Simple and Sensitive Assay for Nucleic Acids Using their Quenching Effect on the Chemiluminescence Reaction Between Luminol and Hydrogen Peroxide with Manganese-Tetrasulfonatophthalocyanine as a New Catalyst

The manganese-tetrasulfonatophthalo-cyanine (MnTSPc) catalyzed luminol-hydrogen peroxide chemiluminescence (CL) system can be quenched in the presence of nucleic acids. A new and highly sensitive CL quenching method for determining nucleic acids in aqueous solutions has been developed. Under optimum conditions, linear relationships were found between the quenched intensity of CL and the concentration of nucleic acids in the range 0.10–2.0µgmL^–1 for calf thymus DNA and 0–1.6µgmL^–1 for fish sperm DNA. The limits of detection were 14.8ngmL^–1 for calf thymus DNA and 21.7ngmL^–1 for fish sperm DNA. The relative standard deviation (RSD) of nine replicate measurements is 1.4% for 1.0µgmL^–1 calf thymus DNA. The method was applied to the analysis of nucleic acids in synthetic samples and the results are satisfactory.Received December 2, 2002; accepted June 2, 2003 published online September 1, 2003     

Selective Spectrophotometric Determination of Aluminium in the Presence of Beryllium and Lanthanide Cations

A spectrophotometric method for the selective determination of Al(III) in the presence of Be and Ln (lanthanide) cations is proposed. It is based on the selective reaction of SXO (Semi-Xylenol Orange) with Al(III) at pH 2.6. The presence of 8%v/v of 1,2-ethanediol serves to stabilize the chelate formed by heating at 100°C for 5min. 0.5mg each of Be(II), Ce(III), La(III), 4mg of Mn(II), 1.2mg of Pb(II), 1mg of Tl(I), 40mg each of Ca(II) and Mg(II) and 1.9mg of sodium dihydrogen phosphate are tolerable. A ligand buffer of HEDTA-Pb is incorporated to further enhance the selectivity of the color reaction. Under the specified conditions SXO reacts with Al(III) to form a 1:1 chelate. Its molar absorptivity at 526nm was found to be 3.3×10⁴Lmol^–1cm^–1. The linear regression equation for 2–20µg of Al(III) is A=0.04458C+0.0112 (where C stands for the concentration of Al(III), µg per 25mL) and correlation coefficient =0.9988. The RSD at the level of 10µg (n=10) and LOQ were found to be 3.5% and 2µg, respectively.     

Voltammetric Behavior of Dopamine at ct-DNA Modified Carbon Fiber Microelectrode

A sub-micrometer thin-layer DNA modified carbon fiber microcylinder electrode was prepared by electrodeposition of ct-DNA at 1.5V (vs. Ag/AgCl). The voltammetric behavior of dopamine (3-hydroxytyramine) was investigated at the modified electrode. It was found that the modified electrode exhibits a highly electrocatalytic activity toward dopamine oxidation. Differential pulse voltammetry was used for determination of dopamine in pH 7.4 phosphate buffer solution. A linear response of the peak current versus the concentration was found in the range of 4×10^–6 to 10^–4molL^–1 at 10^–4molL^–1 AA (ascorbic acid) coexistence (R=0.9959) and the range of 6×10^–5 to 10^–3molL^–1 at 10^–3molL^–1 AA (R=0.9960). The presence of a high concentration of ascorbic acid did not interfere with the determination. The proposed method exhibited good recovery and reproducibility. This method can be applied to the detection of DA in real samples.     

The Fluorescent Reaction Between Quinaldine Red and Nucleic Acids and its Application to Fluorescent Assay of DNA and RNA

The reaction between quinaldine red (QR) and nucleic acids was studied. The free QR alone has no fluorescence in solution. However, it becomes fluorescent after binding to nucleic acids, giving maximum emission at 607nm with maximum excitation at 557nm. Maximum fluorescence intensity is produced in the pH range of 3.2–3.6. Based on the fluorescent reactions, a novel fluorometric method was developed for rapid determination of nucleic acids using QR as the fluorescent probe. Under optimal conditions, the calibration graphs were linear in the range of 0–30.0µgmL^–1 for CT DNA and 0–20.0µgmL^–1 for yeast RNA. The limits of detection were 38ngmL^–1 for CT DNA and 142ngmL^–1 for yeast RNA. Four synthetic samples were determined with satisfaction.Received December 20, 2002; accepted March 27, 2003 Published online August 8, 2003     

Electrochemical Determination of Cinnamtannin B1 with a Pyrolytic Graphite Electrode

Cinnamtannin B1 (trimeric proanthocyanidin), which is identified and isolated from the effective fraction of the root of Lindera aggregata (Sims) Kosterm, is one kind of condensed tannin used as an effective antipyrotic and antitumor agent. Its electrochemical response can be obtained at a pyrolytic graphite electrode. Consequently, an easily performed and sensitive method for the determination of cinnamtannin B1 is developed. The detection limit is estimated to be 1.0×10^–7M with the linear determination range of 2.0×10^–7M to 1.8×10^–6M. Five replicate analyses of 1.0×10^–6M cinnamtannin B1 yields an RSD value of 2.1%. Since the working electrode does not need to be modified with any other species, it is very stable, repeatable and easily treated, and this method therefore potentially useful in real sample analysis.     

Determination of Nucleic Acids Based on Shifting the Association Equilibrium between a Heptamethine Cyanine Dye and Poly-Lysine

A near-infrared (near-IR) fluorescence recovery method for the determination of nucleic acids is presented. This method employs a two-reagent system composed of anionic heptamethines cyanine (HMC) and polycationic poly-lysine. The fluorescence of HMC, with maximum excitation and emission wavelengths at 778 and 804nm, respectively, was quenched by poly-lysine in proper concentration, but recovered by adding nucleic acids. Under optimal conditions, the recovered fluorescence was proportional to the concentration of nucleic acids. The calibration graphs are linear over the range of 5–300ngmL^–1 for herring sperm DNA (FS DNA), 2–100ngmL^–1 for calf thymus DNA (CT DNA) and 5–500ngmL^–1 for snake ovum RNA (SO RNA). The corresponding detection limits are 1.49ngmL^–1 for FS DNA, 0.7ngmL^–1 for CT DNA and 1.61ngmL^–1 for SO RNA, respectively. Four synthetic and three real nucleic acid samples were determined with satisfactory results.     

Solid-Phase Extraction with Slurry Injection of the Resin Into ETAAS for Trace Determination of Thallium in Natural Water

Thallium in natural water samples was determined by electrothermal atomic absorption spectrometry after 1000-fold enrichment by mini solid-phase extraction from a 100-mL sample solution. A Tl-pyrrolidine-1-carbodithioate complex formed in a sample solution of pH 1.6 was extracted on fine particles of a cellulose nitrate resin dispersed in the sample solution. The cellulose nitrate resin was then collected on a membrane filter (25mm^ø) by filtration under suction using a glass funnel with an effective filtration area of 0.64cm². As a result, a circular thin layer of the resin phase with a diameter of 9mm was obtained. Then the resin phase was carved out by an acrylate resin puncher with a 10-mm^ø hole to put it into a sample cup containing 100µL of 10mM HNO₃ containing 0.5mM NaCl. The resin phase was suspended in the solution by ultrasonication. 1000-fold enrichment was thus attained within 15min, and the suspension was delivered to electrothermal atomic absorption spectrometry. The linear calibration graph was obtained in the range of 0–4ng of Tl in 100mL of a sample solution. The detection limit obtained by 3 method was 0.19ng. The proposed method was applied to the determination of Tl in natural water samples. The results showed the concentration of Tl in seawater was 12.1±1.8pgmL^–1 for the calibration graph method and 12.6±1.4pgmL^–1 for the standard addition method. A snowmelt sample contained 20.7±1.0pgmL^–1 of Tl.     

Fluorimetric Determination of Europium Based on the Co-Fluorescence System Eu-Y-Sparfloxacin-SDS

A new room temperature fluorescence enhancement system based on complex formation between europium and sparfloxacin in the presence of yttrium in sodium dodecyl sulfate micelle solution has been discovered. The characteristic fluorescence spectra and the effect factors of the fluorescence intensity of the system have been studied in detail. The study results show that the binary complex formed by europium with sparfloxacin in sodium dodecyl sulfate micelle solution can emit intrinsic fluorescence of the lanthanide ion. In the presence of yttrium, the fluorescence intensity of this system was enhanced about 5-fold compared with that of the system without Y³⁺. Under the optimum conditions selected, the fluorescence intensity is a linear function of the concentration of europium in the range of 5.0×10^–11–5.0×10^–7molL^–1, and the detection limit of europium is 1.0×10^–13molL^–1. The system was used for the determination of trace amounts of europium in rare earth samples with satisfactory results.     

Nickel Ion-Selective PVC Membrane Electrode Based on a New t-Octyl-Calix[6]arene Derivative

This study presents a PVC membrane based on 5,11,17,23,29,35-Hexakis-t-octyl-37,38,39,40,41,42-hexakis(N-phenylthiocarbamoylmethoxy)calix[6]arene which is used as an electroactive substance. The electrode reveals a Nernstian response for Ni²⁺ over a wide pH range with a slope of linear portion (5×10^–6–1×10^–2M, R=0.988) of 29.75±0.2mV·decade^–1 at 25°C. The electrodes sensitivity is enhanced upon the introduction of sodium tetraphenylborate into the membrane as a negatively charged lipophilic additive. The selectivity coefficients of various interfering ions were determined using the fixed interference method (FIM). The membrane reveals good selectivity for nickel ions over the other metal ions investigated. The lifetime of this electrode is about one month. This electrode has been applied to the determination of nickel ions in wastewater from the electroplating industry.     

Antioxidative and antitumour properties of metal(II) solid complexes with 8-acetyl-4-methyl umbelliferone

Six transition metal(II) complexes with 8-acetyl-4-methyl umbelliferone (AMUH), ML2 (L=AMU^–; M=Mn, Co, Ni, Cu, Zn and Cd), have been synthesized and characterized by elemental analysis, electrical conductance, i.r., 1H-n.m.r. and u.v. spectroscopic techniques. The e.s.r. spectra show that CuL2 is anisotropic, with g2.05 and g2.26. The MnL2 species is characterized by a very broad g2 centered resonance without manganese hyperfine structure. The antioxidative and other biological activities of selected complexes were also investigated, indicating that both the ligand and complexes exhibit a scavenging effect on the superoxide radical (O₂ ^–·) and a suppressing effect on the hydroxyl radical (·OH). The suppressing effect on ·OH is greater than the scavenging effect on O₂ ^–·. The Cu and Mn complexes exhibit very significant suppression ratios for both radicals. However, the inhibitory effect on lipid peroxides results show that, in the initial stage, the Cu and Mn complexes exhibit obvious inhibitory effects on lipid peroxides, but after one hour, they begin to accelerate the lipid peroxides. Lower catalytic activity and instability when the complexes react with active oxygen may be responsible for this dual nature. The ligand and four transition metal(II) complexes selectively, obviously, inhibit the growth of HCT-8 and HL-60 tumour cell lines. They also exhibit a minor influence on the proliferation of B cells in higher concentration (10^–5m), but only a weak effect on the proliferation of T cells of the BALB/C (nude rate) spleen cells.     

Application of L-Cysteine-Capped ZnS Nanoparticles in the Determination of Nucleic Acids Using the Resonance Light Scattering Method

Nanometer-sized L-cysteine-capped ZnS particles were synthesized by a colloidal aqueous method. The functionalized nanoparticles are water-soluble and suitable for biological applications. In Tris-HCl buffer solution, nucleic acids combine with cysteine-capped nano-ZnS particles by intermolecular forces to form larger nanoparticles. There are two resonance light scattering peaks at 304.5nm and 373.8nm, respectively. The enhanced RLS is related to the concentration of nucleic acids in the range of 0.04 to 1.2µgmL^–1 for calf thymus DNA and 0.2 to 1.0µgmL^–1 for fish sperm DNA. The detection limits (3) are 19ngmL^–1 for calf thymus DNA and 23ngmL^–1 for fish sperm DNA, respectively. Four synthetic samples were analyzed satisfactorily.     

The Electrochemical Behavior of p-Aminophenol at a ω-Mercaptopropionic Acid Self-Assembled Gold Electrode

A -mercaptopropionic acid (MPA) self-assembled monolayer modified electrode (MPA/SAM/Au) on a gold electrode has been fabricated. The characterization of the MPA/SAM/Au was investigated using attenuated total reflection-fourier transform infrared (ATR-FTIR) and A.C. impedance. The electrochemical behaviors of p-aminophenol (p-AP) were studied at the MPA/SAM/Au by cyclic voltammetry and semi-derivative voltammetry (SDV) in BR buffer solution. The modified electrode shows excellent electrocatalytic activity for the redox of p-AP and accelerates the electron transfer rate. The diffusion coefficient (D) is 4.55×10^–6cm²s^–1. The oxidative peak current increases linearly with the concentration of p-AP in the range of 4.0×10^–88×10^–6molL^–1 and 1.0×10^–52×10^–4molL^–1 by square wave voltammetry response, respectively. The detection limit (three times the signal blank/slope) is up to 1.2×10^–8molL^–1. The modified electrode is able to eliminate the interference of p-benzenediol, o-benzenediol and o-AP at a 40-, 90- or 70-fold concentration of p-AP, and it has been satisfactorily used for the determination of the real sample.     

Determination of Nucleic Acids Based on their Resonance Light Scattering Enhancement Effect on Metalloporphyrin Derivatives

A new method for the determination of nucleic acids at nanogram per mL level is proposed based on the enhanced resonance light scattering (RLS) signal resulting from the interaction of metalloporphyrins with nucleic acids. Under optimum conditions, the weak RLS signal of metalloporphyrin is enhanced by nucleic acids, and the enhanced RLS intensity is proportional to the concentration of nucleic acids. The detection limits of calf thymus DNA were 3.5ngmL^–1, 2.9ngmL^–1 and 1.0ngmL^–1 for three metalloporphyrins, respectively. Synthetic samples were determined with satisfactory results.     

Development of an FI-ICP Method for On-Line Preconcentration and Determination of Platinum

This paper presents a new simple and rapid procedure for the preconcentration and determination of platinum. It is based on the adsorption of the metal ion and preconcentration on a micro-column (3cm×3mm) placed in the injection valve of a flow injection (FI) manifold and packed with 1,5-bis[(2-pyridyl)-3-sulphophenyl-methylene]thiocarbonohydrazide (PSTH) immobilised on an anion-exchange resin (Dowex 1X8-200). The metal was eluted from the column using a solution of 2M HNO₃. Various parameters and chemical variables affecting the preconcentration and determination of this metal by ICP-AES were evaluated. Five variables (sample flow rate, eluent flow rate, nebulizer flow rate, buffer concentration and mixing coil length) were considered as factors in the optimisation process. Interactions between analytical factors, and their optimal levels were investigated using two level factorial and central matrix designs. The optimum conditions established were applied to the determination of platinum by flow injection inductively coupled plasma atomic emission spectrometry (FI-ICP-AES). The method has a linear calibration range of 25 to at least 200ngmL^–1 with a detection limit of 7.4ngmL^–1 (S/N=3) and a throughput of 10 samples h^–1 using 5min. preconcentration time. The precision of the method (RSD) was 3.06% ngmL^–1 at the 50ngmL^–1 level of Pt(IV) and 2.93% at the 150ngmL^–1 level. The accuracy of the method was examined by determining the analyte content in spiked waters and by analysing an automobile catalyst standard reference material. The results show good agreement with the certified value and sufficiently high recoveries.     

Determination of Trace Iron in KH2PO4 Crystal by Derivative Adsorption Chronopotentiometry

A sensitive derivative adsorption chronopotentiometric method was developed to determine trace amounts of Fe³⁺. The influence of preconcentration time, pH of solution and constant reducing current of the peak height is discussed. The detection limit and the linear range are 5.0×10^–10 and 2.0×10^–93.0×10^–6molL^–1, respectively. The method was used to determine trace Fe³⁺ in KH₂PO₄ crystals (KDP).     

The Exciton Model and the Circular Dichroism of Polypeptides

Summary. Exciton coupling of the 190nm ^* transition is an important factor in the CD spectrum of peptides and proteins. The CD spectrum of the -helix is dominated by the exciton effect. The spectrum is sensitive to the direction of the ^* transition dipole moment, especially for short helices. Exciton theory is much less successful in accounting for the CD spectrum of the poly(proline)II (PPII) conformation, an important conformer in collagen and in unordered peptides. Mixing of the ^* transition with high-energy transitions in the peptide backbone and in side chains must be considered to explain the strong negative CD band near 200nm of PPII.     

Solid-Phase Extraction and Spectrophotometric Determination of Trace Amounts of Mercury in Natural Samples

A sensitive and selective solid phase spectrophotometric method for the determination of trace amounts of inorganic mercury is described. Hg²⁺ was sorbed on a silica gel-packed column as an Hg²⁺–N,N-bis(2-mercaptophenyl)ethanediamide (H₂L) complex. The Hg²⁺ complex was eluted from the column using 7mL of acetone. Various parameters including pH, column flow rate, and ligand concentration were optimized. The complex was found to obey Beers law from 2.3 to 73.7µgmL^–1 within the optimum range when the preconcentration factor was two. The effective molar absorption coefficient at 523nm was 1.17×10³Lmol^–1cm^–1 at 523nm. The concentration limits in Beers law dropped from 0.09 to 2.95µgmL^–1 within the optimum range when the preconcentration factor was 50. The relative standard deviation at a concentration level of 5µgmL^–1 Hg²⁺ (9 repetitive determinations) was 1.6%. The detection limits are 0.34µgmL^–1 and 0.015µgmL^–1 when the preconcentration factors are 2 and 50, respectively. The method has been used for routine determination of trace levels of Hg²⁺ in natural waters. The potential application of this method for the removal of Hg²⁺ from natural samples (sea water and lake water) spiked with 100ngmL^–1 of Hg²⁺ was studied. In order to validate the proposed method, LGC 6156 (harbour sediment – extractable metals) was analysed by this method. The results proved that excellent extraction of Hg²⁺ from both natural water samples was obtained by solid phase extraction using N,N-bis(2-mercaptophenyl) ethanediamide.     

Determination of Attogram Quantities of Silver via Quenching of the Solid Substrate Room-Temperature Phosphorescence of Rhodamine B Based on the Catalysis of its Oxidation by Potassium Persulfate

A new method of SS-RTP for the determination of trace silver has been established. This method is based on the fact that Ag⁺, when activated by ,-bipyridyl (bipy) in a media of HAc–NaAc (pH=4.9), can catalyze the reaction of Rhodamine B (RhoB) oxidized by K₂S₂O₈, thus causing the Solid Substrate Room-Temperature Phosphorescence (SS-RTP) of RhoB to be quenched. The activating efficiency of bipy is 6.7 times higher than that of o-phenanthroline (phen). The reduction of the phosphorescence intensity (I_p) of RhoB is directly proportional to the concentration of Ag⁺ ions in the range of 1.6016.0agspot^–1 (0.40µLspot^–1). The regression equation of the working curve can be expressed as I_p=18.78+5.100m_Ag+ (agspot^–1) (r=0.9994, n=6), the detection limit is 0.28agspot^–1. This rapid, accurate and sensitive method has been successfully applied to the determination of trace silver in tea and human hair samples, and the results agree well with the Atomic Absorption Spectroscopy (AAS) method. The mechanism of the catalyzing reaction is also discussed.     

The Dissociation Constants of 1,1′-Bis (pyridinium-4-aldoxime)trimethylene dibromide

Summary. The dissociation constants of the two oxime groups of 1,1-bis(pyridinium-4-aldoxime)trimethylene dibromide (TMB-4) were determined using spectrophotometric data. Two numerical methods were applied to treat the overlapping equilibria. The results obtained by both agreed with each other and their mean values at 25°C corrected for the ionic strength of 0.05moldm^–3 are pK_a1=7.49±0.11 and pK_a2=8.96±0.09. These values were discussed in terms of the pK_as of 1,1-bis(pyridinium-4-aldoxime)oxydimethylene dichloride (Toxogonin), a similar dioxime, which were derived by extrapolation of literature data.     

Determination of Heavy Metal Ions in Chinese Herbal Medicine by Microwave Digestion and RP-HPLC with UV-Vis Detection

A new method for the simultaneous determination of heavy metal ions in Chinese herbal medicine by microwave digestion and reversed-phase high-performance liquid chromatography (RP-HPLC) has been developed. The Chinese herbal medicine samples were digested by microwave digestion. Lead, cadmium, mercury, nickel, copper, zinc, and tin ions in the digested samples were pre-column derivatized with tetra-(4-chlorophenyl)-porphyrin (T₄-CPP) to form the colored chelates which were then enriched by solid phase extraction with C₁₈ cartridge and eluted from the cartridge with tetrahydrofuran (THF). The chelates were separated on a Waters Xterra RP₁₈ column by gradient elution with methanol (containing 0.05molL^–1 pyrrolidine-acetic acid buffer salt, pH=10.0) and THF (containing 0.05molL^–1 pyrrolidine-acetic acid buffer salt, pH=10.0) as mobile phase at a flow rate of 0.5mLmin^–1 and detected with a photodiode array detector in the range of 350–600nm. In the original samples the detection limits of lead, cadmium, mercury, nickel, copper, zinc and tin are 4ngL^–1, 3ngL^–1, 6ngL^–1, 5ngL^–1, 2ngL^–1, 6ngL^–1, and 4ngL^–1, respectively. This method was applied to the determination of lead, cadmium, mercury, nickel, copper, zinc and tin in Chinese herbal medicine samples with good results.