The concentrations of mercury and selenium in thirteen tissues of the fetuses and sucklings of striped dolphin (Stenella coeruleoalba) were examined. A large portion of the mercury in the fetuses and sucklings was present in the methylated form. The total mercury level in the liver of the fetuses was slightly higher than in the other tissues. In halfofthe tissues, the total mercury levels were lower in the sucklings than in the fetuses. Although the selenium level in the fetuses was the highest in the liver, that in the sucklings was slightly higher in the blubber than in other tissues. In most tissues, the selenium level at the suckling stage was higher than at the fetal stage. A highly significant correlation was detected between the total mercury and selenium burdens in the fetuses. Linear relationships were observed between the body weight and total mercury burden and between the body weight and selenium burden of fetuses. The total burden of methylmercury in the term fetuses was around 1%of those in the pregnant females. The mercury contents of fetuses appear not to be affected by those of their mothers.

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Amongphotomixotrophic green calluses tested (N. rustica. N. tobacum L. cv. BY-4 and Samsun), the callus of Samsun had the highest contents of chlorophyll and chloroplast lipids, such as monogalactosyldiglyceride (MGDG), digalactosyldiglyceride (DGDG), sulfoquinovosyldiglyceride (SQDG) and phosphatidylglycerol (PG). However, the chlorophyll and chloroplast lipids in the green callus of Samsunwere still 1/6 and 1/3 of that in the parent leaves, respectively. The relative content of a-linolenate in MGDG, DGDG and SQDG of the green calluses were higher than that of the white calluses. The ratios of hexadecatrienoate in MGDG and hexadecenoate (A³-trans) in PGin the green calluses were trace or less compared with that of the parent leaves. The crude lipids and total fatty acid contents of the chlorophyll deficient leaves (N. tabacum L. cv. Consolation 402 and Dominant Aurea Su/su) were almost the same as those of the normal leaves (cv. BY-4 and Samsun), although the chlorophyll contents of the chlorophyll deficient leaves were 1/3 1/4 of that of the normal leaves. The ratios of chloroplast lipids in the total polar lipids in the chlorophyll deficient leaves were a little lower than that in the normal green leaves, but the former had a slightly higher ratio ofphospholipids such as phosphatidylcholine and phosphatidylethanolamine than the latter. There were few differences in the fatty acid compositions of each individual lipid betweeen both types of leaves.

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Alkaline oxidation of paralytic shellfish toxins with tert-butyl hydroperoxide yielded highly fluorescent derivatives. Even gonyautoxins I and IV and neosaxitoxin, which were nonfluorescing by previously proposed hydrogen peroxide oxidations, were successfully converted to fluorescent compounds. A continuous shellfish toxin analyzer was constructed by incorporating the reaction into a high pressure liquid chromatographic system. The limit of detection was within the range of 0.042.2 nmol for gonyautoxins I V, neosaxitoxin and saxitoxin.

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α-Hydroxyglutarate dehydrogenase (NAD⁺ specific) of Peptococcus aerogenes was purified by manganese chloride treatment, ammoniumsulfate fractionation and chromatographies with DEAE-cellulose, hydroxyapatite and' Sephadex G-200, and then crystallized in a colorless thin plate form by the addition of ammoniumsulfate. The enzyme has a molecular weight of approximately 53, 000 and consists of two subunits identical in molecular weight (about 32, 000). The isoelectric point of the enzyme is pH 3.7±0.1. The enzyme acts almost exclusively on α-ketoglutarate and ahydroxyglutarate. The Michaelis constants for α-ketoglutarate, NADH, α-hydroxyglutarate and NAD⁺ are 0.12, 0.028, 0.67 and 0.077mM, respectively.

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The factors involved in the selective accumulation of ansamitocins P-2, P-3 and P-4 by Actinosynnema pretiosum subsp. pretiosum No. C-15003 (Synonym : Nocardia sp. No. C-15003) were studied. The production of ansamitocin P-2, with a propionyl moiety at the C-3 position of the ansa chain, was stimulated more than 3-fold by the addition of isoleucine, propionate, propionaldehyde and n-propyl alcohol to the fermentation medium. The production of ansamitocin P- 3, with an isobutyryl moiety, was enhanced by the addition ofvaline, isobutyrate, isobutyraldehyde and isobutyl alcohol, and the proportion of P-3 reached more than 90% of the total ansamitocins produced. The production of P-4, with an isovaleryl moiety, was stimulated by leucine, isovalerate, isovaleraldehyde and isoamyl alcohol. The radioactive compounds, which selectively stimulated the production of each ansamitocin component, were preferentially incorporated into their respective acyl moieties of ansamitocins. Based on these results, we propose that the acyl moieties of ansamitocins P-2, P-3 and P-4 are derived not only through catabolic pathways of the respective amino acids, but also directly from alcohols, aldehydes and fatty acids having the same number of carbon atoms as those of the acyl moieties.

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The synthesis of jasmine ketolactone 3, a minor componentof Italian jasmine oil, via the intramolecular Michael reaction is described.

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Emulsions of methyl linoleate monohydroperoxides (18:2-monoHP) and methyl linolenate monohydroperoxides (18:3-monoHP)were incubated with ferrous sulfate and ascorbic acid. Gas chromatography mass spectrometric analysis of the trimethylsilyl and ter/-butyldimethylsilyl derivatives of the reaction products showedthat isomerization and secondary oxidation happen competitively during decomposition of 18:2-monoHP, while the secondary oxidation reaction proceeds preferentially and little isomerization is observed in 18:3-monoHP. It is suggested that 18:3-monoHP is more susceptible to secondary oxidation than 18:2-monoHP because of 18:3 specific secondary oxidation resulting in hydroperoxy-cyclic peroxides and dihydroperoxides. Moreover, an experiment using ¹⁸O₂ has demonstrated that molecular oxygen is scrambled by isomerization and secondary oxidation. It was confirmed that molecular oxygen is attached preferentially to the C-13 position in the 9-monoHPisomer and C-9 position in the 13-monoHP isomer during degradation of 18:2-monoHP.

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Protaminobacter ruber, having a firm cell wall, could be lyzed by treatment with glycine or penicillin. Among amino acids tested, only glycine showed an excellent effect for the lysis. The optimal amount and time of addition ofglycine for the lysis were 6mg per ml culture and 4 to 5hr after initiation of the cultivation, respectively. Aeration was necessary for the lysis as well as the growth of P. ruber. Morphological changes during the lysis of the bacterium were confirmed by electron microscopy. Proteins and δ-ALA dehydratase were excreted into the mediumwith the progress of lysis.

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Fromthe vacuumdistilled volatiles of shoyu, a neutral fraction was obtained. Shoyu was also directly extracted with dichloromethane and then the extract was separated into ten (AJ) fractions. The J fraction was a neutral one. The J fraction was further separated into twelve fractions by liquid column chromatography. All fractions obtained were analyzed by gas chromatography and combined gas chromatography-mass spectrometry. Consequently, 142 components were identified, 82 of which have not been reported previously as volatile constituents of shoyu. The identified compounds were 37 hydrocarbons, 22 alcohols, 22 carbonyls, 22 esters, 12 furans, 6 sulfurous compounds, 1 pyrone, 5 phenols, 1 furanone, 1 acid, 1 lactone and 12 other compounds.Fromthe results of quantitative analysis and organoleptic evaluation, phenylacetaldehyde is considered to be most important in the neutral fraction.

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The fine structures of amylopectin and intermediate material characteristic of amylomaize starch were investigated by chemical and enzymatic means. In comparison with waxy-maize amylopectin, that of amylomaize starch was found to possess a CLapproximately 10 glucose units longer. Unit-chain profiles of waxy and amylomaize amylopectins revealed that the clear difference lay simply in the relative amounts of two unit-chain fractions. By fractionations of debranched /Mimit dextrins, it was demonstrated that the CL of the internal chains in amylomaize amylopectin was 9 glucose units longer than that in waxy-maize amylopectin. In addition, the proportion of maltose and maltotriose fractions in the debranched dextrin for amylomaize amylopectin was noticeably smaller than found for waxy-maize amylopectin. These data suggest a lesser branching frequency of outer branches in amylomaize amylopectin, confirming the previous proposal that this amylopectin has longer inner and outer branches than those of normal amylopectin.
As for amylomaize intermediate material, the average degree of polymerization was estimated to be 250 to 300 glucose units per molecule. It was also indicated that there were 5 or 6 glucose residues corresponding to the non-reducing end in the molecule. The unit-chain profile of the intermediate material implied that this molecule was mainly composed of branches with CL around 50. Moreover, the presence of only small amounts of maltose and maltotriose fractions was demonstrated by the unit-chain distribution of this β-limit dextrin. These findings indicate that amylomaize intermediate material is totally consistent with a branched glucan having a low molecular weight, proposing that this anomalous glucan has such a fine structure that four or five branches with CLaround 50 are linked to a main linear chain of 100 to 150 glucose units.

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The absorbability of polyethylene glycol (PEG), a water-soluble nutritional marker, from the gastrointestinal tract of rat was examined using the [¹⁴C]-labeled compound ([¹⁴C]PEG) having a molecular weight of 4000. Intravenously injected [¹⁴C]PEG was readily excreted and recovered almost completely in the urine and neither hepatic nor renal uptake of the PEG was observed. Intragastrically administered [¹⁴C]PEG was eliminated in the urine with an average recovery of only 0.43±0.13% (Mean+S.D., n= 10) of the dose over 24hr. From the gel column chromatographic profile of the radioactivity excreted in the urine after an oral dose, [¹⁴C]PEG was suggested to be absorbed in two forms, as an original form and as a low molecular weight component. The latter component might be the degraded product of PEG in the gastrointestinal tract. From these results it was confirmed that PEG with a molecular weight of 4000 is a satisfactory marker because of its low absorbability.

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Four kinds ofdisaccharide peptides were isolated from a hydrolysate ofL. plantarum cell walls by combined catalysis by M-l N-acetylmuramidase and AM₃-endopeptidase, which were derived from the cultural broth of Stm. globisporus 1829. The chemical structures of these compounds were determined by enzymatic and chemical analyses to be β-N-acetylglucosaminyl-(1→)-N-acetyl-muramyl-L-alanyl-D-isoglutaminyl-(L)-meso-2, 6-diaminopimelic acid-(D)-amide (GMP₃-A), β-N-acetylglucosaminyl-( 1 ^4)-A^-acetylmuramyl-L-alanyl-D-isoglutaminyl-(L)-meso-2, 6-diaminopimelic acid-(D)-amide-(L)-D-alanine (GMP₄-A), β-N-acetylglucosaminyl-(1→4)-Ar, 6-O-diacetyl muramyl-L-alanyl-D-isoglutaminyl-(L)-meso-2, 6-diaminopimelic acid-(D)-amide (GMP₃-B) and ^-7V-acetylglucosaminyl-( l ->> 4)-7V, 6-O-diacetylmuramyl-L-alanyl-D-isoglutaminyl-(L)-meso-2, 6- diaminopimelic acid-(D)-amide-(L)-D-alanine (GMP₄-B). GMP₃-A and GMP₄-A were obtained in 20 and 10% yields from alkali-treated cell walls of L. plantarum, respectively. These compounds were examined as to their pyrogenicity, and mitogenic and immunoadjuvant activities. The presence of an 0-acetyl group at C-6 of the N-acetylmuramic acid moiety did not contribute to enhancement of these immunobiological activities. The pyrogenicity of GMP₄-A was much less than that of GMP₃-A. The C-terminal D-amino acid of disaccharide peptides appeared to play an important role in the pyrogenicity.

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A polarographic method based on the Brdicka current (the polarographic catalytic hydrogen evolution current produced by proteins in the presence of cobalt salts) was applied to direct titration of subtilisin BP1ST (S.BPINT) with plasminostreptin (PS) at a concentration level of 10^-8 m.The first and second dissociation constants of the S.BPN -PS complex were determined by fitting theoretical curves to the titration data, in which the multiple equilibria involving microscopically distinct forms of S.BPN -PS complex were taken into account. The intrinsic free energy change in the first binding of S.BPN' to dimeric PS was larger than that in the second binding. The dependence of the microscopic dissociation constants of S.BPN'-PS complex on pH indicates the participation of ionizable groups of pK_a 8.0 and 9.4 in the complex formation. The repulsive effect between negatively charged molecules of S.BPN' and PS in the complex formation at elevated pH is suggested.

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In order to reveal the mechanism of the Maillard reaction between proteins and reducing sugars, unmodified and chemically modified lysozymes were incubated with and without glucose at 50°C and 75% relative humidity in the solid state. Incubation of unmodified lysozyme with glucose resulted in browning and polymerization of the protein, and noticeable losses of arginine, lysine, and tryptophan residues. Those changes were little affected by the presence of an oxygen adsorber. Acetylation of lysozyme almost completely prevented those changes, indicating that the reaction of free amino groups of the protein with glucose is essential at the initial stage of these changes.
Incubation of lysozyme the arginine residues of which were maskedwith 1, 2-cyclohexanedione (CHD)resulted in almost the same changes as above even in the absence of glucose. Those changes could be explained as caused by the action of CHD released from the arginine residues. This similarity in the effects on protein of CHD and glucose implies that dicarbonyl compounds are key components at the secondary stage of the Maillard reaction between proteins and reducing sugars.

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Particulate preparations from Agrobacterium radiobacter IFO 12665b1 and Rhizobium phaseoli AHU 1133 have been shown to catalyze the synthesis of (1→2)-β-D-glucan from UDP-D- [¹⁴C]glucose. The (1→2)-β-D-glucans synthesized are suggested to be in a cyclic form without other glycosidic linkages and to consist of a mixture of several components with degrees of polymerization of 17 and more. The enzyme systems from A. radiobacter IFO 12665b1 and R. phaseoli AHU 1133 both required Mn²⁺ and had optimum activities at pH 7.5-8, and their Km values for UDPD-[¹⁴C]glucose were 5×10^-5M and 3.3×10^-5M, respectively.

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Krill aminopeptidase was purified about, 1, 100-fold from an extract of Euphausia superba with DEAE-cellulose column chromatography, Toyopearl HW55, and hydroxyapatite column chromatography. The final preparation was electrophoretically homogeneous. The molecular weight was determined to be 140, 000 by gel filtration and SDS-polyacrylamide disc gel electrophoresis. The optimum pH and optimum temperature were 8.4 and 45°C respectively. Krill aminopeptidase was inhibited by EDTA, Hg⁺⁺ and amastatin, and partially by bestatin, and was activated by Co⁺⁺. Alanyl-p-nitroanilide was hydrolyzed faster than leucyl-p-nitroanilide. Alanyl peptides (di-, tri-, tetra- and hexa-alanyl peptide) were hydrolyzed very fast.
These results suggest that krill aminopeptidase is an alanine aminopeptidase which is activated by cobaltous ion.

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The photobromination of l, 5-anhydro-2, 3-O-isopropylidene-β-D-ribofuranose gave the corresponding (5S)-5-bromo compound. The reduction of the bromide with triphenyltindeuteride gave (5S)-(5-²H₁)-1, 5-anhydro-2, 3-O-isopropylidene-β-D-ribofuranose, with a chiral purity of 76% at C-5, which was converted to (5R)- and (5S)-(5-²H₁)-D-riboses and other ribofuranose derivatives.

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Several analogs of stevioside were synthesized for an examination of their tastes. Hepta-O- acetylsteviolbioside was coupled with tri-0-acetyl-α-D-xylopyranosyl bromide, tri-O-acetyl-β-Larabinopyranosyl bromide, tetra-O-acetyl-α-D-mannopyranosyl bromide, tetra-O-acetyl-α-Lglucopyranosyl bromide, tri-O-acetyl-α-L-rhamnopyranosyl chloride and tri-(9-acetyl-α-Lquinovopyranosyl bromide, respectively, and the resulting coupling products were deacetylated to give 13-O-β-sophorosyl-steviol β-D-xylopyranosyl ester (2), 13-0-β-sophorosyl-steviol a-Larabinopyranosyl ester (3), 13-O-β-sophorosyl-steviol a-D-mannopyranosyl ester (4), 13-0-β- sophorosyl-steviol β-L-glucopyranosyl ester (5), 1 3-O-β-sophorosyl-steviol a-L-rhamnopyranosyl ester (6) and 13-O-β-sophorosyl-steviol β-L-quinovopyranosyl ester (7). Stevioside and 2 -7 were sweeter than sucrose by 255, 160, 285, 285, 210, 200, and 1 10 times on a weight basis, respectively.

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Monoclonal antibodies are now a powerful tool in biology and medicine. Transglutaminase has been implicated in diverse biological functions, and the characteristics of its catalytic action are suitable for applied enzymology. In this study, we produced hybridoma cells which synthesize monoclonal antibodies against guinea pig liver transglutaminase by fusing mouse myeloma cells with spleen cells of mouse immunized with the enzyme protein. Eight hybridoma clones (coded 2F, 4B, 7C, 8B, 8D, 8E, 9F and 1 1C) were selected to produce monoclonal antibodies. The subclass of IgG produced by clone 9F was IgG_2a and those from the seven other clones were all IgG₁ The 9F antibody inhibited transglutaminase activity, but the other antibodies did not. A solid-phase antibody-binding assay showed that of these antibodies, 8D antibody has the highest affinity to the antigen. Transglutaminase protein in crude liver extract was identified with Western blotting analysis using 8Dantibody as the probe.

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The reaction of cyclohexanone diethyl acetal with D-mannitol yielded quantitatively 1, 2 : 5, 6- di-O-cyclohexy idene-D-mannitol (1) and its isomer (2). From 1, 2, 3-O-cyclohexylidene-Dglyceraldehyde (3) was obtained in a quantitative yield without racemization.

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β-Xylosidase was purified 25 fold from a culture filtrate by ammoniumsulfate fractionation, DEAE-Sephadex chromatography, column electrophoresis, gel filtration on Biogel P-100, and isoelectric focusing. The purified β-xylosidase was found to be homogeneous on SDS (sodium dodecyl sulfate) polyacrylamide gel electrophoresis and on disc electrophoresis. A molecular weight of 101, 000 was estimated by chromatography on Sephadex G-200, and 102, 000 was obtained by SDS polyacrylamide gel electrophoresis. The purified β-xylosidase had an isoelectric point at pH 4.45, and contained 4.5% carbohydrate residue. The optimum activity for the enzyme was found to be at pH 4.5 and 55°C. The enzyme activity was inhibited by Hg²⁺, and JV-bromosuccinimide at a concentration of 1 x10^-3 m. The purified enzyme hydrolyzed phenyl β-D-xyloside (K_o= 13.0 sec^-1), p-nitrophenyl β-D-xyloside (K_O= 21.3 sec^-1), o-nitrophenyl β-D-xyloside (K_O=22.2 sec^-1), o- chlorophenyl β-D-xyloside (k_o=20.0 sec^-1), p-methylphenyl β-D-xyloside (k_o=9.0 sec^-1), omethylphenyl β-D-xyloside (k_o= 10.7 sec^-1), β-methoxyphenyl β-D-xyloside (k_o= l0.3 secβ^-1), omethoxyphenylβ β-D-xyloside (k_o= 10.9 sec^-1), xylobiose (k_o=36A sec^-1), xylotriose (k_o=34.5 sec^-1), xylotetraose (k_o= 17.4 sec^-1), and xylopentaose (k_o= 13.0 sec^-1). On enzymic hydrolysis of phenyl β-D-xyloside, the reaction product was found to be β-D-xylose with retention of configuration. The purified β-xylosidase was practically free of α-xylosidase and β-glucosidase activities.

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β-Xylosidase was purified 662 fold from a culture filtrate by ammoniumsulfate fractionation, gel filtration on Biogel P-100, DEAE-Sephadexchromatography, and gel filtration on Sephadex G- 200. With isoelectric focusing, the purified β-xylosidase found to be homogeneous on SDS (sodium dodecyl sulfate) polyacrylamide gel electrophoresis. The molecular weight was estimated by gel filtration to be 240, 000, and 1 16, 000 by SDS polyacrylamide gel electrophoresis. The purified fixylosidase had an isoelectric point at pH 3.25, and contained 4% carbohydrate residue. The optimum pH was found to be in the range of 4.5-5, and the optimum temperature was 55°C. The enzyme activity was inhibited by Hg²⁺, SDS, and 7V-bromosuccinimide at a concentration of 1×10^-3 m, andalso by/7-chloromercuribenzoate at a concentration of 1×10^-4 m. The purified enzyme hydrolyzed phenyl β-D-x loside (k_o = 302.6 sec^-1), p-nitrophenyl β-D-xyloside (k_o =438.9 sec^-1), onitrophenyl β-D-xyloside (K_o=431.0 sec^-1), p-chlorophenyl β-D-xyloside (K_O=207.9 sec^-1), ochlorophenyl β-D-xyloside (K_O=211.8 sec^-1), p-methylphenyl β-D-xyloside (K_O=96.5 sec^-1), omethylphenyl β-D-xyloside (K_O=83. 1 sec^-1), p-methoxyphenyl β-D-xyloside (K_O =99.3 sec^-1), omethoxyphenyl β-D-xyloside (K_O= 100.0 sec^-1), xylobiose (k_o=992A sec^-1), xylotriose (K_o= 1321.9 sec^-1), xylotetraose (K_O=789.7 sec^-1) and xylopentaose (K_O=508.0 sec^-1). On enzymic hydrolysis ofphenyl β-D-xyloside, the reaction product was found to be β-D-xylose with retention of the configuration. The purified β-xylosidase was practically free of α-xylosidase and β-glucosidase activities.

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An enzymatically modified gelatin (EMG-12) produced with covalent attachment of an lleucine n-dodecyl ester w s investigated as to its emulsifying properties under various conditions. The emulsifying activity of EMG-12was not so dependent on temperature as that of Tween-80 (T-80) used as a control. EMG-12and T-80 were investigated as to two parameters: their emulsion stability against coalescence and that against creaming. Both for EMG-12 and T-80, a lower temperature and a higher emulsifier concentration had a more favorable effect on these parameters. It was characteristic that the cream layer of the emulsion prepared with EMG-12held a large amount of entrapped water. Other surface properties of EMG-12, including its molecular area at the oil/water interface, are discussed.

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The stimulation of pancreatic enzymesecretion by diet components was investigated with a pancreatic duct cannula in rats. The infusion of casein into the duodenumresulted in a marked stimulation of the enzymesecretion. Onthe contrary, an aminoacid mixture simulated casein composition had no effect. Someoligopeptides, starch, sucrose, glucose, soybean oil and other commondiet componentsalso did not stimulate pancreatic enzymesecretion. A dose-response relationship was observed between the protein infusion level and the stimulation intensity of pancreatic enzyme secretion. Synthetic substrates for trypsin, tosyl-arginine methyl ester and benzoyl-arginine-p-nitroanilide, and for chymotrypsin, benzoyl-tyrosine ethyl ester, stimulated the enzyme secretion as well as casein; however, a typical substrate for aminopeptidase, leucine-p- nitroanilide, had no effect. These findings suggest that the substrates for pancreatic proteases can act primarily as secretagogues amongmanydiet components.
The ability as a secretagogue was more potent in casein than a-lactalbumin. Native ovalbumin was hardly digested by pancreatin in vitro. However, mild heat-denaturation of this protein considerably improved both its digestibility and ability as a secretagogue. It can be explained that mild heat-denaturation of the protein improves its availability as a substrate for pancreatic protease.
Based on these results, a mechanismby which substrate protein is recognized for pancreatic enzymesecretion in the rat intestine is discussed.

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Degradation of a β-O-4 lignin substructure model dimer by a white rot fungus, Phanerochaete chrysosporium, was investigated using a culture containing H₂¹⁸O, and the following conclusions were made, a) The direct hydrolysis at C_β of the β-O-4 dimer was not involved in formation of arylglycerol. b) About halfofthe oxygen at the benzyl (C_α position of the glycerol was derived from H2O (H₂¹⁸O) and the other half was from the oxygen at the benzyl (C_α) position of the substrate β-O-4 dimer. c) But, the oxygen at the C_α position of the substrate β-O-4 dimer did not migrate to the C_β position of the aryglycerol.

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