Optimized Milling Approaches for Scalable Production of Ritonavir Nanocrystals: from Process Design to Bioperformance Evaluation

The development of nanoformulations aims to overcome the biopharmaceutical limitations associated with conventional drug delivery. Reducing the particle size to the nanometric scale enhances drug solubility, dissolution rate, and bioavailability. In this study, the development and quality control of ritonavir nanocrystals are described by using applied experimental milling approaches. Ritonavir nanosuspensions were initially prepared at a small scale using an Ultra-Turrax Tube Drive, in which 200 and 500 μm beads were identified as the most efficient for particle size reduction. The process was then successfully scaled up by using a bead mill, achieving particle sizes of approximately 300 nm within 30 min, followed by spray-drying. Solid-state characterization by XRD, TGA, DSC, and hot-stage microscopy confirmed that the nanocrystals retained Form II ritonavir throughout processing. The resulting nanocrystals were physically stable and exhibited a marked improvement in dissolution, particularly in a discriminative dissolution medium (0.04 M POE10LE). Process optimization was achieved by balancing bead sizes and steric stabilizers, such as PVP K30 and HPMC. Biodistribution studies using [⁹⁹mTc] radiolabeling (labeling efficiency >90%) showed uptake in the liver, kidneys, and intestines, with notable differences in the cardiac distribution between nanocrystals and nanosuspensions. Pharmacokinetic analysis indicated similar overall distribution profiles, with a transient 4 h peak in blood levels for the nanocrystals. Biochemical analyses suggested formulation-dependent hepatic stress, reflected by increased GGT for the nanosuspension, and alterations in carbohydrate metabolism, including elevated glucose and amylase levels, for the nanocrystal formulation. Overall, ritonavir nanocrystals significantly improved dissolution, and the optimized milling and spray-drying approach represents a robust and scalable strategy to enhance the performance of Class II or IV drugs.