- Email: [email protected]
Experimental Study in Salivary Reaction*
EXPERIMENTAL STUDY IN SALIVARY REACTION * By L. R. GANS, D.D.S., New Haven, Connecticut
(Clinic given before Connecticut State Dental Association, May 1, 1925)
I
T W AS the practice for a long time Brown and Freeman (1 9 1 6 ): “ T he to consider that various abnormal Effect of Variations of the Gastric conditions of the teeth and soft tis Secretions upon the Composition of sues of the oral cavity were caused by Saliva.” or were associated with the presence of Barnes ( 1907): “T he Bactericidal acid in the mouth. In recent years, Action of Saliva.” this view has been challenged re Stern (1 9 0 4 ): “ Changes in the Sali peatedly. Because of the important vary Secretion Affected by Systemic functions which saliva fulfils in the mouth, its reaction may deservedly be Diseases.” T he present paper was undertaken studied carefully. In reviewing the literature relative with the thought in mind of an investi to experimentation and work done on gation to determine the true acidity of salivary reaction and associated influ saliva as distinguished from so-called ences, such works as the following, to acidity as measured by the usual titra gether with the numerous and exhaustive tion procedure and the uses of various studies o f saliva by Gies, Kirk, Pickerill, indicators. Howe, Nielson, Carlson and many Some investigators have reported that others, were read and studied for in saliva is commonly or normally acid, dicators of salivary reaction. and others that it is alkaline in reaction. Prinz (1 9 1 8 ): “ Relationship of As long ago as 1911, Howe1 drew at O ral Secretions and Dental Caries.” tention to these conflicting statements in M arshall (1 9 1 7 ): “ Composition of the earlier literature, and he especially Saliva in Relation to the Incident of pointed out the importance o f using the Dental Caries.” appropriate indicators in measuring the Ferris (1 9 1 9 ): “ M outh Hygiene acidity of saliva. In general, he fpund Controlled by Diet Through Salivary that “ if the majority of text books speak Analysis.” the truth, then acidity is the divergent Graham (1 9 1 9 ): “ Litmus as an In step from normal and its degree is the dicator of Salivary Reaction.” index of the underlying physiological Myers (1 9 1 8 ): “ Salivary Analysis.” alteration.” Since then, numerous in Bloomfield and Huck (1 9 2 0 ): “ Re vestigations and studies have proved that action o f Saliva.” ♦Experimental W ork Conducted Brady Laboratory, Yale University. lour. A. D.A., February, 1926
at
the
1. Howe, P. R .: Indicators in Salivary Analyses, Dent. Cosmos, S3: 320 (M arch) 1911.
222
Cans— Experim ental S tu d y in Salivary Reaction
the conclusions of these earlier workers were not entirely correct. In the past ten years, the methods of measuring hydrogen-ion concentration, resulting in determining the true acidity of the substance examined, have been perfected. W e have, therefore, con sidered it important to undertake a study of the hydrogen-ion concentration of saliva in normal mouths and in the mouths o f persons showing carious teeth, gingivitis, pyorrhea and other pathologic conditions, as will be noted from the accompanying table. No effort o f any degree was exerted in obtaining the saliva, any procedure as used in the works o f nerve stimula tion and inhibition as done by Heidenhain and Langley or psychic stimuli by Pawlow, being eliminated, nor was the saliva dialyzed, to avoid the loss of. carbondioxid and an increase in pH.2 T h e measurements o f the acidity of saliva were conducted colorimetrically. In general, the procedure was as fol lows: A sample of fresh saliva was collected in a clean test tube. One half o f the collected saliva was placed into a second tube, and promptly after diluted with approximately 4 parts of distilled water and the mixture stirred. T he diluted saliva was then distributed into five clean tubes. T he other half was filtered through filter paper, the filtrate being then subdivided in two, one-half distributed in five clean tubes and the other h alf boiled and distributed in five clean tubes. T he unfiltered and filtered, and the filtered boiled prepara tions were then tested with: (a) litmus, (b) phenolphthalein, (c) congo red, (d ) lacmoid and (e ), colorimetrically, with bromthymol blue.
223
Bromthymol blue is that indicator in the Clark and Lubs series which is ap propriate for the measurements of hydrogen-ion concentration (or true acidity) in the zone of reaction within which saliva usually falls. T he theory and technic of measuring true acidity are given at length by C lark.3 In the colorimetric examination, the following procedure was employed: Bromthymol blue, 1 c. c., was used rather than the drop method, as giving accuracy in the amount used each time fo r each test. T he saliva was diluted in the ratio 1: 9 with distilled water, which is exactly the same dilution em ployed by Starr and Bloomfield and Huck.4 Such quantities of saliva were col lected and transferred by means of a pipet to five test tubes containing such amounts of diluted water as to render the proportion o f saliva to water, suc cessively, 1 part saliva to 10 parts water. T hen 0.01 per cent bromthymol blue was added so that the indicator was al ways present in the proportion of 1:10 with respect to the total quantity of diluted saliva. Preliminary to using any of the fore going apparatus, such as test tubes, pipet, and funnels, in the investigation, they were washed and rinsed thoroughly in freshly boiled distilled water. T he color reactions in each tube were then carefully noted, and matched exactly in color the standard buffer solution, rang ing in f B from 6.2 to 7.6. Each sample was tested, heated and unheated, to de termine whether the measured acidity was influenced or determined by the con-
3. C lark: The Determination o f H ydro gen-ion, 1922. 2. The f B is the Sorenson index of true acidity. T he smaller the f B, the higher +. Bloomfield, A. L., and Huck, J . G .: the acidity; and the greater the f n, the Bull. Johns Hopkins Hosp. 31: 118 (A pril) lower the acidity or the greater the alkalinity. 1920.
224
The Journal of the American D ental Association
56 35 50 28 16 15 38 50 24 35 50 32 48 17 28 28 40 50 28 55 50 29 35 î 27 42 24 34
20 23 50 70 18 25 23
20
1
1 1 1 2 12 1 1 4 3 4 9 6 2 14 4
20 52 55 62 35
22 22
I
1 1 1 1
1 1 1
1 1 1 1 1 1 1 1
1 1 1
1 8 20 14
1 1
1 1
1 1 1
1
1
1 1
1 1
1 1 1 1
4
1
1
1 1 4
1
1 1 1 1
15 1 1
1 1
12 1 1
4
8 3 2 5 3 8
1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1
1 1 1 1 1
1
1 1 1 1
1 1 1 1 1
1
1 1 1 1
1
2
1 1
2
13 45 16* 16 50
21* 22
17 27 32 37* 40
*S ma ll a m o u n t of ca rie s.
1 1 1 1 1 1
1 1 1
1 1 1 1 1
1 1 1 1 1
1
1
t V egetarian.
F M M F M M M F M F F M M F F F M M F F F F M M F M F M F M F M M M F F F F M F M F F F M M M F M F M M F M M M F
F M M F M F F M M F M M M M F F F M M F M M F
C o n d itio n s
D evitalized Teeth
ral
Parti al P l a t e s
O
! E d en tulous 1 M ouths
t/3
!
E xten sive | C ar ie s
a t h o lo g ic
1 1 1
1
11
10
P
1 1
1 1
1 1 12
21
45 37 16
1
1 1
29 18 28 24 30 35 40 16
and
1 E xten sive Bridgework
Pyorrhea
! 22
ormal
1
21 28 26 35 45 25 28 27 32 13 35
N
Small A m o u n t of Bridgework
in
G ingivitis
C o n c en tr a tio n
Normal G u m s
ydrogen-Ion
E xtractions
H
4 5 2
2 1 1 1 8 1 1 I 2 1 1 1
1 1 2 3
1 1
1 1 1
1 1
1 3 3 4 2
2 3 4 1 3
1 1
1
..
1 11
i
3
m
Gans— Experim ental Study in Salivary Reaction
centration of the carbonic acid gas which is a normal constituent of saliva. Heat ing the saliva drives off the carbonic acid gas, and should, if anything, de crease the acidity. W e have examined about ninety samples of saliva obtained from persons of all ages and both sexes without a purposive selection through a period of about six months. T hey represent samples o f saliva as one would obtain it at random from the clientele of the usual dental practice, including persons who showed evidences of the conditions listed in the accompanying table. I t is unnecessary to report all of the experiments in detail. In all cases, the saliva, fresh, filtered and filtered boiled, was neutral or slightly acid to litmus, colorless to phenolphthalein, red to congo red and blue to lachmoid. No differences between the filtered and un filtered, boiled and unboiled prepara tions were evident, which indicates the relative unimportance o f the carbon dioxid concentration. T he results with these indicators show the samples of diluted saliva to be in the zone of acid ity which is represented by the range of fH 6.4-7-0 inclusive. O u r results with bromthymol blue are in complete accord with the degrees of acidity suggested by the other indica tors. T h e result is accurate not only in
225
giving the reaction of the saliva (acid or alkaline) but also in giving the de gree o f acidity or alkalinity. T he results are summarized in the table where the of the saliva is indicated according to the condition of the teeth and gums. T he table conclusively shows the magnitude of the field covered in pyorrheic mouths containing all variaties of dental constructions. T he various metals used in restoration, such as amalgam, alloys, synthetics, gold and solders, were taken into consideration in the determination of the final hydrogen-ion concentration. Included also in the consideration of change in hydrogen-ion concentration were food inges tions, mouth washes and toothpastes. It is evident from the data collected in this investigation that: 1. Saliva is nominally slightly acid (pa 6.4 to 7.0). 2. T he normal acidity of saliva is independent of the condition of the teeth and gums. 3. T he acidity shows no constant relation to the occurrence o f ab normalities in the mouth. I am greatly indebted for the out line o f this work and the valuable as sistance in conducting these experiments to Prof. C. Underhill of the Physiolog ical Laboratory of Yale University.
(Clinic given before Connecticut State Dental Association, May 1, 1925)
I
T W AS the practice for a long time Brown and Freeman (1 9 1 6 ): “ T he to consider that various abnormal Effect of Variations of the Gastric conditions of the teeth and soft tis Secretions upon the Composition of sues of the oral cavity were caused by Saliva.” or were associated with the presence of Barnes ( 1907): “T he Bactericidal acid in the mouth. In recent years, Action of Saliva.” this view has been challenged re Stern (1 9 0 4 ): “ Changes in the Sali peatedly. Because of the important vary Secretion Affected by Systemic functions which saliva fulfils in the mouth, its reaction may deservedly be Diseases.” T he present paper was undertaken studied carefully. In reviewing the literature relative with the thought in mind of an investi to experimentation and work done on gation to determine the true acidity of salivary reaction and associated influ saliva as distinguished from so-called ences, such works as the following, to acidity as measured by the usual titra gether with the numerous and exhaustive tion procedure and the uses of various studies o f saliva by Gies, Kirk, Pickerill, indicators. Howe, Nielson, Carlson and many Some investigators have reported that others, were read and studied for in saliva is commonly or normally acid, dicators of salivary reaction. and others that it is alkaline in reaction. Prinz (1 9 1 8 ): “ Relationship of As long ago as 1911, Howe1 drew at O ral Secretions and Dental Caries.” tention to these conflicting statements in M arshall (1 9 1 7 ): “ Composition of the earlier literature, and he especially Saliva in Relation to the Incident of pointed out the importance o f using the Dental Caries.” appropriate indicators in measuring the Ferris (1 9 1 9 ): “ M outh Hygiene acidity of saliva. In general, he fpund Controlled by Diet Through Salivary that “ if the majority of text books speak Analysis.” the truth, then acidity is the divergent Graham (1 9 1 9 ): “ Litmus as an In step from normal and its degree is the dicator of Salivary Reaction.” index of the underlying physiological Myers (1 9 1 8 ): “ Salivary Analysis.” alteration.” Since then, numerous in Bloomfield and Huck (1 9 2 0 ): “ Re vestigations and studies have proved that action o f Saliva.” ♦Experimental W ork Conducted Brady Laboratory, Yale University. lour. A. D.A., February, 1926
at
the
1. Howe, P. R .: Indicators in Salivary Analyses, Dent. Cosmos, S3: 320 (M arch) 1911.
222
Cans— Experim ental S tu d y in Salivary Reaction
the conclusions of these earlier workers were not entirely correct. In the past ten years, the methods of measuring hydrogen-ion concentration, resulting in determining the true acidity of the substance examined, have been perfected. W e have, therefore, con sidered it important to undertake a study of the hydrogen-ion concentration of saliva in normal mouths and in the mouths o f persons showing carious teeth, gingivitis, pyorrhea and other pathologic conditions, as will be noted from the accompanying table. No effort o f any degree was exerted in obtaining the saliva, any procedure as used in the works o f nerve stimula tion and inhibition as done by Heidenhain and Langley or psychic stimuli by Pawlow, being eliminated, nor was the saliva dialyzed, to avoid the loss of. carbondioxid and an increase in pH.2 T h e measurements o f the acidity of saliva were conducted colorimetrically. In general, the procedure was as fol lows: A sample of fresh saliva was collected in a clean test tube. One half o f the collected saliva was placed into a second tube, and promptly after diluted with approximately 4 parts of distilled water and the mixture stirred. T he diluted saliva was then distributed into five clean tubes. T he other half was filtered through filter paper, the filtrate being then subdivided in two, one-half distributed in five clean tubes and the other h alf boiled and distributed in five clean tubes. T he unfiltered and filtered, and the filtered boiled prepara tions were then tested with: (a) litmus, (b) phenolphthalein, (c) congo red, (d ) lacmoid and (e ), colorimetrically, with bromthymol blue.
223
Bromthymol blue is that indicator in the Clark and Lubs series which is ap propriate for the measurements of hydrogen-ion concentration (or true acidity) in the zone of reaction within which saliva usually falls. T he theory and technic of measuring true acidity are given at length by C lark.3 In the colorimetric examination, the following procedure was employed: Bromthymol blue, 1 c. c., was used rather than the drop method, as giving accuracy in the amount used each time fo r each test. T he saliva was diluted in the ratio 1: 9 with distilled water, which is exactly the same dilution em ployed by Starr and Bloomfield and Huck.4 Such quantities of saliva were col lected and transferred by means of a pipet to five test tubes containing such amounts of diluted water as to render the proportion o f saliva to water, suc cessively, 1 part saliva to 10 parts water. T hen 0.01 per cent bromthymol blue was added so that the indicator was al ways present in the proportion of 1:10 with respect to the total quantity of diluted saliva. Preliminary to using any of the fore going apparatus, such as test tubes, pipet, and funnels, in the investigation, they were washed and rinsed thoroughly in freshly boiled distilled water. T he color reactions in each tube were then carefully noted, and matched exactly in color the standard buffer solution, rang ing in f B from 6.2 to 7.6. Each sample was tested, heated and unheated, to de termine whether the measured acidity was influenced or determined by the con-
3. C lark: The Determination o f H ydro gen-ion, 1922. 2. The f B is the Sorenson index of true acidity. T he smaller the f B, the higher +. Bloomfield, A. L., and Huck, J . G .: the acidity; and the greater the f n, the Bull. Johns Hopkins Hosp. 31: 118 (A pril) lower the acidity or the greater the alkalinity. 1920.
224
The Journal of the American D ental Association
56 35 50 28 16 15 38 50 24 35 50 32 48 17 28 28 40 50 28 55 50 29 35 î 27 42 24 34
20 23 50 70 18 25 23
20
1
1 1 1 2 12 1 1 4 3 4 9 6 2 14 4
20 52 55 62 35
22 22
I
1 1 1 1
1 1 1
1 1 1 1 1 1 1 1
1 1 1
1 8 20 14
1 1
1 1
1 1 1
1
1
1 1
1 1
1 1 1 1
4
1
1
1 1 4
1
1 1 1 1
15 1 1
1 1
12 1 1
4
8 3 2 5 3 8
1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1
1 1 1 1 1
1
1 1 1 1
1 1 1 1 1
1
1 1 1 1
1
2
1 1
2
13 45 16* 16 50
21* 22
17 27 32 37* 40
*S ma ll a m o u n t of ca rie s.
1 1 1 1 1 1
1 1 1
1 1 1 1 1
1 1 1 1 1
1
1
t V egetarian.
F M M F M M M F M F F M M F F F M M F F F F M M F M F M F M F M M M F F F F M F M F F F M M M F M F M M F M M M F
F M M F M F F M M F M M M M F F F M M F M M F
C o n d itio n s
D evitalized Teeth
ral
Parti al P l a t e s
O
! E d en tulous 1 M ouths
t/3
!
E xten sive | C ar ie s
a t h o lo g ic
1 1 1
1
11
10
P
1 1
1 1
1 1 12
21
45 37 16
1
1 1
29 18 28 24 30 35 40 16
and
1 E xten sive Bridgework
Pyorrhea
! 22
ormal
1
21 28 26 35 45 25 28 27 32 13 35
N
Small A m o u n t of Bridgework
in
G ingivitis
C o n c en tr a tio n
Normal G u m s
ydrogen-Ion
E xtractions
H
4 5 2
2 1 1 1 8 1 1 I 2 1 1 1
1 1 2 3
1 1
1 1 1
1 1
1 3 3 4 2
2 3 4 1 3
1 1
1
..
1 11
i
3
m
Gans— Experim ental Study in Salivary Reaction
centration of the carbonic acid gas which is a normal constituent of saliva. Heat ing the saliva drives off the carbonic acid gas, and should, if anything, de crease the acidity. W e have examined about ninety samples of saliva obtained from persons of all ages and both sexes without a purposive selection through a period of about six months. T hey represent samples o f saliva as one would obtain it at random from the clientele of the usual dental practice, including persons who showed evidences of the conditions listed in the accompanying table. I t is unnecessary to report all of the experiments in detail. In all cases, the saliva, fresh, filtered and filtered boiled, was neutral or slightly acid to litmus, colorless to phenolphthalein, red to congo red and blue to lachmoid. No differences between the filtered and un filtered, boiled and unboiled prepara tions were evident, which indicates the relative unimportance o f the carbon dioxid concentration. T he results with these indicators show the samples of diluted saliva to be in the zone of acid ity which is represented by the range of fH 6.4-7-0 inclusive. O u r results with bromthymol blue are in complete accord with the degrees of acidity suggested by the other indica tors. T h e result is accurate not only in
225
giving the reaction of the saliva (acid or alkaline) but also in giving the de gree o f acidity or alkalinity. T he results are summarized in the table where the of the saliva is indicated according to the condition of the teeth and gums. T he table conclusively shows the magnitude of the field covered in pyorrheic mouths containing all variaties of dental constructions. T he various metals used in restoration, such as amalgam, alloys, synthetics, gold and solders, were taken into consideration in the determination of the final hydrogen-ion concentration. Included also in the consideration of change in hydrogen-ion concentration were food inges tions, mouth washes and toothpastes. It is evident from the data collected in this investigation that: 1. Saliva is nominally slightly acid (pa 6.4 to 7.0). 2. T he normal acidity of saliva is independent of the condition of the teeth and gums. 3. T he acidity shows no constant relation to the occurrence o f ab normalities in the mouth. I am greatly indebted for the out line o f this work and the valuable as sistance in conducting these experiments to Prof. C. Underhill of the Physiolog ical Laboratory of Yale University.