1. CARBON BLOCK VS. GRANULAR COLUMNS FOR BINDING OF LIVER FAILURE TOXINS
Stephen R. Ash MD FACP and David J. Carr MsChe
HemoCleanse, Inc. and Clarian Arnett Health Lafayette, IN
ESAO 2007, Krems, Austria

2. The Problem
Capacity for high molecular weight and protein-bound toxins has limited carbon’s efficacy in extracorporeal therapy (and other sorbents). Most of the active carbon surface in granules is in the interior, hidden from the flowing stream and protein-bound toxins.
Small particle size allows direct interaction of sorbents with macromolecules and bound toxins. However, particles of 1-10 microns are impossible to directly fabricate into columns.
Sorbent suspensions are difficult to retain during convection at membranes.

3. Adsorption Background
Transport Processes
Bulk Convection
Axial Dispersion
Film Diffusion
Pore Diffusion
Surface Diffusion
Adsorption Processes
Aggregation
Adsorption
Denaturation
Interference
Solid Phase Reaction

4. Diagram courtesy Dr. N.-H. L. Wang
Adsorption Processes Diagram
Diagram courtesy Dr. N.-H. L. Wang

5. Micropores vs. Mesopores

6. Using sorbent regeneration avoids need for large amounts of plasma and sterile replacement fluid, making the system easier to implement and control...

7. But if the sorbents saturate, there is decreasing clearance of hepatic toxins during the treatment. Example: decreasing clearance of bilirubin over time in the MARS system (partly due to column saturation):

8. Further evidence for sorbent capacity limitations
PrometheusTM : “Blood clearances of protein-bound toxins decrease over time. The rate and the efficiency of removal of albumin-bound toxins are interrelated to both the strength of the albumin binding and the saturation of the adsorption columns” (Cl tB 29.3 ± 5.1 vs ± 3.7)
*P. Evenepoel, Y. Vanrenterghem et al., Detoxifying Capacity and Kinetics of Prometheus® - A New Extracorporeal System for the Treatment of Liver Failure , Blood Purification

9. Our Project Goals:
Develop method of screening sorbents for detoxification applications to predict removal of small and protein-bound toxins.
Compare efficacy of toxin removal by mesoporous carbons in several physical forms.

10. Activated Carbons Tested
Description
Surface Area, m2/gram
Bulk Density, g/mL
Preliminary Study
Maxsorb Pellets
1.5mm diameter
2,060
0.31
Maxsorb Powder
25 to 75 μm
Norit A
Powder, 1 to 25 μm
1,700
0.22
Granular
Norit C Gran: ,700 μm
1,400
0.20
HSGD
Synthetic beads, ,000 μm
~1,600
0.10
Block
Immobilized powder
1,300
0.41
Nanofiber
800
0.21

11. HSGD 500 microns 50 microns 10.0 microns 2.0 microns 1.0 microns
Micrographs courtesy Dr. VG Nikolaev

12. Maxsorb Pellets
Carbon Block
Nanofiber

13. Preliminary Study
Bilirubin adsorption of two carbons with similar surface areas was compared as a function of particle size. Maxsorb carbon is commercially available in pellets. It was tested as pellets and as powder after grinding in a mortar and pestle and sieving.
Equilibrium binding of bilirubin in 5% albumin was tested for these carbons. Initial [bilirubin] was up to 12 mg/dL.

14. Bilirubin Adsorption by Activated Carbon
Powdered vs. Granular

15. Results of Preliminary Study Langmuir coefficients for bilirubin
Maximum Capacity, mg/g carbon
Relative Capacity
Binding Constant, mL/mg bilirubin
Relative Binding Constant
Maxsorb Pellets
0.069 1
32.8
17.4
Maxsorb Powder
3.5 51
1.9
1.0
Norit A
20.7
300
4.0
2.1
Powdered carbons had much higher bilirubin capacity than granular carbon.

16. Materials & Methods
Activated carbons were tested as powders in mixed suspension and as columns of beads or immobilized particles. Test conditions were scaled from human clinical application. Isothermal adsorption of 3 compounds from aqueous solution at low concentration ( ppm) was used as a screening criterion: methylene blue (MW 320), albumin (MW 66,000) and blue dextran (MW 2,000,000).
Three carbons with the highest large-molecule adsorption were tested in columns. Adsorption at 37°C and constant pH of bilirubin (MW 585) or cytokines (IL-1β, IL-6, & IL-10) from plasma was tested in a system that recirculated treated plasma to a tank simulating a patient for 10 hours.
Removal efficiency is the final toxin concentration in the tank is expressed as a percentage of the initial tank concentration.

17. Results: Binding of Marker Molecules

18. Results: Binding of Marker Molecules

19. Results: Binding of Marker Molecules

20. Results: Binding of Bilirubin
Granular carbon=near zero

21. Results: Binding of Cytokines
Granular carbon=near zero

22. Results Summary
Methylene blue performance is similar for all the carbons tested.
Carbon adsorbs blue dextran in proportion to its mesoporous character AND to the surface area exposed to flowing fluid.
Carbons with significant blue dextran interaction also remove bilirubin and cytokines from plasma.
Carbon block (powdered) removes bilirubin and cytokines about as well as HSGD, the best clinically tested carbon.

23. Carbon Comparison +++ ++ + -- - With coating Carbon Block HSGD
Carbon Block
HSGD
Nanofiber
Granular
Carbon Density
+++ ++ +
Lack of Fines --
Small Toxin Capacity
Bilirubin Capacity
Cytokine Capacity
Hemoperfusion capable -
With coating
Additional sorbent capable

24. Conclusions
Blue dextran adsorption from aqueous solution is indicative of in-vitro bilirubin and cytokine binding capacities.
Mesoporous carbons with high surface area in contact with flowing fluid are the best candidates for clinically effective sorption of protein-bound toxins.
Examples are HSGD and carbon block (pore size range = 2 to 50 nanometers)
For reasons of density, lack of fines, flexibility, carbon block is a practical and effective choice.

25. Progress towards Carbon Block for Biological Fluid Regeneration
Carbon type, particle size, and size of block
Purity of perfusate-AANSI standards for metals, endotoxin, bacteria
Free of organics-GCMS assay
Case design
Sterilization of product
Priming with sterile fluid
Platform definition-regenerate dialysate, then albumin-dialysate and plasma.

26. Alternative Carbon to Consider: carbide-derived carbon

27. Once the artificial liver is built, how to test it? Rats!
Peritoneal implants 107 Cells in membranes

28. Sorbent-Based Pheresis in the Rat

29. Plasmafilter and Sorbent Reactors

30. Animal Interface

31. Hydraulic Performance

32. Blood cellular and chemical component tests

33. Treatment Results include survival to death or euthanasia by defined criteria
Pheresis with sorbents and/or cells is possible for a rat liver failure model

35. Artificial liver support therapy for patients with fulminant hepatic failure currently used in Japan- TAD, Yoshiba et al.

38. Evidence for sorbent capacity limitations
MARS : The removal efficiency of albumin-bound toxins drops after the initiation of treatment to become insignificant after 6 hours due to both the strength of the albumin binding and the saturation of the adsorption columns*
*P. Evenepoel, Y. Vanrenterghem et al., Detoxifying Capacity and Kinetics of the Molecular Adsorbent Recycling System Contribution of the Different Inbuilt Filters, Blood Purification