Electrochemical Determination of Diffusion Coefficients: Anomalies in Constrained Channels

1. Electrochemical determination of diffusion coefficients: Observations at tissue matrix barriers and major anomalies in constrained channels K Adatia, M Raja, P Vadgama Queen Mary, University of London

2. Fick’s Laws Can we use molecular diffusion as a probe for water structure? Jakubowski 2016

5. Enzyme biosensorsDiffusion control membranes Enzyme Molecular concentration Current Flux

6. d L Recessed tip voltammetry electrodes for biomatrix/H2O isolation Initial condition: C(0<x<L, t=0)=0 Boundary condition: C(x=L, t)=C0 100μm Cotterell Equation

7. Bipartite analytical solution for cylindrical coordinate: Fick’s Laws Normalised time T≥ 0.063 T< 0.063

8. Recessed Lactate electrode Increased recess length Effective enzyme Km D values as predicted for bulk water

9. Tissue matrix • Signalling molecules • 3D scaffolds • Drug delivery Artificial membranes • Biosensors • Haemodialysis • Bioreactors Special case of the blood brain barrier Blood Brain Barrier

10. Track etch polymer membranes 0.4µm polycarbonate (PC) 8.0µm polycarbonate (PC) c) 0.4µm polyethylene terephthalate (PET)

11. 4b) 4a) Gaussian pore distribution – no tortuosity 8µm PC 0.4µm PC 4c) 0.4µm PET

12. Planar electrochemical Cell

14. Cotterell Randles-Sevcik equation

15. Bulk water diffusion coefficients

16. Dialysis membrane permeability ~ bulk water Catechol

17. Pore normalised diffusion coefficients – track etched • PC – PET different –Cannot be wall effects at µm scale • Charge and MW dependent –Cannot be polymer charge ?Zeta potential Large pore PET Small pore PC

18. Bulk H2O normalised diffusion coefficients

19. Dpore/Dwater Ratios

20. Artefact? Amperometryvs CV < PET PC Amp CV CV

21. Surface geometry underestimates pore diameters

22. Permeabiitydetermined by the material • Total pore area 0.4µm PC > 8µm PC > 0.4µm PET • Diffusivities: • 0.4µm PC > 0.4µm PET > 8µm PC PC imposes the greater constraint

23. Pore constrained water - different properties

24. ECM • Types I IV collagen, fibronectin, heparin and chondroitin sulphate • Collagen concentration effects • Charge selective? • Enantioselective? Tendon Cartilage

25. Type I and Type IV Collagen: different roles I IV Sheets Fibres SEM Type IV collagen SEM Type I collagen (5mg/ml)

26. Crosslinked Membrane mats Jakubowski

27. Collagen I concentration effect *** *** ** *** ** *** ** ** *

28. Collagen I vs Collagen IV I IV ** ** ** **

29. Bulk water normalised diffusion - Collagen IModulated microsolute transport (cf track etched) Ratios should be the same for all molecules for a given collagen concentration

30. Membrane charge effect only on ascorbate – Hep, CSO4 *** *** *** *** *** Hep CSO4

31. Bulk water normalised charge selectivity with heparin (H) and chondroitin sulphate CSO4 Ratios all different H CSO4 H2O2 Ascorbate H CSO4

32. Natural cartilage avascular : surprising open system: we cannot reproduce Cartilage 200mg/ml collagen D~10-6cm2 s-1 Dialysis membrane

33. Tendon diffusion two orders lower: a physiological problem for cells 300mg/ml collagen D~10-8cm2 s-1 + Fibre orientation effect

34. Track etched I IV Hep

35. Is the data reliable? Two methods Dialysis membrane H2O Multipoint fit Simulation/data (Dialysis) Simulation/data (Track etch membrane)

36. Conventional surface model A) Triple water phases : (i) Surfacefixed water (non-freezing) (ii) Loose bound ‘gel water’ (freezing) (iii) Bulk water Gel water determines biocompatibility: PEO augments gel water (Tanaka) Quasi-aqueous PEO cushion

37. Conclusions • Porosity doesn’t predict permeability • Unknown effect • Double layer problem • Cell biology problem Acknowledgements: BBSRC - Flowers’ Scholarship (KA)

40. Precision pore analysis possible

41. Cylindrical pore membrane permeability:Simple? 0.4µm polycarbonate 8.0µm polycarbonate 0.4µmpolyethylene terephthalate (PET)