Development and validation of an economical uric acid-Fe3+/Fe2+-ferrozine-based colorimetric assay to estimate uric acid level of pure and biological samples

ABSTRACT

This work presents the development and validation of a simple, rapid, and cost-effective spectrophotometric method for quantitative analysis of uric acid in biological samples. The method relies upon uric acid-led reduction of Fe(III) to Fe(II) of sample/standard solutions which stoichiometrically engages ferrozine to form a magenta-colored complex. Different parameters including pH, metal and chelator concentrations, temperature, etc., were optimized for the maximum intensity and stability of the complex. The uric acid concentrations of synthetic/plasma solutions were determined by comparing the color intensity of Fe(ferrozine)₃ ²⁺ complex produced by test solution with the standard curve formed by known uric acid concentrations. The method was validated in accordance with ICH guidelines and subjected to human plasma analysis. The results obtained were compared with a reference (enzymatic) method which revealed that there was no significant difference between the two methods at 95% confidence level. The method is highly specific, precise, linear, accurate, and robust.     

New spectrofluorimetric and spectrophotometric methods for the determination of the analgesic drug,nalbuphine in pharmaceutical and biological fluids

We describe the first studies of a simple and sensitive spectrofluorimetric and spectrophotometric methods for the analysis of nalbuphine (NLB) in dosage form and biological fluids. The spectrofluorimetric method was based on the oxidation of NLB with Ce(IV) to produce Ce(III) and its fluorescence was monitored at 352 nm after excitation at 250 nm. The spectrophotometric method involves addition of a known excess of Ce(IV) to NLB in acid medium, followed by determination of residual Ce(IV) by reacting with a fixed amount of methyl orange and measuring absorbance at 510 nm. In both methods, the amount of Ce(IV) reacted corresponds to the amount of NLB and measured fluorescence or absorbance were found to increase linearly with the concentration of NLB, which are corroborated by correlation coefficients of 0.9997 and 0.9999 for spectrofluorimetric and spectrophotometric methods, respectively. Different variables affecting the reaction conditions such as concentrations of Ce(IV), type and concentration of acid medium, reaction time, temperature, and diluting solvents were carefully studied and optimized. The accuracy and precision of the methods were evaluated on intra‐day and inter‐day basis. The proposed methods were successfully applied for the determination of NLB in pharmaceutical formulation and biological samples with good recoveries. Copyright © 2012 John Wiley & Sons, Ltd.     

Sub millimetre flexible fibre probe for background and fluorescence free Raman spectroscopy

Using the shifted-excitation Raman difference spectroscopy technique and an optical fibre featuring a negative curvature excitation core and a coaxial ring of high numerical aperture collection cores, we have developed a portable, background and fluorescence free, endoscopic Raman probe. The probe consists of a single fibre with a diameter of less than 0.25 mm packaged in a sub-millimetre tubing, making it compatible with standard bronchoscopes. The Raman excitation light in the fibre is guided in air and therefore interacts little with silica, enabling an almost background free transmission of the excitation light. In addition, we used the shifted-excitation Raman difference spectroscopy technique and a tunable 785 nm laser to separate the fluorescence and the Raman spectrum from highly fluorescent samples, demonstrating the suitability of the probe for biomedical applications. Using this probe we also acquired fluorescence free human lung tissue data.     

High resolution,label‐free photoacoustic imaging of live chicken embryo developing in bioengineered eggshell

Chicken embryos have been proven to be an attractive vertebrate model for biomedical research. They have helped in making significant contributions for advancements in various fields like developmental biology, cancer research and cardiovascular studies. However, a non‐invasive, label‐free method of imaging live chicken embryo at high resolution still needs to be developed and optimized. In this work, we have shown the potential of photoacoustic tomography (PAT) for imaging live chicken embryos cultured in bioengineered eggshells. Laser pulses at wavelengths of 532 and 740 nm were used for attaining cross‐sectional images of chicken embryos at different developmental stages. Cross‐sections along different depths were imaged to gain knowledge of the relative depth of different vessels and organs. Due to high optical absorption of vasculature and embryonic eye, images with good optical contrast could be acquired using this method. We have thus reported a label‐free method of performing cross‐sectional imaging of chicken embryos at high resolution demonstrating the capacity of PAT as a promising tool for avian embryo imaging.     

Alpha-Tocopherol-Induced Regulation of Growth and Metabolism in Plants Under Non-stress and Stress Conditions

Alpha-tocopherol (α-Toc) is a member of the vitamin E family and is lipid soluble. Its biosynthesis is by the reaction of isopentyl diphosphate and homogentisic acid in plastid membranes. The putative biochemical activities of tocopherols are linked with the formation of tocopherol quinone species, which subsequently undergo degradation and recycling within cells/tissues. α-Toc plays a key role in a variety of plant metabolic processes throughout the ontogeny of plants. It can maintain the integrity and fluidity of photosynthesizing membranes. It can also neutralize lipid peroxy radicals, consequently blocking lipid peroxidation by quenching oxidative cations. It preserves membrane integrity by retaining membranous structural components under environmental constraints such as water deficiency, high salt content, toxic metals, high/low temperatures, and radiations. α-Toc also induces cellular signalling pathways within biological membranes. Its biosynthesis varies during growth and developmental stages as well as under different environmental conditions. The current review primarily focuses on how α-Toc can regulate various metabolic processes involved in promoting plant growth and development under stress and non-stress and how it can effectively counteract the stress-induced high accumulation of reactive oxygen species (ROS). Currently, exogenous application of α-Toc has been widely reported as a potential means of promoting resistance in plants to a variety of stressful environments.

     

Correlating the charge transfer efficiency of metallic sulfa-isatins to design efficient NLO materials with better drug designs

BioMetals - The present study presents synthesis, characterization and first principle studies on metal chelates, (1–12), of sulfonamide-isatin reacted ligands (S1–S3). All the products...     

Interferon-γ down-regulates membrane adenylate cyclase activity of a murine macrophage-like cell line (P388D1)

Effects of recombinant murine interferon-γ (rIFN-γ) on the membrane adenylate cyclase of a murine macrophage cell line (P388D₁) were investigated in order to explore the nature of a signal transmitted by IFN-γ receptor. Following the incubation of P388D₁ cells with 40 U/ml of rIFN-γ, the intracellular level of cAMP gradually increased about twofold over the control level within 60 min, and then began to gradually decline to about half the control level by 24 h incubation. The initial rise in cAMP level appeared to be due to the modest activation of adenylate cyclase and not due to the inhibition of cAMP-phosphodiesterase. Later decrease of intracellular cAMP may be due to quantitative down-regulation of the adenylate cyclase system. The basal enzymatic activity of the membrane prepared from P388D₁ cells exposed to IFN-γ for 24 h was found to be reduced to about 20% of that of the control membrane. However, the quality of the adenylate cyclase system appeared unchanged, because the relative degree of the response of the down-regulated membrane adenylate cyclase to prostaglandin PGE₂, NaF, guanylimidodiphosphate (GppNHp), cholera toxin (CT), or forskolin was found to remain unchanged. This quantitative down-regulation of adenylate cyclase must be due to the action of rIFN-γ, since the prior treatment of rIFN-γ with either acid (pH 2) or monoclonal anti-IFN-γ antibody inhibited the ability of IFN-γ to induce the down-regulation. The rIFN-γ-induced down-regulation is a reversible process, since the adenylate cyclase activity of the membrane was found to be restored when the rIFN-γ-exposed cells were cultured for 72 h in the absence of rIFN-γ. In addition, the 48 h-incubation of P388D₁ cells with rIFN-β or IFN-α was found not to significantly affect the membrane adenylate cyclase system.     

The biological effects of vanadyl curcumin and vanadyl diacetylcurcumin complexes: the effect on structure,function and oxidative stability of the peroxidase enzyme,antibacterial activity and cytotoxic effect

Curcumin has multiple pharmacological effects, but it has poor stability. Complexation of curcumin with metals improves its stability. Here, the effects of vanadyl curcumin and vanadyl diacetylcurcumin on the function and structure of horseradish peroxidase enzyme were evaluated by spectroscopic techniques. Cytotoxic effect of the complexes was also assessed on MCF-7 breast cancer, bladder and LNCaP prostate carcinoma cell line. The results showed that the complexes improve catalytic activity of HRP, and also increase its tolerance against the oxidative condition. The result also indicated that the affinity of HRP for hydrogen peroxide substrate decreases, while the affinity increases for phenol substrate. Circular dichroism and fluorescence spectroscopies showed that compactness of the enzyme structure around the catalytic heme group and the distance between the heme group and tryptophan residue decreases after the binding. The antibacterial and cytotoxic results indicated that the complexes have anticancer potential, but they have no considerable antibacterial activity.     

Why Do Intravascular Schistosomes Coat Themselves in Glycolytic Enzymes?

Schistosomes are intravascular parasitic helminths (blood flukes) that infect more than 200 million people globally. Proteomic analysis of the tegument (skin) of these worms has revealed the surprising presence of glycolytic enzymes on the parasite's external surface. Immunolocalization data as well as enzyme activity displayed by live worms confirm that functional glycolytic enzymes are indeed expressed at the host–parasite interface. Since these enzymes are traditionally considered to function intracellularly to drive glycolysis, in an extracellular location they are hypothesized to engage in novel “moonlighting” functions such as immune modulation and blood clot dissolution that promote parasite survival. For instance, several glycolytic enzymes can interact with plasminogen and promote its activation to the thrombolytic plasmin; some can inhibit complement function; some induce B cell proliferation or macrophage apoptosis. Several pathogenic bacteria and protists also express glycolytic enzymes externally, suggesting that moonlighting functions of extracellular glycolytic enzymes can contribute broadly to pathogen virulence. Also see the video abstract here https://youtu.be/njtWZ2y3k_I