Rat livers were perfused for 4 hours, and bile-duct ligation and the addition of bile acids such as chenodeoxycholate (CDCA), ursodeoxycholate (UDCA), deoxy-cholate (DCA) and cholate (CA) into the perfusion medium were conducted 30 min after the start of perfusion. Activities of the following enzymes released into the perfusate were determined at intervals of 30min during the per fusion: lactate dehydrogenase (LDH₅), malate dehydrogenase (MDHs), glutamic-pyruvic transaminase (GPT) and α-hydroxybutyrate dehydrogenase (α-HBD) which are distributed in hepatic soluble fraction, and malate dehydrogenase (MDHm) and glutamic-oxaloacetic transaminase (GOTm) which are distributed in the mitochondrial fraction. The perfusion of livers with bile-duct ligation caused a much higher time-dependent increase of all enzyme activities in the perfusate than that of control livers, in which LDH₅, MDHs and MDHm showed a marked increase in activitis as compared with GPT, α-HBD and GOTm. The perfusion of livers with the medium containing 0.01% CDCA, 0.01% DCA, 0.05% UDCA or 0.05% CA caused the time-dependent increase in activities of all six enzymes in the perfusate just as the case of bile-duct ligation. CDCA and DCA showed a stronger effect on release of enzymes from liver than UDCA and CA.
The present results suggest that the injurious effect of bile-duct ligation on liver cells may be partly due to the cytotoxic effect on bile acids accumulated in the tissue, and that CDCA and DCA have a more toxic effect on liver cells than UDCA and CA, in which these damages by bile acids occur not only in plasma membranes but also in mitochondria. In addition it was indicated that both LDH5 and MDHs, and MDHm are marker enzymes sensitive to the bile acid-induced damage of liver plasma membranes and mitochondria, respectively.

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The serum Lp (a) lipoprotein concentration in patients with hepatobiliary disorders was determined by single radial immunodiffusion, and alterations in concentration during the clinical course were observed. The Lp (a) concentration was significantly decreased in patients with liver cirrhosis or hepatocellular carcinoma (p<0.001), with fulminant hepatitis (p<0.01), and with acute hepatitis or obstructive jaundice (p<0.05). In general, the Lp (a) level did not vary so much if the liver function was stable.
From observation of the clinical course in hepatobiliary disorders, the Lp (a) concentration was found to vary inversely with changes in the GPT level, which is a parameter of liver cell damage. Its concentration roughly paralleled that of other liver function parameters. Therefore, Lp (a) is considered to be a lipoprotein whose concentration is able to change according to the degree of liver cell damage, and the data strongly suggest that Lp (a) is, at least in part, synthesized in liver cells.

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Urinary 17-Ketosteroid Sulfates (17-KS-S), 17-KS Glucuronides (17-KS-G) and 17-Hydroxycorticosteroid Glucuronides (17-OHCS-G) were determined in patients with Cushing's Syndrome, with Addison's Disease (administered with high amounts of cortisol), with nonendocrine diseases etc. and normal adults.
In the above subjects 17-KS-G correlated well with 17-OHCS-G, but 17-KS-S did not correlate: Dissociation of 17-KS-S and the former two excretion was observed.
Measurement of urinary 17-KS-S, consisting of storage form adrenal androgens, Dehydroepiandrosterone-S (DHA-S), Androsterone-S (A-S), etc., is useful not only for diagnosis of adrenocortical diseases, but for determining severity of non-endocrine diseases.

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In order to estimate the hepatotoxicity of bile acids such as chenodeoxycholate (CDCA), deoxycholate (DCA), ursodeoxycholate (UDCA) and cholate (CA), we studied the effect of these bile acids on the release of various cytosolic and mitochondrial enzymes and protein into the incubation medium and on cell mortality in liver cells isolated from adult male rats.
Liver cells showed little change either in the release of cytosolic enzymes such as lactate dehydrogenase (LDH), glutamic-Pyruvic transaminase (GPT) and malate dehydrogenase (MDHs) and mitochondrial enzymes such as malate dehydrogenase (MDHm) and glutamic-oxaloacetic transaminase (GOTm), and protein immediately after addition of bile acids to the medium. When liver cells were incubated with each bile acid at 37°C for 60 min, both hepatic cell mortality and the ratio of enzymes and protein from the cells increased concomitantly with an increase in the concentration of bile acids added. There was a clear difference in the abilty to cause these changes among the four bile acids: COCA>DCA> UDCA>CA. Of hepatic cytosolic and mitochondrial enzymes determined in the medium under these conditions, MDHs showed the highest activity, while LDH and GPT were higher in the ratio of enzyme release than MDHs, MDHm and GOTm. In addition, the ratios of LDH and GPT activities in the medium total activities in the cells increased almost in accordance with cell mortality. It was also found histologically that the plasma membrane and mitochondria of liver cells were damaged by treatment with bile acids at 37°C for 60 min.

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Some substrates for estimation of serum cholinesterase were synthesized from the derivative compounds of benzoic acid and naphthoic acid.
3, 4-dihydroxybenzoyl choline was specific for pseudo-cholinesterase activity and was hardly hydrolyzed in buffer solution. The disappearance of substrate was measeured by ultraviolet spectrophotometry at 340 nm. Both Within-run and day-to-day imprecision were between 0.6 and 1.4%. Addittion of 18 substances to sera did not almost effect the determination of pseudo-cholinesterase activity.
A good correlation was observed between the present method and the Kalow's method using benzoylcholine as substrate.

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A fluorescence high-performance liquid chromatographic (HPLC) method for determinatination of urinary 17-hydroxycorticosteroids (17-OHCS) is described.
Urinary 17-OHCS and 3β, 5α-tetrahydrhydrocortisol (3β, 5α-THF, internal standard) were labelled quantitatively by treatment with dansylhydrazine and the derivatives were clearly separated on Cosmosil 5SL column by use of a mixture of dichloromethane-ethanolwater (88: 5: 7) as a mobile phase. The analytical recovery of added 17-OHCS was almost 100 percent and the reproducibility was also satisfactory. The present method was found to be applicable to urine samples obtained from patients on drug medication such as carbamazepine, which interferes with the determination of urinary 17-OHCS by conventional colo-rimetric methods.
The present method is clinically useful for routine assay of urinary 17-OHCS.

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Furosine (ε-N-(2-furoylmethyl)-L-lysine), which is an acid-hydrolysis product derived from the Amadori compoud fructose-lysine (ε-N-(1-deoxy-D-fructose-1-yl)-L-lysine) was used in determining the nonenzymatic glycation of tissue proteins. The furosine level in the aorta obtained at autopsy was significantly higher in the aged (over 65 years) than in the neonate. A significant positive correlation was found between the visually estimated severity of sclerosis of the aorta and the furosine level in the aorta in the aged.
These results suggest that an increase in nonenzymatic glycation in the human aorta may be an etiological factor of arteriosclerosis.

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Simplified method for extracting leukotrienes (LT's) from cell culture medium was developed, and conditions for their analysis by HPLC were studied. LT, s released into the medium from A23187-stimulated human WBC's and rat alveolar macrophages were extracted and concentrated by passage through a Sep-Pak C₁₈ cartridge after deproteinization with methanol. Human WBC's as well as rat alveolar macrophages secreted only LTC₄ and LTB₄ into the incubation medium. Productions of LTB₄ and LTC₄ were dose-dependently stimulated by calcium ionophre (A23187) at up to 10^-5M (final conc.) Production of LTB₄ reached the maximum value after 15 min incubation and then decreased. On the other hand, the amount of LTC₄ in the medium increased throughout the incubation period of 30min. The appropriate conditions for HPLC analysis on three different ODS columns were determined by changing the ratio of solution A in the eluent (A, acetonitrile; B, Methanol: H₂O: acetic acid/100: 300: 1, pH 5.6, containing 0.08% EDTA) from 29 to 34% with the use of 2 pumps. Retention times were prolonged and peaks of LT's flattened and widened by decreasing the percentage of solution A in the eluent. And by increasing the ratio of solution A, the peaks became sharper and taller and retention times of LT's shortened, however, separation of peaks became incomplete. The best resolution was obtained with a Novapak C₁₈ column and eluent containing 32-33% solution A. Recovery ratios of standard LT's (LTB₄, LTC₄, LTD₄) were more than 95% except for LTE₄ whose recovery was 67.1% through the whole procedures of extraction and HPLC analysis. Relationship between area of peak and amount of LT was linear from 0.5 up to 4ng of each standard sample. We conclude that measurement of LT's by HPLC depends in large part on the quality of the HPLC column, and we obtained the best result by the procedures described here in using a Novapak C₁₈ column.

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