FTIR characterization of the secondary structure of proteins encapsulated within PLGA microspheres.
A commonly used technique for protein encapsulation in microspheres is the double-emulsion method wherein an initial water-in-oil (w/o) emulsion of protein and polymer is formed via sonication, and then a second emulsion (w/o)/w is formed by dispersion in an aqueous phase via homogenization. This approach is often used to produce microspheres of biodegradable poly(lactic-co-glycolic acid) (PLGA). The harsh processing associated with this method can cause denaturation of the encapsulated protein. Herein, we have used Fourier transform infrared (FTIR) spectroscopy to determine the secondary structures of two model proteins, bovine serum albumin (BSA) and chicken egg-white lysozyme, within PLGA microspheres. The alpha-helix content of both proteins in the microspheres was about a third lower than in the lyophilized state, indicating conformational changes upon protein entrapment within the microspheres. BSA microspheres containing the stabilizing excipient trehalose have a higher alpha-helix content than those without excipient, suggesting that trehalose partially prevents the denaturing effects incurred during processing. In addition, BSA released from microspheres is improved by incorporation of trehalose: analysis of the protein released from the microspheres indicates that there is less BSA dimer formation in the trehalose-containing microspheres than in those without trehalose.
Fu
K
K
Harvard-MIT Division of Health Sciences and Technology, Cambridge, MA, USA.
Griebenow
K
K
Hsieh
L
L
Klibanov
A M
AM
LangerR
D000818:Animals; D002417:Cattle; D002645:Chickens; D004355:Drug Stability; D004529:Egg White / Q000032:analysis; D005079:Excipients / Q000737:chemistry; D066298:In Vitro Techniques; D019344:Lactic Acid / Q000737:chemistry*; D008863:Microspheres; D009113:Muramidase / Q000737:chemistry* / Q000187:drug effects / Q000493:pharmacokinetics; D011100:Polyglycolic Acid / Q000737:chemistry*; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011108:Polymers / Q000737:chemistry*; D017433:Protein Structure, Secondary / Q000187:drug effects; D012710:Serum Albumin, Bovine / Q000737:chemistry* / Q000187:drug effects / Q000493:pharmacokinetics; D017550:Spectroscopy, Fourier Transform Infrared; D014199:Trehalose / Q000737:chemistry*
D005079:Excipients; D011108:Polymers; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011100:Polyglycolic Acid; D012710:Serum Albumin, Bovine; D019344:Lactic Acid; D014199:Trehalose; D009113:Muramidase