A novelin vitro release technique for peptide-containing biodegradable microspheres |
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Authors: | Janusz W Kostanski Patrick P DeLuca |
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Institution: | (1) Pharmaceutical Research Institute, Bristol-Myers Squibb Company, New Brunswick, NJ 08903, USA;(2) Faculty of Pharmaceutical Sciences, University of Kentucky College of Pharmacy, Lexington, KY 40536, USA;(3) Oakwood Laboratories, LLC, Oakwood, OH 44146, USA |
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Abstract: | The purpose of this study was to develop and evaluate a dialysisin vitro release technique for peptide-containing poly(d, 1-lactide-coglycolide) (PLGA) microspheres (ms) that would correlate within vitro data. Using a luteinizing hormone- releasing hormone analogue (LHRH), Orntide acetate, solubility and stability were determined
in 0.1 M phosphate buffer (PB), pH 7.4, and in 0.1 M acetate buffer (AB), pH 4.0, with highperformance liquid chromotography
(HPLC), and peptide permeability through a dialysis membrane (molecular weight cut-off 300,000) was determined. Orntide ms
were prepared by a dispersion/solvent extraction/evaporation method and characterized for drug content (HPLC), particle size
distribution (laser diffraction method), and surface morphology (scanning electron microscopy).In vitro release was studied in PB using a conventional extraction method and with a new dialysis method in AB. Gravimetric analyses
of polymer mass loss and matrix hydration, and peptide adsorption to blank PLGA ms (50∶50, Mw 28 022) were carried out in PB and AB upon incubation at 37°C. Serum Orntide and testosterone levels in rats after administration
of Orntide ms were determined by radioimmunoassay. Orntide acetate solubility was influenced by pH; approximately 2.3 mg/mL
dissolved in PB and >18 mg/mL in AB. Stability was pH- and temperature-dependent. The peptide was very stable at pH 4.0, 4°C,
but degraded rapidly at pH 7.4,37°C. Peptide permeability through the dialysis membrane was accelerated by agitation and>95%
equilibrium was reached within 48 hours. The overall release rate was higher with the dialysis method. Mass loss of the Orntide
ms was faster in AB (50% loss in 3 weeks: 95% in 35 days) than in PB (65% in 35 days). In contrast, hydration after 35 days
was 4-fold higher in PB. The nonspecific adsorption to blank ms was greater in PB (128 μ g Orntide/10 mg PLGA) compared with
AB (<5 μ g Orntide/10 mg PLGA). Administration of 30-day Orntide PLGA ms to rats resulted in an initial serum Orntide level
of 21 ng/mL after 6 hours and a Cmax of 87 ng/mL after 6 days. Testesterone levels were suppressed immediately after ms administration (3 mg Orntide/Kg) from
5.2 ng/mL to 0.3 ng/mL (after 24 hours) and remained suppressed for 38 days. Orntide acetate solubility and degradation kinetics
were markedly influenced by pH of the buffer systems and mass loss; matrix hydration, as well as the nonspecific adsorption
to blank ms, was pH-dependent. Thein vitro release profile obtained with the dialysis method in AB correlated well with thein vivo data, therepy providing a more reliable prediction ofin vivo performance. |
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Keywords: | Orntide Microspheres Solubility Stability In vitro release Dialysis PLGA |
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