Journal of Controlled Release 60 (1999) 279–286 Influence of formulation parameters on the characteristics of poly( D,L-lactide-co-glycolide) microspheres containing poly( L-lysine) complexed plasmid DNA a b b b Yilmaz Capan , Byung H. Woo , Sisay Gebrekidan , Shamim Ahmed , Patrick b, * P. DeLuca a Hacettepe University, Faculty of Pharmacy, Department of Pharmaceutical Technology, 06100 Ankara, Turkey b Faculty of Pharmaceutical Sciences, University of Kentucky, College of Pharmacy, Rose Street, Lexington, KY 40536, USA Received 13 January 1999; accepted 9 March 1999 Abstract This study describes the influence of polymer type, surfactant type / concentration, and target drug loading on the particle size, plasmid DNA (pDNA) structure, drug loading efficiency, in vitro release, and protection from DNase I degradation of poly(D,L-lactide-co-glycolide) (PLGA) microspheres containing poly( L-lysine) (PLL) complexed pDNA. PLGA microspheres containing pDNA–PLL were prepared using the water-in-oil-in-water (w–o–w) technique with poly(vinyl alcohol) (PVA) and poly(vinyl pyrrolidone) (PVP) as surfactants in the external aqueous phase. A complex ratio of 1:0.33 (pDNA–PLL, w / w) enhanced the stability of pDNA during microsphere preparation. Higher pDNA–PLL loading efficiency (46.2%) and supercoiled structure (64.9%) of pDNA were obtained from hydrophobic PLGA ( M 31 000) microspheres compared with w hydrophilic PLGA or low-molecular-weight PLGA microspheres. The particle size decreased from 6.6 to 2.2 mm when the concentration of PVA was increased from 1 to 7%. At the same concentration of surfactant, PVA stabilized microspheres showed higher pDNA–PLL loading efficiency (46.2%) than PVP stabilized microspheres (24.1%). Encapsulated pDNA in PLGA microspheres was protected from enzymatic degradation and maintained in the supercoiled form. The pDNA–PLL microspheres showed in vitro release of 95.9 and 84.9% within 38 days from the low-molecular-weight PLGA and hydrophilic PLGA microspheres, respectively, compared to 54.2% release from the hydrophobic, higher-molecular-weight PLGA microspheres. The results suggest loading and release of pDNA–PLL complex can be influenced by surfactant concentration and polymer type.  1999 Elsevier Science B.V. All rights reserved. Keywords: Microsphere; Plasmid DNA; Poly(D,L-lactide-co-glycolide); Poly(L-lysine); DNase 1. Introduction great potential utility for gene replacement therapy, therapeutic applications, and vaccines [1–3]. How- Plasmid DNA (pDNA) has been shown to have ever, the use of pDNA-based pharmaceuticals as drug substances will largely depend on the develop- ment of efficient delivery systems. Free pDNA, *Corresponding author. Tel.: 11-606-257-1831; fax: 11-606- without any delivery system, has been found to be 323-0242. E-mail address: ppdelu1@pop.uky.edu (P.P. DeLuca) highly susceptible to nuclease degradation [4]. It is 0168-3659 / 99 / $ – see front matter  1999 Elsevier Science B.V. All rights reserved. PII: S0168-3659(99)00076-0