|The prevalence of food allergy in children younger than 3 years of age can be up to 8% and in adults about 2% (Sampson 1999). The most important allergens of animal origin are milk, egg, fish, and crustaceae. The frequencies of self-reported allergy to milk, egg, and fish in adults representative of the general population were between 0.5 and 0.7% (Table 1). Allergy to milk and egg is most frequent in young children and has a good prognosis of outgrowing in the first six years of life. In contrast fish and shrimp allergy are not likely to be lost and tend to persist or develop in adulthood (Table 2). The most common symptoms of food allergy are gastrointestinal, cutaneous, and respiratory reactions. Anaphylactic reactions to foods are less frequent. However foods of animal origin are commonly seen among foods inducing anaphylactic reactions (Table 3). Moreover fatal cases of anaphylaxis after ingestion of cow's milk, hen's egg, fish, and shrimp have been reported (Yunginger et al. 1988, Sampson et al. 1992, European Commission 1999).||Table 1: Allergy prevalences to foods of animal origin in the general
adult population of Great Britain (16420 men and women,
> 15 years of age, interview survey) (Emmett et al. 1999)
Table 2: Allergy prevalences to foods of animal origin in children
and adolescents with DBPCFC-proven food allergy (Bock
& Atkinson 1990)
Table 3: Frequency of
foods of animal origin in episodes of food-induced anaphylaxis
(Andre et al. 1994)
(Pumphrey & Stanworth 1996)
(Novembre et al. 1998)
(Kemp et al. 1995)
|Adults and Children
|Adults and Children
|Children (<16 years)
|Adults (12-75 years)
|Crustaceae 17%||Eggs 4.4%||Fish 30%||Crustaceae 29%|
|Fish 13%||Cow's milk 3.3%||Cow's milk 22%||Beef 9%|
|Cow's milk 3.3%||Crustaceae 2.2%||Hen's egg 11%||Pork 4.5%|
|Eggs 1.7%||Fish 2.2%||Goat's milk 4%||Eggs 3.4%|
|Ewe's milk 1.7%||Tuna 1.1%|
Table 4 shows the threshold concentrations for elucidation of symptoms
after ingestion of the offending food as determined by double- blind, placebo
controlled food challenge (DBPCFC). Threshold concentrations ranged from
1 mg to 12 g of protein in the cited studies. They are strongly dependent
on the patient's individual susceptibility and the allergenic potency of
the particular food. According to Moneret-Vautrin et al. (1998) 30% of
40 egg allergic individuals reacted to doses of less than 11 mg, another
30% to doses between 11 and 110 mg, and 30% to 110 mg to 780 mg of hen's
egg white protein. The most potent animal allergen sources seem to be hen's
egg and fish. Although allergic reactions to lower amounts can not be excluded,
the lowest dose elucidating allergic symptoms in DBPCFC was 1 mg of codfish
Table 4: Threshold concentrations for elucidation of symptoms in
|Allergen||Dose at first reaction||Amount of protein*||Reference|
|Cow's milk||a) 0.5 - 8 g (dried milk)
b) up to 206 g
|a) 125 mg - 2 g
b) up to 6.6 g
|Bock et al. 1978|
|Cow's milk||5-52 g||165 mg - 1.65 g||Host & Samuelsson 1988|
|Cow's milk||5 - 250 g||165 mg - 8.25 g||Norgaard & Bindslev-Jensen 1992|
|Hen's egg||25 mg - 8 g (dried egg)||12 mg - 4 g||Bock et al. 1978|
|Hen's egg||50 mg - 50 g||6 mg - 6 g||Norgaard & Bindslev-Jensen 1992|
|Codfish||6 mg - 6.7 g||1 mg - 1 g||Hansen & Bindslev-Jensen 1992|
|Shrimps||4 - 64 g **||700 mg - 12 g||Daul et al. 1988|
|Beef||250 mg - 60 g||48 mg - 11.5 g||Werfel et al. 1997|
|Bovine serum albumin||90 mg**||Fiocchi et al. 1995a|
|Ovine serum albumin||63 mg**||Fiocchi et al. 1995a|
Food allergens of animal origin are generally resistant
to extremes of heat, pH and enzymatic degradation. Resistance to denaturation
and degradation during food processing and passage through the digestive
system enables the allergen to either sensitize the individual or to elucidate
an allergic reaction. Only a few allergens are labile and do not survive
processing. Several allergens of animal origin have been identified
and characterized (Table 5), but little is known about their stability
and the allergenicity of processed foods determined under standardized
During food processing the allergenicity can be altered by various procedures such as storage time, prolonged washing, separation techniques, heating, and texturizing. Moreover, various chemical interactions during the food manufacture between natural food ingredients and food additives can occur. The allergenic potential may be unaffected or decreased or even increased by food processing. Physico-chemical methods may simply reduce the allergen content of a specific product by e.g. extraction, precipitation, or ultrafiltration. The molecular basis of allergen alteration is the inactivation or destruction of IgE-binding epitope structures or the formation of new epitopes or better accessability of cryptic epitopes after denaturation of the native allergen. Heat treatment can induce the loss of the tertiary protein structure and induce aggregation of allergens affecting the conformational structure. In contrast, proteolytic or hydrolytic treatments affect the conformational structure as well as the linear amino acid sequence which may destroy sequential IgE-binding epitopes.
The majority of available studies examined the impact
of heating (dry heating, boiling or cooking) and enzymatic digestion on
native foods or allergen extracts of native foods. For recent reviews on
the alteration of allergenicity by food processing see Moneret-Vautrin
1998 and Hefle 1999. This review summarizes available data on the stability
of cow's milk, hen's egg, fish, crustaceae, and meat allergens during food
processing as well as the allergenic potential of food additives of animal
origin such as lecithins and gelatine.
Table 5: Characterized food allergens of animal origin (Larsen
& Lowenstein 2000)
|Animal Source||Species||WHO/IUIS Allergen Nomenclature||Protein name / family||Molecular mass (kDa)|
|Abalone (snail)||Haliotis midae||Hal m 1||Muscle protein||49|
|Cow's milk||Bos domesticus||Bos d 4
Bos d 5
Bos d 6
Bos d 7
Bos d 8
|Codfish||Gadus callarias||Gad c 1||Parvalbumin||12|
|Hen's egg||Gallus domesticus||Gal d 1
Gal d 2
Gal d 3
Gal d 4
Gal d 5
Serum albumin (alpha-Livetin)
|Lobster||Homarus americanus||Hom a 1*||Tropomyosin|
|Lobster||Panulirus stimpsoni||Pan s 1*||Tropomyosin||34|
|Salmon||Salmo salar||Sal s 1||Parvalbumin||12|
|Shrimp||Metapenaeus ensis||Met e 1||Tropomyosin||34|
|Shrimp||Penaeus aztecus||Pen a 1||Tropomyosin||36|
|Shrimp||Parapenaeus fissurus||Par f 1*||Serum albumin||39|
|Shrimp||Penaeus indicus||Pen i 1||Tropomyosin||34|
|Squid||Todarodes pacificus||Tod p 1||Tropomyosin||38|
Several milk proteins have been identified as allergenic in humans (Besler et al. 2000). The major allergens from cow's milk are constituted by caseins (Bos d 8) and whey proteins beta-lactoglobulin (Bos d 5), alpha-lactalbumin (Bos d 4) and serum albumin (Bos d 6) (see Table 5).
Raw Milk, Processed Milk, and Boiled Milk
Pasteurized and homogenized milk are as allergenic as raw milk, while boiling of milk for 10 min results in partial reduction of its allergenicity. Host & Samuelsson (1988) found comparable allergenic potencies of raw milk, pasteurized milk (75°C, 15s) and pasteurized and homogenized milk (60°C, 175kg/cm2) in 5 cow's milk allergic children by skin prick test (SPT) and DBPCFC. They observed no positive reactions to an extensively hydrolysed casein-based infant formula. Pasteurized and homogenized whole milk was also capable of inducing allergic reactions in 3 adults in DBPCFC (Norgaard & Bindslev-Jensen 1992). Amounts of milk elucidating symptoms during DBPCFC range between 5 g and 250 g (Table 4).
A partial reduction of its allergenicity was observed after boiling of milk (100°C) for 10 min in SPT and dot-immunoblotting. In contrast, boiling for 2 and 5 min, respectively, induced no significant alterations (Norgaard et al. 1996, Werfel et al. 1997). Alpha- lactalbumin and caseins showed a 50% and 66% decrease in IgE-binding in scored crossed radio-immunoelectrophoresis (CRIE) after heating of skimmed milk for 10 min, while beta- lactoglobulin and serum albumin lost their IgE-binding potencies completely (Gjesing et al 1982). Norgaard et al. (1996) confirmed the inactivation of beta- lactoglobulin and serum albumin after boiling milk for 10 min, while caseins still induced positive reactions in SPT.
Studies on allergenicity during the complicated manufacture of cheese have not been performed to date. Although there are more than 600 different kinds of cheese, cheese-making usually involves common procedures like pasteurization of milk, acidification by added starter cultures (change milk sugar into lactic acid), and production of curd by adding rennet (mainly chymosin). Hereby the structure of the caseins is changed to form a solid curd called para- casein. The whey fraction including alpha- lactalbumin and beta- lactoglobulin passes out and is usually lost. The ripening process is brought about through enzyme systems produced by bacteria which have grown in the curd.
Several cases of allergic reactions, including anaphylactic reactions after ingestion of different kinds of cheese (hard cheese, soft cheese, like mozarella and parmesan), have been reported (Wüthrich & Hofer 1986, Fiocchi et al. 1999). Goat's and sheep's cheese induced allergic reactions as well (Wüthrich & Johansson 1995, Umpierrez et al. 1999).
Lactic Acid Fermentation
Fermentation of sterilized cow's milk by meso- and thermophile lactic acid cultures decreased the antigenicity of alpha- lactalbumin and beta- lactoglobulin about 99% employing rabbit antibodies (ELISA). In contrast, the allergenicity of the fermented milk product was hardly affected in skin tests (Jedrychowski & Wroblewska 1999).
Hydrolysis with Digestive Enzymes
Purified milk proteins are more susceptible to enzymatic degradation than proteins in "crude" milk samples. In vitro digestions with duodenal fluid and human trypsin and elastase, respectively, showed the highest degradation rate for caseins followed by beta- lactoglobulin and alpha- lactalbumin, which were hydrolyzed at 100 and 500 times lower rates (Jakobsson et al. 1982, 1983).
Schmidt et al. (1995) combined an in vitro hydrolysis of whey proteins with pepsin (90 min) and a subsequent hydrolysis with a mixture of pancreatic enzymes (pH 7.5 for 150 min). Peptic digestions were performed at pH 2, 3 and 4 simulating pH values of the gastric fluids from adults and small children, respectively. Residual IgE-binding activities of whey proteins after hydrolysis at pH 2 and pH 3 were less than 14%. In contrast after digestion at pH 4 alpha- lactalbumin, serum albumin and bovine immunoglobulin still retained 48%, 58% and 91% of IgE-binding activity (RAST inhibition). Beta- lactoglobulin was hardly affected by pepsin hydrolysis, but was almost completely digested by pancreatic enzymes (Schmidt et al. 1995).
Astwood et al. (1996) found a high stability (>60min) of beta- lactoglobulin against peptic hydrolysis (pH 1.2), while caseins and serum albumin were completely hydrolyzed after 2 min and 30 s, respectively.
Hypoallergenic Infant Formulas
Infant formulas produced by enzymatic hydrolysis of cow's milk proteins are commonly used as milk substitutes in infant nutrition. Hydrolyzed formulas can be classified as either partially hydrolyzed or extensively hydrolyzed. Although the partially hydrolyzed cow's milk is reduced in allergenicity, it still contains full- length allergen and large allergen fragments capable of binding IgE. Therefore only extensively hydrolyzed formulas should be applied in the nutrition of cow's milk allergic infants (Bousquet et al. 1998). Hydrolyzed formulas are produced from caseins or whey proteins by heat denaturation and enzymatic hydrolysis, sometimes combined with ultrafiltration. Extensively hydrolyzed whey protein formulas (Schwartz et al. 1991, Halken et al. 1993) as well as extensively hydrolyzed casein formulas (Sampson et al. 1991) are well tolerated by most cow's milk allergic children (immunoblot, RAST, DBPCFC). Nevertheless, severe adverse reactions have been observed in a few patients. Anaphylactic reactions occurred either after ingestion of extensively hydrolyzed casein formulas (Lifschitz et al. 1988, Amonette et al. 1991, Saylor & Bahna 1991) and extensively hydrolyzed whey formulas (Businco et al. 1989).
In a study with 20 cow's milk allergic children 45% did not tolerate a partially hydrolyzed whey formula, while 10% and 13% did not tolerate extensively hydrolyzed formulas produced from caseins and whey proteins, respectively (Ragno et al. 1993).
Comparing the allergenicity in RAST inhibition and skin tests casein formulas were less allergenic than whey formulas. Of 45 cow's milk allergic children 15% reacted to extensively hydrolyzed whey formula and 2.5% to extensively hydrolyzed casein formula in SPT (Oldaeus et al. 1991). Similarly according to Wahn et al. (1992) extensively hydrolyzed casein formulas had the least residual allergenic activity in skin tests and oral challenges of 6 different, tested formulas.
Traces of native milk proteins could be specifically detected in extensively hydrolyzed formulas using both monoclonal antibodies and serum from allergic subjects in SDS-PAGE immunoblotting (Restani et al. 1995, 1996).
Simplesse, a fat substitute, is made from ultrafiltrated egg white and skimmed milk by heat and high-shear processing. The coagulated nanoparticles are 0.1-3 mm in diameter. Sampson and Cooke (1992) investigated the allergenic potential of Simplesse by SDS-PAGE immunoblotting. No differences in IgE-binding patterns between Simplesse proteins and native egg and cow's milk proteins were observed using 16 sera from egg and/or cow's milk allergic individuals.
A 30-year old woman experienced anaphylactic symptoms after eating a prepackaged bread with salmon (Koppelman et al. 1999). The patient was not sensitized to fish. Detailed examination of the product revealed that a new process for restructured meat had been introduced by the manufacturer. The salmon had been treated with the microbial enzyme transglutaminase to improve the structure of the meat by covalent linkage between added caseins and meat proteins. Assuming 10 to 50 g of salmon were ingested approximately 10 to 50 mg of caseins induced the allergic reactions (Koppelman et al. 1999).
Ovomucoid (Gal d 1), ovalbumin (Gal d 2), ovotransferrin (Gal d 3), and lysozyme (Gal d 4) are the most important allergens in egg white protein (Barkholt et al. 2000). The major allergenic component of the egg yolk is alpha-livetin (identical to chicken serum albumin, Gal d 5) (Table 5).
Raw Eggs, Boiled Eggs and Heated Egg White
Challenges with dilutions of masked fresh, raw hen's egg induced symptoms in DBPCFC in 7 egg allergic adults (Norgaard & Bindslev-Jensen 1992). A dose of 50 mg whole egg induced symptoms of oral allergy syndrome (OAS), diarrhoea, asthma, and conjunctivitis in 2 patients. Doses of 500 mg and 2.5 g induced allergic reactions in 2 other patients. In the other patients doses of 50 g of whole egg were necessary for the elucidation of symptoms. The threshold doses were equivalent to 5 mg to 5 g of egg protein (Table 4).
Ovomucoid and ovalbumin could be detected in soft-boiled (100°C, 3 min) and hard-boiled eggs (100°C, 20 min) by radio- immunoelectrophoresis (RIEP) with rabbit antisera showing reduced, but clear residual antigenicity (Hoffmann 1983).
IgE-binding of egg white decreased about 58% in RAST after heating to 90°C for 10 min (Anet et al. 1985). In DBPCFC a decrease in positive reactions occurred after heating of egg white (90°C, 60 min). Only 55% of egg allergic patients with positive DBPCFC to freeze-dried egg white reacted to heated egg white (Urisu et al. 1997). Heated, ovomucoid depleted egg white was positive in only 6% (DBPCFC).
Egg Protein Additives: Meat Pastes
Leduc et al. (1999) tested the IgE-binding activity of 3 experimental pork meat pastes containing 2% dried egg white. Egg white allergens could be detected by SDS-PAGE immunoblot and EAST in raw and in pasteurized meat pastes (70°C, 2 h), while no IgE-binding to the sterilized paste (115°C, 90 min) was observed.
Hydrolysis with Digestive Enzymes
Ovalbumin and phosvitin resisted peptic digestion at pH 1.2 (>60min), while ovotransferrin was immediately degraded and ovomucoid was degraded after 8 min of treatment (Astwood et al. 1996). In contrast, Urisu et al. (1999) determined significant IgE-binding by pepsin, chymotrypsin, and trypsin digested ovomucoid in DBPCFC positive egg allergic children (RAST inhibition).
Palm et al. (1999) proved the allergenic potential of egg lecithins in a 15-month old girl in DBPCFC. After ingestion of 50 mg of egg lecithins allergic symptoms of the skin occurred within an hour (erythema of neck and shoulders). The protein content of the lecithin preparation was 11.3%.
See cow's milk allergens.
The first food allergen characterized in detail was "Allergen M" (Gad c 1, 12 kDa) from codfish (Aas & Elsayed 1969). Gad c 1 belongs to the family of Ca-binding parvalbumins, which are present in muscle tissue of amphibians and fish. Proteins with cross- reactivity to Gad c 1 have been identified in several fish species: salmon (Sal s 1), tuna, perch, carp, eel, catfish, dogfish, and snapper (James et al. 1997, Bugajska-Schretter et al. 1998). Additional fish allergens are present in the molecular mass range from 15 to 200 kDa (Bugajska-Schretter et al. 1998).
Raw and Boiled Fish
The capability of fresh, raw codfish meat to induce allergic reactions was proven in 10 fish allergic adults by DBPCFC (Table 4). Amounts as low as 6 mg to 6.7 g elucidated allergic symptoms. More severe reactions including anaphylaxis occurred with relatively high amounts of 25-50 g codfish (Hansen & Bindslev-Jensen 1992).
The allergenic potential of boiled fish was confirmed by Bernhisel-Broadbent et al. (1992a) testing 10 different fish species in DBPCFC. SDS-PAGE experiments revealed denaturation of some protein bands and formation of high-molecular bands in boiled fish as compared to protein extracts from raw fish. A relatively strong decrease in IgE-binding to allergens from boiled fish could be observed in immunoblotting. Bernhisel-Broadbent et al. (1992a) found that in vitro tests were frequently not in agreement with the clinical relevance of fish hypersensitivity.
Allergens in codfish, herring, and plaice were heat- stable after boiling for 6 min, 1 h, and 4 h tested with sera from 2 patients with codfish allergy (Hansen et al. 1994). With the exception of mackerel the activity of histamine release was unchanged for all protein extracts from boiled fish. Protein bands > 40 kDa were found to be heat labile, while proteins with lower molecular mass proved to be stable. Still, after 4 h of boiling IgE-binding fish proteins could be detected in SDS-PAGE immunoblot.
Canned Fish Products
Studies of Bernhisel-Broadbent et al. (1992b) indicate that canning fish results in the loss of its allergenic activity. DBPCFC with canned tuna was negative in 18 fish allergic patients as well as with canned salmon in 2 salmon allergic patients.
SDS-PAGE proved a significant loss of distinct protein bands in canned fish as compared to raw and boiled fish. The IgE-binding activity of canned fish was weak in immunoblotting and about 100 to 200 times less as compared to boiled fish in EAST inhibition (Bernhisel-Broadbent et al. 1992b).
Thoroughly washing of fish meat eliminates the water- soluble proteins and retains water- insoluble proteins. Therefore surimi, which is produced from minced, thoroughly washed fish meat from different species, presents an allergen pattern different from native fish species. Allergens in fresh codfish were detected in the range of 13 to 63 kDa, while surimi retained only a 63 kDa allergen in SDS-PAGE (Mata et al. 1994). In SPT 2 out of 6 patients with codfish allergy reacted to surimi, while all 6 sera were positive to surimi in RAST. IgE-binding to codfish extract was inhibited by surimi extracts with maximum inhibition of 60 to 94% (RAST inhibition). Helbling et al. (1992) proved the IgE-binding potential of surimi with sera from fish allergic patients in RAST inhibition. Surimi may contain other ingredients like starch, egg white, and other food additives giving a gelateous structure after heating.
"Hidden" Allergens in Used Fat
Frying foods using the same fat which was previously used for fish may threaten fish allergic subjects. Yunginger et al. (1988) reported a fatal anaphylactic reaction in a fish allergic individual after eating fried potatoes. The fat had been previously used for frying fish.
Hydrolysis with Digestive Enzymes
Aas & Elsayed (1969) examined the stability of the muscle myogene fraction from codfish against extensive enzymatic digestion with trypsin, pepsin, subtilisin, and pronase (48h, 37°C), respectively. After the treatment the allergenic activity in skin testing was lost. In contrast, after digestion with elastase approximately 50% of allergenic activity was retained. Some allergenic activity was present in partial hydrolysed protein fractions after digestion with pronase for 24 h and with trypsin and pepsin for 2 h, respectively.
The family of crustaceae includes shrimps, lobster, crab, and crayfish. The major allergens of crustaceae are tropomyosins which were identified in several shrimp species (Met e 1, Pen a 1, Pen i 1) and lobsters (Pan s 1, Hom a 1) as well as in squids (Tod p 1), which belong to the family of molluscs (Table 5).
The heat stability of shrimp allergens was evidenced by several studies. 30% of 30 patients with a history of shrimp allergy reacted to boiled shrimps after open oral challenge (Daul et al. 1988). Six of these patients were positive in DBPCFC too. Challenge tests were performed with doses of 1 to 16 shrimps corresponding to 4 to 64 g of shrimps (Table 4).
Naqpal et al. (1989) isolated a 34 kDa allergen (Pen i 1) from boiled shrimps which retained its allergenic activity. Daul et al. (1994) detected similar allergenic activity of the major allergen Pen a 1 in shrimp extracts as well as in the cooking water by SDS-PAGE immunoblot.
One patient was described who experienced an anaphylactic reaction after ingestion of boiled shrimps while having a negative SPT to raw shrimps. The evaluation by SPT revealed exclusive sensitivity to boiled shrimps in this particular patient (Rosen et al. 1994).
Yunginger et al. (1988) reported a fatal case of anaphylaxis after ingestion of crab.
Meat allergens are serum albumins (66 kDa), gamma- globulins (160kDa), and actins (Restani et al. 1997) as well as several additional proteins (14, 18, 20, 45 und >60 kDa). Cross-reactivity to whey proteins from milk are due to serum albumins and gamma- globulins. Most recently Ayuso et al. (2000) obtained complete inhibition of IgE-binding to bovine IgG with lamb, venison, and milk. Denatured type I collagen (beef) was identified as the major allergenic component of gelatine.
Poultry and Mammals
According to a recent study (Ayuso et al. 1999) 75% of patients with sensitivity to meat from mammals were sensitive to poultry at the same time (IgE-binding in SDS-PAGE immunoblot). Vice versa only 50% of poultry sensitive patients showed IgE-binding to meat extracts from mammals. Sensitivity in SDS-PAGE immunoblot was associated to clinically relevant poultry meat allergy in 64% of patients, while only 29% of patients with beef sensitivity had a clinically relevant allergy to beef (Ayuso et al. 1999).
Restani et al. (1998) found no IgE-binding to serum albumins from turkey and chicken in 6 children allergic to bovine serum albumin (SDS-PAGE immunoblot).
A case of adverse reactions to raw meat without sensitivity to cooked meat was described by Fisher (1982). A patient experienced anaphylactic symptoms after eating rare-cooked beef, while well-cooked beef was tolerated.
In a DBPCFC study with 11 children (aged 16 months to 14 years) with beef allergy 8 of them reacted against well-cooked beef (brown), while 3 children reacted to rare-cooked meat (red centrally and pink peripherally) (Werfel et al. 1997). Reported adverse reactions were predominantly cutaneous symptoms, whereas severe systemic reactions were not observed.
Heat treatment induces denaturation of meat proteins which lose their state of water solubility. The analysis of allergens by SDS-PAGE immunoblotting revealed a 17.8 kDa and 19 kDa allergen and four additional weakly detected bands (14, 20, 45 and >60 kDa) in minced beef which was heated (85°C) up to 2 h (Werfel et al. 1997). These protein bands were identified in well-cooked minced beef (20 min, up to 80°C) as well. The strongest IgE-binding activity was observed to the 17.8 kDa allergen by sera from patients with positive DBPCFC to cooked beef. Bovine serum albumin and gamma-globulin bands vanished in SDS-PAGE after heating (80°C) of minced beef for 10 and 3 min, respectively. Purified serum albumin was stable for 15 min at 95°C purified, while gamma-globulin was stable for 15 min at 65°C (Werfel et al. 1997).
Seven of 11 children with beef allergy and positive SPT to native bovine serum albumin also showed a sensitivity in SPT to heated bovine serum albumin. And 4 children had positive challenge with heated bovine serum albumin in DBPCFC (Fiocchi et al. 1998).
Restani et al. (1998) found that purified bovine serum albumin was still capable of IgE-binding after treatment at 100° for 10 min. In these conditions, bovine serum albumin produced aggregates but both monomers and polymers were antigenic. It was suggested that milk could protect bovine serum albumin from aggregation with easier denaturation, thus thermostability could depend on the medium.
Of 57 patients with suspected meat allergy to beef, pork, lamb, rabbit, or poultry only 2 showed in vitro reactivity to tropomyosins from pork and chicken, respectively (Ayuso et al. 1999). IgE-binding to allergens from raw meat was stronger than to allergens from heated meat (20 min, 140°C) in 75% of sera with the exception of chicken meat. 6 of 24 sera reacted strongly to proteins in heated chicken meat, while IgE-binding to proteins from raw meat was weak. Several neoallergens were detected in heated chicken meat at a molecular mass range of 14 to 90 kDa. Heat labile allergens with 45 and 150 kDa were identified in raw chicken meat. Heat stable allergens simultaneously detected in raw and heated chicken meat had molecular masses of 17, 20, 24, 28, 31, and 66 kDa in SDS-PAGE.
IgE-binding in SDS-PAGE immunoblots disappeared completely in cooked beef extracts (20 min, 140°C) and raw beef extracts separated under reducing conditions using sera from 12 patients with convincing history of beef allergy and IgE-binding to raw beef extracts indicating heat labile and conformational epitopes, respectively (Ayuso et al. 2000).
Homogenization and Lyophilization
Fiocchi et al. (1998) studied the impact of freeze-drying and homogenization on the allergenicity of beef. 10 children with positive SPT to raw and heated beef and positive DBPCFC with 180 g heated beef (5 min, 100°C) were tested. All children reacted to native bovine serum albumin in SPT and 5 children reacted to native bovine serum albumin in DBPCFC.
Only one child had a positive reaction to freeze-dried beef in SPT, while no positive reaction to freeze-dried beef was observed in DBPCFC. With homogenized beef neither positive SPT nor positive DBPCFC results could be observed in any of the children. In an earlier study (Fiocchi et al. 1995b) weak allergenicity of homogenized and freeze-dried beef and freeze-dried sheep's meat could be observed in 2 of 12 children with clinically relevant beef allergy by SPT.
Hydrolysis of Meat Proteins
Fiocchi et al. (1995b) performed SPT with pepsin digested serum albumins from beef and sheep in 12 children with clinically relevant symptoms of beef allergy and positive SPT to native serum albumin. After 5 min of digestion 4 children showed positive SPT to digested bovine serum albumin, while after 2 and 4 h of pepsin digestion 2 positive SPT were obtained, respectively. Pepsin preparations of ovine serum albumin digested for 5 min elucidated positive reactions in SPT in 3 children, while preparations digested for 2 h elucidated no skin reactions in any of the children. The pepsin digestions all were negative in RAST.
Cases of anaphylaxis to gelatine containing vaccines have been frequently reported in the literature, while reports of allergic events after ingestion of gelatine containing foods are rare. According to a study of Sakaguchi et al. (1995) 7 of 26 children who experienced systemic allergic reactions induced by gelatine containing vaccines showed allergic reactions after ingestion of gelatine containing foods. The cross- reactive potential of gelatines from various mammalian species was proven by RAST inhibition with sera from 12 children with bovine gelatine allergy. In one child a strong sensitization to fish gelatine could be observed (Sakaguchi et al. 1999a). The alpha 2 chain of type I collagen from beef was identified as the major IgE-binding fraction of gelatine in SDS-PAGE immunoblot and RAST (Sakaguchi et al. 1999b).
Wahl and Kleinhans (1989) reported a case of oral allergy syndrome after ingestion of fruit gums containing gelatine. SDS-PAGE immunoblotting revealed allergens in the molecular mass range of 40 to 120 kDa.
Enzymatic digestion with collagenase degraded pork gelatine (heat- denatured collagen) to peptides <10 kDa which retained very low IgE-binding potentail (Sakai et al. 1998).
Table 6: Stability of important food allergens of animal origin
|Allergen||Heat Treatment||Enzymatic Hydrolysis|
|Cow's milk||stable||partially resistant|
Food allergens of animal origin are generally highly resistant to common treatments during food processing, with limitations to some extent as regards meat. The impact of heat and enzymatic treatments on the allergenicity of food allergens are summarized in Table 6. Cow's milk allergens retain their allergenicity after conventional industrial treatments. In order to produce "hypoallergenic" (less allergenic) cow's milk infant formulas, extensive conditions employing hydrolysis, heating, and sometimes ultrafiltration are needed. Allergens from hen's egg are stable to heat and enzymatic digestion nonetheless. Moreover, milk and egg products are common ingredients of many convenience foods and therefore hidden potential allergens in processed foods. The significance of fish and crustaceae as hidden allergens is low, while they are usually stable to heat treatment. The allergenic potential of meat is partially heat stable, while the stability to enzymatic digestion is relatively weak. Gelatine may be important as a hidden allergen, while its prevalence is very low.
The allergic individual is usually advised to avoid all forms of the
food allergen to which he is sensitized. On one hand this situation reflects
the lack of data on the allergenicity of processed foods and food additives.
On the other hand it shows the need for the development of well characterized
less or non-allergenic foods.
Future research should investigate the allergenic potentials of foods, especially cow's milk and hen's egg, throughout manufacturing processes from source to shelf- products. Modern food quality management should take into account standardized food specifications with respect to the state of allergenicity. Clinical evaluations of processed foods with an appropriate number of patients with convincing allergy to the native food allergen are also needed. Ideally food products should be monitored simultaneously by various analytical and diagnostic methods like DBPCFC, SPT, RAST, SDS-PAGE immunoblot, and inhibition tests.