En protokoll for produksjon av Gliadin-cyanoakrylat Nanopartikler for hydrofile belegget

Summary

This article presents a protocol for the production of protein-based nanoparticles that changes the hydrophobic surface to hydrophilic. The produced nanoparticle is an assembly of gliadin-cyanoacrylate diblock copolymers. Spray coating with the produced nanoparticle changes the surface of target material to a hydrophilic surface.

Abstract

Denne artikkelen viser en protokoll for fremstilling av proteinbaserte nanopartikler som endrer den hydrofobe overflaten til hydrofile ved en enkel sprøytebelegg. Disse nanopartikler produseres ved polymerisasjonsreaksjonen av alkyl cyanoakrylat på overflaten av kornprotein (gliadin) molekyler. Alkyl cyanoakrylat er en monomer som polymeriserer raskt ved RT når det påføres på overflaten av materialer. Dens Polymeriseringsreaksjonen initieres av spormengder av svakt basiske eller nukleofile arter på overflaten, inkludert fuktighet. Når polymerisert, de polymeriserte alkyl cyanoakrylater viser en sterk affinitet med objekt materialer fordi cyanatgruppene er ryggraden i poly (alkyl cyanoacrylate). Proteiner også fungere som pådriver for denne polymerisasjon fordi de inneholder amingrupper som kan initiere polymerisasjon av cyanoacrylate. Hvis aggregert protein som blir brukt som en initiator, blir proteinaggregat omgitt av den hydrofobepoly (alkyl cyanoakrylat) kjedene etter polymerisasjonsreaksjonen av alkyl cyanoakrylat. Ved å kontrollere den eksperimentelle tilstand, blir partikler i nanometer serien som produseres. De fremstilte nanopartikler lett adsorberes på overflaten av de fleste materialer som glass, metall, plast, tre, lær og tekstiler. Når overflaten av et materiale sprøytes med den produserte nanopartikkelsuspensjonen og skylt med vann, den micellære struktur av nanopartikler endrer sin konformasjon, og de hydrofile proteiner er eksponert for luft. Som et resultat endrer nanopartikkel-belagt overflate til hydrofil.

Introduction

The goal of this article is to show the protocol for the preparation of nanoparticle suspension that modifies the wetting property of materials by a simple spray. The presented nanoparticle suspension is made from alkyl cyanoacrylate¹ and a cereal protein, gliadin^2,3. During the manufacturing process, protein aggregates are formed in aqueous ethanol⁴. Subsequent reaction with monomer (alkyl cyanoacrylate) produces the nanoparticle that is comprised of a protein core surrounded by linear polymer chains [poly(alkyl cyanoacrylate)]⁵.

Poly(alkyl cyanoacrylate)s are biodegradable and have been used for the production of nanoparticles via emulsion polymerization⁶. This reaction is spontaneously initiated by the hydroxyl groups dissociated from water or by other nucleophilic groups in the reaction medium⁷. In the case of the reaction presented in this article, the amine groups on the surface of protein aggregates initiate the polymerization reaction of alkyl cyanoacrylate monomers^5,8. As a result of this reaction, nanoparticles are formed in the reaction medium. The core of the nanoparticle is protein aggregates and the outer layer is poly(alkyl cyanoacrylate) (PACA) chains. The prepared nanoparticle has a strong affinity on most materials (more precisely, any material which PACA can adsorb to) and adheres onto their surface to form a thin coating on a nanometer scale. A simple spray coating instantly turns the surface of the materials hydrophilic.

Gliadin is one of the main fractions of gluten, which is in the endosperms of wheat. Gliadins are mainly monomeric proteins with molecular weights around 28,000 – 55,000. Non-covalent bonds such as hydrogen bonds, ionic bonds and hydrophobic bonds are responsible for the aggregation of gliadins². Although gliadin is chosen as a reactant in this article, many other proteins can also be used for the same purpose. However, the reaction condition needs to be modified accordingly because the condition for inducing aggregation is dependent on the type of protein to be employed⁸. Compared with other proteins, gliadin is more readily available, purification is simple, and production cost is low. Although ethyl cyanoacrylate (ECA) is chosen as a monomer for the presented reaction, other alkyl cyanoacrylates can also be used for the same reaction. The reason for choosing ECA is that it is readily available at low cost.

Protocol

1. Avfetting Commercial Gliadin Mål 150 ml aceton med en målesylinder og hell i 250 ml erlenmeyerkolbe. Under omrøring med en rørestav på en magnetrører ved romtemperatur, tilsett 30 g kommersiell gliadin pulver. Lukke åpningen i kolben med aluminiumsfolie, og holder på røring O / N i hetten. Filtratet oppløsningen med et filterpapir. Vask filtratet med frisk aceton (ca. 50 ml). La stå i 10 min for å tillate aceton til avløp. Overfør filtratet samm…

Representative Results

Nanopartiklene kan fremstilles på forskjellige reaksjonsbetingelser. Gliadin former aggregere i bredt spekter av etanolinnhold 5. Imidlertid er størrelsen på aggregatene må være så liten som mulig fordi et ekstra lag (f.eks., Polymerisert ECA) legges til denne aggregat, og denne prosess vil gjøre den endelige størrelse større. Dersom den endelige størrelsen på partikkelen er for stor, vil partikkelen være ustabile og vil lett bli utfelt. Derfor, ble 68% va…

Discussion

There are several critical steps in the production of the nanoparticle suspension. If the purified gliadin contains impurities, the reaction with ECA will produce side products. Although these unwanted products can be removed from the reaction medium during the centrifugation stage, it lowers the yield of the major product. If the gliadin solution prepared during experimental step 2.3) does not show clear separation between supernatant and precipitate after two days, the solution needs to stand for longer time. Using fre…

Materials

Ethyl cyanoacrylate (ECA) monomer	K&R International (Laguna Niguel, CA)	I-1605	Any pure ECA can be used.
Gliadin	MGP Ingredients, Inc (Atchison, KS)	Gift from the company	Gliadin can be purchased from Sigma-Aldrich (cat #: G3375-25G). Instead of gliadin, any commercial  gluten can be used.
HCl	Any		Any reagent grade chemical can be used.
Acetone	Any		Any reagent grade chemical can be used.
Methanol	Any		Any reagent grade chemical can be used.
Ethanol (100%)	Any		Any reagent grade chemical can be used.
Filter paper	Any		Any grade filter paper larger than 10 cm can be used.
Cell culture square dish	Any		Any dish larger than 20 cm x 20 cm can be used.
Coffee grinder	Any		Any coffee grinder can be used.
Rotary evaporator	Any		Any rotary evaporator can be used.
Freeze Dryer	Any		Any freeze dryer that can reach – 70°C can be used.
Centrifuge	Any		Any centrifuge that can apply 1000 x g can be used.
Magnetic stirrer	Any		Any magnetic stirrer that can turn spin bar to 1000 RPM can be used.
Dynamic Light Scattering (DLS)	Brookhaven Instruments Corporation	NanoBrook Omni Zeta Potential Analyzer	DLS from any company can be used.
Scanning Electron Microscope (SEM)	Carl Zeiss Inc.		Any SEM can be used.
Dynamic Contact Angle (DCA)	Thermo Cahn Instruments		Any DCA can be used.