Design and total synthesis of (-)-codonopsinine, (-)-codonopsine and codonopsinine analogues by O-(2-oxopyrrolidin-5-yl)trichloroacetimidate as amidoalkylating agent with improved antimicrobial activity via solid lipid nanoparticle formulations

A general strategy towards total synthesis of (-)-codonopsinine, (-)-codonopsine and codonopsinine analogues has been developed from (D)-tartaric acid via the intermediate (3S,4R)-1-methyl-2-oxo-5-(2,2,2-trichloroacetamido)pyrrolidinediacetate (7). α-amidoalkylation studies of 7 with electron rich benzene derivative 8a-g as C-nucleophiles afforded (aryl derivatives) 9a-g. The target compounds 1, 2 and 13c-g were readily obtained from 10a-g via Grignard addition to the homochiral lactam which was produced by deoxygenation using Lewis-acid followed by deacetylation. The synthesized compounds were loaded onto solid lipid nanoparticle formulations (SLNs) prepared by hot emulsification-ultrasonication technique using Compritol as solid lipid and Pluronic f68 as surfactant. SLNs were fully evaluated and the permeation of synthesized compound from SLNs was assayed against non-formulated compounds through dialysis membranes using Franz cell. The data indicated good physical characteristics of the prepared SLNs, sustaining of release profiles and significant improvement of permeation ability when compared to the non-formulated compounds. The antibacterial and antifungal activities of 1, 2 and 13c-g were determined by disc diffusion and microbroth dilution method to determine the minimum inhibitory concentrations (MIC) against seven microorganisms (Staphyloccus aureus, Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Pseudomonas aeruginosa, Acinetobacter baumannii and Candida albicans). The most active compounds against the Gram positive S. aureus were 1, 13C, 13d, and 13g. Also, 13c, 13d, and 13e had antibacterial activity but not 13f against some Gram negative organisms (E. coli, and P. mirabilis). MIC concentrations against P. aeruginosa, and K. pneumoniae were?≥512?μg/ml, while that against A. baumannii was?≥128?μg/ml except for nanoformulae of 13e and 13f that were 16 and 64?μg/ml, respectively. No antifungal activity against Candida albicans was recorded for all compounds and their nanoformulae (MIC?>?1024?μg/ml). SLNs were found to decrease the MIC values for some of the compounds with no effect on the antifungal activity. In conclusion, we demonstrated a novel, straight-forward and economical procedure for the total synthesis of (-)-codonopsinine 1, (-)-codonopsine 2 and codonopsinine analogues 13c-g from simple and commercially available starting materials; d-tartaric acid; with antimicrobial activities against Gram positive and Gram-negative organisms that were improved by SLNs formulations.     

Citric acid production from sugar cane molasses by 2-deoxyglucose-resistant mutant strain ofAspergillus niger

Citric acid production from sugar cane molasses byAspergillus niger NIAB 280 was studied in a batch cultivation process. A maximum of 90 g/L total sugar was utilized in citric acid production medium. From the parental strainA. niger, mutant strains showing resistance to 2-deoxyglucose in Vogal's medium containing molasses as a carbon source were induced by γ-irradiation. Among the new series of mutant strains, strain RP7 produced 120 g/L while the parental strain produced 80 g/L citric acid (1.5-fold improvement) from 150 g/L of molasses sugars. The period of citric acid production was shortened from 10 d for the wild-type strain to 6–7 d for the mutant strain. The efficiency of substrate uptake rate with respect to total volume substrate consumption rate,Q _s (g per L per h) and specific substrate consumption rate,q _s (g substrate per g cells per h) revealed that the mutant grew faster than its parent. This indicated that the selected mutant is insensitive to catabolite repression by higher concentrations of sugars for citric acid production. With respect to the product yield coefficient (Y _p/x), volume productivity (Q _p) and specific product yields (q _p), the mutant strain is significantly (p≤0.05) improved over the parental strain.     

A tyrosine-sulfated CCR5-mimetic peptide promotes conformational transitions in the HIV-1 envelope glycoprotein

The HIV-1 envelope glycoprotein is a trimeric complex of heterodimers composed of a surface glycoprotein, gp120, and a transmembrane component, gp41. The association of this complex with CD4 stabilizes the coreceptor-binding site of gp120 and promotes the exposure of the gp41 helical region 1 (HR1). Here, we show that a 15-amino-acid peptide mimetic of the HIV-1 coreceptor CCR5 fused to a dimeric antibody Fc domain (CCR5mim-Ig) bound two gp120 molecules per envelope glycoprotein complex and by itself promoted HR1 exposure. CCR5mim-Ig also stabilized the association of a CD4-mimetic peptide with the envelope glycoprotein. A fusion of the CD4- and CCR5-mimetic peptides, DM1, bound gp120 and neutralized R5, R5X4, and X4 HIV-1 isolates comparably to CD4, and they did so markedly more efficiently than either peptide alone. Our data indicate that the potency of DM1-Ig derives from its avidity for the HIV-1 envelope glycoprotein trimer and from the bidirectional induction of its receptor-mimetic components. DM1 has significant advantages over other inhibitors that target both coreceptor and CD4-binding sites, and it may serve as a lead for a new class of HIV-1 inhibitor peptides.     

Performance of down-flow hanging sponge (DHS) reactor coupled with up-flow anaerobic sludge blanket (UASB) reactor for treatment of onion dehydration wastewater

In this study, a promising system consisting of up-flow anaerobic sludge blanket (UASB) reactor followed by down-flow hanging sponge (DHS) reactor was investigated for onion dehydration wastewater treatment. Laboratory experiments were conducted at two different phases, i.e., phase (1) at overall hydraulic retention time (HRT) of 11 h (UASB reactor: 6 h and DHS reactor: 5 h) and phase (2) at overall HRT of 9.4 h (UASB reactor: 5.2 h and DHS reactor: 4.2 h). Long-term operation results of the proposed system showed that its overall TCOD, TBOD, TSS, TKN and NH₄N removal efficiencies were 92 ± 5, 95 ± 2, 95 ± 2, 72 ± 6 and 99 ± 1.3%, respectively (phase 1). Corresponding values for the 2nd phase were 85.4 ± 5, 86 ± 3, 87 ± 6, 65 ± 8 and 95 ± 2.8%. Based on the available results, the proposed system could be more viable option for treatment of wastewater generated from onion dehydration industry in regions with tropical or sub-tropical climates and with stringent discharge standards.     

Differences in DNA double strand breaks repair in male germ cell types: lessons learned from a differential expression of Mdc1 and 53BP1

In male germ cells the repair of DNA double strand breaks (DSBs) differs from that described for somatic cell lines. Irradiation induced immunofluorescent foci (IRIF's) signifying a double strand DNA breaks, were followed in spermatogenic cells up to 16 h after the insult. Foci were characterised for Mdc1, 53BP1 and Rad51 that always were expressed in conjecture with gamma-H2AX. Subsequent spermatogenic cell types were found to have different repair proteins. In early germ cells up to the start of meiotic prophase, i.e. in spermatogonia and preleptotene spermatocytes, 53BP1 and Rad51 are available but no Mdc1 is expressed in these cells before and after irradiation. The latter might explain the radiosensitivity of spermatogonia. Spermatocytes from shortly after premeiotic S-phase till pachytene in epithelial stage IV/V express Mdc1 and Rad51 but no 53BP1 which has no role in recombination involved repair during the early meiotic prophase. Mdc1 is required during this period as in Mdc1 deficient mice all spermatocytes enter apoptosis in epithelial stage IV when they should start mid-pachytene phase of the meiotic prophase. From stage IV mid pachytene spermatocytes to round spermatids, Mdc1 and 53BP1 are expressed while Rad51 is no longer expressed in the haploid round spermatids. Quantifying foci numbers of gamma-H2AX, Mdc1 and 53BP1 at various time points after irradiation revealed a 70% reduction after 16 h in pachytene and diplotene spermatocytes and round spermatids. Although the DSB repair efficiency is higher then in spermatogonia where only a 40% reduction was found, it still does not compare to somatic cell lines where a 70% reduction occurs in 2 h. Taken together, DNA DSBs repair proteins differ for the various types of spermatogenic cells, no germ cell type possessing the complete set. This likely leads to a compromised efficiency relative to somatic cell lines. From the evolutionary point of view it may be an advantage when germ cells die from DNA damage rather than risk the acquisition of transmittable errors made during the repair process.     

Immunological responses and viral modulatory effects of vaccination with recombinant modified vaccinia virus Ankara (rMVA) expressing structural and regulatory transgenes of simian immunodeficiency virus (SIVmac32H/J5M)

Background The immunogenicity and protective efficacy of recombinant modified vaccinia virus Ankara (rMVA) vectors expressing structural (gag/pol, env) and regulatory (tat, rev, nef) genes of SIVmac251/32H‐J5 (rMVA‐J5) were assessed. Methods Immunization with rMVA constructs (2.5 × 10⁷ IU) 32, 20 and 8 weeks pre‐challenge was compared with 32 and 20 weeks but with a final boost 8 weeks pre‐challenge with 2 × 10⁶ fixed‐inactivated HSC‐F4 cells infected with SIVmac32H. Controls received rMVA vectors expressing an irrelevant transgene or were naïve challenge controls. All received 10 MID₅₀ SIVmac32H/J5 intravenously. Results Vaccinates immunized with rMVA‐J5 exhibited significant, albeit transient, control of peak primary viraemia despite inconsistent and variable immune responses elicted by vaccination. Humoral and cellular responses to Env were most consistent, with lower responses to Nef, Rev and Tat. Increasing titres of anti‐vaccinia neutralizing antibodies reflected the number and dose of rMVA inoculations. Conclusions Improved combinations of viral vectors are required to elicit appropriate immune responses to control viral replication.     

Placenta growth factor-1 exerts time-dependent stabilization of adherens junctions following VEGF-induced vascular permeability

Increased vascular permeability is an early event characteristic of tissue ischemia and angiogenesis. Although VEGF family members are potent promoters of endothelial permeability the role of placental growth factor (PlGF) is hotly debated. Here we investigated PlGF isoforms 1 and 2 and present in vitro and in vivo evidence that PlGF-1, but not PlGF-2, can inhibit VEGF-induced permeability but only during a critical window post-VEGF exposure. PlGF-1 promotes VE-cadherin expression via the trans-activating Sp1 and Sp3 interaction with the VE-cadherin promoter and subsequently stabilizes transendothelial junctions, but only after activation of endothelial cells by VEGF. PlGF-1 regulates vascular permeability associated with the rapid localization of VE-cadherin to the plasma membrane and dephosphorylation of tyrosine residues that precedes changes observed in claudin 5 tyrosine phosphorylation and membrane localization. The critical window during which PlGF-1 exerts its effect on VEGF-induced permeability highlights the importance of the translational significance of this work in that PLGF-1 likely serves as an endogenous anti-permeability factor whose effectiveness is limited to a precise time point following vascular injury. Clinical approaches that would pattern nature's approach would thus limit treatments to precise intervals following injury and bring attention to use of agents only during therapeutic windows.     

Chromium and Manganese Levels in Biological Samples of Normal and Night Blindness Children of Age Groups (3–7) and (8–12) Years

This study was designed to compare the levels of chromium (Cr) and manganese (Mn) in scalp hair, blood, and urine of night blindness in children age ranged (3–7) and (8–12) years of both genders, comparing them to sex- and age-matched controls. A microwave-assisted wet acid digestion procedure, was developed as a sample pretreatment, for the determination of Cr and Mn in biological samples of night blindness children. The proposed method was validated by using conventional wet digestion and certified reference samples of hair, blood and urine. The digests of all biological samples were analyzed for Cr and Mn by electrothermal atomic absorption spectrometry. The results indicated significantly higher levels of Cr, whilst low level of Mn in the biological samples (blood and scalp hair) of male and female night blindness children, compared with control subjects of both genders. These data present guidance to clinicians and other professional investigating deficiency of Mn and excessive level of Cr in biological samples (scalp hair and blood) of night blindness children.