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THE

POISONOUS SUBSTANCES

(PYRROLIZIDINE ALKALOIDS, PAs)

FOUND IN FIREWEED

Food safety implications

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In this presentation I will be summarising what is known about the poisonous

substances (pyrrolizidine alkaloids, PAs) present in fireweed and how they endanger

the health and productivity of livestock and also pose a threat to the health of

humans. I will be presenting factual information from the literature. Any “opinions”

expressed are my own and not those of any organisations I am associated with.

Here are a few of the plants found in Australian agricultural systems that contain PAs.

Senecio madagascariensis

Fireweeds, species of the genus Senecio, including S. madagascariensis shown

here, are prominent among them.

All regions of the world, from high mountains to sea level, have species of

fireweeds/Senecio. One important factor contributing to their success in many

environments is their ability to produce poisonous PAs that protect them against

many herbivores. For example, there are only a few insects, with a very specific

tolerance for PAs, that can eat these plants.

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Other plants containing PAs include Paterson’s curse/salvation jane (Echium

plantagineum).

Common heliotrope/ potato weed (Heliotropium europaeum).

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Echium vulgare (called blue borage in New Zealand) is another PA species (found

around Cooma),

and another is Amsinkia

.

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It has been estimated that 3% of all flowering plants contain PAs and it has been

shown that pyrrolizidine alkaloids have independently evolved on at least 4 occasions

in different plant families. This demonstrates, from an evolutionary perspective, the

remarkable protection that PAs afford plants that have the ability to make them. All

mammals are poisoned by them and, as I have already said, only certain insects that

have evolved specific mechanisms to avoid poisoning can eat them.

Heliotrine Monocrotaline

Lasiocarpine

Senecionine

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Let me assure all non-chemists that this is the only slide showing chemical

structures. PAs, such as these are a well-established cause of liver failure in both

livestock and humans. The structural features necessary for liver damage and

several other adverse health effects are the bicyclic ring system from which the name

pyrrolizidine is derived, a double bond opposite the nitrogen and ester groups as

shown. This latter part of the structure is highly variable as you can see and is the

reason why there are several hundred PAs known from a wide range of plants – all of

which are poisonous.

[This also one reason that makes them hard to analyse since the analyst must be

able to recognise several hundred different PAs from their mass spectra and nuclear

magnetic resonances and ideally have authentic standards for comparison and this

takes many years of experience and practical guidance from people with experience.

There are also many pitfalls for beginners in isolating PAs for analysis.]

Fireweeds are a major source of a particularly hazardous group of “macrocyclic” PAs.

Senecionine, shown here, is the “macrocyclic” PA on which many of the poisonous

PAs found in fireweeds are derived. I have a sample with me that I will pass around.

Please do not open it.

A great deal is known about the toxicity and chemistry of PAs and how to detect and

quantify them in plants and food items however this knowledge is at present

restricted to a few laboratories worldwide. I will be giving only a brief summary of this

information today. More information can be obtained from the inter-net sites

mentioned at the end of this paper).

Research at CSIRO Livestock Industries, where I worked for many years, has, since

the 1950s and before, made major contributions to knowledge of the toxicity,

chemical structure, and analysis of PAs. Unfortunately much of Australia’s expertise

on PAs has been lost in recent years as people have retired and the last vestige of

expertise in Australia is about to disappear in July this year when the group at CSIRO

Livestock Industries, under Steve Colegate, will be disbanded. CSIRO/Australian

interest in PAs has waned as international concern is growing.

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Mechanism of action

Before describing and giving examples of the diseases caused by PAs, I will briefly

describe their mechanism of toxicity.

PAs are not toxic per se. They are converted into toxic substances by cytochrome

P450 enzymes in the liver. This is why the damage caused by PAs is largely confined

to the liver.

Activation of PAs in the liver

DNA

PA

DNA

DNA

Liver enzymes

PA PA* PA

Cytochromes P450

Protein

Protein

PA

Protein

Activated PA* attaches to DNA and proteins

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It is ironic that the enzymes in the liver that activate PAs are normally involved in

detoxifying harmful substances but in the case of pyrrolizidine alkaloids their action

converts PAs into extremely harmful, chemically reactive substances that

immediately form chemical bonds with liver constituents including DNA and proteins.

From a chemical perspective, the formation of these harmful PA metabolites is

equivalent to injecting mustard gas of the First World War directly into the liver (both

activated PAs and mustard gas are biological alkylating agents).

The activated PAs produced in the liver cells immediately react with genetic material

(that is DNA), and proteins in the liver cells. In fact a portion of the PA molecule is left

attached to liver DNA and proteins. The toxic PA metabolites have two chemically

reactive centres and can form cross-links between strands of DNA. Protein-PA-DNA

cross-links are also generated. As you can imagine, this is very disruptive to the

functioning and viability of the liver.

Pyrrolizidine alkaloids cause:

• Liver damage (hepatotoxins)

• Cancer (carcinogenic)

• Mutations (mutagenic, genotoxic)

• Fetal abnormalities (teratogenic, fetotoxic)

• Lung changes, leading to

• Right heart congestive failure (cor pulmonale)

The DNA-damaging/gene damaging properties of PAs provide a basis for some of

the effects of PAs including the abnormalities they produce in fetuses (teratogenic)

and their cancer causing (carcinogenic) properties. PAs fall into a category of

poisonous substances referred to as “genotoxic carcinogens”, that is direct gene

damaging, cancer causing chemicals.

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PAs are “genotoxic carcinogens”

Unlike many other poisonous substances, e.g. strychnine, they have no threshold of

toxicity

I mention this terminology to indicate that they are different from many other toxins,

strychnine, for example. As you all know strychnine is considered very poisonous but

to be harmful it must exceed a particular level called the lethal dose. If only half a

lethal dose of strychnine is consumed it produces no poisonous effects. Strychnine is

also readily cleared from the body and no damage accumulates. This means that you

can eat half a lethal dose of strychnine every day forever and not be poisoned

(although I do not recommend it).

Genotoxic carcinogens are given a

“Provisional Tolerable Daily Intake”

“Safe” daily intakes can be determined in the case of strychnine and, for example, all

agricultural chemicals that have a “no observable adverse effect” threshold level.

Strychnine and many other poisons have what is called a threshold of poisoning. As

long as this is not exceeded you are safe so that a safe daily intake/dose can be

determined for strychnine and for agricultural chemicals for example. On the other

hand, PAs, and other genotoxic carcinogens, in theory at least, have no safe level.

Genotoxic carcinogens, such as PAs, are given what is referred to as a maximum

provisional tolerable daily intake/dose, as opposed to a safe level.

This is why a Safety Guide published by the World Health Organisation states that:

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“ Prevention of exposure is the only effective method of limiting toxicity due to

PAs. Even low doses over a period of time may present a health risk and

exposure should be avoided or minimized as far as possible.”

World Health Organization (1989).

PAs can have different effects depending on the amount of PAs livestock or people

are exposed to in their diet. These can be referred to as acute, sub-acute and chronic

poisoning and can be considered as time and dose-related stages of poisoning.

Dose-related, progressive PA poisoning

Acute sub-acute chronic

(a lot of PA) (A medium amount ( A small amount

of PA.) of PA, intermittently)

The term “acute” means that the amount consumed is sufficiently large over a short

period of time that clinical poisoning is seen within hours, days or weeks after

exposure. In “chronic” poisoning the clinical disease develops imperceptibly over

months or years and the source of poisoning is therefore not easily recognised.

“Chronic” toxicity is more likely in countries such as Australia where quality

assurance measures are in place that reduce exposure to PAs, at least for some

commodities such as wheat, and, provided that these measures are adhered to, the

levels of PAs in major commodities are likely to be very low so that “acute” PA toxicity

of people is very unlikely in Australia.

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Acute PA poisoning

• Liver enlargement (hepatomegaly)

• Fluid buildup in peritoneal cavity (Ascites)

• Liver tissue around veins disintegrates with bleeding (haemorrhagic

necrosis)

• Death or

• Progression to sub-acute poisoning

In acute PA-poisoning, the cells of the liver around the veins (the site where PAs are

activated) are the immediate site of damaged leading to disintegration of the tissue

and bleeding to produce a condition called haemorrhagic necrosis. This may be the

only abnormality seen in the liver before death occurs.

Sub-acute PA poisoning

• Veins of liver become constricted (veno-occlusive disease)

• Poor blood flow

• Pooling of blood behind vein constrictions

• Breakdown (necrosis) of surrounding liver tissue

More commonly, where poisoning develops over a longer period the effect is less

catastrophic, and physical changes to the veins in the liver can be seen. The veins

become constricted or blocked, leading to poor blood flow, pooling of the blood

behind the constriction, breakdown of the surrounding liver tissue and eventually

cirrhosis indistinguishable in the end from that caused by other things such as over

consumption of alcohol or hepatitis viruses for example.

If exposure to PAs is very low and intermittent the liver experiences intermittent

periods of damage and regeneration. Nodular regeneration may be patchy

throughout the liver and is accompanied by “fibrosis”. The result is a poorly

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functioning liver and, in many cases, eventually leads to cirrhosis and liver failure

without any prior indication of ill health.

Chronic PA poisoning

• Due to long term, low level intake of PAs or

• Final consequence of earlier acute/subacute poisoning or

• Intermittent periods of sub-acute damage and regeneration leading to -

• Nodular regeneration and fibrosis and eventually to -

• Cirrhosis and liver failure, indistinguishable for other causes (alcohol,

hepatitis), and to congestive heart failure, also difficult to attribute to

PAs.

The liver is not the only tissue that can be damaged and changes can be seen in

other organs. Some of the harmful PA metabolites produced in the liver can escape

from the liver and be carried in the blood to the lungs leading to constriction of the

arteries in the lungs and this leads on to congestive heart failure (cor pulmonale).

Pulmonary Arterial Hypertension

Some PAs cause thickening/narrowing of the arteries in the lungs, leading to

effects on the heart and resulting in right heart congestive failure.

As in the case of cirrhosis of the liver, the medical profession can sometimes not be

sure of what causes congestive heart failure. Exposure to PAs in the diet is rarely, if

ever, considered as a possible cause. GPs are generally unaware of PAs and the

potential they have to cause pulmonary arterial hypertension and, more importantly,

they are also generally unaware that PAs can occur at hazardous levels in some

common foods.

The immediate physical and biochemical damage to the liver and sometimes the

lungs and other tissues does not bring to an end the PAs potential for causing harm.

It has been suggested that at least some of the linkages attaching the PA fragments

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to DNA and proteins in the liver are reversible and, over time, the reservoir of PA

fragments bound to the tissues is released as PA* to attack other sites.

Progressive effects:

Leukaemia

Liver

Cancer Lung

Rhabdomysarcoma

etc

Protein

PA PA*

Protein

Indistinguishable

Cirrhosis from other causes

This may be why, after a single exposure to PAs, there can be a long period of

apparently normal liver function and then a sudden deterioration and the

development of cirrhosis and cancer with no obvious cause. The newly released

“active” PA* is considered by some to be the main cancer-causing entity/agent.

Humans are generally believed to be relatively sensitive to pyrrolizidine alkaloid

poisoning compared to rats, guinea pigs and certainly more susceptible than sheep,

goats and cattle for example. So if these livestock species are being poisoned, the

foods they are producing for human consumption, e.g. milk and meat, is suspect.

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Clinical Disease in Humans

• Humans (esp. foetuses and infants) are considered to be particularly

sensitive to PAs.

• Veno-occlusive disease and an established source of PA exposure (e.g.

a herbal tea) viewed as definitive evidence of PA poisoning.

• Cirrhosis and liver failure after long symptom-less period difficult to

associate with low level of PAs in the diet.

• Cancer and congestive heart failure?

Constriction of the veins (veno-occlusive disease) in the liver is the principle and

diagnostic manifestations in people and is considered highly indicative of PA

poisoning if a dietary source of exposure to PAs can be found. However, if there has

been a long period of low level, intermittent exposure to PAs the constriction of the

veins may be obscured by the liver damage and fibrosis and a possible role for PAs

is unlikely to be considered, especially as the liver damage progresses to cirrhosis.

There have been no confirmed cases of cancer in humans from exposure to PAs

although they are proven to cause a number of different cancers in animals and the

mechanism of carcinogenesis (direct damage to DNA) means that humans must also

be at risk. It has been suggested that DNA repair in humans may be more efficient

than in other animals. Not a great safety net in my opinion.

The next slide shows the typical course and outcomes of the clinical disease from an

episode of “acute” poisoning. In this case, widespread PA-poisoning in the West

Indies in the 1940s and 50s (and probably before and since) was due to widely used,

PA-containing traditional herbal medicines. About 20% of those involved in this

particular poisoning incident died rapidly from acute liver damage. About 50%

recovered completely. Twenty percent showed initial clinical recovery but some later

displayed sub-acute veno-occlusive disease. Thirteen percent went from the acute

phase to the sub-acute directly. Of these, a small number went on to complete

recovery while other, after a latent period, sometimes of years in duration, developed

cirrhosis that was fatal.

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In countries like Australia the primary concern is with low levels of PAs in the diet.

Probably too low to induce acute disease but which progresses, after a latent phase,

to produce cirrhosis and liver failure of undetermined cause in a percentage of

individuals, and maybe also to right heart congestive failure or cancer.

Traditional herbal medicines, e.g.

• Symphytum spp. (Comfrey)

• Arnica

• Borago officinalis (Borage)

• Senecio scandens (Qian liguang)

• Tussilago farfara (Kuan donghua)

• Etc.

These are just a few of the many herbal medicines that contain PAs.

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Incidents of acute PA poisoning, large in some cases and restricted to individuals in

others, have been documented in regions of the world where PA-containing

traditional herbal medicines are in use such as Europe, Africa, Asia the USA and, as

previously mentioned, the Caribbean.

Typical case PA poisoning

The mother of a new born child had consumed a herbal tea, bought at a Swiss

pharmacy as an expectorant, during her pregnancy. The child was admitted to

intensive care 5 days after birth and died of liver failure 33 days later. The

death was attributed to PAs (senecionine) in the tea. The mother was not

affected

Roulet et al., 1988

One recent case of PA poisoning occurred in Freiburg, Germany a few years ago

when I was there.

A baby was delivered prematurely by cesarean section due to fetal ascites and

liver enlargement and died shortly after delivery. Liver histology showed veno-

occlusive disease. A cooking spice used daily during the pregnancy contained

PAs. The amount of spice used per day was about 2 g. The level of PAs found

in 2 grams of spice was about 25 µg.

Rasenack et al., 2002

A woman was using a particular spice in her cooking during the period of her

pregnancy. It was later found to contain small quantities of PAs. It was estimated that

the woman was ingesting about 20 to 30 micrograms (millionth of a gram) per day.

Her newborn child had a damaged liver typical of that caused by PAs and died within

a week.

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In November 2007, WHO reported an acute outbreak of PA poisoning caused by

Heliotrope seeds in flour used to make bread in Herat Province of Afghanistan. By

May 2008, 190 people were affected and 17 had died from liver failure.

http://www.Irinnews.org/report.aspx?ReportId=78218

The only thing standing between us and this are grain receival standards on which I

will say more, later.

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Large-scale episodes of acute PA poisoning of humans have occurred when staple

foods such as bread made from PA-contaminated grain are consumed.

Here are a few examples of large-scale, “acute” and “sub-acute” poisoning involving

bread made from contaminated wheat.

Country Year Number affected

(% deaths)

South Africa 1910-1920s Large numbers

Afghanistan 1999 150 deaths

Tajikistan 1992 >4000 (15%)

Afghanistan 1975 >8000 (high)

India 1978 108 (63%)

Uzbekistan 1960s -

USSR (former) 1939 1500 (15%)

“ 1947 Large numbers

From WHO 1988

“Bread poisoning” was common and endemic in the 1920s and 1930s, in southern

Africa, especially among children and younger people. When the cause was

eventually investigated seeds of several fireweeds (Senecio species) were found in

the wheat. Experiments with animals demonstrated that the PAs in these seeds were

the cause of the liver disease known as “bread poisoning.”

Several other regions of the world have endemic liver disease attributable to PA

contamination of grain. There have been a number of large-scale episodes of

poisoning in the former USSR and the Indian subcontinent that continue to the

present day.

Contamination of grain has been the most important cause of both acute and chronic

PA poisoning of humans.

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While acute PA poisoning incidents have been avoided in countries such as

Australia, because there are grain receival standards specifying the maximum level

of foreign seeds in grain for human consumption, we should not be too complacent.

Seeds allowed in Australian wheat for Human use

• 20 heliotrope seeds*/kg *9 micrograms of PAs in one seed

• 100 Paterson’s curse seeds/kg

• Zero fireweed seeds.

• Grain is “cleaned” of these seeds before milling.

• “Cleaned” grain can however still harbour dust containing PAs

• Amount of wheat contaminated by PA-containing seeds, at levels higher

than the receival standard which is “cleaned” prior to delivery?

The seeds of Heliotropium europium and Echium plantagineum are sometime found

in Australian grain. There is a limit of 20 heliotrope seeds per kg of grain (8

seeds/half litre) if it is destined for human consumption.

Pigs and poultry poisoned by contaminated feed

• In 1992/3 100,000 to 200,000 chickens and 1000 to 4000 pigs were

poisoned by commercial feed.

• The feed contained high levels of PAs believed to have come from the

wheat/wheat screenings component.

In 1994, we investigated a large-scale poisoning incidents in Australia caused by the

grain component being used in commercial pig and poultry feed. Heliotrope seeds

contaminated the wheat or wheat screenings used in the feed. Hundreds of

thousands of poultry were poisoned and several thousands of pigs over a few

months.

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Even when a sample of the contaminated wheat involved was “cleaned” of all

heliotrope seeds the grain was found to be still poisonous to pigs that like humans

are particularly sensitive to PAs. This was attributed to the dust remaining in the grain

that could not be removed by the “cleaning” process.

This, and similar incidents are, I believe, a signal that we need to monitor levels of

PAs in foods for human consumption and not relying on counting the number of

contaminating seeds.

A grossly contaminated grain can be completely “cleaned” of seeds but still contain a

significant level of PAs.

Let me give you another Australian example, this time a smaller incident that involved

a single egg producer in South Australia.

Wheat contaminated by Heliotropium europaeum, purchased from a neighbour, was

suspected of poisoning egg-laying chickens. We examined extracts of the wheat and

found the expected pyrrolizidine alkaloids.

This is the fast atom bombardment mass spectrum of the grain extract showing

typical heliotrope PAs.

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We also examined some of the eggs that had been laid by the chickens during and

after the poisoning and were surprised to find not only the expected alkaloids from

Heliotropium europaeum but also a series of other PAs from Paterson’s curse.

How these came to be present in the eggs was never determined but the clear

implication was that they were in a component of the diet of the chickens.

This is an indication that there is sometimes likely to be PA contamination occurring

in our food without our knowledge.

Foods that may become contaminated by PAs

• Grains (wheat, sorghum, barley etc)

• Milk (including human milk)

• Eggs

• Honey and pollen

These are the main foods that may sometimes be contaminated by PAs and in some

cases, such as honey from fireweed or Paterson’s curse, are always contaminated.

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POSSIBLE DIETARY INTAKE OF PAs

Commodity Possible

contamination

(µg PA/kg)

Typical

consumption

PA

Exposure

(µg/day)§

Wheat†* 90

(equivalent to 10

heliotrope seeds/kg)

408g/day

(12-15 year old

males)

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Milk†# 300

(300-800 µg/kg

reported)

Say 500g/day 150

Eggs† 168

(19-168 µg/kg

reported)

Say 1x50g egg/day 8

Honey† 2274

(typical unifloral

Paterson’s curse¶)

18.1g/day

(20-45 year old

women)

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§Dutch food regs. allow max. of 1 µg PA/kg food; German herbal regs.

0.1 µg PA/day, zero for pregnant or nursing women; Austrian and Swiss

herbal regs. zero PAs; Australian recommendation 1 µg PA/kg bw/day. †Only some product likely to be contaminated. ¶Product is always contaminated.

*Wheat is “cleaned“ before milling. # Milk is normally blended, potentially reducing

level of any PA contamination.

This rather complex Table shows typical levels of PAs that could be found in some

foods or, in the case of honey from Paterson’s curse, are always present, and

indicates possible daily dietary exposures compared with tolerable levels for PAs set

by international regulations.

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Wheat

The level of PAs shown in wheat is the equivalent of what would be found if 10

heliotrope seeds were present in one kg. That is half the receival standard for AWS

wheat for human consumption. I have chosen 12-15 yo males as typical consumers

in this case because, if you have ever had one in the family, you will know what high

consumers of breakfast cereals and bread they are!! In fact they apparently consume

on average 408 grams of grain-based products per day.

Milk

PAs are transferred from contaminated fodder into milk of cows, goats etc and there

are many studies in which the milk of experimental animals fed PAs has poisoned

suckling young. Levels of PAs between 300 and 800 micrograms per litre have been

recorded in the milk of cows and goats fed with PA plants and the lower figure is

used here.

There have also been quite a few recorded instances in humans where veno-

occlusive disease, typical of PAs, occurred in breast-fed infants of mothers

consuming PA-containing herbal medicines where there was no history of direct

herbal administration to the infants. Eight cases in Germany and Austria described by

one author (Wurm, 1939) are also thought to have been caused by PAs being

transferred in their mothers’ milk.

A review published in 1990 (Molyneux and James, 1990) on the potential for

pyrrolizidine alkaloid contamination in milk from cows presenting a health risk to

humans, concluded that, because commercial milk from cows comes from many

sources and extensive mixing occurs during processing, pyrrolizidine alkaloids in

commercial milk will be diluted and is unlikely to pose a significant health risk for

most consumers although it could add to the cumulative dietary exposure to these

natural toxicants. Families drinking unblended milk they produced themselves or

bought from a neighbour may be more at risk.

In my view, if a dairy producer is experiencing any signs of PA poisoning in their

cows, the milk should be analysed for PAs by a competent laboratory to ensure its

safety.

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Eggs

The level of PAs in the eggs we analysed was up to 168 micrograms of PAs per kg or

about 8 micrograms in one 50g egg and this is the figure used in the Table.

Honey (and pollen)

Honey from fireweeds have been found to contain up to 4000 micrograms of PAs per

kilogram of honey and a reasonable average value for honey from several popular

PA plants (e.g. for Paterson’s curse, blue heliotrope etc) would be 2000 micrograms

of PAs per kilogram.

I have used, as an example in this Table, an actual uni-floral Paterson’s curse honey,

which contained a typical level of 2274µg of PAs per kg. Honey from all PA plants will

be contaminated by PAs and all honeys produced in areas with PA plants are

flowering will contain a certain amount of PAs. We have analysed honeys labeled as

Eucalypt honey and found significant levels of PAs. Also supermarket honeys, with

no floral attribution, have been found to contain significant amounts of PAs.

A few years ago FSANZ recommended that all Paterson’s curse honey be blended

with another type of honey to dilute the level of PAs. Since Paterson’s curse honey

represents 10 to 20% of the honey produced in Australia and there are several other

types of honey that contain PAs, this ensures that most honeys without floral

attribution sold in Australia are likely to contain some PAs. Blending honeys without

knowing if they contain PAs is fraught with problems (no analyses of PAs are being

undertaken prior to blending).

Unlike the other examples in this Table, approximately this level of contamination will

be present in all Paterson’s curse honeys. Grain, milk and eggs are only intermittently

contaminated, and the first two are also likely to be blended with uncontaminated

product, while honey from PA plants, like Paterson’s curse or fireweeds for example,

will always be contaminated by PAs.

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Another worrying bee product is bee pollen. This photograph shows Paterson’s curse

pollen collected by a honeybee but it could equally be fireweed pollen. Pollen

collected from Paterson’s curse has 1000 times more PAs than are found in

Paterson’s curse honey, that is 1000s of milligrams, not micrograms, per kilogram of

pollen. If an apiarist places a “pollen trap” on the hives, these granules of pollen are

dislodged from the legs of the bees and can be collected. One can buy jars of bee

pollen granules in health food shops in Australia and throughout the world. Being

blue in colour it is easy to see Paterson’s curse pollen in jars of mixed granules on

health food store shelves and I can assure you they are there. Fireweed pollen

granules are much harder to see at a glance because they are yellow or orange in

colour, like most other pollens. Pollen granules, sold as health food supplements, are

therefore a particular concern as a source of PA exposure.

Question:

If we are exposed to low levels of PAs in our foods why are there not more

cases of PA poisoning being diagnosed?

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Possible answer:

“A major reason is that they have not been looked for.”

Huxtable 1989

Specialists, and some GPs, may be aware of PAs in herbal products like comfrey but

they are generally unaware that foods such as Paterson’s curse honey contain PAs

at potentially harmful levels.

If one looks into the medical literature it is quite easy to find many cases of liver

failure of unknown cause. Statements like this, where in this case PA exposure was

suspected, are typical.

“Despite extensive diagnostic examinations, we could not completely elucidate

the aetiology of the liver disease in our case. No history of ingestion of toxic

substances or herbal teas was reported by interviewing the mother.

Nevertheless we cannot rule out that the mother had consumed food or drinks

contaminated with pyrrolizidine alkaloids during pregnancy.”

Sergi et al., 1999

There are many other examples in the medical literature where exposure to PAs in

the diet may be the cause of liver failure but this was not considered and the right

questions were never asked. I am sure that the clinicians involved in this case did not

ask the mother whether she was a regular consumer of honey for example.

Here is another case of liver failure looking for a source of exposure to PAs.

“Despite extensive examinations and diagnostics, the etiology of VOD in our

patient has not been elucidated. Nevertheless, we cannot exclude the

possibility that food or drinks consumed by the mother during pregnancy were

contaminated with pyrrolizidine alkaloids.”

Seibold-Weiger et al., 1997

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Besides my colleagues and I, others are also raising the issue of PAs in foods and

trying to make the medical profession more aware of likely exposure to PAs in a

normal diet. Here is an example relating to clusters of leukemia of unknown etiology.

“Surprisingly, no consideration has been devoted to one potential cause that

would account for many, if not all, of the aspects of these clusters.

….

A coordinated investigation by epidemiologists, toxicologists, and

environmental chemists of a PA-leukemia linkage could prove to be a prudent

investment.”

Daughton 2005

Until very recently there was no monitoring of PAs in food in any country in the

world. Australia does not monitor foods such as grain, dairy products and

honey for PAs.

…………..

There are very few laboratories in the world with experience in analyzing foods

for PAs

There is no monitoring of PAs in the food supply in any country, including Australia

and, as I have indicated, there is a lack of awareness, not only among consumers but

also among health professional, of the possible risk to health from PAs in a normal

diet. There are also very few laboratories in the world able to conduct analyses for

PAs in food.

The cohort in the population most likely to be first to show evidence of PA poisoning

are newborn infants whose mothers have been exposed to PAs during pregnancy.

Hospitals should be made aware of, and should consider this possibility in

determining the cause of liver failure in neonates, especially where there is no other

obvious cause. Experience shows that the mother may show no symptoms of

poisoning at dietary levels of PAs that are fatal for their unborn babies.

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Foetuses and neonates, are unfortunately, likely to be the “canaries in the mine”

indicating PAs in the food supply.

Enhancing the capacity of livestock to destroy PAs.

Before concluding my presentation I would like to digress from my main topic for a

few minutes and talk about a different approach to dealing with fireweed that could

form a part of an integrated control program for this weed.

One approach to the problem of fireweed being considered at this meeting is the

introduction of biological agents to help control the level of the weed. I would like to

spend a few minutes mentioning another potential approach to the problems of

fireweed. That is, to enhance the ability of ruminants, such as sheep and goats, to

destroy PAs and ideally to make some ruminants completely resistant to PAs and

useful as biological control agents for PA plants such as fireweed.

Enhanced destruction of PAs in the rumen

• Peptostreptococcus heliotrinreducans

• 85% destruction of PAs in sheep rumen

• Requires free hydrogen

• Competes for hydrogen with methane producing bacteria

• Inhibition of methane producing bacteria may lead to enhanced PA

destruction

•

http://www.pharmcast.com/Patents/Yr2001/June2001/062601/6251879_Antimethanogenic062601.htm

It has been known since the 1970s, as a result of the work of a colleague of mine at

CSIRO, George Lanigan, that a bacterium which he isolated from the rumen of sheep

and cattle and which he named Peptostreptococcus heliotrinreducans, breaks PAs

into two non-toxic components. This is why sheep and some other ruminants such as

goats and, to a lesser extent, cattle are much more resistant to PAs than mono-

gastric animals such as horses, chickens, pigs and humans. Sheep for example can

destroy around 85% of the PAs they ingest in their rumen, prior to absorption. In

many parts of Australia sheep have traditionally been used to control PA plants

29

however sheep are not completely resistant to PAs and do succumb over about 2

seasons of exposure and their productive life is shortened by several years. Their

new born lambs also suffer liver damage.

George Lanigan found that destruction of PAs by Peptostreptococcus

heliotrinreducans requires free hydrogen to be available in the rumen. Unfortunately,

most of the hydrogen in the rumen is used by other rumen bacteria to produce

methane. Methane-producing bacteria compete for available hydrogen with the PA-

destroying bacterium. George reasoned that if we could stop methane production the

increased availability of hydrogen might increase the efficiency of PA destruction

from 85% to 100% and the animals would then be completely protected and the PAs

they ingested would be completely destroyed. There would also be no PAs left to be

transferred into products such as milk. A fully protected cow, sheep or goat, for

example, would be a very effective agent for controlling PA plants such as fireweed.

In recent years methane production by ruminants has been recognised as a major

contributor to climate change. In fact methane is considered to be a much stronger

green house gas than carbon dioxide. If methane production in ruminants could be

inhibited they would not add to green house gas levels and may, at the same time,

become more effective destroyer of PAs. In modern parlance it would be a win, win

situation. As well as being a greenhouse gas, it has long been recognised that

methane also represents lost production. If the carbon lost as methane was instead

converted into productive growth this would be a win, win, win situation all around!

Funding for such a project could come jointly from greenhouse gas abatement

programs and from weed control programs thus spreading the cost burden.

[For a description of a CSIRO-patented antimethanogen that enhanced the

destruction of heliotrope PAs in the rumen see:

http://www.pharmcast.com/Patents/Yr2001/June2001/062601/6251879_Antimethano

genic062601.htm ]

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PAs and insects

I want to conclude with a few words about insects and PAs. This is an area that I

have worked on, on and off, for more than 40 years and it is the main focus of my

current work at the University of Freiburg in Germany.

This moth is Nyctermera amica, commonly call the magpie moth or the Senecio

moth. It stores PAs in its body. These PAs protect it from being eaten by birds,

spiders and other predators. It lays its eggs exclusively on fireweeds (Senecio spp.)

and its caterpillars acquire PAs from the plants that are then transferred to the adult

moth. PAs are also found in the eggs so that all life stages of the moth are protected

by PAs acquired form the plants (fireweeds) it feeds on.

I should make it clear that, despite its specificity for laying its eggs and eating

Senecio species, Nyctermera is not likely to be useful in controlling fireweed in

Australia. It is a species indigenous to Australia and is clearly not significantly

reducing the density of fireweed here and it also does not discriminate Senecio

madagascariensis from indigenous Senecio spp.

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Nyctermera belongs to the moth family Arctiideae, commonly called tiger moths

because their caterpillars and some of the moths have distinctive yellow or orange

and black striped bodies.

Many other species of tiger moths feed exclusively on PA plants and store PAs in

their bodies for protection against predators. Because they are protected by PAs they

are day-flying and warningly coloured. They also fly in a slow and obvious way to

advertise that they are dangerous to eat.

This is another day-flying tiger moth that is full of PAs acquired from its larval food

plants.

Its common name is the “speckled footman” and its Latin name is Utethiesa

pulcheloides. Its caterpillars feed not on fireweeds but on Paterson’s curse and

common heliotrope. Close relatives of the speckled footman in Queensland,

Utethiesa lotrix, specialise on another genus of PA-containing plants, Crotalaria spp.

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When I first analysed this moth for PAs in 1970 and found that it contained the PAs

present in its food plants I also discovered that the male moths secrete a volatile

substance made from the PAs on these organs, called coremata. The coremata are

used to release pheromones for chemical communication between individual moths.

Later on, a group at Cornell University, working on a related moth, Utethiesa ornatrix,

which feeds exclusively on Crotalaria spp., found that it too stores PAs in its body

and secretes the same PA-derived pheromone. They showed that the coremata are

exposed during courtship and that the volatile, PA-derived substance released on the

coremata demonstrates to the female that the male courting her is efficient at storing

of PAs and is therefore a fit partner. The scientists at Cornell also showed that during

mating the male moth transfers not only sperm to the female but also a package of

PAs to ensure that the eggs she lays are well protected by these toxins against

predators such as ants. The package of PAs transferred to the female moth is said to

be a nuptial gift, a wedding present.

33

The group at Cornell showed that when U. ornatrix moths were put into a spider’s

web the spider did not kill and eat them, instead it snipped its web and releases the

moths, allowing them to fly away unharmed. When the moths were reared on an

artificial diet containing no PAs they had no PAs in their bodies and the spiders

readily ate them, demonstrating that the PAs were protecting the moths.

Conclusions:

Returning to my main theme, PAs are clearly undesirable substances in food and

every effort should be made to keep PA plants, such as fireweed out of agricultural

production systems.

PAs are clearly undesirable substances in food and every effort should be

made to keep PA plants, such as fireweed out of agricultural

production systems.

34

There is reason to be concerned about PA poisoning from consumption of foods

containing low levels of PAs. In Australia I am most concerned with the lack of PA

monitoring of honey and the growing use of pollen food supplements that may

contain pollen granules from PA plants. Grains and dairy products are also possible

dietary sources of low levels of PAs that may contribute to chronic, long-term effects.

I believe the efforts of the organisers of this meeting to control fireweed are to be

commended and supported.

“Prevention of exposure is the only effective method of limiting toxicity due to

PAs. Even low doses over a period of time may present a health risk and

exposure should be avoided or minimized as far as possible.”

World Health Organization (1989).

I will finish with this quote again from a pyrrolizidine alkaloid health and safety guide

published by W.H.O. In my opinion we are not doing enough to avoid or minimize

exposure to PAs in foods.

John Edgar, Bega, May 28-29, 2008.

Further reading: International Programme on Chemical Safety (1988). “Pyrrolizidine Alkaloids”. Environmental Health Criteria 80. W.H.O. Geneva. http://www.inchem.org/documents/ehc/ehc/ehc080.htm International Programme on Chemical Safety (1989). Pyrrolizidine Alkaloids Health and Safety Guide. Health and Safety Guide No. 26 W.H.O. Geneva. http://www.inchem.org/documents/hsg/hsg/hsg026.htm California Environmental Protection Agency (1999) Safe Drinking Water and Toxic Enforcement Act of 1986 (proposition 65) available: http://www.oehha.ca.gov/prop65/pdf/batch3_8.pdf

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German Federal Health Bureau (1992). Bundesanzeiger, June 17, 4805; Dt. Apoth. Ztg, 132: 1406-1408. Australia New Zealand Food Authority (2001). Pyrrolizidine alkaloids in food. A Toxicological review and risk assessment. Technical report series No. 2. http://www.foodstandards.gov.au/_srcfiles/TR2.pdf Huxtable, R.J. (1989). Human health implications of pyrrolizidine alkaloids and the herbs containing them. In Toxicants of Plant Origin Vol.1 Alkaloids, pp 41, P.R. Cheeke ed, CRC Press, Boca Raton. Prakash, A.R., Pereira, T.N., Reilly, P.E.B. and Seawright, A.A. (1999). Pyrrolizidine alkaloids in human diet. Mutation Research 443: 53-67. Edgar, J.A. and Smith, L.W. (1999). Transfer of pyrrolizidine alkaloids into eggs: food safety implications. In Natural and Selected Synthetic Toxins, Biological Implications. Tu, A.T., Gaffield, W. Eds. ACS Symposium Series 745. American Chemical Society. Washington DC. Chapter 8, pp 118-128. Edgar, J.A., Roeder, E. and Molyneux, R.J. (2002). Honey from plants containing pyrrolizidine alkaloids: a potential threat to health. J. Agric. and Food Chem. 50: 2719-2730. Beales KA, Betteridge K, Colegate SM, Edgar JA. Solid phase extraction and LCMS analysis of pyrrolizidine alkaloids in honeys. J Agric Food Chem 2004; 52: 6664-6672. Betteridge K, Cao Y, Colegate SM. An improved method for extraction and LCMS analysis of pyrrolizidine alkaloids and their N-oxides in honey: Application to Echium vulgare honeys. J Agric Food Chem 2005; 53: 1894-1902. Boppré M, Colegate SM, Edgar JA. Pyrrolizidine alkaloids of Echium vulgare honey found in pure pollen. J Agric Food Chem 2005; 53: 594-600. Roulet M, Laurini R, Rivier L, et al. Hepatic veno-occlusive disease in newborn infant of a woman drinking herbal tea. J Pediatr 1988: 112: 433-436. Rasenack R, Müller C, Kleinschmidt M, et al. Veno-occlusive disease in a fetus caused by pyrrolizidine alkaloids of food origin. Fetal Diagn Ther 2003; 18: 223-225. Sergi C, Beedgen B, Linderkamp O, et al. Fatal course of veno-occlusive disease of the liver (endophlebitis hepatica obliterans) in a preterm infant. Pathol Res Pract 1999; 195: 847-851. Seibold-Weiger K, Vochem M, Mackensen-Haen S, et al. Fatal hepatic veno-occlusive disease in a newborn infant. Am J Perinatol 1997; 15: 107-111. Müller-Höcker J, Weiss M, Meyer U, et al. Fatal copper storage disease of the liver in a German infant resembling Indian childhood cirrhosis. Virchows Arch A Pathol Anat Histopathol 1987; 411: 379-385.

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Tollmann R, Neureiter D, Lang T, et al. Late manifestation of Indian childhood cirrhosis in a 3-year-old German girl. Eur J Pediatr 1999; 158: 375-378. Price LA, Walker NI, Clague AE et al. Chronic copper toxicosis presenting as liver failure in an Australian child. Pathol 1996; 28: 316-320. Edgar JA. Pyrrolizidine alkaloids and food safety. Chemistry in Australia 2003;70:4-7. Xia Q, Chou MW, Edgar JA, Doerge DR, Fu PP. Formation of DHP-derived DNA adducts from metabolic activation of the prototype heliotridine-type pyrrolizidine alkaloid, lasiocarpine. Cancer Lett accepted for publication. Fu PP, Xia Q, Lin G, Chou MW. Genotoxic pyrrolizidine alkaloids – mechanisms leading to DNA adduct formation and tumorigenicity. Int J Mol Sci 2002;3:948-64. ABC Health Report. Honey – Nature’s own health food? Think again. Australian Broadcasting Corporation: 2004. Available at: http://www.abc.net.au/rn/talks/8.30/helthrpt/stories/s1037977.htm Afghanistan: WHO confirms “charmak” (heliotrope poisoning) disease in Herat Province in 2007-2008. http://www.irinnews.org/report.aspx?ReportId=78218 Antimethanogen enhancing PA destruction in the rumen. http://www.pharmcast.com/Patents/Yr2001/June2001/062601/6251879_Antimethanogenic062601.htm