Functionalization of Surfaces with Nisin in a Poly[ethylene oxide] brush layer

Matt RyderDr. Joe McGuire – BioEngineeringOSUHHMI Summer ‘07

Hydrophobic Surface

Background – Instances of Infection1

Infection in hospitals is the fourth largest killer in the US

2 million patients contract infections in hospitals each year…about 103,000 die as a result 228 infections/hour…12 deaths

Cost? $30.5 billion each year

Background Three issues with implanted devices

Clot Formation Bacterial Adhesion Cell Proliferation

Currently, methods to counteract include: Loading patients

with heparin or antibiotics.

Background

These issues are directly related, both are initiated by adsorption events.

Hydrophobic Surface

Background – Brush Layer

F108 creates brush layer that protects against protein adsorption. Protein adsorption can result in

very different outcomes, from benign surface coatings, to large clot formation (therefore risk of stroke).

Brush Layer – Protein Repellant

A.

F108 Coated SurfaceBare Surface

F108 Coated SurfaceBare Surface

B.

Background – Nisin

Nisin is a small antimicrobial peptide produced by strains of Lactococcus lactis subsp. lactis. Nisin kills Gram positive bacteria through a multistep process that destabilizes the phospholipid bilayer of the cell and creates transient pores. The efflux of low molecular weight compounds from the cytoplasm and subsequent dissipation of membrane potential rapidly kills the targeted bacterium.

Nisin - Mechanism

Background – Nisin & Brush Layer

Goes against function of brush layer

Hydrophobic Surface Hydrophobic Surface

Background – Product Activity

Hydrophobic Surface

Hypothesis

If a surface can be chemically or functionally modified to adsorb and retain the Lantibiotic Nisin, antimicrobial activity and anti clotting function will be higher as compared to current coating methods.

Procedure

overnight

x # of days

2 days

microspheres

F108

Nisin

Pediococc

us

Results

Optimized concentrations of F108, Nisin and microspheres in solution.

Conducted serial dilution tests to find optimum concentration of Pediococcus.

Researched literature for background knowledge and future effectiveness

Standardized procedures to decrease variability.

Obtained valuable insight on Nisin activity from longevity testing.

Results

10-6 dilution w/o Nisin

10-4 dilution w/ Nisin

Dilution optimized for 30-300 colonies

Results

Results

Future Research

28 day trials with current procedure Blood serum studies to test

longevity of Nisin in physiological conditions

Tests with EGAP rather than F108, a more clinically accepted polymer

in vitro studies using catheters

Acknowledgements

Special Thanks to: Dr. Joe McGuire – Mentor Dr. Christine Kelly Karle Schilke Dr. Jeff Tai – Protocols and instruction Dr. Kevin Ahern – HHMI Program The Howard Hughes Medical Institute

References

1. Committee to Reduce Infectious Diseases http://www.hospitalinfection.org/essentialfacts.shtml

Pictures1. http://www.flickr.com/photos/gaspirtz/384254225/

2. www.altham.com/html/food_hygiene_cartoons.html

3. http://www.sciencestuff.com/prod/L-p-Empty/1001-20

4. http://www.flickr.com/photos/rdbkorn/85401201/

5. http://www.bergoiata.org/fe/favs/Bacteria.jpg

6. www.sigmaaldrich.com/img/assets/4261/micro_7.gif