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  • Brassica napus
  • particle bombardment
  • somaclonal variation
Material
Years
Year
Keywords
  • 1
    Electronic Resource
    Electronic Resource
    Springer
    Sexual plant reproduction 2 (1989), S. 15-17 
    ISSN: 1432-2145
    Keywords: Brassica napus ; Spontaneous androgenesis ; Erucic acid content ; Flower colour
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Seeds of androgenetic origin were obtained among the F1 progenies of two crosses between resynthesized and cultivated forms of Brassica napus. The high-erucic, white-flowered, resynthesized line No7076 acted as the female, and the zero-erucic, yellow-flowered, cultivars ‘Topas’ and ‘Puma’, as males. No androgenetic seeds were obtained in the reciprocal crosses. Resynthesized rape could thus be of potential use for the production of androgenetic plants. Of special interest is the high frequency (21%) of spontaneous androgenesis observed in one of the two crosses. One plant, determined from erucic acid content and flower colour analysis as androgenetic, had a diploid chromosome number. Further knowledge about the genetic control of spontaneous androgenesis in the present material and the origin of the cytoplasm in androgenetic plants are required in order to exploit this phenomenon in practical plant breeding.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    Springer
    Plant cell reports 8 (1989), S. 303-306 
    ISSN: 1432-203X
    Keywords: Ti plasmid ; Virulence ; Brassica napus ; Brassica juncea
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Brassica napus and Brassica juncea were infected with a number of Agrobacterium tumefaciens strains. Tumourigenesis was very rapid and extremely efficient on B. juncea with all but one of the strains. Tumourigenesis on B. napus varied widely. It was very efficient with the nopaline strains, was reduced with the succinamopine strain A281 and was very weak with the octopine strains. The latter observation was confirmed with six different B. napus rapeseed cultivars. The selectivity was due to differences in the virulence of Ti plasmids with B. napus, rather than the tumourigenicity of the T-DNA or virulence of the chromosomal genes associated with the strains. An exception was strain LBA4404. The virulence of the octopine strains was increased by coinfection with more virulent disarmed strains and by induction with acetosyringone.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 77 (1989), S. 489-494 
    ISSN: 1432-2242
    Keywords: Microspore culture ; Spontaneous diploids ; Erucic acid ; Inheritance ; Brassica napus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The levels of erucic acid and other fatty acids in seeds of microspore-derived spontaneous diploid plants from crosses between low and high erucic acid parents were examined. The analysis confirmed that erucic acid is simply inherited and is determined by two genes that act in an additive manner. The effects of the genes for erucic acid on the levels of the other fatty acids was also determined and many significant correlations were found. In particular, erucic acid levels were negatively correlated with oleic acid and linoleic acid levels. The study also illustrates several advantages of using haploidy to analyze the inheritance of agronomically important traits. In particular, the number of phenotypic classes is smaller in androgenic populations and differences between classes are greater than in an F2 population.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 77 (1989), S. 651-656 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Brassica nigra ; Somatic hybrids ; Resistance to Phoma lingam
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Brassica napus and B. nigra were combined via protoplast fusion into the novel hybrid Brassica naponigra. The heterokaryons were identified by fluorescent markers and selected by flow sorting. Thirty hybrid plants were confirmed by isozyme analysis to contain both B. nigra and B. napus chromosomes; of these, 20 plants had the sum of the parental chromosome numbers. A non-random segregation of the chloroplasts was found in the hybrids. Of 14 hybrid plants investigated, all had the B. napus type of chloroplast. The resistance to Phoma lingam found in the B. nigra cultivar used in the fusion experiments was expressed in 26 of the hybrid plants. The hybrids obtained in this study contain all of the three Brassica genomes (A, B and C) and have thus created unique possibilities for genetic exchanges between the genomes. Since most of the plants were fertile as well as resistant to P. lingam, they have been incorporated into conventional rapeseed breeding programs.
    Type of Medium: Electronic Resource
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  • 5
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 77 (1989), S. 721-728 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Introgression ; Adaptation to short day ; Photo-insensitivity ; Oligogenic control
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Among the oleiferous Brassicas, B. napus has the highest seed and oil productivity. As it is a species adapted to the temperate regions, its spring type is either unable to flower or flowers too late in the short — day winter (rabi) season of the subtropics. B. napus (genome AACC) is an amphidiploid between B. campestris (AA) and B. oleracea (CC), and shares one genome with the other allotetraploids B. juncea (AABB) and B. carinata (BBCC). While B. napus lacks ecotypes adapted to the subtropics, the other four species are well represented in this climatic zone. Reciprocal crosses with or without one direct backcross to B. napus have been carried out with the intention of transfering short-day adaptability. The aim was to introgress the A genome of carefully selected early representatives of B. campestris and B. juncea with the corresponding genome in B. napus, and similary the C genome from B. oleracea and B. carinata with the analogous genome in B. napus. B. campestris, B. juncea and the clearly later species, B. oleracea var ‘alboglabra’ and B. carinata, seem to be almost equally effective in introgressing the appropriate earliness necessary for growth in Bangladesh. One backcross sligthly delayed segregation of early types. Convergent crosses did not result in the transgression of earliness, which was unexpected since the inheritance of flowering and maturity indicated a polygenic regulation. This result is partly explained by assuming dominant oligogenic control of the photoperiodic response. Introgression of earliness with the C genome doesn't seem to be necessarily related with the earliness of the donor species. Intergenomic interactions may be important. Interesting new lines were selected with high yield. Thus there is a good probability that Bangladesh will have a new oil crop. As these lines were observed to be early in Sweden as well, they could potentially push rapseed cultivation further north in temperate regions where the growing period is limited by short summers.
    Type of Medium: Electronic Resource
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  • 6
    ISSN: 1432-2242
    Keywords: Agrobacterium-transformation ; Asymmetric protoplast fusion ; Brassica napus ; Brassica nigra
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary With the idea to develop a selection system for asymmetric somatic hybrids between oilseed rape (Brassica napus) and black mustard (B. nigra), the marker gene hygromycin resistance was introduced in this last species by protoplast transformation with the disarmed Agrobacterium tumefaciens strain C58 pGV 3850 HPT. The B. nigra lines used for transformation had been previously selected for resistance to two important rape pathogens (Phoma lingam, Plasmodiophora brassicae). Asymmetric somatic hybrids were obtained through fusion of X-ray irradiated (mitotically inactivated) B. nigra protoplasts from transformed lines as donor with intact protoplasts of B. napus, using the hygromycin resistance as selection marker for fusion products. The somatic hybrids hitherto obtained expressed both hygromycin phosphotransferase and nopaline synthase genes. Previous experience with other plant species had demonstrated that besides the T-DNA, other genes of the donor genome can be co-transferred. In this way, the produced hybrids constitute a valuable material for studying the possibility to transfer agronomically relevant characters — in our case, diseases resistances — through asymmetric protoplast fusion.
    Type of Medium: Electronic Resource
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  • 7
    ISSN: 1432-2242
    Keywords: Brassica napus ; Protoplast fusion ; Triazine resistance ; Cytoplasmic male sterility ; Mitochondrial DNA
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Fusion of leaf protoplasts from an inbred line of Brassica oleracea ssp. botrytis (cauliflower, n=9) carrying the Ogura (R1) male sterile cytoplasm with hypocotyl protoplasts of B. campestris ssp. oleifera (cv “Candle”, n=10) carrying an atrazine-resistant (ATR) cytoplasm resulted in the production of synthetic B. napus (n=19). Thirty-four somatic hybrids were produced; they were characterized for morphology, phosphoglucose isomerase isoenzymes, ribosomal DNA hybridization patterns, chromosome numbers, and organelle composition. All somatic hybrids carried atrazine-resistant chloroplasts derived from B. campestris. The mitochondrial genomes in 19 hybrids were examined by restriction endonuclease and Southern blot analyses. Twelve of the 19 hybrids contained mitochondria showing novel DNA restriction patterns; of these 12 hybrids, 5 were male sterile and 7 were male fertile. The remaining hybrids contained mitochondrial DNA that was identical to that of the ATR parent and all were male fertile.
    Type of Medium: Electronic Resource
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  • 8
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 78 (1989), S. 473-479 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Microspore and anther culture comparison ; Androgenic lines ; Days to flower ; Days to maturity
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A direct comparison of microspore culture and anther culture was made in Brassica napus using F1 crosses of Regent (canola) by Golden (rapeseed), and their reciprocals, as well as a hybrid between Reston and a highly embryogenic, canola-quality breeding line (G231) as donor plants. The study confirmed that microspore culture can be ten times more efficient than anther culture for embryo production. Embryo yields from cultures initiated from the Reston x G231 were four-fold greater than those initiated from the Regent x Golden crosses, and significant differences were also detected among cultures initiated from the different Regent x Golden crosses. These results illustrate the influence that donor plant genotype has on embryo production. However, superior embryogenic potential among donor material was not always coincident with superior plant production. The average haploid-todiploid ratio in microspore-derived regenerates was 2∶1 for the population obtained from the Regent x Golden crosses but 1∶1 for the Reston x G231 cross. For both types of material, the frequency of diploids increased upon repeated cycles of explanting. A field study showed that there were no differences between the populations of anther-derived and microspore-derived spontaneous diploid and doubled haploid lines, with respect to the days required for them to flower or to mature. The information is valuable for canola breeding programs considering the use of haploidy.
    Type of Medium: Electronic Resource
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  • 9
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 78 (1989), S. 161-168 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Transgenic ; Heavy metal tolerance ; Human metallothionein gene ; Ti-plasmid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary A chimeric gene containing a cloned human metallothionein-II (MT-II) processed gene was introduced into Brassica napus and Nicotiana tabacum cells on a disarmed Ti-plasmid of Agrobacterium tumefaciens. Transformants expressed MT protein as a Mendelian trait and in a constitutive manner. Seeds from self-fertilized transgenic plants were germinated on media containing toxic levels of cadmium and scored for tolerance/ susceptibility to this heavy metal. The growth of root and shoot of transformed seedlings was unaffected by up to 100 μM CdCl2, whereas control seedlings showed severe inhibition of root and shoot growth and chlorosis of leaves. The results of these experiments indicate that agriculturally important plants such as B. napus can be genetically engineered for heavy metal tolerance/sequestration and eventually for partitioning of heavy metals in non-consumed plant tissues.
    Type of Medium: Electronic Resource
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  • 10
    Electronic Resource
    Electronic Resource
    Springer
    Theoretical and applied genetics 78 (1989), S. 793-797 
    ISSN: 1432-2242
    Keywords: Brassica napus ; Rapeseed ; Fatty acid composition ; Linolenic acid
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary Results from a diallel mating of two rapeseed lines with distinctly different linolenic acid concentration show that this trait is mainly under control of nuclear genes of the embryo. However, significant differences in reciprocal F1, BC1 and BC2 indicate maternal control, which is realized by interaction between maternal genotype and nuclear genes of the embryo. Additionally, temperature exerts considerable influence on the degree of maternal control. Since no reciprocal differences are detectable in F2, cytoplasmic factors seem not to be involved in the inheritance of linolenic acid concentration. Hypotheses on the physiological nature of maternal control of this trait are discussed.
    Type of Medium: Electronic Resource
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  • 11
    ISSN: 1615-6102
    Keywords: Brassica napus ; Vegetative cell ; Microbodies ; Pollen
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The ultrastructure of the vegetative cell ofBrassica napus tricellular pollen grains, just before anthesis with standard chemical fixation, is reported. The vegetative cell may be regarded as a highly differentiated and metabolically active fat-storage cell. It contains many mitochondria with a well developed internal membrane system, starchless plastids, microbodies, lipid bodies, dictyosomes and numerous vesicles thought to originate from the dictysomes. Rough endoplasmic reticulum organized in stacks of cisternae is also spatially associated with certain organelles, mainly lipid bodies, microbodies and plastids. There are also randomly distributed polyribosome areas. The microbodies are mainly polymorphic in shape and are often observed in contact with lipid bodies. The above spatial relationship implies that the microbodies may have a glyoxysomal function. In the late period of vegetative cell maturation, the microbodies are probably involved in the process of glyconeogenesis in which the conversion of lipid reserves to sugar takes place.
    Type of Medium: Electronic Resource
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  • 12
    ISSN: 1615-6102
    Keywords: Brassica napus ; Cell division ; Male germ unit ; Pollen ; Sperm cells
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary The association of the two sperm cells inBrassica napus pollen following the generative cell division was investigated. The generative cell during division is located in the center of the pollen grain, within the vegetative cell. The space present between the two cells is slightly irregular as seen following standard glutaraldehyde fixation. After completion of mitosis vesicles appear in the equatorial plane, coalescing centripetally to form a cell plate which fuses with the membrane of the generative cell, dividing it in two sperm cells. They are isolated from the vegetative cell by the space between the two cell membranes and are separated from each other by a similar space resulting from the cell plate formed during cytokinesis.
    Type of Medium: Electronic Resource
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  • 13
    ISSN: 1617-4623
    Keywords: Acetolactate synthase ; DNA sequence ; Brassica napus ; Oilseed rape ; Herbicide resistance
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary An acetolactate synthase gene was isolated and characterized from Brassica napus. This B. napus acetolactate synthase gene encodes a deduced polypeptide sequence of 637 amino acids which is 85% homologous to the corresponding proposed gene product from Arabidopsis thaliana. Peptide domains recently associated with herbicide resistance/sensitivity are conserved between the two sequences. From Southern analysis we conclude that the gene isolated is one member of a multigene acetolactate synthase gene family comprising four or five members. A probe spanning the presumptive transit peptide sequence of this gene was shown by Southern analysis to hybridize to a unique sequence in the B. napus genome. This unique probe was used to analyse DNA from B. campestris and B. oleracea, the presumed progenitors of B. napus. On the basis of restriction fragment length polymorphism, we conclude that the B. napus gene isolated here originated in B. campestris. Total acetolactate synthase-homologous transcripts were analysed in a variety of B. napus tissues, and showed preferential accumulation in rapidly growing material. The genomic clone was mutated in vitro at codon 173 to replace a proline residue with serine. This was re-introduced into plants, using Agrobacterium vectors, producing a herbicide-resistant phenotype which is characteristic of the predicted gene product.
    Type of Medium: Electronic Resource
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  • 14
    ISSN: 1573-5060
    Keywords: Diplotaxis erucoides ; Brassica napus ; rapeseed ; ovary culture ; intergeneric hybridization ; cytogenetics ; addition lines ; cytoplasmic male sterility
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Intergeneric hybrids (F1) Diplotaxis erucoides (DeDe) x Brassica napus (AACC) and the first backcross to B. napus (BC1) have been obtained through in vitro culture of excised ovaries. The chromosome numbers of F1 and BC1 plants proved the occurrence of unreduced gametes. The study of metaphase I chromosome pairing showed that autosyndesis in De genome and allosyndesis between De and A/C genomes might exist. The male fertility of the F1 plants was low. Some male-sterile plants were found in F1 and BC1 progeny. The possibilities of creating addition lines B. napus-D. erucoides and of obtaining a new cytoplasmic male sterility in B. napus are discussed.
    Type of Medium: Electronic Resource
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  • 15
    Electronic Resource
    Electronic Resource
    Springer
    Euphytica 42 (1989), S. 135-140 
    ISSN: 1573-5060
    Keywords: Brassica napus ; swede breeding ; inter-plot competition ; dry-matter yield
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Two experiments were done to evaluate the use of single-row plots for assessing dry-matter yield of swedes. In the first experiment the correlation between the dry-matter yields of 20 cultivars in unguarded single-row plots and those in guarded double-row plots was poor (r=0.39). It was improved (r=0.58) by adjusting the single-row plot fresh-weight yields for inter-plot competition, through an analysis of covariance in which the covariate was the mean of the adjacent plot yields, and then estimating the pure stand dry-matter yields. However, the correlation was still lower than that between unguarded double-row plots and the guarded double-row plots (r=0.73), and this latter correlation was also improved (r=0.82) by adjusting the fresh-weight yields of the unguarded plots for inter-plot competition. The second experiment provided an explanation for the partial success of the covariance adjustment. The individual neighbour effects of six cultivars were determined. Doon Major and Ruta Øtofte decreased the yields of adjacent cultivars whereas Dryden and Melfort increased them. Angela and Magres had little effect. The correlation between the dry-matter yield neighbour effects of cultivars and their fresh-weight yields was high (r=−0.81), but not complete. So although competitive ability was associated with root fresh-weight yield, other factors must also be involved. It was concluded that a two-replicate trial with unguarded double-row plots is preferable to a four-replicate trial with unguarded single-row plots for dry-matter yield assessment of early generation families in a pedigree swede breeding programme.
    Type of Medium: Electronic Resource
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  • 16
    Electronic Resource
    Electronic Resource
    Springer
    Euphytica 42 (1989), S. 155-162 
    ISSN: 1573-5060
    Keywords: Rubus laciniatus ; thornless blackberry ; brambles ; tissue culture ; periclinal chimera ; in vitro ; ex vitro ; somaclonal variation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary ‘Thornless Evergreen’ (TE) blackberry (Rubus laciniatus Willd.) is a periclinal chimera in which the epidermis has mutated to thornlessness (designated ‘t’), while the internal portions of the plant remain genetically thorny (tTT). Over 300 ex vitro plants (from shoot tip cultures of TE) were established in the field and observed over two fruiting seasons for various vegetative traits and fruiting characteristics. Both parental types and variants were included in the field plantings. Plants were evaluated for growth habit, vigor, flowering, and fruiting traits. Continuous variation for growth habit, vigor, flower number, and fertility was observed. Some of the variation could be explained in terms of chimeral segregation (pure thornless vs. chimeral types). Tissue culture-induced (somaclonal) variation was assumed to be the source of most of the other variation.
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  • 17
    Electronic Resource
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    Springer
    Euphytica 40 (1989), S. 75-81 
    ISSN: 1573-5060
    Keywords: Brassica juncea ; Indian mustard ; plant generation ; somaclones ; yield ; oil content ; dwarf type ; somaclonal variation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Multiple shoot formation in cotyledonary callus of Indian mustard (Brassica juncea cv. Prakash) was induced on modified MS media supplemented with high cytokinin (kinetin or zeatin) and low IAA concentrations. Complete plants were obtained on prolonged incubation of shoots on the same medium. 6-Benzyladenine alone or in combination with IAA or NAA did not support plantlet regeneration. A total of 71 plants were transferred to greenhouse. The seed, however, could be collected from 37 plants only. The seed was sown in the field to evaluate the material for somaclonal variation in R1 generation. Data were recorded for yield, plant height, number of primary branches, siliqua number, 1,000 seed weight and oil content. Somaclonal lines showed tremendous amount of variation for all the characters studied. A number of plants in this generation showed significantly higher yield and/or other improved agriculturally important characteristics as compared to the control. A line with dwarf plant type was also identified. A number of plants were selected from this generation and carried forward to R2 generation. Most of these lines bred true in R2 generation. The material seems to be very promising for future breeding programmes.
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  • 18
    ISSN: 1573-5060
    Keywords: Brassica napus ; forage rape ; swedes ; self-incompatibility ; S-genes ; hybrid breeding
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary Seed set after self-pollination confirmed that 19 lines of Brassica napus were self-incompatible. Eight lines, H, J, Q, W, X, K, P and Z, were fully cross-compatible. Line R was cross-compatible with these lines but often had a low seed set as female parent. These results are consistent with the activity of nine distinct S-alleles. Line S was cross-incompatible with K, as was V with P, and F with Z. With each of the lines A, E, B, O, G, L and M at least one reciprocal difference was found so that the number of additional distinct S-alleles could not be inferred, but there must be a minimum of seven. Pollen tube counts of intra- and inter-line pollinations using M, B and X confirmed the homozygosity of these lines with respect to self-incompatibility status and the presence of non-reciprocal compatibility between M and B. The results are interpreted in terms of the activity of both B. oleracea and B. campestris S-loci and the implications for hybrid breeding are discussed.
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  • 19
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    Euphytica 42 (1989), S. 99-103 
    ISSN: 1573-5060
    Keywords: Brassica napus ; self-incompatibility
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Summary The interactions of nine S-gene lines of Brassica napus were examined. Seven different S-genes were involved in the crosses, with three lines of different origin having the same S-gene. In general, the reactions on selfing the heterozygotes were as expected from the results of test crosses with the homozygous parent lines. All types of dominance relationships were found, including reversal of dominance in pollen and pistil. Weakening of activity of one or both genes was found in several cases. Background genotype was found have to some effect and reciprocal differences in reactions were also observed.
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  • 20
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    Plant cell, tissue and organ culture 17 (1989), S. 153-158 
    ISSN: 1573-5044
    Keywords: Brassica napus ; spring cultivars ; protoplast culture ; plant regeneration ; cultivar variation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplast cultures were prepared from hypocotyls of ten spring rapeseed cultivars. Protoplasts from all genotypes tested formed calli, and shoots were regenerated from calli of nine of the genotypes at frequencies varying from 15 to 76%. The regenerating cultivars fell into a high regenerating group (〉60% and a low regenerating group 〈25%).
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  • 21
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    Plant cell, tissue and organ culture 18 (1989), S. 129-141 
    ISSN: 1573-5044
    Keywords: Brassica napus ; organogenesis ; plantlet formation ; regeneration ; root segments ; shoot bud differentiation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Root segments obtained from aseptically germinated seedlings of Brassica napus cv. Westar were used to optimize conditions for high-frequency shoot bud differentiation. The presence of low kinetin (0.5 μM) and relatively high indole-butyric acid (1.0 μM) levels facilitated optimum shoot bud differentiation. Modified MS medium (MMS) was superior to the other three basal media tested (MS, B5 and White's). Elevated sodium dihydrogen phosphate levels increased the differentiation of shoot buds. Increasing or decreasing the level of sucrose from 3% reduced the frequency of explants forming shoot buds. Addition of glutamine enhanced both the frequency of responding explants, as well as the number of shoots per responding explant. Root segments from 13-day-old seedlings produced the highest response (58%) in the presence of 100 mg l-1 glutamine. The position of the segment on the main root, size, and the presence or absence of lateral roots altered the morphogenic response. Sealing of the donor seedling cultures with Parafilm® instead of Stretch' n seal® resulted in a higher production of shoot buds, although root segment cultures were not affected by the type of sealing. Spontaneous rooting occurred on all developed shoots.
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  • 22
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    Plant cell, tissue and organ culture 19 (1989), S. 159-165 
    ISSN: 1573-5044
    Keywords: cultivars ; roots ; shoots ; somaclonal variation
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Four tronchuda (Brassica oleracea var. tronchuda Bailey) cultivars were tested for their ability to regenerate in vitro on Murashige & Skoog (MS) medium supplemented with 3 different combinations of α-naphthaleneacetic acid (NAA) and 6-benzylaminopurine (BAP). Explants were either axillary bud-free whole cotyledons or hypocotyls from 7-day-old darkgrown seedlings. The ability to regenerate varied by cultivars, explants and the concentration of growth regulators. Hypocotyl explants of all 4 cultivars, and cotyledon explants of 2 cultivars, developed plantlets within 4 weeks. Hypocotyl explants produced more shoots than cotyledons. Cotyledon explants produced more roots than hypocotyls. Best shoot regeneration was on MS medium supplemented with 2 mgl-1 BAP and 0.1 mgl-1 NAA. ‘Portuguesa’ produced the most shoots. Some regenerants varied in leaf shape and phyllotaxy.
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  • 23
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    Methods in cell science 12 (1989), S. 139-144 
    ISSN: 1573-0603
    Keywords: microinjection ; genetic transformation ; protoplasts ; microspores ; Brassica napus
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Summary This paper describes a general method suitable for the microinjection ofBrassica napus protoplasts, unicellular microspores, and multicellular microspores. By incorporating components taken from other methods, manual operations frequently involved in the microinjection of plant cells have been simplified and microinjection rates increased. The embedding of cells in agarose provides a simple alternative to the variety of sophisticated immobilization strategies devised for different plant cell types thereby reducing the manipulations often involved in the culture of microinjected cells. Use of an automatic microinjector eliminated the operation of fine control systems on manual injectors; however, precision in sample delivery was reduced. Analyses indicate that transformed tissues can be recovered from microinjected protoplasts and microspores at high frequencies.
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  • 24
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    Plant and soil 113 (1989), S. 155-160 
    ISSN: 1573-5036
    Keywords: Brassica napus ; Ca uptake ; cation-anion balance ; rock phosphate ; split pot experiments
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Rape and sunflower were compared with respect to their rock phosphate mobilizing capacities, cationanion balance and uptake of Ca and NO3 at P-starvation. Rape was able to mobilize P from rock phosphate, whereas sunflower was not. When grown on a complete nutrient solution with NO3 as the only nitrogen source, both species took up more nutrient anions than cations. Withholding phosphate from the nutrient solution did not change the uptake pattern of rape, but sunflower took up more nutrient cations than anions at P-starvation, due to a strong decline in NO3 uptake. With both species, Ca uptake was not affected by phosphate in the nutrient solution. In split pot experiments, with rock phosphate supplied spatially separated from other nutrients, rape was still able to mobilize rock phosphate. A high Ca concentration had no effect on this capacity. The results indicated that in our experiments rock phosphate mobilization by rape was not associated with an excess of cation over anion uptake and neither with a high Ca uptake rate.
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  • 25
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    Plant and soil 113 (1989), S. 161-165 
    ISSN: 1573-5036
    Keywords: agar plate technique ; Brassica napus ; citric acid ; malic acid ; phosphate nutrition ; rhizosphere acidification
    Source: Springer Online Journal Archives 1860-2000
    Topics: Agriculture, Forestry, Horticulture, Fishery, Domestic Science, Nutrition
    Notes: Abstract Local rhizosphere acidification by rape as a reaction to P-starvation was visualized by means of an agar plate technique. By means of a modification of this technique local differences in cation-anion uptake and organic acid exudation along intact roots of rape were observed for plants grown on nutrient solution with or without added P. No differences in uptake rates of K-, NO3- and Ca-ions could be detected between P-starved and P-supplied plants. However, exudation of malic and citric acid was distinctly higher in acidified root zones of P-starved plants, coinciding with higher levels of malate in the corresponding root tissue. Organic acid exudation is indicates as the cause of local rhizosphere acidification by rape as a reaction to P-starvation and as a possible mechanism of its phosphate-solubilizing capacity.
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  • 26
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    Plant cell, tissue and organ culture 19 (1989), S. 213-224 
    ISSN: 1573-5044
    Keywords: Brassica napus ; B. oleracea ; rapid-cycling brassica populations ; protoplasts ; regeneration ; maltose
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Protoplasts were isolated from aseptic shoot cultures of commercial cultivars ofBrassica napus, B. oleracea andB. campestris, and from the six ‘rapid-cycling brassica species’. Of the rapid-cycling species, onlyB. napus responded well to the culture conditions used; 2% of protoplasts formed calli and up to 5% of calli regenerated shoots. Regeneration was also achieved from commercial cultivars ofB. napus andB. oleracea. For these two species the plating density, time of dilution with fresh medium and the composition of the shoot-inducing medium were all found to have an important influence on the efficiency of plant regeneration. Both responded better to maltose than to sucrose-based media. Under the optimum conditionsB. napus showed a plating efficiency of 7.8% and shooting efficiency of 17%; forB. oleracea the figures were 2% and 56%, respectively.
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  • 27
    ISSN: 1573-5044
    Keywords: Oryza sativa L. ; somaclonal variation ; grain and quality characters ; protein content
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Variations in some grain and quality characters in progenies of regenerated rice plants were studied. Grain length and weight decreased significantly, yet gel consistency increased. Variations in these quantitative characters of all cultivars studied were consistent, showing the tendencies of the variations. Grain protein contents of the somaclones were higher in one cultivar. Variability of most traits was increased by combining low-dosage radiation and tissue culture.
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  • 28
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    Biomedical Chromatography 3 (1989), S. 46-47 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Different metal salts have been tried as impregnating reagents for developing TLC separation schemes for some antihistamines on silica gel ‘G’ plates, using a new solvent system benzene + dimethyl formamide + acetic acid (30:10:7). Spots were visualized by spraying with a solution of Dragendroff's reagent.
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  • 29
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    Biomedical Chromatography 3 (1989), S. i 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 30
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    Biomedical Chromatography 3 (1989), S. 10-13 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A simplified method for the preparation of wheat germ agglutinin(WGA)-Sepharose 4B by coupling highly purified WGA, prepared by improved affinity chromatography, with BrCN activated Sepharose 4B in a solution of high carbonate buffer is described. The amount of WGA linked to Sepharose 4B was 82.40% (3.07 mg WGA per ml Sepharose 4B). MN blood group antigens of human erythrocyte membranes purified with WGA-Sepharose 4B affinity chromatography showed a single band on sodium dodecyl sulfate/polyacrylamide gel electrophoresis (SDS/PAGE). The yield of the antigens from 400 mL fresh blood was 32-40 mg. The WGA-Sepharose 4B column could be used several times without loss of activity.
    Additional Material: 4 Ill.
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  • 31
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    Biomedical Chromatography 3 (1989), S. 32-34 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A method for the unambiguous characterization of DNA photoproducts has been developed. It does not require radio-labelled DNA, or specialized techniques. In the preliminary step, UV-irradiated DNA is hydrolysed to its constituent bases and photoproducts. The photoproducts are then separated from nucleic acid bases using low pressure ion-exchange chromatography on a Dowex 1 × 8 200-400, and an Amberlite CG-50-H+ column. Further separation and purification of photoproducts is carried out on the HPLC Whatman Partisil ODS2 column, using 15% MeOH. The procedure is simple, reproducible and versatile.
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  • 32
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    Biomedical Chromatography 3 (1989), S. 43-45 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Two new solvent systems, n-hexane + propionic acid (26:5, v/v) and chloroform + acetone (29:3, v/v), for the rapid resolution and identification of an 18-component mixture of phenylthiohydantoin amino acids are reported. Using these systems certain difficult combinations of phenylthiohydantoin amino acids are resolved. Two more solvent systems, viz chloroform+acetic acid (27:3, v/v) and chloroform + methanol (30:4, v/v), are developed to resolve phenylthiodantoin derivatives of aspartic and glutamic acids.
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  • 33
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    Biomedical Chromatography 3 (1989), S. i 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 34
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    Biomedical Chromatography 3 (1989), S. 49-52 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A simple rapid method for simultaneous ethosuximide and phenobarbital assay in brain tissue, serum and urine has been developed. Extraction of samples from brain tissue and serum were performed with dichloromethane at low pH in the presence of an excess of ammonium sulfate. Glucuronide conjugates in urine samples were hydrolyzed by enzymatic cleavage with β-glucuronidase and then extracted with dichloromethane. The extracts were analyzed using a Spherisorb 5 ODS column and a mixture of acetonitrile, methanol and phosphate buffer (21: 24: 55, v/v) as eluent. No interference was encountered and the method is both precise and reproducible.
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  • 35
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    Biomedical Chromatography 3 (1989), S. 136-138 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A high performance liquid chromatographic (HPLC) method for the determination of the anticoccidial and antitheilerial drug halofuginone in bovine plasma was developed. Samples were diluted with acetic acid (10%, v/v) and cleaned up on a Bond Elut C8 column. The analyte was eluted from the extraction column and chromatographed by reversed-phase HPLC using decylamine as a competing-ion reagent. Detection was by UV at 243 nm. Recovery from plasma was 75%, and within-day and between-day coefficients of variation were 5.23 and 6.35% respectively. The specificity and sensitivity of this method (limit of detection in plasma, 1 ng/mL) were sufficiently high to enable us to characterize the time course of the drug in plasma after oral administration of therapeutic doses to cattle.
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  • 36
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    Biomedical Chromatography 3 (1989) 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 37
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A new method based on displacement electrophoresis has been developed for the determination of the total and free concentration of propranolol, a β-adrenergic blocker drug, in plasma. To determine the total concentration the drug is extracted from human plasma into a chloroform + heptane mixture in the presence of ammonia. After evaporation of the solvent mixture the residue is submitted to displacement electrophoresis in a glass tube containing a two-layer polyacrylamide gel. When the propranolol electrophoretically leaves the gel column it is transferred by a buffer flow to the cuvette of a fluorimeter for continuous detection and quantification. The concentration of the free non-protein bound drug can be determined by the same displacement electrophoresis technique following extraction of the plasma sample into the above organic solvent mixture in the absence of ammonia. Alternatively the extraction procedure can be exchanged for dialysis for 1 h. To decrease the risk that large size material might comigrate with propranolol two layers of polyacrylamide are used, one having small pores to retard such material. In addition, we use fluorimetric detection which means that many possible contaminants are not recorded and therefore do not affect a quantitative determination of propranolol.
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  • 38
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    Biomedical Chromatography 3 (1989), S. 255-261 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Analyses of estrogen and progesterone receptors in biopsies of breast carcinoma play a vital role in the selection of patients likely to respond to hormone manipulation. Sucrose density gradient centrifugation has been the reference method in the determination of estrogen receptors in human breast carcinoma cytosols. To reduce assay time and circumvent prolonged manipulation of labile receptor preparations, high performance liquid chromatography techniques in the size-exclusion and ion-exchange modes were compared as potential alternate methods for the rapid separation of receptor isoforms. Multidimensional analyses were performed by reapplying estrogen receptor isoforms obtained from high performance size-exclusion and ion-exchange chromatography to sucrose density gradients and vice versa. This confirmed that the estrogen-binding components identified by high performance liquid chromatography appear to correspond to estrogen receptor species from sucrose density gradients.
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  • 39
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    Biomedical Chromatography 3 (1989), S. 269-271 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A sensitive (2 ng/mL) and specific method for the determination of the benzodiazepine antagonist Ro 15-1788 or flumazenil is described. Following a simple extraction, the compound is analyzed by reversed phase high performance liquid chromatography (HPLC) and detection at 245 nm. The method was applied to plasma specimens collected from patients receiving a single dose of this drug.
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  • 40
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A simple and precise method for the quantitation of epomediol in human plasma and urine is described. Each biological sample is added with the internal standard and applied directly to an Extrelut-1 solid-phase column. After absorption the column is eluted with chloroform and the eluate is evaporated to dryness. The residue, reconstituted in ethanol, is analysed by capillary gas chromatography. No interferences from possible metabolites or endogenous constituents can be noted. The method has been applied to human pharmacokinetic studies: the results of a subacute administration to volunteers are presented.
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  • 41
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Malonyl-CoA is a biochemically important compound, formed by an acetyl-coenzyme A carboxylase catalysed reaction. The stability of this short-chain coenzyme A derivative under various experimental conditions is discussed in this article. High-performance liquid chromatography was used for the analysis of the reaction mixture because of its excellent selectivity and sufficient sensitivity. Several variables were investigated as possible stability-influencing factors: pH, magnesium and buffer concentration, reaction temperature and time. The Plackett-Burman screening design was first used for selecting the most important variables, with which a central composite design was constructed. In this way, a response surface was obtained with the percentage remaining malonyl-CoA as a function of magnesium concentration, reaction temperature and time. The usefulness of this approach is demonstrated by obtaining kinetic data from the mathematical function and by the evaluation of the stopping of reaction procedure in the activity assay of acetyl-coenzyme A carboxylase.
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  • 42
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    Biomedical Chromatography 3 (1989), S. 221-225 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Rabbit kidney (RK-13) and human jejunum and ileum (I-407) cells infected with herpes simplex virus type 1, strain F, were radiolabelled with [14C]glucosamine or [35S]methionine for 24 h. The cells were extracted with 1% Triton X-100 and the extracts were separated by gel filtration high performance liquid chromatography. Monoclonal antibody immunoprecipitation of the fractions collected from the column revealed a monomeric glycoprotein D (gD) of 52 - 56 000 molecular weight from RK-13 cells and two monomeric forms of gD, 54 000 and 58 000 molecular weight, from I-407 cells. Densitometry scanning of the autoradiograms from SDS-PAGE showed gD from the RK-13 host cells to be 98.7% pure with the [35S]methionine label and 97.0% pure with the [14C]glucosamine. On the other hand, gD from the I-407 host cells was only 78.6% with the [35S]methionine label and 96% pure with the [14C]glucosamine. This method could provide a means for the isolation of native gD for structural and immunological studies.
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  • 43
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    Biomedical Chromatography 3 (1989), S. 233-240 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The dinitrophenyl (DNP) derivatives of amino acids have found continual application in protein sequencing since Sanger used them for the first time for the sequencing of insulin. Dansyl derivatives of amino acids have been widely used in protein sequencing because of their fluorescent nature. The success of protein sequencing largely depends upon correct identification of such derivatives. The choice for the method of identification is related to cost, the availability of instrumentation and to the sensitivity needed for the analysis. Thin layer chromatography (TLC) is simple and has several advantages over other chromatographic methods. Therefore the literature after 1972 is reviewed for TLC analysis of dansyl- and DNP-amino acids, the two important amino acid derivatives required for identifying protein sequences. Additionally, the literature on the TLC resolution of enantiomeric mixtures of dansyl amino acids is reviewed. Application of various adsorbents, composition of solvent systems and other experimental conditions together with successful resolution data have been discussed. TLC provides a direct and inexpensive method for the resolution of enantiomers, and is fast becoming a sensitive instrumentalized quantitative analytical technique.
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  • 44
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    Biomedical Chromatography 3 (1989), S. 272-273 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: An HPLC method for the simultaneous detection of six organophosphorus pesticides (Dimethoate, Ethion, Malathion, Phorate, Phosalone and Parathion) on a Zorbex ODS column using methanol + water (80: 20) as solvent is described.
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  • 45
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A novel analytical method for biological polyamines (putrescine, spermidine and spermine) was developed. Polyamines were separated by ion-pair reversed phase chromatography using a polymer-based octadecyl bonded column. A polyamine oxidase immobilized column worked effectively as a post-column reactor to convert polyamines to hydrogen peroxide which was eventually detected by electrochemical oxidation on platinum electrode. This method required neither tedious derivatization nor gradient elution, permitting us to perform simple and rapid analysis of polyamines. The detection limits were 0.3, 0.6, and 4 pmol injected for putrescine, spermidine, and spermine, respectively with a linear range of two to three orders of magnitude. Chromatograms obtained with samples from human urine and rat brain homogenates demonstrated the high sensitivity and selectivity of the method.
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  • 46
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    Biomedical Chromatography 3 (1989), S. 276-278 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Highly purified monomeric human plasma lecithin:cholesterol acyltransferase (LCAT), completely free of apolipoprotein D, has been chromatographed on a MonoQ HR 5/5 anion exchanger. LCAT eluted as symmetrical peaks after 12.8 min and 14.8 min at pH 5.0 and pH 6.0, respectively, using a linear NaCl gradient. The corresponding concentrations of NaCI effecting desorption of LCAT from the anion exchanger were 125 mM and 175 mM. At both pH values human serum albumin eluted earlier and was well separated from the enzyme. Rechromatography of LCAT in the eluates from these experiments at acid pH, on high performance gel filtration, demonstrated absence of aggregation. The nonspecific adsorption during anion exchange chromatography at pH 5.0 and pH 6.0 was negligible, as demonstrated by a linear relationship between injected amounts of LCAT and recorded peak areas for a 2-20 μg protein range. Zone immunoelectrophoresis assay indicated unaltered immunoreactivity of the eluted LCAT.
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  • 47
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The chromatographic behaviour of the membrane-attached variant surface glycoprotein (mfVSG) of bloodstream forms of Trypanosoma brucei brucei AnTat 1.1 A preparations were studied by reversed phase high performance liquid chromatography (RP-HPLC). Among the different preparation procedures used, only the trifluoroacetic acid extraction gave a mfVSG preparation which was eluted from the RP-HPLC column. As well as for the soluble variant surface glycoprotein (sVSG) it was found that different mfVSG forms were eluted at different organic solvent concentrations from the RP-HPLC column. In addition, in preliminary studies, we have attempted to characterize the factors responsible for the trypanosome surface coat assembly. Using chromatographic techniques (RP-HPLC and thin layer chromatography), the results suggest a close relationship between the content of lipid-containing species and the heterogeneity of mfVSG preparation.
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  • 48
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    Biomedical Chromatography 3 (1989) 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 49
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    Biomedical Chromatography 3 (1989), S. ii 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 50
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    Biomedical Chromatography 3 (1989), S. iii 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 51
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    Biomedical Chromatography 3 (1989), S. 153-156 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: An automated column-switching HPLC system is described for the simultaneous determination of midodrine, an alpha-adrenergic stimulating drug, and its active metabolite, ST-1059. Serum or plasma (850 μ L) is directly injected onto a RP 18 (30 μm particle size) pre-column (9 × 4 mm ID) which acts as an on-line liquid-solid extractor and analyte enrichment system. The injection is followed by washing steps. The fraction containing the analytes is transferred onto an analytical RP18 column via step gradient elution where the final analysis is performed. Fluorescence detection is used (λex 290 nm and λem 322 nm), and method detection limits of 0.8 ng/mL plasma were reached. These were sufficiently low to determine the plasma concentration-time profiles for both compounds following oral administration of 2.5 mg and 5 mg midodrine hydrochloride. The assay in serum or plasma was linear in the range of 1 to 15 ng analyte/mL, the recovery was 〉95%, and the reproducibility was sufficient. The assay was rugged and was maintained by routinely changing the home-made, dry packed pre-column every 20th serum injection.
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  • 52
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    Biomedical Chromatography 3 (1989), S. 177-179 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A method has been developed for the separation and measurement of fluvoxamine in human plasma by high performance liquid chromatography. The method uses metapramine as an internal standard and provides a limit of detection of about 1.5 ng/mL for fluvoxamine. At a concentration of 25 ng/mL, fluvoxamine could be measured within a coefficient of variation of ±5.82 of the mean and at 100 ng/mL within a CV of ±2.78 of the mean. The method has been applied to the analysis of plasma from patients undergoing fluvoxamine therapy.
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  • 53
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Conditions for unblocking reversible chemical modifications such as maleylation or citraconylation ‘in situ’ at the N-terminus of proteins after transfer of proteins to immobilon membranes from SDS-PAGE are described. Demaleylation or decitraconylation occurred at 55°C in 70% formic acid (pH 1.50) during 60 min. During the unblocking reaction, Coomassie blue dye was completely removed, resulting in superior high performance liquid chromatographic separation of phenylthiohydantoin-amino acid (PTH-AA) after Edman degradation (automatic gas phase sequencer). The protein fixed on the matrix after demaleylation and removal of Coomassie blue was not degraded. The possible cleavage at the aspartyl-prolyl peptide bonds was considered, but no side reaction was observed. Furthermore, the incubation time in 70% formic acid at 55°C could be reduced to 10 min in the absence of maleylation of the starting material, and this was suitable for the removal of Coomassie blue and the quantification of phenylthiolhydantoin-amino acids (PTH-AAs) by HPLC. The yield from the starting protein through SDS-PAGE, blotting, and Edman degradation to quantitative analysis of PTH-aminoacid(s) by HLPC was established.
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  • 54
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 55
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    Biomedical Chromatography 3 (1989) 
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    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 56
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    Biomedical Chromatography 3 (1989), S. 192-195 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: An HPLC method with direct sample injections and column switching was investigated for the analysis of drugs in tissue homogenates. The appropriate precolumn packings and analytical column packings were surveyed in order to obtain quantitative recovery. The use of a large bore end-fitting filter for the precolumn avoided the interference due to minute tissue particles. A minicolumn was used to trap the analyte or clean up the sample. The method was applicable to hydrophilic as well as hydrophobic drugs in liver, kidney or heart tissues, and has been extensively used in the determination of drugs in centrifugal cell fractions such as the nucleus, mitochondria and microsome.
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  • 57
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    Biomedical Chromatography 3 (1989), S. 196-198 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A quantitative method for the determination of estriol (E3) and creatinine (C) in random urine by high performance liquid chromatography is described. The mobile phase was a mixed solution of methanol and phosphate buffer (0.025 M, pH 6.5) and the detection wavelength was at 205 nm. The method was simple, rapid and accurate. The OCV for E3 and C using this method were 1.7 - 3.4% and 2.2 - 2.5%, respectively. The RCV for E3 and C were 6.2 - 7.0% and 4.5 - 6.9%, respectively. The recoveries were 87 - 104% for E3 and 98 - 103% for C, respectively. The method has been used for clinical determinations.
    Additional Material: 5 Ill.
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  • 58
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Different strategies for HPLC separation, including molecular sieving, ion-exchange, and hydrophobic interaction as well as reversed phase chromatography, were used to study molecular components in human cerebrospinal fluid (CSF). The separations were followed by photodiode-array UV detection, which is a recently developed technique allowing a direct and rapid discrimination between peptides and proteins differing in their content of aromatic amino acids. By the various HPLC techniques in conjunction with diode-array detection it was possible to identify and characterize several protein and peptide components present in CSF. The procedure also allowed quantitative analysis of CSF proteins using minute amounts of the fluid.
    Additional Material: 8 Ill.
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  • 59
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A newly developed method for the simultaneous extraction and quantitation of the unconjugated levels of the catecholamine metabolites vanilmandelic acid (VMA), 3-methoxy-4-hydroxyphenylethylene glycol (MHPG) and homovanillic acid (HVA) in plasma by high performance liquid chromatography with electrochemical detection was modified and applied to studies of human saliva. The assay had a mean coefficient of variation under 3% for each of the metabolites. Levels of plasma VMA, MHPG and HVA were measured in 28 normal subjects and compared to their saliva levels, obtained before and after stimulation by mastication. Significant correlations were found between plasma and saliva MHPG and HVA, but there was no correlation between plasma and saliva VMA. Salivary MHPG and HVA can be reproducibly assayed and may be useful tools for indications of changes in central and peripheral catecholamine metabolism.
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  • 60
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    Biomedical Chromatography 3 (1989), S. 5-9 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A sensitive liquid chromatography method has been developed using electrochemistry for the determination of leukotrienes in biological fluids. Biological specimens are treated with 3,5-dinitrobenzoyl chloride in acetonitrile which undergoes rapid reaction with hydroxyl groups of non-peptidic leukotrienes in the presence of pyridine and with amino groups of peptidic leukotrienes in the presence of potassium tetraborate buffer. The resulting dinitroben-zoate derivatives of leukotrienes are highly electroactive, suitable for reduction or oxidation at moderate potentials by an electrochemical detector. In reductive mode at -0.7 V or oxidative mode at + 1.15 V potentials, the lower limits of detection for leukotriene derivatives were approximately 8±3 pg and 70 ± 16 pg respectively, with a signal-to-noise ratio of 5 to 1. This method was applied to the detection of leukotrienes in plasma, nasal and bronchial fluids of patients with asthma.
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  • 61
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A new capillary GC method is described for the compositional analysis of the three main gangliosides isolated from adult human myometrium. The sample was subjected to methanolysis, acetylation and trimethylsilylation which allows all the constituents to be analyzed simultaneously. The predominant ganglioside was found to be GD3, with GM3 and GT1b the next most abundant.
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  • 62
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A gas chromatographic-mass spectrometric (GC/MS) method for analysis of Δ1-tetrahydrocannabinol (Δ1-THC) in human fat samples is described. The fat sample, obtained from heavy marihuana users 1 week before and 4 weeks after smoking, is homogenized in hexane + 2-propanol, centrifuged, and the supernatant mixed with Lipidex 5000. The solvent is evaporated and the dried gel is packed in a glass column. Δ1 -THC is eluted from the column with methanol + water + acetic acid, diluted with water and the eluent is passed through a bed of Octadecylsilanebonded silica. After washing and drying, the retained Δ1-THC is eluted with hexane, derivatized with N-methyl-N-(t-butyl-dimethysilyl)trifluoroacetamide (MTBSTFA) and finally purified by HPLC on an Octadecyl SI 100 column in methanol. The amount of Δ1-THC is determined by GC/MS, using selected ion monitoring, and a deuterated internal standard. The recovery of Δ1-THC is about 80%, and the concentration of Δ1-THC in the fat samples analysed ranged between 0.4 and 193 ng/g wet tissue.
    Additional Material: 3 Ill.
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  • 63
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The peroxyoxalate chemiluminescence detection of biological thiols combined with high-performance liquid chromatography (HPLC) is described. SH groups of the thiol compounds including glutathione (GSH), cysteine, N-acetylcysteine, cysteamine, and D-penicillamine were labelled with N-[4-(6-dimethylamino-2-benzofuranyl)phenyl]maleimide (DBPM), a specific fluorogenic reagent for SH group. The labelling reaction was carried out at 60°C for 30 min and at pH 8.5 and a sample of the resulting reaction mixture was subjected to HPLC. Five kinds of labelled thiols were separated within 12 min on ODS-80 column (150 × 4.6mm ID; 5 μm) and detected in the ranges from 500 fmol to 2 pmol/100 μL (cysteamine and N-acetylcysteine), to 3 pmol/100 μL (cysteine) and to 5 pmol/100 μL (GSH and D-penicillamine). The lower detection limits were from 7 fmol (cysteamine) to 113 fmol (GSH) per 100 μL (S/N = 2). The method was applied to the determination of thiols in a rat liver. The amounts of glutathione and cysteine were 1.23±0.15 μmol/g (n = 5) and 0.15±0.04 μmol/g (n = 5), respectively.
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  • 64
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    New York, NY : Wiley-Blackwell
    Biomedical Chromatography 3 (1989) 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 65
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A rapid assay employing HPLC with electrochemical detection for catechol-O-methyltransferase (COMT) activity in red blood cells is described. Enzyme activity is determined from erythrocyte lysates using S-adenosyl-L-methionine as methyl donor and 3,4-dihydroxybenzoic acid as substrate. The 3-O- and 4-O-methylated reaction products are measured by high-performance liquid chromatography with electrochemical detection. Human erythrocyte soluble form of COMT had Km values of 6.1 μM and 26.0 μM for S-adenosyl-L-methionine and dihydroxybenzoic acid, respectively. The mean O-methylation ratio for the soluble form of COMT was 5.3. An O-methylation ratio of 15.5 was estimated in the membrane fraction of an erythrocyte pool from three samples. The activities of soluble COMT in erythrocytes of some animal species are also reported. The procedure is easily automated, and a large number of samples can be processed during one working day.
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  • 66
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    Biomedical Chromatography 3 (1989), S. 72-74 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A reversed-phase HPLC system with 88% methanol in 1 M ammonium acetate buffer pH 5.35 as the mobile phase on an ODS column was used to analyse pheophorbides a, a', b, b' and pyropheophorbides a and b. Pyropheophorbides a and b were found in samples obtained from silkworm excrement but not from spinach leaves though the preparation methods used were the same. This difference suggests that chlorophylls undergo metabolism in the body of silkworm to give pyrochlorophyll derivatives.
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  • 67
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    Biomedical Chromatography 3 (1989), S. 82-87 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Hormone-sensitive lipase (HSL), the enzyme controlling the rate of adipose tissue lipolysis and also possibly involved in the regulation of steroidogenesis, has been purified from bovine omental adipose tissue. Partially detergent-solubilized, delipidated and purified HSL was obtained through step-elution at conventional DEAE ion-exchange chromatography, followed by concentration on hydroxylapatite. High performance hydrophobic interaction chromatography (HPHIC) on phenylsilica then resulted in an increase of HSL protein purity from 2% to more than 70%. Final purification of the enzyme to apparent homogeneity (〉95% protein purity), concentration and removal of most of the detergent was obtained by high performance cation exchange chromatography on Mono S. At least 0.5 mg of highly stable HSL was obtained from 5 kg of bovine omental fat within four working days. The purified lipase had a lower specific activity than previously reported for the corresponding rat enzyme but the preparations have proved very useful for enzyme structure studies and as an antigen.
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  • 68
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    Biomedical Chromatography 3 (1989), S. i 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 69
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    Biomedical Chromatography 3 (1989), S. 95-104 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Additional Material: 3 Ill.
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  • 70
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Fluorescein isothiocyanate (FITC)-labelled asialotransferrin and pyridyl aminated oligosaccharides were prepared from asialotransferrin and human milk using affinity chromatography and high performance liquid chromatography (HPLC), respectively. These substances were incubated with galactosidase or sialyltransferase and then examined by lectin affinity HPLC. The elution patterns changed according to the period of incubation and amount of enzyme. This analytical method using lectin affinity HPLC with fluorescence labelled glycoprotein or oligosaccharides as the substrates has great value for detecting these enzyme under the same chromatographic conditions. In addition, differences were noted in the activity of β-galactosidase toward oligosaccharides having the Gal β(1 → 3)GlcNAc or Gal β(1 → 4)GlcNAc structure at reducing termini.
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  • 71
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The isolation and properties of lactoferrin from human breast milk and from neutrophilic granulocytes were investigated. Human breast milk lactoferrin was purified by means of heparin-sepharose or Cibacron Blue affinity chromatography. Quantitative recovery using these two methods was comparable but Cibacron Blue affinity chromatography allowed for isolation of a more homogenous protein. Lactoferrin could only be isolated from human neutrophilic granulocytes by sequential use of antibody affinity followed by non-specific affinity chromatography. Both breast milk lactoferrin and granulocyte lactoferrin were separated into apo and iron-rich species by SDS polyacrylamide gel chromatography. Iron binding is accompanied by a conformational change in tertiary structure associated with more rapid electrophoretic migration. The isoelectric point of both human breast milk lactoferrin and human granulocyte lactoferrin is 5.5 - 6.2. Both types of lactoferrin have similar iron binding properties with release of iron from the one binding site occurring at pH 5.2 - 6.0 while the other binding site holds on to iron down to pH 3.6 - 3.2. Despite the high affinity for iron the percentage saturation of native lactoferrin is low, that for breast milk lactoferrin averaging 12 - 25% and that for granulocyte lactoferrin 〈 10%.
    Additional Material: 7 Ill.
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  • 72
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    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Human trypsin inhibitor (home prepared), lactalbumin, trypsinogen, carbonic anhydrase, and bovine serum albumin were submitted to succinylation and their molecular masses were determined by SDS-PAGE according to the method of Weber and Osborn (1969 J. Biol. Chem. 244, 4406) before and after chemical modification. High estimates of their molecular masses were obtained. The monomer and dimer of arrowhead inhibitor proteinase-B (Chinese vegetable legume) obtained after chemical crosslink(s) were also submitted to SDS-PAGE and their apparent molecular masses were also determined and compared to the native arrowhead inhibitor proteinase-B. Abnormally high estimates of their molecular masses were obtained. Our results agree with those in the literature.
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  • 73
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    Biomedical Chromatography 3 (1989) 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 74
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    Biomedical Chromatography 3 (1989), S. 114-117 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A HPLC method for the quantitative determination of 5-hydroxy-3-indoleacetic acid (5-HIAA) in urine is described. The method is based on ion-pair chromatography, reversed phase (RP) column material and specific fluorimetric detection at 300 nm and 355 nm. Sample preparation and gradient elution were avoided by using a column-switching technique. The sensitivity of the assay was excellent for clinical routine analysis, with a detection limit of 0.2 mg/L 5-HIAA. No endogenous or exogenous interference problems arose. Intra- and interassay precision was good, with observed coefficients of variation of 1.5 to 2.6% and 2.1%, respectively. Recoveries were 93 to 98%. The system described can be used for clinical diagnosis and therapy follow-up of carcinoid tumors. It has been running for over a year without disturbances and with a minimum of technical attendance.
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  • 75
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    Biomedical Chromatography 3 (1989), S. 127-130 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The soluble form of catechol-O-methyltransferase (EC 2.1.1.6.) from rat liver was purified to homogeneity by high-performance anion-exchange chromatography and high-performance gel-filtration chromatography. The specific activity of the final pool was 270 U/mg protein. The purification was 1180-fold and recovery of the enzyme activity was 15%. During this rapid and gentle purification there were no problems with loss of activity, and the estimated half life of the final purified enzyme pool was 5.5 days at +4°C. The only additive used was phenylmethylsulfonylfluoride in the homogenizing buffer.
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  • 76
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Analytical conditions of prepurification extraction and HPLC separation were optimized for determination of urinary serotonin and tryptamine. Under optimal conditions, serotonin, tryptamine and an internal standard were extracted with 15% v/v n-propanol in diethyl ether from urine samples alkalized with a phosphate buffer (0.75 mol/L, pH 10.0), and then they were re-extracted into an HCI solution (0.1 mol/L). Purified indoleamines were simultaneously separated by reversed-phase ion-pair HPLC with native fluorescence detection. Urinary serotonin and tryptamine were selectively determined within about 45 min per sample for the whole procedure. Analytical recovery, reproducibility and detection sensitivity were satisfactory for pursuing time-dependent changes in indoleamine levels. Urinary excretion profiles of serotonin and tryptamine in subjects dosed with L-tryptophan were successfully analyzed by our method.
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  • 77
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    Biomedical Chromatography 3 (1989) 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 78
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    Biomedical Chromatography 3 (1989), S. 241-245 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A practical procedure for isolating and purifying glycopeptides is described, viz. enzymatic hydrolysis - gel permeation chromatography - ion exchange chromatography - reversed phase HPLC. Using this procedure 28 glycopeptides from hen ovalbumin have been isolated some of which hitherto have not been identified. Water was a suitable mobile phase for preparing pure glycopeptides, and control of column temperature was important for good separations and reproducible retention times. Structural confirmation was by fast atom bombardment mass spectrometry.
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  • 79
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    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A method is described to isolate rapidly human hemoglobin-benzo[a]pyrene diol epoxide adducts. A combination of 300 Å pore size C4 reversed phase HPLC to effect separation of adducted protein from native protein, and μ-bore C18 reversed phase HPLC to isolate and partially characterize proteolytic peptide adducts (by UV), was used.
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  • 80
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A simple, accurate and precise procedure for the quantitation of itazigrel (a potent lipophilic inhibitor of collagen and arachidonic acid-induced aggregation being studied for its effects on peripheral vascular disease) from granulated rodent diet is presented. The drug was extracted from rodent diet using methanol + water (80:20) following dissolution of the diet in water. Samples of the supernatant were injected into the HPLC and the eluent was monitored with a fluorescent detector (λex = 320 and λem = 430) to achieve analytical specificity. Interday coefficients of variation of the calibration curve slope were ±6% on standards between 0 and 1000 μg/g. Potency and homogeneity of the drug spiked diet prepared over a 1 year interval at 70,200 and 600 μg/g was 99.3 ± 2.5%, 100 ± 1.8%, and 101 ± 1.9% of label, respectively. Samples prepared for chromatography were stable for 24 h at 20°C, and drug in diet was stable for 102 days when protected from light and stored at 20°C.
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  • 81
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    Biomedical Chromatography 3 (1989), S. i 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 82
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    Biomedical Chromatography 3 (1989), S. 183-185 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: The release and content of the excitatory amino acid neurotransmitters glutamate and aspartate in rat striatum were determined by liquid chromatography/electrochemistry. This determination was based on precolumn off-line derivatization of the amino acids with o-phthaldialdehyde and 2-mercaptoethanol (OPT/2-MCE), and the adducts formed were separated under isocratic conditions and oxidized on a glassy carbon electrode at moderate potential (+0.6 V). The standard and the extracted glutamate when derivatized with OPT/2-MCE produced similar electrochemical and chromatographic characteristics. The detection limit of glutamate was 0.5 pmol. Depolarization induced by the high potassium medium (40 mmol/L) enhanced the release of glutamate and aspartate from superfused rat striatum, whereas the efflux of glutamine remained unchanged. Perfusion (for 60 - 70 min) removed 50 - 80% of the free amino acid content of striatal tissue. The method described here is useful in neurochemical investigations of the brain amino acid neurotransmitters.
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  • 83
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    Biomedical Chromatography 3 (1989), S. 209-212 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A method for the determination of cyanide in human urine has been developed. The method is based on the reaction of cyanide with 2,3-naphthalenedialdehyde and taurine to give a fluorescent product for reversed-phase HPLC separation and fluorometric detection. After centrifugation followed by dilution of urine samples, the specimens could be analysed directly by this method. The recovery of cyanide added to urine at concentration levels of 50 - 1000 pmol/mL was 85 - 96%. The detection limit of cyanide was 30 pmol/mL in urine. The method was successfully applied to the analysis of urine from smokers and nonsmokers. The mean concentrations of cyanide were found to be 215 pmol/mL for the former and 84 pmol/mL for the latter.
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  • 84
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    Biomedical Chromatography 3 (1989), S. i 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
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  • 85
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    Biomedical Chromatography 3 (1989), S. 226-232 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A simple, selective and very sensitive assay is described for the quantification of physostigmine in blood, plasma and urine. The most appropriate solid phase column was selected after a systematic investigation of nine types of phase. The conditions for solid phase extraction were optimized using [3H]physostigmine so that the overall recoveries were 〉90%. Physostigmine was retained on alkaline treated cyanopropyl columns and eluted into the minimum volume of methanol, obviating the need for an evaporation step. Extracted samples were quantified by HPLC with a three electrode coulometric detection system. The limit of detection was 50 pg/mL for a 0.5 mL plasma sample. The precision (CV) for 0.5 mL plasma samples containing 50 pg was 8.1%. Application of the method to plasma, blood and urine samples is presented.
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  • 86
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    Biomedical Chromatography 3 (1989), S. 251-254 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: Meibomian secretions from the hamster were hydrolysed with base and examined as TMS, [2H9]TMS, methyl ester/TMS, picolinyl ester/TMS and nicotinate/TMS derivatives by capillary GC/MS. Over 90 compounds, representing over 89% of the hydrolysed fraction, were identified. Fatty acids with chain lengths from 10 to 32 carbon atoms were found, the most common of these were in the C15 to C18 and in the C25 to C30 regions. Chain types were predominantly iso or anteiso branched, mono-unsaturated (C16 and C18) and straight. Fatty alcohols were mainly from the iso or anteiso series and tended to have longer chain lengths; the major alcohols had anteiso-25 and 27 and iso-26-chains. In these respects the secretions were similar to those reported earlier from other species, although fewer mono-unsaturated compounds with longer chains (C20 to C30 region) were found than in the rat and human. The steroid fraction was characterized by a larger number of compounds than normally present in secretions of this type. The major compound was cholesterol, in common with that in all other examined species except the rabbit.
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  • 87
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    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A simple and precise method was developed for the separation of nucleosides including modified nucleosides and oligonucleotides. Nineteen kinds of nucleosides were completely separated by HPLC using an ODS column (TSK-gel ODS 80TM) and aqueous mobile phases. The RNA molecule was digested by base restrictive RNase (RNase A, RNase T1) and the digests were separated chromatographically into each oligonucleotide. The nucleoside composition of an oligonucleotide was then determined by this analytical system. It is thus possible to fit the oligonucleotide in the original RNA molecule by using modified bases as markers. The reaction site of quinacrine mustard for tRNAPhe (from yeast) could be determined by this analytical system.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
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  • 88
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Biomedical Chromatography 3 (1989), S. 262-265 
    ISSN: 0269-3879
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Medicine
    Notes: A high performance liquid chromatographic method is described for the simultaneous determination of furosemide and bumetanide in horse plasma. The C8 (3 μm) reversed phase column (4.8 × 150 mm) provided clear separation of furosemide and bumetanide with other components present in the horse plasma. The detection limit for both the drugs was 10 ng/mL. Both drugs were stable in plasma (at natural or acidic pH) for up to 24 h. The method is sufficiently sensitive to detect furosemide levels in plasma obtained from horses receiving a therapeutic dose of furosemide.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
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  • 89
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Journal of Chemometrics 3 (1989), S. 601-608 
    ISSN: 0886-9383
    Keywords: Gas phase flow injection analysis ; Membrane-differentiated analysis ; Multicomponent determinations ; Successive linear regression ; Successive regression in fiduciary region ; Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: A membrane-discriminated gas phase analyzer is proposed for multicomponent determinations. Nitrogen gas flows countercurrent through outer and inner channels in a tube-in-tube arrangement. The only communication between the two channels occurs through a 500 μm aperture covered by a porous PTFE membrane. A mixture of organic compounds (up to four components) is injected into the inner channel by a heated backflushed injector and the sample components diffusing into the outer channel are monitored by a flame ionization detector (FID). A calibration set, consisting of pure components, binary, ternary and quaternary mixtures (a total of 64 samples), provides the known data base: temporal profiles of the FID output as a function of sample composition. Although the overall response behavior is not a linearly additive function of individual analyte concentrations, the use of successive inverse multiple linear regression (while continually altering the choice of the calibration samples considered for the forward regression, on the basis of the most recent values of the predicted unknown sample composition) is shown to yield analytical results for unknown samples that are in good agreement with their true values.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
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  • 90
    Electronic Resource
    Electronic Resource
    New York, NY : Wiley-Blackwell
    Journal of Chemometrics 3 (1989), S. 609-609 
    ISSN: 0886-9383
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Type of Medium: Electronic Resource
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  • 91
    Electronic Resource
    Electronic Resource
    Chichester : Wiley-Blackwell
    Biological Mass Spectrometry 18 (1989), S. 174-176 
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: Plasma levels and urinary excretion of mevalonate were reported to be correlated with cholesterol biosynthesis. Evaluation of mevalonate concentration in plasma and urine represents therefore a non-invasive method for studying the modifications of cholesterol synthesis. A method is described here by wich mevalonate in plasma and urine is determined by the selected ion monitoring technique after extraction as mevalonolactone and conversion into the trimethylsilyl ether. Linear responses were obtained in the evaluation of mevalonate added to plasma in the 10-100 ng/ml (r 〉 0.995) and to urine in the 50-1000 ng/ml concentration ranges, respectively. Identity of mevalonate in plasma and urine was confirmed by high-resolution mass spectrometry.
    Additional Material: 2 Tab.
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  • 92
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: Fast atom bombardment (FAB) mass spectrometry was used to characterize the pyochelin complexes of iron and zinc. Ferripyochelin readily forms the [M + H]+ and [2M + H]+ ions in a glycerol matrix. The molecular ion was 2.5 fold more abundant when acetic acid was present. Iron was shown to be present in the iron-pyochelin complex as the ferric ion by wet chemical methods. Differentiation between ferric and ferrous ions by FAB was not successful owing to the FAB reducing environment. Ferric ions were reduced by FAB to ferrous ions in the following model salts - ferric chloride, ferric nitrate and ferric sulfate - when dissolved in either glycerol, thioglycerol or the magic bullet matrix. Ferric and ferrous chloride salts gave virtually identical spectra with a glycerol/acetic acid matrix. Zinc-pyochelin readily forms the [M + H]+ and [2M + H]+ ions, but only when acetic acid was present in the glycerol matrix. The FAB mass spectrum of zinc-pyochelin exhibited various fragmentations which can be used in structural analysis. Linked-scan at constant B/E and accurate mass data were used to characterize the fragmentations of the zinc-pyochelin complex. Metastable analysis allowed the fragmentation pathway of zinc-pyochelin to be determined.
    Additional Material: 7 Ill.
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  • 93
    Electronic Resource
    Electronic Resource
    Chichester : Wiley-Blackwell
    Biological Mass Spectrometry 18 (1989), S. 207-217 
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: The electron impact (EI) mass spectra of 2,6-di-tert-butyl-4-methylphenol (BHT) and certain of its alteration products are described in detail. Accurate mass measurements confirm the elemental compositions of important fragment ions in the EI spectra. Collisionally activated mass spectra are also used to study fragmentation and suggest common ion structures. The reference spectra provide the basis for identifying various alteration products of BHT by capillary gas chromatography/mass spectrometry (GC/MS) without the necessity of isolating individual components. Application of GC/MS is made to three studies: (i) pyrolysis of hydroperoxy-BHT as a potential pathway to alteration products in food; (ii) GC/MS pyrolysis of hydroperoxy-BHT as a model study; and (iii) alteration of BHT in ethanol/water as a food-simulating solvent.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
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  • 94
    Electronic Resource
    Electronic Resource
    Chichester : Wiley-Blackwell
    Biological Mass Spectrometry 18 (1989), S. 363-372 
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: The MI and CA spectra of selected positive and negative ions generated by fast atom bombardment (FAB) of the stereoisomeric aldohexoses can be used to differentiate among these isomers, without prior derivatization or addition of other substances. The main fragmentation routes were traced and compared to those of some 2- and 6-deoxyaldohexoses, which appeared to be suitable model compounds. In both positive and negative ion FAB mass spectrometry, the OH group at the C-2 position plays an important role in the fragmentation reactions of the pseudomolecular [M + glycerol + H]+ and [M - H]- ions.
    Additional Material: 1 Ill.
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  • 95
    Electronic Resource
    Electronic Resource
    Chichester : Wiley-Blackwell
    Biological Mass Spectrometry 18 (1989), S. 387-393 
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: Chicken intestinal calbindin-D28k is an intracellular protein which is believed to have a fundamental role in vitamin D-mediated transport of calcium. A mapping approach based on 252Cf plasma desorption mass spectrometry (PD mapping) was used to screen the DNA-deduced sequence of calbindin-D28k for sequence changes and posttranslational modifications. In the PD mapping experiment, purified calbindin-D28k was cleaved with cyanogen bromide and the resulting peptides were subjected to PD mass spectrometric analysis either as a mixture or as high-performance liquid chromatography isolated fractions. The DNA-derived primary structure of calbindin-D28k was confirmed by rapid PD mass spectral identification of the CNBr peptide fragments, and the nature of the N-terminal blocking group was readily determined to be an acetyl group. The relatively non-destructive nature of the PD mass spectrometric analysis allowed the mapping of the N-terminal peptide through an additional in situ V8 protease enzymatic reaction.
    Additional Material: 6 Ill.
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  • 96
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: A current limitation in the use of fast atom bombardment (FAB) mass spectrometric mapping of peptide mixtures, derived from enzymic digestion of proteins, is that most of the hydrophilic peptides are not observed. However, it has been demonstrated from previous work that esterification of the peptide mixture results in the detection of almost all peptides in FAB mass spectrometry. This strategy of FAB mapping was applied to the protein actinidin, isolated from an Italian variety of Actinidia chinensis. Two of the 12 tryptic peptides in FAB mass spectrometry did not exhibit molecular ions predicted from the known sequence of actinidin isolated from the New Zealand variety of A. chinensis. The two peptides were isolated by high-performance liquid chromatography, subjected to Staphylococcus aureus V8 protease digestion and sequenced by gas-phase microsequencing. Nine changes in amino acid composition were detected using the rapid and powerful combination of FAB mass spectrometric mapping and gas-phase microsequencing.
    Additional Material: 1 Ill.
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  • 97
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: The metabolism of 17α-methyl-17β-hydroxy-2-oxa-5α-androstan-3-one (oxandrolone) in man has been investigated by gas chromatography/mass spectrometry. After oral administration of a 10 mg dose to man, five metabolites were detected in the free fraction of the urinary samples. Oxandrolone, the major compound excreted in urine, was detected within 72 h after administration. During this period 35.8 and 8.4% of the administered dose was excreted as unchanged oxandrolone and 17-epioxandrolone, respectively. In addition, minute amounts of 16α- and 16β- hydroxyoxandrolone and a δ-hydroxy acid resulting from the hydrolysis of the lactone group of oxandrolone were detected in the urine samples 8-60 h after administration. Furthermore, the susceptibility of oxandrolone to hydrolysis was investigated under several pH conditions. Extraction and fractionation of steroidal metabolites was achieved by using C18 and silica Sep Pak™ chromatography. The mass spectra of the metabolites are presented and major fragmentation pathways discussed.
    Additional Material: 10 Ill.
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  • 98
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: An increasing amount of evidence is accumulating to support the proposal that steroidogenesis can occur by a sesterterpene pathway as well as the cholesterol pathway. Key intermediates on the sesterterpene pathway are 23,24-dinor-5-cholen-3β-ol (guneribol) and some of its metabolites, e.g. 23,24-dinor-4-cholen-3-one (guneribone). It has been reported that these intermediates are biosynthesized and converted to steroid hormones by a range of endocrine tissues in vitro. Monitoring the pentafluorobenzyloxime derivatives by negative ion chemical ionization mass spectrometry in the electron capture mode provided evidence for the presence of guneribone in extracts of bovine testicular and human adrenal tumour tissue. Complementary evidence was obtained from gas chromatographic/tandem mass spectrometric data generated on a triple-quadrupole instrument by monitoring daughter ions (in the multiple ion detection, MID mode) of the molecular anion of derivatized guneribone in both standards and tissue extracts. The present findings that sesterterpene pathway intermediates are present as endogenous compounds in tissue extracts, together with the previously reported radiochemical data, give further support to the sesterterpene pathway hypothesis.
    Additional Material: 5 Ill.
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  • 99
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: A gas chromatographic/mass spectrometric analytical procedure is described to determine glycols in plasma as cyclic butyl boronate esters. The method, involving a pre-deproteinization step, required only 0.25 ml of plasma and a short time (20 min) to react with the derivatizing agent (butyl boronic acid). The gas chromatographic separation on a CP Sil 8 CB silica capillary column coupled to a mass detector assured a complete identification of the compounds. The analytical recoveries (〉95%) with low coefficient of variation (4-11%) assured the feasibility of the method over a concentration range from 5 to 1000 μg ml-1 for each glycol. The lower detection limits, namely 1-5 μg ml-1, confirmed the sensitivity of the method.
    Additional Material: 3 Ill.
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  • 100
    Electronic Resource
    Electronic Resource
    Chichester : Wiley-Blackwell
    Biological Mass Spectrometry 18 (1989), S. 603-612 
    ISSN: 0887-6134
    Keywords: Chemistry ; Analytical Chemistry and Spectroscopy
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology
    Notes: The synthesis of tetradeuterated 10-methylundecanoic acid, dideuterated tetracos-2-enoic acid, (1-14C)pristanic acid and (1-14C)tetracos-2-enoic acid using the Favorsky rearrangement are described. They will be used for studies of long-chain fatty acid metabolism in patients. The positions of isotope labelling were determined by mass spectrometry.
    Additional Material: 12 Ill.
    Type of Medium: Electronic Resource
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