Investigation of a green process for the polymerization of catechin

Flavonoids are polyphenolic secondary plant metabolites which possess antioxidant and anti-inflammatory properties. Besides, they have been shown to exhibit increased antioxidant properties in their polymerized form. Catechins are one of the attractive class of flavonoids which belong to the group of flavan-3-ols. Polymerization of catechins have been investigated in numerous studies indicating the requirement of certain amount of organic solvent to provide the solubility of the monomer. However, many research projects have been conducted recently to replace toxic organic contaminants of the processes with environmentally friendly solvents. In this aspect, deep eutectic solvents (DESs) that are regarded as “green solvents” have been studied extensively in various enzyme catalyzed reactions. In the present study, we focused on establishing a green pathway for laccase catalyzed polycatechin synthesis by replacing organic solvent content with DESs as green solvents. For this aim, various parameters were investigated, such as DES types and concentrations laccase amount and reaction time. Consequently, the highest molecular weight polycatechin was obtained using 5% (v/v) B–M, 125?U laccase in 1?hr of reaction time, at 30°C, as 4,354?±?678?g?mol^?1. Corresponding X/XO inhibitory activity and superoxide radical scavenging activities were achieved as, 59 and 50%, respectively.     

Desiccated Syntrichia ruralis shoots regenerate after 20 years in the herbarium

The longest record for a moss withstanding continuous desiccation is 19 years. This report demonstrates that a herbarium specimen of Syntrichia ruralis (Hedw.) F.Weber &; D.Mohr from southern Nevada, USA, has retained its viability for 20 years and 3 months. Fifty-eight shoots of a herbarium specimen collected in 1995 were cleaned and placed into culture using locally collected and sterilized sand. These shoots were kept hydrated and examined daily for 28 days for signs of regeneration. Five sets of three additional shoots from the herbarium specimen were assessed for chlorophyll fluorescence at intervals from 30?min to 8 days post-rehydration. About two-thirds of the shoots were viable: producing regenerative protonemata or shoots directly from the original shoots or leaves, with shoot apices not resuming growth despite most regeneration occurring towards the apex of shoots. Chlorophyll fluorescence measurements were similar to those of dead or severely compromised plant tissues over the first 48?h post-rehydration, with F_v/F_m levels <0.05. However, F_v/F_m levels rose to ～0.35 after 8 days as F_m values dropped, indicative of some viable tissues.     

Stability of Phosphorothioate Oligonucleotides in Aqueous Ammonia in Presence of Stainless Steel

Abstract

Antisense oligonucleotides as modulators of gene expression represent an exciting new drug technology. Oligodeoxyribonucleotide phosphorothioates are now among the most intensively investigated nuclease-resistant antisense analogs, as evidenced by a number of ongoing clinical trials against several disease targets. Structurally, these differ from natural oligonucleotides by the replacement of one of two nonbridging oxygen atoms by a sulfur atom at each internucleotide linkage. Among factors in the successful development of these complex molecules to support broad clinical trials have been advances made in automation, analysis and purification. The large scale synthesis of oligonucleotide phosphorothioates is presently carried out by initial formation of the internucleotide phosphite linkage using solid-phase phosphoramidite chemistry followed by sulfurization. Efficient synthesis of 20-mer oligophosphorothioates has been achieved on 0.15 mole scale with only 1.75-fold excess of amidite synthons. However, as the scale of synthesis increases to meet future market demands, issues related to fast and efficient synthesis, automation, scalability and product purification are also being investigated. One issue has been the protocol for final product deprotection. Since deprotection involves large quantities of saturated aqueous ammonium hydroxide, one might consider use of stainless steel reactors to withstand resulting vapor pressure at 55°C. A recent report,¹ however, discusses the instability of dimer phosphorothioates in aqueous ammonia in the presence of metal ions. As this is potentially an important issue for phosphorothioate oligonucleotide synthesis, we describe herein our findings regarding deprotection of a 20-mer oligodeoxyribonucleotide phosphorothioate with aqueous ammonia during process development studies.     

Use of Phenylacetyl Disulfide (PADS) in the Synthesis of Oligodeoxyribonucleotide Phosphorothioates

Abstract

Oligodeoxyribonucleotide phosphorothioates, where one of the nonbridging oxygen of the internucleotide phosphate is formally replaced by a sulfur atom, are the first class to undergo human clinical trials. Ongoing phosphorothioate clinical trials against several disease targets has necessitated manufacture of very large quantities of oligonucleotide active pharmaceutical ingredient (API). Clinical trial and future market demands have stimulated effort towards developing cost efficient large scale synthesis of these complex bio-molecules. This effort has culminated in the routine synthesis of 20-mer oligodeoxyribonucleotide phosphorothioates at 150 mmole scale using only 1.75-fold molar excess of amidites in less than 8 h total synthesis time.     

Pro-inflammatory cytokines from Kupffer cells downregulate hepatocyte expression of adrenomedullin binding protein-1

Polymicrobial sepsis is characterized by an early, hyperdynamic phase followed by a late hypodynamic phase. Adrenomedullin (AM), a vasodilatory peptide, inhibits this transition from the early phase to the late phase. Adrenomedullin binding protein-1 (AMBP-1) enhances AM-mediated activities. The decrease of AMBP-1 levels in late sepsis reduces the vascular response to AM and produces the hypodynamic phase. Studies have indicated that the administration of LPS downregulates AMBP-1 production in the liver. Since hepatocytes are the primary source of AMBP-1 biosynthesis in the liver, we employed a co-culture strategy using hepatocyte and Kupffer cells to determine whether LPS directly or by increasing pro-inflammatory cytokines from Kupffer cells downregulates AMBP-1 production. Hepatocytes and Kupffer cells isolated from rats were co-cultured and treated with LPS for 24 h. LPS significantly attenuated AMBP-1 protein expression in a dose-dependent manner. Since AMBP-1 is basically a secretory protein, cell supernatants from co-culture cells treated with LPS were examined for AMBP-1 protein levels. LPS treatment caused a dose related decrease in AMBP-1 protein secretion. Similarly, LPS treatment produced a significant decrease in AMBP-1 protein expression in hepatocytes and Kupffer cells cultured using transwell inserts. LPS had no direct effect on AMBP-1 levels in cultured hepatocytes or Kupffer cells alone. To confirm that the observed effects in co-culture were due to the cytokines released from Kupffer cells, hepatocytes were treated with IL-1beta or TNF-alpha for 24 h and AMBP-1 expression was examined. The results indicated that both cytokines significantly inhibited AMBP-1 protein levels. Thus, pro-inflammatory cytokines released from Kupffer cells are responsible for downregulation of AMBP-1.     

Ghrelin inhibits sympathetic nervous activity in sepsis

Our previous studies have shown that norepinephrine (NE) upregulates proinflammatory cytokines by activating alpha(2)-adrenoceptor. Therefore, modulation of the sympathetic nervous system represents a novel treatment for sepsis. We have also shown that a novel stomach-derived peptide, ghrelin, is downregulated in sepsis and that its intravenous administration decreases proinflammatory cytokines and mitigates organ injury. However, it remains unknown whether ghrelin inhibits sympathetic activity through central ghrelin receptors [i.e., growth hormone secretagogue receptor 1a (GHSR-la)] in sepsis. To study this, sepsis was induced in male rats by cecal ligation and puncture (CLP). Ghrelin was administered through intravenous or intracerebroventricular injection 30 min before CLP. Our results showed that intravenous administration of ghrelin significantly reduced the elevated NE and TNF-alpha levels at 2 h after CLP. NE administration partially blocked the inhibitory effect of ghrelin on TNF-alpha in sepsis. GHSR-la inhibition by the administration of a GHSR-la antagonist, [d-Arg(1),d-Phe(5), d-Trp(7,9),Leu(11)]substance P, significantly increased both NE and TNF-alpha levels even in normal animals. Markedly elevated circulating levels of NE 2 h after CLP were also significantly decreased by intracerebroventricular administration of ghrelin. Ghrelin's inhibitory effect on NE release was completely blocked by intracerebroventricular injection of the GHSR-1a antagonist or a neuropeptide Y (NPY)/Y(1) receptor antagonist. However, ghrelin's downregulatory effect on TNF-alpha release was only partially diminished by these agents. Thus ghrelin has sympathoinhibitory properties that are mediated by central ghrelin receptors involving a NPY/Y1 receptor-dependent pathway. Ghrelin's inhibitory effect on TNF-alpha production in sepsis is partially because of its modulation of the overstimulated sympathetic nerve activation.     

Mammary gland differentiation inversely correlates with GDF-8 expression

GDF-8 is recognised as an inhibitor of muscle cell growth and differentiation. Although initially thought to be restricted to muscle cells it is now accepted that GDF-8 expression has a broader tissue distribution. We demonstrate GDF-8 expression in the mouse mammary gland, which is predominantly associated with epithelial cells and displays an inverse correlation to the differentiated state of the gland. Specifically, the highest GDF-8 mRNA levels correlate with periods of maximal ductal growth, diminish as pregnancy progressed and are down-regulated to minimal levels by the onset of lactation as the epithelium differentiates. A similar profile is observed for both GDF-8 protein processing and reflects Smad2/3 phosphorylation profile. However, in contrast to muscle cells, GDF-8 neither reduces proliferation nor induces p21 expression levels in mammary epithelial cells. These data implicate a role for GDF-8 in mammary epithelial cell differentiation and demonstrate that GDF-8 has cell-type specific activities.     

Identification and estimation of endogenous digoxin in biological fluids and tissues by TLC and HPLC.

A procedure for estimation of digoxin in biological samples after adding a known quantity of digoxin followed by extraction, separation by TLC and HPLC is described. The identity of digoxin thus extracted from rat brain has been established by reaction with digoxin antibody and by its inhibition of Na(+)-K+ ATPase activity. The method could be a better substitute to the routine radioimmunoassay as interfering substances are removed by TLC and HPLC.     

Nitrogen-fixing azotobacters from mangrove habitat and their utility as marine biofertilizers

A dearth of information is available for nitrogen-fixing bacteria in coastal mangroves, and hence, the present study has been undertaken to analyse 44 root and associated soil samples, derived from a mangrove habitat of southeast coast of India. The root samples exhibit high counts of total heterotrophic bacteria and azotobacters along with high rates of nitrogen fixation, as compared to the rhizosphere soil samples. Among the plant species, Bruguiera cylindrica records high microbial counts and nitrogen fixation. From the samples analysed, three species of Azotobacter, viz., A. chroococcum, A. virelandii and A. beijerinckii were isolated, purified and identified. These species exhibit high growth, nitrogen fixation and in vitro production of phytohormone (Indole Acetic Acid, IAA) at NaCl salinity of 30 g l⁻¹. The azotobacters, which were inoculated with Rhizophora seedlings, increased significantly the average root biomass up to by 98.2%, the root length by 48.45%, the leaf area by 277.86%, the shoot biomass by 29.49% as compared to controls and they also increased the levels of total chlorophylls and carotenoids up to by 151.0% and 158.73%, respectively. Thus, azotobacterisation is beneficial in raising vigorous seedlings of mangroves in coastal wetlands.     

Low temperature-synthesized MgAl2O4:Eu3+ nanophosphors and their structural validations using density functional theory: photoluminescence,photocatalytic, and electrochemical properties for multifunctional applications

A low temperature-assisted and oxalyl dihydrazide fuel-induced combustion synthesized series of uncalcined MgAl₂O₄:Eu³⁺ nanophosphors showed an average crystallite size of ~20 nm, and bandgap energy (E_g) of 4.50–5.15 eV, and were validated using density functional theory and found to match closely with the experimental values. The photoluminescence characteristic emission peaks of Eu³⁺ ions were recorded between 480 and 680 nm. The nanophosphors excited at 392 nm showed f–f transitions assigned as ⁵D₀→⁷F_J (J = 0, 1, 2, and 3). The optimized MgAl₂O₄ phosphors had Commission Internationale de l'Eclairage coordinates in the red region, a correlated colour temperature of 2060 K, and a colour purity of 98.83%. The estimated luminescence quantum efficiency ( $\eta$) was observed to be ~63% using Judd–Ofelt analysis. Electrochemical and photocatalytic performance were explored and indicated its multifunctional applications. Therefore, MgAl₂O₄:Eu³⁺ nanophosphors could be used for the fabrication of light-emitting diodes, industrial dye degradation, and as electrodes for supercapacitor applications.