IPCS INCHEM Home

    CYANOGENIC GLYCOSIDES

    First draft prepared by
    Dr G. Speijers,
    National Institute of Public Health and Environmental Protection
    Laboratory for Toxicology
    Bilthoven, The Netherlands

    1.  EXPLANATION

         Cyanogenic glycosides are phytotoxins which occur in at least
    2000 plant species, of which a number of species are used as food in
    some areas of the world. Cassava and sorghum are especially
    important staple foods containing cyanogenic glycosides (Conn,
    1979a,b; Nartey, 1980; Oke,1979, 1980; Vennesland  et al., 1982;
    Rosling, 1987). There are approximately 25 cyanogenic glycosides
    known. The major cyanogenic glycosides found in the edible parts of
    plants used for human or animal consumption are summarized in Table
    1. The potential toxicity of a cyanogenic plant depends primarily on
    the potential that its consumption will produce a concentration of
    HCN that is toxic to exposed animals or humans (see Table 2).
    Several factors are important in this toxicity:

         The first aspect is the processing of plant products containing
    cyanogenic glycosides. When the edible parts of the plants are
    macerated, the catabolic intracellular enzyme ▀-glucosidase can be
    released, coming into contact with the glycosides. This enzyme
    hydrolyzes the cyanogenic glycosides to produce hydrogen cyanide and
    glucose and ketones or benzaldehyde. The hydrogen cyanide is the
    major toxic compound causing the toxic effects. Plant products
    (notably cassava), if not adequately detoxified during the
    processing or preparation of the food, are toxic because of the
    release of this preformed hydrogen cyanide.

         The second aspect is the direct consumption of the cyanogenic
    plant. Maceration of edible parts of the plants as they are eaten
    can release ▀-glucosidase. The ▀-glucosidase is then active until
    the low pH in the stomach deactivates the enzyme. Additionally, it
    is possible that part of the enzyme fraction can become reactivated
    in the alkaline environment of the gut. At least part of the
    potential hydrogen cyanide is released, and may be responsible for
    all or part of the toxic effect of cyanogenic glycosides in the
    cases of some foods.

        Table 1:  The occurrence of cyanogenic glycosides in major edible plants
              (Conn, 1979a,b)
                                                                                       

    Cyanogenic                           Plant species
    glycosides               Common name                 Latin name
                                                                                       

    Amygdalin                almonds                     Prunus amygdalus.

    Dhurrin                  sorghum                     Sorghum album,
                                                         Sorghum bicolor.

    Linamarin                cassava                     Manihot esculenta,
                                                         M. carthaginensis
                             lima beans                  Phaseolus lunatus.

    Lotaustralin             cassava                     Manihot carthaginensis
                             lima beans                  Phaseolus lunatus.

    Prunasin                 stone fruits                Prunus species e.g.,
                                                         P. avium, P. padus,
                                                         P. persica, P. macrophylla.

    Taxiphyllin              bamboo shoots               Bambusa vulgaris.
                                                                                       
    
         The third aspect is that the cyanogenic glycosides taken up
    intact with the food are (partly) hydrolyzed by the ▀-glucosidase
    activity of the bacteria of the gut flora of animals or humans
    (Conn, 1979a,b; Oke, 1979, 1980; Nartey, 1980; Rosling, 1987;
    Gonzales & Sabatini, 1989).

         Cyanide, released from a cyanogenic glycoside in food by
    ▀-glucosidase either of plant or from gut microflora origin and
    taken up, follows the known cyanide metabolic pathway and
    toxicokinetics both for animals and man. Cyanide is detoxified by
    the enzyme rhodanese, forming thiocyanate, which is excreted by
    urine (Conn, 1979a,b; Oke, 1979, 1980). Due to several factors
    influencing hydrolysis of cyanogenic glycosides and the confounding
    influence of nutritional status (such as riboflavin, vit. B12,
    sodium, methionine intake) human case studies and epidemiological
    studies of the chronic toxicological effects have shown very
    variable results and were not conclusive. In addition, the data in
    these studies are rarely of a quantitative character (Conn, 1979a,b;
    Oke, 1979, 1980; Nartey, 1980; Rosling, 1987). In several studies
    both in animals and man the toxicity of cyanogenic glycosides is
    often expressed as mg releasable cyanide.

    
    Table 2: Concentration of cyanide in some tropical foodstuffs
             (Summarized in Nartey, 1980).
                                                                        

    Plant/tissue                                mg HCN/kg
                                                                        

    Cassava(bitter)/dried root cortex             2450
    Cassava(bitter)/leaves                         310
    Cassava(bitter)/whole tubers                   395
    Cassava(sweet)/leaves                          468
    Cassava(sweet)/whole tubers                    462
    Sorghum/whole immature plant                  2500
    Bamboo/immature shoot tip                     8000
    Lima beans from Java (coloured)               3120
    Lima beans from Puerta Rico (black)           3000
    Lima beans from Burma (white)                 2100
                                                                        
    
         Because this monograph first discusses the toxicity of cyanide
    as a basis for understanding that of cyanogenic glycosides, a
    modified form of the general monograph format has been used,
    presenting first biological data for cyanide, then that for
    cyanogenic glycosides.

    CYANIDE

    2.  BIOLOGICAL DATA

    2.1  Biochemical aspects

    2.1.1.1   Absorption, distribution, and excretion

         Hydrogen cyanide after oral administration is readily absorbed
    (it is also readily absorbed after inhalation exposure and through
    skin and eyes). After absorption, cyanide is rapidly distributed in
    the body through the blood.

         The concentration of cyanide is higher in erythrocytes than in
    plasma. It is known to combine with iron in both methaemoglobin and
    haemoglobin present in erythrocytes. The cyanide level in different
    human tissues in a fatal case of HCN poisoning has been reported:
    gastric content; 0.03, blood; 0.50, liver; 0.03, kidney; 0.11,
    brain; 0.07, and urine; 0.20 (mg/100 g) (EPA, 1990).

         The pharmacokinetics of 14CN- and S14CN- in rats
    exposed to these agents in diet for 3 weeks was investigated. All
    tissues contained radioactivity 9 h after intraperitoneal injection
    of 14CN-; highest radioactivity was found in the stomach (18%).
    Eighty per cent of this activity was in the form of thiocyanate. At
    this point 25% of the dose had already been eliminated in the urine
    and 4% in the expired air. When S14CN- was given  per os to
    rats with elevated plasma thiocyanate levels due to chronic oral
    exposure to cyanide, most of the activity was eliminated in the
    urine and only small amounts were found in the faeces. This
    indicated the existence of a gastrointestinal circulation of
    thiocyanate (Okoh & Pitt, 1981).

         The excretion of an acute oral dose of 14C-labelled cyanide
    in urine, faeces and expired air was studied in rats (12
    animals/group) pretreated orally for 6 weeks with unlabelled KCN or
    a control diet. Urinary excretion was the main route of elimination
    of 14C-labelled cyanide in these rats, accounting for 83% of the
    total excreted radioactivity at 12 h and 89% of the total excreted
    radioactivity at 24 h. The major metabolite of cyanide excreted in
    urine was thiocyanate, and this metabolite accounted for 71% and 79%
    of the total urinary activity at 12 h and 24 h, respectively. Only
    4% of the mean total activity excreted was found in expired air
    after 12 h, and this value did not change after 24 h. Of the total
    activity in expired air in 24 h, 90% was present as carbon dioxide
    and 9% as cyanide. When these results were compared with those
    observed for control rats, it was clear that the mode of elimination
    of cyanide carbon was altered in neither urine nor breath by the
    chronic intake of cyanide (Okoh, 1983).

         Golden hamsters exposed to cyanide by subcutaneous infusion
    appeared to excrete only a relatively low percentage (10-15%) of the
    dose as thiocyanate in the urine (Doherty  et al., 1982).

         The major defence of the body to counter the toxic effects of
    cyanide is its conversion to thiocyanate mediated by the enzyme
    rhodanese. The conversion of cyanide to the less toxic thiocyanate
    by rhodanese was discovered by Lang (1933).

         Thiosulfate and 3-mercapto-pyruvate can act as sulfur donors,
    but neither free cystine nor cysteine can. The enzyme contains an
    active disulfide group which reacts with the thiosulfate and
    cyanide. The trivial name rhodanese is more widely used than that
    assigned by the Enzyme Commission [thiosulfate-cyanide sulfur
    transferase, EC. 1.8.1.1]; it has been inappropriately called
    rhodanase in several reports. 

         The rhodanese-catalyzed irreversible conversion of cyanide to
    thiocyanate, in the presence of thiosulfate, provides a means for
    the treatment of cyanide poisoning. Since the enzyme, which is
    usually localized in the mitochondria in different tissues, is
    relatively abundant, but in sites which are not readily accessible
    to thiosulfate, the limiting factor for the conversion of cyanide is
    thus thiosulfate (EPA, 1990).

         The overall rate of  in vivo detoxification of cyanide may be
    influenced by several minor reactions. Cystine may directly react
    with cyanide to form 2-imino-thiazolidine-4-carboxylic acid which is
    excreted in saliva and urine. Traces of hydrogen cyanide may be
    found in expired air, saliva, sweat and urine. A minor amount may be
    converted into formic acid which may be excreted in urine or
    participate in the metabolism of one carbon compound. One minor
    detoxification route is the combination of cyanide with
    hydroxycobalamine (vitamin B12) to form cyanocobalamine which is
    excreted in urine and bile. It may be reabsorbed by the intrinsic
    factor mechanism at the level of the ileum allowing effective
    recirculation of vitamin B12. Methaemoglobin effectively competes
    with cytochrome oxidase for cyanide and its formation from
    haemoglobin, effected by sodium nitrite or amylnitrite, is exploited
    in the treatment of cyanide (EPA, 1990). It has been reported that
    other species have lower rhodanese activity than the rat and hence
    the rat may be able to convert cyanide to thiocyanate more easily
    than other species (Himwich & Saunders, 1948).

    2.1.2  Biotransformation

         No information available.

    2.1.3  Effects on enzymes and other biochemical parameters

         Cyanide causes a decrease in the utilization of oxygen in the
    tissues, producing a state of histotoxic anoxia. This occurs through
    inactivation of tissue cytochrome oxidase by cyanide, which combines
    with Fe3+/Fe2+ contained in the enzyme. The enzyme-cyanide
    complex dissociation constant has been found to be 1 * 10-6 and 1
    * 10-4 (moles/l) for the oxidized and reduced form of the enzyme,
    respectively. Thus, the affinity of cyanide for the oxidized form of
    the enzyme is two orders of magnitude higher than for the reduced
    form. However, the rate of reaction of cyanide with the reduced
    enzyme is twice the rate of reaction with the oxidized form.

         Cyanide can inhibit several other metalloenzymes most of which
    contain iron, copper or molybdenum (e.g., alkaline phosphatase,
    carbonic anhydrase), as well as enzymes containing Schiff base
    inter-mediates (e.g., 2-keto-4-hydroxyglutarate aldolase). The
    effect of sublethal doses of cyanide on the metabolism of glucose in
    mice has been studied using radiorespirometric techniques
    (Solomonson, 1981).

         Cyanide causes an increase in blood glucose and lactic acid
    levels and a decrease in the ATP/ADP ratio indicating a shift from
    aerobic to anaerobic metabolism. Cyanide apparently activates
    glycogenolysis and shunts glucose to the pentose phosphate pathway
    decreasing the rate of glycolysis and inhibiting the tricarboxylic
    acid cycle (EPA, 1990).

    2.2  Toxicological studies

    2.2.1  Acute toxicity studies

         Lethal doses of HCN in mg/kg bw were reported for mouse, 3.7;
    dog, 4.0; cat, 2.0 and for cattle and sheep 2.0 (Summarized by Conn,
    1979a).

    
    Table 3. Acute toxicity of cyanide
                                                                                  

                                  Acute toxicity

    Species             Route             LD50             References
                                       (mg/kg bw)
                                                                                  

    Mouse               i.v.           0.99 (HCN)          EPA 1990*
    Rat                 i.v.           0.81 (HCN)          EPA 1990*
    Guinea-pig          i.v.           1.43 (HCN)          EPA 1990*
    Rabbit              i.v.           0.66 (HCN)          EPA 1990*
    Cat                 i.v.           0.81 (HCN)          EPA 1990*
    Dog                 i.v.           1.34 (HCN)          EPA 1990*
    Monkey              i.v.           1.30 (HCN)          EPA 1990*
                                                                                  

    Mouse               s.c.            6.0 (KCN)          WHO, 1965*
                        i.v.            2.5 (KCN)          WHO, 1965*
    Rat                 oral           10-15 (KCN)         WHO, 1965*
                        i.v.            2.5 (KCN)          WHO, 1965*
    Dog                 oral            5.3 (KCN)          WHO, 1965*
                                                                                  

    Rabbit              s.c.           2.2 (NaCN)          WHO, 1965*
    Guinea-pig          s.c.           5.8 (NaCN)          WHO, 1965*
    Dog                 i.v.           2.8 (NaCN)          WHO, 1965*
                                                                                  

    Mouse               oral           598 (NaSCN)         WHO, 1965*
                        i.v.           484 (NaSCN)         WHO, 1965*
    Rat                 oral           765 (NaSCN)         WHO, 1965*
                        i.p.           540 (NaSCN)         WHO, 1965*
                                                                                  

         * as summarized in
    
    2.2.2  Short-term toxicity studies

    2.2.2.1  Rats

         In a 13-week toxicity study male Sprague-Dawley rats
    (approximately 30 rats/group) were administered KCN in the drinking
    water. The dose levels were 40, 80 and 160/140 mg KCN/kg bw/24 h.
    Three control groups were used, respectively a normal drinking water
     ad libitum, a "paired drinking" group (parallel to the high dose

    level KCN) and a group receiving drinking water with 10% ethyl
    alcohol. In addition, one group received drinking water with KCN
    (80 mg/kg bw) and 10% alcohol.

         Behaviour, external appearance, body weight, food consumption
    (daily) and drinking water consumption (twice weekly) were recorded
    frequently. Extensive haematological, clinical chemical (in serum)
    and urine analyses were carried out in 5 animals per group in week 6
    and week 13. Autopsy and macroscopy were performed after 13 weeks
    (approximately 20 animals/group) and 11 organs were weighed.
    Histopathological examination was performed in brain, kidneys,
    heart, liver and testes of these animals. In addition thyroids of
    the control, the "pair drinking" control and the high-dose group
    (160/140 mg/kg bw/day) were examined. There was a clear indication
    that reduced food consumption and body weight in the KCN groups were
    caused by a decrease in water consumption due to decreased
    palatibility. Urinalyses revealed a higher level of protein in the
    animals receiving KCN. The amounts of protein determined showed a
    clear correlation to the increasing doses of KCN, as did the
    drinking water. Several changes in absolute organ weights were seen
    in the 160/140 mg KCN/kg bw/day group. Relative weights of organs
    were very slightly increased in the 40, slightly increased in the
    80 mg and clearly increased in the 160/140 mg KCN/kg bw groups. The
    thymus weight was, however, reduced in the high-dose group.
    Histopathological examination revealed no indication of damage to
    the brain, heart, liver, testes, thyroids nor kidneys due to
    treatment with KCN (Leuschner  et al., 1989b).

    2.2.3  Long-term carcinogenicity studies

         No data on the carcinogenicity of hydrogen cyanide have been
    published. However, anticarcinogenic effects of cyanide have been
    reported. Longevity of mice with transplanted Ehrlich ascites
    tumours and Sarcoma 180 was increased 20 to 70% on i.p. injection of
    sodium cyanide in the dose range 0.75 to 2.0 mg/kg bw (EPA, 1990).

    2.2.4  Reproduction studies

    2.2.4.1  Rats

         A short-term reproductive study (49 day study in adults and 28
    day study in pups) was performed to evaluate the cumulative effects
    of adding 500 mg KCN/kg to cassava root flour-based diet in pregnant
    rats. This meal was prepared from a low-HCN cassava variety (21 mg
    HCN/kg feed).

         High dietary level of KCN did not have any marked effect in
    gestation and lactation performance of female rats. No carry-over
    effect of high cyanide-containing diet fed during gestation was
    observed on lactation performance. The high cyanide-containing diet,

    however, significantly reduced feed consumption and daily growth
    rate of the offspring when fed during post-weaning period. Protein
    efficiency ratio was not only reduced by the cyanide diet during
    post-weaning growth phase but there was an additional carry-over
    effect from gestation. Serum thiocyanate was significantly increased
    in lactating rats and their offspring during lactation and in the
    postweaning growth phase of the pups. No apparent carry-over effect
    was noticed on this parameter. Rhodanese activity in liver and
    kidneys was unaffected by feeding the high cyanide diet during
    gestation, lactation, nor during postweaning growth (Tewe & Maner,
    1981b).

    2.2.5  Special studies on embryotoxicity and teratogenicity

    2.2.5.1  Hamsters

         Pregnant golden hamsters were exposed to sodium cyanide on days
    6-9 of gestation by infusion via subcutaneously implanted osmotic
    minipumps. Cyanide (0.126-0.1295 mmol/kg/h) induced high incidences
    of resorptions and malformations in the offspring. The most common
    abnormalities observed were neural tube defects (Doherty  et al.,
    1982).

    2.2.6  Special studies on the thyroid gland

    2.2.6.1  Rats

         A group of 10 male rats was fed a 10% casein diet containing
    added methionine, vitamin B12, iodine and potassium cyanide
    (1500 mg/kg feed) for nearly one year. Compared to a control group
    not receiving cyanide, depression of body-weight was observed
    throughout the study period, but there were no deaths nor clinical
    signs of toxicity. Depression of both plasma thyroxine and thyroxine
    secretion rate suggestive of depressed thyroid function were evident
    at 4 months but less so after 1 year. At autopsy the animals were
    found to have enlarged thyroids and this may have been the mechanism
    of adaptation. Some differences in the histopathology of the spinal
    cord, notably the white matter, were also found between controls and
    cyanide-treated animals (Philbrick  et al., 1979).

    2.2.6.2  Pigs

         Performance and metabolic and pathological changes were
    evaluated in 48 growing pigs fed different levels of dietary protein
    (9 and 16%), cyanide, and iodine (0 and 0.36 mg iodine/kg feed)
    during 56 days. Protein deficiency reduced urinary iodine excretion
    and the concentrations of protein, protein-bound iodine (PBI) and
    thiocyanate in serum. It also reduced liver rhodanese activity and
    caused a decrease in urinary thiocyanate excretion which was not
    significant. Dietary cyanide increased urinary thiocyanate and

    iodine excretion and serum PBI. Pathological studies showed that
    cyanide treatment had no marked effect on the microanatomy of the
    tissues examined. Dietary iodine deficiency caused histological
    changes in the thyroid gland and bone which suggested a decline in
    metabolic activity. Iodine deficiency caused hyperplastic goitre in
    the experimental animals (Tewe & Maner, 1980).

    2.2.7  Genotoxicity

         Two negative and one marginally positive genotoxicity studies
    for cyanide have been reported. Potassium cyanide was not mutagenic
    in  Salmonella typhimurium strains TA1535, TA1537, TA1538, TA98 nor
    TA100 with or without S-9 liver microsomes. Cyanide was also
    negative in recombinant-assay in  Bacillus subtilis. One study
    reported marginally mutagenic activity of HCN to  Salmonella
     typhimurium strain TA100 in the absence of S-9 mix. In the same
    study no mutagenic activity in strain TA98 with or without S-9 mix
    was observed (EPA, 1990).

         An  in vitro Ames test with HCN in  Salmonella strains
    TA1537, TA1538 and TA98 for detection of frame shift mutation and
    TA1535 and TA100 for base-pair substitutions was performed with and
    without metabolic activation of a S-9 microsome mixture. There was
    no indication of mutagenic properties under these conditions
    (Leuschner  et al., 1983a).

         An  in vivo mutagenicity study in Chinese hamsters detecting
    chromosomal aberrations with HCN orally administered to Chinese
    hamsters was carried out. Preparations of metaphase cells were
    studied for structural chromosome aberrations after 6, 24 and 48 h
    after oral administration of 0.4 mg HCN/kg bw. The incidence of
    aberrations or gaps was within the spontaneous range. Neither
    multiple aberrations nor pulverised metaphases were found. There was
    no indication of mutagenic properties relative to structural
    chromatid or chromosome damage (Leuschner  et al., 1983b).

         A gene mutation assay in cultured Chinese hamster cells, V79
    (genetic marker HGPRT) both in the presence and absence of metabolic
    activation system was carried out with KCN. The duration of the
    exposure with the test substance was 24 h in the experiments without
    S-9 mix and 2 h in the experiments with S-9 mix. The test compound
    dose levels employed were chosen following a preliminary toxicity
    experiment. The dose-levels for the main study were 400, 800, 1000,
    2000 and 3000 Ág/ml without S9 mix and 1000, 2000, 3000, 4000, 6000,
    8000 and 10 000 Ág/ml with S9 mix. KCN was tested up to a high
    cytotoxicity in the absence and presence of metabolic activation.
    Under the present test conditions KCN was negative in the V79
    mammalian cell mutagenicity test (Leuschner  et al., 1989a).

    2.2.8  Special studies on nervous system

         A special study on the behavioural effects of chronic sublethal
    dietary cyanide (KCN; 0.4, 0.7 and 1.2 CN-/kg bw) in juvenile
    swine, mimicked the situation of free CN- intake in Liberia due to
    eating cassava-based foods. There were two clear behavioural trends:
    1) increasing ambivalence and slower response time in reacting to
    various stimuli and 2) an energy conservation gradient influencing
    which specific behaviours would be modified in treated animals.
    Serum SCN- was positively correlated with daily CN- intakes.
    CN- treatment diminished T3 and T4 levels but elevated fasting
    blood glucose values (Collier-Jackson, 1988).

         Neuronal lesions in several animal species have been produced
    by chronic cyanide intoxication either by injection of unbuffered
    alkaline cyanide salts or by inhalation of hydrogen cyanide. The
    neuropathological changes include areas of focal necrosis especially
    around the centrum ovale, corpus striatum, corpus callosum,
    substantia nigra, anterior horn cells, and patchy demyelination in
    the periven-tricular region. In some species, the earliest effects
    may be on the oligodendroglia and hence myelin lesions may precede
    neuronal damage. Bass (1968) showed that in rats chronic cyanide
    intoxication produces myelin loss by its primary effect on glial
    cells followed by breakdown of myelin. Brierly  et al. (1977)
    reported myelin damage and changes in the oligodendroglia in cyanide
    poisoning in rats. Clark (1936) described fatty degeneration of the
    liver and secretory tubules of the kidney in rats subjected to
    chronic cyanide intoxication. In these animal experiments relatively
    large doses of cyanide were given, often sufficient to cause partial
    asphyxia. Therefore it was doubtful whether the neuropathological
    effects demonstrated were due to asphyxia or chronic cyanide
    intoxication so they did not appear to have an obvious parallel to
    human exposure at lower levels. It is noteworthy, however, that
    changes in optic nerves and tracts occur consistently only in
    primates (Ferraro, 1933; Hurst, 1940; Lessell, 1971). In a carefully
    controlled experiment in which small weekly doses of cyanide were
    administered over several months to rats, neuronal degeneration and
    demyelination were reported (Smith  et al., 1963). Williams and
    Osuntokun (1969) found that the demyelination of peripheral nerves
    induced in rodents by cyanide injection bore a striking resemblance
    to the lesions found in biopsy specimen of peripheral nerves of
    Nigerian patients who suffer from tropical neuropathy (reviewed by
    Osuntokun, 1981).

    2.3  Observations in humans

    2.3.1  Acute toxicity studies in humans

         The acute oral lethal dose of HCN for human beings is reported
    to be 0.5-3.5 mg/kg bw corresponding to 1.0-7.0 mg/kg bw of KCN. The
    clinical signs are well described (Montgomery, 1969; Gosselin

     et al., 1976) and include headache, dizziness, mental confusion,
    stupor, cyanosis with twitching and convulsions, followed by
    terminal coma (Conn, 1979a).

         The acute oral lethal dose of HCN for man was reported to be
    60 mg (Sinclair & Jeliffe, 1961). For man the acute oral dose of HCN
    is usually given as 50-90 mg and for potassium cyanide as 200 mg,
    corresponding to 81 and 110 mg HCN respectively (Lehman, 1959). Data
    on the oral lethal dose of cyanide for man in four cases of suicide,
    calculated from the amount of HCN absorbed in the body at the time
    of death, and from the amount of HCN found in the digestive tract,
    differed considerably ((calculated as mg HCN: 1450 (62.5 kg bw),
    556.5 (74.5 kg), 296.7 (50.7 kg) and 29.8 (51 kg)) (Geitler & Baine,
    1938). This corresponds to doses varying from 0.58-22 mg/kg bw (in
    WHO, 1965).

    CYANOGENIC GLYCOSIDES

    2.  Biological data

    2.1  Biochemical aspects

    2.1.1  Absorption, distribution, and excretion

         Wistar rats (4 animals/sex/group) were given 50 mg
    amygdalin/rat respectively, intravenously and orally after an
    overnight fast. During the experiment and the night before they were
    kept in metabolic cages which allowed collection of urine without
    faeces and minimized coprophagy.

         After intravenous and oral administration fractions of the dose
    excreted by the rat as unchanged amygdalin were 70% and 0.8%,
    respectively. The fraction excreted as prunasin after intravenous
    administration was 6.6%, whereas it was 39% of the total dose after
    oral administration (Rauws  et al., 1982).

         In a toxicokinetic study 10 male rats (100-120 g) were given
    50 mg pure linamarin (dissolved in water, volume 0.5 ml) and 6 male
    rats were given water alone by stomach tube. Seven rats dosed with
    50 mg linamarin died within 4 h. In a second trial 6 male rats were
    given 30 mg linamarin. Following dosing, urine and faeces were
    collected after 24, 48 and 72 h, and heparinized blood samples were
    taken from the optic vein or lateral tail vein. Blood was taken
    after 30, 40, 60, 80 and 100 min and at 2, 4, 8, 24 and 48 h.

         No intact linamarin was detected in faeces nor blood of the
    rats dosed with 30 mg (300 mg/kg bw). No linamarin in the faeces of
    the three surviving rats dosed with 50 mg (500 mg/kg bw) was
    detected, however blood and urine were not examined in these
    animals. Linamarin was excreted in the urine at a level of 5.65 mg
    (cumulated after 72 h) along with 0.823 mg of thiocyanate. These
    findings indicate that linamarin was absorbed intact in considerable
    proportion and was partially hydrolyzed (Barrett  et al., 1977).

         In a toxicokinetic study beagle dogs (4 animals/sex/group) were
    administered 500 mg amygdalin in 10 ml solution respectively,
    intravenously and orally after overnight fast. Blood was sampled
    from jugular vein and urine was collected by a funnel. Faeces were
    removed from the funnel.

         The major part of the dose (71%) was recovered in the urines
    collected during 6 h following intravenous amygdalin administration.
    The fraction of the dose excreted by glomerular filtration was
    calculated using the ratio of diatrizoate (which was administered
    simultaneously) clearance to amygdalin clearance, showing that 97%

    of the amount of amygdalin to be expected was recovered from the
    urine. The result of the experiments after intravenous
    administration were analyzed assuming a two-compartment model. The
    distribution T alpha was 0.10 and the elimination T ▀ 0.57. No
    prunasin was detected in urine (detection limit = 0.2% of dose).

         After oral administration of amygdalin a very low maximal
    plasma level is found after approximately 0.75 h. Only 2.3% of the
    amygdalin was systematically available (absolute bioavailablity).
    Prunasin was found in plasma and urine of dogs. In the urine
    collected during 6 h following amygdalin administration only about
    1% of the dose was recovered unchanged and 21% of the dose was
    identified as prunasin (Rauws  et al., 1982).

         In a toxicokinetic study prunasin was administered
    intravenously and orally as 100 mg doses to female dogs (2/group;
    10 kg) after an overnight fast. Blood was sampled from jugular vein
    and urine was collected by a funnel.

         The results of the experiment after intravenous administration
    were analyzed assuming a two-compartment model. The prunasin results
    were compared to those obtained with amygdalin in a earlier
    experiment. The distribution T alpha was 0.08 h and elimination
    T ▀ was approximately 0.64 h.

         Prunasin was absorbed to a large extent after oral
    administration. The absolute bioavailability after oral
    administration was 50% of the administered dose. The volume of
    distribution (0.34 L/kg) and the clearance (0.55 L/kg h) are larger
    than those of amygdalin (respectively 0.19 L/kg and 0.39 L/kg h).
    The oral bioavailability of prunasin is considerably greater,
    whereas amygdalin is only slightly (2.3%) absorbed (Rauws  et al.,
    1983).

         In a toxicokinetic study pure linamarin, at a dose level of
    300 mg/kg bw, was administered in food to a group of Wistar rats
    maintained on vitamin B2-deficient, -sufficient, and -excess diets
    for 5 weeks and to another group of kwashiorkor rats. Free and total
    cyanide, intact linamarin and thiocyanate levels were estimated in
    urine and faeces obtained at 0, 24, 48 and 72 h periods and in blood
    samples obtained 72 h after the compound had been administered.
    There was no detectable cyanide nor intact linamarin in the faecal
    samples. Rats on vitamin B2-sufficient and B2-excess diets excreted
    higher total and free cyanide in urine than the respective vitamin
    B2-deficient groups. Most of the linamarin was degraded after 24 h.
    The rate of breakdown of the glycoside within the first 24 h was
    slowest for zero and half normal vitamin B2 status rats as evidenced
    by appearance of the glycoside in large quantities in the urine. The
    kwashiorkor rats, on the other hand, excreted less thiocyanate than

    the controls. In addition, their control group excreted most of the
    thiocyanate in the first 24 h, whilst the kwashiorkor rats excreted
    most of the thiocyanate in the first 48 h. Dietary protein
    deficiency prolongs the time of metabolism and hence increases the
    toxicity of cyanogenic glycosides in the body (Umoh  et al., 1986).

    2.1.2  Biotransformation

         Strained ruminal fluid was collected from cattle fed five diets
    (concentrate diet, freshly harvested alfalfa, cubed alfalfa, alfalfa
    hay and orchard grass) to determine  in vitro rates of cyanogenesis
    from the glycosides amygdalin, prunasin and linamarin. Rates of
    dissociation for the corresponding aglycones, benzaldehyde
    cyanohydrin and acetone cyanohydrin were also determined. Hydrogen
    cyanide (HCN) in ruminal fluid was determined with a modified method
    of HCN analysis that independently measured the overall rate of
    cyanogenesis and the non-enzymatic dissociation of cyanohydrins, the
    intermediate products in the degradation of cyanogenic glycosides to
    HCN. Rate of dissociation of cyanohydrins in ruminal fluid was
    pH-dependent, with high rates of dissociation (as expressed by the
    rate constant or half-life of reaction) occurring at pH >6 and
    slower rates at pH 5 to 6. Cyanohydrin dissociation was most rapid
    when cattle were fasted for 24 to 48 h and ruminal pH was high; rate
    of dissociation was much slower during feeding and digestion. When
    the glycosides were examined, highest rates of cyanogenesis
    (mg HCN/L/s) were observed after a 24 h postprandial period. The
    rates were highest after feeding hay: 0.019 for amygdalin, 0.033 for
    linamarin and 0.048 for prunasin. Hence cattle are most susceptible
    to poisoning by cyanogenic plants when the pH of ruminal fluid is
    elevated, leading to rapid dissociation, and also when the activity
    of ▀-glucosidase is adequate for rapid hydrolysis of glycosidic
    bonds. Rates of cyanogenesis were higher when ruminal inocula were
    from cattle fed fresh alfalfa or cubed alfalfa hay rather than from
    those fed grain or long hay. Rates of HCN production were lowest
    using inocula from cattle fed grain; rates for the three glycosides
    were negligible at the 3 and 6 h postprandial sampling times. In
    agreement with previous studies, prunasin was degraded in ruminal
    fluid much more rapidly than linamarin or amygdalin (Majak  et al.,
    1989).

         In a comparative metabolism study rates of cyanide liberation
    resulting from hydrolysis of the cyanogenic glycosides linamarin,
    amygdalin and prunasin by a crude ▀-glucosidase prepared from
    hamster caecum were studied  in vitro. In addition, hamster blood
    cyanide and thiocyanate concentrations were determined at 0.5, 1, 2,
    3, and 4 h after oral dose of 0.44 mmol linamarin or amydalin/kg bw.

         Plots of cyanide liberated versus time for linamarin and
    prunasin yielded straight lines, whereas for amygdalin the plot was
    curvilinear; the rate of cyanide release increased with time. At
    10-3 M substrate concentrations, the averaged rates of hydrolysis

    of prunasin, amygdalin and linamarin were 1.39, 0.57 and
    0.13 nmol/min/mg protein, respectively. Lineweaver-Burk plots
    yielded apparent Km and Vmax values of 3.63 * 10-5 M and
    0.13 nmol/min/mg protein, respectively for amygdalin, and 7.33 *
    10-3 M and 1.04 nmol/min/mg protein, respectively for linamarin.

         Blood cyanide concentrations following amygdalin treatment of
    the hamster reached their highest level (130 nmol/ml) 1 h after
    dosing and remained elevated until 3 h after treatment. Blood
    cyanide concentrations following linamarin treatment reached their
    highest level (116 nmol/ml) after 3 h and then declined immediately.
    Area under the blood cyanide concentration-time curve was
    395 nmol h/ml for amygdalin and 318 nmol h/ml for linamarin. The
    results suggest a faster rate of enzymatic hydrolysis and cyanide
    absorption for amygdalin than for linamarin (Frakes & Sharma, 1986).

         In humans, pharmacological studies have shown that amygdalin is
    broken down to HCN, benzaldehyde and glucose by enzymes found in gut
    bacteria, but not intracellularly in humans. Animal and human
    tissues contain no significant concentrations of ▀-glucosidase, the
    only known activating enzyme of hydrolysis of cyanogenic glycosides
     in vivo (Dorr & Paxinas, 1978).

         Cyanogenic glycosides are hydrolyzed by ▀-glucosidase produced
    by intestinal bacteria to glucose, HCN and benzaldehyde or acetone.
    Benzaldehyde is oxidized to benzoic acid (and subsequently to
    hippuric acid) or salicylic acid isomers. Thiocyanate is present in
    body fluids; blood, urine, saliva, sweat and tears (Oke, 1979).

         The cyanide-yielding capacity of insufficiently processed
    cassava probably occurs as linamarin, or an intermediate break-down
    product, from which cyanide may be yielded in the gut by action of
    microbial enzymes. Significant amounts of linamarin are observed in
    the urine after consumption of insufficiently processed cassava as
    well as after consumption of other plants containing linamarin.
    These results indicate that linamarin, if not metabolized in the
    gut, will be absorbed and excreted in the urine without causing
    exposure to HCN. About 80% of ingested cyanide will be turned into
    thiocyanate and is excreted in the urine after a short period
    (Rosling, 1987).

    2.1.3  Effects on enzymes and other biochemical parameters

         No information available.

    2.2  Toxicological studies

    2.2.1  Acute toxicity studies

        Table 4. Acute toxicity studies of cyanogenic glycosides or cyanogenic plant
             tissues
                                                                                       
                                     Acute toxicity

    Species          Sex           Route          LD50            References
                                               (mg/kg bw)
                                                                                       
    Mouse             ?            i.p.        0.1 mmole          Solomonson, 1981
                                               amygdalin/kg

    Rat               ?            i.v.        20 000             Oke, 1979
                                               linamarin

    Rat               ?            oral        450 linamarin      Oke, 1979

                                                                                       
    
         A dose of 25 mg linamarin (250 mg/kg bw) fed to rats
    (100-120 g bw) caused clinical signs of toxicity, including apnoea,
    ataxia and paresis. These symptoms were very marked in the absence
    of methionine supplementation, 50% of these rats died within 4 h. In
    the presence of adequate methionine supplementation, 10% of rats
    died and about 40% showed no signs of toxicity. The activity of
    Na+K+-dependent ATPase was reduced in much the same way as it
    was by the glycoside, digitalis (reviewed by Oke, 1980).

         In a toxicokinetic study 7 out 10 rats (100 g bw) died after
    administration of 50 mg linamarin by stomach tube (Barrett  et al.,
    1977).

         Oral doses of 100, 120 and 140 mg linamarin/kg bw given by
    stomach tube to hamsters (90 g bw) produced signs of cyanide
    intoxication in a large percentage. The signs appeared within 1 h
    after dosing included dyspnoea, hyperpnoea, ataxia, tremors and
    hypothermia. Two animals dosed with 140 mg/kg bw and one animal
    dosed with 120 mg/kg bw died within 2 h of dosing. The signs of
    poisoning were greatly reduced or gone within 3 h after treatment in
    the surviving animals. No relationship between length of
    intoxication and dose was observed (Frakes  et al., 1985).

         Hamsters have been reported to be more susceptible than rats to
    the acute toxic effects of orally administered amygdalin and
    prunasin (Willhite, 1982).

         A species difference in reaction to cyanogenic glycosides is
    observed due to a difference of detoxifying ability due to
    anatomical structure. Ruminants, e.g., cattle and sheep are supposed
    to be more susceptible to the acute toxic effects because of their
    larger flora of microorganisms and considerable quantities of the
    enzyme emulsine which hydrolyzes the glycoside (Oke, 1979).

    2.2.2  Short-term toxicity studies

    2.2.2.1  Rats

         Albino female rats (10 animals/group) were fed  ad libitum one
    of the following diets, A; a normal laboratory diet (control), B; a
    50% gari diet (Nigerian preparation of cassava), C; a raw cassava
    diet, D; a diet containing 5 g KCN/100 g and E; a diet containing
    10 g KCN/100 g during a 14-day period. The 50% gari diet caused no
    significant biochemical nor haematological changes in the female
    rats, whereas for both the raw cassava diet and the KCN diets a
    decrease of Hb, PCV, total serum protein concentration and T4
    concentration was observed. In the 50% gari diet group, and to a
    greater extent, in the other treatment groups, the serum thiocyanate
    levels were increased. The body weight gain was not significantly
    decreased in the 50% gari group, whereas the other treatment groups
    showed instead of gain a loss of body weight (Olusi  et al., 1979).

    2.2.2.2  Guinea-pigs

         In a 24-day toxicity experiment guinea-pigs (8 animals/group)
    were dosed daily with, respectively, laetrile (10 mg amygdalin) and
    8 mg KCN/kg bw with and without ascorbic acid (100 mg). No
    significant effect on the body weight nor liver weight was observed.
    However, treatment with laetrile alone for 4, 16 or 24 days,
    respectively, resulted in a significant increase in urinary levels
    of thiocyanate. The increase was less in animals treated with
    vitamin C. In guinea-pigs treated with 8 mg KCN/kg bw, toxic effects
    were seen as evidenced in slight tremors in 3 of the 8 animals,
    which recovered within 5 min. All animals in the KCN group which
    were supplemented with ascorbate showed severe tremors, motor
    ataxia, bizarre neuromuscular manifestations and rhythmic head
    movements. The toxicity of KCN increased with elevation of vitamin
    C, whereas urinary excretion of thiocyanate decreased (Basu, 1983).

    2.2.2.3  Chickens

         In two feeding experiments (respectively 63 and 56 days) one
    day old broiler chickens (male and female) were fed a diet
    containing 0, 10, 20 or 30% cassava, respectively. The animals were
    studied for haematological and histopathological effects. The
    cassava diet studied in the 1st experiment consisted of a
    high-cyanide-containing cassava root meal (CRM) supplying 300 mg of
    total cyanide/kg, most of it in the form of cyanogenic glycosides.

    The cassava diet in the 2nd experiment also contained cassava
    foliage meal (CFM) supplying 156 mg total cyanide/kg. In the 1st
    experiment 26 chickens per group were used and in the 2nd experiment
    160 chickens were used for the cassava groups and 80 for the control
    group.

         No changes in the haematological parameters due to cassava were
    seen. Addition of up to 30% CRM failed to adversely affect broiler
    survival, performance nor feed efficiency, but the inclusion of CFM
    in the experimental diets increased mortality, decreased weight gain
    and decreased feed efficiency. In both experiments, increased
    quantities of dietary cassava cyanate were associated with increased
    (P < 0.05) blood serum thiocyanate concentrations.
    Histopathological examination of thyroid, liver and kidney revealed
    no appreciable alterations due to the cassava feeding, however there
    was no conclusive evidence of cyanide or thiocyanate effects on
    thyroid activity. Aflatoxin contamination appeared to have
    contributed to the high mortality rate associated with CFM diets.
    The results showed that broilers were tolerant of relatively high
    levels of dietary cyanogenic glycosides (Gomez  et al., 1988).

    2.2.3  Long-term/carcinogenicity studies

         No information available.

    2.2.4  Reproduction studies

    2.2.4.1  Rats

         In a one-generation reproduction study albino female rats (10
    rats/group) were fed  ad libitum one of the following diets: A, a
    normal laboratory diet, B, a 50% gari diet (Nigerian preparation of
    cassava), C, a raw cassava diet, D, a diet containing 5 g KCN/100 g
    and E, a diet containing 10 g KCN/100 g. After 2 weeks rats in each
    group were mated with 5 adult males fed normal diet. Pregnant rats
    from each group were maintained on their respective diets. After
    littering, the newborn rats were studied for postnatal development.
    After 21 days F1 rats were put for another 4 weeks on their
    respective diets.

         The offspring of the rats fed the 50% gari diet had
    significantly lower birth weights and brain weights and never
    attained the same adult weights as those of the controls. The adult
    female rats fed a diet consisting entirely of raw cassava had
    significantly reduced haematological and biochemical parameters
    (Hb, PCV, serum protein and T4 concentration). This diet also
    caused an increased incidence of cannibalism and a significant
    reduction in the frequency of pregnancy, the average number of pups
    per litter and birth weights among these pups. In addition there was
    an increased incidence of neonatal deaths among the offspring which

    also had poor development, reduced brain weights and an increased
    tendency of aggression towards their litter mates. Adult female rats
    fed diets containing 5 and 10 g KCN/100 g laboratory diet survived
    for more than three months but never became pregnant. They developed
    enlarged thyroid glands and tumours of the large intestine. The
    usual content of cyanide in cassava varies from 70 to 500 mg/kg
    which is much less than the levels used in these experiments; thus
    the rats were able to cope with the 50% gari diet and detoxify the
    glycoside present (Olusi  et al., 1979).

    2.2.4.2  Pigs

         General toxicity and reproductive effects were studied for
    cassava in combination with added cyanide. In a 110-day feeding
    experiment 18 pregnant Yorkshire gilts were allocated to three equal
    groups and fed fresh cassava (containing 40.2 HCN/kg) supplemented
    with 0, 250, and 500 mg cyanide (KCN) per kg of fresh cassava
    offered. Serum thiocyanate concentration was slightly but not
    significantly increased in the 500 mg KCN/kg group and serum protein
    bound iodine decreased during gestation in all groups. Fetal serum
    thiocyanate concentration was significantly (p <0.05) higher in the
    group fed 500 mg KCN/kg. A small increase in maternal thyroid weight
    with increasing levels of cyanide was observed. Pathological studies
    showed proliferation of glomerular cells of the kidneys in gilts of
    all groups and reduced activity of the thyroid gland in gilts fed
    500 mg KCN/kg group. Cyanide fed during gestation did not affect
    performance during lactation. Milk thiocyanate and colostrum iodine
    concentrations were significantly higher in the group fed 500 mg
    KCN/kg feed. No effects of cyanide were reported on indices of
    reproduction performance (Tewe & Maner, 1981a).

    2.2.5  Special studies on embryotoxicity and/or teratogenicity

         In a teratogenicity study pregnant hamsters received oral doses
    of 70, 100, 120 or 140 mg linamarin/kg bw or an equivalent volume
    (0.5 ml/100 g) of isotonic saline during the early primitive streak
    stage of gestation (day 8 of gestation). The hamsters were killed on
    the morning of day 15 of pregnancy. Fetuses were removed by
    caesarian section and the numbers of resorption sites, dead fetuses,
    and living fetuses were recorded. Living fetuses were examined for
    gross external malformations and by means of histopathological
    methods for internal malformations.

         A dose of 120 or 140 mg linamarin/kg bw was associated with an
    increased incidence of vertebral and rib anomalies as well as the
    production of encephaloceles in the offspring. These larger doses of
    linamarin also resulted in obvious maternal toxicity (dyspnoea,
    hyperpnoea, ataxia, tremors and hypothermia). Two animals dosed with
    140 mg and one animal dosed 120 mg/kg bw died. In surviving animals
    the signs of poisoning were greatly reduced or gone within 3 h after
    treatment.

         Linamarin treatment had no effect on fetal body weight,
    ossification of skeletons, embryonic mortality, nor litter size.
    Although ingestion of the cyanogenic glycoside was associated with a
    significant teratogenic response, the effects occurred only at doses
    that elicited signs of maternal intoxication (Frakes  et al.,
    1985).

         In a teratogenic study groups of pregnant hamsters
    (8 dams/group) were fed diets consisting of cassava meal:laboratory
    chow (80:20) during days 3-14 of gestation. One low cyanide (sweet)
    cassava meal and one high cyanide (bitter) cassava meal were
    studied. An additional group was fed a diet which resembled cassava
    in nutrtional value, but which lacked cyanogenic glycosides.
    Thiocyanate concentrations in the urine and blood of dams fed
    cassava diets increased significantly. Increased tissue thiocyanate
    concentrations were observed in fetuses recovered from cassava-fed
    dams. Cassava-fed dams gained significantly less weight than did
    control animals and their offspring showed evidence of fetotoxicity.
    Reduced fetal body weight and reduced ossification of sacrocaudal
    vertebrae, metatarsals and sternebrae were associated with cassava
    diets. High cyanide cassava diets were also associated with a
    significant increase in the numbers of runts compared to litters
    from dams fed either low protein or laboratory stock diets (Frakes
     et al., 1986).

    2.2.6  Special studies on the thyroid gland

         In a study cited by Oke (1980), the influence of a 100% cassava
    diet on the thyroid in a 7-day experiment with rats. A significant
    decrease in glandular stores of stable iodine, significantly higher
    thyroid weight and higher thyroidal 131I uptake were observed.
    Each effect is due to a synthetic block in the conversion of
    monoiodothyronine to diiodothyronine.

    2.3  Observations in humans

    2.3.1  Acute toxicity studies in humans

         One to 10 g of amygdalin have been given parenterally in
    humans, apparently without acute toxicity. This indirectly suggests
    that there is no significant metabolism of the intact injected
    glycoside. The cyanide-containing breakdown products possess
    well-defined toxicities, and 50 mg of hydrogen cyanide can be fatal.

         With oral dosing of amygdalin, a toxic potential is manifest.
    ▀-Glucosidase is present in the gastrointestinal lumen, a
    contribution of intestinal microflora. According to Eyerly (1976)
    oral laetrile (amygdalin) could be 40 times more toxic than
    parenterally administered doses. This is probably due to the free
    HCN released by the ▀-glucosidase enzyme present in the gut (Dorr &
    Paxinos, 1978).

         The lethal dose of amygdalin for man when ingested is reported
    to be in the range of 0.02-0.13 mmol/kg bw (Solomonson, 1981).

         If it is assumed that about 100-2000 mg HCN is the lethal dose
    for man, as much as 10-20 kg of Lafun cassava (10-20 mg cyanide/kg)
    will have to be consumed at a sitting to produce toxicity
    (Oke, 1980).

         Well-nourished individuals have ingested 1000 mg or more of
    pure amygdalin every day without any evidence of "side effects"
    (Oke, 1979).

         In a case study an 11-month-old girl was reported accidentally
    to have ingested 1-5 amygdalin tablets (500 mg). The patient became
    listless within one half hour of ingestion and vomited. Breathing
    became irregular and her state of consciousness became altered. An
    hour after ingestion she was in shock and died approximately 72 h
    following ingestion in spite of hospital treatment (Humbert  et al.,
    1977).

         In a case-study a 17-year-old girl suffering from cancer made a
    practice of taking, instead of radiotherapy, four ampoules of
    laetrile (3 g amygdalin) intravenously. One day she swallowed three
    1/2 ampoules of laetrile. Shortly after ingestion, a severe headache
    and dizziness developed, and she collapsed. Laboured breathing
    developed, her pupils became dilated, and she became comatose. All
    symptoms occurred within 8-10 minutes after ingestion. She died 24 h
    after ingestion (Sadoff  et al., 1978).

         In Anatolia (Turkey) 9 cases of cyanide intoxication of
    children due to the ingestion of wild apricot seeds (217 mg
    HCN/100g) were reported. The victims had probably eaten more than 10
    seeds. Also in studies of Jeanin  et al. (1961) and Pijoun (1942)
    poisoning after consuming a relative large amount of peach seeds or
    bitter almonds are reported. Quantitative figures on cyanogenic
    glycoside or cyanide intake are not given (Sayre & Kaymakcalavu,
    1964).

         The toxicity of cassava and cassava processing products was
    until recently assumed to be associated with free cyanide, this was
    50-60 mg which constitutes a lethal dose for an adult man. The
    cyanogenic glycosides were at first thought to be of little
    consequence to mammals if cassava hydrolytic enzymes have been
    inactivated. The possibility of hydrolysis during digestion,
    however, is also important (Cooke & Coursey, 1981).

         In a case-study a 67-year-old woman collapsed after ingestion
    of a slurry of 12 bitter almonds ground up and mixed with water. She
    recovered after treatment in the hospital. The average cyanide
    content was 6.2 mg HCN/bitter almond (Shragg  et al., 1982).

         The consumption of 60 bitter almonds is deadly for an adult.
    For young children, however 5-10 almonds or 10 droplets of bitter
    almond oil are fatal (Askar & Moral, 1983).

    2.3.2  Long-term toxicity studies in humans with cyanogenic
           glycosides and cyanides

         A study to evaluate the possible association of high cyanide
    and low sulfur intake in cassava-induced spastic paraparesis was
    performed. The north-eastern part of Mozambique suffered a severe
    drought in 1981: the only crop available was the most toxic variety
    of cassava and, due to lack of food during the harvest period, the
    roots were eaten after only a few days of sun drying. A field survey
    revealed 1102 cases of spastic paraparesis. In 1982 urine was
    collected from 30 apparently healthy children (age 8.1 years). As
    reference 17 Swedish children (age 8.6 years) were used. In a second
    stage urine was sampled in 1983 (when the nutritional situation was
    improved but still unsatisfactory) from 31 children (9.0 years) in
    the same village and 30 schoolchildren (8.1 years) in a nearby
    district where no cases of paraparesis were seen in 1981 and from 28
    children (7.1 years) of the city who ate virtually no cassava. The
    children from the village had increased thiocyanate and decreased
    inorganic sulfate excretion, indicating high cyanide and low
    sulfur-containing amino acid intake. Children from a neighbouring
    cassava-eating area, where no cases of spastic paraparesis had
    occurred, had lower thiocyanate excretion but higher organic sulfate
    excretion. These results support the hypothesis that the epidemic
    was due to the combined effects of high dietary cyanide exposure and
    sulfur deficiency (Cliff  et al., 1985).

         Several studies were performed, including epidemiological
    studies, on the role of chronic cyanide intoxication caused by the
    consumption of cassava diet in the etiology of tropical (ataxia)
    neuropathy (TAN) in Nigerian populations. As described in over 400
    Nigerian patients the essential neurological components of the
    disease are myelopathy, bilateral optic atrophy, bilateral
    perceptive deafness and polyneuropathy. The initial and most common
    symptoms consist of various forms of paraesthesia and dysaesthesia
    usually starting in the distal part of the lower limbs. The next
    most common finding is blurring or loss of vision. Other common
    symptoms in order of frequency are ataxia, tinnitus, deafness,
    weakness and thinning of the legs.

         In about a third of the patients stomatoglossitis is present,
    additionally motor neurone disease, Parkinson's disease, cerebellar
    degeneration, psychosis and dementia have been associated with the
    disease. TAN affects males and females in all age groups equally,
    but occurs only rarely in children under 10 years. Patients usually
    give histories of almost total dependence on a monotonous diet of
    cassava derivatives; occasional dietary supplements include yam,

    maize, rice, vegetables and animal protein. Analysis of family
    relationships among patients showed no evidence of a genetically
    determined predisposition. The families were usually poor and
    members lived communally.

         Clinical evidence of malnutrition was frequently absent. The
    occurrence of cyanide intoxication in Nigerian patients was
    indicated by the significantly higher cyanide and thiocyanate plasma
    levels and higher excretion of thiocyanate than controls. Hepatic
    rhodanese activity was not different from that in controls and
    histology of liver biopsy specimens showed no abnormality. Total
    plasma vitamin B12 levels are normal or high in patients and healthy
    Nigerians but plasma concentration of cyanocobalamin was highly
    significantly raised in patients. A small proportion of
    cyanocobalamin was found in the liver of patients. Methylmalonic
    acid excretion was normal in patients, indicating that there was
    physiological vitamin B12 adequacy at tissue or cellular level in
    these patients.

         In Nigeria, endemic foci of the disease (in epidemiological
    studies) recognized since the early 1930's, correspond with the
    areas where cassava is intensively cultivated and consumed as the
    major or the sole dietary source of carbohydrate. There was evidence
    of increased exposure to cyanide in members of families where
    multiple cases were found. In biochemical studies no biochemical
    evidence of protein-calorie malnutrition was seen and serum
    transferrin, said to be sensitive index of protein nutritional
    status, was normal. No haematological abnormalities were seen in
    patients. Investigation of patient nutritional status with regard to
    the water-soluble vitamins showed no abnormality, except in
    riboflavin intake. Plasma concentrations of calcium, phosphate,
    sodium, potassium, chloride, bicarbonate, and cholesterol, and tests
    of thyroid, hepatic, and renal functions were normal. Amino acids
    and porphyrobilinogen and other porphyrins were absent in urine.
    There was no biochemical nor other evidence of malabsorption.
    Glucose tolerance tests were normal. Histamin-fast achlorhydria was
    very rare. Serologic tests for syphilis, typhoid, typhus,
    brucellosis and screening for prevalent viral infections gave
    negative or insignificant results. There was no increased prevalence
    of malaria and urinary tract infections were not encountered in
    patients. ECG's were normal. Diminished urinary excretion and
    riboflavin (vitamin B2) and low serum riboflavin and caeruloplasmin
    levels in patients compared with healthy persons were the only
    significant abnormalities found. Riboflavin deficiency is, however,
    widespread in many parts of Nigeria and especially in areas endemic
    for TAN, probably because cassava is a poor source of the vitamin.
    The daily intake of HCN from cassava derivatives in areas in Nigeria
    endemic for TAN may be as high as 50 mg, which is nearly an acutely
    sublethal amount, and can conceivably produce cyanide intoxication.
    This is particularly plausible as cassava root is a major item of

    food, often the main source of carbohydrate, and is poor in
    sulfur-containing amino acids which are essential for detoxification
    of cyanide. A high prevalence (2%) of goitre in populations with a
    high incidence of TAN is seen; this appears to be related to cassava
    diet and high plasma thiocyanate. The effects of riboflavin
    insufficiency may combine with those of chronic cyanide intoxication
    in the etiology of TAN (Osuntokun, 1981).

         Several epidemiological and experimental studies revealed that
    TAN resulted from chronic cyanide poisoning due to the release of
    HCN from cyanogenic glycosides present in certain cassava food
    products. Since TAN invariably occurs among poorly nourished people,
    the condition may result from unidentified nutritional deficiencies
    or excesses as well as cyanide toxicity. Among patients showing the
    TAN syndrome the frequency of goitre was increased (Conn, 1979a,b).

         From both experimental and epidemiological studies there is
    strong, but not conclusive, evidence that cassava toxicity is a
    causative factor in some neurological disorders like TAN and endemic
    spastic paraparesis. As well, different deficiencies such as
    deficiency of sulfur-containing amino acids may play a role. The
    geographical distribution of malnutrition-related diabetes coincides
    with that of cassava consumption. Dietary exposure to cyanide has
    been proposed as a possible cause, but has not been proven. The poor
    nutritional quality of cassava seems a very likely causative factor
    (Rosling, 1987).

         Workers in certain occupations are exposed to HCN additional to
    sources encountered by the general public. These individuals have
    been studied to determine whether chronic cyanide poisoning is a
    clinical entity. Symptoms reported were headache, vertigo, tinnitus,
    nausea, vomiting, and tremors. Although these symptoms are
    sufficiently documented and characteristic, they are transitory in
    that exposure to fresh air causes recovery. These do not seem to
    produce the outward symptoms of TAN, the pathological condition that
    has been attributed in part to exposure to cyanide or cyanogenic
    glycosides in certain preparations of cassava (Conn, 1979b).

         Neurological disorders such as ataxic neuropathy and cretinism
    have been associated directly with the intake of cyanogenic
    glycoside-contaminated diets. There are grounds to suspect that
    cyanogenic glycoside-contaminated foodstuffs such as cassava and
    pulses are directly implicated in acute and chronic cyanide toxicity
    in the tropics. Although correlation between dietary
    cyanogen-contaminants and disease such as TAN, goitre, cretinism and
    mental retardation exists and is based on experimental evidence, the
    mechanism of action of cyanide and thiocyanate on the cellular level
    is not well understood (Nartey, 1980).

         Based on several studies in humans consuming as their main food
    cassava or other food products rich in cyanogenic glycosides, it
    seems that chronic cyanide intoxication in combination with
    deficient intake of riboflavin and/or a poor quality of protein and
    hence methionine deficiency is/are responsible, to a large extent,
    for the etiology of TAN in cassava-eating areas, whereas chronic
    cyanide intoxication in combination with a deficient iodine intake
    is responsible for goitre (Oke, 1979, 1980).

         Epidemiological and experimental studies show that cyanogenic
    glycosides in food products play a important role in the development
    of goitre. Thiocyanate, the detoxification product from the HCN
    derived from cyanogenic products, is responsible for interference
    with thyroid function. Studies on endemic goitre in Africa have
    identified iodine deficiency and an antithyroid activity of cassava
    diets as major etiological factors of the disease (Conn, 1979a,b).

         Extensive studies in Zaire have established that goitre and
    cretinism due to iodine deficiency can be considerably aggravated by
    a continuous dietary cyanide exposure from insufficiently processed
    cassava. This effect is caused by thiocyanate. Thiocyanate has a
    similar size to the iodine molecule and interferes with the iodine
    uptake in the thyroid gland. Thiocyanate levels which can occur
    after exposure to cyanide from cassava can only affect the gland
    when the iodine intake is below 100 micrograms/day, which is
    regarded minimal for normal function. Several studies have shown
    that populations with considerable cyanide exposure from
    inadequately processed cassava are free from goitre as long as their
    iodine intake is sufficient. Populations in northern Zaire with very
    low iodine intake and in addition high thiocynate levels resulting
    from the consumption of cassava products, show very severe endemic
    goitre. When the population was given iodine supplementation, the
    goitre decreased (reviewed by Rosling, 1987).

         In persons who ingested cassava a decreased 131I-uptake by
    the thyroids was seen, confirming the goitrogenous nature of cassava
    (De Lange, 1973 as cited by Oke, 1980).

         In one study more than 50% of workers in a processing factory
    revealed thyromegaly and an increased 131I-uptake. All workers had
    increased blood haemoglobin concentration as well as increased
    lymphocyte counts. Two of the employees developed psychosis,
    although it is difficult to know whether this was cause or effect.
    Abnormal thyroid function has also been found in workers from a
    photographic plant in which a cyanide extracting process was used to
    recover silver from X-ray films (El Ghawabi  et al., 1975).

         Alterations in vitamin B12 and folate metabolism have also been
    noted to play a role, although the significance of these
    observations are not known. Short of having an elevated whole blood

    cyanide level, the laboratory findings of cyanide poisoning are
    fairly non-specific. Perhaps the most valuable clue is the finding
    of a severe metabolic acidosis with a high anion gap. In clinical
    medicine there have been reported only 7 causes of anion gap
    metabolic acidosis. In most of these disorders lactic acidosis is
    present owing to severe tissue underperfusing and hypoxia. In the
    patient with cyanide poisoning tissue hypoxia is universally present
    (Gonzales & Sabatini, 1989).

    3.  COMMENTS AND EVALUATION

         The potential toxicity of a cyanogenic plant depends primarily
    on its capacity to produce a concentration of hydrogen cyanide toxic
    to animals and humans. The release of hydrogen cyanide can occur
    either following maceration of the plant material - this activates
    the intracellular ▀-glucosidase which in turn hydrolyses glycoside -
    or by hydrolysis of glycoside by the ▀-glucosidase produced by the
    microflora of the gut. The level of ▀-glucosidase activity in the
    gut depends on the pH and the bacterial composition. The cyanogenic
    glycoside content of a foodstuff, when known, is usually expressed
    in terms of the amount of cyanide released by acid hydrolysis; exact
    figures for the concentration of the glycosides themselves are very
    rarely given.

         Hydrogen cyanide absorbed from the gut can be detoxified by
    metabolic conversion to thiocyanate; this depends on the presence of
    nutritional factors, such as sulfur-containing amino acids and
    vitamin B12. Acute toxicity results when the rate of absorption of
    hydrogen cyanide is such that the metabolic detoxification capacity
    of the body is exceeded.

         Available reports of toxicological studies lack information on
    the level of intake of cyanogenic glycosides or on the amount of
    hydrogen cyanide potentially released. No long-term toxicity or
    carcinogenicity studies were available to the Committee. However,
     in vitro and  in vivo genotoxicity were negative. Teratogenic and
    adverse reproductive effects attributable to linamarin (cassava) and
    hydrogen cyanide were seen only at doses that also caused maternal
    toxicity.

         The toxic effects of cyanide on the thyroid (via its metabolite
    thiocyanate) depend on the iodine status of the test animals, as
    indicated in the twenty-fifth report of the Committee (Annex 1,
    reference 56).

         On the basis of epidemiological observations, associations have
    been made between chronic exposure to cyanogenic glycosides and
    diseases such as spastic paraparesis, tropical ataxic neuopathy, and
    goitre. However, these observations were confounded by nutritional
    deficiencies, and causal relationships have not been definitely
    established.

         Traditional users of foods containing cyanogenic glycosides
    usually have a basic understanding of the treatment required to
    render them safe for consumption. However, some products are sold
    commercially and are consumed by people who may not be familiar with
    such procedures. The Committee therefore recommended that guidelines

    be developed to provide reliable and sensitive methods for the
    analysis of these foodstuffs for hydrogen cyanide releasable from
    cyanogenic glycosides, in order to ensure that amounts in foods as
    consumed do not present a hazard.

         Because of a lack of quantitative toxicological and
    epidemiological information, a safe level of intake of cyanogenic
    glycosides could not be estimated. However, the Committee concluded
    that a level of up to 10 mg/kg hydrogen cyanide in the Codex
    Standard for Cassava Flour (CAC, 1991) is not associated with acute
    toxicity.

    4.  REFERENCES

    ASKAR, A., & MORAD, M.M. (1983). Lebensmittelvergiftigung 1. Toxine
    in natŘrlichen Lebensmittel.  Alimentia, 19: 59-66.

    BARRETT, M.D., HILL, D.C., ALEXANDER, J.C. & ZITNAK, A. (1977). Fate
    of orally dosed linamarin in the rat.  Can. J. Physiol. Pharmacol.,
    55: 134- 136.

    BASS, N.H. (1968). Pathogenesis of myelin lesions in experimental
    cyanide poisoning: a microchemical study.  Neurology, 18: 167-177,
    as cited in Osuntokun, 1981.

    BASU, T.K. (1983). High-dose ascorbic acid decreases detoxification
    of cyanide derived from amygdalin (laetrile): Studies in
    guinea-pigs.  Can. J. Physiol. Pharmacol., 61: 1426-1430.

    BRIERLEY, J.B., PRIOR, P.F., CALVERLEY, J. & BROWN, A.W. (1977).
    Cyanide intoxication in  Macaca mulatta: Physiological and
    neurological aspects.  J. Neurol. Sci., 31: 133-157, as cited in
    Osuntokun, 1981.

    CAC (1991). Codex Alimentarius, Vol. XII, Suppl 4. Codex Standard
    for Edible Cassava Flour (African Regional Standard). Rome, Food and
    Agriculture Organization of the United Nations (CODEX STAN 176).

    CLARK, A. (1936). Report on effects of certain poisons contained in
    food plants of West Africa upon health of native races.  J. Trop.
     Med. Hyg., 39: 285-295, as cited in Osuntokun, 1981.

    CLIFF, I., LUNDQUIST, P., M─RTENSSON, I., ROSLING, H. & SÍRBO, B.
    (1985). Association of high cyanide and low sulfur intake in
    cassava-induced spastic paraparesis.  Lancet, II: 1211-1213

    COLLIER JACKSON, J. (1988). Behavioral effects of chronic sublethal
    dietary cyanide in an animal model: implications for humans
    consuming cassava  (Manihot esculenta). Journal of the Society for
     the Study of Human Biology, 60: 597-614.

    CONN, E.E. (1979a). Cyanide and cyanogenic glycosides. In Rosenthal,
    G.A. & Janzen, D.H. (eds.), Herbivores: Their interaction with
    secondary plant metabolites, Academic Press, Inc., New York-London,
    pp 387-412.

    CONN, E.E. (1979b). Cyanogenic glycosides. International review of
    biochemistry. In Biochemistry and Nutrition 1A, Neuberger, A., &
    Jukes, T.H. (eds), University Park Press, Baltimore, 27: 21-43.

    COOKE, R.D. & COURSEY, D.G. (1981). Cassava: A major
    cyanide-containing food crop. In Vennesland, B., Conn, E.E.,
    Knowles, W, Westley, J. & Wissing, F. (Eds).  Cyanide in Biology.
    Academic Press, London.

    DOHERTY, P.A., FERN, V.H. & SMITH, R.P. (1982). Congenital
    malformations induced by infusion of sodium cyanide in the Golden
    hamster.  Toxicology and Applied Pharmacology, 64: 456-464.

    DORR, R.T. & PAXINOS, I. (1978). The current status of laetrile.
     Annals of Internal Medicine, 89: 389-397.

    EPA (1990). Summary Review of Health Effects Associated with
    Hydrogen Cyanide, Health Issue Assessment Environmental Criteria and
    Assessment Office, Office of Health and Environmental Assessment
    Office of Research and Development, US Environmental Protection
    Agency Research Triangle Park, North Carolina, USA.

    EYERLY, R.C. (1976). Laetrik; focus on the the facts.  Cancer, 26:
    50-54.

    FERRARO, A. (1933). Experimental toxic encephalomyelopathy (diffuse
    sclerosis following subcutaneous injection of postassium cyanide).
     Archs. Neurol. Psychiat., 29: 1364-1367, as cited in Osuntokun,
    1981.

    FRAKES, R.A., SHARMA, R.P. & WILLHITE, C.C. (1985). Development
    toxicity of the cyanogenic glycoside linamarin in the golden
    hamster.  Teratology, 31: 241- 246.

    FRAKES, R.A., SHARMA, R.P., WILLHITE, C.C. & GOMEZ, G. (1986).
    Effect of cyanogenic glycosides and protein content in cassava diets
    on hamster prenatal development.  Fundamental and Applied
     Toxicology, 7: 191-198.

    FRAKES, R.A., SHARMA, R.P. & WILLHITE, C.C. (1986). Comparative
    metabolism of linamarin and amygdalin in hamsters.  Food Chemical
     Toxicology, 24: 417-420.

    GEITLER, A.O. & BAINE, J.O. (1983). The toxicity of cyanide.  Am. J.
     Med. Sci., 195: 182-198.

    GOMEZ, G., APARICIO, M.A. & WILLHITE, C.C. (1988). Relationship
    between dietary cassava cyanide levels and Brailer performance.
     Nutrition Reports International, 37: 63-75.

    GONZALES, I. & SABATINI, S. (1989). Cyanide poisoning:
    pathophysiology and current approaches to therapy.  The Int. I. of
     Artificial Organs, 12(6): 347-355.

    HIMWICH, W.A. & SAUNDERS, I.P. (1984). Enzymatic conversion of
    cyanide to thiocyanate.  American Journal of Physiology, 58: 348.

    HUMBERT, I.R., TRESS, I.H. & BRAICO, K.T. (1977). Fatal cyanide
    poisoning: accidental ingestion of amygdalin.  JAMA, 238: 482.

    HURST, E.W. (1940). Experimental demyelination of the central
    nervous system.  Aust. J. Exp. Biol. Med. Sci., 18: 201-223, as
    cited in Osuntokun, 1981.

    LANG, K. (1933) Die Rhodanbildung im Tierk÷rper.  Biochem Z., 259:
    243-256.

    LESSELL, S. (1971) Experimental cyanide optic neuropathy.  Archs.
    Ophtal., 84: 194-204, as cited in Osuntokun, 1981.

    LEUSCHNER, F., NEUMANN, B.W., & LIEBSCH, M. (1983a). Mutagenicity
    study of hydrocyanic acid in the Ames  Salmonella/microsome plate
    test  (in vitro). Unpublished study, Laboratory of Pharmacology and
    Toxicology, Hamburg, August 1983, submitted to the WHO by Detia
    Freyberg GmbH.

    LEUSCHNER, F., NEUMANN, B.W., & LIEBSCH, M. (1983b). Mutagenicity
    study of hydrocyanic acid in Chinese hamster (chromosome aberration)
    by oral administration. Unpublished study, Laboratory of
    Pharmacology and Toxicology, Hamburg, August 1983, submitted by
    Detia Freyberg GmbH.

    LEUSCHNER, F. & NEUMANN, B.W. (1989a).  In vitro mutation assay of
    KCN in Chinese hamster cells. Unpublished study, Laboratory of
    Pharmacology and Toxicology, July 1989, submitted by Detia Freyberg
    GmbH.

    LEUSCHNER, F., NEUMANN, B.W., OTTO, H. & MÍLLER, E. (1989b). 13-Week
    toxicity study of potassium cyanide administered to Sprague-Dawley
    rats in the drinking water. Unpublished study, Laboratory of
    Pharmacology and Toxicology, July 1989, submitted by Detia Freyberg
    GmbH.

    MAJAK, W., MCDIARMID, R.E., JAKOBER, K. & CHENG, K.I. (1989).
    Diurnal changes in rates of degradation of cyanogenic glycosides in
    bovine rumen fluid.  Toxicon., 27: 61.

    NARTEY, F. (1980). Toxicological aspects of cyanogenesis in tropical
    foodstuffs in Toxicology in the Tropics. Editors R.L. Smith and E.A.
    Bababumni, Taylor & Francis Ltd, London, 53-73.

    OKE, O.L. (1979). Some aspects of the role of cyanogenic glycosides
    in nutrition.  Wld. Rev. Nutr. Diet, 33: 70-103.

    OKE, O.L. (1980). Toxicity of cyanogenic glycosides.  Food
     Chemistry, 6: 97-109.

    OKOH, P.N. & PITT, G.A.J. (1981). The metabolism of cyanide and the
    gastrointestinal circulation of the resulting thiocyanate under
    conditions of chronic cyanide intake in the rat.  Can. J. Physiol.
     Pharmacol., 60: 381-385.

    OKOH, P.N. (1983). Excretion of 14C-labeled cyanide in rats
    exposed to chronic intake of potassium cyanide.  Toxicology and
     Applied Pharmacology, 70: 335-339.

    OLUSI, S.O., OKE, O.L. & ODUSOTE, A.C. (1979). Effects of cyanogenic
    agents on reproduction and neonatal development in rats.  Biol.
     Neonate, 36: 233-293.

    OSUNTOKUN, B.O. (1981). Cassava diet, chronic cyanide intoxication
    and neuropathy in the Nigerian Africans.  Wld. Rev. Nutr. Diet, 36:
    141-173.

    PHILBRICK, D.I., HOPKINS, I.B., HILL, D.C., ALEXANDER, I.C. &
    THOMSON, R.G. (1979). Effects of prolonged cyanide and thiocyanate
    feeding in rats.  Journal of Toxicology and Environmental Health,
    5: 579-592.

    RAUWS, A.G., OLLING, M. & TIMMERMAN, A. (1982). The pharmacokinetics
    of amygdalin.  Arch. Toxicol., 49: 311-319.

    RAUWS, A.G., OLLING, M. & TIMMERMAN (1983). The pharmacokinetics of
    prunasin, a metabolite of amygdalin.  J. Toxicol. Clin. Toxicol.,
    19: 851-856.

    ROSLING, H. (1987). Cassava toxicity and food security. Ed. Rosling.
     Tryck Kontakt, Uppsala, Sweden, 3-40.

    SADOFF, L., FUCHS, K. & HOLLANDER, I. (1978). Rapid death associated
    with laetrile ingestion.  JAMA, 239: 1532.

    SAYRE, I.W. & KAYMAKCALAVU, S. (1964). Cyanide poisoning from
    apricot seeds among children in Central Turkey.  New. Engl. J. Med.,
    270: 1113-1118.

    SHRAGG, T.A., ALBERTSON, T.E. & FISHER, C.J. (1982). Cyanide
    poisoning after bitter almond ingestion.  The Western Journal of
     Medicine, 136: 65-69.

    SMITH, A.D.M., DUCKETT, S. & WATERS, A.H. (1963) Neuropathological
    changes in chronic cyanide intoxication.  Nature, 200: 179-181, as
    cited in Osuntokun, 1981.

    SOLOMONSON, L.P. (1981). Cyanide as a metabolic inhibitor. In
    Cyanide in Biology, Vennesland, B., Conn, E.E., Knowles, C.J.,
    Westley, J & Wissing, F. Academic Press, London, New York, Toronto,
    11-18.

    TEWE, O.O. & MANER, I.H. (1980). Cyanide, protein and iodine
    interactions in the performance, metabolism and pathology of pigs.
     Research in Veterinary Science, 29: 271-276.

    TEWE, O.O. & MANER, I.H. (1981a). Performance and
    patholophysiological changes in pregnant pigs fed cassava diets
    containing different levels of cyanide.  Research in Veterinary
     Science, 30: 147-151.

    TEWE, O.O. & MANER, I.H. (1981b). Long-term and carry-over effect of
    dietary inorganic cyanide (KCN) in the life cycle performance and
    metabolism of rats.  Toxicological and Applied Pharmacology, 58:
    1-7.

    UMOH, L.B., MADUAGWA, E.N. & AMOLE, A.A. (1986). Fate of ingested
    linamarin in malnourished rats.  Food Chemistry, 20: 1-9.

    VENNESLAND, B., CASTRIC, P.A., CONN, E.E., SOLOMONSON, L.P., VOLINI,
    M. & WESTLEY, I. (1982). Cyanide metabolism.  Fed. Proc., 41(10):
    2639-2648.

    WILLHITE, C.C. (1982). Congenital malformations induced by laetrile.
     Science, 215: 1513-1515.

    WILLIAMS, A.O. & OSUNTOKUN, B.O. (1969). Light and electron
    microscopy of peripheral nerves in tropical ataxic neuropathy.
     Archs. Neurol., 21: 475-492, as cited in Osuntokun, 1981.

    WHO (1965). Evaluation of the hazards to consumers resulting from
    the use of fumigants in the protection of food. Report of the second
    Joint Meeting of FAO Committee on Pesticides in Agriculture and the
    WHO Expert Committee on Pesticides Residues, FAO Meeting Report No
    Pl/1965/10/2: WHO Food series 28.65.


    See Also:
       Toxicological Abbreviations
       CYANOGENIC GLYCOSIDES (JECFA Evaluation)