Alkaloids in food and feed - Conffidence · Alkaloids in food and feed: ... atropine, caffeine,...

24
11/11/2010 1 Alkaloids in food and feed: an “emerging risk” ? Hans van Egmond & Hester van den Top, 22 April 2010

Transcript of Alkaloids in food and feed - Conffidence · Alkaloids in food and feed: ... atropine, caffeine,...

Page 1: Alkaloids in food and feed - Conffidence · Alkaloids in food and feed: ... atropine, caffeine, solanine, nicotine, cocaine ... “Camel belly” in Uzbekistan, ...

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Alkaloids in food and feed:

an “emerging risk” ?

Hans van Egmond & Hester van den Top, 22 April 2010

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de bladeren het giftige klein kruiskruid kan zitten. De discounter Plus maakte

woensdag bekend uit voorzorg de slasoort voorlopig nergens meer te verkopen.

Plus reageerde op een waarschuwing van een Duitse wetenschapper.

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Alkaloids:

Naturally occurring chemical compounds

containing basic nitrogen atoms

Produced by various organisms, mainly plants

Often physiologically active and poisonous

Over 3000 compounds known, including

atropine, caffeine, solanine, nicotine, cocaine

Introduction

EFSA opinions and EU legislation

PA in food: the Afghanistan drama

PA in feed: the ragwort issue

EU - FP7 project CONffIDENCE

Summary and conclusions

Outline of presentation

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Introduction

EFSA opinions and EU legislation

PA in food: the Afghanistan drama

PA in feed: the ragwort issue

EU - FP7 project CONffIDENCE

Summary and conclusions

Outline of presentation

Opinions and limits

Ergot alkaloids

- EFSA opinion for feed, 2005

- EU limits: 0.05 % for ergot in intervention wheat;

0.1% for feedstuffs containing unground cereals

Pyrrolizidine and tropane alkaloids

- EFSA opinions for feed, 2007 and 2008

- EU legislation in feed: ranges from 0.01-0.3 % for

botanical impurities (weed seeds and unground

and uncrushed fruits containing alkaloids)

- EU limits: not (yet) developed

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Ergot alkaloids

Mycotoxins, formed by Clavicipitaceae

Ergot occurs in grasses, grains, sorghum

Adverse effects: convulsions, gangrene

Animal sensitivity: poisoning outbreaks in livestock

Methods of analysis: LC-FLD and LC-MS/MS, but

not (yet) interlaboratory validated

Limited data on carry-over do not point at animal

products as an important source of exposure

Tropane alkaloids

Plant toxins, formed primarily by Solanaceae

Seeds may contaminate plants, e.g. soybean

Humans: accidental exposure rare

Animals: pigs very sensitive (Datura poisoning)

Methods of analysis: HPLC, GC, RIA, CE-MS,

LC-MS/MS, but not (yet) interlaboratory validated

No info on carry-over, traces of scopolamine

found in eggs, no further data on residues

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Pyrrolizidine alkaloids

Toxins formed in common plants, e.g. Senecio spp.

These plants may contaminate food or feed

Re-intro of certain species in nature may increase intoxications in wildlife and grazing animals

Adverse effects in humans and livestock

Methods of analysis: GC-MS, LC-MS/MS, but not (yet) interlaboratory validated

Old data on carry-over point at low amounts of alkaloids in milk, significance unclear

Pyrrolizidine alkaloids (PA)

Increased interest for PA (EU workshop held)

Occurring in 3% of world’s plant species- protect plants against herbivores

More than 350 different components:- liver toxic - liver carcinogenic- mutagenic- treatment not well defined

Intake limit (RIVM): 30 ng/day (carc. riddelliine)

Acute clinical symptoms (WHO): 1 mg/day

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Introduction

EFSA opinions and EU legislation

PA in food: the Afghanistan drama

PA in feed: the ragwort issue

EU - FP7 project CONffIDENCE

Summary and conclusions

Outline of presentation

Outbreak of liver disease in Hirat district

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Hepatic veno-occlusive disease in history

“Bread poisoning” in South Africa in 1920

“Camel belly” in Uzbekistan, 1931 and 1945

“Gulran disease” in Afghanistan, 1974 and 1999

“Gulran disease” was attributed to consumption of bread, made from wheat contaminatedwith charmac

Charmac (Heliotropium sp.)contains pyrrolizidine alkaloids (PA), primarily heliotrine

Gulran district

Approx. 150 km2, approx.

110.000 inhabitants

Remote villages, undulating hills, scanty

vegetation, serving as pasture lands

Inhabitants mostly wheat farmers

who may keep sheep and goats

Diet consists mainly of wheat

bread, occasionally meat

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Outbreak of liver disease in Hirat district

More than 270 people affected

Approx. 50 people died

WHO alarmed, and RIVMcontracted to investigatethe cause of the disease

Hypothesis: PA poisoning

Samples sent to RIVM

Methodology: LC-MS/MS

Suspect: local weed (charmac)

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Results: charmac

Pyrrolizidine alkaloids in charmac

0

200

400

600

800

1000

1200

stem/leaf stem/leaf root seed

mg

/kg

Investigated materials

WeedFlour

Qurut Milk

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Not all samples well-packed

Not all samples well-packed

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Results: wheat flour

Pyrrolizidine alkaloids in flour

0

5

10

15

20

25

30

35

40

45

50

C S C S C S S S C S S S S S C S C

mg

/kg

Results: qurut

Pyrrolizidine alkaloids in qurut

0

100

200

300

400

500

600

700

S S C C S C C C C C S S S

µg

/kg

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Results: milk

Pyrrolizidine alkaloids in milk

0

0,5

1

1,5

2

2,5

3

3,5

Goat 1 Goat 2 Goat 3

µg

/kg

Conclusions of the investigations

High concentrations of PA found in charmac

Same PA were found in wheat flour

In dairy products PA were also found; qurut contained trichodesmine, not present in charmac

Prolonged, regular exposure to contaminated wheat, in combination with low protein diet, likely to be the cause of the disease

Publication about the Hirat poisoning episodein press in Journal of Toxicology

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Introduction

EFSA opinions and EU legislation

PA in food: the Afghanistan drama

PA in feed: the ragwort issue

EU - FP7 project CONffIDENCE

Summary and conclusions

Outline of presentation

Pilot carry-over study ragwort

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Background

Senecio jacobeae: PA

containing weed, occurring

as impurities in hay

Other possible contaminants:

S.vulgaris, S. inaequidens

Consumption by dairy cattle

could lead to PA into milk

Very limited data exist about

carry-over experiment

Carry-over experiment: collection of material

Ragwort (Senecio jacobaea)

Full grown plants near Wageningen Period: June-July 2008 Complete plants air-dried and

homogenised 12.5 kg

Narrow-leaved ragwort (Senecio inaequidens)

Ditto, Ede (3 places) 2.5 kg

Materials of both samples mixed

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Composition feeding material (RIKILT)

0%

5%

10%

15%

20%

25%S

n

Sn

-ox Ir

Ir-o

x

Rt

Rt-

ox

Us

Us-

ox

Rd

Rd

-ox

Sp

Sp

-ox St

St-

ox

AcS

p

AcS

p-o

x

Jb

Jb-o

x Jl

Jl-o

x

Jn

Jn-o

x Jz

DH

Jn Er

Er-

ox

AcE

r

AcE

r-o

x

Sv

Sv-

ox

Sk Ot

On

De

sDo

r

Fs

Fd

Total: 2.34 mg/g (82% N-oxides; 17% tertiary bases)

jacoline

Pilot carry-over study: experimental set-up

Waaiboer farm (ASG, Lelystad) 3 cows (age: > 2 jr), weight: 600-700 kg

exposure: September-October 2008

dosing of ragwort via fistula, directly after milking

Exposure periode: 5 weeks Week 1: clean feed

Week 2: 25 g, twice daily

Week 3: 50 g, twice daily

Week 4: 100 g, twice daily

Week 5: clean feed (“wash out”)

Sampling scheme: milk, twice daily, 2 x 40 ml

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Results: daily milk production (ASG)

0

10

20

30

40

50

60

14-S

ep-0

8

16-S

ep-0

8

18-S

ep-0

8

20-S

ep-0

8

22-S

ep-0

8

24-S

ep-0

8

26-S

ep-0

8

28-S

ep-0

8

30-S

ep-0

8

02-O

ct-0

8

04-O

ct-0

8

06-O

ct-0

8

08-O

ct-0

8

10-O

ct-0

8

12-O

ct-0

8

14-O

ct-0

8

16-O

ct-0

8

18-O

ct-0

8

lite

r

3484

3616

3685

indigestion

Results: total PA in milk (RIVM)

Cow evening milk

0.00

2.00

4.00

6.00

8.00

10.00

12.00

14.00

16.00

15-9

-200

8

17-9

-200

8

19-9

-200

8

21-9

-200

8

23-9

-200

8

25-9

-200

8

27-9

-200

8

29-9

-200

8

1-10

-200

8

3-10

-200

8

5-10

-200

8

7-10

-200

8

9-10

-200

8

11-1

0-20

08

13-1

0-20

08

15-1

0-20

08

17-1

0-20

08

19-1

0-20

08

date

ng

/ml S

um

PA

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Jacoline in milk (RIVM)

cow number 3685 morning and evening

0

2

4

6

8

10

12

6 ok

t 200

8 oc

h

6 ok

t 200

8 av

7 ok

t 200

8 oc

h

7 ok

t 200

8 av

8 ok

t 200

8 oc

h

8 ok

t 200

8 av

9 ok

t 200

8 oc

h

9 ok

t 200

8 av

10 o

kt 2

008 o

ch

10 o

kt 2

008 a

v

11 o

kt 2

008 o

ch

11 o

kt 2

008 a

v

12 o

kt 2

008 o

ch

12 o

kt 2

008 a

v

13 o

kt 2

008 o

ch

13 o

kt 2

008 a

v

date

ng

/ml

0%

5%

10%

15%

20%

25%

Sn

Sn

-ox Ir

Ir-o

x Rt

Rt-

ox

Us

Us-

ox

Rd

Rd

-ox

Sp

Sp

-ox St

St-

ox

AcS

p

AcS

p-o

x

Jb

Jb-o

x Jl

Jl-o

x

Jn

Jn-o

x Jz

DH

Jn Er

Er-

ox

AcE

r

AcE

r-o

x

Sv

Sv-

ox

Sk Ot

On

De

sDo

r

Fs

Fd

jacoline

Succesful transfer experiment

Milk: only tertiairy bases found, no N-oxides

Senkirkine, jacoline, otosenine, jaconine,

florosenine and jacobine found

Jacoline: 81 % of PA in milk, only 1% in weed!

Jacoline transfer: 6-8 % of ingested mount

Conclusions carry-over experiment

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Results: risk assessment (RIVM)

Human intake limit: 30 ng/day

Daily milk consumption: 1 l

Allowed milk concentration: 0.03 ng/ml

Jacoline →milk transfer factor: 0.0021

Allowed jacoline intake of cow: 14.3 μg

Jacoline concentration in weed: 21.8 μg/g

Allowed daily weed intake of cow: 0.66 g/day

Assumptions:- ridelliine intake limit applies to jacoline as well- jacoline is stable upon pasteurisation

Further investigations

Second carry-over study to confirm findings

Better insight in mass balance needed

Production of several dairy products planned

Distribution and stability of PA in dairy products

Analysis of commercial milk products planned

Surveys of animal feeds recommended

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Introduction

EFSA opinions and EU legislation

PA in food: the Afghanistan drama

PA in feed: the ragwort issue

EU - FP7 project CONffIDENCE

Summary and conclusions

Outline of presentation

About CONffIDENCE

LCP Food, Agriculture and Fisheries,and Biotechnology, 2008-2012

Simple, fast, multi-analyte,multi-class detection

Training workshops andeducating modules

Includes WP Biotoxins

alkaloids (pyrrolizidine, tropane, ergot)

Fusarium mycotoxins

phycotoxins

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The alkaloids work package

Alkaloids

PA in honey and feed

TA in feed

EA in feed and cereals

Methods

Multiplex dipsticks

NIR (in line)

Status

Prototypes in development

Multiplex dipsticks for PA, TA and EA alkaloids

jacobine

lycopsamine

CTRL

PA

ergotamine

ergocristine

CTRL

EA

scopolamine

atropine

CTRL

TA

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Conveyor belt

NIR cameraLight source

NIR imaging method to detect ergot

Introduction

EFSA opinions and EU legislation

PA in food: the Afghanistan drama

PA in feed: the ragwort issue

EU - FP7 project CONffIDENCE

Summary and conclusions

Outline of presentation

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Summary and conclusions

Many alkaloids exist, particularly in plants

Alkaloid poisonings: recurring events

EFSA opinions: limits better set for toxins; methods of analysis to be validated; RMs to be developed; data on carry-over to animal products scarce

Small amounts of PA in feed could lead to unacceptable amounts in milk; needs confirmation

CONffIDENCE: a project contributing to the development of rapid tests for alkaloids

Alkaloids in food and feed:

an “emerging risk”

With thanks to Ron Hoogenboom, Patrick Mulder, Gerrit Remmelink,

Ronald Schothorst, Marco Zeilmaker and .... the audience