THE USE OF RADIOACTIVE PHOSPHORUS IN TOOTH METABOLISM STUDIES OF
CARIES RESISTANT AND CARIES SUSCEPTIBLE STRAINS OF ALBINO RATS

By
RAY LOUIS SHIRLEY

A THESIS

Submitted to the School of Graduate Studies of Michigan
State College of Agriculture and Applied Science
in partial fulfillment of the requirements
for the degree of
DOCTOR OF PHILOSOPHY
Department of Chemistry
1949

ACKNOWLEDGMENT

The writer is greatly indebted to Dr. Carl Hoppert, Professor of
Biochemistry, for suggesting the problem and for much advice and en­
couragement throughout the course of the experimental work and for
assisting with the preparation of the manuscript.
To Dr. E. J. Miller, Head of the Department of Agricultural Chemis­
try, the writer is especially indebted for material help in obtaining
the radioactive tracer assay equipment used in this investigation and
for active interest in the work during the entire period of investigation.
The writer wishes to thank Dr. E. J. Benne, Professor of Agricultural
Chemistry (Research), for his friendly criticisms and suggestions through­
out the experimental work.
Thanks are due Dean R. C. Huston, Dean of the Graduate School, and to
Dr. L. L. Quill, Head of the Department of Chemistry, for advise and en­
couragement upon entering the radioactive element tracer field. The
writer appreciates the aid that Dr. Thomas Osgood, Head of the Department
of Physics, gave in procuring the radioactive phosphorus from the Atomic
Energy Commission.
Thanks are due Mr. John Wood, Research Chemist in the Bureau of
Plant Industry, Beltsville, Maryland, for kindly showing the writer the
tracer laboratory equipment used in his investigations. Thanks are due
Dr. C. L. Comar, Biochemist, Department of Agricultural Chemistry, Univer­
sity of Florida, for suggestions concerning available tracer assay equip­
ment.
The writer is especially appreciative to Mr. Robert Stipek, Graduate
Student, for supplying the breeding rats of the susceptible strain; to
Dr. H. R. Hunt, Head of the Department of Zoology, for supplying the breed
ing rats of the resistant strain; to Mr. Leo Klever, Foreman of Caretakers
Chemistry Rat Colony, for indispensable help in caring for the rats during
the investigational period; and to Mr. John Blakeslee, Radio Engineer, for
checking and replacing tubes in the Geiger-Mieller counter when necessary.

**********
********
******

****
**

*

*******************
I . Dedicated
|
* to my wife
j
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TABLE OF CONTENTS
Introduction...................................

Page
1

Facts pertinent to radioactive isotopes and the use of phosphor­
us - 32 isotope as a "tracer”.........
3
Definition of radioactive isotope.............
3
3
Types of emissions..........
Half-life...........................................
4
Types of measuring equipment...... ,...................
4
Equipment required for radioactive phosphorus assay
5
Sample preparation and measurement..................... 6
Counter measurement standardization. ......... »........ 7
Precision of measurements
.........
9
Radioactive principles pertinent toradioanalysis......... 9
Literature review of the application of radioactive phosphorus
to research on the metabolism of teeth.
.......

12

Experimental and Result s.

16

.......

General procedure............................
Weight of teeth.
.........
Per cent ash in the t e e t h . .............
Per cent phosphorus in the teeth..........
Rates of deposition and removal of radioactive phosphorus
in the teeth of rats injected intraperitoneally with the
isotope..............
Occurrence of radioactive phosphorus in the teeth of off­
spring of mothers that were injected intraperitoneally
with the isotope during pregnancy.....................
In vitro adsorption of radioactive phosphorus by the teeth

16
IB
19
20
21
24
25

Discussion.........................

27

Conclusions..................

30

Bibliography...........................

57

INTRODUCTION
In 1931-32 Hoppert et al. (1, 2) of the Department of Chemistry,
Michigan State College, published data demonstrating that dental caries
could be dereloped in rats by feeding diets containing coarsely ground
corn or rice.

Subsequently, a genetic study was undertaken with Hunt,

of the Department of Zoology, with Albino rats (rattus norvegicus) in
which two distinct strains have been developed that vary markedly in
their susceptibility to dental caries (3, 4, 5, 6).
The susceptible and resistant strains have been Inbred through
eighteen and fourteen generations, respectively.

In the case of the sus­

ceptible strain studies have been made to determine the prophylactic
value of various types of treatment both topical and dietary.

These have

included the topical application of fluorides (7), addition of fluoride
to the drinking water (8), the use of diets varying in their content of
carbohydrate and fat (9) and the introduction into the diet of urea plus
urease, and diammonium phosphate (8).
However, until the present study was undertaken, little work had
been done on the chemical constituents of the teeth.

In this investiga­

tion a comparison of the whole teeth of the resistant and susceptible
strains and stock animals has been made with regard to (l) weight of the
teeth, (2) per cent ash in the teeth, (3) per cent phosphorus in the
teeth calculated on the (a) dry weight basis, and (b) in the ash, (4) the
rate of deposition and removal of radioactive phosphorus in the teeth of
rats injected intraperitoneally with the isotope, (5) the occurrence of
labelled phosphorus in the teeth of offspring whose mothers were injected

intraperitoneally with the isotope during pregnancy, and (6) the ad­
sorption of radioactive phosphorus in vitro by the teeth of the differ­
ent strains.
The use of radioactive phosphorus in tooth metabolism studies is
founded on the assumption that stable and radioactive isotopes are chemi­
cally alike and behave in the same manner in the animal organism except
in cases where the radiations are so intense as to produce pathological
tissue changes.
The advantages of using radioactive phosphorus in tooth metabolism
studies are (1) the radioactivity makes it possible to trace the element
easily, (2) purification processes associated with direct chemical ap­
proach methods are not necessary to determine the concentration of the
isotope, (3) the element upon administration may be distinguished from
that already present, thereby making it possible to determine the rate
of upfcake by the teeth of a given dose as well as the subsequent rate of
removal of the element from the tissue.

Limitations in the use of any

radioactive isotope are chiefly the half-life of the isotope, and the
energy of the radiation.

Fortunately, of the radioactive elements none

are freer from these limitations than phosphorus-32.

Phosphorus is par­

ticularly well adapted to the study of dental tissues, since it comprises
about 17 and 12.5 per cant of the dry weight of the enamel and dentin,
respectively (10).

FACTS PERTINENT TO RADIOACTIVE ISOTOPES AND THE USE OF
RADIOACTIVE PHOSPHORUS-32 AS A ”TRACER”
A.

Definition of radioisotope (11):

A radioactive isotope is an isotope

of an element that has the property of emitting alpha or beta particles,
or gamma rays.

When an element emits such particles it undergoes a

change into another element.
two (P

For example, radioactive phosphorus thirty-

) gradually decomposes to stable sulfur as:
P^2 (15p, 17n) — > Beta particle (1.69 menr) + S^(16p, 16n).

5,

Types of emissions (11, 12):

Emissions from radioactive isotopes may

be alpha or beta particles or gamma rays.
tive electrical charge of two units.

The alpha particles have a posi­

They have two protons (from which

they derive their positive electrical charge) and two neutrons.

The alpha

particles are nuclei of helium atoms which contain two protons and two
neutrons.

These particles have very great ionizing power but very small

penetrating power.

They may be stopped by a fraction of a millimeter of

lead.
Gamma rays are uncharged and because of this they can not be de­
flected from their path by either electric or magnetic fields as can both
the alpha or beta particles.

The action of gamma rays is quite similar

to that of x-rays although the gamma rays have a shorter wave length and
a higher frequency than the x-rays which gives them great penetrating
power.
Beta particles are the same as electrons and therefore have a nega­
tive charge of one unit.

The penetrating power of these particles is

about 100 times that of alpha particles.

Radioactive phosphorus-32 which

was used in this investigation emits only beta particles.

C.

Half-life (11» 12)*

The length of time required for a radioisotope

to decay to half its original activity is known as its half-life.
half-lives of different isotopes vary greatly.

The

For example, one of

the isotopes of carbon, C-ll, has a half-life of 21 minutes whereas C-14
has a half-life of about 5000 years.
half-life of 14.3 days.

Radioactive phosphorus-32 has a

The decay rate of any isotope can be conveniently

shown by graphically plotting time against the activity remaining.
Figure 1 shows a decay curve obtained with a standard solution of radio­
active phosphorus in this investigation.

As indicated in the graph the

determined decay rate paralleled very closely the theoretical decay rate.
D.

Types of measuring equipment (11, 12, 13, 14, 15)*

Whereas measure­

ments of stable isotope concentration are normally based on the difference
in physical constants of aggregates containing the tracer, or in spectromebrlc analysis of mass ratios, the concentration of a radioactive tracer
is usually far too small to be measured by such means.

Use is made in­

stead of its emission properties due to its disintegration into other
atoms with accompanying emission of alpha or beta particles or gamma rays.
All radioactivity measuring instruments make use of the secondary
effects of the radiation.

One method of studying the distribution of

radioactive materials, known as the radioautographic method, is the direct
exposure of a photographic plate to a section of the subject in question.
Much more sensitive measurements may be made with electroscopes or
electrometers.

These instruments determine the amounts of ionisation in

a gas produced by radiations from the active materials by measuring the
conductivity of the ionized gas.

In the simple electroscope this is

accomplished by charging two relatively displacable parts alike.

Their

rate of approach to their uncharged positions is dependent on the dis­
charge rate which measures the amount of ionization.

Electrometers use

more elaborate measuring methods, gaining in sensitivity, range, accuracy,
or convenience.
'Where samples are of weak radioactivity or where the greatest pos­
sible accuracy is desired a Geiger-Mueller counter is generally used.
This instrument is sensitive to individual ionizing particles.

In con­

junction with its accompanying circuits the Geiger counter actually
counts and records the number of emitted particlee passing into its tube.
An ionizing emission in the specially gas filled counter tube will pro­
duce a number of free electrons which will be accelerated toward the
positive electrode.

Collisions between these and other gas particles

will free more electrons in "avalanche" fashion with the result that a
measureable electric pulse is produced for each emission that enters the
Geiger-Mieller tube.

Then, either because the tube is "self-quenching" by

virtue of the composition of the gas it contains or by a special quenching
circuit, the pulse is quickly extinguished and the system is ready to
record the next particle.

In this investigation a Bale-Haven dipping-type

Geiger-Mueller tube (16) was used.

This tube is of the non-self-quenching

type and a modified Neher-Harper circuit was attached to the counter by
the Tracerlab, Inc., Boston, for quenching purposes.
E.

Equipment required for radioactive phosphorus assay (13)a The neces­

sary equipment for the routine evaluation of radiotracer phosphorus in­
cludes the Geiger-Mueller tube (in this investigation a dipping-type

tube (16) supplied by the Distillation Products Corp., 755 Ridge Road,
Rochester, N. Y. was used), and a unit for supplying the operating volt­
age for the counter tube, counting the impulses from the tube, and a
timer to measure the time required for the reception of a given number
of these impulses (a compact unit is marketed by Tracerlab, Inc., Boston,
which has. these three capacities and was used in this investigation).

In

addition lead shields may be necessary to protect personnel for certain
operations with concentrated samples.
tact with the active materials.

Tongs help to avoid too close con­

The use of area monitoring equipment is

mandatory, a Tracerlab Monitor Su-3 having been employed in this investi­
gation.
F.

Sample preparation and measurement (15)*

For tracer work it is not

necessary to know the absolute value of the activity of a sample but
only the activity relative to a standard solution of the isotope.

Stand­

ard positions are necessary since the measured activity is partly depen­
dent on the absorption of the rays in the sample itself.

The same

mounting should be used to eliminate deviations due to the difference in
secondary radiations and back-scattering.

The dipping-type tube such as

was used in this investigation modified many of the geometry problems
existing for "dry” tubes.
In this investigation the teeth were ashed in a muffle furnace.

The

ash m s dissolved in 10 ml. of 2.5 N hydrochloric acid and made to volume
in a suitable volumetric flask.

A 5 ml. aliquot of the solution was

delivered to a cylinder-like cup having a flanged top and the dipping-type
Geiger-Mueller tube was then inserted into the cup in a reproducible

manner and the activity of the solution determined.
G.

Counter measurement standardization (15)s

A counter-circuit system

when operating properly should give a voltage-counting rate curve like
that shown in Figure 2.

The important range, called the ’'plateau", is

that portion between A and B in *4iich the counter registers close to
100 per cent of the ionizing particles introduced.

The operating volt­

age should normally lie on the flattest portion of the curve as voltage
fluctions will then be reflected least in the counting rate.

To increase

the lifetime of the counter the operating voltage should be at the low
voltage end of the flattest portion.

When an extinguishing circuit is

employed which is required for a non-self-quenching type of Geiger-Mieller
tube, inflections are sometimes introduced into the plateau curve.

A non­

self-quenching tube was used in this investigation and inflections were
common on the plateau of the voltage-activity curve.

However, a satisfac­

tory operating voltage was obtained without such inflections on the low
voltage side of the plateau.

In practice the plateau region was found

not to be very flat and the higher the activity of the sample the greater
was the slope with the dipping-type tube used in this investigation.

In

order to have comparable results it was found to be of the greatest impor­
tance to maintain the voltage constant for all counting operations.
Preliminary to ordering the radioactive phosphorus from Oak Ridge
the Geiger-Hueller counter assay equipment was tested for reproducibility
and precision by determining the counts per minute of 0.4, 0.02, and
0.004 H solutions of thorium nitrate.

Essentially a straight line was

obtained upon plotting activity versus concentration of thorium nitrate.

Five ml. of 0.4 N solution had an activity of approximately 1500 counts
per minute.

The counter was also demonstrated to give the same total

counts per minute for a bottle of uranium acetate and a bottle of
thorium nitrate as the sum of their individual counbs at a given dis­
tance from the Geiger-Mueller tube, thereby obtaining a value for the
resolving time of the tube.
A.

Corrections.

Measurements, as previously mentioned, were taken

with the source of radiation at a standard position relative to the
counter and were corrected for systematic variations before any attempt
was made at interpretation.

1.

The necessary corrections weret

1.

Resolving time correction.

2.

Background correction.

3.

Efficiency changes with time.

4.

Absorption and geometry corrections.

Resolving time correction*

The Geiger-Mieller counter registers

the passage of individual particles through the tube, but there is a
short interval following each registered count (resolving interval) dur­
ing which it is not able to register such emissions.

In this investiga­

tion the counter was demonstrated to count linearly with increasing con­
centration up to 5000 counts per minute.

Also, to minimise resolving

time error standard sample concentrations were chosen that approximated
the concentrations of the experimental samples.
2.

Background correction*

The background count which is due to

cosmic rays and stray radiations in the laboratory was measured under
the exact conditions of the other measurements, except with the absence

of the radioactive sample.
rate of the sample.

This value was subtracted from the counting

Throughout this investigation the background count

•was always in the range of 12 to 20 counts per minute.
3.

Efficiency changes with time:

Such changes may be corrected

for by counting a standard long-lived source regularly.

After sample

counts and standard counts are corrected for resolving time and back­
ground, their ratio is a meastire of the strength or efficiency of the
calibrated system.
such standards.

Uranium or radium sources are frequently used for

In this investigation solutions of radioactive phos­

phorus which were equivalent to the doses of isotope injected into the
animals were analyzed with the Geiger-Mueller counter at approximately
the same time as the samples of teeth containing the labelled phosphorus
thereby eliminating the necessity for decay and efficiency corrections.
4.

Geometry and absorption corrections:

For comparative studies

as in tracer investigations where absolute values are not sought geometry
and absorption errors should be cancelled by treating the samples and
standards alike during the assay procedure.

This principle was followed

in the investigation.
H.

Precision of measurements (15):

The precision of all measurements

is limited by the statistical accuracy of counting.

The reproducibility

of measurements of samples that were rechecked in the course of this in­
vestigation is snown in Table 1. Satisfactory reproducibility was
obtained on samples that were analyzed as long as 25 days apart.
J.

Radioactive principles pertinent to radioanalysis (17, 18):

equation expressing the basic law of radioactive decay is,

The

dKt
0.693 N
- —
s n— « — t»
dt
T
O
the time, t;

dNt
where — ■ is the absolute disintegrationrate
dt

at

is the number of atoms potentially radioactive which

are present at the time tj and Tg-, the half life, is that interval of
time required to reduce N^. two-fold.

This equation nay be used to

become oriented as to the amounts of radioactive material normally used
in "tracer” investigations.

By substituting Avogadro's number (6.02 X

*)9

10

fi

molecules per mole) and the millicurie count per second 37 x 10 )

into equation (1), equation (2) is obtained ass
(2)

^

= K (me) (T§-) (A).

"6 is the weight in micrograms

of radioactive substance present; (me) is the millicurie strength of the
sample; (T%) is the half-life; and (A) is the atomic or molecular weight
of the radioactive substance in question.

K is a constant whose value

depends upon the time units ofthe half-life.

For convenience, these

numerical values are tabulatedbelow:

_Num9rical_value

Half-1ife, _ expressed_in____
_seconds__min.___hrs.__ da^s^ _2££E£

K

>11
,9
,7
/6
,3
8.8x10 5.5x10 3.2x10 7.7x10
2.8x10

The following four examples illustrate the use of equation (2):
1.

"What weight of P-32 (half-life - 14.3 days) is equivalent to 1 me.?
t

/6

^ = 7.7x10 x 1 x 14.3 x 32 s 0.0035 ndcrograms.
2. A patient with hyperthyroidism is treated with 10 me. of 8 day 1-131.
What weight of iodine was given?
s 7.7 x I'd6'
"X 10 x 8 x 131 a 0.08 ndcrograms of 1-131.

3. A BaCOg (Mol. Wt. a 197) precipitate containing C-14 (half-life -

-10-

5000 yrs.) and -weighing 10 mg. gives a radioassay of 37 counts per
second which is estimated to be equivalent to about 3700 absolute dis­
integrations per second (10^ me.).

What fraction of the sample is

barium radiocarbonate?
'Z
-4
^ - 2.8 x 10 x 10 x 5000 x 197 - 0.28 micrograms.
✓5
Thus, the fraction of BaCO^ containing C-14 is 2.8 x 10 or approximately
4 molecules in 10,000 are potentially radioactive.
4.

If a 10 mg. precipitate of BaSO^ (mol. wt. z 233) contained 10 me.

of 87 day S-35, wnat fraction of the precipitate contains S-55?

,

x6

j

A

x5

- 7.67 x 10 x 10 x 87 x 233 e 1.56 x 10 micrograms.
.9
Thus this fraction is 1.56 x 10 or approximately 6 barium sulfate mole8

cules in 10

contain S-35.

-11-

LITERATURE REVIEW CF THE APPLICATION OP RADIOACTIVE
PHOSPHORUS TO RESEARCH ON THE METABOLISM CF TEETH
One of the first investigations of the metabolism of teeth in which
a radioactive element was used was that of J. Aub (19), who demonstrated
that a naturally occuring radioisotope of lead could be deposited in liv­
ing dental tissues, and that, with the passage of time, such deposits
apparently disappeared.
In 1935 Chievlts and Hevesy (20) employed radioactive phosphorus in
the first study of teeth utilizing an artificial radioactive element.
In 1937 Chievitz and Hevesy (21) added radioactive phosphorus in the form
of disodium hydrogen phosphate to the diets of rats and reported that the
phosphorus quickly found its way into the teeth.

Krogh (22) reported in

a review article that Hevesy had found that a week after an adult rat had
been given radioactive phosphorus 29 per cent of the dose was found in
the skeleton, 3.3 per cent in the Incisors, and 0.2 per cent in the molars
The ratio of radioactive phosphorus to total pnosphorus per mg. of ash of
the incisors was found by Chievitz and Hevesy (21) to be 2.5 times that
of the molars.

These investigators also reported that when the rats were

allowed to live longer the isotope decreased in the various tissues due
to its replacement by phosphorus from the diet.

Manly and Bale (23) gave

radioactive phosphorus by stomach tube to 25 rats weighing 110 to 150
grams and reported that the isotope in the growing incisors showed increas
ing amounts from the tips through the middle and root portions.

These

investigators found that the incisors contained from 2.0 to 3.5 per cent
of marked phosphorus 4 hours after receiving the dose, 4 to 7 per cent two

-12-

days later, and increased slowly up to B per cent in 20 days, the results
being calculated on the basis of per cent dose found per gram of ash in
the teeth.

In contrast the molars contained only 1 per cent of the dose

in 4 hours after receiving the isotope and remained fairly constant from
10 hours up to 8 days after which time the percentage of labelled phos­
phorus decreased slowly.

At the end of 20 days approximately two-thirds

of the maximum amount of labelled phosphorus was still present in the
molars.

The rate of deposition of the radioactive phosphorus during the

first day was about 0.07 per cent of the total phospnorus present, or
about one-tnlrd that deposited in the incisors in the same period.
Hevesy (24) demonstrated that labelled phosphorus is removed rapidly
from the olood of rats.

Only two per cent of the dose remained in the

plasma two hours after injection of the isotope, while 92 per cent was
recovered in the calcified tissues.

Hodge at al. (25, 26), suggested

that the greater part of tne radioactive phosphorus going into the bones
and teeth was probably due to the natural affinity of the apatite struc­
ture for phosphate.

In equilibrium the phosphate content of bone is

1000 times that of the plasma (27).
Manly and Bale (23) determined the retention of labelled phosphorus
in the molars and incisors of 28 rats whose teeth were extracted with
an ethylene glycol solution of 3 per cent potassium hydroxide.

The re­

sults agreed well with previous studies made by Hevesy et al. (28) on
unextracted teeth, thereby indicating that the exchange phenomenon took
place in the inorganic constituents of the teeth.
After consideration of the available evidence from histological,
chemical, and physical studies Chase (29) in 1951 concluded tnat enamel

13-

nust be na lifeless, inert, mostly inorganic substance".

However, since

then radioactive phosphorus nas made possiole a new approach in the
study of this dental tissue.

Hevesy and Armstrong (30) reported tnat,

in vitro, the exchange of labelled phosphorus in the enamel was I qbs
than 10 per cent that in the dentin.

Volker and Sognnaes (31) reported

that some of the inorganic phosphorus in the enamel apparently could be
derived from the saliva.

These workers (32) demonstrated that the sur­

face enamel of cats retained on the average, three times as much marked
phosphorus as the remaining enamel when the isotope was administered
through a stomach tube.

The distribution of the isotope in the various

density fractions of the whole crowns of the teeth of a dog showed a
direct relationship between the density and the concentration of isotope
in the enamel, but an inverse relationship in the dentin.

The surface

enamel of the teeth of a dog also contained the greatest deposit of
labelled phosphorus.

Yfhen the enamel was protected from the saliva by a

plaster covering a reverse in the density-phosphorus-32 relationship was
obtained.

It thus appeared that the surface of the enamel adsorbed

appreciable amounts of inorganic phosphorus from the saliva.
Barmrn and Armstrong (33) covered various parts of human teeth with
collodion and exposed them in vitro to a solution of radioactive phos­
phorus and found that the phosphorus reached the enamel through the.-dentin
and also reached the dentin through the enamel.

Inasmuch as the quantity

passing through the enamel was relatively small compared to that which
passed through the dentin they postulated, that, in vivo, more phosphate
reached the enamel through the blood circulation and the pulp than was

absorbed from the saliva.

They found that the ratio of the activity

of 1 gram of enamel to that of 1 gram of dentin ■was 0.07.

On the basis

of the specific activities of the dental tissues* the rate of phosphorus
exchange in the enamel was approximately 5 per cent of that in the den­
tin.

Armstrong and Barnum (34) gave a male rat* weighing 238 grams* by
6
stomach tube 8 ml. of a solution containing about 0.6 x 10
counts per

minute of calcium-45 isotope and 9 x 10® counts per minute of phosphorus32 isotope and determined the concurrent uptake and retention of both
elements by the teeth.

They found that the uptake and retention of both

the radiocalcium and radiophosphorus stood in the following decreasing
order*

incisor dentin* incisor enamel* molar dentin* and molar enamel.

Pederson and Schmidt-Nielson (35) were unable to demonstrate any
appreciable concentration of radioactive phosphorus in cement-capped
enamel surfaces of several teeth from two young girls who had been given
a dose of the isotope several days previous to the extraction of the
teeth.
Manly and Levy (36) fed labelled phosphorus in the form of disodium
hydrogen phosphate to 31 experimental rats.

One group was mated 2 days

after administration of the dose* and the other group 7 days afterwards.
The data obtained indicated that when the mothers were on an adequate
diet no significant exchange of inorganic phosphorus took place between
her skeleton and the offspring.

EXPERIMENTAL AND RESULTS
General procedure:

In this investigation a comparison was made of the

whole teeth of resistant and susceptible strains and stock Albino rats
with regard to (1) weight, (2) ash content, (3) phosphorus content ex­
pressed as per cent of (a) dry weight of the teeth and (b) of the ash,
(4) rates of deposition and removal of radioactive phosphorus-32 in the
teeth after intraperitoneal injection of the isotope, (5) occurrence of
radioactive phosphorus in the teeth of offspring whose mothers were in­
jected intraperitoneally with the isotope during pregnancy, and (6) the
adsorption of radioactive phosphorus in vitro by the teeth of the differ­
ent strains.
Three rations were used in this investigation.

They were designated

as fine rice, stock, and pnas, respectively, and consisted of the follow­
ing ingredients:
Stock

Fine rice
rice* - 66%
milk powder - 30%
alfalfa meal - 3%
salt - 1%

corn meal - 35%
ground wheat - 25%
w, milk powd, - 20%
linseed oil meal - 1.0%
alfalfa - 6%
brewers yeast - 3%
salt - 1%

Anas
powder, milk
alfalfa meal
salt - 1%

- 79%
- 20%

*The rice was ground so that 2 per cent was held on a 20 mesh
screen and 40 per cent on a 40 mesh screen.
The above rations gave the following results when analyzed for per
cent moisture, ash, and pnosphorus according to the recommendations of
the A,0.A.C, Methods of Analysis:

-16-

Constituent

Fine rice
%

Stock
%

Pmas
%

9.80
4.42
0.37

9.56
4.97
0.47

5.50
9.10
0.66

'

moisture
ash
phosphorus (dry basis)

Approximately 224 rats of the resistant and susceptible strains
and stock colony were compared in the age range of 6 to 60 days in re­
gard to weight of teeth, per cent ash in the teeth, per cent phosphorus
in the teeth calculated on the basis of the dry weight and in the ash.
An individual member of each litter used in the investigation was iso­
lated, fed the fine rice ration, and observed for development of caries
in order to check if the various strains were responding as expected.
The stock colopy rats appeared to be more like the susceptible than the
resistant strain in susceptibility to dental caries.

Rats of the suscept­

ible strain would almost invariably have small cavities developing at 60
days of age and for this reason older rats were not used in this investiga­
tion.

Unpublished data obtained by Hoppert et al. show that the stock

ration is less cariogenic than the fine rice ration, and that the pmas
ration is essentially non-cariogenic.

Both of the developed strains and

the stock rats were compared on the fine rice ration in the age range of
6 to 60 days.

The susceptible strain and stock rats were also compared

on the stock ration in the age range of 6 to 21 days.

A few rats of the

susceptible strain were fed the pmas ration.

The resistant strain was

maintained entirely on the fine rice ration.

The fine rice diet was also

used in all of the tracer studies.

-17-

To obtain the teeth for analysis the rats were killed with ethyl
ether, and the teeth removed from the jam and cleaned of all tissue
with a scalpel and cheesecloth.

The pulp ^

removed from all incisors,

but in the case of the molars the pulp was removed only in rats 20 days
of age or younger.

Although the teeth do not erupt in young rats until

about the 14th to 16th day, eight molars and the four incisors were re­
moved from rats ranging from 6 to 14 days.

In the case of older rats

the four back molars were also included.
The various phases of this investigation listed above will be pre­
sented in turn.
Weight of teeth*

The molars and incisors were dried at least 2

hours (to constant weight) in a hot air oven at 103° C.

The data obtained

with the susceptible and resistant strains and stock rats on the fine rice
ration are presented in Table 2.

Figures 3 and 4 present in graphical

form the data obtained with the molars and incisors, respectively.

The

molars increased in weight linearly up to 20 days of age after which time
the rate of growth gradually decreased, whereas the incisors grew linearly
throughout the investigational period i.e. 6 to 60 days.

The weight of

both the molars and incisors of the resistant strain tended to be slightly
lighter in weight after 30 days of age than those of the susceptible
-strain and stock colony rats.
Thirty rats of the susceptible strain and 19 rats of the stock
colony whose mothers were maintained on the stock ration were investi­
gated in the age range of 6 to 21 days for rate of tooth growth and were
found to give comparable results to rats on the fine rice diet.

Per cent ash in the teebhi

(1) The per cent ash '■as obtained by
o

heating the dried teeth in a muffle furnace overnight at 600

C.

The

data obtained with the susceptible and resistant strains and stock rats
on the fine rice ration at 6 to 60 days of age are shown in Table 2.
Figures 5 and 6 present in graphical form the data obtained on the molars
and incisors* respectively.

The per cent ash in the molars increased

rapidly from approximately 40 per cent at 6 days of age to approximately
77 to BO per cent at 20 days of age after which time the values tended to
be constant throughout the period of observation.

The per cent ash in

the incisors increased from approximately 60 per cent at 6 days of age
to approximately 75 per cent at 20 days after which time it remained
quite constant.

Table 3 shows that the average concentration of ash
*

♦

in the molars from 20 to 60 days of age was 76.7 " 0.9* 78.8 - 0.9* and
*

77.6 " 1.0 per cent for the susceptible* resistant* and stock colony
groups* respectively.

The incisors of the susceptible strain contained

approximately 0.6 per cent less ash than those of the resistant and stock
rats.
(2) The per cent ash in the teeth was also obtained by extracting
the organic matter with an ethylene glycol solution of 3 per cent potas­
sium hydroxide at the boiling temperature according to the procedure of
Crowell et al. (37).
Table 4.

The data obtained by this technique is shown in

The per cent ash was found by this extraction procedure to be

approximately 5 to B per cent higher than by miffle furnace ashing at
C. However* the resistant and susceptible rats of the same age
gave similar values by the extraction method in both molars and incisors.

-19-

Per cent phosphorus in the teeth:

The phosphorus in the teeth was

determined by the volumetric phosphomolybdate method (38).

The ash resi­

dues of the teeth were used for the phosphorus analysis and the results
were calculated on the dry weight basis of the teeth and also on the
basis of the phosphorus in the ash.

The susceptible and resistant strains

and stock animals were compared on the fine rice ration from 6 to 60 days
of age and the data obtained is shown in Table 2.

The data are presented

graphically for the molars and incisors in Figures 7 and 8, respectively,
in which the per cent phosphorus was calculated on the basis of the dry
weight of the teeth, and in Figures 9 and 10, respectively, in which the
per cent phosphorus was calculated on the basis of the ash content of
the teeth.

Figure 7 shows that the per cent phosphorus in the molars on

the dry weight basis increased from 8 to 10 per cent at 6 days of age to
approximately 14 to 15 per cent at 20 days of age at which time the val­
ues became constant.

Figure 8 indicates that the per cent phosphorus in

the incisors on the dry weight basis followed closely the pattern of the
molars.

Figures 9 and 10 show that when phosphorus was expressed in

terms of the per cent of the ash, age

appeared to have little influence.

The somewhat greater variations found in the

case of the very young

animals may have been due to the inherent difficulties in the preparation
of comparable samples of dental tissue.
eight molars of 6 day old rats was as

The aggregate weight of the
littleas 5 mg. in some

cases.

As shown in Table 3 in which average phosphorus values are given for
the age range of 20 to 60 days, the molars of the resistant strain con♦

tained 15.5 “ 0.4 per cent calculated on the dry weight basis as compared

-20-

to 15.0 * 0.2, and 15.1 “ 0.4 per cent for the susceptible and stock
colony rats, respectively.

However, no significant difference was demon­

strated between the 3 groups for phosphorus in the incisors on this basis,
nor in the phosphorus content of the ash of either the molars or incisors.
Thirty-three rats of the susceptible strain and 19 rats of the stock
colony maintained on the stock ration were sacrificed in the ago range of
6 to 21 days, and 14 rats of the susceptible strain on the pmas ration
that were sacrificed in the age range of 40 to 100 days, gave phosphorus
values comparable to those found above in the case of rats fed the fine
rice ration.

Also, a 167 day old resistant rat kept on the fine rice

ration gave similar phosphorus values.

The data obtained indicate that

the phosphorus content of the ash of the teeth is relatively independent
of age, sox, heredity, and diet.
Rate of deposition in and removal of radioactive phosphorus from the
teeths

This phase of the investigation of the role of phosphorus in the

teeth of the specialised strains and stock rats involved two shipments of
radioactive phosphorus from Oak Ridge, Tennessee, after allocation from
the Atomic Energy Commission.

The amount received each time was 1 ml. of

an aqueous solution of phosphoric acid containing 3 millicuries of the
radioactive isotope.

The 1 ml. of solution was diluted before use to

approximately 150 ml. with distilled water, sufficient sodium chloride
added to give a 0.9 per cent solution, and the pH adjusted to approxi­
mately 7.5 with sodium hydroxide.

On the basis of the Oak Ridge assay

report 1 ml. of the diluted solution, hereafter designated as "stock
solution", should have contained approximately 15 to 20 microcuries of

>21-

the isotope and was found to have an activity of approximately 1,600,000
counts per minute on the dipping-type Geiger-Mieller tube used in this
investigation.

The activity determination was made by diluting 1 ml. of

the stock solution plus a few drops of concentrated nitric acid to 2000
ml. with distilled water and using a 5 ml. aliquot for analysis.
One ml. of the stock solution was injected into the peritoneal
cavity of resistant, susceptible, and stock rats at different ages i.e.
at 20, 24, 29, 32, and 35 days of age.

At intervals of a few hours up

to 40 days after injection of the isotope the animals were sacrificed and
the teeth removed.

The molars and incisors were separated, dried at 100°

C., and ashed in a nuffle furnace at 600° C.

The ash was then dissolved

in 10 ml. of approximately 2.5 N hydrochloric acid and transferred to a
suitable volumetric flask from which aliquots were taken for activity
measurements and analysis for total phosphorus.
The data for the weight of teeth, per cent ash, and per cent phos­
phorus is shown in Table 3 (see column headed nage P®5* injected" for
guide to rats that were injected with labelled phosphorus).

The data

obtained for the deposition and removal of the injected radioactive phos­
phorus is presented in Table 4.

The results are expressed in terms of

per cent of injected dose found, per 100 mg. of ash in the teeth, at the
time of sacrifice of the animals.

Figures 11 and 12 show graphically

the results obtained for molars and incisors, respectively, on the resis­
tant and susceptible strains and stock rats that were injected with the
isotope at 20 days of age.

Figures 13 and 14 show graphically the results

for molars and incisors, respectively, on rats that were injected at 32 to

-22-

35 days of age.

Rats of both specialized strains and stock rats rapidly

deposited the isotope in the teeth, approaching a maximum percentage of
the injected dose at 24 to 48 hours after injection.
As shown in Figure 11, the molars of the 20 day old rats deposited
approximately 4 to 7 per cent of the injected isotope at 24 to 48 hours
after receiving the dose.

The per cent deposition increased slightly

up to 15 days, after which time the concentration began to decrease.
Figure 12 shows the per cent of phosphorus-32 found in the incisors of
the rats injected with the isotope at 20 days of age.

These animals had

a deposition of approximately 5 to 9 per cent of the injected isotope in
the incisors within 24 to 48 hours after receiving the dose and tended to
increase slightly up to 5 to 10 days after which time the concentration
decreased gradually to 3 to 4 per cent after 30 to 40 days.
Figure 13 shows graphically the data obtained on the molars of rats
that were injected with the isotope at 32 to 35 days of age.

Both the

specialized strains and stock rats had approximately one per cent of the
dose deposited in the molars by 24 to 48 hours following the injection.
The per cent deposition was relatively constant at 1 to 1.5 per cent
throughout the range of 5 to 15 days after the isotope was administered,
after which time the concentration decreased.

Figure 14 presents graphi­

cally the data obtained on the incisors of rats that were injected with
the isotope at 32 to 35 days of age.

The incisors of the resistant strain

contained slightly more of the isotope than those of the stock and sus­
ceptible strains.

The incisors in general deposited 2 to 3 times more

radioactive phosphorus than the corresponding molars.

-23-

The 35 day old

rats deposited only a third to one-half as nuch of the isotope in their
molars and incisors as the 20 day old rats.
Radioactive phosphorus found in the teeth of offspring of mothers
that were administered the isotope during pregnancy*

In this experiment

pregnant females of the resistant and susceptible strains and stock rats
were injected intraperitoneally during pregnancy with 5 ml. of the stock
isotope solution which contained approximately 90 to 100 microcuries of
radioactive phosphorus.

The offspring were then sacrificed at 6, 9, 14

and 20 days of age and the activity in the molars and incisors determined
after drying at 100° C. in a hot air oven, ashing at 600° C. in a muffle
furnace overnight, and dissolving in dilute hydrochloric acid.

Two females

of each of the specialised strains and stock rats were injected with the
isotope.

However, one resistant strain mother and one stock colony mother

destroyed all of their offspring, whereas the other stock rat mother des­
troyed all but one and the second susceptible strain mother destroyed all
but three of their offspring.

The data obtained are shown in Table 5.

These data indicate that the offspring were normal in regard to weight of
teeth, per cent ash, and per cent phosphorus in the teeth at the age that
they were sacrificed.

An appreciable amount of activity was found in the

teeth of all offspring.

However the longer the gestation period after

the injection of the isotope the lower the percentage found in the teeth
of the offspring.
Fortunately, a comparison of the susceptible and resistant strains
was made possible by the birth of a litter of each type two days after
the females were administered the isotope.

-24-

The data obtained with these

two litters are shown graphically in Figures 15 and 16, for molars and
incisors, respectively.

The per cent of injected isotope found per 100

mg, of ash in the molars of both strains decreased from approximately
5 per cent at 6 days of age to approximately 2,1 per cent at twenty days
of age.

In the case of the incisors the percentage of dose found

decreased from approximately 6 to 7 per cent at 6 days of age to 2,4 per
coot at 20 days of age,

A representative line drawn through the points

on these graphs would be essentially identical for both strains.
In vitro adsorption of radioactive pnosphorus by the teethi

In this

experiment the teeth of the susceptible and resistant strains and stock
colony rats that had been on the fine rice ration were investigated with
regard to in vitro adsorption of radioactive phosphorus.
40, and 60 days of age were used for the study.
similar to that used by Hodge et al, (25),

Hats at 20, 30,

The procedure used wa6

The teebh were extracted with

an ethylene glycol solution of 3 per cent potassium hydroxide to remove
the organic matter.

After filtering the extracting solution the teeth

were ground to medium fineness, and agitated for B hours in 25 ml, of a
neutral aqueous solution of radioactive sodium hydrogen phosphate at 40° C,
The radioactive solution had an activity of approximately 2000 counts per
minute per 5 ml. as determined on the dipping-type Geiger-Mueller tube
used in this investigation.

After exposure the teeth were wasned by de­

cantation and transfer to a Selas crucible with 5 washings of 25 ml. of
distilled water at 40° C.

The ground dental tissue was then dissolved

in approximately 2.5 N hydrochloric acid, diluted to volume in a volu­
metric flask, and an aliquot taken for measurement of the activity

-25-

adsorbed from the radioactive phosphorus solution.

In the case of the

40 day old rats the molars were equally divided and half were ashed in
the nuffle furnace at 600° C. and the capacity of the ash to adsorb
radioactive phosphorus was determined as above.

The data obtained in

these in vitro studies e c t b presented in Table 6.

The teeth of the 60 day

old rats adsorbed appreciably less phosphorus than those of the younger
rats i.e. both the molars and incisors adsorbed radioactive pnosphorus
equivalent to 20 to 30 counts per minute per mg. of ash, whereas the
teeth of the 20, 30 and 40 day old rats adsorbed sufficient isotope to
give 50 to 60 counts per minute in the case of the molars, and 70 to B0
counts per minute in the case of the incisors.

The teeth of the 40 day

old rats that were ashed in the nuffle furnace adsorbed only enough iso­
tope to give approximately lb counts per minute per mg. of ash, in the
case of both the susceptible and resistant strains.

- 26-

DISCUSSION

Very little chemical •work has been done on the teeth of the caries
resistant and caries susceptible strains of Albino rats previous to this
investigation*

Whole teeth only were used in this study in order to

determine whether gross chemical differences existed between the strains.
Circumstances did not permit a separation of the enamel and dentine for
individual study altho it is recognised that more detailed and valuable
information would have been obtained by so doing.
A few ash determinations made by extracting the organic matter from
the teeth with 3 per cent potassium hydroxide in ethylene glycol gave 5
to 6 per cent higher values than those obtained on corresponding teeth
ashed at 600° C.

Bowes and Murray (39) found that ashing at high tempera­

tures decomposed all the combined carbon dioxide in the dentine and about
75 per cent of that in the enamel.

They reported that dry dentine and

dry enamel contained 3.16 and 1*95 per cent of combined carbon dioxide,
respectively.

These investigators also reported that extraction methods

remove approximately 0.4 per cent of the phosphorus as organically com­
bined phosphorus. Further work is obviously necessary to determine what
part of the molars of the specialized strains is responsible for the
small differences in ash and phosphorus encountered in this investigation.
The data obtained in this study indicated that the phosphorus content of
the ash in the whole teeth of the specialized strains and stock rats is
independent of age, sex, and heredity.
The variation in the capacity of members of the same strain to
deposit and remove radioactive phosphorus from the teeth indicates that

-27-

small differences between the strains cannot be determined easily.

The

apparent biological variation encountered in this study is of the same
order as that obtained by Manly and Bale (23) when they gave radioactive
phosphorus by stomach tube to Albino rats and determined the rate of turn­
over of the isotope in the teeth.

In studies of the absorption, distribu­

tion, and excretion of labelled phosphorus in the rat Cohn and Greenberg
(40) administered the isotope by stomach tube and by intraperitoneal in­
jection.

They found both techniques to give appreciable variations in

the isotope content of the various tissues of the body.
The experiment, in which the two specialised strains were compared
in regard to the occurrence of labelled phosphorus in the teeth of the
offspring of mothers that had been administered the isotope during preg­
nancy, indicates that the early phosphorus metabolism of the teeth of the
two strains is very similar.

Sognnaes and Volker (32) demonstrated that

the enamel and dentine of the unerupted teeth of a monkey adsorbed the
same amount of radioactive phosphorus.

Since the teeth were removed from

the young rats in the present experiment in the pre-erupted stage or soon
afterwards, it might be assumed that the enamel and dentine of the two
strains are equivalent in their capacity to assimilate phosphorus at
least in the early period of their tooth development.
Eodge et ai. (25) found that Albino rat teeth adsorbed radioactive
phosphorus in vitro from an aqueous solution in accordance with the
Freundlich adsorption isotherm.

Subsequently, Hodge et al. (41) showed

that when enamel and dentine were exposed to radioactive phosphorus in
vitro fairly rapid adsorption of the isotope occurred in each substance

-28-

over a period of at least 8 hours. For this reason, the teeth were
exposed to the labelled phosphorus in aqueous solution for 8 hours in
the in vitro adsorption tests made in the present investigation.

CONCLUSIONS
The data obtained in this investigation of the whole teeth of two
strains of Albino rats xhat are markedly different in their susceptibil­
ity to dental caries indicate that they vary slightly in the following
respects:
1* Weight of teeth.
2.

Per cent ash in the molars.

3.

Per cent phosphorus in the molars.

They were found to be essentially similar in the following re­
spects:
1.

Per cent ash in the incisors.

2.

Per cent phosphorus in the incisors on
the dry weight basis, and per cent phos­
phorus in the ash of the molars and incisors.

3.

Rate of deposition and removal of radio­
active phosphorus in the teeth when the iso­
tope was administered intraperitonoally.

4.

Occurrence of radioactive phosphorus in the
teeth of offspring of females that were in­
jected intraperitonoally with the isotope
during pregnancy.

5.

In vitro adsorption of the isotope by the
teeth from aqueous solution.

-30-

Table 1. Data showing reproducibility of assay technique for rechecking
per cent radioactive phosphorus in acid solutions of rat teeth ash.
designation
of teeth

Note*

1st analysis
date
% P32

2nd analysis
date % P32

difference
days
% P

Ml

7/11

0.88

8/5

0.89

25

-0.01

2

7/12

0.66

8/5

0.70

24

-0.04

3

7/24

1.16

8/5

1.U9

12

-0.07

4

7/27

0.94

8/9

1.00

13

-0.06

5

7/29

0.76

8/9

0.85

11

-0.09

12

7/23

0.94

8/5

0.87

13

-0.07

13

8/9

1.02

8A2

0.86

3

-0.16

16

7/11

0.95

8/5

0.94

25

-0.01

17

7/12

0.96

8/5

1.08

24

-0.12

18

7/27

1.81

8/9

1.90

13

-0.09

19

7/30

2.42

8/9

2.37

10

-0.05

28

7/23

2.01

8/12

2.15

20

-0.14

29

7/28

2.77

8/12

2.85

15

-0.08

39

7/25

2.00

8/6

2.18

12

-0.18

41

7/25

2.05

8/12

2.03

18

-0.02

42

7/31

4.34

BA2

4.37

13

-0.03

43

8/4

4.44

B/12

4.59

8

-0.15

44

8/6

1.75

B/L2

1.74

6

-0.01

47

8/2

1.70

8/7

1.83

5

-0.13

48

8/7

1.84

8/12

2.03

5

-0.19

51

b/ii

2.79

8/12

2.72

1

-0.07

The above values for % F32 were calculated on the basis of per
cent injected dose found in the teeth.

—31—

Table 2. Weight of teeth, per cent ash, per cent pnosphorus in the teeth, and
per cent phosphorus in the ash of the teeth of the resistant and susceptible
strains and stock rats on the fine feed ration in the age range of 6 to 60 days.
age
Age rat
wt.of
Tit. of teeth
% ash
tfo phosphorus
strain inject, sacrific. rat sex
(mgs)
dry basis
in ash
(days)
(days)
(gas)
mol.
inc. mol. inc. mol, inc. mol, inc.
susc.

tt
tt
tt
tt
it
tt
it
M
tt
It
tt
tt
it
it
tt
tt
«

n
tt
tt
it
tt
it
tt
tt
tt
tt
tt
tt
tt
n
tt
tt
tt

tt
it
tt
tt
tt
tt
tt
tt
tt

-

mm
-

mm
-

-

-

-

am

-

20
20
20
20
20
20
-

-

29
20
20
35
33
-

29
-

20

6
6
6
6
9
9
9
9
9
10
10
10
14
14
14
19
20
20
20
20
20
21
22
25
25
30
30
30
30
30
30
32
34
35
35
37
36
39
39
40
40
40
40
40

lo

15
12
15
17
25
15
13
13

M
F
M
F
F
M
M
M
M

-

-

-

-

11
2e
35
22
39
35
32
46
30
37
37
39
55
50
70
75
77
62
66
70
83
91
98
70
97
96
100
110
127
103
79
119
70

M
F
M
M
F
M
F
M
F
F
F
F
M
M
F
M
F
F
F
F
F
M
M
M
M

F
F
M

F
M
M
F

9.5
6.9
5.6
6.9
14.7
17.7
15.2
11.8
12.1
15.4
15.9
11.5
33.5
34.5
27.5
54.8
48.2
60.7
66.5
35.4
61.2
64.3
70.8
77.4
84.3
92.8
91.1
83.2
84.5
89.3
65.6
80.4
78.2
95.2
100.0
104.2
103.9
95.9
102.0
100.1
102.1
90.5
-

95.8

6.5 48.4
4.9 43.5
4.6 41.1
5.2 55.0
9.9 57.2
9.4 56.5
9.0 54.6
7.7 50.9
7.6 52.1
9.0 67.5
9.4 67.3
7.6 50.4
22.4 74.6
21.1 72.5
18.9 69.9
35.4 77.4
29.6 78.2
38.3 76.9
44.9 77.9
25.6 78.3
42.5 77.3
45.4 78.4
51.6 77.1
61.1 77.1
61.6 76.3
70.6 75.7 76.3
61.1 79.0
76.0 76.9
71.9 77.6
67.8 76.4
62.7 76.9
63.5 77.8
62.2 77.2
85.0 76.1
102.0 77.3
95.4 •76.3
90.6 75.8
94.3 77.5
96.9 77.7
95.9 75.7
88.3 75.8
108.4 mm
97.0 75.7

61.5
55.1
54.4
46.2
68.7
58.5
66.7
62.3
64.5
83.3
85.1
63.2
77.7
73.5
69.5
73.7
72.6
73.9
74.8
75.2
74.4
74.2
75.7
74.8
75.3
-

73.4
77.9
77.6
76.6
74.9
74.5
75. e

76.5
75.9
77.4
75.9
74.4
76.4
77.6
74.9
75.3
75.8
77.3

10.1 13.3
-

mm

-

10.3 14.1 19.1
12.1 15.4 21.1
-

10.9
10.8
10.9
12.0
12.2
10.8
14.4
14.7
-

15.1
15.2
15.1
15.1
16.0
15.0

21.6
-

30.6
22.4
14.6 20.0 21.9
13.5 21.2 21.7
14.1 21.0 21.9
15.3 17.7 18.4
15.7 18.1 18.5
13.7 21.4 21.7
15.4 19.3 19.8
17.9 20.3 24.7
14. 6 19.5 21.1
15.0 19.5 20.6
15.0 19.6 20.3
15.1 19.4 20.1
16.1 20.5 21.4
15.1 19.4 20.3

-

-

-

-

-

-

-

-

-

-

-

-

15.0
15.4
13.1
15.3
13.1
15.1
-

-

14.4 14.6
•

13.4
13.2
15.2
13.2
14.9
15.0
14.3

-

13.4 19.7 21.1
13.7 19.5 20.2
15.9 19.6 20.5
15.8 19.7 20.6
15.6 19.8 20.8
13.6 19.6 20.7

-

-

18.9
-

-

19.5
-

16.1 20.2 21.3
13.5 20.0 20.8
13.8 19.6 20.4
15.8 19.5 20.3
13.4 19.7 20.6
13.3 18.6 20.3
•
15.9
21.0
15.0 18.9 19.4

Continued next page

-32-

20.9

(Table 2 continued)____________________________ ____________
age
age rat wt.of
vrfc.of teeth $ ash '
% phosphorus
strain inject, sacrific. rat seoc
(mgs)
dry basis
in ash
______ (days)
(days)
(gins)
mol.
inc. mol, inc. mol, inc. mol, ino.
BUSC.
«

tt
It
tt
tt
tt
tt
tt
tt

33
_
•
_

35
35
-

2°
-

tt

_

H

mm

tt

-

It

20

resist.
n
tt
n
it
•t
N
It
It
It
II
W
11
tt
«
It
II
It
It
It

mm
mm

—
—

&
•
—
mm
mm
mm
mm

•
mm
mm

—
-

19
19

It

tt
It

tt
It
tt

II
II
II

20
20
20
20
20
20
20
25

'

43
49
49
50
50
50
50
55
55
57
€0
60
60
60

134
145
135
133
136
132
140
132
70
163
146
155
147
85

F 108.4
90.1
H
F 117.4
F 112.6
M 108,8
F 112.0
F 97.7
F 113.7
II 91.8
F 130.5
F mm
F 125.6
F 121.7
M 105.7

123.3
106.9
112.8
135.5
120.6
136.7
139.0
153.3
105.1
168.9
156.4
167.4
I72i3
124.3

5
5
5
6
6
6
9
9
9
9
10
10
10
10
13
14
14
19
19
19
20
20
21
21
21
22
25
25
26

7
7
7
8
10
10
7
12
6
18
17
12
16
17
20
20
22
31
25
22
35
31
28
27
30
-

2.5
2.1
2.4
5.7
F
5.2
M
5.0
M
8.0
F
M 11.5
6.7
M
F 15.9
F 16.8
M 14.7
M 14.5
F 15.1
M 26.9
M 29.4
F 25.4
F 49.1
M 43.2
M 41.3
F 46.6
M 56.5
F 57.8
F 55.4
M 67.4
F 62.4
F 68.3
M 51.0
F 76.3

2.1
2.3
2.2
3.3
4.6
3.3
5.0

-

•

_

3.2
10.5
10.2
8.1
8.8
6.6
16.7
19.3
17.4
33.8
27.3
25.1
34.4
39.4
39.2
39.8
44.2
45.7
51.9
38.8
61.1

'

75.4
76.4
76.7
76.3
77.2

'
14.7 is;6
15.1 15.9 20.0
15.0 15.5 19.7
15.0 16.0 19.5
14.8 15.4 19.3
14.9 15.2 19.3
16.1 19.9
15.1 15.8 20.2
14.3 15.1 19.1
15.1 15.6 18.7
15.7
15.0 15.7 20.0
14.8 15.7 19.5
14.3 15.1 19.0

75.3
75.5
75.1

76.2
75.3
78.7
76.2
76.5
78.5
75.7
76.3
75.3
76.7
75.6
75.8
76.4

40.0
23.8
29.2
43.9
42.4
36.0
48.8
52.2
47.8
58.0
54.8
53.7
52.4
53.0
70.2
72.5
72.2
78.8
78.3
78.7
79.6
80.3
79.3
80.0
78.5
79.5
78.5
77.8
79.6

57.1 14.4 13.3 36.0
52.2 8.2 11.0 34.4
45.5 8.8 12.0 30.3
63,6 6.5 14.3 19.4
58.7 42.5 66.0 9.6 12.6 19.7
20.0
10.4
63.6 10.6 16.5 22.3
71.5 11.9 14.5 20.6
68.6 10.5 13.7 19.2
66.7 10.5 14.1 19.4
64.8 11.0 13.9 21.0
62.1 10.9 13.8 20.7
73.4 14.2 15.4 20.2
74.6 14.5 15.4 20.1
70.7 73.7 15.4 15.1 19.4
mm
72.0 73.7
76.5 15.8 16.0 18.9
77.2 15.3 14.2 18.7
77.1 15.0 14.5 18.9
77.9 15.0 14.5 18.7
76.0 76.4 75.5 73.7 76.7 -

74.7
75.1
76.1

Continued next page

-33-

- ’

20.8
20.3
20.3
20.2
19.9
20.5
20.8
19.8
20.8
20.o
20.7
20.7
19.8
23.3
21.0
26.4
22.5
mm

19.0
25.9
20.3
20.0
21.2
21.4
22.2
21.0
20.6
-

20.5
-

18.8
18.4
18.8
18.4
mm

•
-

(Table 2 continued)___________________________ _____ ____________________
age P*2 age rat Wt.of
Wt.of teeth % ash
% phosphorus
strain inject, sacrific. rat sex
(mgs)
dry basis
in ash
______ (days)
(days)
(gma)
mol.
inc. mol, inc. mol, inc. mol, inc.
resist*
w
n
tt
tt
tt

tt
tt

tt
tt

n
tt
tt
tt
n
tt
tt
tt

tt
tt
tt

25
20
20
20
25
20
20
20
20
20
29
35
35
29
35
25
35
20
20
mm

tt
tt
tt

stock
n
tt

20
_
am

w

-

tt

—

tt
n

_

n

n
n

-

_

w

_

w

_

it

-

n
n

—
_

«

-

n
n
n

-

-

“

27
30
30
30
30
20
20
22
35
35
31
36
37
39
40
40
40
50
50
55
60
60
60
167
6
6
6
6
6
9
9
9
9
9
11
14
14
14
18
19
19
19
20

_
-

55
-

28
33
53
e*
62
84
93
_
mm

77
68
134
122
98
17
13
10
10
14
12
21
23
17
19
-

33
37
31
29
31
36

76.0
78.3 16.1 16.5
75.3 15.7 13.6
15.2 15.7
79.0 75.8 15.6 16.1
79.6 76.5 16.2 16.1
79.6 75.8 16.3 15.8
79.6 76.0 16.1 16.1
78.8 75.3 15.6 15.6
80.7 76.8 15.1 15.2
76.9 75.2 15.2 15.5
mm
78.8 76.2
79.2 76.2
77.0 76.2 15.3 15.6
mm
76.7
16.1
78.2 76.2 16.1 16.1
75.7 74.3 15.4 15.6
78.3 75.4 15.8 15.8
78.5 77.9 14.8 15.3
78.2 77.9 14.8 15.4
78.1 78.3 15.6 16.3
16.1
77.5 78.3 77.0 15.0 15.3
78.6 77.0 15.3 15.3

76.0
83.4
72.9
71.6
62.1
53.8
M 54.4
M 57.8
F 81.1
M 87.7
F 63.6
F 79.3
F 83.7
F 75.8
M
M 83.1
F 78.5
M 91.7
M 96.5
F 98.1
F113.6
F _
M104.2
F119.3

64.7
66,0
66.5
65.6
58.3
36.7
39,0
42.8
79.9
79.0
69.0
75.7
87.5
94.3
100.5
84.2
77.3
116.1
105.1
116.5
154.3
146.4
134.7
228.6

79.2
80.6
79.0

6.7
_ 8.3
- 4.3
F 3.0
5.3
M 12.9
F 12.9
M 14.9
F 18.2
F 12.1
.M 22.2
F 31.9
M 34.6
F 35.1
F 56.2
M 52.8
M 53.4
F 55.6
F 56. B

4.8
5.9
3.6
3.2
4.4
8.5
7.5
8.9
11.3
8.0
14.0
22.0
21.2
22.1
35.0
35.7
35,7
36.4
36.9

46.3
45.8
72.1
46.7
54.7
51.9
54.3
61.1
61.5
50.4
61.7
68.7
74.0
71.2
76.3
77.5
77.7
77.7
78.0

M
F
F
M
F
F

60.4
54.2
91.7
68.B
59.2
65.9
66.7
78.7
82.3
62.5
71.4
71.4
78.3
71.0
72.0
74.2
72.8
75.0
73.7

9.6
9.5
11.6
10.8

12.5
12.3
14.4
13.4

-

-

10.5
10.9
15.4
11.1
10.6
12.4
13.5
14.0
14.2
15.5
15.3
14.4
15.7
15.3

12.2
14.4
14.7
14.9
13.5
14.6
14.8
15.5
14.4
15.0
15.1
14.9
15.5
15.2

Continued next page

- 34-

19.1
19.4

«•
21.0
18.0
21.
21.1
20.9
21.2
20.8
19.7
20.7
20.5
21.0
21.2
21.0
21.0
19.6
19.8
20.8
20.7
19.9
19.9

20.7
20.8
16.1
22.9

20.7
23.6
14.8
19.5

20.0
19.9
19.7
20.3
20.4
20.2
19.8
16.8
19.8
19.8
mm

20.5
20.4
20.3
18.8
18.9
20.0
-

-

20.1
30.1
18.9
18.0
21.0
20.1
19.7
18.9
19.9
20.3
19.7
18.5
20.2
19.6

-

18.6
21.6
18.6
18.1
21.6
20.7
20.8
19.8
20.3
20.8
20.4
20.4
20.7
20.6

(Table 2 continued)
tfo phosphorus
wt.of teeth % ash
age P52 age rat •wt.of
strain inject* sacrific. rat sex
(mgs)
dry basis
in ash
(days)
(days)
(gras)
mol.
inc. mol. inc. mol. inc. mol. inc.
stock
tt
t«
tt
tt
tt

20
20
24
-

tt

-

tt

20
24
20
29
32
24
29

tt
tt
tt
tt
tt

tt
n
tt
tt

mm

35
32

tt

-

tt

-

tt
tt
tt

n

32
35
20
35

20
21

32

99
MM

-

29
30
30
30
30
34
35
34
35
39
39
40
40
42
46
46
47
52
55
55

72
60
54

mt

-

M
F
F
M
F
F
F
M

-

M

-

M
M
F
F
M
F
M
F
F
F
M
F
F
M

77
-

96
87
120
-

132
131
-

134
-

159

53.0
52.4
64. 8
72.8
95.8
79.0
75.8
82.9
84.1
93.6
83.6
87.6
84.5
95.2
90.4
94.4
106.2
118.7
124.1
103.7
108.5
109.0
101.2

-35-

36.3
39.6
46.9
60.7
73.3
64.1
60. B
67.0
80.6
83.6
77.7
84.3
76.0
85.1
99.5
111.9
111.8
144.0
138.8
124.2
135.8
136.0
142.0

80.8
80.0
76.9
78.6
79.0
78.7
77.9

78.5
76.8
74.5
77.5
77.4
76.8
76.8

-

-

77.5
76.7
78.3
77.5
76.5
77.4
76.0

76.9
74.4
75.2
75.5
75.9
77.0
76.6

15.6 15.7 19.3
■
B
15.0 14.9
-

15.3
15.6
15.6
15.5
15.0
15.1
15.0
-

14.2
15.2
14.3
mm
15.0
76.0 78.6 15.3
76.6 76,9 15,3
77.5 77.575.3 76.3 14.3
78.0 77.9 15.6
77.2 77.5 14.5

.

-

15.9
16.0
16.1
16.1
15.9
15.6
15.4
mm

14.9
15.8
14.7
15.7
15.8
16,0
14.9
16.0
15.1

-

19.4
19.7
19.8
20.0
-

19.9
mm

20.5
20.7
20.9
21.0
mm

19.6
19.6

20.4
20.7

-

-

18.5
19.6
18.9

19.5
20.5
19.5

mm

20.1
20.0
19.0
20.0
18.7

mm

20.9
20.8
19.6
20.5
19.4

Table 3. Ash and phosphorus values obtained in the age range of 20 to 60 days.
Strain

No.
Rats

% Ash
Mol.

Incls.

% Phosphorus
dry basis
in ash
Mol.
Incis.
Mol.
Incis.

Susceptible 37

76.710.9

75.811.1

15.0l0.2

15.510.3

19.6+0.3

20.510.4

Resistant

25

78.8+0.9

76.4+0.8

15.510.4

15.511.0

19.6+0.5

20.310.8

Stock

IB

77.6+1.0

76.4ll.O

15.110.4

15.610.4

19.5l0.4

20.3-0.5

-36-

Table 4, Rates of deposition in and removal of radioactive phosphorus
from the teeth of the resistant and susceptible strains and stock rats
that were injected intraperitoneally with the isotope.
strain

stock
tt
tt
tt
n
tt
tt
SU SC *

tt
tt
tt

at time of
injection
age wfc*
(da.) (gm.)
n z'

C'J
«

n
it
*t
n
it
20
«
w
»

tt
tt

tt

tt

II

tt

tt
tt

tt
resist*
tt
n
tt
tt
tt
tt
tt
tt
tt
tt
n
n
tt
n
tt
tt
tt

H

19
n
20
n
n
n
n
n
n
n
n
n
n
it
w

n
n
it

8ex

time after
injection

% inject.dose found/100 mg. ask

(hrs.)

molars

incisors

27
33
32
31
32
33
30

F
F
F
M
F
M
F

24
48
72
240
360
480
840

5.30
6.67
7.05
7.49
5.85
6.97
5.17

3.34
8.78
10.1
9.96
8.32
8.62
3.34

3B
40
41
39
48
40
38
28
34

F
F
M
M
F
F
M
M

24
48
120
240
240
360
480
840
960

4.03
3.99
5.04
6.31
4.11
6.24
3.91
2.82
3.12

8.75
6.98
10.1
12.2
6.99
9.17
5.41
3.14
3.68

22
25
28
33
28
30
34
32
37
28
37
32
35
25
29
28
29
28

M
M
F
M
F
F
F
M
F
H
F
M
M
F
M
M
F
M

2
9
1
4
18
24
48
48
120
120
240
240
360
360
480
720
840
960

6.28
8.25
1.47
3.39
3.34
3.44
4.50
3.70
5.34
5.59
4.04
5.32
7.64
4.85
4.79
3.60
3.84
3.46

7.99
10.6
1.99
3.49
4.97
8.30
8.25
4.52
9.26
10.9
7.58
8.05
8.68
7.38
6.03
3.52
3.47
2.99

M

Continued next page

-37-

(Table 4 continued)
at time of
injection sex
strain
age
irfc.
(da.) (gnu)
stock
tt

24
tt

tt

tt

tt

tt

tt

tt

n

tt

resist.
tt

25
tt

tt

tt

tt

tt

tt

tt

stock
tt

SU S C .
n

resist.
tt

stock

29
tt

29
n

29
tt

32

tt

tt

tt

tt

it

tt

n

tt

tt

tt

tt

resist.
tt

35
35
tt

tt

tt

tt

tt

n

tt

S U S C .
tt

33

tt

35

tt

tt

it

tt

tt

time after % injact.dose found/lOO mg. ask
inj ection
(hrs)

molars

incisors

33
31
30
33
35
31

M
M
F
M
M
F

24
48
72
120
240
360

2.50
3.72
4.27
4.00
3.76
4.02

4.75
8.79
8.45
8.98
6.08
9.08

55
54
44
50
35

F
M
F
F
F

24
48
120
240
360

1.55
1.79
4.13
2.77
3.99

3.41
3.41
8.10
5.52
7.81

52
55

M
F

120
240

1.72
1.69

4.12
4.25

58
74

M
M

120
240

3.00
2.20

6.11
5. 84

53
50

F
F

41
240

1.47
1.54

3.55
4.42

73
66
61
59
65
58
74

M
M
F
F
F
M
M

24
48
72
120
240
360
480

0.97
1.07
1.50
1.31
1.55
1.52
0.82

2.41
2.27
3.90
3.75
4.42
5.06
2.27

42
64
39
55
63

F
F
M
F
M

24
48
120
240
360

1.41
1.06
1.68
1.46
1.19

5.01
3.04
6.23
4.98
4.70

85
90
110
83
90

F
F
M
F
F

120
240
48
360
480

1.10
1.05
0.92
1.39
1.08

3.44
3.12
2.90
3.83
3.77

-38-

Table 5. Radioactive phosphorus found in the teeth of offspring of females of
the resistant and susceptible strains and stock rats that were injected with
the isotope during pregnancy.
wt.
gestat.* age**
sex wt. of teeth
strain time bet. offspr,. of
% ash
% P in
ing. and saerif,, offspr
ash
birth(da.) (da.)
mol.
(gms)
inc. mol. inc. mol. inc.

%inj.dose
found per
100 mg.ash
mol, inc.

S U S C .

2

6

15

F

6.9

4.9 43.5 55.1

n

tt

tt

12

M

5.6

4.6 41.1 54.4

tt

tt

9

17

F

14.7

H

tt

14

22

M

27.5

18.9 69.9 69.5

tt

tt

20

30

F

35.4

25.6 78.3 75.2 20.5 21.4 2.09 2.40

2

6

10

M

5.2

4.6 42.4 58.7

-

-

5.45 7.04

tt

tt

6

10

M

5.0

3.3 36.0 42.5

-

-

7.22

tt

it

9

18

F

15.9

10.5 58.0 71.5 20.6 20.3 3.59 3.47

tt

tt

14

22

F

25.4

17.4 72.2 70.7

tt

tt

20

35

F

46.6

34.4 79.6 76.5 19.6 20.9 2.08 2.36

stock

5

6

10

F

5.3

4.4 54.7 59.2

15

6

15

F

6.9

5.2 55.0 46.2 19.1 30.6 0.11 0.19

tt

tt

9

25

M

17.7

tt

tt

14

35

M

34.5

resist.,

S U S C .

-

-

4.66 6.30

-

5.65 6.40

9.9 57.2 68.7 21.1 22.4 2.73 5.29

9.4 56.5 58.5

-

-

-

••

-

-

-

-

2.60 2.90

2.24 2.60

0.20 0.24

0.96 1.06

21.1 72.5 73.5 20.3 24.7 0.77 0.82

* Abbreviated reading should read "gestation time between injection
and birth (days)."
** Abbreviated reading should read "age offspring sacrificed (days)."

-39-

-

Table 6, In vitro adsorption of radioactive phosphorus by the teeth of the
resistant and susceptible strains and stock rats.
wt.
wt.of teeth
%
ash
adsorp. of ^^(c/in/mg.ash)
of
(®g»)
eth.gly.ex. 600°C. pH efch.gly.ex.ash 600°C.ash
rat sex
_____ (da.)(gma.)
mol.
inc. mol.
inc. mol.____ mol._______ inc.____ mol.
strain age

20

40

M

61.8

43.6 85.1

78.4

-

7

46.8

78.6

-

n

57

M

58.1

40.5 83.8

79.3

-

tt

54.9

73.9

m

tt

50

F

53.8

38.1 62.9

82.2

-

n

56.3

70.3

-

50

75

M

81.6

60.0 85.2

86.5

-

3

54.6

83.0

mm

N

49

F

73.4

59.4 86.2

87.2

-

tt

57.9

77.8

-

n

n

58

F

72.1

61.6 86.7

86.2

-

tt

63.7

81.7

-

SUSC.

40

119

tt

95

SUSC.

stock
«

SUSC.

resist,ft

resist.ft

60

SUSC.

resist,ft it
*

M 114.4*
H

■

93.2*

-

68.3

-

76.5

tt

57.2

-

14.6

-

84.7

-

76.8

tt

54.8

-

16.6

7

29.3

20.3

-

tt

27.2

24.2

-

176 F 120.3

159.8 82.6

84.7

-

151

119.7 84.4

86.5

-

M 105.8

Intact teeth were put into isotope solution during adsorption
period of 8 hours. All other samples of teeth were ground
with a mortar and pestle to fine crystals before tests were
made.

-40-

"Til

Both points are projected
from the zero axis

O O O ** Values obtained with counter
•

•

• = Theoretical values

O

14

21
28
Days after initial count

35

•

42

49

Figure 1. Comparison of the decay rate obtained on a standard solution of rad­
ioactive phosphorus and the theoretical (14.3 day half-life) decay rate.

2000

1500
B
1

&
1000
a
i
>o
*
r
I

W
+»
a
3
o
o
500

1200

1400
Volts

Figure 2. A characteristic voltage-counting rate curve

1600

200

O O ■ Susceptible strain

©

© © = Resistant strain

•

# • ■ Stock rats

•8

120

•

OJ
i

o

•° -

Weight

(mgs)

160

O

©
o©

1

©
8

Cb
o

O
80
o«®

©©

40
%

10

20

30

40
Age (days)

50

60

Figure 3. Weight of the molars of the susceptible and resistant strains and
stock rat3 in the age range of 6 to 60 days.

70

w
200

O O O
160

=» Susceptible strein

Q

O

O ■ Resistant Btrain

•

•

• “ Stock rats

o

«

i
it*.
i

O •
O

120

Weight

(raga)

°o

•

O
d»
°« o

S

©
o

©

80
« $ 8 5 3r > °

40

0

10

20

30

40
Age (days)

50

60

Figure 4. Weight of the incisors of the susceptible and resistant strains
and stock rats in the age range of 6 to 60 days.

70

»

i

50

60

90

e

Per cent ash

80

q

70
I

Q>

O O O «* Susceptible strain
O

60

©

0 #

© ■ Resistant strain
# “ Stock rats

50
o

*
40

«0
10

20

30

40
Age (days)

Figure 5. Per cent ash found in the molars of the susceptible and resistant
strains and stock rats in the age range of 6 to 60 days.

70

90

80

00
O •

70
•

Of

oo

O O O * Susceptible strain
Q

Q

•

•

© ■ Resistant strain

60
I
8

Stock rats

a

50

40
10

20

30

40
Ac® (days)

50

60

70

Figure 6. Per cent ash found in the incisors of the susceptible and resistant
strains and stock rats in the age range of 6 to 60 days.

w

..

18

©

16

©

Per cent phosphorus

•
O O

p

4•

14

©

• 8

°

•

o

O O O - Susceptible strain
.12
—

10 ____

,4

O ©

© - Resistant strain

•

• = Stock rats

•

O

o« ©
©
•
e

10

20

30
40
Age (days)

50

60

70

Figure 7. Per cent pho3T>horu3 found in the molars (dry basis) of the suscept­
ible and resistant strains and stock rats in the age range of 6 to 60 days.

18

16

Per cent phosphorus

O

14

.•&

Oq

©Cfc

I *

•

*

'

r

12

u®
%

>

i

9
O

ft
^

©
Q

o coo

o®
&

OQ
Q

fp

t
-©

_

O

O

O O » Susceptible strain

Q

Q Q «•Resistant strain

•

• # ■ Stock rats

e
Q
10

8

10

20

30

40
Age (days)

50

60

70

Figure 8. Per cent phosphorus found in the incisors (dry basis) of the suscep­
tible and resistant strains and stock rats in the age range of 6 to 60 days.

O

O O * Susceptible strain

Q

G © * Hesistant strain

#

•

Stock rats

o

j3©
I

8

v ,3>,

•

9
e
o

9

9
©

$

s
%

.88

o

•o
o

10

20

30

40
Age (days)

50

60

70

Figure 9. Per cent phosphorus in the ash of the molars of the susceptible and
resistant strains and stock rats in the age range of 6 to 60 dayB.

25

O O 6

= Susceptible strain

O O O

» Resistant strain

#

#

# * Stock rats

Per cent phosphorus

23

8

21

§

o o
•o
°§

•0
19

c©

$
8

ee

eo

17

15

10

20

30

40
Age (days)

50

60

70

Figure 10. Per cent phosphorus found in the ash of the incisors of the suscept­
ible and resistant strains and stock rats in the age range of 6 to 60 days.

10

O O O

XtI
o
cd

■ Susceptible strel)

Q

Q

O ■ assistant strain

•

•

A - Stock rata

u

<0
p.

o
48>*

o

o

ao

GO

TJ
*P

I

cn

O

Q

®

“©-0
-P

3
®
c
(
©4
ex.

i

O

o

0
e

to

o

o

o

12

18
24
Days after injection

30

36

40

Figure 11. Rates of deposition and removal of labelled phosphorus in the molars
of the susceptible and resistant strains and stock rate that were injected intraperitor.eslly
with the Isotope at 20 days of age.

e
o

• o

Per cent injected isotope per 100 mg. ash

10

9
09
09
8

o

o

Susceptible strain

0 9

0

Resistant strain

•••

O

Stock rats

Q
e

e

o
Q
O

8

12

18
24
Days after injection

30

36

o
9

42

Figure 12. Rates of deposition and removal of labelled phosphorus in the incis­
ors of the resistant and susceptible strains and stock rats that were injected intraperitoneally with the Isotope at 20 days of age.

Per cent Injected isotope per 100 rag. ash

w

O O O

■ Susceptible strain

Q 9 9

- Resistant strain

0 0

# = Stock rats

o
o

Q
O

9
12'
Lays after injection

Q_

15

18

21

Figure 13. Hates of deposition and removal of labelled phosphorus in the molars
of the susceptible and resistant strains and stock rata that were injected intraperitoneally with the isotope at 32 to 35 days of age.

W

10

An
efl
o
o

O

O O

0

0

•

8

•

“ Susceptible strain

9 "
•

Resistant strain
■ Stock rats

u

©

P.

»
&
4*
O
©

TJ
©

Q

4»

I

©
«

©

CJ1

I

4»
(3
©
o
f-.
©
S«

o

O

8

9
12
Days after injection

15

18

21

Figure 14. Rates of deposition and removal of labelled phosphorus in the incisors
of the susceptible and resistant strains and stock rats that were'injected intraperitoneally with the isotope at 32 to 35 days of age.

Per cent injected isotope per 100 mg. ash

O O O“

Susceptible strain

Q Q Q»

Resistant strain

O
Q

O
o

o i--------------- ---------------- ---------------- ---------------- ---------------- ---------------- ---------------0

3

6

,

9
12
Age of offspring (days)

15

18

21

figure 15. Per cent labelled phosphorus found in the molars of young rats of
the susceptible and resistant strains whose mothers had been injected intraperitoneally with the isotope during pregnancy.

Q
o

0 O O

= Aisoeptible strain

O

= Resistent strain

©

©

I
en
o>
i
s

0

___________

0

3

6

9

12

15

18

21

;»ge of offspring (days)
Figure 16 . Per cent labelled phosphorus found in the incisors of young rats of
the susceptible and resistant strains whose mothers had been injected intraperitor.eally
with the isotope during pregnancy.

BIBLIOGRAPHY
1.

Hoppert, C. A., Webber, P. A., and Canniff, T. L.s The Production
of Dental Caries in Rats Fed an Adequate Diet, Science 74,
77 (1931).
—

2.

Ibid,

3.

Hunt, H, R., and Hoppert, C. A,*
Genetics 24, 76 (1939).

4.

Ibid.

5.

Ibid. Inheritance of Susceptibility and Resistance
to Caries In
Albino RatB (mas norveglicus),. J. Am. Col. Den. 11, 33 (1944).

J, Dental Research 12, 161 (1932),
Inheritance in Dental Caries,

J. Am. Col. Den. 6, 70 (1939).

6. Hunt, E. R., Hoppert, C. A., and Erwin, W. G.t Inheritance of Sus­
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7. Olson, Kenneth J.t Thesis for M.S. degree, entitled, "A Study of
the Effect of Fluorides on the Development of Dental Caries in
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8.

Stipek, Robert W.: .Thesis for M.S. degree, entitled, nA Study of
the Influence of Diet and Other Factors on the Production of
Dental Caries in a Susceptible Strain of Rats". Michigan State
College, 1948.

9. Olson, Kenneth J.t Thesis for the PhD degree, entitled, "The Influ­
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Carles in a Susceptible Strain of Rats’*. Michigan State College,
1947.
10. Armstrong, W. D., and Brekhus, P. J., J. Biol. Chem., 120, 677-87
(1937).
.■I

11. Cork, J. M«j Test entitled, "Radioactivity and Nuclear Physics'*.
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12.

Hevesy, George, and Faneth, F. A .t Text entitled, " Manual on Radio­
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13. Kamen, Martin D.i "Radioactive Tracers in Biology".
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Academic Press

14. Korf, Serge A .t Text entitled, "Electron and Nuclear Counters.
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(1946).

-57-

15. Wilson, D. Wright, Nier, A. 0. C., and Reimann, Stanley P. t Tart
entitled, "Preparation and Measurement of Isotopic Tracers".
J. W. Edwards, Ann Arbor, Michigan. (1947).
16.

Bale, W. F., Haven, F. L., and LeFevre, M.t Apparatus for the
Rapid Determination of Beta Ray Activity in Solutions.
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17. Tracerlog (Adr. Bulletin of Tracerlab, Inc., Boston, Mass.) No. 4,
p. 9 (1947).
18. Ibid.

No. 5, p. 13 (1947).

19. McCauley, H. Burtons Significance of Radioactive Isotopes in Dental
Research. J. of the Amer. Dental Assoc., 29, 1219-1230. (1942).
20. Chievits, 0., and Hervesy, G.t Radioactive Indicators in the Study
of Phosphorus Metabolism in Rats. Nature, 136, 754-5. (1935).
21. Ibid.

Metabolism of Phosphorus in Animals.
Biol. Medd., IS, No. 9. (1937).

Kgl. Danske Videnskeb.

22. Krogh, A.t Use of Isotopes as Indicators in Biological Research.
Science, 85, 187. (1937).
23. Manly, M. L., and Bale, W. F.t Metabolism of Inorganic Phosphorus
of Rat Bones and Teeth as indicated by Radioactive Isotope.
J. Biol. Chem., 129, 125. (1939).
24. Hevesy, G.t Applications of Isotopes in Biology.
139, 1213. (1939).

J. Chem. Soc.,

25. Hodge, Harold Carpenter, Van Huysen, Grant, Gonner, John F., and
Van Voorhis, Stanlqr N.t Adsorption of Phosphates at 40° by
Enamel, Dentin, Bone, and Hydroxyapatlte as Shown by Radio­
active Isotope. J. Biol. Chem., 138, 451. (1941).
26.

Hodge, H. C. LeFevre,M., and Bale, W. F.tChemical and X-Ray
Dif­
fraction Studies of Calcium Phosphates.
Ind. Eng. Chem., Anal.
Ed., 10, 156. (1938).

■ 27. Hevesy, G.t Applications of Radioactive Indicators in Biology.
Ann. Rev. Biochem., 9, 641.
(1940).
28.

Hevesy, G. C«, Holst, J. J., and Knogh, A.t Investigations on
Exchange of Phosphorus in Teeth Using Radioactive Phosphorus
as Indicator. Kgl. Danske Videusk, Selsk. Biol. Medd., 13,
No. 13, 1937.

-58-

29.

Chase* S. W.s Critical Review of Controversy Concerning Metabolism
in Enamel. J. Amer. Dental Assoc. 18, 697. (1931).

30.

Hevesy,G. C., and Armstrong, W. D.* Exchange of Radiophosphate by
Dental Enamel. Proc. Am. Soc. Biol. Chem., 133, 44. (1940).

31.

Volker, J. F., and Sognnaes, R. F.j Study of Phosphorus Metabolism
in Dental Tissues of the Cat by Use of Radioactive Phosphorus.
J. Dental Research 19, 292. (1940).

32.

Sognnaes, R. F., and Volker, J. F.t Studies on Distribution of
Radioactive Phosphorus in Tooth Enamel of Experimental Animals.
Am. J. Physiology, 133, 112. (1941).

33.

Barnum,C. P., and Armstrong, W. D.t In vivo and in Vitro Exchange
of Phosphorus by Enamel and Dentin, Am. J. Physiology, 135,
478. (1942).

34.

Armstrong, W. D., and Barnum, Cyrus P.t Concurrent Use of Radioactive Isotopes of Calcium and Phosphorus in the Study of the
Metabolism of Calcified Tissues. J. Biol. Chem., 172, 199.
(1948).

35.

Pederson, P. 0., and Schmidt-Ni elsen, B.s Experiment ell o Undersogelser over Fosforstofskifte. 1. Menneskelige Taender med
Anvendelse af Radioaktivt Fosfor som Indikator. Tandlaegebladet 45, 396. (1941).

36. Manly, M. L., and Levy, S. R. t Effect of Pregnancy on Phosphorus
Turnover of Skeleton of Rats Maintained on Normal and Rachitogenic Diets. J. Biol. Chem. 159, 35. (1941).
37.

Crowell, C. D.. Hodge, H. C., and Line, W. R.t Chemical Analysis of
Tooth Samples Composed of Enamel, Dentine and Cementum.
J. Dental Research 14, 251-268. (1934).

38.

Official and Tentative Methods of Analysis of the Association of
Official Agricultural Chemists, p. 127, Sixth Edition. (1945).

39.

Bowes, J. H., and Itarray, M. M.j The Chemical Composition of Teeth.
Biochem. J., 29, 2721. (1935).

40. Cohn, W. E., and Greenberg, D. M.t Studies in Mineral Metabolism
with Aid of Artificial Radioactive Isotopes. 1. Adsorption,
Distribution, and Excretion of Phosphorus. J. Biol. Chem.,
123, 185. (1938).
41.

Johansson, E. Gunnar, Falkenheia, Marlene, and Hodge, Harold
Carpenter* The Adsorption of Phosphates by Enamel, Dentin,
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