To solve metabolic uncertainties on the oxidative degradation of choline, some properties of choline dehydrogenase (EC 1. 1. 99. 1), initial enzyme in the pathway, were investigated. Choline dehydrogenase activity was detected only in the liver and kidney, where the specific activities were almost same, and hence, about 80% of its total activity was found in the former. Subcellular localization was re-confirmed since the activity found in the mitochondrial fraction and the negligible amounts were found in neither purified nuclei nor microsomes.
The enzyme could be solubilized in about 50% yield by successive treatments of rat liver mitochondria with ultrasonication and 2% of Triton X-100, however, once the detergent was deleted from the enzyme preparation so solubilized, the activity decreased considerably rapid. The solubilized enzyme was sta-bilized without disturbing an oxygraphical assay for choline dehydrogenase activity by adding 1/15 (mg) of 5, 5′-dithiobis- (2-nitrobenzoic acid) per mg protein of the preparation.

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When DMA (dimethylamine) was added to the mixtures of 5 to 7% brewed food such as shoyu (soy sauce), miso (soybean paste), wine, vineger or mirin and nitrite at pH 3.6, the formation of NDMA (N-nitroso dimethylamine) was markedly reduced. Especially, addition of shoyu, miso or acid hydro-lysates of proteins was most effective of all for the suppression of nitrosation ranging from 60 to 70%.
A study was made on the mechanism of the suppression of nitrosation by shoyu, using various methods such as analysis on chemical components of shoyu and fractionation of shoyu components with Amberlite CG-120, IR-45 and ultrafiltration.
Occurrence of Van Slyke reaction between α-amino acid and nitrite in the reaction mixture containing DMA, shoyu and nitrite was confirmed by the decrease of the amino acids in the shoyu, with accompanied formation of α-hydroxy carboxylic acid and nitrogen gas during the reaction. Furthermore, reactivity of glutamic acid as a typical amino acid in shoyu against nitrite was found to be much faster than that of DMA at pH 3.6. It was, therefore, concluded from the the results that Van Slyke reaction suppressed nitrosation by competitively interfering with the reaction of DMA to nitrite.

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A new colorimetric determination was devised for estimation of isothiocyanate content in radish root (Raphanus sativus L.) juice. Allylthiourea (AU) and trans-4-methylthio-3-butenyl thiourea (MBU) were prepared from allylisothiocyanate and Raphanus sativus L. cv. Aokubi Miyashige Daikon respectively.
It was observed that both thiourea derivatives turned blue with 25-fold diluted Grote reagent. In the absorption spectra of the colored solutions of AU and MBU, both peaks were observed at 605nm. After examination of several different conditions, new method based on the colorimetry of thiourea derivatives was established as follows. Each mixture of 1ml AU or 1ml MBU and 4ml modified Grote reagent was incubated at 37°C for 45min, and the optical density was determined at 600nm. The absorbancy was proportional to MBU concentration in the range between 20μg/ml and 200μg/ml.
After preliminary experiments, a suitable method was found to determine the isothiocyanate content in radish juice. Juice from radish root was kept at 30°C for 30min. To convert produced isothiocyanates into thiourea derivatives, a mixture of ethanol and aqueous ammonia was added to juice, followed by maintaining at 30°C for 60min. The mixture was neutralized with 50% acetic acid and filtered. An aliquot of the filtrate was employed for the determination of MBU as described previously. The resulting value was calculated as trans-4-methylthio-3-butenyl isothiocyanate.
Using the new method, comparison of the concentration of pungent principles in radish roots was performed, in respect to the differences in varieties and maturities as well as sections of the root.

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Free and combined amino acids in the ethanolic extracts of various spontaneous edible mushrooms were determined by liquid chromatography, prior to the determination of proteinaceous amino acids. The following results were obtained.
1. Total nitrogen in the extracts significantly differed in 28 specimens of spontaneous mushrooms subjected to the present study. They deviated in a range of several hundreds to 6, 000mg per 100g of dry fruiting bodies. Percentage of free amino acid nitrogen to total nitrogen was commonly under 30%. Mushrooms belonging to Tricholomataceae, Agaricaceae and Phylacteriaceae showed considerable superiority in the contents as 700 to 1, 600mg. Nitrogen in combined form was, on the other hand, less than that in free form, ranging from 20 to 400mg.
2. Total free amino acids in these mushrooms varied widely from 10 to 800μmol/g dry materials. In these mushrooms, alanine, glutamic acid, threonine and serine were found commonly to be containedin a high amount as free form. On the contrary, cystine, methionine, tyrosine, phenylalanine, histidine and arginine were significantly low in their levels.
3. Similarity of the pattern of free amino acids found to range within 0.80-0.99. The evidence suggests that the mutual similarity in the free amino acid patterns exists in the species subjected. Rather low similarities were observed, however, in several species, Laccaria laccata, Armillariella mellea, Lentinus edodes and Ramaria spp.
4. The amount of combined amino acid in ethanolic extracts of these mushrooms was less than 30% of the amount of free form, whereas several species were found to contain combined form equal to or more than that of free form. Glutamic acid was the most abundant as combined amino acid and aspartic acid and glycine were also commonly found.

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On the fruiting bodies of most common edible mushrooms in Japan, Pleurotus ostreatus, Pholiota nameko, Tricholoma matsutake, Lyophyllum aggregatum and Flammulina velutipes, free and combined amino acid patterns were examined regarding to the differences between spontaneous and cultivated pecimens and among cultivated ones. The results are as follows.
1. The distribution patterns of free and combined amino acid were quite similar in each species. No difference was found also in the content and distribution of these amino acids among cultivated specimens of P. ostreatus and P. nameko, cultivated industrially at various place, and among spontaneous T. matsutake differed from the harvesting place.
2. The spontaneous specimens of P. ostreatus and P. nameko were found to contain free amino acids less than the cultivated ones contained. Little difference was observed, however, between the spontaneous and cultivated specimens in L. aggregatum and F. velutipes.
3. Free amino acids contained in the pilei of T. matsutake were significantly higher than those in the stipes. The large fruiting bodies of P. ostreatus, cultivated in the authors' laboratory, showed somewhat lower content of the amino acids than the small ones did. P. ostreatus, cultivated in a poor nutrient supplementation, however, contained extremely low level of free amino acids.
4. The following fact was observed in P. nameko throughout its spontaneous and cultivated specimens; little amount of aromatic and basic amino acids was found as free form whereas the amount as combined form was considerably high. The above stated fact should be mentioned as a remarkable nature of this species.

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The diurnal rhythms of intestinal digestive enzymes were observed in streptozotocin-induced diabetic rats. Rats were raised under artificial light (light; 9: 00-21: 00, dark; 21: 00-9: 00) and fed during the dark. Water was given ad libitum.
Diabetic rats showed high concentration of blood glucose and urinary glucose. The body weight gain of diabetic rats was less than that of normal rats. However, the wet weight of intestinal mucosa in diabetic rats was significantly increased as compared with normal rats, especially in the dark.
A typical diurnal rhythm of sucrase activity, specific and total activity, which is low in the light and high in the dark, was observed in normal rats. On the other hand, in diabetic rats, the specific activity of sucrase increased in the light, and was similar or decreased in the dark compared with normal rat, especially in duodenum and jejunum. Therefore, the diurnal variation of specific activity of sucrase was smaller in diabetic rats than in normal rats. However, a typical diurnal rhythm was shown in total activity. Isomaltase activity showed similar pattern as sucrase activity.
Alkaline phosphatase activity showed a diurnal rhythm in normal and diabetic rats.

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Total content of sterols and their composition in the ethanolic extract from frozen Antarctic krills (Euphausia superba) were investigated.
The contents of total sterols and free sterols were determined as 0.31% and 0.27% on wet weight respectively by the use of Zak and his research groups method.
Free sterols were isolated from the ethanol extract as a white crystalline form, using frorisil column chromatography (solvent: hexane containing 15% ether), and subjected to GC-MS analysis. The major component was cholesterol and amounted to 97.4%. Stigmasterol (0.01%), β-sitosterol (0.4%) and other five kinds of sterols (C₂₆-C₂₉) were estimated as the minor components.

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Hydrolysis of glucosyl-sucrose (G₂F) and maltosyl-sucrose (G₃F) was studied with disaccharidases solubilized by papain and purified sucrase-isomaltase complex from rat intestinal mucosa.
Chromatographic profiles on Sephadex G-200 column of papain-solubilized disaccharidases revealed that almost hydrolyzing activities of G₂F and G₃F were detected with maltase activity which is glucoamylase, and a little activity was detected with sucrase-isomaltase activities. Furthermore, it was demonstrated that using purified sucrase-isomaltase complex, G₂F and G₃F were not initially hydrolyzed between glucose and fructose. These results suggest that G₂F and G₃F were initially hydrolyzed from the glucose of non-reducing end by maltase, thereafter, the produced sucrose was finally hydrolyzed to glucose and fructose by sucrase-isomaltase complex.

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