Effects of Dietary Marine Fish Oil on Free Radical-Removing Enzyme Activities and Glutathione in Rat Livers (B. LIVING SCIENCE)

In order to elucidate effects of dietary marine fish oil on free radical generation and scavenging, the changes of activities of radical-removing enzymes [GST, GSH・Px, γ-GTP, SOD and xenobiotic-metabolizing enzyme (MFO)] and glutathione were examined on rat hepatic tissues. The diet consisted of 5% (5GM), 20% (20GM) wheat gluten and 20% casein (control) as a protein source. These diets contained adequate amounts of corn oil, minerals, vitamins and corn starch as the other nutrients. In the case of 5GM and 20GM groups, corn oil in the diets was replaced by marine fish oil (Gadus macrocephalus). Food intake, body weight (body weight gain) and liver weight of the 5GM group were significantly lower than those of the 20GM and the control groups, although there were no differences in these parameters between the 20GM and the control groups. For total hepatic GSH contents, low protein diet group (5GM) showed marked decrease, whereas a significant rise of hepatic GSH content was found in the 20GM group. GST activity of the 5GM reduced significantly, although 20GM group activity did not differ from the control level. In the comparison with dietary corn oil and marine fish oil, there was a different evidence that marine fish oil decreased significantly GST activity in low protein diet group and reversely this oil apparently increased it in 20% protein diet group. Enzyme activities of SOD and γ-GTP were not different between the 5GM, 20GM and control groups. GSH・Px activity of the 5GM group increased significantly and MFO activity of 20GM group decreased markedly

Effects of Dietary Fat and Oils on Body Compositions of the Exercised Rats (B. LIVING SCIENCE)

Nutritional effects of dietary lipids on body compositions of the exercised rats were determined. Body weights of the exercise groups significantly decreased as compared with those of the sedentary groups. Crude lipids, especially triacylglycerol, in the liver and carcass decreased significantly due to loading of endurance exercise. Glycogen reserves in the liver tissues were promoted by feeding of plant-or marine fish-oil diets. Protein contents in the carcass had a tendency to increase in corn oil and marine fish oil diet groups. From above results of reductions of crude lipids and triacylglycerol in the animal body, it is suggested that body fat can be apparently consumed as an energy source of the exercise

穀類におけるシステインプロテアーゼインヒビターの存在(B. 生活科学)

トウモロコシ, アワ, ヒエ, 小麦, 大麦の種子および米糠について, システインプロテアーゼインヒビター活性の有無を検索した。各穀類の1%食塩水抽出液を硫安90%飽和して得た粗調製品は, すべて, パパインに対する阻害活性を示した。最も高い阻害活性はトウモロコシに見られ, つづいてはアワであった。他の穀類に認められた活性は弱いものであった。粗調製品のpH7.5,90℃における熱処理からは, トウモロコシ, アワ, ヒエのインヒビターは熱安定性であり, 小麦, 大麦, 米糠のインヒビターは熱不安定性であることが示された。アワの粗インヒビターを用いた予備的実験から, アワ種子には等電点4.9,分子量13,000の少なくとも一種のタンパク質性システインプロテアーゼインヒビターが存在しており, 本インヒビターは比較的アルカリ条件下で安定であることが示された。Seeds of corn (Zea mays L.), foxtail millet (Setaria italica), barnyard millet (Echinochloa utilis), wheat (Triticum aestivum L.), and barley (Hordeum distichum L.), and bran of rice (Oryza sativa L.) have been examined for cysteine proteinase inhibitor activity using papain as a target enzyme. All of the crude samples prepared from the NaCl extracts of these cereals by ammonium sulfate fractionation were found to contain a substance (s) inhibiting papain. The highest inhibitory activity was shown in corn, followed by foxtail millet, and the other cereals had only weak activity. Heat treatments of the crude samples at 90℃ and pH 7.5 revealed that the inhibitors from corn, foxtail millet, and barnyard millet were thermo-stable, whereas those from wheat, barley, and rice thermo-labile. Preliminary experiments concerning foxtail millet indicated that the grain contained at least a proteinaceous cysteine proteinase inhibitor having an isoelectric point of 4.9 and a molecular weight of 13,000,and this inhibitor was relatively stable under alkaline conditions

Activities of Hexosaminidases and Hexosidases in the Brain and Spinal Cord of Rats (B. LIVING SCIENCE)

In the present study, activities of 4 kinds of hexosaminidases and 5 kinds of hexosidases in rat brain and spinal cord were examined. All hexosaminidases in the brain were significantly higher in activity than those in the spinal cord at p>N-acetyl-β-D-galactosaminidase>>N-acetyl-α-D-galactosaminidase. However activity of N-acetyl-α-D-glucosaminidase could be detected in neither brain nor spinal cord. The most active enzyme in N-acetyl-hexosaminidases was N-acetyl-β-D-glucosaminidase. The most active enzyme in hexosidases was β-galactosidase in both brain and spinal cord. And the activity in the latter organ was significantly higher than that of the former. In hexosaminidases, distribution of each enzyme activity in both organs showed the almost similar pattern, although activities themselves of hexosaminidases were higher in brain than those in spinal cord, showing 83∿84% for β-GluN-ase, 12% for β-GalN-ase and 4∿5% for α-GalN-ase. On hexosidase activity, β-galactosidase occupied 30 to 40% of total hexosidase activity in both organs. Furthermore β-galactosidase activity of the spinal cord was higher in both specific activity itself and distribution ratio than those of the brain

Leiomyosarcoma occurring in CRJ-SD Male Rat fed 20% Casein Diet for Long Period. (B. LIVING SCIENCE)

20%カゼイン飼料長期投与CRJ-SD雄ラットの1例に腫瘍が発症した。このラットの体重変動, 症状及び剖検時ならびに組織学的所見を報告する。43週まで体重585gを示して順調に成長していたラットがその後次第に体重減少を来し, 死亡前17日間で体重が約100gも減少し, それにともなって下痢症状が現われた。死後剖検時に右側後腹壁に30×20×15mmの腫瘤を認めた。この腫瘤は限局性で底部は周囲組織に固着していた。顕微鏡観察で大きなblunt-endedな核をもつ紡錘形の腫瘍細胞がみられ, 核はhyperchromaticな部分とvesicularな部分とが見られ, 細胞の異型性が強かった。また分裂像も認められた。一方腫瘤中心部には変性, 壊死像も観察された。以上所所見からこの腫瘍は後腹膜壁に発症した平滑筋肉腫と考えられる。A tumor was found in one of the rats (Charles-River CRJ-SD strain) fed 20% casein diet for long period. The detail of growth, symptom and histochemical findings were presented here. For about 43 weeks the rat grew well showing 585g of body weight. But thereafter it decreased gradually and diarrheas were sometimes observed with severe weight loss, nearly 100g of loss in the last 17 experimental days. The tumor was found in the right-side region of retroperitoneum. Its dimensions were 30×20×15mm. It was sharply circumscribed and fixed to the adjacent muscular tissues. Microscopically, there were elongated spindle-shaped tumor cells with large blunt-ended nuclei which were hyperchromatic and partly vesicular. Tumor cells were very pleomorphic and mitosis was also observed. The degeneration and necrosis were found in the central portion of the tumor mass, and otherwise, the tumor cells infiltrated into the adjacent muscular tissues. From these results, it was suggested that the tumor would be a leiomyosarcoma arising from retroperitoneum in a male rat

塩酸吸着コーンスターチの湿熱処理により調製した難消化性デキストリンのラットコレステロール代謝に及ぼす影響(B. 生活科学)

The effects of indigestible dextrin obtained by heat-moisture treatment of HCl-adsorbed corn starch on cholesterol metabolism were investigated. Four-week-old male Wistar rats were fed diets containing indigestible dextrin of a 5% level with or without cholesterol for 21 days. A fiber-free diet was used as a control. Serum total cholesterol level was significantly decreased in the rats fed the indigestible dextrin diet without cholesterol compared to the rats fed a fiber-free diet, but there was no difference in fecal excretion of bile acids between the dietary groups. Ingestion of indigestible dextrin significantly increased cecal weight and lowered pH of cecal contents. These results suggest that the hypocholesterolemic effect of indigestible dextrin is not explained by the increase of fecal excretion of bile acids

酸性条件下における米糠トリプシンインヒビターの Asp (19)-Pro (20) 結合の開裂(B. 生活科学)

米糠トリプシンインヒビター(RBTI)分子中のAsp (19)-Pro (20)結合の開裂が酸性pH下で非酵素的に生じた。このことは,pH3.0でのRBTIのインキュベーション,開裂したペプチド結合を有する修飾RBTIの単離,さらに修飾インヒビターの還元およびS-カルボキシメチル化によって得られたペプチドの分析などにより明らかにされた。修飾RBTIのトリプシン阻害活性は未修飾のものに比べ著しく低下しており,これはRBTI分子中のAsp (19)-Pro (20)結合の開裂により阻害活性部位周辺の立体構造に変動が生じたためであると考えられた。The cleavage of Asp (19)-Pro (20) bond in rice bran trypsin inhibitor (RBTI) occurred non-enzymatically at an acid pH. This was proved by the experiments such as incubation of RBTI at pH 3.0,isolation of a modified RBTI containing a cleaved peptide bond, and chemical analyses of peptides obtained by reduction and S-carboxymethylation of the modified inhibitor. The trypsin inhibitory activity of the modified RBTI was dramatically weaker than that of native RBTI, suggesting that the cleavage of the Asp (19)-Pro (20) bond in RBTI caused a conformational change in the structure of a reactive-site region

蛋白質性プロテアーゼインヒビターとトリプシンの会合平衡定数測定(B. 生活科学)

トリプシンとインヒビターの会合平衡定数測定のための改良法を確立した。本法は3段階より成っており, 1段階は, 活性部位滴定や酵素-インヒビター滴定によるトリプシンおよびインヒビターの濃度標定, 2段階は, GreenとWork法での滴定のための遊離酵素活性の正確な測定で, ここでは, 低酵素濃度と高感度基質が用いられる。最後の段階は, マイクロ・コンピューターによるデータの非線型最小自乗分析で, これにより正確に定数を評価できうる。この方法により, ウシおよびブタ・トリプシンと粟種子トリプシンインヒビターIIとの会合平衡定数は, 25℃, pH8.0で, それぞれ3.3×10^M^, 4.6×10^9M^と決定された。An improved procedure for the measurement of association equilibrium constants of trypsins with a protein proteinase inhibitor has been established. The procedure consists of three stages : (a) standardization of trypsin and inhibtor solutions by the active-site titration with p-nitrophenyl-p\u27-guanidinobenzoate and by an enzyme-inhibitor titration at a high concentration (∿10^M), (b) second is the accurate measurement of free enzyme activities for Green and Work titrations, where low enzyme concentrations (∿10^M) and a highly sensitive substrate are used, and (c) nonlinear least-squares analyses of the data by a microcomputer, which make it possible to accurately estimate the constants. By this procedure, the association equilibrium constants of bovine and hog trypsins with foxtail millet trypsin inhibitor II were determined to be 3.3×10^M^ and 4.6×10^9M^, respectively, at 25℃ and pH 8.0

ラット肝臓カテプシン活性に及ぼす食餌タンパク質レベルの影響(B. 生活科学)

幼ラットにカゼイン含量の異なる飼料を与え, 21日間飼育し, 体重増加量, 肝DNA, RNA, タンパク質含量に加えて肝カテプシン活性を測定した。3週間の飼育において, 最大成長は20%およびそれ以上のカゼイン含量の飼料を与えられたラットに認められた。肝カテプシンB, H, Lの総活性は, 飼料中のカゼインレベルが0%から20%に増加すると共に増加してほぼ最大値となった。この値は, 飼料中のカゼインレベルがそれ以上増加してもあまり変らなかった。このカテプシン活性の飼料中カゼインレベルに対する応答は, 肝重量, 肝タンパク質, 肝RNAの応答と類似していた。Male weanling rats were fed diets containing various levels of casein for 21 days and liver cathepsin activities in addition to body-weight gain and hepatic DNA, RNA, and protein contents were measured. After three weeks of feeding, maximum growth was observed in animals fed diets containing 20% or more casein. Total activities of liver cathepsin B, H, and L increased to nearly maximal values as the level of casein increased from O to 20% in the diet and the values did not change greatly with further increasings of dietary protein level. This response was similar to those of liver weight, liver protein, and liver RNA

タンパク質源の異なる飼料で飼育したラットの肝臓におけるプロテアーゼ活性レベル(B. 生活科学)

タンパク質レベル10%, タンパク質源として卵白アルブミン, カゼイン, SPI, グルテン, ゼインの5種を用いる飼料を幼ラットに21日間与え, 体重増加量, 肝タンパク質, DNA, RNA含量, さらに肝臓のプロテアーゼ活性レベルを調べた。21日間の飼育において, アルブミン区が最も良く成長し, つづいてカゼイン, SPI区の順であった。一方, グルテンおよびゼイン区では初体重を維持することができなかった。肝臓の総タンパク質量, 総RNA量については, アルブミン区が最大値を示し, つづいてカゼイン, SPI, グルテン, ゼイン区の順となった。肝総DNA量は, 各区間に有意差は認められなかった。肝リソソーム画分のカテプシン活性レベルは, アルブミン, カゼイン, SPI区で高く, グルテン, ゼイン区で低い傾向が見られた。同様の傾向が肝サイトソール画分のプロテアーゼ活性でも観察された。これらの結果より, 肝臓のプロテアーゼ活性レベルは異なる生理的条件下で変化すること, すなわち, 概して, 栄養価の高いタンパク質で飼育したラットの活性レベルの方が低栄養価のタンパク質で飼育したものより高くなることが示唆された。しかしながら, 活性レベルと栄養価を反映する体重増加量の間には決ずしも明確な相関は見られなかった。Male weanling rats were fed diets which differed in source of protein for 21 days and body-weight gain, hepatic DNA, RNA, and protein contents, and levels of liver proteinase activities were estimated. Albumin from egg white, casein, soybean protein isolate (SPI), gluten, and zein were used as sources of dietary protein. After three weeks of feeding with 10% level of protein diets, the best growth rate was yielded by an albumin-fed group and the second best rate was shown by a casein group, followed by a SPI group. On the other hand, a gluten-fed group and a zein group did not maintain their initial body weights. Liver RNA and protein contents of the albumin, casein, and SPI groups were significantly higher than those of the gluten and zein groups, though hepatic DNA contents were not different among the diet groups. Levels of total liver cathepsin cativities tended to be higher in the albumin, casein, and SPI groups than in the gluten and zein groups. The same tendency as above was also shown in the levels of cytoplasmic proteinase activity in livers. These results suggest that the levels of liver proteinase activities change under a distinct physiological condition. That is to say, the levels were in general higher in rats fed a protein with high nutritional quality than in rats fed a protein with low quality. However, there was not necessarily strict correlation between the activity levels and weight gain