Method For Determination Of Glutathione Research Articles

Construction of core–shell MOF@COF hybrid materials as highly efficient light-responsive oxidase-like nanozymes for colorimetric detection of glutathione

In this work, a novel and light-responsive nanozyme, with enzyme-like characteristics, was synthesized based on core–shell MOF@COF hybrid materials. The donor–acceptor type COF (as shell) synthesized by 1,3,6,8-tetrakis(4-aminophenyl)pyrene (Py, donor) and thieno[3,2-b]thiophene-2,5-dicarbaldehyde (Tt, acceptor) was wrapped on the NH2-Cu-MOF (robust catalyst, as core) through a sequential growth strategy. Surprisingly, with visible-light irradiation, the core–shell Cu-MOF@Py-Tt hybrid materials exhibited extremely high oxidase-like enzymatic catalytic activity attributing to the high separation efficiency of photogenerated carriers in core–shell hybrid materials and the shell with the characteristics of large π-conjugated and D-A electronic structure. With the light-control catalytic activity, Cu-MOF@Py-Tt can form the O2•- that can catalyze the TMB to blue color. Consequently, a light-control, sensitive and rapid colorimetric method for determination of glutathione (GSH) was designed based on light-responsive oxidase mimic of core–shell Cu-MOF@Py-Tt with the linear range of 0.2–70 μM and limit of detection of 0.17 μM. The proposed method demonstrated the future potential of core–shell MOF@COF hybrid materials to obtain excellent nanozyme activity.

Voltammetric method for determination of glutathione on a gold-carbon-containing electrode

Glutathione (GSH) is one of the most important thiol-containing antioxidants involved into various biochemical processes in the human body. Glutathione determination in biological fluids (saliva, urine, serum) and pharmaceutical preparations is rather important for clinical practice. Various analytical methods — spectrophotometry, fluorimetry, high-performance liquid chromatography, NMR spectroscopy, capillary electrophoresis and electrochemical methods — are widely used for this purpose. Electrochemical methods are characterized by easy implementation, low cost and possibility of miniaturization. The electrochemical behavior of reduced (GSH) and oxidized (GSSG) glutathione on a gold-carbon-containing electrode (AuCE) was studied using cathodic voltammetry with different methods of removing oxygen from an electrochemical cell: nitrogen sparging and addition of sodium sulfite (4 mol/dm3). It has been shown that traces of H2O2 that remain in the near-electrode layer on the AuCE even after oxygen removal influence the electrochemical properties of GSH at a cathode sweep of the potential from 0 to –1.8 V: GSH is oxidized by H2O2 to GSSG, the most important product of this reaction is O2. An indirect determination of GSH by the current of oxygen reduction in the Na2SO3 medium in the concentration range from 0.5 × 10–8 to 4.2 × 10–8 mol/dm3 with a detection limit of 2.5 × 10–9 mol/dm3 is proposed. The developed voltammetric method is approved for the determination of GSH in certain pharmaceutical preparations.

Fluorometric determination and imaging of glutathione based on a thiol-triggered inner filter effect on the fluorescence of carbon dots

The authors describe a sensitive method for determination of glutathione (GSH) that is based on a thiol-triggered inner filter effect on the fluorescence of N-doped carbon dots (N-doped CDs). N-doped CDs with a quantum yield as high as 31% were prepared by a one-pot procedure, and 5,5′-dithiobis-(2-nitrobenzoic acid) was employed as a reagent for GSH recognition. The reaction product (5-thio-2-nitrobenzoic acid; TNB) acts as an absorber of the 410-nm light used to photo-excite the N-doped CDs. Hence, the fluorescence of N-doped CDs (peaking at 510 nm) is reduced with increasing concentrations of GSH. As little as 30 nM of GSH can be detected by this method. The approach was successfully applied to (a) food analysis, (b) an investigation of an oxidative stress model, and (c) to live cells imaging. The method does not require the surface of N-doped CDs to be chemically modified, and a linkage between receptor and fluorophore is not needed. In our perception, the method may become a viable tool for the detection and imaging of thiols.

Determination of glutathione based on NiPd nanoparticles mediated with acetaminophen

A new method for determination of glutathione (GSH) was carried out with acetaminophen (AAP) mediation on a NiPd nanoparticle modified electrode.

Glutathione and related thiol compounds. I. Glutathione and related thiol compounds in flour

Abstract The high pressure liquid chromatography method for determination of glutathione in free and protein-bound forms was re-established and has successfully been developed to measure glutathione related thiol compounds, i.e. l -cysteine, γ- l -glutamyl- l -cysteine and l -cysteinyl- l -glycine, in both free and protein-bound forms. The natural levels of those compounds in typical strong, weak flours, and flours from 36 wheat varieties grown in the UK were investigated. The total free and protein-bound glutathione compounds found in the 36 UK varieties was 358±51 and 190±17 nmol/g, respectively. Multiple correlation analysis did not show a clear-cut relationship between the natural level of glutathione and any related thiol compound in either free or protein-bound forms and flour quality attributes, including rheological properties, baking performance, protein content and SDS sedimentation test values. Therefore, it can be suggested that glutathione and related thiol compounds at natural levels do not lead to significant differences in the rheological properties of dough and the baking performance of flour.

Determination of glutathione in single human hepatocarcinoma cells by capillary electrophoresis with electrochemical detection

A method for determination of glutathione (GSH) in single human hepatocarcinoma (HH) cells was described by capillary zone electrophoresis with electrochemical detection at a gold/mercury amalgam micro-disk electrode. When HH cells were washed with the running buffer instead of physiological buffer saline, only one electrophoretic peak for GSH is depicted on the electropherograms of single HH cells. When electroosmotic injection of 0.01 mol/l NaOH for lysing the cell introduced into the capillary, the lysis time can be shorten to 5 s. The whole cell injection and no need of derivatization reaction lead more accurate and precise results. The average amount of GSH in an individual HH cell is 22.3±5.8 fmol (mean±standard deviation), which is consistent with that of its homogenate.

The enhanced electrogenerated chemiluminescence of Ru(bpy)3(2+) by glutathione on a glassy carbon electrode modified with some porphine compounds.

It has been found that the electrochemical activity of glutathione was increased greatly at the glassy carbon electrodes modified with 5,10,15,20-tetraphenylporphine ruthenium(II) carbenyl (RuTPP), meso-tetraphenylporphine copper(II) complex (CuTTP) and hemin. It has been also found that glutathione would enhance the electrogenerated chemiluminescence (ECL) of Ru(bpy)3(2+) at a hemin glassy carbon electrode; the enhanced ECL intensity was linear with the concentration of glutathione in the range of 1 x 10(-7)-1 x 10(-4) mol l-1, based on which method for determination of glutathione has been developed. The detection limit of glutathione was 2 x 10(-8) mol l-1, and the relative standard deviation for 1 x 10(-6) mol l-1 glutathione was 2.7%. The mechanism for this ECL system has been proposed.

Validated High-Performance Liquid Chromatography-Electrochemical Method for Determination of Glutathione and Glutathione Disulfide in Small Tissue Samples

Glutathione (GSH) and glutathione disulfide (GSSG) are biologically important intracellular thiols; alterations in the GSH/GSSG ratio are often used to assess exposure of cells to oxidative stress. Although several methods are available for measuring GSH and GSSG, all have some disadvantages including the need to generate derivatives, the inability to conveniently measure both GSH and GSSG, and a lack of sufficient sensitivity to allow detection in very small samples/cells of extrahepatic tissue. These studies present a rapid, validated HPLC-electrochemical method for determining GSH and GSSG in small samples such as those from microdissected airways of the mouse containing 50–200 μg protein which is suitable for routine use. GSH and GSSG can be measured at levels of 1 and 2 pmol on column, respectively, with acceptable accuracy and precision and without the need to generate derivatives. In microdissected airways from the mouse, the intraday assay coefficient of variation for GSH varied from 4.7 to 5.9% and for GSSG was 4.4 to 5.7%. The interday assay coefficient of variation ranged from 6.0 to 7.6% for GSH and 5.5 to 23% for GSSG. The effects of repeated freezing and thawing on the concentrations of GSH and GSSG indicate that multiple cycles do not significantly alter the GSH or GSSG concentration as the number of cycles increases. Addition of GSH or GSSG to samples increased the peak areas appropriately, without altering the peak shape, retention time, or peak area of the corresponding reduced (oxidized) thiol. The ratio of GSH/GSSG in freeze-clamped liver ranged from 46 to 248, while liver tissue which was homogenized fresh had GSH/GSSG ratios of 62–150. The technique appears to be capable of reproducibly measuring GSH and GSSG in small quantities of nonhepatic tissue.

A specific, sensitive, and rapid method for the determination of glutathione and its application in ocular tissues

A method for specific determination of glutathione (GSH) is described. This method utilizes the enzymatic conjugation of GSH to 1-chloro-2,4-dinitrobenzene through reaction catalyzed by glutathione S-transferase. The recovery of GSH as determined by this method is comparable to that in currently used methods. The method is specific for GSH determination. Other sulfhydryl (-SH) compounds including the protein -SH or β-mercaptoethanol, which are often included in tissue homogenates, do not interfere with GSH determination. Acid extraction of the tissue is not required in this method and comparatively smaller amounts of tissue samples (as little as 20 μl of a 10% w v tissue homogenate) are needed for the analyses. The method when applied for GSH determination in ocular tissues yielded results in agreement with the reported values in literature. Evidence for the sensitivity, accuracy, and convenience of the method is provided by analysing the sample containing GSH in the range of 1–200 nmol by this method.

A high-sensitivity fluorometric high-performance liquid chromatographic method for determination of glutathione and other thiols in cultured melanoma cells, microdialysis samples from melanoma tissue, and blood plasma

A high-performance liquid chromatographic method with fluorometric detection is described which is suitable for determination of glutathione in small samples. Reduced glutathione (GSH) and total glutathione obtained as GSH after reduction with glutathione reductase is derivatized with N-(7-dimethylamino-4-methyl-3-coumarinyl) maleimide (DACM) and subjected to chromatography. The detection limit for the GSH-DACM derivative was 5-10 fmol/injection, and analytical recovery was quantitative. The method is suitable for determination of both reduced and total glutathione in samples from microdialysis of melanoma tumours, and cysteine can be quantified in the same chromatogram. Application is shown also for glutathione determinations in cultured melanoma cells, melanoma homogenates and plasma.

Determination of glutathione in Scots pine needles by high-performance liquid chromatography as its mono-bromobimane derivative

A method for determination of glutathione in its reduced (GSH) and oxidized (GSSG) forms in Scots pine extracts by reversed-phase high-performance liquid chromatography utilizing monobromobimane as the derivatization reagent was developed. The recovery and the precision for GSH in pine needles was high, ca. 99 and 3.6%, respectively. The determination of GSSG showed lower recovery ( ca. 80%) and poorer precision (13.3%). The identity of the putative GSH-bimane derivative was confirmed indirectly by gas chromatography-mass spectrometry. GSH comprised 77% of water-extractable thiols in pine needles, and the ratio GSSG/GSH was low (0.024).