1. Goal 3 was completed by the construction of the water decontamination chamber.
The chamber had a maximum water volume of ~197 mL. The beads with
porphyrins filled ~200mL of volume inside the chamber. Two condenser columns
were used to maintain an average temperature of ~32°C. Two 500 W halogen
lamps were placed on either side of the chamber equidistantly to simulate sunlight.
Medical grade tubing prevented biofilm buildup. A 300 mm steel mesh was used to
allow for water flow and to keep the beads inside the chamber. The rate of flow
could be altered within the range of 0.1 mL/min. to 84 mL/min. The chamber, and
its components, were sealed with silicone. Average contact time of the water with
the porphyrins inside the chamber was ~2.20 minutes. The chamber was
configured to run in a continuous closed loop mode. See Fig. 3.
The research described here focuses on a prototype of a device designed to be
used in sterilization of water contaminated by microorganisms [1]. The device has
potential applications in biomedical field where sterile solutions or potable water is
required, but where other means are not available or impractical. The method of
sterilization described here is based on action of photoactivatable porphyrins
covalently anchored to a solid support. Porphyrins are remarkably efficient light
absorbers throughout practically entire visible spectrum, not just a specific region
such as the UV, which makes them very attractive targets for the abovementioned
applications [2]. Linking them covalently to a solid support dramatically increases
the system’s stability and ease of use. The choice of components of the system
was based on the following principles: it must be inexpensive, easily maintained,
require minimal user input, long lasting, and have the ability to be configured to
meet alternative demands. The device design, application, and efficiency of
process of water photo-decontamination utilizing standardized samples of E. coli
contaminated water are discussed and preliminary results are presented.
The design and execution of the research in question are as follows:
1. Synthesis of 5,10,15,20-meso-para-carboxytetraphenylporphyrin (p-TCPP).
2. Diisopropylcarbodiimide (DIC) mediated conjugation of the porphyrin with the
solid support, and purification of the system, see Fig. 1 & 2.
3. Construction of a small size irradiation flow chamber and charging it with the
porphyrin on solid support heterogeneous photocatalyst. See Fig. 3.
4. Preparation of water samples containing standardized amounts of viable
microorganisms (Escherichia coli Male Strain 3300).
5. Control tests regarding quality of water passing through the system. See Fig. 4.
6. Photo-decontamination experiments (rate of flow, light intensity and time
dependence) and data collection, quantification and interpretation. See Fig. 5 & 6.
Goals 1 - 2 were achieved by standard synthesis of p-TCPP [3,4,5], followed by
activation of porphyrins with DIC and conjugation with amino groups of the solid
support [6], see Fig. 1. Purification of the beads was achieved by flushing the
system with two liters of boiling DI water before every run. The bead size was
confirmed (scanning electron microscope) to be ~530 mm as claimed by the
manufacturer. See Fig. 2.
Solar Powered Water Decontamination
Justin M. Barrett, David R. Williams and Mariusz P. Gajewski*
Arkansas Tech University, Department of Physical Sciences, Russellville AR 72801, USA
Goal 6. The results are summarized. Each experiment altered the time a sample
spent flowing through the system, and all other variables remained constant. The
amount of final colony forming unit (CFU) reduction was tested against the leading
industry standard of water purification systems, in that a system must be able to
perform a >6 log bacterial reduction (99.9999% reduction) from a challenge level of
107 CFU’s per 100 mL of water.
The final bacterial reduction for Experiments 1 and 2 are 2.61×106 and 2.12×106
respectively. Experiment 1 obtained a 97.2612% reduction while Experiment 2
obtained a 91.4224% reduction. After two hours, the amount of additional bacterial
elimination is minimal. See Fig. 5 & 6 and Tables 1 & 2.
Goals 1-6 have been accomplished. The bactericidal properties of the synthesized
porphyrins have been observed. In a two hour time frame, near 99% of the viable
E. coli in solution had been eliminated. The system is capable of eliminating a high
concentration of bacteria per milliliter, however the reduction level is does not meet
the standard of greater than a 6 log bacterial reduction.
Future research will entail testing the effectiveness of eliminating other bacteria,
amoebas, protist, and aquatic fungi. Besides potential applications in the
biomedical field, there is interest in expanding the scope of this project to include
the prospect of decontaminating water for use in developing communities and third
world countries, where clean water or means of purifying it are not readily
available, in hopes to prevent illness from contaminated drinking water.
References: a) picture credit: http://www.energysmartjobs.org/wp-content/uploads/2013/09/Better-Life-With-Renewable-Energy.jpg [1] Jori, G. et al. Journal of Environmental Pathology, Toxicology and Oncology; 2011, 30(3) 261-271.
[2] David Kessel, “Photodynamic Therapy of Neoplastic Disease”, 1990, Vol 1, p.12; [3] P. Rothemund J. Am. Chem. Soc. 1935, 57 (10): 2010–2011. [4] P. Rothemund J. Am. Chem. Soc. 1936, 58 (4): 625–627. [5] Multiauthor, edited by
Kadish, K.M.; Smith, K.M. and Guilard, R. “The Porphyrin Handbook: Synthesis and organic chemistry”, Vol 1. [6] Eric Valeur, E. and Bradley, M.; Chem. Soc. Rev., 2009,38, 606-631.
Figure 5. Experiment 1 – Viable count of E. coli
(×1 mil.) per mL over a 2 hour time frame. Initial
viable count was 2,680,000 bac./mL. Total
bacterial reduction from initial solution was 2.61 x
106 .
a)
Figure 6. Experiment 2 – Viable count of E. coli
(×1 mil.) per mL over a 6 hour time frame. Initial
viable count was 2,320,000 bac./mL. Total
bacterial reduction from initial solution was 2.12 x
106.
INTRODUCTION
GOALS AND METHODS
CONCLUSIONS
RESULTS
Figure 3. Irradiation chamber charged with
beads conjugated with porphyrins, seen here
in purple, with water being pumped inside. The
inlet tube is on the top left of the chamber. The
exit tube is on the bottom right. The bottom
right of the chamber contains a one-way valve
used to periodically and aseptically collect
samples.
Acknowledgements:
This research was supported by ATU ACENRES grant. The authors are thankful to Dr. Scott Kirkconnell (ATU,
Department of Biological Sciences) and Mr. Zac Gleason (Mountain Safety Research) for help with microbiology,
and Mr. Stan Apple (ATU, Department of Mechanical Engineering) for assistance with scanning electron
microscopy.
propionic
acid

coupling with
solid support
Solid
Support
pyrrole
p-carboxybenzaldehyde p-TCPP
solid support conjugated
with porphyrins
DIC/CH2Cl2
Figure 1. Synthetic route leading to polystyrene support with
amino groups covalently linked to p-TCPP molecules.
Figure 2. Polystyrene support with and without covalently bonded porphyrins viewed under optical
and scanning electron microscopes. Note the purple color of the porphyrins on the conjugated
bead, and increased porosity of the systems after the conjugation process. SEM samples were
covered with ~30 nm layer of gold.
Goal 4 was accomplished by inoculating
a 1 L container of TSA broth with E. coli
Male Strain 3300 at ~30 mil. bac./mL to
obtain a stock solution. A 900 mL
solution with a bacterial dilution
containing a viable count of ~2.5 mil.
bac./mL from that stock solution was
prepared per test.
Goal 5 was completed by running four
controls to eliminate any possibility of
other agents killing bacteria, namely
heat or trace chemicals. The control test
results proved that the only active
bactericide was the action of the
porphyrins exposed to light. See Fig. 4.
Figure 4. a) 0.1 mL spread plate of solution with porphyrins, without light exposure at a time of 0 minutes.
b) 0.1 mL spread plate of solution with porphyrins, without light exposure at a time of 60 minutes.
c) 0.1 mL spread plate of solution with porphyrins, with light exposure at a time of 0 minutes.
d) 0.1 mL spread plate of solution with porphyrins, with light exposure at a time of 60 minutes.
Tables 1 and 2. Bacteria count per mL in the solutions for Experiments 1 and 2. Both experiments indicate
~2 x 106 reduction in bacteria count.
Experiment 1
Time Plate Solution Reduction
0 Min. 134 2,680,000
2.61 x 106
(97.2612%)120 Min. 3.67 73,400
Experiment 2
Time Plate Solution Reduction
0 Min. 116 2,320,000
2.12 x 106
(91.4224%)360 Min. 9.95 199,000
FUTURE GOALS
y = -1E-11x5 + 1E-08x4 - 5E-06x3 + 0.0008x2 - 0.0574x + 2.165
0
0.5
1
1.5
2
2.5
0 50 100 150 200 250 300 350 400
y = -1E-09x5 + 5E-07x4 - 8E-05x3 + 0.0058x2 - 0.197x + 2.6754
0
0.5
1
1.5
2
2.5
3
0 20 40 60 80 100 120 140