COPY NOT - Sorghum in the 21st Century...2018/06/14 · VICAM History 1995 Multiple mycotoxin IA columns launched for LC and LC/MS/MS detection 1985 AflaTest ships 1987 2006 Over

©2018 Waters Corporation 1COMPANY CONFIDENTIAL

Monitoring Natural Toxins and Pesticide Residue in the Supply Chain

Technology and Tools that assist in complying to current regulations

Mr. Willem Joubert Chemistry Specialist Waters Chromatography and Mass Spectrometry Division Microsep (Pty) Ltd

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Mycotoxins

A mycotoxin (from the Greek mykes, "fungus" and toxikon, "poison") is a toxic secondary metabolite produced by organisms of the fungus kingdom and is capable of causing disease and death in both humans and other animals. The term 'mycotoxin' is usually reserved for the toxic chemical products produced by fungi that readily colonize crops

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Mycotoxins in history

'St Anthony's fire' was one of the earliest recognised diseases caused by mycotoxins in Europe – Well known by the end of the first millennium. – The result of eating rye contaminated with ergot alkaloids produced by the mould Claviceps purpurea. – Epileptic fits and excruciating burning sensation experienced led pilgrims to the shrine of St Anthony in France in hope of

a cure.

Alimentary Toxic Aleukia in Russia during World War 2 (T-2) Mycotoxins came to prominence in the UK in 1960 (Turkey-X)

– 100 000 turkey and other poultry died just before Christmas – Traced to a toxic contaminant later called aflatoxin present in the groundnut meal forming part of their diet.

This incident illustrated the potential threat to the food chain posed by mycotoxins – Ushered in the modern era of mycotoxin studies.– Mycotoxins were subsequently linked to many historical cases of human and animal deaths and illnesses.

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Mycotoxins in history - continued

"poison in the peanuts“ - Primary Schools Nutrition Programme (PSNP) – South Africa May 2001 (Aflatoxin)

2003 Kenya: acute poisoning, 120 confirmed deaths (Aflatoxin) Dog food aflatoxin outbreak 2006 – USA (Aflatoxin) February–March 2013: Romania, Serbia, Croatia imported into western Europe February 2013: Iowa contamination. 2014 (ongoing): Nepal and Bangladesh, neonatal exposures, found in umbilical cord blood. Dog food recall June 2014 – South Africa (DON)

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Worldwide regulation of Mycotoxins

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Most Common Mycotoxins regulated in Human Foods Worldwide

Mycotoxin Number of countries

Total Aflatoxin 76Aflatoxin B1 61Aflatoxin M1 60Patulin 48Ochratoxin A 37Deoxynivalenol 37Zearalenone 17Fumonisin 6

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FOODSTUFFS, COSMETICS AND DISINFECTANTS ACT, 1972 (ACT 54 OF 1972), Government Notice 1145 (8 October 2004) and 987 (5 September 2016) Restrictions: For the purposes of section 2(1)(b)(i) of the Act, in so far as it is applied to and is applicable to foodstuffs, the following foodstuffs are hereby deemed

to be contaminated, impure or decayed A: Peanuts intended for further processing, which contain more than 15 µg/kg of aflatoxin (total); a. B: All foodstuffs, ready for human consumption, which contain more than 10 µg/kg of aflatoxin, of which aflatoxin B1 is more than 5 µg/kg; b. C: Milk containing more than 0.05 µg/l of aflatoxin M1; D: Wheat, rye, barley and oats which contain more than 0.02% (m/m) of Ergot sclerotia; E: Apple juices and apple juice ingredients in other beverages containing more than 50 µg/l of patulin. F: Cereal grains (wheat, maize and barley) intended for further processing, which contain more than 2000pg /kg of Deoxynivalenol; G: Flour, meal, semolina and flakes derived from wheat, maize or barley, ready for human consumption, which contain more than 1000pg /kg

of Deoxynivalenol; H: Raw maize grain, intended for further processing, which contain more than 4000pg /kg of Fumonisins (B1 + B2), the whole commodity. I: Maize flour and maize meal, ready for human consumption, which contain more than 2000pg /kg of Fumonisins (B1 + B2), the whole

commodity.

Enforcement The sampling plan for the analysis of total aflatoxins in peanuts intended for further processing to be used for enforcement and control in terms of

these regulations shall be in accordance with the provisions laid down by the Joint Food and Agricultural Organization/World Health Organization (FAO/WHO) Food Standards Programme’s Codex Alimentarius Commission, i.e. Sampling Plan for Aflatoxins in Peanuts Intended for Further Processing (CODEX STAN 209 of 1999 as revised in 2001).

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Companies Doing Testing Food Testing (68%) Feed Testing (32%)

Market Segments:Food or Feed Companies

Number of Tests Performed per sector Food Testing (Africa 17%, S. Africa 22%) Feed Testing (Africa 14%, S. Africa 47%)

Food68%

Feed32%

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VICAM History

1995Multiple

mycotoxinIA columns launched

for LC and LC/MS/MS detection

1985

AflaTestships 1987

2006

Over 60% market share in

mycotoxindiagnostics

.

2010Vertu

mycotoxinstrip test

range expanded

2018

M1V, OchraV, ZearalaV,

FGIS-GIPSA approvals,

more commodities,

and more than

mycotoxins

Collaboration between

Harvard and MIT scientists with business

leaders creates VICAM

VICAM acquired by Waters

Corporation

Vertu quantitative lateral flow test strips launched

…and beyond

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Some Africa CompaniesSupport In Africa

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Points of Mycotoxin Monitoring

Harvest• Screening• Confirmation

Storage• Segregation• QC Before

Processing

Finished Products• Laboratory

QC• Prior to Export

or Import

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Where Is Testing Performed?

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Considerations when choosing a testing strategy

Time of Analysis

Labor -Training required- appropriate level of education

Cost - Capital investment and consumables

Testing environment- laboratory or in the field

Is an Official Method required?

Precision, accuracy and limit of detection needed for the method

Information content – qualitative or quantitative, total toxin vs individual toxins

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Vertical Markets - Mycotoxin Testing

ReferenceLab

Govt/Export & Import

Harvest Storage/Handling

Processing Shipment

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Analytical requirements for mycotoxins & volume of testing

Research and metabolomic based analysisAcademic, government, commercial laboratoriesFinished food products, biological samples, speciation, conjugated metabolites

Quantitative laboratory based testing, Commercial & official control laboratories finished foods & feeds, complex matrices,multi-toxin analysis

High volume testing - field tests, short turnaround times

Analysis conducted at receiving points, transit points, processors sites, commercial labs

HR-MS

LC-UV / LC -FLR and LC-MS/MS (QqQ)

Rapid Test kitsFluorometer, Lateral Flow,

Strips

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Good Best

Vertu™

DON-V™

Afla-V™

Fumo-V™

Best With Sophisticated Labs

HPLC/UPLC

LC/MS/MS

ImmunoaffinityPlatform with Series 4 EX Fluorometer

VICAM Solutions - Positioning

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AflaCheck / DonCheck

–Qualitative Strip Test–Yes / No Answer

Afla-V / Afla-V AQUA / DON-V / Fumo-V with Vertu Lateral Flow

–Simple, easy to use, low cost quantitation–Field Testing Alfa Markets–DON markets (Flour milling, pet food, feed)–Better user interface for low end–Small footprint

GOOD - BETTER

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Vicam Immunoaffinity Platform with Series 4 EX Fluorometer

BENEFITS– Convenient - For use with fluorometer or HPLC– Comprehensive - Total readings for all aflatoxins– Durable - Long shelf life; requires no refrigeration– Versatile - For use with a variety of samples– Quick - Less than 10 minutes to isolate toxin*– Wide Range - Detects levels as high as 300 ppb and as low as 0.1 ppb**– Safe - Requires less toxic materials than other methods

Fluorometeric TestsAflaTestAfla BAfla M1 FL+FumoniTestFumoniTest 200OchraTestZearalaTest

BEST

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HR-MS



Strips

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Vicam Immunoaffinity Platform with HPLC / UPLC / LC-MS/MS

BENEFITS

–Exclusive – Specifically for HPLC/UPLC use–Convenient – Uses same methods as other VICAM tests–Comprehensive – Detects Mycotoxins using reference

methods–Durable – Long shelf life; requires no refrigeration–Versatile – Can be used with a variety of samples–Wide Range – Low Detection Limits

BEST with Sophisticated Lab’s

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AOAC Official methods

Official Method Mycotoxin Foodstuff991.31 Aflatoxins Corn, Raw Peanuts, and

Peanut Butter999.07 Aflatoxin B1 and Total

AflatoxinsPeanut Butter, Pistachio Paste, Fig Paste, and Paprika Powder

2000.03 Ochratoxin A Barley

2000.08 Aflatoxin M1 Liquid Milk

2000.09 Ochratoxin A Roasted Coffee

2000.16 Aflatoxin B1 Baby Food

2001.04 Fumonisin Corn and Cornflakes

2001.01 Ochratoxin Wine and Beer

2003.02 Aflatoxin B1 Cattle Feed

2004.10 Ochratoxin Green Coffee

2008.02 Aflatoxins and Ochratoxin Ginseng and Ginger

2013.05 Aflatoxins Olive Oil, Peanut Oil, and Sesame Oil

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CEN Official methodsOfficial Method Mycotoxin Foodstuff

EN12955 aflatoxin cereals, shell-fruits and derived products

EN14123 aflatoxin hazelnuts, peanuts, pistachios, figs, and paprika powder

EN14132 ochratoxin A barley and roasted coffee

EN14133 ochratoxin A wine and beer

EN14352 fumonisin maize based foods

EN14501 aflatoxin milk and milk powder

EN15791 deoxynivalenol animal feeding stuffs

EN15792 zearalenone animal feeding stuffs

EN15835 ochratoxin A cereal based foods for infants and young children

EN15839 ochratoxin A currants, raisins, sultanas, mixed dried fruit and dried figs

EN15850 zearalenone maize based baby food, barley flour, maize flour, polenta, wheat flour and cereal based foods for infants and young children

EN15851 aflatoxin B1 cereal based foods for infants and young children

EN15891 deoxynivalenol cereals, cereal products and cereal based foods for infants and young children

EN15892 ochratoxin A currants, raisins, sultanas, mixed dried fruit and dried

EN16187 fumonisin maize containing foods for infants and young children

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HR-MS



Strips

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LC-MS (Liquid chromatography- Mass spectrometry)

Advantage of MS- It can detect molecules that do not fluoresce or absorb

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Instrument can be used for other analysis besides mycotoxins (such as pesticides)

Good for multiple mycotoxin analysis Good for confirmation of mycotoxins

Must have a laboratory environment Triple Quad (MS/MS) gives best results (lower limits

of detection and confirmation) Best for well trained scientists Need to accurately measure and store mycotoxins

in order to get accurate results Need to use matrix matched calibration standards or

internal calibrators to adjust for matrix affects

Characteristics of Mass Spectrometry

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Total Ion Chromatogram

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Fluorometeric TestsAflaTestAfla BAfla M1 FL+FumoniTestFumoniTest 200OchraTestZearalaTest

• AflaTest• AflaTest WB• Afla WB SR• Afla M1 HPLC• AflaOchra HPLC• AOZ HPLC• Myco6in1• CitriTest HPLC• DONtest• DONtest WB HPLC• DON-NIV WB• FumoniTest• FumoniTest WB

HPLC/UPLC/LC/MS Tests using IAC

Aflatoxins, DON, NIV, T-2, HT-2, OTA, fumonisins, zearalenone

Myco 6in1+ IAC

Qualitative StripsAflaCheckDONCheck

Quantitative StripsAfla-V AflaM1-VDON-VFumo-V

http://vicam.com/products

VICAM technology address all the markets

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http://vicam.com/products


Multi residue pesticides

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Grape

Avocado

Marjoram

Ginger

Ultra Trace Analysis: Sample Complexity

Xevo TQ-S

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Acetonitrile– QuEChERS (includes salting out)

Ethyl acetate– SweEt

Acetone– Mini-Luke or NL (extracted with acetone followed of

partition with dichlorometane / petroleum ether (1:1)

AOAC Official Method 2007.01 and CEN Method EN 15662

Extraction for multi-residue analysis

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Overlay UPLC-MS/MS Chromatogram Post-Spiked at 10 ppb

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Incurred Residues by UPLC-MS/MSName Retention

Time MatchIon Ratio

MatchIncurred

Residue (mg/kg) EU MRLs (mg/kg)

Boscalid 0.006 5

Buprofezin 0.001 1

Fenhexamid 0.036 15

Imidacloprid 0.012 1

Methoxyfenozide 0.018 1

Pyraclostrobin 0.001 1

Trifloxystrobin 0.001 3

Incurred residues detected are well below the EU MRLs

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Ion Source Versatility– Universal source architecture: Enables the widest range of ionisation techniques to be

utilised

– Simple and quick: Allowing you to change between ion source options in minutes

Universal Source Options:UPLC to APGC

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Atmospheric Pressure Gas Chromatography

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Comparison of mass spectra for endrin

APGCEI

Which ion to select as precursor ion for MS/MS?

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Overlay APGC-MS/MS Chromatogram post-spiked at 10 ppb

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Incurred Residues by APGC-MS/MS

Name Retention Time Match

Ion RatioMatch

IncurredResidue (mg/kg)

EU MRLs (mg/kg)

Pyriproxyfen 0.0009 0.05

Tetrahydrophthalimide 0.0003 0.02

2-Phenylphenol 0.0007 0.05

Pyrimethanil 0.008 5

Anthraquinone 0.0004 0.01

Incurred residues detected are well below the EU MRLs

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Improved efficiencies for LC and GC amenable multi-residue pesticides analysis on a single instrument platform

Innovative workflow strategies for MS quantification and to maximizelaboratory efficiency

APGC-MS/MS

UPLC-MS/MS

Summary

− IntelliStart− QuanPedia− TargetLynx XS

− TrendPlot− RADAR− PICsDO N

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Q-IMS-Tof

TQ-S micro

TQD

SQ

Tof

TQ-XS

QTofIMS-QTof

Diverse MS Needs

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Thank you for your Attention!!

Thank you for your attention

[email protected]

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Documents

COPY NOT - Sorghum in the 21st Century...2018/06/14 · VICAM History 1995 Multiple mycotoxin IA columns launched for LC and LC/MS/MS detection 1985 AflaTest ships 1987 2006 Over