Development of an Efficient Single-Step Freeze-Drying Cycle for Protein Formulations

Purpose. An efficient freeze-drying cycle for recombinant human interleukin-1 receptor antagonist (rhIL-lra) formulations, which contained glycine and sucrose as excipients, was developed. Methods. Development was based on characterizing the frozen formulations by thermal analysis and by examining the effect of various lyophilization process parameters on the sublimation rate of ice. Results. Thermal analysis showed that the metastable glass of glycine in frozen formulation could be devitrified by slowly warming the frozen product to –15°C. During drying, the sublimation rate of ice was increased as a linear function of the difference between the vapor pressure of ice at the product temperature (P _O) and the chamber pressure (P _C). Therefore, the product temperature (T_p) was maintained as high as possible at temperatures below Tg of the formulation, in order to maximize the P _O without allowing the collapse of cake. Although various combinations of shelf temperatures and chamber pressures could be used to obtain the same T_p, the combination of higher shelf temperature and lower chamber pressure was used to maximize sublimation rate. Conclusions. A single-step drying cycle was developed to take advantage of these observations. The shelf temperature was set for the secondary drying and the product temperature during primary drying was maintained below T_g by adjusting the chamber pressure. As the sublimation completed, the product temperature increased naturally to the shelf temperature for the secondary drying. This process resulted in successful drying of 1 ml of rhIL-lra formulation to 0.4% moisture content within 6 hours.     

Using dextran of different molecular weights to achieve faster freeze-drying and improved storage stability of lactate dehydrogenase

Freeze-drying of protein formulations is frequently used to maintain protein activity during storage. The freeze-drying process usually requires long primary drying times because the highest acceptable drying temperature to obtain acceptable products is dependent on the glass transition temperature of the maximally freeze-concentrated solution (T_g′). On the other hand, retaining protein activity during storage is related to the glass transition temperature (T_g) of the final freeze-dried product. In this study, dextrans with different molecular weight (1 and 40?kDa) and mixtures thereof at the ratio 3:1, 1:1, and 1:3 (w/w) were used as cryo-/lyoprotectant and their impact on the stability of the model protein lactate dehydrogenase (LDH) was investigated at elevated temperatures (40?°C and 60?°C). The dextran formulations were then compared to formulations containing sucrose as cryo-/lyoprotectant. Because of the higher T_g′ values of the dextrans, the primary drying times could be reduced compared to freeze-drying with sucrose. Similarly, the higher T_g and T_g′ of dextrans relative to sucrose led to benefits during storage which was shown through improved protection of LDH activity.     

Protein Quantity on the Air–Solid Interface Determines Degradation Rates of Human Growth Hormone in Lyophilized Samples

Recombinant human growth hormone (rhGH) was lyophilized with various glass-forming stabilizers, employing cycles that incorporated various freezing and annealing procedures to manipulate glass formation kinetics, associated relaxation processes, and glass-specific surface areas (SSAs). The secondary structure in the cake was monitored by infrared and in reconstituted samples by circular dichroism. The rhGH concentrations on the surface of lyophilized powders were determined from electron spectroscopy for chemical analysis. Glass transition temperature (T_g), SSAs, and water contents were determined immediately after lyophilization. Lyophilized samples were incubated at 323 K for 16 weeks, and the resulting extents of rhGH aggregation, oxidation, and deamidation were determined after rehydration. Water contents and T_g were independent of lyophilization process parameters. Compared with samples lyophilized after rapid freezing, rhGH in samples that had been annealed in frozen solids prior to drying, or annealed in glassy solids after secondary drying retained more native-like protein secondary structure, had a smaller fraction of the protein on the surface of the cake, and exhibited lower levels of degradation during incubation. A simple kinetic model suggested that the differences in the extent of rhGH degradation during storage in the dried state between different formulations and processing methods could largely be ascribed to the associated levels of rhGH at the solid–air interface after lyophilization. © 2014 Wiley Periodicals, Inc. and the American Pharmacists Association.     

Accurate Prediction of Collapse Temperature using Optical Coherence Tomography-Based Freeze-Drying Microscopy

The objective of this study was to assess the feasibility of developing and applying a laboratory tool that can provide three-dimensional product structural information during freeze-drying and which can accurately characterize the collapse temperature (T_c) of pharmaceutical formulations designed for freeze-drying. A single-vial freeze dryer coupled with optical coherence tomography freeze-drying microscopy (OCT–FDM) was developed to investigate the structure and T_c of formulations in pharmaceutically relevant products containers (i.e., freeze-drying in vials). OCT–FDM was used to measure the T_c and eutectic melt of three formulations in freeze-drying vials. The T_c as measured by OCT–FDM was found to be predictive of freeze-drying with a batch of vials in a conventional laboratory freeze dryer. The freeze-drying cycles developed using OCT–FDM data, as compared with traditional light transmission freeze-drying microscopy (LT-FDM), resulted in a significant reduction in primary drying time, which could result in a substantial reduction of manufacturing costs while maintaining product quality. OCT–FDM provides quantitative data to justify freeze-drying at temperatures higher than the T_c measured by LT-FDM and provides a reliable upper limit to setting a product temperature in primary drying.     

Effect of Moisture on the Stability of a Lyophilized Humanized Monoclonal Antibody Formulation

Purpose. To determine the effect of moisture and the role of the glass transition temperature (T_g) on the stability of a high concentration, lyophilized, monoclonal antibody. Methods. A humanized monoclonal antibody was lyophilized in a sucrose/histidine/polysorbate 20 formulation. Residual moistures were from 1 to 8%. T_g values were measured by modulated DSC. Vials were stored at temperatures from 5 to 50°C for 6 or 12 months. Aggregation was monitored by size exclusion chromatography and Asp isomerization by hydrophobic interaction chromatography. Changes in secondary structure were monitored by Fourier transform infrared (FTIR). Results. T_g values varied from 80°C at 1% moisture to 25°C at 8% moisture. There was no cake collapse and were no differences in the secondary structure by FTIR. All formulations were stable at 5°C. High moisture cakes had higher aggregation rates than drier samples if stored above their T_g values. Intermediate moisture vials were more stable to aggregation than dry vials. High moisture samples had increased rates of Asp isomerization at elevated temperatures both above and below their T_g values. Chemical and physical degradation pathways followed Arrhenius kinetics during storage in the glassy state. Only Asp isomerization followed the Arrhenius model above the T_g value. Both chemical and physical stability at T T_g were fitted to Williams-Landel-Ferry (WLF) kinetics. The WLF constants were dependent on the nature of the degradation system and were not characteristic of the solid system. Conclusion. High moisture levels decreased chemical stability of the formulation regardless of whether the protein was in a glassy or rubbery state. In contrast, physical stability was not compromised, and may even be enhanced, by increasing residual moisture if storage is below the T_g value.     

Effect of Product Temperature During Primary Drying on the Long-Term Stability of Lyophilized Proteins

Our objective was to investigate the effect of performing primary drying at product temperatures below and above Tg′ (glass transition temperature of the freeze-concentrated phase) on the long-term stability of lyophilized proteins. Two protective media differing in the nature of the bulking agent used (amorphous or crystalline) were selected. Several lyophilization cycles were performed by using various combinations of shelf temperature and chamber pressure to obtain different values of product temperature during primary drying. The antigenic activity of the proteins was measured after lyophilization and after 6 months of storage at 4°C and 25°C. After 6 months of storage and regardless of the protective medium, the losses of antigenic activity of both toxins increased from 0% when primary drying was performed at a product temperature lower than Tg′ and to 25% when the product temperature was higher than Tg′. The use of partially crystalline systems makes it possible to withstand high primary drying temperatures (above Tg′). However, the shelf life of lyophilized proteins may be decreased when the amorphous phase including the protein and the stabilizing molecule changes to the viscous state.     

Temperature Measurement by Sublimation Rate as a Process Analytical Technology Tool in Lyophilization

Product temperature (T_b) and drying time constitute critical material attributes and process parameters in the lyophilization process and especially during the primary drying stage. In the study, we performed a temperature measurement by the sublimation rate (TMbySR) to monitor the T_b value and determine the end point of primary drying. First, the water vapor transfer resistance coefficient through the main pipe from the chamber to the condenser (C_r) was estimated via the water sublimation test. The use of C_r value made it possible to obtain the time course of T_b from the measurement of pressure at the drying chamber and at the condenser. Second, a Flomoxef sodium bulk solution was lyophilized by using the TMbySR system. The outcome was satisfactory when compared with that obtained via conventional sensors. The same was applicable for the determination of the end point of primary drying. A laboratory-scale application of the TMbySR system was evidenced via the experiment using 220-, 440-, and 660-vial scales of lyophilization. The outcome was not dependent on the loading amount. Thus, the results confirmed that the TMbySR system is a promising tool in laboratory scale.     

Stabilizing Effect of Four Types of Disaccharide on the Enzymatic Activity of Freeze-dried Lactate Dehydrogenase: Step by Step Evaluation from Freezing to Storage

Purpose In order to understand the stabilizing effects of disaccharides on freeze-dried proteins, the enzymatic activity of lactate dehydrogenase (LDH) formulations containing four types of disaccharide (trehalose, sucrose, maltose, and lactose) at two relative humidity (RH) levels (about 0 and 32.8%) was investigated after three processes: freeze-thawing, freeze-drying, and storage at three temperatures (20, 40, and 60°C) above and/or below the glass transition temperature (T _g). Materials and Methods The enzymatic activity was determined from the absorbance at 340 nm, and T _g of the samples was investigated by differential scanning calorimetry. Results At each RH condition, T _g values of sucrose formulations were lower than those of other formulations. Although effects of the disaccharides on the process stability of LDH were comparable, storage stability was dependent on the type of disaccharide. All the formulations were destabilized significantly during storage at temperature above T _g. During storage at temperature below T _g, the LDH activity decreased with increases in the storage temperature and moisture. Maltose and lactose formulations showed significant destabilization with the change of color to browning. Conclusions Taking the storage stability of freeze-dried proteins under the various conditions (temperature and RH) into consideration, trehalose is better suited as the stabilizer than other disaccharides.     

Crystallization of Cyclophosphamide Monohydrate During Lyophilization

The purpose of this study was to investigate the phase behavior of cyclophosphamide (CPA) during various stages of lyophilization, with special emphasis on obtaining crystalline CPA monohydrate (CPA-MH) in the lyophilized product. Subambient differential scanning calorimetry and low-temperature X-ray diffractometry (LTXRD) were used to study the phase behavior of CPA solution (3.7% w/v). In situ lyophilization in LTXRD chamber was used to monitor the phase transitions occurring during the drying stages. Finally, the implications of these findings were confirmed by freeze-drying the aqueous solution in a laboratory-scale freeze-dryer. The results suggested that CPA remains amorphous during freeze concentration, with a T_g' of ?50°C. However, its crystallization as CPA-MH can be induced by annealing the frozen solution between ?5°C and ?10°C. In situ lyophilization in LTXRD showed that the CPA-MH crystallized during annealing, rapidly dehydrated during primary drying, thereby causing structural collapse. The dehydration of CPA-MH can be prevented by lowering the escaping tendency of water molecules from the crystal lattice of CPA-MH by maintaining the chamber pressure to 300, 400, or 500 mTorr. This study highlights the relationship of process parameters used during lyophilization with the solid form of lyophilized CPA.     

Effect of Glass Transition Temperature on the Stability of Lyophilized Formulations Containing a Chimeric Therapeutic Monoclonal Antibody

Purpose. The purpose of this study is to highlight the importance of knowing the glass transition temperature, T_g, of a lyophilized amorphous solid composed primarily of a sugar and a protein in the interpretation of accelerated stability data. Methods. Glass transition temperatures were measured using DSC and dielectric relaxation spectroscopy. Aggregation of protein in the solid state was monitored using size-exclusion chromatography. Results. Sucrose formulation (T_g ~ 59°C) when stored at 60°C was found to undergo significant aggregation, while the trehalose formulation (T_g ~ 80°C) was stable at 60°C. The instability observed with sucrose formulation at 60°C can be attributed to its T_g (~59°C) being close to the testing temperature. Increase in the protein/sugar ratio was found to increase the T_gs of the formulations containing sucrose or trehalose, but to different degrees. Conclusions. Since the formulations exist in glassy state during their shelf-life, accelerated stability data generated in the glassy state (40°C) is perhaps a better predictor of the relative stability of formulations than the data generated at a higher temperature (60°C) where one formulation is in the glassy state while the other is near or above its T_g.     

The freezing step in lyophilization: Physico-chemical fundamentals, freezing methods and consequences on process performance and quality attributes of biopharmaceuticals

Lyophilization is a common, but cost-intensive, drying process to achieve protein formulations with long-term stability. In the past, typical process optimization has focused on the drying steps and the freezing step was rather ignored. However, the freezing step is an equally important step in lyophilization, as it impacts both process performance and product quality.While simple in concept, the freezing step is presumably the most complex step in lyophilization. Therefore, in order to get a more comprehensive understanding of the processes that occur during freezing, the physico-chemical fundamentals of freezing are first summarized. The available techniques that can be used to manipulate or directly control the freezing process in lyophilization are also reviewed. In addition, the consequences of the freezing step on quality attributes, such as sample morphology, physical state of the product, residual moisture content, reconstitution time, and performance of the primary and secondary drying phase, are discussed. A special focus is given to the impact of the freezing process on protein stability.This review aims to provide the reader with an awareness of not only the importance but also the complexity of the freezing step in lyophilization and its impact on quality attributes of biopharmaceuticals and process performance. With a deeper understanding of freezing and the possibility to directly control or at least manipulate the freezing behavior, more efficient lyophilization cycles can be developed, and the quality and stability of lyophilized biopharmaceuticals can be improved.     

Optimization of secondary drying condition for desired residual water content in a lyophilized product using a novel simulation program for pharmaceutical lyophilization

The aim of this study was to optimize the shelf temperature and the drying time, mainly dependent on the residual water content of a lyophilized product using a novel simulation program for the secondary drying of lyophilization. The simulation program was developed based upon heat transfer formulas, two empirical formulas, and a modified Fick’s second law. When a preliminary lyophilization run of secondary drying was carried out, the equilibrium product temperature at the end of secondary drying under various shelf temperatures was accurately predicted by the heat transfer formulas. The apparent diffusion coefficient of water, D_eff, and the apparent equilibrium residual water content, W_e, under the predicted equilibrium product temperature were estimated by two empirical formulas. These estimated D_eff and W_e allow the modified Fick’s second law to predict the residual water content in the lyophilized product. Using the developed simulation program, it was verified that the secondary drying condition to achieve the desired residual water content in the lyophilized product was successfully predicted. Therefore, the simulation program can be used to effectively design the secondary drying condition of lyophilization cycles without a trial and error approach.     

Physical characterisation of formulations for the development of two stable freeze-dried proteins during both dried and liquid storage

The development of stable freeze-dried proteins requires maintaining the physical and biological integrity of the protein as well as increasing the efficiency of the manufacturing process. Our objective was to study the effects of various excipients on both the physical characterisation and the dried and liquid stability of two proteins. Thermo-physical properties of 13 formulations were determined using both differential scanning calorimetry and freeze–drying microscopy. The antigenic activity was evaluated immediately after freeze–drying and after subsequent storage in both dried and liquid state. From the comparison between glass transition (T′_g) and collapse (T_coll) temperatures, we concluded that the collapse temperature was a more relevant parameter than T′_g for freeze–drying cycle development and optimisation. One crystalline formulation composed of 4% mannitol and 1% of sucrose protected efficiently both proteins during subsequent storage in dried state (6 months at 25 °C) and in liquid state (3 months at 4 °C after rehydration). However, the freeze–drying behaviour of this crystalline formulation remained difficult to predict and control. On the other hand, two amorphous formulations composed of 4% of maltodextrin and 0.02% of Tween 80, or 5% of BSA preserved antigenic activity during storage in dried state. The glassy character of these formulations as well as their high collapse temperature values (−9 and −12 °C, respectively) should allow simplification and shortening of freeze–drying process.     

Evaluation of the Physical Stability of Freeze-Dried Sucrose-Containing Formulations by Differential Scanning Calorimetry

Freeze-dried samples of sucrose with buffer salts, amino acids, or dextran have been analyzed with differential scanning calorimetry (DSC) to evaluate the use of DSC thermograms in predicting the physical storage stability. The glass transition temperature, T _g, of the amorphous cake, crystallization, and melting of sucrose are observed with DSC. T _g appeared to be an important characteristic of the physical stability of the amorphous freeze-dried cake. A storage temperature above T _g results in collapse or shrinkage of the cake, which for a sucrose-based formulation, may be accompanied by crystallization of the sucrose. The T _g of the amorphous sucrose is influenced by other components present in the cake. Dextran-40 raised T _g, while the addition of glycine to the formulation lowered T _g. The residual moisture content strongly influences T _g, since water acts as a plasticizer of the system; the higher the moisture content, the lower the T _g and the less physically stable the freeze-dried cake. Crystallization of amorphous sucrose is shown to be inhibited by high molecular weight components or ionic compounds. DSC analysis of freeze-dried cakes proved to be a powerful tool in formulation studies.     

Effects of Solute Miscibility on the Micro- and Macroscopic Structural Integrity of Freeze-Dried Solids

The purpose of this study was to elucidate the effect of solute miscibility in frozen solutions on their micro- and macroscopic structural integrity during freeze-drying. Thermal analysis of frozen solutions containing poly(vinylpyrrolidone) (PVP) and dextran showed single or multiple thermal transitions (T′_g: glass transition temperature of maximally freeze-concentrated solutes) depending on their composition, which indicated varied miscibility of the concentrated noncrystalline polymers. Freeze-drying of the miscible solute systems (e.g., PVP 10,000 and dextran 1060, single T′_g) induced physical collapse during primary drying above the transition temperatures (> T′_g). Phase-separating PVP 29,000 and dextran 35,000 mixtures (two T′_gs) maintained their cylindrical structure following freeze-drying below both of the T′_gs (<?24°C). Primary drying of the dextran-rich systems at temperatures between the two T′_gs (? 20 to ? 14°C) resulted in microscopically disordered “microcollapsed” cake-structure solids. Freeze-drying microscopy (FDM) analysis of the microcollapsing polymer system showed locally disordered solid region at temperatures between the collapse onset (T_c1) and severe structural change (T_c2). The rigid dextran-rich matrix phase should allow microscopic structural change of the higher fluidity PVP-rich phase without loss of the macroscopic cake structure at the temperature range. The results indicated the relevance of physical characterization and process control for appropriate freeze-drying of multicomponent formulations.     

Physical Stability of an Amorphous Sugar Matrix Dried From Methanol as an Amorphous Solid Dispersion Carrier and the Influence of Heat Treatment

An amorphous sugar matrix, after drying from an organic solvent, was investigated for use as a method for dispersing hydrophobic drugs (solid dispersion). However, the amorphous sugar, originally contained in the organic solvent, had a significantly low glass transition temperature (T_g), thus rendering it physically unstable. In this study, we examined the physicochemical properties of a sugar in a dried matrix and in an organic solvent, using α-maltose and methanol as a representative sugar and organic solvent. The apparent molar volume of α-maltose was ∼30% smaller in methanol than in water. The methanol-originated amorphous α-maltose exhibited a much greater degree of hydrogen bonding than the water-originated one. Considering these findings, we conclude that the α-maltose maintained its compact conformation in the dried state and consequently caused the markedly low T_g. Second, it was found that heating under appropriate conditions resulted in an increase in the T_g of the methanol-originated amorphous α-maltose as well as a decrease in the level of hydrogen bonding. The aqueous dissolution of 2 model hydrophobic drugs (indomethacin and ibuprofen) from the solid dispersion was also improved as the result of the heat treatment, whereas, to the contrary, the dissolution of another model drug (curcumin) was lowered.     

A Quantitative Assessment of the Significance of Molecular Mobility as a Determinant for the Stability of Lyophilized Insulin Formulations

Purpose The purpose was to explore a method for quantitatively assessing the contribution of molecular mobility to the chemical reactivity of amorphous solids. Degradation of insulin in lyophilized formulations containing trehalose and poly(vinylpyrrolidone)(PVP) was chosen as a model system, and the temperature- and glass transition temperature (T_g)-dependence of the degradation rate was analyzed to obtain the relative contributions of molecular mobility and that of the chemical activational barrier reflected in the energy of activation.Methods Insulin degradation and dimerization in lyophilized trehalose and PVP formulations were monitored at various relative humidities (6–60% RH) and temperatures (10–60°C) by reverse-phase high-performance liquid chromatography (HPLC) and high-performance size-exclusion chromatography (HP-SEC), respectively. The T_g and fragility parameter of the lyophilized insulin formulations were determined by differential scanning calorimetry (DSC).Results Insulin degradation in the initial stage was describable with first-order kinetics for both of the trehalose and PVP formulations. The temperature- and T_g-dependence of the degradation rate indicated that the reactivity of insulin in the trehalose formulation is affected by molecular mobility at low humidity (12% RH), such that the ratio of the observed rate constant (k′) to the rate constant governed only by the activational barrier (k) was 0.051 at the T_g. At higher humidities, in contrast, the value of k′/k was much higher (0.914, 0.978, and 0.994 for 23% RH, 33% RH, and 43% RH, respectively), indicating that insulin degradation rate is determined predominantly by the activational barrier. For insulin degradation in the PVP formulation at temperatures below T_g, the contribution of molecular mobility to the degradation rate appeared to be negligible, as the extrapolated value of t₉₀ at the T_g exhibited a large difference between the formulations with differing T_g values (because of differing water contents).Conclusions The reactivity of insulin in the trehalose and PVP formulations can be described by an equation including factors reflecting the activational barrier (activation energy and frequency coefficient) and factors reflecting the molecular mobility (T_g, fragility parameter and a constant representing the relationship between the molecular mobility and the reaction rate). Thus, analysis of temperature dependence based on the proposed equation allows quantitative assessment of the significance of molecular mobility as a factor affecting chemical reactivity.     

Stability and surface activity of lactate dehydrogenase in spray‐dried trehalose

The stability of the model protein lactate dehydrogenase (LDH) during spray‐drying and also on subsequent dry storage was examined. Trehalose was used as a carrier. The spray‐drying temperatures T_inlet and T_outlet have a measurable effect on LDH inactivation. Low T_inlet produced the least process inactivation, but gave a high residual moisture content making the protein's storage stability poor. High T_inlet reduced residual moisture and improved storage stability, but at the cost of high process inactivation. As already found for other systems, addition of a surfactant (in this case polysorbate 80) could ameliorate process inactivation of LDH at T_inlet = 150 °C. Surfactant had, however, a deleterious effect on storage stability of LDH, the vital factor being the molar ratio of surfactant/protein in the dried product. By using electron spectroscopy it was shown that LDH has a 10 times higher surface concentration in the dried trehalose particles than expected for a homogeneous distribution. Surface tension measurements at the water/air interface proved that LDH is surface active, although the Gibbs equation appeared to be inapplicable. Calculations of spray‐droplet formation time and drying time indicate than the extent of diffusion‐driven LDH adsorption to the liquid/air interface is sufficient to account for the measured amount of LDH inactivation during spray‐drying. The presence of 0.1% polysorbate 80 to the spray solution prevents LDH from appearing at the surface of the dried particles. As a negative control, the phosphatide Lipoid E 80 does not prevent the appearance of LDH in the surface according to electron spectroscopy and does not therefore prevent LDH inactivation during spray‐drying at T_inlet = 150 °C.     

Significant Drying Time Reduction Using Microwave-Assisted Freeze-Drying for a Monoclonal Antibody

Microwave-assisted freeze-drying (MFD) is a rapid drying process well known in food technology. However, little is known about its application to biologicals. In this study, we investigated the applicability and feasibility of this technology to different monoclonal antibody formulations and the influence on the resulting product properties. Moreover, one of our main objectives was to study if significant reductions in drying times could be achieved. In addition, the effect of the drying process on the accelerated stability of a sucrose-based antibody formulation at 40°C and 25°C over 12 weeks was investigated. MFD resulted in drying time reduction >75%. For all model formulations, cake appearance and solid state properties were found to be comparable to standard lyophilized products. These formulations covered a wider range of lyophilization excipients comprising sucrose and trehalose, semi-crystalline forming solids like mannitol:sucrose mixtures and others like arginine phosphate and a mixture of 2-hydroxypropyl-β-cyclodextrin with sucrose. Moreover, comparable low changes in relative monomer content, the relative amount of soluble aggregates and cumulative particles ≥1 μm per mL were observed over 12 weeks of storage, regardless of the drying technology. This makes MFD a promising innovative alternative for the rapid production of freeze-dried biologicals while maintaining product quality.     

In-Situ Molecular Vapor Composition Measurements During Lyophilization

Purpose

Monitoring process conditions during lyophilization is essential to ensuring product quality for lyophilized pharmaceutical products. Residual gas analysis has been applied previously in lyophilization applications for leak detection, determination of endpoint in primary and secondary drying, monitoring sterilization processes, and measuring complex solvents. The purpose of this study is to investigate the temporal evolution of the process gas for various formulations during lyophilization to better understand the relative extraction rates of various molecular compounds over the course of primary drying.

Methods

In this study, residual gas analysis is used to monitor molecular composition of gases in the product chamber during lyophilization of aqueous formulations typical for pharmaceuticals. Residual gas analysis is also used in the determination of the primary drying endpoint and compared to the results obtained using the comparative pressure measurement technique.

Results

The dynamics of solvent vapors, those species dissolved therein, and the ballast gas (the gas supplied to maintain a set-point pressure in the product chamber) are observed throughout the course of lyophilization. In addition to water vapor and nitrogen, the two most abundant gases for all considered aqueous formulations are oxygen and carbon dioxide. In particular, it is observed that the relative concentrations of carbon dioxide and oxygen vary depending on the formulation, an observation which stems from the varying solubility of these species. This result has implications on product shelf life and stability during the lyophilization process.

Conclusions

Chamber process gas composition during lyophilization is quantified for several representative formulations using residual gas analysis. The advantages of the technique lie in its ability to measure the relative concentration of various species during the lyophilization process. This feature gives residual gas analysis utility in a host of applications from endpoint determination to quality assurance. In contrast to other methods, residual gas analysis is able to determine oxygen and water vapor content in the process gas. These compounds have been shown to directly influence product shelf life. With these results, residual gas analysis technique presents a potential new method for real-time lyophilization process control and improved understanding of formulation and processing effects for lyophilized pharmaceutical products.