Studies of the synthesis of biomarkers. X. Synthesis of 13,17-secodiacholestanes.

13,17-Secodiacholestanes (6) were synthesized from cholesterol (1) in six steps. The key intermediates, (20R)- and (20S)-diacholest-13(17)-enes (3a and 3b), underwent ozonization and reduction to provide (20R)- and (20S)-13,17-secodiacholesta-13,17-dione (5a and 5b), respectively. On Clemmensen reduction, the diones (5a and 5b) yielded the target molecule 6. The structure of an unknown biomarker was shown to be different from the proposed 6 by gas chromatographic/mass spectrometric comparison.     

The glycolipids of Lactobacillus casei A.T.C.C. 7469

1. The lipids were extracted from Lactobacillus casei A.T.C.C. 7469 with chloroform-methanol mixtures. The glycolipids were obtained by chromatography on silicic acid and DEAE-cellulose (acetate form). 2. Hydrolysis of the glycolipids with alkali gave two glycerol glycosides and a mixture of fatty acids. 3. The glycosides were separated and their structures elucidated. The major component was O-alpha-d-galactopyranosyl-(1-->2)-O-alpha-d-glucopyranosyl-(1-->1)-glycerol and the minor component O-alpha-d-glucopyranosyl-(1-->6)-O-alpha-d-galactopyranosyl-(1-->2)-O-alpha-d-glucopyranosyl-(1-->1)-glycerol. 4. Analysis of the fatty acids by gas-liquid chromatography showed that they were predominantly palmitic acid, octadecenoic acid and lactobacillic acid.     

E.s.r. of free radicals in aqueous solutions of substituted pyrimidines.

Reactions of OH radicals with methyl and ethyl derivatives of uracil, cytosine and thymine in aqueous solutions have been investigated. Photolysis of H2O2 was used to generate OH radicals and the radicals on the base derivatives were spin-trapped using t-nitrosobutane and identified with the help of e.s.r. spectroscopy. Addition of OH radicals was found to take place predominantly to the C(5)--C(6) double bond of the bases. H-abstraction from the methyl group occurred in the N(1) methyl derivatives of uracil, cytosine and thymine. Radicals formed by H-abstraction from the methyl group were also detected for 3-methyluracil, thymine, 1-methylthymine and 1-ethylthymine. Introduction of a methyl or ethyl group at the N(1) position of uracil, cytosine and thymine causes an increase in the C(6) proton coupling and a decrease in the N(1) splitting for radicals formed by OH addition at the C(5) position.     

R.b.e. and o.e.r. of extended-Bragg-peak helium ions: survival and development of rat embryos.

Rats were exposed under aerobic or hypoxic conditions to 200-1200 rads of 60Co gamma-rays or extended-Bragg-peak helium ions on the eighth day of gestation. Uterine contents were examined on the twentieth day of gestation. At the 50 per cent embryonic survival level, helium ion r.b.e. was 1(.0) (aerobic) and 1(.2) (hypoxic). Maximum attainable gamma-ray and helium-ion o.e.r.s. were 2(.2) and 1(.7) respectively, indicating an oxygen-effect gain (o.e.g.) of 1(.2). At the 10 per cent survival level helium ion r.b.e. was 1(.1) (aerobic) and 1(.4) (hypoxic). Gamma-ray and helium-ion 0.e.r.s. were 2(.0) and 1(.5) respectively, indicating a helium ion o.e.g. of 1(.3). These data demonstrate that the small fraction of high-LET radiation present in this helium ion beam has a neglible effect on the aerobic r.b.e., but lowers the effective o.e.r. of the beam approximately 25 per cent relative to that of gamma-rays. Helium ions were significantly more effective than gamma-rays in killing embryos under hypoxic conditions, in producing congenital abnormalities under aerobic conditions, and in stunting foetal growth under both conditions.     

DNA of Epstein-Barr virus. VI. Mapping of the internal tandem reiteration.

Epstein-Barr virus (B95-8) DNA consists of short (10 X 10(6)) and long (87 X 10(6)) unique DNA sequences joined by 10 tandem reiterations of a 1.85 X 10(6) DNA segment. The reiterated sequence contains BamI and BglII sites separated by 4 X 10(5). The 4.5 X 10(5) and 14.0 X 10(5) segments generated by cleavage of the reiterated DNA with BamI and BglII contain sequences which hybridize to each other, suggesting that the internal tandemly reiterated sequence has a direct or inverted repeat within it. The opposite ends of the linear, nicked, double-stranded DNA molecule (R. F. Pritchett, S. D. Hayward, and E. D. Kieff, J. Virol. 15:556--569, 1975) consist of from 1 to 12 direct repeats of another 3 X 10(5) sequence (D. Given and E. Kieff, J. Virol. 28:524--542, 1978; D. Given, D. Yee, K. Griem, and E. Kieff, J. Virol. 30:852--862, 1979). There is no homology between the internal reiterated sequence and either terminus. However, part of the internal reiteration (less than 5 X 10(5) is reiterated at two separate locations in the long unique region. The internal reiterations are a source of variation within EBV (B95-8) DNA preparations. Thus, although the majority of molecules contain 10 tandem reiterations, some molecules have 9, 8, 7, 6, 5, 4, or fewer tandem reiterations. A consequence of this variability is that the KpnI A fragment and the EcoRI/Hsul A fragment consist of a family of seven or more fragments differing in the number of tandem internal reiterations. The EcoRI/HsuI A fragment of EBV (W91) DNA is approximately 6 X 10(6) smaller than the largest and dominant EcoRI/HsuI A fragment of EBV (B95-8) DNA. EBV (W91 DNA also differs from EBV (B95-8) DNA by an additional 7 X 10(6) to 8 X 10(6) of DNA in the long unique DNA region (D. Given and E. Kieff, J. Virol. 28:524--542, 1978; N. Raab-Traub, R. Pritchett, and E. Kieff, J. Virol. 27:388--398, 1978). These data suggest the possibility that the smaller number of internal reiterations in EBV (W91) DNA may be a consequence of the additional unique DNA and a restriction in the overall size of EBV DNA.     

Hydrogen bonding of flavoprotein. I. Effect of hydrogen bonding on electronic spectra of flavoprotein.

The effect of hydrogen bonding on the transition energy and the oscillator strength of the isoalloxazine nucleus of flavins was studied by the molecular orbital method. Among the possible hydrogen bondings examined, characteristic spectral shifts were found for the hydrogen bondings at N(1) and N(5) of the nucleus. The hydrogen bonding at N(1) resulted in the shift of the first absorption band towards blue and that of the second one towards red. On the other hand, the hydrogen bonding at N(5) resulted in the shifts of both the first and the second band towards red. The spectral characteristics reported on Clostridium MP and Desulfovibrio vulgaris flavodoxin coincided with the calculated results. The application of the calculated results to D-amino acid oxidase (D-amino acid: oxygen oxidoreductase (deaminating), EC 1.4.3.3) led to the conclusion that hydrogen bonding occurs at O(12), N(3)H, O(14) and N(5) of the isoalloxazine nucleus. The occurrence of hydrogen bondings at O(12), N(3)H, and O(14) is favorable for N(5) of the isoalloxazine nucleus to accept electron from an electron donor.     

Raman spectral studies of nucleic acids. XVII. Conformational structures of polyinosinic acid.

Laser-Raman spectra of poly(rI) show the formation of an ordered complex in aqueous solutions of high ionic strength. This structure exhibits the A-helix geometry, contains stacked bases and is apparently stabilized by specific hydrogen bonding involving hypoxanthine C6=0 groups. Thermal dissociation of the poly(rI) complex (Tm=45 degrees C) yields single-stranded and disordered poly (RI) chains. A disordered structure also occurs for poly (rI) in aqueous solutions of low ionic strength. In oriented films, poly (rI) forms an ordered structure probably the same as that which occurs in solutions of high ionic strength. Raman intensities measured at 815 and 1100 cm-1 in spectra of poly (rI) and poly (rU)-poly (rA)-poly(rU) indicate that the correlation previously established for single- and double-stranded ribopolymer structures is valid also for these multi-stranded structures. X-ray diffraction and model-building studies confirm the A-helix structure.     

The vacuum UV CD spectra of G.G.C triplexes.

Vacuum UV circular dichroism (CD) spectra were measured down to 175 nm for d(C)10, d(G)10, the d(G)10.d(C)10 duplex, and the d(G)10.d(G)10.d(C)10 triplex. A CD difference spectrum was calculated for d(G)10.d(C)10 giving the change in CD induced by forming the duplex from d(G)10 and d(C)10. The d(G)10.d(G)10.d(C)10 CD difference spectrum gave the CD induced by triplex formation from binding of d(G)10 to the d(G)10.d(C)10 duplex. In the near-UV, the d(G)10.d(C)10 and d(G)10.d(G)10.d(C)10 difference spectra resembled the difference spectrum for poly[r(G).r(C)] (Biopolymers 29, 325-333). This similarity may be an indication of similar purine base stacking. The d(G)10.d(G)10.d(C)10 vacuum UV difference spectrum had a negative band at 195 nm and a positive band at 180 nm, making it similar to difference spectra for homopolymer triplexes containing T.A.T and U.A.U triplets (Nucl. Acids Res. 19, 2275-2280). The appearance of these bands in difference spectra should be good indicators of triplex formation. The complementary oligonucleotides c-mycI d(CCCCACCCTCCC) and c-mycII d(GGGAGGGTGGGG) are part of the regulatory sequences of the human c-myc gene. G.G.C rich triplexes formed by binding c-mycII or c-mycIII d(GGGGTGGGTGGG) to the c-mycI.c-mycII duplex had CD difference spectra similar to that of d(G)10.d(G)10.d(C)10 in both the vacuum UV and near UV regions, indicating similar triplet structures.     

Molecular orbital studies on nucleoside analogs. II. Conformation of 6-azapyrimidine nucleosides.

PCILO (Perturbative Configuration Interaction using Localised Orbitals) computations have been carried out for three 6-azapyrimidine nucleosides, 6-azauridine, 6-azacytidine and 6-azathymidine, for both C(2')-endo and C(3')-endo pucker of the sugar ring. The results indicate a syn (chiCN=180 degrees) conformation followed by chiCN=90 degrees and gg conformation for C(3')-endo 6-aza analogs as compareed to the anti (chiCN=0 degrees) and gg conformation preferred by the corresponding pyrimidine nucleosides. For C(2')-endo sugar geometry, 6-azauridine and 6-azacytidine prefer, respectively, chiCN=0 degrees (anti) and phi C(4')-C(5')=60 degrees C (gg) and chiCN-240 degrees (syn) and phi C(4')-C(5')=120 degrees. The corresponding nucleosides, uridine and cytidine, show a preference for syn (chiCN=240 degrees) and gg and anti(chiCN=0 degrees) and gg , respectively. The X-ray crystallographic conformations of 6-azauridine and 6-azacytidine have been attributed to intermolecular hydrogen bonding and crystal packing forces. The results of PMR, CD and ORD studies on 6-azauridine and 6-azacytidine in aqueous solutions are in agreement with the PCILO predictions.     

Syntrophomonas wolfei subsp. methylbutyratica subsp. nov., and assignment of Syntrophomonas wolfei subsp. saponavida to Syntrophomonas saponavida sp. nov. comb. nov

A spore-forming bacterium strain 4J5(T) was isolated from rice field mud. When co-cultured with Methanobacterium formicicum DSM 1535(T), strain 4J5(T) could syntrophically degrade saturated fatty acids with 4-8 carbon atoms, including 2-methylbutyrate. Phylogenetic analysis based on 16S rRNA gene similarity showed that strain 4J5(T) was most closely related to Syntrophomonas wolfei subsp. wolfei DSM 2245(T) (98.9% sequence similarity); however, it differed from the latter in the substrates utilized and its genetic characteristics. Therefore, a new subspecies Syntrophomonas wolfei subsp. methylbutyratica is proposed. The type strain is 4J5(T) (=CGMCC 1.5051(T)=JCM 14075(T)). Furthermore, based on 16S rRNA sequence divergence and substrate utilization, we propose the assignment of Syntrophomonas wolfei subsp. saponavida DSM 4212(T) to Syntrophomonas saponavida sp. nov. comb. nov.     

Application of multilevel models to morphometric data. Part 2. Correlations.

Multilevel organization of morphometric data (cells are "nested" within patients) requires special methods for studying correlations between karyometric features. The most distinct feature of these methods is that separate correlation (covariance) matrices are produced for every level in the hierarchy. In karyometric research, the cell-level (i.e., within-tumor) correlations seem to be of major interest. Beside their biological importance, these correlation coefficients (CC) are compulsory when dimensionality reduction is required. Using MLwiN, a dedicated program for multilevel modeling, we show how to use multivariate multilevel models (MMM) to obtain and interpret CC in each of the levels. A comparison with two usual, "single-level" statistics shows that MMM represent the only way to obtain correct cell-level correlation coefficients. The summary statistics method (take average values across each patient) produces patient-level CC only, and the "pooling" method (merge all cells together and ignore patients as units of analysis) yields incorrect CC at all. We conclude that multilevel modeling is an indispensable tool for studying correlations between morphometric variables.     

OH radical formation by photolysis of aqueous porphyrin solutions. A spin trapping and e.s.r. study

Radical production during the photolysis of deaerated aqueous alkaline solutions (pH 11) of some water-soluble porphyrins was investigated. Metal-free and metallo complexes of tetrakis (4-N-methylpyridyl)porphyrin (TMPyP) and tetra (4-sulphonatophenyl)porphyrin (TPPS4) were studied. Evidence for the formation of OH radicals during photolysis at 615, 545, 435, 408 and 335 nm of Fe(III) TPPS4 is presented. Fe(III) TMPyP, Mn(III) TPPS4 and Mn(III) TMPyP also gave OH radicals but only during photolysis at 335 nm. The method of spin trapping with 5,5-dimethyl-1-pyrroline-1-oxide (DMPO) and 4-pyridyl-1-oxide-N-tert-butylnitrone (POBN) combined with e.s.r. was used for the detection of OH, H and hydrated electrons. With the spin trap DMPO, photolysis generated DMPO-OH adducts under certain conditions but no DMPO-H adducts could be observed. With POBN, no POBN-H adducts were found. The formation of OH was confirmed by studying competition reactions for OH between the spin traps and OH scavengers (formate, isopropanol) and the concomitant formation of the CO-2 adduct and the (CH3)2COH adduct with both DMPO and POBN. The photochemical generation of OH radicals was pH dependent; at pH 7.5 no OH radicals could be detected. Photolysis (615-335 nm) of dicyanocomplexes of the Fe(III) porphyrins did not produce OH radicals. When corresponding Cu(II), Ni(II), Zn(II) and metal-free porphyrins were photolysed at 615 and 335 nm, no OH radicals could be spin trapped. These results tend to associate the well-known phenomenon of photoreduction of Fe(III) and Mn(III) porphyrins with the formation of OH radicals. This process is described mainly as the photoreduction of the metal ion by the ligand-bound hydroxyl ion via an intramolecular process.     

Study on Chemical Constituents of Gentianopsis barbata var. Stennocalyx H. W. Li ex T.N. Ho

Nine compounds were isolated from Gentianopsis barbata var. stennocalyx H. W. Li ex. T.N.Ho. Their structures are identified as 1-hydroxy4, 7, 8-trimethoxyxanthone (Ⅰ), 1, 7-dihydroxy-3, 8-dimethoxyxanthone (Ⅱ), 1, 7, 8-trihydroxy-3-methoxyxanthone (Ⅲ), 1-O-(β-D- xylopyranosyl-(1→6)-β-D-glucopyranosyl)-3, 7, 8-trimethoxyxanthone (Ⅳ), 1-O-(β-D-xylopy- ranosyl- (1→6)-β-D-glucopyranosyl)-7-hydroxy-3, 8-dimethoxyxanthone (Ⅴ), 1-O-(β-D-xylo- pyranosyl-(1→6)-β-D-glucopyranosyl)-7, 8-dihydroxy-3-methoxyxanthone (Ⅵ), luteolin-7-O- β-D-glucoside (Ⅶ), oleanolic acid (Ⅷ) and ursolic acid (Ⅸ) by means of chemical methods and UV, IR and NMR determinations respectively.     

The teichuronic acid from the walls of Bacillus licheniformis A.T.C.C. 9945.

The teichuronic acid of Bacillus licheniformis A.T.C.C. 9945 grown under phosphate limitation was isolated from the cell walls and purified by ion-exchange and Sephadex chromatography. The detailed structure of the polysaccharide was established by methylation analysis, periodate oxidation and partial acid hydrolysis. The polymer is composed of tetrasaccharide repeating units with the structure [GlcA beta(1 leads to 4)GlcA beta(1 leads to 3)GalNAc beta(1 leads to 6)GalNAc alpha(1 leads to 4)n. 13C n.m.r. analysis has confirmed most of the structural features of the polysaccharide and, in particular, the anomeric configurations and linkage positions of substituents. The teichuronic acid from glucose-limited cells was identical with that from cells grown under phosphate limitation.     

Violence and psychosis. I. Risk of violence among psychotic men.

A survey of the records of 1241 men remanded in prison on criminal charges over four months yielded a high prevalence of psychiatric disorder. Of the total prison intake of 2743 men over the same period, 246 (9.0%) showed major symptoms of psychiatric illness and a further 237 (8.6%) symptoms of withdrawal from drugs or alcohol. Symptoms of neurotic disorders were underrecorded, so in terms of diagnosis 237 men (8.7%) were considered to be psychotic. Of these, 166 (70%) were schizophrenic. The influence of affective psychosis was small. The risk of violence among men with schizophrenia was high. Twenty five (9%) non-fatal personal assaults and 24 (21%) offences of damage to property were committed by men with schizophrenia. The presence of mental illness probably influences the decision to remand in custody for some of these offences, but this is unlikely to explain the substantially higher prevalence of schizophrenia among men convicted of homicide (five (11%) ) and arson (six (30%) ) than would be expected in the general population of Greater London (0.1-0.4%). The prevalence of schizophrenia among men convicted of homicide may even be an underestimate, as may the prevalence of affective psychosis and possibly of other psychiatric abnormalities, given the substantial incidence of concurrent suicide in such men.     

Patterns of DNA replication of human chromosomes. II. Replication map and replication model.

Combining higher resolution chromosome analysis and bromodeoxyuridine (BrdU) incorporation, our study demonstrates that: (1) Human chromosomes synthesize DNA in a segmental but highly coordinated fashion. Each chromosome replicates according to its innate pattern of chromosome structure (banding). (2) R-positive bands are demonstrated as the initiation sites of DNA synthesis in all human chromosomes, including late-replicating chromosomes such as the LX and Y. (3) Replication is clearly biphasic in the sense that late-replicating elements, such as G-bands, the Yh, C-bands, and the entire LX, initiate replication after it has been completed in the autosomal R-bands (euchromatin) with minimal or no overlap. The chronological priority of R-band replication followed by G-bands is also retained in the facultative heterochromatin or late-replicating X chromosome (LX). Therefore, the inclusion of G-bands as a truly late-replicating chromatin type or G(Q)-heterochromatin is suggested. (4) Lateral asymmetry (LA) in the Y chromosome can be detected after less than half-cycle in 5-bromodeoxyuridine (BrdUrd), and the presence of at least two regions of LA in this chromosome is confirmed. (5) Finally, the replicational map of human chromosomes is presented, and a model of replication chronology is suggested. Based on this model, a system of nomenclature is proposed to place individual mitoses (or chromosomes) within S-phase, according to their pattern of replication banding. Potential applications of this methodology in clinical and theoretical cytogenetics are suggested.     

The time of fruiting of Ulota Bruchii Hornsch. and U. crispa Brid.

Abstract

For obscure reasons, sporophytes of the dioicous Hamatocaulis vernicosus are currently unknown in France. With the aim of understanding this failure of sporophyte production we investigated (1) sexual phenology, (2) sex ratio pattern at different spatial scales, (3) limitation of sporophyte formation by the availability of only one sex, (4) limitation of sporophyte formation caused by the distance between male and female, (5) location of sporophytes and (6) the consevation implications of this failure of sporophyte production. Principal results show that phenology cannot explain the failure of sporophyte production. At the regional scale, sporophyte formation is prevented by the fact that several mountain ranges have single-sex populations. The production of spore capsules relies on a strict combination of factors at the local scale: occurrence of sexually expressed mixed-sex colonies, a short distance between male and female individuals and favourable environmental factors (light grazing, high water table). In the Massif Central of France spore production is unlikely because of current unfavourable conditions. Fertility patterns appeared useful in a conservation context.     

Properties of feline leukemia virus. III. Analysis of the RNA.

The kinetics of virus labeling was used to study the maturation of viral RNA in the Rickard strain of feline leukemia virus. Viral RNA labeled over differing intervals was characterized by gel electrophoresis and velocity sedimentation in sucrose gradients made up in aqueous buffer and 99% dimethyl sulfoxide. Labeled virus was found within 30 min after adding radioactive uridine to the cells and production of labeled virus reached a maximum at 4 to 5 h after pulse labeling. Native RNA from feline leukemia virus resolved into three size classes when analyzed by electrophoresis on 2.0% polyacrylamide-0.5% agarose gels: a 6.2 x 10(6) to 7.1 x 10(6) mol wt (50 to 60S) class, an 8.7 x 10(4) mol wt (approximately 8S) class, and a 2.5 x 10(4) mol wt (4 to 5S) class. From two experiments during which RNA degradation appeared minimal, these made up to 57 to 76%, 2 to 5%, and 6 to 12%, respectively, of the total RNA. The 8S RNA in feline leukemia virus has not previously been reported. The 50 to 60S RNA from virus harvested after 4 h of labeling electrophoretically migrated faster and sedimented more slowly in sucrose gradients than did the same RNA species harvested after 20 h of labeling. This argues for an intravirion modification of the high-molecular-weight RNA. The large subunits of denatured viral RNA from both 4- and 20-h labeled-viral RNA electrophoretically migrated with an estimated molecular weight of 3.2 x 10(6) but sedimented with 28S ribosomal RNA (1.8 X 10(6) mol wt) when analyzed by velocity sedimentation through 99% dimethyl sulfoxide.     

13C-n.m.r. study of C hordein.

Insoluble xylan was prepared from ground birch (Betula pubescens) pulp by alkali extraction and precipitation with ethanol. The only sugar detected after acid hydrolysis of the preparation was xylose. The insoluble xylan was used as substrate in a nephelometric assay to determine the xylanase (EC 3.2.1.8, 1,4-beta-D-xylan xylanohydrolase and EC 3.2.1.37, 1,4-beta-D-xylan xylohydrolase) activities of Aspergillus and Trichoderma enzymes. The nephelometric method is reliable in evaluating xylanase hydrolysis of insoluble xylan.     

Models of parent-offspring conflict. II. Promiscuity

The population genetics of Trivers (1974) concept of parent-offspring is examined for species in which the effects of the conflict are felt by future half-sibs, as in promiscuous mating systems in which the male shows no parental care. Whether or not a rare conflictor gene will spread in a non-conflictor population depends on f(m) greater than (m + 1)/(0.5m + 1.5) for a dominant gene, and on f(m) greater than 1/4(7 + 3) for a recessive gene; f(m) is the fitness gained by a conflictor relative to a non-conflictor offspring [f(m) greater than 1], and m is the amount of parental investment taken by a conflictor relative to m = 1 for a non-conflictor. The ESS value for conflict (mo) in promiscuous species with zero male parental care has mo = f(mo)/4[df(mo)/dmo]. However, where the male maintains the same harem for several breeding seasons, or where there is promiscuity but both sexes contribute equally to parental care, conditions for conflict are equivalent to monogamy.