So if we were designing a heparinase reactor how would we get the beads containing the immobilized enzyme in a well-mixed state? The paper by Langer’s group, ie. Ameer et al. 1999 (I) looked at fluidization of whole blood in a Taylor-Couette Flow Device The Taylor vortices would facilitate the fluidization of the agarose beads and lead to a well-mixed reactor Taylor vortices are flow instabilities that occur within an annular gap of concentric cylinders when the inner cylinder is rotated above a critical rotation rate

Factors affecting the fluidization of the beads included: Gap space, 3.2 mm and 6.4 mm Rotation rate of inner cylinder Flow rate of blood thru the device

Results and Discussion Immobilization bound 95% of the soluble enzyme and retained 30-40% of the enzyme activity 2. Over a 4 day period of operation no leaching of the enzyme from the agarose was detected 3. Heparin degradation products were found to be nontoxic at [ ]’s up to 100x’s the expected levels during clinical exposure 4. Phase I clinical trials in healthy volunteers showed no acute reactions when exposed to soluble heparinase which was obtained from Flavobacterium heparinum

Narrow gap 90% removal of heparin in 3 minutes

Attempted to reduce blood damage in this initial design by: Increasing the gap width while maintaining annular volume constant Reducing the rotation rate Note that conversion is now about 80% at 3 minutes

Note that Hb release is significantly reduced between design I and design II

Blood cell damage reduced from design I to design II

Constant flow showed about a 23% per pass removal Clinical application needs about 45%

Although Taylor vortices are usually gentle to RBC’s this study showed that when used in combination with the agarose beads hemolysis of the RBC’s is a big problem. So, how would you solve this problem? The next paper by the same group and labelled II illustrates how they attacked this problem Design criteria: efficacy, need a 40-50% per pass heparin conversion Safety, no significant effect on whole blood cells, no hemolysis, no reduction in platelets or white blood cells, and no activation of these cells either 3. stability, stable operation at flowrates up to 300 ml/min 4. simplicity, simple operation and low cost To minimize blood damage, a device that SIMULTANEOUSLY effected plasma separation and the enzyme reaction with the capacity to treat whole blood at high flowrates was designed.

Previous paper Poor mass transfer and loading issues Blood cells and the platelets would not come into contact with the agarose beads and hence minimize hemolysis

Conversion about 34% and below their target of 40-50%, but could be increased by adding more beads

Eliminating contact between the cells and the agarose beads as in this design was an effective way to significantly reduce hemolysis.