There are 92 natural elements; approximately 22 are known to be essential nutrients of the mammalian body and are referred to as micronutrients (Concon, 1988). Among the micronutrients are iron, zinc, copper, manganese, molybdenum, selenium, iodine, cobalt, and even aluminum and arsenic. However, among the 92 elements, lead, cadmium, and mercury are familiar as contaminants (or at least have more specifications setting their limits in food ingredients). The prevalence of these elements as contaminants is not due so much to their ubiquity in nature but rather to their use by humans.
Although the toxicity of lead is well known, lead may also be an essential trace mineral. A lead deficiency induced by feeding rats <50 ppb lead (vs 1000 ppb in controls) over one or more generations produced effects on the hematopoietic system, decreased iron stores in the liver and spleen, and caused decreased growth (Kirchgessner and Reichmayer-Lais, 1981), but apparently not as a result of an effect on iron absorption. Although the toxic effects of lead are discussed elsewhere in this text, it is important to note that its effects are profound (especially in children) and appear to be long-lasting because mechanisms for excretion appear to be inadequate in comparison to those for uptake (Linder, 1991). Foods may become contaminated with lead if they are grown, stored, or processed under conditions that could introduce larger amounts of lead into the food, such as when a root crop is grown in soil that has been contaminated from the past use of leaded pesticides.
Over the years, recognition of the serious nature of lead poisoning in children has caused the World Health Organization (WHO) and FDA to adjust the recommended tolerable total lead intake from all sources of not more than 100 μg/day for infants up to six months old and not more than 150 μg/day for children from six months to two years of age to the considerably lower range of 6 to 18 μg/day as a provisional tolerable range for lead intake in a 10-kg child. The FDA is recommending that lead levels in candy products likely to be consumed frequently by small children do not exceed 0.1 ppm because such levels are achievable under good manufacturing practices and would not pose a significant risk to small children for adverse effects. This recommended maximum level is consistent with the FDA’s longstanding goal of reducing lead levels in the food supply (FDA, 2005).
Initiatives to reduce the level of lead in foods, such as the move to eliminate lead-soldered seams in soldered food cans that was begun in the 1970s and efforts to eliminate leachable lead from ceramic ware glazes, have resulted in a steady decline in dietary lead intake (Shank and Carson, 1992). However, lead remains in the diet as the result of still extant lead water pipe (EPA, 2011) and has triggered recent recalls of containers (lead in stainless steel brandy flasks) and food (lead in turmeric spice and, ironically, in a bubble gum called “Toxic Waste”) (FDA, 2011c, d, and e).
Cadmium is a relatively rare commodity in nature and usually is associated with shale and sedimentary deposits. It is often found in association with zinc ores and in lesser amounts in fossil fuel. Although rare in nature, it is a nearly ubiquitous element in American society because of its industrial uses in plating, paint pigments, plastics, and textiles. Exposure to humans often occurs through secondary routes as a result of dumping at smelters and refining plants, disintegration of automobile tires (which contain cadmium-laden rubber), subsequent seepage into the soil and groundwater, and inhalation of combustion products of cadmium-containing materials. The estimated yearly release of cadmium from automobile tires alone ranges from 5.2 to 6.0 metric tons (Davis, 1970; Lagerwerff and Sprecht, 1971). However, in the absence of an overt environmental contamination, food remains the primary source for cadmium uptake for nonsmokers (EFSA, 2009).
Cadmium, like mercury, can form alkyl compounds, but unlike mercury, the alkyl derivatives are relatively unstable and consumption almost always involves the inorganic salt. Of two historical incidents of cadmium poisoning, one involved the use of cadmium-plated containers to hold acidic fruit slushes before freezing. Up to 13 to 15 ppm cadmium was found in the frozen confection, 300 ppm in lemonade, and 450 ppm in raspberry gelatin. Several deaths resulted. A more recent incident of a chronic poisoning involved the dumping of mining wastes into rice paddies in Japan. Middle-aged women who were deficient in calcium and had multiple pregnancies seemed to be the most susceptible. Symptoms included hypercalciuria; extreme bone pain from osteomalacia; lumbago; pain in the back, shoulders, and joints; a waddling gait; frequent fractures; proteinuria; and glycosuria. The disease was called itai-itai (ouch-ouch disease) as a result of the pain with walking. The victims had a reported intake of 1000 μg/day, approximately 200 times the normal intake of unexposed populations (Yamagata and Shigematsu, 1970). Cadmium exposure has also been associated with cancer of the breast, lung, large intestine, and urinary bladder (Newberne, 1987).
Cadmium absorption is greatest with pulmonary exposure, but it is relatively low (3%–5%) after dietary exposure to humans, although absorption is effected by several factors including nutritional status (eg, low body iron), number of pregnancies, and general health. Cadmium is retained in the kidney and liver in the body, with a very long biological half-life ranging from 10 to 30 years. Cadmium is primarily toxic to the proximal tubular cells of the kidney and tends to accumulate in the cortex of the kidney, eventually leading to renal failure. Bone demineralization occurs with prolonged exposure, as illustrated in the Japanese exposure, which may be secondary to renal dysfunction (EFSA, 2009).
JECFA (2005) investigated cadmium consumption for several countries including the United States and found that the greatest impacts of maximum (allowable) levels (ML) were seen with stem/root vegetables, other vegetables, and molluscs (41%, 68%, and 42%, respectively, when the lowest MLs were used), which represented an insignificant change over a previous intake assessment. Therefore, the current total intake of cadmium was only 40% to 60% of the Provisional Tolerable Weekly Intake (PTWI) of 7 μg/kg bw/week established several years ago, and a slight variation of 1% to 6% due to the use of the proposed new maximum limits is of no significance in terms of risk to human health (JECFA, 2005).
Exposure to elemental mercury is relatively rare, although it was once responsible for an occupational disease of hat manufacturers, as elemental mercury was used for the curing of animal pelts. Inhalation of the mercury fumes led to mental deterioration, subsequently named “mad hatter syndrome.” The effects of mercury were more pronounced following the application of mercurial ointments for treatment of syphilis and gave rise to the pun “a night with Venus is followed by a lifetime with Mercury” (Waldron, 1983).
Of interest to food toxicology is the methyl derivative, methyl mercury, formed by bacterial action in an aquatic environment from anthropogenic and natural sources of elemental mercury. Anthropogenic sources include burning of coal (which contains mercury), chloralkali process, and other sources of elemental mercury discharge into aquatic environments. In the case of Minamata, Japan, there was a direct discharge of methyl mercury into the environment. Methyl mercury exposure may cause neurological paresthesias, ataxia, dysarthria, hearing defects, and death. Developmental delays have been documented in children borne of mothers exposed to methyl mercury (Carrington and Bolger, 1992). Other than direct exposure to methyl mercury, exposure usually comes about as the result of methyl mercury becoming incorporated into the food chain. Near the peak of the food chain, methyl mercury becomes concentrated in fish including, bonito (Sarda spp), halibut (Hippoglossus spp), mackerel (Scomberomorus spp), marlin (Makaira spp), shark (all species), swordfish (Xiphias gladius), and bluefin tuna (Thunnus spp). The selection of these species for monitoring and tolerance setting was based on historical data on levels of methyl mercury found in fish consumed in the United States. The selection was also based on an FDA action level of 1.0 ppm in the edible portion of fish (FDA, 2001). However, the allowable level of mercury depends on whether the mercury was “added,” that is, did the presence of mercury arise from an anthropogenic source (ie, was the fish caught in an area known for mercury discharge or was it naturally present in the environment?) (Hutt et al., 2007).
Halogenated Hydrocarbons (Polychlorinated and Polybrominated Hydrocarbons)
Halogenated hydrocarbons have been with us for some time, and given their stability in water and resistance to oxidation, ultraviolet light, microbial degradation, and other sources of natural destruction, halogenated organics will persist in the environment for some time to come, albeit in minute amounts. However, with the introduction of chlorinated hydrocarbons as pesticides in the 1930s, diseases associated with an insect vector such as malaria were nearly eliminated. In the industrialized world, chlorinated organics brought the promise of nearly universal solvents, and their extraordinary resistance to degradation made them suitable for use as heat-transfer agents and carbonless copy paper, among other uses. As a result of their facile nature, their resulting wide-range uses, and resistance to degradation (and ease of detection), chlorinated hydrocarbons have been found in a wide variety of foods (Table 31-23).
Table 31-23Examples of Chlorinated Hydrocarbons in British Food ||Download (.pdf) Table 31-23 Examples of Chlorinated Hydrocarbons in British Food
| ||CHLORINATED HYDROCARBONS (μg/kg) |
|FOOD ||CHCl3 ||CCl4 ||TCE ||TCEY ||TTCE ||PCE ||HCB ||HCBD ||PerCE |
|Milk ||5.0 ||0.2 || ||0.3 ||— ||— ||1.0 ||0.08 ||0.3 |
|Cheese ||33.0 ||5.0 || ||3.0 ||0.0 ||0.0 ||0.0 ||0.0 ||2.0 |
|Butter ||22.0 ||14.0 || ||10.0 ||— ||— ||— ||2.0 ||13.0 |
|Chicken eggs ||1.4 ||0.5 || ||0.6 ||0.0 ||0.0 ||0.0 ||0.0 ||0.0 |
|Beef steak ||4.0 ||7.0 ||3.0 ||16.0 ||0.0 ||0.0 ||0.0 ||0.0 ||0.9 |
|Beef fat ||3.0 ||8.0 ||6.0 ||12.0 || || || || ||1.0 |
|Pork liver ||1.0 ||9.0 ||4.0 ||22.0 ||0.5 ||0.4 || || ||5.0 |
|Margarine ||3.0 ||6.0 ||— || ||0.8 || || || ||7.0 |
|Tomatoes ||2.0 ||4.5 ||— ||1.7 ||1.0 || ||70.1 ||0.8 ||1.2 |
|Bread (fresh) ||2.0 ||5.0 ||2.0 ||7.0 ||— ||— ||— ||— ||1.0 |
|Fruit drink (canned) ||2.0 ||0.5 ||— ||5.0 || ||0.8 || || ||2.0 |
There have been only a few incidents of mass poisonings via food, two of which involved cooking oil contaminated with polychlorinated organics. The first became known as yusho, or rice oil disease, from rice oil contamination originally thought to be confined to polychlorinated biphenyls (PCBs), but has since been revised to include polychlorinated quarterphenyls and polychlorinated dibenzofurans (PCDFs) (Kanagawa et al., 2008). The poisoning occurred in 1968 in Japan and affected approximately 2000 individuals. The most vulnerable were newborns of poisoned mothers. The liver and skin were the most severely affected. Symptoms included dark brown pigmentation of nails; acne-like eruptions; increased eye discharge; visual disturbances; pigmentation of the skin, lips, and gingiva; swelling of the upper eyelids; hyperemia of the conjunctiva; enlargement and elevation of hair follicles; itching; increased sweating of the palms; hyperkeratotic plaques on the soles and palms; and generalized malaise. Recovery requires several years (Anderson and Sogn, 1984; Guo et al., 2003), although more recent authors claim that recovery is gradual even at the 37-year mark, as some patients exhibit symptoms specific to the disease including, but not limited to, hyperglyceridemia, pulmonary disorders, and intractable headache, which were added to the initial diagnostic criteria (Kanagawa et al., 2008). The second incident occurred in 1979 in Yucheng, Taiwan, which also involved PCB-contaminated cooking oil and exposed a similar number of people as the earlier incident in Japan (Guo et al., 2003). The total intakes of PCBs and PCDFs for Yusho and Yucheng victims were estimated to be 633 and 3.4 mg and 973 and 3.84 mg, respectively (Masuda, 1985).
Polychlorinated biphenyls are not formed naturally and were manufactured for use as insulators in electrical machinery, although manufacturing was banned in the United States in 1977. Despite the persistence of chlorinated hydrocarbons in our environment and their reported ubiquity, their low degree of demonstrable toxicity at ambient levels indicates a relatively low risk to humans. Ames et al. (1987) described a method for interpreting the differing potencies of carcinogens and human exposures: the percentage human exposure dose/rodent potency dose (HERP). Using this method, they claimed that the hazard from trichloroethylene-contaminated water in Silicon Valley or Woburn, Massachusetts, or the daily dietary intake from dichlorodiphenyltrichloroethane (DDT) (or its product, dichlorodiphenyldichloroethylene [DDE]) at a HERP of 0.0003% to 0.004% is considerably less than the hazard presented by the consumption of symphytine in a single cup of comfrey herb tea (0.03%) or the hazard presented by aflatoxin in a peanut butter sandwich (0.03%). The FDA’s authority to set tolerances has been used only once in establishing levels for PCBs (21 CFR 109.15 and 109.30).
Polybrominated biphenyls (PBBs) and polybrominated biphenyl ethers (PDBEs) (the latter are also called brominated diphenyl oxides) were used as fire retardants in, among other things, household furniture, airplane seat upholstery, and children’s pajamas, all of which had been identified as sources of preventable disfiguring burns and tragic deaths. Uses also extended to electrical equipment, paint, and plastics, and PBBs have been manufactured globally by several companies.
Appropriately, one of the names under which PPBs were sold was “FireMaster.” Tragedy struck in Michigan in 1973, when FireMaster was placed in bags labeled “NutriMaster” (a magnesium oxide animal feed supplement). Several farms received feed contaminated by FireMaster and feed consumption and milk production dropped by as much as 50%, but the causative agent remained a mystery. As the result of a persistent dairy farmer and more than one serendipitous event (eg, a chromatograph left on by accident, allowing PBBs to elute after a considerable period of time), PBBs were identified as the causative agent. It was found that approximately 650 lbs of PBB had entered the food chain through milk and other dairy products, as well as beef, pork, sheep, poultry, and egg products (Fries, 1985; MDCH, 2011). Over 500 Michigan farms were quarantined, and approximately 30,000 cattle, 4500 swine, 1500 sheep, and 1.5 million chickens were destroyed, along with over 800 tons of animal feed, 18,000 pounds of cheese, 2500 pounds of butter, 5 million eggs, and 34,000 pounds of dried milk products (MDCH, 2011). The incident was eventually publicized in 1981 as a television movie entitled “Bitter Harvest.” PBB-exposed Michigan residents had a variety of complaints including nausea, abdominal pain, loss of appetite, joint pain, fatigue, and weakness, and although a cause–effect relationship could not be established for these symptoms, there is a fairly strong case for development of acne from exposure to PBBs.
Like PCBs, many distinct isomers are possible of PBB and PBDE, but the three main commercial products are pentabromodiphenyl oxide or ether, octabromodiphenyl oxide or ether, and decabromodiphenyl oxide or ether. Since the Michigan incident, the production of PBBs has declined, but with a concomitant increase in demand for PBDEs. Worldwide demand for PBDEs in 2001 was estimated at 70,000 metric tons. These substances are chemically rugged entities and are very resistant to acids, bases, heat, light, reduction, and oxidation—all advantages for their intended use, but disadvantages when discharged into the environment. Interestingly, PBDEs are reported to occur naturally and are produced by Vibrio spp, in association with sponges of the Dysidea genus. Conversely, some environmental remediation may occur with two strains of Pseudomonas isolated from runoff areas of the Michigan manufacturer.
Absorption of PBDEs is limited with some 80% to 90% of a dose eliminated in the feces unchanged; degree of bromination is inversely proportional to absorption, but once absorbed, PBDEs are persistent and bioaccumulating. PBDEs in rat subchronic studies produced effects in liver, kidney, and thyroid in males and females. Liver enlargement, associated with increased metabolism, was reported at doses of 1 to 10 mg/kg bw. DecaBDE at 2.5% and 5% of the diet to mice and rats was carcinogenic. PBDEs are not genotoxic, but are reported to have reproductive effects in rats and rabbits. Data for humans were reported as being incomplete. Because of the limited amount of data, the JECFA committee could not allocate a Provisional Tolerable Weekly Intake (PTWI) (deBoer et al., 2000; JECFA, 2005).
Nitrosmaines, Nitrosamides, and N-Nitroso Substances
Nitrogenous compounds such as amines, amides, guanidines, and ureas can react with oxides of nitrogen (NOx) to form N-nitroso compounds (NOCs) (Hotchkiss et al., 1992). The NOCs may be divided into two classes: the nitrosamines, which are N-nitroso derivatives of secondary amines, and nitrosamides, which are N-nitroso derivatives of substituted ureas, amides, carbamates, guanidines, and similar compounds (Mirvish, 1975).
Nitrosamines are stable compounds, whereas many nitrosamides have half-lives on the order of minutes, particularly at pH >>6.5. Both classes have members that are potent animal carcinogens, but by different mechanisms. In general, the biological activity of an NOC is thought to be related to alkylation of genetic macromolecules. N-nitrosamines are metabolically activated by hydroxylation at an α-carbon. The resulting hydroxyalkyl moiety is eliminated as an aldehyde, and an unstable primary nitrosamine is formed. The nitrosamine tautomerizes to a diazonium hydroxide and ultimately to a carbonium ion. Nitrosamides spontaneously decompose to a carbonium ion at physiological pH by a similar mechanism (Hotchkiss et al., 1992). This is consistent with in vitro laboratory findings because nitrosamines require S9 for activity and nitrosamides are mutagenic de novo.
NOCs originate from two sources: environmental formation and endogenous formation (Table 31-24). Environmental sources have declined over the last several years but still include foods (eg, nitrate-cured meats) and beverages (eg, malt beverages), cosmetics, occupational exposure, and rubber products (Hotchkiss, 1989). NOCs formed in vivo may actually constitute the greatest exposure and are formed from nitrosation of amines and amides in several areas, including the stomach, where the most favorable conditions exist (pH 2–4), although consumption of H2-receptor blockers or antacids decreases the formation of NOCs.
Table 31-24Sources of Dietary NOCs ||Download (.pdf) Table 31-24 Sources of Dietary NOCs
The use of nitrate and/or nitrite as intentional food additives, both of which are added to fix the color of meats, inhibit oxidation, and prevent toxigenesis.
Drying processes in which the drying air is heated by an open-flame source. NOx is generated in small
amounts through the oxidation of N2, which nitrosates amines in the foods. This is the mechanism for contamination of malted barley products.
NOCs can migrate from food contact materials such as rubber bottle nipples.
NOCs can inhabit spices, which may be added to food
Cooking over open flames (eg, natural gas flame) can result in NOC formation in foods by the same mechanism as drying.
Environmentally, nitrite is formed from nitrate or ammonium ions by certain microorganisms in soil, water, and sewage. In vivo, nitrite is formed from nitrate by microorganisms in the mouth and stomach, followed by nitrosation of secondary amines and amides in the diet. Sources of nitrate and nitrite in the diet are given in Table 31-25. Many sources of nitrate are also sources of vitamin C. Another possibly significant source of nitrate is well water; although the levels are generally in the range of 21 μM, levels of 1600 μM (100 mg/L) have been reported (Hotchkiss et al., 1992). However, on the average, Western diets contain 1 to 2 mmol nitrate/person/day (Hotchkiss et al., 1992). Nitrosation reactions can be inhibited by preferential, competitive neutralization of nitrite with naturally occurring and synthetic materials such as vitamin C, vitamin E, sulfamate, antioxidants such as butylated hydroxytoluene, BHA, and gallic acid, and even amino acids or proteins (Hotchkiss, 1989; Hotchkiss et al., 1992).
Table 31-25Nitrate and Nitrite Content of Food ||Download (.pdf) Table 31-25 Nitrate and Nitrite Content of Food
|VEGETABLES ||NITRATE (ppm) ||NITRITE (ppm) ||MEAT ||NITRATE (ppm) ||NITRITE (ppm) |
|Artichoke ||12 ||0.4 ||Unsmoked side bacon ||134 ||12 |
|Asparagus ||44 ||0.6 ||Unsmoked back bacon ||160 ||8 |
|Green beans ||340 ||0.6 ||Peameal bacon ||16 ||21 |
|Lima beans ||54 ||1.1 ||Smoked bacon ||52 ||7 |
|Beets ||2400 ||4 ||Corned beef ||141 ||19 |
|Broccoli ||740 ||1 ||Cured corned beef ||852 ||9 |
|Brussel sprouts ||120 ||1 ||Corned beef brisket ||90 ||3 |
|Cabbage ||520 ||0.5 ||Pickled beef ||70 ||23 |
|Carrots ||200 ||0.8 ||Canned corn beef ||77 ||24 |
|Cauliflower ||480 ||1.1 ||Ham ||105 ||17 |
|Celery ||2300 ||0.5 ||Smoked ham ||138 ||50 |
|Corn ||45 ||2 ||Cured ham ||767 ||35 |
|Radish ||1900 ||0.2 ||Belitalia (garlic) ||247 ||5 |
|Rhubarb ||2100 ||NR ||Pepperoni (beef) ||149 ||23 |
|Spinach ||1800 ||2.5 ||Summer sausage ||135 ||7 |
|Tomatoes ||58 ||NR ||Ukranian sausage (Polish) ||77 ||15 |
|Turnip ||390 ||NR ||German sausage ||71 ||17 |
|Turnip greens ||6600 ||2.3 || || || |
N-nitrosoproline is the most common nitrosoamine present in humans and is excreted virtually unchanged in the urine. The basal rate of urinary excretion of nitrosoproline, which is claimed to be noncarcinogenic, is 2 to 7 g/day in subjects on a low-nitrate diet (Oshima and Bartsch, 1981).
Epidemiological studies have not provided compelling evidence of a causal association between nitrate exposure and human cancer nor has a causal link been shown between NOCs, preformed in the diet or endogenously synthesized, and the incidence of human cancer (Gangolli, 1999). The cancer risk of nitrite, nitrate, and processed meats is weak and inconclusive, and the risk must be weighed against the health benefits of restoring NO homeostasis via dietary nitrite and nitrate. These benefits should be considered before arriving at any new regulatory or public health guidelines (Milkowski et al., 2010).
Food-borne Molds and Mycotoxins
Molds have served humans for centuries in the production of foods (eg, ripening cheese) and have provided various fungal metabolites with important medicinal uses; they also may produce secondary metabolites with the potential to produce severe adverse health effects, including behavioral changes (Cousins et al., 2005). It is possible that ergot mycotoxins may have exerted a major role in restricting population expansion and only the reduced dependency on rye cereal as the staple food in the sixteenth and seventeenth centuries, arising from the introduction of wheat and potatoes, allowed the steady upward movement in population growth in Europe (CAST, 2003; IFST, 2006a).
Mycotoxins are secondary fungal metabolites (ie, not essential for survival of the mold) secreted into the microenvironment around the mold. Mycotoxins represent a diverse group of chemicals that can occur in a variety of plants used as food, including commodities such as cereal grains (barley, corn, rye, wheat), coffee, dairy products, fruits, nuts, peanuts, and spices. A few mycotoxins also can occur in animal products derived from animals that consume contaminated feeds (eg, milk). However, because commodities are eaten in the greatest amounts, the mycotoxins present in these foods represent the greatest risk (Cousins et al., 2005).
The current interest in mycotoxicosis was generated by a series of reports in 1960–1963 that associated the death of turkeys in England (so-called turkey X disease) and ducklings in Uganda with the consumption of peanut meal feeds containing mold products produced by Aspergillus flavus (Stoloff, 1977). The additional discovery of aflatoxin metabolites (eg, aflatoxin M1 in milk) led to more intensive studies of mycotoxins and to the identification of a variety of these compounds associated with adverse human health effects, both retrospectively and prospectively.
Moldy foods are consumed throughout the world during times of famine, as a matter of taste, and through ignorance of their adverse health effects. Epidemiological studies designed to ascertain the acute or chronic effects of such consumption are few. Data from animal studies indicate that the consumption of food contaminated with mycotoxins has the potential to contribute to a variety of human diseases (Miller, 1991). Reports of acute intoxications are few; however, prolonged exposure to small quantities of mycotoxin may lead to more insidious effects including growth retardation, birth defects, impaired immunity, decreased disease resistance, and tumor formation in humans and decreased production in farm animals (CAST, 2003).
With some exceptions, molds can be divided into two main groups: “field fungi” and “storage fungi.” The former group contains species that proliferate in and under field conditions and do not multiply readily once grain is in storage. Field fungi may be superseded and overrun by storage fungi if conditions of moisture and oxygen allow.
Importantly, the presence of a toxigenic mold does not guarantee the presence of a mycotoxin, which is elaborated only under certain conditions. Further, more than one mold can produce the same mycotoxin (eg, both Aspergillus flavus and several Penicillium species produce the mycotoxin cyclopiazonic acid) (El-Banna et al., 1987; Truckness et al., 1987). Also, more than one mycotoxin may be present in an intoxication; that is, as in the outbreak of turkey X disease, there is speculation that aflatoxin and cyclopiazonic acid both exerted an effect, but the profound effects of aflatoxin would have overshadowed those of cyclopiazonic acid (Miller, 1989). Although there are many different mycotoxins and subgroups (Table 31-26), this discussion will be confined largely to four of the more important: aflatoxins, trichothecenes, fumonisins, and ochratoxin A.
Table 31-26Selected Mycotoxins Produced by Various Molds and Some of Their Effects and Commodities Potentially Contaminated ||Download (.pdf) Table 31-26 Selected Mycotoxins Produced by Various Molds and Some of Their Effects and Commodities Potentially Contaminated
|MYCOTOXIN ||SOURCE ||EFFECT ||COMMODITIES CONTAMINATED |
|Aflatoxins B1, B2, G1, G2 ||Aspergillus flavus, A parasiticus ||Acute aflatoxicosis, carcinogenesis ||Corn, peanuts, and others |
|Aflatoxin M1 ||Metabolite of AFB1 ||Hepatotoxicity ||Milk |
|Fumonisins B1, B2. B3, B4, A1, A2 ||Fusarium verticillioides ||Renal and liver carcinogenesis ||Corn |
|Trichothecenes (for example, T-2, deoxynivalenol, diacetoxyscirpenol) ||Fusarium, Myrothecium ||Hematopoietic toxicity, meningeal hemorrhage of brain, “nervous” disorder, necrosis of skin, hemorrhage in mucosal epithelia of stomach and intestine, emesis, feed refusal, immune suppression. ||Cereal grains, corn |
|Zearalenones ||Fusarium ||Estrogenic effect ||Corn, grain |
|Cyclopiazonic acid ||Aspergillus, Penicillium ||Muscle, liver, and splenic toxicity ||Cheese, grains, peanuts |
|Kojic acid ||Aspergillus ||Hepatotoxic? ||Grain, animal feed |
|3-Nitropropionic acid ||Arthrinium sacchari, A saccharicola, A phaeospermum ||Central nervous system impairment ||Sugarcane |
|Citreoviridin ||Penicillium citreoviride, P toxicarium ||Cardiac beriberi ||Rice |
|Cytochalasins E, B, F, H ||Aspergillus and Penicillium ||Cytotoxicity ||Corn, cereal grain |
|Sterigmatocystin ||Aspergillus versiolar ||Carcinogenesis ||Corn |
|Penicillinic acid ||Penicillium cyclopium ||Nephrotoxicity, abortifacient ||Corn, dried beans, grains |
|Rubratoxins A, B ||Penicillium rubrum ||Hepatotoxicity, teratogenic ||Corn |
|Patulin ||Penicillium patulum ||Carcinogenesis, liver damage ||Apple and apple products |
|Ochratoxin ||Aspergillus ochraceus, A carbonarius, Penicillium verrucosum ||Endemic nephropathy, carcinogenesis ||Grains, peanuts, grapes, green coffee |
|Citrinin ||Aspergillus and Penicillium ||Nephrotoxicity ||Cereal grains |
|Penitrem(s) || ||Tremors, incoordination, bloody diarrhea, death ||Moldy cream cheese, English walnuts, hamburger bun, beer |
|Ergot alkaloids ||Clavicepts purpurea ||Ergotism ||Grains |
Organic foods, produced without the use of insecticides and fungicides, may be more susceptible to mycotoxin contamination than foods produced using conventional agricultural practices. For example, higher levels of ochratoxin were found in organic cereal when compared to nonorganic cereal and cereal products in Spain and Portugal (Juan et al., 2008). The UK Food Standards Agency found several organic maize meal products highly contaminated with fumonisin mycotoxins, whereas conventionally produced maize meal products analyzed concurrently had levels below recommended limits (UK Food Standards Agency, 2003). Because European agriculture faces a growing demand by consumers for organic produce, the European Union has established a project called “safe organic vegetables and vegetable products by reducing risk factors and sources of fungal contaminants throughout the production chain” focusing on organic carrots and reduction of alternaria toxins (EU, 2002).
Among the various mycotoxins, the aflatoxins have been the subject of the most intensive research because of the potent hepatocarcinogenicity and toxicity of aflatoxin B1 in rats. Epidemiological studies conducted in Africa and Asia suggest that it is a human hepatocarcinogen, and various other reports have implicated the aflatoxins in incidences of human toxicity (Krishnamachari et al., 1975; Peers et al., 1976; Kensler et al., 2011).
Generally, aflatoxins occur in susceptible crops as mixtures of aflatoxins B1, B2, G1, and G2, with only aflatoxins B1 and G1 demonstrating carcinogenicity. A carcinogenic hydroxylated metabolite of aflatoxin B1 (termed aflatoxin M1) can occur in the milk from dairy cows that consume contaminated feed. Aflatoxins may occur in a number of susceptible commodities and products derived from them, including edible nuts (peanuts, pistachios, almonds, walnuts, pecans, and Brazil nuts), oil seeds (cottonseed and copra), and grains (corn, grain sorghum, and millet) (Stoloff, 1977). In tropical regions, aflatoxin can be produced in unrefrigerated prepared foods. The two major sources of aflatoxin contamination of commodities are field contamination, especially during times of drought and other stresses, which allow insect damage that opens the plant to mold attack, and inadequate storage conditions. Since the discovery of their potential threat to human health, progress has been made in decreasing the level of aflatoxins in specific commodities in developed countries. For example, in the United States and Western European countries, control measures include ensuring adequate storage conditions and careful monitoring of susceptible commodities for aflatoxin level and the banning of lots that exceed the action level for aflatoxin B1.
Aflatoxin B1 is acutely toxic in all species studied, with an LD50 ranging from 0.5 mg/kg for the duckling to 60 mg/kg for the mouse (Wogan, 1973). Death typically results from hepatotoxicity. Aflatoxin B1 is also highly mutagenic, hepatocarcinogenic, and possibly teratogenic. A problem in extrapolating animal data to humans is the extremely wide range of species susceptibility to aflatoxin B1. For instance, whereas aflatoxin B1 appears to be the most hepatocarcinogenic compound known for the rat, the adult mouse is essentially totally resistant to its hepatocarcinogenicity.
Aflatoxin B1 is an extremely reactive compound biologically, altering a number of biochemical systems. The hepatocarcinogenicity of aflatoxin B1 is associated with its biotransformation to a highly reactive electrophilic epoxide, which forms covalent adducts with DNA, RNA, and protein. Damage to DNA is thought to be the initial biochemical lesion resulting in the expression of the pathological tumor growth (IARC, 2002). Species differences in the response to aflatoxin may be due in part to differences in biotransformation and susceptibility to the initial biochemical lesion (Monroe and Eaton, 1987).
Trichothecenes are toxic sesquiterpenoid compounds that have an epoxide functionality that is apparently crucial for their toxicity (Desjardins et al., 1993). They include many different chemical entities that all contain the trichothecene nucleus (Ellison and Kotsonis, 1975) and are produced primarily by Fusarium, but also by a number of commonly occurring molds, including Myrothecium, Trichothecium, Stachybotrys, and Cephalosporium. The trichothecenes were first discovered during attempts to isolate antibiotics, and although some show antibiotic activity, their toxicity has precluded their use as therapeutic agents. Fusarium head blight, caused by trichothecene-producing Fusarium species, is a destructive disease of cereal grain crops that has a worldwide economic impact (Fouroud and Eudes, 2009). Consumption of contaminated grain has been associated with intestinal irritation in mammals and can lead to feed refusal in livestock and other toxic responses (Eriksen and Pettersson, 2004). There have been many reported cases of trichothecene toxicity in farm animals and a few in humans. One of the most famous cases of presumed human toxicity associated with the consumption of trichothecenes occurred in Russia during 1944 around Orenburg, Siberia. Disruption of agriculture caused by World War II resulted in millet, wheat, and barley being overwintered in the field. Consumption of these commodities resulted in vomiting, skin inflammation, diarrhea, and multiple hemorrhages, among other symptoms. About 10% of the population was affected and mortality rates were as high as 60% in some counties (Ueno, 1977; Beardall and Miller, 1994), and was subsequently identified as alimentary toxic aleukia (CAST, 2003). Trichothecenes are protein synthesis inhibitors known to bind to ribosomes. The acute LD50s of the trichothecenes range from 0.5 to 70 mg/kg, and although there have been reports of possible chronic toxicity associated with certain members of this group, more research will be needed before the magnitude of their potential to produce adverse human health effects is understood (Sato and Ueno, 1977). The extent of toxicity associated with the trichothecenes in humans and farm animals is poorly understood, owing in part to the number of entities in this group and the difficulty of assaying for these compounds (JECFA, 2001).
Fumonisins are mycotoxins produced by Fusarium verticillioides (formerly known as F moniliforme) and several other Fusarium species. Corn products contaminated with F verticilliodes are responsible for agriculturally important diseases in horses and swine (ICPS, 2000) and are actively being evaluated to determine how great a threat they pose to public health. Initial evidence of the involvement of F verticilliodes–produced toxins in human disease was reported by Marasas et al. (1988), who found that an increased incidence of esophageal cancer was associated with the consumption of contaminated corn (maize) by humans in a region in South Africa. Fumonisins have been associated with cancer, reproductive toxicity (neural tube defects), and acute disease outbreaks where low-quality corn is consumed on a regular basis (Cousins et al., 2005). Fumonisins target different organs in different species, but the underlying mechanism is a disruption of lipid metabolism by inhibition of ceramide synthetase, an enzyme integral to the formation of complex lipids for use in membranes (ICPS, 2000; IARC, 2002).
Corn borer insect pests cause damage to the developing grain, which enables spores of the toxin-producing fungi, Fusarium, to germinate. The fungus then proliferates, which leads to ear and kernel rot and the production of potentially hazardous levels of fumonisins. Corn varieties, which express the Bt insect control proteins, have been shown to contain significantly reduced levels of fumonisin because the Bt protein significantly reduces the corn borer–induced tissue damage in corn products (Munkvold et al., 1997, 1999; Masoero et al., 1999; Hammond et al., 2004; Papst et al., 2005; Ostry et al., 2010; Folcher et al., 2010).
This mycotoxin is primarily produced by Aspergillus ochraceus, A carbonarius, and Penicillium verrucosum, and human exposure occurs as the result of contamination of small grains (barley, wheat, and corn), coffee beans, and grapes. The effects of ochratoxin A were discovered as the result of feeding the mycotoxin to pigs, who subsequently drank copious amounts of water, urinated near continuously, and exhibited pain in the area of the kidney. Ochratoxin A is nephrotoxic and carcinogenic in mice and rats. Ochratoxin A is absorbed from the GI tract and enters the enterohepatic circulation; it is also absorbed by the proximal and distal tubules of the kidney. It binds tightly to albumin in the blood and can therefore have a very long serum half-life. Epidemiological evidence indicates nearly half the European population is exposed to ochratoxin A and there is an association of endemic nephropathy and renal tumors in humans in parts of Eastern Europe (CAST, 2003).
Ethyl carbamate or urethane, the ethyl ester of carbamic acid, was used for many years as an intravenous anesthetic until its mutagenic and carcinogenic properties became known. It has since been classified by the International Agency for Research on Cancer (IARC) as “possibly carcinogenic to humans” (Group 2B) and “reasonably anticipated to be a human carcinogen” by the NTP (2004a). The primary use of urethane is as a chemical intermediate in the preparation of amino resins, with lesser uses as a solubilizer in the manufacture of pesticides, fumigants, and cosmetics and as an intermediate for pharmaceuticals and biochemical research. It is allowed in some anticonvulsant drugs at a level of 1 ppm and is still used as a veterinary anesthetic. Urethane has been found in fermented foods and beverages including liquor, wine, beer, bread, soy sauce, and yogurt. Diethylpyrocarbonate, an inhibitor of fermentation, can form ethyl carbamate.
Ethyl carbamate is easily absorbed and undergoes CYP2E1-mediated metabolic activation to vinyl carbamate epoxide, which binds covalently to nucleic acids and proteins, producing adducts. Ethyl carbamate is a multisite carcinogen with a short latency period. Single doses or short-term oral dosing at 100 to 200 mg/kg BW/day has been shown to induce tumors in mice, rats, and hamsters. Intake estimates from food and alcoholic beverages range from a mean of 0.015 μg/kg BW/day to 0.080 μg/kg BW/day for high-end users. The benchmark dose lower confidence limit as set by JECFA is 0.3 mg/kg BW/day, which yields a margin of safety of 20,000 (JECFA, 2005).
Fluorine, in the form of fluoride, is nearly ubiquitous in nature. Primary human exposure is via drinking water, although it is also present in some foods, notably some teas, vegetables, and marine fish. Exposure also occurs via processed food made with fluoridated water or produce washed with fluoridated water (the so-called ‘halo’ effect). The greatest nondietary source is fluoridated toothpaste (NRC, 2006). Fluoride taken in water has a high degree of bioavailability with an absorption of 90%, whereas fluoride taken in food is approximately 50% absorbed. Consumption of fluoride results in uptake by bone and teeth, where enamel crystallites form fluorhydroxyapatite in place of the naturally formed hydroxyapatite; the former being stronger and more acid-resistant than the latter, resisting and even reversing the initiation and progression of dental caries. Ionic fluoride rarely exists in blood, most is trapped by bone tissue, where new bone growth is stimulated and this mechanism has served as the basis of some treatments for osteoporosis (WHO, 1996).
Fluorosis occurs as the result of high fluoride intake and may be complicated by low calcium intake; it is cumulative and endemic to some areas of the world (eg, China and India). Fluorosis is dose-responsive, producing a range of effects from cosmetic (mottling of teeth) to adverse functionality (skeletal fluorosis). Enamel fluorosis occurs as the result of high fluoride consumption prior to tooth eruption (ie, in children up to the age of eight years, exposed to water with a fluoride content of ≥4 mg/L) and can range from a mild discoloration of the tooth surface to severe (brown) staining and pitting of the teeth to the point of enamel loss (NRC, 2006). In skeletal fluorosis, in the asymptomatic, preclinical stage, patients have an increased bone density. Stage 1 skeletal fluorosis is characterized by occasional stiffness, pain in joints, or some osteosclerosis of the pelvis and vertebra. Stage 2 skeletal fluorosis is characterized by sporadic pain, stiffness of joints, and osteosclerosis of the pelvis and spine, although mobility is not severely affected. In Stage 3 (rarely seen in the United States), there may be crippling, dose-related calcification of ligaments, osteosclerosis, exotoses, osteoporosis of long bones, muscle wasting, and neurological effects due to hypercalcification of vertebrae (at this point, bone ash fluoride may be two to three times that of the bones of normal subjects). Although it is agreed that skeletal fluorosis is the result of prolonged exposure to increased amounts of fluoride, because the incidence of crippling skeletal fibrosis continues to be rare even in geographic areas of high exposure, unidentified intervening metabolic or dietary factors may have rendered skeletons more or less susceptible. Other effects attributed to excess fluoride include lower IQs and decreased thyroid function, increased calcitonin activity, increased parathyroid hormone activity, secondary hyperparathyroidism, impaired glucose tolerance, and possible effects on timing of sexual activity (NRC, 2006). The reports of possible carcinogenic effects of fluoride, including those of the bone (osteosarcoma), are tentative and mixed (NRC, 2006; Douglass and Joshipura, 2006).
Guidelines for fluoridation of the public water supply recommend addition at levels of 0.7 to 1.2 mg/L, to achieve target Adequate Intake (AI) levels based on a 2 L water/day intake by adults, with adjustments in warmer regions where water intake is high in the summer months, or where fluoride occurs naturally at high levels (eg, some areas of Colorado, 11.2 mg/L; Oklahoma, 12.0 mg/L; New Mexico, 13.0 mg/L; and Idaho, 15.9 mg/L). Although the essentiality of fluoride has not been described, an AI has been established for various age groups as a balance between caries resistance and possible fluorosis of teeth. For example, the AI for infants is 0.01 mg/day, for adult females and males is 3 and 4 mg/d, respectively, and there is a range of graduated AIs for intervening age groups (IOM, 1997).
Toxins in Fish, Shellfish, and Turtles
There are a number of marine (seafood) toxins (to be distinguished from marine venoms), many of which are not confined to a single species (over 400 species have been associated with ciguatera toxicity). Some of these toxins occur with sporadic frequency and nonpredictability, indicating an environmental influence and can often be traced to the presence of an algae (including dinoflagellates) or commensurate bacteria. However, some marine toxins appear to be specific to a single genus or species and are therefore innate to that taxonomic group.
The “cigua” in ciguatera toxin is derived from the Spanish name for the sea snail Turbo pica in which the symptoms were first reported. Ciguatera and related toxins (scaritoxin and maitotoxin) are ichthyosarcotoxic neurotoxins (anticholinesterase) and are found in 11 orders, 57 families, and over 400 species of fish as well as in oysters and clams. The penultimate toxin (gambiertoxin) is produced by the dinoflagellate Gambierdiscus toxicus, commonly isolated from microalgae growing on or near coral reefs that have ingested the dinoflagellate. The pretoxin appears to pass through the food chain and is biotransformed upon transfer to or by the ingesting fish to the active form, which is consumed by mammals. Other toxins, including palytoxin and okadaic acid, unrelated to gambiertoxin, may be present in ciguarteric fish and may contribute to toxicity. The asymptomatic period is three to five hours after consumption but may last up to 24 hours. The onset is sudden and symptoms may include abdominal pain, nausea, vomiting, and watery diarrhea; muscular aches; tingling and numbness of the lips, tongue, and throat; a metallic taste; temporary blindness; and paralysis. Deaths have occurred. Recovery usually occurs within 24 hours, but tingling may continue for a week or more. The intraperitoneal (i.p.) LD50 of maitotoxin in mice is 50 ng/kg (Bryan, 1984; Liston, 2000).
Palytoxin is produced by the zoanthid soft coral of the genus Palythoa, and fish, crabs, and polychaete worms, living in close association with or eating this mass, may become contaminated with palytoxin. The toxin is not part of the stinging nematocyst of the coral, but may be produced by female polyps and mature eggs of the organism, possibly requiring the presence of symbiotic algae (possibly the dinoflagellate Ostreopsis siamensis). Palytoxin, in various forms, is produced by any number of species, including P tuberculosa in the tropical waters in the Pacific and Japan, P mammilosa and P caribaeorum in the West Indies and Puerto Rico, and in the Bahamas, P vestitus and other Palythoa spp. On occasion, the coral becomes detached from its anchorage and becomes a soft floating mass with a seaweed or moss-like appearance, a very attractive feeding ground for fish. Indigenous peoples of Hawaii knew this as limu-make-o-Hana (the deadly seaweed of Hana) and some are said to have smeared the moss on spear points to enhance their utility as a weapon (Onuma et al., 1999; Tan and Lau, 2000; Tosteson, 2000).
The toxin has been reported in mackerel, parrotfish, and several species of crabs. Victims report a bitter, metallic taste from the meat (most often muscle, liver, ovary, and digestive tract), followed immediately by nausea, vomiting, and diarrhea. Within several hours, symptoms include myoglobinuria, a burning sensation around the mouth and extremities, muscle spasms, dyspnea, and dysphonia. Cause of death may be the result of myocardial injury, although it is known in vitro to be a powerful hemolysin.
Although there are several isoforms and possibly minor toxins associated with palytoxin (depending on the producing species), the predominant action is as a ouabain-sensitive Na+K+ ATPase inhibitor. Unlike ouabain, palytoxin has no effect on H+-, Ca2+-, or H+/K+-transporting ATPases. The toxin is quite effective with intravenous LD50 of 0.078, 0.45, 0.033, and 0.089 μg/kg for monkeys, mice, dogs, and rats, respectively. The standard assay is measured in mouse units (MU), the time taken to kill a mouse weighting 20 grams in four hours following intraperitoneal (i.p.) injection of 0.25 mL (Tan and Lau, 2000; Tosteson, 2000).
Abalone Poisoning (Pyropheophorbide)
Abalone poisoning is caused by abalone viscera poison (located in the liver and digestive gland) of the Japanese abalone, Haliotis discus and H sieboldi, and is unusual in that it causes photosensitization; Hashimoto et al. (2010) report the toxin was also found in the midgut glands of cultured scallops (Patinopecten yessoensis) gathered in early spring in Japan. The toxin, pyropheophorbide a, is stable to boiling, freezing, and salting. The development of symptoms is contingent on exposure to sunlight. The symptoms are of sudden onset and include a burning and stinging sensation over the entire body, a prickling sensation, itching, erythema, edema, and skin ulceration on parts of the body exposed to sunlight (Bryan, 1984; Shiomi, 1999). Paralytic shellfish toxin (PST) has been detected in abalone, probably through consumption of the mossworm, a plankton feeder that also clings to seaweed and some shellfish (Takatani et al., 1997).
Pheophorbide b ethyl ester has been isolated from a Chlorella vulgaris dietary supplement (Chee et al., 2008), and pheophorbide a and phyloerythrin were putatively identified as the causative substance in photosensitization and erythematopurpuric eruptions on the skin of patients with a history of consuming chlorella dietary supplements (Jitsukawa et al., 1984). Pyropheophorbide, pheophorbide, and similar phototoxic agents are breakdown products of chlorophyll.
Dinoflagellate Poisoning (Paralytic Shellfish Poisoning or PSP; Saxitoxin)
The etiological agent in dinoflagellate poisoning is saxitoxin or related compounds and is found in mussels, cockles, clams or soft shell clams, butter clams, scallops, and shellfish broth. Bivalve mussels are the most common vehicles. Saxitoxin, originally isolated from toxic Alaskan butter clams (Saxidomus giganteus) is actually a family of neurotoxins and includes neosaxitin and gonyautoxin one through four. All block neural transmission at the neuromuscular junction by binding to the surface of the sodium channels and interrupting the flow of Na+ ions; apical vesicles (AV) nodal conduction may be suppressed, there may be direct suppression of the respiratory center and progressive reduction of peripheral nerve excitability. Saxitoxin produces parathesia and neuromuscular weakness without hypotension and lacks the emetic and hypothermic action of tetrodotoxin. Moderate symptoms are produced by 120 to 180 μg/person and are reversible within hours or days, whereas 80 μg of purified toxin per 100 g of tissue (0.5–2 mg/person) may be lethal, due to asphyxiation, usually within 12 hours of ingestion. The toxin is an alkaloid and is relatively heat-stable. The toxin is produced by several genera of plankton (Gonyaulax [now known as Alexandrium] catenella, G acatenella, and G tamarensis, Pyrodinium spp, Ptychodiscus brevis, Gymnodinium catenatum, and others), and during red tide blooms may reach 20 to 40 million/mL. Toxic materials are stored in various parts of the body of shellfish. Digestive organs, liver, gills, and siphons contain the greatest concentrations of poison during the warmer months. Distribution is worldwide (Bryan, 1984; Clark et al., 1999; Liston, 2000). The tolerance for Paralytic Shellfish Poisoning (PSP) for clams, mussels, and oysters is 80 μg/100 g meat (Compliance Policy Guideline, 540.250).
Neurotoxic Shellfish Poisoning
Traditionally limited to the coast of Florida, Gymnodinium breve form red tide blooms containing polycyclic ether toxins called brevetoxins (based on the backbone structure of the molecule, they are generally divided into Type 1 or Type 2), with Type 2 the most often found. Brevetoxins bind to voltage-dependent sodium channels and strength of binding varies with the specific affinity of the toxin and thus the relative potency. Symptoms of Neurotoxic Shellfish Poisoning (NSP) include nausea, tingling, and numbness of the oral area, loss of motor control, and severe muscular ache, all of which resolve in a few days and no deaths have been reported, unlike PSP. An additional route of entry for mammals may result from inhalation of aerosolized toxin as the result of the relative ease of lysis of the unarmored G breve organism during the breaking of waves on the shore. Symptoms of this type of exposure are seen as irritation of the throat and upper respiratory tract. A “kill” of nearly 150 manatees was reported during an unprecedented large outbreak of the toxin, although the specific mode of transmission is uncertain. Human exposure is primarily via consumption of filter-feeding organisms, which may concentrate the toxin (Van Dolah, 2000). An FDA Action Level for NSP in food has been set at 0.8 ppm brevetoxin-2 equivalent (SNIC, 2007).
Amnesic Shellfish Poisoning (Domoic Acid)
Consumption of mussels harvested from the area off Prince Edward Island in 1987 resulted in gastroenteritis, and many older consumers or those with underlying chronic diseases experienced neurological symptoms including memory loss. Despite treatment, three patients (71–84 years old) died within 11 to 24 days. The poisoning was attributed to domoic acid produced by the diatom Nitzschia pungens f multiseries (now called Pseudonitzschia multiseries), which had been ingested by the mussels during the normal course of feeding. Occurrence of domoic acid has also been reported in California shellfish and produced by Nitzchia pseudodelicatissima and in anchovies (resulting in pelican deaths) produced by N pseudoseriata (now called Pseudonitzchia australis). Domoic acid has been reported in shellfish in other provinces of Canada, and in the United States, in Alaska, Washington, and Oregon and may be as frequent as PSP toxins. Domoic acid has also been reported in seaweed. Domoic acid was reported in Japan in 1958 and was isolated from the red algae, Chondria armata.
In the Canadian outbreak, mice injected with extracts (as in the PSP assay) died within 3.5 hours. The mice exhibited a scratching syndrome uniquely characteristic of domoic acid that was followed by increasingly uncoordinated movements and seizures until the mice died. Levels of domoic acid >40 μg/g wet weight of mussel meat caused the mouse symptoms (Canadian authorities require cessation of harvesting when levels approach 20 μg/g.). Mice and rats can generally tolerate 30 to 50 mg/kg. Domoic acid is dose-responsive in humans, with no effect at 0.2 to 0.3 mg/kg, mild GI symptoms at 0.9 to 2.0 mg/kg, and the most serious symptoms at 1.9 to 4.2 mg/kg with GI effects and neurological effects, including dizziness, disorientation, lethargy, seizures, and permanent loss of short-term memory. Although rodents appear to be more tolerant, the fatalities in humans were likely associated with underlying illness. Domoic acid is an analog of glutamine, a neurotransmitter, and of kainic acid; the toxicity of all three is similar, as they are excitatory and act on three types of receptors in the CNS with the hippocampus being the most sensitive. Domoic acid may be a more potent activator of kainic acid receptors than kainic acid itself. The stimulatory action may lead to extensive damage of the hippocampus, but less severe injury to the thalamic and forebrain regions (Todd, 1993; Clark et al, 1999; Van Dolah, 2000). An FDA Action Level for amnesic shellfish poisoning has been set at 20-ppm domoic acid, except in the viscera of Dungeness crab, where 30 ppm is permitted (SNIC, 2007).
There are naturally occurring toxins in some species that do not involve marine algae or other environmental influences, but are innate to the particular marine species. The first example is Escolar (Lepidocybium flavobrunneum), and Oilfish or Cocco (Ruvettus pretiosus), a marine fish of the snake mackerel family, which are sometimes sold under the category of “butterfish,” and contain a strong purgative oil, that when consumed can cause diarrhea known as gempylid fish poisoning, gempylotoxism, or keriorrhea (FDA, 2010a). The toxin consists of wax esters (C32, C34, C36, and C38 fatty acid esters), the primary component of which is C34H66O2 (Ukishima et al., 1987); these constitute a substantive portion of the lipid present in these fish (14%–25% by weight). Escolar oil contains >90% wax esters (Nichols et al., 2001). Ingestion of fish containing wax esters in large amounts, coupled with their indigestibility and low melting point, results in diarrhea (Berman et al., 1981). No tolerances have been established, and the FDA recommends avoidance of these fish (Dolan et al., 2010; FDA, 2010a).
A second example of an innate toxin is tetramine found in the salivary glands of Buccinum, Busycon, or Neptunia spp, a type of whelk or sea snail that is distributed in temperate and tropic waters and has long been a food source for humans. These whelks are associated with a heat-stable neurotoxin, tetramine, which upon ingestion by humans causes, among other symptoms, eyeball pain, headache, dizziness, abdominal pain, ataxia, tingling in the fingers, nausea, and diarrhea (Reid et al., 1988; Kim et al., 2009). Power et al. (2002) report that the highest concentration of tetramine is in the salivary gland (up to 6530 μg/g), but varies according to season. Reid et al. (1988) reported levels of 37.5 μg tetramine/g of salivary gland tissue (Reid et al, 1988). Because the whelk is a predator of bivalves, it is assumed the toxin is used for food procurement (Power et al., 2002). Although the FDA recommends removal of the salivary gland to avoid possible intoxication (FDA, 2010b), tetramine is present in other tissues, albeit at lesser concentrations (Anthoni et al., 1989; Dolan et al., 2010).
The third and last example is the meat of the Greenland shark (Somniosus microcephalus) and the related member of the dogfish family, the pacific sleeper shark (Somniosus pacificus), is known to be poisonous to both man and dogs. The causative agent is trimethylamine oxide, which breaks down to trimethylamine in the gut, probably by enteric bacteria. The result is absorption of trimethylamine, which acts as a neurotoxin, producing ataxia in both humans and dogs. However, the flesh may be consumed if boiled several times with changes of water, or as the Inuit people prepare it, by burying it in the ground and allowing the meat to go through several freezing and thawing cycles (Anthoni et al., 1991; Benz et al., 2004; Idboro, 2008; Dolan et al., 2010).
Microbiological Agents—Preformed Bacterial Toxins
Although the United States likely has the safest and cleanest food supply in the world, most food-related illnesses in the United States result from microbial contamination. Food-borne disease outbreaks are tracked by the Centers for Disease Control and Prevention (CDC) in Atlanta, Georgia. The CDC reports that there are approximately 400 outbreaks of food-borne disease per year involving 10,000 to 20,000 people. However, the actual frequency may be as much as 10 to 200 times as high because (1) an outbreak is classified as such only when the source can be identified as affecting two or more people and (2) most home poisonings are mild or have a long incubation time and are therefore not connected to the ingested food, go unreported, and are often felt to be only a “24-hour bug.” Naturally, because of differences in virulence and opportunity, some species are more likely than others to cause outbreaks.
If all the microbiological food-borne health concerns could be divided into two categories—poisonings and infections—the former would include chemical poisonings and intoxications, which may have a plant, animal, or microbial origin. In the infections category, food acts as a vector for organisms that exhibit their pathogenicity once they have multiplied inside the body. Infections include the two subcategories: enterotoxigenic infections (with the release of toxins following colonization of the GI tract) and invasive infections in which the GI tract is penetrated and the body is invaded by organisms.
There are a number of food toxins of microbial origin; however, discussion in this chapter will be limited to preformed bacterial toxins—that is, those toxins elaborated by bacteria concomitant to their residence and growth in or on the food prior to ingestion. Importantly, the bacteria need not be present for the intoxication to take place because the bacteria may have been killed by heat while the toxin survives. Bacterial toxins may be divided on the basis of activity: emetic toxins (ie, Bacillus cereus), which produce their effect by binding to specific receptors in the duodenum, neurotoxins (whose action is self-explanatory), and enterotoxins, which are protein toxins having action on the enteric cells of the intestine. Enterotoxins can be subdivided into cytotoxic enterotoxins which disrupt the cell membrane or other vital functions of the cell and cytotonic enterotoxins, which enter the epithelial cell and cause diarrhea without direct membrane disruption or cell death (Granum, 2006). Bacterial toxins may also be divided on the basis of their origin: an endotoxin is generally a lipopolysaccharide membrane constituent released from a dead or dying Gram-negative bacteria, and these toxins are nonspecific and stimulate inflammatory responses from macrophages including, but not limited to prostaglandins, thromboxanes, interleukins, and other mediators of immunity; exotoxins, which are synthesized and released (usually by Gram-positive bacteria) and are not an integral part of the organism, but may enhance its virulence. Some bacteria, such as Shigella spp, Staphylococcus aureus, or Escherichia coli, can elaborate both endotoxin and exotoxin.
Clostridium botulinum, C butyricum, and C baratti
Food botulism rarely causes illness because the confluence of conditions required for its occurrence—Clostridia in the presence of low acidity, high water activity, absence of preservatives, ambient temperature, and anaerobic environment—such a combination rarely occurs in foods, but botulinum poisoning remains important, the result of its potency (Sobel et al., 2004). All Clostridia are Gram-positive, spore-forming anaerobes. Botulism is a product of the toxins: A (the predominant form in the United States), B (the predominant form in Europe), E (the predominant form in Northern latitudes), and F that may be produced by one or more strains of C botulinum, C butyricum (Type E only), and C baratti; toxins C and D cause botulism in animals. Type G has not caused any human cases. C botulinum toxins are categorized as Group I to IV on the basis of toxin produced; additionally, Group I is proteolytic in culture (liquefying egg white, gelatin, and other solid proteins). The toxin is elaborated in foods, wounds, and infant gut and is neurotoxic, interfering with acetylcholine at peripheral nerve endings. Botulinum neurotoxins induce blockage of voluntary motor and autonomic cholinergic neuromuscular junctions, which prevent motor fiber stimulation. Clinical illness is characterized by cranial nerve palsies, followed by descending flaccid muscle paralysis, which can involve the muscles of respiration. Although ptosis and dysarthria may be mistaken for signs of encephalopathy, patients are fully alert, and the results of a sensory examination are normal. Recovery often takes weeks to months (Sobel et al., 2004). Although the spores are among the most heat-resistant, the toxins are heat-labile (the toxin may be rendered harmless at 80°C–100°C for five–10 minutes). Botulinum toxins are large zinc metalloproteins of ~150,000 Da, composed of two parts, a 50,000-Da piece, the catalytic subunit, and the 100,000-Da piece containing an N-terminal translocation domain and a C-terminal binding domain. The structural features are similar to tetanus toxin. For Types B, D, F, and G (and tetanus toxin), the target protein is vesicle-associated membrane protein (VAMP/synaptobrevin), a protein associated with the synaptic vesicle. Types A and E cleave a protein associated with the presynaptic membrane, ANAP25. Botulinum toxin C cleaves SNAP25 and syntaxin, another protein involved in exocytosis. Although intracellular mechanisms of botulinum and tetanus toxins are similar, symptoms are different because different populations of neurons are targeted. The symptoms may include respiratory distress and respiratory paralysis that may persist for six to eight months. The case fatality rate in the United States is 4% (Sobel et al., 2004) and the poison is fatal in three to 10 days; a lethal dose is approximately 1 ng.
Current methods for detecting botulinum toxin include a mouse bioassay and an enzyme-linked immunosorbent assay. The mouse bioassay is the accepted standard, where the mouse is injected with a lethal dose, the signs of which should develop in eight hours and, if not, the mouse is observed for four days. The mouse bioassay can also be used to differentiate between the toxin types by mixing neutralizing antibodies with the sample, prior to injection. Determining which mice survive following specific combinations of toxin and antisera, determines the specific toxin type. The absolute amount of toxin detected in the mouse bioassay is not well defined but is thought to be 10 to 20 pg/mL for type A (Barr et al., 2005).
Sources and reservoirs for Clostridia include soil, mud, water, and the intestinal tracts of animals. Foods associated with botulinum toxin include improperly canned low-acid foods (green beans, corn, beets, asparagus, chili peppers, mushrooms, spinach, figs, baked potato, cheese sauce, beef stew, olives, and tuna). The toxin also may occur in smoked fish, fermented food (seal flippers, salmon eggs), and improperly home-cured hams. An increasing source of poisonings is from the use of flavored oils or oil infusion, most typically in garlic-in-oil preparations. In 1993, FDA required acidification of such preparations to prevent the growth of Clostridia (FDA, 2005a). Whereas a proteolytic strain of C botulinum (Group I) may cause the food to appear and smell “spoiled” (by-products include isobutyric acid, isovaleric acid, and phenylpropionic acid), this is not the case with nonproteolytic strains, many of which can flourish and elaborate toxin at temperatures as low as 3°C (Loving, 1998; Belitz and Grosch, 1999; Crane, 1999; Lund and Peck, 2000).
The successful use of nitrates in meat to prevent spoilage by C botulinum resulted in the petitioning of FDA by the USDA to have sodium and potassium nitrate approved for use by “prior sanction” (21 CFR 181.33). The mechanism of nitrates is believed to be due to an inactivation by nitric oxide of iron-sulfur proteins such as ferrodoxin and pyruvate oxidoreductase within the germinated cells. The activity is dependent on the pH and is proportional to the level of free HNO2; 100 mg nitrate/kg of meat is necessary for the antimicrobial effect, although this effect can be enhanced with ascorbates and chelating agents. Other antibacterials that prevent C botulinum include nisin (used in cheese spreads), parabens, phenolic antioxidants, polyphosphates, and carbon dioxide (Belitz and Grosch, 1999; Lund and Peck, 2000).
Unlike C botulinum, the primary reservoir for C perfringens is the intestinal tract of warm-blooded animals (including humans). Most incidences of C perfringens food poisoning are associated with the consumption of roasted meat that has been contaminated with intestinal contents at slaughter, followed by roasting and inadequate storage, allowing C perfringens growth and enterotoxin Clostridium perfringens enterotoxin (CPE) to be elaborated (although some CPE may actually be released during a “second-sporulation” process in the stomach of the victim). Virtually all food poisoning is produced by type A strain, although a particularly severe form (a necrotic enteritis called “pig-bel” among indigenous peoples of the New Guinea highlands or in Germany known as “Darmbrand”) is produced by type C strain, which has a mortality rate of 15% to 25% even with treatment. The toxin is normally trypsin-sensitive, but people with low intakes of protein or who consume trypsin-inactivating foods (eg, sweet potatoes) are more at risk than carnivorous people with normal trypsin levels (Granum and Brynestad, 1999; Granum, 2006).
CPE is enterotoxic and follows an ordered series of events, first causing cellular ion permeability, followed by macromolecular (DNA, RNA) synthesis inhibition, morphological alteration, cell lysis, and villi tip desquamation and severe fluid loss. This is manifested by abdominal cramping, and diarrhea occurs within eight to 16 hours, although symptoms are of short duration, one day or less. Foods associated with C perfringens poisoning include cooked meat or poultry, gravy, stew, and meat pies. C perfringens is also associated with the production of another 11 toxins, including those associated with gas gangrene (Hobbs et al., 1953; Hauschild, 1971; Walker, 1975; Hobbs, 1976; Crane, 1999; Labbe, 2000).
Bacillus cereus is also a Gram-positive, spore-forming rod, but is an aerobe or facultative anaerobe. Bacillus cereus is a causative agent of emetic or diarrheagenic exo- and enterotoxins elaborated in food. The emetic thermostable toxin (surviving 259°F for 90 minutes) is called cerulide (a small cyclic peptide, 1.2 kDa, that acts on 5-HT3 receptors stimulating the vagus afferent nerve) and is produced by serotypes 1, 3, and 8; it is also resistant to pH and proteolysis, but is not antigenic. The diarrheagenic thermolabile toxin (surviving 133°F for 20 minutes) is produced by serotypes 1, 2, 6, 8, 10, and 19 and may also be produced in situ in the lower intestine of the host. The diarrheal form may actually consist of three toxins, one of which is hemolytic (Granum, 2006). Reservoirs are soil and dust. Foods associated with this organism and its toxic properties include boiled and fried rice (principally the emetic form), while the diarrheal form has a wider occurrence and may be found in meats, stews, pudding, sauces, dairy products, vegetable dishes, soups, and meat loaf (Goepfert et al., 1972; Gilbert, 1979; Bryan, 1984; Crane, 1999; Granum and Lund, 1997). The foods associated with the two types somewhat reflect the geographic distribution of the types, as the emetic type predominates in Japan, whereas in North America and Europe, the diarrhea type is most often seen.
Evidence is accumulating that other species of Bacillus may elaborate food toxins, including Bt, B subtilis, B licheniformis, and B pumilis (Crane, 1999; Granum and Baird-Parker, 2000; Granum, 2006). A notable exception is B cereus var toyoi, a naturally occurring, nontoxigenic, and nonpathogenic strain. B toyoi has been tested in a variety of systems, including conventional toxicity studies and tests for enterotoxicity and genotoxicity, and determined to be safe for its intended use in animal feed as a probiotic to promote digestive health (Williams et al., 2009). B toyoi has been approved for addition to swine, bovine, poultry, and rabbit feed in the European Communities.
Staphylococcal intoxication includes staphyloenterotoxicosis and staphylococcus food poisoning. S aureus produces a wide variety of exoproteins, including toxic shock syndrome toxin-1 (TSST-1), the exfoliative toxins ETA and ETB, leukocidin, and the staphylococcal enterotoxins (SE) (SEA, SEB, SECn,17 SED, SEE, SEG, SHE, and SEI). TSST-1 and the SE are also known as pyrogenic toxin superantigens on the basis of their biological characteristics. There is a relatively wide degree of molecular diversity among SE toxins and this is thought to be the result of adaptation to allow for a broad range of potential hosts (Monday and Bohach, 1999). Some, but not all, SE require Zn++ for superantigen activity. Although enterotoxemia only develops from ingestion of large amounts of SE, emesis is produced as the result of stimulation of the putative SE receptors in the abdominal viscera, followed by a cascade of inflammatory mediator release. All the SE toxins share a number of properties: an ability to cause emesis and gastroenteritis in primates, superantigenicity, intermediate resistance to heat and pepsin digestion, and tertiary structural similarity, including an intramolecular disulfide bond. Induction of emesis separates the SE toxins from TSST-1, but the induction of emesis is not directly correlated to superantigen activity (Granum, 2006). The exact link between superantigenicity and lethality by the SE toxins and TSST-1 is not known, but may be dependent upon cytotoxicity for certain cells, possibly in the kidneys, liver, or vascular endothelium (Monday and Bohach, 1999). Sources of Staphylococcus include nose and throat discharges, hands and skin, infected cuts, wounds, burns, boils, pimples, acne, and feces. The anterior nares of humans are the primary reservoirs. Other reservoirs include mastitic udders of cows and ewes (responsible for contamination of unpasteurized milk) and arthritic and bruised tissues of poultry. Foods usually are contaminated after cooking by persons cutting, slicing, chopping, or otherwise handling them, and then keeping them at room temperature for several hours or storing them in large containers. Foods associated with staphylococcal poisoning include cooked ham; meat products, including poultry and dressing; sauces and gravy; cream-filled pastry; potatoes; ham; poultry; fish salads; milk; cheese; bread pudding; and generally high-protein leftover foods (Cohen, 1972; Bryan, 1976, 1984; Minor and Marth, 1976; Crane, 1999; Dinges et al., 2000).
Although E coli does not produce a preformed toxin, it deserves mention because of the overwhelming publicity the emergent strain O157:H7 has received (H and O refer to flagellar antigens and virulence markers). There are four categories of E coli associated with diarrheal disease: enteropathogenic, enterotoxigenic, enteroinvasive, and Vero cytotoxin–producing E coli (VTEC). The classification VTEC also includes “shiga-like toxin”–producing E coli and “shiga toxin”–producing E coli (STEC). Enterohemorrhagic E coli (EHEC) refers to those strains producing bloody diarrhea and are a subset of VTEC. The reference to shiga toxin is the result of the clinical similarity of the bloody diarrhea caused by EHEC to that caused by Shigellae. Each of the diseases presented by the four categories is also associated with one or more toxins (Willshaw et al., 2000). The symptoms of STEC infections vary, but commonly include severe stomach cramps, diarrhea (often bloody), and vomiting, and about 5% to 10% of those diagnosed with STEC infections develop hemolytic uremic syndrome, a potentially life-threatening complication because the kidneys may stop working (CDC, 2010).
Cattle are a significant reservoir of E coli; therefore, it is logical that most outbreaks in the United States have been associated with hamburgers and other beef products, although raw vegetables (often fertilized with manure) and unpasteurized apple cider and juice have been reported as sources of outbreaks. Outbreaks in Europe are more often associated with contamination of recreational waters (swimming pools, lakes, etc). Other sources of contamination include person-to-person contact (especially in families and among institutionalized persons) and contact with farm animals especially following educational farm visits (Karch et al., 1999).
The subject of organic food has increasingly captured the public interest. As a result, numerous studies have been done comparing the nutritional and health benefits of organic and conventional foods. When currently available literature was reviewed, there was no compelling evidence that there are nutrition-related health effects from the consumption of organically produced foods (Dangour et al., 2010, Gueguen and Pascal, 2010).
Within this issue, there is a debate concerning the use of organic fertilizers (eg, cow manure) in organic and conventional farming, which may contain E coli O157:H7 (Stephenson, 1997). Data reported to the US CDC in 1996, and tabulated in a CDC document entitled “Clusters/Outbreaks of E coli O157:H7 reported to CDC in 1996,” show that approximately 10% of all E coli O157:H7 infections reported that year were from organically grown lettuce, although organic foods apparently account for less than 1% of the total food supply.
At the basis of the potential problem is the use of inadequately treated manure for fertilizer. Human cases of E coli O157:H7 infection have been reported from consumption of contaminated lettuce, potatoes, radish sprouts, alfalfa sprouts, cantaloupe, and unpasteurized apple cider and juice (Karch et al., 1999). Adequate treatment of manure requires composting the manure for a minimum of three months during which the heap must reach a temperature of 60°C and although this may be adequate to kill vegetative pathogens, it will not destroy spore-formers such as Clostridium perfringens or C botulinum. Survival of viruses and protozoa during composting is not known (Anonymous, 1999).
Bovine Spongiform Encephalopathy
Bovine spongiform encephalopathy (BSE) was first identified in Great Britain in 1986. BSE is a neurological disease classified as a transmissible spongiform encephalopathy (TSE) and is similar to TSEs in other species including scrapie (sheep and goats), transmissible mink encephalopathy (ranch-bred mink), chronic wasting disease (CWD) (mule deer and elk), exotic ungulate encephalopathy (captive exotic bovoids such as bison, orynx, kudu), and feline spongiform encephalopathy (domestic cats and zoo Felidae). TSEs among humans include kuru, Creutzfeldt–Jakob Disease (CJD) and “new variant” CJD (nvCJD), and Gerstmann–Sträussler–Scheinker syndrome (to be distinguished from CJD by an earlier onset and that it tends to run in families). There is compelling epidemiological and laboratory evidence of a causal association between the BSE outbreak in cattle in Great Britain and the new human prion disease nvCJD (CDC, 2011).
Clinically, these diseases present neurological deterioration and wasting, with the incubation period and interval from clinical onset to inexorable death determined by the dose of infective agent, its virulence, and the genetic makeup of the victim. The incubation of BSE in cattle is generally four to five years (range of 20 months–18 years) and an interval of 1 to 12 months from presentation of clinical signs to death. Characteristic histological lesions in the brain and spinal cord are vacuolation and “spongiform” changes. BSE fibrils (long strands of host glycoprotein called prion protein [PrP]) in spinal cord preparations may be seen with electron microscopy following detergent extraction and proteinase K digestion. BSE/scrapie tissues with highest infectivity are brain and spinal cord, followed by retina, spleen, tonsil lymph nodes, distal ileum, and proximal colon. The infective agent can be transferred using preparations of neural tissue from infected animals across species barriers. The most effective method of transfer is direct injection into the brain or spinal cord, but transfer has been reported with intraperitoneal injection and oral dosing. Vertical transfer (mother to offspring) has been reported among domestic cattle, and lateral transfer through biting or injury (especially among mink) has also been reported. Indirect transmission of CWD has been reported recently (Miller et al., 2004); CWD of mule deer (Odocoileus hemionus) can be transmitted from environments contaminated by excreta 2.2 years earlier or decomposed carcasses ~1.8 years earlier.
It is generally agreed that the infective agent is likely a variant of scrapie (endemic to sheep) and was transferred to cattle from rendered sheep via inadequately processed meat and bone meal protein supplement. There is strong evidence and general agreement that the outbreak was amplified and spread throughout the UK cattle industry by feeding rendered (contaminated) bovine meat- and-bone meal to young calves. Disputes have arisen about other details of BSE and its relationship to other TSEs and effects in humans because of an expectation of conformation by BSE to historical principles of disease transmission. Recently, the human susceptibility to sheep and goat passaged-BSE prions was evaluated using transgenic mice expressing PrP and evidence was provided suggesting that humans might be equally or more susceptible to sheep or goat BSE agent compared to that of cattle (Padilla et al. 2011).
Responsive to concerns about transmission, new enhanced BSE-related feed bans went into effect in Canada in 2007 and in the United States in 2009. The enhanced bans prohibit most proteins, which include potentially BSE infectious tissues from all animal feeds, pet foods, and fertilizers not just from cattle feed as required by bans instituted in 1997 (CDC, 2011).
The currently most accepted theory is that the infective agent is a modified form of a normal cell surface component known as prion protein PrPc (α-helix form), which when introduced into an organism causes a conversion of PrPc into a likeness of itself (ie, the isoform), but then designated as the pathogenic form, PrP* or PrPsc for scrapie or PrPres for protease resistant (β-pleated sheet form) (Flechsig and Weissmann, 2004; Frosch et al., 2005). The agent does not possess nucleic acid. The pathogenic form of the protein, PrP*, is both less soluble and more resistant to enzyme degradation than the normal form. The protein is resistant to heat, antimicrobials, ultraviolet-, or ionizing radiation, and is not consistently inactivated with alcohol, formaldehyde, glutaraldehyde, or sodium hydroxide. Phenol and sodium hypochlorite disinfection have had variable success.
Investigators have concluded that the agent in nvCJD and BSE is the same strain and may be the same agent in feline spongiform encephalopathy and exotic ungulate encephalopathy. Although this information might indicate a simple mode of transmission, workers with the highest potential incidence of exposure to BSE or TSE (sheep farmers, butchers, veterinarians, cooks, and abattoir workers) do not have an unusually high incidence of nvCJD (Prusiner, 1991; Collee, 2000). Likewise, hemophilic patients have not reflected an increased incidence of nvCJD, although CJD transmission has been documented as the result of injections of human growth hormone or gonadotrophin (derived from human pituitary gland), implantation of dura mater and corneas, and even infected electroencephalographic (EEG) electrodes and neurosurgical instruments (Prusiner, 1994; Lee et al., 1998; Collee, 2000).
Substances Produced by Cooking or Processing
Tolerances cannot be set for contaminants that are produced as a result of an action taken by the consumer because the home is out of the jurisdiction of FDA. An example of this type of contaminant is heterocyclic amines (HCAs), which are generated during cooking. HCAs were discovered serendipitously by Japanese investigators who, while examining the mutagenicity of smoke generated by charred foods, found that the extracts of the charred surfaces of the meat and fish were quantitatively more mutagenic than could be accounted for by the presence of polycyclic aromatic hydrocarbons (Sugimura et al., 1989). Collectively, there are more than 20 HCAs. They are formed as a result of high-temperature cooking of proteins (especially those containing high levels of creatinine) and carbohydrates. Normally, as a result of such heating, desirable flavor components are formed, for example, pyrazines, pyridines, and thiazoles. Intermediates in the formation of these substances are dihydropyrazines and dihydropyridines, which in the presence of oxygen form the flavor components; however, in the presence of creatinine, HCAs are formed (Table 31-27) (Chen and Chiu, 1998; Schut and Snyderwine, 1999).
Table 31-27Amounts of Heterocyclic Amines in Cooked Foods ||Download (.pdf) Table 31-27 Amounts of Heterocyclic Amines in Cooked Foods
| ||AMOUNT (ng/g) IN COOKED FOOD |
|SAMPLE ||IQ ||MeIQx ||4,8-DiMeIQx ||Trp-P-1 ||Trp-P-2 |
|Broiled beef ||0.19 ||2.11 || ||0.21 ||0.25 |
|Fried ground beef ||0.70 ||0.64 ||0.12 ||0.19 ||0.21 |
|Broiled chicken || ||2.33 ||0.81 ||0.12 ||0.18 |
|Broiled mutton || ||1.01 ||0.67 || ||0.15 |
|Food-grade beef extract || ||3.10 || || || |
These substances are rapidly absorbed by the GI tract, are distributed to all organs, and decline to undetectable levels within 72 hours. HCAs behave as electrophilic carcinogens (Table 31-28). They are metabolized first by N-hydroxylation followed by further activation by O-acetylation or O-sulfonation to react with DNA. DNA adducts are formed with guanosine in various organs, including the liver, heart, kidney, colon, small intestine, forestomach, pancreas, and lung. Unreacted substances are subject to phase II detoxication reactions and are excreted via the urine and feces. In vitro, HCAs require metabolic activation, with some requiring O-acetyltransferase and others not. Although much of the mutagenicity testing has been carried out in TA98 and TA100, these substances are mutagenic in mammalian cells both in vitro and in vivo, Drosophila, and other strains of Salmonella (Munro et al., 1993; Skog et al., 1998; Sugimura and Wakabayashi, 1999).
Table 31-28Mutagenicity and Carcinogenicity of Heterocyclic Amines ||Download (.pdf) Table 31-28 Mutagenicity and Carcinogenicity of Heterocyclic Amines
| || ||CARCINOGENICITY |
|HCA ||NUMBER OF REVERTANTS n/g (STRAIN TA98) ||SPECIES ||STATISTICALLY SIGNIFICANT TUMORS |
|MeIQ ||47,000,000 ||Mouse ||Liver, forestomach |
| || ||Rat ||Zymbal gland, oral cavity, colon, skin, mammary gland |
|IQ ||898,000 ||Mouse ||Liver, forestomach, lung |
| || ||Rat ||Liver, mammary gland, Zymbal gland |
| || ||Monkey ||Liver, metastasis to lungs |
|MeIQx ||417,000 ||Mouse ||Liver, lung, lymphoma, leukemia |
| || ||Rat ||Liver, Zymbal gland, clitoral gland, skin |
|Glu-P-1 ||183,000 ||Mouse ||Liver, blood vessels |
| || ||Rat ||Liver, small and large intestine, brain, clitoral gland, Zymbal gland |
|Glu-P-2 ||930 ||Mouse ||Liver, blood vessels |
| || ||Rat ||Liver, small and large intestine, Zymbal gland, brain, clitoral gland |
|DiMeIQx ||126,000 ||No data || |
|Trp-P-1 ||8990 ||Mouse ||Liver |
| || ||Rat ||Liver, metastasis to lungs |
|Trp-P-2 ||92,700 ||Mouse ||Liver, lung |
| || ||Rat ||Liver, clitoral gland |
|PhIP ||1800 ||Mouse ||Liver, lung, lymphoma |
| || ||Rat ||Colon, mammary gland |
Prior to 2002, when Swedish investigators detected acrylamide in food (Table 31-29), it was of interest only to specialists in worker safety, as this chemical is an important intermediate in the manufacture of polyacrylamides. Although there are many industrial and manufacturing uses of polyacrylamides, the bulk of production are used as chemical flocculants for water treatment, oil recovery, and in construction of dam foundations, tunnels, and sewers—consumer exposure is largely incidental (NTP, 2004b). End users are exposed to polyacrylamide, which is not toxic as long as the monomer is not present. Acrylamide (the monomer) was known to be a neurotoxin, creating morphological changes in peripheral nerves at doses as low as 1 mg/kg BW/day. Much more is now known about acrylamide and its primary metabolite, glycidamide, which is produced from acrylamide by the enzyme CYP2E1; both acrylamide and glycidamide will form adducts with hemoglobin. Acrylamide is absorbed rapidly and extensively (23%–48% of the administered dose to rodents) from the GI tract. Both acrylamide and glycidamide are largely eliminated as mercapturic acid conjugates. Repeated dosing of acrylamide (in drinking water at 21 mg/kg BW/day for 40 days) produces morphological changes in the brain areas critical for learning, memory, and other cognitive functions (ie, cerebral cortex, thalamus, and hippocampus) (JECFA, 2005). Acrylamide was classified as “probably carcinogenic to humans” (IARC Group 2A) by the IARC (IARC, 1994) and “reasonably anticipated to be a human carcinogen” by the NTP (2004b).
Table 31-29Representative Concentrations of Acrylamide in Several Foods (JECFA, 2005) ||Download (.pdf) Table 31-29 Representative Concentrations of Acrylamide in Several Foods (JECFA, 2005)
|FOOD ||MEAN CONC. (μg/kg) ||REPORTED MAXIMUM (μg/kg) |
|Cereal-based products || || |
Breads and rolls
Pastry and cookies
| || |
|Roots and tubers || || |
| || |
|Coffee || || |
| || || |
|Vegetables || || |
| || || |
|Infant formula || || |
|Baby food (biscuits) || || |
Acrylamide is formed in foods that are high in carbohydrate, but low in protein, which are subjected to processing temperatures of at least 120°C. These high-temperature processing conditions are largely the same as required for the Maillard reaction, which imparts a toasted, or baked (ie, “crust”), flavor to breads, toast, and other baked goods, breaded meats and vegetables for sautéing or frying, and production of French fries and potato chips. Most acrylamide is formed in the final stages of baking, grilling, or frying as the moisture content of the food falls and the surface temperature rises (JECFA, 2005). The presence of ammonium bicarbonate as a leavening agent increases the formation of acrylamide.
A critical element is the presence of asparagine, and amino acids competing with asparagine in the Maillard reaction, which reduces the levels of acrylamide in the final product. Strategies for mitigation focus on reduction of asparagine through use of asparaginase, breeding and selection of low-asparagine plants, and prolonged yeast fermentation; alternatively, processing temperature could be lowered (JECFA, 2005).
Acrylamide intake estimates range from 0.3 to 2.0 μg/kg BW/day for the average population, with the 90th to 97th percentile at 0.6 to 3.5 μg/kg BW/day and the 99th percentile at up to 5.1 μg/kg BW/day. Primary sources include french fries (1%–30%), potato chips (6%–46%), coffee (13%–39%), pastry and cookies (10%–20%), and bread and rolls/toasts (10%–30%). The NOEL for morphological change in nerves and for reproductive effects is 200 and 2000 μg/kg BW/day, respectively (JECFA, 2005).
Furan was once known only as an industrial chemical intermediate in the synthesis of polymers used to prepare temperature-resistant structural laminates and to prepare copolymers used in machine dishwashing products. Furan has recently been found to occur in a number of foods that undergo heat treatment, such as canned and jarred foods, including baby food. It is considered by IARC (1995) to be possibly carcinogenic to humans. According to the NTP, furan is hepatotoxic and shows clear evidence of carcinogenicity in both sexes and both species of mice and rats (NTP, 1993). Furan is produced in a variety of experimental systems, including heating of sugars (eg, glucose, lactose, fructose, xylose, rhamnose), heating sugars in the presence of amino acids or protein (eg, alanine, cysteine, casein), and thermal degradation of vitamins (ascorbic acid, dehydroascorbic acid, thiamine) (FDA, 2004).
Furan has been found in a small number of heat-treated foods, including coffee, canned meat, baked bread, cooked chicken, sodium caseinate, filberts (hazelnuts), soy protein isolate, hydrolyzed soy protein, rapeseed protein, fish protein concentrate, and caramel (FDA, 2004). Very little information has been developed on furan levels in food.
Miscellaneous Contaminants in Food
Despite normal precautions taken to protect ourselves from known toxins, some toxins have a propensity to appear unexpectedly in unfamiliar places. Cases in point here include honey poisoning, as documented by Xenophon, who when describing the “Retreat of the Ten Thousand” from Asia back to Greece, the soldiers entered an area rich in honeycombs and seized upon the food. Xenophon noted that the soldiers then “went off their heads” appearing to first be drunk, but then in a state a delirium—a description fitting what we know today about grayanotoxins from mountain laurel and other species.
Mountain laurel (Kalima spp), rhododendron, and azaleas (Rhododendron spp) all possess grayanotoxins of which there may be as many as 60, but the most potent are grayanotoxins I and III (Gunduz et al., 2008). The toxins are present in the shoots, leaves, twigs, and flowers. Honey made from flowers of these plants is toxic to humans, and after an asymptomatic period of four to six hours, salivation, malaise, vomiting, diarrhea, tingling of the skin, muscular weakness, headache, visual difficulties, coma, and convulsions occur. Symptoms are proportional to dose, and atropine administration is indicted as a primary treatment modality (Gunduz et al., 2008). Life-threatening bradycardia and arterial hypotension may occur. Needless to say, beekeepers maintain apiaries well away from these species of plants.
A similar poisoning can occur with oleander (Nerium oleander and N indicum), where honey made from the flowers, meat roasted on oleander sticks, or milk from a cow that eats the foliage can produce prostrating symptoms. The oleander toxin consists of a series of cardiac glycosides: thevetin, convallarin, steroidal, helleborein, ouabain, and digitoxin. Sympathetic nerves are paralyzed; the cardiotoxin stimulates the heart muscles similar to the action of digitalis, and gastric distress ensues (Anderson and Sogn, 1984; VonMalottki and Wiechmann, 1996). Although certainly as possible as the story captured by Xenophon, the stories about scouts sickened from roasting hot dogs on oleander sticks have not been documented and are likely apocryphal.
Closer to home and prevalent in the Midwest in the 18th and 19th centuries, was a mysterious scourge called “milk sickness” also known as “puking fever,” “sick stomach,” “the slows,” and “the trembles”; thousands of people have been reported as dying, including Abraham Lincoln’s mother, Nancy Hanks Lincoln. In humans, milk sickness is characterized by loss of appetite, listlessness, weakness, vague pains, muscle stiffness, vomiting, abdominal discomfort, constipation, foul breath, and finally, coma. For many years, the origin of milk sickness was unknown because there was nothing comparable in Europe (origin of most of the pioneers) and the outbreaks were sporadic. It was not recognized until the late 19th and early 20th century, that white snakeroot (Ageratina altissima née Eupatorium rugosum) and rayless goldenrod (Bigelowia spp, Haplopappus heterophyllus, and Isocoma pluriflora), when eaten by cattle, were the source. The sporadic nature of outbreaks became clear when it was realized that cattle would consume these plants in over-grazed pasture or in years of drought; additionally, the toxin levels in plants can vary considerably, making identification of the source of poisonings difficult. Tremetol or tremetone is the toxic agent and consists of a mixture of sterols and derivatives of methyl ketone benzofuran. The three major benzofuran ketones are tremetone, dehydrotremetone, and 3-oxyangeloyl-tremetone (Panter and James, 1990; Lee et al., 2009; NPS, 2010; Dolan et al., 2010).