NITROGEN AND PHOSPHORUS OF EARLY, SMOOTH-SURFACE

CARIES OF ENAMEL
THOMAS B. COOLIDGE
Walter G. Zoller Memorial Dental Clinic and Department of Biochemistry,
University of Chicago, Chicago, Ill.
T HIS paper presents nitrogen and phosphorus analyses of sound human
enamel and of the enamel of early, smooth-surface caries before changes of
surface contour have occurred.
Analyses of enamel have been reviewed by Leicester' and nitrogen analyses
of enamel by Losee and Hess.2 The novel feature of the present analyses is
believed to lie in the small amount (1 mg.) of material required for analysis by
the methods used, so that comparison is possible between the composition of
various enamel and of noncarious enamel from the same tooth. The analyses
were made with the purpose of gaining information on the early stages of the
processes producing smooth-surface caries of the enamel.
EXPERIMENTAL
Freshly extracted human teeth were placed in water covered with toluene
and stored 1 to 6 weeks. The teeth were rinsed with water, scrubbed with soap
and a hard bristle brush, scraped free of debris, scrubbed again, and stored
from a few days to a month in distilled water covered with toluene. Before
analysis the teeth were dried with absorbent tissue paper. Sound enamel was
ground in successive portions from areas of about 8 square millimeters. Carious
portions were selected which were colorless, completely homogeneous in appear-
ance, and without change in contour from that of the surrounding enamel.
Some were gingival but most were interproximal. Dr. J. R. Blayney confirmed
the clinical diagnosis of several of these areas. Carious material, about a milli-
gram in weight, was removed with a delicate hand instrument. Hard white
spots (hypoplasia?) of enamel of similar size were removed with a diamond
wheel.
All samples were weighed to a precision of about 0.01 mg. immediately
after removal. They were then ashed with 2.5 ml. of 1ON H204 and with
H202 by the method of Koch and McMeekin.3 Aliquots were analyzed for
nitrogen by the method of Conway4 and for phosphorus by the method of
Gomori.5 Nitrogen analyses were accepted when duplicates within 2 per cent
were obtained; phosphorus analyses when duplicates were within 3 per cent.
The probable error of the analyses was obtained from 30 analyses of separately
weighed samples from 5 pooled lots of normal enamel.
Received for publication, June 26, 1950; revised by author, Sept. 26, 1950.
97
98 COOLIDGE J. D. Res.
February, 1951

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Volume 30 NITROGEN AND PHOSPHORUS ANALYSES OF ENAMEL 99
Number I

RESULTS
The probable error of the nitrogen analyses, including weighing, was 0.95
per cent; that of the phosphorus analyses, 1.4 per cent. The results of the
analyses are given in Table I. Layer 1 represents the layer of enamel removed
first; Layer 2 that removed second, etc., all "layers" being removed from ap-
parently normal areas.
DISCUSSION
The analyses show that there is a large variation in the nitrogen and phos-
phorus content of enamel from different teeth and from different areas of the
same tooth. They show no consistent differences between normal enamel, various
enamel, and hard white spots of enamel. The outermost layers of normal enamel
show a higher nitrogen content in general than deeper layers as might have been
anticipated from the appearance of a surface membrane on decalcification of
teeth with mineral acids.
It is to be noted that a loss of phosphorus from enamel in the various process
would not be entirely reflected in phosphorus analyses of the removed material
were there either (1) a decrease in density of the enamel associated with caries,
or (2) a more rapid loss of water, before weighing from the removed various
enamel than from normal enamel. Insofar as a decrease in density was due to
replacement of apatite by water, such a decrease would not entirely eliminate
a drop in phosphate as measured in removed material. Loss of weight of the
various material during weighing has not been observed. The phosphorus
analyses do not show a large general removal of phosphate in early smooth
surface caries of the enamel.
With similar considerations as to density change and water loss obtaining,
the figures for nitrogen indicate that any nitrogenous material lost by proteolysis
and diffusion is balanced by replacement of nitrogen from some such source as
saliva.
The phosphorus to nitrogen ratios are not subject to the above considerations
as to density and water loss. They give no evidence for a preponderance of
either phosphorus or nitrogen removal in the type of caries examined.
SUMMARY
Phosphorus and nitrogen analyses of normal enamel and of the enamel of
early smooth surface caries show no large general differences in phosphorus or
nitrogen content.
All the analyses were perf ormed by Miss A. E. I. Ebisch. The teeth were supplied
through the courtesy of Dr. Eli Olech of the University of Illinois College of Dentistry.
REFERENCES
1. Leicester, H. M.: Biochemistry of the Teeth, St. Louis, 1949, The C. V. Mosby Company.
2. Losee, F. L., and Hess, W. C.: J. D. Res. 28: 512, 1949.
3. Koch, F. C., and McMeekin, T. L.: J. Am. Chem. Soc. 46: 2066, 1924.
4. Conway, E. J.: Microdiffusion Analysis, London, 1947, C. Lockwood.
5. Gomori, G.: J. Lab. & Clin. Med. 27: 955, 1942.