A multipurpose capacitive biosensor for assay and quality control of human immunoglobulin G

We report a flow‐injection biosensor system with a capacitive transducer for assay and quality control of human immunoglobulin G (hIgG). The sensing platform is based on self‐assembled monolayers (SAMs) of carboxylic acid terminated alkyl‐thiols with covalently attached concanavalin A. The electrochemical characteristics of the sensor surface were assessed by cyclic voltammetry using a permeable redox couple (potassium ferricyanide). The developed biosensor proved capable of performing a sensitive label‐free assay of hIgG with a detection limit of 1.0 µg mL^?1. The capacitance response depended linearly on hIgG concentration over the range from 5.0 to 100 µg mL^?1, in a logarithmic plot. Typical measurements were performed in 15 min and up to 18 successive assays were achieved without significant loss of sensitivity using a single electrode. In addition, the biosensor can detect hIgG aggregates with concentrations as low as 0.01% of the total hIgG content (5.0 µg mL^?1). Hence, it represents a potential post‐size‐exclusion chromatography–UV (post‐SEC–UV) binding assay for in‐process quality control of hIgG, which cannot be detected by SEC–UV singly at concentrations below 0.3% of the total hIgG content. Biotechnol. Bioeng. 2009; 104: 312–320 © 2009 Wiley Periodicals, Inc.     

A novel fluorescent sensor based on electrosynthesized benzene sulfonic acid‐doped polypyrrole for determination of Pb(II) and Cu(II)

To develop conducting organic polymers (COPs) as luminescent sensors for determination of toxic heavy metals, a new benzene sulfonic acid‐doped polypyrrole (PPy‐BSA) thin film was electrochemically prepared by cyclic voltammetry (CV) on flexible indium tin oxide (ITO) electrode in aqueous solution. PPy‐BSA film was characterized by FTIR spectrometry, X‐ray photoelectron spectroscopy (XPS) and scanning electron microscopy (SEM). The optical properties of PPy‐BSA were investigated by ultraviolet (UV)‐visible absorption and fluorescence spectrometry in dimethylsulfoxide (DMSO) diluted solutions. PPy‐BSA fluorescence spectra were strongly quenched upon increasing copper(II) ion (Cu²⁺) and lead(II) ion (Pb²⁺) concentrations in aqueous medium, and linear Stern–Volmer relationships were obtained, which indicated the existence of a main dynamic fluorescence quenching mechanism. BSA‐PPy sensor showed a high sensitivity for detection of both metallic ions, Cu²⁺ and Pb²⁺, with very low limit of detection values of 3.1 and 18.0 nM, respectively. The proposed quenching‐fluorimetric sensor might be applied to the determination of traces of toxic heavy metallic ions in water samples.     

Electro-optical behaviour of CuFe2O4@rGO nanocomposite for nonenzymatic detection of uric acid via the electrochemical method

Uric acid (UA) is a blood and urine component obtained as a metabolic by-product of purine nucleotides. Abnormalities in UA metabolism cause crystal deposition as monosodium urate and lead to various diseases such as gout, hyperuricemia, Lesch–Nyhan syndrome, etc. Monitoring these diseases requires a rapid, sensitive, selective, and portable detection approach. Therefore, this study demonstrates the hydrothermal synthesis of CuFe₂O₄/reduced graphene oxide (rGO) nanocomposite for selective detection of UA. After the nanocomposite synthesis, characterization was performed by X-ray diffraction spectroscopy, Fourier transform infrared spectroscopy, Raman spectroscopy, X-ray photoelectron spectroscopy, UV–visible spectrometry, atomic force spectroscopy, scanning electron microscopy, and electrochemical analysis. Furthermore, from the electrochemical analysis using cyclic voltammetry (CV), kinetic studies were carried out by varying the scan rate to obtain the diffusion coefficient, surface concentration, and rate of charge transfer to achieve a calibration curve that indicates the quasi reversible nature of the fabricated electrode with a linear regression coefficient of oxidation (R²: 0.9992) and reduction (R²: 0.9971) peaks. Moreover, the fabricated nonenzymatic amperometric sensor to detect UA with a linearity (R²: 0.9989) of 1–400 μM was highly sensitive (2.75 × 10⁻⁴ mAμM⁻¹ cm⁻²) and had a lower limit of detection (0.01231 μM) at pH 7.5 in phosphate-buffered saline solution. Therefore, the CuFe₂O₄/rGO/ITO-based nonenzymatic sensor could detect interfering agents and spiked real bovine serum samples with higher sensitivity and selectivity for UA detection.     

A 31P NMR Study of the Reaction of Adenosine 3′ 5′-Cyclic Monophosphate with 2, 4, 6-Triisopropylbenzenesulfonyl Chloride

Abstract

Detection limits for the minor component in binary mixtures of Ado/AraA, Ado/XyloA, and Urd/dUrd depend strongly on the combined concentration of analytes. Limiting concentrations (in which ≤1% of the minor component was detected) were about two orders of magnitude lower with HPLC (UV detection) than with ¹H NMR and TLC (UV detection) with these nucleosides (ε_max 10 000–15 000). Minimum molar percentages of minor components detected in the 0.1–10 mM range were 0.25–1% with HPLC (UV), 1–2% with ¹H NMR, and ～2% with TLC (UV).     

Specific and Rapid Detection of Lily symptomless virus and Arabis mosaic virus in Lily by Dual IC‐RT‐PCR

Lily symptomless virus (LSV) and Arabis mosaic virus (ArMV) cause severe losses of quantity and quality of lily flower and bulb production. Specificity, sensitivity and speed of detection methods for viruses need to be improved greatly to prevent LSV and ArMV from spreading from infected lilies. A dual IC‐RT‐PCR procedure for detection was developed in which the antibodies of LSV and ArMV were mixed and the mixture used to coat the PCR tubes. The particles of the two viruses were captured by the respective antibodies. Interference by other RNA viruses in infected lily was eliminated in the RT‐PCR. Also, an RNA extraction step was omitted. The dual IC‐RT‐PCR products of LSV and ArMV were 521 bp and 691 bp, respectively. The specificity of the method was validated; only LSV and ArMV of four viruses were detected by dual IC‐RT‐PCR. The sensitivity of the detection method is 1 mg leaf tissue and higher than DAS‐ELISA due to enrichment by dual immunocapture.     

Facile synthesis of fluoro,methoxy, and methyl substituted ferrocene-based urea complexes as potential therapeutic agents

In the present work, the synthesis, characterization (FT-IR, multinuclear (¹H and ¹³C) NMR, AAS, Raman, and elemental analysis), DNA binding (cyclic voltammetry, UV–Vis spectroscopy and viscometry), and in vitro biological assessment of nine new ferrocene-based ureas are reported. The desulphurization of ferrocenyl thioureas to the corresponding oxo analogues using aqueous sodium hydroxide and mercuric chloride led to the ferrocenyl ureas (F1–F9) in high yields. The DNA binding studies performed by cyclic voltammetry and UV–Vis spectroscopy produced results that are in close agreement with one another for the binding constants (K) and an electrostatic mode of interaction was observed. The nature and the extent of interaction with DNA was further investigated by viscometry. The DFT/B3LYP method was used to determine the charge distribution and HOMO/LUMO energies of the optimized structure. The DFT calculated HOMO and LUMO energies correlate well with the experimentally determined redox potential values. The synthesized ferrocenyl derivatives exhibited good scavenging activity against 1,1-diphenyl-2-picrylhydrazyl radical (DPPH). These complexes were also scanned for their in vitro cytotoxicity against human carcinoma cell line THP-1 (leukemia cells). The results showed a moderate level of cytotoxicity against the subjected cancer cell line as compared with the standard chemotherapeutic drug (cisplatin).     

2-Amino-5-substituted-1,3,4-oxadiazole as chemosensor for Ni(II) ion detection: antifungal,antioxidant, DNA binding,and molecular docking studies

An oxadiazole derivative 2 was prepared by condensation reaction through cyclization of semicarbazone in the presence of bromine; the structural confirmation was supported by ¹H and ¹³C nuclear magnetic resonance (NMR) spectroscopy, Fourier transform-infrared spectroscopy, and liquid chromatography-mass spectrometry. Its sensing ability towards Ni²⁺ ion was examined showing a binding constant of 1.04 × 10⁵ compared with other suitable metal cations (Ca²⁺, Co²⁺, Cr³⁺, Ag⁺, Pb²⁺, Fe³⁺, Mg²⁺, and K⁺) using ultraviolet–visible (UV–vis) and fluorescence spectroscopic studies. The minimum concentration of Ni²⁺ ions and limit of detection was found to be 9.4 μM. A job's plot gave the binding stoichiometry ratio of oxadiazole derivative 2 vs Ni²⁺ ions as 2:1. Furthermore, the intercalative binding mode of oxadiazole derivative 2 with calf thymus DNA was supported by ultraviolet–visible (UV–vis) and fluorescent light, viscosity, cyclic voltammetry, time-resolved fluorescence, and circular dichroism measurements. The molecular docking result gave the binding score for oxadiazole derivative 2 as −6.5 kcal/mol, which further confirmed the intercalative interaction. In addition, the antifungal activity of oxadiazole derivative 2 was also screened against several fungal strains (C. albicans, C. glabrata, and C. tropicalis) by broth dilution and disc diffusion methods. In antioxidant studies, the oxadiazole derivative 2 showed potential scavenging activity against 2,2-diphenyl-1-picrylhydrazyl and H₂O₂ free radicals.     

Application of Co-naphthalocyanine (CoNPc) as alternative cathode catalyst and support structure for microbial fuel cells

Co-naphthalocyanine (CoNPc) was prepared by heat treatment for cathode catalysts to be used in microbial fuel cells (MFCs). Four different catalysts (Carbon black, NPc/C, CoNPc/C, Pt/C) were compared and characterized using XPS, EDAX and TEM. The electrochemical characteristics of oxygen reduction reaction (ORR) were compared by cyclic voltammetry (CV) and linear sweep voltammetry (LSV). The Co-macrocyclic complex improves the catalyst dispersion and oxygen reduction reaction of CoNPc/C. The maximum power of CoNPc/C was 64.7 mW/m² at 0.25 mA as compared with 81.3 mW/m² of Pt/C, 29.7 mW/m² of NPc/C and 9.3 mW/m² of carbon black when the cathodes were implemented in H-type MFCs. The steady state cell, cathode and anode potential of MFC with using CoNPc/C were comparable to those of Pt/C.     

Assessment of peanut allergen Ara h1 in processed foods using a SWCNTs-based nanobiosensor

The goals of this research were to develop a rapid single-walled carbon nanotube (SWCNT)-based biosensor and to employ it to commercial food products for Ara h1 detection. The SWCNT-based biosensor was fabricated with SWCNTs immobilized with antibody (pAb) through hybridization of 1-pyrenebutanoic acid succinimidyl ester (1-PBASE) as a linker. The resistance difference (ΔR) was calculated by measuring linear sweep voltammetry (LSV) using a potentiostat. Resistance values increased as the concentration of Ara h1 increased over the range of 1 to 10⁵ ng/L. The specific binding of anti-Ara h1 pAb to antigen including Ara h1 was confirmed by both indirect ELISA kit and biosensor assay. The biosensor was exposed to extracts prepared from commercial processed food containing peanuts, or no peanuts, and could successfully distinguish the peanut containing foods. In addition, the application of present biosensor approach documented the precise detection of Ara h1 concentrations in commercially available peanut containing foods.     

Genosensor for rapid,sensitive, specific point-of-care detection of H1N1 influenza (swine flu)

A 5′ amine group-linked haemagglutinin (HA) gene-specific probe was attached over the surface of a working electrode to develop a rapid, specific, and sensitive point of care detection assay for H1N1 (swine flu) in human respiratory nasal swabs. The probe was attached with a cysteine covered screen-printed gold electrode via 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS). The electrochemical assay was performed using differential pulse voltammetry with the use of the redox indicator methylene blue for the detection of different concentrations of the single-stranded viral genome. The developed genosensor showed high sensitivity for H1N1 influenza virus with a detection limit of 0.002 ng/6 μL of viral nucleic acid in the sample. Samples were analysed by quantitative real-time Polymerase Chain Reaction as well as by conventional PCR. The genosensor showed high specificity, as no cross-reaction was observed with the heterologous nucleic acid of different pathogens (Salmonella typhi, Neisseria meningitides, and Streptococcus pyogenes) and human DNA, and it was specific for H1N1 with a sensitivity of ∼49 μA cm⁻² ng^-1. Genosensor is based on a very simple methodology that can be followed based on its easy-to-access approach. It is quick and could be used as a point-of-care test for the detection of influenza virus within 30 min.     

Electrochemical detection of honokiol and magnolol in traditional Chinese medicines using acetylene black nanoparticle-modified electrode

Introduction – Honokiol and magnolol are the active components of Magnolia officinalis, which is a widely used traditional Chinese medicine. Their simultaneous analysis is, therefore, important for the quality control of the product. Objective – To establish a simple, sensitive and rapid electrochemical method for the simultaneous detection of honokiol and magnolol based on the remarkable enhancement effect of acetylene black nanoparticle (AB). Methodology – The AB‐modified electrode was prepared via solvent evaporation. The electrochemical response of honokiol and magnolol was investigated using cyclic voltammetry. The simultaneous detection was performed with differential pulse voltammetry. The method was validated in terms of linearity, sensitivity, precision and accuracy. Results – The linear range for honokiol is 0.5–300 µg/L, and the limit of detection (LOD) is 0.25 µg/L (9.4 × 10^?10 mol/L). For magnolol, the linear range is 10–250 µg/L, and the LOD is 5 µg/L (1.88 × 10^?8 mol/L). Conclusion – The new method was successfully used to determine honokiol and magnolol in a traditional Chinese medicine called Ageratum liquid. Copyright © 2010 John Wiley & Sons, Ltd.     

Preparation of a manganese titanate nanosensor: Application in electrochemical studies of captopril in the presence of para-aminobenzoic acid

This study reports the synthesis and characterization of a novel nanostructure-based electrode for electrochemical studies and determination of captopril (CP). At first manganese titanate nanoceramics were synthesized by the sol–gel method. The structural evaluations of the pure nanopowders were investigated by different techniques such as X-ray diffraction (XRD), transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Then it was used to prepare a new nanostructured manganese titanate carbon paste electrode (MnTiO₃/CPE). The characterization of the modified sensor was carried out by comprehensive techniques such as electrochemical impedance spectroscopy (EIS), SEM, and voltammetry. Subsequently, the modified electrode was used for CP catalytic oxidation in the presence of para-aminobenzoic acid (PABA) as a mediator. The results showed that PABA has high catalytic activity for CP oxidation. The electrochemical behavior of CP was studied by cyclic voltammetry (CV), linear sweep voltammetry (LSV), chronoamperometry (CHA), and differential pulse voltammetry (DPV) techniques. Under the optimized conditions, the catalytic oxidation peak current of CP showed two linear dynamic concentration ranges of 1.0 × 10⁻⁸ to 1.0 × 10⁻⁷ and 1.0 × 10⁻⁷ to 1.0 × 10⁻⁶, with a detection limit of 1.6 nM (signal/noise = 3), using the DPV technique. Finally, the proposed method was successfully applied for determination of CP in pharmaceutical and biological samples.     

Amperometric biosensor based on tyrosinase-conjugated polysaccharide hybrid film: selective determination of nanomolar neurotransmitters metabolite of 3,4-dihydroxyphenylacetic acid (DOPAC) in biological fluid

The amperometric detection of neurotransmitters metabolite of 3,4-dihydroxyphenylacetic acid (DOPAC) was achieved at a tyrosinase-chitosan composite film-modified glassy carbon (GC) electrode. The optimal conditions for the preparation of the biosensor were established. This bio-composite film was characterized by scanning electron microscopy (SEM) and Fourier transformed infrared (FT-IR) spectra, suggesting that chitosan covalently connected to chitosan chains. Electrochemical characterization of the bio-hybrid membrane-covered electrodes were also performed in 0.05 M phosphate buffer solution (pH 6.52) containing neurotransmitters or their derivatives by using cyclic voltammetry (CV), linear sweep voltammetry (LSV), square wave voltammetry (SWV) and amperometry. This simply-prepared protein-polysaccharide hybrid film provides a microenvironment friendly for enzyme loading. The sensor was operated at -0.15 V with a short response time. The current linearly increased with the increasing concentration of DOPAC over the concentration of 6 nM-0.2 mM. The lower detection limit for DOPAC is 3 nM (S/N=3). The sensitivity of the sensor is 40 microA mM(-1). A physiological level of neurotransmitters and their derivatives including dopamine, l-dopa, adrenaline, noradrenaline and homovanillic acid as well as ascorbic acid, uric acid and acetaminophen do not affect the determination of DOPAC.     

Selection in vitro for UV-tolerant sugar beet (Beta vulgaris) somaclones

With a reduced stratospheric ozone concentration, the generation of UV-tolerant plants may be of particular importance. Among different crop plants there is large variation in sensitivity to UV-B radiation. This study was undertaken to investigate the possibilities of using somaclonal variation and selection in vitro for improving UV-B tolerance in sugar beet (Beta vulgaris L.). Sugar beet callus was exposed to UV radiation (280–320 nm, 0.863–5.28 kJ m^-2 day^-1, unweighted) and resultant shoots were selected from surviving cells. After establishment of the plants, they were grown under either visible radiation (114 μmol m^-2 s^-1 PAR) or with the addition of UV radiation (6.3 kJ m^-2 day^-1 biologically effective UV-B). Screening of regenerants in vivo for tolerance to UV radiation was undertaken 10 months after termination of the UV selection pressure. Screening was done visually and by using a number of physiological parameters, including chlorophyll fluorescence induction, ultraweak luminescence, pigment analysis and total content of UV-screening pigments. A clear difference between the unselected and the UV-selected somaclones was observed when visually studying the UV damage and other leaf injury. The observations were supported by the ultraweak luminescence measurements. Unselected plants showed significantly greater damage when subjected to subsequent UV radiation as compared to the selected plants. The clones subjected to UV selection pressure displayed a significantly higher concentration of UV-screening pigments under subsequent UV radiation. The unselected plants under subsequent UV treatment showed a lower carotenoid concentration when compared to selected plants. However, no significant difference between treatments was found for chlorophyll a/b, or F/F_max, a measure of photosynthetic quantum yield.     

Energy from algae using microbial fuel cells

Bioelectricity production from a phytoplankton, Chlorella vulgaris, and a macrophyte, Ulva lactuca was examined in single chamber microbial fuel cells (MFCs). MFCs were fed with the two algae (as powders), obtaining differences in energy recovery, degradation efficiency, and power densities. C. vulgaris produced more energy generation per substrate mass (2.5 kWh/kg), but U. lactuca was degraded more completely over a batch cycle (73 ± 1% COD). Maximum power densities obtained using either single cycle or multiple cycle methods were 0.98 W/m² (277 W/m³) using C. vulgaris, and 0.76 W/m² (215 W/m³) using U. lactuca. Polarization curves obtained using a common method of linear sweep voltammetry (LSV) overestimated maximum power densities at a scan rate of 1 mV/s. At 0.1 mV/s, however, the LSV polarization data was in better agreement with single‐ and multiple‐cycle polarization curves. The fingerprints of microbial communities developed in reactors had only 11% similarity to inocula and clustered according to the type of bioprocess used. These results demonstrate that algae can in principle, be used as a renewable source of electricity production in MFCs. Biotechnol. Bioeng. 2009;103: 1068–1076. © 2009 Wiley Periodicals, Inc.     

Highly selective and sensitive ‘on–off’ fluorescent chemosensor for Fe3+ ions crafted by benzofuran moiety in both experimental and theoretical methods

A benzofuran glycinamide-based chemosensor, 3-(2-([4-fluorobenzyl]amino)acetamido)benzofuran-2-carboxamide ( BGA ) was developed and synthesized for the selective and sensitive detection of Fe³⁺ ions. The photophysical properties of the probe BGA were studied using UV–visible light absorption and fluorescence spectrophotometers. The chemosensor BGA showed a marked ‘on–off’ fluorescence response towards Fe³⁺ ions in the presence of other metal ions in DMSO/H₂O solution (9/1, v/v). The very low limits of detection (LOD) were calculated to be 10 nM and 43 nM using UV–visible light absorption and fluorescence spectrophotometers, respectively. Job's plot analysis revealed the formation of a BGA -Fe³⁺ complex with a 1:1 binding stoichiometry ratio using UV–visible light spectroscopy. The sensing mechanism was also demonstrated using density functional theory calculation.     

Enhancing dopamine detection using a glassy carbon electrode modified with MWCNTs, quercetin, and Nafion

Glassy carbon (GC) electrode was modified using multi-wall carbon nanotubes (MWCNTs), quercetin (Q) and Nafion^® in this sequence. The thus modified electrode was used for the detection of dopamine (DA) in the presence of equimolar ascorbic acid (AA). It is demonstrated in this study that MWCNTs can increase the current response of DA by five-fold and Q can reduce the oxidation overpotential of DA by about 60 mV, compared to these parameters obtained with a bare GC electrode. It is also shown that a layer of Nafion^® can virtually eliminate the interference of AA for the detection of DA. The GC/MWCNTs/Q/Nafion^® electrode (hereafter also called composite electrode) shows a current density of about 900 μA cm⁻² for DA, compared to the value of 80 μA cm⁻² of the GC electrode and to the value of 390 μA cm⁻² of the GC/MWCNTs electrode. The 11-fold enhancement in the sensitivity of the GC electrode for DA determination is attributed to the composite modification of the electrode, and is substantiated through various cyclic voltammetric experiments. Cyclic voltammetry (CV) and linear sweep voltammetry were used to characterize the electrodes. Calibration curves of batch and flow systems were obtained by amperometry for the detection of DA. Additionally, the composite modified electrode was tested with a human serum sample for the determination of DA and was found to be promising at our preliminary experiments.     

Electrochemiluminescence of an electrocatalytic action of etimicin on Tris(2,2′‐bipyridyl)ruthenium(II) immobilized in Nafion modified carbon paste electrode

A sensitive electrochemiluminescence (ECL) detection of etimicin at Tris(2,2′‐bipyridyl)ruthenium(II) [Ru(bpy)₃²⁺]–Nafion modified carbon paste electrodes was developed. The immobilized Ru(bpy)₃²⁺ shows good electrochemical and photochemical activities. Electrochemical and electrochemiluminescence characterizations of the modified carbon electrodes were made by means of cyclic voltammetry and electrochemical impendence spectroscopy. The modified electrode showed an electrocatalytic response to the oxidation of etimicin, producing a sensitized ECL signal. The ECL sensor showed a linear response to etimicin in the range of 8.0–160.0 ng mL^?1 with a detection limit of 6.7 ng mL^?1. This method for etimicin determination possessed good sensitivity and reproducibility with a coefficient of variation of 5.1% (n = 7) at 100 ng mL^?1. The ECL sensor showed good selectivity and long‐term stability. Its surface could be renewed quickly and reproducibly by a simple polish step. Copyright © 2009 John Wiley & Sons, Ltd.     

Electrochemical studies of biocatalytic anode of sulfonated graphene/ferritin/glucose oxidase layer-by-layer biocomposite films for mediated electron transfer

In this study, a bioanode was developed by using layer-by-layer (LBL) assembly of sulfonated graphene (SG)/ferritin (Frt)/glucose oxidase (GOx). The SG/Frt biocomposite was used as an electron transfer elevator and mediator, respectively. Glucose oxidase (GOx) from Aspergillus niger was applied as a glucose oxidation biocatalyst. The electrocatalytic oxidation of glucose using GOx modified electrode increases with an increase in the concentration of glucose in the range of 10–50 mM. The electrochemical measurements of the electrode was carried out by using cyclic voltammetry (CV) at different scan rates (20–100 mV s⁻¹) in 30 mM of glucose solution prepared in 0.3 M potassium ferrocyanide (K₄Fe(CN)₆) and linear sweep voltammetry (LSV). A saturation current density of 50 ± 2 mA cm⁻² at a scan rate of 100 mV s⁻¹ for the oxidation of 30 Mm glucose is achieved.     

VI. Measurement of proenkephalin A-derived peptides in biological tissues by high pressure liquid chromatography coupled with amperometric electrochemical detection

We describe an analytic method for the separation and quantitation of a number of proenkephalin A-derived peptides using high pressure liquid chromatography coupled with amperometric electrochemical detection (HPLC-AECD). Initially, we coupled our HPLC separation system with AECD in series with a UV detector for additional confirmation of peak specificity. AECD provided a 10⁶ - fold increase in sensitivity over UV detection for these peptides. In addition to Met-enkephalin (ME), ME-Arg, ME-Arg-Phe, ME-Arg-Gly-Leu, Leu-enkephalin (LE) and LE-Arg (Dyn 1–6), we separated and detected the sulfoxides of ME and its extended peptides. Subsequently, we used minor modifications of the isocratic mobile phase to separate and detect enkephalin-related peptides with greater sensitivity and shorter chromatographic run times; each of these mobile phases was used to separate and detect two to three peptides. We have applied this HPLC-AECD methodology to quantitate ME, ME-Arg-Phe, ME-Arg-Gly-Leu and LE in pheochromocytoma tumors.