Per the specifications outlined in ICH Q6B, we offer analytical services in support of protein drug development to determine physical and chemical instabilities at different stages. The molecular structure of protein is delicate and highly sensitive to environmental changes and stresses. Ensuring native-like higher order structure in a biologic is essential because the overall conformation defines the drug’s stability, biological activity, efficacy and safety.
We apply numerous techniques (e.g., freeze-drying and spray drying) for solid state protein formulation development and to tailor test to your specific needs.
We evaluate the physical stability of protein in various formulation excipients such as ionic strength, buffers, pH, surfactants, sugars, salts, antioxidants and amino acids on the physicochemical properties of the protein.
- Gel permeation chromatography and size exclusion chromatography, coupled with a static-light-scattering detector, to detect and quantify protein aggregation
- Nano-DSC to monitor conformational stability of biologics in their formulations; detecting structural alterations (unfolding/aggregation) in a biologic in the form of a Tm (denaturation temperature) shift or a change in the shape of the endothermic peak (∆H and ∆Hv for domain and subunit organization)
- Dynamic light scattering (DLS) or quasi-elastic light scattering to evaluate the aggregation state; nondestructive DLS determines size distribution of particles in the diameter range of 1 nm to 2 µm
Lyophilization Process Development and Optimization
We offer small-scale lyophilization services to screen common excipients for protein stability.
- Protein formulation in compatible excipients based on the physicochemical properties of the protein
- Temperature-modulated differential scanning calorimetry to determine the glass transition temperature of the formulation in the frozen state (Tg’) and freeze-drying microscopy to determine the collapse temperature
- Optimal shelf temperatures and chamber pressures for primary and secondary drying
- Characterization of the finished lyophilized product using X-ray powder diffraction and polarized light microscopy to assess crystallinity; thermogravimetry and DSC for thermal properties; scanning electron microscopy for particle morphology; Karl Fischer titration for water content; dynamic vapor sorption/desorption for hygroscopicity testing and solid state NMR; FTIR and Raman spectroscopy for chemical and physical fingerprinting
- Evaluation of physical stability (unfolding/aggregation) of protein upon lyophilization by SEC, nano-DSC, DLS, CD and HPLC
Real-Time and Accelerated Stability Studies
Storage stability of proteins correlates with the degree of retention of native structure of proteins and the level of hydration during drying. Environmental conditions such as temperature, pH, ionic strength, oxygen and protease content can cause denaturation, aggregation, degradation and chemical modification (oxidation and deamidation). We offer advanced analytical methodologies for stability evaluation that ensure structural integrity and physicochemical identity.
- Physical stability assessment: Aggregation state evaluation by SEC and DLS, conformational change by nano-DSC and solid state characterization by various analytical techniques including XRPD, TG, DSC, SEM, SSNMR, FTIR and Raman
- Chemical stability assessment: Ultra-high resolution Q-TOF mass spectrometer and solids/liquids NMR spectrometer in conjunction with techniques such as peptide mapping, MALDI-TOF MS, amino acid analysis and Edman N-terminal sequencing for accurate mass and primary sequence of proteins and peptides