This study aims to examine the throughput of nanoparticles through a Cladophora cellulose based virus removal filter. The effect of Brownian motion and flow velocity on the retention of 5 nm gold nanoparticles, 12.8 nm dextran nanoparticles and 28 nm ΦX174 bacteriophages was examined through MATLAB simulations and filtration experiments. Modeling of Brownian motion at different flow velocities was performed in MATLAB by solving the Langevin equation for particle position and velocity for all three types of particles. The motion of all three particle types was shown to be constrained at local flow velocities of 1∙10-2 m/s or greater. The constraint was greatest for ΦX174 bacteriophages, followed by dextran particles and then gold particles as a result of particle diameter. To verify the effect experimentally, virus removal filters were prepared with a peak pore width of 23 nm. Filtration experiments were performed at different flux values where gold and dextran particles did not exhibit any difference in retention between fluxes. However, a significant amount of gold and dextran particles were removed by the filter despite being smaller than the measured pore size. A decrease in retention with filtrated volume was observed for both particle types. Filtration of ΦX174 bacteriophages exhibited a difference in retention at different fluxes, where all bacteriophages where removed at a higher flux. The results from both simulations and experiments suggest that the retentive mechanism in filtering is more complex than what can be described only by size exclusion sieving, Brownian diffusion and hydrodynamic constraint of particles.

The studies in the present thesis explored the feasibility of using nanocellulose-based filters in virus removal filtration of plasma-derived proteins.   In Paper I, two-step nanofiltration of commercially available human serum albumin (HSA) product, which was diluted to 10 g L-1 by phosphate buffer saline (PBS) and adjusted pH to 7.4, was performed to remove soluble protein aggregates and reduce filter fouling. The two-step filtration of HSA employed nanocellulose-based filters of varying thickness, i.e. 11 μm and 22 μm filters.  The removal of HSA aggregates during filtration through 11 μm pre-filters dramatically improves the flow properties of the 22 μm filter, enabling high protein throughput and high virus clearance. A distribution of pore sizes between 50 nm and 80 nm, which is present in the 11 μm filter and is absent in the 22 μm filter, plays a crucial part in removing the HSA aggregates. With respect to virus filtration, 1 bar constant trans-membrane pressure filtration shows poor removal ability of ΦX174 bacteriophage (28 nm), i.e., log10 reduction value (LRV) ≤ 3.75, while that at 3 bar and 5 bar achieves LRV[MOU1] [LW2]  > 5 model virus clearance and overall rapid filtration. Removal of protein aggregates during bioprocessing of HSA products is key to improving the filtration flux, which makes it possible to apply virus removal filtration for HSA to ensure its virus safety.   In Paper II, nanofiltration of human plasma-derived intravenous immuno-globulin (IVIG) intermediate (11.26 g L-1, pH 4.9) was carried out to demonstrate high product recovery and high model virus clearance. Virus removal filtration of industrial-grade human IVIG was achieved using 33μm filters at both low (60 Lm-2) and high (288 Lm-2) volumetric load. No changes in IVIG structure were detected and high product recovery was recorded. High virus clearance (LRV ≥ 5-6) was achieved for the small-size model viruses (ΦX174 and MS2 bacteriophages) during the load volume of 60 Lm-2. Side-by-side comparisons with commercial virus removal filters suggest that the nanocellulose-based filter paper presents great potential for industrial bioprocessing of plasma-derived IVIG.   In Paper III, process analytical technology (PAT) approach was employed to identify the critical filter parameters, e.g. thickness, basis weight, pore size, and flux, affecting model virus removal efficiency using filters produced by different hot presses.  The quality parameters were analyzed with ANOVA and Shewhart charts. Compared with other studied parameters, the hydraulic flux appears as the most relevant final product quality attribute of the nanocellulose-based filter paper to reflect the virus removal efficiency. In particular, a 15% higher flux may be associated with a 0.5-1.0 log10 reduced virus clearance (p=0.007). The results are highlight the importance of continued systematic studies in quality assurance using statistical process control tools  [MOU1]Define LRV  [LW2]Defined in the line above