a decrease in mineralization. While the free EDCs showed a total mineralization of 34 to 45 %, the nano complexed EDCs were only mineralized to 12 to 15 %. Since no effects of the nanomaterial on the biological soil activity were observed, we conclude that the reduced EDC mineralization is directly linked to their interaction with the CNPs. Since additionally the EDC adsorption to CNPs reduced the EDC uptake by soil microorganism, we assume that CNPs generally form more or less recalcitrant aggregates which likely protect the associated EDCs from degradation.

  4. DEVELOPMENT OF CHEMICAL METHODS TO CHARACTERIZE EXPOSURE TO EDCS IN THE NEUSE RIVER BASIN

    EPA Science Inventory

    To develop a quantitative health and environmental risk assessment of endocrine disrupting compounds (EDCs), information on exposures is essential. A full exposure assessment has complex requirements that require preliminary information to direct further research in this area....

  5. Effect of Transpiration on Plant Accumulation and Translocation of PPCP/EDCs

    PubMed Central

    Dodgen, Laurel K; Ueda, Aiko; Wu, Xiaoqin; Parker, David R; Gan, Jay

    2015-01-01

    The reuse of treated wastewater for agricultural irrigation in arid and hot climates where plant transpiration is high may affect plant accumulation of pharmaceutical and personal care products (PPCPs) and endocrine disrupting chemicals (EDCs). In this study, carrot, lettuce, and tomato plants were grown in solution containing 16 PPCP/EDCs in either a cool-humid or a warm-dry environment. Leaf bioconcentration factors (BCF) were positively correlated with transpiration for chemical groups of different ionized states (p < 0.05). However, root BCFs were correlated with transpiration only for neutral PPCP/EDCs (p < 0.05). Neutral and cationic PPCP/EDCs showed similar accumulation, while anionic PPCP/EDCs had significantly higher accumulation in roots and significantly lower accumulation in leaves (p < 0.05). Results show that plant transpiration may play a significant role in the uptake and translocation of PPCP/EDCs, which may have a pronounced effect in arid and hot climates where irrigation with treated wastewater is common. PMID:25594843

  6. Effect of transpiration on plant accumulation and translocation of PPCP/EDCs.

    PubMed

    Dodgen, Laurel K; Ueda, Aiko; Wu, Xiaoqin; Parker, David R; Gan, Jay

    2015-03-01

    The reuse of treated wastewater for agricultural irrigation in arid and hot climates where plant transpiration is high may affect plant accumulation of pharmaceutical and personal care products (PPCPs) and endocrine disrupting chemicals (EDCs). In this study, carrot, lettuce, and tomato plants were grown in solution containing 16 PPCP/EDCs in either a cool-humid or a warm-dry environment. Leaf bioconcentration factors (BCF) were positively correlated with transpiration for chemical groups of different ionized states (p < 0.05). However, root BCFs were correlated with transpiration only for neutral PPCP/EDCs (p < 0.05). Neutral and cationic PPCP/EDCs showed similar accumulation, while anionic PPCP/EDCs had significantly higher accumulation in roots and significantly lower accumulation in leaves (p < 0.05). Results show that plant transpiration may play a significant role in the uptake and translocation of PPCP/EDCs, which may have a pronounced effect in arid and hot climates where irrigation with treated wastewater is common. PMID:25594843

  7. Collagen-hyaluronic acid scaffolds for adipose tissue engineering.

    PubMed

    Davidenko, N; Campbell, J J; Thian, E S; Watson, C J; Cameron, R E

    2010-10-01

    Three-dimensional (3-D) in vitro models of the mammary gland require a scaffold matrix that supports the development of adipose stroma within a robust freely permeable matrix. 3-D porous collagen-hyaluronic acid (HA: 7.5% and 15%) scaffolds were produced by controlled freeze-drying technique and crosslinking with 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride. All scaffolds displayed uniform, interconnected pore structure (total porosity approximately 85%). Physical and chemical analysis showed no signs of collagen denaturation during the formation process. The values of thermal characteristics indicated that crosslinking occurred and that its efficiency was enhanced by the presence of HA. Although the crosslinking reduced the swelling of the strut material in water, the collagen-HA matrix as a whole tended to swell more and show higher dissolution resistance than pure collagen samples. The compressive modulus and elastic collapse stress were higher for collagen-HA composites. All the scaffolds were shown to support the proliferation and differentiation 3T3-L1 preadipocytes while collagen-HA samples maintained a significantly increased proportion of cycling cells (Ki-67+). Furthermore, collagen-HA composites displayed significantly raised Adipsin gene expression with adipogenic culture supplementation for 8 days vs. control conditions. These results indicate that collagen-HA scaffolds may offer robust, freely permeable 3-D matrices that enhance mammary stromal tissue development in vitro. PMID:20466086

  8. Porous ovalbumin scaffolds with tunable properties: a resource-efficient biodegradable material for tissue engineering applications.

    PubMed

    Luo, Baiwen; Choong, Cleo

    2015-01-01

    Natural materials are promising alternatives to synthetic materials used in tissue engineering applications as they have superior biocompatibility and promote better cell attachment and proliferation. Ovalbumin, a natural polymer found in avian egg white, is an example of a nature-derived material. Despite the availability and reported biocompatibility of ovalbumin, limited research has been carried out to investigate the efficacy of ovalbumin-based scaffolds for adipose tissue engineering applications. Hence, the current study was carried out to investigate the effect of different crosslinkers on ovalbumin scaffold properties as first step towards the development of ovalbumin-based scaffolds for adipose tissue engineering applications. In this study, highly porous three-dimensional scaffolds were fabricated by using three different crosslinkers: glutaraldehyde, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and 1,4-butanediol diglycidyl ether. Results showed that the overall scaffold properties such as morphology, pore size and mechanical properties could be modulated based on the type and concentration of crosslinkers used during the fabrication process. Subsequently, the efficacy of the different scaffolds for supporting cell proliferation was investigated. In vitro degradation was also carried on for the best scaffold based on the mechanical and cellular results. Overall, this study is a demonstration of the viability of ovalbumin-based scaffolds as cell carriers for soft tissue engineering applications. PMID:25158688

  9. Radioprotection provides functional mechanics but delays healing of irradiated tendon allografts after ACL reconstruction in sheep.

    PubMed

    Seto, Aaron U; Culp, Brian M; Gatt, Charles J; Dunn, Michael

    2013-12-01

    Successful protection of tissue properties against ionizing radiation effects could allow its use for terminal sterilization of musculoskeletal allografts. In this study we functionally evaluate Achilles tendon allografts processed with a previously developed radioprotective treatment based on (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide) crosslinking and free radical scavenging using ascorbate and riboflavin, for ovine anterior cruciate ligament reconstruction. Arthroscopic anterior cruciate ligament (ACL) reconstruction was performed using double looped allografts, while comparing radioprotected irradiated and fresh frozen allografts after 12 and 24 weeks post-implantation, and to control irradiated grafts after 12 weeks. Radioprotection was successful at preserving early subfailure mechanical properties comparable to fresh frozen allografts. Twelve week graft stiffness and anterior-tibial (A-T) translation for radioprotected and fresh frozen allografts were comparable at 30 % of native stiffness, and 4.6 and 5 times native A-T translation, respectively. Fresh frozen allograft possessed the greatest 24 week peak load at 840 N and stiffness at 177 N/mm. Histological evidence suggested a delay in tendon to bone healing for radioprotected allografts, which was reflected in mechanical properties. There was no evidence that radioprotective treatment inhibited intra-articular graft healing. This specific radioprotective method cannot be recommended for ACL reconstruction allografts, and data suggest that future efforts to improve allograft sterilization procedures should focus on modifying or eliminating the pre-crosslinking procedure. PMID:23842952

  10. Mapping the surface-exposed regions of papaya mosaic virus nanoparticles.

    PubMed

    Rioux, Gervais; Majeau, Nathalie; Leclerc, Denis

    2012-06-01

    In general, the structure of the papaya mosaic virus (PapMV) and other members of the potexviruses is poorly understood. Production of PapMV coat proteins in a bacterial expression system and their self-assembly in vitro into nanoparticles is a very useful tool to study the structure of this virus. Using recombinant PapMV nanoparticles that are similar in shape and appearance to the plant virus, we evaluated surface-exposed regions by two different methods, immunoblot assay and chemical modification with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide or diethyl-pyrocarbonate followed by mass spectrometry. Three regions were targeted by the two techniques. The N- and C-termini were shown to be surfaced exposed as expected. However, the region 125-136 was revealed for the first time as the major surface-exposed region of the nanoparticles. The presence of linear peptides at the surface was finally confirmed using antibodies directed to those peptides. It is likely that region 125-136 plays a key role in the lifecycle of PapMV and other members of the potexvirus group. PMID:22524169

  11. Formulation and characterisation of antibody-conjugated soy protein nanoparticles--implications for neutralisation of snake venom with improved efficiency.

    PubMed

    Renu, Kadali; Gopi, Kadiyala; Jayaraman, Gurunathan

    2014-12-01

    The present study reports the formulation of soy protein nanoparticles and its conjugation to antivenom. The conditions for nanoparticle formation were optimised by considering particle size, count rate, stability and zeta potential. The smallest particle size of 70.9 ± 0.9 nm with a zeta potential of -28.0 ± 1.4 mV was obtained at pH 6.2, with NaOH 5.4 % and 28 μg/mg glutaraldehyde. The nanoparticle was conjugated with antisnake venom immunoglobulins (F(ab')2 fragments) using 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide. TEM analysis indicated the increased size of particle to 600 nm after conjugation to antivenom. Further, in vitro studies indicated that conjugated antibodies inhibited the activity of protease, phospholipase and hyaluronidase enzymes of Bungarus caeruleus venom more efficiently than the free antivenom. This is the first report on the use of protein nanoparticles for conjugating snake venom antibodies and their implications for neutralising snake venom enzymes with increased efficiency. PMID:25185504

  12. Thermostable trypsin conjugates immobilized to biogenic magnetite show a high operational stability and remarkable reusability for protein digestion

    NASA Astrophysics Data System (ADS)

    Pečová, M.; Šebela, M.; Marková, Z.; Poláková, K.; Čuda, J.; Šafářová, K.; Zbořil, R.

    2013-03-01

    In this work, magnetosomes produced by microorganisms were chosen as a suitable magnetic carrier for covalent immobilization of thermostable trypsin conjugates with an expected applicability for efficient and rapid digestion of proteins at elevated temperatures. First, a biogenic magnetite was isolated from Magnetospirillum gryphiswaldense and its free surface was coated with the natural polysaccharide chitosan containing free amino and hydroxy groups. Prior to covalent immobilization, bovine trypsin was modified by conjugating with α-, β- and γ-cyclodextrin. Modified trypsin was bound to the magnetic carriers via amino groups using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysulfosuccinimide as coupling reagents. The magnetic biomaterial was characterized by magnetometric analysis and electron microscopy. With regard to their biochemical properties, the immobilized trypsin conjugates showed an increased resistance to elevated temperatures, eliminated autolysis, had an unchanged pH optimum and a significant storage stability and reusability. Considering these parameters, the presented enzymatic system exhibits properties that are superior to those of trypsin forms obtained by other frequently used approaches. The proteolytic performance was demonstrated during in-solution digestion of model proteins (horseradish peroxidase, bovine serum albumin and hen egg white lysozyme) followed by mass spectrometry. It is shown that both magnetic immobilization and chemical modification enhance the characteristics of trypsin making it a promising tool for protein digestion.

  13. Application of suspension array for simultaneous detection of four different mycotoxins in corn and peanut.

    PubMed

    Wang, Ying; Ning, Baoan; Peng, Yuan; Bai, Jialei; Liu, Ming; Fan, Xianjun; Sun, Zhiyong; Lv, Zhiqiang; Zhou, Caihong; Gao, Zhixian

    2013-03-15

    Mycotoxins are highly toxic contaminants in foodstuffs and feedstuffs. The study presents a novel suspension array technology for quantifying four mycotoxins, namely, aflatoxin B1, deoxynivalenol, T-2 toxin, and zearalenone, in corn and peanut. Using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride, the complete antigens of the mycotoxins became attached to the microspheres with viable activity. The optimal concentrations of each antibody and biotin-rabbit anti-goat IgG were obtained through chessboard titration. The four mycotoxins were detected simultaneously and quantitatively in corn and peanut using indirect competitive immunoassay. Multi-channel standard curves with appropriate logistic correlation (R(2)>0.9819) were respectively plotted. The broad working ranges with three to four orders of magnitude were calculated, and limits of detection at the pg level were found to be better than those obtained using high-performance liquid chromatography. The recovery rates in the actual samples generally ranged from 80.16% to 117.65%, with an intra-assay coefficient of variation lower than 15%, which indicated high accuracy and repeatability. A suspension array method for the simultaneous detection of the four mycotoxins within 4h was successfully developed using minimal samples; the method was proven to have high throughput, flexibility, accuracy and reproducibility. The approach could detect multiple contaminants in actual samples. PMID:23017676

  14. Identification of AFB1-interacting proteins and interactions between RPSA and AFB1.

    PubMed

    Zhuang, Zhenhong; Huang, Yaling; Yang, Yanling; Wang, Shihua

    2016-01-15

    A method using immobilized affinity chromatography (IAC) was developed to screen for aflatoxin B1 (AFB1)-binding proteins. AFB1 and bovine serum albumin (BSA) coupled protein (BSA-AFB1) was prepared using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride. The resulting coupled compound was immobilized onto PVDF transfer membranes, which were then incubated with total protein from mouse liver. AFB1-binding proteins were eluted, after non-specific washing, by specific elution, and the eluted proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Two candidate AFB1-binding proteins were identified by liquid chromatography-tandem mass spectrometry as the 40S ribosomal protein SA (RPSA) and a putative uncharacterized protein. RPSA and AFB1 interactions were further analyzed by ELISA in vitro and laser confocal immunofluorescence analysis in vivo. The results from ELISA and immunofluorescence showed that RPSA efficiently bound AFB1 in vitro and in vivo. This study's conclusion laid the foundation for further exploration of the role of AFB1-binding proteins in AFB1 toxicology towards hepatocytes and the entry pathway of AFB1 into hepatocytes. PMID:26372695

  15. Application of quantum-dots for analysis of nanosystems by either utilizing or preventing FRET

    NASA Astrophysics Data System (ADS)

    Kim, Joong H.; Chaudhary, Sumit; Stephens, Jared P.; Singh, Krishna V.; Ozkan, Mihrimah

    2005-04-01

    We have developed conjugates with quantum-dots (QDs) for the purpose of analysis of nanosystems that are organic or inorganic in nature such as DNA and carbon nanotubes. First, by employing Florescence Resonant Energy Transfer (FRET) principles, a hybrid molecular beacon conjugates are synthesized. For water- solubilization of QDs, we modified the surface of CdSe-ZnS core-shell QD by using mercaptoacetic acid ligand. This modification does not affect the size of QDs from that of unmodified QDs. After linking molecular beacons to the carboxyl groups of the modified QDs using 1-Ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride, hybrid molecular beacons are prepared as a DNA probe. After hybridization with specific target DNA and non-specific target DNA, the hybrid conjugates show high specificity to the target DNA with 5-fold increase in the intensity of fluorescence. By developing atomic model of the conjugates, we calculated with 8 numbers of molecular beacons on a single quantum dots, we could increase the efficiency of FRET up to 90%. In other hands, for application of quantum dots to the carbon nanotubes, FRET is a barrier. Thus, after employing 1 % sodium-dodecyl-sulfonate (SDS), single-walled carbon nanotubes are decorated with QDs at their outer surface. This enables fluorescent microscopy imaging of single-walled carbon nanotubes which is a more common technique than electron microscopy. In summary, QDs can be used for analysis or detection of both organic and inorganic based nanosystems.

  16. Design of a versatile chemical assembly method for patterning colloidal nanoparticles

    NASA Astrophysics Data System (ADS)

    Choi, J. H.; Adams, S. M.; Ragan, R.

    2009-02-01

    Poly(methyl methacrylate) (PMMA) domains in phase-separated polystyrene-b-poly(methyl methacrylate) (PS-b-PMMA) diblock copolymer thin films were chemically modified for controlled placement of solution synthesized Au nanoparticles having a mean diameter of 24 nm. Colloidal Au nanoparticles functionalized with thioctic acid were immobilized on amine functionalized PMMA domains on the PS-b-PMMA template using 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride linking chemistry and N-hydroxy sulfosuccinimide stabilizer. Atomic force microscopy and scanning electron microscopy images demonstrated immobilization of Au nanoparticles commensurate with PMMA domains. Nanoparticles form into clusters of single particles, dimers, and linear chains as directed by the PMMA domain size and shape. Capillary forces influence the spacing between Au nanoparticles on PMMA domains. Inter-particle spacings below 3 nm were achieved and these assemblies of closely spaced nanoparticle clusters are expected to exhibit strong localized electromagnetic fields. Thus, these processes and material systems provide an experimental platform for studying resonantly enhanced excitations of surface plasmons as a function of material and geometric structure as well as utilization in catalytic applications.

  17. Synthesis and characterization of Pseudomonas aeruginosa alginate-tetanus toxoid conjugate.

    PubMed

    Kashef, Nasim; Behzadian-Nejad, Qorban; Najar-Peerayeh, Shahin; Mousavi-Hosseini, Kamran; Moazzeni, Mohammad; Djavid, Gholamreza Esmaeeli

    2006-10-01

    Chronic infection with Pseudomonas aeruginosa is the main proven perpetrator of lung function decline and ultimate mortality in cystic fibrosis (CF) patients. Mucoid strains of this bacterium elaborate mucoid exopolysaccharide, also referred to as alginate. Alginate-based immunization of naïve animals elicits opsonic antibodies and leads to clearance of mucoid P. aeruginosa from the lungs. Alginate was isolated from mucoid P. aeruginosa strain 8821M by repeated ethanol precipitation, dialysis, proteinase and nuclease digestion, and chromatography. To improve immunogenicity, the purified antigen was coupled to tetanus toxoid (TT) with adipic acid dihydrazide (ADH) as a spacer and 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDAC) as a linker. The reaction mixture was passed through a Sepharose CL-4B column. The resulting conjugate was composed of TT and large-size alginate polymer at a ratio of about 3 : 1; it was non-toxic and non-pyrogenic, and elicited high titres of alginate-specific IgG. Antisera raised against the conjugate had high opsonic activity against the vaccine strain. The alginate conjugate was also able to protect mice against a lethal dose of mucoid P. aeruginosa. These data indicate that an alginate-based vaccine has significant potential to protect against chronic infection with mucoid P. aeruginosa in the CF host. PMID:17005795

  18. Optimization of the conjugation method for a serogroup B/C meningococcal vaccine.

    PubMed

    Fukasawa, Lucila O; Schenkman, Rocilda P F; Perciani, Catia T; Carneiro, Sylvia M; Dias, Waldely O; Tanizaki, Martha M

    2006-11-01

    A conjugate meningococcal vaccine against serogroup B/C consisting of capsular PS (polysaccharide) from serogroup C conjugated to OMV (outer membrane vesicle) from serogroup B would be a very useful vaccine in regions where there is a prevalence of both serogroups, for example in Brazil. For this purpose, the conjugation method that uses ADHy (adipic acid dihydrazide) as spacer and a carbodi-imide derivative, EDAC [1-ethyl-3-(3-dimethylaminopropyl)carbodi-imide], as catalyser was optimized looking for synthesis yield and maintenance of the antigenicity of both components. The best synthesis conditions preserving the vaccine immunogenicity resulted in a final yield of approx. 17%. Immunogenicity of the vaccine was highest when 10% of the sialic acid residues of the PS were occupied by the ADHy spacer. Sterilization of the conjugate by filtration through a 0.22-microm-pore-size membrane resulted in a low recovery of protein and PS (approximately 50%), although the vaccine immunogenicity was maintained. Using gamma irradiation on freeze-dried sample, it was possible to maintain the integrity of OMV structure and, consequently, its ability to induce bactericidal antibodies. PMID:16776648

  19. Pseudomonas aeruginosa PAO-1 Lipopolysaccharide-Diphtheria Toxoid Conjugate Vaccine: Preparation, Characterization and Immunogenicity

    PubMed Central

    Najafzadeh, Faezeh; Shapouri, Reza; Rahnema, Mehdi; Rokhsartalab Azar, Shadi; Kianmehr, Anvarsadat

    2015-01-01

    Background: Treatment of Pseudomonas aeruginosa PAO-1 infections through immunological means has been proved to be efficient and protective. Objectives: The purpose of this study was to produce a conjugate vaccine composed of detoxified lipopolysaccharide (D-LPS) P. aeruginosa and diphtheria toxoid (DT). Materials and Methods: Firstly, LPS was purified and characterized from P. aeruginosa PAO1 and then detoxified. D-LPS was covalently coupled to DT as a carrier protein via amidation method with adipic acid dihydrazide (ADH) as a spacer molecule and 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide (EDAC) as a linker. The molar ratio of LPS to DT in the prepared conjugate was 3:1. The immunogenicity of D-LPS-DT conjugate vaccine in mice model was evaluated as well. Results: The conjugate was devoid of endotoxin activity and 0.125 U/mL of D-LPS was acceptable for immunization. D-LPS-DT conjugate was nonpyrogenic for rabbits and nontoxic for mice. Mice immunization with D-LPS-DT conjugate vaccine elicited the fourfold higher IgG antibody compared to D-LPS. Anti-LPS IgG antibody was predominantly IgG1 subclass and then IgG3, IgG2a and IgG2b, respectively. Conclusions: Vaccine based on the conjugation of P. aeruginosa PAO-1 LPS with DT increased anti-LPS antibodies and had a significant potential to protect against Pseudomonas infections. PMID:26301059

  20. Synthesis, characterization and immunological properties of Escherichia coli 0157:H7 lipopolysaccharide- diphtheria toxoid conjugate vaccine

    PubMed Central

    Rokhsartalab-Azar, Shadi; Shapouri, Reza; Rahnema, Mehdi; Najafzadeh, Faezeh

    2015-01-01

    Background and Objective: Escherichia coli O157:H7, an emerging pathogen, causes severe enteritis and the extraintestinal complication of hemolytic-uremic syndrome. The goal of this study was to evaluate the conjugate of E. coli O157: H7 lipopolysaccharide (LPS) with diphtheria toxoid (DT) as a candidate vaccine in mice model. Material and Methods: LPS from E. coli O157:H7 was extracted by hot phenol method and then detoxified. Purified LPS was coupled to DT with adipic acid dihydrazide (ADH) as a spacer and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDAC) as a linker. The coupling molar ratio of LPS to DT was 3:1. Clinical evaluation of E. coli O157:H7 LPS-DT conjugate was also performed. Results: The conjugate was devoid of endotoxin activity and indicated 0.125 U/ml of D-LPS. Mice immunization with D-LPS DT conjugate elicited fourfold higher IgG antibody in comparison to D-LPS. Also, in vivo protection of mice with conjugate provided high protection against the LD50 of E. coli O157:H7, which indicated a good correlation with the IgG titer. Conclusion: Our results showed that the suggested vaccine composed of E. coli O157:H7 LPS and DT had a significant potential to protect against E. coli infections. PMID:26668702

  1. Films prepared from electrosterically stabilized nanocrystalline cellulose.

    PubMed

    Yang, Han; Tejado, Alvaro; Alam, Nur; Antal, Miro; van de Ven, Theo G M

    2012-05-22

    Electrosterically stabilized nanocrystalline cellulose (ENCC) was modified in three ways: (1) the hydroxyl groups on C2 and C3 of glucose repeat units of ENCC were converted to aldehyde groups by periodate oxidation to various extents; (2) the carboxyl groups in the sodium form on ENCC were converted to the acid form by treating them with an acid-type ion-exchange resin; and (3) ENCC was cross-linked in two different ways by employing adipic dihydrazide as a cross-linker and water-soluble 1-ethyl-3-[3-(dimethylaminopropyl)] carbodiimide as a carboxyl-activating agent. Films were prepared from these modified ENCC suspensions by vacuum filtration. The effects of these three modifications on the properties of films were investigated by a variety of techniques, including UV-visible spectroscopy, a tensile test, thermogravimetric analysis (TGA), the water vapor transmission rate (WVTR), and contact angle (CA) studies. On the basis of the results from UV spectra, the transmittance of these films was as high as 87%, which shows them to be highly transparent. The tensile strength of these films was increased with increasing aldehyde content. From TGA and WVTR experiments, cross-linked films showed much higher thermal stability and lower water permeability. Furthermore, although the original cellulose is hydrophilic, these films also exhibited a certain hydrophobic behavior. Films treated by trichloromethylsilane become superhydrophobic. The unique characteristics of these transparent films are very promising for potential applications in flexible packaging and other high-technology products. PMID:22482733

  2. Fish collagen/alginate/chitooligosaccharides integrated scaffold for skin tissue regeneration application.

    PubMed

    Chandika, Pathum; Ko, Seok-Chun; Oh, Gun-Woo; Heo, Seong-Yeong; Nguyen, Van-Tinh; Jeon, You-Jin; Lee, Bonggi; Jang, Chul Ho; Kim, GeunHyung; Park, Won Sun; Chang, Wonseok; Choi, Il-Whan; Jung, Won-Kyo

    2015-11-01

    An emerging paradigm in wound healing techniques is that a tissue-engineered skin substitute offers an alternative approach to create functional skin tissue. Here we developed a fish collagen/alginate (FCA) sponge scaffold that was functionalized by different molecular weights of chitooligosaccharides (COSs) with the use of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride as a cross-linking agent. The effects of cross-linking were analyzed by Fourier transform infrared spectroscopy. The results indicate that the homogeneous materials blending and cross-linking intensity were dependent on the molecular weights of COSs. The highly interconnected porous architecture with 160-260μm pore size and over 90% porosity and COS's MW driven swelling and retention capacity, tensile property and in vitro biodegradation behavior guaranteed the FCA/COS scaffolds for skin tissue engineering application. Further improvement of these properties enhanced the cytocompatibility of all the scaffolds, especially the scaffolds containing COSs with MW in the range of 1-3kDa (FCA/COS1) showed the best cytocompatibility. These physicochemical, mechanical, and biological properties suggest that the FCA/COS1 scaffold is a superior candidate that can be used for skin tissue regeneration. PMID:26306410

  3. Optical mechanical refinement of human amniotic membrane by dehydration and cross-linking.

    PubMed

    Tanaka, Yuji; Kubota, Akira; Yokokura, Shunji; Uematsu, Masafumi; Shi, Dong; Yamato, Masayuki; Okano, Teruo; Quantock, Andrew J; Nishida, Kohji

    2012-10-01

    The aim of this study was to develop a method for refining the optical and mechanical properties of human amniotic membrane (AM) to provide ophthalmic transparent implants for use during severe donor cornea shortages. AM was allowed to gradually dehydrate at 4-8 °C with and without chemical cross-linking. To improve the transparency of AM, a simple dehydration process using a refrigerator at 4-8 °C overnight was examined. For further improvements, dehydrated AM was then cross-linked with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxy-succimide before rehydration. Light transmittance and tensile strength of individual specimens were evaluated. Light transmittance of AM improved from 50.9-77.7% at 550 nm by this simple low temperature dehydration process. Its high light transmittance was partially maintained at 70.1%, even after rehydration with normal saline. Interestingly, chemically cross-linked AM showed a significantly greater light transmittance of 81.5% under wet conditions. In addition, tensile strength was significantly increased after cross-linking from 2.32 N/mm(2) (native tissue) to 11.78 N/mm(2) (cross-linked specimens). Light transmittance and tensile strength were successfully improved by these approaches, including low temperature dehydration with and without chemical cross-linking. The use of refined AM could be feasible for use in current and future ophthalmic treatments. PMID:22489071

  4. Preparation and characterization of self-assembled nanoparticles based on low-molecular-weight heparin and stearylamine conjugates for controlled delivery of docetaxel

    PubMed Central

    Kim, Dong-Hwan; Termsarasab, Ubonvan; Cho, Hyun-Jong; Yoon, In-Soo; Lee, Jae-Young; Moon, Hyun Tae; Kim, Dae-Duk

    2014-01-01

    Low-molecular-weight heparin (LMWH)–stearylamine (SA) conjugates (LHSA)-based self-assembled nanoparticles were prepared for intravenous delivery of docetaxel (DCT). 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysuccinimide were used as coupling agents for synthesis of LHSA conjugates. The physicochemical properties, in vitro antitumor efficacy, in vitro cellular uptake efficiency, in vivo antitumor efficacy, and in vivo pharmacokinetics of LHSA nanoparticles were investigated. The LHSA nanoparticles exhibited a spherical shape with a mean diameter of 140–180 nm and a negative surface charge. According to in vitro release and in vivo pharmacokinetic test results, the docetaxel-loaded LHSA5 (LMWH:SA =1:5) nanoparticles exhibited sustained drug release profiles. The blank LHSA nanoparticles demonstrated only an insignificant cytotoxicity in MCF-7 and MDAMB 231 human breast cancer cells; additionally, higher cellular uptake of coumarin 6 (C6) in MCF-7 and MDAMB 231 cells was observed in the LHSA5 nanoparticles group than that in the C6 solution group. The in vivo tumor growth inhibition efficacy of docetaxel-loaded LHSA5 nanoparticles was also significantly higher than the Taxotere®-treated group in the MDAMB 231 tumor-xenografted mouse model. These results indicated that the LHSA5-based nanoparticles could be a promising anticancer drug delivery system. PMID:25525355

  5. Label-free photoelectrochemical immunosensor for sensitive detection of Ochratoxin A.

    PubMed

    Yang, Jiaojiao; Gao, Picheng; Liu, Yixin; Li, Rongxia; Ma, Hongmin; Du, Bin; Wei, Qin

    2015-02-15

    A general label-free photoelectrochemical (PEC) platform was manufactured by assembly of CdSe nanoparticles (NPs) sensitized anatase TiO2-functionalized electrode via layer-by-layer (LBL) strategy. CdSe NPs were assembled on anatase TiO2-functionalized electrode through dentate binding of TiO2 NPs to -COOH groups. Ascorbic acid (AA) was used as an efficient electron donor for scavenging photogenerated holes under visible-light irradiation. The photocurrent response of the CdSe NPs modified electrode was significantly enhanced as a result of the band alignment of CdSe and TiO2 in electrolyte. Ochratoxin A (OTA), as model analyte, was employed to investigate the performance of the PEC platform. Antibodies of OTA were immobilized on CdSe sensitized electrode by using the classic 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride coupling reactions between -COOH groups on the surfaces of CdSe NPs and -NH2 groups of the antibody. Under the optimized conditions, the photocurrent was proportional to OTA concentration range from 10pg/mL to 50ng/mL with detection limit of 2.0pg/mL. The employed PEC platform established a simple, fast and inexpensive strategy for fabrication of label-free biosensor, which might be widely applied in bioanalysis and biosensing in the future. PMID:25173733

  6. Relevance of fiber integrated gelatin-nanohydroxyapatite composite scaffold for bone tissue regeneration

    NASA Astrophysics Data System (ADS)

    Halima Shamaz, Bibi; Anitha, A.; Vijayamohan, Manju; Kuttappan, Shruthy; Nair, Shantikumar; Nair, Manitha B.

    2015-10-01

    Porous nanohydroxyapatite (nanoHA) is a promising bone substitute, but it is brittle, which limits its utility for load bearing applications. To address this issue, herein, biodegradable electrospun microfibrous sheets of poly(L-lactic acid)-(PLLA)-polyvinyl alcohol (PVA) were incorporated into a gelatin-nanoHA matrix which was investigated for its mechanical properties, the physical integration of the fibers with the matrix, cell infiltration, osteogenic differentiation and bone regeneration. The inclusion of sacrificial fibers like PVA along with PLLA and leaching resulted in improved cellular infiltration towards the center of the scaffold. Furthermore, the treatment of PLLA fibers with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide enhanced their hydrophilicity, ensuring firm anchorage between the fibers and the gelatin-HA matrix. The incorporation of PLLA microfibers within the gelatin-nanoHA matrix reduced the brittleness of the scaffolds, the effect being proportional to the number of layers of fibrous sheets in the matrix. The proliferation and osteogenic differentiation of human adipose-derived mesenchymal stem cells was augmented on the fibrous scaffolds in comparison to those scaffolds devoid of fibers. Finally, the scaffold could promote cell infiltration, together with bone regeneration, upon implantation in a rabbit femoral cortical defect within 4 weeks. The bone regeneration potential was significantly higher when compared to commercially available HA (Surgiwear™). Thus, this biomimetic, porous, 3D composite scaffold could be offered as a promising candidate for bone regeneration in orthopedics.

  7. Phosphatidylcholine covalently linked to a methacrylate-based monolith as a biomimetic stationary phase for capillary liquid chromatography.

    PubMed

    Moravcová, Dana; Carrasco-Correa, Enrique Javier; Planeta, Josef; Lämmerhofer, Michael; Wiedmer, Susanne K

    2015-07-10

    In this study a strategy to immobilize phospholipids onto a polymer-based stationary phase is described. Methacrylate-based monoliths in capillary format (150×0.1mm) were modified by soybean phosphatidylcholine through 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide coupling to obtain stationary phases suitable to mimic cell surface membranes. The covalent coupling reaction involves the phosphate group in phospholipids; therefore, the described methodology is suitable for all types of phospholipids. Immobilization of soy bean phosphatidylcholine on the monolith was confirmed by attenuated total reflectance Fourier transform infrared spectroscopy and gas chromatography-mass spectrometry of the fatty alcohol profile, generated upon reductive cleavage of the fatty acyl side chains of the phospholipid on the monolith surface with lithium aluminium hydride. The prepared stationary phases were evaluated through studies on the retention of low-molar mass model analytes including neutral, acidic, and basic compounds. Liquid chromatographic studies confirmed predominant hydrophobic interactions between the analytes and the synthesized stationary phase; however, electrostatic interactions contributed to the retention as well. The synthesized columns showed high stability even with fully aqueous mobile phases such as Dulbecco's phosphate-buffered saline solution. PMID:26024990

  8. Folic acid-CdTe quantum dot conjugates and their applications for cancer cell targeting

    SciTech Connect

    Suriamoorthy, Preethi; Zhang, Xing; Hao, Guiyang; Joly, Alan G.; Singh, S.; Hossu, Marius; Sun, Xiankai; Chen, Wei

    2010-12-01

    In this study, we report the preparation,luminescence, and targeting properties of folic acid- CdTe quantum dot conjugates. Water-soluble CdTe quantum dots were synthesized and conjugated with folic acid using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide-N-hydroxysuccinimide chemistry. The in-fluence of folic acid on the luminescence properties of CdTe quantum dots was investigated, and no energy transfer between them was observed. To investigate the efficiency of folic acid-CdTe nanoconjugates for tumor targeting, pure CdTe quantum dots and folic acid-coated CdTe quantum dots were incubated with human naso- pharyngeal epidermal carcinoma cell line with positive expressing folic acid receptors (KB cells) and lung cancer cells without expression of folic acid receptors (A549 cells). For the cancer cells with positive folate receptors (KB cells), the uptake for CdTe quantum dots is very low, but for folic acid-CdTe nanoconjugates, the uptake is very high. For the lung cancer cells without folate receptors (A549 cells), the uptake for folic acid- CdTe nanoconjugates is also very low. The results indicate that folic acid is an effective targeting molecule for tumor cells with overexpressed folate receptors.

  9. Hyaluronic acid as an internal wetting agent in model DMAA/TRIS contact lenses.

    PubMed

    Weeks, Andrea; Luensmann, Doerte; Boone, Adrienne; Jones, Lyndon; Sheardown, Heather

    2012-11-01

    Model silicone hydrogel contact lenses, comprised of N,N-dimethylacrylamide and methacryloxypropyltris (trimethylsiloxy) silane, were fabricated and hyaluronic acid (HA) was incorporated as an internal wetting agent using a dendrimer-based method. HA and dendrimers were loaded into the silicone hydrogels and cross-linked using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide chemistry. The presence and location of HA in the hydrogels was confirmed using X-ray photoelectron spectroscopy and confocal laser scanning microscopy, respectively. The effects of the presence of HA on the silicone hydrogels on hydrophilicity, swelling behavior, transparency, and lysozyme sorption and denaturation were evaluated. The results showed that HA increased the hydrophilicity and the equilibrium water content of the hydrogels without affecting transparency. HA also significantly decreased the amount of lysozyme sorption (p < 0.002). HA had no effect on lysozyme denaturation in hydrogels containing 0% and 1.7% methacrylic acid (MAA) (by weight) but when the amount of MAA was increased to 5%, the level of lysozyme denaturation was significantly lower compared to control materials. These results suggest that HA has great potential to be used as a wetting agent in silicone hydrogel contact lenses to improve wettability and to decrease lysozyme sorption and denaturation. PMID:21750182

  10. Development and evaluation of buccoadhesive tablet for selegiline hydrochloride based on thiolated polycarbophil.

    PubMed

    Wasnik, Mangesh N; Godse, Rutika D; Nair, Hema A

    2014-05-01

    Selegiline hydrochloride (SHCl), a monoamine oxidase B inhibitor, is used as an adjunct in the therapy of Parkinson's disease. This study is concerned with the preparation and evaluation of mucoadhesive buccal tablet for controlled systemic delivery of SHCl. Buccal absorption of selegiline can bypass its first-pass metabolism and improve bioavailability accompanied by greatly reduced metabolite formation, which is potentially of enhanced therapeutic value in patients with Parkinson's disease. Polycarbophil-cysteine (PCP-cys) conjugate, which is a thiolated derivative of the mucoadhesive polymer polycarbophil, was synthesized by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride-mediated amide bond coupling. Tablets of SHCl based on native and thiolated polycarbophil were prepared. The prepared tablets were evaluated for drug content, swelling behavior, mucoadhesive strength, in vitro drug release, ex vivo permeation and in vitro cytotoxicity. PCP-cys tablets showed enhanced mucoadhesion and retarded drug release compared to polycarbophil tablets. Permeation data of SHCl from matrices prepared using the PCP-cys polymer revealed a significantly higher value of apparent permeability in comparison to polycarbophil, which supported the information in literature that thiolation imparts permeation enhancing properties to mucoadhesive polymers. In vitro cytotoxicity studies on PCP-cys using L-929 mouse fibroblast cell line indicated that conjugation with cysteine does not impart any apparent toxicity to polycarbophil. The results from the study indicate that the buccal delivery of SHCl using thiolated polycarbophil tablet could provide a way for improved therapy of Parkinson's disease. PMID:24517570

  11. Use of Protein Cross-Linking and Radiolytic Labeling To Elucidate the Structure of PsbO within Higher-Plant Photosystem II.

    PubMed

    Mummadisetti, Manjula P; Frankel, Laurie K; Bellamy, Henry D; Sallans, Larry; Goettert, Jost S; Brylinski, Michal; Bricker, Terry M

    2016-06-14

    We have used protein cross-linking with the zero-length cross-linker 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide, and radiolytic footprinting coupled with high-resolution tandem mass spectrometry, to examine the structure of higher-plant PsbO when it is bound to Photosystem II. Twenty intramolecular cross-linked residue pairs were identified. On the basis of this cross-linking data, spinach PsbO was modeled using the Thermosynechococcus vulcanus PsbO structure as a template, with the cross-linking distance constraints incorporated using the MODELLER program. Our model of higher-plant PsbO identifies several differences between the spinach and cyanobacterial proteins. The N-terminal region is particularly interesting, as this region has been suggested to be important for oxygen evolution and for the specific binding of PsbO to Photosystem II. Additionally, using radiolytic mapping, we have identified regions on spinach PsbO that are shielded from the bulk solvent. These domains may represent regions on PsbO that interact with other components, as yet unidentified, of the photosystem. PMID:27203407

  12. A simple preparation of a stable CdS-polyacrylamide nanocomposite: structure, thermal and optical properties.

    PubMed

    Bach, Long Giang; Islam, M Rafiqul; Hong, Seong-Soo; Lim, Kwon Taek

    2013-11-01

    A facile approach was employed for the preparation of stable luminescent nanocomposites of CdS quantum dots (QDs) and polyacrylamide (PAM) through the cross coupling chemistry. Initially, CdS QDs were synthesized in a well controlled manner using 3-mercaptopropionic acid as a capping agent. Then, carboxylic acid groups on CdS QDs were chemically incorporated into PAM matrices with the assistance of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride and N-hydroxysulfosuccinimide coupling agents. FT-IR analysis was used to investigate the chemical incorporation of CdS QDs in PAM matrices via the covalent protocol. The XPS elemental mapping studies further suggested the formation of CdS-PAM nanocomposites. FE-SEM and TEM images were engaged to study the morphologies, and distribution of CdS QDs in the PAM networks. The physical structure of the CdS-PAM nanocomposites was investigated by XRD analysis. Thermal stability of the nanocomposites was observed to be enhanced in compare to PAM as evidenced from TGA data. The UV-vis and photoluminescence studies of the CdS-PAM nanocomposites suggested their promising optical applications. PMID:24245319

  13. Permethylated-β-Cyclodextrin Capped CdTe Quantum Dot and its Sensitive Fluorescence Analysis of Malachite Green.

    PubMed

    Cao, Yujuan; Wei, Jiongling; Wu, Wei; Wang, Song; Hu, Xiaogang; Yu, Ying

    2015-09-01

    In the present work, the CdTe quantum dots were covalently conjugated with permethylated-β-cyclodextrin (OMe-β-CD) using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride as cross-linking reagent. The obtained functional quantum dots (OMe-β-CD/QDs) showed highly luminescent, water solubility and photostability as well as good inclusion ability to malachite green. A sensitive fluorescence method was developed for the analysis of malachite green in different samples. The good linearity was 2.0 × 10(-7)-1.0 × 10(-5) mol/L and the limit of detect was 1.7 × 10(-8) mol/L. The recoveries for three environmental water samples were 92.0-108.2 % with relative standard deviation (RSD) of 0.24-1.87 %, while the recovery for the fish sample was 94.3 % with RSD of 1.04 %. The results showed that the present method was sensitive and convenient to determine malachite green in complex samples. Graphical Abstract The analytical mechanism of OMe-β-CD/QDs and its linear response to MG. PMID:26250058

  14. Mucoadhesive 4-carboxybenzenesulfonamide-chitosan with antibacterial properties.

    PubMed

    Suvannasara, Phruetchika; Juntapram, Kotchakorn; Praphairaksit, Nalena; Siralertmukul, Krisana; Muangsin, Nongnuj

    2013-04-15

    The mucoadhesive property of chitosan, especially in an acidic (1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride as a coupling agent were investigated. The 0.2:1 (w/w) ratio 4-CBS:chitosan revealed a 20-fold stronger mucoadhesion to mucin type II than the native chitosan in the simulated gastric fluid (SGF; pH 1.2), and a swelling ratio after 1 h in water, SGF and simulated intestinal fluid (pH 7.4) of about 2.9-, 3.0- and 3.4-fold higher than that of chitosan, respectively. In tissue culture, the 4-CBS-chitosan, like chitosan, were found to be non-cytotoxic to the Vero, KB, MCF-7 and NCI-H187 cell lines but showed potential antibacterial activity against Escherichia coli and Staphlyococcus aureus as model gram-negative and gram-positive bacteria, respectively. PMID:23544535

  15. Endocrine-Disrupting Chemicals (EDCs): In Vitro Mechanism of Estrogenic Activation and Differential Effects on ER Target Genes

    PubMed Central

    Li, Yin; Luh, Colin J.; Burns, Katherine A.; Arao, Yukitomo; Jiang, Zhongliang; Teng, Christina T.; Tice, Raymond R.

    2013-01-01

    Background: Endocrine-disrupting chemicals (EDCs) influence the activity of estrogen receptors (ERs) and alter the function of the endocrine system. However, the diversity of EDC effects and mechanisms of action are poorly understood. Objectives: We examined the agonistic activity of EDCs through ERα and ERβ. We also investigated the effects of EDCs on ER-mediated target genes. Methods: HepG2 and HeLa cells were used to determine the agonistic activity of EDCs on ERα and ERβ via the luciferase reporter assay. Ishikawa cells stably expressing ERα were used to determine changes in endogenous ER target gene expression by EDCs. Results: Twelve EDCs were categorized into three groups on the basis of product class and similarity of chemical structure. As shown by luciferase reporter analysis, the EDCs act as ER agonists in a cell type– and promoter-specific manner. Bisphenol A, bisphenol AF, and 2-2-bis(p-hydroxyphenyl)-1,1,1-trichloroethane (group 1) strongly activated ERα estrogen responsive element (ERE)-mediated responses. Daidzein, genistein, kaempferol, and coumestrol (group 2) activated both ERα and ERβ ERE-mediated activities. Endosulfan and kepone (group 3) weakly activated ERα. Only a few EDCs significantly activated the “tethered” mechanism via ERα or ERβ. Results of real-time polymerase chain reaction indicated that bisphenol A and bisphenol AF consistently activated endogenous ER target genes, but the activities of other EDCs on changes of ER target gene expression were compound specific. Conclusion: Although EDCs with similar chemical structures (in the same group) tended to have comparable ERα and ERβ ERE-mediated activities, similar chemical structure did not correlate with previously reported ligand binding affinities of the EDCs. Using ERα-stable cells, we observed that EDCs differentially induced activity of endogenous ER target genes. PMID:23384675

  16. Identification of Edc3p as an enhancer of mRNA decapping in Saccharomyces cerevisiae.

    PubMed Central

    Kshirsagar, Meenakshi; Parker, Roy

    2004-01-01

    The major pathway of mRNA decay in yeast initiates with deadenylation, followed by mRNA decapping and 5'-3' exonuclease digestion. An in silico approach was used to identify new proteins involved in the mRNA decay pathway. One such protein, Edc3p, was identified as a conserved protein of unknown function having extensive two-hybrid interactions with several proteins involved in mRNA decapping and 5'-3' degradation including Dcp1p, Dcp2p, Dhh1p, Lsm1p, and the 5'-3' exonuclease, Xrn1p. We show that Edc3p can stimulate mRNA decapping of both unstable and stable mRNAs in yeast when the decapping enzyme is compromised by temperature-sensitive alleles of either the DCP1 or the DCP2 genes. In these cases, deletion of EDC3 caused a synergistic mRNA-decapping defect at the permissive temperatures. The edc3Delta had no effect when combined with the lsm1Delta, dhh1Delta, or pat1Delta mutations, which appear to affect an early step in the decapping pathway. This suggests that Edc3p specifically affects the function of the decapping enzyme per se. Consistent with a functional role in decapping, GFP-tagged Edc3p localizes to cytoplasmic foci involved in mRNA decapping referred to as P-bodies. These results identify Edc3p as a new protein involved in the decapping reaction. PMID:15020463

  17. Comparison of Electronic Data Capture (EDC) with the Standard Data Capture Method for Clinical Trial Data

    PubMed Central

    Walther, Brigitte; Hossin, Safayet; Townend, John; Abernethy, Neil; Parker, David; Jeffries, David

    2011-01-01

    Background Traditionally, clinical research studies rely on collecting data with case report forms, which are subsequently entered into a database to create electronic records. Although well established, this method is time-consuming and error-prone. This study compares four electronic data capture (EDC) methods with the conventional approach with respect to duration of data capture and accuracy. It was performed in a West African setting, where clinical trials involve data collection from urban, rural and often remote locations. Methodology/Principal Findings Three types of commonly available EDC tools were assessed in face-to-face interviews; netbook, PDA, and tablet PC. EDC performance during telephone interviews via mobile phone was evaluated as a fourth method. The Graeco Latin square study design allowed comparison of all four methods to standard paper-based recording followed by data double entry while controlling simultaneously for possible confounding factors such as interview order, interviewer and interviewee. Over a study period of three weeks the error rates decreased considerably for all EDC methods. In the last week of the study the data accuracy for the netbook (5.1%, CI95%: 3.5–7.2%) and the tablet PC (5.2%, CI95%: 3.7–7.4%) was not significantly different from the accuracy of the conventional paper-based method (3.6%, CI95%: 2.2–5.5%), but error rates for the PDA (7.9%, CI95%: 6.0–10.5%) and telephone (6.3%, CI95% 4.6–8.6%) remained significantly higher. While EDC-interviews take slightly longer, data become readily available after download, making EDC more time effective. Free text and date fields were associated with higher error rates than numerical, single select and skip fields. Conclusions EDC solutions have the potential to produce similar data accuracy compared to paper-based methods. Given the considerable reduction in the time from data collection to database lock, EDC holds the promise to reduce research-associated costs

  18. Uptake and Accumulation of Four PPCP/EDCs in Two Leafy Vegetables

    PubMed Central

    Dodgen, LK; Li, J; Parker, D; Gan, JJ

    2013-01-01

    Many pharmaceutical and personal care products (PPCPs) and endocrine-disrupting chemicals (EDCs) are present in reclaimed water, leading to concerns of human health risks from the consumption of food crops irrigated with reclaimed water. This study evaluated the potential for plant uptake and accumulation of four commonly occurring PPCP/EDCs, i.e., bisphenol A (BPA), diclofenac sodium (DCL), naproxen (NPX), and 4-nonylphenol (NP), by lettuce (Lactuca sativa) and collards (Brassica oleracea) in hydroponic culture, using 14C-labeled compounds. In both plant species, plant accumulation followed the order of BPA > NP > DCL > NPX and accumulation in roots was much greater than in leaves and stems. Concentrations of 14C-PPCP/EDCs in plant tissues ranged from 0.22±0.03 to 927± 213 ng/g, but nearly all 14C-residue was non-extractable. PPCP/EDCs, particularly BPA and NP, were also extensively transformed in the nutrient solution. Dietary uptake of these PPCP/EDCs by humans was predicted to be negligible. PMID:23911624

  19. Uptake and accumulation of four PPCP/EDCs in two leafy vegetables.

    PubMed

    Dodgen, L K; Li, J; Parker, D; Gan, J J

    2013-11-01

    Many pharmaceutical and personal care products (PPCPs) and endocrine-disrupting chemicals (EDCs) are present in reclaimed water, leading to concerns of human health risks from the consumption of food crops irrigated with reclaimed water. This study evaluated the potential for plant uptake and accumulation of four commonly occurring PPCP/EDCs, i.e., bisphenol A (BPA), diclofenac sodium (DCL), naproxen (NPX), and 4-nonylphenol (NP), by lettuce (Lactuca sativa) and collards (Brassica oleracea) in hydroponic culture, using (14)C-labeled compounds. In both plant species, plant accumulation followed the order of BPA > NP > DCL > NPX and accumulation in roots was much greater than in leaves and stems. Concentrations of (14)C-PPCP/EDCs in plant tissues ranged from 0.22 ± 0.03 to 927 ± 213 ng/g, but nearly all (14)C-residue was non-extractable. PPCP/EDCs, particularly BPA and NP, were also extensively transformed in the nutrient solution. Dietary uptake of these PPCP/EDCs by humans was predicted to be negligible. PMID:23911624

  20. Polish Society of Endocrinology Position statement on endocrine disrupting chemicals (EDCs).

    PubMed

    Rutkowska, Aleksandra; Rachoń, Dominik; Milewicz, Andrzej; Ruchała, Marek; Bolanowski, Marek; Jędrzejuk, Diana; Bednarczuk, Tomasz; Górska, Maria; Hubalewska-Dydejczyk, Alicja; Kos-Kudła, Beata; Lewiński, Andrzej; Zgliczyński, Wojciech

    2015-01-01

    With the reference to the position statements of the Endocrine Society, the Paediatric Endocrine Society, and the European Society of Paediatric Endocrinology, the Polish Society of Endocrinology points out the adverse health effects caused by endocrine disrupting chemicals (EDCs) commonly used in daily life as components of plastics, food containers, pharmaceuticals, and cosmetics. The statement is based on the alarming data about the increase of the prevalence of many endocrine disorders such as: cryptorchidism, precocious puberty in girls and boys, and hormone-dependent cancers (endometrium, breast, prostate). In our opinion, it is of human benefit to conduct epidemiological studies that will enable the estimation of the risk factors of exposure to EDCs and the probability of endocrine disorders. Increasing consumerism and the industrial boom has led to severe pollution of the environment with a corresponding negative impact on human health; thus, there is great necessity for the biomonitoring of EDCs in Poland. PMID:26136137

  1. Transformation and removal pathways of four common PPCP/EDCs in soil.

    PubMed

    Dodgen, L K; Li, J; Wu, X; Lu, Z; Gan, J J

    2014-10-01

    Pharmaceutical and personal care products (PPCPs) and endocrine disrupting chemicals (EDCs) enter the soil environment via irrigation with treated wastewater, groundwater recharge, and land application of biosolids. The transformation and fate of PPCP/EDCs in soil affects their potential for plant uptake and groundwater pollution. This study examined four PPCP/EDCs (bisphenol A, diclofenac, naproxen, and 4-nonylphenol) in soil by using (14)C-labeling and analyzing mineralization, extractable residue, bound residue, and formation of transformation products. At the end of 112 d of incubation, the majority of (14)C-naproxen and (14)C-diclofenac was mineralized to (14)CO2, while a majority of (14)C-bisphenol A and (14)Cnonylphenol was converted to bound residue. After 112 d, the estimated half-lives of the parent compounds were only 1.4-5.4 d. However a variety of transformation products were found and several for bisphenol A and diclofenac were identified, suggesting the need to consider degradation intermediates in soils impacted by PPCP/EDCs. PMID:24997388

  2. Transformation and Removal Pathways of Four Common PPCP/EDCs in Soil

    PubMed Central

    Dodgen, LK; Li, J; Wu, X; Lu, Z; Gan, JJ

    2014-01-01

    Pharmaceutical and personal care products (PPCPs) and endocrine disrupting chemicals (EDCs) enter the soil environment via irrigation with treated wastewater, groundwater recharge, and land application of biosolids. The transformation and fate of PPCP/EDCs in soil affects their potential for plant uptake and groundwater pollution. This study examined four PPCP/EDCs (bisphenol A, diclofenac, naproxen, and 4-nonylphenol) in soil by using 14C-labeling and analyzing mineralization, extractable residue, bound residue, and formation of transformation products. At the end of 112 d of incubation, the majority of 14C-naproxen and 14C-diclofenac was mineralized to 14CO2, while a majority of 14C-bisphenol A and 14C-nonylphenol was converted to bound residue. After 112 d, the estimated half-lives of the parent compounds were only 1.4 – 5.4 d. However a variety of transformation products were found and several for bisphenol A and diclofenac were identified, suggesting the need to consider degradation intermediates in soils impacted by PPCP/EDCs. PMID:24997388

  3. BIODEGRADABILITY OF SELECTED EDCS UNDER REDOX CONDITIONS TYPICAL OF WASTEWATER TREATMENT AND SEDIMENTS

    EPA Science Inventory

    A number of emerging chemicals being detected in the environment are now gaining attention for having possible endocrine disrupting capabilities. These endocrine disrupting chemicals (EDCs) have been shown to have adverse affects on the endocrine system of fish and wildlife. But ...

  4. NMR-based Metabolomics for Studying Toxicity, Compensation, and Recovery in Small Fish Exposed to EDCs

    EPA Science Inventory

    Determining the impact(s) on fish and other aquatic organisms of exposure to endocrine disrupting compounds (EDCs) is critical for determining the risks that these chemicals pose. However, to accurately evaluate these risks, beyond simply measuring a “before and after exposure” ...

  5. Executive Summary to EDC-2: The Endocrine Society's Second Scientific Statement on Endocrine-Disrupting Chemicals.

    PubMed

    Gore, A C; Chappell, V A; Fenton, S E; Flaws, J A; Nadal, A; Prins, G S; Toppari, J; Zoeller, R T

    2015-12-01

    This Executive Summary to the Endocrine Society's second Scientific Statement on environmental endocrine-disrupting chemicals (EDCs) provides a synthesis of the key points of the complete statement. The full Scientific Statement represents a comprehensive review of the literature on seven topics for which there is strong mechanistic, experimental, animal, and epidemiological evidence for endocrine disruption, namely: obesity and diabetes, female reproduction, male reproduction, hormone-sensitive cancers in females, prostate cancer, thyroid, and neurodevelopment and neuroendocrine systems. EDCs such as bisphenol A, phthalates, pesticides, persistent organic pollutants such as polychlorinated biphenyls, polybrominated diethyl ethers, and dioxins were emphasized because these chemicals had the greatest depth and breadth of available information. The Statement also included thorough coverage of studies of developmental exposures to EDCs, especially in the fetus and infant, because these are critical life stages during which perturbations of hormones can increase the probability of a disease or dysfunction later in life. A conclusion of the Statement is that publications over the past 5 years have led to a much fuller understanding of the endocrine principles by which EDCs act, including nonmonotonic dose-responses, low-dose effects, and developmental vulnerability. These findings will prove useful to researchers, physicians, and other healthcare providers in translating the science of endocrine disruption to improved public health. PMID:26414233

  6. INFLUENCE OF ENDOCRINE DISRUPTING COMPOUNDS (EDCS) ON MAMMARY GLAND DEVELOPMENT AND TUMOR SUSCEPTIBILITY

    EPA Science Inventory

    Influence of Endocrine Disrupting Compounds (EDCs) on Mammary Gland Development and Tumor Susceptibility.

    Suzanne E. Fenton1, and Jennifer Rayner1,2

    1 Reproductive Toxicology Division, NHEERL/ORD, U.S. EPA, Research Triangle Park, NC, and 2 Department of Environmen...

  7. New enolate-carbodiimide rearrangement in the concise synthesis of 6-amino-2,3-dihydro-4-pyridinones from homoallylamines.

    PubMed

    Kuznetsov, N Yu; Tikhov, R M; Godovikov, I A; Khrustalev, V N; Bubnov, Yu N

    2016-05-01

    Three-step synthesis of 6-amino-2,3-dihydro-4-pyridinones from homoallylamines involving NBS-mediated cyclization of N-(3-butenyl)ureas to 6-(bromomethyl)-2-iminourethanes, dehydrohalogenation and a novel rearrangement as a key step has been developed. The scope and limitations of the method, as well as the mechanism of the rearrangement, supported by kinetic studies and the isolation of N-(1-adamantyl)carbodiimide, are discussed. The final products, imino-analogues of well known piperidine-2,4-diones, are promising building blocks in the synthesis of bio-/pharmacological compounds. PMID:27080757

  8. EDC-2: The Endocrine Society's Second Scientific Statement on Endocrine-Disrupting Chemicals.

    PubMed

    Gore, A C; Chappell, V A; Fenton, S E; Flaws, J A; Nadal, A; Prins, G S; Toppari, J; Zoeller, R T

    2015-12-01

    The Endocrine Society's first Scientific Statement in 2009 provided a wake-up call to the scientific community about how environmental endocrine-disrupting chemicals (EDCs) affect health and disease. Five years later, a substantially larger body of literature has solidified our understanding of plausible mechanisms underlying EDC actions and how exposures in animals and humans-especially during development-may lay the foundations for disease later in life. At this point in history, we have much stronger knowledge about how EDCs alter gene-environment interactions via physiological, cellular, molecular, and epigenetic changes, thereby producing effects in exposed individuals as well as their descendants. Causal links between exposure and manifestation of disease are substantiated by experimental animal models and are consistent with correlative epidemiological data in humans. There are several caveats because differences in how experimental animal work is conducted can lead to difficulties in drawing broad conclusions, and we must continue to be cautious about inferring causality in humans. In this second Scientific Statement, we reviewed the literature on a subset of topics for which the translational evidence is strongest: 1) obesity and diabetes; 2) female reproduction; 3) male reproduction; 4) hormone-sensitive cancers in females; 5) prostate; 6) thyroid; and 7) neurodevelopment and neuroendocrine systems. Our inclusion criteria for studies were those conducted predominantly in the past 5 years deemed to be of high quality based on appropriate negative and positive control groups or populations, adequate sample size and experimental design, and mammalian animal studies with exposure levels in a range that was relevant to humans. We also focused on studies using the developmental origins of health and disease model. No report was excluded based on a positive or negative effect of the EDC exposure. The bulk of the results across the board strengthen the evidence

  9. Solving the puzzling competition of the thermal C2–C6 vs Myers–Saito cyclization of enyne-carbodiimides

    PubMed Central

    Rana, Anup; Cinar, Mehmet Emin; Samanta, Debabrata

    2016-01-01

    Summary The mechanism of the thermal cyclization of enyne-carbodiimides 7a–c has been studied computationally by applying the DFT method. The results indicate that enyne-carbodiimides preferentially follow the C2–C6 (Schmittel) cyclization pathway in a concerted fashion although the Myers–Saito diradical formation is kinetically preferred. The experimentally verified preference of the C2–C6 over the Myers–Saito pathway is guided by the inability of the Myers–Saito diradical to kinetically compete in the rate-determining trapping reactions, either inter- or intramolecular, with the concerted C2–C6 cyclization. As demonstrated with enyne-carbodiimide 11, the Myers–Saito channel can be made the preferred pathway if the trapping reaction by hydrogen transfer is no more rate determining. PMID:26877807

  10. Preparation of peptide-functionalized gold nanoparticles using one pot EDC/sulfo-NHS coupling.

    PubMed

    Bartczak, Dorota; Kanaras, Antonios G

    2011-08-16

    Although carbodiimides and succinimides are broadly employed for the formation of amide bonds (i.e., in amino acid coupling), their use in the coupling of peptides to water-soluble carboxylic-terminated colloidal gold nanoparticles remains challenging. In this article, we present an optimization study for the successful coupling of the KPQPRPLS peptide to spherical and rodlike colloidal gold nanoparticles. We show that the concentration, reaction time, and chemical environment are all critical to achieving the formation of robust, peptide-coated colloidal nanoparticles. Agarose gel electrophoresis was used for the characterization of conjugates. PMID:21728291

  11. Rhenium-Catalyzed Synthesis of 1,3-Diiminoisoindolines via Insertion of Carbodiimides into a C-H Bond of Aromatic and Heteroaromatic Imidates.

    PubMed

    Wang, Zijia; Sueki, Shunsuke; Kanai, Motomu; Kuninobu, Yoichiro

    2016-05-20

    The rhenium-catalyzed synthesis of 1,3-diiminoisoindolines and their related compounds from aromatic or heteroaromatic imidates and carbodiimides are reported via C-H bond activation. This reaction is the first example of a transition-metal-catalyzed insertion of carbodiimides into an aromatic or heteroaromatic C-H bond and a novel method for synthesizing 1,3-diiminoisoindolines and their related compounds. Unsymmetrical 1,3-diiminoisoindolines were easily obtained using this method. The reaction proceeded in good to excellent yield using a variety of substrates. PMID:27153181

  12. CHANGES IN GENE AND PROTEIN EXPRESSION IN ZEBRAFISH (DANIO RERIO) FOLLOWING EXPOSURE TO ENVIRONMENTALLY-RELEVANT ENDOCRINE DISRUPTING COMPOUNDS (EDCS)

    EPA Science Inventory

    Endocrine-disrupting chemicals (EDCs) are increasingly being reported in waterways worldwide and have been shown to affect fish species by disrupting numerous aspects of development, behavior, reproduction, and survival. Furthermore, new data have suggested that the reduced repr...

  13. A New Methodology for the Detection of Low-Abundance Species in the Ism: Detection of Interstellar Carbodiimide (HNCNH)

    NASA Astrophysics Data System (ADS)

    McGuire, Brett A.; Loomis, Ryan A.; Charness, Cameron M.; Corby, Joanna F.; Blake, Geoffrey A.; Hollis, Jan M.; Lovas, Frank J.; Jewell, Philip R.; Remijan, Anthony J.

    2013-06-01

    We present the first interstellar detection of carbodiimide (HNCNH) in observations towards Sgr B2(N) using data from the publicly available Green Bank Telescope PRebiotic Interstellar MOlecular Survey project. Recent laboratory work predicts an abundance of HNCNH of ˜10% of the abundance of its tautomer, cyanamide (NH_2CN), or ˜ 2× 10^{13} cm^{-2} in Sgr B2(N). Given this abundance at LTE conditions, the strongest rotational transitions of HNCNH have intensities at or below the noise level of current observations of this source. A thermal population of HNCNH is therefore likely undetectable. Instead, HNCNH is identified via maser emission features at centimeter wavelengths. This detection presents a new methodology for the detection of low-abundance species and further demonstrates the power of cm-wave observations to make definitive identifications based on a small number of observed features.

  14. Controlling coupling reaction of EDC and NHS for preparation of collagen gels using ethanol/water co-solvents.

    PubMed

    Nam, Kwangwoo; Kimura, Tsuyoshi; Kishida, Akio

    2008-01-01

    To control the crosslinking rate of the collagen gel, ethanol/water co-solvent was adopted for the reaction solvent for the collagen microfibril crosslinking. Collagen gel was prepared by using EDC and NHS as coupling agents. Ethanol did not denaturate the helical structure of the collagen and prevented the hydrolysis of EDC, but showed the protonation of carboxylate anions. In order to control the intra- and interhelical crosslink of the collagen triple helix, variations of the mole ratio of carboxyl group/EDC/NHS, and of the ethanol mole concentration were investigated. Increase in the EDC ratio against the carboxyl group increased the crosslinking rate. Furthermore, an increase in the ethanol mole concentration resulted in an increase of the crosslinking rate until ethanol mole concentration was 0.12, but showed gradual decrease as the ethanol mole concentration was further increased. This is because the adsorption of solvent by the collagen gel, protonation of carboxylate anion, and hydrolysis of EDC is at its most optimum condition for the coupling reaction when the ethanol mole concentration is 0.12. The re-crosslinking of the collagen gel showed an increase in the crosslinking rate, but did not show further increase when the coupling reaction was executed for the third time. This implied that the highest possible crosslinking rate for the intra- and interhelical is approximately 60% when EDC/NHS is used. PMID:18023082

  15. Deletion analysis of LSm, FDF, and YjeF domains of Candida albicans Edc3 in hyphal growth and oxidative-stress response.

    PubMed

    Kim, Eung-Chul; Kim, Jinmi

    2015-02-01

    Candida albicans is an opportunistic fungal pathogen whose responses to environmental changes are associated with the virulence attributes. Edc3 is known to be an enhancer of the mRNA decapping reactions and a scaffold protein of cytoplasmic processing bodies (P-bodies). Recent studies of C. albicans Edc3 suggested its critical roles in filamentous growth and stress-induced apoptotic cell death. The edc3/edc3 deletion mutant strain showed increased cell survival and less ROS accumulation upon treatment with hydrogen peroxide. To investigate the diverse involvement of Edc3 in the cellular processes, deletion mutations of LSm, FDF, or YjeF domain of Edc3 were constructed. The edc3-LSmΔ or edc3-YjeFΔ mutation showed the filamentation defect, resistance to oxidative stress, and decreased ROS accumulation. In contrast, the edc3-FDFΔ mutation exhibited a wild-type level of filamentous growth and a mild defect in ROS accumulation. These results suggest that Lsm and YjeF domains of Edc3 are critical in hyphal growth and oxidative stress response. PMID:25626365

  16. Occurrence and spatial distribution of EDCs and related compounds in waters and sediments of Iberian rivers.

    PubMed

    Gorga, Marina; Insa, Sara; Petrovic, Mira; Barceló, Damià

    2015-01-15

    The environmental presence of chemicals capable of affecting the endocrine system has become a matter of scientific and public concern after certain endocrine disruptor compounds (EDCs) have been detected in the aquatic environment. In this work, 31 different EDCs and related compounds (suspect EDCs) belonging to different contaminant classes were studied: 10 estrogens, natural and synthetic, 8 alkylphenolic compounds, bisphenol A, triclosan and triclorocaraban, 4 parabens, 2 benzotriazoles, 3 organophosphorous flame retardants and the chemical marker caffeine, in river water and sediment of four Iberian rivers (Ebro, Llobregat, Júcar and Guadalquivir). An extensive sampling has been undertaken in two monitoring campaigns (2010 and 2011). A total of 77 samples of water and 75 sediments were collected. For this propose two different multiresidue analytical methods were applied, using the automated online EQuan/TurboFlow™ liquid chromatography coupled to mass spectrometry detection in tandem. In terms of concentrations the compounds found at the highest average concentrations were: nonylphenol monocarboxylate (NP1EC), tolyltriazole (TT), tris(chloroisopropyl)phosphate (TCPP) found at average concentrations above 100 ng/L, followed by 1H-benzotriazole and tris(butoxyethyl)phosphate (TBEP) found at average concentration higher than 50 ng/L. Natural and synthetic hormones were found at low levels not exceeding 16 ng/L and 7 ng/g for water and sediment, respectively, however they contributed to more than 80% of the total estrogenicity of the samples (expressed as the equivalents of estradiol EEQ, ng/L). Regarding the spatial distribution of these contaminants, the Llobregat river was found to be the most contaminated river basin, having sites near the mouth of the river the ones with the highest contaminant load. In the Ebro river basin several hot spots were identified and Júcar showed to be the least contaminated. Overall, the study confirmed the presence of

  17. One-dimensional plate impact experiments on the cyclotetramethylene tetranitramine (HMX) based explosive EDC32

    NASA Astrophysics Data System (ADS)

    Burns, Malcolm J.; Gustavsen, Richard L.; Bartram, Brian D.

    2012-09-01

    Eight one-dimensional plate impact experiments have been performed to study both the Shock to Detonation Transition and Hugoniot state in the cyclotetramethylene tetranitramine (HMX) based explosive EDC32. The experiments covered shock pressures ranging from 0.59 to 7.5 GPa with sustained shocks, double shocks, and short pulse shocks. Experiments were instrumented with embedded magnetic particle velocity gauges. Results include; (1) wave profiles of particle velocity vs. time vs. depth in the explosive, (2) time-distance coordinates for onset of detonation vs. initial shock pressure (aka the Pop-plot), (3) a reactants Hugoniot, and (4) measurement of the Hugoniot Elastic Limit of 0.22.GPa.

  18. Bridged bis(amidinate) lanthanide aryloxides: syntheses, structures, and catalytic activity for addition of amines to carbodiimides.

    PubMed

    Tu, Jing; Li, Wenbo; Xue, Mingqiang; Zhang, Yong; Shen, Qi

    2013-04-28

    Various lanthanide aryloxide complexes supported by bridged bis(amidinate) ligand L, LLnOAr(DME) (L = Me3SiNC(Ph)N(CH2)3NC(Ph)NSiMe3, DME = dimethoxyethane, Ln = Y, Ar = 2,6-(Me)2C6H3 (1), 2,6-((i)Pr)2C6H3 (2), 2,6-((t)Bu)2-4-(Me)C6H2 (3); Ar = 2,6-((t)Bu)2-4-(Me)C6H2, Ln = Nd (4), Sm (5), Yb (6)) were synthesized, and complexes 1, 2 and 4–6 were characterized by single crystal X-ray diffraction. All the complexes are efficient precatalysts for catalytic addition of amines to carbodiimides. The catalytic activity is influenced by lanthanide metals and the aryloxide groups (Nd (4) Sm (5) < Y (3) Yb (6) and -2,6-(Me)2C6H3 < -2,6-((i)Pr)2C6H3 < -2,6-((t)Bu)2-4-(Me)C6H2). The catalytic addition reaction with 3 showed a good scope of substrates. The mechanism investigation revealed the real active intermediate being the monoguanidinate complexes supported by an aryloxide and an amidine-functionalized amidinate group, L′Ln[O2,6-((t)Bu)2-4-(Me)C6H2][RNCNHRN(Ar′)] (L′ = Me3SiNHC(Ph)N(CH2)3NC(Ph)NSiMe3, R = (i)Pr, Ar′ = phenyl, Ln = Yb (8), Y (11); R = Cy, Ar′ = phenyl, Ln = Yb (10), Y (12); R = (i)Pr, Ar′ = 4-ClC6H4, Ln = Yb (9)), which were isolated from the reactions of 6 (or 3) with amine and carbodiimide in a molar ratio of 1:1:1 and structurally characterized. The Ln-active group in the present precatalyst is a Ln–amidinate species, not the Ln–OAr group. PMID:23459864

  19. The classic EDCs, phthalate esters and organochlorines, in relation to abnormal sperm quality: a systematic review with meta-analysis

    PubMed Central

    Wang, Chao; Yang, Lu; Wang, Shu; Zhang, Zhan; Yu, Yongquan; Wang, Meilin; Cromie, Meghan; Gao, Weimin; Wang, Shou-Lin

    2016-01-01

    The association between endocrine disrupting chemicals (EDCs) and human sperm quality is controversial due to the inconsistent literature findings, therefore, a systematic review with meta-analysis was performed. Through the literature search and selection based on inclusion criteria, a total of 9 studies (7 cross-sectional, 1 case-control, and 1 pilot study) were analyzed for classic EDCs (5 studies for phthalate esters and 4 studies for organochlorines). Funnel plots revealed a symmetrical distribution with no evidence of publication bias (Begg’s test: intercept = 0.40; p = 0.692). The summary odds ratios (OR) of human sperm quality associated with the classic EDCs was 1.67 (95% CI: 1.31–2.02). After stratification by specific chemical class, consistent increases in the risk of abnormal sperm quality were found in phthalate ester group (OR = 1.52; 95% CI: 1.09–1.95) and organochlorine group (OR = 1.98; 95% CI: 1.34–2.62). Additionally, identification of official data, and a comprehensive review of the mechanisms were performed, and better elucidated the increased risk of these classic EDCs on abnormal sperm quality. The present systematic review and meta-analysis helps to identify the impact of classic EDCs on human sperm quality. However, it still highlights the need for additional epidemiological studies in a larger variety of geographic locations. PMID:26804707

  20. The classic EDCs, phthalate esters and organochlorines, in relation to abnormal sperm quality: a systematic review with meta-analysis

    NASA Astrophysics Data System (ADS)

    Wang, Chao; Yang, Lu; Wang, Shu; Zhang, Zhan; Yu, Yongquan; Wang, Meilin; Cromie, Meghan; Gao, Weimin; Wang, Shou-Lin

    2016-01-01

    The association between endocrine disrupting chemicals (EDCs) and human sperm quality is controversial due to the inconsistent literature findings, therefore, a systematic review with meta-analysis was performed. Through the literature search and selection based on inclusion criteria, a total of 9 studies (7 cross-sectional, 1 case-control, and 1 pilot study) were analyzed for classic EDCs (5 studies for phthalate esters and 4 studies for organochlorines). Funnel plots revealed a symmetrical distribution with no evidence of publication bias (Begg’s test: intercept = 0.40 p = 0.692). The summary odds ratios (OR) of human sperm quality associated with the classic EDCs was 1.67 (95% CI: 1.31-2.02). After stratification by specific chemical class, consistent increases in the risk of abnormal sperm quality were found in phthalate ester group (OR = 1.52 95% CI: 1.09-1.95) and organochlorine group (OR = 1.98 95% CI: 1.34-2.62). Additionally, identification of official data, and a comprehensive review of the mechanisms were performed, and better elucidated the increased risk of these classic EDCs on abnormal sperm quality. The present systematic review and meta-analysis helps to identify the impact of classic EDCs on human sperm quality. However, it still highlights the need for additional epidemiological studies in a larger variety of geographic locations.

  1. The TFPI-2 Derived Peptide EDC34 Improves Outcome of Gram-Negative Sepsis

    PubMed Central

    Papareddy, Praveen; Kalle, Martina; Sørensen, Ole E.; Malmsten, Martin; Mörgelin, Matthias; Schmidtchen, Artur

    2013-01-01

    Sepsis is characterized by a dysregulated host-pathogen response, leading to high cytokine levels, excessive coagulation and failure to eradicate invasive bacteria. Novel therapeutic strategies that address crucial pathogenetic steps during infection are urgently needed. Here, we describe novel bioactive roles and therapeutic anti-infective potential of the peptide EDC34, derived from the C-terminus of tissue factor pathway inhibitor-2 (TFPI-2). This peptide exerted direct bactericidal effects and boosted activation of the classical complement pathway including formation of antimicrobial C3a, but inhibited bacteria-induced activation of the contact system. Correspondingly, in mouse models of severe Escherichia coli and Pseudomonas aeruginosa infection, treatment with EDC34 reduced bacterial levels and lung damage. In combination with the antibiotic ceftazidime, the peptide significantly prolonged survival and reduced mortality in mice. The peptide's boosting effect on bacterial clearance paired with its inhibiting effect on excessive coagulation makes it a promising therapeutic candidate for invasive Gram-negative infections. PMID:24339780

  2. Covalent immobilization of molecularly imprinted polymer nanoparticles on a gold surface using carbodiimide coupling for chemical sensing.

    PubMed

    Kamra, Tripta; Chaudhary, Shilpi; Xu, Changgang; Montelius, Lars; Schnadt, Joachim; Ye, Lei

    2016-01-01

    One challenging task in building (bio)chemical sensors is the efficient and stable immobilization of receptor on a suitable transducer. Herein, we report a method for covalent immobilization of molecularly imprinted core-shell nanoparticles for construction of robust chemical sensors. The imprinted nanoparticles with a core-shell structure have selective molecular binding sites in the core and multiple amino groups in the shell. The model Au transducer surface is first functionalized with a self-assembled monolayer of 11-mercaptoundecanoic acid. The 11-mercaptoundecanoic acid is activated by treatment with carbodiimide/N-hydroxysuccinimide and then reacted with the core-shell nanoparticles to form amide bonds. We have characterized the process by studying the treated surfaces after each preparation step using atomic force microscopy, scanning electron microscopy, fluorescence microscopy, contact angle measurements and X-ray photoelectron spectroscopy. The microscopy results show the successful immobilization of the imprinted nanoparticles on the surface. The photoelectron spectroscopy results further confirm the success of each functionalization step. Further, the amino groups on the MIP surface were activated by electrostatically adsorbing negatively charged Au colloids. The functionalized surface was shown to be active for surface enhanced Raman scattering detection of propranolol. The particle immobilization and surface enhanced Raman scattering approach described here has a general applicability for constructing chemical sensors in different formats. PMID:26397901

  3. Comparison of the properties of collagen-chitosan scaffolds after γ-ray irradiation and carbodiimide cross-linking.

    PubMed

    Chen, Zihao; Du, Tianming; Tang, Xiangyu; Liu, Changjun; Li, Ruixin; Xu, Cheng; Tian, Feng; Du, Zhenjie; Wu, Jimin

    2016-07-01

    The property of collagen-chitosan porous scaffold varies according to cross-linking density and scaffold composition. This study was designed to compare the properties of collagen-chitosan porous scaffolds cross-linked with γ-irradiation and carbodiimide (CAR) for the first time. Eleven sets of collagen-chitosan scaffolds containing different concentrations of chitosan at a 5% increasing gradient were fabricated. Fourier transform infrared spectroscopy was performed to confirm the success of cross-linking in the scaffolds. The scaffold morphology was evaluated under scanning electron microscope (SEM). SEM revealed that chitosan was an indispensable material for the fabrication of γ-ray irradiation scaffold. The microstructure of γ-ray irradiation scaffold was less stable than those of alternative scaffolds. Based upon swelling ratio, porosity factor, and collagenase degradation, γ-ray irradiation scaffold was less stable than CAR and 25% proportion of chitosan scaffolds. Mechanical property determines the orientation in γ-irradiation and CAR scaffold. In vitro degradation test indicated that γ-irradiation and CAR cross-linking can elevate the scaffold biocompatibility. Compared with γ-ray irradiation, CAR cross-linked scaffold containing 25% chitosan can more significantly enhance the bio-stability and biocompatibility of collagen-chitosan scaffolds. CAR cross-linked scaffold may be the best choice for future tissue engineering. PMID:27122297

  4. Bioavailability of endocrine disrupting chemicals (EDCs): Liposome-water partitioning and lipid membrane permeation

    NASA Astrophysics Data System (ADS)

    Kwon, Jung-Hwan

    The bioavailability of endocrine disrupting chemicals (EDCs) is a function of a number of parameters including the ability of the chemical to partition into organic tissue and reach receptor sites within an organism. In this dissertation, equilibrium partition coefficients between water and lipid membrane vesicles and artificial lipid membrane permeability were investigated for evaluating bioavailability of aqueous pollutants. Structurally diverse endocrine disrupting chemicals were chosen as model compounds for partitioning experiments and simple hydrophobic organic chemicals were used for the evaluation of a parallel artificial membrane device developed to mimic bioconcentration rates in fish. Hydrophobic interactions represented by octanol/water partition coefficients (KOWs) were not appropriate for estimating lipid membrane/water partition coefficients (Klipws) for the selected EDCs having a relatively large molar liquid volume (MLV) and containing polar functional groups. Correlations that include MLV and polar surface area (PSA) reduce the predicted value of log K lipw, suggesting that lipid membranes are less favorable than 1-octanol for a hydrophobic solute because of the changes in membrane fluidity and the amount of cholesterol in the lipid bilayers. These results suggested that KOW alone has limited potential for estimating K lipw, and MLV or PSA may be used as additional descriptors for developing quantitative structure-activity relationships (QSARs). The poor correlations between KOW and Klipw observed in this research may be due to the highly organized structure of lipid bilayers. Measured thermodynamic constants demonstrated that the entropy contribution becomes more dominant for more organized liposomes having saturated lipid tails. This implies that entropy-driven partitioning process makes Klipw different from KOW especially for more saturated lipid bilayer membranes. In the parallel artificial membrane system developed, a membrane filter

  5. Improved fibronectin-immobilized fibrinogen microthreads for the attachment and proliferation of fibroblasts

    PubMed Central

    Rajangam, Thanavel; An, Seong Soo A

    2013-01-01

    The aim of this study was to fabricate fibrinogen (Fbg) microfibers with different structural characteristics for the development of 3-D tissue-engineering scaffolds. Fabricated Fbg microfibers were investigated for their biomolecule encapsulation, cell adhesion, and proliferations. Microfibers with three different concentrations of Fbg (5, 10, and 15 wt%) were prepared by a gel solvent-extraction method using a silicone rubber tube. Fbg microfibers were covalently modified with fibronectin (FN) by using water-soluble 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide as the cross-linking agent. Fbg microfibers were characterized by their FN cross-linking properties, structural morphology, and in vitro degradation. Furthermore, FN/Fbg microfibers were evaluated for cell attachment and proliferation. The bio-compatibility and cell proliferation of the microfibers were assessed by measuring adenosine triphosphate activity in C2C12 fibroblast cells. Cell attachment and proliferation on microfibers were further examined using fluorescence and scanning electron microscopic images. FN loading on the microfibers was confirmed by fluorescence and infrared spectroscopy. Surface morphology was characterized by scanning electron microscopy, and showed highly aligned nanostructures for fibers made with 15 wt% Fbg, a more porous structure for fibers made with 10 wt% Fbg, and a less porous structure for those made with 5 wt% Fbg. Controlled biodegradation of the fiber was observed for 8 weeks by using an in vitro proteolytic degradation assay. Fbg microfibers with highly aligned nanostructures (15 wt%) showed enhanced biomolecule encapsulation, as well as higher cell adhesion and proliferation than another two types of FN/Fbg fibers (5 and 10 wt%) and unmodified Fbg fibers. The promising results obtained from the present study reveal that optimal structure of Fbg microfibers could be used as a potential substratum for growth factors or drug release, especially in wound healing and

  6. Enzymatic specificities and modes of action of the two catalytic domains of the XynC xylanase from Fibrobacter succinogenes S85.

    PubMed Central

    Zhu, H; Paradis, F W; Krell, P J; Phillips, J P; Forsberg, C W

    1994-01-01

    The xylanase XynC of Fibrobacter succinogenes S85 was recently shown to contain three distinct domains, A, B, and C (F. W. Paradis, H. Zhu, P. J. Krell, J. P. Phillips, and C. W. Forsberg, J. Bacteriol. 175:7666-7672, 1993). Domains A and B each bear an active site capable of hydrolyzing xylan, while domain C has no enzymatic activity. Two truncated proteins, each containing a single catalytic domain, named XynC-A and XynC-B were purified to homogeneity. The catalytic domains A and B had similar pH and temperature parameters of 6.0 and 50 degrees C for maximum hydrolytic activity and extensively degraded birch wood xylan to xylose and xylobiose. The Km and Vmax values, respectively, were 2.0 mg ml-1 and 6.1 U mg-1 for the intact enzyme, 1.83 mg ml-1 and 689 U mg-1 for domain A, and 2.38 mg ml-1 and 91.8 U mg-1 for domain B. Although domain A had a higher specific activity than domain B, domain B exhibited a broader substrate specificity and hydrolyzed rye arabinoxylan to a greater extent than domain A. Furthermore, domain B, but not domain A, was able to release xylose at the initial stage of the hydrolysis. Both catalytic domains cleaved xylotriose, xylotetraose, and xylopentaose but had no activity on xylobiose. Bond cleavage frequencies obtained from hydrolysis of xylo-alditol substrates suggest that while both domains have a strong preference for internal linkages of the xylan backbone, domain B has fewer subsites for substrate binding than domain A and cleaves arabinoxylan more efficiently. Chemical modification with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide methiodide and N-bromosuccinimide inactivated both XynC-A and XynC-B in the absence of xylan, indicating that carboxyl groups and tryptophan residues in the catalytic site of each domain have essential roles. Images PMID:8021170

  7. A reinforced composite structure composed of polydiacetylene assemblies deposited on polystyrene microspheres and its application to H5N1 virus detection

    PubMed Central

    Dong, Wenjie; Luo, Jing; He, Hongxuan; Jiang, Long

    2013-01-01

    In this study, we immobilized polydiacetylene vesicles (PDAVs) onto the surface of polystyrene (PS) microspheres (1 μm in diameter) by using both electrical charge and conjugated forces to form a reinforced composite structure. These reinforced complexes could be easily washed, separated by centrifugation, and resuspended by gentle agitation. After passing through a narrow 200 μm-diameter channel, the composite structures maintained their original shape, demonstrating their resilience and potential for use in microfluidic technologies. The number of PDAVs in the composite structure could be mediated by changing the extent of layer deposition, which affected the sensitivity of detection. It showed that PDAVs did not change their blue color after addition of detecting probes such as anti-H5N1, which was of great importance in the fabrication and modification of stable color-changeable biosensors based on PDAVs. By conjugating anti-H5N1 antibodies to the PS@PDAV via N -hydroxysuccinimide and 1-ethyl3-(3-dimethylaminopropyl) carbodiimide chemistry, a stable blue complex, anti-H5N1 microsphere (PS@PDAV-anti-H5N1) was formed. A target antigen of H5N1 (HAQ [H5N1 strain A/ environment/Qinghai/1/2008{H5N1} in clade 0]) was detected by PS@PDAV-anti-H5N1. At an optimal PDAV deposition level of three layers, the limit of detection was determined to be approximately 3 0 ng/mL of HAQ by using optical spectrum measurement and visual inspection, meeting the needs of fast and simple color-changeable detection. However, a much lower limitation of detection (1 ng/mL) was able to be obtained using laser-scanning confocal microscopy, which could be compared with the results obtained with other sophisticated equipment. PMID:23403826

  8. Enhanced compatibility of chemically modified titanium surface with periodontal ligament cells

    NASA Astrophysics Data System (ADS)

    Kado, T.; Hidaka, T.; Aita, H.; Endo, K.; Furuichi, Y.

    2012-12-01

    A simple chemical modification method was developed to immobilize cell-adhesive molecules on a titanium surface to improve its compatibility with human periodontal ligament cells (HPDLCs).The polished titanium disk was immersed in 1% (v/v) p-vinylbenzoic acid solution for 2 h to introduce carboxyl groups onto the surface. After rinsing with distilled deionized water, the titanium disk was dipped into 1.47% 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide solution containing 0.1 mg/ml Gly-Arg-Gly-Asp-Ser (GRGDS), human plasma fibronectin (pFN), or type I collagen from calf skin (Col) to covalently immobilize the cell-adhesive molecules on the titanium surface via formation of peptide bonds. X-ray photoelectron spectroscopy analyses revealed that cell-adhesive molecules were successfully immobilized on the titanium surfaces. The Col-immobilized titanium surface revealed higher values regarding nano rough characteristics than the as-polished titanium surface under scanning probe microscopy. The number of HPDLCs attached to both the pFN- and Col-immobilized titanium surfaces was twice that attached to the as-polished titanium surfaces. The cells were larger with the cellular processes that stretched to a greater extent on the pFN- and Col-immobilized titanium surfaces than on the as-polished titanium surface (p < 0.05). HPDLCs on the Col-immobilized titanium surfaces showed more extensive expression of vinculin at the tips of cell projections and more contiguously along the cell outline than on the as-polished, GRGDS-immobilized and pFN-immobilized titanium surfaces. It was concluded that cell-adhesive molecules successfully immobilized on the titanium surface and improved the compatibility of the surface with HPDLCs. The Col-immobilized titanium surface could be used for forming ligament-like tissues around titanium dental implants.

  9. 2,2'Dithiodinicotinyl ligands: Key to more reactive thiomers.

    PubMed

    Menzel, Claudia; Silbernagl, Jennifer; Laffleur, Flavia; Leichner, Christina; Jelkmann, Max; Huck, Christian W; Hussain, Shah; Bernkop-Schnürch, Andreas

    2016-04-30

    The aim of this study was to establish a novel type of preactivated thiomers exhibiting a comparatively higher reactivity with mucus and consequently improved mucoadhesive properties. In order to achieve this goal, the dimeric form of 2-mercaptonicotinic acid (MNA-MNA) was directly attached to the polymeric backbone of chitosan (CHI) via amide bond formation mediated by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDAC) used as a coupling reagent. The remaining free amino groups were in the following reacted with succinic anhydride (Succ) in order to obtain a uniformly anionically charged polymer (CHI-Succ-MNA-MNA). Within this study, different coupling rates of up to 170 μmol MNA-MNA per gram polymer were achieved. The attachment of the dimeric ligand resulted in a preactivated thiomer with a comparatively more reactive disulfide substructure due to the additional nitrogen atom in conjugation over the aromatic moieties. Furthermore, the obtained polymer is entirely preactivated and thus prevented against undesired oxidation reactions. Kinetic studies of disulfide exchange reactions showed a 3.8-fold higher reactivity of CHI-Succ-MNA-MNA in comparison to a state-of-the-art preactivated thiomer. Within rheological measurements, CHI-Succ-MNA-MNA with a coupling rate of 170 μmol (CHI-Succ-MNA-MNA 170) led to a 5.7-fold higher mucus viscosity than the non-thiolated control polymer (CHI-Succ) indicating a rheological synergism due to mucoadhesive properties. These results were confirmed by a second mucoadhesion study, which showed a significantly prolonged retention time of CHI-Succ-MNA-MNA on the small intestinal mucosa compared to CHI-Succ (P<0.02). Accordingly, the double preactivation seems to be a promising strategy in order to obtain entirely preactivated polymers with enhanced mucoadhesive properties. PMID:26972378

  10. Endosomal proteolysis of glucagon at neutral pH generates the bioactive degradation product miniglucagon-(19-29).

    PubMed

    Authier, Francois; Cameron, Pamela H; Merlen, Clemence; Kouach, Mostafa; Briand, Gilbert

    2003-12-01

    We have investigated the proteolytic mechanisms of glucagon degradation within hepatic endosomes at neutral pH before lumen acidification. Hepatic endosomes incubated at neutral pH rapidly degraded native glucagon into 13 intermediate products, one of which corresponded to the bioactive fragment glucagon-(19-29) (miniglucagon). The serine protease inhibitor phenylmethylsulfonyl fluoride as well as the nonspecific protease inhibitor bacitracin inhibited the endosomal degradation of glucagon at pH 7. In purified endosomal fractions, miniglucagon endopeptidase was undetectable as evaluated by immunoblotting, and immunoprecipitation with antibodies to insulin-degrading enzyme, cathepsins B and D, or furin failed to remove the endosomal neutral glucagonase activity. Incubation of endosomal fractions and [125I]iodoglucagon with the zero-length bifunctional cross-linker 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide resulted in specific labeling of a 170-kDa polypeptide. The labeling was completely inhibited by unlabeled glucagon (IC50 value, 5 x 10-7 m) and bacitracin (IC50 value, 1 microg/ml), suggesting that it may correspond to a bacitracin-sensitive glucagon-degrading enzyme. Treatment of the 125I-labeled 170-kDa cross-linked polypeptide with N-glycanase demonstrated that the cross-linked complex contained approximately 30 kDa of N-linked oligosaccharides. Specific cross-linking of the 170-kDa polypeptide was also observed using [125I]Tyr12-miniglucagon as the radioligand. Together, these data suggest that the 170-kDa glycoprotein represents a novel glucagon-degrading activity that could mediate glucagon proteolysis within endosomes before the acidification step and generate the bioactive (19-29) miniglucagon peptide. PMID:12959981

  11. Hyaluronidase-sensitive nanoparticle templates for triggered release of HIV/AIDS microbicide in vitro.

    PubMed

    Agrahari, Vivek; Zhang, Chi; Zhang, Tao; Li, Wenjing; Gounev, Todor K; Oyler, Nathan A; Youan, Bi-Botti C

    2014-03-01

    This study was designed to test the hypothesis that a triggered release of a topical microbicide (tenofovir) from hyaluronic acid nanoparticles (HA-NPs) can be achieved under the influence of hyaluronidase (HAase) enzyme. A fractional factorial experimental design was used to examine the factors [molar concentrations of adipic acid dihydrazide (X1) and 1-ethyl-3-[3-dimethylaminopropyl] carbodiimide hydrochloride (X2), volume of acetone (X3) and reaction time (X4)] influencing the responses, Y1; particle mean diameter: PMD (nanometers: nm), Y2; polydispersity index: PDI and Y3; zeta (ζ) potential: (millivolts). The amide bond formation between HA and ADH after cross-linking was confirmed by FT-IR and (13)C-NMR analyses. These NPs were also characterized for cytotoxicity on a human vaginal epithelial cell line and L. crispatus. When formulated with factors X1; 2.49 mM, X2; 9.96 mM, X3; 60 mL, X4; 6 h, HA-NPs exhibited a spherical shape with PMD, PDI, ζ potential, encapsulation efficiency, and drug loading of 70.6 ± 4.1 nm, 0.07 ± 0.02, -38.2 ± 2.8 mV, 51.8 ± 2.4% w/w and 26.1 ± 1.2% w/w, respectively, (n = 3). Unlike for HA based gel, HAase significantly triggered the drug release and HA degradation from the NPs after 24 h (~90% w/w and 65% w/w, respectively); whereas, in its absence, these values were ~39% w/w and 26% w/w, respectively. The NPs were non-cytotoxic to human vaginal VK2/E6E7, End1/E6E7 cells and Lactobacillus crispatus. These data highlight the potential of HAase-sensitive HA-NPs templates for the controlled and vaginal delivery of anti-HIV/AIDS microbicides. PMID:24343770

  12. In vivo real-time bioimaging of hyaluronic acid derivatives using quantum dots.

    PubMed

    Kim, Jiseok; Kim, Ki Su; Jiang, Ge; Kang, Hyungu; Kim, Sungjee; Kim, Byung-Soo; Park, Moon Hyang; Hahn, Sei Kwang

    2008-12-01

    The effect of chemical modification of hyaluronic acid (HA) on its distribution throughout the body was successfully visualized in nude mice through real-time bioimaging using quantum dots (QDots). Adipic acid dihydrazide modified HA (HA-ADH) was synthesized and conjugated with QDots having carboxyl terminal ligands activated with 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide and N-hydroxysulfosuccinimide. The formation of HA-QDot conjugates could be confirmed by gel permeation chromatography, fluorometry, transmission electron microscopy, and zeta-size analysis. According to the real-time bioimaging of HA-QDot conjugates after subcutaneous injection to nude mice, the fluorescence of HA-QDot conjugates with a near infrared wavelength of 800 nm could be detected up to 2 months, whereas that with an emission wavelength of 655 nm disappeared almost completely within 5 days. The results can be ascribed to the fact that near-infrared light has a high penetration depth of about 5-6 cm in the body compared to that of about 7-10 mm for visible light. Thereby, using QDots with a near-infrared emission wavelength of 800 nm, the distribution of HA-QDot conjugates throughout the body was bioimaged in real-time after their tail-vein injection into nude mice. HA-QDot conjugates with 35 mol% ADH content maintaining enough binding sites for HA receptors were mainly accumulated in the liver, while those with 68 mol% ADH content losing much of HA characteristics were evenly distributed to the tissues in the body. The results are well matched with the fact that HA receptors are abundantly present in the liver with a high specificity to HA molecules. PMID:18690665

  13. Conjugation of an antibody to cross-linked fibrin for targeted delivery of anti-restenotic drugs.

    PubMed

    Thomas, Anita C; Campbell, Julie H

    2004-12-10

    There is an urgent need to treat restenosis, a major complication of the treatment of arteries blocked by atherosclerotic plaque, using local delivery techniques. We observed that cross-linked fibrin (XLF) is deposited at the site of surgical injury of arteries. An antibody to XLF, conjugated to anti-restenotic agents, should deliver the drugs directly and only to the site of injury. An anti-XLF antibody (H93.7C.1D2/48; 1D2) was conjugated to heparin (using N-succinimidyl 3-(2-pyridyldithio)propionate), low molecular weight heparin (LMWH) (adipic acid dihydrazide) and rapamycin (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide/N-hydroxysuccinimide), and the conjugates purified and tested for activity before use in vivo. Rabbits had their right carotid arteries de-endothelialised and then given a bolus of 1D2-heparin, 1D2-LMWH or 1D2-rapamycin conjugate or controls of saline, heparin, LMWH, rapamycin or 1D2 (+/-heparin bolus) and sacrificed after 2 or 4 weeks (12 groups, n=6/group). Rabbits given any of the conjugates had minimal neointimal development in injured arteries, with up to 59% fewer neointimal cells than those given control drugs. Rabbits given 1D2-heparin or 1D2-LMWH had an increased or insignificant reduction in luminal area, with positive remodelling, while the medial and total arterial areas of rabbits given 1D2-rapamycin were not affected by injury. Arteries exposed to 1D2-heparin or 1D2-rapamycin had more endothelial cells than rabbits given control drugs. Thus, XLF-antibodies can site-deliver anti-restenotic agents to injured areas of the artery wall, where the conjugates can influence remodelling, re-endothelialisation and neointimal cell density, with reduced neointimal formation. PMID:15567502

  14. A Novel Europium Chelate Coated Nanosphere for Time-Resolved Fluorescence Immunoassay

    PubMed Central

    Shen, Yifeng; Xu, Shaohan; He, Donghua

    2015-01-01

    A novel europium ligand 2, 2’, 2’’, 2’’’-(4, 7-diphenyl-1, 10-phenanthroline-2, 9-diyl) bis (methylene) bis (azanetriyl) tetra acetic acid (BC-EDTA) was synthesized and characterized. It shows an emission spectrum peak at 610 nm when it is excited at 360 nm, with a large Stock shift (250 nm). It is covalently coated on the surface of a bare silica nanosphere containi free amino groups, using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride and N-Hydroxysuccinimide. We also observed an interesting phenomenon that when BC-EDTA is labeled with a silica nanosphere, the chelate shows different excitation spectrum peaks of about 295 nm. We speculate that the carboxyl has a significant influence on its excitation spectrum. The BC-EDTA/Eu3+coated nanosphere could be used as a fluorescent probe for time-resolved fluorescence immunoassay. We labeled the antibody with the fluorescent nanosphere to develop a nanosphere based hepatitis B surface antigen as a time-resolved fluorescence immunoassay reagent, which is very easy to operate and eliminates potential contamination of Eu3+ contained in the environment. The analytical and functional sensitivities are 0.0037 μg/L and 0.08 μg/L (S/N≥2.0) respectively. The detection range is 0.08-166.67 μg/L, which is much wider than that of ELISA (0.2-5μg/L). It is comparable to the commercial dissociation-enhanced lanthanide fluoro-immunoassay system (DELFIA) reagents (0.2-145μg/L). We propose that it can fulfill clinical applications. PMID:26056826

  15. Determination of small halogenated carboxylic acid residues in drug substances by high performance liquid chromatography-diode array detection following derivatization with nitro-substituted phenylhydrazines.

    PubMed

    Hou, Desheng; Fan, Jingjing; Han, Lingfei; Ruan, Xiaoling; Feng, Feng; Liu, Wenyuan; Zheng, Feng

    2016-03-18

    A method for the determination of small halogenated carboxylic acid (HCA) residues in drug substances is urgently needed because of the potential of HCAs for genotoxicity and carcinogenicity in humans. We have now developed a simple method, involving derivatization followed by high performance liquid chromatography-diode array detection (HPLC-DAD), for the determination of six likely residual HCAs (monochloroacetic acid, monobromoacetic acid, dichloroacetic acid, 2-chloropropionic acid, 2-bromopropionic acid and 3-chloropropionic acid) in drug substances. Different nitro-substituted phenylhydrazines (NPHs) derivatization reagents were systematically compared and evaluated. 2-Nitrophenylhydrazine hydrochloride (2-NPH·HCl) was selected as the most suitable choice since its derivatives absorb strongly at 392 nm, a region of the spectrum where most drug substances and impurities absorb very weakly. During the derivatization process, the commonly used catalyst, pyridine, caused rapid dechlorination or chlorine substitution of α-halogenated derivatives. To avoid these unwanted side reactions, a reliable derivatization method that did not use pyridine was developed. Reaction with 2-NPH·HCl using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride as coupling agent in acetonitrile-water (70:30) at room temperature for 2h gave complete reaction and avoided degradation products. The derivatives were analyzed, without any pretreatment, using gradient HPLC with detection in the near visible region. Organic acids commonly found in drug substances and other impurities did not interfere with the analysis. Good linearity (r>0.999) and low limits of quantitation (0.05-0.12 μg mL(-1)) were obtained. The mean recoveries were in the range of 80-115% with RSD <5.81% except for 3-CPA in ibuprofen which was 78.5%. The intra- and inter-day precisions were expressed as RSD <1.98% and <4.39%, respectively. Finally, the proposed method was successfully used for the residue

  16. Applying graphene oxide nano-film over a polycarbonate nanoporous membrane to monitor E. coli by infrared spectroscopy.

    PubMed

    Singh, Krishna Pal; Dhek, Neeraj Singh; Nehra, Anuj; Ahlawat, Sweeti; Puri, Anu

    2017-01-01

    Nano-biosensors are excellent monitoring tools for rapid, specific, sensitive, inexpensive, in-field, on-line, and/or real-time detection of pathogens in foods, soil, air, and water samples. A variety of nano-materials (metallic, polymeric, and/or carbon-based) were employed to enhance the efficacy, efficiency, and sensitivity of these nano-biosensors, including graphene-based materials, especially graphene oxide (GO)-based materials. GO bears many oxygen-bearing groups, enabling ligand conjugation at the high density critical for sensitive detection. We have fabricated GO-modified nano-porous polycarbonate track-etched (PCTE) membranes that were conjugated to an Escherichia coli-specific antibody (Ab) and used to detect E. coli. The random distribution of nanopores on the PCTE membrane surface and the bright coating of the GO onto the membrane were confirmed by scanning electron microscope. Anti-E. coli β-gal Abs were conjugated to the GO surface via 1-ethyl-3,3-dimethylaminopropyl carbodiimide hydrochloride-N-hydroxysuccinimide chemistry; antibody coating was confirmed by the presence of a characteristic IR peak near 1600cm(-1). A non-corresponding Ab (anti-Pseudomonas) was used as a negative control under identical conditions. When E. coli interacted anti-E.coli β-gal with Ab-coated GO-nano-biosensor units, we observed a clear shift in the IR peak from 3373.14 to 3315cm(-1); in contrast, we did not observe any shift in IR peaks when the GO unit was coated with the non-corresponding Ab (anti-Pseudomonas). Therefore, the detection of E. coli using the described GO-nano-sensor unit is highly specific, is highly selective and can be applied for real-time monitoring of E. coli with a detection limit between 100μg/mL and 10μg/mL, similar to existing detection systems. PMID:27391314

  17. Role of the Dinitrogenase Reductase Arginine 101 Residue in Dinitrogenase Reductase ADP-Ribosyltransferase Binding, NAD Binding, and Cleavage

    PubMed Central

    Ma, Yan; Ludden, Paul W.

    2001-01-01

    Dinitrogenase reductase is posttranslationally regulated by dinitrogenase reductase ADP-ribosyltransferase (DRAT) via ADP-ribosylation of the arginine 101 residue in some bacteria. Rhodospirillum rubrum strains in which the arginine 101 of dinitrogenase reductase was replaced by tyrosine, phenylalanine, or leucine were constructed by site-directed mutagenesis of the nifH gene. The strain containing the R101F form of dinitrogenase reductase retains 91%, the strain containing the R101Y form retains 72%, and the strain containing the R101L form retains only 28% of in vivo nitrogenase activity of the strain containing the dinitrogenase reductase with arginine at position 101. In vivo acetylene reduction assays, immunoblotting with anti-dinitrogenase reductase antibody, and [adenylate-32P]NAD labeling experiments showed that no switch-off of nitrogenase activity occurred in any of the three mutants and no ADP-ribosylation of altered dinitrogenase reductases occurred either in vivo or in vitro. Altered dinitrogenase reductases from strains UR629 (R101Y) and UR630 (R101F) were purified to homogeneity. The R101F and R101Y forms of dinitrogenase reductase were able to form a complex with DRAT that could be chemically cross-linked by 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide. The R101F form of dinitrogenase reductase and DRAT together were not able to cleave NAD. This suggests that arginine 101 is not critical for the binding of DRAT to dinitrogenase reductase but that the availability of arginine 101 is important for NAD cleavage. Both DRAT and dinitrogenase reductase can be labeled by [carbonyl-14C]NAD individually upon UV irradiation, but most 14C label is incorporated into DRAT when both proteins are present. The ability of R101F dinitrogenase reductase to be labeled by [carbonyl-14C]NAD suggested that Arg 101 is not absolutely required for NAD binding. PMID:11114923

  18. Biotinylated recombinant human erythropoietins: Bioactivity and utility as receptor ligand

    SciTech Connect

    Wojchowski, D.M.; Caslake, L. )

    1989-08-15

    Recombinant human erythropoietin labeled covalently with biotin at sialic acid moieties has been prepared, and has been shown to possess high biological activity plus utility as a receptor ligand. Initially, the effects on biological activity of covalently attaching biotin to erythropoietin alternatively at carboxylate, amino, or sialic acid groups were compared. Biotinylation of erythropoietin at carboxylate groups using biotin-amidocaproyl hydrazide plus 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide led to substantial biological inactivation, although biotinylated molecules retained detectable activity when prepared at low stoichiometries. Biotinylation at amino groups using sulfosuccinimidyl 6-(biotinamido) hexanoate resulted in a high level of biological inactivation with little, if any, retention of biological activity, regardless of labeling stoichiometries. Biotinylation at sialic acid moieties using periodate and biotinamidocaproyl hydrazide proceeded efficiently (greater than 95% and 80% labeling efficiencies for human urinary and recombinant erythropoietin, respectively) and yielded stably biotinylated erythropoietin molecules possessing comparably high biological activity (ie, 45% of the activity of unmodified hormone). Utility of recombinant biotin-(sialyl)-erythropoietin (in combination with 125I-streptavidin) in the assay of cell surface receptors was demonstrated using two distinct murine erythroleukemia cell lines, Friend 745 and Rauscher Red 1. The densities and affinities of specific hormone binding sites were 116 +/- 4 sites, 3.3 +/- 0.4 nmol/L kd and 164 +/- 5 sites, 2.7 +/- 0.4 nmol/L kd, respectively. It is predicted that the present development of biotin-(sialyl)-erythropoietin as a chemically and biologically stable, bioactive ligand will assist in advancing an understanding of the regulated expression and physicochemistry of the human and murine erythropoietin receptors.

  19. CCHCR1 interacts with EDC4, suggesting its localization in P-bodies

    SciTech Connect

    Ling, Y.H.; Wong, C.C.; Li, K.W.; Chan, K.M.; Boukamp, P.; Liu, W.K.

    2014-09-10

    Coiled‐coil alpha‐helical rod protein 1 (CCHCR1) is suggested as a candidate biomarker for psoriasis for more than a decade but its function remains poorly understood because of the inconsistent findings in the literature. CCHCR1 protein is suggested to be localized in the cytoplasm, nucleus, mitochondria, or centrosome and to regulate various cellular functions, including steroidogenesis, proliferation, differentiation, and cytoskeleton organization. In this study, we attempted to find a consensus between these findings by identifying the interaction partners of CCHCR1 using co-immunoprecipiation with a stable cell line expressing EGFP-tagged CCHCR1. Out of more than 100 co-immunoprecipitants identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS), the enhancer of mRNA-decapping protein 4 (EDC4), which is a processing body (P-body) component, was particularly found to be the major interacting partner of CCHCR1. Confocal imaging confirmed the localization of CCHCR1 in P-bodies and its N-terminus is required for this subcellular localization, suggesting that CCHCR1 is a novel P-body component. As P-bodies are the site for mRNA metabolism, our findings provide a molecular basis for the function of CCHCR1, any disruption of which may affect the transcriptome of the cell, and causing abnormal cell functions. - Highlights: • We identified CCHCR1 as a novel P-body component. • We identified EDC4 as the major interacting partner of CCHCR1. • N-terminus of CCHCR1 protein is required for its P-bodies localization.

  20. INTEGRATED IN VITRO TESTING AND QSAR MODEL DEVELOPMENT TO RANK AND PRIORITIZE CHEMICALS FOR TIERED ENDOCRINE DISRUPTING CHEMICAL (EDC) TESTING

    EPA Science Inventory

    The primary objective of this research is to develop and validate an integrate suite of in vitro bioassays that may serve as the basis for models to rank and prioritize large inventories of chemicals for EDC activity. The assays will be developed using an aquatic species and foc...

  1. Developing Alliances to Improve Health and Education: Reflections of Leaders from EDC's Health and Human Development Programs

    ERIC Educational Resources Information Center

    Education Development Center, Inc, 2005

    2005-01-01

    Over the past two decades, the division of Health and Human Development Programs at Education Development Center, Inc. (HHD/EDC), has often played a catalytic and facilitative role to create and manage alliances, which address particular challenges, such as improving the health of students and school staff, protecting worker safety, or promoting…

  2. Degradation of the endocrine disrupting chemicals (EDCs) carbamazepine, clofibric acid, and iopromide by corona discharge over water.

    PubMed

    Krause, Holger; Schweiger, Bianca; Schuhmacher, Jörg; Scholl, Saskia; Steinfeld, Ute

    2009-04-01

    Common wastewater treatment plants often do not eliminate endocrine disrupting chemicals (EDCs). Aqueous solutions of three EDCs were treated with an enhanced corona discharge technology. The three EDCs were clofibric acid, a blood lipid regulator, carbamazepine, an antiepileptic drug, and iopromide, a contrast media. To simulate real conditions, EDC solutions containing landfill leachate were also used. In our setup, two barrier electrodes provided an atmospheric pressure corona discharge over a thin water film, in which the counter-electrode was submerged. Clofibric acid, carbamazepine, and iopromide were effectively removed from a single solution. After a treatment of 15min, there were no traces of iopromide estrogen activity either as a single substance or as degradation products when using an E-Screen Assay. Continuous treatment was compared with pulsed treatment using carbamazepine solutions mixed with pretreated landfill leachate. Best degradation results were achieved with a 500 W continuous duty cycle treatment. Counter-electrodes from materials such as boron doped diamond (BDD), titanium iridium oxide, and iron were investigated for their influences on the process effectivity. Significant improvements were achieved by using an enclosed reactor, BDD electrodes, and circulating only a fresh air or argon/air mixture as cooling gas through the barrier electrodes. PMID:19150730

  3. A systematic analysis of DMTMM vs EDC/NHS for ligation of amines to hyaluronan in water.

    PubMed

    D'Este, Matteo; Eglin, David; Alini, Mauro

    2014-08-01

    The activation of carboxyl groups with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride and N-hydroxysuccinimide (EDC/NHS) for amide formation is the standard method for amine ligation to hyaluronan (HA), and a very well established wide-ranging bioconjugation method. In this paper we compare 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM) to EDC/NHS activation chemistry for HA ligation using an array of substrates including small, large and functional molecules. For all the substrates tested DMTMM yields were superior at parity of feed ratio. DMTMM chemistry resulted effective also in absence of pH control, which is essential for EDC/NHS conjugation. Overall our results demonstrate that DMTMM is more efficient than EDC/NHS for ligation of amines to HA and does not require accurate pH control or pH shift during the reaction to be effective. DMTMM-mediated ligation is a new promising chemical tool to synthesize HA derivatives for biomedical and pharmaceutical applications. PMID:24751270

  4. TRANSGENERATIONAL (IN UTERO/LACTATIONAL) EXPOSURE PROTOCOL TO INVESTIGATE THE EFFECTS OF ENDOCRINE DISRUPTING COMPOUNDS (EDCS) IN RATS

    EPA Science Inventory

    This protocol is designed to evaluate the effects of Endocrine Disrupting Compounds (EDCs) through fetal (transplacental) and/or neonatal (via the dam's milk) exposure during the critical periods of reproductive organogenesis in the rat. Continued direct exposure to the F1 pups...

  5. METABOLOMICS AS A TOOL FOR DISCRIMINATING AMONG ADAPTIVE, COMPENSATORY, AND TOXIC RESPONSES UPON EXPOSURE OF SMALL FISH TO EDCS

    EPA Science Inventory

    Determining the impact(s) of exposure on aquatic organisms by endocrine disrupting compounds (EDCs) is essential for determining the risks that these chemicals pose. However, to accurately evaluate these risks, beyond simply measuring a before and after exposure snapshot, resear...

  6. Detection of target DNA using photo-reactive protoporphyrin moeity on a nanocomposite substrate

    NASA Astrophysics Data System (ADS)

    Das, Sumana; Mishra, Madhusmita; Vasireddi, Ramakrishna; Roy Mahapatra, D.

    2014-03-01

    Detection of pathogens from infected biological samples through conventional process involves cell lysis and purification. The main objective of this work is to minimize the time and sample loss, as well as to increase the efficiency of detection of biomolecules. Electrical lysis of medical sample is performed in a closed microfluidic channel in a single integrated platform where the downstream analysis of the sample is possible. The device functions involve, in a sequence, flow of lysate from lysis chamber passed through a thermal denaturation counter where dsDNA is denatured to ssDNA, which is controlled by heater unit. A functionalized binding chamber of ssDNA is prepared by using ZnO nanorods as the matrix and functionalized with bifunctional carboxylic acid, 16-(2-pyridyldithiol) hexadecanoic acid (PDHA) which is further attached to a linker molecule 1-ethyl-3-(3-dimethylaminopropyl) (EDC). Linker moeity is then covalently bound to photoreactive protoporphyrin (PPP) molecule. The photolabile molecule protoporphyrin interacts with -NH2 labeled single stranded DNA (ssDNA) which thus acts as a probe to detect complimentary ssDNA from target organisms. Thereafter the bound DNA with protoporphyrin is exposed to an LED of particular wavelength for a definite period of time and DNA was eluted and analyzed. UV/Vis spectroscopic analysis at 260/280 nm wavelength confirms the purity and peak at 260 nm is reconfirmed for the elution of target DNA. Quantitative and qualitative data obtained from the current experiments show highly selective detection of biomolecule such as DNA which have large number of future applications in Point-of-Care devices.

  7. Stabilization of collagen nanofibers with l-lysine improves the ability of carbodiimide cross-linked amniotic membranes to preserve limbal epithelial progenitor cells

    PubMed Central

    Lai, Jui-Yang; Wang, Pei-Ran; Luo, Li-Jyuan; Chen, Si-Tan

    2014-01-01

    To overcome the drawbacks associated with limited cross-linking efficiency of carbodiimide modified amniotic membrane, this study investigated the use of l-lysine as an additional amino acid bridge to enhance the stability of a nanofibrous tissue matrix for a limbal epithelial cell culture platform. Results of ninhydrin assays and zeta potential measurements showed that the amount of positively charged amino acid residues incorporated into the tissue collagen chains is highly correlated with the l-lysine-pretreated concentration. The cross-linked structure and hydrophilicity of amniotic membrane scaffolding materials affected by the lysine molecular bridging effects were determined. With an increase in the l-lysine-pretreated concentration from 1 to 30 mM, the cross-linking density was significantly increased and water content was markedly decreased. The variations in resistance to thermal denaturation and enzymatic degradation were in accordance with the number of cross-links per unit mass of amniotic membrane, indicating l-lysine-modulated stabilization of collagen molecules. It was also noteworthy that the carbodiimide cross-linked tissue samples prepared using a relatively high l-lysine-pretreated concentration (ie, 30 mM) appeared to have decreased light transmittance and biocompatibility, probably due to the influence of a large nanofiber size and a high charge density. The rise in stemness gene and protein expression levels was dependent on improved cross-link formation, suggesting the crucial role of amino acid bridges in constructing suitable scaffolds to preserve limbal progenitor cells. It is concluded that mild to moderate pretreatment conditions (ie, 3–10 mM l-lysine) can provide a useful strategy to assist in the development of carbodiimide cross-linked amniotic membrane as a stable stem cell niche for corneal epithelial tissue engineering. PMID:25395849

  8. Results of the TOP Study: Prospectively Randomized Multicenter Trial of an Ex Vivo Tacrolimus Rinse Before Transplantation in EDC Livers

    PubMed Central

    Pratschke, Sebastian; Arnold, Hannah; Zollner, Alfred; Heise, Michael; Pascher, Andreas; Schemmer, Peter; Scherer, Marcus N.; Bauer, Andreas; Jauch, Karl-Walter; Werner, Jens; Guba, Markus; Angele, Martin K.

    2016-01-01

    Background Organ shortage results in the transplantation of extended donor criteria (EDC) livers which is associated with increased ischemia-reperfusion injury (IRI). Experimental studies indicate that an organ rinse with the calcineurin inhibitor tacrolimus before implantation protects against IRI. The tacrolimus organ perfusion study was initiated to examine the effects of ex vivo tacrolimus perfusion on IRI in transplantation of EDC livers. Methods A prospective randomized multicenter trial comparing ex vivo perfusion of marginal liver grafts (≥2 EDC according to Eurotransplant manual) with tacrolimus (20 ng/mL) or histidine-tryptophane-ketoglutarate solution (control) was carried out at 5 German liver transplant centers (Munich Ludwig-Maximilians University, Berlin, Heidelberg, Mainz, Regensburg) between October 2011 and July 2013. Primary endpoint was the maximum alanine transaminase (ALT) level within 48 hours after transplantation. Secondary endpoints were aspartate transaminase (AST), prothrombine ratio, and graft-patient survival within an observation period of 1 week. After an interim analysis, the study was terminated by the scientific committee after the treatment of 24 patients (tacrolimus n = 11, Control n = 13). Results Tacrolimus rinse did not reduce postoperative ALT peaks compared with control (P = 0.207; tacrolimus: median, 812; range, 362-3403 vs control: median, 652; range, 147-2034). Moreover, ALT (P = 0.100), prothrombine ratio (P = 0.553), and bilirubin (P = 0.815) did not differ between the groups. AST was higher in patients treated with tacrolimus (P = 0.011). Survival was comparable in both groups (P > 0.05). Conclusions Contrary to experimental findings, tacrolimus rinse failed to improve the primary endpoint of the study (ALT). Because 1 secondary endpoint (AST) was even higher in the intervention group, the study was terminated prematurely. Thus, tacrolimus rinse cannot be recommended in transplantation of EDC livers. PMID:27500266

  9. Clean photodecomposition of 1-methyl-4-phenyl-1H-tetrazole-5(4H)-thiones to carbodiimides proceeds via a biradical

    PubMed Central

    Alawode, Olajide E.; Robinson, Colette; Rayat, Sundeep

    2010-01-01

    The photochemistry of 1-methyl-4-phenyl-1H-tetrazole-5(4H)-thione (1a) and 1-(3-methoxyphenyl)-4-methyl-1H-tetrazole-5(4H)-thione (1b) was studied in acetonitrile at 254 and 300 nm which involves expulsion of dinitrogen and sulfur to form the respective carbodiimides 5a – b as sole photoproducts. Photolysis of the title compounds in the presence of 1,4-cyclohexadiene trap led to the formation of respective thioureas, providing strong evidence for the intermediacy of a 1,3-biradical formed by the loss of dinitrogen. In contrast, a trapping experiment with cyclohexene provided no evidence to support an alternative pathway of photodecomposition involving initial desulfurization followed by loss of dinitrogen via the intermediacy of a carbene. Triplet sensitization and triplet quenching studies argue against the involvement of a triplet excited state. While the quantum yields for the formation of the carbodiimides 5a – b were modest, and showed little change on going from a C6H5 (1a) to mOMeC6H4 (1b) substituent on the tetrazolethione ring, the highly clean photodecomposition of these compounds to a photostable end product makes them promising lead structures for industrial, agricultural and medicinal applications. PMID:21142194

  10. Reactions of titanocene bis(trimethylsilyl)acetylene complexes with carbodiimides: an experimental and theoretical study of complexation versus C-N bond activation.

    PubMed

    Haehnel, Martin; Ruhmann, Martin; Theilmann, Oliver; Roy, Subhendu; Beweries, Torsten; Arndt, Perdita; Spannenberg, Anke; Villinger, Alexander; Jemmis, Eluvathingal D; Schulz, Axel; Rosenthal, Uwe

    2012-09-26

    The reaction of the low valent metallocene(II) sources Cp'(2)Ti(η(2)-Me(3)SiC(2)SiMe(3)) (Cp' = η(5)-cyclopentadienyl, 1a or η(5)-pentamethylcyclopentadienyl, 1b) with different carbodiimide substrates RN═C═NR' 2-R-R' (R = t-Bu; R' = Et; R = R' = i-Pr; t-Bu; SiMe(3); 2,4,6-Me-C(6)H(2) and 2,6-i-Pr-C(6)H(3)) was investigated to explore the frontiers of ring strained, unusual four-membered heterometallacycles 5-R. The product complexes show dismantlement, isomerization, or C-C coupling of the applied carbodiimide substrates, respectively, to form unusual mono-, di-, and tetranuclear titanium(III) complexes. A detailed theoretical study revealed that the formation of the unusual complexes can be attributed to the biradicaloid nature of the unusual four-membered heterometallacycles 5-R, which presents an intriguing situation of M-C bonding. The combined experimental and theoretical study highlights the delicate interplay of electronic and steric effects in the stabilization of strained four-membered heterometallacycles, accounting for the isolation of the obtained complexes. PMID:22891968

  11. Characterization of emissions of dioxins and furans from ethylene dichloride (EDC), vinyl chloride (VCM) and polyvinylchloride (PVC) facilities in the United States.

    SciTech Connect

    Carroll, W.F. Jr.; Borrelli, F.E.; Garrity, P.J.; Jacobs, R.A.; Lewis, J.W.; McCreedy, R.L.; Weston, A.F.; Ledvina, J.C.

    1997-12-31

    Members of The Vinyl Inst., under the auspices of its Dioxin Characterization Program have analyzed for potential dioxin/furan (PCDD/F) concentrations in polyvinylchloride (PVC) resins, treated wastewater effluent, ethylene dichloride (EDC) product and wastewater sludge at EDC, vinyl chloride (VCM) and PVC facilities. No 2,3,7,8-TCDD was detected in any sample analyzed under the program to date. Results from wastewater sludge analysis are pending. Trace concentrations (low pg/g) of PCDD/F were detected in only a few samples of PVC resins and ethylene dichloride (EDC) product. Treated wastewater contained low ppq concentrations of PCDD/F. All concentrations are expressed as Toxicity Equivalents (TEQ). Extrapolation of these data shows that the contribution of EDC/VCM/PVC manufacturing via these media constitutes less than 1 percent of the US annual dioxin emission to the environment.

  12. Allergen Quantification by Use of Electrostatic Dust Collectors (EDCs): Influence of Deployment Time, Extraction Buffer, and Storage Conditions on the Results.

    PubMed

    Sander, Ingrid; Lotz, Anne; Zahradnik, Eva; Raulf, Monika

    2016-08-01

    Sampling of endotoxin, beta-glucan, or allergens on electrostatic dust collectors (EDCs) is a convenient method for exposure assessment. However, especially for allergens few experiments on validation of this method concerning deployment time or storage and extraction procedure have been performed. The aim of study was to optimize the EDC procedure for sampling of allergens in indoor environments. EDCs were placed in households or day-care centers and after extraction, allergens were quantified by six immunoassays detecting mite antigens (Domestic mites DM, Dermatophagoides pteronyssinus Dp, Tyrophagus putrescentiae Tp) or the main allergens from cat (Fel d 1), dog (Can f 1) and mouse (Mus m 1). For 20 EDC holders, deployment times of cloths were varied between 7 and 28 days, 36 EDCs were used to test reproducibility, and for 28 EDCs extraction buffers were varied (with or without 0.05% Tween 20, borate, or phosphate buffer). The influence of storage of cloths at room temperature (2-629 days) or extracts at -80°C (7-639 days), and variation of extract storage temperature (-20°C and -80°C) for long time storage (1.5 years) on the outcome of allergen quantification were tested for about 150 EDCs. The allergens on EDC cloths increased proportionally with deployment time, and allergen loads on parallel sampled tissues were significantly correlated (P < 0.0001, Pearson of log-transformed values 0.91-0.99). Extraction without Tween reduced all results (P < 0.0001, -51% DM, -85% Dp, -60% Tp, -99% Fel d 1, -86% Can f 1, -52% Mus m 1), and borate buffer resulted in lower yields of Mus m 1 (-53%), DP (-45%), and Tp (-27%) than phosphate buffer. Storage of cloths at room temperature significantly decreased Can f 1 levels (P < 0.0001, -4.8% loss for every 30 days), whereas storage of extracts at -80°C decreased DM results (P < 0.0001, -1.2% loss for every 30 days). Extracts stored at -20°C gave at mean 12% higher DM results compared to extracts stored at -80°C for 1

  13. Interactions of fish gelatin and chitosan in uncrosslinked and crosslinked with EDC films: FT-IR study

    NASA Astrophysics Data System (ADS)

    Staroszczyk, Hanna; Sztuka, Katarzyna; Wolska, Julia; Wojtasz-Pająk, Anna; Kołodziejska, Ilona

    2014-01-01

    Films based on fish gelatin, chitosan and blend of fish gelatin and chitosan before and after cross-linking with EDC have been characterized by FT-IR spectroscopy. The FT-IR spectrum of fish gelatin film showed the characteristic amide I, amide II and amide III bands, and the FT-IR spectrum of chitosan film confirmed that the polymer was only a partially deacetylated product, and included CH3sbnd Cdbnd O and NH2 groups, the latter both in their free -NH2 and protonated -NH3+ form. Analysis of FT-IR spectra of two-component, fish gelatin-chitosan film revealed the formation not only of hydrogen bonds within and between chains of polymers, but also of electrostatic interactions between -COO- of gelatin and -NH3+ of chitosan. Modification with EDC provided cross-linking of composites of the film. New iso-peptide bonds formed between activated carboxylic acid groups of glutamic or aspartic acid residue of gelatin and amine groups of gelatin or/and chitosan.

  14. Simultaneously photocatalytic treatment of hexavalent chromium (Cr(VI)) and endocrine disrupting compounds (EDCs) using rotating reactor under solar irradiation.

    PubMed

    Kim, Youngji; Joo, Hyunku; Her, Namguk; Yoon, Yeomin; Sohn, Jinsik; Kim, Sungpyo; Yoon, Jaekyung

    2015-05-15

    In this study, simultaneous treatments, reduction of hexavalent chromium (Cr(VI)) and oxidation of endocrine disrupting compounds (EDCs), such as bisphenol A (BPA), 17α-ethinyl estradiol (EE2) and 17β-estradiol (E2), were investigated with a rotating photocatalytic reactor including TiO₂ nanotubes formed on titanium mesh substrates under solar UV irradiation. In the laboratory tests with a rotating type I reactor, synergy effects of the simultaneous photocatalytic reduction and oxidation of inorganic (Cr(VI)) and organic (BPA) pollutants were achieved. Particularly, the concurrent photocatalytic reduction of Cr(VI) and oxidation of BPA was higher under acidic conditions. The enhanced reaction efficiency of both pollutants was attributed to a stronger charge interaction between TiO₂ nanotubes (positive charge) and the anionic form of Cr(VI) (negative charge), which are prevented recombination (electron-hole pair) by the hole scavenging effect of BPA. In the extended outdoor tests with a rotating type II reactor under solar irradiation, the experiment was extended to examine the simultaneous reduction of Cr(VI) in the presence of additional EDCs, such as EE2 and E2 as well as BPA. The findings showed that synergic effect of both photocatalytic reduction and oxidation was confirmed with single-component (Cr(VI) only), two-components (Cr(VI)/BPA, Cr(VI)/EE2, and Cr(VI)/E2), and four-components (Cr(VI)/BPA/EE2/E2) under various solar irradiation conditions. PMID:25698573

  15. The activation of the decapping enzyme DCP2 by DCP1 occurs on the EDC4 scaffold and involves a conserved loop in DCP1

    PubMed Central

    Chang, Chung-Te; Bercovich, Natalia; Loh, Belinda; Jonas, Stefanie; Izaurralde, Elisa

    2014-01-01

    The removal of the 5′-cap structure by the decapping enzyme DCP2 and its coactivator DCP1 shuts down translation and exposes the mRNA to 5′-to-3′ exonucleolytic degradation by XRN1. Although yeast DCP1 and DCP2 directly interact, an additional factor, EDC4, promotes DCP1–DCP2 association in metazoan. Here, we elucidate how the human proteins interact to assemble an active decapping complex and how decapped mRNAs are handed over to XRN1. We show that EDC4 serves as a scaffold for complex assembly, providing binding sites for DCP1, DCP2 and XRN1. DCP2 and XRN1 bind simultaneously to the EDC4 C-terminal domain through short linear motifs (SLiMs). Additionally, DCP1 and DCP2 form direct but weak interactions that are facilitated by EDC4. Mutational and functional studies indicate that the docking of DCP1 and DCP2 on the EDC4 scaffold is a critical step for mRNA decapping in vivo. They also revealed a crucial role for a conserved asparagine–arginine containing loop (the NR-loop) in the DCP1 EVH1 domain in DCP2 activation. Our data indicate that DCP2 activation by DCP1 occurs preferentially on the EDC4 scaffold, which may serve to couple DCP2 activation by DCP1 with 5′-to-3′ mRNA degradation by XRN1 in human cells. PMID:24510189

  16. Site-directed immobilization of antibody using EDC-NHS-activated protein A on a bimetallic-based surface plasmon resonance chip

    NASA Astrophysics Data System (ADS)

    Sohn, Young-Soo; Lee, Yeon Kyung

    2014-05-01

    The characteristics of a waveguide-coupled bimetallic surface plasmon resonance (WcBiM SPR) sensor using (3-dimethylaminopropyl)-3-ethylcarbodiimide(EDC)-N-hydroxysuccinimide(NHS)-activated protein A was investigated, and the detection of IgG using the EDC-NHS-activated protein A was studied in comparison with protein A and a self-assembled monolayer (SAM). The WcBiM sensor, which has a narrower full width at half maximum (FWHM) and a steeper slope, was selected since it leads to a larger change in the reflectance in the intensity detection mode. A preparation of the EDC-NHS-activated protein A for site-directed immobilization of antibodies was relative easily compared to the engineered protein G and A. In antigen-antibody interactions, the response to IgG at the concentrations of 50, 100, and 150 ng/ml was investigated. The results showed that the sensitivity of the WcBiM sensor using the EDC-NHS-activated protein A, protein A, and SAM was 0.0185 [%/(ng/ml)], 0.0065 [%/(ng/ml)], and 0.0101 [%/(ng/ml)], respectively. The lowest detectable concentrations of IgG with the EDC-NHS-activated protein A, protein A, and SAM were 4.27, 12.83, and 8.24 ng/ml, respectively. Therefore, the increased sensitivity and lower detection capability of the WcBiM SPR chip with the EDC-NHS-activated protein A suggests that it could be used in early diagnosis where the trace level concentrations of biomolecules should be detected.

  17. Comparison between two different methods of immobilizing NGF in poly(DL-lactic acid-co-glycolic acid) conduit for peripheral nerve regeneration by EDC/NHS/MES and genipin.

    PubMed

    Hsieh, Shu-Chih; Tang, Cheng-Ming; Huang, Wen-Tao; Hsieh, Ling-Ling; Lu, Chun-Mei; Chang, Chen-Jung; Hsu, Shan-hui

    2011-12-15

    For surface modification and nerve regeneration, chitosan, followed by nerve growth factor (NGF), was immobilized onto the interior surface of poly (lactic acit-co-glycolic) conduits, using EDC/NHS/MES system (EDCs) and genipin (GP). Four new conduits were, therefore, obtained and named by immobilizing order-EDCs/EDCs, GP/EDCs, EDCs/GP, and GP/GP groups. The immobilized methods used were evaluated and compared, respectively. The researchers found that the EDCs- and GP-cross-linked chitosan displayed higher hydrophilic than pure poly (DL-lactic acid-co-glycolic acid) (PLGA) in water contact angle experiment, which meant the cell compatibility was improved by the modification. Scanning electron microscopic observations revealed that the GP-cross-linking of chitosan greatly improved cell compatibility while cultured rat PC12 cells were flatter and more spindle-shaped than EDCs-cross-linked chitosan. The results concerning the GP-cross-linked chitosan revealed significant proliferation of the seeded cells relative to pure PLGA films, as determined by counting cells and MTT assay. The NGF was released from the modified conduits in two separate periods--an initial burst in 5 days and then slow release from day 10 to day 40. The GP/EDCs group had the highest NGF value among all groups after the 5th day. Finally, the controlled-release conduits were used to bridge a 10 mm rat sciatic nerve defect. Six weeks following implantation, morphological analysis revealed the highest numbers of myelinated axons in the midconduit and distal regenerated nerve in GP/EDCs group. Therefore, the results confirm that GP/EDCs groups with good cell compatibility and effective release of NGF can considerably improve peripheral nerve regeneration. PMID:21953828

  18. Effect of carbodiimide-derivatized hyaluronic acid gelatin on preventing postsurgical intra-abdominal adhesion formation and promoting healing in a rat model.

    PubMed

    Yuan, Fang; Lin, Long-Xiang; Zhang, Hui-Hui; Huang, Dan; Sun, Yu-Long

    2016-05-01

    Adhesions often occur after abdominal surgery. It could cause chronic pelvic pain, intestinal obstruction, and infertility. A hydrogel biomaterial, carbodiimide-derivatized hyaluronic acid gelatin (cd-HA gelatin), has been successfully used to reduce adhesion formation after flexor tendon grafting. This study investigated the efficacy of cd-HA gelatin in preventing postsurgical peritoneal adhesions in a rat model. The surgical traumas were created on the underlying muscle of the abdominal wall and the serosal layer of the cecum. The wounds were covered with or without cd-HA gelatin. Animals were euthanized at day 14 after surgery. Adhesion formation was assessed with adhesion degree and adhesion breaking strength. The healing of abdominal wall was evaluated with biomechanical testing and histological analysis. The adhesions occurred in all rats (n = 12) without cd-HA gelatin treatment. The application of cd-HA gelatin significantly reduced the adhesion rate from 100% to 58%. The decrease of adhesion breaking strength also manifested that cd-HA gelatin could reduce postsurgical intra-abdominal adhesion formation. Moreover, it was found that cd-HA gelatin was a safe material and could promote tissue healing. The cd-HA gelatin hydrogel could reduce the formation of intra-abdominal adhesions without adversely effects on wound healing. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1175-1181, 2016. PMID:26749008

  19. 2-pyrazinylnitrene and 4-pyrimidylnitrene. Ring expansion to 1,3,5-triazacyclohepta-1,2,4,6-tetraene and ring opening to (2-Isocyanovinyl)carbodiimide.

    PubMed

    Addicott, Chris; Wong, Ming Wah; Wentrup, Curt

    2002-11-29

    Tetrazolo[1,5-a]pyrazine/2-azidopyrazine 9T/9A undergo photolysis in Ar matrix at cryogenic temperatures to yield 1,3,5-triazacyclohepta-1,2,4,6-tetraene 21 as the first observable intermediate, and 1-cyanoimidazole 11 and (2-isocyanovinyl)carbodiimide 22 as the final products. The latter tautomerizes to 2-(isocyanovinyl)cyanamide 23 on warming to 40 K. The same intermediate 21 and the same final products are obtained on matrix photolysis of the isomeric tetrazolo[1,5-c]pyrimidine/4-azidopyrimidine 24T/24A. These photolysis results as well as those of the previously reported thermal ring contraction of (15)N-labeled 2-pyrazinyl- and 4-pyrimidylnitrenes to 1-cyanoimidazoles can all be rationalized in terms of selective ring opening of 21 or nitrine 10 to a nitrile ylide zwitterion 28 prior to formation of the final products, 11 and 22. The results are supported by high-level ab initio and DFT calculations (CASPT2-CASSCF(6,6), G3(MP2), and B3LYP/6-31+G) of the energies and IR spectra of the intermediates and products. PMID:12444636

  20. Mechanical and structural properties underlying contraction of skeletal muscle fibers after partial 1-ethyl-3-[3-dimethylamino)propyl]carbodiimide cross-linking.

    PubMed Central

    Bershitsky, S; Tsaturyan, A; Bershitskaya, O; Mashanov, G; Brown, P; Webb, M; Ferenczi, M A

    1996-01-01

    We show prolonged contraction of permeabilized muscle fibers of the frog during which structural order, as judged from low-angle x-ray diffraction, was preserved by means of partial cross-linking of the fibers using the zero-length cross-linker 1-ethyl-3-[3-dimethylamino)propyl]carbodiimide. Ten to twenty percent of the myosin cross-bridges were cross-linked, allowing the remaining 80-90% to cycle and generate force. These fibers displayed a well-preserved sarcomeric order and mechanical characteristics similar to those of intact muscle fibers. The intensity of the brightest meridional reflection at 14.5 nm, resulting from the projection of cross-bridges evenly spaced along the myofilament length, decreased by 60% as a relaxed fiber was deprived of ATP and entered the rigor state. Upon activation of a rigorized fiber by the addition of ATP, the intensity of this reflection returned to 97% of the relaxed value, suggesting that the overall orientation of cross-bridges in the active muscle was more perpendicular to the filament axis than in rigor. Following a small-amplitude length step applied to the active fibers, the reflection intensity decreased for both releases and stretches. In rigor, however, a small stretch increased the amplitude of the reflection by 35%. These findings show the close link between cross-bridge orientation and tension changes. Images FIGURE 1 FIGURE 6 PMID:8874020

  1. USING 1H-NMR METABOLOMICS TO CHARACTERIZE ALTERED METABOLIC PROFILES IN ZEBRAFISH (DANIO RERIO) EXPOSED TO THE MODEL EDCS 17 ALPHA-ETHINYLESTRADIOL (EE2) AND FADROZOLE

    EPA Science Inventory

    Elevated levels of endocrine-disrupting chemicals (EDCs) have been reported in waterways worldwide and have been shown to affect numerous aspects of development, behavior, reproduction, and survival in various fish species. We have examined the effects of the synthetic steroid 17...

  2. Possible strategies for EDC testing in the future: exploring roles of pathway-based in silico, in vitro and in vivo methods

    EPA Science Inventory

    Current methods for screening, testing and monitoring endocrine-disrupting chemicals (EDCs) rely relatively substantially upon moderate- to long-term assays that can, in some instances, require significant numbers of animals. Recent developments in the areas of in vitro testing...

  3. Construction of amphiphilic segments on polypropylene nonwoven surface and its application in removal of endocrine disrupting compounds (EDCs) from aqueous solution

    NASA Astrophysics Data System (ADS)

    Liu, Kai; Wei, Junfu; Zhou, Xiangyu; Liu, Nana

    2015-05-01

    The amphiphilic segments on polypropylene nonwoven (PP nonwoven) surface were constructed using the ultraviolet (UV) irradiation graft polymerization for the removal of endocrine disrupting compounds (EDCs) with different polarity from aqueous solution. The stearyl acrylate (SA) as hydrophobic functional monomer was introduced onto the surface of PP nonwoven fabric at first stage and then the hydroxyethyl acrylate (HEA) as hydrophilic functional monomer was introduced subsequently. The effect of functional monomer concentration and UV irradiation time on grafting ratio was studied and discussed. The novel amphiphilic structure was designed and constructed based on adsorption capacity for the target micropollutants. The structure and composition of the amphiphilic adsorption materials were characterized by Fourier transform infrared spectrometer (FTIR), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM) and contact angle (CA). The adsorption behaviors for EDCs of the amphiphilic adsorption materials were studied and the results indicated that the adsorption capacity and adsorption rate were superior to single SA grafted PP nonwoven (PP-g-SA) and single HEA grafted PP nonwoven (PP-g-HEA). The novel amphiphilic adsorption material was efficient for the removal of EDCs with different polarity and could be utilized as a potential adsorption material for removing EDCs from aqueous solution.

  4. 1H-NMR METABOLOMICS ANALYIS OF ZEBRAFISH (DANIO RERIO) EXPOSED TO THE ENVIRONMENTALLY-RELEVANT EDC 17 ALPHA-ETHINYLESTRADIOL (EE2)

    EPA Science Inventory

    Elevated levels of endocrine-disrupting chemicals (EDCs) have been reported in waterways worldwide and have been shown to affect numerous aspects of development, behavior, reproduction, and survival in various fish species. We have examined the effects of the synthetic steroid 1...

  5. Simultaneous determination of PPCPs, EDCs, and artificial sweeteners in environmental water samples using a single-step SPE coupled with HPLC-MS/MS and isotope dilution.

    PubMed

    Tran, Ngoc Han; Hu, Jiangyong; Ong, Say Leong

    2013-09-15

    A high-throughput method for the simultaneous determination of 24 pharmaceuticals and personal care products (PPCPs), endocrine disrupting chemicals (EDCs) and artificial sweeteners (ASs) was developed. The method was based on a single-step solid phase extraction (SPE) coupled with high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and isotope dilution. In this study, a single-step SPE procedure was optimized for simultaneous extraction of all target analytes. Good recoveries (≥ 70%) were observed for all target analytes when extraction was performed using Chromabond(®) HR-X (500 mg, 6 mL) cartridges under acidic condition (pH 2). HPLC-MS/MS parameters were optimized for the simultaneous analysis of 24 PPCPs, EDCs and ASs in a single injection. Quantification was performed by using 13 isotopically labeled internal standards (ILIS), which allows correcting efficiently the loss of the analytes during SPE procedure, matrix effects during HPLC-MS/MS and fluctuation in MS/MS signal intensity due to instrument. Method quantification limit (MQL) for most of the target analytes was below 10 ng/L in all water samples. The method was successfully applied for the simultaneous determination of PPCPs, EDCs and ASs in raw wastewater, surface water and groundwater samples collected in a local catchment area in Singapore. In conclusion, the developed method provided a valuable tool for investigating the occurrence, behavior, transport, and the fate of PPCPs, EDCs and ASs in the aquatic environment. PMID:23708627

  6. Intramolecular [2+2+2] cycloaddition of bis(propargylphenyl)carbodiimides: synthesis of L-shaped π-extended compounds with pyrrolo[1,2-a][1,8]naphthyridine corner units.

    PubMed

    Otani, Takashi; Saito, Takao; Sakamoto, Ryota; Osada, Hiroyuki; Hirahara, Akihito; Furukawa, Naoki; Kutsumura, Noriki; Matsuo, Tsukasa; Tamao, Kohei

    2013-07-14

    L-shaped π-extended penta-, hexa-, and heptacycles with a pyrrolo[1,2-a][1,8]naphthyridine junction were prepared from N,N'-bis[2-(2-alkyn-1-yl)phenyl]carbodiimides or their naphthyl analogs via Rh(I)-catalyzed intramolecular [2+2+2] cycloaddition and dehydrogenative aromatization. These L-shaped compounds emit sky-blue, yellow-green, or golden-orange fluorescence, with high quantum yields. PMID:23736526

  7. The S. pombe mRNA decapping complex recruits cofactors and an Edc1-like activator through a single dynamic surface

    PubMed Central

    Wurm, Jan Philip; Overbeck, Jan; Sprangers, Remco

    2016-01-01

    The removal of the 5′ 7-methylguanosine mRNA cap structure (decapping) is a central step in the 5′–3′ mRNA degradation pathway and is performed by the Dcp1:Dcp2 decapping complex. The activity of this complex is tightly regulated to prevent premature degradation of the transcript. Here, we establish that the aromatic groove of the EVH1 domain of Schizosaccharomyces pombe Dcp1 can interact with proline-rich sequences in the exonuclease Xrn1, the scaffolding protein Pat1, the helicase Dhh1, and the C-terminal disordered region of Dcp2. We show that this region of Dcp1 can also recruit a previously unidentified enhancer of decapping protein (Edc1) and solved the crystal structure of the complex. NMR relaxation dispersion experiments reveal that the Dcp1 binding site can adopt multiple conformations, thus providing the plasticity that is required to accommodate different ligands. We show that the activator Edc1 makes additional contacts with the regulatory domain of Dcp2 and that an activation motif in Edc1 increases the RNA affinity of Dcp1:Dcp2. Our data support a model where Edc1 stabilizes the RNA in the active site, which results in enhanced decapping rates. In summary, we show that multiple decapping factors, including the Dcp2 C-terminal region, compete with Edc1 for Dcp1 binding. Our data thus reveal a network of interactions that can fine-tune the catalytic activity of the decapping complex. PMID:27354705

  8. Systemic Administration of Human Bone Marrow-Derived Mesenchymal Stromal Cell Extracellular Vesicles Ameliorates Aspergillus Hyphal Extract-Induced Allergic Airway Inflammation in Immunocompetent Mice

    PubMed Central

    Cruz, Fernanda F.; Borg, Zachary D.; Goodwin, Meagan; Sokocevic, Dino; Wagner, Darcy E.; Coffey, Amy; Antunes, Mariana; Robinson, Kristen L.; Mitsialis, S. Alex; Kourembanas, Stella; Thane, Kristen; Hoffman, Andrew M.; McKenna, David H.; Rocco, Patricia R.M.

    2015-01-01

    An increasing number of studies demonstrate that administration of either conditioned media (CM) or extracellular vesicles (EVs) released by mesenchymal stromal cells (MSCs) derived from bone marrow and other sources are as effective as the MSCs themselves in mitigating inflammation and injury. The goal of the current study was to determine whether xenogeneic administration of CM or EVs from human bone marrow-derived MSCs would be effective in a model of mixed Th2/Th17, neutrophilic-mediated allergic airway inflammation, reflective of severe refractory asthma, induced by repeated mucosal exposure to Aspergillus hyphal extract (AHE) in immunocompetent C57Bl/6 mice. Systemic administration of both CM and EVs isolated from human and murine MSCs, but not human lung fibroblasts, at the onset of antigen challenge in previously sensitized mice significantly ameliorated the AHE-provoked increases in airway hyperreactivity (AHR), lung inflammation, and the antigen-specific CD4 T-cell Th2 and Th17 phenotype. Notably, both CM and EVs from human MSCs (hMSCs) were generally more potent than those from mouse MSCs (mMSCs) in most of the outcome measures. The weak cross-linking agent 1-ethyl-3-[3-dimethylaminopropyl]carbodiimide hydrochloride was found to inhibit release of both soluble mediators and EVs, fully negating effects of systemically administered hMSCs but only partly inhibited the ameliorating effects of mMSCs. These results demonstrate potent xenogeneic effects of CM and EVs from hMSCs in an immunocompetent mouse model of allergic airway inflammation and they also show differences in mechanisms of action of hMSCs versus mMSCs to mitigate AHR and lung inflammation in this model. Significance There is a growing experience demonstrating benefit of mesenchymal stromal cell (MSC)-based cell therapies in preclinical models of asthma. In the current study, conditioned media (CM) and, in particular, the extracellular vesicle fraction obtained from the CM were as potent as the

  9. Identification of the Binding Region of the [2Fe-2S] Ferredoxin in Stearoyl-Acyl Carrier Protein Desaturase

    PubMed Central

    Sobrado, Pablo; Lyle, Karen S.; Kaul, Steven P.; Turco, Michelle M.; Arabshahi, Ida; Marwah, Ashok; Fox, Brian G.

    2008-01-01

    Stearoyl-acyl carrier protein desaturase (Δ9D) catalyzes the O2 and 2e- dependent desaturation of stearoyl-acyl carrier protein (18:0-ACP) to yield oleoyl-ACP (18:1-ACP). The 2e- are provided by essential interactions with reduced plant-type [2Fe-2S] ferredoxin (Fd). We have investigated the protein-protein interface involved in the Fd-Δ9D complex by use of chemical cross-linking, site-directed mutagenesis, steady-state kinetic approaches and molecular docking studies. Treatment of the different proteins with 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide and N-hydroxysuccinimide revealed that carboxylate residues from Fd and lysine residues from Δ9D contribute to the cross-linking. The single substitutions of K60A, K56A, and K230A on Δ9D decreased the kcat/KM for Fd by 4-, 22- and 2,400-fold, respectively, as compared to wt Δ9D and a K41A substitution. The double substitution K56A/K60A decreased the kcat/KM for Fd by 250-fold, while the triple mutation K56A/K60A/K230A decreased the kcat/KM for Fd by at least 700,000-fold. These results strongly implicate the triad of K56, K60 and K230 of Δ9D in the formation of a catalytic complex with Fd. Molecular docking studies indicate that electrostatic interactions between K56 and K60 and carboxylate groups on Fd may situate the [2Fe-2S] cluster of Fd near to W62, a surface residue that is structurally conserved in both ribonucleotide reductase and mycobacterial putative acyl-ACP desaturase DesA2. Owing to the considerably larger effects on catalysis, K230 appears to have other contributions to catalysis arising from its positioning in helix-7 and its close spatial location to the diiron center ligands E229 and H232. These results are considered in the light of the presently available models for Fd-mediated electron transfer in Δ9D and other protein-protein complexes. PMID:16605252

  10. Preparation and preclinical evaluation of experimental group B streptococcus type III polysaccharide-cholera toxin B subunit conjugate vaccine for intranasal immunization.

    PubMed

    Shen, X; Lagergård, T; Yang, Y; Lindblad, M; Fredriksson, M; Holmgren, J

    2000-11-22

    Streptococcus group B (GBS) is usually carried asymptomatically in the vaginal tract of women and can be transferred to the newborn during parturition. Serum antibodies to the capsular polysaccharide (CPS) can prevent invasive diseases, whereas immunity acting at the mucosal surface may be more important to inhibit the mucosal colonization of GBS and thus the risk of infection for the newborn. We prepared different GBS type III CPS-protein conjugate vaccines and evaluated their systemic and mucosal immunogenicity in mice. GBS type III CPS was conjugated to tetanus toxoid (TT) or recombinant cholera toxin B subunit (rCTB) either directly or to rCTB indirectly via TT. The conjugation was performed by different methods: (1) CPS was coupled to TT with 1-ethyl-3 (3-dimethylaminopropyl)-carbodiimide (EDAC), using adipic acid dihydrazide (ADH) as a spacer; (2) CPS was conjugated with rCTB using reductive amination; or, (3) N-succinimidyl 3-(2-pyridyldithio) propionate (SPDP) was used to bind rCTB to the TT of the CPS-TT conjugate. Mice were immunized with these conjugates or purified CPS by subcutaneous (s.c.) and intranasal (i. n.) routes. Antibodies to GBS III in serum, lungs and vagina were measured with ELISA. All of the CPS-protein conjugates were superior to unconjugated CPS in eliciting CPS-specific immune responses in serum and mucosal tissue extracts. The conjugates, when administrated s.c., induced only IgG responses in serum, lung and vagina, while i.n. vaccination also elicited IgA responses in the lungs and vagina. The CPS-TT conjugate administrated i.n. induced a strong serum IgG, but only a weak mucosal IgA response, while the CPS-rCTB conjugate elicited high IgG as well as IgA antibodies in the lungs after i.n. immunization. GBS III CPS-TT conjugated with rCTB produced a strong systemic and local anti-CPSIII response after i.n. administration. Co-administration of CT as adjuvant enhanced the anti-CPS systemic and mucosal immune responses further after i

  11. Development of antibody directed nanoparticles for cancer therapy

    NASA Astrophysics Data System (ADS)

    Ivkov, R.; DeNardo, S. J.; Meirs, L. A.; Natarajan, A.; DeNardo, G. L.; Gruettner, C.; Foreman, A. R.

    2007-02-01

    The pharmacokinetics, tumor uptake, and biologic effects of inductively heating 111In-chimeric L6 (ChL6) monoclonal antibody (mAb)-linked iron oxide nanoparticle (bioprobes) by externally applied alternating magnetic fields (AMF) were studied in athymic mice bearing human breast cancer HBT 3477 xenografts. In addition, response was correlated with calculated total deposited heat dose. Methods: Using 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide HCl, 111In-7,10-tetraazacyclododecane-N, N',N'',N'''-tetraacetic acid-ChL6 was conjugated to the carboxylated polyethylene glycol on dextran-coated iron oxide 20-nm particles, one to two mAbs per nanoparticle. After magnetic purification and sterile filtration, pharmacokinetics, histopathology, and AMF/bioprobe therapy were done using 111In-ChL6 bioprobe doses (20 mcg/2.2 mg ChL6/ bioprobe), i.v. with 50 mcg ChL6 in athymic mice bearing HBT 3477; a 153 kHz AMF was given 72 hours postinjection for therapy with amplitudes of 1,300, 1,000, or 700 Oe. Weights, blood counts, and tumor size were monitored and compared with control mice receiving nothing, or AMF, or bioprobes alone. Results: 111In-ChL6 bioprobe binding in vitro to HBT 3477 cells was 50% to 70% of that of 111In-ChL6. At 48 hours, tumor, lung, kidney, and marrow uptakes of the 111In-ChL6 bioprobes were not different from that observed in prior studies of 111In-ChL6. Significant therapeutic responses from AMF/bioprobe therapy were shown compared with no treatment. In addition, greatest therapeutic benefit was observed for the 700 Oe treatment cohort. Toxicity was only seen in the 1,300 Oe AMF cohort, with 4 of 12 immediate deaths associated with skin erythema and petechiae. Conclusion: This study shows that mAb-conjugated nanoparticles (bioprobes), when given i.v., escape into the extravascular space and bind to cancer cell membrane antigen.Thus, bioprobes can be used in concert with externally applied AMF to deliver thermoablative cancer therapy. Therapeutic benefit

  12. Improved drug targeting of cancer cells by utilizing actively targetable folic acid-conjugated albumin nanospheres.

    PubMed

    Shen, Zheyu; Li, Yan; Kohama, Kazuhiro; Oneill, Brian; Bi, Jingxiu

    2011-01-01

    Folic acid-conjugated albumin nanospheres (FA-AN) have been developed to provide an actively targetable drug delivery system for improved drug targeting of cancer cells with reduced side effects. The nanospheres were prepared by conjugating folic acid onto the surface of albumin nanospheres using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDAC) as a catalyst. To test the efficacy of these nanospheres as a potential delivery platform, doxorubicin-loaded albumin nanospheres (DOX-AN) and doxorubicin-loaded FA-AN (FA-DOX-AN) were prepared by entrapping DOX (an anthracycline, antibiotic drug widely used in cancer chemotherapy that works by intercalating DNA) into AN and FA-AN nanoparticles. Cell uptake of the DOX was then measured. The results show that FA-AN was incorporated into HeLa cells (tumor cells) only after 2.0h incubation, whereas HeLa cells failed to incorporate albumin nanospheres without conjugated folic acid after 4.0h incubation. When HeLa cells were treated with the DOX-AN, FA-DOX-AN nanoparticles or free DOX, cell viability decreased with increasing culture time (i.e. cell death increases with time) over a 70h period. Cell viability was always the lowest for free DOX followed by FA-DOX-AN4 and then DOX-AN. In a second set of experiments, HeLa cells washed to remove excess DOX after an initial incubation for 2h were incubated for 70h. The corresponding cell viability was slightly higher when the cells were treated with FA-DOX-AN or free DOX whilst cells treated with DOX-AN nanoparticles remained viable. The above experiments were repeated for non-cancerous, aortic smooth muscle cells (AoSMC). As expected, cell viability of the HeLa cells (with FA receptor alpha, FRα) and AoSMC cells (without FRα) decreased rapidly with time in the presence of free DOX, but treatment with FA-DOX-AN resulted in selective killing of the tumor cells. These results indicated that FA-AN may be used as a promising actively targetable drug delivery system to improve drug

  13. Integrated testing of an electrochemical depolarized CO2 concentrator /EDC/ and a Bosch CO2 reduction subsystem /BRS/. [in spaceborne oxygen reclamation system

    NASA Technical Reports Server (NTRS)

    Schubert, F. H.; Clark, D. C.; Quattrone, P. D.

    1976-01-01

    An oxygen reclamation system (ORS) in a spacecraft has the task to revitalize the spacecraft atmosphere by recovering the elementary oxygen from metabolically produced carbon dioxide and water vapor. Life support subsystems which can form such an ORS are an electrochemical depolarized carbon dioxide concentrator (EDC), a Bosch carbon dioxide reduction subsystem (BRS), and an oxygen generation subsystem (OGS). A total recovery of the oxygen from metabolically generated carbon dioxide can be obtained with the aid of system composed of the considered three subsystems. Attention is given to the control concept which assures an integrated operation of the EDC, BRS, and OGS. A description is presented of the test results obtained during 86 days of testing.

  14. Melatonin-Mediated Intracellular Insulin during 2-Deoxy-d-glucose Treatment Is Reduced through Autophagy and EDC3 Protein in Insulinoma INS-1E Cells

    PubMed Central

    Kim, Han Sung; Han, Tae-Young

    2016-01-01

    2-DG triggers glucose deprivation without altering other nutrients or metabolic pathways and then activates autophagy via activation of AMPK and endoplasmic reticulum (ER) stress. We investigated whether 2-DG reduced intracellular insulin increased by melatonin via autophagy/EDC3 in insulinoma INS-1E cells. p-AMPK and GRP78/BiP level were significantly increased by 2-DG in the presence/absence of melatonin, but IRE1α level was reduced in 2-DG treatment. Levels of p85α, p110, p-Akt (Ser473, Thr308), and p-mTOR (Ser2481) were also significantly reduced by 2-DG in the presence/absence of melatonin. Mn-SOD increased with 2-DG plus melatonin compared to groups treated with/without melatonin alone. Bcl-2 was decreased and Bax increased with 2-DG plus melatonin. LC3II level increased with 2-DG treatment in the presence/absence of melatonin. Intracellular insulin production increased in melatonin plus 2-DG but reduced in treatment with 2-DG with/without melatonin. EDC3 was increased by 2-DG in the presence/absence of melatonin. Rapamycin, an mTOR inhibitor, increased GRP78/BiP and EDC3 levels in a dose-dependent manner and subsequently resulted in a decrease in intracellular production of insulin. These results suggest that melatonin-mediated insulin synthesis during 2-DG treatment involves autophagy and EDC3 protein in rat insulinoma INS-1E cells and subsequently results in a decrease in intracellular production of insulin. PMID:27493704

  15. Removal of selected endocrine disrupting chemicals (EDCs) and pharmaceuticals and personal care products (PPCPs) during ferrate(VI) treatment of secondary wastewater effluents.

    PubMed

    Yang, Bin; Ying, Guang-Guo; Zhao, Jian-Liang; Liu, Shan; Zhou, Li-Jun; Chen, Feng

    2012-05-01

    We investigated the removal efficiencies of 68 selected endocrine disrupting chemicals (EDCs) and pharmaceuticals and personal care products (PPCPs) spiked in a wastewater matrix by ferrate (Fe(VI)) and further evaluated the degradation of these micropollutants present in secondary effluents of two wastewater treatment plants (WWTPs) by applying Fe(VI) treatment technology. Fe(VI)treatment resulted in selective oxidation of electron-rich organic moieties of these target compounds, such as phenol, olefin, amine and aniline moieties. But Fe(VI) failed to react with triclocarban, 3 androgens, 7 acidic pharmaceuticals, 2 neutral pharmaceuticals and erythromycin-H(2)O.Thirty-one target EDCs and PPCPs were detected in the effluents of the two WWTPs with concentrations ranging from 0.2 ± 0.1 ng L(-1) to 1156 ± 182 ng L(-1).Fe(VI) treatment resulted in further elimination of the detected EDCs and PPCPs during Fe(VI) treatment of the secondary wastewater effluents. The results from this study clearly demonstrated the effectiveness of Fe(VI) treatment as a tertiary treatment technology for a broad spectrum of micropollutants in wastewater. PMID:22342241

  16. EDCs, estrogenicity and genotoxicity reduction in a mixed (domestic + textile) secondary effluent by means of ozonation: a full-scale experience.

    PubMed

    Bertanza, G; Papa, M; Pedrazzani, R; Repice, C; Mazzoleni, G; Steimberg, N; Feretti, D; Ceretti, E; Zerbini, I

    2013-08-01

    WWTP (wastewater treatment plant) effluents are considered to be a major source for the release in the aquatic environment of EDCs (Endocrine-Disrupting Compounds), a group of anthropogenic substances able to alter the normal function of the endocrine system. The application of conventional processes (e.g. activated sludge with biological nitrogen removal) does not provide complete elimination of all these micropollutants and, consequently, an advanced treatment should be implemented. This experimental work was conducted on the tertiary ozonation stage of a 140,000 p.e. activated sludge WWTP, treating a mixed domestic and textile wastewater: an integrated monitoring, including both chemical (nonylphenol, together with the parent compounds mono- and di-ethoxylated, and bisphenol A were chosen as model EDCs) and biological (estrogenic and genotoxic activities) analyses, was carried out. Removal efficiencies of measured EDCs varied from 20% to 70%, depending on flow conditions (ozone dosage being 0.5 gO3/gTOC). Biological tests, furthermore, displayed that the oxidation stage did not significantly reduce (only by 20%) the estrogenicity of the effluent and revealed the presence and/or formation of genotoxic compounds. These results highlight the importance of the application of an integrated (biological+chemical) analytical procedure for a global evaluation of treatment suitability; poor performances recorded in this study have been attributed to the presence of a significant industrial component in the influent wastewater. PMID:23648445

  17. Mutations in DCPS and EDC3 in autosomal recessive intellectual disability indicate a crucial role for mRNA decapping in neurodevelopment

    PubMed Central

    Ahmed, Iltaf; Buchert, Rebecca; Zhou, Mi; Jiao, Xinfu; Mittal, Kirti; Sheikh, Taimoor I.; Scheller, Ute; Vasli, Nasim; Rafiq, Muhammad Arshad; Brohi, M. Qasim; Mikhailov, Anna; Ayaz, Muhammad; Bhatti, Attya; Sticht, Heinrich; Nasr, Tanveer; Carter, Melissa T.; Uebe, Steffen; Reis, André; Ayub, Muhammad; John, Peter; Kiledjian, Megerditch; Vincent, John B.; Jamra, Rami Abou

    2015-01-01

    There are two known mRNA degradation pathways, 3′ to 5′ and 5′ to 3′. We identified likely pathogenic variants in two genes involved in these two pathways in individuals with intellectual disability. In a large family with multiple branches, we identified biallelic variants in DCPS in three affected individuals; a splice site variant (c.636+1G>A) that results in an in-frame insertion of 45 nucleotides and a missense variant (c.947C>T; p.Thr316Met). DCPS decaps the cap structure generated by 3′ to 5′ exonucleolytic degradation of mRNA. In vitro decapping assays showed an ablation of decapping function for both variants in DCPS. In another family, we identified a homozygous mutation (c.161T>C; p.Phe54Ser) in EDC3 in two affected children. EDC3 stimulates DCP2, which decaps mRNAs at the beginning of the 5′ to 3′ degradation pathway. In vitro decapping assays showed that altered EDC3 is unable to enhance DCP2 decapping at low concentrations and even inhibits DCP2 decapping at high concentration. We show that individuals with biallelic mutations in these genes of seemingly central functions are viable and that these possibly lead to impairment of neurological functions linking mRNA decapping to normal cognition. Our results further affirm an emerging theme linking aberrant mRNA metabolism to neurological defects. PMID:25701870

  18. Cloning of circadian rhythmic pathway genes and perturbation of oscillation patterns in endocrine disrupting chemicals (EDCs)-exposed mangrove killifish Kryptolebias marmoratus.

    PubMed

    Rhee, Jae-Sung; Kim, Bo-Mi; Lee, Bo-Young; Hwang, Un-Ki; Lee, Yong Sung; Lee, Jae-Seong

    2014-08-01

    To investigate the effect of endocrine disrupting chemicals (EDCs) on the circadian rhythm pathway, we cloned clock and circadian rhythmic pathway-associated genes (e.g. Per2, Cry1, Cry2, and BMAL1) in the self-fertilizing mangrove killifish Kryptolebias marmoratus. The promoter region of Km-clock had 1 aryl hydrocarbon receptor element (AhRE, GTGCGTGACA) and 8 estrogen receptor (ER) half-sites, indicating that the AhRE and ER half sites would likely be associated with regulation of clock protein activity during EDCs-induced cellular stress. The Km-clock protein domains (bHLH, PAS1, PAS2) were highly conserved in five additional fish species (zebrafish, Japanese medaka, Southern platyfish, Nile tilapia, and spotted green pufferfish), suggesting that the fish clock protein may play an important role in controlling endogenous circadian rhythms. The promoter regions of Km-BMAL1, -Cry1, -Cry2, and -Per2 were found to contain several xenobiotic response elements (XREs), indicating that EDCs may be able to alter the expression of these genes. To analyze the endogenous circadian rhythm in K. marmoratus, we measured expression of Km-clock and other circadian rhythmic genes (e.g. Per2, Cry1, Cry2, and BMAL1) in different tissues, and found ubiquitous expression, although there were different patterns of transcript amplification during different developmental stages. In an estrogen (E2)-exposed group, Km-clock expression was down-regulated, however, a hydroxytamoxifen (TMX, nonsteroid estrogen antagonist)-exposed group showed an upregulated pattern of Km-clock expression, suggesting that the expression of Km-clock is closely associated with exposure to EDCs. In response to the exposure of bisphenol A (BPA) and 4-tert-octyphenol (OP), Km-clock expression was down-regulated in the pituitary/brain, muscle, and skin in both gender types (hermaphrodite and secondary male). In juvenile K. marmoratus liver tissue, expression of Km-clock and other circadian rhythmic pathway

  19. Rapid resolution liquid chromatography-tandem mass spectrometry method for the determination of endocrine disrupting chemicals (EDCs), pharmaceuticals and personal care products (PPCPs) in wastewater irrigated soils.

    PubMed

    Chen, Feng; Ying, Guang-Guo; Yang, Ji-Feng; Zhao, Jian-Liang; Wang, Li

    2010-10-01

    A multiresidue analytical method was developed for the determination of 9 endocrine disrupting chemicals (EDCs) and 19 pharmaceuticals and personal care products (PPCPs) including acidic and neutral pharmaceuticals in water and soil samples using rapid resolution liquid chromatography-tandem mass spectrometry (RRLC-MS/MS). Solid phase extraction (SPE), and ultrasonic extraction combined with silica gel purification were applied as pretreatment methods for water and soil samples, respectively. The extracts of the EDCs and PPCPs in water and soil samples were then analyzed by RRLC-MS/MS in electrospray ionization (ESI) mode in three independent runs. The chromatographic mobile phases consisted of Milli-Q water and acetonitrile for EDCs and neutral pharmaceuticals, and Milli-Q water containing 0.01 % acetic acid (v/v) and acetonitrile: methanol (1:1, v/v) for acidic pharmaceuticals at a flow rate of 0.3 mL/min. Most of the target compounds exhibited signal suppression due to matrix effects. Measures taken to reduce matrix effects included use of isotope-labeled internal standards, and application of matrix-match calibration curves in the RRLC-MS/MS analyses. The limits of quantitation ranged between 0.15 and 14.08 ng/L for water samples and between 0.06 and 10.64 ng/g for solid samples. The recoveries for the target analytes ranged from 62 to 208 % in water samples and 43 to 177 % in solid samples, with majority of the target compounds having recoveries ranging between 70–120 %. Precision, expressed as the relative standard deviation (RSD), was obtained less than 7.6 and 20.5 % for repeatability and reproducibility, respectively. The established method was successfully applied to the water and soil samples from four irrigated plots in Guangzhou. Six compounds namely bisphenol-A, 4-nonylphenol, triclosan, triclocarban, salicylic acid and clofibric acid were detected in the soils. PMID:20818522

  20. Association screening in the Epidermal Differentiation Complex (EDC) identifies an SPRR3 repeat number variant as a risk factor for eczema.

    PubMed

    Marenholz, Ingo; Rivera, Vladimir A Gimenez; Esparza-Gordillo, Jorge; Bauerfeind, Anja; Lee-Kirsch, Min-Ae; Ciechanowicz, Andrzej; Kurek, Michael; Piskackova, Tereza; Macek, Milan; Lee, Young-Ae

    2011-08-01

    The genetically determined impairment of the skin barrier is a primary cause of eczema. As numerous genes essential for an intact epidermis reside within the epidermal differentiation complex (EDC), we screened the National Center for Biotechnology Information (NCBI) database for putatively functional polymorphisms in the EDC genes and tested them for association with eczema. We identified 20 polymorphisms with predicted major impact on protein function. Of these, 4 were validated in 94 eczema patients: a nonsense mutation in FLG2 (rs12568784), a stop codon mutation in LCE1D (rs41268500), a 24-bp deletion in SPRR3 (rs28989168), and a frameshift mutation in S100A3 (rs11390146). The minor allele frequencies were 15.1, 6.1, 47.2, and 0.4%, respectively. Association testing of the validated polymorphisms in 555 eczema patients and 375 controls identified a significant effect of rs28989168 (SPRR3) on eczema. The association was replicated in another 1,314 cases and 1,322 controls, yielding an overall odds ratio of 1.30 (95% confidence interval 1.12-1.51; P=0.00067) for a dominant mode of inheritance. Small proline-rich proteins (SPRRs) are crossbridging proteins in the cornified cell envelope (CE), which provides the main barrier function of stratified squamous epithelia. The SPRR3 variant associated with eczema carried an extra 24-bp repeat in the central domain, which may alter the physical properties of the CE. PMID:21490620

  1. Competing risks and the development of adaptive management plans for water resources: Field reconnaissance investigation of risks to fishes and other aquatic biota exposed to endocrine disrupting chemicals (edcs) in lake mead, Nevada USA

    USGS Publications Warehouse

    Linder, G.; Little, E.E.

    2009-01-01

    The analysis and characterization of competing risks for water resources rely on a wide spectrum of tools to evaluate hazards and risks associated with their management. For example, waters of the lower Colorado River stored in reservoirs such as Lake Mead present a wide range of competing risks related to water quantity and water quality. These risks are often interdependent and complicated by competing uses of source waters for sustaining biological resources and for supporting a range of agricultural, municipal, recreational, and industrial uses. USGS is currently conducting a series of interdisciplinary case-studies on water quality of Lake Mead and its source waters. In this case-study we examine selected constituents potentially entering the Lake Mead system, particularly endocrine disrupting chemicals (EDCs). Worldwide, a number of environmental EDCs have been identified that affect reproduction, development, and adaptive behaviors in a wide range of organisms. Many EDCs are minimally affected by current treatment technologies and occur in treated sewage effluents. Several EDCs have been detected in Lake Mead, and several substances have been identified that are of concern because of potential impacts to the aquatic biota, including the sport fishery of Lake Mead and endangered razorback suckers (Xyrauchen texanus) that occur in the Colorado River system. For example, altered biomarkers relevant to reproduction and thyroid function in fishes have been observed and may be predictive of impaired metabolism and development. Few studies, however, have addressed whether such EDC-induced responses observed in the field have an ecologically significant effect on the reproductive success of fishes. To identify potential linkages between EDCs and species of management concern, the risk analysis and characterization in this reconnaissance study focused on effects (and attendant uncertainties) that might be expressed by exposed populations. In addition, risk reduction

  2. PROVIDING A BETTER UNDERSTANDING OF THE SCIENCE UNDERLYING THE EFFECTS, EXPOSURE, ASSESSMENT, AND MANAGEMENT OF EDCS: DOES MILD HYPOTHYROIDISM INDUCED BY ENVIRONMENTAL CONTAMINANTS IRREVERSIBLY ALTER CNS FUNCTION IN THE JUVENILE AND ADULT ANIMAL?

    EPA Science Inventory

    SUMMARY: The NTD research project on Endocrine-Disrupting Chemicals (EDC) is focused on the effects of thyroid hormone (TH) deficiencies on the developing brain and is one component of a larger NHEERL research program evaluating androgen, estrogen, and thyroid-disrupting chemical...

  3. The solid state structure and reactivity of NbCl(5) x (N,N'-dicyclohexylurea) in solution: evidence for co-ordinated urea dehydration to the relevant carbodiimide.

    PubMed

    Aresta, Michele; Dibenedetto, Angela; Stufano, Paolo; Aresta, Brunella Maria; Maggi, Sabino; Pápai, Imre; Rokob, Tibor András; Gabriele, Bartolo

    2010-08-14

    NbCl(5) x (N,N'-dicyclohexylurea) 1a owns a distorted octahedral structure due to intramolecular NH...Cl bonding. The unit cell contains four units which are intermolecularly NH...Cl and NH...N bonded. An extended intramolecular network of H-bonding (N-H...Cl, CH...Cl, CH...N) causes the 3D self assembling of the units. Upon addition of base, the HCl release from 1a is observed with the transfer to Nb of the O-atom of the carbonylic function of the starting urea which is converted into the relevant carbodiimide CyN=C=NCy 4. The latter is quantitatively released by adding an excess of NEt(3) at 308 K (py and DBU are less efficient) with formation of the known NbOCl(3)(NEt(3))(2), isolated in quantitative yield. Increasing the temperature leads to a loss in selectivity as the formed DCC undergoes further reactions. At 350 K, the isocyanate CyN=C=O has been isolated in 60% yield besides a mixture of Nb-complexes. DFT calculations have been coupled to IR and NMR experiments for characterizing possible reaction intermediates and the behaviour of 1a. Several other MCl(x) species (ScCl(3), YCl(3), LaCl(3), TiCl(4), TaCl(5), AlCl(3), SnCl(4)) have been shown to be able to co-ordinate DCU but not all of them promote the conversion of urea into DCC. PMID:20544121

  4. Reproductive Toxicology Testing with EDCS

    EPA Science Inventory

    An introduction to reproductive toxicology: the basic approaches to testing chemicals for adverse effects using multigenerational studies with rats and how the regulatory agencies used the data in risk assessments. Case studies were presented of how endocrine or genomic data were...

  5. Chains of [RE6] octahedra coupled by (NCN) links in the network structure of RE2Cl(CN2)N. Synthesis and structure of two novel rare earth chloride carbodiimide nitrides with structures related to the RE2Cl3 type.

    PubMed

    Srinivasan, Radhakrishnan; Ströbele, Markus; Meyer, H-Jürgen

    2003-06-01

    Solid state metathesis reactions between RECl(3) (RE = La, Ce) and Li(2)CN(2) at 800 degrees C have led to the discovery of the rare earth chloride carbodiimide nitrides La(2)Cl(CN(2))N (1) and Ce(2)Cl(CN(2))N (2), respectively. Single crystal X-ray diffraction analyses revealed that 1 and 2 crystallize isotypic in an orthorhombic system (Cmmm, Z = 4, a = 13.3914(8) A, b = 9.6345(7) A, c = 3.9568(2) A for 1 and a = 13.340(1) A, b = 9.5267(8) A, c = 3.9402(5) A for 2). The crystal structures of 1 and 2 contain linear chains of edge-sharing octahedra built from rare earth metal atoms. Similar to [M(6)X(8)] type clusters, the [RE(6)] octahedra are capped by eight nitrogen atoms above their faces, of which four are from N(3-) ions and the other four are from (CN(2))(2-) ions. The chains are interconnected by bridging (CN(2))(2-) to form a three-dimensional network with Cl(-) ions in linear channels. Compounds 1 and 2 are surprisingly stable toward air and water. They have been characterized by thermal analysis, infrared spectroscopy, and magnetic susceptibility studies. PMID:12767174

  6. Synthesis and mass-spectrometric characterization of human serum albumins modified by covalent binding of two non-steroidal anti-inflammatory drugs: tolmetin and zomepirac.

    PubMed Central

    Zia-Amirhosseini, P; Ding, A; Burlingame, A L; McDonagh, A F; Benet, L Z

    1995-01-01

    Human serum albumins modified by covalently bound tolmetin or zomepirac were synthesized as models for similar products formed in vivo from acyl glucuronides. Activated esters of both drugs were prepared with 1-ethyl-3-(3-dimethylaminopropyl)-carbodi-imide, and then allowed to react with human serum albumin. Tryptic digests of both protein products were analysed by HPLC to identify peptides containing covalently bound drugs, and binding sites on albumin were identified by high-performance tandem MS. Three binding sites were common to both products, i.e. lysine-195, -199 and -351. Three further modified residues were identified for the tolmetin-albumin product, i.e. aspartic acid 1, and lysine-524 and -536. PMID:7487878

  7. Molecular Endpoints and Mixtures of EDCs in Fish

    EPA Science Inventory

    Microarray technology is a relatively novel tool in ecotoxicology and is beginning to be used for exposure and/or hazard characterization for ecological risk assessment. To develop a basis for this type of analysis, fathead minnows (Pimephales promelas) were treated with two bin...

  8. NATIONAL SCREENING SURVEY OF EDCS IN MUNICIPAL WASTEWATER TREATMENT FACILITIES

    EPA Science Inventory

    In 2002 and 2003 the USEPA's Office of Research and Development asked Regional EPA inspectors, state EPA inspectors and municipal plant operators to collect four gallons effluent, either as a grab or composite sample, from up to 50 wastewater treatment plants (WWTP), and ship the...

  9. Amine coupling through EDC/NHS: a practical approach.

    PubMed

    Fischer, Marcel J E

    2010-01-01

    Surface plasmon resonance (SPR) is one of the leading tools in biomedical research. The challenge in its use is the controlled positioning of one of the components of an interaction on a carefully designed surface. Many attempts in interaction analysis fail due to the non-functional or unsuccessful immobilization of a reactant onto the complex matrix of that surface. The most common technique for linking ligands covalently to a hydrophilic solid surface is amine coupling via reactive esters. In this chapter detailed methods and problem discussions will be given to assist in fast decision analysis to optimize immobilization and regeneration. Topics in focus are different coupling techniques for small and large molecules, streptavidin-biotin sandwich immobilization, and optimizing regeneration conditions. PMID:20217613

  10. Luteinizing hormone-releasing hormone targeted poly(methyl vinyl ether maleic acid) nanoparticles for doxorubicin delivery to MCF-7 breast cancer cells.

    PubMed

    Varshosaz, Jaleh; Jahanian-Najafabadi, Ali; Ghazzavi, Jila

    2016-08-01

    The purpose of this study was to design a targeted anti-cancer drug delivery system for breast cancer. Therefore, doxorubicin (DOX) loaded poly(methyl vinyl ether maleic acid) nanoparticles (NPs) were prepared by ionic cross-linking method using Zn(2+) ions. To optimise the effect of DOX/polymer ratio, Zn/polymer ratio, and stirrer rate a full factorial design was used and their effects on particle size, zeta potential, loading efficiency (LE, %), and release efficiency in 72 h (RE72, %) were studied. Targeted NPs were prepared by chemical coating of tiptorelin/polyallylamin conjugate on the surface of NPs by using 1-ethyl-3-(3-dimethylaminopropyl) carboiimid HCl as cross-linking agent. Conjugation efficiency was measured by Bradford assay. Conjugated triptorelin and targeted NPs were studied by Fourier-transform infrared spectroscopy (FTIR). The cytotoxicity of DOX loaded in targeted NPs and non-targeted ones were studied on MCF-7 cells which overexpress luteinizing hormone-releasing hormone (LHRH) receptors and SKOV3 cells as negative LHRH receptors using Thiazolyl blue tetrazolium bromide assay. The best results obtained from NPs prepared by DOX/polymer ratio of 5%, Zn/polymer ratio of 50%, and stirrer rate of 960 rpm. FTIR spectrum confirmed successful conjugation of triptorelin to NPs. The conjugation efficiency was about 70%. The targeted NPs showed significantly less IC50 for MCF-7 cells compared to free DOX and non-targeted NPs. PMID:27463791

  11. The conformational state of polyphenol oxidase from field bean (Dolichos lablab) upon SDS and acid-pH activation.

    PubMed

    Kanade, Santosh R; Paul, Beena; Rao, A G Appu; Gowda, Lalitha R

    2006-05-01

    Field bean (Dolichos lablab) contains a single isoform of PPO (polyphenol oxidase)--a type III copper protein that catalyses the o-hydroxylation of monophenols and oxidation of o-diphenols using molecular oxygen--and is a homotetramer with a molecular mass of 120 kDa. The enzyme is activated manyfold either in the presence of the anionic detergent SDS below its critical micellar concentration or on exposure to acid-pH. The enhancement of kcat upon activation is accompanied by a marked shift in the pH optimum for the oxidation of t-butyl catechol from 4.5 to 6.0, an increased sensitivity to tropolone, altered susceptibility to proteolytic degradation and decreased thermostability. The Stokes radius of the native enzyme is found to increase from 49.1+/-2 to 75.9+/-0.6 A (1 A=0.1 nm). The activation by SDS and acid-pH results in a localized conformational change that is anchored around the catalytic site of PPO that alters the microenvironment of an essential glutamic residue. Chemical modification of field bean and sweet potato PPO with 1-ethyl-3-(3-dimethylaminopropyl)carbodi-imide followed by kinetic analysis leads to the conclusion that both the enzymes possess a core carboxylate essential to activity. This enhanced catalytic efficiency of PPO, considered as an inducible defence oxidative enzyme, is vital to the physiological defence strategy adapted by plants to insect herbivory and pathogen attack. PMID:16393141

  12. Electronic and magnetic structure of transition-metal carbodiimides by means of GGA+U theory.

    PubMed

    Xiang, Hongping; Dronskowski, Richard; Eck, Bernhard; Tchougréeff, Andrei L

    2010-11-25

    The electronic structures and magnetic properties of MNCN (M = Fe, Co, and Ni) have been investigated by density-functional theory including explicit electronic correlation through an ad hoc Coulomb potential (GGA+U). The results evidence CoNCN and NiNCN as type-II anti-ferromagnetic semiconductors (that is, intralayer ferromagnetic and interlayer anti-ferromagnetic), in accordance with experimental observations. Just like the prototype MnNCN, the MNCN phases, with M = Ni and Co, thus resemble the corresponding MO monoxides with respect to their magnetic and transport properties. By contrast, FeNCN remains (semi)metallic even upon applying a strong Coulomb correlation potential. This, most probably, is in contradiction with its observed optical transparency and expected insulating behavior and points toward a serious density-functional theory problem. PMID:21038908

  13. The conformational state of polyphenol oxidase from field bean (Dolichos lablab) upon SDS and acid-pH activation

    PubMed Central

    Kanade, Santosh R.; Paul, Beena; Rao, A. G. Appu; Gowda, Lalitha R.

    2006-01-01

    Field bean (Dolichos lablab) contains a single isoform of PPO (polyphenol oxidase) – a type III copper protein that catalyses the o-hydroxylation of monophenols and oxidation of o-diphenols using molecular oxygen – and is a homotetramer with a molecular mass of 120 kDa. The enzyme is activated manyfold either in the presence of the anionic detergent SDS below its critical micellar concentration or on exposure to acid-pH. The enhancement of kcat upon activation is accompanied by a marked shift in the pH optimum for the oxidation of t-butyl catechol from 4.5 to 6.0, an increased sensitivity to tropolone, altered susceptibility to proteolytic degradation and decreased thermostability. The Stokes radius of the native enzyme is found to increase from 49.1±2 to 75.9±0.6 Å (1 Å=0.1 nm). The activation by SDS and acid-pH results in a localized conformational change that is anchored around the catalytic site of PPO that alters the microenvironment of an essential glutamic residue. Chemical modification of field bean and sweet potato PPO with 1-ethyl-3-(3-dimethylaminopropyl)carbodi-imide followed by kinetic analysis leads to the conclusion that both the enzymes possess a core carboxylate essential to activity. This enhanced catalytic efficiency of PPO, considered as an inducible defence oxidative enzyme, is vital to the physiological defence strategy adapted by plants to insect herbivory and pathogen attack. PMID:16393141

  14. Modified fabrication process of protein chips using a short-chain self-assembled monolayer.

    PubMed

    Jang, Ling-Sheng; Keng, Hao-Kai

    2008-04-01

    In previous work a short chain SAM, 4,4-Dithiodibutyric Acid (DTBA) was found to be a thin monolayer in protein chips. However, obtaining uniform fluorescent intensity remains difficult because water-soluble carbodiimides (EDC) in an aqueous system cause the hydrolysis of N-hydroxysuccinimide ester (NHS esters). The hydrolysis of NHS esters reduces coupling yields and therefore reduces the fluorescent intensity of protein chips. The NHS can increase the stability of active intermediate resulting from the reaction of EDC and NHS, but the ratio of the concentration of EDC to that of NHS strongly affects this stability. The effects of the solvents used in the washing step are studied to solve this problem. The results reveal that PBST (PBS + 5% Tween20) is more effective in reducing the hydrolysis of NHS esters than deionized water. Additionally, the effects of 3:1 and 5:2 EDC/NHS ratios on the chips are examined. The 3:1 EDC/NHS ratio yields a higher fluorescent intensity than the 5:2 ratio. The effects on the chips of dissolving EDC in DI water, DI water + 0.1 M MES and alcohol are also investigated. The results show that alcohol provides higher fluorescent intensity than other solvents and the reaction time of 4 h yields a high fluorescent intensity with 3:1 EDC/NHS ratio. A modified fabrication process of protein chips using 4,4-DTBA is developed. In this work, 160 mM 4,4-DTBA is used as a self-assembled monolayer in the fabrication of protein chips. Experiments to characterize 4,4-DTBA are performed by contact angle goniometry and Fourier transform infrared spectroscopy (FTIR). Furthermore, the immobilized protein A-FITC (fluorescein isothiocyanate) is adopted in fluorescent assays. PMID:17849186

  15. Detection of magnetic-labeled antibody specific recognition events by combined atomic force and magnetic force microscopy

    NASA Astrophysics Data System (ADS)

    Hong, Xia; Liu, Yanmei; Li, Jun; Guo, Wei; Bai, Yubai

    2009-09-01

    Atomic force (AFM) and magnetic force microscopy (MFM) were developed to detect biomolecular specific interaction. Goat anti-mouse immunoglobulin (anti-IgG) was covalently attached onto gold substrate modified by a self-assembly monolayer of thioctic acid via 1-ethyl-3-[3-(dimethylamino) propyl] carbodiimide (EDC) activation. Magnetic-labeled IgG then specifically adsorbed onto anti-IgG surface. The morphological variation was identified by AFM. MFM was proved to be a fine assistant tool to distinguish the immunorecognized nanocomposites from the impurities by detection of the magnetic signal from magnetic-labeled IgG. It would enhance the understanding of biomolecular recognition process.

  16. Metabolomics in Small Fish Toxicology: Assessing the Impacts of Model EDCs

    EPA Science Inventory

    Although lagging behind applications targeted to human endpoints, metabolomics offers great potential in environmental applications, including ecotoxicology. Indeed, the advantages of metabolomics (relative to other ‘omic techniques) may be more tangible in ecotoxicology because...

  17. USING BIOASSAYS TO EVALUATE THE PERFORMANCE OF EDC RISK MANAGEMENT METHODS

    EPA Science Inventory

    In Superfund risk management research, the performance of risk management techniques is typically evaluated by measuring "the concentrations of the chemicals of concern before and after risk management efforts. However, using bioassays and chemical data provides a more robust und...

  18. Curriculum Profiles: A Resource of the EDC K-12 Science Curriculum Dissemination Center

    ERIC Educational Resources Information Center

    Education Development Center, Inc, 2005

    2005-01-01

    The purpose of this document is to provide useful information for teachers and school systems engaged in the process of examining and choosing science curriculum materials appropriate for their settings. The curriculum profiles include summaries of selected programs available for K?12 science curriculum programs. Each profile describes a number of…

  19. PREDICTING ER BINDING AFFINITY FOR EDC RANKING AND PRIORITIZATION: MODEL II

    EPA Science Inventory

    The training set used to derive a common reactivity pattern (COREPA) model for estrogen receptor (ER) binding affinity in Model I (see Abstract I in this series) was extended to include 47 rat estrogen receptor (rER) relative binding affinity (RBA) measurements in addition to the...

  20. APPROACHES TO EXTRAPOLATING EFFECTS OF EDCS ACROSS BIOLOGICAL LEVELS OF ORGANIZATION IN FISH

    EPA Science Inventory

    A challenge in ecological risk assessments is to obtain, in a resource-effective manner, information that provides insight both into chemical mode/mechanism of action (MOA) and adverse effects in individual animals, which are indicative of potential population-level responses. T...

  1. CONCENTRATED ANIMAL FEEDING OPERATIONS AS A SOURCE OF EDCS AND THEIR MANAGEMENT

    EPA Science Inventory

    In the United States, there is an estimated 376,000 animal feed operations, generating approximately 128 billion pounds of waste each year. A facility is an animal feed operation (AFO) if animals are stabled/confined, or fed/maintained, for 45 days or more within any 12-month per...

  2. An Evaluation of the EDC Role in the Bilingual Transitional Clusters of the Boston Public Schools.

    ERIC Educational Resources Information Center

    Cline, Marvin G.; Joyce, John F.

    The report documents the early stages and reflections of some of the initial staff of the Boston Bilingual Clusters program designed for public school instruction for Puerto Rican immigrant children. Conceived as a transitional educational experience, the program seeks to effect the diagnosis and remediation of school problems and unmet needs in…

  3. Cross-species Extrapolation of EDC Toxicity: Consequences for Screening Programs

    EPA Science Inventory

    Many structural and functional aspects of the vertebrate hypothalamic-pituitary-gonadal (HPG) axis are known to be highly conserved, but the full significance of this from a toxicological perspective has received comparatively little attention. High-quality data generated through...

  4. Reaction of protein and carbohydrates with EDC for purpose of making products with unique functional properties

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Prior research from this laboratory has demonstrated the feasibility of using chemical and enzymatic treatments on protein and carbohydrate waste products for the purpose of making fillers to enhance the properties of leather. These treatments (microbial transglutaminase, genipin, and polyphenols i...

  5. Alternatives to in vivo Tests to Detect Endocrine Disrupting Chemicals (EDCs) in Fish and Amphibians

    EPA Science Inventory

    A significant amount of current research in risk assessment of chemicals is targeted to evaluate alternative test methods that may reduce, replace or refine the use of animals, while ensuring human and environmental health and safety. In 2009, the US EPA began implementation of t...

  6. Diagnostic Assessment of the Ecological Risk of EDCs in Complex Mixtures

    EPA Science Inventory

    Although it is important to be able to forecast the potential endocrine toxicity of chemical mixtures that could enter aquatic environments, in many instances there is a need to determine possible effects of endocrine-active chemicals already present in complex environmental mixt...

  7. 3-D QSARS FOR RANKING AND PRIORITIZATION OF LARGE CHEMICAL DATASETS: AN EDC CASE STUDY

    EPA Science Inventory

    The COmmon REactivity Pattern (COREPA) approach is a three-dimensional structure activity (3-D QSAR) technique that permits identification and quantification of specific global and local steroelectronic characteristics associated with a chemical's biological activity. It goes bey...

  8. Improving the assessment of endocrine disrupting chemical (EDC) effects on puberty

    EPA Science Inventory

    During puberty, key developmental events occur that are critical for normal adult male and female reproductive maturation. Recent studies raised concern that exposure to environmental chemicals alter the normal progression through puberty and lead to impaired reproductive functio...

  9. Verification of ELISA Test Kits to Determine EDCs in Animal Feedlot Runoff

    EPA Science Inventory

    The key points discussed in this presentation are: (1) Climate change will very likely exacerbate the already stressed water supply in the U.S. Southeast, the only region that has experienced precipitation decline over >100 years; (2) The effect in precipitation change is locati...

  10. Reaction of protein and carbohydrates with EDC for making unique biomaterials

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Prior research from this laboratory has demonstrated the feasibility of using chemical and enzymatic treatments on protein and carbohydrate waste products for the purpose of making fillers to enhance the properties of leather. These treatments (microbial transglutaminase, genipin, and polyphenols i...

  11. Evaluation of estrogenic activity and measurement of EDCs in wastewater treatment plants

    NASA Astrophysics Data System (ADS)

    Lee, B. C.; Jung, J. Y.; Kim, H. K.

    2006-10-01

    Correlations between estrogenic activity and DOC/UV260 ratio in wastewater treatment processes were investigated to propose a simple, reliable and comprehensive indicator for the presence of estrogenic substances. Contrary to this, when short-term bioassays such as the E-SCREEN, receptor binding and reporter gene expression assays are used for detecting estrogenic activity in the wastewater sample, they require a long time, at least a few days. The major factors contributing to the estrogenic activity were found to be 17β-estradiol (E2) and estrone (El). A good relationship between the DOC/ UV260 ratio and the concentration of estrogens (El and E2) in the effluent of the activated sludge process was found: the E2 concentration increased as the DOC/UV260 ratio increased while the El concentration decreased. The relative estrogenic activity and DOC/UV260 ratio showed a good correlation (R2=0.84) for all sewage samples except the ozonized samples in the sewage treatment plants. This study shows that the estrogenic compounds are hard to be mineralized by the conventional biological processes. Advanced oxidation processes are required to further remove estrogenic substances in the secondary effluent. By analysis of DOC and UV260, the estrogenic activity in the wastewater can be rapidly estimated.

  12. Identification of Catalytic Amino Acid Residues by Chemical Modification in Dextranase.

    PubMed

    Ko, Jin-A; Nam, Seung-Hee; Kim, Doman; Lee, Jun-Ho; Kim, Young-Min

    2016-05-28

    A novel endodextranase isolated from Paenibacillus sp. was found to produce isomaltotetraose and small amounts of cycloisomaltooligosaccharides with a degree of polymerization of 7-14 from dextran. To determine the active site, the enzyme was modified with 1-ethyl-3-[3- (dimethylamino)-propyl]-carbodiimide (EDC) and α-epoxyalkyl α-glucosides (EAGs), an affinity labeling reagent. The inactivation followed pseudo first-order kinetics. Kinetic analysis and chemical modification using EDC and EAGs indicated that carboxyl groups are essential for the enzymatic activity. Three Asp and one Glu residues were identified as candidate catalytic amino acids, since these residues are completely conserved across the GH family of 66 enzymes. Replacement of Asp189, Asp340, or Glu412 completely abolished the enzyme activity, indicating that these residues are essential for catalytic activity. PMID:26907761

  13. Functionalization of poly(ε-caprolactone) surface with lactose-modified chitosan via alkaline hydrolysis: ToF-SIMS characterization.

    PubMed

    Tortora, Luca; Concolato, Sofia; Urbini, Marco; Giannitelli, Sara Maria; Basoli, Francesco; Rainer, Alberto; Trombetta, Marcella; Orsini, Monica; Mozetic, Pamela

    2016-06-01

    Functionalization of poly(ε-caprolactone) (PCL) was performed via hydrolysis and subsequent grafting of lactose-modified chitosan (chitlac) at two different degrees of derivatization (9% and 64%). Time of flight secondary ion mass spectrometry (ToF-SIMS) and multivariate analysis (principal component analysis) were successfully applied to the characterization of PCL surface chemistry, evidencing changes in the biopolymer surface following base-catalyzed hydrolysis treatment. ToF-SIMS analysis also confirmed positive EDC/NHS-catalyzed (EDC: N-ethyl-N'-(3-(dimethylamino)propyl)carbodiimide; NHS: N-hydroxysuccinimide) immobilization of chitlac onto activated PCL surface, with formation of amide bonds between PCL surface carboxyl groups and amine residues of chitlac. Yield of grafting reaction was also shown to be dependent upon the lactosilation degree of chitlac. PMID:26905217

  14. An electrochemical immunosensor based on covalent immobilization of okadaic acid onto screen printed carbon electrode via diazotization-coupling reaction.

    PubMed

    Hayat, Akhtar; Barthelmebs, Lise; Sassolas, Audrey; Marty, Jean-Louis

    2011-07-15

    In this work, an electrochemical method based on the diazonium-coupling reaction mechanism for the immobilization of okadaic acid (OA) on screen printed carbon electrode was developed. At first, 4-carboxyphenyl film was grafted by electrochemical reduction of 4-carboxyphenyl diazonium salt, followed by terminal carboxylic group activation by N-hydroxysuccinimide (NHS), N-(3-dimethylaminopropyle)-N'-ethyle-carbodiimide hydrochloride (EDC). Hexamethyldiamine was then covalently bound by one of its terminal amine group to the activated carboxylic group. The carboxyl group of okadaic acid was activated by EDC/NHS and then conjugated to the second terminal amine group on other side of the hexamethyldiamine through amide bond formation. After immobilization of OA, an indirect competitive immunoassay format was employed to detect OA. The immunosensor obtained using this novel approach allowed detection limit of 1.44 ng/L of OA, and was also validated with certified reference mussel samples. PMID:21645734

  15. Injectable hydrogel provides growth-permissive environment for human nucleus pulposus cells.

    PubMed

    Priyadarshani, Priyanka; Li, Yongchao; Yang, ShangYou; Yao, Li

    2016-02-01

    Degeneration of intervertebral discs (IVDs) results in an overall alteration of the biomechanics of the spinal column and becomes a major cause of low back pain. In this study, an injectable hydrogel composite is fabricated and characterized as a potential scaffold for the treatment of degenerated IVDs. Crosslinking of type II collagen-hyaluronic acid (HA) hydrogel with 1-ethyl-3(3-dimethyl aminopropyl) carbodiimide (EDC) increases the gel stability against collagenase digestion and reduces water uptake in comparison with non-crosslinked gel. Cell viability assay exhibits the proliferation of human nucleus pulposus (HNP) cells in hydrogels. The cells in non-crosslinked gel and the gel crosslinked with a low concentration of EDC (0.1 mM) show superior cell viability and morphology compared with cells in gels crosslinked with higher concentration of EDC. Quantitative PCR assay demonstrates the gene expression of extracellular matrix (ECM) by cells cultured in the gels. The expression of ECM genes by HNP cells in the gels demonstrated the phenotypic change of the cells. This study suggests that the type II collagen-HA hydrogel and crosslinked hydrogel (0.1 mM EDC) are permissive matrix for the growth of HNP cells and can be potentially applied in NP repair. PMID:26422588

  16. Covalent modification of the amine transporter with N,N prime -dicyclohexylcarbodiimide

    SciTech Connect

    Suchi, R.; Stern-Bach, Y.; Gabay, T.; Schuldiner, S. )

    1991-07-02

    N,N{prime}-Dicyclohexylcarbodiimide (DCC) has been previously shown to inhibit the amine transporter from chromaffin granules. A study of the mechanism of inhibition is present together with the demonstration of covalent modification of the protein. DCC inhibits binding of R1 (reserpine) and R2 (tetrabenazine) types of ligands to the transporter as well as transport. Lignads of the R2 type, but not those of the R1 type, protect against inhibition of all the reactions by DCC, i.e., accumulation of serotonin, binding if reserpine (R1 ligand), and binding of ketanserine (R2 ligand). The ability of a given R2 ligand to protect the transporter correlates well with its binding constant. Water-soluble carbodiimides, such as 1-ethyl-3-(3-(diethylamino)propyl)carbodiimide (EDC), do not have any effect on the catalytic activity of the transporter. A flourescent hydrophobic analogue of DCC, N-cyclohexyl-N{prime}-(4-(dimethylamino)-{alpha}-naphthyl)carbodiimide (NCD-4), inhibits at about the same concentration range as DCC. ({sup 14}C)DCC labels several polypeptides in the chromaffin granule membranes. Labeling of a polypeptide with an apparent M{sub r} of 80K is inhibited in the presence of R2 ligands. The labeled polypeptide copurifies with the recently identified and isolated transporter.

  17. The effects of decellularization and cross-linking techniques on the fatigue life and calcification of mitral valve chordae tendineae.

    PubMed

    Gunning, Gillian M; Murphy, Bruce P

    2016-04-01

    In cases of severely diseased mitral valves (MV), the required treatment is often valve replacement. Bioprosthetic and stentless replacement valves are usually either fully or partially composed of animal derived tissue treated with a decellularization process, a cross-linking process, or both. In this study, we analysed the effects of these treatments on the fatigue properties of porcine MV chordae tendineae (CT), as well as on the calcification of the CT using an in vitro technique. CT were tested in 4 groups; (1) native, (2) decellularized (DC), (3) decellularized and cross-linked with glutaraldehyde (DC-GTH), and (4) decellularized and cross-linked with 1-ehtyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)(DC-EDC). CT were tested in both uniaxial tension, and in fatigue at 10MPa peak stress (1Hz). The cycles to failure (mean±SD) for the four groups are as follows; Native- 53,397±55,798, DC- 28,013±30,634, DC-GTH- 97,665±133,556, DC-EDC- 318,601±322,358. DC-EDC CT were found to have a slightly longer fatigue life than the native and DC groups. The DC-EDC group also had a marginally lower dynamic creep rate, meaning those CT elongate more slowly. After in vitro calcification, X-ray microtomography was used to determine relative levels of calcification. The DC-EDC and DC-GTH groups had the lowest volume of calcific deposits. Under uniaxial testing, the ultimate tensile strength (UTS) of the DC-GTH CT was statistically significantly reduced after calcification, while the UTS was relatively unchanged for the DC-EDC group. Overall, these results indicate that a treatment of decellularization plus cross-linking with EDC may improve the fatigue life of porcine CT, reduce the rate of elongation, and help the CT resist the negative effects of calcification. This may be a preferable treatment in the preparation of porcine MVs for the replacement of diseased MVs. PMID:26875146

  18. Removal of phthalate esters from water using immobilized lipase on chitosan beads.

    PubMed

    Dulazi, Ashura A; Liu, Hui

    2011-10-01

    Lipase immobilized on chitosan beads was investigated as a possible efficient and cost-effective method of removing phthalate esters (PAEs) from water, under the hypothesis that the immobilized lipase could degrade PAEs and the amino group of chitosan could adsorb the degraded products. Three immobilization methods, namely crosslinking using glutaraldehyde (GLA), activation with 1-ethyl-3(3-dimethyl-aminopropyl) carbodiimide hydrochloride (EDC) and a binary method using both GLA and EDC, were compared. The results showed that lipase immobilized by crosslinking with GLA exhibited a higher degradation ability for PAEs with lower molecular weights. It removed 100% of dimethyl phthalate and 93.86% of diethyl phthalate. On the other hand, lipase immobilized by EDC activation exhibited efficient degradation of PAEs with higher molecular weights, such as diethylhexyl phthalate (removal efficiency 100%). However, lipase immobilized by the binary method unexpectedly showed less than 50% removal efficiency for all five PAEs. The PAE solutions treated with different immobilized lipases were characterized using high-performance liquid chromatography, and the mechanism of the removal of PAEs by the lipase immobilized on chitosan beads using different methods is further discussed. PMID:22329134

  19. Decoration of silk fibroin by click chemistry for biomedical application.

    PubMed

    Zhao, Hongshi; Heusler, Eva; Jones, Gabriel; Li, Linhao; Werner, Vera; Germershaus, Oliver; Ritzer, Jennifer; Luehmann, Tessa; Meinel, Lorenz

    2014-06-01

    Silkfibroin (SF) has an excellent biocompatibility and its remarkable structure translates into exciting mechanical properties rendering this biomaterial particularly fascinating for biomedical application. To further boost the material's biological/preclinical impact, SF is decorated with biologics, typically by carbodiimide/N-hydroxysuccinimide coupling (EDC/NHS). For biomedical application, this chemistry challenges the product risk profile due to the formation of covalent aggregates, particularly when decoration is with biologics occurring naturally in humans as these aggregates may prime for autoimmunity. Cu(I)-catalyzed azide-alkyne cycloaddition (CuAAC; click chemistry) provides the necessary specificity to avoid such intermolecular, covalent aggregates. We present a blueprint outlining the necessary chemistry rendering SF compatible with CuAAC and with a particular focus on structural consequences. For that, the number of SF carboxyl groups (carboxyl-SF; required for EDC/NHS chemistry) or azido groups (azido-SF; required for click chemistry) was tailored by means of diazonium coupling of the SF tyrosine residues. Structural impact on SF and decorated SF was characterized by Fourier transform infrared spectroscopy (FTIR). The click chemistry yielded a better controlled product as compared to the EDC/NHS chemistry with no formation of inter- and intramolecular crosslinks as demonstrated for SF decorated with fluorescent model compounds or a biologic, fibroblast growth factor 2 (FGF2), respectively. In conclusion, SF can readily be translated into a scaffold compatible with click chemistry yielding decorated products with a better risk profile for biomedical application. PMID:24576682

  20. EDC testing in the future: Exploring roles of pathway-based in silico, in vitro and in vivo methods

    EPA Science Inventory

    Many thoroughly validated, robust tests with both mammalian and non-mammalian models have been developed to identify chemicals with the potential to impact endocrine pathways associated with the hypothalamic-pituitary-gonadal (HPG) and thyroidal axes. In the US, for example, the...

  1. Induction of vitellogenin gene expression in adult male fathead minnows for select EDCs in 48-hour exposures

    EPA Science Inventory

    Endocrine disrupting chemicals have been shown to be present in surface waters, sediments and sludge, and are known to induce vitellogenin gene liver transcripts in male fathead minnows. The purpose of our study was to establish the lowest concentrations of estrogenic chemicals ...

  2. COMPARISON OF THE EFFECTS OF ENDOCRINE DISRUPTING CHEMICALS (EDCS) ON AROMATASE (CYP19) ACTIVITY IN RATS AND FISH

    EPA Science Inventory

    Aromatase, a product of the CYP19 gene, is an important enzyme during steroidogenesis that catalyzes the conversion of androstenedione and testosterone to estrone and estradiol. It is expressed in gonadal and extragonadal tissues in all vertebrates, and is critical for the homeos...

  3. Optimization of Surface Roughness Parameters of Al-6351 Alloy in EDC Process: A Taguchi Coupled Fuzzy Logic Approach

    NASA Astrophysics Data System (ADS)

    Kar, Siddhartha; Chakraborty, Sujoy; Dey, Vidyut; Ghosh, Subrata Kumar

    2016-06-01

    This paper investigates the application of Taguchi method with fuzzy logic for multi objective optimization of roughness parameters in electro discharge coating process of Al-6351 alloy with powder metallurgical compacted SiC/Cu tool. A Taguchi L16 orthogonal array was employed to investigate the roughness parameters by varying tool parameters like composition and compaction load and electro discharge machining parameters like pulse-on time and peak current. Crucial roughness parameters like Centre line average roughness, Average maximum height of the profile and Mean spacing of local peaks of the profile were measured on the coated specimen. The signal to noise ratios were fuzzified to optimize the roughness parameters through a single comprehensive output measure (COM). Best COM obtained with lower values of compaction load, pulse-on time and current and 30:70 (SiC:Cu) composition of tool. Analysis of variance is carried out and a significant COM model is observed with peak current yielding highest contribution followed by pulse-on time, compaction load and composition. The deposited layer is characterised by X-Ray Diffraction analysis which confirmed the presence of tool materials on the work piece surface.

  4. Substitution of synthetic chimpanzee androgen receptor for human androgen receptor in competitive binding and transcriptional activation assays for EDC screening

    EPA Science Inventory

    The potential effect of receptor-mediated endocrine modulators across species is of increasing concern. In attempts to address these concerns we are developing androgen and estrogen receptor binding assays using recombinant hormone receptors from a number of species across differ...

  5. Fabrication of chondroitin sulfate-chitosan composite artificial extracellular matrix for stabilization of fibroblast growth factor.

    PubMed

    Mi, Fwu-Long; Shyu, Shin-Shing; Peng, Chih-Kang; Wu, Yu-Bey; Sung, Hsing-Wen; Wang, Pei-Shan; Huang, Chi-Chuan

    2006-01-01

    The development of a novel, three-dimensional, macroporous artificial extracellular matrix (AECM) based on chondroitin sulfate (ChS)-chitosan (Chito) combination is reported. The composite AECM composed of ChS-Chito conjugated network was prepared by a homogenizing interpolyelectrolyte complex/covalent conjugation technique through co-crosslinked with N,N-(3-dimethylaminopropyl)-N'-ethyl carbodiimide (EDC) and N-hydroxysuccinimide (NHS). In contrast to EDC/NHS, two different reagents, calcium ion and glutaraldehyde, were used to react with ChS or Chito for the preparation of ChS-Chito composites containing crosslinked ChS or Chito network in the matrix. The stability and in vitro enzymatic degradability of the glutaraldehyde-, EDC/NHS-, and Ca2+ -crosslinked ChS-Chito composite AECMs were all investigated in this study. The results showed that crosslinking improved the stability of prepared ChS-Chito AECMs in physiological buffer solution (PBS) and provided superior protective effect against the enzymatic hydrolysis of ChS, compared with their non-crosslinked counterpart. Because ChS was a heparin-like glycosaminoglycan (GAG), the ChS-Chito composite AECMs appeared to promote binding efficiency for basic fibroblast growth factor (bFGF). The bFGF releasing from the ChS-Chito composite AECMs retained its biological activity as examined by the in vitro proliferation of human fibroblast, depending on the crosslinking mode for the preparation of these composite AECMs. Histological assay showed that the EDC/NHS-crosslinked ChS-Chito composite AECM, after incorporated with bFGF, was biodegradable and could result in a significantly enhanced vascularization effect and tissue penetration. These results suggest that the ChS-Chito composite AECMs fabricated in this study may be a promising approach for tissue-engineering application. PMID:16224775

  6. Determination of the cross-linking effect of adipic acid dihydrazide on glycoconjugate preparation.

    PubMed

    Bystrický, S; Machová, E; Malovíková, A; Kogan, G

    1999-11-01

    The cross-linking effect of adipic acid dihydrazide (ADH) on polysaccharide derivatization can be evaluated by applying combination of elemental analysis and colorimetric assay. Elemental analysis is used for estimation of total ADH bound to polysaccharide and a colorimetric trinitrobenzene sulfonic acid assay is used to determine the part of ADH not involved in cross-linking. The difference of values expressed as molar ratios (per repeating unit) provides information on the amount of ADH involved in cross-linking the polysaccharides. Carboxymethylated polysaccharides were derivatized with different amounts of ADH to test the procedure. Analytical results showed that excess of ADH in the reaction only slightly decreased the cross-linking. The number of carboxyl groups remained unmodified even at high excess of ADH and high concentration of carbodiimide (EDC) coupling reagent. PMID:11003553

  7. Effect of Gelatin amount on Properties of NANO-BCP/GEL Scaffolds

    NASA Astrophysics Data System (ADS)

    Bakhtiari, Leila; Hossainalipour, Seyed Mohamad; Rezaie, Hamid Reza

    Biodegradable polymers and bioactive ceramics are being combined in a variety of composite materials for tissue engineering scaffolds. Porous nano-biphasic calcium phosphate/gelatin structure was prepared by freeze-drying method. Pre-pores were created by using naphthalene with different particle sizes. Stabilization of gelatin network matrix carried out by EDC (N-(3-dimethyl aminopropyl)-N‧-ethyl carbodiimide hydrochloride) with cross-linking method. Three different amount of gelatin (2, 6 and 10 mg/ml) solution were used to study the effect of gelatin amount on properties of the scaffold. Microstructural properties of scaffolds were characterized by scanning electron microscopy (SEM). Fourier transform infrared spectroscopy (FTIR) was used to determine the chemical composition of scaffold. Also the morphology and bending strength were investigated.

  8. The stabilization and targeting of surfactant-synthesized gold nanorods

    NASA Astrophysics Data System (ADS)

    Rostro-Kohanloo, Betty C.; Bickford, Lissett R.; Payne, Courtney M.; Day, Emily S.; Anderson, Lindsey J. E.; Zhong, Meng; Lee, Seunghyun; Mayer, Kathryn M.; Zal, Tomasz; Adam, Liana; Dinney, Colin P. N.; Drezek, Rebekah A.; West, Jennifer L.; Hafner, Jason H.

    2009-10-01

    The strong cetyltrimethylammonium bromide (CTAB) surfactant responsible for the synthesis and stability of gold nanorod solutions complicates their biomedical applications. The critical parameter to maintain nanorod stability is the ratio of CTAB to nanorod concentration. The ratio is approximately 740 000 as determined by chloroform extraction of the CTAB from a nanorod solution. A comparison of nanorod stabilization by thiol-terminal PEG and by anionic polymers reveals that PEGylation results in higher yields and less aggregation upon removal of CTAB. A heterobifunctional PEG yields nanorods with exposed carboxyl groups for covalent conjugation to antibodies with the zero-length carbodiimide linker EDC. This conjugation strategy leads to approximately two functional antibodies per nanorod according to fluorimetry and ELISA assays. The nanorods specifically targeted cells in vitro and were visible with both two-photon and confocal reflectance microscopies. This covalent strategy should be generally applicable to other biomedical applications of gold nanorods as well as other gold nanoparticles synthesized with CTAB.

  9. Polyaniline-carbon nanotube composite film for cholesterol biosensor.

    PubMed

    Dhand, Chetna; Arya, Sunil K; Datta, Monika; Malhotra, B D

    2008-12-15

    Nanocomposite film composed of polyaniline (PANI) and multiwalled carbon nanotubes (MWCNT), prepared electrophoretically onto indium tin oxide (ITO)-coated glass plate, was used for covalent immobilization of cholesterol oxidase (ChOx) via N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) chemistry. Results of linear sweep voltammetric measurements reveal that ChOx/PANI-MWCNT/ITO bioelectrode can detect cholesterol in the range of 1.29 to 12.93 mM with high sensitivity of 6800 nA mM(-1) and a fast response time of 10 s. Photometric studies for ChOx/PANI-MWCNT/ITO bioelectrode indicate that it is thermally stable up to 45 degrees C and has a shelf life of approximately 12 weeks when stored at 4 degrees C. The results of these studies have implications for the application of this interesting matrix (PANI-MWCNT) toward the development of other biosensors. PMID:18817744

  10. The Adhesion and Neurite Outgrowth of Neurons on Poly(D-lysine)/Hyaluronan Multilayer Films.

    PubMed

    Shi, Haifei; Sheng, Guoping

    2016-06-01

    Poly(D-lysine)/hyaluronan (PDL/HA) films were prepared using layer-by-layer assembly technique and chemically cross-linked with a water soluble carbodiimide (EDC) in combination with N-hydroxysuccinimide (NHS) through formation of amide bonds. Quartz crystal microbalance with dissipation (QCM-D) was used to follow the cross-linking reaction. Atomic force measurement, ellipsometry, and Fourier transform infrared (FTIR) spectroscopy were performed to study the chemical structure, topography, thickness and mechanical properties of the cross-linked films. QCM-D and Frictional force study were used to reveal the viscoelasticity of the films after cross-linking treatment. The stability of the films was studied via incubating the films in physiological environment. Finally, the neurons were used to evaluate the interaction between films and cells. The results indicated that the neurons were preferably proliferating and outgrowth neurite on cross-linked films while uncross-linked films are highly cell resistant. PMID:27427590

  11. Epoxy Cross-Linked Collagen and Collagen-Laminin Peptide Hydrogels as Corneal Substitutes

    PubMed Central

    Koh, Li Buay; Islam, Mohammad Mirazul; Mitra, Debbie; Noel, Christopher W.; Merrett, Kimberley; Odorcic, Silvia; Fagerholm, Per; Jackson, William. Bruce; Liedberg, Bo; Phopase, Jaywant; Griffith, May

    2013-01-01

    A bi-functional epoxy-based cross-linker, 1,4-Butanediol diglycidyl ether (BDDGE), was investigated in the fabrication of collagen based corneal substitutes. Two synthetic strategies were explored in the preparation of the cross-linked collagen scaffolds. The lysine residues of Type 1 porcine collagen were directly cross-linked using l,4-Butanediol diglycidyl ether (BDDGE) under basic conditions at pH 11. Alternatively, under conventional methodology, using both BDDGE and 1-Ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC)/N-hydroxysuccinimide (NHS) as cross-linkers, hydrogels were fabricated under acidic conditions. In this latter strategy, Cu(BF4)2·XH2O was used to catalyze the formation of secondary amine bonds. To date, we have demonstrated that both methods of chemical cross-linking improved the elasticity and tensile strength of the collagen implants. Differential scanning calorimetry and biocompatibility studies indicate comparable, and in some cases, enhanced properties compared to that of the EDC/NHS controls. In vitro studies showed that human corneal epithelial cells and neuronal progenitor cell lines proliferated on these hydrogels. In addition, improvement of cell proliferation on the surfaces of the materials was observed when neurite promoting laminin epitope, IKVAV, and adhesion peptide, YIGSR, were incorporated. However, the elasticity decreased with peptide incorporation and will require further optimization. Nevertheless, we have shown that epoxy cross-linkers should be further explored in the fabrication of collagen-based hydrogels, as alternatives to or in conjunction with carbodiimide cross-linkers. PMID:24956085

  12. Capsular polysaccharide from Mycoplasma mycoides subsp. mycoides shows potential for protection against contagious bovine pleuropneumonia.

    PubMed

    Mwirigi, Martin; Nkando, Isabel; Olum, Moses; Attah-Poku, Samuel; Ochanda, Horace; Berberov, Emil; Potter, Andrew; Gerdts, Volker; Perez-Casal, Jose; Wesonga, Hezron; Soi, Reuben; Naessens, Jan

    2016-10-01

    Contagious Bovine Pleuropneumonia (CBPP) is a severe respiratory disease caused by Mycoplasma mycoides subsp. mycoides (Mmm) which is widespread in Africa. The capsule polysaccharide (CPS) of Mmm is one of the few identified virulence determinants. In a previous study, immunization of mice against CPS generated antibodies, but they were not able to prevent multiplication of Mmm in this model animal. However, mice cannot be considered as a suitable animal model, as Mmm does not induce pathology in this species. Our aim was to induce antibody responses to CPS in cattle, and challenge them when they had specific CPS antibody titres similar or higher than those from cattle vaccinated with the live vaccine. The CPS was linked to the carrier protein ovalbumin via a carbodiimide-mediated condensation with 1-ethyl-3(3-imethylaminopropyl) carbodiimide (EDC). Ten animals were immunized twice and challenged three weeks after the booster inoculation, and compared to a group of challenged non-immunized cattle. When administered subcutaneously to adult cattle, the vaccine elicited CPS-specific antibody responses with the same or a higher titre than animals vaccinated with the live vaccine. Pathology in the group of immunized animals was significantly reduced (57%) after challenge with Mmm strain Afadé compared to the non-immunized group, a figure in the range of the protection provided by the live vaccine. PMID:27496744

  13. In vitro and in vivo evaluation of bone morphogenetic protein-2 (BMP-2) immobilized collagen-coated polyetheretherketone (PEEK)

    NASA Astrophysics Data System (ADS)

    Du, Ya-Wei; Zhang, Li-Nan; Ye, Xin; Nie, He-Min; Hou, Zeng-Tao; Zeng, Teng-Hui; Yan, Guo-Ping; Shang, Peng

    2015-03-01

    Polyetheretherketone (PEEK) is regarded as one of the most potential candidates of biomaterials in spinal implant applications. However, as a bioinert material, PEEK plays a limited role in osteoconduction and osseointegration. In this study, recombinant human bone morphogenetic protein-2 (rhBMP-2) was immobilized onto the surface of collagen-coated PEEK in order to prepare a multi-functional material. After adsorbed onto the PEEK surface by hydrophobic interaction, collagen was cross-linked with N-(3-dimethylaminopropyl)-N'-ethyl carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). EDC/NHS system also contributed to the immobilization of rhBMP-2. Water contact angle tests, XPS and SEM clearly demonstrated the surface changes. ELISA tests quantified the amount of rhBMP-2 immobilized and the release over a period of 30 d. In vitro evaluation proved that the osteogenesis differentiation rate was higher when cells were cultured on modified PEEK discs than on regular ones. In vivo tests were conducted and positive changes of major parameters were presented. This report demonstrates that the rhBMP-2 immobilized method for PEEK modification increase bioactivity in vitro and in vivo, suggesting its practicability in orthopedic and spinal clinical applications.

  14. Efficient protein immobilization on polyethersolfone electrospun nanofibrous membrane via covalent binding for biosensing applications.

    PubMed

    Mahmoudifard, Matin; Soudi, Sara; Soleimani, Masoud; Hosseinzadeh, Simzar; Esmaeili, Elaheh; Vossoughi, Manouchehr

    2016-01-01

    In this paper we introduce novel strategy for antibody immobilization using high surface area electrospun nanofibrous membrane based on ethyl-3-(3-dimethylaminopropyl)-carbodiimide/N-hydroxysuccinimide (EDC/NHS) coupling chemistry. To present the high performance of proposed biosensors, anti-staphylococcus enterotoxin B (anti-SEB) was used as a model to demonstrate the utility of our proposed system. Polymer solution of polyethersolfone was used to fabricate fine nanofibrous membrane. Moreover, industrial polyvinylidene fluoride membrane and conventional microtiter plate were also used to compare the efficiency of antibody immobilization. Scanning electron microscopy images were taken to study the morphology of the membranes. The surface activation of nanofibrous membrane was done with the help of O2 plasma. PES nanofibrous membrane with carboxyl functional groups for covalent attachment of antibodies were treated by EDC/NHS coupling agent. The quantity of antibody immobilization was measured by enzyme-linked immuno sorbent assay (ELISA) method. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) spectroscopy was performed to confirm the covalent immobilization of antibody on membrane. Atomic force microscopy, scanning electron microscopy and invert fluorescence microscopy were used to analyze the antibody distribution pattern on solid surfaces. Results show that oxygen plasma treatment effectively increased the amount of antibody immobilization through EDC/NHS coupling chemistry. It was found that the use of nanofibrous membrane causes the improved detection signal of ELISA based biosensors in comparison to the standard assay carried out in the 96-well microtiter plate. This method has the potential to improve the ELISA-based biosensor and we believe that this technique can be used in various biosensing methods. PMID:26478348

  15. Simultaneous monitoring of seven phenolic metabolites of endocrine disrupting compounds (EDC) in human urine using gas chromatography with tandem mass spectrometry.

    PubMed

    Schmidt, Lukas; Müller, Johannes; Göen, Thomas

    2013-02-01

    A gas chromatographic-tandem mass spectrometric (GC-MS/MS) method for the simultaneous determination of the three well-known endocrine disruptors, bisphenol A, daidzein and genistein, as well as of four human pesticide metabolites which are supposed to have proper endocrine activity or which are metabolites of endocrine-disrupting compounds, viz., 1- and 2-naphthol, 2-isopropoxyphenol and 3,5,6-trichloropyridinol, has been developed and validated. The method involves enzymatic cleavage of the conjugates using β-glucuronidase/arylsulfatase followed by solid-phase extraction and derivatisation with N-tert-butyldimethylsilyl-N-methyltrifluoroacetamide. Isotopically labelled internal standards were used for all analytes, to achieve best analytical error correction. The method proved to be both sensitive and reliable in human urine with detection limits ranging from 0.1 to 0.6 μg/L for all analytes. Precision and repeatability was determined to range from 1 to 15 %. Compared with other published analytical procedures, the present method enables the simultaneous determination of a couple of phenolic agents with competitive or improved analytical reliability. Thus, the present method is suitable for a combined monitoring of the exposure to prominent xenobiotics with effects on the human endocrine system (bisphenol A, carbaryl, chlorpyrifos, chlorpyrifos-methyl, naphthalene, propoxur, triclopyr) and phytoestrogens (daidzein, genistein) in population studies. PMID:23241820

  16. Testing Of An Ultraviolet (UV)-Transparent Polymer-Based Passive Sampler for Rapid, Ultra-Low-Cost EDC Screening Applications

    EPA Science Inventory

    A new passive sampling method with rapid low-cost spectral detection has recently been developed. The method makes use of an ultraviolet (UV)-transparent polymer which serves as both a concentrator for dissolved compounds, and an optical cell for UV spectral detection. Because ...

  17. High throughput adjustable 96-well plate assay for androgen receptor binding: a practical approach for EDC screening using the chimpanzee AR.

    PubMed

    Hartig, P C; Cardon, M C; Blystone, C R; Gray, L E; Wilson, V S

    2008-09-26

    The issue as to whether natural and man-made chemicals interfere with endocrine function has raised concerns. This interference could be biologically significant even at very low doses if the chemicals interact deleteriously with hormone receptors at low concentrations. Therefore, the United States Environmental Protection Agency (USEPA) Office of Coordination and Policy (OSCP) requested that a nonhuman mammalian androgen receptor binding assay be developed for possible use in their Endocrine Disruptor Screening Program (EDSP). Ideally, this assay would be high throughput, not use animals as a source of receptor protein, easily deployed throughout the scientific community, utilize reagents available to both the public and private sector, and have the potential for future automation. We developed a highly modified 96-well plate assay which meets these criteria. It employs a baculovirus expressed recombinant primate androgen receptor which is publically available and exploits the unique ability of some mammalian androgen receptors to remain biologically active after guanidine hydrochloride (GdnHCl) solubilization. This GdnHCl treated receptor remains soluble and requires no additional purification prior to use. We provide a very detailed description of the assay protocol itself, and similarly detailed method for producing and solubilizing the receptor. PMID:18691642

  18. Alternatives to in vivo tests to detect endocrine disrupting chemicals (EDCs) in fish and amphibians – interactions with estrogens, androgens, and thyroid hormones

    EPA Science Inventory

    Endocrine disruption is considered a highly relevant endpoint for environmental risk assessment of chemicals, plant protection products, biocides and pharmaceuticals. Therefore, screening for endocrine disruption – with focus on vertebrates (fish and amphibians) and estrogen, and...

  19. An investigation into the extent and bilogcal impacts of endocrine disrupting chemicals (EDCs) in a highly effluent-dominated river in New England

    EPA Science Inventory

    The Assabet River in central Massachusetts is a heavily effluent-dominated river and during low-flow conditions, is composed almost entirely of waterwater effluent (i.e., up to 95%). The U.S EPA Regional New England Laboratory and the U.S. EPA Office of Research and Development ...

  20. Chitosan-tethered poly(acrylonitrile-co-maleic acid) hollow fiber membrane for lipase immobilization.

    PubMed

    Ye, Peng; Xu, Zhi-Kang; Che, Ai-Fu; Wu, Jian; Seta, Patrick

    2005-11-01

    A protocol was used to prepare a dual-layer biomimetic membrane as support for enzyme immobilization by tethering chitosan on the surface of poly(acrylonitrile-co-maleic acid) (PANCMA) ultrafiltration hollow fiber membrane in the presence of 1-ethyl-3-(dimethylaminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxylsuccin-imide (NHS). The chemical change of the chitosan-modified PANCMA membrane surface was confirmed with Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy. Lipase from Candida rugosa was immobilized on this dual-layer biomimetic membrane using glutaraldehyde (GA), and on the nascent PANCMA membrane using EDC/NHS as coupling agent. The properties of the immobilized enzymes were assayed and compared with those of the free one. It was found that both the activity retention of the immobilized lipase and the amount of bound protein on the dual-layer biomimetic membrane (44.5% and 66.5 mg/m2) were higher than those on the nascent PANCMA membrane (33.9% and 53.7 mg/m2). The kinetic parameters of the free and immobilized lipases, Km and Vmax, were also assayed. The Km values were similar for the immobilized lipases, while the Vmax value of the immobilized lipase on the dual-layer biomimetic membrane was higher than that on the nascent PANCMA membrane. Results indicated that the pH and thermal stabilities of lipase increased upon immobilization. The residual activity of the immobilized lipase after 10 uses was 53% on the dual-layer biomimetic membrane and 62% on the nascent PANCMA membrane. PMID:15919112

  1. Nanofibrous poly(acrylonitrile-co-maleic acid) membranes functionalized with gelatin and chitosan for lipase immobilization.

    PubMed

    Ye, Peng; Xu, Zhi-Kang; Wu, Jian; Innocent, Christophe; Seta, Patrick

    2006-08-01

    Nanofibrous membranes with an average diameter of 100 and 180 nm were fabricated from poly(acrylonitrile-co-maleic acid) (PANCMA) by the electrospinning process. These nanofibrous membranes contain reactive groups which can be used to covalently immobilize biomacromolecules. Two natural macromolecules, chitosan and gelatin, were tethered on these nanofibrous membranes to fabricate dual-layer biomimetic supports for enzyme immobilization in the presence of 1-ethyl-3-(dimethyl-aminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxyl succinimide (NHS). Lipase from Candida rugosa was then immobilized on these dual-layer biomimetic supports using glutaraldehyde (GA), and on the nascent PANCMA fibrous membrane using EDC/NHS as coupling agent, respectively. The properties of the immobilized lipases were assayed. It was found that there is an increase of the activity retention of the immobilized lipase on the chitosan-modified nanofibrous membrane (45.6+/-1.8%) and on the gelatin-modified one (49.7+/-1.8%), compared to that on the nascent one (37.6+/-1.8%). The kinetic parameters of the free and immobilized lipases, K(m) and V(max), were also assayed. In comparison with the immobilized lipase on the nascent nanofibrous membrane, there is an increase of the V(max) value for the immobilized lipases on the chitosan- and gelatin-modified nanofibrous membranes. Results also indicate that the pH and thermal stabilities of lipases increase upon immobilization. The residual activities of the immobilized lipases are 55% on the chitosan-modified nanofibrous membrane and 60% on the gelatin-modified one, after 10 uses. PMID:16584770

  2. Affinity labeling of the folate-methotrexate transporter from Leishmania donovani

    SciTech Connect

    Beck, J.T.; Ullman, B. )

    1989-08-22

    An affinity labeling technique has been developed to identify the folate-methotrexate transporter of Leishmania donovani promastigotes using activated derivatives of the ligands. These activated derivatives were synthesized by incubating folate and methotrexate with a 10-fold excess of 1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide (EDC) for 10 min at ambient temperature in dimethyl sulfoxide. When intact wild-type (DI700) Leishmania donovani or preparations of their membranes were incubated with a 0.4 {mu}M concentration of either activated ({sup 3}H)folate or activated ({sup 3}H)methotrexate, the radiolabeled ligands were covalently incorporated into a polypeptide with a molecular weight of approximately 46,000, as demonstrated by SDS-polyacrylamide gel electrophoresis. No affinity labeling of a 46,000-dalton protein was observed when equimolar concentrations of activated radiolabeled ligands were incubated with intact cells or membranes prepared from a methotrexate-resistant mutant clone of Leishmania donovani, MTXA5, that is genetically defective in folate-methotrexate transport capability. Time course studies indicated that maximal labeling of the 46,000-dalton protein occurred within 5-10 min of incubation of intact cells with activated ligand. These studies provide biochemical evidence that the folate-methotrexate transporter of Leishmania donovani can be identified in crude extracts by an affinity labeling technique and serve as a prerequisite to further analysis of the transport protein by providing a vehicle for subsequent purification of this membrane carrier. Moreover, these investigations suggest that the affinity labeling technique using EDC-activated ligands may be exploitable to analyze other cell surface binding proteins in Leishmania donovani, as well as in other organisms.

  3. Microfluidic enzymatic-reactors for peptide mapping: strategy, characterization, and performance.

    PubMed

    Wu, Huiling; Zhai, Jianjun; Tian, Yuping; Lu, Haojie; Wang, Xiaoyan; Jia, Weitao; Liu, Baohong; Yang, Pengyuang; Xu, Yunmin; Wang, Honghai

    2004-12-01

    The design and characterization of two kinds of poly(dimethylsiloxane)(PDMS) microfluidic enzymatic-reactors along with their analytical utility coupled to MALDI TOF and ESI MS were reported. Microfluidic devices integrated with microchannel and stainless steel tubing (SST) was fabricated using a PDMS casting technique, and was used for the preparation of the enzymatic-reactor. The chemical modification was performed by introducing carboxyl groups to PDMS surface based on ultraviolet graft polymerization of acrylic acid. The covalent and physical immobilization of trypsin was carried out with the use of the activation reagents 1-ethyl-3-(3-dimethyl aminopropyl)carbodiimide(EDC)/N-hydroxysuccinimide (NHS) and a coupling reagent poly(diallyldimethylammonium chloride)(PDDA), respectively. The properties and success of processes of trypsin immobilization were investigated by measuring contact angle, infrared absorption by attenuated total reflection spectra, AFM imaging and electropherograms. An innovative feature of the microfluidic enzymatic-reactors is the feasibility of performing on-line protein analysis by embedded SST electrode and replaceable tip. The lab-made devices provide an excellent extent of digestion of several model proteins even at the fast flow rate of 3.5 microL min(-1) for the EDC/NHS-made device and 0.8 microL min(-1) for the PDDA-made device, which afford very short residence times of 5 s and 20 s, respectively. In addition, the lab-made devices are less susceptive to memory effect and can be used for at least 50 runs in one week without noticeable loss of activity. Moreover, the degraded PDDA-made device can be regenerated by simple treatment of a HCl solution. These features are the most required for microfluidic devices used for protein analysis. PMID:15570370

  4. Immobilization of Active Bacteriophages on Polyhydroxyalkanoate Surfaces.

    PubMed

    Wang, Chanchan; Sauvageau, Dominic; Elias, Anastasia

    2016-01-20

    A rapid, efficient technique for the attachment of bacteriophages (phages) onto polyhydroxyalkanoate (PHA) surfaces has been developed and compared to three reported methods for phage immobilization. Polymer surfaces were modified to facilitate phage attachment using (1) plasma treatment alone, (2) plasma treatment followed by activation by 1-ethyl-3-(3-(dimethylamino)propyl)carbodiimide hydrochloride (EDC) and N-hydroxysulfosuccinimide (sulfo-NHS), (3) plasma-initiated acrylic acid grafting, or (4) plasma-initiated acrylic acid grafting with activation by EDC and sulfo-NHS. The impact of each method on the surface chemistry of PHA was investigated using contact angle analysis and X-ray photoelectron spectroscopy. Each of the four treatments was shown to result in both increased hydrophilicity and in the modification of the surface functional groups. Modified surfaces were immersed in suspensions of phage T4 for immobilization. The highest level of phage binding was observed for the surfaces modified by plasma treatment alone. The change in chemical bond states observed for surfaces that underwent plasma treatment is suspected to be the cause of the increased binding of active phages. Plasma-treated surfaces were further analyzed through phage-staining and fluorescence microscopy to assess the surface density of immobilized phages and their capacity to capture hosts. The infective capability of attached phages was confirmed by exposing the phage-immobilized surfaces to the host bacteria Escherichia coli in both plaque and infection dynamic assays. Plasma-treated surfaces with immobilized phages displayed higher infectivity than surfaces treated with other methods; in fact, the equivalent initial multiplicity of infection was 2 orders of magnitude greater than with other methods. Control samples - prepared by immersing polymer surfaces in phage suspensions (without prior plasma treatment) - did not show any bacterial growth inhibition, suggesting they did not bind

  5. Label-free detection of rheumatoid factor using YbYxOy electrolyte-insulator-semiconductor devices.

    PubMed

    Pan, Tung-Ming; Lin, Ting-Wei; Chen, Ching-Yi

    2015-09-01

    In this study, we investigated the effect of yttrium content on the structural properties and sensing characteristics of YbYxOy sensing membranes for electrolyte-insulator-semiconductor (EIS) sensors to detect the rheumatoid factor (RF). The YbYxOy EIS device prepared at the 60 W plasma condition exhibited a higher sensitivity of 65.77 mV/pH, a lower hysteresis voltage of ∼1 mV, and a smaller drift rate of 0.14 mV/h than did those prepared at the other conditions. We attribute this behavior to the optimal yttrium content in the YbYxOy film forming a smooth surface. Furthermore, we used a novel YbTixOy EIS biosensor to measure the RF antigen in human serum because of its rapid and label-free detection. Two different techniques were used for the immobilization of RF antibody onto the surface of an YbTixOy EIS sensor. The RF antibody was directly immobilized on the EIS surface modified with 3-aminopropyltriethoxysilane (APTES) followed by glutaraldehyde (GA). In contrast, a mixture of 1-ethyl-3-(3-dimethylamino-propyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS) solution was used to functionalize the carboxyl groups at the tail of RF antibodies. RF antibodies functionalized with the active NHS esters were covalently immobilized on the APTES-modified YbTixOy surface. The immobilized RF antibodies on the EIS that are functionalized with the EDC and NHS exhibit higher (41.11mV/pCRF) for detection of serum RF antigen in the range 10(-7) to 10(-3) M, compared to traditional antibody immobilization technique via APTES and GA linkage. The YbTixOy EIS biosensor is a promising analytical tool for RF antigen monitoring due to its good sensitivity, stability and repeatability. PMID:26388391

  6. Three-dimensional organization of the archaeal A1-ATPase from Methanosarcina mazei Gö1.

    PubMed

    Coskun, Unal; Radermacher, Michael; Müller, Volker; Ruiz, Teresa; Grüber, Gerhard

    2004-05-21

    A modified isolation procedure provides a homogeneous A(1)-ATPase from the archaeon Methanosarcina mazei Gö1, containing the five subunits in stoichiometric amounts of A(3):B(3):C:D:F. A(1) obtained in this way was characterized by three-dimensional electron microscopy of single particles, resulting in the first three-dimensional reconstruction of an A(1)-ATPase at a resolution of 3.2 nm. The A(1) consists of a headpiece of 10.2 nm in diameter and 10.8 nm in height, formed by the six elongated subunits A(3) and B(3). At the bottom of the A(3)B(3) complex, a stalk of 3.0 nm in length can be seen. The A(3)B(3) domain surrounds a large cavity that extends throughout the length of the A(3)B(3) barrel. A part of the stalk penetrates inside this cavity and is displaced toward an A-B-A triplet. To investigate further the topology of the stalk subunits C-F in A(1), cross-linking has been carried out by using dithiobis[sulfosuccinimidylpropionate] (DSP) and 1-ethyl-3-(dimethylaminopropyl)-carbodiimide (EDC). In experiments where DSP was added the cross-linked products B-F, A(x)-D, A-B-D, and A(x)-B(x)-D were formed. Subunits B-F, A-D, A-B-D, and A-B-C-D could be cross-linked by EDC. The subunit-subunit interaction in the presence of DSP was also studied as a function of nucleotide binding, demonstrating movements of subunits C, D, and F during ATP cleavage. Finally, the three-dimensional organization of this A(1) complex is discussed in terms of the relationship to the F(1)- and V(1)-ATPases at a resolution of 3.2 nm. PMID:14988401

  7. Triple-helical collagen hydrogels via covalent aromatic functionalization with 1,3-Phenylenediacetic acid

    PubMed Central

    Tronci, Giuseppe; Doyle, Amanda; Russell, Stephen J.; Wood, David J.

    2016-01-01

    Chemical crosslinking of collagen is a general strategy to reproduce macroscale tissue properties in physiological environment. However, simultaneous control of protein conformation, material properties and biofunctionality is highly challenging with current synthetic strategies. Consequently, the potentially-diverse clinical applications of collagen-based biomaterials cannot be fully realised. In order to establish defined biomacromolecular systems for mineralised tissue applications, type I collagen was functionalised with 1,3-Phenylenediacetic acid (Ph) and investigated at the molecular, macroscopic and functional levels. Preserved triple helix conformation was observed in obtained covalent networks via ATR-FTIR (AIII/A1450 ~ 1) and WAXS, while network crosslinking degree (C: 87-99 mol.-%) could be adjusted based on specific reaction conditions. Decreased swelling ratio (SR: 823-1285 wt.-%) and increased thermo-mechanical (Td: 80-88 °C; E: 28-35 kPa; σmax: 6-8 kPa; εb: 53-58 %) properties were observed compared to state-of-the-art carbodiimide (EDC)-crosslinked collagen controls, likely related to the intermolecular covalent incorporation of the aromatic segment. Ph-crosslinked hydrogels displayed nearly intact material integrity and only a slight mass decrease (MR: 5-11 wt. %) following 1-week incubation in either PBS or simulated body fluid (SBF), in contrast to EDC-crosslinked collagen (MR: 33-58 wt. %). Furthermore, FTIR, SEM and EDS revealed deposition of a calcium-phosphate phase on SBF-retrieved samples, whereby an increased calcium phosphate ratio (Ca/P: 0.84-1.41) was observed in hydrogels with higher Ph content. 72-hour material extracts were well tolerated by L929 mouse fibroblasts, whereby cell confluence and metabolic activity (MTS assay) were comparable to those of cells cultured in cell culture medium (positive control). In light of their controlled structure-function properties, these biocompatible collagen hydrogels represent attractive

  8. Fabrication and characterization of ovalbumin films for wound dressing applications.

    PubMed

    Shojaee, Mozhgan; Navaee, Fatemeh; Jalili-Firoozinezhad, Sasan; Faturechi, Rahim; Majidi, Mohammad; Bonakdar, Shahin

    2015-03-01

    A great number of people suffer from burning injuries all around the world each year. Applying an appropriate wound dressing can promote new tissue formation, prevent losing water and inhibit invasion of infectious organisms. In this study, egg white with a long standing history, as a homemade remedy, was fabricated as a wound dressing for burn injuries. For this reason, ovalbumin films were cross-linked by 1-ethyl-3-3-dimethyl aminopropyl carbodiimide hydrochloride (EDC) with different concentrations (1, 5 and 10mM) using three concentrations of ethanol. Physical-chemical characterizations including Fourier transform infrared spectroscopy (FTIR), gas transmission rate (GTR), tensile mechanical tests, water uptake and degradation rate were performed on the samples. The sample with 5mM crosslinking agent at 70% ethanol was considered as the optimized one with 417kPa of ultimate tensile strength, 64% elongation at break and 230% water uptake. In addition, biological evaluations conducted by MTT and live/dead assay indicated no sign of cyto-toxicity for all the samples. Moreover, scanning electron microscopy (SEM) showed that the fibroblast cells were well spread on the sample with the formation of filopodia. In conclusion, modified ovalbumin can be applied as the base material for fabrication of wound dressing and skin care products. PMID:25579909

  9. Enhanced biocatalytic esterification with lipase-immobilized chitosan/graphene oxide beads.

    PubMed

    Lau, Siaw Cheng; Lim, Hong Ngee; Basri, Mahiran; Fard Masoumi, Hamid Reza; Ahmad Tajudin, Asilah; Huang, Nay Ming; Pandikumar, Alagarsamy; Chia, Chin Hua; Chia, Chi Hua; Andou, Yoshito

    2014-01-01

    In this work, lipase from Candida rugosa was immobilized onto chitosan/graphene oxide beads. This was to provide an enzyme-immobilizing carrier with excellent enzyme immobilization activity for an enzyme group requiring hydrophilicity on the immobilizing carrier. In addition, this work involved a process for the preparation of an enzymatically active product insoluble in a reaction medium consisting of lauric acid and oleyl alcohol as reactants and hexane as a solvent. This product enabled the stability of the enzyme under the working conditions and allowed the enzyme to be readily isolated from the support. In particular, this meant that an enzymatic reaction could be stopped by the simple mechanical separation of the "insoluble" enzyme from the reaction medium. Chitosan was incorporated with graphene oxide because the latter was able to enhance the physical strength of the chitosan beads by its superior mechanical integrity and low thermal conductivity. The X-ray diffraction pattern showed that the graphene oxide was successfully embedded within the structure of the chitosan. Further, the lipase incorporation on the beads was confirmed by a thermo-gravimetric analysis. The lipase immobilization on the beads involved the functionalization with coupling agents, N-hydroxysulfosuccinimide sodium (NHS) and 1-ethyl-(3-dimethylaminopropyl) carbodiimide (EDC), and it possessed a high enzyme activity of 64 U. The overall esterification conversion of the prepared product was 78% at 60 °C, and it attained conversions of 98% and 88% with commercially available lipozyme and novozyme, respectively, under similar experimental conditions. PMID:25127038

  10. Simple and sensitive progesterone detection in human serum using a CdSe/ZnS quantum dot-based direct binding assay.

    PubMed

    Oh, Sung-Duk; Duong, Hong Dinh; Rhee, Jong Il

    2015-08-15

    In this study, we developed a CdSe/ZnS quantum dot (QD)-based immunoassay for use in determining the presence of progesterone (P4) in human serum. Hydrophilic QDs were conjugated to anti-progesterone antibody (P4Ab) via ethyl-3-(dimethylaminopropyl)carbodiimide (EDC) and N-hydroxysuccinimide (NHS) as coupling reagents. After purification, the P4Ab-QD conjugates were immobilized onto the wells of a 96-well microtiter plate, and a direct-binding immunoassay based on the binding of P4 to immobilized P4Ab-QD conjugates had a detection limit of 0.21 ng/ml and a sensitivity of 1.37 ng/ml, with a linear range of 0.385 to 4.55 ng/ml. The proposed immunoassay was successfully used to determine the P4 concentration in real human serum, and the results showed a good correlation with the accredited radioimmunoassay (RIA). PMID:25963894

  11. Fabricating an Amperometric Cholesterol Biosensor by a Covalent Linkage between Poly(3-thiopheneacetic acid) and Cholesterol Oxidase

    PubMed Central

    Nien, Po-Chin; Chen, Po-Yen; Ho, Kuo-Chuan

    2009-01-01

    In this study, use of the covalent enzyme immobilization method was proposed to attach cholesterol oxidase (ChO) on a conducting polymer, poly(3-thiopheneacetic acid), [poly(3-TPAA)]. Three red-orange poly(3-TPAA) films, named electrodes A, B and C, were electropolymerized on a platinum electrode by applying a constant current of 1.5 mA, for 5, 20 and 100 s, respectively. Further, 1-ethyl-3-(3-dimethylamiopropyl)carbodiimide hydrochloride (EDC · HCl) and N-hydroxysuccinimide (NHS) were used to activate the free carboxylic groups of the conducting polymer. Afterwards, the amino groups of the cholesterol oxidase were linked on the activated groups to form peptide bonds. The best sensitivity obtained for electrode B is 4.49 mA M−1 cm−2, with a linear concentration ranging from 0 to 8 mM, which is suitable for the analysis of cholesterol in humans. The response time (t95) is between 70 and 90 s and the limit of detection is 0.42 mM, based on the signal to noise ratio equal to 3. The interference of species such as ascorbic acid and uric acid increased to 5.2 and 10.3% of the original current response, respectively, based on the current response of cholesterol (100%). With respect to the long-term stability, the sensing response retains 88% of the original current after 13 days. PMID:22573987

  12. Antibody conjugated graphene nanocomposites for pathogen detection

    NASA Astrophysics Data System (ADS)

    Sign, Chandan; Sumana, Gajjala

    2016-04-01

    Graphene oxide (GO), due to its excellent electrochemical properties and large surface area, known to be highly suitable material for biosensing application. Here, we report in situ synthesis of silver nanopaticles (AgNPs) onto the GO sheets for the electrochemical detection of Salmonella typhimurium (S.typhimurium). The GO-AgNPs composites have been deposited onto the indium tin oxide (ITO) coated glass substrate by the electrophoretic deposition technique. Carbodiimide coupling (EDC-NHS) has been used for the immobilization of antibodies of Salmonella typhimurium (anti-S.typhimurium) for detection of S.typhimurium. The electron microscopy and UV-visible studies reveal successful synthesis GO-AgNPs composites while FT-IR studies suggest the proper immobilization of anti-S.typhi. The cyclic voltammetry (CV) has been utilized for detection using anti-S.typhi/GOAgNPs/ITO based immunoelectrode as a function of S.typhimurium concentration. The fabricated immunosensor shows improved sensitivity of 33.04 μACFU-1mLcm-2 in a wide detection range of 101 to 106 CFUmL-1. This immunosensor may be utilized for the detection of other food borne pathogens like aflatoxin and E.coli also.

  13. Preparation and characterization of cross-linked collagen-phospholipid polymer hybrid gels.

    PubMed

    Nam, Kwangwoo; Kimura, Tsuyoshi; Kishida, Akio

    2007-01-01

    2-methacryloyloxyethyl phosphorylcholine (MPC)-immobilized collagen gel was developed. Using 1-ethyl-3-(3-dimethyl aminopropyl)-1-carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS), we cross-linked a collagen film in 2-morpholinoethane sulfonic acid (MES) buffer (EN gel). EN gel was prepared under both pH 4.5 and pH 9.0 in order to observe changes in cross-linking ability. To cross-link MPC to collagen gel, poly(MPC-co-methacrylic acid) (PMA) having a carboxyl group side chain was chosen. E/N gel was added to the MES buffer having pre-NHS activated PMA to make MPC-immobilized collagen gel (MiC gel). MiC gel was prepared under both acidic and alkaline conditions to observe the changes in the cross-linking ability of PMA. X-ray photoelectron spectroscopy showed that the PMA was cross-linked with collagen under both acidic and alkaline conditions. Differential scanning calorimetry (DSC) results showed that the shrinkage temperature increased for the MiC gels and that the increase would be greater for the MiC gel prepared under alkaline conditions. The data showed that swelling would be less when the MiC gel was prepared under alkaline conditions. The biodegradation caused by collagenase was suppressed for the MiC gel prepared under alkaline conditions due to stable inter- and intrahelical networks. PMID:16959313

  14. Immobilization of alkaline phosphatase on microporous nanofibrous fibrin scaffolds for bone tissue engineering.

    PubMed

    Osathanon, Thanaphum; Giachelli, Cecilia M; Somerman, Martha J

    2009-09-01

    Alkaline phosphatase (ALP) promotes bone formation by degrading inorganic pyrophosphate (PP(i)), an inhibitor of hydroxyapatite formation, and generating inorganic phosphate (P(i)), an inducer of hydroxyapatite formation. P(i) is a crucial molecule in differentiation and mineralization of osteoblasts. In this study, a method to immobilize ALP on fibrin scaffolds with tightly controllable pore size and pore interconnection was developed, and the biological properties of these scaffolds were characterized both in vitro and in vivo. Microporous, nanofibrous fibrin scaffolds (FS) were fabricated using a sphere-templating method. ALP was covalently immobilized on the fibrin scaffolds using 1-ethyl-3-(dimethylaminopropyl)carbodiimide hydrochloride (EDC). Scanning electron microscopic observation (SEM) showed that mineral was deposited on immobilized alkaline phosphatase fibrin scaffolds (immobilized ALP/FS) when incubated in medium supplemented with beta-glycerophosphate, suggesting that the immobilized ALP was active. Primary calvarial cells attached, spread and formed multiple layers on the surface of the scaffolds. Mineral deposition was also observed when calvarial cells were seeded on immobilized ALP/FS. Furthermore, cells seeded on immobilized ALP/FS exhibited higher osteoblast marker gene expression compared to control FS. Upon implantation in mouse calvarial defects, both the immobilized ALP/FS and FS alone treated group had higher bone volume in the defect compared to the empty defect control. Furthermore, bone formation in the immobilized ALP/FS treated group was statistically significant compared to FS alone group. However, the response was not robust. PMID:19501906

  15. Enhanced Biocatalytic Esterification with Lipase-Immobilized Chitosan/Graphene Oxide Beads

    PubMed Central

    Lau, Siaw Cheng; Lim, Hong Ngee; Basri, Mahiran; Fard Masoumi, Hamid Reza; Ahmad Tajudin, Asilah; Huang, Nay Ming; Pandikumar, Alagarsamy; Chia, Chi Hua; Andou, Yoshito

    2014-01-01

    In this work, lipase from Candida rugosa was immobilized onto chitosan/graphene oxide beads. This was to provide an enzyme-immobilizing carrier with excellent enzyme immobilization activity for an enzyme group requiring hydrophilicity on the immobilizing carrier. In addition, this work involved a process for the preparation of an enzymatically active product insoluble in a reaction medium consisting of lauric acid and oleyl alcohol as reactants and hexane as a solvent. This product enabled the stability of the enzyme under the working conditions and allowed the enzyme to be readily isolated from the support. In particular, this meant that an enzymatic reaction could be stopped by the simple mechanical separation of the “insoluble” enzyme from the reaction medium. Chitosan was incorporated with graphene oxide because the latter was able to enhance the physical strength of the chitosan beads by its superior mechanical integrity and low thermal conductivity. The X-ray diffraction pattern showed that the graphene oxide was successfully embedded within the structure of the chitosan. Further, the lipase incorporation on the beads was confirmed by a thermo-gravimetric analysis. The lipase immobilization on the beads involved the functionalization with coupling agents, N-hydroxysulfosuccinimide sodium (NHS) and 1-ethyl-(3-dimethylaminopropyl) carbodiimide (EDC), and it possessed a high enzyme activity of 64 U. The overall esterification conversion of the prepared product was 78% at 60°C, and it attained conversions of 98% and 88% with commercially available lipozyme and novozyme, respectively, under similar experimental conditions. PMID:25127038

  16. Investigation of terpinolene + ozone or terpinolene + nitrate radical reaction products using denuder/filter apparatus

    NASA Astrophysics Data System (ADS)

    Harrison, Joel C.; Wells, J. R.

    2013-12-01

    Terpinolene's (1-methyl-4-(propan-2-ylidene)cyclohexene) reaction with ozone or the nitrate radical was investigated using a denuder/filter apparatus in order to characterize gas-phase and particulate reaction products. Identification of the reaction products (i.e., aldehydes, ketones, dicarbonyls and carboxylic acids) was made using two derivatization methods; O-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine (PFBHA) to derivatize the carbonyl products or 3-Ethyl-1-[3-(dimethylamino)propyl]carbodiimide hydrochloride (EDC) and 2,2,2-trifluoroethylamine hydrochloride (TFEA) to derivatize the carboxylic acid products. Proposed carbonyl products for ozonolysis of terpinolene are: 4-methylcyclohex-3-en-1-one, 2-hydroxy-4-methylcyclohex-3-en-1-one, glyoxal, methyl glyoxal, 3-oxobutanal, and 6-oxo-3-(propan-2-ylidene)heptanal. Proposed carbonyl products for nitrate radical reaction of terpinolene are: 2-hydroxy-4-methylcyclohex-3-en-1-one, glyoxal, methyl glyoxal, and 4-oxopentanal. No carboxylic acid products were detected with either oxidizing reactant.

  17. Application of DNA Hybridization Biosensor as a Screening Method for the Detection of Genetically Modified Food Components

    PubMed Central

    Tichoniuk, Mariusz; Ligaj, Marta; Filipiak, Marian

    2008-01-01

    An electrochemical biosensor for the detection of genetically modified food components is presented. The biosensor was based on 21-mer single-stranded oligonucleotide (ssDNA probe) specific to either 35S promoter or nos terminator, which are frequently present in transgenic DNA cassettes. ssDNA probe was covalently attached by 5′-phosphate end to amino group of cysteamine self-assembled monolayer (SAM) on gold electrode surface with the use of activating reagents – water soluble 1-ethyl-3(3′-dimethylaminopropyl)-carbodiimide (EDC) and N-hydroxy-sulfosuccinimide (NHS). The hybridization reaction on the electrode surface was detected via methylene blue (MB) presenting higher affinity to ssDNA probe than to DNA duplex. The electrode modification procedure was optimized using 19-mer oligoG and oligoC nucleotides. The biosensor enabled distinction between DNA samples isolated from soybean RoundupReady® (RR soybean) and non-genetically modified soybean. The frequent introduction of investigated DNA sequences in other genetically modified organisms (GMOs) give a broad perspectives for analytical application of the biosensor.

  18. Development of an indirect competitive ELISA for simultaneous detection of enrofloxacin and ciprofloxacin*

    PubMed Central

    Zhang, Hai-tang; Jiang, Jin-qing; Wang, Zi-liang; Chang, Xin-yao; Liu, Xing-you; Wang, San-hu; Zhao, Kun; Chen, Jin-shan

    2011-01-01

    Modified 1-ethyl-3-(3-dimethylaminopropy) carbodiimide (EDC) method was employed to synthesize the artificial antigen of enrofloxacin (ENR), and New Zealand rabbits were used to produce anti-ENR polyclonal antibody (pAb). Based on the checkerboard titration, an indirect competitive enzyme-linked immunosorbent assay (ELISA) standard curve was established. This assay was sensitive and had a linear range from 0.6 to 148.0 μg/kg (R 2=0.9567), with the half maximal inhibitory concentration (IC50) and limit of detection (LOD) values of 9.4 μg/kg and 0.2 μg/kg, respectively. Of all the competitive analogues, the produced pAb exhibited a high cross-reactivity to ciprofloxacin (CIP) (87%), the main metabolite of ENR in tissues. After optimization, the matrix effects can be ignored using a 10-fold dilution in beef and 20-fold dilution in pork. The overall recoveries and coefficients of variation (CVs) were in the ranges of 86%–109% and 6.8%–13.1%, respectively. It can be concluded that the established ELISA method is suitable for simultaneous detection of ENR and CIP in animal tissues. PMID:22042652

  19. Carbon nanodots as molecular scaffolds for development of antimicrobial agents.

    PubMed

    Ngu-Schwemlein, Maria; Chin, Suk Fun; Hileman, Ryan; Drozdowski, Chris; Upchurch, Clint; Hargrove, April

    2016-04-01

    We report the potential of carbon nanodots (CNDs) as a molecular scaffold for enhancing the antimicrobial activities of small dendritic poly(amidoamines) (PAMAM). Carbon nanodots prepared from sago starch are readily functionalized with PAMAM by using N-ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS). Electron microscopy images of these polyaminated CNDs show that they are approximately 30-60nm in diameter. Infrared and fluorescence spectroscopy analyses of the water-soluble material established the presence of the polyamidoaminated moiety and the intrinsic fluorescence of the nanodots. The polyaminated nanodots (CND-PAM1 and CND-PAM2) exhibit in vitro antimicrobial properties, not only to non-multidrug resistant bacteria but also to the corresponding Gram-negative multidrug bacteria. Their minimum inhibitory concentration (MIC) ranges from 8 to 64μg/mL, which is much lower than that of PAMAM G1 or the non-active PAMAM G0 and CNDs. Additionally, they show synergistic effect in combination with tetracycline or colistin. These preliminary results imply that CNDs can serve as a promising scaffold for facilitating the rational design of antimicrobial materials for combating the ever-increasing threat of antibiotic resistance. Moreover, their fluorescence could be pertinent to unraveling their mode of action for imaging or diagnostic applications. PMID:26923697

  20. Development of a Specific Latex Agglutination Test to Detect Antibodies of Enterovirus 71.

    PubMed

    Qin, Bo; Zhang, Jianhua; Xie, Wenhao; Liu, Xuehong; He, Tingting; Chen, Jinkun; Dong, Xuejun

    2015-10-01

    A latex agglutination test (LAT) was developed for the rapid detection of antibodies against the VP1 or VP1 proteins of Enterovirus 71 (EV71). The proteins of interest including prokaryotically expressed VP1 and two strains of anti-VP1 monoclonal antibody (McAb) against EV71 were covalently linked to carboxylated latex using ethyl-dimethyl-amino-propyl carbodiimide (EDC) to prepare sensitized latex beads. LAT was evaluated by an enzyme-linked immunosorbent assay (ELISA) as a reference test. The VP1-LAT showed a sensitivity of 87.0%, specificity of 88.9%, and an agreement ratio of 90.0% in detecting VP1 in 100 serum samples from experimentally infected mice, whereas these values were 86.8, 96.7, and 93.3%, respectively, for 608 clinical human serum samples. The VP1-LAT has advantages over other assays in terms of low cost, rapidity, chemical stability, high sensitivity, repeatability, and specificity. The LAT established in the present study is a rapid and simple test suitable for field monitoring of antibodies against VP1-EV71. PMID:26363276

  1. Generation of anti-trenbolone monoclonal antibody and establishment of an indirect competitive enzyme-linked immunosorbent assay for detection of trenbolone in animal tissues, feed and urine.

    PubMed

    Zhang, Yuanyang; He, Fangyang; Wan, Yuping; Meng, Meng; Xu, Jing; Yi, Jian; Wang, Yabin; Feng, Caiwei; Wang, Shanliang; Xi, Rimo

    2011-01-15

    Trenbolone (TRE) is a steroid used by veterinarians on livestock to increase appetite and body weight. The use of TRE has been restricted because of its harmful side effect for consumers. To effectively control TRE residue in food and food product, a rapid and convenient immunoassay was developed by preparing an anti-TRE monoclonal antibody. The immunogen and coating antigen were prepared by coupling TRE hapten with carrier proteins via 1-ethyl-3-(dimethylaminopropyl)carbodiimide hydrochloride (EDC) method. The optimized method gave an average IC(50) value of 0.323 ng mL(-1) towards TRE and an average detection limit (LOD) of 0.06 ng mL(-1), which is much lower than the maximum residue levels (2.0 ng g(-1)) accepted by the Joint FAO/WHO Expert Committee on Food Additives (JECFA). The specificity of the antibody was evaluated by measuring cross-reactivity of six structurally related compounds, including 19-nortestosterone (9.7%), testosterone (0.13%), methyltestosterone (<0.01%), methandrostenolone (<0.01%), (+)-dehydroisoandrosterone (<0.001%) and β-estradiol (<0.001%). The recovery rates of the test in detection of TRE-fortified animal tissue, urine and animal feed samples were in the range of 81.3-89.4%, while the intra- and inter-assay coefficients of variation were less than 12.0%. PMID:21147313

  2. Preparation and characterization of carboxymethylated carrageenan modified with collagen peptides.

    PubMed

    Fan, Lihong; Tong, Jun; Tang, Chang; Wu, Huan; Peng, Min; Yi, Jiayan

    2016-01-01

    The preparation of carboxymethyl κ-carrageenan collagen peptide (CMKC-COP) was via an imide-bond forming reaction between carboxyl groups in carboxymethyl κ-carrageenan (CMKC) and amino groups in collagen peptide in the presence of 1-ethyl-(dimethylaminopropyl) carbodiimide (EDC) and N-hydroxy sulfosuccinimide (NHS). CMKC-COP products were verified with infrared spectroscopy (FT-IR). The results of degree of substitution (DS) of CMKC-COP was presented, which are depended on reaction time, molar ratio of collagen peptide to carboxymethyl κ-carrageenan and reaction temperature. The optimal reaction conditions were studied by means of single factor experiment. Also MTT assay was applied to evaluate the effects of CMKC-COP on proliferation of chick embryo fibroblasts. The animal experiment results indicated that the wound covered with CMKC-COP were completely filled with new epithelium within 2 weeks without any significant adverse side reactions. Therefore, the CMKC-COP showed the potentiality to repair skin in cosmetic, biomedical and pharmaceutical fields. PMID:26526172

  3. (Au/PANA/PVAc) nanofibers as a novel composite matrix for albumin and streptavidin immobilization.

    PubMed

    Golshaei, Rana; Guler, Zeliha; Sarac, Sezai A

    2016-03-01

    A novel electrospun nanofiber mat (Au/PANA/PVAc) consists of (Gold/Poly Anthranilic acid) (Au/PANA) core/shell nanostructures as a support material for protein immobilization that was developed and characterized by electrochemical impedance spectroscopy. In the core/shells, PANA served carboxyl groups (-COOH) for covalent protein immobilization and Au enhanced the electrochemical properties by acting as tiny conduction centers to facilitate electron transfer. Covalent immobilization of albumin and streptavidin as model proteins onto the (Au/PANA/PVAc) nanofibers was carried out by using 1-ethyl-3-(dimethyl-aminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxyl succinimide (NHS) activation. PVAc nanofibers were compared with Au/PANA/PVAc nanofibers before and after protein immobilization. The successful covalent binding of both albumin and streptavidin onto (Au/PANA/PVAc) nanofibers was confirmed by FTIR-ATR, Electron Microscopy/Energy-Dispersive X-ray Spectroscopy SEM/EDX and Electrochemical impedance spectroscopy (EIS). The nanofibers became resistive due to protein immobilization and the higher charge transfer resistance was observed after higher amount of protein was immobilized. PMID:26706530

  4. Additive manufacturing of collagen scaffolds by three-dimensional plotting of highly viscous dispersions.

    PubMed

    Lode, Anja; Meyer, Michael; Brüggemeier, Sophie; Paul, Birgit; Baltzer, Hagen; Schröpfer, Michaela; Winkelmann, Claudia; Sonntag, Frank; Gelinsky, Michael

    2016-03-01

    Additive manufacturing (AM) allows the free form fabrication of three-dimensional (3D) structures with distinct external geometry, fitting into a patient-specific defect, and defined internal pore architecture. However, fabrication of predesigned collagen scaffolds using AM-based technologies is challenging due to the low viscosity of collagen solutions, gels or dispersions commonly used for scaffold preparation. In the present study, we have developed a straightforward method which is based on 3D plotting of a highly viscous, high density collagen dispersion. The swollen state of the collagen fibrils at pH 4 enabled the homogenous extrusion of the material, the deposition of uniform strands and finally the construction of 3D scaffolds. Stabilization of the plotted structures was achieved by freeze-drying and chemical crosslinking with the carbodiimide EDC. The scaffolds exhibited high shape and dimensional fidelity and a hierarchical porosity consisting of macropores generated by strand deposition as well as an interconnected microporosity within the strands as result of the freeze-drying process. Cultivation of human mesenchymal stromal cells on the scaffolds, with and without adipogenic or osteogenic stimulation, revealed their cytocompatibility and potential applicability for adipose and bone tissue engineering. PMID:26924825

  5. Chemical modification of chitosan film via surface grafting of citric acid molecular to promote the biomineralization

    NASA Astrophysics Data System (ADS)

    Liu, Yang; Shen, Xin; Zhou, Huan; Wang, Yingjun; Deng, Linhong

    2016-05-01

    We develop a novel chitosan-citric acid film (abbreviated as CS-CA) suitable for biomedical applications in this study. In this CS-CA film, the citric acid, which is a harmless organic acid has been extensively investigated as a modifying agent on carbohydrate polymers, was cross-linked by 1-Ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) onto the surface of chitosan (CS) film. Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) confirms the graft copolymerization of the modified chitosan film (CS-CA). Surface wettability, moisturizing performance, the capacity of mineralization in vitro and biocompatibility of the films were characterized. After modification, this CS-CA film has good hydrophilicity. It is very evident that the citric acid grafting treatment significantly promotes the biomineralization of the chitosan based substrates. Cell experiments show that the MC3T3-E1 osteoblasts can adhere and proliferate well on the surface of CS-CA film. This CS-CA film, which can be prepared in large quantities and at low cost, should have potential application in bone tissue engineering.

  6. A simple enzyme-substrate localized conjugation method to generate immobilized, functional glutathione S-transferase fusion protein columns for affinity enrichment.

    PubMed

    Coughlin, John; Masci, Allyson; Gronke, Robert S; Bergelson, Svetlana; Co, Carl

    2016-07-15

    Immobilized protein receptors and enzymes are tools for isolating or enriching ligands and substrates based on affinity. For example, glutathione S-transferase (GST) is fused to proteins as a tag for binding to its substrate glutathione (GSH) linked to solid supports. One issue with this approach is that high-affinity interactions between receptors and ligands require harsh elution conditions such as low pH, which can result in leached receptor. Another issue is the inherent nonspecific chemical conjugation of reactive groups such as N-hydroxysuccinimide (NHS) that couple lysines to solid supports; the nonspecificity of NHS may result in residue modifications near the binding site(s) of the receptor that can affect ligand specificity. In this study, a simple conjugation procedure is presented that overcomes these limitations and results in immobilized GST fusion proteins that are functional and specific. Here, the affinity of GST for GSH was used to generate an enzyme-substrate site-specific cross-linking reaction; GSH-Sepharose was preactivated with 1-ethyl-3-(dimethylaminopropyl)carbodiimide (EDC) and then incubated Fc gamma receptor IIIa (FcγRIIIa)-GST. The immobilized FcγRIIIa-GST more specifically bound glycosylated immunoglobulin G1s (IgG1s) and was used to enrich nonfucosylated IgG1s from weaker binding species. This technique can be used when modifications of amino acids lead to changes in activity. PMID:27063248

  7. Amperometric creatinine biosensor based on covalently coimmobilized enzymes onto carboxylated multiwalled carbon nanotubes/polyaniline composite film.

    PubMed

    Yadav, Sandeep; Kumar, Ashok; Pundir, C S

    2011-12-15

    A mixture of commercial creatinine amidohydrolase (CA), creatine amidinohydrolase (CI), and sarcosine oxidase (SO) was coimmobilized covalently via N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxy succinimide (NHS) chemistry onto carboxylated multiwalled carbon nanotube (c-MWCNT)/polyaniline (PANI) nanocomposite film electrodeposited over the surface of a platinum (Pt) electrode. A creatinine biosensor was fabricated using enzyme/c-MWCNT/PANI/Pt as working electrode, Ag/AgCl as reference electrode, and Pt wire as auxiliary electrode connected through potentiostat. The enzyme electrode was characterized by scanning electron microscopy (SEM), Fourier transform infrared (FTIR) spectroscopy, and electrochemical impedance spectroscopy (EIS). The biosensor detected creatinine levels as low as 0.1 μM, estimated at a signal-to-noise ratio of 3, within 5s at pH 7.5 and 35°C. The optimized biosensor showed a linear response range of 10 to 750 μM creatinine with sensitivity of 40 μA/mM/cm(2). The fabricated biosensor was successfully employed for determination of creatinine in human serum. The biosensor showed only 15% loss in its initial response after 180 days when stored at 4°C. PMID:21906581

  8. Electrochemical detection of uric acid via uricase-immobilized graphene oxide.

    PubMed

    Omar, Muhamad Nadzmi; Salleh, Abu Bakar; Lim, Hong Ngee; Ahmad Tajudin, Asilah

    2016-09-15

    Measurement of the uric acid level in the body can be improved by biosensing with respect to the accuracy, sensitivity and time consumption. This study has reported the immobilization of uricase onto graphene oxide (GO) and its function for electrochemical detection of uric acid. Through chemical modification of GO using 1-ethyl-3-(dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysulfosuccinimide (NHS) as cross-linking reagents, the enzyme activity of the immobilized uricase was much comparable to the free enzyme with 88% of the activity retained. The modified GO-uricase (GOU) was then subjected to electrocatalytic detection of uric acid (UA) via cyclic voltammetry (CV). For that reason, a glassy carbon electrode (GCE) was modified by adhering the GO along with the immobilized uricase to facilitate the redox reaction between the enzyme and the substrate. The modified GOU/GCE outperformed a bare electrode through the electrocatalytic activity with an amplified electrical signal for the detection of UA. The electrocatalytic response showed a linear dependence on the UA concentration ranging from 0.02 to 0.49 mM with a detection limit of 3.45 μM at 3σ/m. The resulting biosensor also exhibited a high selectivity towards UA in the presence of other interference as well as good reproducibility. PMID:27402177

  9. Immobilization of lambda exonuclease onto polymer micropillar arrays for the solid-phase digestion of dsDNAs.

    PubMed

    Oliver-Calixte, Nyoté J; Uba, Franklin I; Battle, Katrina N; Weerakoon-Ratnayake, Kumuditha M; Soper, Steven A

    2014-05-01

    The process of immobilizing enzymes onto solid supports for bioreactions has some compelling advantages compared to their solution-based counterpart including the facile separation of enzyme from products, elimination of enzyme autodigestion, and increased enzyme stability and activity. We report the immobilization of λ-exonuclease onto poly(methylmethacrylate) (PMMA) micropillars populated within a microfluidic device for the on-chip digestion of double-stranded DNA. Enzyme immobilization was successfully accomplished using 3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS) coupling to carboxylic acid functionalized PMMA micropillars. Our results suggest that the efficiency for the catalysis of dsDNA digestion using λ-exonuclease, including its processivity and reaction rate, were higher when the enzyme was attached to a solid support compared to the free solution digestion. We obtained a clipping rate of 1.0 × 10(3) nucleotides s(-1) for the digestion of λ-DNA (48.5 kbp) by λ-exonuclease. The kinetic behavior of the solid-phase reactor could be described by a fractal Michaelis-Menten model with a catalytic efficiency nearly 17% better than the homogeneous solution-phase reaction. The results from this work will have important ramifications in new single-molecule DNA sequencing strategies that employ free mononucleotide identification. PMID:24628008

  10. Conjugation of glucose oxidase onto Mn-doped ZnS quantum dots for phosphorescent sensing of glucose in biological fluids.

    PubMed

    Wu, Peng; He, Yu; Wang, He-Fang; Yan, Xiu-Ping

    2010-02-15

    Integrating various enzymes with nanomaterials provides various nanohybrids with new possibilities in biosensor applications. Furthermore, the enzymatic activity and stability are also improved due to the large surface area of nanomaterials. Here we report the conjugation of glucose oxidase (GOD) onto phosphorescent Mn-doped ZnS quantum dots (QDs) using 1-ethyl-3-(3-dimethylaminopropy)carbodiimide (EDC)/N-hydroxysuccinimide (NHS) as coupling reagents for glucose biosensing based on the effective quenching of the room temperature phosphorescence (RTP) of Mn-doped ZnS QDs by the H(2)O(2) generated from GOD-catalyzed oxidation of glucose. The obtained bioconjugate not only provided improved enzymatic performance with Michaelis-Menten constant of 0.70 mM but also favored biological applications because the phosphorescent detection mode avoided the interference from autofluorescence and scattering light from the biological matrix. In addition, the GOD-conjugated Mn-doped ZnS QDs showed better thermal stability in the temperature range of 20-80 degrees C. The GOD-Mn-doped ZnS QDs based RTP sensor for glucose gave a detection limit of 3 microM and two linear ranges from 10 microM to 0.1 mM and from 0.1 to 1 mM. The developed biosensor was successfully applied to the determination of glucose in real serum samples without the need for any complicated sample pretreatments. PMID:20092317

  11. Photoabsorption and resonance energy transfer phenomenon in CdTe-protein bioconjugates: an insight into QD-biomolecular interactions.

    PubMed

    Vinayaka, Aaydha C; Thakur, Munna S

    2011-05-18

    Luminescent quantum dots (QDs) possess unique photophysical properties, which are advantageous in the development of new generation robust fluorescent probes based on Forster resonance energy transfer (FRET) phenomena. Bioconjugation of these QDs with biomolecules create hybrid materials having unique photophysical properties along with biological activity. The present study is aimed at characterizing QD bioconjugates in terms of optical behavior. Colloidal CdTe QDs capped with 3-mercaptopropionic acid (MPA) were conjugated to different proteins by the carbodiimide protocol using N-(3-dimethylaminopropyl)-N-ethylcarbodiimide hydrochloride (EDC) and a coupling reagent like N-hydroxysuccinimide (NHS). The photoabsorption of these QD-protein bioconjugates demonstrated an effective coupling of electronic orbitals of constituents. A linear variation in absorbance of bioconjugates at 330 nm proportionate to conjugation suggests a covalent attachment as confirmed by gel electrophoresis. A red shift in the fluorescence of bovine serum albumin (BSA) due to conjugation inferred a decrease in Stokes shift and solvent polarization effects on protein. A proportionate quenching in BSA fluorescence followed by an enhancement of QD fluorescence point toward nonradiative dipolar interactions. Further, reduction in photobleaching of BSA suggests QD-biomolecular interactions. Bioconjugation has significantly influenced the photoabsorption spectrum of QD bioconjugates suggesting the formation of a possible protein shell on the surface of QD. The experimental result suggests that these bioconjugates can be considered nanoparticle (NP) superstructures for the development of a new generation of robust nanoprobes. PMID:21452896

  12. Optimized multimodal nanoplatforms for targeting α(v)β3 integrins.

    PubMed

    Bolley, Julie; Lalatonne, Yoann; Haddad, Oualid; Letourneur, Didier; Soussan, Michael; Pérard-Viret, Joelle; Motte, Laurence

    2013-12-01

    Magnetic Resonance Imaging (MRI) using contrast agents is a very powerful technique for diagnosis in clinical medicine and biomedical research. The synthesis of ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles targeting αvβ3 integrins and acting as new MRI contrast agents seems to be a promising way for cancer diagnosis. Indeed, it is well established that αvβ3 integrin plays a key role in tumor angiogenesis acting like a receptor for the extracellular matrix proteins like vitronectin, fibronectin through the arginine-glycine-aspartic acid (RGD) sequence. Up-regulation of αvβ3 has been found to be associated with a wide range of cancers, making it a broad-spectrum tumor-marker. In this study, USPIO nanocrystals were synthesized and surface passivated with caffeic acid. The large number of the carboxylic acid functions at the outer surface of the nanoplatforms was used for the covalent coupling of Rhodamine123, polyethylene glycol (PEG) and cyclic RGD. Soluble carbodiimide (EDC) and N-hydroxysuccinimide (NHS) were used to crosslink carboxylic acid with the amino group of the ligands. We examined the design of the nanoplatforms with each individual entity and then the combination of two and three of them. Several methods were used to characterize the nanoparticle surface functionalization and the magnetic properties of these contrast agents were studied using a 1.5 T clinical MRI scanner. The affinity towards integrins was evidenced by surface plasmon resonance and solid-phase receptor-binding assay. PMID:24154564

  13. Metal-organic coordination-enabled layer-by-layer self-assembly to prepare hybrid microcapsules for efficient enzyme immobilization.

    PubMed

    Wang, Xiaoli; Jiang, Zhongyi; Shi, Jiafu; Liang, Yanpeng; Zhang, Chunhong; Wu, Hong

    2012-07-25

    A novel layer-by-layer self-assembly approach enabled by metal-organic coordination was developed to prepare polymer-inorganic hybrid microcapsules. Alginate was first activated via N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxy succinimide (NHS) coupling chemistry, and subsequently reacted with dopamine. Afterward, the dopamine modified alginate (Alg-DA) and titanium(IV) bis(ammonium lactato) dihydroxide (Ti(IV)) were alternatively deposited onto CaCO3 templates. The coordination reaction between the catechol groups of Alg-DA and the Ti(IV) allowed the alternative assembly to form a series of multilayers. After removing the templates, the alginate-titanium hybrid microcapsules were obtained. The high mechanical stability of hybrid microcapsules was demonstrated by osmotic pressure experiment. Furthermore, the hybrid microcapsules displayed superior thermal stability due to Ti(IV) coordination. Catalase (CAT) was used as model enzyme, either encapsulated inside or covalently attached on the surface of the resultant microcapsules. No CAT leakage from the microcapsules was detected after incubation for 48 h. The encapsulated CAT, with a loading capacity of 450-500 mg g(-1) microcapsules, exhibited desirable long-term storage stability, whereas the covalently attached CAT, with a loading capacity of 100-150 mg g(-1) microcapsules, showed desirable operational stability. PMID:22724538

  14. GlyHisGlyHis immobilization on silicon surface for copper detection

    NASA Astrophysics Data System (ADS)

    Sam, Sabrina; Gouget-Laemmel, Anne Chantal; Chazalviel, Jean-Noël; Ozanam, François; Gabouze, Noureddine

    2013-03-01

    Hybrid nanomaterials based on organic layer covalently grafted on semi-conductor surfaces appear as promising systems for innovative applications, especially in sensor field. In this work, we focused on the functionalization of silicon surface by the peptide GlyHisGlyHis, which forms stable complexes with metal ions. This property is exploited to achieve heavy metals recognition in solution. The immobilization was achieved using multi-step reactions: GlyHisGlyHis was anchored on a previously prepared carboxyl-terminated silicon surface using N-ethyl-N'-(3-dimethylaminopropyl)-carbodiimide (EDC)/N-hydroxysuccinimide (NHS) coupling agents. This scheme is compatible with the mild conditions required for preserving the probe activity of the peptide. At each step of the functionalization, the surface was monitored by infrared spectroscopy Fourier transform (FTIR) in ATR (attenuated total reflexions) geometry and by atomic force microscopy (AFM). Electrochemical behaviour of such prepared electrodes was carried out in the presence of copper ions by means of cyclic voltammetry. The recorded cyclic voltammograms showed a surface reversible process corresponding to the Cu2+/Cu+ couple in the complex Cu-GlyHisGlyHis immobilized on the silicon surface. Copper ions concentrations down than μM where detected. These results demonstrate the potential role of peptide-modified silicon electrodes in developing strategies for simple and fast detection of toxic metals in solution.

  15. Investigation of terpinolene + ozone or terpinolene + nitrate radical reaction products using denuder/filter apparatus

    PubMed Central

    Harrison, Joel C.; Wells, J.R.

    2015-01-01

    Terpinolene’s (1-methyl-4-(propan-2-ylidene)cyclohexene) reaction with ozone or the nitrate radical was investigated using a denuder/filter apparatus in order to characterize gas-phase and particulate reaction products. Identification of the reaction products (i.e., aldehydes, ketones, dicarbonyls and carboxylic acids) was made using two derivatization methods; O-(2,3,4,5,6-pentafluorobenzyl)hydroxylamine (PFBHA) to derivatize the carbonyl products or 3-Ethyl-1-[3-(dimethylamino)propyl]carbodiimide hydrochloride (EDC) and 2,2,2-trifluoroethylamine hydrochloride (TFEA) to derivatize the carboxylic acid products. Proposed carbonyl products for ozonolysis of terpinolene are: 4-methylcyclohex-3-en-1-one, 2-hydroxy-4-methylcyclohex-3-en-1-one, glyoxal, methyl glyoxal, 3-oxobutanal, and 6-oxo-3-(propan-2-ylidene)heptanal. Proposed carbonyl products for nitrate radical reaction of terpinolene are: 2-hydroxy-4-methylcyclohex-3-en-1-one, glyoxal, methyl glyoxal, and 4-oxopentanal. No carboxylic acid products were detected with either oxidizing reactant. PMID:26527171

  16. An improved method for covalently conjugating morpholino oligomers to antitumor antibodies.

    PubMed

    He, Jiang; Liu, Guozheng; Dou, Shuping; Gupta, Suresh; Rusckowski, Mary; Hnatowich, Donald

    2007-01-01

    Whether for conventional pretargeting, amplification pretargeting, or affinity enhancement pretargeting, it will be necessary to conjugate an antitumor antibody as the first injectate. This laboratory is investigating phosphorodiamidate morpholinos (MORFs) for pretargeting, and accordingly we are examining methods of attaching MORFs to antitumor antibodies that provide at least one group per molecule (gpm) without adversely influencing antibody properties. The aim of this investigation was to evaluate the commercial Hydralink for the conjugation of the anti-CEA MN14 antibody with an 18 mer amine-derivatized MORF. The conjugation was approached in both directions by first reacting MN14 with the NHS derivatives of 4-hydrozinonicotinate acetone hydrazone (SANH) or 4-formylbenzoate (SFB) and then combining with MORF that was previously reacted with SFB or SANH to yield MN14(SANH)-MORF and MN14(SFB)-MORF respectively. The storage stability, immunoreactive fraction, and the biodistribution in normal mice were compared for both conjugates. Thereafter, MN14(SANH)-MORF was used in a pretargeting study in tumored nude mice, and the results were compared to that obtained historically with MN14-MORF prepared by carbodiimide (EDC) coupling. Both new methods of conjugation provided between 1 and 2 gpm compared to 0.2 achieved previously by EDC. Furthermore, by repeat SE HPLC with and without CEA, both showed an unimpaired immunoreactive fraction. MN14(SANH)-MORF tolerated long-term storage best. More importantly, when labeled by hybridization with 99mTc-labeled complementary MORF (99mTc-cMORF), the biodistribution of MN14(SANH)-MORF was more favorable than that of MN14(SFB)-MORF in normal mice with lower liver (5.7 vs 9.4 %ID/g at 18 h) and spleen (3.5 vs 8.4 %ID/g) accumulations and higher blood levels (4.8 vs 3.4 %ID/g). Accordingly, only MN14(SANH)-MORF was used in a pretargeting study in tumored mice. When targeted with 99mTc-cMORF and at 2 days postinjection of antibody

  17. Interface control document between the NASA Goddard Space Flight Center (GSFC) and Department of Interior EROS Data Center (EDC) for LANDSAT-D. Partially processed multispectral scanner High Density Tape (HDT-AM)

    NASA Technical Reports Server (NTRS)

    1982-01-01

    The format of the HDT-AM product which contains partially processed LANDSAT D and D Prime multispectral scanner image data is defined. Recorded-data formats, tape format, and major frame types are described.

  18. EDSP Tier 2 test (T2T) guidances and protocols are delivered, including web-based guidance for diagnosing and scoring, and evaluating EDC-induced pathology in fish and amphibian

    EPA Science Inventory

    The Agency’s Endocrine Disruptor Screening Program (EDSP) consists of two tiers. The first tier provides information regarding whether a chemical may have endocrine disruption properties. Tier 2 tests provide confirmation of ED effects and dose-response information to be us...

  19. Interface control document between the NASA Goddard Space Flight Center (GSFC) and Department of Interior EROS Data Center (EDC) for LANDSAT-D. Thematic mapper high resolution 241 mm film

    NASA Technical Reports Server (NTRS)

    1982-01-01

    The 241 mm photographic product produced by the Goddard Space Flight Center Data Management System for LANDSAT-D is described. Film type and format, image dimensions, frame ID, gray scale, resolution patterns, registration marks, etc. are addressed.

  20. Fabrication and Characterization of Spongy Denuded Amniotic Membrane Based Scaffold for Tissue Engineering

    PubMed Central

    Taghiabadi, Ehsan; Nasri, Sima; Shafieyan, Saeed; Jalili Firoozinezhad, Sasan; Aghdami, Nasser

    2015-01-01

    Objective As a biological tissue material, amniotic membrane (AM) has low immunogenicity and to date has been widely adopted in clinical practice. However, some features such as low biomechanical consistency and rapid biodegradation is limited the application of AM. Therefore, in this study, we fabricated a novel three-dimensional (3D) spongy scaffold made of the extracellular matrix (ECM) of denuded AM. Due to their unique characteristics which are similar to the skin, these scaffolds can be considered as an alternative option in skin tissue engineering. Materials and Methods In this experimental study, cellular components of human amniotic membrane (HAM) were removed with 0.03% (w/v) sodium dodecyl sulphate (SDS). Quantitative analysis was performed to determine levels of Glycosaminoglycans (GAGs), collagen, and deoxyribonucleic acid (DNA). To increase the low efficiency and purity of the ECM component, especially collagen and GAG, we applied an acid solubilization procedure hydrochloridric acid (HCl 0.1 M) with pepsin (1 mg/ml). In the present experiment 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC)/N-hydroxysuccinimide (NHS) cross linker agent was used to improve the mechanical properties of 3D lyophilized AM scaffold. The spongy 3D AM scaffolds were specified, by scanning electron microscopy, hematoxylin and eosin (H&E) staining, a swelling test, and mechanical strength and in vitro biodegradation tests. Human fetal fibroblast culture systems were used to establish that the scaffolds were cytocompatible. Results Histological analysis of treated human AM showed impressive removal of cellular components. DNA content was diminished after treatment (39 ± 4.06 μg/ml vs. 341 ± 29.60 μg/ml). Differences were observed between cellular and denude AM in matrix collagen (478 ± 18.06 μg/mg vs. 361 ± 27.47 μg/mg).With the optimum concentration of 1 mM NHS/EDC ratio1:4, chemical cross-linker agent could significantly increase the mechanical

  1. Multi-component nanofibrous scaffolds with tunable properties for bone tissue engineering

    NASA Astrophysics Data System (ADS)

    Jose, Moncy V.

    Bone is a highly complex tissue which is an integral part of vertebrates and hence any damage has a major negative effect on the quality of life. Tissue engineering is regarded as an ideal route to resolve the issues related to the scarcity of tissue and organ for transplantation. Apart from cell line and growth factors, the choice of materials and fabrication technique for scaffold are equally important. The goal of this work was to develop a multi-component nanofibrous scaffold based on a synthetic polymer (poly(lactic-co-glycolide) (PLGA)), a biopolymer (collagen) and a biomineral (nano-hydroxyapatite (nano-HA)) by electrospinning technique, which mimics the nanoscopic, chemical, and anisotropic features of bone. Preliminary studies involved fabrication of nanocomposite scaffolds based on PLGA and nano-HA. Morphological and mechanical characterizations revealed that at low concentrations, nano-HA acted as reinforcements, whereas at higher concentrations the presence of aggregation was detrimental to the scaffold. Hydrolytic degradation studies revealed the scaffold had a little mass loss and the mechanical property was maintained for a period of 6 weeks. This study was followed by evaluation of a blend system based on PLGA and collagen. Collagen addition provides hydrophilicity and the necessary cell binding sites in PLGA. The structural characterization revealed that the blend had limited interactions between the two components. The mechanical characterization revealed that with increasing collagen concentration, there was a decline in mechanical properties. However, crosslinking of the blend system, with carbodiimide (EDC) resulted in improving the mechanical properties of the scaffolds. A multi-component system was developed by adding different concentrations of nano-HA to a fixed PLGA/collagen blend composition (80/20). Morphological and mechanical characterizations revealed properties similar to the PLGA/HA system. Cyto-compatibility studies revealed

  2. Optimized multimodal nanoplatforms for targeting αvβ3 integrins

    NASA Astrophysics Data System (ADS)

    Bolley, Julie; Lalatonne, Yoann; Haddad, Oualid; Letourneur, Didier; Soussan, Michael; Pérard-Viret, Joelle; Motte, Laurence

    2013-11-01

    Magnetic Resonance Imaging (MRI) using contrast agents is a very powerful technique for diagnosis in clinical medicine and biomedical research. The synthesis of ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles targeting αvβ3 integrins and acting as new MRI contrast agents seems to be a promising way for cancer diagnosis. Indeed, it is well established that αvβ3 integrin plays a key role in tumor angiogenesis acting like a receptor for the extracellular matrix proteins like vitronectin, fibronectin through the arginine-glycine-aspartic acid (RGD) sequence. Up-regulation of αvβ3 has been found to be associated with a wide range of cancers, making it a broad-spectrum tumor-marker. In this study, USPIO nanocrystals were synthesized and surface passivated with caffeic acid. The large number of the carboxylic acid functions at the outer surface of the nanoplatforms was used for the covalent coupling of Rhodamine123, polyethylene glycol (PEG) and cyclic RGD. Soluble carbodiimide (EDC) and N-hydroxysuccinimide (NHS) were used to crosslink carboxylic acid with the amino group of the ligands. We examined the design of the nanoplatforms with each individual entity and then the combination of two and three of them. Several methods were used to characterize the nanoparticle surface functionalization and the magnetic properties of these contrast agents were studied using a 1.5 T clinical MRI scanner. The affinity towards integrins was evidenced by surface plasmon resonance and solid-phase receptor-binding assay.Magnetic Resonance Imaging (MRI) using contrast agents is a very powerful technique for diagnosis in clinical medicine and biomedical research. The synthesis of ultrasmall superparamagnetic iron oxide (USPIO) nanoparticles targeting αvβ3 integrins and acting as new MRI contrast agents seems to be a promising way for cancer diagnosis. Indeed, it is well established that αvβ3 integrin plays a key role in tumor angiogenesis acting like a receptor for

  3. Exploitation of lectinized lipo-polymerosome encapsulated Amphotericin B to target macrophages for effective chemotherapy of visceral leishmaniasis.

    PubMed

    Gupta, Pramod K; Asthana, Shalini; Jaiswal, Anil K; Kumar, Vivek; Verma, Ashwni K; Shukla, Prashant; Dwivedi, Pankaj; Dube, Anuradha; Mishra, Prabhat R

    2014-06-18

    We have designed lectin functionalized Lipo-polymerosome bearing Amphotericin B (Lec-AmB-L-Psome) for specific internalization via lectin receptors overexpressed on infected macrophages of mononuclear phagocytic system (MPS) for the effective management of intramacrophage diseases such as visceral leishmaniasis. The lipo-polymerosome composed of glycol chitosan-stearic acid copolymer (GC-SA25%) and model lipid cholesterol was surface-functionalized with lectin by the EDC/NHS carbodiimide coupling method. Our designed Lec-AmB-L-Psome showed >2-fold enhanced uptake and significantly higher internalization in macrophages as compared to AmB-L-Psome. Importantly, pharmacokinetic and organ distribution studies illustrate significantly higher accumulation of Lec-AmB-L-Psome in MPS especially in liver, spleen, and lung as compared to AmB-L-Psome, Ambisome, and Fungizone. The IC50 value demonstrated that Lec-AmB-L-Psome has 1.63, 2.23, and 3.43 times higher activity than AmB-L-Psome (p < 0.01), Ambisome (p < 0.05), and Fungizone (p < 0.05), respectively. Additionally, the Lec-AmB-L-Psome showed significantly higher splenic parasite inhibition (78.66 ± 3.08%) compared to Fungizone and Ambisome that caused only 56.54 ± 3.91% (p < 0.05) and 66.46 ± 2.08% (p < 0.05) parasite inhibition, respectively, in Leishmania-infected hamsters. The toxicity profile revealed that Lec-AmB-L-Psome is a safe delivery system with diminished nephrotoxicity which is a limiting factor of Fungizone application. Taken together, these studies suggest that this surface functionalized self-assembled Lec-AmB-L-Psome can introduce a new platform to specifically target macrophages for effective management of intramacrophage diseases. PMID:24842628

  4. Intra-articular delivery of kartogenin-conjugated chitosan nano/microparticles for cartilage regeneration.

    PubMed

    Kang, Mi Lan; Ko, Ji-Yun; Kim, Ji Eun; Im, Gun-Il

    2014-12-01

    We developed an intra-articular (IA) drug delivery system to treat osteoarthritis (OA) that consisted of kartogenin conjugated chitosan (CHI-KGN). Kartogenin, which promotes the selective differentiation of mesenchymal stem cells (MSCs) into chondrocytes, was conjugated with low-molecular-weight chitosan (LMWCS) and medium-molecular-weight chitosan (MMWCS) by covalent coupling of kartogenin to each chitosan using an ethyl(dimethylaminopropyl) carbodiimide (EDC)/N-hydroxysuccinimide (NHS) catalyst. Nanoparticles (NPs, 150 ± 39 nm) or microparticles (MPs, 1.8 ± 0.54 μm) were fabricated from kartogenin conjugated-LMWCS and -MMWCS, respectively, by an ionic gelation using tripolyphosphate (TPP). The in vitro release profiles of kartogenin from the particles showed sustained release for 7 weeks. When the effects of the CHI-KGN NPs or CHI-KGN MPs were evaluated on the in vitro chondrogenic differentiation of human bone marrow MSCs (hBMMSCs), the CHI-KGN NPs and CHI-KGN MPs induced higher expression of chondrogenic markers from cultured hBMMSCs than unconjugated kartogenin. In particular, hBMMSCs treated with CHI-KGN NPs exhibited more distinct chondrogenic properties in the long-term pellet cultures than those treated with CHI-KGN MPs. The in vivo therapeutic effects of CHI-KGN NPs or CHI-KGN MPs were investigated using a surgically-induced OA model in rats. The CHI-KGN MPs showed longer retention time in the knee joint than the CHI-KGN NPs after IA injection in OA rats. The rats treated with CHI-KGN NPs or CHI-KGN MPs by IA injection showed much less degenerative changes than untreated control or rats treated with unconjugated kartogenin. In conclusion, CHI-KGN NPs or CHI-KGN MPs can be useful polymer-drug conjugates as an IA drug delivery system to treat OA. PMID:25241157

  5. In situ detection of salicylic acid binding sites in plant tissues.

    PubMed

    Liu, Jing-Wen; Deng, Da-Yi; Yu, Ying; Liu, Fang-Fei; Lin, Bi-Xia; Cao, Yu-Juan; Hu, Xiao-Gang; Wu, Jian-Zhong

    2015-02-01

    The determination of hormone-binding sites in plants is essential in understanding the mechanisms behind hormone function. Salicylic acid (SA) is an important plant hormone that regulates responses to biotic and abiotic stresses. In order to label SA-binding sites in plant tissues, a quantum dots (QDs) probe functionalized with a SA moiety was successfully synthesized by coupling CdSe QDs capped with 3-mercaptopropionic acid (MPA) to 4-amino-2-hydroxybenzoic acid (PAS), using 1-ethyl-3-(3-dimethyllaminopropyl) carbodiimide (EDC) as the coupling agent. The probe was then characterized by dynamic light scattering and transmission electron microscopy, as well as UV/vis and fluorescence spectrophotometry. The results confirmed the successful conjugation of PAS to CdSe QDs and revealed that the conjugates maintained the properties of the original QDs, with small core diameters and adequate dispersal in solution. The PAS-CdSe QDs were used to detect SA-binding sites in mung bean and Arabidopsis thaliana seedlings in vitro and in vivo. The PAS-CdSe QDs were effectively transported into plant tissues and specifically bound to SA receptors in vivo. In addition, the effects of the PAS-CdSe QDs on cytosolic Ca(2+) levels in the tips of A. thaliana seedlings were investigated. Both SA and PAS-CdSe QDs had similar effects on the trend in cytosolic-free Ca(2+) concentrations, suggesting that the PAS-CdSe QDs maintained the bioactivity of SA. To summarize, PAS-CdSe QDs have high potential as a fluorescent probe for the in vitro/in vivo labeling and imaging of SA receptors in plants. PMID:24833131

  6. Effect of Surgical Technique on Corneal Implant Performance

    PubMed Central

    Ljunggren, Monika Kozak; Elizondo, Rodolfo A.; Edin, Joel; Olsen, David; Merrett, Kimberley; Lee, Chyan-Jang; Salerud, Göran; Polarek, James; Fagerholm, Per; Griffith, May

    2014-01-01

    Purpose Our aim was to determine the effect of a surgical technique on biomaterial implant performance, specifically graft retention. Methods Twelve mini pigs were implanted with cell-free, 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC)/N-hydroxysuccinimide (NHS) cross-linked recombinant human collagen type III (RHCIII) hydrogels as substitutes for donor corneal allografts using overlying sutures with or without human amniotic membrane (HAM) versus interrupted sutures with HAM. The effects of the retention method were compared as well as the effects of collagen concentration (13.7% to 15% RHCIII). Results All implanted corneas showed initial haze that cleared with time, resulting in corneas with optical clarity matching those of untreated controls. Biochemical analysis showed that by 12 months post operation, the initial RHCIII implants had been completely remodeled, as type I collagen, was the major collagenous protein detected, whereas no RHCIII could be detected. Histological analysis showed all implanted corneas exhibited regeneration of epithelial and stromal layers as well as nerves, along with touch sensitivity and tear production. Most neovascularization was seen in corneas stabilized by interrupted sutures. Conclusions This showed that the surgical technique used does have a significant effect on the overall performance of corneal implants, overlying sutures caused less vascularization than interrupted sutures. Translational Relevance Understanding the significance of the suturing technique can aid the selection of the most appropriate procedure when implanting artificial corneal substitutes. The same degree of regeneration, despite a higher collagen content indicates that future material development can progress toward stronger, more resistant implants. PMID:24749003

  7. Electrochemical label-free and reagentless genosensor based on an ion barrier switch-off system for DNA sequence-specific detection of the avian influenza virus.

    PubMed

    Kurzątkowska, Katarzyna; Sirko, Agnieszka; Zagórski-Ostoja, Włodzimierz; Dehaen, Wim; Radecka, Hanna; Radecki, Jerzy

    2015-10-01

    This paper concerns the development of genosensors based on redox-active monolayers incorporating (dipyrromethene)2Cu(II) and (dipyrromethene)2Co(II) complexes formed step by step on a gold electrode surface. They were applied for electrochemical determination of oligonucleotide sequences related to avian influenza virus (AIV) type H5N1. A 20-mer probe (NH2-NC3) was covalently attached to the gold electrode surface via a reaction performed in the presence of ethyl(dimethylaminopropyl)carbodiimide / N-hydroxysuccinimide (EDC/NHS) between the amine group present in the probe and carboxylic groups present on the surface of the redox-active layer. Each modification step has been controlled with Osteryoung square-wave voltammetry. The genosensor incorporating the (dipyrromethene)2Cu(II) complex was able to detect a fully complementary single-stranded DNA target with a detection limit of 1.39 pM. A linear dynamic range was observed from 1 to 10 pM. This genosensor displays good discrimination between three single-stranded DNA targets studied: fully complementary, partially complementary (with only six complementary bases), and totally noncomplementary to the probe. When the (dipyrromethene)2Co(II) complex was applied, a detection limit of 1.28 pM for the fully complementary target was obtained. However, this genosensor was not able to discriminate partially complementary and totally noncomplementary oligonucleotide sequences to the probe. Electrochemical measurements, using both types of genosensors in the presence of different supporting electrolytes, were performed in order to elaborate a new mechanism of analytical signal generation based on an ion barrier "switch-off" system. PMID:26359972

  8. NeutrAvidin Functionalization of CdSe/CdS Quantum Nanorods and Quantification of Biotin Binding Sites using Biotin-4-Fluorescein Fluorescence Quenching.

    PubMed

    Lippert, Lisa G; Hallock, Jeffrey T; Dadosh, Tali; Diroll, Benjamin T; Murray, Christopher B; Goldman, Yale E

    2016-03-16

    We developed methods to solubilize, coat, and functionalize with NeutrAvidin elongated semiconductor nanocrystals (quantum nanorods, QRs) for use in single molecule polarized fluorescence microscopy. Three different ligands were compared with regard to efficacy for attaching NeutrAvidin using the "zero-length cross-linker" 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC). Biotin-4-fluorescene (B4F), a fluorophore that is quenched when bound to avidin proteins, was used to quantify biotin binding activity of the NeutrAvidin coated QRs and biotin binding activity of commercially available streptavidin coated quantum dots (QDs). All three coating methods produced QRs with NeutrAvidin coating density comparable to the streptavidin coating density of the commercially available quantum dots (QDs) in the B4F assay. One type of QD available from the supplier (ITK QDs) exhibited ∼5-fold higher streptavidin surface density compared to our QRs, whereas the other type of QD (PEG QDs) had 5-fold lower density. The number of streptavidins per QD increased from ∼7 streptavidin tetramers for the smallest QDs emitting fluorescence at 525 nm (QD525) to ∼20 tetramers for larger, longer wavelength QDs (QD655, QD705, and QD800). QRs coated with NeutrAvidin using mercaptoundecanoicacid (MUA) and QDs coated with streptavidin bound to biotinylated cytoplasmic dynein in single molecule TIRF microscopy assays, whereas Poly(maleic anhydride-alt-1-ocatdecene) (PMAOD) or glutathione (GSH) QRs did not bind cytoplasmic dynein. The coating methods require optimization of conditions and concentrations to balance between substantial NeutrAvidin binding vs tendency of QRs to aggregate and degrade over time. PMID:26722835

  9. Effect of cross-linking reagents for hyaluronic acid hydrogel dermal fillers on tissue augmentation and regeneration.

    PubMed

    Yeom, Junseok; Bhang, Suk Ho; Kim, Byung-Soo; Seo, Moo Seok; Hwang, Eui Jin; Cho, Il Hwan; Park, Jung Kyu; Hahn, Sei Kwang

    2010-02-17

    A novel, biocompatible, and nontoxic dermal filler using hyaluronic acid (HA) hydrogels was successfully developed for tissue augmentation applications. Instead of using highly reactive cross-linkers such as divinyl sulfone (DVS) for Hylaform, 1,4-butanediol diglycidyl ether (BDDE) for Restylane, and 1,2,7,8-diepoxyoctane (DEO) for Puragen, HA hydrogels were prepared by direct amide bond formation between the carboxyl groups of HA and hexamethylenediamine (HMDA) with an optimized carboxyl group modification for effective tissue augmentation. The HA-HMDA hydrogels could be prepared within 5 min by the addition of HMDA to HA solution activated with 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC) and 1-hydroxybenzotriazole monohydrate (HOBt). Five kinds of samples, a normal control, a negative control, a positive control of Restylane, adipic acid dihydrazide grafted HA (HA-ADH) hydrogels, and HA-HMDA hydrogels, were subcutaneously injected to wrinkled model mice. According to the image analysis on dorsal skin augmentation, the HA-HMDA hydrogels exhibited the best tissue augmentation effect being stable longer than 3 months. Furthermore, histological analyses after hematoxylin-eosin (H&E) and Masson's trichrome staining revealed the excellent biocompatibility and safety of HA-HMDA hydrogels. The dermal thickness and the dermal collagen density in wrinkled mice after treatment with HA-HMDA hydrogels for 12 weeks were comparable to those of normal mice. Compared with HA-DVS hydrogels and Restylane, the excellent tissue augmentation by HA-HMDA hydrogels might be ascribed to the biocompatible residues of amine groups in the cross-linker of HMDA. The HA-HMDA hydrogels will be investigated further as a novel dermal filler for clinical applications. PMID:20078098

  10. Evidence of Liquid Crystal-Assisted Abiotic Ligation of Nucleic Acids

    NASA Astrophysics Data System (ADS)

    Fraccia, Tommaso P.; Zanchetta, Giuliano; Rimoldi, Valeria; Clark, Noel A.; Bellini, Tommaso

    2015-06-01

    The emergence of early life must have been marked by the appearance in the prebiotic era of complex molecular structures and systems, motivating the investigation of conditions that could not only facilitate appropriate chemical synthesis, but also provide the mechanisms of molecular selection and structural templating necessary to pilot the complexification toward specific molecular patterns. We recently proposed and demonstrated that these functions could be afforded by the spontaneous ordering of ultrashort nucleic acids oligomers into Liquid Crystal (LC) phases. In such supramolecular assemblies, duplex-forming oligomers are held in average end-to-end contact to form chemically discontinuous but physically continuous double helices. Using blunt ended duplexes, we found that LC formation could both provide molecular selection mechanisms and boost inter-oligomer ligation. This paper provides an essential extension to this notion by investigating the catalytic effects of LC ordering in duplexes with mutually interacting overhangs. Specifically, we studied the influence of LC ordering of 5'-hydroxy-3'-phosphate partially self-complementary DNA 14mers with 3'-CG sticky-ends, on the efficiency of non-enzymatic ligation reaction induced by water-soluble carbodiimide EDC as condensing agent. We investigated the ligation products in mixtures of DNA with poly-ethylene glycol (PEG) at three PEG concentrations at which the system phase separates creating DNA-rich droplets that organize into isotropic, nematic LC and columnar LC phases. We observe remarkable LC-enhanced chain lengthening, and we demonstrate that such lengthening effectively promotes and stabilizes LC domains, providing the kernel of a positive feedback cycle by which LC ordering promotes elongation, in turn stabilizing the LC ordering.

  11. Synthesis and psychobiological evaluation of modafinil analogs in mice

    PubMed Central

    2013-01-01

    Background and the purpose of the study Modafinil, a novel wake-promoting agent with low potential for abuse and dependence, has a reliable structure to find some novel derivatives with better activity and lower potential for abuse and risk of dependency. This study was designed to evaluate psychobiological activity of some novel N-aryl modafinil derivatives. Methods Seven novel N-aryl modafinil derivatives were synthesized through three reactions: a) preparation of benzhydrylsulfanyl acetic acid through reaction of benzhydrol with thioglycolic acid, b) formation of desired amide by adding the substituted aniline to activated acid with EDC (1-ethyl-3-(3-dimethyl amino propyl) carbodiimide). This reaction was catalyzed by HOBt (N- hydroxylbenzotriazole), and c) oxidation of sulfur to sulfoxide group with H2O2. Then, their psychobiological effect on the performance of male albino mice were compared to that of modafinil as following: wakefulness by determining the effects of derivatives on phenobarbital-induced loss of the righting reflex (LOPR); exploratory activity by measuring activity in the open field test (OFT); depression by measuring immobility time (IT) during forced swimming test (FST) and the anxiogenic and anxiolytic like effects by using elevated plus-maze test (EPM). All tests were videotaped and analyzed for the frequency and duration of the behaviors during the procedures. Conclusions 2-(Benzhydrylsulfonyl)-N-(4-chlorophenyl)acetamide (4c) showed comparable result in LOPR test. However, all analogs were found to be stimulant except 2-(benzhydrylsulfinyl)-N-phenylacetamide (4a). Also 4c led the most exploratory activity in mice among derivatives. FST results showed that 4a had the longest IT while modafinil, 2-(benzhydrylsulfinyl)-N-(3-chlorophenyl) acetamide (4b) and 2-(benzhydrylsulfinyl)-N-(4-ethylphenyl) acetamide (4d) had the shortest IT. In EPM, all derivatives showed anxiogenic-like behavior since they decreased open arms time and open arms

  12. The Competing Effects of Hyaluronic and Methacrylic Acid in Model Contact Lenses.

    PubMed

    Weeks, Andrea; Subbaraman, Lakshman N; Jones, Lyndon; Sheardown, Heather

    2012-01-01

    The aim of this study was to determine the influence of hyaluronic acid (HA) on lysozyme sorption in model contact lenses containing varying amounts of methacrylic acid (MAA). One model conventional hydrogel (poly(2-hydroxyethyl methacrylate) (pHEMA)) and two model silicone hydrogels (pHEMA, methacryloxypropyltris(trimethylsiloxy)silane (pHEMA TRIS) and N,N-dimethylacrylamide, TRIS (DMAA TRIS)) lens materials were prepared with and without MAA at two different concentrations (1.7 and 5%). HA, along with dendrimers, was loaded into these model contact lens materials and then cross-linked with 1-ethyl-3-(3-dimethylamino propyl)-carbodiimide (EDC). Equilibrium water content (EWC), advancing water contact angle and lysozyme sorption on these lens materials were investigated. In the HA-containing materials, the presence (P < 0.05) and amount (P < 0.05) of MAA increased the EWC of the materials. For most materials, addition of MAA reduced the advancing contact angles (P < 0.05) and for all the materials, the addition of HA further improved hydrophilicity (P < 0.05). For the non-HA containing hydrogels, the presence (P < 0.05) and amount (P < 0.05) of MAA increased lysozyme sorption. The presence of HA decreased lysozyme sorption for all materials (P < 0.05). MAA appears to work synergistically with HA to increase the EWC in addition to improving the hydrophilicity of model pHEMA-based and silicone hydrogel contact lens materials. Hydrogel materials that contain HA have tremendous potential as hydrophilic, protein-resistant contact lens materials. PMID:21477462

  13. An electrochemical sulfite biosensor based on gold coated magnetic nanoparticles modified gold electrode.

    PubMed

    Rawal, Rachna; Chawla, Sheetal; Pundir, Chandra Shekhar

    2012-01-15

    A sulfite oxidase (SO(X)) (EC 1.8.3.1) purified from Syzygium cumini leaves was immobilized onto carboxylated gold coated magnetic nanoparticles (Fe(3)O(4)@GNPs) electrodeposited onto the surface of a gold (Au) electrode through N-ethyl-N'-(3-dimethylaminopropyl) carbodiimide (EDC)-N-hydroxy succinimide (NHS) chemistry. An amperometric sulfite biosensor was fabricated using SO(X)/Fe(3)O(4)@GNPs/Au electrode as working electrode, Ag/AgCl as standard and Pt wire as auxiliary electrode. The working electrode was characterized by Fourier Transform Infrared (FTIR) Spectroscopy, Cyclic Voltammetry (CV), Scanning Electron Microscopy (SEM) and Electrochemical Impedance Spectroscopy (EIS) before and after immobilization of SO(X). The biosensor showed optimum response within 2s when operated at 0.2V (vs. Ag/AgCl) in 0.1 M Tris-HCl buffer, pH 8.5 and at 35 °C. Linear range and detection limit were 0.50-1000 μM and 0.15 μM (S/N=3) respectively. Biosensor was evaluated with 96.46% recovery of added sulfite in red wine and 1.7% and 3.3% within and between batch coefficients of variation respectively. Biosensor measured sulfite level in red and white wines. There was good correlation (r=0.99) between red wines sulfite value by standard DTNB (5,5'-dithio-bis-(2-nitrobenzoic acid)) method and the present method. Enzyme electrode was used 300 times over a period of 4 months, when stored at 4 °C. Biosensor has advantages over earlier biosensors that it has excellent electrocatalysis towards sulfite, lower detection limit, higher storage stability and no interference by ascorbate, cysteine, fructose and ethanol. PMID:22035973

  14. Spectroscopy and microscopy characterization of spiropyran auto-assemble device

    NASA Astrophysics Data System (ADS)

    Ortiz Ramírez, A.; Delgado Macuil, R.; Zaca Moran, P.; Rojas López, M.

    2013-11-01

    There are many studies focuses on self-assembled films characterization, photochromic material (spiropyran) had been electrosattached to different enzymes, and in this work we used these spiropyran characteristics to immobilized glucose oxidase. The aim of this work is establish a protocol to build an optical device that shows absorption changes when an enzyme is immobilized and characterized the devise through FTIR and UV/Vis spectroscopy and Atomic Force and Scanning Electron Microscopy. We use different methods such as self-assembled monolayers and the activation reaction by carbodiimide. Corning of quartz was used as matrix, clean solution (1:1 MeOH:HCl, H2SO4) was used to prepared the matrix, 1% silane solution was prepared to functionalized the surface, and a solution prepared with spiropyran (C19H18N2O3), EDC (C5H11N=C=NC6H11), NHS (C4H5NO3), HEPES (C8H18N2O4S) and glucose oxidase from Aspergillus niger was the last step to immobilized this enzyme. Three steps to build it, the first step was cleaned the matrix, the next step was functionalized the surface with silane material, this step allows to modified the surfaces, to prepared to attach the spiropyran. The bonds found from the FTIR spectra to silanized step and the functionalized step (spiropyran attached). That indicated that FTIR and AFM techniques are available to characterized and identify absorption, morphology of a devise modified with spiropyran, that allows attach some biological material, in this case glucose oxidase, the absorption changes and morphological changes are the evidence of immobilization successful.

  15. Gold and Hydroxyapatite Nano-Composite Scaffolds for Anterior Cruciate Ligament Reconstruction: In Vitro Characterization.

    PubMed

    Smith, S E; White, R A; Grant, D A; Grant, S A

    2016-01-01

    Current anterior cruciate ligament (ACL) graft replacement materials often fail due to the lack of biological integration. While many newly developed extracellular matrix based scaffolds show good biocompatibility they often do not entice cellular remodeling and the rebuilding of a functional ligament. We have proposed the conjugation of gold nanoparticles (AuNP) and hydroxyapatite nanoparticles (nano-HAp) to acellular tissue to enhance cell attachment and proliferation while maintaining an improved degradation resistance and open microstructure. We are the first to investigate the double conjugation of AuNP and nano-HAp onto decellularized tissue to improve the tissue remodeling response. Decellularized porcine diaphragm was crosslinked with two types of nano-HAp and amine-functionalized AuNP with 1-ethyl-3-(3-dimethlaminopropyl) carbodiimide (EDC) crosslinker. Scaffolds were characterized using electron microscopy, differential scanning calorimetry, and fibroblast assays. Results demonstrated that scaffolds with nano-HAp have increased thermal stability at low levels of crosslinking. The open microstructure of the scaffold was not compromised allowing for cell migration while still providing increased degradation resistance. The addition of < 200 nm nano-HAp decreased cell viability compared to scaffolds without nanoparticles, but the addition of AuNP to scaffolds showed enhanced cell viability in the presence of < 200 nm nano-HAp. The addition of < 40 nm nano-HAp showed an increase in cell viability compared to scaffolds crosslinked without nanoparticles. It is concluded that attaching AuNP and < 40nm nano-HAp to extracellular matrices may improve overall properties. PMID:27398580

  16. Immobilized magnetic beads based multi-target affinity selection coupled with high performance liquid chromatography-mass spectrometry for screening anti-diabetic compounds from a Chinese medicine "Tang-Zhi-Qing".

    PubMed

    Tao, Yi; Chen, Zhui; Zhang, Yufeng; Wang, Yi; Cheng, Yiyu

    2013-05-01

    We developed an approach for screening bioactive compounds from botanical drug using multiple target-immobilized magnetic beads coupled with high performance liquid chromatography-mass spectrometry. This novel approach was called magnetic beads based multi-target affinity selection-mass spectrometry (MT-ASMS). It can enrich and identify different types of ligands from mixture extracts. Multiple targets (maltase, invertase, lipase) were immobilized on the magnetic beads by covalent linkage using 1-(3-dimethyl-aminopropyl)-3-ethyl-carbodiimide (EDC) and N-hydroxysuccinimide (NHS) as reaction reagents, respectively. The properties of enzyme conjugated magnetic beads were characterized using transmission electron microscopy, X-ray diffractometer and vibration sample magnetometer. Several factors including pH, ion strength, incubation time and temperature were optimized using three known ligands (caffeic acid, ferulic acid, and hesperidin). The established MT-ASMS approach was applied to screening for ligands from a Chinese medicine "Tang-Zhi-Qing", which was used to treat type II diabetes in China. Seven bound compounds were identified via liquid chromatography-mass spectrometry (LC/MS). Five active compounds including 2,3,4,6-tetra-O-galloyl-D-glucose, 1,2,3,4-tetra-O-galloyl-D-glucose, 1,2,3,4,6-penta-O-galloyl-d-glucose, quercetin-3-O-β-D-glucuronide and quercetin-3-O-β-D-glucoside were identified and their activities were validated by conventional inhibitory assay. Our findings suggested that the proposed approach is efficient in screening compounds with multiple activities from extracts of botanical drugs. PMID:23501439

  17. Effect of disulfiram pretreatment on the tissue distribution, macromolecular binding, and excretion of (U-1,2-14C)dichloroethane in the rat

    SciTech Connect

    Igwe, O.J.; Que Hee, S.S.; Wagner, W.D.

    1986-01-01

    The effect of disulfiram (DSF) pretreatment on the distribution of (14C)ethylene dichloride (EDC) in selected organs and/or tissues of control and EDC-pretreated rats was studied. The presence of EDC metabolites and their binding to an acid-insoluble extract of the tissues, as well as purified protein and DNA, were evaluated. Dietary DSF was found to modulate the distribution, excretion, and macromolecular binding of EDC and/or its metabolites at 4 and 24 hr following ip administration. The urinary excretion of (14C)EDC metabolites was not affected by subchronic inhalation exposure to nonradiolabeled EDC. However, DSF pretreatment increased the fat deposition of EDC and decreased the urinary excretion of its metabolites. DSF also increased the binding of EDC metabolites to DNA and decreased the binding to protein in the liver, kidneys, spleen, and testes. However, prior exposure to EDC alone increased the binding of its metabolites to DNA in the kidneys only.

  18. Developmental exposure to a mixture of two mechanistically distinct antiandrogens results in cumulative adverse reproductive effects in adult male rats

    EPA Science Inventory

    Typically, toxicological studies have focused on the adverse effects from exposure to single chemicals. However, endocrine disrupting chemicals (EDCs) are detected in the environment as mixtures. Empirical evidence suggests that mixtures of EDCs with the same mechanism of action...

  19. Thyroid Nodules

    MedlinePlus

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  20. Hormone Abuse Prevention and What You Need to Know

    MedlinePlus

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    MedlinePlus

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  18. Hormones and Obesity

    MedlinePlus

    ... y Cuidadores Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ... Women's Health Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ...

  19. Myth vs. Fact: Adrenal Fatigue

    MedlinePlus

    ... y Cuidadores Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ... Women's Health Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ...

  20. Menopause

    MedlinePlus

    ... y Cuidadores Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ... Women's Health Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ...

  1. Primary Ovarian Insufficiency

    MedlinePlus

    ... y Cuidadores Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ... Women's Health Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ...

  2. USE OF THE LABORATORY RAT AS A MODEL IN ENDOCRINE DISRUPTOR SCREENING AND TESTING

    EPA Science Inventory

    The screening and testing program the US Environmental Protection Agency is currently developing to detect endocrine-disrupting chemicals (EDCs) is described. EDCs have been shown to alter the following activities: hypothalamic-pituitary-gonadal [HPG] function; estrogen, androge...

  3. SCREENING AND TESTING FOR ENDOCRINE DISRUPTION IN FISH - BIOMARKERS AS "SIGNPOSTS," NOT "TRAFFIC LIGHTS," IN RISK ASSESSMENT

    EPA Science Inventory

    Biomarkers provide important tools for addressing the impacts of endocrine disrupting chemicals (EDCs) in fish. Presently, biomarkers are best used as mechanistic "signposts" rather than as "red traffic lights" in the environmental risk assessment of EDCs. In field studies, bio...

  4. STANDARDIZATION AND VALIDATION OF PROPOSED PROTOCOLS FOR IN VITRO SCREENING ASSAYS AND QSAR FOR ESTROGEN RECEPTOR AND ANDROGEN RECEPTOR

    EPA Science Inventory

    Screening EDCs for androgenic and antiandrogenic activities was recommended by the EDSTAC Committee in it Final Report. This research will develop in vitro approaches to assess estrogen receptor binding, develop cell lines that stably express estrogen receptor for screening EDC...

  5. SHORT TERM FATHEAD MINNOW REPRODUCTION ASSAY GENERATES RESPONSE PROFILES BROADLY CONSISTENT WITH MECHANISMS OF ACTION

    EPA Science Inventory

    Small fish tests have been a recommended component of regulatory programs proposed for EDCs ... the EPA plans to use this assay to help fulfill screening and testing requirements associated with the current EDC regulatory program.

  6. EFFECTIVE RISK MANAGEMENT OF ENDOCRINE DISRUPTING CHEMICALS USING DRINKING WATER TREATMENT PROCESSES

    EPA Science Inventory

    The conventional drinking water treamtent processes of coagulation, flocculation, and filtration as well as specialized treatment processes have been examined for their capacity to remove endocrine disrupting chemicals (EDCs). A groupf od EDCs including 4-nonylphenol, diethylphth...

  7. Endocrine-Disrupting Compounds in Aquatic Ecosystems.

    EPA Science Inventory

    Endocrine disrupting chemicals (EDCs) are a ubiquitous issue of concern in our aquatic systems. Commonly detected EDCs include natural and synthetic hormones, surfactants, plasticizers, disinfectants, herbicides and metals. The potency of these chemicals varies substantially, as ...

  8. Find an Endocrinologist

    MedlinePlus

    ... y Cuidadores Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ... Women's Health Hormones and Health Journey Through the Endocrine System Endocrine Disrupting Chemicals (EDCs) Endocrine Glands and Types ...

  9. Electrochemical carbon dioxide concentrator: Math model

    NASA Technical Reports Server (NTRS)

    Marshall, R. D.; Schubert, F. H.; Carlson, J. N.

    1973-01-01

    A steady state computer simulation model of an Electrochemical Depolarized Carbon Dioxide Concentrator (EDC) has been developed. The mathematical model combines EDC heat and mass balance equations with empirical correlations derived from experimental data to describe EDC performance as a function of the operating parameters involved. The model is capable of accurately predicting performance over EDC operating ranges. Model simulation results agree with the experimental data obtained over the prediction range.

  10. Destruction of estrogens using Fe-TAML/peroxide catalysis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Endocrine disrupting chemicals (EDCs) impair living organisms by interfering with hormonal processes controlling cellular development. Reduction of EDCs in water by an environmentally benign method is an important green chemistry goal. One EDC, 17 alpha-ethinylestradiol (EE2), the active ingredien...

  11. Using the Health Belief Model to Illustrate Factors That Influence Risk Assessment during Pregnancy and Implications for Prenatal Education about Endocrine Disruptors

    ERIC Educational Resources Information Center

    Qiu, Xing; Chen, Shaw-Ree; Barrett, Emily S.; Velez, Marissa; Conn, Kelly; Heinert, Sara

    2014-01-01

    Endocrine disrupting chemicals (EDCs) such as Bisphenol A (BPA) and phthalates are ubiquitous in our environment and a growing body of research indicates that EDCs may adversely affect human development. Fetal development is particularly susceptible to EDC exposure, and prenatal care providers are being asked to educate women about the risks of…

  12. DEVELOPING TOOLS FOR EVALUATING RISK MANAGEMENT OF ENDOCRINE DISRUPTING CHEMICALS

    EPA Science Inventory

    The goal of endocrine disrupting chemical (EDC) risk management (RM) is to minimize the release of EDCs into the environment or to minimize the exposure of humans or wildlife to EDCs already present in the environment. RM research projects may involve: substituting more innocuous...

  13. PROGRAM FOR THE IDENTIFICATION AND REPLACEMENT OF ENDOCRINE DISRUPTING CHEMICALS

    EPA Science Inventory

    A computer software program is being developed to aid in the identification and replacement of endocrine disrupting chemicals (EDC). This program will be comprised of two distinct areas of research: identification of potential EDC nd suggstions for replacing those potential EDC. ...

  14. Monitoring of environmental phenolic endocrine disrupting compounds in treatment effluents and river waters, Korea.

    PubMed

    Ko, Eun-Joung; Kim, Kyoung-Woong; Kang, Seo-Young; Kim, Sang-Don; Bang, Sun-Baek; Hamm, Se-Yeong; Kim, Dong-Wook

    2007-10-15

    The last two decades have witnessed growing scientific and public concerns over endocrine disrupting compounds (EDCs) that have the potential to alter the normal structure or functions of the endocrine system in wildlife and humans. In this study, the phenolic EDCs such as alkylphenol, chlorinated phenol and bisphenol A were considered. They are commonly found in wastewater discharges and in sewage treatment plant. In order to monitor the levels and seasonal variations of phenolic EDCs in various aquatic environments, a total of 15 water samples from the discharged effluent from sewage and wastewater treatment plants and river water were collected for 3 years. Ten environmental phenolic EDCs were determined by GC-MS and laser-induced fluorescence (LIF). GC-MS analysis revealed that most abundant phenolic EDCs were 4-n-heptylphenol, followed by nonlyphenol and bisphenol A during 2002-2003, while 4-t-butylphenol and 4-t-octylphenol were newly detected in aquatic environments in 2004. The category of phenolic EDCs showed similar fluorescence spectra and nearly equal fluorescence decay time. This makes it hard to distinguish each phenolic EDC from the EDCs mixture by LIF. Therefore, the results obtained from LIF analysis were expressed in terms of the fluorescence intensity of the total phenolic EDCs rather than that of the individual EDC. However, LIF monitoring and GC-MS analysis showed consistent result in that the river water samples had lower phenolic EDCs concentration compared to the effluent sample. This revealed a lower fluorescence intensity and the phenolic EDCs concentration in summer was lower than that in winter. For the validation of LIF monitoring for the phenolic EDCs, the correlation between EDCs concentration acquired from GC-MS and fluorescence intensity from LIF was obtained (R=0.7379). This study supports the feasibility of the application of LIF into EDCs monitoring in aquatic systems. PMID:19073088

  15. Orientation and density control of bispecific anti-HER2 antibody on functionalized carbon nanotubes for amplifying effective binding reactivity to cancer cells

    NASA Astrophysics Data System (ADS)

    Kim, Hye-In; Hwang, Dobeen; Jeon, Su-Ji; Lee, Sangyeop; Park, Jung Hyun; Yim, Dabin; Yang, Jin-Kyoung; Kang, Homan; Choo, Jaebum; Lee, Yoon-Sik; Chung, Junho; Kim, Jong-Ho

    2015-03-01

    cotinine tandem antibody. This new approach provides an effective control over antibody orientation and density on the surface of carbon nanotubes through site-specific binding between the anti-cotinine domain of the bispecific tandem antibody and the cotinine group of the functionalized carbon nanotubes. The developed synthetic carbon nanotube/bispecific tandem antibody conjugates (denoted as SNAs) show an effective binding affinity against HER2 that is three orders of magnitude higher than that of the carbon nanotubes bearing a randomly conjugated tandem antibody prepared by carbodiimide chemistry. As the density of a tandem antibody on SNAs increases, their effective binding affinity to HER2 increases as well. SNAs exhibit strong resonance Raman signals for signal transduction, and are successfully applied to the selective detection of HER2-overexpressing cancer cells. Electronic supplementary information (ESI) available: Materials; synthesis of carboxymethylated phenoxy dextran (CM-PhO-dex); synthesis of the SWNT bioconjugate prepared by EDC coupling; NMR results; Raman Instrument for detection of cancer cells with SNAs; NIR fluorescence spectrophotometer; quantification of the bispecific tandem antibody bound to the SWNT; all supplementary figures, table and scheme. This material is available from the Wiley Online Library or from the author. See DOI: 10.1039/c4nr07305c

  16. Direct action of endocrine disrupting chemicals on human sperm.

    PubMed

    Schiffer, Christian; Müller, Astrid; Egeberg, Dorte L; Alvarez, Luis; Brenker, Christoph; Rehfeld, Anders; Frederiksen, Hanne; Wäschle, Benjamin; Kaupp, U Benjamin; Balbach, Melanie; Wachten, Dagmar; Skakkebaek, Niels E; Almstrup, Kristian; Strünker, Timo

    2014-07-01

    Synthetic endocrine disrupting chemicals (EDCs), omnipresent in food, household, and personal care products, have been implicated in adverse trends in human reproduction, including infertility and increasing demand for assisted reproduction. Here, we study the action of 96 ubiquitous EDCs on human sperm. We show that structurally diverse EDCs activate the sperm-specific CatSper channel and, thereby, evoke an intracellular Ca(2+) increase, a motility response, and acrosomal exocytosis. Moreover, EDCs desensitize sperm for physiological CatSper ligands and cooperate in low-dose mixtures to elevate Ca(2+) levels in sperm. We conclude that EDCs interfere with various sperm functions and, thereby, might impair human fertilization. PMID:24820036

  17. Direct action of endocrine disrupting chemicals on human sperm

    PubMed Central

    Schiffer, Christian; Müller, Astrid; Egeberg, Dorte L; Alvarez, Luis; Brenker, Christoph; Rehfeld, Anders; Frederiksen, Hanne; Wäschle, Benjamin; Kaupp, U Benjamin; Balbach, Melanie; Wachten, Dagmar; Skakkebaek, Niels E; Almstrup, Kristian; Strünker, Timo

    2014-01-01

    Synthetic endocrine disrupting chemicals (EDCs), omnipresent in food, household, and personal care products, have been implicated in adverse trends in human reproduction, including infertility and increasing demand for assisted reproduction. Here, we study the action of 96 ubiquitous EDCs on human sperm. We show that structurally diverse EDCs activate the sperm-specific CatSper channel and, thereby, evoke an intracellular Ca2+ increase, a motility response, and acrosomal exocytosis. Moreover, EDCs desensitize sperm for physiological CatSper ligands and cooperate in low-dose mixtures to elevate Ca2+ levels in sperm. We conclude that EDCs interfere with various sperm functions and, thereby, might impair human fertilization. PMID:24820036

  18. Possible relationship between endocrine disrupting chemicals and hormone dependent gynecologic cancers.

    PubMed

    Dogan, Selen; Simsek, Tayup

    2016-07-01

    The effects of the natural and synthetic estrogens have been studied for a long time but the data regarding estrogen related chemicals (endocrine disrupting chemicals, EDCs) and their effects on reproductive system are scarce. EDCs are hormone like agents that are readily present in the environment, which may alter the endocrine system of humans and animals. Approximately 800 chemicals are known or suspected to have the potential to function as EDC. Potential role of EDCs on reproductive disease has gained attention in medical literature in recent years. We hypothesize that exposure to low doses of EDCs in a chronic manner could cause hormone dependent genital cancers including ovarian and endometrial cancer. Long term exposure to low concentrations of EDCs may exert potentiation effect with each other and even with endogenous estrogens and could inhibit enzymes responsible for estrogen metabolism. Exposure time to these EDCs is essential as we have seen from Diethylstilbestrol experience. Dose-response curves of EDCs are also unpredictable. Hence mode of action of EDCs are more complex than previously thought. In the light of these controversies lower doses of EDCs in long term exposure is not harmless. Possibility of this relationship and this hypothesis merit further investigation especially through in vivo studies that could better show the realistic environmental exposure. With the confirmation of our hypothesis, possible EDCs could be identified and eliminated from general use as a public health measure. PMID:27241264

  19. Endocrine-disrupting compounds in reclaimed water and residential ponds and exposure potential for dislodgeable residues in turf irrigated with reclaimed water.

    PubMed

    Sidhu, Harmanpreet S; Wilson, Patrick C; O'Connor, George A

    2015-07-01

    Endocrine-disrupting chemicals (EDCs) occur in reclaimed water (RW), which may serve as an exposure source for humans. The presence of EDCs in RW used to irrigate turf and in nearby water-retention ponds was determined. In addition, the total dislodgeable mass of each EDC was determined after irrigation (using RW) to simulate exposure of a 3-year-child playing in turf grass recently irrigated with RW. Five EDCs (estrone, 17β-estradiol, 17α-ethynylestradiol, bisphenol A, and 4-n-nonylphenol) were quantified in 28 samples of RWs (wastewater-treatment plant effluents) and 88 samples from residential surface water-retention ponds. St. Augustine variety of turf grass was irrigated with spiked RW to study dislodgement of the five EDCs overtime using a drag-sled method. Grass clippings were analyzed to relate masses of EDC on grass with masses dislodged. EDCs were detected in both RW and ponds at ng/L concentrations. Maximum EDC masses were dislodged immediately after irrigation. Dislodged masses of estrone and 17β-estradiol are two separate EDCs, 17β-estradiol and 17α-ethynylestradiol decreased rapidly and were lower than detection limits 4 h after application. Dislodged bisphenol-A and nonylphenol decreased more slowly but were not detected 6 h after application. Avoiding contact with recently irrigated turf grass should decrease the risks of exposure to these EDCs. PMID:25758534

  20. Transcriptional analysis of endocrine disruption using zebrafish and massively parallel sequencing

    PubMed Central

    Baker, Michael E.; Hardiman, Gary

    2014-01-01

    Endocrine disrupting chemicals (EDCs) including plasticizers, pesticides, detergents and pharmaceuticals, affect a variety of hormone-regulated physiological pathways in humans and wildlife. Many EDCs are lipophilic molecules and bind to hydrophobic pockets in steroid receptors, such as the estrogen receptor and androgen receptor, which are important in vertebrate reproduction and development. Indeed, health effects attributed to EDCs include reproductive dysfunction (e.g., reduced fertility, reproductive tract abnormalities and skewed male/female sex ratios in fish), early puberty, various cancers and obesity. A major concern is the effects of exposure to low concentrations of endocrine disruptors in utero and post partum, which may increase the incidence of cancer and diabetes in adults. EDCs affect transcription of hundreds and even thousands of genes, which has created the need for new tools to monitor the global effects of EDCs. The emergence of massive parallel sequencing for investigating gene transcription provides a sensitive tool for monitoring the effects of EDCs on humans and other vertebrates as well as elucidating the mechanism of action of EDCs. Zebrafish conserve many developmental pathways found in humans, which makes zebrafish a valuable model system for studying EDCs especially on early organ development because their embryos are translucent. In this article we review recent advances in massive parallel sequencing approaches with a focus on zebrafish. We make the case that zebrafish exposed to EDCs at different stages of development, can provide important insights on EDC effects on human health. PMID:24850832

  1. MODIS land data at the EROS data center DAAC

    USGS Publications Warehouse

    Jenkerson, C.B.; Reed, B.C.

    2001-01-01

    The US Geological Survey's (USGS) Earth Resources Observation Systems (EROS) Data Center (EDC) in Sioux Falls, SD, USA, is the primary national archive for land processes data and one of the National Aeronautics and Space Administration's (NASA) Distributed Active Archive Centers (DAAC) for the Earth Observing System (EOS). One of EDC's functions as a DAAC is the archival and distribution of Moderate Resolution Spectroradiometer (MODIS) Land Data collected from the Earth Observing System (EOS) satellite Terra. More than 500,000 publicly available MODIS land data granules totaling 25 Terabytes (Tb) are currently stored in the EDC archive. This collection is managed, archived, and distributed by EOS Data and Information System (EOSDIS) Core System (ECS) at EDC. EDC User Services support the use of MODIS Land data, which include land surface reflectance/albedo, temperature/emissivity, vegetation characteristics, and land cover, by responding to user inquiries, constructing user information sites on the EDC web page, and presenting MODIS materials worldwide.

  2. Zebrafish (Danio rerio) as a model organism for investigating endocrine disruption.

    PubMed

    Segner, Helmut

    2009-03-01

    Endocrine-disrupting compounds (EDCs) are widespread in the aquatic environment and can cause alterations in development, physiological homeostasis and health of vertebrates. Zebrafish, Danio rerio, has been suggested as a model species to identify targets as well as modes of EDC action. In fact, zebrafish has been found useful in EDC screening, in EDC effects assessment and in studying targets and mechanisms of EDC action. Since many of the environmental EDCs interfere with the sex steroid system of vertebrates, most EDC studies with zebrafish addressed disruption of sexual differentiation and reproduction. However, other targets of EDCs action must not be overlooked. For using a species as a toxicological model, a good knowledge of the biological traits of this species is a pre-requisite for the rational design of test protocols and endpoints as well as for the interpretation and extrapolation of the toxicological findings. Due to the genomic resources available for zebrafish and the long experience with zebrafish in toxicity testing, it is easily possible to establish molecular endpoints for EDC effects assessment. Additionally, the zebrafish model offers a number of technical advantages including ease and cost of maintenance, rapid development, high fecundity, optical transparency of embryos supporting phenotypic screening, existence of many mutant strains, or amenability for both forward and reverse genetics. To date, the zebrafish has been mainly used to identify molecular targets of EDC action and to determine effect thresholds, while the potential of this model species to study immediate and delayed physiological consequences of molecular interactions has been instrumentalized only partly. One factor that may limit the exploitation of this potential is the still rather fragmentary knowledge of basic biological and endocrine traits of zebrafish. Information on species-specific features in endocrine processes and biological properties, however, need to be

  3. Removal characteristics of endocrine-disrupting chemicals by laccase from white-rot fungi.

    PubMed

    Sei, Kazunari; Takeda, Tomoaki; Soda, Satoshi O; Fujita, Masanori; Ike, Michihiko

    2008-01-01

    Laccase from 5 white-rot fungal strains (4 Trametes and 1 Pycnoporus strains) were evaluated in the removal spectra with/without mediators against 11 EDCs. Purified laccase from Trametes sp. was also used to reveal the precise degradation spectra and degradation profiles in time course against 20 EDCs with/without mediators. In addition, effectivity of laccase for the purification of complex EDCs contamination was evaluated combining several EDCs. The removal characteristics among tested strains were almost the same and crude/purified laccase could remove various EDCs. Bis(4-hydroxyphenyl)sulfone, diethylhexylphthalate (DEHP), pyrene (PY), anthracene, 3,5-dichlorophenol and pentachlorophenol could not be removed by laccase. DEHP and PY could not be removed even with mediators. Vanillin and vanillic acid revealed to be possible naturally occurring mediators. Laccase-mediator system could expand the degradation spectrum and enhance the EDCs removal ratio and rate. When complex mixtures of EDCs were treated with laccase, the removal ratio was enhanced in comparison to that of single application. Some coexisting EDCs could act as mediators. Thus, the availability of laccase and the effectivity of a mediator on EDCs treatment were indicated. PMID:18161558

  4. Endocrine disrupting chemicals targeting estrogen receptor signaling: Identification and mechanisms of action

    PubMed Central

    Shanle, Erin K.; Xu, Wei

    2011-01-01

    Many endocrine disrupting chemicals (EDCs) adversely impact estrogen signaling by interacting with two estrogen receptors (ERs): ERα and ERβ. Though the receptors have similar ligand binding and DNA binding domains, ERα and ERβ have some unique properties in terms of ligand selectivity and target gene regulation. EDCs that target ER signaling can modify genomic and non-genomic ER activity through direct interactions with ERs, indirectly through transcription factors like the aryl hydrocarbon receptor (AhR), or through modulation of metabolic enzymes that are critical for normal estrogen synthesis and metabolism. Many EDCs act through multiple mechanisms as exemplified by chemicals that bind both AhR and ER, such as 3-methylcholanthrene. Other EDCs that target ER signaling include phytoestrogens, bisphenolics, and organochlorine pesticides and many alter normal ER signaling through multiple mechanisms. EDCs can also display tissue-selective ER agonist and antagonist activities similar to selective estrogen receptor modulators (SERMs) designed for pharmaceutical use. Thus, biological effects of EDCs need to be carefully interpreted because EDCs can act through complex tissue-selective modulation of ERs and other signaling pathways in vivo. Current requirements by the U.S. Environmental Protection Agency require some in vitro and cell-based assays to identify EDCs that target ER signaling through direct and metabolic mechanisms. Additional assays may be useful screens for identifying EDCs that act through alternative mechanisms prior to further in vivo study. PMID:21053929

  5. The Use of Electronic Data Capture Tools in Clinical Trials: Web-Survey of 259 Canadian Trials

    PubMed Central

    Jonker, Elizabeth; Sampson, Margaret; Krleža-Jerić, Karmela; Neisa, Angelica

    2009-01-01

    Background Electronic data capture (EDC) tools provide automated support for data collection, reporting, query resolution, randomization, and validation, among other features, for clinical trials. There is a trend toward greater adoption of EDC tools in clinical trials, but there is also uncertainty about how many trials are actually using this technology in practice. A systematic review of EDC adoption surveys conducted up to 2007 concluded that only 20% of trials are using EDC systems, but previous surveys had weaknesses. Objectives Our primary objective was to estimate the proportion of phase II/III/IV Canadian clinical trials that used an EDC system in 2006 and 2007. The secondary objectives were to investigate the factors that can have an impact on adoption and to develop a scale to assess the extent of sophistication of EDC systems. Methods We conducted a Web survey to estimate the proportion of trials that were using an EDC system. The survey was sent to the Canadian site coordinators for 331 trials. We also developed and validated a scale using Guttman scaling to assess the extent of sophistication of EDC systems. Trials using EDC were compared by the level of sophistication of their systems. Results We had a 78.2% response rate (259/331) for the survey. It is estimated that 41% (95% CI 37.5%-44%) of clinical trials were using an EDC system. Trials funded by academic institutions, government, and foundations were less likely to use an EDC system compared to those sponsored by industry. Also, larger trials tended to be more likely to adopt EDC. The EDC sophistication scale had six levels and a coefficient of reproducibility of 0.901 (P< .001) and a coefficient of scalability of 0.79. There was no difference in sophistication based on the funding source, but pediatric trials were likely to use a more sophisticated EDC system. Conclusion The adoption of EDC systems in clinical trials in Canada is higher than the literature indicated: a large proportion of

  6. An overview of dioxin-like compounds, PCB, and pesticide exposures associated with sexual differentiation of neuroendocrine systems, fluctuating asymmetry, and behavioral effects in birds.

    PubMed

    Ottinger, Mary Ann; Lavoie, Emma T; Abdelnabi, Mahmoud; Quinn, Michael J; Marcell, Allegra; Dean, Karen

    2009-10-01

    Dioxin, polychlorinated biphenyls (PCBs), and pesticides impact neural systems in birds due to interference with sexual differentiation. Early endocrine disrupting chemical (EDC) effects may delay maturation and have long-term effects on lifetime reproduction, especially in precocial birds that complete sexual differentiation prior to hatch. Semi-altricial and altricial species appear more resilient to EDC effects and show a gradient in sensitivity, especially in the neuroplastic song system. Embryonic steroid exposure occurs via maternally deposited steroids followed by embryo produced hormones; EDCs potentially affect these developing systems. As such, EDCs can impact lifelong fitness by acting on neural systems that regulate reproduction, metabolism, and behavior. PMID:19953400

  7. Endocrine disrupting chemicals as potential risk factor for estrogen-dependent cancers.

    PubMed

    Rutkowska, Aleksandra Z; Szybiak, Aleksandra; Serkies, Krystyna; Rachoń, Dominik

    2016-08-01

    Civilization, industrialization, and urbanization create an environment where humans are continuously exposed to endocrine disrupting chemicals (EDCs). Some of breast cancers and endometrial cancer, which are the most common female malignant neoplasms, are estrogen-dependent tumors. Prolonged exposure to estrogens or substances with estrogenic properties may be a risk factor for their development. This paper aimed to discuss the potential adverse effect of EDCs on human health, including the role of EDCs in hormone-dependent carcinogenesis. A review of literature regarding the sources of environmental exposure to EDCs and molecular mechanisms of their action was performed. We analyzed the possible mechanisms of how these substances alter the function of the endocrine system, resulting in adverse health effects. Hundreds of substances with endocrine disrupting potential have been identified in our environment. There is accumulating evidence linking exposure to EDCs with the development of mammary and endometrial cancer. By interacting with steroid receptors, EDCs can impact the cellular processes potentially leading to carcinogenesis. There are also data showing the effect of EDCs on immune dysfunction. During lifespan, people are usually exposed to a mixture of various EDCs, which complicates the assessment of individual substances or compounds implicated in cancer development. As the prevalence of hormone-dependent tumors among women continues to increase, their successful prevention is of human benefit. Institutions representing medicine, science, industry, and governments should develop joint strategies to decrease exposure to EDC, and thus to reduce the risk of hormonedependent tumors in women. PMID:27509913

  8. Facile kinetic induction of a dihydropyridide to pyrrolide ring contraction.

    PubMed

    Carbery, David R; Hill, Michael S; Mahon, Mary F; Weetman, Catherine

    2016-04-14

    Reactions between magnesium 1,4-dihydropyridide or 1,2-dihydro-iso-quinolide derivatives and carbodiimides, RN[double bond, length as m-dash]C[double bond, length as m-dash]NR, generally result in Mg-N insertion and formation of guanidinate complexes. More sterically perturbed systems with N-aryl carbodiimide substitution, however, follow a divergent course of reaction initiating heterocyclic ring contraction and pyrrolide formation under unprecedentedly mild conditions. PMID:26309116

  9. ENDOCRINE DISRUPTERS: A REVIEW OF SOME SOURCES, EFFECTS, AND MECHANISMS OF ACTIONS ON BEHAVIOR AND NEUROENDOCRINE SYSTEMS

    PubMed Central

    Frye, C.; Bo, E.; Calamandrei, G.; Calzà, L.; Dessì-Fulgheri, F.; Fernández, M.; Fusani, L.; Kah, O.; Kajta, M.; Le Page, Y.; Patisaul, H.B.; Venerosi, A.; Wojtowicz, A.K.; Panzica, G.C.

    2011-01-01

    Some environmental contaminants interact with hormones and may exert adverse consequences due to their actions as endocrine disrupting chemicals (EDCs). Exposure in people is typically due to contamination of the food chain, inhalation of contaminated house dust, or occupational exposure. EDCs include pesticides and herbicides (such as diphenyl-dichloro-trichloroethane, DDT, or its metabolites), methoxychlor, biocides, heat stabilizers and chemical catalysts (such as tributyltin, TBT), plastic contaminants (e.g. bisphenol A, BPA), pharmaceuticals (i.e. diethylstilbestrol, DES; 17alpha-ethynilestradiol, EE2), or dietary components (such as phytoestrogens). The goal of this review is to address sources, effects and actions of EDCs, with an emphasis on topics discussed at the International Congress on Steroids and the Nervous System. EDCs may alter reproductively-relevant or non-reproductive, sexually-dimorphic behaviors. In addition, EDCs may have significant effects on neurodevelopmental processes, influencing morphology of sexually-dimorphic cerebral circuits. Exposure to EDCs is more dangerous if it occurs during specific “critical periods” of life, such as intrauterine, perinatal, juvenile or puberty periods, when organisms are more sensitive to hormonal disruption, than in other periods. However, exposure to EDCs in adulthood also can alter physiology. Several EDCs are xenoestrogens, may alter serum lipid concentrations, or metabolism enzymes that are necessary for converting cholesterol to steroid hormones, ultimately altering production of E2 and/or other steroids. Finally, many EDCs may have actions via, or independent of, classic actions at cognate steroid receptors. EDCs may have effects through numerous other substrates, such as the aryl hydrocarbon receptor (AhR), the peroxisome proliferator-activated receptor (PPAR) and retinoid X receptor (RXR), signal transduction pathways, calcium influx, and/or neurotransmitter receptors. Thus, EDCs, from varied

  10. Reproductive health and the environment: Counseling patients about risks.

    PubMed

    Haruty, Bella; Friedman, Julie; Hopp, Stephanie; Daniels, Ryane; Pregler, Janet

    2016-05-01

    Endocrine-disrupting chemicals (EDCs) are associated with reproductive complications such as infertility, pregnancy complications, poor birth outcomes, and child developmental abnormalities, although not all chemicals of concern are EDCs. Pregnant patients and women of childbearing age need reasonable advice about environmental contaminants and reproductive health. PMID:27168513

  11. Nonmonotonic Dose Response Curves (NMDRCs) Common after Estrogen or Androgen Signaling Pathway Disruption: Fact or Falderal?

    EPA Science Inventory

    EDCs appear to induce some effects that do not appear to display a threshold (apparent Linear No Threshold responses) NMDRCs for EDCs Biologically plausible Occur frequently in vitro, but these are generally not relevant to in vivo effects and do not occur at low concentrations ...

  12. Environmental factors and puberty timing: Expert panel research needs

    EPA Science Inventory

    An expert panel reviewed the literature on endocrine disrupting chemicals (EDCs), body size and puberty. The panel concluded that available experimental animal and human data support a possible role of EDCs and body size in relation to alterations in pubertal onset and progressio...

  13. Differences in Uptake, Metabolism and Clearance of Atrazine and Tamoxifen in a Fish and a Rat Species

    EPA Science Inventory

    Atrazine and tamoxifen are known endocrine-disrupting chemicals (EDCs) that have metabolites exhibiting biological activities that are equally or more potent than the parent compound. To evaluate if uptake, metabolism and clearance of such EDCs is a concern in interspecies extrap...

  14. Differences in Uptake, Metabolism and Clearance ofAtrazine and Tamoxifen in a Fish and a Rat Species

    EPA Science Inventory

    Atrazine and tamoxifen are known endocrine-disrupting chemicals (EDCs) that have metabolites exhibiting biological activities that are equally or more potent than the parent compound. To evaluate if uptake, metabolism and clearance of such EDCs is a concern in interspecies extrap...

  15. Modulation of estrogenic effects by environmental temperature and food availability

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Endocrine-disrupting chemicals (EDCs), in combination with environmental influences, interfere with endocrine function in humans and wildlife. Estrogens are a type of EDC that may alter the hypothalamic-pituitary-gonadal axis in male fathead minnows, Pimephales promelas. The impact of estrogens on P...

  16. REVIEW OF EVIDENCE: ARE ENDOCRINE DISRUPTING CHEMICALS IN THE AQUATIC ENVIRONMENT IMPACTING FISH POPULATIONS

    EPA Science Inventory

    In this paper, evidence from the current literature is presented that addresses either of two questions: 1) do EDCs in the aquatic environment have the potential to impact the reproductive health and survival of various fish species, and 2) are EDCs in the aquatic environment act...

  17. OVERVIEW OF THE INTRAMURAL RISK MANAGEMENT RESEARCH PROGRAM

    EPA Science Inventory

    This presentation will provide a summary of the risk management portion of ORD's endocrine disrupting chemicals (EDCs) research program, including its motivation, goals, planning efforts and resulting research areas.

    In an emerging research area like EDCs, risk management ...

  18. Histopathologic Effects of Estrogens on Marine Fishes

    EPA Science Inventory

    Endocrine-disrupting chemicals (EDCs), such as estrogens estradiol (E2) and ethinylestradiol (EE2) have been reported to affect fish reproduction. This study histologically compared and evaluated effects of EDCs in two species of treated fish. Juvenile male summer flounder (Paral...

  19. Obesogens: an emerging threat to public health.

    PubMed

    Janesick, Amanda S; Blumberg, Bruce

    2016-05-01

    Endocrine disrupting chemicals (EDCs) are defined as exogenous chemicals, or mixtures of chemicals, that can interfere with any aspect of hormone action. The field of endocrine disruption is historically rooted in wildlife biology and reproductive endocrinology where EDCs are demonstrated contributors to infertility, premature puberty, endometriosis, and other disorders. Recently, EDCs have been implicated in metabolic syndrome and obesity. Adipose tissue is a true endocrine organ and, therefore, an organ that is highly susceptible to disturbance by EDCs. A subset of EDCs, called "obesogens," promote adiposity by altering programming of fat cell development, increasing energy storage in fat tissue, and interfering with neuroendocrine control of appetite and satiety. Obesity adds more than $200 billion to US healthcare costs and the number of obese individuals continues to increase. Hence, there is an urgent, unmet need to understand the mechanisms underlying how exposures to certain EDCs may predispose our population to be obese. In this review, we discuss the history of obesogen discovery from its origins in reproductive biology to its latest role in the transgenerational inheritance of obesity in mice. We discuss the development of adipose tissue in an embryo, maintenance of adipocyte number in adults, how EDC disruption programs stem cells to preferentially make more adipocytes, the mechanisms by which chemicals can permanently alter the germline epigenome, and whether there are barriers to EDCs in the gametes. PMID:26829510

  20. ENDOCRINE DISRUPTING COMPOUNDS: PROCESSES FOR REMOVAL FROM DRINKING WATER AND WASTEWATER

    EPA Science Inventory

    Although the list of potentially harmful substances is still being compiled and more sophisticated laboratory tests for detection of endocrine disrupting chemicals (EDCs) are being developed, an initial list of known EDCs has been made and an array of drinking water and wastewate...

  1. Sex Differentiation as a Target of Endocrine Disrupting Compounds in Early Life Stage Fathead Minnows (Pimephales promelas)

    EPA Science Inventory

    The occurrence of endocrine disrupting chemicals (EDCs) in concentrated animal feed operation (CAFO) waste, and the potential effects of these chemicals on aquatic ecosystems have been of recent concern. There is evidence that exposure to EDCs during enhanced windows of sensitiv...

  2. Assessing Risks of Endocrine-disrupting Chemicals: A Scientific Odyssey

    EPA Science Inventory

    In the mid-90s there was a marked increase in public awareness of, and concern for, endocrine-disrupting chemicals (EDCs). There have been a number of purported impacts of EDCs on both human and wildlife health; however, in some instances it has been challenging to relate observ...

  3. Patterns of estrogen occurrence in sewage treatment effluent (STPE) from a university campus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In order to evaluate the extent to which sewage treatment plant effluent (STPE) irrigation reduces or eliminates the entrance of endocrine disrupting compounds (EDCs) into groundwater, studies on the fate and transport of selected EDCs in the Penn State spray-irrigation system are on-going. This stu...

  4. DEVELOPMENT OF A SMALLMOUTH BASS QUANTITATIVE REAL-TIME PCR ASSAY TO MEASURE VTG GENE INDUCTION IN MALE FISH FROM THE SOUTH BRANCH OF THE POTOMAC RIVER

    EPA Science Inventory

    A high incidence of intersex bass, primarily male smallmouth bass with previtellogenic oocytes, exists in the south branch of the Potomac River. Exposures to endocrine disrupting chemicals (EDCs) may be the cause of these abnormalities. Potential sources of EDCs to the river are ...

  5. Designing, Teaching, and Evaluating Two Complementary Mixed Methods Research Courses

    ERIC Educational Resources Information Center

    Christ, Thomas W.

    2009-01-01

    Teaching mixed methods research is difficult. This longitudinal explanatory study examined how two classes were designed, taught, and evaluated. Curriculum, Research, and Teaching (EDCS-606) and Mixed Methods Research (EDCS-780) used a research proposal generation process to highlight the importance of the purpose, research question and…

  6. FATHEAD MINNOW AND PEARL DACE PILOT AT CANADIAN EXPERIMENTAL LAKES AREA

    EPA Science Inventory

    There is increasing concern about the potential impact of EDCs on aquatic organisms. Among the EDCs found in aquatic habitats are synthetic estrogens, which are used in contraceptives and other pharmaceuticals. These chemicals enter waterways through sewage treatment plants and s...

  7. HIGH-THROUGHPUT CHEMICAL SCREENING USING PROTEIN PROFILING OF FISH PLASMA

    EPA Science Inventory

    Compounds that affect the hormone system, referred to as "endocrine-disrupting chemicals" (EDCs), cause human and animal health problems. It is necessary to test putative EDC chemicals for such deleterious effects, though current testing methodologies are time/animal intensive an...

  8. Effects of Xenoestrogen and Androgen Mixtures on Ovarian Transcriptome of the Fathead Minnow

    EPA Science Inventory

    Endocrine disrupting chemicals (EDCs), such as the estrogens ethinylestradiol (EE2) and bisphenol A (BPA), and androgens like 17â-trenbolone (TRB) can occur as mixtures in aquatic environments. To date, however, most studies with EDCs in fish have focused on their effects as indi...

  9. MANAGING ENDOCRINE DISRUPTING CHEMICALS USING EXISTING AND INNOVATIVE WASTEWATER TREATMENT TECHNOLOGIES

    EPA Science Inventory

    Research has shown that wastewater (WW) can be a significant source of endocrine disrupting chemicals (EDCs) to the environment. WW treatment (WWT) may include centralized wastewater treatment plants (WWTPs) or smaller on-site WWT technologies. EDCs found in WWT effluents (aqueou...

  10. WASTEWATER TREATMENT AND ITS MANAGEMENT OF ENDOCRINE DISRUPTING CHEMICALS

    EPA Science Inventory

    Research has shown that wastewater treatment (WWT) can be a significant source of endocrine disrupting chemicals (EDCs) to the environment. WWT can include centralized wastewater treatment plants (WWTPs) or on-site WWT technologies. EDCs found in WWT effluents (aqueous and biosol...

  11. EVALUATION OF THE REMOVAL OF ESTROGENS THROUGH THE COAGULATION PROCESS

    EPA Science Inventory

    A number of estrogenic compounds have been shown to be present in surface waters in the U.S. These compounds have the potential to act as potent endocrine disrupting chemicals (EDCs), leading to a growing concern over the possible presence of EDCs in finished drinking waters. Con...

  12. EVALUATION OF THE REMOVAL OF ESTROGENS THROUGH THE COAGULATION PROCESS

    EPA Science Inventory

    A number of estrogenic compounds have been shown to be present in surface waters in the U.S. These compounds have the potential to act as endocrine disrupting chemicals (EDCs), leading to concern over the possible presence of EDCs in finished drinking waters. Consequently, it is ...

  13. EVALUATION OF THE REMOVAL OF ESTROGENS THROUGH THE COAGULATION PROCESS - PAPER

    EPA Science Inventory

    A number of estrogenic compounds have been shown to be present in surface waters in the U.S. These compounds have the potential to act as potent endocrine disrupting chemicals (EDCs), leading to a growing concern over the possible presence of EDCs in finished drinking waters. ons...

  14. EVALUATION OF THE REMOVAL OF ESTROGENS THROUGH THE COAGULATION PROCESS

    EPA Science Inventory

    A number of estrogenic compounds have been shown to be present in surface waters in the U.S. These compounds have the potential to act as potent endocrine disrupting chemicals (EDCs), leading to a growing concern over the possible presence of EDCs in finished drinking waters. C...

  15. THE INSTABILITY OF ESTROGENIC CHEMICALS DURING LABORATORY STATIC EXPOSURE CONDITIONS WITH MALE FATHEAD MINNOWS

    EPA Science Inventory

    Endocrine disrupting chemicals (EDCs) such as Para-nonylphenol (NP), estradiol (E2), estrone (E1), estriol (E3) and ethynylestradiol (EE2) are shown to be ubiquitous in surface waters, sediments and sludge. These EDCs are known to induce vitellogenin gene (Vg) expression in male...

  16. ENDOCRINE-DISRUPTING CONTAMINANTS AND REPRODUCTION IN VERTEBRATE WILDLIFE.

    EPA Science Inventory

    The fields of toxicology, endocrinology, and reproductive physiology recently have combined resources to study the effects of endocrine-disrupting contaminants (EDCs) in wildlife populations. EDCs include a wide variety of chemicals that are only related by the ability to disrupt...

  17. Using fetal endocrine and genomic signatures to predict the relative potency of phthalate esters and their effects on postnatal development of the male rat reproductive tract

    EPA Science Inventory

    The first part of this presentation will address concerns expressed by some scientist that the screening and testing protocols for endocrine disrupting chemicals (EDCs) are 1) unable to adequately detect the low dose effects of EDCs, 2) they are unable to define the shape of the ...

  18. EVALUATION OF THE REMOVAL OF ESTROGENS FOLLOWING CHLORINATION

    EPA Science Inventory

    A number of estrogenic compounds have been shown to be present in surface waters in the U.S. These compounds have the potential to act as potent endocrine disrupting chemicals (EDCs). Although there has not yet been a determination of risks posed by EDCs in finished drinking wat...

  19. Impacts of 17α-ethynylestradiol exposure on metabolite profiles of zebrafish (Danio rerio) liver cells

    EPA Science Inventory

    Endocrine disrupting chemicals (EDCs) that are frequently detected in bodies of water downstream from sewage treatment facilities can have adverse impacts on fish and other aquatic organisms. To properly assess risk(s) from EDCs, tools are needed that can establish linkages from ...

  20. A Comparison of Pathology Found in Three Marine Fish Treated with Endocrine Disrupting Compounds

    EPA Science Inventory

    Endocrine-disrupting chemicals (EDCs), such as the estrogen estradiol (E2) have been reported to affect fish reproduction. This study histopathologically compared and evaluated the effect of EDCs in three species of treated fish. Juvenile male summer flounder (Paralichthys dentat...

  1. A PERSPECTIVE ON THE RISK ASSESSMENT PROCESS FOR ENDOCRINE-DISRUPTIVE EFFECTS ON WILDLIFE AND HUMAN HEALTH

    EPA Science Inventory

    The topic of endocrine disrupting chemicals (EDCs) presents significant issues to the risk assessment process. . . We have a working definition of an EDC, that provides a starting point for considering what chemicals are of concern. We also have an understanding of the important ...

  2. Exposure to Endocrine Disrupting Chemicals and Male Reproductive Health

    PubMed Central

    Jeng, Hueiwang Anna

    2014-01-01

    Endocrine disrupting chemicals (EDCs) can interfere with normal hormonal balance and may exert adverse consequences on humans. The male reproductive system may be susceptible to the effects of such environmental toxicants. This review discusses the recent progress in scientific data mainly from epidemiology studies on the associations between EDCs and male reproductive health and our understanding of possible mechanisms associated with the effects of EDCs on male reproductive health. Finally, the review provides recommendations on future research to enhance our understanding of EDCs and male reproductive health. The review highlights the need for (1) well-defined longitudinal epidemiology studies, with appropriately designed exposure assessment to determine potential causal relationships; (2) chemical and biochemical approaches aimed at a better understanding of the mechanism of action of xenoestrogens with regard to low-dose effects, and assessment of identify genetic susceptibility factors associated with the risk of adverse effects following exposure to EDCs. PMID:24926476

  3. Targeting testis-specific proteins to inhibit spermatogenesis: lesson from endocrine disrupting chemicals

    PubMed Central

    Wan, HT; Mruk, Dolores D; Wong, Chris KC; Cheng, C Yan

    2014-01-01

    Introduction Exposure to endocrine disrupting chemicals (EDCs) has recently been linked to declining fertility in men in both developed and developing countries. Since many EDCs possess intrinsic estrogenic or androgenic activities, thus, the gonad is one of the major targets of EDCs. Areas covered For the past 2 decades, studies found in the literature regarding the disruptive effects of these EDCs on reproductive function in human males and also rodents were mostly focused on oxidative stress-induced germ cell apoptosis, disruption of steroidogenesis, abnormal sperm production and disruption of spermatogenesis in particular cell adhesion function and the blood–testis-barrier (BTB) function. Herein, we highlight recent findings in the field illustrating testis-specific proteins are also targets of EDCs. Expert opinion This information should be helpful in developing better therapeutic approach to manage ECD-induced reproductive toxicity. This information is also helpful to identify potential targets for male contraceptive development. PMID:23600530

  4. Epigenetic effects of endocrine-disrupting chemicals on female reproduction: An ovarian perspective

    PubMed Central

    Zama, Aparna Mahakali; Uzumcu, Mehmet

    2010-01-01

    The link between in utero and neonatal exposure to environmental toxicants, such as endocrine-disrupting chemicals (EDCs) and adult female reproductive disorders is well established in both epidemiological and animal studies. Recent studies examining the epigenetic mechanisms involved in mediating the effects of EDCs on female reproduction are gathering momentum. In this review, we describe the developmental processes that are susceptible to EDC exposures in female reproductive system, with a special emphasis on the ovary. We discuss studies with select EDCs that have been shown to have physiological and correlated epigenetic effects in the ovary, neuroendocrine system, and uterus. Importantly, EDCs that can directly target the ovary can alter epigenetic mechanisms in the oocyte, leading to transgenerational epigenetic effects. The potential mechanisms involved in such effects are also discussed. PMID:20609371

  5. A framework for understanding the relationship between externally and internally directed cognition.

    PubMed

    Dixon, Matthew L; Fox, Kieran C R; Christoff, Kalina

    2014-09-01

    Externally directed cognition (EDC) involves attending to stimuli in the external environment, whereas internally directed cognition (IDC) involves attending internally to thoughts, memories and mental imagery. To date, most studies have focused on the competition or trade-offs between these modes of cognition. However, both EDC and IDC include a variety of cognitive states that differ along multiple dimensions. These dimensions may influence the way in which EDC and IDC relate to each other. In this review, we present a novel framework that considers whether cognitive resources are oriented externally or internally, and also whether a given cognitive state involves intentional (i.e., voluntary) or spontaneous (i.e., involuntary) processing. Within this framework, we examine the conditions under which EDC and IDC are expected to either compete, or co-occur without interference. We argue that EDC and IDC are not inherently antagonistic, but when both involve higher levels of intentionality they are increasingly likely to compete, due to the capacity limitations of intentional processing. In contrast, if one or both involve spontaneous processing, EDC and IDC can co-occur with minimal interference given that involuntary processes are not subject to the same capacity constraints. A review of the brain regions implicated in EDC and IDC suggests that their neural substrates are partially segregated and partially convergent. Both EDC and IDC recruit the lateral prefrontal cortex (PFC) during intentional processing, and may therefore compete over the processes and representational space it supports. However, at lower levels of intentionality, EDC and IDC rely on largely distinct neural structures, which may enable their co-occurrence without interference. The proposal that EDC and IDC can in some cases co-occur, provides a framework for understanding the complex mental states that underlie theory of mind, creativity, the influence of self-evaluative processing on

  6. Evaluation of Water Treatment Methods for Endocrine Disrupting Compounds

    NASA Astrophysics Data System (ADS)

    Thomas, S. M.; Murray, K. E.

    2006-05-01

    Endocrine disrupting compounds (EDCs) have caught recent attention as one of the major concerns in the environment. They are known to interfere with the activity of growth-related hormones and usually, as a result, cause disruption in normal functioning of the body. The compounds currently classified as EDCs range from a variety of both natural and synthetic organic compounds and also some heavy metals. Most of these compounds are used in household, pharmaceutical, industrial, agricultural activities, the consumption or usage of which increases with population. There is a lack of detailed chemical and biological analysis as to what concentrations each of these EDCs pose harmless to the environment because of the large number of the suspected compounds. However, several published reports have established that endocrine disruption is observed in aquatic species due to chronic exposure to concentrations of some EDCs as low as a few ng/l. Conventional water treatment facilities do not usually suffice to remove EDCs in concentrations below 1 ng/l. Available technologies for removal of EDCs include adsorption, degradation and membrane treatment. The removal rates, however, are dependant on the properties of the compound, such as molecular weight, water- octanol partition coefficient and vapor pressure; physiochemical conditions of the matrix such as, redox and temperature conditions; type and dose of degrading agent and the concentration of the EDCs. Since, EDCs comprise a vast variety of compounds, their response to each of these treatment methods will be different and hence it is plausible that a single treatment technique will not be sufficient to remove the EDCs to very low concentrations. Based on our review of existing water treatment methods, we believe that a sequential treatment technique that consists of an adsorption, a degradation and finally a fine membrane treatment, each optimized for favorable, efficient and inexpensive removal may be required to remove

  7. Dietary exposure to endocrine disrupting chemicals in metropolitan population from China: a risk assessment based on probabilistic approach.

    PubMed

    He, Dongliang; Ye, Xiaolei; Xiao, Yonghua; Zhao, Nana; Long, Jia; Zhang, Piwei; Fan, Ying; Ding, Shibin; Jin, Xin; Tian, Chong; Xu, Shunqing; Ying, Chenjiang

    2015-11-01

    The intake of contaminated foods is an important exposure pathway for endocrine disrupting chemicals (EDCs). However, data on the occurrence of EDCs in foodstuffs are sporadic and the resultant risk of co-exposure is rarely concerned. In this study, 450 food samples representing 7 food categories (mainly raw and fresh food), collected from three geographic cities in China, were analyzed for eight EDCs using high performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS). Besides estrone (E1), other EDCs including diethylstilbestrol (DES), nonylphenol (NP), bisphenol A (BPA), octylphenol (OP), 17β-estradiol (E2), 17α-ethinylestradiol (EE2), and estriol (E3) were ubiquitous in food. Dose-dependent relationships were found between NP and EE2 (r=0.196, p<0.05), BPA (r=0.391, p<0.05). Moreover, there existed a correspondencebetween EDCs congener and food category. Based on the obtained database of EDCs concentration combined with local food consumption, dietary EDCs exposure was estimated using the Monte Carlo Risk Assessment (MCRA) system. The 50th and 95th percentile exposure of any EDCs isomer were far below the tolerable daily intake (TDI) value identically. However, the sum of 17β-estradiol equivalents (∑EEQs) exposure in population was considerably larger than the value of exposure to E2, which implied the underlying resultant risk of multiple EDCs in food should be concern. In conclusion, co-exposure via food consumption should be considered rather than individual EDCs during health risk evaluation. PMID:26025473

  8. Investigation and remediation of a 1,2-dichloroethane spill. Part 1: Short and long-term remediation strategies

    SciTech Connect

    Sehayek, L.; Vandell, T.D.; Sleep, B.E.; Lee, M.D.; Chien, C.

    1999-06-30

    Release of an estimated 150,000 gallons of 1,2-dichloroethane (EDC) from a buried pipeline into a ditch and surrounding soil resulted in shallow subsurface contamination of a Gulf Coast site. Short-term remediation included removal of EDC DNAPL (dense nonaqueous phase liquid) by dredging and vacuuming the ditch, and by dredging the river where the ditch discharged. EDC saturation in shallow impacted sediments located beneath the ditch was at or below residual saturation and these sediments were therefore left in place. The ditch was lined, backfilled, and capped. Long-term remediation includes EDC DNAPL recovery and hydraulic containment from the shallow zone with long-term monitoring of the shallow, intermediate, and deep aquifers. Ground water, DNAPL, and dissolved phase models were used to guide field investigations and the selection of an effective remedial action strategy. The DNAPL modeling was conducted for a two-dimensional vertical cross section of the site, and included the three aquifers separated by two aquitards with microfractures. These aquitards were modeled using a dual porosity approach. Matrix and fracture properties of the aquitards used for DNAPL modeling were determined from small-scale laboratory properties. These properties were consistent with effective hydraulic conductivity determined from ground water flow modeling. A sensitivity analysis demonstrated that the vertical migration of EDC was attenuated by dissolution of EDC into the matrix of the upper aquitard. When the organic/water entry pressure of the aquitard matrix, or the solubility of EDC were decreased to unrealistically low values, EDC DNAPL accumulated in the aquifer below the upper aquitard. EDC DNAPL did not reach the regional (deepest) aquifer in any of the cases modeled. The limited extent of vertical EDC migration predicted is supported by ground water monitoring conducted over the four years since the spill.

  9. Combination of UV absorbance and electron donating capacity to assess degradation of micropollutants and formation of bromate during ozonation of wastewater effluents.

    PubMed

    Chon, Kangmin; Salhi, Elisabeth; von Gunten, Urs

    2015-09-15

    In this study, the changes in UV absorbance at 254 nm (UVA254) and electron donating capacity (EDC) were investigated as surrogate indicators for assessing removal of micropollutants and bromate formation during ozonation of wastewater effluents. To measure the EDC, a novel method based on size exclusion chromatography followed by a post-column reaction was developed and calibrated against an existing electrochemical method. Low specific ozone doses led to a more efficient abatement of EDC than of UVA254. This was attributed to the abatement of phenolic moieties in the dissolved organic matter (DOM), which lose their EDC upon oxidation, but are partially transformed into quinones, which still absorb in the measured UV range. For higher specific ozone doses, the relative EDC abatement was lower than the relative UVA abatement, which can be explained by the oxidation of UV absorbing moieties (e.g. non-activated aromatic compounds), which contribute less to EDC. The abatement of the selected micropollutants (i.e., 17α-ethinylestradiol (EE2), carbamazepine (CBZ), atenolol (ATE), bezafibrate (BZF), ibuprofen (IBU), and p-chlorobenzoic acid (pCBA)) varied significantly depending on their reactivity with ozone in the examined specific ozone dose range of 0-1.45 mgO3/mgDOC. The decrease of EE2 and CBZ with high ozone reactivity was linearly proportional to the reduction of the relative residuals of UVA254 and EDC. The abatement of ATE, BZF, IBU, and pCBA with intermediate to low ozone reactivities was not significant in a first phase (UVA254/UVA254,0 = 1.00-0.70; EDC/EDC0 = 1.00-0.56) while their abatement was more efficient than the degradation of the relative residual UVA254 and much more noticeable than the degradation of the relative residual EDC in a second phase (UVA254/UVA254,0 = 0.70-0.25; EDC/EDC0 = 0.56-0.25) because the partially destroyed UV absorbing and electron donating DOM moieties become recalcitrant to ozone attack. Bromate formation was

  10. Prenatal Exposures to Multiple Thyroid Hormone Disruptors: Effects on Glucose and Lipid Metabolism.

    PubMed

    Molehin, Deborah; Dekker Nitert, Marloes; Richard, Kerry

    2016-01-01

    Background. Thyroid hormones (THs) are essential for normal human fetal development and play a major role in the regulation of glucose and lipid metabolism. Delivery of TH to target tissues is dependent on processes including TH synthesis, transport, and metabolism. Thyroid hormone endocrine disruptors (TH-EDCs) are chemical substances that interfere with these processes, potentially leading to adverse pregnancy outcomes. Objectives. This review focuses on the effects of prenatal exposures to combinations of TH-EDCs on fetal and neonatal glucose and lipid metabolism and also discusses the various mechanisms by which TH-EDCs interfere with other hormonal pathways. Methods. We conducted a comprehensive narrative review on the effects of TH-EDCs with particular emphasis on exposure during pregnancy. Discussion. TH imbalance has been linked to many metabolic processes and the effects of TH imbalance are particularly pronounced in early fetal development due to fetal dependence on maternal TH for proper growth and development. The pervasive presence of EDCs in the environment results in ubiquitous exposure to either single or mixtures of EDCs with deleterious effects on metabolism. Conclusions. Further evaluation of combined effects of TH-EDCs on fetal metabolic endpoints could improve advice provided to expectant mothers. PMID:26989557

  11. Endocrine-disrupting chemicals in the Pearl River Delta and coastal environment: sources, transfer, and implications.

    PubMed

    Xu, Weihai; Yan, Wen; Huang, Weixia; Miao, Li; Zhong, Lifeng

    2014-12-01

    A study was conducted to investigate the occurrence and behavior of six endocrine-disrupting chemicals (EDCs) in sewage, river water, and seawater from the Pearl River Delta (PRD). The six EDCs under study were 4-nonylphenol (NP), bisphenol A (BPA), 17α-ethynylestradiol (EE2), estrone (E2), 17β-estradiol (E2), and estriol (E3). These EDCs, predominated by BPA, were found in high levels in the influents and the effluents of sewage treatment plants in the area. The relatively high concentrations (0.23-625 ng/L) of the EDCs detected in the receiving river water suggested that the untreated sewage discharge was a major contributor. The EDCs detected in eight outlets of the Pear River and the Pear River Estuary were in the ranges of 1.2-234 and 0.2-178 ng/L, respectively. The estrogen equivalents in the aquatic environments under study ranged from 0.08 to 4.5 ng/L, with E1 and EE2 being the two predominant contributors. As the fluxes of the EDCs from the PRD region to the nearby ocean are over 500 tons each year, the results of this study point to the potential that Pearl River is a significant source of the EDCs to the local environment there. PMID:24817613

  12. Electrochemical carbon dioxide concentrator subsystem development

    NASA Technical Reports Server (NTRS)

    Koszenski, E. P.; Heppner, D. B.; Bunnell, C. T.

    1986-01-01

    The most promising concept for a regenerative CO2 removal system for long duration manned space flight is the Electrochemical CO2 Concentrator (EDC), which allows for the continuous, efficient removal of CO2 from the spacecraft cabin. This study addresses the advancement of the EDC system by generating subsystem and ancillary component reliability data through extensive endurance testing and developing related hardware components such as electrochemical module lightweight end plates, electrochemical module improved isolation valves, an improved air/liquid heat exchanger and a triple redundant relative humidity sensor. Efforts included fabrication and testing the EDC with a Sabatier CO2 Reduction Reactor and generation of data necessary for integration of the EDC into a space station air revitalization system. The results verified the high level of performance, reliability and durability of the EDC subsystem and ancillary hardware, verified the high efficiency of the Sabatier CO2 Reduction Reactor, and increased the overall EDC technology engineering data base. The study concluded that the EDC system is approaching the hardware maturity levels required for space station deployment.

  13. An investigation of common crosslinking agents on the stability of electrospun collagen scaffolds.

    PubMed

    Huang, Gloria Portocarrero; Shanmugasundaram, Shobana; Masih, Pallavi; Pandya, Deep; Amara, Suwah; Collins, George; Arinzeh, Treena Livingston

    2015-02-01

    Electrospinning is a widely used processing method to form fibrous tissue engineering scaffolds that mimic the structural features of the native extracellular matrix. Electrospun fibers made of collagen have been sought because it is a natural structural protein that supports cell attachment and growth. Yet, conventional solvents used to electrospin collagen can result in the loss of hydrolytic stability and fiber morphology of the scaffold. This study evaluated the effect of commonly used synthetic and natural crosslinking agents, genipin, glutaraldehyde, N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC), and EDC with N-hydroxysulfosuccinimide (EDC-NHS), on electrospun collagen. Crosslinked collagen scaffolds were assessed for structural integrity in an in vitro immersion study for up to 3 months. Their cytocompatibility was evaluated by human mesenchymal stem cell morphology and proliferation. Our results showed that dimensional stability and cytocompatibility of crosslinked electrospun collagen scaffolds are dependent on the type of crosslinking agent used. Collagen scaffolds treated with EDC and EDC-NHS were structurally stable and retained fiber structure for up to 3 months and were cytocompatible. Therefore, EDC and EDC-NHS are favorable crosslinking agents for electrospun collagen that can be utilized in tissue engineering applications. PMID:24828818

  14. Occurrence and fate of selected endocrine-disrupting chemicals in water and sediment from an urban lake.

    PubMed

    Wu, Chenxi; Huang, Xiaolong; Lin, Juan; Liu, Jiantong

    2015-02-01

    Occurrence of five endocrine-disrupting chemicals (EDCs)-bisphenol-A (BPA), triclosan (TCS), 17α-ethinyl estradiol (EE2), benzophenone-3, and 4-methylbenzylidene camphor-were monitored in East Lake, the largest urban lake in China. Other than EE2, all selected EDCs were detected at least once in the lake water with concentrations ≤89.1 ng/L. EDCs were detected with greater occurrence in spring than in other seasons. In lake sediment, TCS was detected at the greatest concentration (30.9 ng g(-1)), whereas BPA and EE2 were not detected. Spatial distribution of the EDCs in the lake water and the lake sediment showed similar patterns, and greater EDC residuals were found from those sites with known wastewater input. The linear adsorption coefficients (K d) varied from 17.9 to 1,017 L kg(-1) and were related to the octanol-water partition coefficient (K ow) values of the compounds. Photodegradation was a major process removing the EDCs from the lake water, and the presence of dissolved organic material and NO3 (-) in the lake water can accelerate the photodegradation process. Degradation of the EDCs in the lake sediment was relatively slow and occurred mainly due to the microbial processes. All compounds were found more persistent under anoxic conditions than under oxic conditions. PMID:25298153

  15. Effects of Endocrine Disruptor Compounds, Alone or in Combination, on Human Macrophage-Like THP-1 Cell Response.

    PubMed

    Couleau, N; Falla, J; Beillerot, A; Battaglia, E; D'Innocenzo, M; Plançon, S; Laval-Gilly, P; Bennasroune, A

    2015-01-01

    The aim of the present study was to evaluate the immunological effects on human macrophages of four endocrine disruptor compounds (EDCs) using the differentiated human THP-1 cell line as a model. We studied first the effects of these EDCs, including Bisphenol A (BPA), di-ethylhexyl-phthalate (DEHP), dibutyl phthalate (DBP) and 4-tert-octylphenol (4-OP), either alone or in combination, on cytokine secretion, and phagocytosis. We then determined whether or not these effects were mediated by estrogen receptors via MAPK pathways. It was found that all four EDCs studied reduced strongly the phagocytosis of the differentiated THP-1 cells and that several of these EDCs disturbed also TNF-α, IL-1 β and IL-8 cytokine secretions. Furthermore, relative to control treatment, decreased ERK 1/2 phosphorylation was always associated with EDCs treatments-either alone or in certain combinations (at 0.1 μM for each condition). Lastly, as treatments by an estrogen receptor antagonist suppressed the negative effects on ERK 1/2 phosphorylation observed in cells treated either alone with BPA, DEHP, 4-OP or with the combined treatment of BPA and DEHP, we suggested that estrogen receptor-dependent pathway is involved in mediating the effects of EDCs on human immune system. Altogether, these results advocate that EDCs can disturb human immune response at very low concentrations. PMID:26133781

  16. Effects of Endocrine Disruptor Compounds, Alone or in Combination, on Human Macrophage-Like THP-1 Cell Response

    PubMed Central

    Couleau, N.; Falla, J.; Beillerot, A.; Battaglia, E.; D’Innocenzo, M.; Plançon, S.; Laval-Gilly, P.; Bennasroune, A.

    2015-01-01

    The aim of the present study was to evaluate the immunological effects on human macrophages of four endocrine disruptor compounds (EDCs) using the differentiated human THP-1 cell line as a model. We studied first the effects of these EDCs, including Bisphenol A (BPA), di-ethylhexyl-phthalate (DEHP), dibutyl phthalate (DBP) and 4-tert-octylphenol (4-OP), either alone or in combination, on cytokine secretion, and phagocytosis. We then determined whether or not these effects were mediated by estrogen receptors via MAPK pathways. It was found that all four EDCs studied reduced strongly the phagocytosis of the differentiated THP-1 cells and that several of these EDCs disturbed also TNF-α, IL-1 β and IL-8 cytokine secretions. Furthermore, relative to control treatment, decreased ERK 1/2 phosphorylation was always associated with EDCs treatments—either alone or in certain combinations (at 0.1 μM for each condition). Lastly, as treatments by an estrogen receptor antagonist suppressed the negative effects on ERK 1/2 phosphorylation observed in cells treated either alone with BPA, DEHP, 4-OP or with the combined treatment of BPA and DEHP, we suggested that estrogen receptor-dependent pathway is involved in mediating the effects of EDCs on human immune system. Altogether, these results advocate that EDCs can disturb human immune response at very low concentrations. PMID:26133781

  17. Metabolic disruption in context: Clinical avenues for synergistic perturbations in energy homeostasis by endocrine disrupting chemicals

    PubMed Central

    Sargis, Robert M

    2015-01-01

    The global epidemic of metabolic disease is a clear and present danger to both individual and societal health. Understanding the myriad factors contributing to obesity and diabetes is essential for curbing their decades-long expansion. Emerging data implicate environmental endocrine disrupting chemicals (EDCs) in the pathogenesis of metabolic diseases such as obesity and diabetes. The phenylsulfamide fungicide and anti-fouling agent tolylfluanid (TF) was recently added to the list of EDCs promoting metabolic dysfunction. Dietary exposure to this novel metabolic disruptor promoted weight gain, increased adiposity, and glucose intolerance as well as systemic and cellular insulin resistance. Interestingly, the increase in body weight and adipose mass was not a consequence of increased food consumption; rather, it may have resulted from disruptions in diurnal patterns of energy intake, raising the possibility that EDCs may promote metabolic dysfunction through alterations in circadian rhythms. While these studies provide further evidence that EDCs may promote the development of obesity and diabetes, many questions remain regarding the clinical factors that modulate patient-specific consequences of EDC exposure, including the impact of genetics, diet, lifestyle, underlying disease, pharmacological treatments, and clinical states of fat redistribution. Currently, little is known regarding the impact of these factors on an individual’s susceptibility to environmentally-mediated metabolic disruption. Advances in these areas will be critical for translating EDC science into the clinic to enable physicians to stratify an individual’s risk of developing EDC-induced metabolic disease and to provide direction for treating exposed patients. PMID:27011951

  18. Prenatal Exposures to Multiple Thyroid Hormone Disruptors: Effects on Glucose and Lipid Metabolism

    PubMed Central

    Molehin, Deborah

    2016-01-01

    Background. Thyroid hormones (THs) are essential for normal human fetal development and play a major role in the regulation of glucose and lipid metabolism. Delivery of TH to target tissues is dependent on processes including TH synthesis, transport, and metabolism. Thyroid hormone endocrine disruptors (TH-EDCs) are chemical substances that interfere with these processes, potentially leading to adverse pregnancy outcomes. Objectives. This review focuses on the effects of prenatal exposures to combinations of TH-EDCs on fetal and neonatal glucose and lipid metabolism and also discusses the various mechanisms by which TH-EDCs interfere with other hormonal pathways. Methods. We conducted a comprehensive narrative review on the effects of TH-EDCs with particular emphasis on exposure during pregnancy. Discussion. TH imbalance has been linked to many metabolic processes and the effects of TH imbalance are particularly pronounced in early fetal development due to fetal dependence on maternal TH for proper growth and development. The pervasive presence of EDCs in the environment results in ubiquitous exposure to either single or mixtures of EDCs with deleterious effects on metabolism. Conclusions. Further evaluation of combined effects of TH-EDCs on fetal metabolic endpoints could improve advice provided to expectant mothers. PMID:26989557

  19. Endocrine Aspects of Environmental "Obesogen" Pollutants.

    PubMed

    Nappi, Francesca; Barrea, Luigi; Di Somma, Carolina; Savanelli, Maria Cristina; Muscogiuri, Giovanna; Orio, Francesco; Savastano, Silvia

    2016-01-01

    Growing evidence suggests the causal link between the endocrine-disrupting chemicals (EDCs) and the global obesity epidemics, in the context in the so-called "obesogenic environment". Dietary intake of contaminated foods and water, especially in association with unhealthy eating pattern, and inhalation of airborne pollutants represent the major sources of human exposure to EDCs. This is of particular concern in view of the potential impact of obesity on chronic non-transmissible diseases, such as type 2 diabetes, cardiovascular disease, and hormone-sensitive cancers. The key concept is the identification of adipose tissue not only as a preferential site of storage of EDCs, but also as an endocrine organ and, as such, susceptible to endocrine disruption. The timing of exposure to EDCs is critical to the outcome of that exposure, with early lifetime exposures (e.g., fetal or early postnatal) particularly detrimental because of their permanent effects on obesity later in life. Despite that the mechanisms operating in EDCs effects might vary enormously, this minireview is aimed to provide a general overview on the possible association between the pandemics of obesity and EDCs, briefly describing the endocrine mechanisms linking EDCs exposure and latent onset of obesity. PMID:27483295

  20. Effect of deployment time on endotoxin and allergen exposure assessment using electrostatic dust collectors.

    PubMed

    Kilburg-Basnyat, Brita; Metwali, Nervana; Thorne, Peter S

    2015-01-01

    The electrostatic dust collector (EDC) is a passive dust sampling device for exposure assessment of airborne endotoxin and possibly allergens. EDCs consist of a non-conducting plastic folder holding two or four electrostatic cloths of defined area. The sampling time needed to achieve detectable and reproducible loading for bioaerosols has not been systematically evaluated. Thus, in 15 Iowa farm homes EDCs were deployed for 7-, 14-, and 28-day sampling periods to determine if endotoxin and allergens could be quantified and if loading rates were uniform over time, i.e. if loads doubled from 7 to 14 days or 14 to 28 days and quadrupled from 7 to 28 days. Loadings between left and right paired EDC cloths were not significantly different and were highly correlated for endotoxin, total protein, and cat (Fel d1), dog (Can f1), and mouse (Mus m1) allergens (P < 0.001). EDCs performed especially well for endotoxin sampling with close agreement between paired samples (Pearson r = 0.96, P < 0.001). Endotoxin loading of the EDCs doubled from 7- to 14-day deployments as hypothesized although the loading rate decreased from 14 to 28 days of sampling with only a 1.38-fold increase. Allergen exposure assessment using EDCs was overall less satisfactory. Although there was reasonable agreement between paired samples, only exposures to cat, dog, and mouse allergens were reliable and these only at the longer deployment times. PMID:25187036