Dynamical Transition in Dehydrated Proteins.

Terahertz time-domain spectroscopy and differential scanning calorimetry were used to study the role of the dynamics of biomolecules decoupled from solvent effects. Lyophilized sucrose exhibited steadily increasing absorption with temperature as anharmonic excitations commenced as the system emerged from a deep minimum of the potential energy landscape where harmonic vibrations dominate. The polypeptide bacitracin and two globular proteins, lysozyme and human serum albumin, showed a more complex temperature dependence. Further analysis focused on the spectral signature below and above the boson peak. We found evidence of the onset of anharmonic motions that are characteristic for partial unfolding and molecular jamming in the dry biomolecules. The activation of modes of the protein molecules at temperatures comparable to the protein dynamical transition temperature was observed in the absence of hydration. No evidence of Fröhlich coherence, postulated to facilitate biological function, was found in our experiments.

Towards simultaneous determination of tablet porosity and height by terahertz time-domain reflection spectroscopy.

The quality control of pharmaceutical tablets is still based on testing small sample numbers using at- and off-line testing methods. Traditional in-process controls, such as tablet mass, height, mechanical strength, and disintegration time are time- and resource-consuming and poorly suited to support an effective transition towards continuous manufacturing. Another suitable parameter to monitor during production would be tablet porosity. Porosity can be linked to mechanical strength and disintegration but typically requires knowledge of tablet dimensions and mass. Tablet porosity measurements based on terahertz time-domain spectroscopy (THz-TDS) offer a fast and non-destructive approach to in-process control testing for physical tablet properties. This study presents THz-TDS reflection measurements as an alternative to the previously reported transmission setup. It is shown that the proposed method can determine porosity based on the reflected amplitude from the tablet surface, but also allows for precise determination of tablet height in the same measurement. The tablet mass can be estimated by combining the height and porosity measurements. This opens up for the opportunity to determine the tablet's mechanical strength by using the possible correlation to the determined porosity.

Terahertz time-domain spectroscopy for the investigation of tablets prepared from roller compacted granules.

Roller compaction before tableting is a common unit operation to increase the processability of powders. Terahertz time-domain spectroscopy (THz-TDS) has recently been introduced as a potential process analytical technology (PAT) for measuring tablet porosity based on the refractive index of the tablet. Tablet porosity is a governing parameter for tablet disintegration and dissolution. The first aim of this study was to investigate tablets prepared from roller-compacted materials with THz-TDS to explore its usefulness for particle size evaluation of granules in tablets. Secondly, the impact of roller compaction and granule size before tablet compression on the established THz-TDS based measurement of tablet porosity was investigated. Microcrystalline cellulose and α-lactose monohydrate were roller compacted separately at five specific compaction forces (2, 4, 8, 12, and 16 kN cm-1) and fractionated into three size fractions. Tablets were prepared from the fractionated and unfractionated granules at twelve tableting pressures and subjected to THz-TDS transmission measurements. It was possible to use the scattering behaviour of the tablets at terahertz frequencies to describe the granulated materials' particle size changes during tableting. At the same time, prediction of porosity was impaired due to the deviation of the refractive index in strongly scattering samples. A correction method was introduced in which the porosity error was corrected based on the tablet's scattering behaviour, resulting in an improved prediction of tablet porosity. In conclusion, THz-TDS is considered a promising technique for the process monitoring of tableting based on its sensitivity to porosity and particle size changes within the tablet non-destructively, with a possible application as part of an in-process control strategy of the tableting of granulated or non-granulated materials.

The effect of mAb and excipient cryoconcentration on long-term frozen storage stability - Part 1: Higher molecular weight species and subvisible particle formation.

Cryoconcentration upon large-scale freezing of monoclonal antibody (mAb) solutions leads to regions of different ratios of low molecular weight excipients, like buffer species or sugars, to protein. This study focused on the impact of the buffer species to mAb ratio on aggregate formation after frozen storage at -80 °C, -20 °C, and - 10 °C after 6 weeks, 6 months, and 12 months. An optimised sample preparation was established to measure Tg' of samples with different mAb to histidine ratios via differential scanning calorimetry (DSC). After storage higher molecular weight species (HMWS) and subvisible particles (SVPs) were detected using size-exclusion chromatography (SEC) and FlowCam, respectively. For all samples, sigmoidal curves in DSC thermograms allowed to precisely determine Tg' in formulations without glass forming sugars. Storage below Tg' did not lead to mAb aggregation. Above Tg', at -20 °C and - 10 °C, small changes in mAb and buffer concentration markedly impacted stability. Samples with lower mAb concentration showed increased formation of HMWS. In contrast, higher concentrated samples led to more SVPs. A shift in the mAb to histidine ratio towards mAb significantly increased overall stability. Cryoconcentration upon large-scale freezing affects mAb stability, although relative changes compared to the initial concentration are small. Storage below Tg' completely prevents mAb aggregation and particle formation.

Terahertz time-domain spectroscopy for powder compact porosity and pore shape measurements: An error analysis of the anisotropic bruggeman model.

Terahertz time-domain spectroscopy (THz-TDS) is a novel technique which has been applied for pore structure analysis and porosity measurements. For this, mainly the anisotropic Bruggeman (AB-EMA) model is applied to correlate the effective refractive index (n eff) of a tablet and the porosity as well as to evaluate the pore shape based on the depolarisation factor L. This paper investigates possible error sources of the AB-EMA for THz-TDS based tablet analysis. The effect of absorption and tablet anisotropy - changes of pore shape with porosity and density distribution - have been investigated. The results suggest that high tablet absorption has a negligible effect on the accuracy of the AB-EMA. In regards of tablet anisotropy the accuracy of the porosity determination is not impaired significantly. However, density distribution and variations in the pore shape with porosity resulted in an unreliable extraction of the tablet pore shape. As an extension of the AB-EMA a new concept was introduced to convert the model into bounds for L. This new approach was found useful to investigate tablet pore shape but also the applicability of the AB-EMA for an unknown set of data.

Non-destructive quantification of fragmentation within tablets after compression from scattering analysis of terahertz transmission measurements.

Material deformation behaviour has a critical impact on tablet formation. Fragmentation is one of the key mechanisms affecting the strength of a final compact, however, quantitative methods for estimating fragmentation are often complex, destructive and time-consuming. The purpose of this study was to investigate the applicability of terahertz time-domain spectroscopy (THz-TDS) to quantify fragmentation upon tableting. Up to five size fractions of microcrystalline cellulose (MCC), dibasic calcium phosphate (DCP), and lactose monohydrate (lactose) in the range of <125 µm up to the range of 355-500 µm were compressed into tablets and analysed with THz-TDS. The effective refractive index and absorbance spectra of whole tablets were measured in transmission, and the optical properties were clearly affected by fragmentation upon compression. The scattering observed from the absorbance spectra was fitted into a power law equation (y = AνB). It was observed that up to pressures of 50 MPa the values of parameter A that were extracted from the power law fit decreased exponentially with increasing compression pressure. For higher compression pressures the value of A remained constant. This observation was more pronounced for DCP, followed by lactose and then MCC and the effect was more pronounced for larger compared to smaller initial particles. The non-destructive measurements correlated with previously obtained results based on particle size distribution measurements of the particles before compression and those obtained from destructive analysis of tablets. The terahertz method can resolve similar differences in fragmentation behaviour upon compression compared to the particle size analysis but requires no sample preparation.