Grantee Research Project Results

Final Report: Nanosensor for Detection of Saxitoxin

EPA Grant Number: GR832382
Title: Nanosensor for Detection of Saxitoxin
Investigators: Gawley, Robert E.
Institution: University of Arkansas
EPA Project Officer: Hahn, Intaek
Project Period: August 1, 2005 through July 31, 2008
Project Amount: $340,000
RFA: Greater Research Opportunities: Research in Nanoscale Science Engineering and Technology (2004) RFA Text | Recipients Lists
Research Category: Nanotechnology , Safer Chemicals

Objective:

The objective of this research project is to: design and prepare nanoscale sensors for the detection of saxitoxin and other paralalytic shellfish toxins (PSTs).

Approach:

We will base our work on the principles of molecular recognition and photoinduced electron transfer, and the techniques of chemical synthesis, from the molecular to macromolecular scale. We have previously developed a small molecule chemosensor that is selective for saxitoxin over several organic analytes and three common cations found in environmental samples. Using this work as a foundation, we will develop a new generation of fluorescent sensor moieties that are selective for saxitoxin and avoid interference by constituents of the shellfish matrix or other environmental constituents. The sensor moiety will then be incorporated into a nanometer-sized dendrimer to improve sensitivity. To evaluate selectivity, we will screen our sensors against shellfish extracts that have been found to be both toxic and nontoxic by mouse bioassay, and certify the results using HPLC analysis. Finally, we will attach the dendritic sensors to solid support to conduct a preliminary evaluation of a visual sensor.

Summary/Accomplishments (Outputs/Outcomes):

For the past several years, we have been investigating a class of crown ethers having a pendant fluorophore for the detection of saxitoxin (Figure 1).^1-3 We have investigated several aromatic groups for the fluorescence response, including the anthracene,^1,2 coumarin,³ and acridine⁴ fluorophores shown in Figure 1. The binding constants for these compounds to saxitoxin is in the range of 10⁴ – 10⁵ M^–1.

Figure 1. Prototype crown ether chemosensors for saxitoxin. The buffer used is tetrabutylammonium

phosphate.

In 2007, we reported a visible chemosensor in the Journal of Organic Chemistry (Figure 2).⁵

Figure 2. Visible chemosensors for saxitoxin.

In the final year of this project (no cost extension), we focussed on two things, both relating to detection of saxitoxin on self-assembled monolayers. In one project, we successfully showed that the c-lobe of saxiphilin can be successfully immobilized on a streptavidin-modified surface through a biotin linker, and that the dynamics of toxin binding are comparable to that previously observed in solution. As part of this effort, we reported our methods for the recombinant preparation of saxiphilin c-lobe, its complete characterization by mass spectrometry, and the dynamics as determined by surface plasmon resonance. This work was published in Toxicon.

The second goal of the final year was to prepare derivatives of the anthracyl methyl crown and the boron dipyrrin crown that are suitable for assembly into a self-assembled monolayer. The compounds indicated in Figure 3 have been prepared, but we are still working on the incorporation of these species into monolayers suitable for incorporation into a detection device. The effort to incorporate these species onto a quartz surface is being supported by STTR grants from NIH and NSF.

Figure 3. Modified chemosensors for incorporation into self-assembled monolayer.

Students Penny Lewis and Ryan Farris have worked on this project, as well as postdocs Hua Mao. Undergraduate Jennifer Pharr is a coauthor in the paper listed below.

References

(1) Gawley RE, Zhang Q, Higgs PI, Wang S, Leblanc RM. Anthracylmethyl crown ethers as fluorescence

sensors of saxitoxin. Tetrahedron Lett. 1999;40:5461-5465. Corrigendum 6135.

(2) Gawley RE, Pinet S, Cardona CM, Datta PK, Ren T, Guida WC, Nydick J, Leblanc RM.

Chemosensors for the marine toxin saxitoxin. J Am. Chem. Soc. 2002;124:13448-13453.

(3) Kele P, Orbulescu J, Calhoun TL, Gawley RE, Leblanc RM. Coumaryl crown ether based

chemosensors: selective detection of saxitoxin in the presence of sodium and potassium ions.

Tetrahedron Lett. 2002;43:4413-4416.

(4) Gawley RE, Shanmugasundaram M, Thorne JB, Tarkka RM. Selective detection of saxitoxin over

tetrodotoxin using acridinylmethyl crown ether chemosensor. Toxicon 2005;45:783-787.

Corrigendum 746, 477.

(5) Gawley RE, Mao H, Haque MM, Thorne JB, Pharr JS. Visible fluorescence chemosensor for

saxitoxin. J. Org. Chem. 2007;72:2187-2191.

Expected Results:

The research described herein will lead to a photochemical method that could provide a practical alternative to mouse bioassay for the detection of the pathogen saxitoxin in environmental samples. At present, detection of saxitoxin and its congeners in seafood is done by mouse bioassay. Chemical means of detection such as liquid chromatography/mass spectrometry (LC/MS) require considerable technical skill and expensive equipment. Such techniques are not amenable to high-throughput or remote analysis. We anticipate that this project will afford nanoscale sensors that could be attached to a solid support for detection of saxitoxin.

Journal Articles on this Report : 3 Displayed | Download in RIS Format

Publications Views
Other project views:	All 3 publications	3 publications in selected types	All 3 journal articles

Publications
Type	Citation	Project	Document Sources
Journal Article	Kele P, Orbulescu J, Gawley RE, Leblanc RM. Spectroscopic detection of saxitoxin:an alternative to mouse bioassay. Chemical Communications 2006;(14):1494-1496.	GR832382 (2006) GR832382 (Final)	Abstract from PubMed
Journal Article	Lewis P, Fritsch I, Gawley RE, Henry R, Knight A, Lay Jr. JO, Liyanage R, McLachlin J. Dynamics of saxitoxin binding to saxiphilin c-lobe reveals conformational change. Toxicon 2008;51(2):208-217.	GR832382 (Final)	Abstract from PubMed Full-text: ScienceDirect Exit Other: ScienceDirect PDF Exit
Journal Article	Mao H, Thorne JB, Pharr JS, Gawley RE. Effect of crown ether ring size on binding and fluorescence response to saxitoxin in anthracylmethyl monoazacrown ether chemosensors. Canadian Journal of Chemistry 2006;84(10):1273-1279.	GR832382 (2006) GR832382 (Final) R829599 (Final)	Abstract: Canadian Journal of Chemistry Abstract Exit

Supplemental Keywords:

, Sustainable Industry/Business, RFA, Scientific Discipline, Ecosystem Protection/Environmental Exposure & Risk, TREATMENT/CONTROL, Water, Technology for Sustainable Environment, Sustainable Environment, Environmental Chemistry, Engineering, Chemistry, & Physics, Monitoring/Modeling, New/Innovative technologies, Environmental Engineering, Environmental Monitoring, Technology, nanoscale sensors, environmental sustainability, innovative technologies, aquatic ecosystem, nano engineering, marine toxins, chemical composition, environmentally applicable nanoparticles, nanotechnology, nanosensors, nanoengineering, saxitoxin detection, analytical chemistry, aquatic toxins, chemical sensors

Progress and Final Reports:

Original Abstract

2006 Progress Report

2007

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