Impact of additives on drug particles during liquid antisolvent crystallization and subsequent freeze-drying

The impact of single or combinations of additives on the generation of nanosuspensions of two poorly water-soluble active pharmaceutical ingredients (APIs), fenofibrate (FF) and dalcetrapib (DCP), and their isolation to the dry state via antisolvent (AS) crystallization followed by freeze-drying was explored in this work. Combinations of polymeric and surfactant additives such as poly(vinyl alcohol) or hydroxypropyl methyl cellulose and sodium docusate were required to stabilize nanoparticles (∼200−300 nm) of both APIs in suspension before isolation to dryness. For both FF and DCP, multiple additives generated the narrowest, most stable particle size distribution, with the smallest particles in suspension, compared with using a single additive. An industrially recognized freeze-drying process was used for the isolation of these nanoparticles to dryness. When processed by the liquid AS crystallization followed by freeze-drying in the presence of multiple additives, a purer monomorphic powder for FF resulted than when processed in the absence of any additive or in the presence of a single additive. It was noted that all nanoparticles freeze-dried in the presence of additives had a flat, flaky habit resulting in large surface areas. Agglomeration occurred during freeze-drying, resulting in micron-size particles. However, after freeze-drying, powders produced with single or multiple additives showed similar dissolution profiles, irrespective of aging time before drying, thus attenuating the advantage of multiple additives in terms of size observed before the freeze-drying process.</p

The biocompatibility of mesoporous silicates

Micro- and nano-mesoporous silicate particles are considered potential drug delivery systems because of their ordered pore structures, large surface areas and the ease with which they can be chemically modified. However, few cytotoxicity or biocompatibility studies have been reported, especially when silicates are administered in the quantities necessary to deliver low-potency drugs. The biocompatibility of mesoporous silicates of particle sizes similar to 150 nm, similar to 800 nm and similar to 4 mu m and pore sizes of 3 nm, 7 nm and 16 nm, respectively, is examined here. In vitro, mesoporous silicates showed a significant degree of toxicity at high concentrations with mesothelial cells. Following subcutaneous injection of silicates in rats, the amount of residual material decreased progressively over 3 months, with good biocompatibility on histology at all time points. In contrast, intra-peritoneal and intra-venous injections in mice resulted in death or euthanasia. No toxicity was seen with subcutaneous injection of the same particles in mice. Microscopic analysis of the lung tissue of the mice indicated that death may be due to thrombosis. Although local tissue reaction to mesoporous silicates was benign, they caused severe systemic toxicity. This toxicity might be mitigated by modification of the materials. (C) 2008 Elsevier Ltd. All rights reserved

Injectable in situ cross-linking hydrogels for local antifungal therapy

Invasive fungal infections can be devastating, particularly in immunocompromised patients, and difficult to treat with systemic drugs. Furthermore, systemic administration of those medications can have severe side effects. We have developed an injectable local antifungal treatment for direct administration into existing or potential sites of fungal infection. Amphotericin B (AmB), a hydrophobic, potent, and broad-spectrum antifungal agent, was rendered water-soluble by conjugation to a dextran-aldehyde polymer. The dextranaldehyde-AmB conjugate retained antifungal efficacy against Candida albicans. Mixing carboxymethylcellulose-hydrazide with dextran-aldehyde formed a gel that cross-linked in situ by formation of hydrazone bonds. The gel provided in vitro release of antifungal activity for 11 days. and contact with the gel killed Candida for three weeks. There was no apparent tissue toxicity in the murine peritoneum and the gel caused no adhesions. Gels produced by entrapment of a suspension of AmB in CMC-dextran without conjugation of drug to polymers did not release fungicidal activity, but did kill on contact Injectable systems of these types, containing soluble or insoluble drug formulations, could be useful for treatment of local antifungal infections, with or without concurrent systemic therapy. (C) 2009 Elsevier Ltd. All rights reserved

Investigation of the particle growth of fenofibrate following antisolvent precipitation and freeze-drying

Submicron to small-micron-sized particles of the hydrophobic drug, fenofibrate, were prepared by controlled crystallization in order to influence its dissolution behavior. An antisolvent precipitation process successfully generated particles (200-300 nm) which matched the size and dissolution behavior of a commercial wet-milled formulation of the drug. Although the preparation of submicron-sized particles was straightforward, retaining their size in suspension and during isolation was a challenge. Additives were employed to temporarily stabilize the suspension, and extend the time window for isolation of the submicron particles. Precipitated particles were isolated primarily by immediate freeze-drying, but drying stresses were found to destabilize the fragile submicron system. The growth pathway of particles in suspension and during oven and freeze-drying were compared. Although the growth pathways appeared considerably different from a visual morphological perspective, an investigation of the electron diffraction patterns and the inner-particle surfaces showed that the growth pathways were the same: molecular addition by Ostwald ripening. The observed differences in the time-resolved particle morphologies were found to be a result of the freeze-drying process

New solid forms of griseofulvin: a solvate and a relict polymorph related to reported solvates

 This work presents two new solid forms, a polymorph and a solvate, of the antifungal active pharmaceutical ingredient griseofulvin (GSF). The novel forms were characterized by powder X-ray diffraction, differential scanning calorimetry, and thermogravi-metric analysis, and their crystal structures were determined by single-crystal X-ray diffraction. The new polymorphic form (GSF Form VI) was obtained upon drying at room temperature the GSF-acetonitrile solvate. GSF Form VI is a relict structure related to reported solvates of GSF. Thermal stability studies show that Form VI is metastable and monotropically related to the stable GSF Form I. The new GSF-n-butyl acetate solvate was obtained by crystallization from an n-butyl acetate solution. The stoichiometry of the n-butyl acetate solvate is 1:0.5. The solvate loses the solvent from the crystal lattice at a temperature between 363.15 and 374.15 K </p

Quantitative TEM analysis of a hexagonal mesoporous silicate structure

TEM analysis of mesoporous materials is generally undertaken to give qualitative results. Accurate quantitative analysis is demonstrated in this study. A systematic image analysis of a powder form of a hexagonal mesoporous material known as KIT-6 is conducted using a transmission electron microscope (TEM). Three types of image contrast typically appear in this material (a hexagonal honeycomb structure, wide and narrow parallel lines). The honeycomb face is used to characterise this material in terms of a conventional 2-D hexagonal structure and the d-spacings for the (100) and (110) planes are experimentally measured in varying focus conditions. A tilting experiment is conducted to determine how the angle of tilt affects the line spacing and their visibility. Tilting has very little effect on the line spacing, whereas it affects the visibility of both the wide and narrow lines by limiting an angle range of visibility. The hexagonal lattice structure parameter determined by TEM method is found to be similar to 7% lower than that calculated by low-angle X-ray diffraction. Thus we conclude that TEM data can be used to determine the geometry and dimensions of hexagonal mesoporous silica materials, with a small error in the hexagonal lattice parameter

Impact of excipients and seeding on the solid-state form transformation of indomethacin during liquid antisolvent precipitation

Long-acting injectables are a unique drug formulation strategy, providing a slow and sustained release of active pharmaceutical ingredients (APIs). In this study, a novel approach that combines liquid antisolvent precipitation with seeding to obtain a stable form of the API indomethacin while achieving the desired particle size distribution is described. It was proven that when a metastable form of indomethacin was initially nucleated, the rate of its transformation to the stable form was influenced by the presence of excipients and seeds (17.10 ± 0.20 μm), decreasing from 48 to 4 h. The final particle size (D50) of the indomethacin suspension produced without seeding was 7.33 ± 0.38 μm, and with seeding, it was 5.61 ± 0.14 μm. Additionally, it was shown that the particle size distribution of the seeds and the time point of seed addition were critical to obtain the desired solid-state form and that excipients played a crucial role during nucleation and polymorphic transformation. This alternative, energy-efficient bottom-up method for the production of drug suspensions with a reduced risk of contamination from milling equipment and fewer processing steps may prove to be comparable in terms of stability and particle size distribution to current industrially accepted top-down approaches.</p

Protecting bactofencin A to enable its antimicrobial activity using mesoporous matrices

There is huge global concern surrounding the emergence of antimicrobial resistant bacteria and this is resulting in an inability to treat infectious diseases. This is due to a lack of new antimicrobials coming to the market and irresponsible use of traditional antibiotics. Bactofencin A, a novel antimicrobial peptide which shows potential as an antibiotic, is susceptible to enzyme degradation. To improve its solution stability and inherent activity, bactofencin A was loaded onto a traditional silica mesoporous matrix, SBA-15, and a periodic mesoporous organosilane, MSE. The loading of bactofencin A was considerably higher onto SBA-15 than MSE due to the hydrophilic nature of SBA-15. While there was no detectable peptide released from SBA-15 into phosphate buffered saline and only 20% of the peptide loaded onto MSE was released, the loaded matrices showed enhanced activity compared to the free peptide during in vitro antimicrobial assays. In addition, the mesoporous matrices were found to protect bactofencin A against enzymatic degradation where results showed that the SBA-15 and MSE with loaded bactofencin A exposed to trypsin inhibited the growth of S. aureus while a large decrease in activity was observed for free bactofencin upon exposure to trypsin. Thus, the activity and stability of bactofencin A can be enhanced using mesoporous matrices and these matrices may enable its potential development as a novel antibiotic. This work also shows that in silico studies looking at surface functional group and size complementarity between the peptide and the protective matrix could enable the systemic selection of a mesoporous matrix for individual bacteriocins with potential antimicrobial therapeutic properties

Impact of propeptide cleavage on the stability and activity of a streptococcal immunomodulatory C5a peptidase for biopharmaceutical development

Posttranslational modifications of proteins can impact their therapeutic efficacy, stability, and potential for pharmaceutical development. The Group AStreptococcus pyogenesC5a peptidase (ScpA) is a multi-domain protein composed of an N-terminal signal peptide, a catalytic domain (including propeptide), three fibronectin domains, and cell membrane-associated domains. It is one of several proteins produced by Group AS. pyogenesknown to cleave components of the human complement system. After signal peptide removal, ScpA undergoes autoproteolysis and cleaves its propeptide for full maturation. The exact location and mechanism of the propeptide cleavage, and the impact of this cleavage on stability and activity, are not clearly understood, and the exact primary sequence of the final enzyme is not known. A form of ScpA with no autoproteolysis fragments of propeptide present may be more desirable for pharmaceutical development from a regulatory and a biocompatibility in the body perspective. The current study describes an in-depth structural and functional characterization of propeptide truncated variants of ScpA expressed inEscherichia colicells. All three purified ScpA variants, ScpA, 79ΔPro, and 92ΔPro, starting with N32, D79, and A92 positions, respectively, showed similar activity against C5a, which suggests a propeptide-independent activity profile of ScpA. CE-SDS and MALDI top-down sequencing analyses highlight a time-dependent propeptide autoproteolysis of ScpA at 37 °C with a distinct end point at A92 and/or D93. In comparison, all three variants of ScpA exhibit similar stability, melting temperatures, and secondary structure orientation. In summary, this work not only highlights propeptide localization but also provides a strategy to recombinantly produce a final mature and active form of ScpA without any propeptide-related fragments.</p

Investigation of the solid-state polymorphic transformations of piracetam

The solid-state polymorphic transformations of 2-oxo-1-pyrrolidine acetamide (Piracetam) were investigated using a combination of off-line and on-line techniques; Differential Scanning Calorimetry (DSC), High Temperature X-ray Diffraction (HT-XRD), thermal analysis and Hot Stage Optical Microscopy. Form II and Form III were each observed to transform directly to Form I upon heating, with Form II transforming at a slightly lower temperature. The transformation of both polymorphs to Form I was observed to cause physical cracking of the crystals as well as changing the optical properties. Form I consistently transformed to Form II when cooled. The molecular rearrangements required for the transformation from Form I to Form II were found to be more energetically favourable than those required for the transformation to Form III. The transformation from the metastable Form II to the stable Form III was not observed in the solid state, while the Form II – Form III transition temperature was found to be higher than the transition temperature of both polymorphs to Form I