Studies on Paper-analysis in the Field of Balneology. (II) 1. A New Method for the Determination of Cobalt in Blood. 2. Effects of Radioactive Thermal Bathing and Internal Use of Vitriol Water upon Cobalt Levels in Blood

1. A new method for the determination of cobalt in blood. The author proposed a new method for the determination of cobalt in blood by means of paper-chromatography. The procedure is follows. In a Kjeldahl colben, 5 m!. of blood is taken, and turned into wet ash with 3 ml. of HNO(3), 0.5 ml. of H(2)SO(4) and 1.5 ml. of HClO(4) and the ash aqueus solution is then neutralized with ammonium hydroxid, the neutralization being indicated by the development of color of 0.1% of p-nitrophenol (one drop) added to the solution. After addition of 2 ml. of 40% ammonium citrate, 0.5 ml. of 20% Na(2)S(2)O(3), 3～5 drops of H(2)O(2) and 1 ml. of o-nitrosoresorcine monomethyl ether (hereafter N.R.M.E.), the mixture is left for at least 15 minutes and the aqueus solution is taken in a stoppered test tube (inside diameter: 1.3 cm., hight: 16.5 cm.). The mixture is shaken with 2 ml. of carbon tetrachloride. Carbon tetrachloride is shaken with 5 ml. of 20% Na(2)CO(3) solution. Then the excess of N.R.M.E. is removed and carbon tetrachloride is concentrated on water bath to 0.3～0.5 ml. and all CCl(4) is used for a determination sample. The sample is placed on the paper (Toyo-filter paper No. 50 or 51A) in a thin line at a distance of 3 cm. from the edge, and the spot is developed with a mixture of 20 ml. of CCl(4) and 1 ml. of 90% ethylalcohoI. After development for 30～40 min., an orange-colored linear spot appears at a distance of 9～10 em. from the origin point. Spot intensity is measured at 460 mμ with Natsume's densitometer and quantitative estimation is made comparing with the standard graph. 2. Effects of radioactive thermal spring bathing and internal use of vitriol water upon cobalt levels in blood. As the author's method above-mentioned is able to determine the cobalt level in a small amount of blood, it is very convenient in investigating cobalt metabolism in the field of balneology. The author examined the changes of cobalt levels in blood after radioactive thermal bathing as well as after internal use of acid vitriol water. a. By the author's method cobalt in normal human blood was 0.4～3.6γ/100ml. in man (average: 1.6±0.4γ/100ml.) and 0.2～2.2γ/100ml. in woman (average: 0.7±0.3γ/100ml.). b. The cobalt levels in blood were measured by the author's method before and 5, 30, 60 minutes after the radioactive thermal single bathing, and before and 1, 2, 3, 4 weeks after a series of radioactive thermal baths (Rn-content: 10～30 Mache, 42～3°C., for 10 minutes). The cobalt levels in the blood of healthy subjects showed no significant change after the thermal single bathing, but those of patients with rheumatoid arthritis showed a slight increase on 3rd or 7th day and a fall on 2nd week of a series of radioactive thermal baths and then tended to return to the initial levels during the 3rd and 4th weeks of spa treatment. c. Thirty ml. of Yanahara mineral water (an acid vitriol water, pH: 2.2) was diluted with plain water to 200 ml. (Co(++)content: ca.3γ) and administered to healthy fasting subjects orally. Blood samples were taken from the cubital vein before and 1, 3, 6 hours after the intake of the vitriol water. The cobalt levels in the blood showed an increase of 14% of the initial levels at 1 and 3 hours and a decrease of 22% at 6 hours after the drinking of Yanahara mineral water.