HARMFUL ALGAL

TOXINS DETECTION

FOR REGULATORY

PURPOSE

GIRES USUP

Universiti Kebangsaan Malaysia

Vilarino et al. (2013) Detection of aquatic algal toxins has become critical for the protection

of human health. During the last 5 years, techniques such as optical, electrochemical, and piezoelectric biosensors or fluorescent-microsphere-based assays have been developed for the detection of aquatic algal toxins, in addition to optimization of existing techniques, to achieve higher sensitivities, specificity, and speed or multidetection. New toxins have also been incorporated in the array of analytical and biological methods. The impact of the former innovation on this field is highlighted by recent changes in legal regulations, with liquid chromatography-mass spectrometry becoming the official reference method for marine lipophilic toxins and replacing the mouse bioassay in many countries. This review summarizes the large international

effort to provide routine testing laboratories with fast, sensitive, high-throughput, multitoxin, validated methods for the screening of seafood, algae, and water samples.

TOPICS

Toxicities and toxins

Current status of detection methods

Issues and challenges

Should we ‘reinvent the wheel’?

How much are we willing to pay to keep our seafood

safe?

Paralytic shellfish poisoning

and PSTs

Regulatory status Regulated: Yes

Regulatory limit: 800 ug/kg STX eq (600 in some countries)

Current methods:

MBA with 0.1M HCl (15min) (EC) No 2074/2005

LOD: 370 ug/kg STX eq

HPLC-FLD (Lawrence method) (EC) No 1664/2006

LOQ: 10-80 ug/kg STX eq for individual analogs

RBA: LOQ in the femto-molar range STX eq

Standardized methods:

AOAC method 959.08 (MBA)

AOAC method 2005.06 (HPLC-FLD)

AOAC OMA-2011.27 (RBA)

Potential MBA replacement

for PSTS

Diarrheic shellfish poisoning and toxins

Regulatory status

Regulated: Yes

Regulatory limit:

OA: 160 ug/kg OA eq

Pect-2: 160 ug/kg OA eq

AZA-1: 160 ug/kg AZA eq

YESSO: 1000 ug/kg YTX eq

Current methods:

MBA or Rat bioassay with acetone extraction (24h)

(EC) 2074/2005

Standardized methods:

None yet

Potential MBA replacement for DSP toxins

Amnesic Shellfish Poisoning

and toxins

Regulatory status

Ciguatera Fish Poisoning and Toxins

Regulatory status

Note: EU and WHO suggested limit of 0.01 ug/kg P-CTX1 eq as safety limit for human

consumption

Issues and Challenges Toxin standards

Equipment

Personnel

Transportation

Operational budget

How much are we willing to

invest? Equipment

LC/MS/MS

Scintillation counter

Associated extraction consumables

Manpower

Training of dedicated personnel

Operational costs and sustainability

Possible cost-sharing between government and industry

Anal Bioanal Chem. 2013 Sep;405(24):7719-32. doi: 10.1007/s00216-013-7108-6.

Epub 2013 Jul 3.

Innovative detection methods for aquatic algal toxins and their presence in the food

chain.

Vilariño N1, Louzao MC, Fraga M, Rodríguez LP, Botana LM.

Author information

Abstract

Detection of aquatic algal toxins has become critical for the protection of human

health. During the last 5 years, techniques such as optical, electrochemical, and

piezoelectric biosensors or fluorescent-microsphere-based assays have been

developed for the detection of aquatic algal toxins, in addition to optimization of

existing techniques, to achieve higher sensitivities, specificity, and speed or

multidetection. New toxins have also been incorporated in the array of analytical and

biological methods. The impact of the former innovation on this field is highlighted by

recent changes in legal regulations, with liquid chromatography-mass spectrometry

becoming the official reference method for marine lipophilic toxins and replacing the

mouse bioassay in many countries. This review summarizes the large international

effort to provide routine testing laboratories with fast, sensitive, high-throughput,

multitoxin, validated methods for the screening of seafood, algae, and water

samples.

PMID: 23820950 [PubMed - indexed for MEDLINE]

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Molecular Interactions

Interactomics

Signal Transduction

Cell Adhesion

Antibody Binding

Endogenous Receptor-Ligand

Interactions

DNA Binding Proteins

Drug – Target Interactions

What can be studied?

Scope of Technique

Proteins

Nucleic Acids Lipids & membrane molecules

Carbohydrates

Small-Molecule Drugs

Whole Cells

Viruses Bacteria

Biomolecular Interaction Analysis: BIAcore

Detection principle: Surface Plasmon Resonance: SPR

One binding partner (LIGAND) immobilised on chip

Other (ANALYTE) injected: microfluidics

PC collects binding data in real time

Chip is regenerated to remove analyte

Cycle is repeated

Comprehensive information

Molecular interactions in real time:

Detect

Yes/No

Identify

Specificity

Binding partners

Characterize

Affinity

Kinetics

Epitope mapping

Concentration

Thermodynamics

Mass Spec Link-Up

Versatility

• Range of chip surfaces

• Range of coupling chemistries for different needs

Surface Plasmon Resonance

• Critical angle of polarised light total internal reflection

• Results in excitation of surface plasmons in the gold

• Creates evanescent wave field dissipates into sample matrix

• Intensity of reflected light depletes

• Critical angle changes with changes in sample matrix (binding)

• Change in angle is converted to resonance signal directly proportional to mass

bound at surface