Purpose: Ice-nucleation during the freezing step in a typical lyophilization process is stochastic and associated with prolonged primary drying and high intra- and inter- batch heterogeneity. To this end, techniques that allow control of the ice-nucleation during freezing step of lyophilization has recently garnered a lot of interest. In the current research, we explored the significance of process parameters of a controlled ice-nucleation lyophilization employing the pressurization-depressurization technique on the primary drying efficiency and product quality.
Methods: A Lyostar 3™ (SP Scientific, PA, USA) freeze-dryer equipped with pressurization-depressurization ControLyo® technology was used. Mass flow profile during sublimation and desorption were obtained by an in-line tunable diode laser absorption spectrometer (TDLAS) from Physical Sciences Inc. (Andover, MA, USA). The pressure difference between the pirani pressure gauge and capacitance manometer was used to control primary drying end-point.
Two model formulations were employed in the study; an amorphous formulation with low product resistance and a crystalline formulation with relatively high product resistance. For each model formulation, a fixed primary drying and secondary drying cycle was employed. The impact of nucleation temperature on process performance and product quality were first evaluated at nucleation temperatures of -5°C, -7°C and -10°C. A conventional freezing protocol (Uncontrolled ice-nucleation) with and without an annealing step were used as controls. In addition, the impact of process parameters such as hold time prior to induction of nucleation, post nucleation cooling rate, hold time and temperature post ice nucleation were studied. The response parameters were ice crystal size, dry layer resistance during ice sublimation, duration of primary drying and ensuing product moisture content and porosity. Moreover, using the measured parameters obtained from the on-line PAT tools, critical product parameters such as the product batch average temperature, dry layer thickness, ice thickness and dry layer resistance were estimated using a steady state model. These parameters were then assessed individually for their sensitivity to variance in nucleation temperature as well as their ability to distinguish a successful and failed CIN process.
Results: As expected application of controlled ice-nucleation resulted in increase in ice crystal size and subsequently a decrease in primary drying time. An inverse correlation was observed between the degree of super-cooling and product moisture content. The average moisture content for products nucleated at -5°C, -7°C and -10°C were 3.1%, 2.0% and 1.6% respectively. Further induction of ice-nucleation by the pressurization- depressurization technique resulted in concurrent ice nucleation with less than 1°C variance in nucleation temperature. However, ice nucleation occurred over a 20-minute period with nucleation temperature variance over 7°C in the conventional uncontrolled nucleation lyophilization. Interestingly, conducting CIN at -7°C in a completely amorphous model product resulted in a higher sublimation rate and a shorter primary drying time than corresponding process at -5°C. This indicates the need for a more robust design of a CIN process rather than mere decreasing the degree of super cooling (Figure 1). This was corroborated by the lower product resistance and higher pore size observed in resulting products from CIN -7°C. Addition of extra hold time at the nucleation temperature post induction of ice-nucleation resulted in an increase in mass flux and about 15% decrease in primary drying time. However, it was found associated with a relative increase in product moisture content from 2.4±0.1% to 3.2±0.05%. Also, decreasing the post ice nucleation shelf cooling rate from 1°C to 0.1°C enhanced the ice crystal structure and resulted in about 10% decrease in primary drying time. The evaluated PAT tools could be employed in distinguishing a failed and successful CIN process. Further, for the model formulation and process employed in this study, the sensitivity of the measured and predicted process and product parameters directly correlated with the variance in the degree of supercooling.
Conclusion: The nucleation temperature is a critical parameter for a process employing controlled ice nucleation temperature. However, the associated advantage of shortening the primary drying time can only be attained through a robust design of the freezing step taking into consideration the key process parameters such as hold time (pre-and post ice nucleation), the shelf ramp rate post ice nucleation and the intended primary drying process.
Ann-Marie Ako-Adounvo– Howard University, Maryland
Leanna Hengst– Contractor, FDA/CDER/OTR/DPQR, Maryland
Yifan Wang– US Food and Drug Administration, Silver Spring
Sau Lee– US FOOD AND DRUG ADMINISTRATION
Celia N. Cruz– Director, DPQR, FDA/CDER/OTR/DPQR, Silver Spring, Maryland
Muhammad Ashraf– Supervisory Chemist, US Food and Drug Administration, Silver Spring, Maryland