Hongmei Shi

Highly selective and sensitive determination of several antioxidants in human breast milk using high-performance liquid chromatography based on Ag(III) complex chemiluminescence detection

Ascorbic acid (AA), uric acid (UA) and glutathione (GSH) are the most important water-soluble antioxidants. The concentrations of GSH and glutathione disulfide (GSSG) and their molar ratio are the indicators of oxidative stress. Little is known about the contents of UA, GSH and GSSG in human milk; a reliable and sensitive method to monitor the concentrations of the four compounds simultaneously in human milk is of critical importance. A new method for separation and quantification of these water-soluble antioxidants by HPLC coupled with Ag(III) chemiluminescence detector has been developed in this work with better recoveries. The antioxidants contents were determined in different times of lactation utilizing this method. The results show that the levels of AA, UA, GSH and GSH/GSSG of human colostrum are significantly higher than those of mature milk (P<0.05). It is concluded that colostrum contains more water-soluble antioxidants than mature milk.

Kinetics and mechanism of oxidation of the drug intermediate 1-(2-Hydroxyethyl)piperidine by Bis(hydrogenperiodato)argentate(III)

1-(2-Hydroxyethyl)piperidine (HEP) is involved in many drugs and drug leads, but its oxidation mechanisms are poorly understood. The oxidation of HEP by bis(hydrogenperiodato)-argentate(III) ([Ag(HIO6)2]5-;) in aqueous alkaline medium was shown, by electrospray ionization mass spectrometry (ESI-MS), to generate piperidine and formaldehyde as the major products. The reaction was monitored spectrophotometrically in the 25.0 to 40.0 oC range revealing that the oxidation followed a first-order kinetics in [Ag(III)] and a fractional-order in [HEP]. A rate law and a reaction mechanism were proposed based on the study of the dependency of the pseudo-first-order rate constants, kobsd, on [OHˉ] and on [IO4ˉ]tot (total concentration of periodate). The mechanism involves the formation of a periodato-Ag(III)-HEP ternary complex, whose decomposition generates Ag(I) by means of two pathways: one independent and another facilitated by OHˉ. The reaction rate constants and associated equilibrium constants as well as the activation parameters of the rate-determining steps were calculated.

Trace level determination of 5-hydroxytryptamine and its related indoles in amniotic fluid by gas chromatography–mass spectrometry

&NA; 5‐hydroxytryptamine (5‐HT) and its derivatives are endogenously active substances involved in multiple physiological and pathological processes. A novel method of detetermining 5‐hydroxyindole ethanol (5‐HTOL), 5‐hydroxyindole acetic acid (5‐HIAA), 5‐hydroxytryptophan (5‐HTP) and 5‐HT in amniotic fluid by gas chromatography‐mass spectrometry (GC–MS) was established based on a modified method of derivatization by silanization, in combination with solid‐phase extraction pretreatment. Good linearity was achieved in the tested calibration range. The limits of detection (LOD) were 0.05, 0.08, 0.56, 0.43 &mgr;g/L for 5‐HTOL, 5‐HIAA, 5‐HTP and 5‐HT, respectively. Accuracy (92.4‐103.3) and precision (RSD < 5.4%) for all analytes was also determined. Then the method was used to analyze samples of amniotic fluid from 12 patients carrying foetuses with trisomy 21 and 12 healthy controls. Compared with normal fetuses, the levels of 5‐HTOL, 5‐HTP and 5‐HT in the amniotic fluid were significantly altered in the fetuses with trisomy 21 (P < 0.01); the level of 5‐HIAA showed no significant difference between the two groups (P>0.05). This is a rapid, sensitive and reliable method for the determination of 5‐HTOL, 5‐HIAA, 5‐HTP and 5‐HT, and the study provide both potential trisomy 21 markers and elucidation of the physiological and pathological roles of 5‐HT. Graphical abstract Figure. No caption available. Highlights5‐HT and its related indoles play important physiological roles.A novel GC–MS method of detecting 5‐HTOL, 5‐HIAA, 5‐HTP and 5‐HT in amniotic fluid was established.5‐HT and its related indoles in the T21 had been explored by the proposed method.The sensitive method may be used in studies focusing on clinical pathogenesis of T21.

Investigation of a novel Ag(III) chemiluminescence system and its mechanism for determination of uric acid in human urine

A simple and sensitive method based on flow-injection chemiluminescence analysis coupled with luminol-Ag(III) complex was developed for the determination of uric acid (UA) in human urine. The method was based on the chemiluminescence (CL) reaction of luminol with Ag(III) in alkaline solution. UA in the urine can dramatically enhance CL intensities. In optimum conditions, the relative CL intensity had a linear relationship with UA concentration in the range of 1.0 × 10-8 to 5.0 × 10-6 mol L-1 with a detection limit of 2.0 × 10-9 mol L-1 (3σ). The relative standard deviation (n = 11) was 0.71% for 1.0 × 10-7 mol L-1 UA. Possible reaction mechanisms for the CL system were suggested based on the UV absorption spectra, CL spectra, and the free radical trapping experiment performed in this work.

Development of a derivatization method for the quantification of hydrogen sulfide and its application in vascular calcification rats

Hydrogen sulfide (H2S) plays major functional and structural roles in diverse physiological functions and the pathogenesis of a variety of disorders in biological matrices. The significance of H2S has prompted the development of sensitive and selective methods to determine its concentration in biological samples. The fluorescent reagent monobromobimane (MBB) has been widely used to measure various thiol-containing species through alkylation. MBB may prevent the oxidation of sulfide and the reaction of sulfide with several different species (such as superoxide radicals, hydrogen peroxide and peroxynitrite). An isomers of MBB, 3-(bromomethyl)-2, 6, 7-trimethyl-1H, 5H-pyrazolo [1,2-a] pyrazole-1, 5-dione (MMB), is cheaper than MBB and its use in the analysis of H2S has not previously been reported. In the present study, we compared the derivatization reactions of hydrogen sulfide with MMB and MBB and developed a sensitive method to quantify H2S in blood. In our method, H2S was incubated in the dark with excess MMB in 0.1M Tris-HCl buffer (pH 10.1) at 50°C for 120min. 50μL aliquots of the derivatized product were analyzed using HPLC system with gradient elution of 0.1% (v/v) formic acid-acetonitrile. The limit of detection for the derivatized product was 0.03nmol/mL. The derivatization reaction was suitable for detecting low concentrations of H2S. The derivate product is stable over time, permitting batch storage and analysis.