Natural and synthetic biodegradable polymers have been investigated for controlled drug release. Poly(3-hydroxybutyrate) can be produced by bacteria and is remarkable for this application due to its excellent biocompatibility and biodegradability. The objective of this work was to study different drug-entrapment and emulsification methods for the obtaining of doxycycline-loaded PHB micro- and nano-spheres. The micro-/nano-particles were prepared by polymer precipitation via dialysis, simple emulsion (O/W) or multiple emulsion (W1/O/W2) applying solvent evaporation in the last two cases. This was carried out either by ultrasonication, dripping and/or high speed stirring. Different processing conditions were varied in order to evaluate their influence on morphology, size, and drug entrapment capabilities. The highest drug loading was obtained by single emulsion with high speed stirring. In the case of multiple emulsion, the combination of ultrasound with high speed stirring resulted in the most elevate process yield and drug loading capability.