Molecules2013, 18(12), 15501-15518; doi:10.3390/molecules181215501 (doi registration under processing) - published online 12 December 2013 Show/Hide Abstract
Abstract: The organophosphorous hydrolase (PTE) from Brevundimonas diminuta is capable of degrading extremely toxic organophosphorous compounds with a high catalytic turnover and broad substrate specificity. Although the natural substrate for PTE is unknown, its loop remodeling (loop 7-2/H254R) led to the emergence of a homoserine lactonase (HSL) activity that is undetectable in PTE (kcat/km values of up to 2 × 104), with only a minor decrease in PTE paraoxonase activity. In this study, homology modeling and molecular dynamics simulations have been undertaken seeking to explain the reason for the substrate specificity for the wild-type and the loop 7-2/H254R variant. The cavity volume estimated results showed that the active pocket of the variant was almost two fold larger than that of the wild-type (WT) enzyme. pKa calculations for the enzyme (the WT and the variant) showed a significant pKa shift from WT standard values (ΔpKa = 3.5 units) for the His254residue (in the Arg254 variant). Molecular dynamics simulations indicated that the displacement of loops 6 and 7 over the active site in loop 7-2/H254R variant is useful for N-acyl-L-homoserine lactone (C4-HSL) with a large aliphatic chain to site in the channels easily. Thence the expanding of the active pocket is beneficial to C4-HSL binding and has a little effect on paraoxon binding. Our results provide a new theoretical contribution of loop remodeling to the rapid divergence of new enzyme functions.
Molecules2013, 18(12), 15490-15500; doi:10.3390/molecules181215490 (doi registration under processing) - published online 12 December 2013 Show/Hide Abstract
Abstract: pH-Zone refining counter-current chromatography was successfully applied to the preparative isolation and purification of six alkaloids from the ethanol extracts of Uncaria macrophylla Wall. Because of the low content of alkaloids (about 0.2%, w/w) in U. macrophylla Wall, the target compounds were enriched by pH-zone refining counter-current chromatography using a two-phase solvent system composed of petroleum ether–ethyl acetate–isopropanol–water (2:6:3:9, v/v), adding 10 mM triethylamine in organic stationary phase and 5 mM hydrochloric acid in aqueous mobile phase. Then pH-zone refining counter-current chromatography using the other two-phase solvent system was used for final purification. Six target compounds were finally isolated and purified by following two-phase solvent system composed of methyl tert-butyl ether (MTBE)–acetonitrile–water (4:0.5:5, v/v), adding triethylamine (TEA) (10 mM) to the organic phase and HCl (5 mM) to aqueous mobile phase. The separation of 2.8 g enriched total alkaloids yielded 36 mg hirsutine, 48 mg hirsuteine, 82 mg uncarine C, 73 mg uncarine E, 163 mg rhynchophylline, and 149 mg corynoxeine, all with purities above 96% as verified by HPLC Their structures were identified by electrospray ionization-mass spectrometry (ESI-MS) and 1H-NMR spectroscopy.
Molecules2013, 18(12), 15474-15489; doi:10.3390/molecules181215474 (doi registration under processing) - published online 12 December 2013 Show/Hide Abstract
Abstract: Collagen hydrolysate (CH) has been reported to exhibit a positive effect on bone. In the present study, the in vitro effects of CH (<3 kDa) were examined and the in vivo experiments confirmed the positive effects of CH in ovariectomized (OVX) rats. Bone mineral density (BMD) was examined by DXA analysis. Scanning electron microscopic analysis and quantitative 3D-color backscattered electrons imaging analysis were performed on the lumbar vertebrae. CH increased osteoblastic cell proliferation and alkaline phosphatase activity in a dose-dependent manner. Collagen synthesis and collagen, type1, alpha1 (COL1A1)gene expression were also increased by CH treatment. Furthermore, CH-induced COL1A1 gene expression was completely abolished by extracellular signal-regulated kinase (ERK) inhibitor, suggesting the involvement of ERK/MAPK signaling for transcriptional effects on COL1A1 expression. OVX rats supplemented with CH showed osteoprotective effects as the BMD levels were increased compared with control. Moreover, CH prevented the trabecular bone loss induced by OVX and improved the microarchitecture of lumbar vertebrae. CH administration dose-dependently reduced the serum procollagen type I N-terminal propeptide level, which was elevated by OVX. The present study suggests that CH isolated in this study is a promising alternative to current therapeutic agents for the management of osteoporosis.
Molecules2013, 18(12), 15464-15473; doi:10.3390/molecules181215464 (doi registration under processing) - published online 12 December 2013 Show/Hide Abstract
Abstract: The variation of six biogenic amines (BAs) and total viable count (TVC) in common carp (Cyprinus carpio) stored in ice with 0, 4 and 8 h delay before icing was evaluated in a period of 4 days. Delayed icing led to significant (p < 0.05) increases in TVC throughout the period of storage and showed a good correlation with BAs content. The obtained data showed that putrescine and cadaverine were predominant in all samples and it was indicated that they could be proper indicators to determine the carp quality. Spermidine and spermine increased slightly toward the end of storage and the levels of dangerous BAs (histamine and tyramine) were under the limit over the period. As a result, it is indicated that delaying time affects on formation of BAs and the effect in samples with 8 h delay was significantly (p < 0.05) more than those with 0 and 4 h delay.
Molecules2013, 18(12), 15448-15463; doi:10.3390/molecules181215448 (doi registration under processing) - published online 12 December 2013 Show/Hide Abstract
Abstract: Cancer is commonly diagnosed in dogs over the age of 10 and is a leading cause of death due to the lack of effective drugs. Flavonoids possess antioxidant, anti-inflammatory and anticarcinogenic properties and have been studied as chemopreventive agents in human cancer therapy. However, the literature on dogs is sparse. In this study, we analyzed the effect of nine flavonoids on cell viability, DNA damage and topoisomerase IIa/IIb gene expression in a canine tumor cell line (DH82). Apigenin, luteolin, trans-chalcone and 4-methoxychalcone showed the highest degree of cytotoxicity in the absence of considerable DNA damage, whereas genistein exhibited low cytotoxicity but induced a high level of DNA damage. These five flavonoids inhibited topoisomerase IIa and IIb gene expression to variable extents and with variable specificity. Genistein exerted a lower inhibitory effect on the two topoisomerases than luteolin and apigenin. trans-Chalcone and 4-methoxychalcone exerted greater inhibition of topoisomerase IIa expression than topoisomerase IIb. The differences in the effects between genistein and luteolin and apigenin might be explained by the position of ring B, whereas the more specific effect of chalcones on topoisomerase IIa might be due to their open chain structure.
Abstract: Solubilization of single walled carbon nanotubes (SWNTs) in aqueous gel phases formed by reversible, G-quadruplex self-assembly of guanosine monophosphate (GMP) alone or with guanosine (Guo) is described. Unlike other media and methods for aqueous solubilization of SWNTs, the guanosine gels (“G-gels”) are found to readily disperse high (>mg/mL) concentrations of individual rather than bundled SWNTs. SWNT dispersions in GMP alone precipitate in several hours and re-form upon shaking; however, dispersions in the binary GMP/Guo gels are indefinitely stable. Increasing GMP or KCl concentration in the binary gels increased the relative abundance of large diameter and semi-conducting SWNTs. Different gel compositions also displayed different selectivities toward SWNTs of different chiralities. These results indicate a strong connection between the self-assembled G-gels and the dimensions and structures of the SWNTs that they solubilize.