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    ALPHA-METHYLBENZYL ALCOHOL

    First draft prepared by
    Ms E. Vavasour, Toxicological Evaluation Division
    Bureau of Chemical Safety, Food Directorate
    Health and Welfare Canada
    Ottawa, Ontario, Canada

    1  EXPLANATION

         alpha-Methylbenzyl alcohol has not previously been evaluated by
    the Committee.  This substance is used as a flavouring agent in
    foods and beverages; it also occurs naturally in a variety of foods
    at levels of up to 1.3 ppm (Maarse and Visscher 1991).

    2  BIOLOGICAL DATA

    2.1  Biochemical aspects

    2.1.1  Absorption, distribution, and excretion

         Following a single oral dose of 460 mg/kg bw in rabbits,
    alpha-methylbenzyl alcohol was rapidly excreted in the urine.  
    Within 24 hours, 82% of the dose was retrieved as urinary
    metabolites (Smith et al. 1954).

    2.1.2  Biotransformation

         Metabolites in the urine of rabbits following a single oral
    dose of 460 mg/kg bw alpha-methylbenzyl alcohol were
    alpha-methylbenzyl alcohol glucuronide (50%), hippuric acid (30%)
    and mandelic acid (1-2%) (Smith et al. 1954).  These results in the
    rabbit were cited in a 1959 review, with the observation that both
    optical forms of the alcohol behaved similarly with respect to
    biotransformation (Williams, 1959).

         Hopkins  et al. (1972) reported that a small proportion of a
    dose of alpha-methylbenzyl alcohol in rats was excreted in the urine
    as acetophenone.

         Rats displayed substrate stereoselectivity in the metabolism of
    racemic mixtures of alpha-methylbenzyl alcohol.  The R(+) isomer was
    largely excreted as the glucuronide while the S(-) isomer underwent
    further oxidative metabolism (Testa and Jenner 1976).

    2.2  Toxicological studies

    2.2.1  Acute studies

         The results of acute toxicity studies with alpha-methylbenzyl
    alcohol are summarized in Table 1.

    Table 1.  Acute toxicity studies with alpha-methylbenzyl alcohol

                                                               

    Animal      Route        LD50        Reference
                          (mg/kg bw)
                                                               

    Mouse       Oral       1 2501        Dieter 1990
    Rat         Oral         400         Smyth & Carpenter 1944
    Rat         Oral       1 2501        Dieter 1990
                                                               

    1Value is a LDlow

    2.2.2  Short-term toxicity studies

    2.2.2.1  Mice

         Groups of male and female B6C3F1 mice (4 or 5/sex/group), 6-8
    weeks old, received 0, 125, 250, 500, 1 000, or 2 000 mg/kg bw/dy
    alpha-methylbenzyl alcohol in corn oil by gavage, 5 days a week over
    16 days (12 doses).  The animals were observed twice daily and
    weighed on day 1, after one week and at the end of the study.  All
    animals were necropsied.  Histological examinations were performed
    on two males and two females in the 500 mg/kg bw/dy dose group and
    on one male and one female in the control group.  In the 1 000 and
    2 000 mg/kg bw/dy groups, 16/18 mice died within 3 days.  No
    treatment-related histopathological lesions were observed in the
    500 mg/kg bw/dy group (Dieter 1990).

         In a 13-week study, alpha-methylbenzyl alcohol was administered
    by gavage in corn oil to six groups of B6C3F1 mice (10/sex/group)
    at dose levels of 0, 47, 94, 188, 375 or 750 mg/kg bw/dy for 5
    days/week.  Animals were observed twice daily and body weights
    recorded once weekly.  A necropsy was performed on each animal and
    the liver weights were recorded.  Histopathological examinations
    were performed on all animals in the vehicle control groups, all
    animals dying on test and on all male and female mice in the
    750 mg/kg bw/dy group.  No deaths occurred which were related to the
    effects of the test material.  At the two top dose levels, mice
    exhibited laboured breathing, ataxia and lethargy for up to 30
    minutes after dosing.  Body weights were not affected by
    administration of the test material.  Absolute and relative liver
    weights were significantly elevated from controls in both male and
    female groups receiving 187.5 mg/kg bw, but no dose-response
    relationship was evident.  No treatment-related histopathological
    lesions were observed in the 750 mg/kg bw/dy dose group (Dieter
    1990).

    2.2.2.2  Rats

         Groups of male and female F344/N rats (5/sex/group), 6-7 weeks
    old, received 0, 125, 250, 500, 1 000, or 2 000 mg/kg bw/dy
    alpha-methylbenzyl alcohol in corn oil by gavage, 5 days a week over
    16 days (12 doses).  The animals were observed twice daily and
    weighed on day 1, after one week and at the end of the study.  All
    animals were necropsied.  Histological examinations were performed
    on two males and two females in the 1 000 mg/kg bw/dy dose group and
    on one male and one female in the control group.  In the 2 000 mg/kg
    bw/dy group, 2/5 male and 4/5 female rats died before the end of the
    study.  Laboured breathing and lethargy were noted in one rat of
    each sex immediately after dosing in this group.  The body weights
    of male and female rats in the top three dose groups were lower than
    those of the respective vehicle controls.  In the highest dose
    group, body weight decrements from controls were 21% and 15%,

    respectively, for males and females.  Haemorrhage in the
    gastrointestinal tract was observed in one female and two male rats
    in this group.  No treatment-related histopathological lesions were
    observed in the two male and two female rats in the 1 000 mg/kg
    bw/dy group (Dieter 1990).

         alpha-Methylbenzyl alcohol in corn oil was administered by
    gavage to six groups of F344/N rats (10/sex/group) at dose levels of
    0, 93, 187, 375, 750, or 1 500 mg/kg bw, 5 days/week for 13 weeks. 
    Animals were observed twice daily and body weights recorded once
    weekly.  A necropsy was performed on each animal.  The livers were
    weighed at this time.  Histopathological examinations were performed
    on all animals in the vehicle control groups, all animals dying on
    test and on all male and female rats in the 1 500 mg/kg bw/dy group. 
    Histopathological examination of the spleen was carried out in male
    rats in the 375 mg/kg bw/dy group and in male and female rats in the
    750 mg/kg bw/dy group.  In the 1 500 mg/kg bw/dy dose group, 1/10
    male and 3/10 females died as a result of treatment with the test
    material.  Body weights of male rats in the 1 500 mg/kg bw/dy groups
    were significantly decreased at the end of the study compared with
    vehicle controls.  Throughout the study, rats receiving 750 or 1 500
    mg/kg bw/dy exhibited ataxia, rapid breathing and lethargy for up to
    30 minutes after dosing.  Relative liver weights were significantly
    elevated in all the treated female groups and in the 3 top male dose
    groups.  The absolute liver weights were also elevated in comparison
    with the controls in these groups, with statistical significance in
    all but one of the treated female groups (187 mg/kg bw) and none of
    the male groups.  No pathological changes of the liver were noted
    during histopathological examination of the high-dose group.  A
    minimal-to-mild increase in brown pigment, characteristic of
    haemosiderin, was noted in the macrophages of the spleen in 10/10 of
    the male rats receiving 750 mg/kg bw, 9/10 receiving 1 500 mg/kg bw.
    and in none receiving 375 mg/kg bw.  In the female rats, a similar
    pigment was seen in 6/10 receiving 1 500 mg/kg bw. and none
    receiving 750 mg/kg bw/dy (Dieter 1990).

    2.2.3  Long-term toxicity/carcinogenicity studies

    2.2.3.1  Mice

         A carcinogenicity study was conducted with B6C3F1 mice in
    which groups of 49 or 50 mice of each sex were administered 0, 375
    or 750 mg/kg bw/dy of alpha-methylbenzyl alcohol in corn oil by
    gavage, 5 days/week, for 103 weeks.  Mice were observed twice daily. 
    Body weights were recorded weekly for the first 12 weeks and once a
    month for the rest of the study.  Necropsy was performed on all
    animals at termination and those dying on test.  Histopathological
    examination of 33 tissues and organs was conducted in all mice from
    all groups.

         There was a greater than 10% reduction in body weight gain in
    the high-dose males and females, but administration of
    alpha-methylbenzyl alcohol had no effect on survival.  There were no
    increases which were attributable to administration of the test
    material in neoplastic or nonneoplastic lesions of the kidney or any
    of the other tissues.  The NOEL for this study was 375 mg/kg bw/day
    based on the reduction in body weight gain at the next higher dose
    (Dieter 1990).

    2.2.3.2  Rats

         Male and female F344/N rats, 50/sex/group, received 0, 375 or
    750 mg/kg bw/dy alpha-methylbenzyl alcohol in corn oil by gavage 5
    days/week for 103 weeks.  Rats were observed twice daily.  Body
    weights were recorded weekly for the first 12 weeks and once a month
    for the rest of the study.  Necropsy was performed on all animals at
    termination and those dying on test.  Histopathological examination
    of 36 tissues and organs was conducted in all rats from all groups.

         There was a greater than 10% reduction in body weight gains in
    both treated male groups and the high-dose females.  Survival of
    rats in these groups was also significantly reduced.  At study
    termination, male rat survival was 35/50, 8/50 and 1/50 for the
    control low-dose and high-dose groups, respectively;  for the female
    rats, survival was 34/50, 26/50 and 11/50, respectively.  A cluster
    of accidental deaths in both groups of treated males and in the
    high-dose females during weeks 48 to 53 were considered to have been
    gavage-related even though the animals had not been handled
    differently from the other groups and no other predisposing factors
    could be found.

         Chronic progressive nephropathy was a common age-related
    finding in both male and female rats.  Treatment with either dose of
    alpha-methylbenzyl alcohol resulted in an increased severity of this
    lesion in the male, but not the female rats and also contributed to
    lowered survival in these groups.  A statistically significant
    increase was noted in the combined incidence of adenomas and
    adenocarcinomas of the kidney in male rats at both doses (control,
    1/49; low-dose, 13/41; high-dose, 14/28 [the denominator represents
    the number of animals alive at week 81 when the first kidney tumour
    was observed]).  In fact, of these neoplasms, only one
    adenocarcinoma was detected and it was found in a low-dose animal. 
    Because renal tubular cell neoplasms are often late-appearing, and
    were not visible macroscopically at necropsy, multiple sections of
    the kidney were used to evaluate the incidence of these lesions. 
    The incidence of tubular cell hyperplasia in the male rats was 1/49,
    7/41 and 6/28 for the control, low-dose and high-dose animals,
    respectively.  There was no indication that the renal toxicity was
    mediated through alpha2u-globulin, since no compound-related
    increases in hyaline droplet formation were observed in the short-
    term studies described above (data not presented).  [However,

    immunohistochemical analysis for detection of alpha2u-globulin is a
    more sensitive technique to preclude this as a mechanism for the
    genesis of these neoplasms.]  Parathyroid hyperplasia, calcification
    of the heart and glandular stomach, and fibrous osteodystrophy of
    the bone were observed at markedly increased incidences in the low-
    dose male rats.  These changes were believed to be secondary to a
    mineral imbalance caused by renal toxicity.  The incidence of
    hyperplasia of the transitional epithelium overlying the renal
    pelvis also was significantly increased in the low-dose male rats
    (control - 3/50;  low dose - 20/50;  high dose - 4/50;  females:
    1/50, 0/49, 0/50).  Transitional cell papillomas of the urinary
    bladder were observed in 1/47 high-dose male rats and 2/48 high-dose
    female rats and in none of the other groups.  The historical
    incidence for this lesion in both sexes of F344/N rats is 0.2%.  The
    incidences of epithelial hyperplasia of the urinary bladder did not
    show an increase in the treated rats (male: control - 3/48; low dose
    - 4/46; high dose - 1/47;  female: 0/49; 1/47; 0/48).

         The incidence of several other lesions was increased in treated
    groups compared with controls: in males (centrilobular necrosis of
    the liver, inflammation of the forestomach, suppurative inflammation
    of the nasal cavity, acute inflammation of the salivary glands); or
    in both sexes (congestion of the lung, haemorrhage and foreign
    material in the lung).  A no-effect level was not observed in this
    study since reduced survival and body weight gain and an increased
    incidence of kidney adenomas were noted in the male rats at both
    doses administered in this study (Dieter 1990).

    2.2.4  Reproduction Studies

         No information available

    2.2.5  Special studies on teratogenicity

         alpha-Methylbenzyl alcohol was applied dermally to the skin of
    pregnant Sprague-Dawley rats at doses of 0, 0.14, 0.43 and 1.40
    ml/kg bw/dy on days 6-15 of gestation.  With a specific gravity of
    1.01, these doses were equivalent within 1% to the same doses in
    g/kg bw.  At the high dose, the test material exhibited an increased
    incidence of embryolethality, growth retardation and malformations
    (anophthalmia, microphthalmia, ventricular septal defects, defects
    and irregularities affecting the thorax, kinky tail, defects of the
    thoracic ribs and occurrence of cervical ribs) as well as maternal
    toxicity.  The NOEL in this study was 0.43 g/kg bw/dy.  Since only
    summary data from this study were available in preparation of this
    monograph, it was not possible to determine the independence of the
    teratogenic and embryotoxic effects from maternal toxicity (USEPA
    1976, cited Dieter, 1990).

    2.2.6  Special studies on genotoxicity

         The results of genotoxicity studies with alpha-methylbenzyl
    alcohol are summarized in Table 2.

    2.3  Observations in humans

         Contact sensitization tests using 8% alpha-methylbenzyl alcohol
    in petrolatum produced no sensitization reactions in 25 volunteers
    (Kligman 1966, 1973).

         No information was available concerning the effects of
    ingestion of this flavouring in humans.

        Table 2. Results of genotoxicity assays on alpha-methylbenzyl alcohol

                                                                                                                     

    Test System                Test Object               Concentration of       Results          Reference
                                                         alpha-methylbenzyl
                                                         alcohol
                                                                                                                     

    Ames test1                 S. typhimurium            33-6 666 g/ml         Negative         Dieter 1990
                               TA98, TA100,
                               TA1535, TA1537

    DNA damage/growth          E. coli                   50 l/plate            Negative         Fluck et al. 1976
    inhibition

    Yeast mutation             S. sake                   0.20%                  Positive         Kojima et al. 1976

    Mammalian cell             Mouse L5178Y/TK+/-        62.5-1 200 g/ml       Positive         Dieter 1990
    mutation                   lymphoma cells
    (trifluorothymidine
    resistance)

    Chromosomal aberration1    Chinese hamster ovary     1 000-3 000 g/ml      Positive (+S9)   Dieter 1990
                               cells

    Sister chromatid           Chinese hamster ovary     33.3-1 000 g/ml                        Dieter 1990
    exchange1                  cells
                                                                                                                     

    1Both in the presence and absence of metabolic activation.
    
    3  COMMENTS

         Following oral administration this compound is rapidly excreted
    in the urine.  The principal metabolites have been identified as
    hippuric acid and the glucuronide conjugate.

         The Committee reviewed a series of acute, short-term, and long-
    term toxicity studies in which alpha-methylbenzyl alcohol was
    administered in corn oil by gavage in rats and mice.  In addition,
    the Committee evaluated a dermal teratogenicity study in rats and a
    battery of  in vitro genotoxicity tests, which included bacterial
    and mammalian point mutation and mammalian chromosomal aberration
    studies.

         In the short-term toxicity studies, high rates of mortality
    were associated with dose levels of 1 000 mg/kg bw/day and above in
    the mouse and 2 000 mg/kg bw/day in the rat.

         An increase in haemosiderin deposits in macrophages of the
    spleen was noted at 750 mg/kg bw/day in male rats during a 13-week
    study.  This effect was not, however, noted during a 2-year study. 
    A dose-related increase in liver weight that was noted in both male
    and female rats in the 13-week study could not be assessed in the
    2-year study since liver weights were not recorded.  However, an
    increase in the incidence of centrilobular necrosis was observed in
    male rats at both dose levels (375 and 750 mg/kg bw/day) in the
    latter study.

         In the long-term toxicity studies, the body weight gains were
    at least 10% higher in untreated controls in male and female mice
    and in female rats at doses of 750 mg/kg bw/day and in male rats at
    375 and 750 mg/kg bw/day.  Long-term survival of the rats, but not
    the mice, was significantly reduced by oral administration of
    alpha-methylbenzyl alcohol at 375 or 750 mg/kg bw/day.  
    Exacerbation of age-related nephropathy was noted at both dose
    levels in male rats.  The incidence of renal tubular cell
    hyperplasia and adenomas was also statistically-significantly
    increased in these rats than in untreated controls.  A NOEL was not
    observed in this study.

         A teratogenicity study in rats in which alpha-methylbenzyl
    alcohol was administered dermally showed a range of teratogenic and
    embryotoxic effects at the highest dose of 1400 mg/kg bw/day. 
    Maternal toxicity was also demonstrated at this dose.  Because
    alpha-methylbenzyl alcohol was administered dermally, the extent of
    absorption and metabolism were likely to differ from absorption and
    metabolism by the oral route, hence comparability of dose levels was
    uncertain.  The NOEL in this study was 430 mg/kg bw/day.

         alpha-Methylbenzyl alcohol was mutagenic in eukaryotic cells
    (mammalian and yeast), but not in prokaryotic cells (bacteria) and
    induced chromosomal aberrations in mammalian cells  in vitro.

         The Committee noted that alpha-methylbenzyl alcohol
    administered by gavage in corn oil was associated with a higher
    incidence of renal tubular cell adenomas in male rats than in
    untreated contols, but not in female rats or in mice, at dose levels
    at or exceeding the MTD and in the presence of factors that
    exacerbated a high incidence of age-related chronic progressive
    nephropathy.

    4  EVALUATION

         The intake of this compound is extremely low.  On the basis of
    the available evidence, the Committee concluded that the higher
    incidence of benign neoplasms in the kidney of male rats is not
    relevant to humans.  In view of the limited database, the Committee
    concluded that the available data could be used to set an ADI by
    application of a 1 000-fold safety factor to the minimal-effect
    level of 93 mg/kg bw/day with respect to liver weight increase in
    the absence of associated pathology in the 13-week study in rats. 
    On this basis, an ADI of 0-0.1 mg/kg bw/day was allocated for
    alpha-methylbenzyl alcohol.

    5.  REFERENCES

    DIETER, M.P. (1990).  Toxicology and carcinogenesis studies of
    alpha-methylbenzyl alcohol in F344/N rats and B6C3F1 mice (gavage
    studies).  National Toxicology Program (NTP), Technical Report
    Series No. 369, NIH Publication No. 89-2824, Department of Health
    and Human Services, Research Triangle Park, NC 27709, USA.

    FLUCK, E.R., POIRIER, L.A. & RUELIUS, H.W. (1976).  Evaluation of a
    DNA polymerase-deficient mutant of  E. coli for the rapid detection
    of carcinogens.   Chem. Biol. Interact., 15, 219-231. (Cited
    Dieter, 1990).

    HOPKINS, R.P., BORGE, P.A. & CALLAGHAN, P. (1972).  Dehydrogenation
    of DL-methylphenylcarbinol in the rat.   Proc. Biochem. Soc. 127,
    26P-27P.

    KLIGMAN, A.M. (1966).  The identification of contact allergens by
    human assay, III.  The maximization test.  A procedure for screening
    and rating contact sensitizers.   Journal of Investigative
     Dermatology, 47, 393. (Cited Opdyke, 1974).

    KLIGMAN, A.M. (1973).  Report to Research Institute for Fragrance
    Materials.  9 October. (Cited Opdyke, 1974).

    KOJIMA, M., KATOHGI, Y. & HATAE, K. (1976).  Induction of a
    respiration-deficient mutant of  Saccharomyces sake by phenyl
    propanols and related compounds.   Hakko Kogaku Zasshi (J. Ferment.
     Technol.), 54, 11-15. (Cited Dieter, 1990).

    MAARSE, H. & VISSCHER, C.A. (1991).  Volatile Compounds in Food. 
    Qualitative and Quantitative Data.  Supplement 2.  TNO
    Biotechnology and Chemistry Institute.  Zeist, The Netherlands.

    OPDYKE, D.L.J. (1974).  Styrallyl alcohol.   Food and Cosmetics
     Toxicology, 12 (Suppl.), (Special Issue - Monographs on Fragrance
    Raw Materials).  December 1974.

    SMITH, J.N., SMITHIES, R.H. & WILLIAMS, R.T. (1954).  The metabolism
    of alkylbenzenes.  Stereochemical aspects of the biological
    hydroxylation of ethylbenzene to methylphenylcarbinol. 
     Biochemistry, 56, 320-324. (Cited Dieter, 1990).

    SMYTH, H.F. & CARPENTER, C.P. (1944).  The place of the range
    finding test in the industrial toxicology laboratory.   J. Ind. Hyg.
     Toxicol. 26, 269-273. (Cited Dieter, 1990).

    TESTA, B. & JENNER, P. (1976).  Drug Metabolism: Chemical and
    Biochemical Aspects.  New York: Marcel Dekker, Inc., p. 247.
    (Cited Dieter, 1990).

    U.S. Environmental Protection Agency (USEPA) (1976).  Organic
    Compounds Identified in Drinking Water in the United States.  USEPA,
    Cincinnati. (Cited Dieter, 1990).

    WILLIAMS, R.T. (1959).  Detoxication Mechanisms.  The Metabolism
    and Detoxication of Drugs, Toxic Substances and Other Organic
    Compounds, 2nd ed., p. 321.  Chapman & Hall Ltd., London. (Cited
    Opdyke, 1974).


    See Also:
       Toxicological Abbreviations
       alpha-METHYLBENZYL ALCOHOL (JECFA Evaluation)