To elucidate the effect of aging on the vitamin B₁₂ ( B₁₂) transport in enterocytes, young and old (3-4 months and over 1.5 years, respectively) female rats were studied. Two units of rat intrinsic factor (IF), saturated with ⁵⁷Co-labeled cyanocobalamin were orally administered, and the amount of B₁₂ absorbed into each subcellular fraction of enterocytes was assayed. Concentration of endogenous B₁₂ in each subcellular fraction was also studied. Absorption of radioactive B₁₂ in the lysosomal fraction was maximum between 2 and 4 hr in each age group. In the older rats, the amount absorbed was lower than in the young rats. The older rats showed a significantly lower value of endogenous B₁₂ in the mitochondria. It has already been reported by us that there exist two B₁₂ binders in enterocytes: lysosomal and microsomal binders. The concentrations of lysosomal and microsomal binders as well as B₁₂ uptake in the mitochondria were significantly lower in the older rats than in the young rats. These data might help to explain the lower B₁₂ absorption in the lysosomal fraction and lower B₁₂ contents in the mitochondrial fraction.

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The dietary effect of ω-3 type highly polyunsaturated fatty acid concentrate (PUFA mix) was investigated on cholesterol and triglyceride levels in the serum and liver, and high density lipoprotein cholesterol (HDL-cholesterol) levels in the serum. The PUFA mix prepared from squid liver oil contained about 75% of total ω-3 type fatty acids. In rats fed the normal diet to which 300 PUFA mix was added, the levels of triglyceride, total cholesterol and phospholipid in the serum markedly decreased as compared to rats fed a 3% methyl-oleate diet. However, lipid peroxide values in the liver and serum increased in rats fed PUFA mix-diets. In the hypercholesterolemic rats, a 5% PUFA mix-diet caused growth retardation and a corresponding reduction in food intake. Lipid peroxide in the liver and serum were more elevated in rats fed diets containing 1, 3 and 5% PUFA mix than in rats fed diets containing 5 oleate or 500 linoleate. In all the rats on PUFA mix-diets, there were depression of serum total cholesterol and elevation of serum HDLcholesterol. The ratio of HDL-cholesterol to total cholesterol increased in proportion to the amounts of dietary PUFA mix. Total cholesterol level in the liver was depressed after PUFA mix feeding.

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The effect of long-term (7-9 months) feeding of a diet containing iodine-enriched eggs, which had considerably higher amounts of iodine than ordinary eggs, was studied on the lipid metabolism in rats. A laboratory powder chow was added at 100 (w/w) level with ordinary egg powder (Ordinary egg diet as control: 35.1μg iodine/100g diet) or iodineenriched egg powder (Iodine-enriched egg diet: 392.2μg iodine/100g diet). The animals were meal-fed twice a day and allowed to take daily voluntary running exercise in wheels for 24 hr. The long-term feeding with iodineenriched eggs did not influence body weight gain, food intake, voluntary running activity, weights of several organs and adipose tissues, and lipid concentrations in several organs and tissues. However, the serum lipid levels were decreased after the feeding of iodine-enriched eggs. A decrease in triacylglycerol (TG) level was significant (p<0.05). The long-term feeding with iodine-enriched eggs resulted in a reduced hepatic-intestinal TG production rate and also an enhanced TG removal rate from blood. Furthermore, skeletal muscle and adipose tissue lipoprotein lipase activity was significantly elevated with the feeding of iodine-enriched eggs. These results suggest that the lowering effect of blood TG level with feeding of iodine-enriched eggs may be mediated by either a reduced hepaticintestinal TG production or an enhanced peripheral removal of circulating TG. Serum total iodine concentration was 5 times higher in the iodineenriched egg fed rats as compared to controls, but the serum concentrations of thyroid stimulating hormone and thyroid hormones as well as the thyroid total iodine content were not different between the two dietary groups of animals. Thus, the present study did not demonstrate any relationship between the hypotriacylglycerolemic effect of iodineenriched eggs and the serum levels of thyroid relating hormones.

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To identify the synergistic hypocholesterolemic substance found in soybean oil unsaponifiable matter, rats were fed diets containing various fractions of the unsaponifiable matter prepared by silicic acid column chromatography. The plasma cholesterol level of the group fed the alcohol fraction, which mainly consisted of triterpene alcohols, was significantly lower and the effect was synergistic with soysterol. So the effect of cycloartenol and 24-methylenecycloartanol, which are main constituents of triterpene alcohols in soybean oil unsaponifiable matter, was investigated. Both compounds were prepared from γ-oryzanol (ferulate) and were added (0.05 %) respectively to the experimental diet containing O.5% cholesterol and 100 soysterol. It was observed that both cycloartenol and 24-methylenecycloartanol in combination with soysterol greatly reduced the plasma cholesterol and enhanced cholesterol excretion. This suggests that the hypocholesterolemic activity of dietary vegetable oils may account for not only their fatty acid compositions and sterol contents but also the synergistic hypocholesterolemic effect of triterpene alcohols.

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A possibility whether an oral feeding of citrate, which has been reported to inhibit phosphofructokinase in vitro, following exercise to exhaustion could increase the rate of glycogen repletion in liver and soleus muscle was tested in treadmill running trained (experiments 1 and 2) and swimming trained rats (experiment 3). An exhaustive running or swimming was loaded at the end of the experiments, resulting in a significant reduction in liver and soleus muscle glycogen stores. The feeding of 1.0 and 0.5g of citrate per kg of body weight just after the exhaustive running could significantly increase the liver glycogen repletion during a 2-hr recovery period, but this was not observed in soleus muscle (experiment 1). As compared with a single feeding of 3.3g of glucose per kg of body weight, a mixed feeding of 0.5g of citrate and 3.0g of glucose after an exhaustive running (experiment 2) and swimming (experiment 3) could significantly enhance the repletion of glycogen stores in both liver and soleus muscle. These results clearly indicate that the postexercise feeding of citrate can stimulate the glycogen repletion in liver and skeletal muscle during an early period of recovery.

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After 2 days of oral administration of autoxidized linseed oil (AOLO; peroxide value 1, 000meq/kg, 0.3ml/rat/day), ultraweak chemiluminescence (CL) spontaneously emitted from rat liver homogenate increased up to about three times accompanying the accumulation of thiobarbituric acid (TBA) reactive products. This liver CL was quenched effectively by the in vitro addition of free radical scavengers, e.g. butyl hydroxytoluene, d-α-tocopherol and 2, 5-diphenylfuran, while superoxide dismutase, catalase and D-mannitol gave only negligible inhibitions on AOLO-induced liver CL. The luminescence spectrum recorded by a filter spectral analyzing method gave definitive two emission peaks at 585 and 635 nm and broad emission band around 500-560 nm. The spectrum was similar to that reported for a singlet molecular oxygen. In the experiment of successive AOLO administration (0.3ml/rat/day) for 10 days, a large increase in CL intensities and TBA reactants in various tissues was observed, especially on the 2nd day to the 6th day of feeding. Among them, the enhancement of CL and the accumulation of TBA reactants in liver and kidney were inhibited significantly by the oral administration of supplemental d-α-tocopherol (5mg/rat/day). These results suggest that AOLO-induced tissue spontaneous CL originated in the occurrence of free radical species which are possibly derived from chain reactions of tissue lipid peroxidation caused by the AOLO administration to rats.

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Effects of a toxic lectin from Kintoki beans (Phaseolus vulgaris, cultivar, Kintoki) on four types of animal cells were investigated. The cells used in this study were mouse L 929, human HeLa S3, and tendon and liver cells from chick embryo. The Kintoki bean lectin agglutinated these cells not only in suspension but also in monolayer, resulting in the marked growth inhibition of these cells. The incorporations of ³H-thymidine and ³H-leucine into trichloroacetic acid insoluble fraction of the cells were considerably inhibited by this lectin. There was, however, some lag period before the revelation of the inhibition. Kintoki bean lectin bound to these cells within 1 hr. The type of the lectin receptor seemed uniform for each cell type and the number of the binding sites per cell was different from cell type to cell type. When Kintoki bean lectin was removed from the culture medium, these cells slowly recovered back to normal growth.

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Feeding of starch or casein to rats that had been starved for 4 days resulted in marked increase in duodenal alkaline phosphatase activity in a few hours. In rats with a duodenal blind loop, the enzyme was induced by feeding starch, but not casein.
Conditions leading to hyperglycemia, such as an intragastric administration and an intravenous infusion of glucose, induced duodenal alkaline phosphatase to the same extent and in the same period as starch feeding. Subcutaneous injection of insulin alone did not induce the enzyme. In chemically sympathectomized rats, the enzyme was induced by starch feeding. In vagotomized or hexamethonium-treated rats, the enzyme was not induced by starch feeding or intragastric administration of glucose solution. On the contrary, subcutaneous injection of carbachol alone induced the enzyme. These results suggest that stimulation of the parasympathetic nervous system via hyperglycemia is important for induction of duodenal alkaline phosphatase by starch feeding. The mechanism of enzyme induction by casein feeding is still unknown.

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Studies were made on memory of the rhythmic change in activity Qf duodenal alkaline phosphatase in rats. During starvation, the peak enzyme activity decreased gradually disappearing in 4 days. The peak activity of duodenal alkaline phosphatase was retained by feeding starch diet for 4 days instead of starvation for 4 days, but not by feeding casein diet for the same period. Starvation for one day after feeding casein diet for 4 days resulted in disappearance of the peak activity. However, the peak activity was still retained after one day of starvation after 4 days on starch diet. Therefore, starch feeding appears to be important in the memory of the rhythmic change in activity of duodenal alkaline phosphatase.

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