High-performance liquid chromatography is a promising alternative for determining the G+ C content of bacterial deoxyribonucleic acid (DNA). The method which we evaluated involves enzymatic degradation of the DNA to nucleosides by P1 nuclease and bovine intestinal mucosa alkaline phosphatase, separation of the nucleosides by high-performance liquid chromatography, and calculation of the G + C content from the apparent ratios of deoxyguanosine and thymidine. Because the nucleosides are released from the DNA at different rates, incomplete degradation produces large errors in the apparent G+ C content. For partially purified DNA, salts are a major source of interference in degradation. However, when the salts are carefully removed, the preparation and degradation of DNA contribute little error to the determination of G+C cmtent. This method also requires careful selection of the chromatographic conditions to ensure separation of the major nucleosides from the nucleosides of modified bases and precise control of the flow rates. Both of these conditions are achievable with standard equipment and C18 reversed-phase columns. Then the method is precise, and the relative standard deviations of replicate measurements are close to 0.1%. It is also rapid, and a single measurement requires about 15 min. It requires small amounts of sample, and the G+C content can be determined from DNA isolated from a single bacterial colony. It is not affected by contamination with ribonucleic acid. Because this method yields a direct measurement, it may also be more accurate than indirect methods, such as the buoyant density and thermal denaturation methods. In addition, for highly purified DNA, the extent of modification can be determined.The measurement of G + C content is one of the most common measurements performed on bacteria. Its greatest values are probably its usefulness as a taxonomic marker and its usefulness in distinguishing phenotypically similar microorganisms. G + C content is also a fundamental property of cellular deoxyribonucleic acid (DNA) and is correlated with the amino acid composition of proteins, codon usage in messenger ribonucleic acid, auxotrophy for specific bases, and other properties of general biological interest (1, 8, 9, 19,25,26,29,36). Therefore, interest in this parameter is not limited to microbial taxonomists.Although the G + C content of DNA has a precise analytical meaning, the techniques for determining this parameter are often indirect and lack a high degree of precision. For instance, the standard deviations of the most widely used methods, the buoyant density centrifugation and thermal denaturation methods, are usually about k1.0 and k0.4 mol% G+C, respectively (5,20,21,23,28,34). The accuracy of both of these methods is also affected by the purity of the DNA and the presence of modified bases (20, 28). The other methods which are occasionally used, the chemical modification and ultraviolet absorption methods, also present difficulties under certain experimental conditions (3, Recently, high-pe...

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Potent antifungal cyclic lipopeptides, burkholdines (Bk), were isolated from a culture of Burkholderia ambifaria 2.2N. Bk-1229 (1) and Bk-1097 (2) are octapeptides comprised of nonproteinogenic amino acids, including -hydroxytyrosine, -hydroxyasparagine, and a new fatty acyl amino acid. 1 and 2 are fungicidal against a panel of fungi with potencies 2-60-fold better than amphotericin B control.

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Bioassay-guided fractionation of the anti-inflammation fractions of the Red Sea sponges Scalarispongia aqabaensis and Callyspongia siphonella yielded two new sterols from chloroform fractions of methanol extracts, namely scalaristerol (5α,8α-dihydroxycholest-6-en-3β-ol) (1) from Scalarispongia aqabaensis, and callysterol (ergosta-5,11-dien-3β-ol) (2) from Callyspongia siphonella. Structure determination was based on extensive NMR studies and mass spectrometry. The antiinflammatory activity of compounds 1 and 2 was assessed using the rat-hind paw edema method and by study of their effect on the release of O2- and TXB2 from LPS-activated rat neonatal microglia.

A new latrunculin, oxalatrunculin B (3), was isolated from Red Sea sponge Negombata corticata. Extensive spectroscopic analysis revealed an unprecedented heterocycle in which the rare thiazolidinone ring found in latrunculins was oxidized with three additional oxygens. An actin polymerization inhibition assay agreed with MM-PBSA free energy calculations that 3 binds more weakly than latrunculin B to actin. Significant antifungal and anticancer activity of 3 was found, suggesting an alternate target in addition to actin for latrunculin bioactivity.Latrunculins A (1) and B (2) (Fig. 1) were first isolated from Negombata magnifica (Keller) (formerly Latrunculia magnifica), discovered from the Red Sea. 1 Among the characteristic features of these natural products is the presence of a macrocyclic lactone ring of 14 or 16 carbon atoms and a 2-thiazolidinone moiety (Figure 1). 2,3 In vitro experiments revealed that the latrunculins disrupt actin polymerization, 4 a possible target for cancer. 5A recent study has also reported antimigratory and antiangiogenic activity of latrunculins, adding to their possible utility in the control of cancer. 6 In addition to significant ichthyotoxic and cytotoxic properties, the latrunculins are also antiviral against herpes simplex type 1 virus (HSV-1). 7Several new latrunculins including natural, synthetic or semisynthetic analogs have been reported recently. 6,8 We report here the structure assignment and bioactivity of a unique new jkhalifa_99@yahoo.com, rjd@olemiss.edu, mthamann@olemiss.edu. As further evidence for this, combustion analysis on 3 showed 53.6% carbon, 6.51% hydrogen, 3.16% nitrogen, 28.6% oxygen and 6.34% sulfur. 13 C NMR indicated that the carbon skeleton of 3 consisted of 2 methyl carbons, 7 methylenes, 7 methines and 4 quaternary carbon atoms, which is identical to the carbon multiplicities of 2. The 13 C NMR chemical shifts for the 14-membered and tetrahydropyran rings were in close agreement with those of 2 revealing that additional oxygenation must occur at the heteroatoms. Significant differences were found in the chemical shifts of the thiazolidinone ring. The C-17 resonance shifted downfield from δ28.5 to δ49.9 indicating the introduction of a neighboring oxygen atom. C-16 shifted slightly upfield so the only possible position for oxidation was at the sulfur atom. C-18 also moved upfield from δ175.3 to δ157.3. The chemical shift of the carbonyl carbon of a 5-membered lactam ring will move upfield around 15ppm when the lactam nitrogen is hydroxylated. 9,10 It has also been shown that oxygenation of the sulfur atom adjacent to the carbonyl of a 5-membered thiazolidinone will move chemical shifts upfield by ~10 ppm. 11 This suggested that two of the three new oxygen atoms are at the S and the last is assigned as the hydroxyl group at N. Further evidence for this was the disappearance of the H-N at δ6.1. In addition the amide IR resonance in 2 is absent in 3 and a new resonance at 1756 cm −1 appeared, indicating the presence of a CONOH system. 11 The OH...

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A new, simple, sensitive, rapid, and accurate isocratic RP-HPLC method was developed and validated for simultaneous analysis of the principal antioxidants of Nigella sativa, i.e., thymoquinone (TQ), carvacrol (CR), and its isomer thymol (THY), in different phytopharmaceuticals. The mobile phase was water-methanol (40 + 60, v/v) at a flow rate of 1.5 mL/min. Quantification was achieved with UV detection at 254 nm, based on peak area. The method was validated for linearity, accuracy, precision, selectivity, and robustness. The proposed method is stability-indicating for determination of TQ in the presence of its degradants. The LOD and LOQ (microg/mL) were, respectively, 0.006 and 0.021 for TQ, 0.002 and 0.006 for CR, and 0.027 and 0.090 for THY. The mean recoveries measured at three concentrations were higher than 99%, with RSD <2%. This analytical method is suitable for quality control of the marker substances in this widely used natural protective and curative remedy.