rapid mycotoxin analysis for the field waters technology … · 2017. 8. 7. · ©2017 waters...

©2017 Waters Corporation 1COMPANY CONFIDENTIAL

Rapid Mycotoxin Analysis For The Field

Waters Technology Conference 2017

Patricia Jackson

VICAM, A Waters Business


Mycotoxins

• Mykes: Greek for fungus/mold

• Toxicum: Latin for poison/toxin


Mycotoxins AFLATOXINSB1, B2, G1, G2, M1

DEOXYNIVALENOL FUMONISINSB1, B2, B3

OCHRATOXIN A

T-2/HT-2 ZEARALENONE

Selected MoldsThat Produce Toxins

Aspergillus flavus,Aspergillus parasiticus

Fusarium graminearum

Fusarium verticillioides

Aspergillus ochraceusPenicillium verrucosum

Fusarium and other moldspecies

Fusarium graminearum

Foods Susceptibleto Contamination

Maize, groundnuts, nuts,cottonseed, copra, spices, milk, wheat, oats, barley, and rice

Maize, wheat, barley,malted barley, and oats

Maize and other cerealgrains

Maize, wheat, barley, beer,oats, sorghum, dried vine fruits, wine, coffee, and cocoa

Maize, wheat, barley, oats,rice, sorghum, and other cereal grains

Maize, wheat, barley,grain, and sorghum

Health Effects Liver cancer and damageImmunosup-pressionDecreased milk and egg production

Damage to digestivetract, bone marrow, spleen, reproductive organsWeight loss, vomiting, and feed refusal

Cancer in ratsBrain decay in horses Lung congestion in pigsHuman Esophageal Cancer

Kidney damage andcancerImmunosup-pression

Skin and oral lesionsin livestock and humans Alimentary toxic aleukia in humansConsidered 10x more toxic than DON

Negatively impactsreproduction, fetal development, and the health of newbornsCauses feminization in animals at 1 ppm


• Tree Nuts

• Peanuts

• Grain

• Wine

• Coffee

• Flour Milling

• Cereals

• Feed

• Oats

• Ethanol

• Dairy

• Rice

• Botanicals

• Spices

• Snack Foods

• Pet Food

Food & Agricultural Products Affected by

Mycotoxin Contamination


Why Field Based Mycotoxin Detection Matters:

Harvest

•Screening

•Confirmation

Storage

• Segregation

• QC Before Processing

Finished Products

• Laboratory QC

•Prior to Export or Import


Technologies Commonly Used to Detect Mycotoxins

LC-MSUPLC-MS-MS

HPLCor UPLC

Quantitative Strip tests orImmunoaffinitycolumns with Fluorometer

QualitativeStrip tests

Higher Complexity/CostLower Complexity/Cost

Raw Commodities

Buying Points

Groundnut Producer/Shellers

Grain & Feed

Finished Product Testing

Import/Export

Research

Independent Laboratory


Rapid Methods vs. Laboratory Methods

Rapid Methods Laboratory Methods

Extraction Extraction

Simple to Use Training, Expertise Needed

Result in minutes Method Dependent, Batch Sample Runs

Less Sensitive Highly Sensitive

Lower Setup Cost Capital Investment

Little Training/Expertise Required

Well Trained Technician


Rapid Field-Ready Testing Approaches

• Fluorometer

• Qualitative Screening

• Quantitative Strip Tests


Qualitative Lateral Flow Strip Tests

• AflaCheck : 10ppb or 20ppb screening level

with one strip

• DONCheck: 1 part per million (ppm) screening

level


AflaCheck Strip Test Procedure

Peanut or Corn

SampleMeasure 5g Add 10mL 70% MeOH

(10ppb cut off)

Shake 1 min.

Let settleDilute sample 1:1

with water; add strip

Positive

Negative


DONCheck Strip Test Procedure

Measure 2g Add 500 uL

Additive + 12

mL Water

Shake 1 min.

Let settle

3 min.

Add 250 uL sample to

strip test vial, insert strip

Wait 3-10 Min

Positive

Corn, Wheat or

Barley Sample

Negative


Quantitative Strip Tests


Quantitative Strip Tests with Vertu Reader

Vertu Reader

• 5-10 Minute Development Time

• Eliminate Organic Solvent Use

• Simple User Interface

• Limited Hands-On Time

• Reduce Test Error


• Solvent-free extraction

• Results in 5-15 minutes*

• Simple To Use

• Print/Download Results

*after extraction**limits of detection will vary based on procedure. † with dilution

AQUA Strip Test Methods – Solvent Free, Sustainable

• Sensitive

• Long Shelf Life

• Real Time Data

• Print or Download Results


Barcode Scanner: Vertu Reader

Calibration


Quantitative Strip Tests:

Afla M1-VAfla M1-V HS


NEW AQUA Solvent-Free Extraction


Afla-V AQUA HS (Groundnuts)

• Results in 5 minutes*

• LOD: 2 ppb

• Range: 0 to 100 ppb

Procedure:

Weigh 10 g

Sample, add 50

mL AQUA Premix

Vortex 2 min,

Filter,

Apply 100uL to

Strip

After 5 Min,

Place in Vertu

Reader, Analyze


Fumo-V AQUA for Corn


• LOD: 0.2 ppm**

• Range - 0 to 100 ppm† (1st Strip: 0-20 ppm; 2nd: 20-100 ppm)

*after extraction**limits of detection will vary based on procedure. † with dilution

Weigh 5g

Sample add 25

mL AQUA Premix

Vortex 2 min,

Filter,

Apply 100uL to

Strip

After 5 Min,

Place in Vertu

Reader, Analyze

Procedure:


DON-V for Wheat and Corn


• LOD: 0.2 ppm

• Range: 0.2-10 Parts per million (ppm)

•

Procedure:

5g Sample + 20mL

DI Water,

Vortex 2 min

Filter,

Dilution 1

200 uL Water +

100 uL Filtrate.

mix well.

Dilution 2

Add 100 uL

Diluent to 100 uL

sample.

Mix well,

Incubate strip

test and solution

for 2 min @ 40º

C,

Apply 100 uL

sample mixture

to strip test

Allow strip to

develop 5 min;

Place in Vertu

reader, analyze

for DON level.


Ochra-V AQUA for Wheat, Green Coffee


• LOD: 2 ppb

•Range: 0-30 Parts per billion (ppb)

Procedure:

Weigh 5g

Sample add 25

mL AQUA Premix

Vortex 2 min,

Filter,

Apply 100uL to

Strip

After 5 Min,

Place in Vertu

Reader, Analyze


Afla M1-V for Raw and Whole Milk

• No sample prep

• Meets US Maximum of 500 ppt (0.5 ppb)

• Results in 8 minutes

• LOD: 100 ppt

• Range: 100-750 Parts per trillion (ppt)

Add 200 uL Cold

Milk to Strip Test

Vial

Insert Strip Test

and Incubate 8

minutes

Place in Vertu

Reader, Analyze

Procedure:


Coming Soon….Afla M1-V High Sensitivity

• Results in 8 minutes

• Meets EU Reg Limit of 50 ppt

• LOD: 25 ppt

• Range: 0-200 Parts per trillion (ppt)

Add 200 uL Cold

Milk to Strip Test

Vial

Insert Strip Test

and Incubate 8

minutes

Place in Vertu

Reader, Analyze

Procedure:


Immunoaffinity Column with Fluorometer


Fluorometer Calibration


Inside a Mycotoxin Immunoaffinity Column

Support

Bead

MonoclonalAntibody

PBSBuffer


Mycotoxins Impurities

Inside a Mycotoxin Immunoaffinity

Column cont…



Column cont…


Impurities

Mycotoxins bound to monoclonal antibodies

Mycotoxins


Column cont…


Column Wash

http://www.google.com/url?sa=i&rct=j&q=&esrc=s&source=images&cd=&cad=rja&uact=8&docid=wr5wpiD5RJzXfM&tbnid=Vw7WD_93ss3NuM:&ved=0CAUQjRw&url=http://www.askipedia.com/category/earth-geography/page/2/&ei=MaXGU_uqFfK_sQSSlICYCA&bvm=bv.71126742,d.cWc&psig=AFQjCNG6WH8NdSdQRWhLFnB71PVRXxKAhQ&ust=1405613657387235




















Column cont…Mycotoxins bound to monoclonal antibodies


Methanol elution




















Purified mycotoxin(s) in methanol


Get results by…

Obtaining Results With the Fluorometer….


Mycotoxins Methods Using Fluorometer

- AflaTest IA Column

-Total Afatoxins {B1+B2+G1+G2}

-6 Minute Test Time

-Range 0 – 1000 parts per billion (maize)

- LOD < 3 parts per billion (ppb)

- FumoniTest IA Column

-15 Minute Test Time

- Range: 0 – 10 parts per million (ppm) Corn/Milo

- LOD: 0.25ppb (corn), 0.50 (Milo)

- OchraTest IA Column

- 15-20 Minute Test Time

- 0-100 parts per billion (ppb) for Corn

- LOD: 2 ppb


Mycotoxins By Fluorometer

-ZearalaTest IA Colum for Corn, Milo and 17% Protein Poultry Feed

- 15-20 Minutes Test Time

- Range: 0-2 parts per million (ppm)

- LOD: 0.1 ppm

-Afla M1 FL+ for Liquid Milk

- 25 Min to Purify Sample

- LOD 12.5 ppt

- Range: 0 – 200 ppt

- Afla B in Corn, Grains and Feeds

- 10-15 Minutes Test Time

- LOD 1 ppb

- Range: 0 – 300 ppb


VICAM Fluorometer Method Approvals

AOAC International (Association of Official Analytical Communities)

USDA-GIPSA (United States Department of Agriculture- Grain Inspection Packers and Stockyard administration) Certificate of Conformance

Government Approvals and Collaborations across 100 countries


Immunoaffinity Column w/Fluorometer

USDA-GIPSA Certified Official Methods

Accepted methods for interstate and exported grain, peanut and tree nut shipments

Methods for total aflatoxins (B1 + B2 + G1 + G2), fumonisins (B1 + B2 + B3), ochratoxin A and zearalenone

Excellent correlation with LC methods

Rapid test times (6-20 minutes)

Simple to perform

Field and laboratory ready


Immunoaffinity Column Sample Cleanup

Offering immunoaffinity columns used with both benchtop Fluorometerand/or LC system to determine mycotoxin levels to ensure food safety.


Thank you – Questions?

rapid mycotoxin analysis for the field waters technology … · 2017. 8. 7. · ©2017 waters...

Documents