SAXITOXIN

1
SAXITOXIN

• A Marine and Freshwater Toxin

2
Toxicity

• Associated with Red tides
• Origin
• Dinoflagellates
• Gonyaulax tamarensis
• Gonyaulax catenella
• Cyanobacteria
• Cylindrospermopsis raciborskii
• Anabaena spiroides
• Major transvectors are organisms feeding on
  producers
• Molluscs, crabs, snails, etc.

3
Type and properties

• Tetrahydropurine
• Guanidinium
• Heterocyclic
• Hygroscopic
• Water soluble
• Thermally stable
• MW of 299g

4
STX and its derivatives
STX has a total of about 20 different derivatives
5
Mechanism of Action

• Absorbed through mucosae and excreted in urine
• Blocks nerve
• signal
• transmission

6
Sodium Ion Channel

• STX binds to receptor site extracellularly
• Electrostatic attraction involving 7,8,9
  guanidinium
• Hydrogen-bonding involving C-12 hydroxyls
• Located on a subunits
• 4 homologous domains and 6 transmembrane segments
• P-loop between S5 and S6
• S4 contains positive aas
• EEDD outer and DEKA inner form selectivity filter
  and binding site

7
Nerve Propagation
8
Potassium Ion Channel

• STX acts internally
• Human ether-a-go-go-related gene
• hERG
• Associated with potassium ion current
• Four a subunits and 6 transmembrane segments
• Four domains surround central pore
• STX binds 11
• Stabilizes the closed state

9
Voltage Activation Curves
10
hERG
11
Clinical Diagnoses

• Symptoms appear with 2 hours of ingestion
• Paralysis
• Weakness
• Headache
• Nausea and vomiting
• Death within 12 hours, depending on severity
• Recovery in 2 to 7 days
• No lasting side affects
• Paralytic Shellfish Toxins (PSPs)

12
STX Studies

• Classic form is Mouse Bioassay
• Limit 0.4µg/g shellfish meat
• Non-specific for STX
• Enzyme Immunoassays
• Developed by Johnson and Mulberry (1966)
• Formaldehyde condensation
• Antitoxin produced in rabbits
• Antiserum STX used in hemagluttination and
  bentonite flocculation tests
• Toxin precipitated
• Specific for STX, but only qualitatively

13
More STX Studies

• Fluorimetric determination
• Acidic extraction and alkaline oxidation
• Spectrophotometric
• Limit .004µg/g meat
• Could not detect different PSPs
• Crown ethers
• Anthracyl group serves as a fluorescent probe
• STX binds through C-8 and C-12

14
General Structure
15
STX Docked In Crown
16
Last But Not Least

• HPLC
• Bonded cyano column
• Alkiline oxidation using periodic acid and sodium
  phosphate buffer
• Six individual PSPs identified and quantified
• Limit 0.2fg/g

17
Natural Detection

• Saxiphilin (SAX)
• Compound isolated from N.A. bullfrog
• Rana catesbeiana
• Soluble protein
• Detected using tritiated-STX (H3-STX)
• Specific and high affinity for STXs
• Derivatives of STXs and SAXs used as radioligands
• Measure sodium density channels
• Does not bind STX competitor TTX

18
Saxiphilin (cont.)

• Structure
• N-lobe and C-lobe
• Binding site for STX on C-lobe
• Cloning and sequencing cDNA
• 90-92 kDa protein
• Related to transferrin family
• Purpose and action still unknown
• Maybe a species-specific detoxification mechanism
• STX has high affinity for SAX

19
Conclusion

• STXs are dangerous marine and freshwater toxins
• Biosynthesis still unknown
• Valuable for medical research
• Nervous system disorder
• Implicated for use in chemical warfare
• No cure known

20
References

• Edwards, Neil. "Saxitoxin...from food poisoning
  to chemical warfare...." The Chemical
  Laboratories. 18 Aug. 1998. University of Sussex
  at Brighton. 14 Nov. 2005 http//www.bris.ac.uk/De
  pts/Chemistry/MOTM/stx/saxi1.htm.
• "Environmental Health Criteria 37 Aquatic
  (Marine and Freshwater) Biotoxins." International
  Programme on Chemical Safety. 1984. World Health
  Organization. 14 Nov. 2005 lthttp//www.inchem.org/
  documents/ehc/ehc/ehc37.htmgt.
• Fleming, Lora E. "Paralytic Shellfish Poisoning."
  NIEGS Marine and Freshwater Biomedical Sciences.
  14 Nov. 2005 lthttp//www.whoi.edu/science/B/redtid
  e/illness/psp.htmlgt.
• Gawley, R.E., Pinet, S., Cardona, C.M., Datta,
  P.K., Ren, T., Guida, W.C., Nydick, J., and
  Leblanc, R.M. J. Am. Chem. Soc., 124, 45, 13448 -
  13453, 2002, 10.1021/ja027507p.
• Goldin, Alan L. "Mechanisms of Sodium Channel
  Inactivation." Current Opinion in Neurolbiology
  13, 284-290 (2003). 2 Dec 2005 lthttp//www.biol.sc
  .edu/vogt/courses/neuro/pdf/Na-channel-inactivati
  on-Goldin-CurrOpNeuro-2003.pdfgt.
• Jixin Wang, Joseph J. Salata, and Paul B.
  Bennett. Saxitoxin Is a Gating Modifier of hERG
  K Channels. J. Gen. Physiol., 121 583-598
  published online before print as
  10.1085/jgp.200308812.
• Krishnan, Gomathi, Maria A. Morabito, and Edward
  Moczydlowski. "Expression and characterization
  of Flag-epitope- and hexahistidine-tagged
  derivatives of saxiphilin for use in detection
  and assay of saxitoxin." Toxicon 39, 291-301
  (2001). 27 Nov 2005 lthttp//people.clarkson.edu/e
  moczydl/niallpaper1.pdfgt.