1. WELCOME
Food Safety and Toxicology
Presentation on Toxicants Generated During
Thermal Processing of Food
(Acrylamide)
Presented By,
Basavaraj Gani Sadashivappa
2. INRODUCTION
Acryl amide is a thermally induced toxicant present in different
processed foods in varying amount. Due to its detrimental
effect on human health, it has become a major concern in
public health and food safety. This presentation aims to
review and focuses on the mechanisms of acryl amide
formation, the effects of different processing parameters
such as pH, temperature, time, types and the amount of raw
materials, its toxicity level, and its detection methods in
complex food systems. Toxicity levels of acryl amide have
been found to be neurotoxin and carcinogenic. Food safety
authorities including Codex Alimentarius Commission are in
the process of reviewing their standards to fix the limit of
acryl amide in processed foods.
3. Continue..
• Acryl amide is a highly reactive unsaturated amide.
• Chemical formula is C3H5NO and named as prop-2-enamide
in IUPAC system of nomenclature.
• It is a white odourless crystalline solid, highly soluble in
water, ethanol, ether and chloroform.
• Non-thermal decomposition of acryl amide produces
ammonia whereas thermal decomposition produces carbon
monoxide, carbon dioxide, and oxides of nitrogen
5. MECHANISM OF ACRYLAMIDE
FORMATION
•
The main ingredients that are responsible for acryl amide formation in
foods are carbohydrates especially reducing sugars and asparagine.
• Maillard reaction has been considered as the major reaction pathways for
acryl amide formation while processing of food.
• Three major hypotheses are proposed to describe the mechanism of
formation of acryl amide in foods.
1. The first mechanism explains the direct formation of acryl amide from
amino acids such as alanine, asparagine and glutamine and from
methionine
2. The second mechanism involves the formation of acrylamide via the
formation of acrolein
3. The most explained pathway for acrylamide formation is via Maillard
reaction between reducing sugars and the amino acid asparagine.
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•
•
•
High temperature cooking such as frying, roasting, or baking is most
likely to cause acrylamide formation.. Acrylamide is found in significant
amount in plant based foods such as potato products, grain products
or coffee.
The amount of acrylamide found in dairy, meat, and fish products is
negligible.
Generally acrylamide is more likely to accumulate when cooking is done
for longer periods or at higher temperatures. It is generally formed at
temperatures higher than 120ᵒC and increases with increasing frying
and baking temperatures.
7. A pathway of Acrylamide formation from amino
acid and reducing sugar
Asparagine + Glucose
(HEAT)
Schiff’s base
(DECARBOXYLATION)
Amadori re-arrangement
(B-ELIMINATION)
Acryl amide
8. Alternative routes for formation of
acrylamide
Fatty Acids
Acrolein
Glycerol
Aspartic acid
Carnosine
B-Alanine
Acrylic Acid
Acryl Amide
Serine
Cysteine
Pyruvic acid
9. Sources in Human Nutrition
Food commodities
Maximum acryl amide content in PPM
In EU
In US FDA
Breads
12-3200
<10-364
Crackers and biscuits
<30-2000
26-504
Toasted cereals
<30-3200
52-266
French fries
85-1104
20-1325
Vegetables
10-<50
<10-70
Meats
<30-64
<10-116
Candy and desserts
<20-110
<10-909
Coffee and tea
170-700
175-351
Dairy products
10-100
<10-43
Infant foods
40-120
<10-130
10. Absorption of Acryl Amide
•
Absorption of Acrylamide has traditionally been researched using the oral and
intraperitoneal modes of introduction into the body with relatively little research into
inhalation exposure. Absorption is closely linked with the mode of environmental exposure
to the human body. Acryl amide's half-life in the open air is relatively short in comparison to
its other modes of transportation.
•
Studies conducted involving oral exposures in rats have yielded results showing that
Acrylamide is readily absorbed into the GI tract .
•
A comparison of oral vs. intraperitoneal exposure was conducted showing that systemic
absorption following oral exposure was less versus that of intraperitoneal exposure.
•
Oral exposure has been the primary focus of Acryl amide's main entry into the human and
thus a heavy focus of tissue research into its toxicology upon GI tract tissues has taken place.
•
Another tissue susceptible to Acrylamide absorption is in nervous tissue of the peripheral
nervous system. High uptake levels of Acrylamide in the motor nerve terminus and optic
nerve are two areas, which have been shown to result in degeneration of the distal axon
terminal. Essentially, this process of dying back allows more acrylamide to enter the cell
causing further intoxication. Earliest neuromuscular changes can be seen in the pacinian
corpuscles followed by the muscle spindles and finally the motor nerve terminus.
•
In the optic nerve, mid diameter axons are most prone to acrylamide absorption in the optic
tract allowing for neuronal degeneration in tested primates.
11. Continue..
•
Acrylamide metabolism can have 2 possible pathways in route to the excretion of
the compound. Pathway 1 involves utilizing the Cytochrome P450 metabolic
enzyme to transform Acrylamide to an easier excrete able Gylcidamide. Pathway 2
involves the conjugation of Acrylamide to Glutathione (GST) there by forming Nacetyl-S- (3-amino-3-oxopropyl) Cysteine as the final product.
•
Differentiation between pathways 1 versus pathway 2 is dependant on the species
absorption of Acrylamide. Area specific metabolism in rats has shown that the
liver, kidney, brain, and erythrocyte GST has a significant higher affinity for binding
Acrylamide with the liver being 3 times more prone to Acrylamide binding versus
the other mentioned target organs. Solubility of the compound and its method of
absorption is highly dependant on the GST pathway for conjugation to a watersoluble product. The conversion by GST to Gylcidamide results in the chemical
creation of a radical epoxide, which can express toxicity independently from other
neuropathies associated with Acrylamide.
•
Studies have been conduct in rats, which indicate that some species of rats may in
fact inhibit GST conjugation thus allowing for increased amounts of the toxic
product Gylcidamide.
12. Toxicity
•
Human exposure to acrylamide primarily comes from dermal contact with solid
monomer and inhalation of dust and vapour in the occupational setting
• Humans may be exposed to acrylamide through the ingestion of drinking water
that is contaminated with acrylamide or the intake of acrylamide from food.
• Major health effects of acrylamide are skin irritation such as redness and peeling
of the skin of palms and neuropathy regarding the central nervous system and the
peripheral nervous system. Acute and sub-acute intoxication with a large dose by
ingestion by water contaminated with acrylamide can cause severe symptoms like:
1. Acute toxicity : Acrylamide is a skin and respiratory tract irritant in humans.
Reported oral LD50 values are in the range of 159 mg/kg to 300 mg/kg body
weight in rats the central nervous system and polyneuropathy may appear later
2. Chronic Toxicity : Acrylamide is a human neuro-toxicant. Adverse effects in rats
administered small amounts of acrylamide include general systemic toxicity and
haematological changes. Acrylamide is also a neuro-toxicant to animals.
13. a. In rats, repeated oral administration of acrylamide at doses of 20 mg/kg bw/day
and above produced peripheral neuropathy, At 5 mg/kg bw/day in a 90-day study in
rats, peripheral lesions occurred and slight changes in peripheral nerve tissue
occurred.
b. In monkeys, clinical signs of peripheral neuropathy occurred at doses of 10 mg/kg
bw/day for up to 12 weeks.
c. Carcinogenicity : Although inadequate evidence is available from human studies,
several laboratory animal studies have shown that acrylamide causes a variety of
tumours in rats and mice. Acrylamide has been classified by the U.S. EPA as a B2, a
probable human carcinogen, by IARC as a 2B, a possible human carcinogen, and by
ACGIH as an A3, confirmed animal carcinogen with unknown relevance to human.
d. Genotoxicity : Acrylamide causes chromosomal aberrations, dominant lethality,
chromatic exchanges and unscheduled DNA synthesis in various in vitro and in vivo
systems. When administered at a level of 500 ppm in the diet for 3 weeks in mice,
acrylamide caused a high frequency of chromatic exchanges.
e. Neurotoxicity : Acrylamide is a neurotoxin by either oral (in animals) or inhalation
exposure (in humans and in animals). Toxic effects are central and peripheral
neuropathy causing drowsiness, hallucinations and distal numbness. Recovery is
possible after cessation of exposure. EPA has derived an oral reference dose (RfD)
of 0.0002 mg/kg/day for acrylamide, based on adverse nervous system effects in
laboratory animals.
14. Determination of Acryl Amide
• Various methods are available to determine the acrylamide content
of food. The most common methods utilize either GC-MS or LC-MS
• Water is often used as the extraction solvent. Solid-phase
extraction (SPE) is typically use to prepare samples. SPE phases
that have been used for this method include graphitized carbon
black, mixed mode anion and cation exchange, and polymeric
materials.
• The GC-MS method of determining acrylamide content in food is
well established. A water extract of the food is brominated to form
2,3-dibromopropionamide, a derivative of acrylamide with
enhanced GC properties.
• The derivative is then analyzed by GC-MS using Methacrylamide as
an internal standard . Detection limits for the GC-MS method are
typically in the 5–10 µg/kg range
15. Steps in Acryl amide analysis by GC/MS
Method
•
Acryl amide extracted with water, test portion homogenized and acidified
to pH 4-5
•
Addition of carrez 1 and carrez solution
•
Extraction with Ethyl acetate hexane (80:20), Filtration over Na2So4
•
Clean up with flurosile elution of acrylamide with acetone
•
Evaporation residue taken up in ethyl acetate, tri ethylamine added
•
Filtration, injection into GC-MS
•
Identification and Quantification
16. FDA Action Plan for Acrylamide in Food
WHO/FAO Recommendations
• In June 2002, the World Health Organization (WHO) and the Food
and Agriculture Organization (FAO) convened an expert
consultation on acrylamide. The consultation, in which three FDA
experts participated, concluded that the presence of acrylamide in
food is a major concern, and recommended more research on
mechanisms of formation and toxicity. Both the WHO/FAO
consultation and the FDA have recommended that people continue
to eat a balanced diet rich in fruits and vegetables. The WHO/FAO
consultation advised that food should not be cooked excessively,
i.e., for too long a time or at too high a temperature, but also
advised that it is important to cook all food thoroughly--particularly
meat and meat products--to destroy food borne pathogens
(bacteria, viruses, etc.) that might be present.
17. Further actions
• FDA will develop and revise regulatory options as additional
knowledge is gained on acrylamide in food. Many of the items
in the action plan are geared toward achieving that end.
• FDA will encourage industry to adopt feasible, practical, and
safe processes that are successful at reducing acrylamide, if
needed.
• FDA will develop and revise consumer messages about
dietary choices and cooking methods, as additional
knowledge is gained about acrylamide in food. FDA
understands the importance of consumer messages that can
assist consumers in making informed dietary choices. Any
adjustments in consumer messages would consider the
totality of the science.
