Arylglycerol-gamma-Formyl Ester as an Aromatic Ring Cleavage Product of Nonphenolic beta-O-4 Lignin Substructure Model Compounds Degraded by Coriolus versicolor

4-Ethoxy-3-methoxyphenylglycerol-gamma-formyl ester (compound IV) was identified as a degradation product of both 4-ethoxy-3-methoxyphenylglycerol-beta-syringaldehyde ether (compound I) and 4-ethoxy-3-methoxyphenylglycerol-beta-2,6-dimethoxyphenyl ether (compound II) by a ligninolytic culture of Coriolus versicolor. An isotopic experiment with a C-labeled compound (compound II') indicated that the formyl group of compound IV was derived from the beta-phenoxyl group of beta-O-4 dimer as an aromatic ring cleavage fragment. However, compound IV was not formed from 4-ethoxy-3-methoxyphenylglycerol-beta-guaiacyl ether (compound III). gamma-Formyl arylglycerol (compound IV) could be a precursor of 4-ethoxy-3-methoxyphenylglycerol (compound VI), because 3-(4-ethoxy-3-methoxyphenyl)-1-formyloxy propane (compound VII) was cleaved to give 3-(4-ethoxy-3-methoxyphenyl)-1-propanol (compound VIII) by C. versicolor. 4-Ethoxy-3-methoxyphenylglycerol-beta,gamma-cyclic carbonate (compound V), previously found as a degradation product of compound III by Phanerochaete chrysosporium (T. Umezawa, and T. Higuchi, FEBS Lett., 25:123-126, 1985), was also identified from the cultures with compound I, II, and III and degraded to give the arylglycerol (compound VI). An isotopic experiment with C-labeled compounds II' and III' indicated that the carbonate carbon of compound V was derived from the beta-phenoxyl groups of beta-O-4 substructure.     

New acylated triterpenoid saponins from Maesa lanceolata

Ten new acylated triterpenoid saponins were isolated from the leaves of Maesa lanceolata. For their structure elucidation extensive use was made of homo- and heteronuclear 2D NMR techniques such as COSY, NOESY, HSQC and HMBC. All saponins identified contained the same tetraglycosidic side chain, but the triterpenoid moiety showed a variable esterification pattern. Monoester, diester and triester derivatives were present. Maesasaponin I was a 21-monoester derivative, i.e. ?3 beta-O-[alpha-L-rhamnopyranosyl-(1-->2)-beta-D-galactopyranosyl- (1-->3)]-[beta-D-galactopyranosyl-(1-->2)]-beta-D-glucuronopyranosyl+ ++?-21 beta-angeloyloxy-13 beta, 28-oxidoolean-16 alpha, 22 alpha, 28 alpha-triol. Maesasaponins III, IV3, V3 and VI2 had an additional acetyl, propanoyl, n-butanoyl and angeloyl substituent, respectively, in position 22. Maesasaponins II, IV2, V2, VI3 and VII1 were characterised as the 16-acetyl derivatives of maesasaponins I, III, IV3, V3 and VI2, respectively. Structures of saponins previously reported in M. lanceolata had to be revised.     

The biosynthesis of highly branched N-glycans: studies on the sequential pathway and functional role of N-acetylglucosaminyltransferases I, II, III, IV, V and VI

At least 6 N-acetylglucosaminyltransferases (GlcNAc-T I, II, III, IV, V and VI) are involved in initiating the synthesis of the various branches found in complex asparagine-linked oligosaccharides (N-glycans), as indicated below: GlcNAc beta 1-6 GlcNAc-T V GlcNAc beta 1-4 GlcNAc-T VI GlcNAc beta 1-2Man alpha 1-6 GlcNAc-T II GlcNAc beta 1-4Man beta 1-4-R GlcNAc T III GlcNAc beta 1-4Man alpha 1-3 GlcNAc-T IV GlcNAc beta 1-2 GlcNAc-T I where R is GlcNAc beta 1-4(+/- Fuc alpha 1-6)GlcNAcAsn-X. HPLC was used to study the substrate specificities of these GlcNAc-T and the sequential pathways involved in the biosynthesis of highly branched N-glycans in hen oviduct (I. Brockhausen, J.P. Carver and H. Schachter (1988) Biochem. Cell Biol. 66, 1134-1151). The following sequential rules have been established: GlcNAc-T I must act before GlcNAc-T II, III and IV; GlcNAc-T II, IV and V cannot act after GlcNAc-T III, i.e., on bisected substrates; GlcNAc-T VI can act on both bisected and non-bisected substrates; both Glc-NAc-T I and II must act before GlcNAc-T V and VI; GlcNAc-T V cannot act after GlcNAc-T VI. GlcNAc-T V is the only enzyme among the 6 transferases cited above which can be essayed in the absence of Mn2+. In studies on the possible functional role of N-glycan branching, we have measured GlcNAc-T III in pre-neoplastic rat liver nodules (S. Narasimhan, H. Schachter and S. Rajalakshmi (1988) J. Biol. Chem. 263, 1273-1281). The nodules were initiated by administration of a single dose of carcinogen 1,2-dimethyl-hydrazine.2 HCl 18 h after partial hepatectomy and promoted by feeding a diet supplemented with 1% orotic acid for 32-40 weeks. The nodules had significant GlcNAc-T III activity (1.2-2.2 nmol/h/mg), whereas the surrounding liver, regenerating liver 24 h after partial hepatectomy and control liver from normal rats had negligible activity (0.02-0.03 nmol/h/mg). These results suggest that GlcNAc-T III is induced at the pre-neoplastic stage in liver carcinogenesis and are consistent with the reported presence of bisecting GlcNAc residues in N-glycans from rat and human hepatoma gamma-glutamyl transpeptidase and their absence in enzyme from normal liver of rats and humans (A. Kobata and K. Yamashita (1984) Pure Appl. Chem. 56, 821-832).     

Transformation of arylamines into direct-acting mutagens by reaction with nitrite

Arylamines including aniline (I), 1-naphthylamine (II), 2-naphthylamine (III), 2-aminofluorene (IV), 1-aminoanthracene (V) and 1-aminopyrene (VI) were treated with 4 equivalent amounts of nitrite at pH3 and 37 degrees C for 4 h. The reaction mixtures of I, IV, V and VI showed mutagenicity to Salmonella typhimurium TA98 and TA100 strains without metabolic activation. The numbers of His+ revertant colonies to TA98 strain were 110/0.05 mumole I, 970/0.055 mumole IV, 620/0.10 mumole V and 870/0.02 mumole VI. These arylamines were converted into mutagens with diazoquinone, diazonium and nitro functions depending on their structures. The mutagen from I was p-diazoquinone (I2). The mutagen from IV was highly unstable fluorene-2-diazonium salt (IV1). The mutagens from V were N3O3-introduced anthracene (V1-1) and 1-nitroanthracene (V2), and those from VI were unidentified nitro-introduced compound (VI1) and 1-nitropyrene (VI2).     

Arylglycerol-γ-Formyl Ester as an Aromatic Ring Cleavage Product of Nonphenolic β-O-4 Lignin Substructure Model Compounds Degraded by Coriolus versicolor

4-Ethoxy-3-methoxyphenylglycerol-γ-formyl ester (compound IV) was identified as a degradation product of both 4-ethoxy-3-methoxyphenylglycerol-β-syringaldehyde ether (compound I) and 4-ethoxy-3-methoxyphenylglycerol-β-2,6-dimethoxyphenyl ether (compound II) by a ligninolytic culture of Coriolus versicolor. An isotopic experiment with a ¹³C-labeled compound (compound II′) indicated that the formyl group of compound IV was derived from the β-phenoxyl group of β-O-4 dimer as an aromatic ring cleavage fragment. However, compound IV was not formed from 4-ethoxy-3-methoxyphenylglycerol-β-guaiacyl ether (compound III). γ-Formyl arylglycerol (compound IV) could be a precursor of 4-ethoxy-3-methoxyphenylglycerol (compound VI), because 3-(4-ethoxy-3-methoxyphenyl)-1-formyloxy propane (compound VII) was cleaved to give 3-(4-ethoxy-3-methoxyphenyl)-1-propanol (compound VIII) by C. versicolor. 4-Ethoxy-3-methoxyphenylglycerol-β,γ-cyclic carbonate (compound V), previously found as a degradation product of compound III by Phanerochaete chrysosporium (T. Umezawa, and T. Higuchi, FEBS Lett., 25:123-126, 1985), was also identified from the cultures with compound I, II, and III and degraded to give the arylglycerol (compound VI). An isotopic experiment with ¹³C-labeled compounds II′ and III′ indicated that the carbonate carbon of compound V was derived from the β-phenoxyl groups of β-O-4 substructure.     

Synthesis of modified fragments of fibrinogen and their effect on the activity of proteolytic enzymes

New analogues of the Gly-Pro-Arg and Arg-Gly-Asp fragments of fibrinogen were synthesized: Gly-Pro-Arg-Pro (I), Gly-Pro-Arg-Pro-Met-OMe (II), Gly-Pro-Arg-Pro-Phe (III), Gly-Pro-Arg-Pro-Asp (IV), Gly-Pro-Arg-Pro-Glu (V), and Arg-Asn-Trp-Asp (VI). Their effect on the activity of proteases of various types was studied with the method of lysis of fibrin plates. All the peptides were found to inhibit plasmin activity (by 60-85%) and the gamma-subunit of nerve growth factor (by 55-93%). Tetrapeptide (VI) proved to be an effective inhibitor of tissue activator of plasminogen and the gamma-subunit of nerve growth factor (by 96 and 93%, respectively). The peptides exerted practically no effect on the activity of urokinase and moderately inhibited the activity of streptokinase [(III), (IV), and (VI)], papain [(I), (II), (IV), and (VI)], subtilisin [(V) and (VI)], alpha-chymotrypsin [(III), (V), and VI)], and Bacillus subtilis metalloprotease (VI). They inhibit trypsin [except for (I) and (III)] when applied on fibrin plates at a concentration of 1 x 10(-2) M, while, at a concentration of 1 x 10(-3) M, (I) and (II) induced an increase in proteolytic activity by 35 and 47%, respectively.     

Oligosaccharide structures of isolated human colonic mucin species

Purified human colonic mucin contains six distinct components which may be separated by DEAE-cellulose chromatography. Past studies defined the structure of oligosaccharide side chains from the most abundant species III, IV, and V which elute at intermediate salt concentrations. In these studies the structures of oligosaccharide side chains liberated from the remaining early and late eluting species I, II, and VI were determined after isolation by sequential conventional and high performance liquid chromatography through combination of gas chromatography, methylation analysis, and sequential glycosidase digestion. Mucin species I, II, and VI contained a less varied array of discrete oligosaccharide structures than that observed in the major mucin components. Mucin species I and II contained five and 10 structures, respectively, which account for 68 and 71% of total oligosaccharide content in these fractions. The predominant oligosaccharides of mucin species I included three neutral structures: a disaccharide GlcNAc beta (1-3)GalNAc-ol, a trisaccharide Gal beta (1-4)GlcNAc beta (1-3)GalNAc-ol, and a tetrasaccharide GlcNAc beta (1-4)Gal beta (1-4)GlcNAc beta (1-3)GalNAc-ol as well as two acidic components representing the sialylated forms of two of these oligosaccharides. Mucin species II contained these same oligosaccharides as well as four additional acidic structures, notably a disaccharide Neu alpha (2-6)GalNAc-ol and a hexasaccharide Gal beta (1-4)GlcNAc beta (1-3)Gal beta (1-4)GlcNAc beta (1-3) (NeuAc alpha (2-6))-GalNAc-ol, not identified in any other mucin species. The late eluting mucin species VI contained at least five discrete neutral oligosaccharides and six major acidic structures. While the majority of these structures had been previously isolated from the earlier eluting mucin species IV and V, species VI also contained di- and trisialylated oligosaccharides not identified in other mucin species. In conjunction with earlier studies of the major mucin species III, IV, and V, these data define the range of oligosaccharide structures present in human colonic mucin. These studies demonstrate that human colonic mucin possesses species with characteristic and distinguishable combinations of oligosaccharides which reflect variations of common core structures.     

Microbiological degradation of bile acids. Metabolites formed from 3-(3a alpha-hexahydro-7a beta-methyl-1,5-dioxoindan-4 alpha-yl) propionic acid by Streptomyces rubescens.

1. The metabolism of 3-(3a alpha-hexahydro-7a beta-methyl-1,5-dioxoindan-4 alpha-yl)propionic acid (III), which is a possible precursor of 2,3,4,6,6a beta, 7,8,9,9a alpha,9b beta-decahydro-6a beta-methyl-1H-cyclopenta[f]quinoline-3,7-dione (II) formed from cholic acid (I) by streptomyces rubescens, was investigated by using the same organism. 2. This organism effected amide bond formation, reduction of the carbonyl groups, trans alpha beta-desaturation and R-oriented beta-hydroxylation of the propionic acid side chain and skeleton cleavage, and the following metabolites were isolated as these forms or their derivatives: compound (II), 1,2,3,4 a beta,-5,6,6a beta,7,8,9a alpha,9b beta-dodecahydro-6a beta -methylcyclopental[f][1]benzopyran-3,7-dione (IVa), (1R)-1,2,3,4a beta,5,6,6a beta,7,8,9.9a alpha,9b beta-dodecahydro-1-hydroxy-6a beta-methylcyclopenta[f][1]benzopyran-3,7-dione (IVb), (E)-3-(3aalpha-hexahydro-5 alpha-hydroxy-7a beta-methyl-l-oxo-indan-4 alpha-yl)prop-2-enoic acid (V), (+)-(5R)-5-methyl-4-oxo-octane-1,8-dioic acid (VI), 3-(4-hydroxy-5-methyl-2-oxo-2H-pyran-6-yl)propionic acid (VII) and 3-(3a alpha-hexahydro-1 beta-hydroxy-7a beta-methyl-5-oxoindan-4 alpha-yl)propionic acid (VIII). The metabolites (IVb), (V), (VI) and (VII) were new compounds, and their structures were established by chemical synthesis. 3. The question of whether these metabolites are true degradative intermediates is discussed, and a degradative pathway of compound (III) to the possible precursor of compound (VII), 7-carboxy-4-methyl-3,5-dioxoheptanoyl-CoA (IX), is tentatively proposed. The further degradation of compound (IX) to small fragments is also considered.     

Lignin degradation byPhanerochaete chrysosporium: Metabolism of a phenolic phenylcoumaran substructure model compound

The degradation of a lignin substructure model compound, 5-formyl-3-hydroxymethyl-2-(4-hydroxy-3,5-dimethoxyphenyl)-7-methoxycoumaran (I), in ligninolytic culture of a white-rot wood decay fungus,Phanerochaete chrysosporium, was investigated. It was found that I was hydroxylated or dehydrogenated in its coumaran ring to give 2-(5-formyl-2-hydroxy-3-methoxyphenyl)-3-hydroxypropiosyringone (II) and two coumarones, 5-formyl-3-hydroxymethyl-2-(4-hydroxy-3,5-dimethyoxyphenyl)-7-methoxycoumarone (V) and 3,5-diformyl-2-(4-hydroxy-3,5-dimethoxyphenyl)-7-methoxycoumarone (VI), II was further converted to 2,6-dimethoxy-p-benzoquinone (IV), syringic acid (III), and 5-carboxyvanillic acid (VIII). These metabolic products were identified by mass spectrometric comparison with the authentic compounds. A proposed pathway for the degradation of I is presented on the basis of these metabolic products. The degradation could be catalyzed mainly by phenol-oxidizing enzymes.Non-Standard Abbreviations TLC thin layer chromatography     

The in vivo metabolism of 7 beta, 17-dimethyltestosterone-6,7-3H.

7 beta, 17-Dimethyltestosterone (17 beta-hydroxy-7 beta, 17-dimethyl-4-androsten-3-one) (I) was given to three subjects in oral doses of 400 mg per day for ten days. The initial dose contained the steroid tritiated in the 6 and 7 positions. Plasma levels and urinary excretion patterns were followed in all three subjects. Isolations were done on the urine, plasma, and stools of one patient. From the urine 7 beta, 17-dimethyl- 5 alpha-androstane-3 beta,17 beta-diol (VI) was isolated from the nonhydrolyzed fractions. Unchanged (I), 7 beta,17-dimethyl-5 beta-androstane-3 alpha,17 beta-diol (III) and 7 beta, 17-dimethyl-5 beta-androstane-3 beta,17 beta-diol (IV) were isolated from the nonhydrolyzed and enzyme-hydrolyzed fractions. 7 beta,17-dimethyl-5 alpha-androstane-3 alpha,17 beta-diol (V) was isolated from the enzymatic fractions. From the stools were isolated unchanged (I), (III), (IV), (V), and (VI). Unchanged (I) and its 5 alpha-dihydro derivative (17 beta-hydroxy-7 beta,17-dimethyl-5 alpha-androstan-3-one) (II) were identified in the plasma. The total recovery of radioactivity in the one patient on whom the isolations were done was 57%; 40% from the urine and 17% from the stools.     

Oxidation of phenolic arylglycerol beta-aryl ether lignin model compounds by manganese peroxidase from Phanerochaete chrysosporium: oxidative cleavage of an alpha-carbonyl model compound.

Manganese peroxidase (MnP) oxidized 1-(3,5-dimethoxy-4-hydroxyphenyl)-2-(4-(hydroxymethyl)-2-methoxyphenoxy) -1,3-dihydroxypropane (I) in the presence of MnII and H2O2 to yield 1-(3,5-dimethoxy-4-hydroxyphenyl)- 2-(4-(hydroxymethyl)-2-methoxyphenoxy)-1-oxo-3-hydroxypropane (II), 2,6-dimethoxy-1,4-benzoquinone (III), 2,6-dimethoxy-1,4-dihydroxybenzene (IV), 2-(4-(hydroxymethyl)-2-methoxyphenoxy)-3-hydroxypropanal (V), syringaldehyde (VI), vanillyl alcohol (VII), and vanillin (VIII). MnP oxidized II to yield 2,6-dimethoxy-1,4-benzoquinone (III), 2,6-dimethoxy-1,4-dihydroxybenzene (IV), vanillyl alcohol (VII), vanillin (VIII), syringic acid (IX), and 2-(4-(hydroxymethyl)-2-methoxyphenoxy)-3-hydroxypropanoic acid (X). A chemically prepared MnIII-malonate complex catalyzed the same reactions. Oxidation of I and II in H2(18)O under argon resulted in incorporation of one atom of 18O into the quinone III and into the hydroquinone IV. Incorporation of one atom of oxygen from H2(18)O into syringic acid (IX) and the phenoxypropanoic acid X was also observed in the oxidation of II. These results are explained by mechanisms involving the initial one-electron oxidation of I or II by enzyme-generated MnIII to produce a phenoxy radical. This intermediate is further oxidized by MnIII to a cyclohexadienyl cation. Loss of a proton, followed by rearrangement of the quinone methide intermediate, yields the C alpha-oxo dimer II as the major product from substrate I. Alternatively, cyclohexadienyl cations are attacked by water. Subsequent alkyl-phenyl cleavage yields the hydroquinone IV and the phenoxypropanal V from I, and IV and the phenoxypropanoic acid X from II, respectively. The initial phenoxy radical also can undergo C alpha-C beta bond cleavage, yielding syringaldehyde (VI) and a C6-C2-ether radical from I and syringic acid (IX) and the same C6-C2-ether radical from II. The C6-C2-ether radical is scavenged by O2 or further oxidized by MnIII, subsequently leading to release of vanillyl alcohol (VII). VII and IV are oxidized to vanillin (VIII) and the quinone III, respectively.     

Effect of some new cAMP analogs on cAMP-dependent protein kinase isoenzymes.

1. Ten new cAMP analogs were synthesized by replacing the purine ring with with indazole, benzimidazole or benztriazole and/or their nitro and amino derivatives. 2. Each analog proved effective in activating cAMP-dependent protein kinase I (PK-I) purified from rabbit skeletal muscle and cAMP-dependent protein kinase II (PK-II) from bovine heart and chasing 8-[3H]cAMP bound to regulatory subunits in the half-maximal effective concentrations of 2 x 10(-8)-8 x 10(-6) M. 3. The N-1-beta-D-ribofuranosyl-indazole-3'5'-cyclophosphate(I) proved a very poor chaser and activator of both isoenzymes, but when indazole was attached at its N-2 to ribose (IV) or when its H at C-4 (equivalent to the position of amino-group in adenine) was substituted by an amino-(III) or especially nitro-group (II) its efficiency was dramatically increased. 4. Analogs containing benztriazole ring proved as powerful as cAMP irrespective of the presence of substituents (VII-X). 5. Benzimidazole derivatives with amino-(VI) or nitro-group (V) activated PK-II 3 and 20 times better than PK-I. 6. Attaching of ribose to N-2 of indazole or benztriazole increased the affinity to PK-II 10 and 4 times, respectively. 7. Chasing efficiency of cAMP analogs at half-saturating [3H]cAMP tended to correlate with activating potency only for PK-I but at saturating [3H]cAMP concentration for both isoenzymes. 8. On the basis of synergistic activation with 8-Br-cAMP a site 2-selective binding of nitro-benzimidazole (V) and unsubstituted benztriazole (VII) derivatives to PK-II is suggested.     

The postnatal histology of the epididymis in buffalo (Bubalus bubalis).

The epididymis of buffalo is differentiated into 6 regions. The head represents the first 3 regions, the body the 4th region and the tail the 5th and 6th region. At 3-30 weeks, the epithelium is simple high columnar with few basal cells in region I, simple low columnar with few basal cells in region II, pseudostratified low columnar in regions III, IV, V, and pseudostratified low columar in region VI. As age advances, the epithelium increases in height and shows a tendency toward advanced pseudostratification. The basal cells are greater in number in regions III, IV and V than in the other regions of the epididymis. The epithelial cells contain stereocilia in region I at 3 weeks and regions III, IV and V at 30 weeks, whereas they are absent in regions II and VI. The tubules in region I are the smallest in diameter while the tubules in region VI have the largest diameter.     

Mutagenicity of the anticancer drug, caracemide, and related compounds for salmonella

Caracemide, MeCON(CONHMe)(OCONHMe) (I), is a novel anticancer drug. Since it was derived from acetohydroxamic acid (II), a known mutagen, its potential metabolites and related compounds were synthesized and tested for mutagenicities in S. typhimurium TA98 and TA100. These compounds were: MeNHCONH(OCONHMe) (III), MeCONH(OCONHMe) (IV), MeCONOH(CONHMe) (V), MeNHCOONH2 X HCl (VI), MeNHCONHOH (VII), MeNHCOON(CONHMe)2 (VIII), and NOH(CONHMe)2 (IX). The mutagenicities in the absence of rat liver homogenate were: (VI) much greater than (IV) greater than (II), (III), (V). The other compounds were not mutagenic. (I) was mutagenic only in the presence of rat liver homogenate. The doses required to demonstrate mutagenicities of these compounds were from 0.05 to 5 mumoles/plate. The major hydrolytic products at 25 degrees C, pH 7, were (III), (IV), and (V) from (I); (II) and (III) from (IV); and (II), (III), (VII) and MeNHCONH(OCOMe) (X) from (V). (III) was stable at pH 7. Treatment of (IV) with HCl yielded (VI). Hydrolysis of (III) or (V) with ammonia yielded (VII). These results suggest that caracemide may be activated enzymatically or nonenzymatically by deacetylation or decarbamoylation, and its anticancer activity may be related to the reactivity of its metabolites with DNA. The synthetic procedures and characterizations of new compounds (IV), (V) and (X) are described.     

Specific and nonspecific glucanases from Trichoderma viride

Six endoglucanases (Endo I, II, III, IV, V, and VI), three exoglucanases (Exo I, II, and III), and a beta-glucosidase (beta-gluc I) isolated from a commercial cellulase preparation of Trichoderma viride origin were examined as to their activities on xylan ex oat spelts. Endo I, II, and III as well as Exo II and III showed no activity toward xylan and were classified as specific glucanases. Less specificity was found for the endoglucanases Endo IV, V, and VI, Exo I, and beta-gluc I, whose enzymes were able to hydrolyze xylan. With respect to product formation these xylanolytic cellulases fit the classification of xylanases generally accepted in the literature. Kinetic experiment with xylan, CM-cellulose, and p-nitrophenyl-beta-D-glucoside revealed that Endo IV, V, an VI and Exo I prefer to hydrolyze beta-1, 4-D-glucosidic linkages. beta-Gluc I showed no clear substrate preference.     

Further studies on proctolin analogues modified in position 2 of the peptide chain and their influence on heart-beat frequency of insects

Seven proctolin analogues (I-VII) modified in position 2 of the peptide chain by Phe (p-guanidino) (I), Phe (p-OEt) (II), Tyr (3'-NH2) (III), Tyr (3'-NO2) (IV), Afb (p-OH) (V) (Afb = 3-amino-4-phenyl-L-butyric acid), Afb (p-NH2) (VI), Afb (p-NO2) (VII), and the tetrapeptide Tyr (3'-NH2)-Leu-Pro-Thr (VIII) were synthesized by the classic liquid-phase method. The biological effects of the peptides were investigated in cardioexcitatory tests on two insect species, the cockroach Periplaneta americana L., and the yellow mealworm, Tenebrio molitor L. Within physiological concentrations (10(-9)-10(-7) M) peptides II, III, and IV stimulated the heart action of P. americana like proctolin itself. Under identical conditions, in the case of T. molitor, only peptide III showed cardiostimulatory properties, whereas other compounds (including II and IV) were inactive at concentrations up to 10(-7) M. Results reported here reflect, with reference to the analogues I-VII, selective recognition of receptors on myocardium of both insect species. The tetrapeptide VIII revealed a weak deacceleratory effect on P. americana and T. molitor heart action.     

Microbial conversion of pregna-4,9(11)-diene-17alpha,21-diol-3,20-dione acetates by Nocardioides simplex VKM Ac-2033D

The conversion of pregna-4,9(11)-diene-17alpha,21-diol-3,20-dione 21-acetate (I) and 17,21-diacetate (VI) by Nocardioides simplex VKM Ac-2033D was studied. The major metabolites formed from I were identified as pregna-1,4,9(11)-triene-17alpha,21-diol-3,20-dione 21-acetate (II) and pregna-1,4,9(11)-triene-17alpha,21-diol-3,20-dione (IV). Pregna-4,9(11)-diene-17alpha,21-diol-3,20-dione (III) and pregna-1,4,9(11)-triene-17alpha,20beta,21-triol-3-one (V) were formed in minorities. Biotransformation products formed from VI were pregna-1,4,9(11)-triene-17alpha,21-diol-3,20-dione 17,21-diacetate (VII), pregna-1,4,9(11)-triene-17alpha,21-diol-3,20-dione 21-acetate (II), pregna-1,4,9(11)-triene-17alpha,21-diol-3,20-dione (IV), pregna-1,4,9(11)-triene-17alpha,21-diol-3,20-dione 17-acetate (VIII), pregna-1,4,9(11)-triene-17alpha,20beta,21-triol-3-one (V). The conversion pathways were proposed including 1(2)-dehydrogenation, deacetylation, 20beta-reduction and non-enzymatic migration of acyl group from position 17 to 21. The conditions providing predominant accumulation of pregna-1,4,9(11)-triene-17alpha,21-diol-3,20-dione 21-acetate (II) from I and pregna-1,4,9(11)-triene-17alpha,21-diol-3,20-dione 17-acetate (VIII) from VI in a short-term biotransformation were determined.     

Anticholinesterase activity of potential therapeutic 5-(1,3,3-trimethylindolinyl) carbamates.

Six N-alkyl and N-aryl 5-(1,3,3-trimethylindolinyl) carbamates were synthesized and studied for their structure-activity relationships in inhibiting eel acetylcholinesterase (AChE). The carbamates were 5-(1,3,3-trimethylindolinyl)N,N-dimethylcarbamate (Cui Xing Ning) (I), 5-(1,3,3-trimethylindolinyl)N,N-diethylcarbamate (IV), 5-(1,3,3-trimethylindolinyl)N-ethylcarbamate (III), 5-(1,3,3-trimethylindolinyl)N,N-diethylcarbamate (IV), 5-(1,3,3-trimethylindolinyl)N-heptylcarbamate (V), and 5-(1,3,3-trimethylindolinyl)N-(3-chlorophenyl)carbamate (VI). The inhibition studies were carried out at 25.0 degrees C at pH 7.60. The rank order of the ki values for eel AChE inhibition is II > V > I > III > VI > IV. Compound II has a greater affinity for the enzyme than any irreversible inhibitor cited in the literature (Kd = 7.14 x 10(-8) M). Our findings should aid in the application of these carbamates (1) for counteracting the cholinergic problems associated with various diseases, and (2) for developing potential pretreatment compounds for organophosphate poisoning.     

Subunit arrangement in beef heart complex III

Beef heart mitochondrial complex III was separated into 12 polypeptide bands representing 11 different subunits by using the electrophoresis conditions described by Sch?gger et al. [(1986) Methods Enzymol. 126, 224-237]. Eight of the 12 polypeptide bands were identified from their NH2-terminal sequences as obtained by electroblotting directly from the NaDodSO4-polyacrylamide gel onto a solid support. The topology of the subunits in complex III was explored by three different approaches. (1) Protease digestion experiments of submitochrondrial particles in the presence and absence of detergent showed that subunits II and VI are on the M side of the inner membrane and subunits V and XI on the C side. (2) Labeling experiments with the membrane-intercalated probes [125I]TID and arylazidoPE indicated that cytochrome b is the predominant bilayer embedded subunit of complex III, while the non-heme iron protein appears to be peripherally located. (3) Cross-linking studies with carbodiimides and homobifunctional cleavable reagents demonstrated that near-neighbor pairs include subunits I+II, II+VI, III+VI, IV+V, V+X, and reagents demonstrated that near-neighbor pairs include subunits I+II, II+VI, III+VI, IV+V, V+X, and VI+VII. The cytochrome c binding site was found to include subunits IV, VIII, and X. The combined data are used to provide an updated model for the topology of beef heart complex III.     

A comparative cytogenetic study of the rice (<Emphasis Type="Italic">Oryza sativa</Emphasis> L.) autotetraploid restorers and hybrids

We studied pollen fertility, seed set and cytogenetic characteristics of restorer lines and F1 hybrids of autotetraploid rice. T4002, T4063, T461A × T4002 and T461A × T4063 showed significantly higher pollen fertility and seed set than T4132 and T461A × T4132. Meiotic pairing configurations of T4002, T4063, T4132, T461A × T4002, T461A × T4063 and T461A × T4132 were 0.05 I + 19.96 II (9.89 rod + 10.07 ring) + 0.01 III + 2.00 IV, 0.11 I + 19.17 II (8.90 rod + 10.37 ring) + 0.09 III + 2.26 IV + 0.01 VI, 1.34 I + 9.46 II (4.50 rod + 4.96 ring) + 0.80 III + 6.02 IV + 0.09 VI + 0.09 VIII, 0.02 I + 14.36 II (6.44 rod + 7.91 ring) + 0.01 III + 4.80 IV + 0.01 VIII, 0.06 I + 17.67 II (11.01 rod + 6.67 ring) + 0.06 III + 3.10 IV + 0.01 VI and 1.11 I + 11.31 II (5.80 rod + 5.51 ring) + 0.41 III + 5.63 IV + 0.03 VI + 0.03 VIII, respectively. Configuration 16 II + 4 IV and 12 II + 6 IV occurred in the highest frequency among the autotetraploid restorers and hybrids. Meiotic chromosome behaviors were less abnormal in the tetraploids with high seed set than those with low seed set. The hybrids had fewer frequencies of bivalents, univalents, trivalents and multivalents than the restorers, but higher frequency of quatrivalents than the restorers at MI. The frequency of univalents at MI had the most impact on pollen fertility and seed set, i.e., pollen fertility decreased with the increase of univalents. The secondary impact factors were trivalents and multivalents, and bivalents and quatrivalents had no effect on pollen fertility and seed set. The correlative relationship between pollen fertility and cytogenetic behaviors could be utilized to improve seed set in autotetraploidy breeding.