MINERAL OILS (FOOD-GRADE), PARAFFIN WAXES AND MICROCRYSTALLINE WAXES
First draft prepared by
Ms F D Pollitt, Dr S Barlow and Ms S O'Hagan
Health Aspects of the Environment and Food (Medical) Division
Department of Health, London, United Kingdom
Explanation
Biological data
Toxicological studies
Short-term toxicity studies
Observations in humans
Comments
Evaluation
References
1. EXPLANATION
Food grade mineral oils were last evaluated by the Committee at
its thirty-seventh meeting (Annex 1, reference 94). At that meeting,
it reconsidered two 90-day feeding studies in Fischer 344 (F344) rats
given both oleum-treated and hydrogenated mineral oil (the "Shell"
studies, Baldwin et al., 1992). In the first study, haematological
changes and deposition of mineral oil in the liver, spleen and lymph
nodes were reported; in the second, deposition in the liver, spleen
and lymph nodes was again reported, but no haematological
investigations were conducted. The Committee considered that both
effects required further investigation and recommended that an
adequate long-term feeding study should be performed using food-grade
mineral oils representative of those in commercial use. The temporary
ADI "not specified" was extended.
Petroleum jelly was last evaluated at the thirty-third meeting
(Annex 1, reference 83). At that meeting, the Committee reiterated its
concern about whether the chemical composition of petroleum jelly in
current use met the specifications for the materials tested in
toxicological studies submitted to it. It concluded that, for newer
formulations of petroleum jelly, new specifications were required and
that adequate long-term, mutagenicity, and reproductive
toxicity/teratogenicity studies should be completed. No ADI was
allocated.
Paraffin wax and microcrystalline wax were last evaluated by the
Committee at its thirty-ninth meeting (Annex 1, reference 101). At
that meeting, the Committee prepared separate specifications for these
waxes and concluded that, because these specifications limit the
number of waxes that can be used for food applications as compared
with those tested in previous studies, previous long-term toxicity
studies were suitable for evaluating the safety of hydrocarbon waxes
in current use. The Committee reviewed the results of extraction and
migration tests on waxes and wax-bearing products, information on the
absorption and metabolism of hydrocarbon waxes, a long-term feeding
study in Sprague-Dawley rats and a series of 180-day feeding
studies in rats, and concluded that petroleum-derived paraffin and
microcrystalline waxes were non-toxic and non-carcinogenic. A group
ADI "not specified" was established for micro-crystalline wax and
paraffin wax for the uses indicated in the specifications (chewing-gum
base, protective coating, defoaming agent and surface-finishing
agent). The Committee was informed that a 90-day study on hydrocarbon
waxes made both by newer processes and by traditional methods was
under way, and asked to be informed of the results when they became
available.
Since the previous evaluations, additional toxicological data
have become available on a range of mineral oils, microcrystalline wax
and paraffin wax representative of materials currently in use. These
data are summarized and discussed in the following monograph addendum.
2. BIOLOGICAL DATA
2.1 Toxicological studies
2.1.1 Short-term toxicity studies
2.1.1.1 Rats
A range of mineral oils and waxes were administered in the diet
to groups of F344 rats (20/sex) at levels of 0.002%, 0.02%, 0.2% or 2%
for 90 days (equivalent to 2, 20, 200 or 2000 mg/kg bw/day). The oils
used in the study were N10(A), P15(H), N15(H), N70(H), N70(A) and
P100(H)1. This range of oils was chosen to cover the variables
previously identified as potential influences on the toxicity of
mineral oils (viz. viscosity, method of refining and oil type). The 3
waxes tested were a hydrotreated, low-melting point, paraffinic wax
(LMPW); a hydrotreated, high-melting point, microcrystalline wax
(HMPW) and a clay treated, microcrystalline wax (high sulphur wax,
HSW). Groups of rats (60/sex) were fed the control diet for the same
period of time. A further group of rats (20/sex) were fed diets
containing 2% coconut oil. These animals comprised the main study.
Additional groups of 10 males and 10 females were fed diets
containing 2% test article or 2% coconut oil for 90 days followed by a
28-day period on control diet. Groups of 30 males and 30 females were
fed control diet for the same period of time. These animals comprised
the reversal study.
Further additional groups of 5 male and 5 female rats received
control diet, diet containing 2% test article or 2% coconut oil for 90
days (tissue level study). Further groups of 5 male and 5 female rats
received control diet, diet containing 2% test article or 2% coconut
oil for 90 days followed by a 28-day period on control diet (tissue
level reversal study).
1 Oils may be obtained from crude oil sources of naphthenic
(N) or of paraffinic (P) origin and by either the conventional
acid (oleum)-treatment process (A) or the hydrogenation or
hydrotreatment process (H). Their viscosity ranges from
10-100 centistokes (cSt) (10-100 mm2/s). Thus a P100(H) oil
refers to a paraffinic oil with a viscosity of 100 cSt
produced by the hydrogenation process and a N10(A) oil to a
naphthenic oil with a viscosity of 10 cSt produced by the
acid-treatment process.
All animals were monitored for body weight, food intake and
clinical condition. Main study and reversal study animals were subject
to a full necropsy. End of test haematology and clinical chemistry
parameters were measured. All tissues from the high-dose and control
groups were processed and examined by light microscopy, as were
limited tissues from all intermediate-dose animals. A limited list of
tissues was taken from the tissue level and tissue level reversal
animals, weighed and the level of mineral hydrocarbon material present
determined. All increases or decreases in parameters or incidence of
lesions cited below refer to comparisons with the concurrent control
group, unless otherwise stated.
Body weights were largely unaffected by any of the treatments in
this study. All treatments apart from the HSW caused statistically
significant increases in food consumption in male rats in the main
study at the 2% level. In females, increased food intake was observed
in the 2% N15(H), N70(H) and N70(A) oil and HMPW groups only, although
there were intermittent increases in food intake in females receiving
coconut oil. In the reversal study (dosed at the 2% level only), food
intake was increased in all male groups, including the HSW group,
throughout the study. In the reversal period, intake returned to
normal by day 105 in all groups with the exception of the N70(A) oil
group. Among females, there were intermittent increases in food intake
throughout the study and the reversal period in all groups including
the coconut oil group. In general, the increases did not differ from
controls by more than 10%.
Increased organ weights were recorded in a number of organs,
particularly the kidney, liver, spleen, lymph node and caecum. The
weight changes were generally more marked in female rats than in
males.
Two main types of haematological changes were observed. The first
was characterized by an increased WBC count, often accompanied by
increases in the neutrophil, lymphocyte and monocyte population and a
decrease in platelet level. Such changes were seen in both male and
female rats receiving 2% N15(H), P15(H) and N10(A) oils, LMPW and HSW
(females only). Similar changes were observed at the 0.2% level in
both sexes receiving LMPW. The second type of change was characterized
by decreased RBC and haemoglobin levels and was reported in most
instances in females, especially those receiving 2% N15(H), N70(H),
P15(H) and N10(A) oils and LMPW and 0.2% P15(H) and N10(A) oils. This
latter change seemed to be readily reversible because, apart from the
LMPW group, no effect was seen in the reversal study. In males
receiving LMPW, decreased haemoglobin and mean corpuscular haemoglobin
(MCH) levels were seen in both the main and reversal studies. There
were no significant changes in haematological parameters in rats
receiving coconut oil.
Raised serum liver enzyme levels (ALT, ASAT and gamma-GT were
seen with the N15(H), N70(A), P15(H), P100(H) and N10(A) oils in
female rats and with LMPW in both female and male rats. There was a
slight but statistically significant decrease in alkaline phosphatase
levels in a number of treatment groups. A lower albumin:globulin ratio
seen in some groups generally reflected decreased total protein and
albumin levels. There was a slight but statistically significant
higher glucose levels in all but the highest dose group of many of the
female treatment groups. Males showed a much more limited range of
effects than females. Reversal animals of both sexes given LMPW had
statistically significantly increased levels of serum liver enzymes.
Gamma-GT was also higher than control in the female N10(A) oil group
after the reversal period. Some other clinical chemistry parameters
remained altered in the reversal study, particularly in the LMPW,
N15(H) and N10(A) oil groups.
Serum vitamin E levels were measured in the tissue level and
tissue level reversal studies (2% test article). All the oils
significantly reduced the serum vitamin E concentration in both male
and female rats (to approximately 30-75% of control). This may simply
be due to a reduction in bioavailability of the vitamin resulting from
its preferential solubility in the oils. Serum vitamin E was
significantly increased in males and females receiving LMPW and in
females receiving coconut oil (to approximately 130% of control). The
only groups to show an effect of treatment after the 4-week reversal
period were the male groups treated with N70(H) and N10(A) oils.
Liver granulomas, occasionally accompanied by centrilobular
vacuolation, were seen in female groups receiving N15(H), N70(H),
N70(A), P15(H) and N10(A) oils at the highest dose level and in both
sexes in the LMPW group at the 2% and 0.2% levels. The granulomas were
focal and consisted of collections of macrophages, some with necrotic
cells, surrounded by inflammatory cells and variable fibrosis. In
general, one month after cessation of treatment, granulomas were still
present in livers of animals from these groups although in some cases
a reduction in the severity of the lesion was evident.
In the mesenteric lymph nodes, lesions were seen which comprised
focal collections of macrophages, often in the cortical region.
The macrophages were vacuolated and some had a yellowish-brown
pigmentation of varied intensity. The focal collections of macrophages
were classified as histiocytosis. They were not homogeneously
distributed but were often restricted to one node or even part of one
node, with some areas severely affected and others relatively free of
any effect. In view of the uneven distribution, the author commented
that recording the presence of this lesion was subject to chance
variation related to selection of tissue and sectioning and that this
may have led to an under-reporting of the incidence of lesion.
Histiocytosis was increased compared to controls in males given N
10(A), N70(A) and N70(H) oils at levels down to 0.2%, P15(H) oil and
LMPW at levels down to 0.02%, and in those given N15(H) oil at levels
down to 0.002%. It was seen in females given any of the N15(H),
N70(A), N70(H), P15(H) or N10(A) oils down to 0.02% and in those given
LMPW at all dose levels. In each case the severity increased with dose
and the effect was more severe in females than in males. There were
also significant levels of histiocytosis in the same treatment
groups in the reversal study. HSW, HMPW and P100(H) oil were
indistinguishable from controls at any dose level in either sex.
Macrophage accumulation in Peyers patches was observed in both
male and female animals receiving 0.2 and 2% LMPW. Among females,
macrophage infiltration of the lamina propria was also observed at the
2% level.
In the heart, focal inflammatory lesions were observed within the
cusps of the mitral valve. The lesion was characterized by an
increased cellularity of the valve with destruction of the fibrous
core. The increased cellularity was composed of a mixed population of
macrophages, plasma cells and lymphocytes. Pyknotic nuclei and cell
debris were scattered throughout the lesions. In many lesions mitotic
figures could be identified. The lesion was seen in 11/20 males and
11/20 females in the 2% LMPW groups, and in 5/20 females in the 0.2%
LMPW group. The lesion was present in 1/60 male control animals and in
occasional males from the high-dose treatment groups. Following a
28-day reversal period there was still a significantly increased
incidence of this lesion in both male and female animals treated with
LMPW, although the incidences were lower than at the end of the
treatment period.
Tissue levels of mineral hydrocarbons were determined in the
kidney, liver, mesenteric lymph node and perirenal fat in the tissue
level study. Increased levels were seen in both males and females in
one or more of these tissues for all oils and waxes except the HSW and
HMPW. A decrease in hepatic levels was seen following the 28-day
reversal period (from 16% for the N70(H) oil to 76% for the N70(A)
oil). Little reduction was seen in the mesenteric lymph nodes or in
the kidneys.
Livers of female rats in the control and 2% dose groups from the
main and reversal studies (excluding the coconut oil group) were
subsequently analyzed for content of hydrocarbons of different chain
length. Four animals/group were used from the control, P15(H) oil
and LMPW groups but only one animal/group from the other groups.
Chromatograms of liver extracts of the control, reversal P100(H) oil
group, main and reversal HMPW groups and main and reversal HSW groups
showed no oil or wax profile. The authors concluded that, for the
other test articles, the composition of the liver deposits was
comparable but not identical to the original material. For the LMPW,
the relative content of higher boiling hydrocarbon specimens in the
liver deposit was found to have further increased after the 28-day
reversal period. Also, the isoparaffin content of the residues was
relatively higher compared to the original test material. The authors
suggested that mobilization takes place and that the rate depends on
molecular weight and structure of the hydrocarbon: the lower molecular
weight, linear molecules being metabolized fastest. It was further
concluded from these results that no hydrocarbons with carbon numbers
over C35 passed the intestinal membranes.
The NOELs in this study were as follows: N15(H) oil and LMPW:
less than 0.002%, equivalent to < 2 mg/kg bw/day; N70(H), N70(A),
P15(H) and N10(A) oils: 0.002%, equivalent to 2 mg/kg bw/day; P100(H)
oil, HSW and HMPW: 2%, equivalent to 2000 mg/kg bw/day (Worrell,
1992).
In a further 90-day study in F344 rats, a P70(H) oil was
administered in the diet to groups of 20 male and 20 female rats at
levels of 0.002%, 0.02%, 0.2% or 2% for 90 days. Groups of 20 males
and 20 females were fed control diet for the same period. These
animals comprised the main study. A further 5 male and 5 female rats
received control diet or diet containing 2% test article for 90 days
to provide tissue for measurement of mineral hydrocarbon levels
(tissue level study). Study parameters monitored were as for Worrell,
1992, above, except that histopathological examination was limited to
the liver and mesenteric lymph nodes of rats in the main study.
There were no treatment-related effects on body weight. Food
consumption was increased by approximately 10% in both sexes at the 2%
level in the main study. Increased spleen weights were seen in male
rats at the 0.2 and 2% levels in the main study only; in female rats,
increased liver, spleen and kidney weights were seen at the 2% level
in the main study but in the tissue level study only increased liver
weights were seen.
No treatment-related changes were seen in haematological
parameters. Raised ALAT, gamma-GT and ALP levels were seen
intermittently in male and female dose groups but there was no clear
dose-related effect. The only significant histopathological finding
was an increased incidence of pigmented macrophages in the lymph nodes
of male rats receiving the oil at the 2% level.
Residues of mineral hydrocarbon material were found in the liver
and mesenteric lymph nodes of both male and female rats and in the
kidneys of female rats in the tissue level study. In this study, the
NOEL for the P70(H) oil was considered to be 0.2% since only minor
effects, of doubtful biological significance, were seen at this dose
level (Brantom, 1993).
In a further 90-day study in F344 rats, the test materials were
three mineral hydrocarbon waxes: a paraffinic, high-melting point,
intermediate wax (also referred to as wax 64); a blended wax
containing a 50:50 mixture of the low-melting point and high-melting
point waxes used in the study by Worrell, 1992 (microcrystalline/
paraffin wax blend); and the LMPW tested by Worrell, 1992. The
intermediate wax and the blended wax were fed in the diet to groups of
20 male and 20 female rats at levels of 0.02, 0.2 or 2%. The LMPW was
fed at 2% only. Groups of 20 male and 20 female rats received control
diet only. These animals comprised the main study. Further groups of
10 male and 10 female rats were fed either control diet only, 2%
intermediate wax, 2% blended wax or 2% LMPW for 90 days, followed by
an 85- or 86-day reversal period in which they received control diet
only. This was the reversal study. Further groups of 5 male and 5
female rats received the same dose levels as in the reversal study for
96-97 days to provide tissue for measurement of mineral hydrocarbon
content (tissue level study). Samples of liver were also taken from 5
female animals/group in the reversal study for measurement of mineral
hydrocarbon levels (pooled sample). Methodology was as in the study by
Worrell, 1992, except that histopathological examination was limited
to liver and lymph nodes, any abnormal tissue, and to the mitral valve
of the hearts from 5 male and 5 female animals from the control, 2%
intermediate wax, 2% blended wax and 2% LMPW groups in the main and
reversal studies.
There were no significant treatment-related effects on body
weight. Food intake was increased in male and female rats receiving 2%
LMPW and 2% blended wax in the main study and in female rats receiving
0.2% blended wax.
Increased liver, mesenteric lymph node and/or spleen weights were
seen with all waxes in the 0.2% or 2% groups. Increased mesenteric
lymph node weight was also seen in the 0.02% intermediate wax group.
In the reversal study, the weights of these organs were still
frequently slightly increased in all treated groups.
Among haematology parameters, there were some sporadic decreases
in haemoglobin and MCH concentrations. In male rats, increased
monocyte and neutrophil levels and decreased lymphocyte levels were
recorded in the groups receiving 2% blended wax or LMPW. In females,
monocyte levels were markedly increased at all dose levels in the
intermediate wax groups, and at 2% blended wax and LMPW. Increases in
eosinophil and neutrophil counts also occurred. Platelet levels were
decreased in both male and females at all but the lowest dose level.
In the reversal study, no effects were seen in male rats but, in
female rats, WBC levels were raised and platelet levels remained
decreased.
Transaminase levels were increased in male and female rats
receiving the intermediate and blended waxes at the 0.2 and 2% levels
and in male rats receiving 0.02% intermediate wax. Large (1-2 fold)
increases in transaminase levels were seen in the LMPW groups. These
enzymes remained elevated in both sexes receiving blended wax and LMPW
in the reversal study. Large increases were recorded in gamma-GT
levels in female rats only in the 2% blended wax and LMPW groups in
the main study and in all wax groups in the reversal study.
At histopathological examination, increased incidences of
granuloma and/or microgranuloma were seen in the liver in all the 0.2%
and 2% dose groups, often with an increased incidence of centrilobular
or periportal vacuolation. Increased incidences of microgranuloma
persisted in the reversal study in all groups except the females
receiving intermediate wax. Significant levels of histiocytosis were
seen in the cervical lymph nodes of male and female rats receiving the
blended wax or LMPW at 2% and in female rats receiving 2% intermediate
wax in the main study. In the reversal study, the incidence remained
significantly increased only in female rats receiving intermediate wax
or LMPW. In the mesenteric lymph nodes, histiocytosis was seen in all
dose groups in the main study and the incidence remained significantly
increased compared to controls in all dose groups in the reversal
study.
The lesion in the mitral valve, reported in the study by Worrell,
1992, was also seen in this study. Birefringent material, identified
as mineral hydrocarbon material, was also seen in the mitral valve in
certain animals in this study. The incidences among those rats
examined (2% dose level only) are shown in Table 1.
Tissue levels of mineral hydrocarbons were determined in the
liver, kidneys, spleen and mesenteric lymph nodes in the tissue level
study. Markedly increased levels were seen in both males and females
in all tissues examined except in the kidneys of males fed
intermediate wax. In the reversal study in female animals, hepatic
levels of mineral hydrocarbons were considerably less (84%, 80% and
91% less for the LMPW, intermediate wax and blended wax groups,
respectively). No NOEL could be identified in this study for any of
the waxes tested (Brantom & Coatsworth, 1993; Freeman et al.,
1989; Simpson & Smith, 1992).
A further 90-day study was carried out on oil P15(H), the
objectives of which were to reproduce liver and lymph node findings
attributed to some of the mineral oils in previous studies and to
provide insight into the comparative sensitivities of different rat
strains to a representative oil. Only female rats were used as these
had shown the greatest sensitivity to mineral oils in previous
studies.
Groups of 35 female F344 rats received either control diet or
diet containing 2% P15(H) oil. A further group of 15 F344 rats
received diet containing 0.2% P15(H) oil. Groups of 15 female CD
Sprague-Dawley rats received control diet, 0.2 or 2% P15(H) oil. In
the F344 rat control and high-dose groups, interim sacrifices of 10
animals/group were carried out at 30 or 61 days. Five animals/group
were used for mineral hydrocarbon analysis at end of test (tissue
level study). Methodology was similar to that in the study by Worrell,
1992, except that histopathological examination was limited to liver,
mesenteric lymph nodes and any abnormal tissues.
Table 1. Incidence of effects of different waxes on the mitral valve
(2% dose level)
Control Intermediate Blended LMPW
Main study
Mitral valve lesion M 0/5 0/5 4/5 5/5
F 1/5 2/5 5/5 5/5
Bifringent material M 0/5 0/5 5/5 5/5
F 0/5 5/5 5/5 5/5
Reversal study
Mitral valve lesion M 0/5 2/5 4/5 5/5
F 0/5 3/5 5/5 5/5
Bifringent material M 0/5 1/5 0/5 0/5
F 0/5 1/5 0/5 3/5
There were no consistent, treatment-related effects on
body-weight gain nor on food consumption in either strain. Increased
liver weights were seen in all F344 rat groups receiving P15(H) oil,
and increased mesenteric lymph node and spleen weights were also
recorded in rats dosed at the 2% level. There were no such effects in
Sprague-Dawley rats.
WBC counts were increased in F344 rats in the 2% group at days 30
and 61 but not at the end of test (EOT). The proportion of neutrophils
was increased and that of lymphocytes decreased in the 2% F344 rat
group at day 61 and at EOT. Decreased RBC counts, haemoglobin and
haematocrit values were also seen in F344 rats in the 2% group at day
61 and EOT, and in the 0.2% group at EOT. No treatment-related
haematological changes were seen in Sprague-Dawley rats. F344 rats
receiving 0.2% or 2% P15(H) oil showed an increase in gamma-GT at EOT.
Intermittent increases in cholesterol levels were also seen in F344
rats but no effects were seen on clinical chemistry parameters in
Sprague-Dawley rats.
Among F344 rats, microgranulomas were present in the livers of
rats fed 2% P15(H) oil at day 61 and at EOT but not at day 30, and in
rats fed 0.2% at EOT. Treatment-related effects in the mesenteric
lymph nodes were microgranulomas (in the 0.2% group at EOT, 2% group
at day 61 and EOT), histiocytosis (2% group at day 30) and focal
infiltrates of eosinophils (2% group at day 30). Among Sprague-Dawley
rats, there was a slightly increased incidence of minimal, multifocal,
chronic inflammation in the 2% dose group at EOT. In contrast to F344
rats, discrete microgranulomas were not observed in either dose group.
Tissue levels of mineral hydrocarbons were determined in the
liver, mesenteric lymph node, kidney and spleen. Increased levels were
seen in particular in the liver, with a dose-related increase compared
to controls in both strains of rats. The level of mineral hydrocarbon
material was equivalent in the 2% Sprague-Dawley and the 0.2% F344
groups. Mineral hydrocarbon levels in the mesenteric lymph node were
raised to a similar extent in F344 and Sprague-Dawley rats. No
accumulation of mineral hydrocarbon was seen in the spleen or kidney.
In this study, Sprague-Dawley rats were clearly less susceptible
to the toxic effects of the test oil than F344 rats, although mineral
hydrocarbon material was absorbed by both strains. No NOEL was
apparent in F344 rats; the NOEL in Sprague-Dawley rats was probably 2%
(Exxon Biomedical Sciences, 1993).
2.2 Observations in humans
In a case report, tissues samples were taken at autopsy from a
55-year old man who died from coronary failure. Small granulomas were
seen in the axillary and apical lung areas which, on microscopic
examination, showed the presence of giant macrophage cells containing
'needle-like' inclusions consisting of long-chain n-alkanes. Similar
crystalline inclusions were found in other organs, mainly in
reticulo-endothelial cells. No significant difference was found in the
amounts of neutral lipids and polar lipids in control and patient
tissues. However. analysis by GLC identified the presence of abnormal
compounds in lipid extracts from the patient tissues, which had the
same GLC retention time as n-alkanes with 29, 31 and 33 carbon chain
length. These n-alkanes were detected in most of the tissues examined
except brain, but were not detected in control organs. In the organs
studied, the highest concentrations of n-alkanes were found in lung
granuloma and in lumbo-aortic lymph nodes. Intermediate levels were
found in the liver and adrenal glands; very low levels were detected
in the aorta, myocardium and adipose tissue. The amount of alkanes
detected by GLC correlated with the histopathological findings. It was
concluded that these alkanes were characteristic of vegetable
cuticular waxes and, therefore, their source was deemed to be dietary.
The patient was known to have consumed about one kg of unpeeled apples
per day (for 18 years) corresponding to about 10 mg of n-alkanes daily
(Salvayre et al, 1988; Duboucher et al, 1988).
3. COMMENTS
At the present meeting, the Committee reviewed the results of
four recent 90-day studies in F344 rats on a range of mineral oils and
waxes representative of materials currently in use. The materials
tested were as follows:
Mineral oils
N10(A), N15(H), P15(H), N70(A), N70(H), P70(H) and P100(H).
Paraffin waxes
Low-melting-point wax (LMPW) and intermediate-melting-point wax
(IMPW).
Microcrystalline waxes
High-melting-point wax (HMPW) and high-sulfur wax (HSW).
The dose levels used were 0.002%, 0.02%, 0.2% or 2% in the diet
(equivalent to 20, 200, or 2000 mg/kg bw/day), except for the IMPW,
for which the lowest dose level was 0.02%, equivalent to 20 mg/kg
bw/day.
In these studies, neither HSW nor HMPW accumulated in any tissues
or produced any effects. The P100(H) oil produced no effects but did
accumulate in the liver to a small extent at the highest dose level.
The P70(H) oil accumulated in the liver, kidney and mesenteric lymph
nodes when administered at the highest dose level, but the only
treatment-related effect, which was seen at this level only, was
described in the study report as an increased incidence of pigmented
macrophages in the lymph nodes. With all the other materials, there
was evidence of accumulation of test material and effects indicative
of a reaction to a foreign body at one or more dose levels. The types
of effects seen were similar in nature and included focal
histiocytosis; increases in the weight of the liver, lymph nodes,
spleen and kidney; granulomas or microgranulomas in the liver;
haematological changes typical of a mild, chronic inflammatory
reaction; and biochemical changes indicative of mild hepatic damage.
These effects were similar to those seen in the two 90-day "Shell"
studies with mineral oil considered by the Committee at its
thirty-seventh meeting.
The incidence of inflammatory lesions in the mitral valve of the
heart was significantly increased in rats fed 0.2% or 2% LMPW and
occasionally in rats fed other materials. Such lesions were also seen
occasionally in control rats. The Committee considered that the
significance and treatment-related incidence of these lesions could be
clarified by a re-examination of the histological data on all treated
and concurrent control groups in the recent studies and in historical
controls.
The Committee considered that, although the types of effects seen
were essentially foreign body reactions, it was possible that a
prolonged inflammatory response of this type could result in
functional changes in the immune system and that this aspect required
further investigation.
The NOELs in these studies are given in Table 2. Except for the
P70(H) oil and those materials showing no effects, the NOELs were
based on an increased incidence of histiocytosis in the lymph nodes at
the next highest dose level. For the P70(H) oil, the NOEL was based on
an increased incidence of pigmented macrophages in the lymph nodes at
the 2% dose level.
Two of the studies included a reversal period of either 28 or
approximately 90 days, but most of the toxicological effects were
still evident at the end of this period. The Committee noted that
there was limited evidence that the severity of some of these effects
had decreased during this phase, although it appeared that a period
longer than 90 days would be required in order to determine whether
the effects produced by these materials are fully reversible.
The Committee recalled that much earlier rat studies on mineral
oils and waxes, of duration ranging from 90 days to lifetime and
utilizing strains of rat other than F344, had shown no adverse
toxicological effects. Although many of these studies had design
and/or reporting deficiencies, the lack of significant findings in
other strains suggested to the Committee that the F344 rat may be
especially sensitive. In one of the studies considered at the meeting,
the responses of F344 and Sprague-Dawley rats to P15(H) oil were
compared, and it was found that the latter were less susceptible to
the toxic effects of the test materials, although absorption and organ
weight and histopathological changes were evident in both strains.
The Committee also noted the limited data available on mineral
hydrocarbons from human studies, which showed that mineral oil-induced
lesions similar to those seen in rats have been identified in human
tissues (Annex 1, reference 102). Vacuoles of accumulated mineral oil
have been found in the liver, spleen and lymph nodes. Some studies
reported the presence of an accompanying inflammatory or granulomatous
reaction, while others reported no tissue reaction to the accumulated
material. However, none of the studies contained detailed information
about the individuals' history of use of liquid paraffin as a medicine
nor their dietary intake. It was therefore not possible to assess the
level of intake of mineral oil associated with hydrocarbon
accumulation in humans.
Food grade mineral oils and paraffin and microcrystalline waxes
are complex mixtures of hydrocarbons and other materials. It was
evident from the results of the new 90-day studies that the absorption
and subsequent toxicity of these materials are associated with their
physical properties rather than the crude oil source or refining
method. The oils and waxes with a high NOEL contain a higher
proportion of hydrocarbon components of high relative molecular mass
(high carbon number) and have higher viscosities than those with a low
NOEL, which contain a greater proportion of hydrocarbon components of
low molecular weight (low carbon number). Mineral hydrocarbons are
specified, on the whole, by means of ranges of physical and chemical
parameters. The results of the studies considered at the present
meeting were therefore applicable not only to the particular materials
tested but also to other mineral hydrocarbons having physical and
chemical parameters falling within the same ranges.
4. EVALUATION
The Committee allocated ADIs for oils and waxes as set out in
Table 2.
A temporary group ADI of 0-0.01 mg/kg bw was allocated to mineral
oils falling within the specifications for the N10(A), N15(H), P15(H),
N70(A) and N70(H) oils and a temporary ADI of 0-1 mg/kg bw was
allocated to mineral oils meeting the specification of the P70(H) oil.
The Committee requires information about the compositional factors in
mineral oils that influence the absorption and toxicology for review
in 1998. It also requires a study of at least 1 year's duration on one
of these materials in F344 rats, which should include an assessment of
immune function at appropriate time periods and an investigation of
the kinetics of accumulation of the material, and particularly whether
a plateau is reached. A reversal period of 1 year should also be
included, in order to determine whether the granulomatous hepatic
lesions observed in rats in the 90-day studies are fully reversible.
The results of this study, together with all relevant background data,
including the physical and chemical parameters of the materials
tested, should be submitted for review in 1998.
For those materials that caused no adverse effects at the highest
level tested, the Committee concluded that, although the NOEL was
derived from a study of only 90 days' duration, the dose administered
was sufficiently high to provide reassurance that accumulation of
these substances was unlikely to occur following longer-term
administration. An ADI of 0-20 mg/kg bw could therefore be established
for mineral oils meeting the specifications for the P100(H) oil and a
group ADI1 of 0-20 mg/kg bw could be established for waxes meeting
the specifications for the HSW and HMPW.
Insufficient information was available for the Committee to
re-evaluate petroleum jelly This substance should be considered at a
future meeting.
Table 2. NOELs and ADIs for mineral oils, paraffin waxes and
microcrystalline waxes tested in 90-day studies in F334
rats.
Substance NOEL ADI (mg/kg bw)
(mg/kg bw/day)
LMPW < 2 ADI withdrawn1
IMPW < 2 ADI withdrawn1
N10(A) oil 2 0-0.012
N15(H) oil < 2 0-0.012
P15(H) oil 2 0-0.012
N70(A) oil 2 0-0.012
N70(H) oil 2 0-0.012
P70(H) oil 200 0-13
P100(H) oil 2000 0-20
HSW 2000 0-204
HMPW 2000 0-204
1 Previous ADI "not specified"
2 Temporary group ADI
3 Temporary ADI
4 Group ADI
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