Actual changes in tissue partial pressures of each gas and tissue perfusion with serial artificial CO₂ bathing were evaluated by means of medical mass spectrometry using 5 rabbits.
An artificial CO₂ bath was prepared by adding “BUB”-KAO, a 50g sodium hydrogencarbonate and succinic acid tablet producing fine CO₂ bubbles in water of constant temperature 20-litre tub at 36-37°C.
Regional tissue perfusion volume was determined on the basis of a clearance curve for Argon tissue partial pressure which was monitored by an on-line computer system with mass spectrometry.
Increase in subcutaneous tissue PCO₂ changed from 27% to 10%, and in PO₂ from 12% to 5% on average by serial bathing every day for 4 weeks, on the other hand tissue perfusion volume was increased in 3 out of 5 cases; from 20.85±3.56 (X±SE) ml/100g/min, SD=6.71, p<0.05 to 25.23±8.00 (X±SE) ml/100g/min, SD=6.96, p<0.05.
CO₂ has been shown to be locally absorbed through the skin resulting in the elevation of subcutaneous tissue CO₂ partial pessure which decreases depending on the tissue perfusion irrespective of metabolic processes. It is, therefore, assumed that the elimination of absorbed constituents by serial bathings is not caused by a decrease in the percutaneous absorption rate but an increase in washing out rate by the improved tissue perfusion.

Artificial CO₂-bath was prepared with a tablet (50g): made from sodium bicarbonate and succinic acid, putting simply in plain water bath tub of 100-200 litre at 40C.
The effect of artificial CO₂-bath was studied in relation to the change in blood lactic acid which is thought to be indicative of the physical fatigue.
It has been clearly demonstrated experimentally using rats that the blood lactic acid after exercise is significantly reduced as compared both to a plain and an artificial Na₂ SO₄-NaHCO₃ bathing.
A favorable effect of the artificial CO₂-bath was also confirmed clinically by a relief from the stiff pain following maximal abdominal muscle exercise.

Anti-inflammation effect of artificial CO₂-bathing was confirmed on carrageenin induced edema experimentally using rats.
Artificial CO₂-bath was prepared with carbonated water by high pressure CO₂ bubbling and with a 50g tablet, made from sodium bicarbonate and succinic acid, putting simply in plain water.
Carrageenin induced edema was suppressed by plain water bathing, and was further enhanced at lower temperature.
It has been definitely shown that the edema is significantly suppressed with CO₂-bathing as compared to plain bathing due possibly to improved tissue perfusion.

Sodium succinate is remained behind following artificial CO₂ bathing, prepared with a 50g sodium hydrogencarbonate and succinic acid tablet producing fine bubbles in water.
2NaHCO₂+(CH₂)₂→(COOH)₂→(CH₂)₂(COONa)₂+H₂O+CO₂
Na₂CO₃+(CH₂)₂(COOH)₂→(CH₂)₂(COONa)₂+H₂O+CO₂
Balneotherapeutic effect of sodium succinate was evaluated experimentally using 10 rabbits by means of medical mass spectrometry.
Subcutaneous PO₂ and PCO₂ did not change during the bathing and tissue perfusion of 21.30±3.48ml/100g/min (mean±standard error), SD=4.94, p<0.05 was evaluated at 36-37°C, demonstrating no significant difference compared with that of 20.85±3.56ml/100g/min, SD=6.71, p<0.05 by plain water bathing.
These data showed clearly that the balneotherapeutic effects of an artificial CO₂ bathing prepared with a CO₂-tablet, “BUB”-KAO, was not from organic salt of sodium succinate but from CO₂.

It has been well recognized that no known method can produce as strong or as effective a bath as can be given with natural waters.
The method of bubbling gas through the bath water is thought to be almost entirely ineffective; the solubility of CO₂-gas in water at 40°C is about 10%, but is much further enhanced about 50% by bubbling through an airstone, making fine bubbles.
Tablet form of the CO₂-bath preparation, made from bicarbonate and succinic acid, produces fine CO₂ bubbles so that about 80% of water solubility at 40°C can be obtained.
A CO₂-bath preparation, 50g tablet, can produce 100ppm in maximum concentration in a bath of 150 liter at 40°C, maintaining the level of more than 60ppm for 2 hours.
In order for the safety to estimate the escaped CO₂, a large quantity of the CO₂-bath preparation, 20 and 50 tablets, was used in a relatively air tight room; CO₂ concentration reached the maximum level of 1.9 and 6.6% respectively with a tendency of rapid falling, so that the critical complications could rarely be happened.

Increase in dermal blood flow by the artificial CO₂-bathing was confirmed by means of a Thermocouple flow meter and by a Laser doppler velocimeter.
The artificial CO₂-bath was prepared with 50g tablet, made from sodium bicarbonate and succinic acid, putting simply in plain water at 38°C.
Dermal blood flow was increased nearly 5-fold by the simple bathing, and was further enhanced 1.3-fold by the artificial CO₂-bathing.
It has been definitely shown by the artificial CO₂-bathing that an increase in oral, finger tip, and forehead temperature and transepidermal water loss is significant compared to the plain bathing, so that the thermal effect equivalent to carbon-dioxated spring will be obtained.

It was found that no effect of increase in dermal blood flow depended solely on HCO₃^- or CO₃^2-, but on CO₂ gas dissolved in water.
The artificial CO₂ bath was prepared with sodium bicarbonate and citric acid, mixing simply in plain water at 38-40°C.
Thermal efficacy was confirmed by a rise in temperature of oral, finger tip and forehead respectively, and by a high transepidermal water loss (TWL) in consequence of increase in dermal blood flow, compared to a plain bathing.
A remarkable effect in artificial CO₂ bathing of 400-800ppm has been well-known clinically, however, a substantial increase in dermal blood flow has observed with artificial CO₂-bathing of 59.8ppm or greater in this study, so that a CO₂-bathing using the preparation would be popularized.