Vamsi

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Biomineralization, Magnetic nanoparticles

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Vamsi

  1. 1. “ Ferric Iron Nanoparticles Formation Mediated By Negatively Charged Polypeptides” Vamsi Krishna Aluru, Dr Robin S. Hissam Department of Chemical Engineering, West Virginia University, Morgantown, WV November 9, 2010 AICHE 2010 Annual Meeting
  2. 2. <ul><li>Background </li></ul><ul><li>Objective </li></ul><ul><li>Research </li></ul><ul><li>Results </li></ul><ul><li>Conclusions </li></ul><ul><li>Future Work </li></ul><ul><li>Acknowledgement </li></ul>Outline AICHE 2010 Annual Meeting
  3. 3. Background <ul><li>Biomineralization </li></ul><ul><ul><ul><li>Examples: Silicates in algae, carbonates in invertebrates, </li></ul></ul></ul><ul><ul><ul><li> magnetite in magnetotactic bacteria </li></ul></ul></ul><ul><li>Magnetotactic Bacteria </li></ul>Transmission Electron photograph of magnetotactic bacterium and bacterial magnetic particles A : Magnetospirillum magneticum AMB-1 * B : Desulfovibrio magneticum RS-1 ** * Matsunaga T et al. Appl Microbiol Biotechnol, 1991 ** Sakaguchi T Matsunaga T et al. Nature, 1993 AICHE 2010 Annual Meeting
  4. 4. Magnetic Particles <ul><li>Two types </li></ul><ul><ul><li>Iron oxides (Fe 3 O 4 ) and iron sulfides (Fe 3 S 4 , FeS 2 ) </li></ul></ul><ul><li>All magnetic particles are of size 35-120 nm </li></ul><ul><li>Morphology of crystals </li></ul><ul><ul><li>cuboidal, </li></ul></ul><ul><ul><li>elongated prismatic and </li></ul></ul><ul><ul><li>arrowhead-shaped </li></ul></ul><ul><li>Stable single magnetic domains </li></ul><ul><li>(SMD) </li></ul>http://scienceblogs.com/afarensis/2006/01/24/more_on_magnetic_bacteria/ AICHE 2010 Annual Meeting
  5. 5. Applications of Magnetic particles <ul><li>Magnetic drug delivery system </li></ul><ul><li>DNA extraction </li></ul><ul><li>Magnetic Resonance Imaging (MRI) </li></ul>*Hypothetical magnetic drug delivery system * Pankhurst Q A et al. J.Phys. 2003 AICHE 2010 Annual Meeting Magnetic Nanoparticles Tumor DDS Diagnosis Therapy Hyperthermia MRI MI sensor
  6. 6. Synthesis of Magnetic Particles <ul><li>Magnetosome vesicle formation </li></ul><ul><ul><ul><ul><li>Invagination of cytoplasmic membrane </li></ul></ul></ul></ul><ul><li>Uptake of iron into vesicle </li></ul><ul><li>Biomineralization </li></ul>Fe +3 Fe +2 +1 vesicle AICHE 2010 Annual Meeting
  7. 7. Magnetosome <ul><li>Magnetosome </li></ul><ul><ul><ul><li>Cell organelle with magnetite crystal and an outer cover of lipid bilayer membrane </li></ul></ul></ul><ul><li>Membrane Composition </li></ul><ul><ul><ul><li>Phospholipids, fatty acids and proteins </li></ul></ul></ul><ul><li>Proteins are specific to membrane and are responsible for crystal growth </li></ul><ul><li>Arakaki et al. characterized a number of proteins in Magnetospirillum magneticum strain AMB-1 </li></ul>AICHE 2010 Annual Meeting
  8. 8. Biomineralization Protein-Mms6 <ul><li>All proteins isolated had hydrophobic N-terminal region and a hydrophilic C-terminal region </li></ul><ul><li>Magnetite nanoparticles with similar morphology to bacterial magnetite particles were synthesized in presence of recombinant Mms6 expressed in E.coli </li></ul>* Electron micrographs of magnetic particles synthesized in the presence (A) and absence (B) of Mms6. *Arakaki A et al. J. Biol. Chem 2003 AICHE 2010 Annual Meeting
  9. 9. Mms6 Protein <ul><li>Hydrophilic C-terminal region of Mms6 contains amino acid residues with hydroxyl or carboxyl groups </li></ul><ul><li>Arakaki et al. proposed that Mms6 acts as a template for magnetic nanoparticle synthesis </li></ul>AICHE 2010 Annual Meeting G A V Y A Y M K S R D I E S A Q S D E E V E L R D
  10. 10. Advantages of Biomimetics <ul><li>Chemical processes </li></ul><ul><ul><li>High temperature, pressure and pH </li></ul></ul><ul><ul><li>Low yield , difficult to reproduce </li></ul></ul><ul><ul><li>Often produce toxic byproducts </li></ul></ul><ul><li>Biomimetics is a natural way to produce nanoparticles of similar morphology to those produced in magnetotactic bacteria </li></ul>AICHE 2010 Annual Meeting
  11. 11. Objective <ul><li>Central hypothesis </li></ul><ul><ul><ul><li>Effect of hydroxyl and carboxyl groups on size and shape of nucleated particles </li></ul></ul></ul><ul><li>Aim </li></ul><ul><li>To design peptides with different functional groups </li></ul><ul><li>To inspect the morphology of crystals formed with different peptides </li></ul>AICHE 2010 Annual Meeting
  12. 12. Design of Peptides AICHE 2010 Annual Meeting Name Peptide VA1 M G A V Y A Y G K S R S I Y S A Q S Y S S V S L R Y (OH) VA2 M G A V D A D G K E R D I D E A Q E D E E V D L R D (COOH) VA3 M G S E D A Y G S Y D E S Y D E Q S E D Y E S Y Y S (Combination) VAMms6 M G A V Y A Y G K S R D I E S A Q S D E E V E L R D
  13. 13. <ul><li>Background </li></ul><ul><li>Objective </li></ul><ul><li>Research </li></ul><ul><li>Results </li></ul><ul><li>Conclusions </li></ul><ul><li>Future Work </li></ul><ul><li>Acknowledgement </li></ul>AICHE 2010 Annual Meeting
  14. 14. SDS-PAGE Protein purification profile of VA-Mms6, VA1 and VA2, 12.5 % separating gel, ran @100V: Lane 1) Protein ladder, Lane 2) VA-Mms6, Lane 3) VA1 and Lane 4) VA2   AICHE 2010 Annual Meeting 37 kD 25 kD 20 kD 15 kD 10 kD VA-Mms6 VA1 VA2 1 2 3 4
  15. 15. Western Blot Western Blot Stained with anti-His antibody: Lane1) purified His-tagged VA-Mms6; Lane 2) purified His-tagged VA1; Lane 3) purified His-tagged VA2 Courtesy: Dr. Yogesh Kulkarni AICHE 2010 Annual Meeting
  16. 16. Experiment <ul><li>Co-precipitation reactions were carried out using all peptides </li></ul><ul><li>100µl of 0.6M FeCl 3 and 0.3M FeCl 2 were added to 10µl of peptide solution ( 5mg/ml) </li></ul><ul><li>0.1N NaOH was titrated against above reaction mixture under constant nitrogen flow </li></ul><ul><li>The color of solution turned from pale yellow to black </li></ul><ul><li>All solutions were degassed with argon prior to reaction </li></ul><ul><li>Particles were allowed to grow for 7 days and concentrated at the bottom using neodymium-boron magnet. </li></ul>AICHE 2010 Annual Meeting
  17. 17. Scanning Electron Microscopy (SEM) (a) SEM image of iron particles synthesized in the presence of VA-Mms6 (b) EDS of iron particles synthesized in the presence of VA-Mms6   AICHE 2010 Annual Meeting (c) SEM image of iron particles synthesized in the presence of VA2 (d) SEM image of iron particles synthesized in the presence of VA3 (c) (d)
  18. 18. Transmission Electron Microscopy (TEM) TEM images of magnetite particles synthesized in presence of : (a) VA2 (b) Mms6   VA2 *Mms6 Courtesy: Dr. Chaoying Ni, University of Delaware *Arakaki A et al. J. Biol. Chem 2003 AICHE 2010 Annual Meeting Selected Area Electron Diffraction (SAED) patterns of iron oxide particles synthesized in the presence of VA2 (a) (b)
  19. 19. Atomic Force Microscopy (AFM) (a) (b) (c) AFM image of magnetite particles synthesized in the presence of (a) VA-Mms6, (b) VA2, (c) VA3 Courtesy: Srikanth Raghavan, WVU AICHE 2010 Annual Meeting
  20. 20. X-ray Photoelectron Spectroscopy (XPS) XPS spectra of magnetite particles synthesized in the presence of VA2, VA3, VA-Mms6 and XPS spectra of standard magnetite particles 711 eV 724.4 eV VA-Mms6 Courtesy: Srikanth Raghavan, WVU AICHE 2010 Annual Meeting
  21. 21. DLS AICHE 2010 Annual Meeting Size distribution of magnetite particles synthesized in the presence of (a) VAMms6, (b) VA2, and (c) VA3 Size (d,nm) Size (d,nm) Size (d,nm) Number (%) Number (%) Number (%) a) b) c) Peak Diameter VAMms6 88 nm VA2 62 nm VA3 41 nm
  22. 22. AICHE 2010 Annual Meeting
  23. 23. Magnetic Force Microscopy (MFM) MFM image of magnetite particles synthesized in presence of VA2 (a) topography (b) phase   (a) (b) Courtesy: Srikanth Raghavan, WVU AICHE 2010 Annual Meeting Individual particles of size range 40 nm
  24. 24. Conclusions <ul><li>Magnetite nanoparticles were synthesized with the aid of VA-Mms6, VA2 and VA3 </li></ul><ul><li>Discrete particles of size range 60 nm were observed in the presence of VA2 </li></ul><ul><li>Magnetite particles synthesized in the presence of VA2 showed stable magnetic domain </li></ul><ul><li>Carboxyl group peptide VA2 showed a variation in size and morphology of particles in comparison to VA-Mms6 and VA3 </li></ul>AICHE 2010 Annual Meeting
  25. 25. Future Work <ul><li>In the near future characterization of particles synthesized in the presence of PEG-VA2 and PEG-VA3 should be carried out </li></ul><ul><li>Film coating of the iron particles with a biocompatible polymer like PEG will allow the particles to be used as drug delivery agents. </li></ul><ul><li>Functionalized Magnetite nanoparticles can even be used as contrast agents in MRI </li></ul>AICHE 2010 Annual Meeting
  26. 26. Preliminary Work <ul><li>Iron particles were synthesized in the presence of polyethylene glycol (PEG) coupled to the peptides VA2 and VA3 </li></ul>(a) (b) SEM image of magnetite particles synthesized in presence of (a) PEG-VA2 (b) PEG-VA3 AICHE 2010 Annual Meeting
  27. 27. Acknowledgement <ul><li>Dr. Robin Hissam </li></ul><ul><li>The Hissam Group </li></ul><ul><ul><ul><li>Avram Siegel, Jaclyn Kokx, Logan May, and Andrew Graves </li></ul></ul></ul><ul><li>Dr. Charter D. Stinespring and Srikanth Raghavan </li></ul><ul><li>Dr. Rajesh Naik (AFRL) </li></ul><ul><li>Liviu Magean and Adrienne McGraw </li></ul><ul><li>Dr. Chaoying Ni, Dr. Thomas Epps, and Julie N. L. Albert (University of Delaware) </li></ul><ul><li>WVU Research Corporation for funding </li></ul>AICHE 2010 Annual Meeting
  28. 28. Thank You AICHE 2010 Annual Meeting

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