The organic volatiles of the mouth air, expired air and vapor from saliva were assayed by the gas chromatograph equipped with a flame ionization detector (Shimazu model GC-4APEE).
For the detection of amines the enrichment device was employed introducing a solid selective absorbor.
The mouth air was repeatedly breathed through the absorbor and the collected amines were released and expelled by the addition of alkaline. It was condensed in a U-tube immersed in liquid nitrogen using helium as a carrier gas.
The enriched samples evaporated from the warmed U-tube was introduced to the gas chromatograph.
As for the column materials for low-molecular amines the mixed coating with 5% tetraethylenpentamine and 15% diglycerol on Chromosorb W, HMDS was most sensitive but too sensitive for the practical use as being accompanied with many non-identifiable peaks. From the standpoint of reliability Chromosorb 103 was most satisfactory.
Regardless of the enrichment procedures only three very flat peaks were detected on the chromatogram using Chromosorb 103.
The components were too diluted for identifying amines. Therefore the evidence of amines in the oral cavity was not demonstrated in periodontitis as well as in healthy state.
The reason of unsatisfactory results might be attributed to the inefficiency of the enrichment procedure for amines, resulting the reduction of sensitivity down to one hundredth of the direct injection of liquid sample. It appeared to suggest the absorbance of samples on the wall of long tubings.
Surveying other sampling techniques, the direct sampling of air from the oral cavity was found most sensitive and reproducible. The air collected into the pre-warmed syringe was directly injected in the column packed with Chromosorb 101.
The distinct presence of ethanol and aceton was demonstrated in all subjects.
The presence of methanol, though seldom found, was always related to the drinking of alcoholics on the previous day.
The ethanol concentration of non-drinking subjects was higher in the oral cavity than in expired air, suggesting the local biogenesis in the mouth.
The organic volatiles from incubated saliva demonstrated a relatively dense unidentified component with the retention time between methanol and ethanol. The component appeared to be more condensed in subjects with offensive breath.
The ethanol concentration of the oral cavity was always high in alcohol-sensitive subjects while very low in resistant one after a hour from drinking the same volume of whisky.
The concentration of methanol was higher after drinking Japanese Sake (rice wine) than in case of whisky.
In the volatile from saliva following the drinking of Japanese Sake, many unidentified components appeared, suggesting the presence of more complicated metabolites.
After the work of anesthetizing many rats for experiments, ethylether was detected in expired air, reflecting the influence of inhaled air. The concentration in the oral cavity was one half or one third of that in expired air. Even after 24 hours, ethylether was still detected in the breath.
The suffer containing organic volatiles such as mercaptans were not detected in mouth air of patients by gas chromatography using the column packed with 30% tricresyl phosphate on Celite 545.
Therfore the volatiles most suspected as the cause of offensive breath, that is amines and mercaptans, were both negative in the oral cavity in situ by the present technique of gas chromatography.

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Various methods by means of which to eliminate or prevent the deposition of dental calculi have been reported and, as a preventive measure, the application of chemicals is being tried. As a step in this direction, the author investigated into the effects of a few proteolytic enzymes on the saliva so as to measure fluctuation in the amounts of Ca both in the supernatant and sedimentation and also to evaluate the proteolytic action of these enzymes on saliva protein in vitro condition.
At the same time, with use of a modified calculus depositing apparatus devised by Yankelowitz and others he mixed the saliva with a variety of proteolytic enzymes and quantitatively measured the deposits thus produced.
As a result of these studies, the author arrived at the conclusions as follow.
1. When fluctuational changes in the amount of Ca both in the supernatant and sedimentation of the saliva mixed with various proteolytic enzymes were observed in the pasage of time, it was found that Ca in the supernatant registered a decrease in the first one hour but suddenly increased in the six hours, beyond which the increase was gradual. On the other hand, Ca in the sedimentation increased proportionally till six hours and the increase then because quite gradual.
2. In terms of effects of the proteolytic enzymes on the amounts of Ca both in the supernatant andd artifically induced calculi, Ca in the supernatant was measured to increase chronologically in proportion to the amount of Ca in the latter.
On the other hand, Ca in the supernatant registered a decrease chronologically when a variety of proteolytic enzymes were added to it.
Rates of these proteolytic enzymes in preventing the dental calculi were in the range of 86% to 53%.
3. The effect of proteolytic enzymes on saliva protein was observed by measuring the amount of none-protein substances produced by supplement of these enzymes into saliva at intervals of one hour, six hours and twenty-four hours. As a result, it was found the amount of none-protein substances increased suddenly at the first one hour, beyond which, the increase was gradual, or no change was noted. On the other hand, in the case of Lysozyme the degree of increase was greater as compared to other enzymes.
On the strength of these experimental data, it may be supposed that, when a proteolytic enzyme is mixed with the saliva, Ca contained in the enzyme will be wrapped up by the protective glutamin action of the saliva protein and, though it deposits itself for a time, this action will be gradually decomposed in the passage of time, thus Ca in the absolute state being gradually transferred from the sediment to the supernatant.
This supposition is endorsed by another experiment in which the addition of a proteolytic enzyme apparently destroyed the agglutinating property of the saliva, with the result that a lesser amount of Ca adhered itself to it.
As a result of these studies, it is to be concluded that there exists a good possibility for the prevention of calculi from depositing on the dental surfaces by the application of proteolytic enzymes.

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