Prepared by the Fifty-first meeting of the Joint FAO/WHO
    Expert Committee on Food Additives (JECFA)

    World Health Organization, Geneva, 1999
    IPCS - International Programme on Chemical Safety


    First draft prepared by
    E.J. Vavasour
    Chemical Hazard Assessment Division, Bureau of Chemical Safety, Food
    Directorate, Health Protection Branch, Health Canada, Ottawa, Ontario,

          Biological data 
              Biochemical aspects 
                   Effects on enzymes and other biochemical 
              Toxicological studies 
                   Short-term studies of toxicity 
                   Developmental toxicity 
                   Special studies 
                        Promotion of carcinogenesis 
              Observations in humans 
                   Case studies 
                   Food challenges 
                   Prevalence studies 
          Comments and evaluation 


         Sulfur dioxide and sulfites were evaluated at the sixth, eighth,
    ninth, and seventeenth meetings of the Committee (Annex 1, references
    6, 8, 11, and 32). An ADI of 0-0.7 mg/kg bw was allocated at the
    seventeenth meeting to sulfur dioxide and to sulfur dioxide
    equivalents arising from sodium and potassium metabisulfite, sodium
    sulfite, and sodium hydrogen sulfite. At subsequent meetings (Annex 1,
    references 41, 47, and 62), potassium and calcium hydrogen sulfite and
    sodium thiosulfate were included in the group ADI.

         At its thirtieth meeting, the Committee retained the ADI of 0-0.7
    mg/kg bw allocated to this group of compounds (Annex 1, reference 73).
    The ADI was based on long-term studies in rats, including a
    three-generation study of reproductive toxicity, with a NOEL of 0.25%
    sodium metabisulfite in the diet (supplemented with thiamine, as
    treatment of foods with sulfites reduces their thiamine content),
    equivalent to 70 mg/kg bw per day of sulfur dioxide equivalents. At
    higher doses (> 1%), local irritation of the stomach was observed,
    with inflammatory changes and hyperplasia, and occult blood was
    detected in the faeces at even higher doses. The histopathological
    changes were limited to the stomach; the incidence of neoplasms was

    not increased at any site or at any dose. A safety factor of 100 was
    used. Similar local changes in the stomach were observed in pigs fed
    thiamine-supplemented diets to which sodium metabisulfite was added.
    The Committee also reviewed case studies and challenge tests for
    idiosyncratic sensitivity to sulfiting agents and noted the
    life-threatening nature of the adverse effects in some cases. It
    recommended that, where a suitable alternative method of preservation
    exists, its use should be encouraged, particularly in those
    applications in which the use of sulfites may lead to high acute
    intake. The Committee also reiterated the view expressed at the
    twenty-seventh meeting (Annex 1, reference 62, section 2.4) that
    appropriate labelling is the only feasible means of protecting
    individuals who cannot tolerate certain food additives. The Committee
    recommended that the situation with regard to the prevalence of
    idiosyncratic adverse reactions and the relative toxicity of free and
    bound sulfur dioxide be kept under review. It also requested
    information on the chemical forms of sulfur dioxide in food.


    2.1  Biochemical aspects

    2.1.1  Biotransformation

         The metabolic disposition of sulfite was studied in isolated,
    perfused rat liver and in isolated rat hepatocytes. In particular, the
    kinetics of metabolism of sulfite ion to inorganic sulfate ion and the
    effect of sulfite ion on the concentrations of the endogenous,
    low-molecular-mass thiol-containing molecules glutathione and cysteine
    were considered to be of interest. Sulfite was rapidly converted to
    sulfate by isolated hepatocytes at concentrations ranging from 200
    µmol/L to 2 mmol/L. The rate of conversion was linear and quantitative
    over this range. These results confirm the presence of substantial
    concentrations of sulfite oxidase in liver. Cytotoxicity was not seen,
    even at the highest concentration of sulfite. Perfusion of isolated
    rat liver with 1 mmol/L sulfite resulted in nearly 98% extraction of
    sulfite from the perfusate during the first 3 min of single-pass
    perfusion. At the same time, sulfate ion was shown to accumulate
    rapidly in the perfusate, reaching 830 µmol/L by 5 min and nearly 100%
    conversion within 30 min. Perfusion of the liver also resulted in
    release of glutathione into the perfusate in a rapid phase in which 11
    µmol/L glutathione accumulated over 5 min and a slow phase in which 17
    µmol/L glutathione accumulated over 60 min. Free cysteine was not
    dected in the perfusate at any time. Incubation of isolated
    hepatocytes with 0.5, 1, or 2 mmol/L sulfite resulted in a
    time-dependent increase in the amount of free glutathione, but not
    cysteine, associated with the cells. The authors speculated that
    glutathione was released from mixed disulfides of protein and
    low-molecular-mass thiols and from oxidized glutathione in hepatocytes
    and perfused liver, and was not synthesized  de novo. Sulfitolysis of
    disulfides by sulfite ion at physiological pH was demonstrated by

    incubation of sulfite with oxidized glutathione, resulting in a time-
    and concentration-dependent release of glutathione from the disulfide
    (Sun et al., 1989).

    2.1.2  Effects on enzymes and other biochemical parameters

         In studies of glutathione  S-sulfonate, a well-documented
    reaction product of sulfite with oxidized glutathione, and the enzyme
    glutathione  S-transferase from rat liver and lung and human
    tumour-derived lung cells, glutathione  S-sulfonate was a strong
    competitive inhibitor of both cytosolic and microsomal glutathione
     S-transferase in all cell lines. Glutathione  S-sulfonate
    subsequently underwent reduction to oxidized glutathione or
    glutathione. These results indicate that the glutathione pathway may
    be vulnerable to the effects of sulfur dioxide through its reaction
    with oxidized glutathione, which depletes the glutathione supply
    (Leung et al., 1985).

         The activities of some intestinal enzymes were measured in female
    rats exposed to sodium metabisulfite in the diet. Groups of 10 female
    Wistar rats weighing 70-100 g received diets containing 0, 0.5, or
    2.5% sodium metabi-sulfite  ad libitum for five weeks. After
    sacrifice, the entire small intestine was removed and homogenized. The
    activities of maltase, sucrase, lactase, and alkaline phosphatase were
    assayed in crude intestinal homogenates and in brush-border membrane
    fractions. Feeding with 2.5% metabisulfite resulted in small increases
    in the activities of maltase and lactase in the homogenates, which did
    not reach statistical significance. The activities of all three
    disaccharidases in the brush-border fractions were statistically
    significantly increased at the high dose. Alkaline phosphatase
    activity was significantly increased in both the crude homogenates and
    the brush-border fractions from rats receiving the high dose. The
    authors considered the possibility that the increased activity of the
    enzymes involved in carbohydrate digestion was due to a decrease in
    their degradation; however, since the increase in alkaline phosphatase
    activity was probably due to an increase in enzyme concentrations,
    this could also be a factor in the increased activity of the
    disaccharides (Rodriguez-Vieytes et al., 1994).

         The possibility that the adverse effects attributed to ingestion
    of sulfites, including bronchoconstriction, are mediated through
    effects on nitric oxide as part of the parasympathetic pathway was
    tested in a series of experiments  in vitro. Sulfites were shown to
    react in solution with nitric oxide and with biological carriers of
    nitric oxide  (S-nitrosylated bovine serum albumin and
    S-nitrosoglutathione) and to interfere with the effects of nitric
    oxide in biological processes such as inhibition of platelet
    aggregation. The results suggest that sulfites interfere with the
    biological effects of nitric oxide at physiologically realistic
    concentrations (Harvey & Nelsestuen, 1995).

    2.2  Toxicological studies

    2.2.1  Short-term studies of toxicity


         The subchronic toxicity of free inorganic sulfite (as sodium
    metabisulfite) and acetaldehyde hydroxysulfonate, a major bound form
    of sulfite in beer and wine, was evaluated after their addition to the
    drinking-water of normal and sulfite oxidase-deficient rats. Groups of
    eight female Wistar rats weighing 175-199 g were assigned to one of 14
    groups receiving sodium metabisulfite or acetaldehyde hydroxysulfonate
    in the drinking-water for eight weeks at doses of 0, 7, 70, or 350
    (three weeks)/175 (five weeks) mg/kg bw per day of sulfur dioxide
    equivalents. The study included both normal rats and rats that had
    been made sulfite oxidase-deficient by the addition of 200 ppm
    tungsten to their drinking-water in the form of sodium tungstate. The
    animals were observed daily throughout the experiment; body weights
    were measured weekly, food consumption twice per week, and water
    consumption three times per week throughout the study. Blood and urine
    samples were collected before and every two weeks during the
    experiment for determination of haemoglobin and plasma protein,
     S-sulfonates and sulfite, and urinary sulfite and thiosulfate. At
    the end of the study, the rats were killed and autopsied. Liver
    samples were collected for determination of thiamine content and
    sulfite oxidase activity, the latter as an indication of the
    effectiveness of tungsten treatment. Histopathological examination was
    conducted on the aortic arch, heart, representative portions of the
    liver, left kidney, left lung, ovaries with fallopian tubes,
    oesophageal-forestomach junction, forestomach, fundic area of the
    stomach, pylorus-duodenal junction, mid- and distal duodenum, jejunum,
    ileum, caecum, and mid-colon.

         Tungstate treatment effectively obliterated hepatic sulfite
    oxidase activity; untreated animals had an average value of 350
    units/g liver. The overall health of the animals was not affected by
    treatment, except that sulfite oxidase-deficient rats receiving either
    of the sulfite treatments had dried blood around their noses four to
    five weeks after the beginning of treatment, whereas sulfite
    oxidase-deficient controls did not. This effect was attributed to
    respiratory distress related to the lung oedema noted at necropsy. The
    rats were not deficient in thiamine. After eight weeks of treatment,
    the body weights of the sulfite oxidase-deficient rats receiving the
    highest dose of metabisulfite were significantly lower than those of
    controls. All groups of rats receiving metabisulfite consumed more
    feed, when calculated as grams per kilogram bodyweight per day, than
    either the respective control groups or those receiving acetaldehyde
    hydroxysulfonate, although no dose-response relationship was apparent.
    A dose-related decrease in water consumption was noted in the sulfite
    oxidase-deficient rats, which reached statistical significance in
    those receiving metabisulfite but not in those receiving acetaldehyde
    hydroxysulfonate. Haemoglobin and plasma protein concentrations were
    not affected by treatment. 

         Urinary sulfite was found at low concentrations or was
    undetectable in rats with normal sulfite oxidase activity, even when
    they were treated with sulfite in the drinking-water, indicating
    rapid, efficient metabolism of sulfite by this enzyme. Sulfite was
    detected in the urine of sulfite oxidase-deficient rats even before
    sulfite treatment was initiated. The urinary sulfite concentrations
    increased in the enzyme-deficient rats after initiation of treatment
    with either the free or the bound forms of sulfite; however, the
    sulfite concentrations were variable, and no correlation with dose or
    length of treatment was apparent, although rats treated with
    acetaldehyde hydroxysulfonate tended to excrete more free sulfite than
    did metabisulfite-treated rats. The authors suggested that the lower
    water intake of the latter groups may have contributed to this
    phenomenon. By contrast, the plasma sulfite concentrations were low
    and variable in all groups. This effect was attributed to the ability
    of the sulfite ion to react with many biological compounds to form
     S-sulfonates, possibly by sulfitolysis of disulfide bonds in
    proteins and free cysteine. Plasma  S-sulfonate concentrations were
    generally lower in rats with normal sulfite oxidase activity, but even
    the enzyme-deficient control rats had elevated plasma  S-sulfonate
    concentrations, indicating that these substances are formed
    endogenously. There was no dose-response relationship between plasma
     S-sulfonate concentration and administered sulfite; however, these
    substances were present at higher concentrations in enzyme-deficient
    rats treated with free sulfite than in those treated with acetaldehyde
    hydroxysulfonate or in controls. The urinary thiosulfate
    concentrations were higher in sulfite oxidase-deficient than in normal
    rats given either sulfite treatment or in controls. As for urinary
    sulfite and plasma  S-sulfonates, no general increase in urinary
    thiosulfate was observed with increasing dose of administered sulfite.
    The urinary thiosulfonate concentrations were low and variable in the
    enzyme-deficient rats, although they were still higher than in normal
    rats, suggesting that this is a minor pathway for excretion of excess

         The only finding at gross necropsy was white patches in the lungs
    of the sulfite oxidase-deficient rats receiving sulfite treatment.
    Histopathological examination revealed lesions in the fore- and
    glandular stomachs of both normal and sulfite oxidase-deficient rats
    receiving the highest dose (350/175 mg/kg bw per day) of bound or free
    sulfite. The enzyme-deficient rats had the most severe lesions. These
    included moderate hyperkeratosis of the forestomach and alterations of
    the fundic portion of the stomach, including a thinner fundic mucosa,
    distention of the basal portion of the fundic glands, chief-cell
    hypertrophy along the fundic glands with varying amounts of apical
    acidophilic granules and fewer parietal and mucous cells (less evident
    in acetaldehyde hydroxysulfonate-treated rats), clusters of aberrant
    fundic glands with hyperplastic chief cells (metabisulfite-treated
    rats only), and damage to the surface epithelium and gastric pits,
    with oedema and sloughing, resulting in bleeding (acetaldehyde
    hydroxysulfonate-treated rats only). In addition, pathological changes
    in the liver were seen in the acetaldehyde hydroxy-sulfonate-treated
    rats, affecting sulfite oxidase-deficient animals receiving the

    intermediate and high doses and normal animals receiving the high
    dose. The changes were described as cytoplasmic vacuoles within
    hepatic parenchymal cells. It could not be determined whether the
    vacuoles contained lipid or glycogen. In enzyme-deficient rats
    receiving the high dose, the death of about 0.5% of the parenchymal
    cells was seen, which was more prevalent in extensively vacuolated
    cells. The authors speculated that the liver lesions were due to the
    effects of free acetaldehyde. They concluded that the toxicity of
    acetaldehyde hydroxysulfonate is equivalent to that of free sulfite
    and that although sulfite oxidase-deficient rats were more sensitive
    to the effects of sulfite the no-effect levels were identical. The
    NOEL was 70 mg/kg bw per day of sulfur dioxide equivalents (Hui et
    al., 1989).

    2.2.2  Genotoxicity

         Sodium bisulfite, as a 1:3 mixture of NaHSO3 and Na2SO3, was
    tested at concentrations of 0.05-1 mmol/L in human peripheral
    lymphocytes  in vitro. Positive results were obtained in assays for
    chromosomal aberrations, micronucleus formation, and sister chromatid
    exchange (Meng & Zhang, 1992).

    2.2.3  Developmental toxicity


         Groups of 12 or 13 pregnant Wistar rats (body weights not
    specified) received potassium metabisulfite in the diet on days 7-14
    of gestation at concentrations of 0, 0.1, 1, or 10%. Two-thirds of the
    rats receiving 0, 0.1, and 10% and all those given 1% were killed on
    day 20 of gestation for examination of the fetuses; the remaining rats
    were allowed to deliver their litters and to rear their offspring for
    15 weeks.

         The body-weight gain of dams receiving 10% potassium
    metabisulfite was markedly depressed during the treatment period: the
    animals lost weight during this period but by day 20 had nearly
    compensated by faster body-weight gain. The body-weight gain of the
    other treated animals was similar to that of controls. The actual
    intakes of metabisulfite were 0, 130, 1300, and 2900 mg/kg bw per day,
    respectively, corresponding to 0, 75, 760, and 1700 mg/kg bw per day
    of sulfur dioxide equivalents. There were more resorptions and fetal
    deaths in the treated groups than in controls, but the difference was
    not statistically significant and was not dose-related. There was no
    difference in the number of live fetuses per dam or in the sex ratio
    of the fetuses. The fetal body weights in dams given 10% were
    significantly lower than in controls. No skeletal or external
    anomalies were detected in any group; delayed ossification and some
    skeletal variations were noted in all of the groups but the incidence
    was not related to dose. Several fetuses with visceral anomalies were
    found in all groups, both control and treated, with no relationship to
    dose. The number of liveborn pups, the live birth index (number of
    liveborn pups divided by number of implantation sites), and the

    survival rate of offspring on day 4 after birth were decreased and the
    number of stillborn pups increased in dams receiving the 10% dose that
    were allowed to bring their litters to term in comparison with
    controls. None of these differences was statistically significant. The
    total litter loss between day 0 and day 4 in one dam at 10% had a
    large effect on the survival index for the whole group. Survival and
    body-weight gain were similar in all groups during the remainder of
    the preweaning period and after weaning. The adverse effects on pup
    birth weights observed with the 10% dietary dose were probably due to
    the drastic maternal body-weight loss during the treatment period. No
    teratogenic effects were observed, and there were no persistent
    effects on pup survival or body-weight gain. The NOEL was 760 mg/kg bw
    per day of sulfur dioxide equivalents (Ema et al., 1985).

         Groups of 10-12 pregnant Wistar rats (mean body weight, 240 g)
    were fed diets containing 0, 0.32, 0.63, 1.25, 2.5, or 5% sodium
    sulfite heptahydrate in the diet  ad libitum on days 8-20 of
    gestation. Satellite groups of four pregnant rats received 0, 0.32, or
    5% in the diet on the same days. Body weights, food consumption, and
    clinical signs of toxicity were recorded daily. On day 20 of
    gestation, rats in the main group were sacrificed and the uteri opened
    and examined for the presence and position of resorptions, viable and
    non-viable fetuses, and implantation sites. Live fetuses were weighed,
    sexed, and examined for external abnormalities. Half of the fetuses
    were subjected to visceral examination, and the other half were
    prepared for skeletal examination. Dams in the satellite group were
    allowed to deliver and rear their litters to weaning. The average
    daily intake of sodium sulfite heptahydrate, in order of ascending
    dose, was 0, 300, 1100, 2100, and 3300 mg/kg bw per day (0, 80, 280,
    530, and 840 mg/kg bw per day of sulfur dioxide equivalents). 

         Both food intake and body-weight gain during treatment were
    reduced in the animals receiving the 5% diet. Food intake was also
    reduced in the groups receiving the two lowest doses, with no apparent
    dose-response relationship. The total number of implantations was
    higher in dams at the three highest doses than in the controls or rats
    at the two lower doses, although the difference was not statistically
    significant. The percentage of intrauterine deaths (resorptions and
    dead fetuses) was also increased over that in controls in dams at the
    two highest doses. As a result, the litter sizes in all groups were
    comparable. The fetal body weights were statistically significantly
    lower in all treated groups than in controls, with the exception of
    dams at 2.5%. No external, skeletal, or visceral anomalies were
    observed in fetuses in any group, although the incidences of several
    fetal skeletal variations, such as lumbar rib, hypoplastic 13th rib,
    and delayed ossification of the skull and sternum, were slightly
    increased in some of the sodium sulfite-treated groups in a weakly
    dose-related manner. Dilatation of the renal pelvis and of the lateral
    ventricle were also observed, with no significant dose-response
    relationship. None of these effects was statistically significant.
    Dams that brought their litters to term had normal weight gain over
    the lactation period. Treatment had no effect on litter parameters
    (number of liveborn pups, live birth index, survival rate at days 7

    and 28  post partum, or pup body weight on day 21). The authors noted
    that maternal toxicity occurred only at the highest dose (5% of the
    diet); however, since mild growth retardation was seen at all doses
    and may have caused the slight increase in developmental variations,
    the LOEL for the study was 80 mg/kg bw per day of sulfur dioxide
    equivalents (Itami et al., 1989).

    2.2.4  Special studies  Nephrotoxicity

         The effect of continuous consumption of metabisulfite on rat
    kidney cells was studied by assaying the activities of selected
    enzymes before and during its administration. Groups of 18 male albino
    rats (strain not indicated) received distilled water or sodium
    metabisulfite solution orally by syringe to deliver a dose of 5 mg/kg
    bw daily (equivalent to 3.4 mg/kg bw of sodium dioxide equivalents)
    for 1, 3, 7, 9, 11, 13, or 15 days. A subset of the rats were
    sacrificed 24 h after each dose, and blood and kidneys were collected.
    An additional 18 rats were held in metabolism cages for daily
    collection of urine; 12 of these rats were fed sodium metabisulfite
    solution every 24 h for 15 days, with daily urine collection, and the
    remaining six rats served as controls and received 1 ml distilled
    water. The activities of alkaline phosphatase, acid phosphatase,
    lactic dehydrogenase, and glutamate dehydrogenase were measured in
    serum, renal tissue, and urine. Protein excretion was also measured.

         The activity of alkaline phosphatase in renal tissue was reduced
    immediately after the first dose of metabisulfite, reaching a level
    that was one-third that of the controls and persisting over the two-
    week treatment period; there were concomitant increases in the
    activity of this enzyme in serum and urine. Lactate dehydrogenase
    activity was also reduced in renal tissue, starting after five days of
    treatment and reaching a value about one-half that of controls by the
    end of the experiment; concomitant increases were seen in the activity
    of this enzyme in urine but not in serum. The activity of neither acid
    phosphatase nor glutamate dehydrogenase was affected by treatment.
    Urinary excretion of total protein was increased by nearly 10-fold by
    the end of the experiment. The authors concluded that chronically
    ingested sulfite can damage renal-cell membranes but is quickly
    inactivated after entering the cell, as indicated by the leakage of
    two enzymes found in the cell membrane or the cytosol, and does not
    affect enzymes located in other cell organelles (Akanji et al., 1993).  Promotion of carcinogenesis

          N-Methyl- N'-nitro- N-nitrosoguanidine (MNNG) was
    administered at a concentration of 100 mg/L in the drinking-water of
    male Wistar rats given a diet supplemented with 10% sodium chloride,
    for eight weeks. After this treatment, the animals received standard
    diet and drinking-water to which potassium metabisulfite had been
    added at a concentration of 1% for 32 weeks. Rats initiated with MNNG
    and then treated with metabisulfite had a higher incidence of

    adenocarcinomas of the glandular stomach, located in the pyloric
    region, than initiated controls. Lesions of the gastric mucosa were
    found in sulfite-treated animals, regardless of whether they had
    received prior treatment with MNNG. The lesions were described as
    diffuse, deep gastric pits with clearly increased numbers of mucous
    neck cells in the fundic mucosa (Takahashi et al., 1986).

    2.3  Observations in humans

    2.3.1  Case studies

         In a double-blind, placebo-controlled trial, a 49-year-old woman
    with a history of allergic rhinitis and who was a steroid-dependent
    asthmatic received a sulfite challenge of 5 mg by capsule, solution,
    or subcutaneously, which resulted in a drop in forced expiratory
    volume in one second (FEV1). Skin-prick tests with sulfite produced
    inconsistent reactivity, but reproducible reactions were elicited by
    the intradermal technique. The skin reactivity could be transferred
    via a heat-sensitive principle in the serum. Tests for histamine
    release from leukocytes with sulfite gave inconsistent results. The
    authors suggested that skin-sensitizing antibodies mediate
    sulfite-sensitive asthma through involvement of immunoglobulin (Ig) E
    (Simon & Wasserman, 1986).

         Three patients from a larger group suspected of being sensitive
    to ingested sulfites were shown to have allergic reactions. These
    subjects ranged in age from 22-55 and all were female. Two had
    steroid-dependent asthma, and all three had experienced extreme
    reactions in response to foods eaten in restaurants and to wine. One
    subject had experienced similar reactions after inhalation of
    potassium metabisulfite during wine-making. Their serum IgE
    concentrations were elevated, and skin tests (prick or intradermal)
    for reaction to potassium metabisulfite were positive. Similar
    responses to those to ingestion of food were provoked by oral
    challenge with potassium metabisulfite at doses of 1-50 mg. The
    responses consisted of urticaria or angio-oedema, asthma, headache,
    rhinoconjunctivitis, abdominal pain, and anaphylaxis. A passive
    transfer test (heat-sensitive principles in serum) was positive in
    both subjects in whom it was conducted, confirming that an
    IgE-mediated mechanism was involved in these two individuals (Yang et
    al., 1986).

         Six girls and two boys aged two to six were enrolled in a study
    for sensitivity to food additives and found to be sensitive to sulfite
    in an oral provocation test. The primary manifestation was urticaria
    with or without accompanying angio-oedema (Botey et al., 1987).

         A 31-year-old steroid-dependent woman with asthma who had begun
    to experience severe episodes of bronchospasm of sudden onset five
    years previously, sometimes to restaurant meals, was tested for
    sensitivity to sulfites. She had a significant reduction in pulmonary
    function (> 20% drop in FEV1) in response to a single-blind, oral
    challenge with 25 mg sodium metabisulfite in acidic solution, tidal

    inhalation of non-nebulized sodium metabisulfite solution (10 mg/ml),
    and a double-blind, oral challenge with 10 mg potassium metabisulfite
    in capsules. Skin prick and intradermal challenge with potassium
    metabisulfite also produced positive responses, as did intradermal
    challenge with acetaldehyde hydroxysulfonate. The patient also had a
    significant drop in FEV1 in response to a double-blind oral challenge
    with sulfited lettuce and mushrooms but not with sulfited shrimp.
    Sulfited potatoes produced an inconsistent response (Selner et al.,

         A fatal anaphylactic reaction was reported in a 33-year-old man
    with chronic steroid-dependent asthma. He had been treated previously
    for an acute asthma attack after consumption of dried apricots; in a
    subsequent incident, he had developed dizziness, nausea, and dyspnoea
    shortly after eating a salad in a restaurant. The man had then began
    to avoid foods known to contain sulfite. The anaphylactic reaction was
    precipitated by only a few sips of white wine, which was found to
    contain sulfites at a concentration of 92 ppm (Tsevat et al., 1987).

         An analysis of reports of adverse reactions to sulfites to the
    Adverse Reaction Monitoring System at the Center for Food Safety and
    Applied Nutrition of the US Food and Drug Administration showed that
    the food items most frequently associated with adverse effects were
    salad-bar items, non-salad-bar fresh fruit and vegetables, wine, and
    seafood. The most frequently reported symptoms were those associated
    with asthmatic or allergic reactions (difficulty in breathing,
    wheezing, difficulty in swallowing, hives, itching, local swelling)
    and with gastrointestinal distress (diarrhoea, vomiting and nausea,
    abdominal pain, and cramps). Most (74%) of the reporting consumers
    were female. Of those reporting severe reactions, 25% reported
    difficulty in breathing. A small percentage of the reports were of
    death after exposure to sulfites. In August 1986, the US Food and Drug
    Administration banned the use of sulfiting agents on fresh fruits and
    vegetables, other than potatoes and grapes, as a result of the
    accumulating evidence that sulfiting agents are a hazard to public
    health (Tollefson, 1988).

         A 38-year old female patient with a family history of allergy and
    elevated serum total IgE, who suffered severe bronchospasm in response
    to a number of stimuli, including pharmaceutical preparations, foods,
    and beverages containing sulfite, was subjected to a double-blind oral
    challenge procedure with sodium metabisulfite. The oral challenge
    clearly resulted in moderate obstruction of the central airways and
    lesser obstruction of the peripheral airways, which correlated well
    with the increased plasma concentrations of sulfites. The authors
    suggested that the rapid onset of symptoms and the lack of atopy
    markers to sulfites, as established by negative results in tests for
    skin hypersensitivity, are consistent with mediation of effects
    through the parasympathetic system (Acosta et al., 1989).

         A group of patients with chronic asthma and histories suggestive
    of sulfite sensitivity was studied in order to determine whether the
    ingestion of aqueous potassium metabisulfite was associated with
    degranulation of mast cells, as measured by the release of the
    neutrophil chemotactic factor of anaphylaxis. Single-blind, oral
    aqueous challenges of up to a maximum of 200 mg potassium
    metabisulfite were given to 13 patients. Serum samples were obtained
    from all the patients before the challenge and for 180 min afterwards.
    The samples were tested for the presence of neutrophil chemotactic
    factor by both a 51Cr microchamber chemotaxis assay and a leukocyte
    polarization technique. Six of the patients gave positive responses,
    as indicated by a fall in FEV1 > 20% of prechallenge values. No
    significant increase in the amount of neutrophil chemotactic factor
    was detected in post-challenge serum samples from any patient. Skin
    testing with potassium metabisulfite in 10 of the patients yielded
    uniformly negative reactions. The authors concluded that sensitivity
    to aqueous metabisulfite is not associated with mast-cell
    degranulation in patients in whom the result of a skin test is
    negative (Sprenger et al., 1989).

         Evidence for an IgE-mediated reaction to oral challenge with
    metabisulfite was reported in a 34-year-old woman with a long-standing
    history of allergic rhinitis, nasal polyposis, and recurrent
    sinusitis, who had reported reactions to restaurant meals on several
    occasions. Various allergies had been confirmed by skin-prick testing
    and did not include foods eaten at restaurants. Her serum IgE
    concentrations were within the normal range. Repeated oral challenges
    with sodium metabisulfite resulted in a consistent spectrum of
    reactions with doses of 50 mg by gelatin capsule or 1 mg in lemonade,
    which consisted of nasal congestion, profuse rhinorrhoea, swelling of
    the face and lips, and urticarial reactions on the hands, antecubital
    fossae, soles of the feet, and midriff. Close inspection revealed
    swelling of the nasal polyps. These symptoms were resolved after
    administration of epinephrine. Pulmonary function was unaffected in
    this patient after the metabisulfite challenge. Subsequent oral
    challenges with 10 mg metabisulfite at six months produced similar
    results. Skin-prick testing induced a positive reaction to
    metabisulfite, as did a basophil histamine release test. This case of
    adverse reaction to metabisulfite is one of the few in which
    convincing evidence was found for an IgE-mediated mechanism. The
    authors indicate that the case is somewhat unusual in that the patient
    was not asthmatic and asthma was not part of the clinical reaction;
    while it is probably quite rare, IgE-mediated sulfite sensitivity
    obviously exists (Sokol & Hydick, 1990).

         A two-year old boy in whom asthma had been diagnosed at the age
    of two months experienced attacks of wheezing in association with
    ingestion of foods containing sulfites. He also had a history of
    adverse food reactions (hives) to peanuts and milk and showed
    immediate skin test hypersensitivity to inhaled allergens. In order to
    confirm that sulfites were the precipitating agent in the attacks of
    wheezing, a double-blind oral challenge test was conducted with
    graduated doses of powdered potassium metabisulfite mixed with apple

    sauce. Reaction to the challenge was assessed by measuring pulse,
    respiratory rate, and blood pressure. A reaction to a dose of 25 mg
    was observed after 2 min, consisting of an increase in all the
    measured parameters and accompanied by cough, profuse perspiration,
    and wheezing. No reaction was observed to challenges with placebo. On
    a separate occasion, intradermal testing with metabisulfite resulted
    in a wheal-and-flare reaction comparable to that with histamine,
    although the result of previous skin-prick tests had been negative.
    The significance of this result was considered equivocal in light of
    the fact that an identical response had been obtained in a control
    without asthma who did not have a history of wheezing after ingestion
    of foods containing sulfites (Frick & Lemanske, 1991).

         A series of oral challenges was carried out with metabisulfite in
    a 22-year-old woman with a history of seasonal rhinoconjunctivitis due
    to sensitivity to grass pollen, who had experienced episodes of
    urticaria and angio-oedema affecting the face, neck, and upper chest
    and difficulty in speaking (dysphonia), without respiratory
    difficulties or gastrointestinal symptoms, after ingestion of a number
    of grape-based beverages, salad-bar items, and shellfish. The subject
    had a normal blood count, including eosinophils, total serum IgE at
    the high end of the normal range, and negative results in skin-prick
    tests for several foods and positive results for grass pollen.
    Skin-prick and intradermal tests showed no sensitivity to potassium
    metabisulfite. Pulmonary function was normal. A positive response was
    induced with a 25-mg challenge, with urticaria on the face and upper
    thorax, nasal itching, rhinorrhoea, and dysphonia. The symptoms were
    resolved by the administration of epinephrine. A challenge one week
    later with 25 mg potassium metabisulfite 30 min after administration
    of chromolyn sodium induced a similar positive response. Two months
    later, the same challenge given 90 min after administration of 5000 µg
    cyanocobolamin (vitamin B12) induced the same symptoms as seen
    previously, but they were less intense. Consequently, the patient's
    urticaria was considered to have been sulfite-mediated. The authors
    indicated that the clinical manifestations in the subject suggested an
    IgE-mediated mechanism, but the results of skin tests were negative
    and the oral challenge with metabisulfite was not inhibited by prior
    administration of cromolyn sodium. The authors were unable to identify
    a pathogenic mechanism (Belchi-Hernandez et al., 1993).

         Three further cases of adverse reactions to ingested sulfites
    were reported. The first was in a 45-year-old woman who had had
    bronchial asthma and rhinitis during the previous year and who
    developed anaphylactoid syndrome during a restaurant meal, with flush,
    generalized urticaria, severe wheezing, and dyspnoea, leading to
    shock. The blood eosinophil count was elevated, but the serum IgE was
    within the normal range. A scratch test with sodium metabisulfite
    solution resulted in a strongly positive reaction, and the diagnosis
    was an IgE-mediated allergy to sulfites in an asthma patient. 

         The second case involved a 36-year-old woman with a history of
    intrinsic asthma and rhinitis, who experienced generalized itching,
    feelings of heat, tachycardia, angio-oedema of the face and lips, and
    chest tightness while eating a cafeteria meal. A prick test for sodium
    bisulfite gave negative results. An oral challenge test with sodium
    metabisulfite in capsules resulted in a decreased FEV1 when a dose of
    25 mg was reached. This was accompanied by nasal congestion, a
    sensation of heat in the head, redness of the fingers, and shortness
    of breath. The diagnosis was an anaphylactoid reaction due to sulfite

         In the third case, the patient was a 37-year-old man who had
    experienced recurrent, severe episodes of acute urticaria,
    angio-oedema, and dyspnoea, mainly during the night, at least once
    every three months for the previous year. The IgE level was somewhat
    elevated (180 kU/L). Single-blind challenge tests were performed
    during a symptom-free period. One hour after a single dose of 50 mg
    sodium metabisulfite, urticaria appeared on the hands, with
    progression and generalization, despite treatment with antihistamines
    and cortisone. A biopsy taken 5 h after the appearance of urticaria
    indicated a leukocytoclastic vasculitis with eosinophilia, confirming
    the diagnosis of sulfite-induced urticarial vasculitis (Wüthrich 1993;
    Wüthrich et al., 1993).

         A 25-year-old man experienced numerous episodes of itchy
    erythematous rash and swelling in the face within minutes of consuming
    various brands of wine and a particular brand of beer. When an oral
    challenge test was performed with the beer, 100 ml were found to
    provoke the same reaction, accompanied by a statistically significant
    increase in serum histamine, as detected by radioimmunoassay.
    Identical symptoms were observed after a dose of 10 mg sodium
    metabisulfite. No specific IgE or IgG antibodies were detected in the
    patient's serum by a radioimmunosorbent test. The beer was found to
    contain 3-4 mg/L of sulfite. As the metabisulfite challenge provoked
    the same symptoms as those encountered by the patient when he ingested
    the particular brand of beer, sulfite was considered to be the most
    likely causative agent. The authors suggested that the dose-dependence
    and the release of histamine in the absence of specific antibodies
    indicate a diagnosis of intolerance rather than allergy (Gall et al.,

         A 53-year old woman was given a single-blind oral challenge test
    with 200 mg sodium bisulfite after having experienced several episodes
    of periorbital oedema in response to eye drops. Intramuscular
    treatment of the oedema with dexamethasone containing metabisulfite
    exacerbated the periorbital oedema and erythema, which was relieved by
    administration of antihistamine medications. The patient had no
    history of rhinoconjunctivitis or lower respiratory symptoms, and
    negative responses were elicited to 80 common inhaled allergens in
    skin-prick tests. Twelve hours after ingestion of sodium bisulfite,
    periorbital erythemous oedema developed in her left eye, which
    disappeared with antihistamine medication. Avoidance of

    sulfite-containing foods and medications was found to prevent
    recurrences (Park & Nahm, 1996).

    2.3.2  Food challenges

         Three men and five women aged 25-55, with asthma, who had been
    identified as sulfite-sensitive on the basis of double-blind capsule
    or beverage challenges, were challenged with various sulfited foods.
    Positive responses were measured by reductions in the FEV1. The first
    challenge was conducted double-blind with sulfited lettuce; subsequent
    challenges comprised fresh mushrooms, dried apricots, white grape
    juice, dehydrated potatoes, and shrimp. Four subjects reacted to the
    lettuce challenge, and three of these reacted to one or more of the
    other sulfited foods. Several of these individuals had positive
    responses to skin-prick and intradermal tests with the bound and free
    forms of sulfite, suggesting an immunological basis for their
    hypersensitivity to sulfites. One subject had a history of reacting to
    acidic solutions of sulfite and not to capsules. This patient reacted
    only to sulfited lettuce, which, because of its composition, contains
    no bound sulfites. The remaining four subjects failed to react to any
    of the food challenges. The authors noted the variation in the
    responses to sulfited foods other than lettuce. In addition, the
    likelihood of a reaction to a particular sulfited food was not
    correlated with the amount of sulfite ingested from that food (Taylor
    et al., 1988). 

         A double-blind placebo-controlled study with sulfited lettuce was
    performed with four women and one man, aged 25-71 with asthma, who had
    previously given positive responses to sulfite capsule challenges.
    Four of the subjects had a history of atopy to various aeroallergens.
    All of the patients reacted positively to sulfite-treated lettuce, as
    indicated by a reduction in pulmonary function. Two of these
    experienced severe reactions. Other symptoms experienced were flushing
    and itching around the mouth, throat, and skin. None of the patients
    responded to placebo-treated lettuce (Howland & Simon, 1989).

         A study was conducted to determine whether asthma attacks
    precipitated in 10 males and eight females aged 12-23 years
    immediately after consumption of pickled onions were due to the
    presence of sulfite. All had a history of allergy to airborne
    allergens, but only two were steroid-dependent. Most of them did not
    react to other potential sulfite-containing foods such as salads,
    vinegar, fruit juices, beer, cider, grape juice, shellfish, and
    several pickled vegetables, nor did they react to raw or cooked onion.
    A single-blind challenge test was conducted with solutions of
    metabisulfite in lemon juice, the pH being adjusted to 4.2 and 3.3 to
    deliver doses up to 150 mg in stepwise increments. A result was
    considered positive when the drop in FEV1 was > 20% A positive
    response to metabisulfite at pH 4.2 was observed in six of the
    patients and at pH 3.3 in four others. Skin-prick tests with
    metabisulfite, pickled onions, the preserving liquid, and raw onions
    gave negative results in all patients. Seven of the patients
    subsequently consented to undergo a challenge with pickled onions:

    three gave a positive response. All of these patients had reacted to
    one of the metabisulfite challenges. The sulfite concentration in the
    pickled onions in question was determined to be 800-1200 ppm, much
    higher than in other varieties of pickled onion (< 200 ppm) and
    exceeding the permitted concentration (100 ppm). Decreasing the pH of
    the metabisulfite solution resulted in a 2.5-fold increase in the
    amount of sulfur dioxide released. The authors suspected that
    inhalation of sulfur dioxide was the mechanism for precipitation of
    the asthma attacks (Gastaminza et al., 1995).

    2.3.3  Prevalence studies  Adults

         In a prospective single-blind screening study, 120
    non-steroid-dependent and 83 steroid-dependent patients with asthma
    were challenged orally with capsules of potassium metabisulfite. On
    the basis of a > 20% reduction in FEV1 within 30 min of the oral
    challenge, five of the non-steroid-dependent and 16 of the
    steroid-dependent patients were judged to have had a positive
    response. The non-steroid-dependent and seven of the steroid-dependent
    responders were re-challenged with increasing doses of potassium
    metabisulfite capsules in a double-blind study. In this protocol, a
    considerable number of the patients who gave positive reactions in the
    single-blind study did not react: positive responses were seen in one
    of the five non-steroid-dependent patients and three of seven
    steroid-dependent patients. The authors therefore estimated the
    prevalence of sulfite sensitivity in their overall study population to
    be 3.7%. They considered this figure to be an overestimate, since
    there was a larger percentage of steroid-dependent patients in their
    study population than in the general population of asthma patients.
    Less than 1% of the non-steroid-dependent asthmatics were affected,
    while the figure was closer to 8% in the steroid-dependent group,
    indicating that the latter are at greater risk for sulfite sensitivity
    (Bush et al., 1986). 

         Oral challenge tests were given to 44 non-atopic patients with
    steroid-dependent bronchial asthma (14 males, 30 females; aged 14-74),
    with incre-mental doses of sodium metabisulfite in solution and in
    capsules for the higher doses. The participants had no clinical
    history of sulfite sensitivity. A single-blind challenge protocol was
    used for 22 of the patients and a double-blind protocol for the
    remaining 22 patients; those who reacted in the single-blind challenge
    were re-tested in a double-blind protocol. A positive response was
    considered to be a reduction of > 20% in the FEV1. Of six
    initially positive responses, only two were confirmed by double-blind
    testing. These results suggest that the prevalence of sulfite
    sensitivity is 4.5% in steroid-dependent asthma patients (Prieto et
    al., 1988).

         A study was undertaken among adult Melanesians with recurrent
    asthma in Papua-New Guinea to determine whether common food additives
    had precipitated asthma in this population. Two female and five male
    patients, including three who were steroid-dependent, with a mean age
    of 34 years (range, 22-50), received sodium metabisulfite dissolved in
    freshly squeezed, pure orange juice, or placebo, in a double-blind
    procedure. The solution was swilled around the mouth for 5 s before
    being swallowed. Graduated doses of 5-200 mg metabisulfite were used
    for the challenge. A positive response was considered to be a decrease
    of > 20% in FEV1. Positive responses were demonstrated in three of
    the patients, including one who was steroid-dependent, all after the
    200-mg dose (Timberlake et al., 1992). 

         False-positive results may be generated in asthma patients by
    oral challenge owing to the fact that hyperactive airways and
    bronchial hypersensitivity may be exacerbated by the common practice
    in trials of withholding all bronchodilators on the day of challenge.
    The author was unable to establish cross-sensitivity to sulfites in 70
    patients with documented sensitivity to aspirin, and none of 33
    sulfite-sensitive patients were found to be sensitive to aspirin. It
    was suggested that accounts of cross-sensitivity of sulfite-sensitive
    patients to aspirin and to other food additives had not been confirmed
    by oral challenges (Simon, 1994).

         The incidence of delayed hypersensitivity reactions to sulfites
    was investigated in a series of 2894 patients with eczematous
    dermatitis. The patients were subjected to a series of patch tests,
    starting with sodium metabisulfite, and positive reactions to sodium
    metabisulfite were elicited in 1.7% (50 patients). Patients with a
    positive reaction were subsequently tested with sodium sulfite, sodium
    bisulfite, and potassium metabisulfite. Those who reacted to sodium
    metabisulfite also reacted to potassium metabisulfite and sodium
    bisulfite, while only two had positive reactions to sodium sulfite.
    Prick tests and intradermal tests with a sodium metabisulfite solution
    were carried out on 20 patients, and an oral challenge with doses of
    30 and 50 mg sodium metabisulfite was administered to five patients.
    None of the subjects who reacted to the patch test reacted to either
    the prick or intradermal tests, and no flare-ups of dermititis or
    patch site were precipitated by the oral challenge (Vena et al.,
    1994).  Children

         Sodium metabisulfite was administered to 29 children with chronic
    asthma in a single-blind challenge. The children ranged in age from
    5.5 to 14 years and were essentially evenly divided by sex; 28 of the
    29 were atopic for airborne allergens. A positive response was judged
    by a reduction in pulmonary function. The patients were challenged
    both with capsules at doses up to 100 mg and citric acid solution
    containing sodium metabisulfite at doses up to 50 mg. Metabisulfite
    hypersensitivity was detected in 19 children; all reacted to
    metabisulfite in solution, and none responded when it was administered
    in capsule form. Most of the responders experienced immediate

    reactions, which consisted initially of a burning sensation in the
    throat, tight cough, wheezing, and signs of respiratory distress.
    Seven of the 19 had a history which suggested sensitivity to
    metabisulfite in foods. The authors considered that the lack of
    response to metabisulfite in capsules and the rapid onset of bronchial
    symptoms suggested that inhalation of sulfur dioxide was the trigger
    (Towns & Mellis, 1984).

         The prevalence of sulfite sensitivity was determined in a group
    of 51 children with chronic asthma, who were recruited from an allergy
    clinic. They ranged in age from five to 16 years, with a mean age of
    10.3 years. Nearly 90% of the children were atopic, and 18% were
    steroid-dependent. Potassium metabisulfite was administered in
    lemonade in an open challenge, in graduated doses of 0.5-100 mg.
    Eighteen of the 51 reacted with a decrease > 20% in FEV1 after
    drinking the metabisulfite solution. Placebo challenges given to 14 of
    the 18 subjects resulted in one positive reaction. There was no
    difference in the prevalence of sulfite sensitivity between
    steroid-dependent and non-steroid-dependent subjects. All of the
    patients who reacted to metabisulfite solution but only two-thirds of
    those who did not react were reported to be sensitive to smog
    (Friedman & Easton, 1987).

         Six children aged five months to 14 years with a history of
    severe atopic dermatitis were tested for their reactions to sodium
    metabisulfite in a double-blind, placebo-controlled challenge, as part
    of a larger study of foods and food additives. The test compound was
    administered by nasogastric tube. Two of the children reacted
    positively to the challenge, the reactions consisting of exacerbation
    of isolated pruritis and redness of the skin. No respiratory symptoms
    were seen (Van Bever et al., 1989).

         Bronchorestrictive airway responses were evaluated in a group of
    35 boys and 21 girls aged 6-14 years with chronic asthma, by open
    challenge after ingestion of incremental doses of potassium
    metabisulfite solution or capsules. All but two of the children had a
    history of allergy to various agents, but none showed allergy to
    metabisulfite in a skin-prick test. When the challenge was given with
    capsules, positive responses (reduction in pulmonary function) were
    noted in four of the children, only at the maximum challenge of 200
    mg. Two children who had not reacted to the challenge with
    encapsulated metabisulfite responded positively to metabisulfite
    solution. None had systemic symptoms (flushing, tingling, and/or
    faintness) or hypotensive symptoms after metabisulfite challenge. All
    the patients who reacted positively to either challenge had severe to
    moderate bronchial reactivity to methacholine, while those who did not
    react had only mild reactivity to methacholine (Boner et al., 1990).

         The prevalence of sensitivity to sulfite was determined in a
    group of 15 boys and five girls with steroid-dependent asthma. Only
    one child was suspected of intolerance to this agent. A single-blind
    oral challenge test was performed with increasing doses of sodium
    metabisulfite solution. The solution was held in the mouth for 10 s

    before being swallowed, and FEV1 was evaluated 10 min later. Children
    who did not react were re-tested for up to 90 min to evaluate possible
    late reactions. A double-blind challenge was performed on those who
    reacted. The single-blind challenge resulted in positive responses in
    six of the children; four of these cases were confirmed in the double-
    blind study. The authors postulated a prevalence of 20% among children
    with steroid-dependent asthma (Sanz et al., 1992).

         The prevalence of sensitivity to sulfite was determined in 14
    girls and 23 boys, aged 5-13 years, with chronic asthma. Five of the
    children were steroid-dependent. A double-blind challenge was
    conducted, in which the children received a dose of 66 mg sodium
    metabisulfite (equivalent to 41 mg sulfur dioxide) dissolved in apple
    juice; a control group of 22 children received pure apple juice.
    Response to the challenge was determined on the basis of a 10% or
    > 20% drop in FEV1. Eight of the 37 children experienced a drop of
    > 20%, consistent with the rates in boys only, in girls only, and
    in steroid-dependent asthma patients only (Steinman et al., 1993).


         The Committee identified two main issues in the toxicological
    evaluation of sulfites: general toxicity and idiosyncratic
    intolerance. The latter does not appear to be related to sulfite
    oxidase deficiency in humans. Since general toxicity and idiosyncratic
    intolerance appeared to be unrelated, they were considered separately.

     General toxicity

         Studies performed since the thirtieth meeting have demonstrated
    the quantitative conversion of sulfite ion to sulfate ion. Release of
    reduced glutathione by both isolated hepatocytes and perfused liver is
    probably due to sulfitolysis of oxidized glutathione by sulfite ion.
    The other product of sulfitolysis, glutathione  S-sulfonate, was
    found to be a strong competitive inhibitor of both microsomal and
    cytosolic glutathione  S-transferase in rat lung and liver cells and
    human lung tumour cells. These results suggest that sulfite ion could
    interfere with the glutathione pathway through its reaction with
    oxidized glutathione. Dietary administration of 2.5% metabisulfite to
    rats for five weeks increased the content of three disaccharidases
    involved in carbohydrate metabolism and of alkaline phosphatase in the
    brush-border membrane of small-intestinal cells.

         The short-term toxicity of bound and free forms of sulfite
    (acetaldehyde hydroxysulfonate and sodium metabisulfite, respectively)
    administered in drinking-water was assessed in normal rats and rats
    made sulfite oxidase-deficient by administration of tungstate ion,
    also in drinking-water, for eight weeks. The sulfite oxidase-deficient
    rats were considered to be a better model for humans than conventional
    rats, which have an estimated 10-20 times higher concentration of this
    enzyme in the liver. Acetaldehyde hydroxysulfonate is a major bound
    form of sulfite in wines and other fermented foods and beverages and
    is considered to be a very stable form of bound sulfite. The doses of

    bound and free sulfite chosen were calculated to deliver the same dose
    in terms of sulfur dioxide equivalents (7, 70, or 350 mg/kg bw per
    day, the highest dose being reduced to 175 mg/kg bw per day after
    three weeks). The effects on body weight, food consumption, and water
    intake were strongest in rats receiving free rather than bound sulfite
    and in enzyme-deficient as compared with normal rats. The
    concentrations of sulfite and thiosulfate in urine and of
     S-sulfonates in plasma were elevated in untreated, sulfite
    oxidase-deficient rats. Administration of bound sulfite to these rats
    resulted in increased excretion of sulfite in the urine, while
    administration of free sulfite was associated with increases in the
    concentrations of urinary thiosulfate and plasma  S-sulfonates. The
    measured concentrations were variable and were not clearly related to
    dose. Histopathological lesions of the forestomach and glandular
    stomach were detected in all groups of rats treated at the high dose.
    The severity and frequency of the gastric lesions were greater in the
    sulfite oxidase-deficient rats. Hepatic lesions were observed in
    animals receiving acetaldehyde hydroxysulfonate, and the NOEL was
    lower in the sulfite oxidase-deficient rats than in normal rats (7 and
    70 mg/kg bw per day of sulfur dioxide equivalents, respectively). The
    effect may have been due to acetaldehyde formed after dissociation.
    The results showed that the gastric toxicity of bound sulfites, in the
    form of acetaldehyde hydroxysulfonate, is equivalent to that of free
    sulfite: while the enzyme-deficient rats were more sensitive at doses
    above the NOEL, the NOEL was the same as that in normal rats.

         The results of three assays for genotoxicity  in vitro were
    reviewed, which confirmed the previously noted observation that
    sulfites are genotoxic  in vitro but not  in vivo. This conclusion
    is consistent with the high reactivity of sulfite and its rapid
    inactivation in mammals. Two studies of teratogenicity involving
    dietary administration of sulfite were available: neither was
    completely satisfactory, as treatment did not cover the entire period
    of organogenesis. Maternal toxicity was demonstrated in both studies
    at doses equivalent to 840 mg/kg bw per day of sulfur dioxide
    equivalent or higher. No teratogenic effects were noted at intakes up
    to this concentration.

         The results of a study of the effects of ingested sulfite on the
    rat kidney demonstrated that administration of sodium metabisulfite by
    gavage at a dose of 5 mg/kg bw per day (3.4 mg/kg bw per day of sulfur
    dioxide equivalent) for up to two weeks reduced alkaline phosphatase
    activity in renal tissue and concomitantly increased the serum and
    urinary concentrations of the enzyme; the effects occurred after the
    first dose. The activity of lactate dehydrogenase was likewise
    decreased in renal tissue and was increased in urine but not in serum.
    Urinary excretion of protein was increased 10-fold by the end of two
    weeks. These effects would appear not to be related to treatment, as
    sulfite is quickly and quantitatively metabolized to sulfate and no
    sulfite was detected in urine, nor were lesions of the kidney detected
    after eight weeks in normal rats receiving 20-50 times higher doses of
    sulfite in drinking-water. 

         The previous NOEL in studies of animals that ingested sulfite was
    confirmed in an eight-week study in which the NOEL for gastric lesions
    was 70 mg/kg bw per day of sulfur dioxide equivalents in both normal
    and sulfite oxidase-deficient rats, whether they were fed free sulfite
    or a stable and prevalent form of bound sulfite, acetaldehyde
    hydroxysulfonate. The gastric effects of sulfite reported in rats and
    pigs arise from local irritation. These effects would therefore be
    dependent on concentration rather than dose, and a numerical ADI might
    not be appropriate. The Committee considered that the role of
    acetaldehyde in the reported hepatic effects of acetaldehyde
    hydroxysulfonate should be resolved before a re-evaluation of the
    safety could be completed. In view of these reservations, the
    previously established ADI of  0-0.7 mg/kg bw was maintained.

     Idiosyncratic reactions in humans

         A number of human case studies were available in which suspected
    hypersensitivity to sulfites based on adverse reactions to foods was
    confirmed by administration of single- or double-blind oral challenges
    with sulfites in solution or capsule form. While many of the cases
    involved individuals with chronic asthma whose response was primarily
    respiratory, a number of reports described adverse allergic-type
    responses in people without asthma, which did not involve respiratory
    symptoms. Some of these individuals had chronic allergic conditions.
    The responses were consistent with self-reported reactions documented
    by the United States Food and Drug Administration's Adverse Reaction
    Monitoring System. Evidence for an allergic basis for the adverse
    reactions to free and bound sulfites was provided in a number of the
    case reports; in studies in which only skin-prick testing had been
    attempted, allergy could not be ruled out, as further testing had not
    been done. Challenges with various sulfited food commodities also
    resulted in positive responses. In a double-blind study, sulfited
    lettuce elicited the most consistent responses; sulfite in
    sulfite-treated lettuce is considered to be present mostly as free
    sulfite. While the mechanism by which sulfite ingestion precipitates
    idiosyncratic adverse reactions has not been established, some
    evidence was available that the adverse effects of ingested sulfites
    are mediated through effects of nitric oxide on parasympathetic

         The prevalence of sulfite sensitivity was determined in groups of
    steroid-dependent and non-steroid-dependent adults and children with
    asthma. In adults, the prevalence was 4-8% among steroid-dependent
    asthmatic patients and appeared to be less than 1% among non-steroid
    dependent patients. The prevalence in the total adult asthmatic
    population has been estimated to be 4%, but the prevalence in
    asthmatic children was higher, approximately 20-30%, after
    double-blind challenges to both steroid-dependent and
    non-steroid-dependent asthmatic children. 

         The Committee reiterated the recommendation made at its thirtieth
    meeting (Annex 1, reference 73) that, when a suitable alternative
    method of preservation exists, its use should be encouraged,
    particularly in those applications (e.g. control of enzymic browning
    in fresh salad vegetables) in which the use of sulfites may lead to
    high levels of acute intake and which have most commonly been
    associated with life-threatening adverse reactions. The Committee
    considered that appropriate labelling would help in alerting
    individuals who cannot tolerate sulfites.

         The intake of sulfites is assessed in "Evaluation of National
    Assessments of Intake of Sulfites".


    Acosta, R., Granados, J., Mourelle, M., Perez-Alvarez, V. & Quezada,
    E. (1989) Sulfite sensitivity: Relationship between sulfite plasma
    levels and bronchospasm: case report.  Ann. Allergy, 62, 402-405.

    Akanji, M.A., Olagoke, O.A. & Oloyede, O.B. (1993) Effect of chronic
    consumption of metabisulphite on the intergrity of the rat kidney
    cellular system.  Toxicology, 81, 173-179.

    Belchi-Hernandez, J., Florido-Lopez, F., Estrada-Rodriguez, J.L.,
    Martinez-Alzamora, F., Lopez-Serrano, C. & Ojeda-Casas, J.A. (1993)
    Sulfite-induced urticaria.  Ann. Allergy, 71, 230-232.

    Boner, A.L., Guarise, A., Vallone, G., Fornari, A., Piacentini, F. &
    Sette, L. (1990) Metabisulfite oral challenge: Incidence of adverse
    responses in chronic childhood asthma and its relationship with
    bronchial hyperactivity.  Allergy Clin. Immunol., 85, 479-483.

    Botey, J., Cozzo, M., Eseverri, J.L. & Marin, A. (1987) Sulfites and
    skin pathology in children.  Allergol. Immunopathol., 15, 365-367. 

    Bush, R.K., Taylor, S.L., Holden, K., Nordlee, J.A. & Busse, W.W.
    (1986) Prevalence of sensitiviy to sulfiting agents in asthmatic
    patients.  Am. J. Med., 81, 816-820.

    Ema, M., Itami, T. & Kanoh, S. (1985) Effect of potassium
    metabisulfite on pregnant rats and their offspring.  J. Food Hyg. 
     Soc. Jpn, 26, 454-459.

    Frick, W.E. & Lemanske, R.F. (1991) Oral sulfite sensitivity and
    provocative challenge in a 2 year old.  J. Asthma, 28, 221-224.

    Friedman, M.E. & Easton, J.G. (1987) Prevalence of positive
    metabisulfite challenges in children with asthma.  Pediatr. Asthma 
     Aller. Immunol., 1, 53-59.

    Gall, H., Boehncke, W.-H. & Gietzen, K. (1996) Intolerance to sodium
    metabisulfite in beer.  Allergy, 51, 516-517. 

    Gastaminza, G., Quirce, S., Torres, M., Tabar, A., Echechipía, S.,
    Muñoz, D. & Fernandez de Corres, L. (1995) Pickled onion-induced
    asthma: A model of sulfite-sensitive asthma?  Clin. Exp. Allergy, 25,

    Harvey, S.B. & Nelsestuen, G.L. (1995) Reaction on nitric oxide and
    its derivatives with sulfites: A possible role in sulfite toxicity.
     Biochim. Biophys. Acta, 1267, 41-44.

    Howland, W.C., III & Simon, R.A. (1989) Sulfite-treated lettuce
    challenges in sulfite-sensitive subjects with asthma.  J. Allergy 
     Clin. Immunol., 83, 1079-1082. 

    Hui, J.Y., Beery, J.T., Higley, N.A. & Taylor, S.L. (1989) Comparative
    subchronic oral toxicity of sulphite and acetaldehyde
    hydroxysulphonate in rats.  Food Chem. Toxicol., 27, 349-359.

    Itami, T., Ema, M., Kawasaki, H. & Kanoh, S. (1989) Evaluation of
    teratogenic potential of sodium sulfite in rats.  Drug Chem. 
     Toxicol., 12, 123-135.

    Leung, K.-H., Post, G.B. & Menzel, D.B. (1985) Glutathione
     S-sulfonate, a sulfur dioxide metabolite, as a competitive inhibitor
    of glutathione  S-transferase, and its reduction by glutathione
    reductase.  Toxicol. Appl. Pharmacol., 77, 388-394.

    Meng, Z. & Zhang, L. (1992) Cytogenetic damage induced in human
    lymphocytes by sodium bisulfite.  Mutat. Res., 298, 63-69.

    Park, H.-S. & Nahm, D. (1996) Localized periorbital edema as a
    clinical manifestation of sulfite sensitivity.  J. Korean Med. Sci., 
    11, 356-357.

    Prieto, L., Juyol, M., Paricio, A., Martinez, M.A., Palop, J. &
    Castro, J. (1988) Oral challenge test with sodium metabisulfite in
    steroid-dependent asthmatic patients.  Allergol. Immunopathol., 16,

    Rodriguez-Vieytes, M., Martinez-Sapiña, J., Taboada-Montero, C. &
    Lama-Aneiros, M.A. (1994) Effect of sulfite intake on intestinal
    enzyme activity in rats.  Gastroenterol. Clin. Biol., 18, 306-309.

    Sanz, J., Martorell, A., Torro, I., Cerda, J-C. & Alvarez, V. (1992)
    Intolerance to sodium metabisulfite in children with steroid-dependent
    asthma.  J. Invest. Allergol. Clin. Immunol., 2, 36-38.

    Selner, J., Bush, R., Nordlee, J., Wiener, M., Buckley, J., Koepke,
    J.W.. & Taylor, S. (1987) Skin reactivity to sulfited foods in a
    sulfite-sensitive asthmatic.  J. Allergy Clin. Immunol., 79, 241.

    Simon, R.A. (1994) Oral challenges to detect aspirin and sulfite
    sensitivity in asthma.  Allerg. Immunol., 26, 216-218.

    Simon, R.A. & Wasserman, S.I. (1986) IgE mediated sulfite sensitive
    asthma.  J. Allergy Clin. Immunol., 77, 157.

    Sokol, W.N. & Hydick, I.B. (1990) Nasal congestion, urticaria, and
    angioedema caused by an IgE-mediated reaction to sodium metabisulfite.
     Ann. Allergy, 65, 233-238.

    Sprenger, J.D., Altman, L.C., Marshall, S.G., Pierson, W.E. & Koenig,
    J.Q. (1989) Studies of neutrophil chemotoactic factor of anaphylaxis
    in metabisulfite sensitivity.  Ann. Allergy, 62, 117-121.

    Steinman, H.A., Le Roux, M. & Potter, P.C. (1993) Sulphur dioxide
    sensitivity in South African asthmatic children.  S. Afr. Med. J., 
    83, 387-390. 

    Sun, Y., Cotgreave, I., Lindeke, B. & Moldéus, P. (1989) The
    metabolism of sulfite in liver: Stimulation of sulfate conjugation and
    effects on paracetamol and allyl alcohol toxicity.  Biochem. 
     Pharmacol., 38, 4299-4305.

    Takahashi, M., Hasegawa, R., Furukawa, F. Toyoda, K., Sato, H. &
    Hayashi, Y. (1986) Effects of ethanol, potassium metabisulfite,
    formaldehyde and hydrogen peroxide on gastric carcinogenesis in rats
    after initiation with N-methyl-N'-nitro-N-nitrosoguanidine.  Jpn. J. 
     Cancer Res. (Gann), 77, 118-124.

    Taylor, S.L., Bush, R.K., Selner, J.C., Nordlee, J.A., Wiener, M.B.,
    Holden, K., Koepke, J.W. & Busse, W.W. (1988) Sensitivity to sulfited
    foods among sulfite-sensitive subjects with asthma.  J. Allergy 
     Clin. Immunol., 81, 1159-1167.

    Timberlake, C.M., Toun, A.K. & Hudson, B.J. (1992) Precipitation of
    asthma attacks in Melanesian adults by sodium metabisulphite.  PNG 
     Med. J., 35, 186-190.

    Tollefson, L. (1988) Monitoring adverse reactions to food additives in
    the US Food and Drug Administration.  Regul. Toxicol. Pharmacol., 8,

    Towns, S.J. & Mellis, C.M. (1984) Role of acetyl salicylic acid and
    sodium metabisulfite in chronic childhood asthma.  Pediatrics, 73,

    Tsevat, J., Gross, G.N. & Dowling, G.P. (1987) Fatal asthma after
    ingestion of sulfite-containing wine.  Ann. Intern. Med., 102, 263.

    Van Bever, H.P., Docx, M. & Stevens, W.J. (1989) Food and food
    additives in severe atopic dermatitis.  Allergy, 44, 588-594.

    Vena, G.A. Foti, C. & Angelini, G. (1994) Sulfite contact allergy.
     Contact Dermatitis, 31, 172-175.

    Wüthrich, B. (1993) Adverse reactions to food addtives.  Ann. 
     Allergy, 71, 379-384.

    Wüthrich, B., Kägi, M.K. & Hafner, J. (1993) Disulfite-induced acute
    intermittent urticaria with vasculitis.  Dermatology, 187, 290-292.

    Yang, W.H., Purchase, E.C.R. & Rivington, R.N. (1986) Positive skin
    tests and Prausnitz-Küstner reactions in metabisulfite-sensitive
    subjects.  J. Allergy Clin. Immunol., 78, 443-449.

    See Also:
       Toxicological Abbreviations