Fabrication of PLG microspheres with precisely controlled and monodisperse size distributions

https://doi.org/10.1016/S0168-3659(01)00289-9Get rights and content

Abstract

The size distribution of biodegradable polymer microspheres critically impacts the allowable routes of administration, biodistribution, and release rate of encapsulated compounds. We have developed a method for producing microspheres of precisely controlled and/or monodisperse size distributions. Our apparatus comprises spraying a polymer-containing solution through a nozzle with (i) acoustic excitation to produce uniform droplets, and (ii) an annular, non-solvent carrier stream allowing further control of the droplet size. We used this apparatus to fabricate poly(d,l-lactide-co-glycolide) (PLG) spheres. The acoustic excitation method, by itself, produced uniform microspheres as small as 30 μm in diameter in which ≥95% of the spheres were within 1.0–1.5 μm of the average. The carrier stream method alone allowed production of spheres as small as ∼1–2 μm in diameter from a 100-μm diameter nozzle, but generated broader size distributions. By combining the two devices, we fabricated very uniform spheres with average diameters from ∼5 to >500 μm. Furthermore, by discretely or continuously varying the experimental parameters, we fabricated microsphere populations with predefined size distributions. Finally, we demonstrate encapsulation and in vitro release of a model drug compound, rhodamine B. In summary, our apparatus provides unprecedented control of microsphere size and may allow development of advanced controlled-release delivery systems.

Introduction

Rapid advances in biotechnology have led to the discovery of numerous protein and peptide therapeutics, many of which have recently reached the marketplace or are currently under regulatory review by the United States Food and Drug Administration [1]. Unlike traditional small-molecule drugs, however, proteins and peptides generally cannot be administered orally; injection or infusion is most often required. Further, because of their fragility and short in vivo half-lives, encapsulation of proteins in biodegradable polymeric devices, from which the drug can be delivered, locally or systemically, for a prolonged period of time, has been a promising and intensely studied solution to these problems [2], [3], [4]. Biodegradable microspheres comprising a variety of polymers have been the most studied devices due to relatively simple fabrication and facile administration to a variety of locations in vivo through a syringe needle.

Several methodologies for microsphere fabrication have been described in the literature including precipitation [5], spraying [6], [7], phase separation, and/or emulsion techniques [8], [9], [10]. The emulsion and spraying approaches have been commonly used both at the bench and industrial scales [11], [12]. Sphere size and size distribution are reproducible but often poorly controllable. Standard deviations equal to 25–50% of the mean diameter are not uncommon.

Control of sphere size and size distribution has several important implications for controlled-release drug delivery. For example, there typically is an ideal sphere size that provides a desired release rate and route of administration. Spheres that are ‘too small’ exhibit poor encapsulation efficiency, may migrate from the site of injection, and may exhibit undesirably rapid release of their payload. Spheres that are ‘too large’ may not easily pass through a syringe needle. Thus, the typically polydisperse spheres generated by conventional fabrication techniques must be filtered or sieved to isolate particles within the desired size range, and the polymer and drug composing spheres outside that range are wasted. Further, precise size control may lead to advanced delivery systems not possible with polydisperse spheres. Uniform microspheres approximately 1–5 μm in diameter would be ideal for passive targeting of professional antigen-presenting cells (APCs) such as macrophages and dendritic cells [13], [14]. Similarly, microspheres 10–20 μm in diameter could be used to target the tortuous capillary bed of tumor tissues by chemo-embolization [15]. A system capable of precise microsphere fabrication, therefore, is needed and could allow the optimal size for such applications to be identified and provide an efficient route to commercial manufacture and clinical implementation.

A long-sought goal for controlled-release drug delivery technologies is the ability to precisely control the release rate of encapsulated compounds, and microsphere size is a major determinant of release kinetics. Larger spheres generally release encapsulated compounds more slowly and over longer time periods, other properties (polymer molecular weight, initial porosity, drug distribution within the sphere, etc.) being equal [16]. A constant (i.e. zero-order) release rate is often preferred [17], [18], while variable drug release rates can be beneficial for many important indications [19]. For example, intermittent high doses of antibiotics may alleviate evolution of resistance in bacteria, and discontinuous administration of vaccines often enhances the immune response [20], [21].

Methods to control drug release rate include (i) choice of polymer chemistry (anhydrides, esters, etc.) and comonomer ratios, (ii) conjugating the drug to the polymer [22], (iii) varying the microsphere formulation parameters, and thus the physical characteristics of the resulting particles [23], [24], [25], [26], [27], and (iv) manipulating the sphere size and distribution [16], [28], [29]. The success of the latter studies was limited by the relatively broad microsphere size distributions.

In recent years, there have been several reports of the fabrication of biodegradable polymer microspheres with controlled, uniform size [16], [23], [29], [30], [31]. However, none of these methods was successful in generating particles in a size range appropriate for drug delivery (∼1–100 μm) while maintaining narrow size distributions. In addition, these previous methods appear to be difficult to scale-up for commercial applications.

We have developed a methodology, based on spraying a polymer solution through a small orifice, for fabricating polymeric microspheres. Here we report the capabilities of two techniques, individually and in combination, for generating monodisperse microspheres with precisely controlled sizes from ∼1 to >500 μm in diameter. We have further demonstrated the capability of the apparatus to fabricate predefined particle size distributions via continuous variation of the process parameters.

Section snippets

Materials

Poly(d,l-lactide-co-glycolide) polymers (50:50 lactic acid/glycolic acid; [i.v.]=0.15–0.63 dl/g corresponding to Mw 6,000–52,000) were obtained from Birmingham Polymers. Poly(vinyl alcohol) (PVA; 88% hydrolyzed) was obtained from Polysciences, Inc. Rhodamine B chloride was obtained from Sigma. HPLC grade dichloromethane (DCM) and dimethylsulfoxide were purchased from Fisher Scientific.

Preparation of microspheres

Five- to 20-ml quantities of 5% w/v PLG dissolved in DCM were prepared. In some cases, rhodamine B (1, 3 or 5%

Description and theory of the microsphere generating apparatus

The main apparatus (Fig. 1A), which provides fabrication of monodisperse microparticles, is based on passing a solution containing the sphere material, and any drug to be encapsulated, through a small nozzle or other orifice (20 μm to a few millimeters in diameter) to form a smooth, cylindrical jet. To controllably break the jet into droplets, the nozzle is vibrated by a piezoelectric transducer driven by a wave generator at a frequency tuned to match the flow rate and the desired drop size

Discussion

The sizes of biodegradable polymer microspheres have several critical implications for controlled-release drug delivery. For example, sphere size impacts allowable routes of administration and the final disposition of the spheres in the body. Importantly, microsphere diameter is a determining factor of drug release rates and controlled manipulation of the size may provide a means to tailor release rate profiles. Finally, sphere size can be used to passively target the delivery vehicles for

Conclusions

We have developed a microsphere fabrication process capable of reproducibly generating monodisperse particles as well as populations exhibiting pre-defined size distributions. Using acoustic excitation, we can precisely control microsphere diameter within a range from ∼1- to 10-times the orifice diameter (Table 1). By further employing an annular, non-solvent carrier stream, the sphere-forming liquid can be ‘focused’ into a fine jet, reducing particle size by two orders of magnitude. The

Acknowledgements

We wish to thank Karen Gibbs of Applied Engineering Materials for her assistance with the particle sizing and Dave Garland of Valpey Fisher for donating piezoelectric transducers. This work was funded in part by the University of Illinois Campus Research Board.

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