Cho-Chun Hu

Quantitation of branched‐chain amino acids in ascites by capillary electrophoresis with light‐emitting diode‐induced fluorescence detection

Branched‐chain amino acids (BCAAs) are one of the important biomarkers for monitoring liver disease such as hepatitis or hepatoma. In this communication, we present the determination of the concentrations of BCAA in ascites by CE light‐emitted diode‐induced fluorescence (LEDIF) using 1.5% m/v poly(ethylene oxide) (average Mv: ∼8 000 000 g/mol) that was prepared in 10 mM sodium tetraborate solution (pH 9.3). Naphthalene‐2,3‐dicarboxaldehyde was used to derivatize 15 amino acids (AAs) to form naphthalene‐2,3‐dicarboxaldehyde (NDA)‐AA derivatives prior to CE analysis. The separation of 15 NDA‐AA derivatives was accomplished within 15 min, with RSD values of <5.8% (within‐day) and 7.4% (between‐days) with respect to their migration times. The limits of detection for the tested BCAAs ranged from 10.6 to 10.9 nM. We determined the concentrations of three BCAAs – leucine, isoleucine and valine – in ascites by applying a standard addition method, with recovery percentages ranging from 93.9 to 111%. The results obtained from this CE‐LEDIF method is in good agreement with those by a gold standard method using an AA analyzer. We have found that the concentrations of the three BCAAs in ascites obtained from patients suffering from liver diseases were lower than those from healthy individuals. Our approach is highly efficient, sensitive, and cost‐effective, which holds great potential for the diagnosis of liver diseases.

Oxidized multiwalled carbon nanotubes decorated with silver nanoparticles for fluorometric detection of dimethoate

A novel method for the detection of dimethoate based on the peroxidase-like activity of silver-nanoparticles-modified oxidized multiwalled carbon nanotubes (AgNPs/oxMWCNTs) has been developed. The synthesized AgNPs/oxMWCNTs showed excellent peroxidease-like catalytic activity in hydrogen peroxide-Amplex red (AR) system (AR is oxidized to resorufinat, with the resorufin fluorescence at 584nm being used to monitor the catalytic activity). After dimethoate was added to AgNPs/oxMWCNTs, the interaction between dimethoate and the AgNPs inhibited the catalytic activity of AgNPs/oxMWCNTs. The decrease in fluorescence was used for the detection of dimethoate in the range of 0.01-0.35μgmL-1 (R2=0.998) with a detection limit of 0.003μgmL-1 (signal/noise=3). This method exhibited good selectivity for the detection of dimethoate even in the presence of high concentration of other pesticides. Consequently, the method was applied to measure the concentration of dimethoate residue in lake water and fruit, thus obtaining satisfactory results.

Using electrospray ionization mass spectrometry to explore the interactions among polythymine oligonucleotides, ethidium bromide, and mercury ions.

We have used electrospray ionization mass spectrometry (ESI-MS) and fluorescence and circular dichroism (CD) spectroscopy to explore the binding of ethidium bromide (EthBr) to non-self-complementary polythymine (polyT) strands in the absence and presence of Hg2+ ions. In the gas phase, ESI-MS revealed that Hg2+ ions have greater affinity, through T-Hg2+-T coordination, toward polyT strands than do other metal ions. These findings are consistent with our fluorescence and CD results obtained in solution; they revealed that more T33-EthBr-Hg2+ complexes existed upon increasing the concentrations of Hg2+ ions (from 0 to 50 µM). Surprisingly, the ESI-MS data indicated that the Hg2+ concentration dependence of the interaction between T33 and EthBr is biphasic. Our ESI-MS data revealed that the T33-EthBr-Hg2+ complexes formed with various stoichiometries depending on their relative concentrations of the components and the length of the DNA strand. When the concentrations of T33/EthBr/Hg2+ were 5/5/2. 5 µM and 5/10/7. 5 µM, 1:1:1 and 1:1:2 T33-EthBr-Hg2+ complexes were predominantly formed, respectively. Thus, Hg2+-induced DNA conformational changes clearly affect the interactions between DNA and EthBr.

Capillary electropherograms for restriction fragment length polymorphism of Helicobacter pylori.

Rapid identification of Helicobacter pylori strains is of importance for diagnosis and then treatment of duodenal and gastric ulcers. We developed a CE approach for the analysis of RFLP of the PCR products of urease (UreAB) gene and flagellin A (FlaA) gene fragments. Prior to CE analysis, the 2.4‐kbp UreAB and 1.5‐kbp FlaA PCR products were digested with the restriction enzymes HaeIII and HhaI, respectively. The DNA fragments were then separated by CE in conjunction with laser‐induced fluorescence detection using poly(ethylene oxide) in the presence of electroosmotic flow. The DNA fragments range in sizes 259–1831 bp and 12–827 bp for UreAB and FlaA restriction fragments, respectively. Of 27 samples, the CE approach provided five and ten different RFLP patterns of the HaeIII and HhaI digests. The RFLP of PCR products of the two genes allow great sensitivity of identification of H. pylori strains. When compared with slab gel electrophoresis, the present CE approach provides advantages of rapidity (within 6 min per run), simplicity, and automation. The preliminary results have shown great practicality of the CE approach for screening H. pylori strains.

Highly sensitive sensing of hydroquinone and catechol based on β-cyclodextrin-modified carbon dots

In the proposed study, an efficient method for a carbon dot@β-cyclodextrin (C-dot@β-CD)-based fluorescent probe was developed for the analyses of catechol (CC) and hydroquinone (HQ) at trace levels in water samples. The properties of C-dot@β-CD nanocomposites were characterized by Fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM) and X-ray photoelectron spectroscopy (XPS). The sensing behaviors of C-dot@β-CD toward CC and HQ were investigated by fluorescence spectroscopy. Based on the host–guest chemistry between C-dot@β-CD and phenolic compounds, which can quench C-dot@β-CD fluorescence, the prepared C-dot@β-CD nanocomposites could be used for the sensitive and selective detection of CC or HQ across a wide linear range (0.1 to 10 μM) with detection limits of 47.9 and 20.2 nM, respectively. These results showed that the synthesized C-dot@β-CD nanocomposite exhibited strong fluorescence and high degree of water solubility and thus, it is suitable for use as a nanoprobe for detecting CC or HQ in real water samples.

Gold-nanoparticle-based fluorescent “turn-on” sensor for selective and sensitive detection of dimethoate

A simple approach to fabricate a highly selective and sensitive dimethoate probe was developed based on Rhodamine B (RB)-functionalized gold nanoparticles (AuNPs). The quenching of RB fluorescence in the presence of AuNPs in the solution, mediated by fluorescence resonance energy transfer, was observed. In the presence of dimethoate, the fluorescence intensity of the RB-AuNP solution is gradually recovered when dimethoate molecules displace RB molecules on the surface of the AuNPs, which significantly increased the fluorescence intensity of RB. Fluorescence is proportional to the dimethoate concentration in the range of 0.005-1.0 ppm (R2 = 0.989), and the LOD was 0.004 ppm. The recoveries of dimethoate in water and fruit samples were 86-116% with a good RSD (< 9.3%). Because of its high sensitivity, excellent selectivity, and convenient fabrication process, this method is a promising candidate for dimethoate screening.

Analysis of steroid hormones in human saliva by matrix-assisted laser desorption/ionization mass spectrometry

This work describes a simple and rapid method for the quantitative analysis of four steroid hormones, including testosterone, progesterone, hydrocortisone and cortisone in human saliva samples by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry using nanomaterials as the matrix. The optimal pretreatment processes of saliva samples as well as MALDI conditions for efficient desorption/ionization of these steroid hormones are systematically explored. Parameters, including the concentration and pH level of ammonium citrate buffer, the nature and concentration of the MALDI matrix, laser energy, and the sample extraction method, were systematically investigated in this study. The limits of detection at a signal to noise ratio of 3 for testosterone, progesterone, hydrocortisone and cortisone provided by the proposed approach are 6.0, 6.0, 6.0, and 3.0 μM, respectively, with satisfactory quantitative linearity (R2 > 0.985) and recovery (89.3–111.2%) for the four analytes. Our technique provides the advantages of sensitivity, repeatability, speed, and simplicity for the quantitation of the four steroid hormones in saliva samples. This method holds great potential for high-throughput biological screening and disease detection.