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    -CYCLODEXTRIN

    First draft prepared by
    Ms F D Pollitt
    Health Aspects of the Environment and Food (Medical) Division
    Department of Health
    London, England

    Explanation
    Biological data
    Toxicological studies
         Short-term toxicity studies
         Long-term toxicity/carcinogenicity studies
         Reproductive toxicity studies
         Special studies on interactions with bile
           acids and lipophilic nutrients
    Comments
    Evaluation
    References

    1.  EXPLANATION

         -Cyclodextrin was previously evaluated at the forty-first
    meeting of the Committee (Annex 1, reference 107), at which time a
    temporary ADI of 0-6 mg/kg bw was allocated, based on a NOEL of 2.5%
    in the diet (equal to 1230 mg/kg bw/day) in study in dogs and a safety
    factor of 200. The results of an ongoing 1-year oral toxicity study in
    dogs and information on the effects of -cyclodextrin on the
    bioavailability of lipophilic nutrients were required by 1995. These
    data, along with the results of a 1-year toxicity study in rats, a
    3-generation reproductive toxicity study in rats and carcinogenicity
    studies in mice and rats were available for review at the present
    meeting, and are summarized in the following monograph addendum.

    2.  BIOLOGICAL DATA

    2.1  Toxicological studies

    2.1.1  Short-term toxicity studies

    2.1.1.1  Rats

         In a 52-week study, groups of 20 male and 20 female CrI:CD(SD)BR
    rats were fed diets containing 0, 1.25, 2.5 or 5% -cyclodextrin (>
    99% pure). Treatment commenced at 6 weeks of age. The rats were housed
    5 to a cage and food and water were available  ad libitum. Animals
    were observed daily for clinical signs. Body weight and food intake
    were recorded weekly. Haemato-logical (including RBC, WBC, haemoglobin
    and haematocrit), plasma biochemical (including protein, ASAT, ALAT,
    ALP, gamma-GT, sodium, potassium, calcium and triglycerides) and urine
    analyses (including pH and protein) were undertaken during weeks 13,
    26, 39 and prior to termination at 52 weeks. Food was withdrawn
    overnight prior to sampling and water was also withdrawn overnight
    prior to urine sampling. Major organs were weighed. At termination,
    full macroscopic and microscopic examination of tissues and organs was
    undertaken.

         During the course of the study, 4 deaths were recorded, 3 males
    from the control group and 1 female which had received diet containing
    1.25% -cyclodextrin. None of these deaths were attributed to
    treatment. No abnormal clinical signs were observed.

         Since the dietary concentration was fixed throughout the study
    period, the intakes of -cyclodextrin, expressed as mg/kg bw/day,
    decreased. The intakes of -cyclodextrin (mg/kg bw/day) at the
    beginning and end of the study are shown in Table 1.

    Table 1. Variation of intake in the 52-week study (HRC, 1994a)
                                                                        

    % -Cyclodextrin              Intake, mg/kg bw/day
    in diet
                                                                        

                              Male                    Female
                                                                        

                        Week 1    Week 52       Week 1       Week 52
                                                                        

    1.25                1506      395           1593         565
    2.5                 3292      797           3227         1146
    5                   6106      1639          6358         2391
                                                                        

         In males, intake had decreased in week 52 to approximately 25% of
    that at week 1 and, in females, to approximately 33%. The mean intake
    in the 1.25% group over the course of the study was 650 mg/kg bw/day
    and 860 mg/kg bw/day in male and female rats, respectively.

         There was no statistically significant effect of treatment on
    body weight during the course of the study, although the body-weight
    gains in the high-dose males and females were slightly reduced in
    comparison to controls (4% and 7%, respectively).

         No effects due to -cyclodextrin were observed on haematological
    or urinalysis parameters. In the biochemical investigations, ALAT and
    ASAT were increased significantly compared to controls at week 13
    in both males and females in the high- and mid-dose groups. The
    concentrations of these enzymes continued to be increased in both
    males and females in these groups throughout the study except at week
    39, when the increase was only observed in the high-dose females. At
    week 13, significant increases in OCT levels were observed in mate and
    female animals receiving 5% -cyclodextrin and in males receiving 2.5%
    -cyclodextrin. Increased OCT concentrations were also observed at
    other times throughout the study, but these differences were not
    always statistically significant when compared with control values. At
    week 52, significant increases were observed in males receiving 5 and
    2.5% -cyclodextrin.

         Triglyceride concentrations tended to be reduced compared to
    controls from week 26 onwards in the high-dose male rats and from
    week 39 in high-dose female rats, the difference occasionally
    reaching statistical significance. There was a tendency to reduced
    -lipoprotein concentrations at the high dose and, occasionally, the
    mid dose but the biological significance of this was unclear since
    statistically significantly lower levels than in controls were also
    observed in predose high-dose animals and, in males, in mid-dose blood
    samples.

         Analysis of the urine for -cyclodextrin at weeks 13 and 52
    demonstrated the presence of a low level of unchanged compound.
    Analysis of the faeces showed virtual absence of -cyclodextrin.

         Analysis of organ weights at 52 weeks revealed no statistically
    significant differences between treated groups and controls. No
    effects attributable to -cyclodextrin were observed at macroscopic
    examination of the tissues. However, histological examination revealed
    treatment-related changes in the liver and kidney. In the liver,
    significantly increased incidences of single cell necrosis,
    centrilobular hepatocyte enlargement and portal inflammatory cell
    infiltration were seen in both males and females fed 5% -cyclodextrin
    compared to controls. Females from this same dose group were also
    observed to have an increased incidence of focal basophilic

    hepatocytes. Males fed 2.5% -cyclodextrin had an increased incidence
    of portal inflammatory cell infiltration, while females of this group
    showed an increased incidence of single cell necrosis and focal
    basophilic hepatocytes. Examination of the kidneys revealed an
    increased incidence of pigment in the epithelium of the cortical
    tubules in females fed 2.5% or 5% -cyclodextrin, but no treatment-
    related changes were observed in males. No other treatment-related
    abnormalities were observed in any of the other tissues examined. The
    NOEL in this study was 1.25% in the diet, equal to 650 mg/kg bw/day
    (HRC, 1994a).

    2.1.1.2  Dogs

         The oral toxicity of -cyclodextrin (purity >99%) was studied in
    beagle dogs for a period of 52 weeks. The animals were acclimatized
    for a minimum of 4 weeks during which time they were inoculated and
    received anthelmintic treatment. Following acclimatization, the dogs
    were divided into 4 groups each of 4 males and 4 females and were
    administered by inclusion in the diet, 0%, 0.62%, 1.25%, or 5.0%
    -cyclodextrin. The age of the animals at the start of treatment was
    35-36 weeks and their body weights ranged from 8.4-11.8 kg. The dogs
    were fed once daily and had free access to water. They were housed in
    kennels capable of accommodating 2 animals of the same sex and dose
    group. However, dogs were segregated for the assessment of clinical
    signs and food consumption which were recorded daily. Body weights
    were recorded weekly. Ophthalmic, haematological (including
    haematocrit, haemoglobin and blood cell counts), plasma biochemical
    (including protein, urea, inorganic salts, ALAT, ASAT and gamma-GT)
    and urine (including volume, pH and protein) analyses were undertaken
    after 13, 26, 39 and 52 weeks of treatment. Urine and faecal samples
    were collected during weeks 13 and 52 for toxicokinetic purposes.
    Serum vitamin A, D and E were determined at 13, 26, 39 and 52 weeks
    while vitamins A and E were determined in aliquots of liver at
    termination of the study. Major organs were weighed but the caecum was
    not routinely weighed. A full microscopic examination of H and E
    stained sections was performed. Liver sections were also stained with
    Oil Red O (for fat) and PAS (for glycogen).

         There were no deaths during the course of this study. All animals
    receiving -cyclodextrin showed a slightly higher incidence of
    intermittent liquid faeces compared to controls. Body-weight gain in
    both sexes in the high-dose group was reduced in comparison with
    controls, although the difference was not statistically significant.
    One female from the high-dose group showed a weight loss of 0.9 kg
    from week 31, although food consumption was, for the most part,
    maximal. The average intakes of -cyclodextrin over the study
    period were equal to 230, 470 and 1900 mg/kg bw/day for the low-,
    intermediate- and high-dose groups, respectively. There was no effect
    of treatment on food intake, haematological measurements or the
    results of ophthalmic examination.

         There was a statistically significant increase in plasma
    potassium levels in females in the mid- and high-dose dogs at weeks 39
    and 52 compared to controls. However, there were no differences in
    plasma biochemistry parameters between the control and treated groups
    which were considered to be conclusively related to treatment. Other
    differences which were statistically significant showed a lack of
    consistency and/or dose-relationship or could be attributed to
    abnormal group mean values for control dogs.

         Urinalysis revealed an increase in protein concentration in males
    fed 5.0% -cyclodextrin. Although at week 13 this was due to one high
    individual value, a trend developed over the remainder of the
    treatment period with 3/4 males showing elevated urinary protein
    concentrations at week 52. Fractionation of the proteins revealed most
    of the protein to be globular fractions with statistically
    significantly increased globulin levels usually mirroring those of
    increased total protein. This was considered to be an effect
    of treatment. There was also an increase in urinary protein
    concentrations in high-dose females compared to controls, achieving
    statistical significance at week 26, but this effect was not observed
    at termination of the study. Urinary calcium concentration was also
    increased in the high-dose male group from week 13 onwards. The same
    trend was also observed in females from week 13 onwards, but again was
    not observed at week 52. Urinary gamma-GT and N-acetyl--D-glucos-
    aminidase (NAG) levels were measured at week 52 only. The gamma-GT
    level was higher than for the control group in mid- and high-dose
    males, but the difference was only statistically significant for the
    high-dose group. NAG levels were also raised in these groups compared
    to controls but the differences were not statistically significant nor
    was there a dose-related effect. No similar effect was seen in
    females.

         Toxicokinetic analysis demonstrated the presence of unchanged
    -cyclodextrin in urine and, to a lesser extent, in faeces. During the
    24 h that the dogs were in the metabolism cages for collection of
    urine, the amount of -cyclodextrin excreted varied from 1.3 to 6.2%
    of the daily dose on week 13, and from 1.4 to 3.3% of the daily dose
    on week 52 (group mean data). There was no effect of treatment on
    serum vitamin A, D or E levels nor on vitamin A and E levels in the
    liver.

         At necropsy, group mean thymus weights in all treated male groups
    were statistically significantly higher than in the control group. No
    such effect was seen in females and no histological changes were
    observed in the thymus. This effect was therefore not considered to be
    related to treatment. In high-dose males, group mean prostate weight
    was statistically significantly reduced compared to the control group.
    No treatment-related abnormalities were recorded at histopathological

    examination in this or any other tissue examined at termination of
    this study. The Committee concluded that the NOEL in this study was
    1.25%, equal to 470 mg/kg bw/day, based on urinary effects in males
    and the slightly reduced body-weight gain seen at the high-dose level
    (HRC, 1994b).

    2.1.2  Long-term toxicity/carcinogenicity studies

    2.1.2.1  Mice

         -cyclodextrin (purity > 86.6%) was administered in the diet
    to groups of Crl-CD-I(ICR)BR mice (52/sex/dose) at dose levels of
    0, 25, 75, 225 or 675 mg/kg bw/day. Treatment commenced at about
    5 weeks of age and continued for 93 weeks (males) or 105 weeks
    (females). The mice were housed 4 to a cage and food and water were
    available  ad libitum. All animals were observed daily for clinical
    signs.

         There were no treatment-related effects on survival nor on the
    incidence or type of clinical signs. Body weights were recorded weekly
    for the first 13 weeks of treatment, biweekly until week 73 and weekly
    thereafter. Food consumption was measured weekly for the first 13
    weeks and biweekly thereafter, and water intake on week 1 and monthly
    thereafter for the control and high-dose groups only. There was no
    effect of treatment on body-weight gain, food consumption or food
    conversion ratio. There was a general trend to lower water consumption
    by the high-dose group than by the controls, but the difference was
    not statistically significant.

         At 12 and 18 months and prior to terminal sacrifice, blood was
    taken from the tail of all animals and smears for differential white
    blood cell counts were prepared. Data were presented for the control
    and high-dose groups only. A number of sporadic, statistically
    significant differences were seen at 12 and 18 months between the
    control and high-dose groups, but no such differences were seen in the
    blood samples taken prior to terminal kill. These sporadic cases of
    statistically significant findings were not considered to be
    treatment-related.

         All mice dying during the course of the study, killed in moribund
    condition, or surviving to terminal sacrifice were subjected to a
    gross necropsy. In the control and high-dose groups, histopathological
    examination was conducted on all organs. At 25, 75 and 225 mg/kg
    bw/day, only abnormalities and major organs were examined
    histopathologically. With the exception of one high-dose male,
    treatment-related lesions were seen only in decedent animals.
    Macroscopically, these lesions comprised: fluid, soft, mucoid or foamy
    and yellowish contents in caecum, colon and rectum; lumen of colon
    distended and mucosa oedematous. Microscopically, these lesions
    comprised: serious atrophy of fat pads in the abdominal cavity; active

    desquamation of surface epithelium in the caecum, colon and/or rectum;
    flattening of the mucosa, atrophy of intestinal glands and/or acute
    catarrhal typhlitis in the caecum; and mucosa covered by mucous
    secretion containing exfoliated epithelial cells in the colon and/or
    rectum. One or more of these symptoms were seen in the following
    animals where death was considered to be caused by treatment: 1 male
    at 75 mg/kg bw/day, 3 males and 2 females at the high dose. In other
    animals exhibiting these symptoms (1 male at 75 mg/kg bw/day, 1 male
    at 225 mg/kg bw/day, 4 males and 4 females at the highest dose tested)
    the cause of death was considered to be an incidental spontaneous
    lesion. The high-dose male animal sacrificed at terminal kill and
    whose symptoms were considered to be treatment-related exhibited
    caecal mucosal flattening associated with atrophy of the intestinal
    glands. No other treatment-related non-neoplastic lesions were seen in
    this study. The NOEL based on the inflammatory changes seen in the
    lower gastrointestinal tract was 25 mg/kg bw/day.

         Among the neoplastic findings, uterine endometrial polyps
    occurred as follows in the female mice: 0/51 in controls, 3/48 in the
    low-dose group, 2/52 at 75 mg/kg bw/day, 3/52 at 225 mg/kg bw/day and
    3/51 in the high-dose group. This increased incidence of uterine
    endometrial polyps showed no dose-related trend and it was concluded
    that this was not a treatment-related effect. Phaeochromocytoma of the
    adrenal gland was seen in 1/52 high-dose males and 1/51 high-dose
    females, and in 1/52 females at 75 mg/kg bw/day. No phaeochromocytomas
    were seen in the control animals, nor in 228 historical control
    animals from the same laboratory. The authors concluded that this was
    not a treatment-related effect. Although an increased incidence of
    phaeochromocytoma has been seen in rats fed polyol sweeteners which,
    like -cyclodextrin, are poorly digestible substances, they were not
    seen in mice fed similarly high levels of polyols. In view of the lack
    of association in mice and the lack of a dose-related trend in the
    incidence of this tumour in this study, the Committee agreed with the
    authors' conclusion. There were no other neoplastic findings of note
    (Gur  et al., 1993a).

    2.1.2.2  Rats

         -cyclodextrin (> 86.6% pure) was administered in the diet to
    groups of 50 male and female Fischer 344 rats at concentrations
    sufficient to provide nominal doses of 0, 25, 75,225 or 675 mg/kg
    bw/day. Treatment commenced at 4-5 weeks of age and continued for 122
    weeks (males) or 130 weeks (females). The rats were housed 5 to a cage
    and food and water were available  ad libitum. All animals were
    observed daily for clinical signs. Body weights were recorded weekly
    for the first 15 weeks of treatment and biweekly thereafter.

         Achieved dosages (mg/kg bw/day) in the first 2 weeks were only
    approximately half the nominal level but were close to the required
    level for the rest of the study. There were no treatment-related
    effects on survival nor on the incidence or type of clinical signs.
    There was no effect of treatment on body-weight gain, food consumption
    or food conversion ratio. Water consumption in the high-dose female
    rats was slightly lower than in controls throughout the study. The
    authors did not consider this to be a treatment-related effect.

         At 12, 18 and 24 months and prior to terminal sacrifice, blood
    was taken from the tail of all animals and smears for differential
    white blood cell counts were prepared. Data were presented for the
    control and high-dose groups only. There was no adverse effect of
    treatment.

         All rats dying during the course of the study, killed in moribund
    condition, or surviving to terminal sacrifice were subjected to gross
    necropsies. In the control and high-dose groups, histopathological
    examination was conducted on all organs. In the other groups, only
    abnormalities and major organs were examined histopathologically.
    There was no effect of treatment on organ weight nor on the incidence
    of macroscopic lesions or non-neoplastic microscopic lesions.

         Historical control data for the F344 rats was provided from the
    testing facility for both life-span studies (n = 60 of each sex) and
    2-year carcinogenicity studies (n = 305 of each sex) and from the US
    National Toxicology Program for life-span studies (n = 529 of each
    sex) and 2-year studies. The incidence of a few types of neoplasm
    showed a trend of statistical significance with treatment but the
    findings were not considered to be related to treatment with
    -cyclodextrin because of the lack of a clear dose-relationship and/or
    because the incidence in treated groups was well within the background
    rate for this strain of rat. These included: testicular interstitial
    cell tumours, subcutaneous fibroma, and endometrial stromal polyp and
    sarcoma combined by disease severity. The incidence of parathyroid
    adenoma also reached a low trend of significance in males and in the
    data combined by sex (males: 0, 0, 3.6, 6.9 and 7.5%; combined: 0, 2,
    2, 6.5 and 5.6% for the control, 25, 75, 225 and 675 mg/kg bw/day
    groups, respectively). The incidences in the 225 and 675 mg/kg bw/day
    dose male groups exceeded those reported for all groups of historical
    controls but, given the lack of a clear dose-relationship in the data
    combined by sex, this finding is considered unlikely to be associated
    with treatment. The finding of a renal neoplasms (one adenoma and one
    carcinoma) in two rats from the high-dose male group and none in any
    other group was not statistically significant. The incidence was high
    compared with background control data from the testing facility but
    within the range seen in the NTP studies. There was no increase in
    renal hyperplasia in treated groups (Gur  et al., 1993b).

    2.1.3  Reproductive toxicity studies

    2.1.3.1  Rats

         In a 3-generation reproductive toxicity study, -cyclodextrin
    (> 99% pure) was administered in the diet at dose levels of 0, 1.25,
    2.5 or 5% to groups of 30 male and 30 female Ico:OFA.SD.(IOPS Caw)
    rats. The parental (P) generation males and females were maintained on
    these diets for 10 and 2 weeks respectively before pairing and during
    the gestation and lactation periods of three successive mating
    periods. Male rats were approximately 6-week old at the start of
    treatment and female rats were approximately 9-week old. Two
    subsequent generations, comprised of 25 males and 25 females, randomly
    selected from the F1b and F2b litters, were treated with dietary
    concentrations of 0, 0.31, 0.62 or 1.25% -cyclodextrin: the dose
    levels were reduced 3 weeks before mating of the F1 generation to
    confirm the definition of a NOEL.

         The F1 and F2 generations were each mated twice and allowed
    to raise their offspring to weaning. The study was terminated with a
    third mating phase of the F2 animals with caesarean examination of
    the pregnant females and soft tissue and skeletal examination of the
    fetuses.

         The offspring from the third mating of the P generation (F1c
    animals) were used to investigate the effects of vitamin D
    supplementation on pup growth, because vitamin D deficiency was
    thought to be responsible for effects on pup growth. The mated females
    were divided into 2 sub-groups and 1 sub-group was given 500 IU
    vitamin D3 diluted 1/80 in 90% ethanol i.p. on day 2 of lactation.
    Milk and blood samples were taken from 5 females with viable foetuses
    from each sub-group on day 15  post-partum for clinical chemistry
    analyses.

         Animals were housed 5 to a cage during the premating periods, 1
    male and 1 female per cage during the mating period, 1 female (and its
    litter) to a cage during gestation and lactation, and with all male or
    female litter-mates in the same cage after weaning from 3 to 6 weeks
    of age. Litters were culled to 10 pups each on day 4  post-partum, to
    yield, where possible 5 males and 5 females. Food and water were
    available  ad libitum throughout. All animals were observed daily for
    clinical signs of toxicity. Body weight and food consumption were
    monitored during the respective pre-mating, gestation and lactation
    periods. Fertility and reproductive performance of the P, F1 and
    F2 generations were assessed by evaluation of mating performance,
    duration of gestation, parturition and viability, growth and
    development of pups.

         Pups which had not been selected for mating were submitted to a
    gross necropsy examination on or soon after day 21  post-partum. At
    terminal necropsy, the adult males and females of each generation were
    given a macroscopic examination; selected organs were weighed and
    selected tissues were examined histopathologically.

         Because -cyclodextrin was fed at a fixed rate in the diet,
    intake on a mg/kg bw/day basis varied considerably during the study.
    This was particularly marked in the first pre-mating period and, in
    females, during lactation, when intakes varied 2-3-fold within each
    dose group.

     P Generation

         Body-weight gains in male groups receiving -cyclodextrin were
    slightly lower than in the control group, in a dose-related manner, at
    most times, but the differences were not statistically significant.
    Conversely, body-weight gain in the high-dose female group was
    statistically significantly higher than in controls during the
    premating period but statistically significantly less during the first
    lactation. During the first and second lactations, females receiving
    5% -cyclodextrin consumed statistically significantly less food than
    the other groups; a trend to lower food consumption was also seen at
    times in the 1.25 and 2.5% female groups.

         There was no effect of treatment on mating performance nor on
    gestation at any of the three matings. There were consistently more
    dams with stillborn young in the treated groups. However, although the
    proportion of pups which were stillborn was also consistently higher
    in the treated groups, there was no dose-relationship and this finding
    can be attributed to an abnormally low incidence of stillbirths in
    controls. Litter size overall was large, averaging 14-15 live young.
    Pup viability during lactation was not affected by treatment but mean
    pup weight was statistically significantly reduced in the high-dose
    group from about day 7 to day 14  post-partum until weaning at all
    three matings. Eye opening was slightly delayed in the high-dose group
    compared to controls at all three matings but this can be attributed
    to lower pup body weight in the second half of lactation. Other
    markers of pup development, i.e. the day of occurrence of pinna
    unfolding and incisor eruption, and testing for auditory and pupil
    reflexes at weaning, were not adversely affected by treatment. The
    administration of vitamin D on day 2 of lactation had no effect on any
    parameter. There were no clear differences in clinical chemistry
    parameters of the maternal blood and milk following treatment with
    -cyclodextrin.

     F1  Generation

         Body weights of intermediate- and high-dose animals were
    statistically significantly lower than controls at the start of this
    phase but began to recover when the doses of -cyclodextrin were
    reduced, although body weights in the high-dose group remained lower
    than other groups during both mating periods. Food consumption showed
    a similar trend although the differences from controls were only
    statistically significant in the high-dose group during gestation
    after the second mating. There was no effect of treatment on female
    body weight or food consumption during gestation or lactation after
    the first mating. After the second mating, high-dose female body
    weights were statistically significantly less than controls on days 1
    and 14 of lactation only. There was no effect of treatment on
    reproductive performance or litter parameters, nor on pup viability,
    body-weight gain or development.

     F2  Generation

         There was no effect of treatment at any of the three matings on
    paternal or maternal body-weight gain or food consumption. No adverse
    effect was seen on reproductive performance, pup viability,
    body-weight gain or development. A slightly higher proportion of dams
    with stillbirths and of pups which were stillborn was seen in the
    intermediate- and high-dose groups at the first and second matings but
    there was no clear dose-response and this was not considered to be a
    treatment-related effect. The incidence of malformations and anomalies
    in fetuses delivered by Caesarian section after the third mating were
    similar in dosed and control groups. The NOEL in this study was 1.25%
    -cyclodextrin in the diet, equal to a mean achieved -cyclodextrin
    intake which ranged from 560 - 2900 mg/kg bw/day over the different
    phases of the study (Pharmakon Europe, 1994; and Personal
    communication from Dr Ph. Olivier, Roquette Frres, Lestrem, France to
    F. D. Pollitt, Department of Health, UK, submitted to WHO by F. D.
    Pollitt).

    2.1.4  Special studies on interactions with bile acids and
           lipophilic nutrients

         An  in vitro study was carried out, using NMR spectrometry, to
    evaluate interactions of -cyclodextrin with key bile acids, including
    sodium lithocholate, which are present in human bile and to study the
    competition between bile acids and the lipophilic vitamins A and D3
    for -cyclodextrin inclusion.

         It was found that, in water solutions, sodium cholate, sodium
    taurocholate, sodium chenodeoxycholate and sodium lithocholate give
    1:1 complexes with -cyclodextrin, the affinity of sodium cholate and
    sodium taurocholate for -cyclodextrin being weaker than that of
    sodium chenodeoxycholate or sodium lithocholate i.e. the affinity

    increased with an increase in the hydrophobicity of the bile acid. The
    complex of vitamin A with -cyclodextrin was shown to have a low
    solubility in water. When sodium cholate or sodium taurocholate
    entered into competition with vitamin A in the presence of a
    sub-stoichiometric quantity of -cyclodextrin, there was incomplete
    complexation of bile acids, confirming that competition occurs between
    these acids and vitamin A. However, when sodium lithocholate or sodium
    chenodeoxycholate were mixed with a vitamin A/-cyclodextrin complex
    in solution, the NMR spectrum of the mixture was identical to that of
    the lithocholate--cyclodextrin complex, showing that vitamin A does
    not interact with -cyclodextrin in the presence of these bile acids.
    Similar results were seen with vitamin D3. It was concluded that these
    findings, together with the fact that the quantity of bile acids
    present during digestion is far higher than the ingested amounts of
    vitamins, indicated that depletion of lipophilic vitamins will not
    occur when -cyclodextrin is used as an ingredient in foods or orally
    administered drugs (Comini  et al., 1994).

    3.  COMMENTS

         The Committee considered that the further toxicity studies
    confirmed the low systemic toxicity of -cyclodextrin. The NOEL in the
    1-year studies in rats and dogs was 1.25% in the diet, equal to 650
    and 470 mg/kg bw/day, respectively. At higher doses there were minor
    changes in blood biochemical and/or urine analytical parameters and,
    in the rat, minor necrotic and inflammatory changes in the liver and
    an increased incidence of pigment in conical tubular epithelium in the
    kidney. The NOEL was also 1.25% in the diet (equal to between 560 and
    2900 mg/kg bw/day, depending on the stage of the study) in the
    3-generation reproductive toxicity study in rats, in which the only
    adverse effect seen at higher doses was impaired pup growth during
    lactation, which was probably secondary to reduced food consumption
    and body-weight gain in the dams at this dose level.

         In the carcinogenicity study in mice, -cyclodextrin caused
    inflammatory changes in the lower gastrointestinal tract, which were
    considered to be the cause of death of some animals. The lowest dose
    level at which this occurred was 75 mg/kg bw/day (1/52 male affected)
    and the NOEL was 25 mg/kg bw/day. The Committee considered that these
    lesions probably represent a species-specific reaction to
    -cyclodextrin in some mice. No such effects were seen either in the
    carcinogenicity study in rats, in which the concentration of
    -cyclodextrin in the diet was about three times that in the diet of
    mice or in shorter-term studies in rats and dogs at far higher
    -cyclodextrin concentrations in the diet. No treatment-related
    neoplastic lesions were observed in the carcinogenicity studies.

         An  in vitro study indicated that -cyclodextrin is unlikely to
    deplete lipophilic vitamins when used as an ingredient in food, since
    it has a higher affinity for bile acids than for vitamins A and D3.
    The fact that laboratory animals experienced no apparent nutritional
    deficiencies affecting clinical status or survival following lifetime
    exposure to high levels of -cyclodextrin in the diet also provides
    reassurance that the use of this additive will not adversely affect
    the nutritional status of humans.

    4.  EVALUATION

         The Committee allocated an ADI of 0-5 mg/kg bw for
    -cyclodextrin, based on the NOEL of 1.25% in the diet (equal to
    470 mg/kg bw/day) in the 1-year study in dogs and a safety factor
    of 100.

    5.  REFERENCES

    COMINI, S., OLIVIER, P., RIOTTOT, M., & DUHAMEL, D. (1994).
    Interaction of -cyclodextrin with bile acids and their competition
    with vitamins A and D3 as determined by 1H-NMR spectrometry.
     Clinica Chimica Actam, 228: 181-194.

    GUR, E., NYSKA, A., & WANER, T. (1993a). -cyclodextrin: Oncogenicity
    study in the mouse by dietary administration. LSRI project
    no. CHS/066/BCD. Unpublished report from Life Science Research Israel
    Ltd, Ness Ziona 70 451, Israel. Submitted to WHO by Roquette Frres,
    Lestrem, France.

    GUR, E., NYSKA, A., & WANER, T. (1993b). -cyclodextrin: Oncogenicity
    study in the rat by dietary administration. LSRI project
    no. CHS/065/BCD. Unpublished report from Life Science Research Israel
    Ltd, Ness Ziona 70 451, Israel. Submitted to WHO by Roquette Frres,
    Lestrem, France.

    HRC (1994a). Beta-cyclodextrin: Toxicity to rats by dietary
    administration for 52 weeks. Unpublished report no. ROQ 4/931090 from
    Huntingdon Research Centre Ltd, Huntingdon, Cambridgeshire, UK.
    Submitted to WHO by Roquette Frres, Lestrem, France.

    HRC (1994b). Beta-cyclodextrin: Toxicity to dogs by repeated dietary
    administration for 52 weeks. Unpublished report no. ROQ 3/931848 from
    Huntingdon Research Centre Ltd, Huntingdon, Cambridgeshire, UK.
    Submitted to WHO by Roquette Frres, Lestrem, France.

    Pharmakon Europe (1994). Beta-cyclodextrin: Three generation oral
    (dietary administration) reproduction toxicity study in the rat. Study
    no. 430/006. Unpublished report from Pharmakon Europe, L'Arbresle,
    France. Submitted to WHO by Roquette Frres, Lestrem, France.
    


    See Also:
       Toxicological Abbreviations
       Beta-Cyclodextrin (WHO Food Additives Series 32)
       beta-CYCLODEXTRIN (JECFA Evaluation)