Volatile monocarbonyl compounds of carrot roots at various stages of maturity

Summary Volatile carbonyl compounds of carrot varietyFeonia Hunderup S-64 at various stages of maturity were analysed as their 2,4-DNPH's by a combined TLC-GLC-MS method. Twenty-three different carbonyl compounds were identified, of which undecanal, buten-2-al, methylbutenal, pentan-2-one, 6-methyl-5-hepten-2-one and 5-methylfurfural have not previously been found in carrot. During maturation the content of acetaldehyde and acetone increased significantly, while that of methylbutenal decreased.

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Die leichtflüchtigen Monocarbonylverbindungen der Karotten in verschiedenen Stufen der Reife

Zusammenfassung Die leichtflüchtigen Carbonylverbindungen der Karotten Varietät Feonia Hunderup S-64 wurden in verschiedenen Stufen der Reife als 2,4-DNPH mit der kombinierten DC-GC-MS-Methode analysiert. Dreiundzwanzig verschiedene Carbonylverbindungen wurden nachgewiesen. Von diesen sind Undecanal, 2-Butenal, Methylbutenal, 2-Pentanon, 6-Methyl-5-hepten-2-on und 5-Methylfurfural noch nicht in Karotten aufgefunden worden. Während des Reifens nahm der Gehalt an Acetaldehyd und Aceton bedeutend zu. Der Gehalt an Methylbutenal verminderte sich.

     

Identification of residues 99, 220, and 221 of human cytochrome P450 2C19 as key determinants of omeprazole activity

Human P450 2C19 is selective for 4'-hydroxylation of S-mephenytoin and 5-hydroxylation of omeprazole, while the structurally homologous P450 2C9 has low activity toward these substrates. To identify the critical amino acids that determine the specificity of human amino acids that determine the specificity of human P450 2C19, we constructed chimeras of p450 2C9 replacing various proposed substrate binding sites (SRS) with those of P450 2C19 and then replaced individual residues of P450 2C19 and then replaced individual residues of P450 2C9 by site-directed mutagenesis. The 339 NH2-terminal amino acid residues (SRS-1-SRS-4) and amino acids 160-383 (SRS-2-SRS-5) of P450 2C19 conferred omeprazole 5-hydroxylase activity to P450 2C9. In contract, the COOH terminus of P450 2C19 (residues 340-490 including SRS-5 and SRS-6), residues 228-339 (SRS-3 and SRS-4) and residues 292-383 (part of SRS-4 and SRS-5) conferred only modest increases in activity. A single mutation Ile99 --> His increased omeprazole 5-hydroxylase to approximately 51% of that of P450 2C19. A chimera spanning residues 160-227 of P450 2C19 also exhibited omeprazole 5-hydroxylase activity which was dramatically enhanced by the mutation Ile99 --> His. A combination of two mutations, Ile99 --> His and Ser200 --> Pro, converted P450 2C9 to an enzyme with a turnover number of omeprazole 5-hyrdroxylation, which resembled that of P450 /c19. Mutation of Pro221 --> Thr enhanced this activity. Residue 99 is within SRS-1, but amino acids 220 and 221 are in the F-G loop and outside any known SRS. Mutation of these three amino acids did not confer significant S-mephenytoin 4'-hydroxylase activity to P450 2C9, although chimeras containing SRS-1-SRS-4 and SRS-2-SRS-5 of P450 2C19 exhibited activity toward this substrate. Our results thus indicate that amino acids 99, 220, and 221 are key residues that determine the specificity of P450 2C19 for omeprazole.     

INCUBATION OF BARLEY KERNEL SECTIONS WITH PURIFIED CELL WALL DEGRADING ENZYMES

The effects of purified Trichoderma reesei endo-xylanase, XYN II and endoglucanases, EG I and EG II on the cell walls of embedded barley cross sections were studied. The cell walls were stained with Calcofluor which is specific for (1–3)(1–4)-β-D-glucans (mixed-linked β-glucans) in histochemical applications. Whereas endosperm cell walls remained highly fluorescent after incubation with buffer, the aleurone cell walls were only weakly fluorescent. Both EG I and EG II were effective in degrading the outer endosperm cell walls, but had little or no effect on the inner endosperm and aleurone cell walls. Corresponding sections incubated with XYN II showed highly intensive fluorescence of aleurone cell walls suggesting that the surface of alaurone cell walls contains a high level of xylan which is removed by xylanase treatment exposing a layer rich in mixed linked β-glucans. This indicates that arabinoxylans and β-glucans in the aleurone cell walls of barley exist as separate, macroscopic layers. Xylanase did not have any Influence on the microstructure of endosperm cell walls which are rich in mixed linked β-glucans. However, incubation with EG II after xylanase treatment caused the aleurone cell walls to become very thin.     

Stability of DNA Origami Nanostructures in Physiological Media: The Role of Molecular Interactions

Programmable, custom-shaped, and nanometer-precise DNA origami nanostructures have rapidly emerged as prospective and versatile tools in bionanotechnology and biomedicine. Despite tremendous progress in their utilization in these fields, essential questions related to their structural stability under physiological conditions remain unanswered. Here, DNA origami stability is explored by strictly focusing on distinct molecular-level interactions. In this regard, the fundamental stabilizing and destabilizing ionic interactions as well as interactions involving various enzymes and other proteins are discussed, and their role in maintaining, modulating, or decreasing the structural integrity and colloidal stability of DNA origami nanostructures is summarized. Additionally, specific issues demanding further investigation are identified. This review – through its specific viewpoint – may serve as a primer for designing new, stable DNA objects and for adapting their use in applications dealing with physiological media.     

Analysis of butter fat triacylglycerols by supercritical fluid chromatography/electron impact mass spectrometry

Triacylglycerols (TAGs) from butter fat isolated by solvent extraction were analyzed by use of a capillary column supercritical fluid chromatograph (SFC) combined with a flame ionization detector or a double focusing mass spectrometer. The chromatographic separation was achieved by using a dimethyldiphenylpolysiloxane phase (DB-5) to bundle up the TAGs with the same carbon number. The ratio of TAGs with varying degree of unsaturation in each SFC peak was determined by using the selected ion monitoring of the molecular ions with electron impact mode. The discrimination between the fatty acids at the position sn-2 and positions sn-1/3 in a triacylglycerol molecule was demonstrated by monitoring the ions [M - RCO2CH2]+ from reference compounds.     

Effect of enzymatic interesterification on the melting point of tallow-rapeseed oil (LEAR) mixture

To reduce the melting point of a tallow-rapeseed oil mixture, the triglyceride composition of the mixture was altered by enzymatic interesterification in a solvent-free system. The interesterification and hydrolysis were followed by melting point profiles and by free fatty acid determinations. The degree of hydrolysis was linearly related to the initial water content of the reaction mixture. The rate of the interesterification reaction was influenced by the amount of enzyme but not much by temperature between 50 and 70°C. The melting point reduction achieved by interesterification depended on the mass fractions of the substrates: the lower the mass fraction of tallow, the larger the reduction of the melting point.     

Lipase-catalyzed transesterification of rapeseed oil and 2-ethyl-1-hexanol

Lipase-catalyzed transesterification (alcoholysis) of lowerucic acid rapeseed oil and 2-ethyl-1-hexanol without an additional organic solvent was studied in stirred batch reactors. Of a number of commercially available enzymes investigated, the best results were obtained with aCandida rugosa lipase. The optimal transesterification conditions were an oil/alcohol molar ratio of 1∶2.8, a minimum of 1.0% (w/w) added water, and with a temperature of 37–55°C. Under the optimal conditions, a nearly complete conversion was obtained in one hour with 14.6% (w/w) lipase, whereas 0.3% (w/w) lipase required 10 h for similar results. The enzyme was inactivated at 60°C.