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    INTERNATIONAL PROGRAMME ON CHEMICAL SAFETY

    WORLD HEALTH ORGANIZATION





    SAFETY EVALUATION OF CERTAIN 
    FOOD ADDITIVES



    WHO FOOD ADDITIVES SERIES: 42





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





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

    SODIUM CARBOXYMETHYL CELLULOSE, ENZYMATICALLY HYDROLYSED

    First draft prepared by
    Dr J.B. Greig
    Department of Health, Skipton House, London, United Kingdom

          Explanation
          Biological data
              Biochemical aspects
                   Absorption, distribution, and excretion
              Toxicological studies
                   Short-term studies of toxicity
          Comments
          Evaluation
          References


    1.  EXPLANATION

         Enzymatically hydrolysed sodium carboxymethyl cellulose is a
    water-soluble dietary fibre with a relative molecular mass range of
    800-10 000 Da, which is lower than that of sodium carboxymethyl
    cellulose itself; for the same concentration, solutions of
    enzymatically hydrolysed material have a lower viscosity than
    carboxymethyl cellulose. It acts mainly as a stabilizer with
    fat-extending properties. It can be used in formulating low-fat and
    reduced-fat foods and soft drinks.

         Enzymatically hydrolysed sodium carboxymethyl cellulose is
    prepared from regular, food-grade carboxymethyl cellulose by partial
    enzymic hydrolysis under mildly acidic conditions with a food-grade
    cellulase enzyme from  Trichoderma longibrachiatum. Carboxymethyl
    cellulose itself was allocated an ADI 'not specified' at the
    thirty-fifth meeting of the Committee (Annex 1, reference 88). The
    cellulase enzyme was allocated an ADI 'not specified' at the
    thirty-ninth meeting (Annex 1, reference 101). Enzymatically
    hydrolysed sodium carboxymethyl cellulose has not been evaluated
    previously by the Committee.

         At the present meeting the Committee reviewed two studies in
    which the properties of enzymatically hydrolysed sodium carboxymethyl
    cellulose were compared with those of the parent material,
    carboxymethyl cellulose. 


    2.  BIOLOGICAL DATA

    2.1  Biochemical aspects

    2.1.1  Absorption, distribution, and excretion

         The absorption and excretion of enzymatically hydrolysed sodium
    carboxymethyl cellulose and carboxymethyl cellulose, each

    radiolabelled with 14C in the carboxymethyl group, in rats were
    compared over five days. The distribution of residual radioactivity in
    the body was then determined. The radiolabelled materials were diluted
    with 'Solka Floc' carboxymethyl cellulose or carboxymethyl cellulose
    hydrolysate to provide materials with appropriate specific activity.
    After a pilot study, two groups of four male and four female
    conventional Wistar Wag/Rij rats were adapted for 14 days to diets
    containing 5% of either material. The rats then received the
    appropriate radiolabelled material at a dose of about 500 mg/kg bw,
    equivalent to approximately 10 µCi per rat. Urine and faeces were
    collected at intervals up to 120 h, and expired carbon dioxide was
    collected at intervals up to 48 h. After 120 h, the rats were killed,
    and 21 organs or tissues were collected, with the contents of the
    gastrointestinal tract and the residual carcass; the blood was
    separated into plasma and cells. 

         Little difference was seen between the two compounds in the
    quantity of radiolabel in excreta: faecal excretion accounted for most
    of the admini- stered dose (range of group means, 90-99%), with lesser
    amounts in urine (1.3-2.0%) and expired air (0.6-0.9%). Peak
    expiration of 14C-carbon dioxide occurred within the first 2 h after
    dosing; the authors suggest that this was due to degradation of
    low-molecular-mass compounds. Slightly more radiolabel was retained in
    the carcasses of animals that received enzymatically hydrolysed
    material than in those given carboxymethyl cellulose. A similar effect
    was seen in individual organs, tissues, and body fluids, the largest
    amounts being detected in fat, skin, muscle, liver, small intestine,
    blood cells, and plasma. The tissue concentrations of both
    enzymatically hydrolysed and parent carboxymethyl cellulose were
    slightly higher in the liver and adrenals than in other tissues.

         The relative molecular mass distribution of radiolabelled faecal
    extracts was compared with that of the corresponding dosing solution
    by gel permeation chromatography. The relative molecular masses of the
    main peaks in the dosing solution differed from those in the faecal
    extracts from animals treated with carboxymethyl cellulose but not for
    animals treated with enzymatically hydrolysed material. The authors
    concluded that the carboxymethyl cellulose was partially degraded
    during passage through the gastrointestinal tract, resulting in the
    formation of fragments similar in size to the preparation of
    enzymatically hydrolysed sodium carboxymethyl cellulose. They were
    unable to exclude the possibility that some labelled macromolecules
    had been persorbed (van Ommen, 1993; Bär et al., 1995a).

    2.2  Toxicological studies

    2.2.1  Short-term studies of toxicity

     Rats

         Groups of 20 male and 20 female Crl:WI(WU)BR rats, about eight
    weeks old, were fed either basal diet containing 10% added wheat
    starch (controls) or diets containing 2.5, 5, or 10% enzymatically

    hydrolysed sodium carboxymethyl cellulose or carboxymethyl cellulose
    in place of the equivalent amount of wheat starch. The diets were
    administered for 91-95 days to males and 98-102 days to females.
    Ophthalmoscopic analyses were conducted in week 13. Haematological
    examination was carried out on blood from 10 rats of each sex in each
    group on day 85 (males) or 89 (females). Nineteen clinical chemical
    parameters were analysed, including glucose levels on day 88 for males
    and day 95 for females, while the other measurements were made at
    necropsy.

         One treated female rat died during the course of the experiment
    from causes unassociated with administration of the diet. Rats fed 10%
    of either material had diarrhoea but were otherwise healthy. At the
    end of the experiment, the mean body weight of female rats fed the 10%
    diets was significantly lower than that of the control group.
    Dose-related increases in water intake were seen in weeks 4, 8, and
    11. The food intake of animals at the higher doses tended to be
    greater than that of rats at lower doses, although the conversion
    efficiency of animals at 10% was only slightly lower. The mean food
    intake of males and females given 10% carboxymethyl cellulose were 19
    and 14 g per rat per day, equal to 6200 and 6800 mg/kg bw per day; the
    corresponding figures for males and females given 10% enzymatically
    hydrolysed material were 19 and 14 g per rat per day, equal to 5900
    and 6600 mg/kg bw per day. 

         Ophthalmoscopic examination showed no treatment-related changes
    in rats at the high doses. The only significant changes seen on
    haematological examination were slight reductions in haemoglobin
    concentration and haematocrit in females fed 10% enzymatically
    hydrolysed sodium carboxymethyl cellulose. Clinical chemistry showed
    some sporadic increases and a suggestion of a dose-related increase in
    alkaline phosphatase and alanine aminotransferase activity with both
    enzymatically hydrolysed and parent carboxymethyl cellulose, which was
    more obvious in male rats. Urinary excretion of cations, particularly
    sodium, and of citrate was altered in animals of each sex fed either
    preparation and was associated with increased urine volume and pH and
    with decreased urine density. Semiquantitative observations and
    microscopic findings in the urine were unaffected in any manner
    consistent with compound- or dose-related toxicity. The absolute and
    relative weights of both the full and empty caecum, noted as caecal
    enlargement at autopsy, were significantly increased in all treated
    animals, with evidence of a dose-response relationship. The Committee
    accepted that these changes are consistent with the incorporation in
    the diet of an indigestible fibre which has a bulking effect (Bär,
    1997; Newberne et al., 1988).

         The observations in sections of the gastrointestinal tract taken
     post mortem were consistent with the increased fluid intake of
    animals at the high dose and were accompanied by histopathological
    changes. The relative kidney weights were significantly higher in all
    animals at the high dose, again with a clear dose-response
    relationship. Statistically significant histopathological changes in
    the kidney included papillomatous urothelial hyperplasia and pelvic

    mineralization, generally graded only slight or very slight, in males
    treated with the highest dose of enzymatically hydrolysed sodium
    carboxymethyl cellulose and those given the lowest dose of
    carboxymethyl cellulose. The incidence of simple and papillary
    epithelial hyperplasia in the urinary bladder, generally only of
    slight grade, was higher in males than in females and in animals
    receiving the 10% dose of either material  (Bär et al., 1995b; Til,
    1992). These changes were attributed to an effect of the concentration
    of sodium in the diet, which was about fourfold higher than in the
    basal diet (Lalich et al., 1974; Bär, 1997).


    3.  COMMENTS

         The Committee reviewed two studies in which the properties of
    enzymatically hydrolysed sodium carboxymethyl cellulose were compared
    with those of the parent material, carboxymethyl cellulose. The first
    was a 90-day study of toxicity in which male and female rats received
    diets containing 0, 2.5, 5, or 10% of either material. The second was
    a comparative study of absorption, distribution, and excretion of
    these two materials in rats.

         Histopathological changes in the kidney and urinary bladder were
    reported in the 90-day study in rats receiving either enzymatically
    hydrolysed carboxymethyl cellulose or parent compound, which were
    associated with changes in kidney weight, increased urine volume, and
    increased excretion of some ionic species in the urine. Caecal
    enlargement was also reported. For comparison, the Committee
    considered the studies of the toxicity of carboxymethyl cellulose that
    had been reviewed at its thirty-fifth meeting. It noted that, in early
    studies that appeared to be comparable with regard to species, dietary
    concentration of carboxymethyl cellulose, and duration of exposure, no
    changes in organ weight or histopathological appearance were found
    that were comparable to those reported from the 90-day study reviewed
    at the present meeting.

         Since the morphological changes in the kidney occurred in males
    receiving the high dose of enzymatically hydrolysed carboxymethyl
    cellulose and the low dose of carboxymethyl cellulose, the Committee
    was not convinced that the response was of toxicological concern. The
    observed epithelial hyperplasia of the urinary bladder was generally
    graded slight or very slight and occurred in male rats fed a diet
    containing 10% of either carboxymethyl cellulose. The Committee
    concluded that this represents a response to a high intake of sodium
    ions by rats of the strain used in the new study. Administration of
    additional sodium (approximately four times that available in the diet
    of the control group) could have led to the observed histopathological
    changes and changes in urinary excretion. Also, the caecal
    enlargement, diarrhoea, and other secondary changes were considered to
    be the consequence of the accumulation of poorly absorbed,
    water-soluble material in the caecum and colon and to be of no
    toxicological significance. The NOEL for enzymatically hydrolysed
    carboxymethyl cellulose was equal to 6000 mg/kg bw per day.

         The metabolic study showed that passage of carboxymethyl
    cellulose through the gastrointestinal tract results in partial
    breakdown of the bonds between monomeric units, to result in a
    material with a relative molecular mass equivalent to that of
    enzymatically hydrolysed carboxymethyl cellulose. The amounts of
    radiolabel excreted in the faeces, urine, and breath after
    administration of radiolabelled versions of the two products were
    quantitatively almost identical, providing additional evidence that
    the end-products of digestion of carboxymethyl cellulose and
    enzymatically hydrolysed carboxymethyl cellulose are similar in rats.


    4.  EVALUATION

         The Committee concluded that these similarities are consistent
    with no toxicologically significant difference between carboxymethyl
    cellulose and enzymatically hydrolysed carboxymethyl cellulose.
    Therefore, the Committee included the latter in the group ADI 'not
    specified' with ethyl cellulose, ethyl hydroxyethyl cellulose,
    hydroxypropyl cellulose, hydroxypropyl methyl cellulose, methyl
    cellulose, methyl ethyl cellulose, and sodium carboxymethyl cellulose.


    5.  REFERENCES

    Bär, A., van Ommen, B. & Timonen, M. (1995a) Metabolic disposition in
    rats of regular and enzymatically depolymerized sodium
    carboxymethylcellulose.  Food Chem. Toxicol., 33,  901-907. 

    Bär, A., Til, H.P. & Timonen, M. (1995b) Subchronic oral toxicity
    study with regular and enzymatically depolymerized sodium
    carboxymethyl cellulose in rats.  Food Chem. Toxicol., 33,  909-917. 

    Bär, A. (1997) Sodium carboxymethyl cellulose, enzymatically
    hydrolyzed (CMC-ENZ). Unpublished dossier for the safety assessment of
    CMC-ENZ from Bioresco Ltd, Binningen, Switzerland. Submitted to WHO by
    Bioresco Ltd, Binningen, Switzerland.

    Lalich, J.J., Paik, W.C.W. & Pradhan, B. (1974) Epithelial hyperplasia
    in the renal papilla of rats: Induction in animals fed excess sodium
    chloride.  Arch. Pathol., 97,  29-32. 

    Newberne, P.M., Conner, M.W. & Estes, P. (1988) The influence of food
    additives and related materials on lower bowel structure and function.
     Toxicol. Pathol., 16,  184-197. 

    van Ommen, B. (1993) A comparative disposition study with
    [14C]-CMC-hydrolysate and [14C]-CMC in rats. Unpublished report No. V
    92.473 from TNO Nutrition and Food Research, Zeist, Netherlands.
    Submitted to WHO by Bioresco Ltd, Binningen, Switzerland.

    Til, H.P. (1992) Comparative sub-chronic (3-month) feeding study with
    sodium carboxymethyl cellulose hydrolysate and sodium carboxymethyl
    cellulose in rats. Unpublished report No. V 92.015 from TNO Nutrition
    and Food Research, Zeist, Netherlands. Submitted to WHO by Bioresco
    Ltd, Binningen, Switzerland.
    


    See Also:
       Toxicological Abbreviations