title: Extraction of PLGA-microencapsulated proteins using a two-immiscible liquid phases system containing surfactants.
abstract: OBJECTIVE
  The extraction of proteins from PLGA/PLA microspheres by a two-immiscible liquid phases system with the addition of surfactants was investigated.
  
  
  METHODS
  First, the extraction without surfactants and the interaction between proteins (IFN-α2b and EGF) and empty microspheres (PLGA or PLA) was studied. Next, proteins stability in presence of different surfactants was evaluated by: (1) bicinchoninic acid protein assay, (2) reversed phase-high performance liquid chromatography, and (3) enzyme-linked immunosorbent assay. Then, proteins were extracted with PBS/dichloromethane including selected surfactants and characterized by the above mentioned techniques, biological activity tests, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and electrospray ionization mass spectrometry.
  
  
  RESULTS
  Without surfactants, protein recovery was only 27-43% for IFN-α2b and 58-73% for EGF. Protein content in solutions incubated with blank microspheres decreased to 66% for IFN-α2b and 86% for EGF. It was only possible to quantify the EGF and IFN-α2b in the same manner as in PBS alone when the surfactant added was Pluronic F-68 and SDS, respectively. Addition of these surfactants allowed the complete isolation of both biomolecules from the microspheres. The extraction procedure did not affect the encapsulated proteins.
  
  
  CONCLUSIONS
  Proteins can be quantitatively extracted, without changes, from PLGA/PLA microspheres using PBS/dichloromethane system that include an appropriate surfactant.
authors.affiliation: Formulation Development Department Center for Genetic Engineering and Biotechnology, PO Box 6162, Havana, Cuba. vivian.saez@cigb.edu.cu
authors.forename: Vivian
authors.initials: V
authors.lastname: Saez
authors.forename: José A
authors.initials: JA
authors.lastname: Ramón
authors.forename: Liurdis
authors.initials: L
authors.lastname: Caballero
authors.forename: Raymersy
authors.initials: R
authors.lastname: Aldana
authors.forename: Elián
authors.initials: E
authors.lastname: Cruz
authors.forename: Carlos
authors.initials: C
authors.lastname: Peniche
authors.forename: Rolando
authors.initials: R
authors.lastname: Paez
mesh_terms: D016475:3T3 Cells; D000818:Animals; D000998:Antiviral Agents / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D004339:Drug Compounding; D004815:Epidermal Growth Factor / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D006801:Humans; D000077190:Interferon alpha-2; D016898:Interferon-alpha / Q000737:chemistry / Q000302:isolation & purification* / Q000494:pharmacology; D019344:Lactic Acid / Q000737:chemistry*; D051379:Mice; D008863:Microspheres; D011100:Polyglycolic Acid / Q000737:chemistry*; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D055550:Protein Stability; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D013501:Surface-Active Agents / Q000737:chemistry*
chemical_list: D000998:Antiviral Agents; D000077190:Interferon alpha-2; D016898:Interferon-alpha; D011994:Recombinant Proteins; D013501:Surface-Active Agents; D000077182:Polylactic Acid-Polyglycolic Acid Copolymer; D011100:Polyglycolic Acid; D019344:Lactic Acid; D004815:Epidermal Growth Factor