Studies on the interaction between imidacloprid and human serum albumin: Spectroscopic approach

The interaction between imidacloprid (IMI) and human serum albumin (HSA) was investigated using fluorescence and UV/vis absorption spectroscopy. The experimental results showed that the fluorescence quenching of HSA by IMI was a result of the formation of IMI–HSA complex; static quenching was confirmed to result in the fluorescence quenching. The apparent binding constant K_A between IMI and HSA at three differences were obtained to be 1.51 × 10⁴, 1.58 × 10⁴, and 2.19 × 10⁴ L mol^?1, respectively. The thermodynamic parameters, ΔH° and ΔS° were estimated to be 28.44 kJ mol^?1, 174.76 J mol^?1 K^?1 according to the van’t Hoff equation. Hydrophobic interactions played a major role in stabilizing the complex. The distance r between donor (HSA) and acceptor (IMI) was obtained according to fluorescence resonance energy transfer. The effect of IMI on the conformation of HSA was analyzed using synchronous fluorescence spectroscopy CD and three-dimensional fluorescence spectra, the environment around Trp and Tyr residues were altered.     

Novel amphiphilic triblock copolymer based on PPDO,PCL, and PEG: Synthesis,characterization, and aqueous dispersion

Amphiphilic triblock copolymer, poly(p-dioxanone-co-caprolactone)-block-poly(ethylene oxide)-block-poly(p-dioxanone-co-caprolactone) (PPDO-co-PCL-b-PEO-b-PPDO-co-PCL) was synthesized by ring opening polymerization (ROP) of p-dioxanone and ɛ-caprolactone initiated through the hydroxyl end of poly(ethylene glycol) (PEG) in the presence of stannous 2-ethyl hexanoate [Sn(oct)₂] as a catalyst. Polymerization and structural features of the polymers were analyzed by different physicochemical techniques (GPC, ¹H NMR, ¹³C NMR, FT-IR, DSC and TGA). The splitting of ¹H NMR resonance at δ 2.3 and δ 4.1 ppm reveals the random copolymerization. Polymeric nanoparticles were prepared in phosphate buffer (pH 7.4) by co-solvent evaporation technique at room temperature (25 °C). Existence of hydrophobic domains as cores of the micelles were characterized by ¹H NMR spectroscopy and further confirmed with fluorescence technique using pyrene as a probe. Critical micelle concentration (CMC) of the polymer in phosphate buffer (pH. 7.4) was decreased from 2.3 × 10⁻³ to 7.6 × 10⁻⁴ g/L with the fraction of PCL. Polymeric nanoparticles observed by atomic force microscopy (AFM) were uniform and spherical, with smooth textured of around 50–30 nm diameter. Dynamic light scattering (DLS) and electrophoretic light scattering (ELS) measurements showed a monodisperse size distribution of around 113–90 nm hydrodynamic diameters and negative zeta (ζ) potential (−4 to −14 mV), respectively. The investigations for the polymeric nanoparticles in aqueous medium showed that the composition of the hydrophobic segment of amphiphilic block copolymer makes a significant influence on its physicochemical characteristics.     

The role of water in the thermodynamics of drug binding to cyclodextrin

The thermodynamic parameters, Δ_BG^∘, Δ_BH^∘, Δ_BS^∘, and Δ_BC_p, of the drugs flurbiprofen (FLP), nabumetone (NAB), and naproxen (NPX) binding to β-cyclodextrin (βCD) and to γ-cyclodextrin (γCD) in 0.10 M sodium phosphate buffer were determined from isothermal titration calorimetry (ITC) measurements over the temperature range from 293.15 K to 313.15 K. The heat capacity changes for the binding reactions ranged from −(362 ± 48) J · mol⁻¹ · K⁻¹ for FLP and −(238 ± 90) J · mol⁻¹ · K⁻¹ for NAB binding in the βCD cavity to 0 for FLP and −(25.1 ± 9.2) J · mol⁻¹ · K⁻¹ for NPX binding in the larger γCD cavity, implying that the structure of water is reorganized in the βCD binding reactions but not reorganized in the γCD binding reactions. Comparison of the fluorescence enhancements of FLP and NAB upon transferring from the aqueous buffer to isopropanol with the maximum fluorescence enhancements observed for their βCD binding reactions indicated that some localized water was retained in the FLP–βCD complex and almost none in the NAB–βCD complex. No fluorescence change occurs with drug binding in the larger γCD cavity, indicating the retention of the bulk water environment in the drug–γCD complex. Since the specific drug binding interactions are essentially the same for βCD and γCD, these differences in the retention of bulk water may account for the enthalpically driven nature of the βCD binding reactions and the entropically driven nature of the γCD binding reactions.     

Fluorescence and DNA-binding spectral studies of neodymium(III) complex containing 2,2′-bipyridine, [Nd(bpy)2Cl3·OH2]

The interaction of [Nd(bpy)₂Cl₃·OH₂], where bipy is 2,2′-bipyridine, with DNA has been studied by absorption, emission, and viscosity measurements. [Nd(bpy)₂Cl₃·OH₂] showed absorption decreasing in charge transfer band with increasing of DNA. The binding constant, K_b has been determined by absorption measurement and found to be (1.5 ± 0.1) × 10⁵ M^?1. The fluorescent of [Nd(bpy)₂Cl₃·OH₂] has been investigated in detail. The interaction was also studied by fluorescence quenching technique. The results of fluorescence titration revealed that DNA had the strong ability to quenching the intrinsic fluorescence of Nd(III) complex at 327 nm. The binding site number n, apparent binding constant K_b and the Stern–Volmer quenching constant K_SV have been determined. Thermodynamic parameters have been calculated according to relevant fluorescent data and Van’t Hoff equation. Characterization of bonding mode has been studied. The results suggested that the major interaction mode between [Nd(bpy)₂Cl₃·OH₂] and DNA was groove binding.     

An enzymatic glucose/O2 biofuel cell: Preparation,characterization and performance in serum

This study demonstrates a new kind of single-walled carbon nanotubes (SWNT)-based compartment-less glucose/O₂ biofuel cell (BFC) with glucose dehydrogenase (GDH) and bilirubin oxidase (BOD) as the anodic and cathodic biocatalysts, respectively, and with poly(brilliant creysl blue) (BCB) adsorbed onto SWNT nanocomposite as the electrocatalyst for the oxidation of NADH. The prepared GDH-polyBCB-SWNT bioanode exhibits an excellent electrocatalytic activity toward the oxidation of glucose biofuel; in 0.10 M phosphate buffer containing 20 mM NAD⁺ and 100 mM glucose, the oxidation of glucose commences at −0.25 V and the current reaches its maximum of 310 μA/cm² at −0.05 V vs. Ag/AgCl. At the BOD-SWNT biocathode, a high potential output is achieved for the reduction of O₂ due to the direct electron transfer property of BOD at the SWNTs. In 0.10 M phosphate buffer, the electrocatalytic reduction of O₂ is observed at a high potential of 0.53 V vs. Ag/AgCl with an electrocatalytic current plateau of ca. 28 μA/cm² at 0.45 V under ambient air and ca. 102 μA/cm² under O₂-saturated atmosphere. In 0.10 M phosphate buffer containing 10 mM NAD⁺ and 40 mM glucose under O₂-saturated atmosphere, the power density of the assembled SWNT-based glucose/O₂ BFC reaches 53.9 μW/cm² at 0.50 V. The performance and the stability of the glucose/O₂ BFC are also evaluated in serum. This study could offer a new route to the development of new kinds of enzymatic BFCs with a high performance and provide useful information on future studies on the enzymatic BFCs as in vivo power sources.     

Electroreduction of O2 at a mediated Melanocarpus albomyces laccase cathode in a physiological buffer

We report on oxygen reduction in a physiological buffer solution (0.05 M phosphate buffer containing dissolved O₂, 0.1 M NaCl, pH 7.4, 37 °C) by Melanocarpus albomyces laccase, co-immobilized with [Os(2,2’-bipyridine)₂(polyvinylimidazole)₁₀Cl]^+/2+ as a mediator, on glassy carbon electrodes. Such oxygen cathodes yielded current densities of 3.8 mA cm⁻² at 0.2 V vs. Ag/AgCl, the largest current density reported to date for a mediated laccase cathode in physiological buffer solutions, showing promise for development of biocatalytic fuel cell prototypes.     

A simple and sensitive spectrophotometric method for the determination of trace amounts of nitrite in environmental and biological samples using 4-amino-5-hydroxynaphthalene-2,7-disulphonic acid monosodium salt

A very simple, sensitive, fairly selective and rapid spectrophotometric method for the determination of trace amounts of nitrite has been described. This method is based on the diazotized intramolecular coupling of electrophilic diazonium cation with the phenolic group of 4-amino-5-hydroxynaphthalene-2,7-disulphonic acid monosodium salt (AHNDMS) in a phosphate buffer solution of pH 7.5. The cyclic product has a purple color with maximum absorbance at 560 nm and is stable for 6 h. Optimum reaction conditions and other important analytical parameters for the maximum color development were established. Beer's law was found to obey for nitrite in the concentration range of 0.1–1.6 μg ml^?1 with molar absorptivity of 2.6 × 10⁴ l mol^?1 cm^?1 and Sandell's sensitivity of 0.0075 μg ml^?1. The effect of interfering ions on the determination is described. The recommended method was applied for the determination of nitrite in different water, soil and human saliva samples. The performance of the recommended method was evaluated in terms of Student's t-test and variance ratio F-test, which indicated the significance of proposed method over the reference method.     

Study on the interaction of 3,3-bis(4-hydroxy-1-naphthyl)-phthalide with bovine serum albumin by fluorescence spectroscopy

The interaction between 3,3-bis(4-hydroxy-1-naphthyl)-phthalide (NPP) and bovine serum albumin (BSA) have been studied by fluorescence spectroscopy. The binding of NPP quenches the BSA fluorescence. By the fluorescence quenching results, it was found that the binding constant K = 5.30 × 10⁴ L mol⁻¹, and number of binding sites n = 0.9267. In addition, according to the synchronous fluorescence spectra of BSA, the results showed that the fluorescence spectra of BSA mainly originate from the tryptophan residues. Finally, the distance between the acceptor NPP and BSA was estimated to be 1.94 nm using Föster's equation on the basis of fluorescence energy transfer. The interaction between NPP and BSA has been verified as consistent with the static quenching procedure and the quenching mechanism is related to the energy transfer.     

Binding of the bioflavonoid robinetin with model membranes and hemoglobin: Inhibition of lipid peroxidation and protein glycosylation

Recent years have witnessed burgeoning interest in plant flavonoids as novel therapeutic drugs targeting cellular membranes and proteins. Motivated by this scenario, we explored the binding of robinetin (3,7,3′,4′,5′-pentahydroxyflavone, a bioflavonoid with remarkable ‘two color’ intrinsic fluorescence properties), with egg yolk phosphatidylcholine (EYPC) liposomes and normal human hemoglobin (HbA), using steady state and time resolved fluorescence spectroscopy. Distinctive fluorescence signatures obtained for robinetin indicate its partitioning (K_p = 8.65 × 10⁴) into the hydrophobic core of the membrane lipid bilayer. HbA–robinetin interaction was examined using both robinetin fluorescence and flavonoid-induced quenching of the protein tryptophan fluorescence. Specific interaction with HbA was confirmed from three lines of evidence: (a) bimolecular quenching constant K_q ? diffusion controlled limit; (b) closely matched values of Stern–Volmer quenching constant and binding constant; (c) τ₀/τ = 1 (where τ₀ and τ are the unquenched and quenched tryptophan fluorescence lifetimes, respectively). Absorption spectrophotometric assays reveal that robinetin inhibits EYPC membrane lipid peroxidation and HbA glycosylation with high efficiency.     

Electronic structures,photophysical properties,and electrochemistry of ruthenium(II)(bpy)2 pyridylimidazole complexes

The properties of Ru^II complexes involving the imidazole moiety are discussed. Complexes [Ru(bpy)₂(L)]²⁺ [bpy = 2,2′-bipyridine, L = 2-(2′-pyridyl)imidazole (2-pimH) and 4-(2′-pyridyl)imidazole (4-pimH)] have been synthesized and fully characterized. Reduction potentials are 0.76 V vs. Fc⁺/Fc⁰ for both complexes in acetonitrile solution, and the deprotonated complexes undergo irreversible electrochemical oxidation at 0.38 V vs. Fc⁺/Fc⁰. Density functional theory (DFT) calculations suggest that oxidation of the protonated complexes is primarily metal-based and that of the deprotonated complexes is ligand-centered. The pK_a of the 4-pimH complex was found to be 9.7 ± 0.2; the pK_a of the 2-pimH complex is 7.9 ± 0.2. Luminescence lifetimes (L = 4-pimH, 277 ns; 2-pimH, 224 ns; 4pim^?, 40 ns; 2pim^?, 34 ns in 5% methanol/water solution) combined with quantum yield data and acid–base behavior suggest that the non-coordinated imidazole nitrogen tunes deactivation pathways.     

Buffer standards for the physiological pH of the zwitterionic compound of 3-(N-morpholino)propanesulfonic acid (MOPS) from T = (278.15 to 328.15) K

This paper reports the pH values of five NaCl-free buffer solutions and 11 buffer compositions containing NaCl at I = 0.16 mol · kg⁻¹. Conventional pa_H values are reported for 16 buffer solutions with and without NaCl salt. The operational pH values have been calculated for five buffer solutions and are recommended as pH standards at T = (298.15 and 310.15) K after correcting the liquid junction potentials. For buffer solutions with the composition m₁ = 0.04 mol · kg⁻¹, m₂ = 0.08 mol · kg⁻¹, m₃ = 0.08 mol · kg⁻¹ at I = 0.16 mol · kg⁻¹, the pH at 310.15 K is 7.269, which is close to 7.407, the pH of blood serum. It is recommended as a pH standard for biological specimens.     

Buffer standards for the physiological pH of N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine (TRICINE) from T = (278.15 to 328.15) K

The pH values of two buffer solutions without NaCl and seven buffer solutions with added NaCl, having ionic strengths (I = 0.16 mol · kg⁻¹) similar to those of physiological fluids, have been evaluated at 12 temperatures from T = (278.15 to 328.15) K by way of the extended form of the Debye–Hückel equation of the Bates–Guggenheim convention. The residual liquid junction potentials (δE_j) between the buffer solutions of TRICINE and saturated KCl solution of the calomel electrode at T = (298.15 and 310.15) K have been estimated by measurement with a flowing junction cell. For the buffer solutions with the molality of TRICINE(m₁) = 0.06 mol · kg⁻¹, NaTRICINE(m₂) = 0.02 mol · kg⁻¹, and NaCl(m₃) = 0.14 mol · kg⁻¹, the pH values at T = 310.15 K obtained from the extended Debye–Hückel equation and the inclusion of the liquid junction correction are 7.342 and 7.342, respectively. These are in excellent agreement. The zwitterionic buffer TRICINE is recommended as a secondary pH standard in the region for clinical application.     

Absorption and emission spectroscopic characterisation of the LOV2-His domain of phot from Chlamydomonas reinhardtii

The absorption and emission behaviour of flavin mononucleotide (FMN) in the wild-type light, oxygen and voltage sensitive domain LOV2 of the photoreceptor phot from the green alga Chlamydomonas reinhardtii is studied. Actually a LOV2-His protein (LOV2 domain bound at N-terminal to 15 His aminoacids via a Gly aminoacid) expressed in an Escherichia coli strain is investigated. For fresh samples stored in the dark an initial fluorescence quantum yield of ϕ_F = 0.12 ± 0.01 and an effective fluorescence lifetime of τ_F = 2.4 ± 0.1 ns are determined. Blue-light photo-excitation generates an intermediate photoproduct (flavin-C(4a)-cysteinyl adduct with absorption peak at 390 nm) resulting in an intensity-dependent fluorescence quenching. In the aqueous solutions at pH 8 approximately 3.8% of the FMN molecules are not bound to the protein binding pocket, whereas 96.2% are non-covalently bound. Even at high-intensity light excitation at 428 nm a fraction of about 7% of the non-covalently bound FMN remains non-converted to an FMN-Cys adduct because of photo-induced back-relaxation of the adduct to non-covalently bound FMN. Two holo-LOV2-His conformations with different adduct recovery time constants are revealed by spectrally and temporally resolved fluorescence and absorption measurements: A fraction of about 48% forms FMN-Cys adducts with a fast recovery time constant of τ_Ad,f = 19 ± 2 s in the dark, and the rest forms adducts with a slow recovery time constant of τ_F,s = 5.5 ± 1 min. Prolonged blue light irradiation of the flavin-C(4a)-cysteinyl adducts reduces their ability to recover back in the dark to non-covalently bound FMN (photo-induced permanent adduct formation). Numerical simulations of the intensity-dependent absorption depletion reveals a quantum yield of intermediate photo-adduct formation of ϕ_Ad = 0.9 ± 0.1. Simulation of the adduct absorption dynamics gives a quantum yield of photo-induced adduct back-relaxation of ϕ_Ad,b = 0.15 ± 0.01 and a quantum yield of photo-induced permanent adduct formation of ϕ_Ad,p = (2.6 ± 0.5) × 10⁻⁴.     

Spontaneous formation of an electroactive co-polymeric film derived from nordihydroguaiaretic acid and 4,4′-diaminobibenzyl

Binary solution of nordihydroguaiaretic acid (NDGA) and 4,4′-diaminobibenzyl (DABB) undergoes rapid oxidation by ambient oxygen to form a thin film of poly-NDGA-co-DABB on the surface of the reaction chamber and on immersed substrates. Electrochemistry of thus formed films was studied in 0.1 M sulfuric acid and in phosphate buffer (pH 7.4). Electrochemical behavior of the co-polymeric film is characterized by two redox couples, the predominant one being observed at more negative potentials comparing to parent NDGA i.e. 0.28 vs. 0.49 V (vs. Ag/AgCl) in 0.5 M H₂SO₄. The peak potentials were shifted toward lower values with solution pH at the rate of 59 mV/pH unit indicating a 2e/2H⁺ transition as expected for quinone-containing films. The poly-NDGA-co-DABB film exhibits catalytic activity toward electroreduction of nitrite to nitric oxide in acidic electrolytes. This reaction can be used to quantify nitrite in a broad concentration range with low detection limit (0.3 μM, S/N = 3).     

Thermochemistry of the biochemical reaction: { pyrophosphate(aq) + H2O(l) = 2 phosphate(aq)}

Calorimetric enthalpies of reaction have been measured for the overall biochemical reaction{pyrophosphate(aq)  +  H₂O(l)  =  2phosphate (aq)} . The reaction was catalyzed by alkaline phosphatase and, to simplify the thermochemistry, was carried out in the absence of Mg ^2 + (aq). Measurements were performed with phosphate buffer ( pH  =  7.19 and 7.94), PIPES buffer ( pH  =  7.13), and HEPES buffer ( pH  =  7.86). The results of these measurements were analyzed by using an equilibrium model. These calculations lead to the standard molar enthalpy changeΔ_rH_m^o =  − (17.3  ±  0.6)kJ·mol ^− 1 (temperature T =  298.15 K and ionic strengthI =  0) for the reference reaction{HP₂O₇^3 − (aq)  +  H₂O(l)  =  2HPO₄^2 − (aq)  +  H ⁺ (aq)} . Values of the apparent equilibrium constantK^′ for the overall biochemical reaction from the literature were also analyzed by using the equilibrium model in order to obtain what is believed to be a reliable value for the equilibrium constantK =  4.7 · 10 ^− 4 for the reference reaction. The values ofK and Δ_rH_m^o for the reference reaction have been used together with values from the CODATA tables to calculate standard molar formation properties for the pyrophosphate species.     

Thermochemistry of uracil and thymine revisited

Thermochemical properties of uracil and thymine have been evaluated using additional experiments. Standard (p⁰ = 0.1 MPa) molar enthalpies of formation in the gas phase at T = 298.15 K for uracil −(298.1 ± 0.6) and for thymine −(337.6 ± 0.9) kJ · mol⁻¹ have been derived from energies of combustion measured by static bomb combustion calorimetry and molar enthalpies of sublimation determined using the transpiration method. The G3 and G4 quantum-chemical methods were used for calculations of theoretical gaseous enthalpies of formation being in very good agreement with the re-measured experimental values.     

Steady-state and laser flash photolysis studies on photochemical formation of 4-tert-butyl-4′-methoxydibenzoylmethane from its derivative via the Norrish Type II reaction in solution

A photochemical formation process of avobenzone (AB; 4-tert-butyl-4′-methoxydibenzoylmethane) from 1,1-(4-tert-butybenzoyl)(4′-methoxybenzoyl)butane (PrAB) is studied by steady-state and laser flash photolysis in solution. The quantum yield of the formation via the triplet state of PrAB is determined to be 0.23 in degassed acetonitrile at 295 K. The Arrhenius plots of the decay rate of triplet PrAB show that photoelimination proceeds with an activation energy of 6.0 kcal mol^?1 and the frequency factor of 4.6 × 10¹⁰ s^?1.     

Photochemical vapor generation of carbonyl for ultrasensitive atomic fluorescence spectrometric determination of cobalt

UV photochemical vapor generation (photo-CVG) as sample introduction was first adapted for determination of ultratrace cobalt by atomic fluorescence spectrometry (AFS). Cobalt volatile species can be generated when the buffer system of formic acid and formate containing Co (II) is exposed to UV radiation. The generated gaseous products were separated from liquid phase within a gas–liquid separator and then transported to AFS for determination of cobalt. Factors affecting the efficiency of photo-CVG were investigated in detail, including type and concentration of low molecular weight (LMW) organic acid, buffer system, UV irradiation time, reaction temperature, carrier gas flow rate and hydrogen flow rate. With 4% (v/v) HCOOH and 0.4 mol L^− 1 HCOONa buffer solution, 150 s irradiation time and 15 W low pressure mercury lamp, a generation efficiency of 23–25% was achieved. A limit of detection (LOD) of 0.08 ng mL^− 1 without any pre-concentration procedure and a precision of 2.2% (RSD, n = 11) at 20 ng mL^− 1 were obtained under the optimized conditions. The proposed method was successfully applied in the analysis of several simple matrix real water samples.     

Carbazole-modified blue light-emitting copolymers with the backbones integrated by diphenyloxadiazole,fluorene, and triphenylamine

Two new blue light-emitting polymers, poly{[2,5-bis(4-phenylene)-1,3,4-oxadiazole]-[9,9-dihexylfluorene-2,7-diyl]-[N-(4-(9H-carbazol-9-yl)phenyl)-N,N-bis(p-phenylene)aniline]} (POFPA) and poly{[2,5-bis(4-phenylene)-1,3,4-oxadiazole]-[9,9-dihexylfluorene-2,7-diyl]-[4-(3,6-(di-9H-carbazol-9-yl)-9H-carbazol-9-yl)-N,N-bis(p-phenylene)-aniline]} (POFCPA), were synthesized by Suzuki coupling reactions. By GPC analysis against a linear polystyrene standard POFPA and POFCPA were found to have M_n of 1.68 × 10⁴ and 3.70 × 10³, respectively. In contrast to POFPA, the main absorption peak of POFCPA in dilute toluene solution was blue-shifted by Δλ = 26 nm owing to its backbone of relatively shorter π-conjugation length and more carbazole units in side chain. The absolute fluorescence quantum yield (Φ_f) of POFCPA in dilute toluene solution was determined as 73%, much higher than that of POFPA (Φ_f ≈ 58.9%) measured under the same conditions. An electroluminescence device based on POFCPA displays a stable blue emission having color coordinates of (0.15, 0.20), a maximum brightness of 4762 cd/m², and a maximum current efficiency of 1.79 cd/A. By using this polymer as the host material doped with 1 wt.% 4,4′-bis[2-(4-(N,N-diphenylamino)phenyl)vinyl]biphenyl, the achieved highest brightness, maximum current efficiency and maximum power efficiency are 13,613 cd/m², 3.38 cd/A, and1.84 lm/W, respectively.     

Analysis of catecholamines and related compounds in one whole metabolic pathway with high performance liquid chromatography based on derivatization

The simultaneous determination of Tyr, catecholamines and their metabolites in one whole metabolic pathway using high performance liquid chromatography coupled with fluorescence detection (HPLC–FLD) based on pre-column derivatization has been achieved successfully. 1,3,5,7-Tetramethyl-8-(N-hydroxysuccinimidyl butyric ester)-difluoroboradiaza-s-indacene (TMBB-Su), a highly reactive fluorescent reagent synthesized in our previous work, was used for the labeling of tyrosine (Tyr), l-3,4-dihydroxyphenylalanine (l-DOPA), dopamine (DA), norepinephrine (NE), epinephrine (E) and metanephrine (MN). Derivatization conditions including reagent concentration, buffer, reaction temperature and reaction time were also investigated to improve the derivatization efficiency and thus the sensitivity of the detection. The separation of the derivatives was obtained on C₁₈ column with the mobile phase of 20 mM pH 3.5 citric acid (H₃Cit)–sodium hydrogen phosphate (Na₂HPO₄) buffer and methanol. Good linearities with correlation coefficients square (R²) greater than 0.998 in the corresponding concentration ranges were observed and the detection limits (S/N = 3) were found in the range from 0.10 to 0.40 nM (l-DOPA: 1.45 nM). The proposed method has been applied to the detection of catecholamines and related compounds in mice liver and brain samples without tedious extraction or purification procedure, which exhibits excellent selectivity and sensitivity in the analysis of complex samples. This work provides an alternative approach in the metabolic research of catecholamines and is helpful for the study of catecholamine metabolism.