Investigation of Cyanocobalamin Interferences to an Electrochemical Based Hemoglobin Test System

The structure of vitamin B-12 with a cobalt ion in the center is similar to the structure of heme with an iron center in the hemoglobin. The suspicion that the presence of vitamin B-12 in venous blood samples caused interference in readings of hemoglobin levels when using the BeneCheck hemoglobin test system, an electrochemical based strip and meter, was investigated in this study. Venous blood samples with 5 mg/dL of vitamin B-12 induced a positive error of 13% in the hemoglobin measurement, while 10 mg/dL of vitamin B-12 caused a positive error of 30% in the hemoglobin measurement. However, 1 mg/dL of vitamin B-12, a concentration much higher than that of venous blood concentrations in patients under B-12 treatment, did not cause an error in the hemoglobin measurement. These results suggest that the BeneCheck hemoglobin test system is suitable for clinical application for patients under B-12 treatment because the dosage is below 1 mg/dL.     

A novel liquid chromatography/tandem mass spectrometry based depletion method for measuring red blood cell partitioning of pharmaceutical compounds in drug discovery

A novel liquid chromatography/tandem mass spectrometry (LC/MS/MS)-based depletion method for measuring compound partitioning between human plasma and red blood cells (RBC) in a drug discovery environment is presented. Conventionally, RBC partitioning is determined by separate measurements of drug concentrations in equilibrating plasma and whole blood or RBC using separate standards prepared in their respective matrices, i.e., in plasma and whole blood or RBC lysates. The process is very tedious, labor-intensive, and difficult to automate. In addition, interferences from the heme and other highly abundant cellular composites make the measurement of the drug concentration in whole blood or RBC inevitably variable even with a highly specific LC/MS/MS method. Therefore, there is an imminent need to develop a straightforward and fast method to assess the partitioning of drug-like compounds in RBC.This work describes an LC/MS/MS-based depletion assay that measures the compound concentration in plasma that has been equilibrating with RBC. Compounds were spiked into fresh human whole blood and plasma respectively to a final concentration of 500 nM. Both the spiked whole blood and plasma control were incubated at 37 degrees C for up to 60 min. During the time course, aliquots of plasma and whole blood from both incubation mixtures were sampled at 10 and 60 min. The whole blood samples were centrifuged to yield the plasma. The plasma samples from both incubations were extracted using a protein precipitation method, and analyzed using LC/MS/MS under the multiple-reaction monitoring (MRM) mode. The RBC partitioning ratio was calculated using the analyte peak area responses of the plasma samples through an equation deduced in this work.The method was first tested using two commercial compounds, phenoprobamate and acetazolamide, to determine the optimal incubation conditions and the concentration dependency of the assay. The assay reproducibility was also assessed by three inter-day assays for phenoprobamate. This method was further evaluated using 20 commercial compounds of different classes with a wide range of RBC partitioning coefficients and the results were compared with those reported in the literature. Excellent correlation (R2=0.9396) was found between the measured and literature values. In addition, several proprietary compounds were assayed using both the new and traditional methods and the measured partitioning ratios from the two methods are equivalent. The experiments in this work demonstrate that the LC/MS/MS-based depletion method can provide direct and accurate measurement of RBC partitioning for compounds in drug discovery.     

Utilization of Geant4 Monte Carlo simulation method for studying attenuation of photons in normal and heavy concretes at high energy values

The mass attenuation coefficients, μ/ρ, the linear attenuation coefficients, μ and the mean free path, MFP of normal and heavy concretes; ordinary, hematite-serpentine, ilmenite-limonite, basalt-magnetite, ilmenite, steel-scrap and steel-magnetite have been investigated using Geant4 Monte Carlo method at photon energies 1.5, 2, 3, 4, 5 and 6 MeV. The mass attenuation coefficients, the linear attenuation coefficient and the mean free path for the concretes were found dependent upon chemical compositions, density and the photon energy. The linear attenuation coefficient values for the selected concretes increase with the density and decrease with the energy. The mean free path thickness for the all the concretes decrease with increase in iron content and increase with increase the photon energy. The Geant4 Monte Carlo simulation method results have been compared with experimental and theoretical XCOM data, and showed a very good agreement. Our investigation validates the utilization of the Geant4 Monte Carlo simulation method for high energy photon interactions.     

Guidelines for routine measurement of blood hemoglobin oxygen affinity. IFCC Scientific Division, Committee on pH, Blood Gases, and Electrolytes.

Two methods for the routine determination of blood hemoglobin oxygen affinity are described. Both methods use whole blood and do not require special equipment, tonometry, or special gas mixtures. The first method consists of a one-point determination of p 50, and requires only 200 muL to 400 muL of whole blood, therefore making it suitable for the pediatric population. The second method uses multiple points, thereby establishing both the shape and position of the hemoglobin oxygen equilibrium curve between 10 and 99% oxygen saturation. Interpretation of p 50 is discussed in relation to evaluation of patients with hemoglobinopathies and as a parameter in estimating availability of oxygen to the tissues.     

Quantitative and qualitative analysis of a microfluidic DNA extraction system using a nanoporous AlO(x) membrane

A nanoporous aluminium oxide membrane was integrated into a microfluidic system designed to extract hgDNA (human genomic DNA) from lysed whole blood. The effectiveness of this extraction system was determined by passing known concentrations of purified hgDNA through nanoporous membranes with varying pore sizes and measuring the amount of hgDNA deposited on the membrane while also varying salt concentration in the solution. DNA extraction efficiency increased as the salt concentration increased and nanopore size decreased. Based on these results, hgDNA was extracted from whole blood while varying salt concentration, nanopore size and elution buffer to find the conditions that yield the maximum concentration of hgDNA. The optimal conditions were found to be using a low-salt lysis solution, 100 nm pores, and a cationic elution buffer. Under these conditions the combination of flow and ionic disruption were sufficient to elute the hgDNA from the membrane. The extracted hgDNA sample was analysed and evaluated using PCR (polymerase chain reaction) to determine whether the eluted sample contained PCR inhibition factors. Eluted samples from the microfluidic system were amplified without any inhibition effects. PCR using extracted samples was demonstrated for several genes of interest. This microfluidic DNA extraction system based on embedded membranes will reduce the time, space and reagents needed for DNA analysis in microfluidic systems and will prove valuable for sample preparation in lab-on-a-chip applications.     

Study of the mass attenuation coefficients and effective atomic numbers in some gemstones

The total mass attenuation coefficients for natural beryl, corundum, garnet, pearl, and tourmaline gemstones were measured at 81, 356.5, 661.6, 1173.2, and 1332.5?keV photon energies. The samples were irradiated with ¹³³Ba, ¹³⁷Cs and ⁶⁰Co radioactive point sources using gamma ray transmission method. Total atomic and electronic cross-sections, effective atomic numbers and electron densities were determined experimentally and theoretically. The experimental values were compared with the calculated values for all samples. The calculations were extended for total photon interactions in a wide energy range (1?keV?C100?GeV) using WinXCom program of the most commonly irradiated gemstones with different sources of ionized radiation. The values of these parameters have been found to vary with photon energy and chemical composition of the gemstones. All variations of these parameters against energy are shown graphically for total photon interactions.     

Simultaneous Molecular Recognition of Interleukins 2 and 4, and of Tumour Necrosis Factor-α in Biological Samples

Two 3D stochastic sensors based on single-walled carbon nanotubes and multi-walled carbon nanotubes decorated with gold nanoparticles and modified with 5,10,15,20-tetrakis (pentafluorophenyl)-21H,23H-porphyrin iron (III) chloride were constructed and utilized for the simultaneous molecular recognition of IL-2, IL-4, and TNF-α in biological samples. The proposed sensors present low limits of quantification, high sensitivity, and wide concentration ranges. Recoveries values higher than 98.2 % of analytes from whole blood and brain tumour tissue samples were obtained.     

A comparative study of 632.8 and 532 nm laser irradiation on some rheological factors in human blood in vitro

The effects of laser irradiation with 632.8 and 532 nm on rheological properties of blood were comparatively studied in vitro. Under the irradiation condition of 30 mW, laser irradiation of blood samples using a spot diameter of 5 mm with each laser, showed promising results in the modulation of hemorheological properties. When blood samples from patients with abnormally high values of erythrocyte sedimentation rate (ESR) were irradiated, the values of ESR were lowered statistically by either of the 632.8 or 532 nm lasers. The laser irradiation reduced blood viscosities at different shear rates (10-110 S(-1)) for the hyper-viscosity blood samples. Laser irradiation increased the electrophoretic mobility (EPM) of erythrocytes when the values of the sample's EPM were abnormally slow. The erythrocyte deformability was enhanced by laser irradiation when the deformability of the sample from the patients was originally poor. For verifying the improvement of laser irradiation on erythrocyte deformability, the typical erythrocyte samples with poor deformability were produced by the pre-treatment of the erythrocytes with Ca(2+). The deformability of these erythrocyte samples was also improved after laser irradiation. These results suggest that membrane-bound hemoglobin (Hbm) might be the initial site of the interaction, since Hbm is the main cause of poor deformability when erythrocytes were treated with Ca(2+). In all experiments including ESR, blood viscosity, EPM and erythrocyte deformability, the 532 nm laser demonstrated more efficient effects on modulating rheological properties than 632.8 nm laser. This wavelength effect is consistent with the absorption spectrum of hemoglobin, reflecting that hemoglobin may be one of the action targets under laser irradiation.     

Capillary electrophoretic analysis of whole blood samples for hemoglobin-based oxygen carriers without the use of immunoprecipitation

Hemoglobin-based oxygen carriers (HBOCs) are blood substitutes, synthesized by polymerizing hemoglobin, which are being developed and investigated as alternatives to blood for medical purposes. However, due to their ability to increase the oxygen carrying capacity when taken by healthy individuals, HBOCs have been used as a doping agent among endurance athletes and are included in the World Anti-Doping Agency's Prohibited List. To maintain the fairness of competitions and continue the battle against doping, it is essential to be able to detect HBOCs if present in an athlete's blood. To achieve this goal, it is necessary to differentiate HBOCs from the native hemoglobin and to do so in a cost and time effective manner. We have developed a rapid capillary zone electrophoresis (CZE), UV absorbance, method capable of detecting HBOCs, in whole blood samples, at levels below those considered necessary to provide a performance enhancement. Our approach to the analysis for HBOCs utilizes the whole blood sample, not just the plasma, and does not require the use of immunoprecipitants to ensure accurate analysis. By lysing the red blood cells and using centrifugal filtration, followed by our CZE separation, we are able to effectively distinguish between native hemoglobin and HBOCs. Through this method, we have been able to reliably detect concentrations of HBOCs at the equivalent of 5.5 g/L, the equivalent to a 3.5% increase in blood hemoglobin concentration for an athlete.     

Tomography and elemental analysis using neutrons in transmission

Expressions for the calculation of minimum detectable mass and length fractions of elements, compounds and mixtures in a sample using neutron attenuation coefficients are derived, based on previous work done with photon attenuation coefficients. These expression allow quantitative information about elemental concentration to be extracted from neutron transmission measurements and neutron transmission tomography. Calculations are carried out for the detectable mass fraction of hydrogen in a number of sample matrices of industrial interest and of elements in a water matrix highlighting the differences with photon attenuation measurements. Results are presented for three neutron energies, cold (0.001 eV), thermal (0.025 eV) and fast (14 MeV).     

Synthesis,characterization and X‐ray shielding properties of polypyrrole/lead nanocomposites

Novel radiation shielding nanocomposites based on a conducting polymer were fabricated and investigated to determine their abilities in attenuation of X‐rays. Polypyrrole/Pb nanocomposites were prepared through chemical reduction of lead salt by a facile solution‐phase method using t‐BuOLi‐activated LiH and in situ chemical polymerization of pyrrole in the presence of dodecyl benzene sulfonic acid as dopant and surfactant and iron chloride as the oxidant. The morphology, composition, and electrical conductivity of resulting products were characterized by scanning electron microscopy, transmission electron microscopy, X‐ray diffraction analysis, energy‐dispersive X‐ray spectroscopy, fourier transform infrared spectroscopy, and standard four‐wire technique, respectively. In order to evaluate capability of nanocomposites in radiation shielding, X‐ray photon interaction parameters such as linear attenuation coefficient, attenuation percentage, and half‐value thickness were determined for the samples with different Pb loadings and thicknesses, at photon energies of 13.95, 17.74, 20.08, 26.34, and 59.50 keV. The investigation was carried out to explore the potential of polypyrrole/Pb nanocomposites as thin and light‐weight radiation shielding materials. Copyright © 2015 John Wiley & Sons, Ltd.     

高效液相色谱-非特异性同位素稀释质谱联用技术定量分析全血中总血红蛋白含量

建立了基于同位素稀释(ID)的高效液相色谱(HPLC)-电感耦合等离子体质谱(ICP-MS)联用技术,通过测量血红蛋白(HGB)中铁(Fe)元素含量实现了全血中总HGB准确定量的策略.该策略包括2种方法,第一种方法为在线HPLC-ID-ICP-MS定量技术:全血中HGB经液相色谱分离后,通过柱后在线添加浓缩54Fe同位素稀释剂的方式,与液相色谱洗脱液经三通混合后进入ICP-MS在线测量54Fe/56Fe的同位素比值,并根据同位素稀释质谱法公式及蛋白中Fe的含量计算HGB的浓度.针对全血样品中存在的其它主要含铁蛋白[如转铁蛋白(Tf)],在实现色谱完全分离的基础上,通过在线优化改变54Fe稀释剂流速实现了HGB和Tf的同时定量.在另一种方法中,先对全血样品进行消解并通过ID-ICP-MS测定其中Fe的总量,然后根据全血样品的HPLC-ICP-MS分析结果,通过计算HGB质谱峰面积占总峰面积的比值从而得到HGB中Fe含量占总Fe含量的比例,实现了其中HGB的准确定量. 2种定量方法得到的结果分别为(115.3±2.4) mg/g和(115.5±2.1) mg/g,结果吻合良好,方法的检出限...     

An enzymatic method for the rapid measurement of the hemoglobin A1c by a flow-injection system comprised of an electrochemical detector with a specific enzyme-reactor and a spectrophotometer

A flow-injection analytical (FIA) system, comprised of an electrochemical detector with a fructosyl-peptide oxidase (FPOX-CET) reactor and a flow-type spectrophotometer, was proposed for the simultaneous measurement of glycohemoglobin and total hemoglobin in blood cell. The blood cell samples were hemolyzed with a surfactant and then treated with protease. In the first stage of operation, total hemoglobin in digested sample was determined spectrophotometrically. In the second stage, fructosyl valyl histidine (FVH) released from glycohemoglobin by the selective proteolysis was determined specifically using the electrochemical detector with the FPOX-CET reactor. The FIA system could be automatically processed at an analytical speed of 40 samples per hour. The proposed assay method could determine selectively only the glycated N-terminal residue of β-chain in glycohemoglobin and total hemoglobin in blood cell. The enzymatic hemoglobin A_1c (HbA_1c) value calculated by the concentration ratio of the FVH to total hemoglobin, was closely correlated with the HbA_1c values certified by the Japan Diabetic Society (JDS) and the International Federation of Clinical Chemistry (IFCC).     

Analysis of photon interaction parameters as function of soil composition

In the present work the total mass attenuation coefficients (μ/ρ) for some soils collected from the Southeast and South of Brazil were measured at 59.5 (²⁴¹Am) and 661.6 keV (¹³⁷Cs) photon energies. The experimental values of the soils μ/ρ were compared with XCOM program calculations and GEANT4 Monte Carlo simulations. Total atomic and electronic cross-sections, effective atomic and electron numbers of all soil samples were calculated in a wide energy range (1 keV–100 GeV). The values of these parameters have been found to vary with photon energy and chemical composition of the soil. The variations of these parameters with energy are shown graphically for total photon interactions. The results showed that loamy soils have low photon attenuation parameters than clayey ones for the region of 59.5 keV.     

Green Determination of Total Iron in Water by Digital Image Colorimetry

A green iOS digital image colorimeter was fabricated for the determination of total iron in natural water. A mobile application operating on an iOS device was designed using Euclidean distance theory to perform iron determination. The application records the components of the color by storing red, green, and blue values as well as calculating the hue, saturation, brightness, and gray values using standard color theory. The component values obtained from the images of orange solutions of iron(II) complex with 1,10-phenanthroline were collected in a database and used for evaluation of the total iron concentration in water samples. Under the optimal conditions for solid-phase extraction used for sample preconcentration, the enrichment factor was 8.18. The iron concentration was determined by this device for concentrations from 0.01 to 1.0?ppm. The iOS digital image colorimeter achieved a limit of quantitation of 0.1?ppm. The accuracy and precision of the iOS digital image colorimeter were validated using a certified reference material, riverine water. The instrumentation was used for the analysis of natural water samples.     

Study of non-validity of mixture rule near K-absorption edges by X-ray spectrometric technique

X-ray spectrometric technique has been described to determine the X-ray mass attenuation coefficient, μ/ρ, of X-rays employing HPGe X-ray detector and radioactive sources. The photon intensity is measured by gating the channel of the spectrometer at FWHM/photo peak. Using the technique the “best value” values of μ/ρ were obtained for those thicknesses which lie in the transmission (T) range 0.5 ≥ T ≥ 0.02. Total attenuation cross sections for other elements and lead compounds were measured at photon energies from 17 to 88 keV to study the Bragg’s additivity law near the absorption edge of the lead. The measured values of mass attenuation coefficient values are compared with theoretical values obtained using Winxcom (programme). This study suggests that measured mass attenuation coefficient values at and near absorption edges differ from the theoretical value by about 17–23%.     

Mössbauer spectroscopic studies of tungsten ores

Tungsten ores from Degana, Rajashthan State, India are studied using Mössbauer spectroscopy. The low grade ore exhibits two types of mineral assemblages as per chemical states of iron. The relation between ferrous/ferric ratio and tungsten concentration also supports the existence of two types of mineral assemblages. The Mössbauer spectroscopy of concentrate ore samples has shown the absence of iron. The energy dispersive X-ray fluorescence spectroscopy on the contrary has evidenced the presence of iron in concentrate ore samples but at lower concentration on comparison with low grade ores. These differences in behaviour in the above spectroscopic studies may be additionally due to higher attenuation of -radiations and higher atomic weight material in the ore concentrates.     

Photon and proton activation analysis of iron and steel standards using the internal standard method coupled with the standard addition method

The internal standard method coupled with the standard addition method has been applied to photon activation analysis and proton activation analysis of minor elements and trace impurities in various types of iron and steel samples issued by the Iron and Steel Institute of Japan (ISIJ). Samples and standard addition samples were once dissolved to mix homogeneously, an internal standard and elements to be determined and solidified as a silica-gel to make a similar matrix composition and geometry. Cerium and yttrium were used as an internal standard in photon and proton activation, respectively. In photon activation, 20 MeV electron beam was used for bremsstrahlung irradiation to reduce matrix activity and nuclear interference reactions, and the results were compared with those of 30 MeV irradiation. In proton activation, iron was removed by the MIBK extraction method after dissolving samples to reduce the radioactivity of⁵⁶Co from iron via⁵⁶Fe(p,n)⁵⁶Co reaction. The results of proton and photon activation analysis were in good agreement with the standard values of ISIJ.     

Gamma-ray shielding and structural properties of barium–bismuth–borosilicate glasses

The attenuation coefficients of barium–bismuth–borosilicate glasses have been measured for gamma-ray photon energies of 662, 1173 and 1332 keV using a narrow beam transmission geometry. These coefficients were then used to obtain the values of mass attenuation coefficients, effective atomic number, effective electron density and mean free path. Good agreement has been observed between experimental and theoretical values of these parameters. From the obtained results it is reported here that from the barium–bismuth–borosilicate glasses are better shields to gamma-radiations in comparison to the standard radiation shielding concretes from the shielding point of view. The molar volume, FTIR and acoustic investigations have been used to study the structural properties of the prepared glass system. The obtained results reveal that the formation of non-bridging oxygens occurs at higher concentration of Bi₂O₃.     

Enzymatic determination of urea in undiluted whole blood by flow injection analysis using an ammonium ion-selective electrode

A flow-injection system is described for the assay of urea in undiluted whole blood. Urea is quantified by means of an ammonium ion-selective electrode covered with a membrane with covalently immobilized urease. The enzymatically generated ammonium ion is directly related to the urea concentration. Interference from potassium is reduced by adjusting the potassium ion concentration in the carrier stream and in the aqueous calibration solutions to 4.0 mM; it can be eliminated by measuring the potassium ion concentration in the sample separately and applying a mathematical correction for the K⁺ contribution to the signal. The linear measuring range is 1–40 mM urea, with an injection frequency of 40 h^?1 and a standard deviation of 1% for whole blood samples. The result of the measurement is obtained within 25 s from the time of injection. Vatiations in the hematocrit level of the sample have no effect on the measurement. The results obtained by the flow-injection method are in excellent agreement with those found routinely at a local hospital. The sensor is stable for more than 25 days.