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    CHLORAMPHENICOL

    EXPLANATION

         Chloramphenicol is an antibiotic originally isolated from the
    soil bacterium Streptomyces venezuelae. It has the following
    chemical structure:

    CHEMICAL STRUCTURE 1

         Chloramphenicol was previously considered at the twelfth meeting
    of the Committee (Annex 1, reference 17), at which time it was
    determined that acceptable levels in food could not be established.

    BIOLOGICAL DATA

    Biochemical aspects

    Absorption, distribution, and excretion

         In dogs, chloramphenicol is rapidly and extensively absorbed
    after oral administration of 50 mg/kg b.w., giving plasma levels of
    16.5 g/l 2 hours after dosing (Watson, 1972; Watson 1977a). Similar
    findings have been observed in rabbits given oral doses of 16 mg/kg
    chloramphenicol (Cid  et al., 1983). Dietary bran markedly 
    increased the rate and extent of absorption of orally administered
    chloramphenicol palmitate in pigs, but it is not known if the
    absorption of chloramphenicol itself would be affected in this way
    (Bueno  et al., 1984). Experiments with everted mouse small intestine
    sacs resulted in absorption of chloramphenicol at similar rates over
    different regions of the small intestine. Various chemicals and
    metallic ions had no effect on the rate of absorption, and penetration
    of the serosal and mucosal surfaces was similar indicating passive
    absorption of chloramphenicol from the gut (Chakrabarti & Banerjee,
    1977).

         In adult human beings, absorption of chloramphenicol is rapid and
    extensive after an oral dose. Serum levels were 20 - 40 mg/l after a
    2 g dose (29 mg/kg b.w.) and 40 - 60 mg/l after a 4 g dose (57 mg/kg
    b.w.) (Yunis, 1973a).

         Chloramphenicol is also well absorbed by infants and neonates
    after oral administration. Serum (peak) concentrations of 20 - 24 mg/l
    were noted after oral doses of 40 mg/kg b.w. in neonates. Infants
    given 26 mg/kg b.w. were found to have peak concentrations of 14 mg/l
    (Mulhall  et al., 1983). Available evidence and theoretical
    considerations suggest that chloramphenicol may be percutaneously
    absorbed in human beings (Guy  et al., 1985a, b).

         Chloramphenicol is distributed to most major organs in newborn
    pigs. After i.v. administration of 0.52 mg/kg b.w. 14C-chloram-
    phenicol, many tissues showed higher levels of chloramphenicol than
    were found in the blood as soon as 5 minutes after administration.
    These tissues included the lungs, liver, kidneys, adrenal cortex,
    myocardium, pancreas, thyroid, spleen, and skeletal muscle. The
    levels remained higher than in blood for up to 8 hours after dosing.
    At 4 and 8 hours after administration, levels in the brain were
    higher than in blood. However, there was no apparent affinity for
    chloramphenicol in the bone marrow over the period of the experiment
    (8 hours), where levels did not reach those noted in serum (Appelgren
     et al., 1982).

         Chloramphenicol in human beings, regardless of the route of
    administration, extensively distributes, although the levels in
    tissues depend on the route, the levels being the highest after oral
    or i.v. administration. The compound has been found in the heart,
    lungs, kidneys, liver, spleen, pleural fluid, seminal fluid, ascitic
    fluid, and saliva. (Ambrose, 1984; Yunis 1973a; Gray, 1955). It binds
    extensively to proteins in both adults and neonates, although binding
    in the latter is less than in the former (Kurz  et al., 1977).

         Chloramphenicol can cross the placenta in human beings (Ambrose,
    1984). After oral doses of 1 or 2 g chloramphenicol to pregnant women,
    chloramphenicol was detected in the placenta after 1.5 - 2.5 hours,
    indicating some potential for the drug to reach the fetus (Ross
     et al., 1950).

         In human beings with normal renal/hepatic function the volume of
    distribution is around 0.7 1.4 1/kg. These values do not deviate
    markedly in patients with hepatic dysfunction or with renal
    impairment. Overall, these values suggest extensive distribution in
    body tissues. (Ambrose, 1984; Burke  et al., 1980; Slaughter
     et al., 1980). Similar values were noted with infants and young
    children given the sodium succinate derivative of chloramphenicol,
    which is converted  in vivo to chloramphenicol (Sack  et al., 1980).

         Chloramphenicol and its metabolites are excreted in the urine of
    rats after oral dosing; up to 70% of an oral dose may he excreted in
    this way (Glazko  et al., 1949; Glazko  et al., 1952). Limited data
    suggest that chloramphenicol may be excreted in the bile of rats
    following administration; around 0.4% of an i.m. dose of 40 mg/kg was
    detected in the bile after 4 hours (Kunii  et al., 1983). There are
    no readily available data for biliary excretion after oral
    administration. In new-born pigs, the majority of an i.v. dose of
    chloramphenicol was excreted in the urine, while a small quantity was
    excreted in the bile (Appelgren  et al., 1982). Liver damage may
    delay body-clearance of chloramphenicol, at least in the mini-pig
    (Kroker, 1985). Following t.v. administration to goats, 69% of the
    dose was excreted in the urine after 12 hours (Javed  et al., 1984).

         Chloramphenicol has been found to be excreted in other body
    fluids after administration to animals. It has been detected at levels
    up to 6 mg/l in the tears of cattle given i.v. doses of 50 mg/kg
    chloramphenicol; it was detected in the tears as soon as 1 hour after
    dosing (Punch  et al., 1985). Chloramphenicol was detected in the
    milk of goats after an i.v. dose of 100 mg/kg, with maximum levels
    1 hour after administration. (Roy  et al., 1986). Administration of
    10 mg/kg chloramphenicol i.m. to cattle resulted in peak levels in
    milk of around 1 mg/l 6 hours after injection. However, after oral
    administration, no chloramphenicol was detected in milk.
    (De Corte-Baeten & Debackere, 1976).

         Nine patients with evidence of chloramphenicol-induced bone
    marrow suppression showed greatly reduced plasma clearance times
    compared with 9 others who showed no evidence of toxicity. In the
    "toxic" group, 5 had liver disease, 2 had pyelonephritis, and 2 showed
    neither liver disease nor renal disease. In the "non-toxic" group, 1
    had liver disease, 2 had renal disease, while 6 had neither. Six hours
    after an i.v. dose of 500 mg chloramphenicol succinate, the blood
    level in the "toxic" group was 4.5 µg/ml (2.8 - 6.9 µg/ml), while in
    the "non-toxic" group the mean level was 1.2 µg/ml (0 - 2.3 ug/ml).
    Similarly, after 8 hours the mean level in the "toxic" group was
    3.5 µg/ml (2.1 - 5.2 ug/ml), while in the "non-toxic" group a mean
    level of 0.7 µg/ml (0 - 2.5 µg/ml) was noted. Such findings suggest
    that patients susceptible to the bone-marrow effects of
    chloramphenicol may clear the drug from the blood at a slower rate
    than those who are not susceptible (Shurland & Weisburger, 1969).

         Chloramphenicol administered to human beings is excreted
    primarily in the urine (90%), up to 15% as the parent compound and the
    remainder as metabolites, including conjugated derivatives
    (Burke  et al., 1980; Ambrose, 1984; Yunis, 1973a). Glomerular
    excretion is thought to be the major mechanism of excretion
    (Glazko  et al., 1949).

         Renal clearance varies depending on age. In one study, clearance
    in neonates (less than 6 months of age) was 0.46 - 9.76 1/hour, while
    in infants aged 6 months to 2.5 years values in the range 1.8 - 2.1
    1/hour were reported. In two subjects aged over 2.5 years, values of
    6.03 and 9.59 1/hour were noted (Burckart  et al., 1983). Similar
    variations with age have been noted in other studies (Mulhall  et al.,
    1983; Sack  et al., 1980; Kauffman  et al., 1981; Burckart  et al.,
    1982; Rajchgot  et al., 1983). Renal clearance takes longer in
    patients with renal insufficiency than in normal patients (Brasfield
     et al., 1983). However, these differences are not marked (Smith &
    Weber, 1983). It has been recommended that dosage adjustments need not
    be made in patients with renal insufficiency or in anephric subjects
    (Van Scoy & Wilson, 1983).

         As with experimental animals, chloramphenicol is excreted in
    human milk. Up to 1.3% of an administered dose may be excreted in this
    way. (Vorherr, 1974). After a single, oral dose of chloramphenicol, a
    peak milk concentration of 3 mg/l has been reported. This peak was
    reached around 2 hours after administration, with a drop to nearly
    pre-dosing levels by 8 hours post-dose (Plomp  et al., 1983). Other
    workers have reported similar findings (Knowles, 1965; Matsuda, 1984).

    Biotransformation

         Early studies indicate that the major metabolite of
    chloramphenicol in the rat was the glucoronide conjugate, which was
    found along with free chloramphenicol after oral dosing
    (Glazko  et al., 1950; Glazko  et al., 1952). In  in vitro studies
    it has been demonstrated that the glucuronide is the main metabolic
    product in isolated rat hepatocytes exposed to chloramphenicol
    (Siliciano  et al., 1978). Glucuronidation of chloramphenicol was
    elevated  in vitro in hepatocytes obtained from phenobarbital-
    pretreated rats. This increase correlated with differential induction
    of UDP-glucuronyltransferase in hepatocytes of rats pretreated with
    phenobarbital (Ullrich & Bock, 1984).

         Several metabolites of chloramphenicol have been identified in
    rat urine. In addition to free chloramphenicol (1) and the glucoronide
    (2), the oxamic acid (3), alcohol (4), and base (5) derivatives have
    been noted in the urine of rats given i.m. doses of [3H]-chlo-
    ramphenicol (1R,2R-1- p-nitrophenyl(2,2-dichloroacetamido-
    1,3-(1-3H)-propandediol). The acetylarylamine (6) and arylamine
    metabolites (7) have also been detected. These metabolites are shown
    in Table 1.

    Table 1  Metabolites of chloramphenicol in the rat
                                                                        

         Compound                      R1           R2            R3
                                                                        

    1. chloramphenicol                 NO2          COCHCl2       OH

    2. glucuronide conjugate           NO2          COCHCl2       C6H9O7

    3. oxamic acid derivative          NO2          COCHO2H       OH

    4. alcohol derivative              NO2          COCH2OH       OH

    5. base derivative                 NO2          H             OH

    6. acetylarylamine derivative      NHCOCH3      COCHCl2       OH

    7. arylamine derivative            NH2          COCHCl2       OH
                                                                        

         Based upon recovered radioactivity, the major metabolites
    appeared to be chloramphenicol base (approx. 26%) and the
    acetylarylamine derivative (19.1%). The other metabolites were
    recovered in the 8-15% range, except for the arylamine derivative,

    which represented approximately 4% of the recovered radioactivity
    (total identified, 93.4% of administered radioactivity; total
    collected, 95.9% of administered radioactivity (Bories  et al.,
    1983).

         An  in vitro study using perfused rat liver and rat liver
    microsomes indicated that the arylamine derivative may undergo
    N-oxidation to form nitrosochloramphenicol by way of the N-hydroxy
    derivative, which may then be conjugated with glutathione
    (Ascherl  et al., 1985).

         There are few data available on the metabolism of chloramphenicol
    in other species. In dogs, chloramphenicol, chloramphenicol base, and
    chloramphenicol glucuronide conjugate appeared to be the major
    metabolites (Glazko  et al., 1950). Chloramphenicol, the glucuronide
    conjugate, and the oxamic acid, acetylarylamine, arylamine, and base
    derivatives were noted in the urine of goats given i.m. injections of
    chloramphenicol (Bories  et al., 1983).  In vitro studies with pig
    liver showed the activity of UDP-glucuronyltransferase in this species
    to be similar to that in rats, suggesting that glucuronidation may be
    a significant pathway of biotransformation of chloramphenicol in pigs.
    The same experiments with sheep and cattle liver preparations showed
    that they have much lower glucuronyl transfer activities (25% and 14%,
    respectively). This may indicate that glucuronidation plays a less
    important role in these species (Smith  et al., 1984).

         In human beings, 93% of an oral dose of chloramphenicol was
    excreted in the urine within 24 hours, and it seems likely that the
    major excretion product was the glucuronide conjugate. Approximately
    48% of the chloramphenicol excreted in the urine within 8 hours after
    oral dosing was the glucuronide conjugate; only 6% was excreted as the
    parent compound and 4% as the base derivative (Baselt, 1982;
    Nakagawa  et al., 1975). The alcohol derivative has been detected in
    the urine of neonates (Dill  et al., 1960). More recent experiments
    have confirmed the presence of the glucuronide conjugate and base
    derivative as the major metabolites after an oral dose of 500 mg
    chloramphenicol (Bories  et al., 1983).

         Human liver has the potential to reduce chloramphenicol. In 10
    livers studied, nitro reductase activity was observed, which was
    dependent on NADPH. Thus, human livers may have the capacity to
    convert the nitro group of chloramphenicol to the amine, with the
    further possibility of nitroso formation. Esters of chloramphenicol,
    for example, the succinate, are converted to chloramphenicol  in vivo
    (Salem et  al., 1981).

    Effects on enzymes and other biochemical parameters

         Chloramphenicol is known to increase the plasma levels of certain
    drugs (e.g., paracetamol and phenytoin) and to prolong barbiturate
    sleeping time, which is indicative of an effect on drug metabolizing
    enzymes (Halpert & Neal, 1981; Nair  et al., 1981; Aravindaksham &
    Cherian, 1984).

         The major contribution to these effects may arise from
    chloramphenicol's ability to act as a suicide substrate in the
    inactivation of cytochrome P-450, possibly by the binding of the
    oxamic acid derivative to lysine residues of the cytochrome molecule
    (Halpert & Neal, 1981; Halpert, 1981; Halpert, 1982). Of the various
    isozymes of cytochrome P-450, those induced by phenobarbital appear to
    be the most sensitive to chloramphenicol. Rats were pretreated with
    various inducers of cytochrome P-450 (phenobarbital, ß-naphthoflavone,
    pregnenolone, 16-x-carbonitrile, and clofibrate) and were then
    injected i.p. with 300 mg/kg b.w. chloramphenicol. The isozymes of
    cytochrome P-450 induced by phenobarbital were inhibited to some
    degree by chloramphenicol administration, but those forms induced by
    the other compounds investigated were not affected. Cytochrome
    P-450-c, the form present in non-pretreated rats, was the most
    susceptible isozyme  in vivo and  in vitro (Halpert  et al., 1985).

         Following i.v. or i.p. administration of 100 mg/kg b.w.
    chloramphenicol to rats, inhibition of the conversion of n-hexane to
    2-hexanol by rat liver or lung microsomes occurred. There was no
    effect on the conversion of n-hexane to l-hexanol, while only liver
    microsomes were markedly inhibited in the formation of 3-hexanol
    (Naesland & Halpert, 1984; Naesland  et al., 1983).

         Chloramphenicol prevented the methanol-potentiated toxicity of
    carbon tetrachloride in rats, possibly by deactivation of cytochrome
    P-450 (Brabec  et al., 1982). Phenobarbital-induced cytochrome P-450
    appears to be responsible for the dechlorination of chloramphenicol
    and the formation of chloramphenicol aldehyde in rat liver microsomes.
    The dechlorination may be enhanced by glutathione. The significance of
    these findings is not clear at the present time (Morris  et al.,
    1982; Morris  et al., 1983).

         Subcutaneous administration of up to 100 mg/kg b.w. chloram-
    phenicol to rats inhibited hepatic N-demethylase, glucose-6-
    phosphate dehydrogenase, and carboxylesterase (Hapke  et al., 1977).
    However, no effects were observed on monoamine oxidase in the liver,
    brain, or heart of rabbits given 60 mg/kg b.w. chloramphenicol i.m.
    for 5 days. Similarly, no effect on monoamine oxidase activity was
    seen in  in vitro experiments where rabbit liver preparations were
    preincubated with chloramphenicol (Ali, 1985).

         Several studies have demonstrated an effect of chloramphenicol on
    mitrochondrial protein synthesis.  In vitro, chloramphenicol
    inhibited mitochondrial protein synthesis in rat liver and rabbit bone
    marrow. The effect was similar to that noted with tetracycline (Summ
     et al., 1976). Nitrosochloramphenicol inhibited rat mitochondrial
    DNA polymerase  in vitro, whereas the arylamine derivative and
    chloramphenicol itself did not (Lim  et al., 1984). Nitroschlor-
    amphenicol inhibited the transport of NAD-linked substrates into
    mitochondria, but it had no effect on FAD-linked substrates. It
    inhibited ATP formation and completely blocked the transport of
    protons out of the mitochondria (Abou-Khalil  et al., 1982).
    Chloramphenicol inhibited the incorporation of leucine into protein in
    mitochondria from erythroid and myeloid rumour cells and in cells from
    normal, myeloid, and erythroid hyperplastic bone marrow of rabbits
     in vitro (Abou-Khalil  et al., 1980, 1981). In mice given
    chloramphenicol, the so-called megamitochondria that were produced
    were deficient in cytochrome oxidase, ATP synthetase, and cytochrome b
    activities, which is indicative of inhibited protein synthesis
    (Wagner & Rafel, 1977). Subcutaneous administration of 500 mg/kg b.w.
    chloramphenicol succinate to partially hepatectomized rats did not
    significantly inhibit protein synthesis. However, liver cytochrome c
    oxidase was strongly inhibited in these rats (Kroon & Vries, 1969).

         Intramuscular administration of chloramphenicol to rats inhibited
    mitochondrial monoamine oxidase activity in the liver, brain, and
    kidney (Banerjee & Basu, 1978). When chloramphenicol was given daily
    for 6 days at doses of 100 mg/kg b.w. i.p., marked reductions in the
    activities of liver kynurenine hydrolase, knynurenine amino-
    transferase, ß-glucuronidase, and acid ribonuclease were observed,
    indicating possible effects on drug metabolizing enzymes and on
    protein biosynthesis. Pyridoxal phosphokinase activity was increased
    (Akhnoukh  et al., 1982).

         In human blood, nitrosochloramphenicol is bound to albumin
     in vitro. In red blood cells, it rapidly forms adducts with
    glutathione and also with the -SH groups of haemoglobin. Moreover, it
    is rapidly reduced to the N-hydroxy compound, although further
    reduction to the amine is very slow. Overall, the data suggest that
    the nitroso derivative would be rapidly detoxified in the blood before
    reaching the bone marrow (Eyer  et al., 1984). Nitrosochloramphenicol,
    but not chloramphenicol, induced methaemoglobinaemia in haemolyzed
    human blood  in vitro (Lim & Yunis, 1982).

         To summarize, chloramphenicol is well absorbed in both animals
    and human beings after oral administration and widespread distribution
    occurs. Urinary excretion is rapid in animals and man, a major
    metabolite being the glucuronide conjugate. Animals and human beings

    produce a range of urinary metabolites;  in vitro experiments suggest
    that nitrosochloramphenicol may be an important metabolite in human
    beings. It is not known if this compound is produced  in vivo in man
    or other species.

    Toxicological studies

    Special studies on carcinogenicity

         In 1982 and 1987, IARC concluded that adequate tests for
    carcinogenicity of chloramphenicol in animals were not available
    (IARC, 1982; IARC, 1987). These conclusions confirmed an earlier
    opinion by IARC workers (Tomatis  et al., 1978). The two studies
    below by Robin  et al., and by Sanguineti  et al., were included in
    the IARC evaluations.

         Four groups of Balb/c × AF1 male mice, 45 per group, were given
    i.p. injections of either 0.5 mg busulphan on days 1, 15, 29 and 43
    (2 groups) or vehicle (acetone plus distilled water, 2 groups. After
    20 weeks, by which time 78 mice remained alive in the busulphan-
    treated groups and 88 in the vehicle groups (the rest having
    died of injection complications), one of the busulphan groups and one
    of the vehicle groups was selected for treatment with chloramphenicol,
    while the others served as controls and were given vehicle only (0.9%
    sodium chloride). Mice in the treatment groups received i.p.
    injections of 2.5 mg chloramphenicol 5 days per week for 5 weeks.
    Sacrifice of the mice followed a complicated schedule depending on the
    appearance of lymphomas, but all animals had been sacrificed by day
    350. The following incidences of lymphoma were noted: busulphan/
    chloramphenicol, 13/37; busulphan/vehicle, 4/35; chloramphenicol/
    vehicle 2/42; vehicle/vehicle, 0/41. The authors thought that this
    suggested that busulphan and chloramphenicol increased the frequency
    and accelerated the onset of lymphomas. It also provided some
    evidence that chloramphenicol alone might induce lymphomas in this
    animal model, but the duration of the experiment along with other
    limitations of the experimental design, particularly the dosing
    regime, prevent any other conclusions from being drawn (Robin
     et al., 1981).

         In a study which was reported in abstract form only, in which
    chloramphenicol was administered in the drinking water, an increased
    incidence of lymphomas in two strains of mice and of hepatocellular
    carcinoma in one strain were noted (Sanguineti  et al., 1983).

         Another study investigated the effect of chloramphenicol on
    cirrhosis and hepatocellular carcinoma induced by the carcinogen
    N-2-fluorenyldiacetamide in rats. A group of 25 male Wistar rats was
    given a diet containing 0.05% N-2-fluorenyldiacetamide (equivalent to
    25 mg/kg b.w./day) plus 2% chloramphenicol (equivalent to 1000 mg/kg
    b.w./day) for 4 weeks. Another group of 20 rats received a diet

    containing only the 0.05% N-2-fluorenyldiacetamide. A "rest" week was
    then followed by another 4 weeks on the respective diets, followed by
    another "rest" week and then 6 weeks of dietary administration of the
    substances. After this the animals given the carcinogen only were
    returned to the control diet, but those given the combined regime were
    continued on this diet for another 2 weeks. Animals were sacrificed at
    46 (carcinogen only) or 46-55 weeks (carcinogen plus chloramphenicol).
    Of 12 animals examined which were given the carcinogen-only diet, 100%
    had cirrhosis and 75% had hepatocellular carcinoma, whereas of 22
    animals examined which were given the combination treatment, only one
    had cirrhosis and hepatic rumours. Thus, chloramphenicol had a
    protective effect on the induction of hepatic rumours by
    N-2-fluorenyldiacetamide (Puron & Firminger, 1965).

    Special studies on haematological effects

         Groups of 18-21 57B1/10ScSnPh mice were given 4.78 Gy of
    X-irradiation and were then treated three times daily with 160 or
    320 mg/kg b.w. chloramphenicol succinate by s.c. injection over 3 or 5
    days, beginning 10 days after the X-ray treatment. Two groups of
    control animals were given chloramphenicol and no irradiation or
    irradiation and no chloramphenicol. Animals were examined 4, 8, and 21
    days after initiation of chloramphenicol treatment (14, 18, and 21
    days after irradiation). No effects on the level of erythrocytes in
    non-irradiated mice occurred; those given chloramphenicol showed
    similar values to non-treated controls. The level of erythrocytes in
    irradiated animals was significantly lower than in non-irradiated
    animals (30% reduction 14 days after irradiation) but there was
    improvement with time (26% reduction 18 days and 15% reduction 21 days
    after irradiation), while irradiated animals given chloramphenicol
    showed lower erythrocyte levels than those that were irradiated only;
    at 14, 18, and 21 days post irradiation, irradiated mice given
    chloramphenicol had erythrocyte levels 8%, 17%, and 4.5% lower,
    respectively, than animals given irradiation alone, indicating that
    chloramphenicol had a deleterious effect on bone marrow recovery after
    X-irradiation (Vacha  et al., 1981).

         Normal mice and those with residual bone marrow damage following
    busulphan administration were investigated for the effects of
    chloramphenicol. Female Balb/c mice were injected with busulphan to
    create bone marrow damage using a method that had been validated
    previously by the authors. Groups of mice, some having busulphan-
    induced damage and others untreated, were then given drinking water
    containing 0.5 g/dl chloramphenicol succinate; groups of 5 mice were
    then killed at various intervals up to and including 150 days after
    chloramphenicol treatment. No effects were seen in the bone marrow
    of mice given chloramphenicol alone nor in mice pretreated with
    busulfan. However, animals given both busulphan and chloramphenicol
    displayed a progressive fall in the number of pluripotential stem
    cells and granulocytic precursor cells (Morley  et al., 1976).

         Similar effects were not seen in another study where busulphan-
    treated mice were given drinking water containing 0.5 g/dl
    chloramphenicol for six weeks. There was no effect on the colony-
    forming ability of bone marrow or spleen cells. This study used
    an identical busulphan regime to that described above (Pazdernik &
    Corbett, 1980).

         In cells of lethally X-irradiated mice given 10 mg
    chloramphenicol i.p. on days 2-8 or on days 7-12 after irradiation,
    mitochondrial swelling was noted in early erythroblasts, but not in
    intermediate or late types. The mitochondria showed reductions in the
    number of cristae (Miura  et al., 1980).

         A similar investigation in mice given 4.78 Gy X-irradiation and
    300 mg/kg b.w. of chloramphenicol succinate showed that dividing bone
    marrow cells had decreased entry into S-phase of the cell cycle. The
    affected cells were mainly of the erythroid type. The same types of
    effect were also noted in non-irradiated mice given chloramphenicol
    (Benes  et al., 1980).

         The femora of mice given 500 mg/kg b.w. chloramphenicol for 6
    days by injection (route unspecified) which were then implanted into
    untreated syngeneic mice showed greatly decreased colony formation
    when the bone marrow was subsequently injected into lethally
    irradiated mice (Nara  et al., 1982).

         No haematological effects, including aplastic anaemia, were seen
    in mice given 40 mg/kg nitrosochloramphenicol for 10 days followed by
    sacrifice 6 weeks after the last injection (Siegel & Krishna, 1980;
    abstract only).

         Groups of 6 male Sprague-Dawley rats were each given 50 mg/kg
    b.w. chloramphenicol succinate by i.v. infusion. Half of the animals
    were subjected to liver resection 15 minutes after infusion, while the
    others were sacrificed at this time. Bleeding time and blood loss were
    significantly increased in resected animals given chloramphenicol
    compared with controls (bleeding time: 500 seconds in treated animals,
    300 seconds in controls; blood loss 2.2 g in treated animals, 0.9 g in
    controls). No effects on haemoglobin or haematocrit were observed
    following infusion or liver resection (Bengmark  et al., 1981).

         Four cats were given daily i.m. injections of 50 mg/kg b.w.
    chloramphenicol for 21 days. Two untreated cats served as controls;
    all 6 animals had recently recovered from experimental infectious
    feline enteritis. Animals given chloramphenicol became very ill, with
    severe loss of appetite developing within 7 days. All four developed
    diarrhoea toward the end of the experiment (they were sacrificed on

    day 21); one was killed  in extremis. Marrow examination was carried
    out, which revealed vacuolation of the myeloid and erythroid
    precursors and of some lymphocytes. There were no significant changes
    in peripheral red cell numbers, but white cell counts were much
    reduced (Penny  et al., 1967).

         A group of 6 cats was given chloramphenicol orally at a dose of
    125 mg/kg b.w./day for 14 days and then observed for another 3 weeks.
    Another group, formerly intended as controls, were dosed with 60 mg/kg
    b.w./day chloramphenicol in the same manner. Signs of toxicity
    included CNS depression, dehydradation, loss of appetite, body weight
    loss, diarrhoea, and vomiting. Blood and bone marrow samples were
    obtained both prior to and after chloramphenicol treatment. The major
    findings related to chloramphenicol administration were severe bone
    marrow suppression with marrow hypoplasia, prevention of maturation of
    erythroid cells, and inhibition of mitosis in the marrow. Vacuolation
    of lymphocytes, and early myeloid and erythroid cells occurred. The
    effects were most severe in cats given 120 mg/kg b.w. chloramphenicol.
    On cessation of treatment the bone marrow suppression resolved (Watson
    & Middleton, 1978).

         In a similar study, 5 cats were given 50 mg/kg b.w./day
    chloramphenicol orally for 21 days. CNS depression, appetite loss, and
    weight loss were noted. Examination of peripheral blood was conducted
    both before and after dosing. This revealed lower platelet numbers
    after 1 week, and a lower neutrophil count after 3 weeks of
    administration. One animal developed lymphocytopenia after 1 week and
    neutropenia after 3 weeks. At the end of treatment, the bone marrow
    was found to have vacuolated early myeloid cells and lymphocytes, with
    reduced myeloid maturation and reduced marrow cellularity. No test for
    recovery was conducted (Watson, 1980).

         Twenty dogs were given oral doses of chloramphenicol for 14 days.
    They were dosed in the following manner: 6 dogs, 225 mg/kg b.w./day;
    4 dogs each, 175 or 125 mg/kg b.w./day; and 3 dogs each at 275 or
    75 mg/kg b.w. day. Signs of toxicity included a decline in the rate of
    weight gain and hypophagia. No changes in erythrocyte counts,
    reticulocyte counts, haemoglobin concentration, packed cell volume, or
    differential leukocyte counts occurred, but bone marrow examination of
    the dogs given 225 or 275 mg/kg/b.w./day revealed suppression of
    erythropoiesis. In dogs given 275 mg/kg b.w./day chloramphenicol,
    decreased mitotic activity and a reduced rate of granulocyte formation
    was also evident. No dogs showed bone marrow vacuolation
    (Watson, 1977).

         After i.v. administration of a single dose of 50 mg/kg b.w.
    chloramphenicol succinate to five mongrel dogs, platelets were
    obtained from blood at 0, 30, 60, 120, 180, and 240 minutes and at 24
    hours after dosing. Protein synthesis as determined by the rate of
    incorporation of 3H-leucine was inhibited, with maximum depression
    (940% of control values) occurring at 30 - 240 minutes after dosing
    (Agam  et al., 1976).

         Neonatal Holstein calves (1 day of age at the beginning of the
    experiment) given "an adequate" intake of colostrum were given
    chloramphenicol by several methods. These were, i.v. as a 25 mg/kg
    b.w. bolus at 1, 7, 14, and 28 days of age, i.v. as a 25 mg/kg b.w.
    injection every 12 hours until 150 mg/kg b.w. had been delivered, and
    as a 25 mg/kg b.w. bolus i.v., i.m., or s.c. alternately, with 1 week
    between the doses. No effects on haematological parameters were
    observed, and bone marrow aspirates showed no evidence of suppression
    or toxicity (Burrows  et al., 1984).

         A similar lack of effect on the bone marrow was noted in a study
    of cross-breed calves given doses of 9, 20, or 60 mg/kg b.w.
    chloramphenicol daily for 6 weeks (Mitema, 1982).

         In another study in Holstein calves in which chloramphenicol was
    given orally at a dose of 100 mg/kg b.w./day over 10 days, bone marrow
    suppression did occur. Similar results were noted in over 50 calves
    investigated over a period of time. Changes included partial aplasia
    to almost complete aplasia of the marrow, with loss in cellularity of
    erythrocytes, white cells, and megakaryocytes. Occasionally only fat
    deposits were noted with lymphocytic infiltrations. Lower blood levels
    of chloramphenicol were attained after oral intake than following i.v.
    injection, but the toxic effects on the marrow were greater after oral
    dosing (Krishna  et al., 1981).

         In in  vitro experiments, chloramphenicol and its postulated
    metabolite, nitrosochloramphenicol, have shown adverse effects on bone
    marrow cells. Chloramphenicol caused dose-related inhibition of
    erythroid and granulocytic colony forming units obtained from LAF1
    mice. The lowest concentration used (5 µg/ml) caused some degree of
    inhibition of erythroid cells, while the highest concentration
    (60 µg/ml) produced complete inhibition (Yunis, 1977). Similar effects
    were noted in a separate study (Hara  et al., 1978).

         Chloramphenicol and nitrosochloramphenicol inhibited DNA
    synthesis in rat bone marrow cells  in vitro. This was reversible
    with chloramphenicol, but not with the nitroso compound. Similarly,
    the nitroso compound, but not chloramphenicol, bound irreversibly to
    bone marrow cells (Gross  et al., 1982). However, in another
     in vitro study, chloramphenicol and nitrosochloramphenicol had no
    effects on mouse haematopoietic precursor cells (Pazdernik & Corbett,
    1979).

    Special studies on mutagenicity

         The antibiotic activity of chloramphenicol is not thought to
    involve any type of reaction with bacterial genetic material. It
    appears to inhibit protein synthesis by binding to the 50S subunit of
    the 70S ribosome, inhibiting the formation of the peptide bond during
    protein synthesis (Smith & Weber, 1983, Gilman  et al., 1985).

         Chloramphenicol has been shown to cause DNA strand breaks in
    bacterial cells and to inhibit DNA synthesis in lymphocytes and in a
    phage of E. coli (Amati, 1970; Dewse, 1977; Jackson  et al., 1977).
    However, chloramphenicol provided resistence to UV-induced damage in
    E. coli B/r (Doudney & Rinaldi, 1985).

         Nitrosochloramphenicol, which is a potent inducer of DNA strand
    breaks in bacterial cells, resulted in strand breakage and loss of
    helix, bringing about rapid degradation of isolated E. coli DNA
     in of helix, bringing about rapid degradation of isolated E. coli
    DNA  in vitro (Skolimowski  et al., 1981, 1983). Nitrosochlor-
    amphenicol, but not chloramphenicol, produced inhibition of DNA
    synthesis and caused DNA strand breaks in E. coli. (Murray
     et al., 1982; Yunis, 1984).

         Chloramphenicol has been tested in a variety of assays for
    mutagenic activity. Most of these assays are well established using
    well validated methods, but some, such as colicine induction, the
    induction of tandem genetic duplications in S. typhimurium, and
    tests using snails, are less well known and are unvalidated. The
    results of these tests are presented in Table 2.

         In general chloramphenicol gave negative results in bacterial
    reverse mutation assays, in assays for DNA repair in bacteria, in the
    dominant lethal test in rodents and D. melanogaster, in the
    CHO/HGPRT test, in a test for sister chromatid exchange in human
    lymphocytes, in the micronucleus test, and in a test for DNA binding.
    Tests for inhibition of growth in E. coli were also negative and,
    moreover, chloramphenicol has sometimes been used as a negative
    control in this assay (McCoy  et al., 1980a; McCoy  et al., 1980b;
    Rosenkranz, 1977; Rosenkranz  et al., 1974; Braun  et al., 1977;
    Braun  et al., 1982). There were isolated exceptions to these
    negative results. However, the only type of test that gave
    consistently positive results was the test for induction of
    chromosomal aberrations. Chromosomal anomalies were noted in human
    lymphocytes treated  in vitro with chloramphenicol, in mouse bone
    marrow in animals given chloramphenicol at doses of 50 mg/kg i.p. or
    i.m., and in F1-generation mouse liver.

         The failure of chloramphenicol to give positive results in the
    Ames test has been attributed by one group of authors to its bacterial
    toxicity. In their study, the D(-)-threo isomer of chloramphenicol
    gave negative results in the Ames test, as observed previously in
    several other studies; it was also toxic to the bacterial tester
    strains used, TA100 and TA1535. However, the L(+)-threo isomer,
    which is not used therapeutically, was much less toxic and could be
    tested at higher concentrations; with the isomer, a dose-related
    mutagenic response was noted in both tester strains (Jackson  et al.,
    1977).

         The failure of chloramphenicol to give positive results in most
    types of commonly used mutagenicity tests, with the exception of those
    examining the induction of chromosome aberrations, has been recognized
    by other reviewers (Waters  et al., 1983; Garrett  et al., 1984), as
    has the failure of the substance to produce mutations  in vivo
    (Holden, 1982).

         Chloramphenicol has been reported to enhance the mutagenicity of
    N-methyl-N-nitro-N-nitrosoguanidine in bacterial assays (Sklar &
    Strauss, 1980; Baltz & Stonesifer, 1985).

    Special study on ocular toxicity

         No toxic effects were seen in groups of three rabbits following
    vitrectomy when solutions containing 10 or 20 µg/ml chloramphenicol
    were infused into the eyes as vitreous replacements. Histological and
    electroretinographical examinations yielded normal results 2 weeks
    after infusion. Following an infusion of a 50 µg/ml solution,
    electroretinography was normal after 2 weeks, but abnormal retinal
    histology, which was not described but was said to be widespread and
    generalized, was noted (Stainer  et al., 1977).

    Special studies on ototoxicity

         Solutions of 0.5% chloramphenicol introduced into the bullae of
    the ears of guinea pigs produced no effects on hearing, but solutions
    containing 1 - 5% chloramphenicol produced moderate hearing loss at a
    variety of frequencies. Similar findings were noted when the
    electrical responses of the hair cells were measured after
    chloramphenicol administration into the bullae. Intratympanic
    administration to guinea pigs of a 1% solution of chloramphenicol
    produced a moderate degree of hair cell loss in the organ of Corti,
    with severe inflammation of the mucosa of the middle ear. Introduction
    of a solution containing 8 or 16 mg chloramphenicol through a hole in
    the bullae of the ears of guinea pigs followed by sacrifice 3, 6, 9,
    or 24 hours later resulted in severe destruction of hair cells and
    supporting cells in the basal turns of the organ of Corti. The effects
    were similar regardless of the dose or the time of sacrifice after
    dosing (Proud  et al., 1968; D'Angelo  et al., 1967; Patterson &
    Gulick, 1963; Morizono & Johnstone, 1975; Parker & James, 1978).

        Table 2.  Results of mutagenicity assays with chloramphenicol
                                                                                              

    Test System         Test Object          Concentration     Results    Reference
                                                                                              

    Ames test           S. typhimurium       30 µg/plate       -          Brem et al.,
                        TA1530, TA1535                                    1974
                        TA1538

                        S. typhimurium       0.17-24 µg/ml     +          Mitchell et al.,
                        TA98                                              1980

                        S. typhimurium       Not given         -          Heddle & Bruce,
                        TA1535, TA1537                                    1977

                        S. typhimurium       30 µg/plate       -          Rosenkranz et al.,
                        TA98, TA100                                       1976

                        S. typhimurium       < 4.5 nmole       -          McCann et al.,
                        TA98, TA1535                                      1975
                        TA1538

    Bacterial           E. coli              27 µg/ml          +          Mitchell &
    mutation assay      CM891                                             Gilbert, 1984

    CHO/HGPRT           Chinese hamster      Not given                    Augustine et al.,
    mutation assay      ovary cells                                       1982

    Gene mutation       D. melanogaster      Not given         +a         Narda & Gupta,
                                                                          1972
                                                                          (abstract only)

    Dominant lethal     D. melanogaster      Not               -          Nasrat et al.,
                                                                          1977

                        Mouse (101×C3H)F1    2×1.5 g/kg        -          Ehling, 1971
                        Mouse (ICR/Ha        333 mg/kg         -          Epstein & Shafner,
                        Swiss)                                            1968
                        Mouse (ICR/Ha        333 & 666 mg/kg   -          Epstein et al.,
                        Swiss)                                            1972

    DNA repair          B. subtilis          2.5×10-3 mg/      -          Sekizawa &
                        H17, M45             disc                         Shibamoto, 1982

                        B. subtilis          Not given         -          Suter & Jaeger,
                        H17, M45                                          1982
                                                                                              

    Table 2.  Results of mutagenicity assays with chloramphenicol (cont'd)
                                                                                              

    Test System         Test Object          Concentration     Results    Reference
                                                                                              

                        E. coli              Not given         +          Suter & Jaeger,
                        AB1157/JC5547                                     1982
                        AB1157/JC2921
                        AB1157/JC2926
                        AB1157/JC5517

                        E. coli WP2                                       Mitchell et al.,
                        uvrA+recA+,uvrA-     Not given         -          1980
                        recA-
                        trp-/trp+            Not given         -
                        A2Cs/A2Cr            3-48 µg/ml        +

                        E. coli B/r          100 µg/ml         -          Masek, 1977

                        E. coli K12          > 30 µg/ml        -          Mamber et al.,
                        (SOS chromotest)                                  1986

    Preferential        E. coli K12          5-20 µg/          -          Hyman et al.,
    inhibition                               plate                        1980

                        E. coli              30 µg/plate       -          Brem et al.,
                        Pol A+, Pol A-                                    1974

                        E. coli              10 µg/plate       -          McCoy et al.,
                        Pol A+, Pol A1-                                   1980a,b

                        E. coli              30 µg/plate       -          Longnecker et al.,
                        Pol A+, Pol A-                                    1974

                        E. coli              30 µg/disc        -          Slater et al.,
                        Pol A+, Pol A-                                    1971

    Gene conversion     S. cerevisiae        Not given         -          Mitchell et al.,
                        D4                                                1980

    Sister chromatid    Human lymphocytes    200 µg            -          Pant et al., 1976
    exchange            (in vitro)
                                                                                              

    Table 2.  Results of mutagenicity assays with chloramphenicol (cont'd)
                                                                                              

    Test System         Test Object          Concentration     Results    Reference
                                                                                              

    Chromosomal         Zea mays             30 µg/ml          -          Verma & Lin, 1978
    aberrations
                        Human lymphocytes    10-40 µg/ml       +          Mitus & Coleman,
                        (in vitro)                                        1970
                        Human lymphocytes    Not given         +          Goh, 1979
                        (in vitro)
                        Human lymphocytes    200 µg            +          Pant et al., 1976

                        Mouse, bone          50 mg/kg b.w.     +          Manna & Bardham,
                        marrow               3×50 mg/kg        +          1977
                                             b.w., 8h
                                             intervals
                        Mouse, F1 liverb     50 mg/kg b.w.     +          Manna & Roy, 1979

    Micronucleus        Tradescantia         0.1 - 5mM         -          Ma et al., 1984
    test                paludosa
                        Mouse                Not given (5      -          Heddle & Bruce,
                        (CH3×C57)F1          daily doses)                 1977

    DNA binding         E. coli              100-1000 µM       -          Kubinski et al.,
    assay                                                                 1981

    Enhancement of      Syrian hamster       0.7-5mM           +          Hatch et al.,
    SA7 virus cell      embryo cells/                                     1986
    transformation      simian adenovirus
                        SA7

    Aneuploidy          Hordeum vulgare      300 µg/ml         +          Yoshida et al.,
    induction                                                             1972

    Colicine            S. typhimurium       0.1-60 µg/        -          Ben-Gurion, 1978
    induction           TA1537 REN           plate
                                                                                              

    Table 2.  Results of mutagenicity assays with chloramphenicol (cont'd)
                                                                                              

    Test System         Test Object          Concentration     Results    Reference
                                                                                              

    Increased           Snail                Not given         -          Xie, 1985
    embryonal length

    Induction of        S. typhimurium       0.155-3.1 µM      -          Pall & Hunter,
    tandem genetic      TR4179, TT1984                                    1985
    duplication
                                                                                              

    a    Very weak positive response.
    b    One male mouse was given 50 mg/kg chloramphenicol i.m. and then was mated
         with 4 untreated females. Mice (3) were sacrificed on days 12, 16, and 18
         of gestation. The remaining mouse was allowed to litter normally and the
         young were sacrificed when 7 days old. Livers of fetuses and neonates were
         removed and examined.
             When groups of 3 - 9 female Sprague-Dawley rats were given
    80 mg/kg b.w. chloramphenicol in the drinking water for 10 days with
    or without exposure to short duration high intensity noise,
    ototoxicity was noted as revealed by reductions in the electrical
    output of the cochlea. Noise exposure alone also reduced the output,
    but noise and chloramphenicol together resulted in a severe effect
    (Henley  et al., 1984). Similar effects were reported in abstract
    form in another study in rats (Henley, 1985).

         Chloramphenicol was given to guinea pigs (number unspecified) as
    a single i.v. dose of 400 mg/kg b.w. The threshold for the Preyer
    reflex was measured after administration and at 10, 20, and 30
    minutes, 1, 2, 3, 4, and 5 hours, and then daily for 7 days. There was
    no change in the Preyer reflex with noise of 1 and 8 KHz and no loss
    of hair cells, indicating no ototoxic response in this study
    (Beaugard  et al., 1979; Beaugard  et al., 1981).

    Special study on effects on sleep

         Oral doses of 160 250 mg/kg b.w. chloramphenicol suppressed
    paradoxical sleep in cats. After a dose of 330 mg/kg b.w. paradoxical
    sleep was suppressed for 24 hours, at which time there was also a
    depression of slow wave sleep (Petitjean  et al., 1975).

    Special study on spermatogenesis

         A group of male rats was treated daily with 100 mg/kg b.w.
    chloramphenicol succinate for 8 days (route and animal numbers not
    stated), after which they were sacrificed and the testes examined
    histologically. Examination revealed total or incomplete inhibition of
    spermatogonial divisions with "perturbed meiosis". No other details
    were provided (Timmermans, 1974).

    Special studies on teratogenicity

    Monkeys

         Chloramphenicol at doses of up to 200 mg per monkey
    (10 mg/kg b.w.) had no effect on the development of Macaca mulatta
    when given for 6 to 17 days at various intervals between the 65th and
    95th days of gestation (Courtney  et al., 1967; Courtney & Valerio,
    1974).

    Rabbits

         Groups of 5 - 8 pregnant mixed breed rabbits were given 500,
    1000, or 2000 mg/kg b.w./day chloramphenicol by garage on days 6 - 15,
    6 - 9, or 8 -11 of gestation, respectively. Historical control data
    collected over the previous 4 year period, using 192 rabbits, were
    used for comparison. Excess fetal deaths did not occur in rabbits
    given 500 mg/kg b.w./day chloramphenicol, but in the mid- and
    high-dose groups 25% and 58% fetal deaths were noted, respectively,
    compared with 10% in the historical controls. There were no excess
    incidences of fetal malformations, but delayed ossification was noted
    in fetuses from dams given chloramphenicol (Fritz & Hess, 1971).

    Chicks

         Chick eggs, less than 3 days old, were treated with 0.1 ml of
    chloramphenicol solution in distilled water. The chloramphenicol, at
    doses of 0.5 or 1.0 mg/egg, was injected into the albumen  via the
    airsac. The major anomaly observed was vesiculation of the heart and
    trunk resulting from the inhibition of differentiation of the
    splanchnopleure; this effect was most severe following 16 - 19 hours
    of incubation (36 - 57% in the 0.5 mg/egg group and 23 - 47% in the
    1.0 mg/egg group). Unfortunately, no control data were provided
    (Blackwell, 1962).

         Chloramphenicol also had adverse effects on development in a
    separate study in which fertilized eggs with embryos at the 14- or
    20-somite stages were explanted and exposed to chloramphenicol at
    concentrations of 0, 200, or 300 µg/ml for 22 - 24 hours. The major
    defects noted were those of the neural tube (failure to close) and
    forebrain. There was also evidence of an inhibition of haemoglobin
    formation (Billet  et al., 1965).

    Rats

         Pregnant Sprague-Dawley rats (5 - 15 per group) were given gavage
    doses of 500, 1000, 1500, or 2000 mg/kg b.w./day chloramphenicol over
    various stages of gestation. In addition, single gavage doses of
    2000 mg/kg b.w. were given to pregnant rats on days 5, 6, 7, 8, 9, or
    10 of gestation. A group of 553 historical control rats was used for
    comparison. Even the lowest daily dose of chloramphenicol on days 5 -
    15 of gestation resulted in significant embryo/fetal deaths (63%) when
    compared with historical controls (23%), whereas doses of 2000 mg/kg
    b.w./day on days 15 - 17 or 2000 mg/kg b.w. on days 5, 6, or 7 had no
    effect. Single doses of 2000 mg/kg b.w. on days 8, 9, or 10 of
    gestation resulted in 45% fetolethality, but the most sensitive period
    appeared to be days 9 - 15. For example, fetal deaths occurred 100% of
    the time when 2000 mg/kg b.w./day was given over days 9 - 11 of
    gestation. The highest incidences of anomalies, umbilical hernia, were
    noted at 2000 mg/kg b.w./day when given over days 6 - 8 of gestation
    (36%) and at 2000 mg/kg b.w. on day 8 (11%). High incidences of
    delayed ossification were seen in fetuses from dams given 1000 mg/kg
    b.w./day chloramphenicol on days 7 - 12 or 2000 mg/kg b.w./day on days
    11 - 13 (Fritz & Hess, 1971).

         The effects of chloramphenicol on pre-implantation embryos in the
    rat were investigated by treating groups of 7 pregnant Sprague-Dawley
    rats with 250 mg/kg b.w. chloramphenicol i.p. on either day 3 or 5 of
    gestation. The animals were killed on day 5 of pregnancy and the
    uterine contents examined. No effects on the number of blastocysts per
    female were noted, but administration on day 3 of pregnancy
    significantly reduced the number of cells per blastocyst. A reduction
    was also seen when the compound was given on day 14, but these results
    were not statistically significant (Giavini  et al., 1979).

         Pregnant Sprague-Dawley rats (number unspecified) were given 3%
    dietary chloramphenicol, equivalent to 1500 mg/kg b.w./day, from days
    0 to 20. The number of resorptions (% of total implants) was elevated
    in rats treated with chloramphenicol (31.4 - 57.0%) compared with
    controls (4.7%), while fetal weight in treated rats was only 50% of
    controls. Placental weight was also much reduced, while the number of
    live fetuses was greatly reduced by chloramphenicol. The authors then
    examined the effects of chloramphenicol when given on specific days of
    pregnancy (0 - 2, 0 - 3, 0 - 4, etc., up to 0 - 12). The major effects
    on implantations, resorptions, number of live fetuses, and fetal
    weights as described above were seen in the 0 - 8 to the 0 - 12
    feeding schedules, suggesting an effect on implantation (or effects on
    embryos soon after implantation). A large proportion of fetuses (71%)
    had edema and there was an increased incidence of wavy ribs (7%) and
    fused ribs (7%) in chloramphenicol-treated groups compared with
    controls (zero incidences in all cases in controls) (Hackler  et al.,
    1975).

         Chloramphenicol was investigated for its effects on avoidance
    learning in rats. Four groups of 15 pregnant Wistar rats each were
    treated as follows: in one group 50 mg/kg b.w. chloramphenicol
    succinate was given s.c. on days 7 - 21 of gestation. In two other
    groups, 50 or 100 mg/kg b.w. was given s.c. to pups for the first 3
    days after birth; the fourth group served as controls. No effects on
    pregnancy, litter size, fetal weight, post-natal weight gain, or
    incidence of gross malformations were seen. When 60 days old, animals
    were selected for conditioned-learning study and were then examined
    for avoidance learning at days 5, 10, 15, and 20 from the start of the
    conditioning procedure. Pups from mothers given chloramphenicol
    succinate and those given the substance as neonates showed a marked
    and statistically significant impairment of avoidance learning at all
    four times. The effects were generally worse in pups given the
    substance post-natally than in those from dams administered it during
    gestation, but the differences were only slight (Bertolini & Poggioli,
    1981).

    Mice

         Groups of 8 pregnant albino mice were given oral chloramphenicol
    at doses of 25, 50, 100, or 200 mg/kg b.w. in 10 ml of distilled water
    over the third trimester of pregnancy for 5 - 7 days. Animals were
    allowed to give birth and the young were tested for conditioned
    avoidance response, electroshock seizure threshold, and performance in
    open-field tests at days 30, 38, and 42. No gross abnormalities were
    seen in any of the progeny. Dose-related effects were seen in all
    three elements of the test, with progeny from chloramphenicol-treated
    dams showing a reduced learning ability, higher brain seizure
    threshold, and poorer performance in the open-field test (Al Hachin &
    Al-Baker, 1974).

         Groups of 7 - 19 pregnant CD1 mice were given by garage 500,
    1000, or 2000 mg/kg b.w./day chloramphenicol on days 5 - 15, 6 - 12,
    or 8 - 10 of gestation, respectively. Historical control data,
    collected over the previous four-year period using 307 mice, were used
    for comparison. In the 1000 and 2000 mg/kg b.w./day dose groups, 71
    and 100% embryo/fetal deaths occurred, respectively, compared with 24%
    in controls and 31% in mice given 500 mg/kg b.w./day chloramphenicol.
    The only defects observed were a low incidence of fused sternebrae and
    elevated incidences of delayed ossification in fetuses from dams given
    1000 mg/kg b.w./day chloramphenicol (Fritz & Hess, 1971).

    In vitro

         Chloramphenicol and several other chemicals were tested in a
    mouse embryo limb bud cell culture test system which itself was being
    validated as part of the study. In all, 22 known mouse teratogens and
    5 non-teratogens were investigated in the system, which makes use of
    high-density cultures of mouse embryo limb bud cells that can

    differentiate and synthesize an extracellular matrix of sulfated
    proteoglycans. The end-point of the test considers incorporation of
    radiolabel (3H-thymidine) and the growth and synthesis of ocular
    protein and cartilage proteoglycan. Chloramphenicol gave a positive
    result in this test, with the maximum active concentration being
    5 µg/ml. [The test was around 89% predictive; no false positives were
    seen and the false negative rate was about 15% (Guntakatta  et al.,
    1984).]

         Another  in vitro test made use of the differentiation
    characteristics of rat embryo midbrain and limb bud cells, and the
    inhibition of differentiation by teratogens. Chloramphenicol gave a
    weak response, resulting in inhibition at concentrations in excess of
    50 µg/l, compared with 10 µg/l or less seen with several known
    teratogens, e.g., captan, colchicine, and parbendazole (Flint & Orton,
    1984).

    Acute toxicity

         Four groups of pregnant and non-pregnant mice were given i.v.
    doses (unspecified) of chloramphenicol. No signs of toxicity were
    reported. The LD50 value for non-pregnant mice was calculated as
    1530 (1260 - 1840) mg/kg b.w., while that for pregnant mice was
    1210 mg/kg b.w. (no confidence limits cited) (Beliles, 1972).

    Short-term studies

         No information available.

    Long-term studies

         See "Special studies on carcinogenicity".

    Observations in human beings

         Chloramphenicol is known to produce two major adverse effects in
    human beings. One of these is a generally irreversible and often fatal
    aplastic anaemia and the other is a reversible bone marrow
    suppression.

    Aplastic anaemia

         Aplastic anaemia is the most dangerous effect produced by
    chloramphenicol, although its occurrence is rare. It is usually fatal
    (Benestad, 1979). Numerous publications have appeared, most of them
    case reports describing the development of aplastic anaemia.

         An investigation into the incidence of chloramphenicol-induced
    aplastic anaemia in Hamburg suggested that the incidence was 1/11500
    with a death rate 1/18500. In the period 1965 - 70, 29 cases were
    reported, while in 1971, 3 cases were reported. Total doses in 18
    cases were in the range of 10 to 100 g, with most individuals having
    received 11 - 30 g. Onset was from 14 days (rare) to 4 - 6 months
    after chloramphenicol therapy (Hausmann & Skrandies, 1974).

         In a study in Israel in 1985, aplastic anaemia incidence was
    7.1/106 in males and 8.7/106 in females. Chloramphenicol was
    thought to account for up to 25% of cases. Aplastic anaemia usually
    took up to 12 months to develop after chloramphenicol treatment
    (Modan  et al., 1975).

         In 1969 in California, aplastic anaemia in chloramphenicol-
    treated patients was said to be 13 times more frequent than in
    the general population. Most individuals had been given oral
    doses, but in some, aplastic anaemia had occurred after i.m.
    administration. Doses were often on the order of 250 mg thrice daily
    to a total of 3 g or 250 mg four times daily to a total of 5 g. The
    majority of patients were in the 50 - 80 age group, but it was
    observed in a 15-year-old boy given a total of 3 g chloramphenicol and
    in a 37-year-old female given a total of 6 g over a month. In both
    cases onset occurred 3 - 4 months after treatment ended
    (Wallerstein  et al., 1969).

         A series of reports from Sweden in the 1970s suggested that the
    incidence of aplastic anaemia was around 80 in 1.2 million. Only 4 or
    5 of these were thought to be due to chloramphenicol treatment,
    placing the risk at 1 in 20000 (Bottiger & Westerholm, 1972;
    Bottiger & Westerholm, 1973; Bottiger, 1978; Bottiger, 1979).

         Of 108 cases of aplastic anaemia reported in Istanbul, 4 were
    thought to be due to chloramphenicol (Aksoy  et al., 1984).

         A total of 380 cases of aplastic anaemia were seen at a hospital
    in Paris in the years 1971 - 1983; 194 adult cases were considered to
    be due to chemical agents. In the period 1971 - 1977, 18/104 cases
    were attributed to chloramphenicol, whereas chloramphenicol accounted
    for 2/36 and 2/52 cases of aplastic anaemia in the periods 1977 - 1980
    and 1980  - 1983, respectively, suggesting a decreasing incidence, at
    least in the area of France under investigation (Najean & Baumelon,
    1984).

         In the period 1975 - 1980, 9 children with aplastic anaemia were
    seen at the Medical School, Gadjah Mada University (Yogyakarta). Two
    were idiopathic, but at least 3 could be attributed to treatment with
    chloramphenicol (Widayat  et al., 1983).

         A study of 40 cases of aplastic anaemia by members of the
    Association of Clinical Pathologists in the USA revealed that 27 were
    probably due to chloramphenicol. Of these, 18 had exceeded 10 g or
    more (up to 250 g) in total dosage, while 8 had received 10 g or less.
    One, an infant, had received less than 2 g chloramphenicol. Onset was
    usually 1 - 3 months after drug cessation. Route of administration and
    its duration were not specified (Sharp, 1963).

         In 1954, of 539 cases of aplastic anaemia from 37 states in the
    USA, 55 were attributable to chloramphenicol. In general there was a
    female preponderance and usually the effect developed 1 - 6 months
    after the drug was withdrawn (Welch  et al., 1954).

         In a study in Iraq, however, a male preponderance of 3:1 over
    females was noted. Of 60 patients with aplastic anaemia at the
    University of Baghdad Teaching Hospital, 12 were associated with
    chloramphenicol treatment (Al-Moudhiry, 1978).

         One study in the Netherlands examined cases of blood dyscrasias
    taken from the literature along with unpublished cases from adverse
    drug reaction reporting systems in the United Kingdom, Denmark,
    Sweden, and the Netherlands. To these were added cases from hospitals
    in Northeast Switzerland. A total of 641 cases of chloramphenicol-
    induced blood dyscrasias were identified. These included:
    thrombocytopenia (21), agranulocytosis (51), hypoplastic anaemia (39),
    bone marrow suppression (39), acute leukaemia (27), and aplastic
    anaemia (464). Of the 464 cases of aplastic anaemia, 335 (72%) had
    proved fatal. There appeared to be a female preponderance (261 cases;
    56%). However, because this cannot be related to the total number
    treated and therefore to those who did not acquire the condition, the
    incidence cannot be determined (Meyler  et al., 1974). The results
    confirmed those of earlier work (Polak  et al., 1972).

         In Italy, it has been claimed that in view of the ease of
    availability of chloramphenicol, the incidence of aplastic anaemia
    appears to be low. For example, in 1971 there were 10 reports of fatal
    side effects due to antibiotics; in 1972 there were 3. None were
    attributable to chloramphenicol. During the period 1973 - 1975 there
    were no reports of fatal cases (Preziosi  et al., 1981).

         For the period 1959 - 1969, 172 cases of aplastic anaemia were
    reported in 15 hospitals in Northeast Switzerland, and 44 of these
    individuals had been treated with chloramphenicol. The smallest total
    dose incriminated was 3 g, while the highest was 315 g
    (Keiser & Bucheggar, 1973).

         In children aged 0 - 14 years in Denmark, the total number of
    registered cases of aplastic anaemia was 39 for the years 1967 - 1982,
    giving an annual incidence of 22/106. Probable causes were
    identified for 21 of the 39 cases, and 2 of these were attributed to
    chloramphenicol treatment (Clausen, 1986).

         It was stated in 1983 that the probable overall incidence of
    chloramphenicol-induced aplastic anaemia is somewhere between 1 in
    20000 and 1 in 40000 (Venning, 1983).

         In Japan, chloramphenicol appeared to be more dangerous to the
    elderly population than to other groups. However, the investigation
    was probably biased in that only fatal cases were examined
    (Mizuno  et al., 1982). Similar findings were made in an earlier
    study (Shimizu  et al., 1979).

         For the years 1961 - 1965, 35 cases of aplastic anaemia were
    reported in Colombia, and of these 10 had had past exposure to
    chloramphenicol. Several others were said to have had a "strong
    suspicion" of exposure to the drug. Mortality was 60% (Sarasti, 1970).

         An age preponderance was noted in an analysis of 21 patients with
    aplastic anaemia, 8 of whom had been treated with chloramphenicol
    (Perez  et al., 1981).

         The minimum dose of chloramphenicol associated with the
    development of aplastic anaemia is not known with certainty. The
    literature citations often state only the total dose. Where cases
    developed after several doses, it is not possible to say if aplastic
    anaemia would also have occurred had only a single dose been given. Of
    15 cases reviewed in a report in 1974, total doses of 4.5 to 80 g had
    been given, with the usual levels being 8 to 14 g. As an example, a
    46-year-old woman was given 15 g of chloramphenicol over 10 days.
    Aplastic anaemia developed after 2 months (Hellriegel & Gross, 1974).

         Total doses of 6.5 to 60 g chloramphenicol had been given to 7
    individuals with aplastic anaemia reported in 1971 (Hodgkinson, 1971).

         A 27-year-old woman given 30 g chloramphenicol i.v. over 12 days
    developed aplastic anaemia 3 months later (Alavi, 1983).

         One individual with aplastic anaemia had undergone previous
    treatment 19 years earlier with a 500 mg initial dose followed by
    250 mg chloramphenicol 4 times daily. At the age of 23 years, he was
    given 750 mg chloramphenicol succinate (62 mg/kg b.w./day) i.v. every
    6 hours for 12 days for a brain abscess. Bone marrow suppression
    ensued and by the twelfth day of drug administration aplastic anaemia
    had developed. The patient eventually died (Daum  et al., 1979).

         A 26-year-old woman was diagnosed as being anaemic during the
    fifth month of pregnancy and in the sixth month she developed a skin
    infection. She was given 8 g chloramphenicol. She developed aplastic
    anaemia and died 8 days after giving birth. Bone marrow aspiration
    confirmed aplastic anaemia. (Suda  et al., 1978).

         Several cases of aplastic anaemia have been reported in children.
    One 7-year-old boy had been given chloramphenicol palmitate in June
    and August of 1959. The dose or route of administration were not
    specified. He developed aplastic anaemia 4 - 5 months later and died
    (Leiken  et al., 1961). A 6-year-old girl was given 25 mg/kg b.w./day
    chloramphenicol for 10 days by an unspecified route. She developed
    aplastic anaemia, apparently immediately after treatment and then
    acute myeloblastic leukaemia (Awaad  et al., 1975). A 4-year-old girl
    was given oral chloramphenicol (dose not stated) for 1 week. After 2
    months she developed aplastic anaemia (Young  et al., 1979). Another
    case of a 4-year-old girl given chloramphenicol was reported more
    recently. Chloramphenicol was given i.v. for 1 week followed by oral
    therapy for 8 weeks. The initial dose was 75 mg/kg b.w./day given
    every 6 hours, but this was adjusted after 3 weeks to 37 mg/kg
    b.w./day. Some months later she developed aplastic anaemia
    (Lepow, 1986). In one recent case the patient was a neonate born at 30
    weeks gestation weighing 1.7 kg. At day 20 after birth she developed
    suspected meningitis and was given 4 doses of 50 mg/kg b.w./day
    chloramphenicol. She developed aplastic anaemia; epidermal necrolysis
    of the skin and biliary cholestasis were found at necropsy
    (White  et al., 1986).

         A 72-year-old male patient was given a 3-week course of
    chloramphenicol (dose and route unspecified) and aplastic anaemia
    ensued within 4 months (Howell  et al., 1975).

         Aplastic anaemia is usually associated with oral intake
    (Matthews  et al., 1980). It has been reported that in 149 cases of
    chloramphenicol-induced aplastic anaemia, 85% followed oral dosing,
    14% followed parental administration, and 3% occurred after rectal
    administration (Plaut & Best, 1982).

         Topical administration of chloramphenicol has been followed by
    aplastic anaemia. In one case, a 73-year-old woman died of aplastic
    anaemia less than 2 months after beginning ophthalmic chloramphenicol
    treatment with a 0.5% solution (3 - 4 times daily) and a 1% ointment
    (once per day to the right eye) (Fraunfelder & Bagby, 1982). Several
    other similar cases have been reported (Abrams  et al., 1980;
    Rosenthal & Blackman, 1965; Carpenter, 1975; Issaragrisil &
    Piankijagum, 1985; Korting & Kifle, 1985). In one case the total dose
    was estimated to be 32 mg of chloramphenicol (Carpenter, 1975).

         One occupational case probably involved inhalation and skin
    contact. It occurred in a shepherd applying a chloramphenicol spray to
    the feet of sheep for treatment of foot rot. He had treated the
    animals twice a week with the spray, which contained 10 g of the drug
    in 100 ml of solution, for two years. Each dose contained 10 mg
    (Del Giacco  et al., 1981).

         A link between the administration of chloramphenicol and the
    development of liver disease and aplastic anaemia is known to exist.
    After five patients aged 4 to 63 years were given chloramphenicol,
    liver disease (as evidenced by jaundice, icterus, and elevated serum
    enzymes and bilirubin) was subsequently noted. Aplastic anaemia
    eventually developed (Hodgkinson, 1973). A similar case has been
    reported in a 15-year-old boy given 250 mg chloramphenicol i.v. every
    6 hours. Liver enzymes became elevated after 18 days and the liver
    itself was tender. Aplastic anaemia developed (Caslae  et al., 1982).

         The mechanism of chloramphenicol-induced aplastic anaemia is not
    understood. There may be a genetic element involved, as the effect has
    been seen in families and in identical twins exposed to chloramphenicol
    (Flach, 1982; Nagao & Maner, 1969; Silver & Zuckerman, 1980; Yunis,
    1978c). The main target may be the haemopoietic pluripotential stem
    cells of the marrow (Vincent, 1986; Silver & Zuckerman, 1980;
    Benestad, 1974; Appelbaum & Fefer, 1981). There may also be the
    possibility of initial damage to the marrow micro-environment (Camitta
     et al., 1982). Failure to note this effect in human beings with
    drug-induced aplastic anaemia makes the latter unlikely (Samson
     et al., 1972; Vincent, 1986).  In vitro studies with human bone
    marrow suggest that both the erythroid and granulocytic series are
    sensitive to chloramphenicol (Nara  et al., 1982; Hara  et al., 
    1978; Yunis, 1977). The erythroid series in particular seemed sensitive
    to chloramphenicol, with inhibition occurring at 10 mg/l compared with
    50 mg/l for the granulocytic series (Yunis, 1977).

         The problem as to whether bone marrow cells are more sensitive to
    chloramphenicol in patients with aplastic anaemia induced by the drug
    is not clear, as reports on the subject are conflicting (Yunis  et al.,
    1973a; Howell  et al., 1975; Kern  et al., 1975). It has been claimed,
    based on the results of animal studies, that individuals who are
    sensitive to chloramphenicol-induced aplastic anaemia may have
    residual bone marrow damage brought about by exposure to other agents
    (Morley  et al., 1976).

         The metabolite or metabolites responsible for the induction of
    aplastic anaemia in human beings is unknown, but nitroso-
    chloramphenicol has been implicated (Murray & Yunis, 1981; Nagai &
    Kanamuru, 1978). Nitrosochloramphenicol can be formed by the
    reduction of chloramphenicol in human liver  in vitro (Salem  et al.,
    1981). This substance is known to be toxic to human bone marrow cells
     in vitro and, moreover, is more toxic than chloramphenicol itself
    (Yunis  et al., 1980a, b). However, nitrosochloramphenicol is not
    myelotoxic to mice  in vivo (Krishna  et al., 1981). It does,
    however, cause DNA strand breakage  in vitro, and inhibit DNA
    synthesis (Gross  et al., 1982; Skolimowski  et al., 1983). Both

    chloramphenicol and nitrosochloramphenicol are taken up rapidly by
    cells, at least as demonstrated by a human transformed lymphoblastoid
    cell line (Raji cells), but the nitroso-compound covalently binds to
    these cells and to bone marrow cells 15 times more tightly than does
    chloramphenicol (Hurray & Yunis, 1981). Photochemical decomposition of
    chloramphenicol may result in potentially myeloblastic derivatives,
    which may be hazardous in ophthalmic solutions (de Vries  et al.,
    1984). Another possible mechanism involves the immune system (and even
    autoimmune damage), but there are no convincing data to support this
    hypothesis. Chloramphenicol  in vitro inhibited lymphocyte
    transformation in human material (Burgio  et al., 1974) and
    chloramphenicol reduction products, including nitrosochloramphenicol,
    were suppressive to antigen-reactive cells in mice (Pazdernik &
    Corbett, 1980).

         In summary, chloramphenicol induces aplastic anaemia in
    susceptible individuals, but no dose-response relationship has been
    identified. The mechanism may involve nitrosochloramphenicol, but this
    has not been proven. The nature of the mechanism is unknown.

    Bone marrow suppression

         Reversible bone marrow suppression has been reported in patients
    given chloramphenicol by a number of routes. The effect is thought to
    be an unlikely event at plasma levels under 20 mg/l, and it is said to
    occur in most patients at levels in excess of 25 mg/l. Generally, it
    occurs within days of administration (Lery  et al., 1978; Benestad,
    1979). One study showed that oral doses likely to result in
    suppression were usually of the order of 2 - 3 g/day or 30 - 50 mg/kg
    b.w./day; durations of dosing were generally 1 - 17 days, most being 5
    - 10 days. Plasma levels of chloramphenicol at 2 - 3 hours and 6 - 8
    hours varied, most being in excess of 25 mg/l at 2 - 3 hours and in
    excess of 30 mg/l at 6 - 8 hours. However, some were below 15 mg/l at
    both times. Of the 17 patients studied, 11 had liver disease, while 3
    of the remainder had renal disease. Bone marrow suppression in a
    female of 21 years of age was attributed to the administration of
    1.5 g/day of chloramphenicol for 18 days. The condition resolved on
    cessation of therapy (Parashar  et al., 1972). In 6 anaemic subjects
    given up to 60 mg/kg b.w./day chloramphenicol, reticulocyte response
    to vitamin B12 or iron dextran was halted and delayed (Saidi
     et al., 1961).

         Maturation arrest and cytoplasmic valuation of erythroid elements
    were seen in the marrow (Rosenbach  et al., 1960; Bartlett, 1982).

         One reported case of bone marrow suppression was in a
    three-year-old girl with cystic fibrosis given 70 mg/kg b.w./day
    chloramphenicol for 2.5 months. The suppression was accompanied by
    physical growth depression and hair loss. All these conditions
    resolved on cessation of treatment (Kapp  et al., 1977).

         The toxic effects of chloramphenicol on the bone marrow were
    lessened in one group of patients by co-administration of
    phenylalanine (Ingall  et al., 1965). The reason for this, and its
    relationship to the pathogenesis of bone marrow suppression, is
    unknown. The mechanism of suppression is thought to involve inhibition
    of mitrochondrial protein biosynthesis in bone marrow cells.  In vitro
    studies showed that 10 mg/l chloramphenicol severely inhibited protein
    synthesis in bone marrow cells, although ten times this concentration
    was required to inhibit mitochondrial respiratory functions (Yunis,
    1973a, b; Yunis, 1978a, b; Martelo  et al., 1969). As a result of
    mitochondrial dysfunction, ferrochelatase activity is suppressed in
    erythroid precursors.  In vitro studies show inhibition of haem
    synthesis with concentrations of chloramphenicol of 10 mg/l
    (Becker  et al., 1974). It seems likely that the major effect of
    chloramphenicol in the pathogenesis of reversible bone marrow
    suppression is a block on hame biosynthesis arising as a result of
    mitochondrial dysfunction (Yunis & Salem, 1980). Consequently,
    co-administration of phenylalaninine may in some way compensate for
    the inhibitory effects of chloramphenicol on protein synthesis.

    Effects on platelets

         In  in vitro experiments, chloramphenicol was shown to decrease
    human platelet aggregation. Unfortunately, no  in vivo studies are
    available (Cronber  et al., 1984, Djaldetti, 1983; Agam  et al.,
    1976).

    Carcinogenicity

         IARC concluded in 1982 and 1987 that there was limited evidence
    for the carcinogenicity of chloramphenicol to human beings based on
    studies describing cases of leukaemia (IARC, 1982; IARC, 1987).

         One study described a 24-year-old male given chloramphenicol for
    typhoid fever. Leukaemia followed aplastic anaemia. A chromosome
    translocation (t 1:7) was discovered on karyotyping. No other details
    were provided, except that similar translocations were noted in 6
    other leukaemia patients known to have had exposure to leukaemogenic
    substances or radiation (Scheres  et al., 1985).

         Acute myeloid leukaemia developed in a 6-year-old girl who
    developed aplastic anaemia after being given 25 mg/kg b.w./day
    chloramphenicol for 10 days. The leukaemia developed 6 months after
    the aplastic anaemia (Awaad  et al., 1975).

         Leukaemia developed in a 38-year-old woman given a total of 8 g
    of chloramphenicol 8 years before which resulted in aplastic anaemia.
    A 57-year-old man who had taken chloramphenicol for 8 years (about
    175 g total dose) developed aplastic anaemia followed by leukaemia
    within a year, while a 61-year-old man who was treated with
    chloramphenicol developed aplastic anaemia and leukaemia (Brauer &
    Dameshek, 1967).

         An 80-year-old man with a bacterial infection of the feet was
    given orally 250 mg/day chloramphenicol, for 7 days. He developed
    acute leukaemia 5 months later. The only other drugs given for his
    infection were penicillin and a topical zinc ointment (Humphries,
    1968).

         A 14-year-old girl given 250 mg chloramphenicol for 4.5 days
    developed aplastic anaemia after approximately 1 year followed by
    acute leukaemia 18 months later (Seaman, 1969).

         Aplastic anaemia and leukaemia may occur together at diagnosis. A
    28-year-old man developed reversible bone marrow depression after
    being given approximately 31 g of chloramphenicol. Five years later he
    was found to have aplastic anaemia and acute leukaemia (Schmitt-Graft,
    1981). Other similar cases have been reported (Adamson & Seiber, 1981;
    Forni & Vigliani, 1974; Meyer & Boxer, 1973).

         Of 641 cases of chloramphenicol-induced blood dyscrasias, 464
    cases of aplastic anaemia and 27 cases of leukaemia were identified
    (Meyler  et al., 1974).

         From 151 cases of blood dyscrasias induced by drugs, 3 cases of
    leukaemia attributable to chloramphenicol were noted (Fraumeni, 1967).

         The majority of cases of leukaemia associated with chlo-
    ramphenicol therapy were acute myeloid leukaemia (Godner  et al.,
    1973).

         The role of chloramphenicol in leukaemogenesis is not known. Some
    studies have revealed abnormal karyotypes in affected patients
    (Scheres  et al., 1985; Goh, 1971; Cohen & Huang, 1973), but it is
    not known if these abnormalities were induced directly by
    chloramphenicol or were merely a feature of the disease. Aplastic
    anaemia, whether induced by chemicals or idiopathic, is known to be
    followed by leukaemia in some cases (Milner & Geary, 1979).
    Cytogenetic abnormalities are common. Fanconi's anaemia, for example,
    is often followed by leukaemia and cytogenetic abnormalities are seen
    in this state. Chloramphenicol is known to induce cytogenetic
    abnormalities in  in vitro test systems, but it is not known if those
    seen in leukaemia occurring after chloramphenicol-induced aplastic
    anaemia are due to the same effects.

    Cardiovascular effects

         Chloramphenicol is known to induce a state often referred to as
    the "Grey Baby Syndrome" or "Grey Syndrome". In general, this is a
    state of cardiovascular collapse that commences on days 2 to 9
    following the beginning of chloramphenicol administration. The main
    features include failure to feed, vomiting, abdominal distension,
    cyanosis, flaccidity, shock, and a fall in body temperature. Death is
    said to occur in 60% of cases, and the syndrome generally occurs where
    the dose of chloramphenicol exceeds 25 mg/kg b.w./day (Meyler &
    Herkeimer, 1968). Metabolic acidosis may be a presenting feature, and
    plasma levels of chloramphenicol in excess of 30 µg/ml are usually
    associated with the development of the syndrome (Evans & Kleiman,
    1986; Mulhall  et al., 1983; Craft  et al., 1974). In several cases
    the doses producing the syndrome were 50 mg/kg b.w./day (Fripp
     et al., 1983; Kraskinski  et al., 1982; Biancaniello  et al.,
    1981; Haile, 1977) and in one case a daily dose of 100 mg/kg b.w. was
    given (Stevens  et al., 1981). A grey-type syndrome has been reported
    in a 16-year-old girl given 1 g chloramphenicol i.v. every 6 hours
    (95 mg/kg b.w./day) for what was thought to be Rocky Mountain spotted
    fever. This dose was later changed to 3.75 g/day i.v. The syndrome
    began to develop after the second day of this regime, but it resolved
    with drug withdrawal and supportive treatment (Brown, 1982). The
    mechanism is unknown, but experiments with isolated pig hearts suggest
    that effects on mitochondria may be important (Werner  et al., 1985).

    Allergenic contact dermatitis

         Allergenic contact dermititis has been reported in case report
    following the use of chloramphenicol topical preparation (Rudzki  et al.,
    1976; Schewach-Millet & Shapiro, 1985; Van Joost  et al.,1986;
    Strick, 1983; Fraki  et al., 1985).

         Two cases of occupational dermatitis in oculists were attributed
    to the use of chloramphenicol-containing preparations (Rebandel &
    Rudzki, 1986). In one study of 330 patients with contact allergy, 10%
    showed positive patch tests to chloramphenicol, while in another
    investigation of 620 patients, 1.7% save positive reactions
    (Blondeel  et al., 1978; Rudzki & Kelniewska, 1970). Others have
    reported similar findings (Forck, 1971).

    Ocular toxicity

         Several cases of ocular toxicity following prolonged treatment
    with chloramphenicol have been reported. This has often been described
    as an optic neuritis with scotomata and failing vision. Retrobulbar
    neuritis may be observed. It has been seen in cases of cystic
    fibrosis, although this may reflect the choice of chloramphenicol as a
    treatment in this condition rather than a specific sensitivity in this

    group. Total doses are often in the 80 - 250 g range given over
    several months. One patient developed bilateral optic neuritis after
    being given 6 g chloramphenicol per day for 6 weeks (approximately
    250 g total). Peripheral neuritis may accompany the ocular effects
    (Wilson, 1962; Steidl, 1965; Malbrel  et al., 1977; Joy  et al.,
    1960; Lasky  et al., 1953; Charache  et al., 1977; Chang  et al.,
    1966; Cocke  et al., 1966; Godel  et al., 1980; Wallenstein &
    Snyder, 1952; Huang  et al., 1966; Walker, 1961; Fraunfelder & Meyer,
    1984).

    Ototoxicity

         Deafness following chloramphenicol therapy has occasionally been
    noted, although the reports have been complicated by administration of
    other potentially ototoxic drugs. In one case, a 2.5-year-old boy was
    given 125 mg/kg b.w./day chloramphenicol for 26 days with no other
    drug treatment. He developed deafness, which persisted beyond
    cessation of drug administration (Gargye & Dutta, 1959; Ajodhia & Dix,
    1976; Nilges & Norther, 1971; Jones & Hanson, 1977).

    Effects on testes

         A 61-year-old man with pneumonia was treated with ampicillin and
    chloramphenicol. He later developed aplastic anaemia after 12 days and
    died of widespread sepsis 10 days after drug withdrawal. Necropsy
    revealed loss of germinal epithelium, with only Sertoli cells
    remaining. The testes were of normal size. As the patient was
    unmarried and also exposed to ampicillin, the authors concluded that
    there was no direct proof of chloramphenicol-induced testicular
    toxicity. However, in view of the normal gross appearance of the
    testes and the low toxicity of ampicillin, chloramphenicol seems the
    most likely cause of the effects noted in this case. No dose levels
    were quoted (Sheehan & Sweeny, 1982).

    Teratogenicity

         Chloramphenicol and/or tetracyclines have been implicated in the
    genesis of cleft-lip in a study of 599 affected children. However,
    because of the nature of the epidemiological study, any effects of
    tetracyclines cannot be separated from those of chloramphenicol and it
    cannot be stated with any certainty that chloramphenicol is a human
    teratogen (Saxen, 1975).

    COMMENTS

         Human exposure to chloramphenicol can give rise to aplastic
    anaemia, a rare but often fatal condition. The Committee concluded
    that no dose-response relationship could be established for this
    effect. The mechanism for the pathogenesis of aplastic anaemia is
    unknown, and no suitable animal model exists.

    EVALUATION

         Because a no-effect level for aplastic anaemia could not be
    established, and therefore it was not possible to give an assurance
    that residues in foods of animal origin would be safe for sensitive
    subjects, an ADI could not be allocated for chloramphenicol.

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    See Also:
       Toxicological Abbreviations
       Chloramphenicol (WHO Food Additives Series 53)
       Chloramphenicol (WHO Food Additives Series 33)
       CHLORAMPHENICOL (JECFA Evaluation)
       Chloramphenicol (IARC Summary & Evaluation, Volume 50, 1990)