Cellular NADH and NADPH Conformation as a Real-Time Fluorescence-Based Metabolic Indicator under Pressurized Conditions

Cellular conformation of reduced pyridine nucleotides NADH and NADPH sensed using autofluorescence spectroscopy is presented as a real-time metabolic indicator under pressurized conditions. The approach provides information on the role of pressure in energy metabolism and antioxidant defense with applications in agriculture and food technologies. Here, we use spectral phasor analysis on UV-excited autofluorescence from Saccharomyces cerevisiae (baker’s yeast) to assess the involvement of one or multiple NADH- or NADPH-linked pathways based on the presence of two-component spectral behavior during a metabolic response. To demonstrate metabolic monitoring under pressure, we first present the autofluorescence response to cyanide (a respiratory inhibitor) at 32 MPa. Although ambient and high-pressure responses remain similar, pressure itself also induces a response that is consistent with a change in cellular redox state and ROS production. Next, as an example of an autofluorescence response altered by pressurization, we investigate the response to ethanol at ambient, 12 MPa, and 30 MPa pressure. Ethanol (another respiratory inhibitor) and cyanide induce similar responses at ambient pressure. The onset of non-two-component spectral behavior upon pressurization suggests a change in the mechanism of ethanol action. Overall, results point to new avenues of investigation in piezophysiology by providing a way of visualizing metabolism and mitochondrial function under pressurized conditions.     

Thermal,spectral and biological characterisation of copper(II) complexes with isoniazid-based hydrazones

Journal of Thermal Analysis and Calorimetry - A series of complexes [Cu(L)(CH3COO)2]·nH2O (L: isonicotinic acid 2-(2-hydroxy-8-substituted-tricyclo[7.3.1.02.7]tridec-13-ylidene)-hydrazones)...     

Simultaneous determination of colchicine and febuxostat in rat plasma: Application in a rat pharmacokinetic study

A selective, sensitive and rapid LC–MS/MS method has been developed and validated as per US Food and Drug Administration regulatory guidelines for the simultaneous quantitation of colchicine and febuxostat in rat plasma. Colchicine and febuxostat were extracted from the rat plasma using 10% tert-butyl methyl ether in ethyl acetate using colchicine-d₆ as an internal standard (IS). The chromatographic separation of colchicine, febuxostat and the IS was achieved using a mobile phase comprising 5 mm ammonium formate and 0.025% formic acid in acetonitrile (20:80, v/v) in isocratic mode on an Eclipse XDB-C₁₈ column. The injection volume and flow rate were 5.0 μl and 0.9 ml/min, respectively. Colchicine and febuxostat were detected by positive electrospray ionization in multiple reaction monitoring mode using transition pairs (Q1 → Q3) of m/z 400.10 → 358.10 and 317.05 → 261.00, respectively. The assay was linear in the ranges of 0.25–254 and 2.60–622 ng/ml for colchicine and febuxostat, respectively. The inter- and intra-day precision values were 0.58–13.0 and 1.03–4.88% for colchicine and febuxostat, respectively. No matrix or carryover effects were observed during the validation. Both analytes were stable on the bench-top, in the autosampler and in storage (freeze–thaw cycles and long-term storage at −80 ° C). A pharmacokinetic study in rats was performed to show the applicability of the validated method.     

Melatonin Rescues the Dendrite Collapse Induced by the Pro-Oxidant Toxin Okadaic Acid in Organotypic Cultures of Rat Hilar Hippocampus

The pro-oxidant compound okadaic acid (OKA) mimics alterations found in Alzheimer’s disease (AD) as oxidative stress and tau hyperphosphorylation, leading to neurodegeneration and cognitive decline. Although loss of dendrite complexity occurs in AD, the study of this post-synaptic domain in chemical-induced models remains unexplored. Moreover, there is a growing expectation for therapeutic adjuvants to counteract these brain dysfunctions. Melatonin, a free-radical scavenger, inhibits tau hyperphosphorylation, modulates phosphatases, and strengthens dendritic arbors. Thus, we determined if OKA alters the dendritic arbors of hilar hippocampal neurons and whether melatonin prevents, counteracts, or reverses these damages. Rat organotypic cultures were incubated with vehicle, OKA, melatonin, and combined treatments with melatonin either before, simultaneously, or after OKA. DNA breaks were assessed by TUNEL assay and nuclei were counterstained with DAPI. Additionally, MAP2 was immunostained to assess the dendritic arbor properties by the Sholl method. In hippocampal hilus, OKA increased DNA fragmentation and reduced the number of MAP2(+) cells, whereas melatonin protected against oxidation and apoptosis. Additionally, OKA decreased the dendritic arbor complexity and melatonin not only counteracted, but also prevented and reversed the dendritic arbor retraction, highlighting its role in post-synaptic domain integrity preservation against neurodegenerative events in hippocampal neurons.     

A New Variant of Emissive RNA Alphabets

A new fluorescent ribonucleoside alphabet (^mthN) consisting of pyrimidine and purine analogues, all derived from methylthieno[3,4-d]pyrimidine as the heterocyclic core, is described. Large bathochromic shifts and high microenvironmental susceptibility of their emission relative to previous alphabets derived from thieno[3,4-d]pyrimidine (^thN) and isothiazole[4,3-d]pyrimidine (^tzN) scaffolds are observed. Subjecting the purine analogues to adenosine deaminase, guanine deaminase and T7 RNA polymerase indicate that, while varying, all but one enzyme tolerate the corresponding ^mthN/^mthNTP substrates. The robust emission quantum yields, high photophysical responsiveness and enzymatic accommodation suggest that the ^mthN alphabet is a biophysically viable tool and can be used to probe the tolerance of nucleoside/tide-processing enzymes to structural perturbations of their substrates.     

Electrospun poly(lactic acid) nanofibers loaded with silver sulfadiazine/[Mg–Al]‐layered double hydroxide as an antimicrobial wound dressing

Poly(lactic acid) (PLA) is a versatile, bioabsorbable, and biodegradable polymer with excellent biocompatibility and ability to incorporate a great variety of active agents. Silver sulfadiazine (SDZ) is an antibiotic used to control bacterial infection in external wounds. Aiming to combine the properties of PLA and SDZ, hydrotalcite ([Mg–Al]‐LDH) was used as a host matrix to obtain an antimicrobial system efficient in delivering SDZ from electrospun PLA scaffolds intended for wound skin healing. The structural reconstruction method was successfully applied to intercalate silver sulfadiazine in the [Mg–Al]‐LDH, as evidenced by X‐ray diffraction and thermogravimetric analyses. Observations by scanning electron microscopy revealed a good distribution of SDZ‐[Mg–Al]‐LDH within the PLA scaffold. Kinetics studies revealed a slow release of SDZ from the PLA scaffold due to the intercalation in the [Mg–Al]‐LDH. In vitro antimicrobial tests indicated a significant inhibitory effect of SDZ‐[Mg–Al]‐LDH against Escherichia coli and Staphylococcus aureus. This antibacterial activity was sustained in the 2.5‐wt% SDZ‐[Mg–Al]‐LDH–loaded PLA nanofibers, which also displayed excellent biocompatibility towards human cells. The multifunctionality of the PLA/SDZ‐[Mg–Al]‐LDH scaffold reported here is of great significance for various transdermal applications.     

One-pot three-component tandem reaction: Synthesis of aryl/alkyl cyanamides libraries and their further conversion into tetrazole derivatives

We have developed methodology for the synthesis of aryl/alkyl cyanamides from amines in one-pot four steps reaction using cheap, readily available and air stable copper source as catalyst under mild reaction conditions. We have also studied the application of cyanamides. In this connection, we could construct aryl tetrazolamine from cyanamides using click reaction     

Progress in drug delivery to the central nervous system by the prodrug approach

This review describes specific strategies for targeting to the central nervous system (CNS). Systemically administered drugs can reach the brain by crossing one of two physiological barriers resistant to free diffusion of most molecules from blood to CNS: the endothelial blood-brain barrier or the epithelial blood-cerebrospinal fluid barrier. These tissues constitute both transport and enzymatic barriers. The most common strategy for designing effective prodrugs relies on the increase of parent drug lipophilicity. However, increasing lipophilicity without a concomitant increase in rate and selectivity of prodrug bioconversion in the brain will result in failure. In these regards, consideration of the enzymes present in brain tissue and in the barriers is essential for a successful approach. Nasal administration of lipophilic prodrugs can be a promising alternative non-invasive route to improve brain targeting of the parent drugs due to fast absorption and rapid onset of drug action. The carrier-mediated absorption of drugs and prodrugs across epithelial and endothelial barriers is emerging as another novel trend in biotherapeutics. Several specific transporters have been identified in boundary tissues between blood and CNS compartments. Some of them are involved in the active supply of nutrients and have been used to explore prodrug approaches with improved brain delivery. The feasibility of CNS uptake of appropriately designed prodrugs via these transporters is described in detail.