Procyanidins B-l and B-3 from azuki beans showed remarkable scavenging activities, especially to hydrophilic radicals. The inhibition rate constants for the scavenging of hydrophilic peroxyl radicals generated from 2, 2'-azobis(2-amidinopropane)hydrochloride in aqueous media were calculated at 37°C to be k_inh = 6.0×10⁴M^-1s^-1, 5.9×10⁴M^-1s^-1, 2.7×10⁴M^-1s^-1, 5.0×10⁴M^-1s^-1 and 1.1 × 10⁵M^-1s^-1, and the stoichiometric factors as n=8.4, 8.0, 3.7, 1.2 and 1.7 for procyanidin B-1, procyanidin B-3, (+)-catechin, ascorbic acid and α-tocopherol, respectively. Thus, the dimeric procyanidin molecule could trap eight radicals. The production and structures of procyanidin B-3 radicals are discussed on the basis of the ESR spectra observed by autoxidation of procyanidin B-3 in an aqueous alkaline methanol solution.

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Kinetic analysis was done on a chitinase (EC 3.2.1.14) purified from the liver of a prawn, Penaeus japonicus, using N-acetylchitooligosaccharides (GlcNAc_n, n=2 to 6), p-nitrophenyl N-acetylchitooligosaccharides (pNp-GlcNAc_n, n =1 to 5), and colloidal chitin as the substrates. The enzyme hydrolyzed GlcNAc₄ to two molecules of GlcNAc₂, GlcNAc₅ to GlcNAc₂ plus GlcNAc₃, and GlcNAc₆ by two ways to GlcNAc₂ plus GlcNAc₄ (87%), and two molecules of GlcNAc₃ (13%). Neither GlcNAc₂ nor GlcNAc₃ was hydrolyzed. The Km and k_cat were 0.249mM and 3.38sec^-1 for GlcNAc₄, 0.018mM and 2.67sec^-1 for GlcNAc₅, and 0.005mM and 2.72sec^-1 for GlcNAc₆, respectively. The cleavage patterns of p-nitrophenyl N-acetylchitooligosaccharides were different from those of the corresponding N-acetylchitooligosaccharides. The enzyme hydrolyzed colloidal chitin to produce mainly GlcNAc₂ and a trace of GlcNAc₃. Allosamidin inhibited prawn chitinase in a competitive way with a Ki of 0.1 μM and 1C₅₀ of 0.14μM. These results suggest that prawn chitinase is an endo-type chitinolytic enzyme, but different from insect chitinase or yam chitinase in the substrate specificity and cleavage pattern.

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The plant protein thaumatin has been expressed and secreted in Aspergillus oryzae. The thaumatin II cDNA was expresed via a Saccharomyces cerevisiae glyceraldehyde 3-phosphate dehydrogenase (GPD) promoter, and its secretion is apparently directed into the medium by the native plant signal sequence. The S. cerevisiae GPD promoter is properly regulated in A. oryzae; the addition of glucose stimulates the production of thaumatin. A 22-kDa immunoreactive protein indistinguishable from mature thaumatin is present in the culture medium. A slightly larger, 25-kDa protein is observed in cell extracts and may represent the unprocessed form of the protein. Maximum accumulation of the extracellular immunoreactive protein is observed after two days and no further production is observed.

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To produce Aspergillus oryzae alkaline protease (Alp) in an osmophilic yeast Zygosaccharomyces rouxii, we constructed an expression plasmid consisting of the Z. rouxii glyceraldehyde-3-phosphate dehydrogenase (GAPDH) promoter, the prepro-Alp cDNA of A. oryzae, the whole sequence of Z. rouxii plasmid pSR1, and the G418 resistant gene. The resulting plasmid, when introduced into Z. rouxii cells, directed the secretion of a large amount (about 300 mg/1) of Alp into the culture medium. The N-terminus and specific activity of the enzyme were identical to those of A. oryzae Alp.

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A method for isolation of egg yolk immunoglobulin, IgG, a livetin protein, was investigated. Several natural gums (carrageenan and xanthan gum) were found to be effective for removal of yolk lipoprotein as a precipitate. The effect was pronounced with λ-carrageenan and the lipid content in the supernatant after removal of the resulting precipitate was less than 0.4% of that of egg yolk. IgY remained in this supernatant, with a yield of 86%, and was isolated by chromatography on a column of DEAE-Sephacel followed by salting-out with sodium sulfate. IgY thus isolated was almost pure (98%) and the yield was 70 to 100 mg per egg.

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After retort pasteurization, the change in the aroma pattern of green, oolong and black tea drinks was investigated.
By pasteurization, such volatile compounds as leaf alcohol and its esters had deteriorated. Others such as linalool and geraniol were liberated from non-volatile precursors. It was revealed that such changes in the aroma compounds destroy the aroma balance of each tea drink, causing the off-flavor called retort smell.
From enzymatic experiments, it was clarified that the precursors of mono-terpene alcohols induced during pasteurization are those water-soluble glucosides dissolved in the green tea extract.

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Five kinds of proteins of Saccharomyces cerevisiae were recently purified to a homogeneous state and identified as yeast cell surface antigens (TLAa, TLAb, TLAc, TLAd, and TLAe), but their physiological functions remained uncertain. In this paper, TLAa was identified as a yeast enolase (EC 4.2.1.11) from the following evidence: (1) molecular weights and amino acid compositions of both proteins were similar, (2) the N-terminal 22 amino acid sequences of both proteins were the same (3), TLAa had enolase activity, and (4) the authentic yeast enolase gave a positive reaction with anti-TLAa serum in the Ouchterlony immunodiffusion test and the immunoblotting test. Two isoproteins of TLAa (enolase) were separated by non-denatured polyacrylamide gel electrophoresis and detected by immunoblotting with anti-TLAa serum. The contents of the two isoproteins varied depending on the following culture conditions: glucose starvation, growth in the presence of non-fermentable carbon sources, and growth in media containing sodium chloride and 2-deoxyglucose. The contents did not vary, however, with heat shock treatment or with growth in media containing sodium azide, tunicamycin, or sorbitol. These results showed that TLAa was a cytoplasmic antigen, and that its synthesis was regulated by some environmental stresses.

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Antifungal activity was detected in the crude saponin fraction obtained from the bottom cut of Asparagus officinalis L. This activity was specific to certain fungi, for example Candida, Cryptococcus, Trichophyton, Microspomm and Epidermophyton. Attempts were made to isolate the active principles from this fraction; in this way a new saponin (AS-1) was isolated, and its structure was estimated to be 3-O-[{β-D-glucopyranosyl(1 →2)}{β-D-xylopyranosyl(1→4)-β-D-glucopyranosyl]-(25S), 5β-spirostan-3β-ol. The minimum inhibitory concentration (MIC) ranged from 0.5μg/ml to more than 8μg/ml depending upon the nature of the fungi. On the basis of the work carried out here, it is probable that asparagus will contain additional antifungal principles.

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Various Japanese foods were treated with 22 mM nitrite at pH 3 for 1 hr at 37°C. Ethyl acetate extracts of the nitrosation mixtures were examined for their total NC content and mutagenicity to Salmonella typhimurium TA 100 to search for foods which formed a considerable amount of N-nitroso compounds (NCs) but showed no mutagenicity.
Most of the foods (32/35) formed NCs at levels in the range of 22-4758μmol N-NO/kg after the nitrite treatment. Among these foods, fermented products had a high level of NCs (mean = 952 μmol N-NO/kg), vegetables, cereals and fruits had a moderate level of NCs (mean = 225 μmol N-NO/kg), and meat and fish had a low level of NCs (mean = 99 μmol N-NO/kg). Only five kinds of food, i.e., soy sauce, salted Chinese cabbage, cabbage, Japanese radish and sausage, showed mutagenicity to Salmonella typhimurium TA 100. These results imply that considerably large amounts of various precursors of non-mutagenic NCs are present in foods, especially in some fermented products and citrus fruits.

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A Pseudomonas strain capable of using pyrazinamide as the sole source of nitrogen was isolated from soil. An aromatic amidase from the bacterium was purified 400-fold to homogeneous on polyacrylamide gel electrophoresis. The enzyme had a molecular weight of 43, 000 by gel filtration on Sephadex G-150 and consisted of two identical subunits. The isoelectric point was at 4.45. Among the compounds tested, pyrazinamide (relative activity, 100%), nicotinamide (60%), and 5-methylpyrazinamide (3.4%) were hydrolyzed at considerable rates. Benzamide, picolinamide, and isonicotinamide were not substrates. Apparent Km of the enzyme for pyrazinamide and nicotinamide were 5.6×10^-5M and below 5×10^-6M, respectively. The enzyme was not able to hydrolyze aliphatic amides. The enzyme was most active between pH 6.5 and 10 and 75°C, and was stable between pH 5.5 and 8.5 and below 45°C.

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A new type of immobilized halophilic nuclease H, Ca-enzyme complex, from a moderate halophile, Micrococcus varians subsp. halophilus was demonstrated. The 4-day cultivation of the bacterium in 3M NaCl CM medium at 25°C was favorable for our purpose, producing less 5'-nucleotidase, but flocculation of the cells grown at 25°C was poorer than that of the cells grown at 30°C. On the other hand, almost all of the nuclease H was precipitated to form Ca-enzyme complex by addition of 70 mM Ca²⁺ to the supernatant of the broth. The nuclease H activity of the Ca-enzyme complex was increased by preferential adsorption of exogenous nuclease H. A column bioreactor packed with this enhanced Ca-enzyme complex was used for 5'-mononucleotide production by continuous RNA degradation.

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Blood ethanol concentrations are decreased by the oral administration of ethanol with acetic acid. The slowing of gastric emptying is supposed to be one of the mechanisms for the blood ethanol-lowering effect. In the present study, to further clarify the contribution of gastric emptying to this effect, we investigated the blood ethanol concentration, and the amounts of ethanol remaining in the stomach and in the small intestine, after orally administering ethanol with acetic acid or various salts of acetic acid. This was done under two conditions in which the osmolarity or concentration of the acetate ion in each administered solution was equalized.
The blood ethanol-lowering effect of acetic acid was weakened by replacing acetic acid with sodium, potassium, or calcium acetate. There was a good negative correlation between the blood ethanol concentration and the amount of ethanol remaining in the stomach one hour after administering the sample solutions. The amount of ethanol remaining in the small intestine was not increased in the acetic acid group. From these results, the slowing of gastric emptying is considered to be the major mechanism for the blood ethanol-lowering effect of acetic acid, and is presumed to be controlled by cations of the solution, and not by the acetate ion.

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The structures of di-O-α-maltosyl β-cyclodextrins ((G₂)₂-β-CDs), which were produced from α-maltosylfluoride (α-G₂F) and cyclodextrin (CD) by the transfer action of debranching enzymes, were examined by the enzymic method using Bacillus subtilis saccharifying a-amylase (BSA). (G₂)₂-β-CD was converted to (G₁)₂-β-CD by treatment with glucoamylase before the examination. BSA completely hydrolyzed (G₁)₂-β-CD to produce glucose, 6³-O-α-glucosylmaltotriose, and 6³, 6⁵-di-O-α-glucosyl maltopentaose. (G₂)₂-β-CD was the mixture of 6^A, 6^C-di-O-α-maltosyl β-CD and 6^A, 6^D-di-O-α-maltosyl β-CD. The ratio of A, C/A, D in (G₂)₂-β-CD synthesized with Pseudomonas isoamylase and Aerobacter pullulanase were 40:60-45:55 and 30:70, respectively.
The content of 6^A, 6^C-di-O-α-maltosyl γ-CD in (G₂)₂-γ-CD synthesized by isoamylase was about 35%.

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The cuticular constituent of the bulb mite Rhizoglyphus robini is revealed to be hexyl 2-formyl-3-hydroxybenzoate (1) [hexyl rhizoglyphinate as the trivial name]. Structural proof of this novel ester relied on MS, GC-FTIR and NMR spectral data. The total synthesis of compound 1 was achieved in two steps, first by formylating methyl 3-hydroxybenzoate with hexamethylenetetramine in 75% polyphosphoric acid to give methyl 2-formyl-3-hydroxybenzoate, and then by a TsOH-catalyzed transesterification of the product with hexanol in benzene. Much higher antifungal activity than that of citral is demonstrated for compound 1, although its activity is lower than that of acaridials.

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A novel human hepatoblastoma cell line (HuH-6KK) with a high growth rate in a serum-free medium without extracellular matrix was developed from an original one, HuH-6 c!5 (HuH). The original HuH cells (38 passages) did not proliferate well in RPMI 1640 medium containing 20% fetal calf serum (FCS). The HuH cells (HuH-6KK) with a high growth rate were selected by culturing them in an enriched RDF containing 20% FCS and 0.01% mucous polysaccharide (spirulinan) isolated from a blue-green alga, Spirulina subsalsa. The HuH-6KK cells showed a rapid growth in serum-free eRDF medium containing insulin, transferrin, ethanolamine, and selenite (eRDF-ITES medium) without fibronectin. The proliferation of the original HuH cells was also observed in the eRDF-ITES medium, but the growth was slow compared with the HuH-6KK cells. In the medium without ITES, the growth of the HuH-6KK and original HuH cells was slow. Among the ITES ingredients, insulin promoted the growth of HuH-6KK cells the most.

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The hydrolytic activity of the purified yeast lipase OF 360 was greatly decreased by heating above 45°C and the residual activity after heating at 50°C for 15 min was about one-tenth of the original. Treatment of the lipase with a bifunctional reagent, dimethyl suberimidate (DMS), increased the heat stability. Stabilization by DMS treatment was supposed to be caused by non-covalent interaction between the lipase and DMS through electrostatic and/or hydrophobic forces. In addition, analysis of FT-IR spectra of the DMS- and non-treated lipases suggested that the main skeleton of the treated lipase was little perturbed by heating.

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Restriction-reduced mutants were isolated from Streptomyces rosa subsp. notoensis KA301 and S. tanashiensis strain Kala which produce the benzoisochromanequinone antibiotics nanaomycin and kalafungin, respectively. The mutants of S. rosa, which can be transformed with a multi-copy plasmid and in which the actinophage Pal6 can propagate, were selected. They were transformed with a single-copy plasmid propagated in S. lividans TK24, and with its modified plasmid propagated in the mutant at higher efficiency. The mutants of S. tanashiensis were selected by their capability to be transformed with a multi-copy plasmid. The efficiency of transformation with a single-copy plasmid propagated in S. lividans TK24 was low, but was much increased by heating the protoplasts at 42°C for ISmin prior to the transformation. These mutants derived from both strains probably lack at least one of their restriction systems.

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Spore germination promoter(s) for the cellular slime mold Dictyostelium discoideum was partially purified from the culture medium of Klebsiella aerogenes using Amberlite XAD-2 and molecular sieve gel column chromatography. The active components were found both outside and inside of a dialyzing bag and were separated into several fractions on a Sephadex G-75 gel column. The treatment of partially purified promoter(s) with NaIO₄ oxidation resulted in the complete loss of the activity. The promoter(s) was very unstable against heat treatment at pH 12. Several enzymes, such as proteinase K, RNase, DNase, lipase, and zymolyase could not inactivate the promoter(s). Interestingly, the spore germination depended on phosphate ion at 25-100 mM and proteose peptone or yeast extract in addition to the promoter(s).

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Changes in the contents of defensive substances against the active oxygen in water-stressed spinach plants were examined. The contents of ascorbate peroxidase (AP; EC 1.11.1.7), glutathione reductase (GR; EC 1.6.4.2) and α-tocopherol increased remarkably in water-stressed spinach leaves, while those of superoxide dismutase (SOD; EC 1.15.1.1), dehydroascorbate reductase (EC 1.8.5.1), ascorbate and glutathione changed little. The content of α-tocopherol in chloroplast thylakoid membranes isolated from water-stressed leaves was higher than that from normal leaves. It is, therefore, conceivable that GR, AP and α-tocopherol might be related to the tolerance of plants to water deficiency.

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A derivative of methoxypolyethylene glycol (mPEG₂) was covalently attached to ricin D, and some properties of the resulting conjugate (niPEG₂-ricin D) were studied. The conjugate was prepared by modification of ricin D with the TV-hydroxysuccinimide ester of mPEG₂ [Yamasaki et al., Agric. Biol. Chem., 52, 2125 (1989)], and purified by DEAE-cellulose chromatography, followed by gel filtration on a Sephadex G-75 column. Analytical data indicated that in the conjugate, 9 mol of mPEG₂ with an average molecular weight of 10, 000 were covalently attached to the ricin D molecule. The cytoagglutinating activity of mPEG₂-ricin D for Sarcoma 180 ascites tumor cells was 11% of that of native ricin D. While ricin D was insolubilized in frozen acidic solution at - 20°C accompanied by the loss of the cytoagglutinating activity irrespective of the coexistence of free mPEG₂, the conjugate was not insolubilized in frozen acidic solution and its cytoagglutinating activity was completely retained. An acetylated derivative of ricin D in which 7 amino groups/mol were modified with acetic anhydride was insolubilized in frozen solution similar to native ricin D. The insusceptibility of mPEG₂-ricin D to freeze-insolubilization may be due to the covalent attachment of the amphipathic copolymer, mPEG₂, to the amino groups of ricin D, but not due to the decrease in the positive charge by modification of amino groups.

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We purified calmodulin to apparent homogeneity from Tora-mame (Phaseolus vulgaris) seeds, one of the Japanese cultivars of the common bean. Electrophoretically, the purified Tora-mame calmodulin migrated faster than bovine brain calmodulin in the presence or absence of Ca²⁺. The estimated molecular weight of Tora-mame calmodulin was 14, 200 in the presenc of Ca²⁺ and 18, 200 in the absence of Ca²⁺. Like other plant calmodulin, Tora-mame calmodulin lacked tryptophan and contained 1 mol each of tyrosine and half cystine per mol of protein. However, Tora-mame calmodulin has fewer acidic amino acid residues than other plant calmodulins did. Rat brain phosphodiesterase (Ca²⁺-PDE) was stimulated by Tora-mame calmodulin, but the concentration of Tora-mame calmodulin, giving the half-maximal activation of Ca²⁺-PDE, was higher than that of bovine calmodulin, and the maximal activity of Ca²⁺-PDE obtained with Tora-mame calmodulin was lower than that obtained with bovine protein. The Ki value of W-7, a calmodulin antagonist, for Tora-mame calmodulin (17 μM) was larger than that for bovine calmodulin (8 μM). These observations suggest that hydrophobic regions of Tora-mame calmodulin exposed by Ca²⁺-induced conformational changes were slightly different from those of bovine calmodulin.

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Regarding the transport of absorbed sterols to endoplasmic reticulum in intestinal mucosal cells, transfer of cholesterol and sitosterol from rat intestinal brush border membranes to phospholipid liposomes was investigated. Transfer of sitosterol from brush border membranes to liposomes was significantly slower than that of cholesterol. Sitosterol transfer increased dose-dependently with the addition of sterol carrier protein₂ (SCP₂), but only a transient increase was observed with cholesterol. The results suggest that a slower rate of transfer of sitosterol than cholesterol from brush border membranes to the intracellular site of use may be one mechanism by which sitosterol transport into lymph is delayed, as observed previously in the rat intestine. SCP₂ may not essentially participate in the transfer of absorbed cholesterol.

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Transfructosylation of the β-fructofuranosidase I from Arthrobacter sp. K-l was investigated. This enzyme catalyzed both transfructosylation and hydrolytic action, when it was incubated with sucrose alone. But in the presence of a suitable acceptor such as D-xylose and lactose, the enzyme catalyzed mostly transfructosylation and transferred the fructose residue preferentially to the acceptor. The enzyme had broad acceptor specificities. D-Xylose, D-galactose, L-sorbose, D- and L-fucose, D- and L-arabinose, maltose, isomaltose, cellobiose, lactose, meiibiose, xylobiose, maltotriose, methyl β-glucoside, and galactoside were efficient acceptors in the transfructosylation. On the other hand, D-ribose, L-rhamnose, D-mannose, 2-deoxy-D-glucose, D-galactosamine, D-galacturonic acid, and 1-kestose were not efficient acceptors. Various primary alcohols, polyhydric alcohols including some sugar alcohols, and some glycosides acted as acceptors, but secondary alcohols with one hydroxyl group such as 2-propanol and 2-butanol were not effective as acceptors.

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It has been proved in our laboratory that rats fed ozonated casein that did not contain aromatic amino acid residues besides a very small amount of phenylalanine residues had no significant unfavorable change compared with those fed casein and that ozonated casein supplemented with amino acids lost by ozonolysis has nearly the same biological value as that of casein. But some unusual ninhydrin-positive substances existed in urine of rats fed ozonatd casein. Two of them, β-aspartylurea (major component) and 4-methyl-proline (minor), were identified. It was confirmed that β-aspartylurea was formed during ozonolysis of casein in aq. solution that contained urea to solubilize casein and, therefore, present originally in ozonated casein ingested by rats. The origin of 4-methylproline is still obscure.

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A lignan mixture from sesame salad oil containing episesamin and sesamin as major components was fed to rats. Lignans at the dietary level of approximately 0.2% tended to decrease plasma and liver cholesterol levels with an accompanying increase in the fecal excretion of neutral steroids, particularly when the dietary fat source was evening primrose oil containing γ-linolenic acid. There was a decreasing trend in the specific activity of Δ5-desaturase in liver microsomes whereas that of Δ6-desaturase tended to increase, in particular in rats fed with safflower oil. The proportion of dihomo-y-linolenate increased in response to the reduction of Δ5-desaturation activity, and that of docosapentaenoate (n-6) decreased in liver phosphatidylcholine in both groups of rats, suggesting that tignans interfered with various steps of linoleate metabolism. However, the production by the aorta of prostacyclin and by platelets of thromboxane A2 was not influenced by lignans. Thus, episesamin and/or sesamin functioned as a regulator of cholesterol and linoleate metabolism in rats.

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The β-l, 3-glucanase (l, 3-β-D-glucan glucanohydrolase, EC 3.2.1.6) gene from Flavobacterium dormitator var. glucanolyticae was cloned into Escherichia coli C600 with a vector plasmid, pBR322. The E. coli cells carrying a recombinant plasmid, pKUβGl (8.2kb), showed a high β-l, 3-glucanase activity and a lytic activity on viable yeast cells. These activities were found in the periplasmic space of E. coli clone cells. Southern hybridization analysis showed that the cloned gene was derived from F. dormitator chromosomal DNA. The gene products were purified from the periplasmic fraction of E. coli by ammonium sulfate fractionation and ion-exchange chromatography. The purified enzymes were demonstrated to be identical with a lytic endo-β, 3-glucanase II and a nonlytic endoβ-l, 3-glucanase I from F. dormitator from their enzymological and immunological properties. In the E. coli cells, endo-β-l, 3-glucanase I was also formed by a proteolytic digestion of endo-β-l, 3-glucanase II during the cultivation as in F. dormitator. Thus, the only endo-β, 3-glucanase II was coded for in the cloned gene.

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Acremoauxin A, a new fungal auxin derivative produced by A. roseum I 4267, was synthesized from indole, lactic acid, and D-mannitol. (+)-2-(3-Indolyl)propionic acid was prepared from its synthetic racemate by biological resolution using the acremoauxin-producing fungus. The synthetically confirmed structure of acremoauxin A was 1-O-[(2S)-(+)-2-(3-indolyl)propionyl]-D-arabitol (1).

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Recombinant human BSF-2 (B cell stimulatory factor 2/Interleukin 6; IL-6) proteins were purified from CHO and NIH3T3 cell cultures and characterized. The lectin binding patterns suggested that the proteins have N-linked oligosaccharide(s) with tri-antennary structure of bisecting GlcNAc. Their N-termini were highly heterogeneous; at least five closely related N-termini were detercted. This N-terminal heterogeneity was not generated in the cell culture because no processing activity was found in the culture medium.

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An efficient method for protoplast formation of Rhizopus niveus, a filamentous fungus with high capability to secrete enzymes out of the cells, was established using three lytic enzymes in combination, and a plasmid with a selection marker for transformation was constructed by transcriptional fusion of a Rhizopus promoter with the G418 resistance gene derived from Tn903. Then the protoplasts were mixed with the plasmid DNA in the presence of polyethylene glycol 4000 and CaCl₂. G418-resistant colonies were obtained, indicating that transformation was successful. Southern blot analysis of DNA from a transformant shows that the introduced DNA was present not only integrated into the host chromosome but also replicating extrachromosomalh, and that it was rearranged with host DNA during the course of cell cultivation. This is the first report of a transformation system for Rhizopus. This system will open the possibility for breeding R. niveus at the molecular level.

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KS-504a inhibited bovine brain calmodulin-dependent cyclic nucleotide phosphodieaterase (CaM-PDE) with an IC₅₀ value of 122μM. The inhibition was reversed by a high concentration of calmodulin. Calmodulin-independent activities of the enzyme were not affected by the compound at the same concentration ranges. Ca²⁺-dependent interaction of the compounds with calmodulin was shown using hydrophobic fluorescence probes. These data indicated that the compound exerted its effects on CaM-PDE by interacting with calmodulin. KS-504a also inhibited other calmodulindependent enzymes at different concentration ranges; myosin light chain kinase was inhibited at the lowest concentrations with an IC₅₀ value of 6.3μM. The inhibition mechanism was competetive with respect to calmodulin and non-competetive to ATP.

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γ-Tocopherol was reacted with an alkylperoxyl radical at 37°C in benzene, 2, 2'-azobis(2, 4-dimethylvaleronitrile) being used to generate the alkylperoxyl radical. The reaction products of γ-tocopherol were isolated by reverse-phase high performance liquid chromatography, and their structures were characterized. The products were tocored, four stereoisomers of 8a-(l-cyano-l, 3-dimethyl)butylperoxy-γ-tocopherone, a mixture of four stereoisomers of 8a-(l-cyano-l, 3-dimethyl)-butylperoxy-5-(γ-tocopheroxy)-γ-tocopherone, two atropisomers of 5-(γ-tocopherol-5-yl)-γ-tocopherol, and 5-(γ-tocopheroxy)-γ-tocopherol. From the reaction products of γ-tocopherol and its dimers, the reaction mechanism of γ-tocopherol with an alkylperoxyl radical is discussed.

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Dietary protein-dependent differences in the linoleic acid desaturation in rats were studied in terms of the differences in the amino acid composition. Soybean protein and its amino acid mixture decreased the linoleate desaturation index, the ratio of (20:3 n-6 + 20:4 n-6)/18:2 n-6, in the liver microsomal phospholipids compared to the casein counterparts, accompanying an increase in the ratio of cholesterol/phospholipid. The Δ6-desaturase activity of the liver microsomes was lower in the soybean protein group than in the casein group. The arginine supplementation to the casein diet tended to decrease but the lysine supplementation to the soybean protein diet tended to increase the activity, although the response pattern of the desaturation index was not necessarily paralleled in the latter situation. The effect of cystine supplementation to the casein diet was also inconclusive. Therefore, the arginine level of dietary protein, rather than the lysine/arginine ratio and the cystine content, seemed to be more important in the regulation of linoleic acid desaturation.

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