T h e Distribution and Storage of
Fluorine in the Tissues of
the Laying Hen*
(Received May 23, 1935)
T
HE ingestion of small quantities of
fluorides with the diet causes an accumulation of fluorine in the tissues of the
animal and ultimately chronic toxicosis.
Sonntag (1916) increased the F content of
the bones of dogs from a value below 0.3
percent in normal animals to 1.73 percent
in NaF fed animals. Brandl and Tappeiner
(1891) examined microscopically the bones
of a young growing dog and found crystals
deposited which resembled the crystalline
fluoride CaF2. Bethke, et al (1929), and
Kick (1932) have shown the increased F
storage in the bones of pigs following F intake as NaF or rock phosphate. Brandl and
Tappeiner (1891), as well as Trebitsch
(1927), further demonstrated the deposition
of F in the tissues of animals fed small
amounts of fluorine in the diet. The work
of Tolle and Maynard (1931) upon rats
showed a relatively large increase in bone
ash from normal to NaF and rock phosphate
fed animals.
Chang, Phillips, Hart, and Bohstedt
(1934) investigated the storage and deposition of fluorine in the tissues and organs of
dairy cows fed various levels of raw rock
phosphate over a long period of time. They
found fluorine in all normal tissue examined
and the greatest quantity of fluorine accompanied Ca and P deposition. The bones and
teeth contained large amounts of fluorine,
* Published with the permission of the Director
of the Wisconsin Agricultural Experiment Station.
whereas the soft tissues such as the kidney,
liver, heart, muscle, etc., contained relatively
small quantities. Fluorine in the form of rock
phosphate was added to the grain mixture,
and when the fluorine content was 0.088
percent of the grain mixture the animals
receiving the supplement showed an increase
of 16 to 24 times the fluorine stored in normal teeth and bones.
The toxic effects of fluorine and its cumulative manifestations depend upon the susceptibility of the species, the amount and
period of fluorine ingestion, and the carrier
of fluorine, i.e., the toxicity of certain forms
of fluorine has been shown to be of the following order: fluosilicates, NaF, rock phosphate, and CaF2.
It is known that various physiological
manifestations occur following the ingestion of fluorides over an extended period of
time. It is the object of this study to present the analytical results of an investigation to determine the fluorine distribution
and storage in fluorine poisoned laying hens
that had been fed various levels of fluorine
in the form of rock phosphate and phosphatic limestone.
EXPERIMENTAL
The laying hens when received were 28
to 30 months of age. They had been on
rations previously described by Halpin and
Lamb (1932) of this station. The allotment
and mineral supplements used are given below:
[154]
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KENNETH HAMAN, PAUL H. PHILLIPS, AND J. G. HALPIN
Department of Agricultural Chemistry and Poultry Husbandry,
University of Wisconsin, Madison
MARCH,
1936.
VOL.
XV,
No.
155
2
Lot I—Basal ration; Lot II—Basal ration plus
3 % bone meal (.005%F); Lot III—Basal ration
plus 1% rock phosphate (.035%F); Lot IV—Basal
ration plus 2% rock phosphate (.070%F); Lot V—
Basal ration plus 3 % rock phosphate (.105% F ) ;
Lot VI—Basal ration plus 3 % phosphatic limestone
(.040%F); Lot VII—Basal ration plus 5% phosphatic limestone (0.68%F).
TABLE 1.—Summary of data showing the storage of fluorine tn the tissues of fluorine poisoned hens. Results
given in mgms. F per 100 gs. dry material
Lot No.
No. of Hens
Per Lot
Kidney
Liver
Bone
Muscle
Fat
I
II
III
IV
V
VI
VII
8
8
9
8
8
5
6
0.65
0.88
1.45
1.58
1.58
0.61
1.93
0.60
0.90
0.89
1.27
1.55
0.69
0.94
61.25
89.99
337.44
672.58
835.28
325.53
515.78
0.46
0.60
0.69
1.00
1.23
0.48
0.70
0.09
0.09
0.14
0.15
0.18
0.09
0.14
hens of each lot. Composite samples of fat,
feathers, and thyroid were obtained for each
lot and individual samples from each hen
obtained for the liver, kidney, tibia, and
thigh muscles. The fat was separated from
connective tissue by heating in a drying oven
at 120°C. for 12 hours. The feathers were
thoroughly washed and rinsed with distilled
water. Other samples were dried in the drying room for 6 to 7 days previous to grinding and storing. The bones were crushed and
extracted with hot alcohol to remove the fat.
The samples of fat were saponified with hot
alkali in alcoholic solution previous to ashing.
Fluorine analyses were made according
to the method of Willard and Winter (1933).
This method was found to be reliable for
biological materials, when the following procedure was used. The sample was first dried
and ground. A weighed quantity in a platimum crucible was then treated with 1:4
The reagents were analyzed at intervals
and the small quantity of F present was
deducted from the titration of the samples.
The accuracy of the method was checked by
adding known amounts of fluorine to a given
sample and conducting the mixture through
the usual procedure and obtaining recoveries. Care must be observed to obtain an ash
entirely free from carbon, and the size of the
original sample must not be allowed to exceed the limit determined by all the inorganic salts remaining in solution during the
distillation.
RESULTS
The results obtained are summarized in
Table I. The presence of fluorine was demonstrated in all the normal tissues studied except the thyroid and washed feathers.
Demonstrable quantities of fluorine could
not be detected in the thyroid gland because
of the limited amount of sample. Fluorine
was not present in feathers in excess of 1
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The bone meal supplement contained 0.16
percent fluorine, the rock phosphate contained 3.52 percent fluorine, and the phosphatic limestone contained 1.35 percent
fluorine.
Samples were obtained from five to nine
Na 2 C0 3 in water solution and dried thoroughly at a low temperature on an ordinary
hot plate. The sample was ashed for 15
to 20 minutes, thoroughly moistened with
redistilled water, dried as before and again
ashed. This procedure was repeated until a
white ash was obtained. The ash was then
taken up in 1:1 HC1, transferred to a distilling flask and distilled at 125-150°C. with
H 2 S0 4 and glass beads in the presence of
H 2 0. The remainder of the method follows
the original outline of Willard and Winter
(1933).
156
SCIENCE
that of the rock phosphate of lot III, and
fluorine deposition was very similar.
Lot VII was supplemented with 5 percent
phosphatic limestone. The fluorine level of
this lot approximated the rock phosphate
level of lot IV.
DISCUSSION
The tolerance of poultry for fluorine under experimental conditions has been shown
by Halpin and Lamb (1932) to be greater
than that of rats, swine, or cattle. By comparing the results obtained from the hens to
the results of Chang, Phillips, Hart, and
Bohstedt (1934) for dairy cows, it becomes
apparent that the retention of fluorine in
the hen is decidedly less than in dairy cattle.
The inclusion of 0.088 percent of fluorine
in the grain mixture fed to the cattle in the
previously mentioned study resulted in an
increase of 16 to 25 times the normal storage
of fluorine in the bones. The bones of the
hens indicated an increase of 13 to 14 times
normal deposition with the ingestion of
0.1056 percent fluorine in the mineral supplement of the ration. The retention of
fluorine in the soft tissues was likewise comparatively greater for the cows than for the
hens.
The smaller retention of fluorine in the
tissues of chickens may be due to a reduced
absorption in the intestinal tract or to more
effective elimination. The decreased retention provides an explanation of the greater
tolerance of this species as compared to some
of the other domestic farm animals.
The relation of F level in the food to the
F content of eggs has been discussed in a
separate publication from this laboratory.
SUMMARY
All normal tissues of the laying hen, with
the possible exception of washed feathers
and thyroid, contain small amounts of
fluorine. Bones have the highest fluorine
content while the more active tissues, i.e.,
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part in 50,000. Normal bone showed an
average of 61.25 mgm. F per 100 gms. of
the dry material. Of the soft tissues examined kidney and liver contained practically
equal amounts of F. Averages of 0.65 mgm.
F for kidney and 0.60 mgm. F for liver
were obtained. The muscle contained 0.46
mgm. F and fat 0.09 mgm. F per 100 gms.
Lot II was fed a supplement of 3 percent
steamed bone meal. The bone meal contained 0.16 percent fluorine and for this
level only a slight increase in fluorine deposition and storage was indicated. The largest
deposition occurred in the bone with 89.99
mgm. F per 100 gms., while the softer tissues
showed an increase of 0.10 mgm. to 0.30
mgm. over the normal storage. No increase
was noted for F storage in the fat.
Lot III was fed a 1 percent rock phosphate supplement, and in this group a substantial increase in F storage occurred. The
bone showed an increase to 337.44 mgm. F
per 100 gm. The F storage in the soft tissues
did not rise so markedly and only a slight
increase of fluorine occurred in the body fat.
Lots IV and V were supplemented with
2 percent and 3 percent rock phosphate respectively. In lot IV the increase of storage
in the bones was approximately 11 times the
normal deposition, and in lot V the increase
was approximately 13 times normal. The
accumulation of F in the soft tissues increased from 2 to 3 times. Average results
for lot V (the highest rock phosphate level)
indicate the storage of 835.28 mgm. F per
100 gms. in the bones of the hens, accumulation of 1.58 mgm. and 1.55 mgm. F for
the kidney and liver respectively and 1.23
mgm. in the muscle. The fat deposition increased to 0.18 mgm. F per 100 gms. No
significant amount of fluorine could be
demonstrated in the washed feathers of the
hens upon this high level.
Lot VI was supplemented with 3 percent
phosphatic limestone. This supplement contained a fluorine level which approximated
POULTRY
MARCH,
1936.
VOL.
XV,
No.
REFERENCES
Bethke, R. M., C. H. Kick, B. H. Edington, and
O. H. Wilder, 1929. The effect of feeding sodium
157
fluoride and rock phosphate on bone development in swine. Am. Soc. Anim. Prod., Proc. p.
29-34.
Brandl, J., and H. Tappeiner, 1891. Ueber die
Ablagerung von Fluorverbindungen im Organismus nach Ftitterung mit Fluornatrium. Zeischr.
f. Biol. 28, 518-540.
Chang, C. Y., P. H. Phillips, E. B. Hart, and G.
Bohstedt, 1934. The effect of feeding raw rock
phosphate on the fluorine content of the organs
and tissues of dairy cows. Jour. Dairy Sci. 17,
695-701.
Halpin, J. G., and A. R. Lamb, 1932. The effect
of ground phosphate rock fed at various levels
on the growth of chicks and on egg production.
Poul. Sci. 11, 5-14.
Kick, C. R., 1932. The specific effect of fluorine in
the rations of farm animals. Ph.D. Thesis, University of Wisconsin.
Willard, H. H., and O. B. Winter, 1933. Volumetric
method for determination of fluorine. Ind. Eng.
Chem., Anal. Ed. S, 7-11.
Sonntag, G., 1915. Uber ein Verf«^hren zur Bestimmung des Fluorgehalts von Knochen und Z'ahnen
normaler und mit Fluoriden gefutterter Hunde.
Arb. Kais. Gesundh. 50, 307-336.
Tolle, C , and L. A. Maynard, 1928. Phosphatic
limestone and other rock products as mineral
supplements. Am. Soc. Anim. Prod., Proc. p.
15-22.
Trebitsch, F., 1927. Fluorgehalt der Zahne.
Biochem. Ztschr. 191, 234-241.
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kidney and liver, are much lower. Muscle
has a normal content of F below that of the
mentioned internal organs, and fat is very
low in fluorine.
Additions of raw rock phosphate or phosphatic limestone supplements to the diets of
laying hens cause increased storage of
fluorine in the tissues. The addition of
0.10S6 percent of fluorine fed as raw rock
phosphate supplement increased the storage
of fluorine 13 to 14 times the normal in the
bone and from 2 to 3 times the normal in
the soft tissues.
The introduction of fluorine containing
minerals into the rations of laying hens results in storage of fluorine which was found
to be proportional to the level fed. This was
found to hold true for both rock phosphate
and phosphatic limestone. Storage of fluorine in the bones of the chicks on the two
types of carriers was in close agreement
when equivalent quantities of fluorine were
incorporated in the ration.
2