Heterogeneous Crystallization Inside Microporous Polymer Particles

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Heterogeneous Crystallization Inside Microporous Polymer Particles as a Process Intensification Technology for the Manufacture of Drug Formulations Jing Ling, and Keith Chadwick Org. Process Res. Dev., Just Accepted Manuscript • Publication Date (Web): 28 Apr 2017 Downloaded from http://pubs.acs.org on April 29, 2017

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Organic Process Research & Development

Heterogeneous Crystallization Inside Microporous Polymer Particles as a Process Intensification Technology for the Manufacture of Drug Formulations

Jing Ling, and Keith Chadwick* Department of Industrial and Physical Pharmacy, College of Pharmacy, Purdue University, West Lafayette, Indiana, United States

* Address correspondence to this author at the Department of Industrial and Physical Pharmacy, College of Pharmacy, Purdue University, West Lafayette, Indiana 47907-2091, United States; Tel/Fax: +1 765-496-2775, +1 765-494-6545; email: [email protected]

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Abstract Current pharmaceutical manufacturing practices lack the type of rapid advancements achieved through the quick adoption of technical innovations as has been done in other manufacturing sectors. There is significant interest from both academia and the pharmaceutical industry in developing new technologies that accelerate process optimization in a cost effective manner. Process Intensification Technologies (PITs) improve efficiency and reduce costs by reducing the number of unit operations required to manufacture a drug formulation. Recent studies have shown the potential of directly crystallizing drug compounds inside porous excipients as a PIT for pharmaceutical manufacturing. However, these studies have primarily focused on developing techniques for crystallizing the drug inside the excipients and maximizing drug loading. There is little understanding of how the excipient particles themselves influence the crystallization process or how this technology compares with other drug encapsulation processes. This investigation considers two other drug encapsulation processes, direct crystallization onto nonporous particles and molecular adsorption of the drug. We discuss the direct crystallization of acetaminophen and sulfathiazole inside micro-porous particles of alginate and carboxymethyl cellulose. The polymer chemistry and available surface area have a significant impact on drug loading and secondary nucleation in the bulk solution. Direct crystallization offers higher drug loadings compared to other drug encapsulation technologies. This study highlights the potential of direct crystallization inside excipients as a PIT for pharmaceutical manufacturing.

Keywords: pharmaceutical manufacturing; process intensification technology; heterogeneous nucleation; direct crystallization; porous particles.



1. Introduction Current pharmaceutical manufacturing practices lack the type of rapid advancements achieved through the quick adoption of technical innovations as has been done in other manufacturing sectors such as electronics, fine chemicals and foods. Recent studies have estimated manufacturing costs to be as high as 30% of total global sales for manufacturers of brand-name pharmaceuticals and 60% for generics.1 In the past decade, leaders in both the pharmaceutical industry and the FDA recognized that pharmaceutical drug development and manufacturing needed to evolve from an art form into one based in science and engineering, and to adopt “quality by design” (QbD) techniques.2 The ultimate goal is to develop new and efficient technologies that improve both manufacturing and the quality of pharmaceutical products. Continuous manufacturing and process intensification technology (PIT) are two strategies that have received considerable research investment over the past decade.3-7 The utilization of crystallization based PITs has potential benefits over other comparable technologies including; the simplicity of the process, formulation stability, improved drug loading, and the potential to perform purification of the API simultaneously.8-13 Moreover, crystallization processes can be designed to control the crystal form and consequently, the physicochemical properties of the API.14, 15 Heterogeneous crystallization based PITs have begun to receive a significant amount of attention, including direct crystallization in/on excipient particles, emulsion crystallization, and drop printing drug solutions on polymer thin films.8, 16, 17 In particular, direct crystallization into excipients has many potential benefits such as, simplicity of the process and capability to scale up. In a recent study, Eral et al. heterogeneously crystallized APIs inside hydrogels, achieving ~30% drug loadings for hydrophilic drugs and


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~80% for hydrophobic drugs.18 However, the hydrogel particles were required to reside in a saturated solution of the hydrophilic drug for 3 days to ensure equilibrium partitioning of the drug into the hydrogel. In addition, the need to create a suitable emulsion within the hydrogel particles in order to achieve high loadings of hydrophobic drugs added complexity to the manufacturing process. To address those potential issues, in a previous study we proposed a PIT based on direct crystallization into micro-porous polymer particles.19 Figure 1 summarizes this proposed downstream manufacturing platform and compares it to a traditional downstream pharmaceutical manufacturing process.20 In the proposed platform engineered microporous particles are added to the crystallizer prior to supersaturation generation. Crystallization is then carried out using established approaches, such as cooling, evaporation or anti-solvent addition. The drug crystallizes inside the particles, which are then filtered and dried. Subsequently, the drug loaded particles are either compressed into tablets or filled into capsules resulting in the final drug product. The potential benefits of directly crystallizing the drug within micro-porous particles are: (1) improved efficiency in manufacturing through removal of unit operations such as blending, granulation and milling, and (2) the ability to manufacture novel drug/excipient formulations. In the study by Acevedo et al., Process Analytical Technology (PAT) was implemented to control the crystallization of acetaminophen (AAP) into micro-porous alginate particles.19 The application of PAT allowed for greater control of drug distribution within the particles and increase the uniformity of the formulation.



Figure 1. Traditional versus Proposed Process Intensification Technology Manufacturing Platforms

However, there are still challenges to overcome in transforming direct crystallization of drugs into microporous particles into a viable technology for implementation in pharmaceutical manufacturing: (1) understanding the impact of polymer chemistry on drug loading, (2) eliminating bulk crystallization (drug crystallization occurring in the solution), and (3) demonstrating the effectiveness of this technique in producing high drug loading formulations compared to other established drug loading technologies. Herein, we discuss the effect of drug/polymer interactions and available polymer surface area on crystallization behavior. Moreover, we evaluate the drug loading capability of direct crystallization into micro-porous polymer particles against two other established encapsulation techniques: crystallization on nonporous particles and molecular adsorption of drug into porous particles. Finally, we assess the effect of particle surface area on the elimination of bulk crystallization. AAP and sulfathiazole (STZ) were used as model drug compounds in this study. Sodium alginate (ALG) and sodium carboxymethyl cellulose (CMC) were chosen as the micro-porous polymer particles based on (1) the chemistry of the polymer and its ability to form favorable interactions with drug and (2) the ease of manufacturing porous particles. In this report, we demonstrate that polymer chemistry, drug encapsulation process, and surface area of the polymer particles were all critical factors for achieving high drug loading inside pharmaceutically acceptable excipient particles, as well as for enabling a significant reduction in secondary nucleation. 2. Materials and Methods 2.1. Materials


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Sodium alginate (Na ALG) (viscosity 5.0 – 40.0 cps), sodium carboxymethyl cellulose (Na CMC) (2500 – 6000 cps), calcium chloride (CaCl2), aluminum chloride (AlCl3), Pluronic F108 (F108), sodium bicarbonate, and acetaminophen (AAP) were purchased from Sigma (St. Louis, MO, USA). Sulfathiazole (STZ) was obtained from Alfa Aesar. (Haverhill, MA, USA). Glacial acetic acid was received from Amresco (Solon, OH, USA). Sodium phosphate monobasic monohydrate and sodium phosphate dibasic anhydrous were purchased from Macron Fine Chemicals (Center Valley, PA, USA). 2.2. Methods 2.2.1. Preparation of Porous Calcium Alginate (Ca ALG) and Aluminum Carboxymethyl cellulose (Al CMC) Particles The methods for producing ALG and CMC particles were adapted from the work of Eiselt et al. 21

An aqueous alginate solution was prepared by stirring a mixture of 5% (w/w) Na ALG, 0.54%

(w/w) F108, and 0.9% (w/w) sodium bicarbonate. The alginate solution was homogenized at 5000 rpm until it had completely converted into a foam. Porous alginate particles were then formed by extruding drops of the foam from a pipette into a 0.1 M CaCl2 solution containing 10% (v/v) acetic acid under constant stirring for 5 min. The particles were collected, washed with deionized water, and freeze dried using a Labconco FreeZone freeze drier (Kensas City, MO, USA). For CMC particles, a mixture of 1.6% (w/w) Na CMC, 0.81% (w/w) F108, and 0.9% (w/w) sodium bicarbonate was dissolved in water. Droplets of the resulting solution were then extruded into an aqueous solution of 0.1 M AlCl3 with 10% (v/v) acetic acid. The droplets were exposed to the AlCl3 solution for 30 seconds while being constantly stirred. The particles were then transferred into a separate 0.1 M AlCl3 solution and incubated for an additional 15 min. The particles were collected, washed with deionized water, and freeze dried.



2.2.2. Preparation of Non-porous Ca ALG and Al CMC Particles Non-porous Ca ALG and AL CMC particles were prepared by extruding a Na ALG and Na CMC solutions (5%) into a 0.1 M CaCl2 and AlCl3 solution, respectively, stirred at 500 rpm. The particles were collected, washed with deionized water, and freeze-dried. 2.2.3. Preparation of Drug-loaded Particles by Direct Crystallization For AAP, a 30 mL saturated solution (saturated with respect to crystal Form I) in ethanol at 30 °C was prepared (277 mg/g) in a 100 mL crystallizer. Fifty porous ALG particles were weighed and then added into the solution and stirred with a magnetic stirring bar at 500 rpm for 1 hour. The structure of the porous particles during the crystallization process was robust. No attrition or annealing were observed under such experimental or scale up conditions. 19 For the rapid cooling crystallizations, the solution was quench cooled to 25 °C with supersaturation of 1.1 (σ = c* / ceq). Once crystallization was observed in the bulk solution, the temperature of the crystallizer was maintained at 25 oC for one hour to allow the system to reach equilibrium. The particles were then decanted from the solution and air-dried. The weight of the particles was monitored until no change was observed, at which time the final weight was recorded. For STZ, a saturated solution (saturated with respect to Form III) in ethanol at 45°C was prepared (8.75mg/g). Thereafter the same protocol used for AAP was followed, with the exception that the solution was cooled to 15°C, resulting in a supersaturation of 2.75. For non-porous particles, the same crystallization process was used. 2.2.4. Preparation of Drug-loaded Particles by Adsorption The drugs were loaded into the microporous particles by suspending 50 particles in saturated ethanolic solutions of either AAP or STZ at 30 and 45°C, respectively, for ~2 hrs. Thereafter, the particles were decanted, dried and weighed.


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2.2.5. Particle size measurements The diameters (n=30) of ALG and CMC particles were measured using a Tool Shop digital Vernier caliper (Menards Inc., Eau Claire, WI, USA). 2.2.6. Particle Morphology Images of the surface and cross-sectional morphologies of the particles were produced using scanning electron microscopy (SEM). The particles were mounted on the holder and coated with platinum under vacuum using a Cressington sputter coater (208HR, Cressington Scientific Instruments, Watford, United Kingdom). The coated samples were then analyzed using a Nova NanoSEM (FEI, OR, USA) with an accelerating voltage of 5kV. 2.2.7. Characterization of true density and porosity of beads The true density of the polymer beads, as well as percentage porosity, were obtained using pycnometry (Micromeritics GeoPyc 1360 V1.03, Micrometrics Limited, Dunstable, United Kingdom) in conjunction with helium displacement pycnometry (Micromeritics AccuPyc 1330 V3.00, Micrometrics Limited, Dunstable, United Kingdom). Three batch samples were analyzed to obtain statistically relevant data. 2.2.8. Drug loading analysis The drug loading inside the particles was determined by gravimetric analysis. The weight of particles was determined before (m1) and after the loading process (m2). The elimination of bulk crystallization was evaluated by measuring the encapsulation efficiency, which is defined as the percentage of the total drug crystal yield present in the particles. For each drug/polymer system, the drug loading and encapsulation efficiency determination was carried out on four different batches of particles (50 particles each). The drug loading and the encapsulation efficiency were calculated as follows:



(1)

(2)

where m1 is the weight of blank alginate particles, m2 is the gross weight of AAP loaded alginate particles, and m3 is the total weight of drug crystallized at the end of the crystallization process.

Results 3.1. Engineering ALG and CMC particles Porous ALG and CMC particles were manufactured by foam extrusion.21-23 Figure 2 shows the surface and cross-sectional SEM images of ALG and CMC microporous particles. Both ALG and CMC particles exhibited internal and surface porosity with an interconnected pore structure. The interior of the particles exhibited greater interconnectivity and porosity than the surface. The average size of the ALG and CMC particles was 4.4 ± 0.1 mm and 3.5 ± 0.2 mm, respectively.


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Figure 2. SEM images showing (A) surface and (B) internal morphology of ALG particles, and (C) surface and (D) internal morphology of CMC particles.

3.2. Direct Crystallization of AAP and STZ into Microporous ALG and CMC particles Based on the propensity to form favorable interaction with polymeric particles, AAP and STZ were selected as model compound. AAP was directly crystallized into ALG and CMC particles. AAP crystals were confirmed to nucleate and grow inside both the ALG and CMC particles (Figure 3). PXRD analysis was used to determine that these crystals were the thermodynamically stable polymorph Form I (see Supporting Information). SEM analysis of the AAP loaded particles was performed to qualitatively assess the distribution of drug throughout the particles. Heterogeneous crystallization was observed on the external, as well as interior surfaces of the particles for both drugs. As shown in Figure 3a, the distribution of AAP crystals in the ALG particles was heterogeneous, with a greater density of crystals in the pores close to the surface compared to the core. For the CMC particles, the distribution was more homogeneous with



significant crystallization observed throughout the particles (Figure 3d). However, the crystal density still appeared greater closer to the surface. The polymorphism of the STZ crystals inside the particles could not be determined due to low drug loading (Figure. 6). For STZ, SEM analysis of both the ALG and CMC showed crystallization on the internal surfaces of the particles (Figure. 4). Furthermore, STZ crystals were observed in greater density closer to the surface for both ALG and CMC particles, with no crystals visible in the center (Figure. 4b, d). A more detailed analysis of the 3D drug distribution by established techniques, such as EDX and Raman mapping, could not be carried out for a variety of reasons: (1) the same elemental constituents in drug and polymer (AAP), (2) low concentration of drug crystals within the polymer matrix (STZ) and (3) special resolution of the instrument (Raman mapping)19.

Figure 3. SEM of the internal structure and morphology of AAP loaded ALG and CMC particles. Crosssectional view of AAP loaded ALG particles at (A) 71x magnification, and (B, C) 2500x magnifications for the edge and center of the particles, respectively. Cross-sectional view of AAP loaded CMC at (D) 75x magnification, and (E, F) 2500x magnification for the edge and center of the particles, respectively.


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Figure 4. SEM of internal structure and morphology of STZ loaded ALG and CMC particles. Crosssectional view of STZ loaded ALG particles at (A) 57x and (B) 2500x. Cross-sectional view of STZ loaded CMC at (C) 73x and (D) 2500x

3. 4. Effect of Encapsulation Method on Drug loading of ALG and CMC particles The drug loading capability of different drug encapsulation methods was determined. The encapsulation methods used were as follows: (1) direct crystallization into micro-porous particles, (2) direct crystallization on non-porous particles and (3) adsorption of molecular drug into micro-porous particles. The drug loading results are summarized in Table 1.

Table 1. Drug loading of AAP in ALG/CMC microporous particles by cooling crystallization and adsorption

Drug/Polymer System

Drug Loading (%w/w) Crystallization Porous Particles

Adsorption on Non-Porous Particles Porous Particles



AAP /ALG AAP/CMC

71.0±4.8 65.1±2.5

36.8 ± 7.8 51.9±7.5

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58.8±9.0 51.7±4.0

It was found that direct crystallization into microporous particles achieved higher drug loadings than the other techniques (Table 1), with drug loadings of up to ~70%. For the molecular drug adsorption method, the drug loading (~59%) is limited by the available surface area for adsorption to occur. Once all the surface area is depleted, no further drug adsorption can take place. Whereas, for direct crystallization in microporous particles, the drug loading is determined by both the available surface area and internal free volume. The available surface area dictates the amount of heterogeneous nucleation that may occur, whereas the internal free volume of the pores determines the extent of crystal growth. The ability to utilize the internal free volume by direct crystallization allows for higher drug loadings. In the case of crystallization on non-porous particles, the drug loading is significantly lower compared to crystallization in microporous particles due to (1) significantly reduced surface area on which heterogeneous nucleation can occur and (2) attrition of crystals grown on the surface of the particles. Direct crystallization inside microporous particles protects crystals from attrition taking place at the external surface and may delay or prevent secondary nucleation from occurring allowing for further crystal growth inside the particles. Furthermore, we have also compared the drug loadings achievable by direct crystallization in microporous particles to those reported using other methods in previous studies. These studies showed that when encapsulating hydrophilic drugs in particles using the capture of preformed crystals or adsorption, the maximum drug loadings observed were between 15 to 30%.18, 24-27 Even when using a modified method designed in our laboratory to encapsulate preformed crystals into a hydrogel matrix, the maximum drug loading of AAP achieved was 44.8 ± 3.8%.28


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We hypothesize that the reason encapsulation of preformed crystals in hydrogel matrices yields lower drug loadings than direct crystallization in microporous hydrogel particles is due to particle stability during manufacturing. Specifically, stable particles cannot form with increasing drug loading.28 Patel et al. suggest that higher loadings of preformed crystals could not be achieved because increasing the crystal density within the hydrogel solution beyond a critical limit prior to ionotropic gelation reduces the amount of cross-linking within the hydrogel particle to the point at which it becomes unstable. In the case of direct crystallization, stable hydrogel particles of ALG and CMC have already been produced prior to crystallization occurring. Therefore, high densities of crystals already present inside the particles cannot influence particle stability. Finally, in a recent study by Eral et al., APIs were heterogeneously crystallized inside nanoporous hydrogel particles. In that study they could only achieve ~30% drug loadings for AAP.18 Though the hydrogel particles were incubated in a AAP solution for days, the drug loading was only ~40% of that achievable by direct crystallization into microporous particles. We hypothesize that this is due to the greater internal free volume inside the microporous particles. That is, greater internal space allowing greater crystal growth is important to achieving high drug loadings. 3. 5. Effect of Surface Chemistry in Drug loading of ALG and CMC particles In previous studies of heterogeneous nucleation, the molecular functionality of both the surface and the solute have been shown to play an important role. Both nucleation kinetics and crystal form are highly sensitive to the intermolecular interactions between surface and nucleating crystal plane.29, 30 Surfaces that have functional groups capable of forming strong intermolecular interactions with the solute make heterogeneous nucleation more favorable.30 We hypothesize that surfaces containing functional groups also present in the drug will result in higher drug



loadings. It is expected that the formation of favorable intermolecular interactions between the drug and polymer particles will lead to preferential crystallization inside the particles and reduce bulk crystallization in the solution. To test this hypothesis, we have studied the crystallization of AAP and STZ in ALG and CMC particles. ALG and CMC both have functional groups also present in AAP, namely, the hydroxyl group (OH), whereas STZ does not share a functional group match with either polymer (Figure 5). Figure 6 shows that AAP has significantly higher drug loadings in both ALG and CMC microporous particles (71.0±4.8% and 65.1±2.5% for ALG and CMC porous particles, respectively) compared to STZ (15.2±5.8% and 7.6±6.4% for ALG and CMC porous particles, respectively).

Figure 5. Chemical Structure of AAP, STZ, Na ALG, and Na CMC


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Figure 6. Drug loading of AAP/STZ in ALG/CMC particles by cooling crystallization and adsorption In order to prove that functional group matching was responsible for the difference in drug loadings of AAP and STZ in the particles, it was necessary to eliminate other factors that could be responsible. First, we examined encapsulation efficiency, which describes what percentage of the total crystal yield (mass of crystals) of AAP and STZ was crystallized inside the ALG and CMC particles. Due to (1) the differences in solubility of AAP and STZ in ethanol and (2) the crystallization conditions selected for this study, the theoretical crystal yield for AAP is 4.5 times greater than STZ. If the encapsulation efficiency of each drug in the particles were 100%, then the difference in drug loadings could be explained by the difference in drug yield. In order to rule this out as a possibility, the experimental encapsulation efficiencies for AAP and STZ in ALG and CMC particles were measured. Table 2 shows that the encapsulation efficiency for AAP in the particles was higher than that of STZ, where the encapsulation efficiencies for STZ in ALG and CMC particles was 7.5% and 0.97%, respectively. This demonstrated that the low crystal yield of STZ was not responsible for the low drug loadings observed in ALG and CMC particles. Thereafter, we examined whether or not drug permeability may have influenced drug loading. However, this was ruled out as a possibility as the particles have an interconnected pore structure meaning that the drug does not have to permeate through the hydrogel in order to access the



interior of the particles. As further demonstration that functional group matching between drug and surface influences drug loading, we studied the nucleation density of AAP and STZ on thin films of CMC. ALG could not be tested as it was not possible to manufacture viable thin films. By using thin films, the issue of drug permeability is removed as a factor. If AAP and STZ have the same heterogeneous crystallization propensity on CMC surfaces, it would be expected that similar nucleation densities of AAP and STZ would be observed on the thin films. Figure 7 shows that crystallization of AAP on CMC results in a significantly higher nucleation density compared to STZ. These data provide strong evidence that the difference in drug loadings for AAP and STZ in ALG and CMC particles is due to the ability of the drugs to interact with the ALG and CMC surfaces. STZ is more hydrophobic than AAP and as a consequence cannot form the same favorable intermolecular interactions with hydrophilic groups present in polymers such as ALG and CMC that are required for favorable heterogeneous nucleation. STZ is limited in its ability to adsorb onto the ALG and CMC surfaces and crystallize within the porous structure, resulting in lower drug loading.

Figure 7. Heterogeneous Crystallization on CMC films (A) AAP crystals (B) STZ crystals 3. 5. Impact of Particle Surface Area on Encapsulation Efficiency


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The purpose of this part of the study was (1) to compare the encapsulation efficiency of heterogeneous crystallization in/on porous and non-porous particles and (2) to investigate the effect of increasing the available surface area of microporous particles on drug encapsulation efficiency. Heterogeneous nucleation onto solid excipient particles has been previously studied in pharmaceutical manufacturing and formulation research and development.8 Ideally, the surfaces of the particles should be completely covered by the drug once the crystallization process is complete. However, results showed minimal coverage of the surfaces by drug crystals. This is likely caused by hydrodynamic forces in solution induced by stirring agitation applied during the pharmaceutical crystallization processes. After heterogeneous nucleation occurs, attrition results in crystal nuclei breakage from the particle surfaces, which may lead to the extensive secondary nucleation in the bulk solution. Low encapsulation efficiency of the drug onto or into the particles negates the potential advantages of manufacturing formulations by heterogeneous crystallization. We hypothesized that increasing the available surface area for direct internal crystallization would result in greater encapsulation efficiency and decrease bulk/secondary nucleation. The theoretical crystal yield of the AAP crystallizing during our experiments was 1.596g. The encapsulation efficiency of 50 non-porous ALG particles was only 0.9% (Table 2). In contrast, for the microporous particles, when adding 50 ALG particles to the crystallizer, the encapsulation efficiency was 22.8%. This represents a 22.1% reduction in secondary nucleation when using porous over non-porous particles. A decrease in secondary nucleation was also observed for CMC, however, it was significantly less than that of ALG, 8.3%. A similar trend was observed for STZ when using microporous ALG particles compared to non-porous particles. The decrease in STZ bulk/secondary nucleation was 4.9%. There was no



difference in encapsulation efficiency for CMC, possibly due to the inability of STZ to crystallize on CMC, irrespective of the geometry of the surface. Table 2. AAP and STZ encapsulation efficiency of ALG, CMC porous particles and ALG/CMC nonporous particles by direct crystallization

Drug/Polymer System AAP /ALG AAP/CMC STZ/ALG STZ/CMC

Encapsulation Efficiency Porous Particles 22.8 ± 5.4 17.0 ± 4.0 7.5 ± 5.8 0.97 ±1.0

Non-Porous Particles 0.9 ± 0.2 9.5 ± 2.8 2.8±1.9 1.5±1.1

ALG micro-porous particles have a greater internal volume than the corresponding CMC microporous particles. ALG particles have a porosity of 95.6 ± 0.5%, while CMC particles have a porosity of 93.9 ± 0.2%. As such, it is hypothesized that ALG microporous particles can provide more free volume in which crystal growth of AAP and STZ can occur, leading to greater encapsulation efficiency. Furthermore, higher AAP encapsulation efficiency for non-porous particles was observed for CMC than ALG. Though the chemistry for the microporous and nonporous particles are the same, the surface area of these particles may play an important role in drug loading capacity. The CMC non-porous particles have a larger diameter than the corresponding ALG particles (Figure 8), providing more surface area on which the drug can nucleate. Therefore, the effect of particle surface area on encapsulation efficiency was also investigated. The available surface area for nucleation was increased by increasing the number of particles added to the crystallizer. The encapsulation efficiency (Eq. 2) increased as a function of the number of microporous particles added during the direct crystallization process as illustrated in Figure 9. It was possible to significantly reduce bulk/secondary nucleation by increasing the available surface area available. Doubling the surface area by doubling the number of particles,


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led to a 29.9% decrease in secondary nucleation. While it is theoretically possible to eliminate all secondary nucleation through the addition of more particles, practically this strategy is limited by the number of particles it is feasible to add to the crystallizer. In order to further increase the encapsulation efficiency and completely eliminate secondary nucleation, it might be expected that the solution is to engineer the microporous particles with greater surface area. However, the tradeoff for creating particles with greater surface area is to reduce the internal free volume of the particle. As such, the available space for crystal growth, which is also a critical factor for drug loading and encapsulation efficiency, would be reduced. Therefore, in order to further eliminate/reduce secondary nucleation, other strategies must be considered. Finally, it was also observed that for AAP there was significant crystallization on the surfaces of both the ALG and CMC porous particles (Figure 10 (A, B)). It is possible that surface crystallization inhibits the free drug in solution access to the core of the particles, reducing the amount of further crystallization in the interior of the particles and limiting encapsulation efficiency. However, surface crystallization was not observed for STZ (Figure 10 (C, D)). This suggests that while surface crystallization may contribute to limiting drug loading and encapsulation efficiency in certain cases, it is not the predominant factor. As such, investigations are currently underway on a number of chemistry and processing based strategies ensuing from this study, in order to address the issue of secondary nucleation and further improving encapsulation efficiency.



Figure 8. SEM Image of Non-porous (A) ALG and (B) CMC particle

Figure 9. Encapsulation efficiency of AAP as the function of the number of particle.


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Figure 10. SEM of the surface structure and morphology of AAP and STZ loaded ALG and CMC particles. (A) AAP loaded ALG particles at 75x magnification, (B) AAP loaded CMC particles at 72x magnification, (C) STZ loaded ALG particles at 73x magnification, and (D) STZ loaded CMC particles at 73x magnification.

4. Conclusion In this study, we have shown how direct crystallization of drugs into microporous excipient particles can be used as a potential PIT to manufacture formulations with high drug loading. It has been demonstrated that this technology offers higher drug loadings compared to two other established drug encapsulation processes. Selecting polymers with functional groups conducive to favorable interactions with the drug molecule is critical to promoting heterogeneous crystallization within the particles and for maximizing drug loading. Finally, both internal free volume and available surface area play an important role in maximizing drug loading,



encapsulation efficiency and reducing secondary nucleation. While the use of heterogeneous crystallization for producing novel drug formulations and improving the efficiency of manufacturing through the elimination of multiple unit operations has become a growing area of research, little consideration has been given to the role of the excipient. Our research demonstrates the necessity of considering the chemistry and material properties of the excipient particles in engineering viable processes for producing formulations with optimized drug loading using heterogeneous crystallization as a PIT technology. 5. Funding Sources This work was supported by Purdue University, College of Pharmacy, West Lafayette, Indiana, USA 6. Supplementary Information The Supplementary information provides the crystal form of AAP in the polymer particles.


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Reference (1) Basu, P.; Joglekar, G.; Rai, S.; Suresh, P.; Vernon, J., Analysis of Manufacturing Costs in Pharmaceutical Companies. J Pharm Innov 2008, 3, (1), 30-40. (2) McKenzie, P.; Kiang, S.; Tom, J.; Rubin, A. E.; Futran, M., Can pharmaceutical process development become high tech? AIChE Journal 2006, 52, (12), 3990-3994. (3) Buchholz, S., Future manufacturing approaches in the chemical and pharmaceutical industry. Chemical Engineering and Processing: Process Intensification 2010, 49, (10), 993-995. (4) Zhao, H.; Shao, L.; Chen, J.-F., High-gravity process intensification technology and application. Chemical Engineering Journal 2010, 156, (3), 588-593. (5) Leuenberger, H., New trends in the production of pharmaceutical granules: batch versus continuous processing. European Journal of Pharmaceutics and Biopharmaceutics 2001, 52, (3), 289-296. (6) Plumb, K., Continuous Processing in the Pharmaceutical Industry: Changing the Mind Set. Chemical Engineering Research and Design 2005, 83, (6), 730-738. (7) Singh, R.; Ierapetritou, M.; Ramachandran, R., An engineering study on the enhanced control and operation of continuous manufacturing of pharmaceutical tablets via roller compaction. Int J Pharm 2012, 438, (1–2), 307-326. (8) Quon, J. L.; Chadwick, K.; Wood, G. P. F.; Sheu, I.; Brettmann, B. K.; Myerson, A. S.; Trout, B. L., Templated Nucleation of Acetaminophen on Spherical Excipient Agglomerates. Langmuir 2013, 29, (10), 3292-3300. (9) Lee, Y. H.; Mei, F.; Bai, M. Y.; Zhao, S.; Chen, D. R., Release profile characteristics of biodegradable-polymer-coated drug particles fabricated by dual-capillary electrospray. Journal of controlled release : official journal of the Controlled Release Society 2010, 145, (1), 58-65. (10) Verreck, G.; Chun, I.; Rosenblatt, J.; Peeters, J.; Dijck, A. V.; Mensch, J.; Noppe, M.; Brewster, M. E., Incorporation of drugs in an amorphous state into electrospun nanofibers composed of a water-insoluble, nonbiodegradable polymer. Journal of Controlled Release 2003, 92, (3), 349-360. (11) Singh, M. N.; Hemant, K. S.; Ram, M.; Shivakumar, H. G., Microencapsulation: A promising technique for controlled drug delivery. Research in pharmaceutical sciences 2010, 5, (2), 65-77. (12) Breitenbach, J., Melt extrusion: from process to drug delivery technology. European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik e.V 2002, 54, (2), 107-17. (13) Hu, X.; Liu, S.; Zhou, G.; Huang, Y.; Xie, Z.; Jing, X., Electrospinning of polymeric nanofibers for drug delivery applications. Journal of Controlled Release 2014, 185, 12-21. (14) Lovette, M. A.; Browning, A. R.; Griffin, D. W.; Sizemore, J. P.; Snyder, R. C.; Doherty, M. F., Crystal Shape Engineering. Industrial & Engineering Chemistry Research 2008, 47, (24), 9812-9833. (15) Abu Bakar, M. R.; Nagy, Z. K.; Saleemi, A. N.; Rielly, C. D., The Impact of Direct Nucleation Control on Crystal Size Distribution in Pharmaceutical Crystallization Processes. Crystal Growth & Design 2009, 9, (3), 1378-1384. (16) Leon, R. A. L.; Wan, W. Y.; Badruddoza, A. Z. M.; Hatton, T. A.; Khan, S. A., Simultaneous Spherical Crystallization and Co-Formulation of Drug(s) and Excipient from Microfluidic Double Emulsions. Crystal Growth & Design 2014, 14, (1), 140-146.



(17) Hsu, H.; Adigun, O. O.; Taylor, L. S.; Murad, S.; Harris, M. T., Crystallization of acetaminophen on chitosan films blended with different acids. Chem Eng Sci 2015, 126, 1-9. (18) Eral, H. B.; López-Mejías, V.; O’Mahony, M.; Trout, B. L.; Myerson, A. S.; Doyle, P. S., Biocompatible Alginate Microgel Particles as Heteronucleants and Encapsulating Vehicles for Hydrophilic and Hydrophobic Drugs. Crystal Growth & Design 2014, 14, (4), 2073-2082. (19) Acevedo, D. A.; Ling, J.; Chadwick, K.; Nagy, Z. K., Application of Process Analytical Technology-Based Feedback Control for the Crystallization of Pharmaceuticals in Porous Media. Crystal Growth & Design 2016, 16, (8), 4263-4271. (20) Schaber, S. D.; Gerogiorgis, D. I.; Ramachandran, R.; Evans, J. M. B.; Barton, P. I.; Trout, B. L., Economic Analysis of Integrated Continuous and Batch Pharmaceutical Manufacturing: A Case Study. Industrial & Engineering Chemistry Research 2011, 50, (17), 10083-10092. (21) Eiselt, P.; Yeh, J.; Latvala, R. K.; Shea, L. D.; Mooney, D. J., Porous carriers for biomedical applications based on alginate hydrogels. Biomaterials 2000, 21, (19), 1921-1927. (22) Wang, C.; Liu, H.; Gao, Q.; Liu, X.; Tong, Z., Alginate–calcium carbonate porous microparticle hybrid hydrogels with versatile drug loading capabilities and variable mechanical strengths. Carbohydrate Polymers 2008, 71, (3), 476-480. (23) J. Mallick, D. S., D. M. Kar, J. Makwana, Alginate Beads of Ibuprofen for Oral Sustained Drug Delivery: an in vitro Evaluation. IJPCBS 2013, 3, 595-602. (24) Wang, S. B.; Chen, A. Z.; Weng, L. J.; Chen, M. Y.; Xie, X. L., Effect of drug-loading methods on drug load, encapsulation efficiency and release properties of alginate/poly-Larginine/chitosan ternary complex microcapsules. Macromolecular bioscience 2004, 4, (1), 2730. (25) Yu, C. Y.; Jia, L. H.; Cheng, S. X.; Zhang, X. Z.; Zhuo, R. X., Fabrication of microparticle protein delivery systems based on calcium alginate. Journal of microencapsulation 2010, 27, (2), 171-7. (26) Whitehead, L.; Collett, J. H.; Fell, J. T., Amoxycillin release from a floating dosage form based on alginates. Int J Pharm 2000, 210, (1-2), 45-9. (27) Gaudio, P. D.; Auriemma, G.; Mencherini, T.; Porta, G. D.; Reverchon, E.; Aquino, R. P., Design of alginate-based aerogel for nonsteroidal anti-inflammatory drugs controlled delivery systems using prilling and supercritical-assisted drying. J Pharm Sci 2013, 102, (1), 185-94. (28) Patel, M. A.; AbouGhaly, M. H.; Schryer-Praga, J. V.; Chadwick, K., The effect of ionotropic gelation residence time on alginate cross-linking and properties. Carbohydr Polym 2017, 155, 362-371. (29) Diao, Y.; Myerson, A. S.; Hatton, T. A.; Trout, B. L., Surface Design for Controlled Crystallization: The Role of Surface Chemistry and Nanoscale Pores in Heterogeneous Nucleation. Langmuir 2011, 27, (9), 5324-5334. (30) Chadwick, K.; Chen, J.; Myerson, A. S.; Trout, B. L., Toward the Rational Design of Crystalline Surfaces for Heteroepitaxy: Role of Molecular Functionality. Crystal Growth & Design 2012, 12, (3), 1159-1166.


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Heterogeneous Crystallization Inside Microporous Polymer Particles

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