The thermal conversion of 1 α-hydroxyprevitamin D₃ (1α-OH-previtamin D₃) diacetate to 1 α-hydroxyvitamin D₃ (1α-OH-vitamin D₃) diacetate was investigated in five solvents. The fraction of 1α-OH-vitamin D₃ diacetate was calculated from the HPLC peak areas (UV detection) of 1α-OH-previtamin D₃ diacetate and 1α-OH-vitamin D₃ diacetate. When 1α-OH-previtamin D₃ diacetate was dissolved in ethanol, benzene, toluene, isopropyl ether, or n-hexane, and heated at 60°C, the yield of 1α-OH-vitamin D₃ diacetate increased during the first 4h, and reached an equilibrium level after 8.5h. Differences in the ratio of 1α-OH-previtamin D₃ diacetate to 1α-OH-vitamin D₃ diacetate at thermal equilibrium, and in the rate of the thermal isomerization were observed among these five solvents. Molecular mechanics (MM) calculations were performed in order to estimate solvent effects on conformation for 1α-OH-previtamin D₃ diacetate and 1α-OH-vitamin D₃ diacetate. The sol-vent effect was treated by specifying a dielectric constant representative of each of the three solvents: ethanol (polar), n-hexane (nonpolar), and benzene (aromatic). The dielectric constants used were 24.3 for ethanol, 1.5 for n-hexane, and 2.3 for benzene. It is suggested that the confor-oration of 1α-OH-vitamin D₃ diacetate is stabilized in polar solvent. However, the order of conformational stability when solvent effects are included in the calculations is: ethanol> benzene>n-hexane. This order does not follow the experimental results. The proton NMR chemical shifts of 1α-OH-vitamin D₃ diacetate are different in deuterated n-hexane, ethanol, and benzene. The downfield shift of the C-6 vinyl proton of 1α-OH-vitamin D₃ diacetate, when compared to the chemical shift in benzene, is 0.15 and 0.11ppm relative to the chemical shift in n-hexane and ethanol, respectively, and that of the C-7 proton was 0.30 and 0.33 ppm, respectively. No significant proton shift of 1α-OH-previtamin D₃ diacetate is recorded in these three solvents. To account for the increased ratio of 1α-OH-vitamin D₃diacetate to 1α-OH-previtamin D₃ diacetate ratio in benzene, we suggest that 1α-OH-vitamin D3 diacetate may be stabilized via specific solute-solvent interactions in benzene.

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The cytokinetics and growth effects of retinnic acid (RA) (13-cis-RA, and all-trans-RA) were determined in human embryonic palatal mesenchymal (HEPM) cells using cell culture and sister chromatid differential staining. The lowest concentration tested that showed growth inhibition was 1.0×10^-4 M for 13-cis-RA, and 6.8×10^-5 M for all-trans-RA after 40 h of treatment. When HEPM cells were grown in the presence of BrdU (5-bromodeoxyuridine) for 44 h, approximately 70% of the control cells were in metaphase in the 3rd replication cycle (M3). The percentage of cells in M3 decreased in the presence of RA with or without a metabolic activation system (S-9), thus indicating a longer cell proliferation time for the RA-treated cells. The estimate of cell proliferation time in 13-cis-RA-treated cells (1.0×10^-4 M) was approximately 25 h at the 2nd cell cycle compared with 20 h in the control cells. When HEPM cells were exposed for 16 h to [³H] -thymidine, labeling indices in 13-cis-RA-treated cells were significantly reduced even at 1.0×10^-6 M. Both 13-cis-RA and all-trans-RA caused concentration-dependent decreases in [³H] -thymidine incorporation into HEPM cells. These results suggest that retinoic acids or their major metabolites interfere with DNA synthesis and decrease the proliferation of HEPM cells.

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1. Vitamin E content in the adipose tissue was examined in rats with and without vitamin E deficiency. With the progression of vitamin E depletion, the more rapid decrease in tocopherol concentration was observed in brown adipose tissue (BAT) than in white adipose tissue (WAT), and the rate of decrease of tocopherol was approximately three times faster in BAT than in WAT. After the intramuscular administra-tion of 10mg/kg of all-rac-tocopheryl acetate twice a week for two weeks to vitamin E-deficient rats, a similar pattern of increase was observed in the tocopherol concentrations of BAT and WAT, although the rate of increase was slower in WAT than in BAT. 2. Changes of tocopherol concentration in BAT and WAT were investigated in normo-nourished rats with hyperlipemia produced by the intramuscular injection of Triton WR-1339 for 7 days. A marked increase in tocopherol concentration was observed in both BAT and WAT in the late period of hyperlipemia, with the increase being greater in WAT. 3. The fatty acid composition of adipose tissue was compared between rats with and without vitamin E deficiency. No significant differences were observed in BAT and WAT between the two groups. 4. The glucose uptake of WAT was not altered in vitamin E-deficient rats when compared with control rats.

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S-Alkylcysteine α, β-lyase [EC 4.4.1.6] was purified to more than 90% homogeneity from the cell extract of Pseudomonas putida ICR 3640. The enzyme has a molecular weight of about 195, 000, and is composed of six subunits identical in molecular weight (37, 000). Pyridoxal 5'-phosphate is required as a cofactor. The enzyme catalyzes the α, β-elimination of S-methyl-L-cysteine and its analogs such as S-ethyl-L-cysteine, L-djenkolate, Se-methyl-DL-selenocysteine, and O-methyl-L-serine. However, S-methyl-D-cysteine, L-methionine, and L-norvaline were inert. The enzyme catalyzes also the β-replacement reaction of the thiomethyl group of S-methyl-L-cysteine with various thiols to yield the corresponding S-substituted cysteines. In addition to S-methyl-L-cysteine, Se-methyl-DL-selenocysteine and O-methyl-L-serine also serve as substrates in the β-replacement reaction.

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To elucidate the synthesis of cobalamin coenzymes in view of comparative biochemistry, tissue distribution of activity of aquacob-alamin reductase [EC 1.6.99.8] catalyzing the reduction of hydroxo-cobalamin to cob (II) alamin was studied in some vertebrates. Activity of NADH- or NADPH-linked aquacobalamin reductase (or both) was found in some tissues of monkey, rat, cow, hog, chicken, fish, and frog. The liver homogenates of these vertebrates contained both NADH- and NADPH-linked enzymes. Monkey and rat livers accounted for more than 80% of total activities of both NADH- and NADPH-linked en-zymes. Ratios of activities of the liver NADH- and NADPH-linked enzymes were above 1.0 in some land animals (monkey, rat, cow, and hog) but below 1.0 in water animals (sea and freshwater fishes). These different ratios are presumably due to the difference in the metabolisms of cobalamin and/or in the other cellular components between land and water animals.

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Thymidylate synthase and thymidine kinase, which cata-lyze the formation of thymidylate via the de novo and salvage pathways, respectively, are rate-determining enzymes in DNA synthesis. The in-creases in the activities of hepatic thymidylate synthase and thymidine kinase were significantly suppressed at 24 h after 70% partial hepatectomy in rats that had been administered cycloheximide. Concomitantly, other regenerative parameters such as the liver weight and contents of protein, RNA, and DNA were also significantly reduced in 24-h regenerating liver of cycloheximide-treated rats. When actinomycin D was injected, the activity of thymidine kinase, the liver weight, and contents of protein, RNA, and DNA were completely depressed in 24-h regenerating liver. However, the activity of thymidylate synthase in actinomycin D-administered rats rose to the level similar to the control (70% partially hepatectomized). The immunoblotting assay showed that thymidylate synthase is newly synthesized during liver regeneration after partial hepatectomy without being affected with actinomycin D.

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1. Tocopherol concentrations in plasma, red blood cells (RBCs), and buccal mucosal cells were examined in newborn infants (before feeding), children (2-15 years old), and adults. Tocopherol concentrations in adults and newborn infants showed the greatest dif-ference in plasma and the smallest difference in RBC. Buccal cell toco-pherol concentrations in adults were 2. 3 times higher than those in infants. The majority of newborn infants had RBC tocopherol levels below the normal limit (115μg/100ml of packed cells). Also, more than one third of Buccal cell tocopherol levels determined in newborn infants were below a level of 15ng/mg protein, which was determined as the lower limit of normal in healthy children on the basis of the testing of 97 samples. 2. After administration of a daily dose of 600mg of RRR-alpha-tocopherol for three months to young adults, tocopherol levels in buccal mucosal cells reached a level of more than 4 times the basal level after rising throughout the 3-month period, while levels in RBC and plasma showed less than a 3-fold increase and reached a maximum within one month. Buccal cell tocopherol levels showed a poor correlation to the RBC and plasma tocopherol levels, while the tocopherol/lipid ratio was closely correlated with RBC and plasma tocopherol levels. After a single dose of 600mg of RRR-alpha-tocopherol, RBC and plasma tocopherol concentrations reached a maximum within 24 hours, while buccal cell tocopherol did so 4 to 6 days later. 3. Very obese children and hyperlipemic obese children showed lower buccal cell tocopherol levels, accompanied by lower RBC tocopherol levels and higher plasma tocopherol levels, when compared with non-obese children and normolipemic obese children.

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Despite the fact that high-density lipoprotein cholesterol (HDLG) is a part of total cholesterol (TC), the serum level of this portion has been reported to have no or only a weak relationship to the TC level. The present study assessed the relationship between HDLC and TC considering alcohol consumption, cigarette smoking, and body mass index (BMI) in 366 male workers classified into three groups by the habitual physical exercise. The results showed the different effects of alcohol consumption, cigarette smoking, and BMI on the level of HDLC among these three groups, and alcohol consumption lowered the LDLC level only in the exercise group. The closest relationship between HDLC and TC was seen in the exercise group, even after taking other factors into account. The result suggests that the HDLG level must be evaluated relative to the level of TC. As an indicator of serum lipid patterns the validity of the ratio of HDLG to TC (HDLCJTC) was discussed.

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Casein was modified by glucose, diacetyl, or hexanal at 50°C, RH 75% for 1, 7, or 11 days. The chemical changes and digesti-bility in vitro of these nondialyzable caseins were investigated. The effects of these nondialyzable caseins supplemented with lost amino acids, on rats were studied by pair-feeding for 2 months. It was observed that internal organs such as liver, spleen, kidney, stomach, small intestine, cecum, colon and rectum were mostly unchanged. Biochemical values such as hematocrit, cholesterol, triglyceride, GPT, and GOT were also unchang-ed. However, the quantity of leucocytes was increased and serum glucose was decreased by feeding rats with modified caseins. Significant decrease in weight gain of rats fed with modified casein was observed, and the rate of decrease depended on the degree of modification of casein by carbonyl compounds. From these results, we supported the suggestion that some inhibitory or antinutritional compounds might be formed during the modification of casein by carbonyl compounds.

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Radiolabeled tracer [³H] very low density lipoprotein (VLDL)-triglyceride (TG) and non-tracer (Triton WR 1339) methods were used to determine VLDL-TG kinetics in normal rats and in rats given either a 10% glucose or a 10% fructose drinking solution for 16 h ad libitum. The carbohydrate-fed rats were hypertriglyceridemic com-pared to control animals. VLDL-TG was endogenously prelabeled with [³H] glycerol in control, glucose- and fructose-fed rats, and injected into identically treated recipients. Triton WR 1339, a potent inhibitor of VLDL-TG catabolism, was injected into the same animal 30 min after the end of the tracer method to measure TG secretion rate (TGSR). The tracer and non-tracer methods showed that fructose-fed rats had a sig-nificantly lower fractional catabolic rate (FCR) than either control or glucose-fed rats. In contrast, glucose-fed rats had a TGSR greater than control without a reduction in FCR. For all treatments, TG concentra-tion correlated with FCR for the tracer method and with TGSR for the non-tracer method. There was good correlation of TGSR determined by the tracer and non-tracer method for control rats despite the substantial difference in the absolute values. No such relationship was observed in carbohydrate-fed rats. Control rats were made hypertriglyceridemic by Intralipid infusion. A lower FCR was observed when determined by the tracer method, but this was not observed by the triton method. These results suggest that TG kinetics determined by the tracer and the triton methods cannot be readily correlated, especially in hypertriglyceridemic state. However, the two kinetic studies both suggested that overproduc-tion of VLDL-TG in glucose-fed rats and impaired VLDL-TG catabo-lism in fructose-fed rats were the primary cause for their hypertriglycer-idemic, respectively.

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The oxidation products of both types of mixtures of α- tocopherol (α-Toc) and Ttocopherol (γ-Toc) or α-Toc and δ-tocopherol (δ-Toc) during autoxidation of methyl linoleate were isolated and iden-tified. The structures of the oxidation products were characterized by UV, IR, ¹H and ¹³C NMR, and mass spectrometry. The 5-[2-(α-tocopherol-5'-yl)ethyl]-a-tocopherylquinone, 5-[2-(α-tocopherol-5'-yl)ethyl]-8a-hy-droxy-α-tocopherone, and O-[8-(5-ethoxymethyl-7-methyltocol)methyl]-α-tocopherol were obtained from both types of the mixtures as the oxida-tion products derived from α-Toc. γ-Toc diphenyl ether dimer (γ-TED) and γ-Toc biphenyl dimer (γ-TBD) were identified from the mixture of α-Toc and y Toc, and δ-Toc diphenyl ether dimer (δTED) from the mixture of α-Toc and δ-Toc. However, no oxidation product composed of both α-Toc and γ-Toc or δ-Toc was detected in oxidation products of both types of the mixtures. These results support the facts that, at first, oxidation of α-Toc proceeds during autoxidation of lipids and then γ-Toc or δ-Toc decomposes after approximate consumption of α-Toc.

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The influence of fish oil and safflower oil contained in the common Japanese diet as the main dietary polyunsaturated fatty acid source on plasma fatty acids in ten female student volunteers (21-22 years old) was investigated. The subjects were divided into two groups and fed the experimental diets for five days. The total daily fat intake in the fish diet and safflower oil diet was 54.4g and 56.2g, respectively, and the fat derived from fish and safflower oil was 16 g and 23 g, respectively. The proportion of linoleic acid was reduced in the plasma of subjects fed the fish diet and increased in the plasma of subjects fed the safflower oil diet. The plasma levels of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) were significantly elevated in the fish diet group. The ratio of EPA/arachidonic acid (AA) was higher, and those of n-6/n-3 and n-9/n-3 were lower in the plasma of subjects fed the fish diet when compared to the results obtained from plasma of subjects fed the safflower oil diet. From these results, it seems likely that fish oil in the common Japanese diet is a favorable source of plasma EPA and DHA even in such short term supplementation and with such a small amount of daily consumption.

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Preparing vegetables with heat the contents of their constituents will change to a various extend. Particularly the water-soluble and the heat-sensitive vitamins are affected. At an early stage the vitamin C losses were investigated, because of vitamin C's indicating function for oxidations and leaching-out processes (1, 2, 7, 11-13, 15, 17). The degree of vitamin losses is influenced by various factors, for example the type of food, variety of vegetables, the way of cutting, preparation, duration and method of cooking. The influence of the various cooking methods with regard to the losses of certain water-soluble vitamins will be discussed.

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To clarify the discrepancy between values of thiamin intake reported in national nutrition survey in Japan and judgment which was concluded by medical and biochemical examination in our field survey, thiamin of various daily foods were analyzed pre and post cooking in the various cooking methods, the following results were obtained. (1) The thiamin contents in cooked daily meals were 50-60 percent of the calculated values on an average. (2) The cooking losses of thiamin were particularly large in rice and green vegetables. (3) The loss of thiamin largest in boiling, followed by baking, parching and frying. (4) High temperature, pH, and chlorine on the public water acclerated thiamin losses. (5) The decrease of thiamin in cooked foods is caused by both of getting away of thiamin from foods and cleavage of thiamin of foods.

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To clarify the cooking losses of minerals (sodium, potassium, phosphorus, calcium, magnesium, iron, zinc, manganese, copper), various food materials were analyzed before and after cooking, and the following results were obtained. (1) The mineral contents of cooked foods in mass cooking were on an average about 60-70 percent of those in raw or uncooked foods. (2) Cooking losses were particularly high in minerals of vegetables. (3) Among various cooking methods, loss of mineral was largest in squeezing after boil and in soaking in water after thin slice, followed by parching, frying and stewing. (4) Cooking losses of minerals in meals cooked in home brought about the similar results as those by the mass cooking procedures. (5) The measures to prevent cooking loss are (a) eating the boiled food with the soup, (b) addition of small amount of salt (about 1% NaCl) in boiling, (c) avoidance of too much boiling, (d) selection of a cooking method causing less mineral loss (stewing, frying or parching).

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Three main parameters influence the nutritional quality of food preparation: the choice of raw materials, the recipe and composition of a meal, and the preparation process. In catering systems the temperature and time history during preparation and distribution, i.e. systems like cook-chill, cook-freeze, or warm-holding, need particular attention with regard to some sensitive nutrients, e.g. vitamins C, B1, folic acid. If the main parameters and influencing factors as described are taken into account, it should not be difficult to produce food in catering with high nutritional quality.

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The growing proportion of processed foods in the average daily diet in industrialized countries is considered as a challenge to the food industry, not only to provide more variety to the food assortment, but also to cope with high nutritional quality standards. Industrial handling of food therefore includes the monitoring of numerous operations and conditions from the agricultural source, through processing, packaging and distribution. Various types of food raw materials are processed by different kinds of technological treatments, the main objectives being to guarantee wholesomeness, taste, nourishment and convenience. New processing methods combined with the cold chain distribution reduce micronutrient losses to a certain extent. We shall examine some of the critical technological operations with regard to losses of vitamins and minerals in representative examples of foods like milk, leafy vegetables, potatoes, etc. and compare the classical food preservation with more modern food handling. With the increasing demand of pre-prepared fresh, refrigerated or frozen foods, the contribution of the maintained micronutrients is of interest, especially with reference to modern eating habits, i.e. lowering the total food intake and improving the nutrient density in the normal diet plan.

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In the first part the reactions and interactions of protein with macroconstituents of our food during processing are exposed from the chemical point of view. The reactions involving only protein (formation of isopeptides, of lysinoalanine, racemization) and the interactions with carbohydrates (Maillard reaction), oxidized lipids and polyphenols are briefly presented. Emphasis is put on the Maillard reaction since it is the most frequent reaction occurring during food processing and storage. The key compound rendering lysine unavailable in processed and stored foodstuffs is N^ε-fructoselysine (FL). Its oxidative degradation product, N^ε-carboxymethyllysine (CML) is found in variable but significant amounts in heat processed proteins. An interesting newer finding is that tryptophan can participate in a Maillard reaction with its indole-NH-group.
In the second part an overview is given on the impact these reactions have on the two components of protein nutritive value, namely digestibility and biological value. Again, most examples will be related to the Maillard reaction. Protein digestibility may be reduced by the modification of the protein molecule (blocking of active amino acid side-chains, establishment of crosslinks) or by the formation of compounds that inhibit digestive enzymes. (Inhibition of aminopeptidase by an advanced Maillard derivative of lysine). Biological value may be diminished by the loss of essential amino acids and/or their reduced specific availability. Ion-exchange chromatography of the protein hydrolyzate is the method of choice to determine amino acid losses. It also provides some clues for the type of processing damage by the presence of unusual amino acids in the chromatogramme (e.g. furosine, lysinoalanine). Global amino acid bioavailability is defined. It is of a complex nature and can only be truely determined in a bioassay in the animal. Specific availability of an amino acid is linked to particular structural features. Thus, specific lysine availability is determined by the presence of a free or “reactive” ε-amino group. This is the basis for the analytical methods for available lysine.
In the third part, the practical application of this knowledge to processed foods is shown using milk and vegetable protein as examples. Figures for the reduction in available lysine (blocked lysine) in different milk products processed according to conventional procedures are given and discussed. More subtile effects of milk processing on milk digestibility and stomach emptying are mentioned. The effects on protein nutritional value of extrusion-cooking of legume seeds and cereal flours are, then, presented. Whereas sulphur amino acid bioavailability and protein digestibility in legumes are improved by appropriate extrusion-cooking, extensive lysine loss and nutritional damage can take place when cereal flours are extruded under severe conditions.
It is concluded that, nowadays, extensive knowledge of the reaction mechanisms, appropriate analytical methods and flexible processing operations are available to prevent to a large extent protein nutritional losses.

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Transglutaminase-catalyzed incorporation of L-lysine into wheat gliadin rendered the lysine-fortified protein poorly digestible in the in vitro tests. In rat feeding tests, however, the luminal leavings and excreta collected after administration of the [₁₄C]lysine-fortified gliadin contained less than one-tenth of the radioactivity originally administered to rats. The enzymes, γ-glutamylamine cyclotransferase and 5-oxoprolinase, known to occur in animal kidney are at least in part responsible for the observed high availability of isopeptide bound lysine. A novel enzyme which is capable of directly hydrolyzing the cross-linked isopeptide into component amino acids and peptides, “N^ε-(γ-glutamyl)lysine hydrolase” was found in the isolated microorganisms which can use the synthesized N^ε-(γ-glutamyl)lysine as their only source of carbon and nitrogen. The enzyme(s) appear to be effectively used for improving digestibility and availability of protein matrixes formed in normal metabolism and by heat and/or shear treatment commonly used in food processing.

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