Urinary 6β-hydroxycortisol excretion was confirmed in the manner previously described.
1) The 24-hour excretion of 6β-hydroxycortisol increased in the patients with Cushing's syndrome, in the third trimester of normal pregnancy, after ACTH stimulation and in the patients with adrenal carcinoma maintained with o, p′-DDD.
2) 6β-Hydroxycortisol excretion was within the normal range in the patients with hyperthyroidism.
3) The patients with liver diseases excreted 6β-hydroxycortisol at various levels, but the decreased excretion of 68-hydroxycortisol after administration of 60mg of cortisol occurred in the patients with acute hepatitis. Thit seemed to demonstrate that there was the latent reduction of production of 6β-hydroxycortisol in the liver in patients with acute hepatitis.
4) After administration of 3g of metopirone urinary excretion of 0-hydroxycortisol decreased in the normal subjects, in the patients with Cushing's syndrome and in the subjects maintained with 60mg of cortisol. Metopirone appeared to inhibit 6β-hydroxylation of cortisol in the liver, just as it inhibited 11β-hydroxylation of 11-desoxycortisol in the adrenal.
5) Using the 24-hour excretion of 6β-hydroxycortisol as an index, it was demonstrated that 60mg of cortisol perorally administrated were _excreted within 24 hours even in the patients with liver diseases in whom the prolonged half-life of cortisol had been indicated.
6) Uncorrected values for 6β-hydroxycortisol excretion were within the range from 900 to 1200 μg/24 hr in normal subjects to whom 60mg of cortisol were administrated perorally.
7) The 24-hour excretion of 6β-hydroxycortisol after administration of 60mg of cortisol increased in the subjects maintained with 600 mg/day of cyclandelate, as compared to the control level. Maintenance with both cyclandelate (600 mg/day) and acenocoumarin (2-4 mg/day) gave a further rise to the 24-hour execretion of 68-hydroxycortisol after administration of 60mg of cortisol. Cyclandelate and acenocoumarin appeared to be hydroxylated in the liver and to induce activity of the hepatic hydroxylases that metabolize these drugs and steroids. (Original is written in Japanese.)

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Even when the diagnosis of primary aldosteronism is obvious before operation, it is very difficult to make certain of the location of the adenoma. In this study a direct comparison of aldosterone levels in 13 patients with primary aldosteronism was made among blood samples obtained from bilateral adrenal or renal veins by percutaneous transfemoral vein catheterization during adrenal venography.
The concentration of aldosterone in the venous blood from the uninvolved adrenal glands averaged 56.6 ng per 100 ml of plasma, whereas the concentration in the blood from tumor-bearing adrenal gland was over 790 ng per 100 ml of plasma. The aldosterone levels in blood from the affected glands did not correlate with the size of the adenoma.
In 6 cases it was impossible to obtain blood samples from the adrenal veins, however, the renal-vein blood could be more easily withdrawn than adrenal-vein blood. The ratio of aldosterone levels in left to right renal-vein blood was 1.65 to 5.47 in 4 cases with a left adrenal adenoma. In 6 cases with right adrenal adenoma no marked difference in aldosterone levels was found between the left and right renal veins. On the other hand, the ratio of aldosterone in blood obtained from the inferior vena cava above the entry of the adrenal vein, to that in blood from the left renal vein, was 1.21 to 3.00 in 6 cases with a right adrenal adenoma, and was below 1.0 in 4 cases with a left adrenal adenoma.
Thus aldosterone-producing adenomas were correctly located before operation in all 13 patients with primary aldosteronism by comparison of aldosterone concentrations in blood obtained by percutaneous catheterization of the adrenal or renal veins. This procedure identified very small functional adenoma which could not be demonstrated radiographically, or seen or palpated at surgery. Therefore, it is concluded that differential aldosterone measurement after percutaneous bilateral adrenal- or renal-vein catherization is a definitive test for the localization of an aldosterone-producing adenoma in ambiguous cases.

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Recent studies have indicated that the sympathetic nervous system plays an important role in the release of renin from the kidney. It has also been shown that many of the signs and symptoms of hyperthyroidism are due to increased sympathetic activity and that these manifestations may be ameliorated by adrenergic blocking agents. It was, therefore, of interest to see whether the increased sympathetic activity of hyperthyroidism was associated with increased levels of renin, angiotensin and aldosterone in plasma.
The mean plasma renin activity (PRA) and plasma angiotensin II (A-II) of hyperthyroidism, 2.22 ± 1.67 ng/ml/h, and 98.7±64.6 pg/ml, respectively, was significantly greater than that of normal subjects, 1.70±0.90 ng/ml/h and 53.2±29.3 pg/ml, respectively. The PRA and A-II levels were not correlated with basal metabolic rate, protein-binding iodine and ¹³¹I uptake. Plasma aldosterone levels (PAL) have shown a high levels of 13.5±13.7 ng/dl in hyperthyroidism. On 2-hour upright position after furosemide administration, PRA increased significantly from 2.00±4.03 to 11.34±8.73 ng/ml/h (<0.005) and PAL from 13.7± 14.1 to 33.9±30.5 ng/dl.
From these results, it was assumed that the elevated levels of PRA, A-II and PAL in thyreotoxicosis could be manifested either by sympathetic stimulation via renal nerves or by the sodium-excreting effect of thyroid hormone.

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A 46-year-old housewife was admitted to Tohoku University Hospital on June 2, 1972, because of fatigability, palpitation, weightloss and sweating. She had been administered orally 3.0 mCi of radioactive iodide under the diagnosis of hyperthyroidism on November, 1971.
Physical examination ; Body weight 43.3 kg, body length 146.5 cm. Blood pressure was 128/80, pulse rate 104 and regular. Skin was warm and moist. Exophthalmos and eye signs were not observed. Thyroid gland was slightly enlarged and systolic murmur was audible on it. No abnormalities were found in physical examination of heart, lung and abdomen. Reflexes were normal. Mild finger tremor was noted.
Laboratory findings showed no abnormalities, except that electrocardiogram showed a sinus tachycardia.
Thyroid function tests ; Both BMR and thyroidal 24-hour¹³¹I-uptake were high, + 30% and 52%, respectively. Resin sponge ¹³¹I-T₃-uptake was 38%, serum total and free T₄ were 11.8 μg/dl and 5.1 ng/dl, binding capacity of TBG and TBPA were 17.0 and 254 μg/dl, respectively (all these values were within normal range). T₃ suppression test was negative. Plasma TSH was not detectable by radioimmunoassay. Normal plasma TSH response was observed by the intravenous injection of 500 μg of TRH. Radioimmunoassay of T₃ revealed elevated total and free T₃, their values were 365 ng/dl and 2,800 pg/dl, respectively.
Under the diagnosis of T₃-thyrotoxicosis, daily dose of 10 drops of Lugol's solution was administered to the patient. Twelve days after the commencement of the therapy, BMR, serum total T₄ and T₃ reduced to normal range, +17%, 6.8 μg/dl and 50 ng/dl, respectively. No signs of hyper- and hypothyroidism had been observed in this patient four months after the continuous administration of Lugol's solution.
The findings that plasma TSH was not detectable might show that T₃-hypersecretion was not caused by TSH stimulation in this patient. But the responsiveness of plasma TSH to TRH may indicate that the suppressed TSH secretion was imporving by radioactive iodine admidistration to normal.
There is a report that radioiodine therapy for T₃-thyrotoxicosis seems to induce hypothyroidism easily (Ivy, H.K. et al. : Arch. Int. Med., 128 : 529, 1971). In this respect, inorganic iodide therapy for T₃-thyrotoxicosis seems to be reasonable, because the inorganic iodide reduces the serum T₃/T₄ ratio effectively without causing any irreversible damage to the thyroid gland.

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Human placental lactogen (hPL) with immunological properties similar to those of human pituitary growth hormone (hGH) was initially isolated from human placental tissue by Josimovich and MacLaren. It was demonstrated that hPL possessed prolactinlike activity was capable of stimulating the pigeon crop sac. When immunological investigations with the hPL were conducted a cross reaction between it and an antiserum of hGH was noted.
In recent years a number of workers including Cohen et al., Friesen, Florini et al., Turtle et al. and Catt et al. described techniques for the purification and isolation of hPL.The substance was generally prepared from placental tissue by procedures involving extraction with aqueous alkali followed by precipitation with ammonium sulphate. Subsequently the extract was purified by ion-exchange chromatography using DEAE-cellulose and gel filtration on Sephadex, and they used the pigeon crop sac assay or the immunocrossreactivity to and hGH serum as an index of hPL during purification of the substance.However, in general, the pigeon crop sac assay often showed nonspecific reaction; moreover there was a possibility for the substance to be contaminated with other placental proteins as judged by the immunocrossreactivity only.
We must take into consideration that the materials used in the method reported previously were not homogeneous. Namely, the purification and isolation of hPL was very difficult.
In this paper the author describes the details of a new procedure of purification of hPL from the term placenta, and he uses two indices concerning hPL at every steps of the purification procedure.
Namely, the biological one was the specific lipolytic activity that Tojo, Mochizuki, and Murata had already reported on experiments with rat epididymal fat pads and the immunological one was the crossreactivity against anti hGH serum. Biological potency of hPL was particularly well correlated with the crossreactivity against anti hGH. As the hPL was easily denaturalized during its purification process in acidic fluid, all purification procedures were carried out at 4°C in a cold room, pH of the buffer was consisiently employed for all the procedures in order to prevent deamidation and degeneration.
For protein determination in effluent fractions from columns, either the method of Lowry was used or the ultraviolet absorption of the fractions was read at 278 mμ.
Their conductivity was measured with the direct reading meter, model CDM-2d manufactured by Radiometer, Copenhagen.
The results of individual steps are discussed below.
Extraction of crude hPL : Crude hPL was isolated by the salting out method of Mochizuki's from the alkaline homogenate solution of the term placenta and was used as the starting material for further purification.
Gel filtration on G-50 : The concentrated solution was filtered on Sephadex G-50 in order to equilibrate it for chromatography on DEAE-C. Weak lipolytic activity was excluded from the gel and it appeared in the effluent between 900 ml and 1600 ml, and its precipitation line was faint on Ouchterlony double immunodiffusion.
Ion exchange chromatography : Crude extracts containing hPL were chromatographed on DEAE-C under the conditions of absorption and elution devised for the stepwise or the linear gradient method. Most of the biological activity placed on the column was found in the absorbed protein, and the biologically active protein was recovered clearly at the high conductivity by the linear gradient elution method. Under these conditions, fraction of the lipolytic activity was quantitatively recovered between 8 and 16.0 mMHO, the peak of activity emerging from the column as the gradient reached a conductivity of 12.0 mMHO.
So, it was suggested that hPL had a strong acidic charge. This fraction was purified further on G-100.
Gel filtration : The active fractions from DEAE-C chromatography were filtered by upward flow in Sephadex G-100 column.

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There is a phenomenon in radioimmunoassay that the binding of ¹²⁵I-ACTH with its antibody is increased with an increase of unlabeled ACTH, when a very small amount of ¹²⁵I-ACTH is added to a relatively high concentration of antisera. We have studied further on this paradoxical binding phenomenon in order to clarify the occurrence of the phenomenon.
Both macroglobulin and globulin fractions of antisera obtained by gel filtration gave similar paradoxical curves. Prolongation of the incubation period or incubation with constant shaking did not affect the paradoxical binding phenomenon.
However, the paradoxical binding phenomenon disappeared and the conventional stancard curve was observed when radioimmunoassay was performed with the first piece (Fab fraction) of papain-treated IgG fraction of an antiserum. The paradoxical phenomenon also disappeared after repeated immunization in an animal.
These findings suggest the presence of a kind of allosteric effect in antigen-antibody reaction similar to that in enzyme-substrate interaction, when some specific antiserum is used.
This phenemenon can be used to develop a sensitive and reliable radioimmunoassay for ACTH.

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