Norepinephrine-Fe(III)-ATP Ternary Complex and Its Relevance to Parkinson's Disease.

The aberrant autoxidation of norepinephrine (NE) in the presence of oxygen, which is accelerated by Fe(III), has been linked to the pathogenesis of the Parkinson's disease (PD). Adenosine triphosphate (ATP), as a neurotransmitter whose release can be stimulated by tissue damage and oxidative stress, is co-stored and often co-released with NE in presynaptic terminals. We have shown previously that ATP inhibits the iron-catalyzed dopamine oxidation, thereby decreasing the production of certain neurotoxins such as 6-hydroxydopamine. Whether ATP plays a similar role in Fe(III)-catalyzed NE oxidation and how it maintains the NE stability have not been investigated. Here, we studied the coordination in a ternary complex among NE, Fe(III), and ATP, and found that Fe(III) is coordinated as a octahedral center by NE and ATP. Voltammetry and mass spectrometry were employed to examine this ternary complex's modulation of the NE autoxidation. NE-Fe(III)-ATP plays a protective role to modulate the autoxidation and Fe(III)-catalyzed oxidation of NE. The ternary complex can be detected in the substantia nigra (SN), locus coeruleus (LC), and striatum regions of C57BL/6 wild-type mice. In contrast, the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mouse brains displayed a significant decrease of the ternary complex in the SN region and an increase in the LC and striatum areas. We posit that the ternary complex is produced by noradrenergic neurons as a protective regulator against neuronal damage and oxidative stress, contributing to the lower vulnerability of LC neurons with respect to that of SN neurons.

An improved Bathocuproine assay for accurate valence identification and quantification of copper bound by biomolecules.

Copper is an essential metal in all organisms. Reliably quantifying and identifying the copper content and oxidation state is crucial, since the information is essential to understanding protein structure and function. Chromophoric ligands, such as Bathocuproine (BC) and its water-soluble analog, Bathocuproinedisulfonic acid (BCS), preferentially bind Cu(I) over Cu(II), and therefore have been widely used as optical probes to determine the oxidation state of copper bound by biomolecules. However, the BCS assay is commonly misused, leading to erroneous conclusions regarding the role of copper in biological processes. By measuring the redox potential of Cu(II)-BCS2 and conducting UV-vis absorption measurements in the presence of oxidizable amino acids, the thermodynamic origin of the potential artifacts becomes evident. The BCS assay was improved by introducing a strong Cu(II) chelator EDTA prior to the addition of BCS to prevent interference that might arise from Cu(II) present in the sample. The strong Cu(II) chelator rids of all the potential errors inherent in the conventional BCS assay. Applications of the improved assay to peptides and protein containing oxidizable amino acid residues confirm that free Cu(II) no longer leads to artifacts, thereby resolving issues related to this persistently misused colorimetric assay of Cu(I) in biological systems.

One-step, room temperature, colorimetric melamine sensing using an in-situ formation of silver nanoparticles through modified Tollens process.

We have developed a rapid, sensitive, one-step, and selective colorimetric detection method for melamine (MEL) in milk powder based upon an in-situ formation of silver nanoparticles (AgNPs) through modified Tollens process at room temperature. The triazine ring N atoms of MEL molecule were strategically designed to complex the Ag(+) through electron donor-acceptor interaction. During the AgNPs formation procedure, the MEL molecule, which has been covalently bonded with the Ag(+) ions, was adsorbed to the surface of as-prepared AgNPs, resulting in the aggregation of the adjacent AgNPs with detectable decreases of absorption signal. The concentration of MEL can be determined with the naked eye or a UV-vis spectrometer at which the yellow-to-brown color change associated with aggregate enhancement takes place. This method enables rapid (less than 30 min) and sensitive (limit of detection, LOD, 10 nM) detection, and it was also able to discriminate MEL from sixteen other milk relevant coexisting compounds. This assay does not utilize organic cosolvents, enzymatic reactions, light-sensitive dye molecules, lengthy protocols, or sophisticated instrumentation thereby overcoming some of the limitations of conventional methods.

A simple and sensitive assay of gallic acid based on localized surface plasmon resonance light scattering of silver nanoparticles through modified Tollens process.

A simple, low toxic, sensitive strategy based on the localized surface plasmon resonance light scattering (LSPR-LS) properties of silver nanoparticles (AgNPs) is introduced for the detection of gallic acid (GA). It was found that the silver ammonium complex, [Ag(NH(3))(2)](+)(aq), could be reduced in the alkaline medium by GA at room temperature; this reaction formed dispersed AgNPs. Transmission electron microscopy analyses were performed to ascertain the formation of AgNPs. UV-visible spectra revealed the localized surface plasmon resonance (LSPR) absorption at 410 nm corresponding to the LSPR of AgNPs. On these basis, we could quantify the GA concentration in the range of 4 × 10(-7) - 5 × 10(-6) mol L(-1) in the optimized experimental conditions. This method was used for determining the concentration of GA in artificial samples with satisfactory results. The detailed mechanism underlying this special phenomenon was elucidated.

A localized surface plasmon resonance light scattering-based sensing of hydroquinone via the formed silver nanoparticles in system.

In this contribution, a simple strategy for the detection of hydroquinone (HQ) is proposed based on the localized surface plasmon resonance light scattering (LSPR-LS) of the silver nanoparticles (AgNPs) formed through the modified silver mirror reaction. The redox reaction between HQ and silver ammonia occurred in the coexistence of sodium hydroxide and ammonia at room temperature, where silver ammonia was reduced by HQ and resulted in the formation of AgNPs without adding the AgNPs seeds. The formed AgNPs were demonstrated to be monodisperse and uniform by transmission electron microscopy (TEM) image. We also studied the localized surface plasmon resonance absorption (LSPR-A) and LSPR-LS spectra using both a UV-vis spectrophotometer and a common spectrofluorometer, and obtained a good agreement between experiments. By carefully optimizing the amount of NaOH and ammonia of the reaction conditions, we were able to obtain the highest net intensity of LSPR-LS on the concentrations of HQ. On the basis of experimental studies, the LSPR-LS intensity enhanced linearly over the range 0.4-2.5 µmol L(-1) with the corresponding limits of determination (3σ) of 70.6 nmol L(-1). With that, the present approach was applied to detect HQ in water samples with satisfactory results.

Chemiluminescence determination of cefotaxime sodium with flow-injection analysis of cerium (IV)-rhodamine 6G system and its application to the binding study of cefotaxime sodium to protein with on-line microdialysis sampling.

A simple, sensitive and rapid flow-injection chemiluminescence (CL) method has been developed for the determination of cefotaxime sodium based on the chemiluminescence reaction of cefotaxime sodium with ceric sulfate and rhodamine 6G in nitric acid solution. The concentration of cefotaxime sodium was proportional with the CL intensity in the range of 4×10(-8)-8×10(-6) mol L(-1). The detection limit (signal-to-noise ratio=3) was 1×10(-8) mol L(-1). Coupled to the technique of on-line microdialysis sampling, this method was successfully applied to study cefotaxime sodium-protein interaction. The drug and protein were mixed in different molar ratios in Ringer's solution, pH 7.4, and incubated at 37°C in a water bath. The microdialysis probe was utilized to sample the mixed solution at a perfusion rate of 5 µL min(-1) and the recovery of cefotaxime sodium under experimental condition was 16.2%. The data obtained by the present Microdialysis-Flow Injection Analysis-CL method was analyzed with the Scatchard analysis and Klotz plot. The estimated association constant (K) and the number of the binding sites (n) on one of BSA molecule were 5.94×10(4) M(-1) and 1.29 (Klotz equation), respectively.

Chemiluminescence microflow injection analysis system on a chip for the determination of uric acid without enzyme.

A new microflow injection analysis (microFIA) system on a chip coupled with chemiluminescence (CL) for the non-enzymatic determination of uric acid is described. The microFIA system produced by using two transparent poly(methylmethacrylate) (PMMA) chips measured 50 x 40 x 5 mm, the microchannels, etched by CO2 laser, were 200 microm wide and 100 microm deep, and the volume of the reaction area (RA) was about 1.2 microL. The injection pump, with accurate time control, monitored all reagents, including the sample. The uric acid was sensed by the chemiluminescence reaction between luminol and ferricyanide. The linear range of the uric acid concentration was 0.8-30 mg/L and the detection limit was 0.5 mg/L (S/N = 3). The relative standard deviation was 4.42% for 5 mg/L uric acid (n = 8). The proposed method has been successfully applied to the non-separation determination of uric acid in human serum and urine.

Flow-injection chemiluminescence detection for studying protein binding of terbutaline sulfate with on-line microdialysis sampling.

The binding of terbutaline sulfate to bovine serum albumin was studied in vitro using the technique of microdialysis sampling combined with flow-injection chemiluminescence analysis (FIA-CL). In the presence of formaldehyde, terbutaline sulfate can be oxidized by KMnO(4) to produce high chemiluminescence emission in sulfate acid media. The concentration of terbutaline sulfate is proportional with the CL intensity in the range of 1 x 10(-7)-2 x 10(-5) mol l(-1) with a detection limit of 3 x 10(-8) mol l(-1). The drug and protein were mixed in different molar ratios in 0.067 mol l(-1) phosphate buffer, pH 7.4, and incubated at 37 degrees C in a water bath. The microdialysis probe was utilized to sample the mixed solution at a perfusion rate of 5 microl min(-1) and the dialytic efficiency of terbutaline sulfate under the experimental conditions was 26.3%. The data obtained by proposed microdialysis flow-injection chemiluminescence method was analyzed with Scrathard analysis and Klotz plot. The estimated association constant (K) and the number of the binding site (n) on one molecule of BSA by Scrathard analysis were 4.11 x 10(4) l mol(-1) and 1.06, respectively. The proposed system proved that FIA-CL coupled with on-line microdialysis sampling is a simple and reliable technique for the study of drug-protein interaction.