Micro/nano Structures Constructured Based On Polypeptides

Polypeptides prepared via N‐carboxyanhydride (NCA) ring‐opening polymerization have receivedintensive concern for decades. Up to date, these polypeptides and their block copolymers have beenreported to be applied as micro/nano functional carriers, bio‐compatible materials and responsivegels due to their unique properties. However, most of the reported works focused on linearpolypeptides and block polypeptide copolymers. In this dissertation, a series of hyperbranchedpolypeptide with different cores and arms were prepared, and some micro/nano structures based onpolypeptides were further constructed. The studies included the synthesis and characterization of thehyperbranched polypeptide copolymers (HPCs), the mineralization of calcium carbonate directed bypolypeptides, polypeptides modification of multiwalled carbon nanotubes and silica nanoparticles bygraft‐from approach, and the gold nanoparticles stabilized by polypeptides and their chargecomplexes. The details and key conclusions are summarized as follows:(1) The preparation of hyperbranch cored, polypeptide armed block copolymers and the study oftheir biocompatibilities.In this chapter, hydrophilic hyperbranched polyethyleneimide (PEI) was used as initiator toprepare four kinds of HPCs with different hydrophobic polypeptide arms by ring‐openingpolymerization of N‐ε‐carbobenzoxy‐L‐Lysine NCA (ZLys‐NCA),γ‐benzyl‐L‐glutamate NCA (BLG‐NCA),β‐benzyl‐L‐aspartate NCA (BAsp‐NCA) and phenylalanine NCA (Phe‐NCA). Three kinds of HPCs withdifferent hydrophilic polypeptide arms can also be obtained after deprotection. Their structures werecharacterized by FT‐IR and 1H NMR. The molecular weights and EI/peptide contents were calculatedwith the assistance of 1H NMR.Hydrophobic hyperbranched polyester Boltorn H₂0/40 were reacted with N‐butoxycarbonyl‐Lphenylalaninein the presence of dicyclohexylcarbodiimide, and then the butoxycarbonyl wasdeprotected to obtain the amino‐terminated hyperbranched polyester Boltorn H₂0‐NH₂ /H40‐NH₂.The two compounds initiated ZLys‐NCA and BAsp‐NCA ring‐opening polymerization, and then theprotective groups were removed to prepare three kinds of hydrophobic cored, hydrophilic armed HPCs.They were also characterized by FT‐IR and 1H NMR, and the molecular weights, peptide contents wereobtained by GPC and 1H NMR.The biocompatibility tests of some HPCs showed that the cytotoxicities were increased withpositive charge content. HPCs with good biocompatibility should contain proper amounts of Aspblock or Glu block, which carries carboxyl side groups.(2) The study of calcium carbonate mineralization directed by poly‐L‐Lysine.In this chapter, the novel calcium carbonate (CaCO₃) morphology, twin‐sphere with an equatorialgirdle, had been obtained under the control of poly(L‐lysine) (PLys) through gas‐diffusion method. TEM images showed the"girdle"width is 2³ nm, and the surface of the twin‐sphere are covered withfractal crystal pieces. The effect of the concentration of calcium ion and PLys, the reaction time, andthe initial pH value were investigated, and various interesting morphologies, including twin‐sphere,discus‐like, hexagonal plate and hallow structure were observed by using scanning electronicmicroscopy (SEM). Laser microscopic Raman (LM‐Raman) spectroscopy studies indicated that all theseCaCO₃ are vaterite.This work found that PLys with side amine groups can control the mineralization of CaCO₃ just likethe acidic polypeptides. A reasonable process for forming the morphologies was discussedpreliminarily. At first, the anisotropic vaterite nanocrystals were formed under the direction of thePLys chains which contained partlyα‐helix, then the nanocrystals assembled to form a nuclear plate,and the growth from both sides of the nuclear plate resulted in the twin‐sphere. With theenvironmental pH value increasing to stronger alkaline, the conformation change of PLys chains leadto the CaCO₃ tangential oriented growth on the sphere surface and vertical oriented growth on theequatorial zone. Finally, the twin‐sphere with equatorial girdle morphology was generated. Itsuggested that alkaline polypeptide or protein may also play a suitable role in the biomineralizationprocess in nature.(3) The study of calcium carbonate mineralization directed by hyperbranched polypeptidecopolymers.In this chapter, various calcium carbonate morphologies had been obtained via gas‐diffusionmethod by the direction of PEI‐PGlu HPCs with different PGlu section length. When the template wasPEI‐PGlu1, which contained short PGlu section, the products were mainly fusiform calcite crystals.When the template was PEI‐PGlu2, which contained medium PGlu section, the morphologies ofcalcium carbonate were significantly affected by the concentration of Ca2+. At lower Ca2+concentration, the morphology of the calcium carbonate was elongated rhombohedral crystal. Withthe Ca2+concentration increasing, the morphology of the calcium carbonate turned to transitionalcrystal posses both the characters of elongated rhombohedral and twin‐sphere. If theCa2+concentration were further increased, the twin‐sphere calcium carbonate macrocrystal wasobtained. Finally, the calcium carbonate macrocrystal showed sphere morphology at highest Ca2+concentration. All these macrocrystals were determined calcite by XRD and LM‐Raman spectra. Themorphology of calcium carbonate could also be affected by Mg2+ or other mixed solvents.PEI‐PGlu served as"3D nanoreactor"in the mineralization of calcium carbonate. PEI section couldabsorb CO₂; PBLG section could conjugate with Ca2+, and"crosslink"with them to construct thetemplate. Both the two sections could affect the morphology of calcium carbonate. A possiblemechanism was described as following: the initial templates are some PEI‐PGlu molecules crosslinkedby Ca2+. The templates absorb CO₂ themselves via PEI section to generate the CaCO₃ nanocrystal. Themorphologies of the templates are determined by the length of PGlu section. PEI‐PGlu1, whichcontains short PGlu section, can only form the template to generate fusiform calcite crystals;PEI‐PGlu2, which contains medium PGlu section, can form various templates to construct CaCO₃morphologies from elongated rhombohedra crystal to sphere macrocrystal relying on the concentration of Ca²⁺. PEI‐PGlu3, which contains long PGlu section and tends to absorb Ca²⁺in onemolecular rather than share Ca²⁺ inter molecules, is a template for produce spherical macrocrystals.(4) Polypeptide modification of inorganic nanomaterials by"Graft From"approach.Covalent surface functionalization of carbon nanotubes with polypeptides is promising in possiblemedical applications. This work presented a graft‐from approach to perform the polypeptidemodification of multiwalled carbon nanotubes (MWNTs). The raw MWNTs were amine‐functionalizedat first, then MWNT‐NH₂ was used as the initiator to initiate the ring‐opening polymerization ofBLG‐NCA, resulting in the polypeptide‐grafted MWNTs. FTIR, XPS and TGA data demonstrated that thefunctionalization was successful. The TEM images of the products showed that the thickness of thepolypeptide shell of the PBLG‐MWNT was about 4.5²2 nm. TGA analysis showed that thepolypeptide contents were 3.74⁶.04 g PBLG/g MWNTs. The average graft lengths are 8.13¹3.2 BLGsections. The polypeptide modified carbon naaotubes showed better solubility in polar solutions.According to the facile route developed here, the functionalized carbon nanotubes with other types ofpolypeptides can be fabricated easily using the corresponding NCAs.Silica nanospheres were modified by coupling agent to obtained SiO₂‐NH₂ at first, then theSiO₂‐NH₂ initiated ZLys‐NCA ring‐opening reaction, resulting in the polypeptide grafted silicananosphere SiO₂‐PZLys. TEM images showed that the thickness of the polypeptide shell is about14.7²6.8nm. Polypeptide contents were calculated 0.23⁰.36 g PZlys/g SiO₂ by TGA.The results of TGA and TEM showed that the grafted polypeptides can not increase linearly withthe NCA feed ratio. It investigated that the grafting reaction was not living polymerization. Besides,equal weight of MWNTs can graft more polypeptide than SiO₂ nanoparticles, because that the specificarea of the MWNTs is much larger than SiO₂ nanoparticles, and the MWNTs are hollow.(5) The study of gold nanoparticles stabilized by polypeptides.The mercaptoethylamine trifluoroacetate initiated ZLys‐NCA ring‐opening polymerization toobtain thiol terminated polypeptide PZLys‐SH. PZLys‐SHs with different molecular weights were usedto prepare gold nanoparticles (GNPs) Au@PZLys‐SH in DMF with HAuCl4 by the reduction of NaBH4.UV‐Vis spectra indicated that the size of GNPs grew up with the amount of HAuCl4. TEM imagesshowed the diameter of GNPs was 5¹0nm and the lattice of GNPs were perfect. GNPs stabilized byPZly‐SH1, Au@PZly‐SH1 may self‐assemble to small vesicles with tens of nanometers diameter inwater. Besides, Au@PZly‐SH can also combine with single walled carbon nanotubes (SWNTs) treatedby nitric acid because the Au@PZly‐SH possess positive charges, and the SWNTs possess carboxyl. TEMimages confirmed that the GNPs in the complexes are most absorbed on the SWNTs bundles, or thecross block of the SWNTs.HAuCl4 was reduced in situ by PLys in aqueous solution for preparing the GNPs stabilized by PLys,Au@PLys. The diameter of the Au@PLys is about 10 nm. Au@PLys can combine with hyperbranchedpolypeptide copolymer H40‐Phe‐PAsp, which contains carboxyl in arms, and form complex micelles.The diameters of the micelles are increasing with the adding of Au@PLys firstly, and then decreasewith the adding of Au@PLys. It contributes to the added positive charges carried by Au@PLys attractthe carboxyl in the micelle shell. Au@PLys can also bind with HsDNA to build complex micelles, the diameters of the micelles are increased from about 150nm to about 700nm with the adding ofAu@PLys. TEM images showed the two kinds of micelles contained several GNPs, and the diameters ofthe micelles are accorded with the results of dynamic light scatting. The two kinds of complex micellesshould be applied in biomedical in the future.