A spectrophotometric method for the determination of free fatty acid in serum using acyl-coenzyme A synthetase and acyl-coenzyme A oxidase

A mixture of Ti(IV) and 4-(2-pyridylazo)resorcinol was found to be useful in the spectrophotometric determination of trace amounts of hydrogen peroxide. The absorbance at 508 nm was proportional to the concentration of hydrogen peroxide added. The reagent was successfully applied to the assay of free fatty acid in serum through the combined use of acyl-CoA synthetase and acyl-CoA oxidase. The latter enzyme produces H₂O₂. As a result, hydrogen peroxide was produced through the enzymatic oxidation of free fatty acid. It was possible to determine free fatty acid in 50 μl of serum at concentrations ranging from 0.02 to 1.5 mm. The coefficient of variation was less than 3% at concentrations ranging from 0.1 to 1.5 mm. In the present method, there is the advantage of minimal influence from reducible substances as well as greater simplicity and accuracy.     

Colorimetric determination of sulphate in freshwater with a chromate reagent

A colorimetric determination of sulphate in freshwater with BaCrO₄ as reagent is described. We consider the method to be simple, fast and precise (sd = 0.6% at 0.7 mM; N = 10). The method is valid in the range of 0.00025 M up to 0.002 M; lower concentrations can be measured by omitting a dilution step. For higher concentrations a smaller sample should be diluted. The chromate spectrum and its ionic distribution as function of pH and dilution are given.     

Sensitive spectrofluorometric method for the determination of ascorbic acid in pharmaceutical nutritional supplements using acriflavine as a fluorescence reagent

An easily performed, specific, sensitive, rapid, reliable and inexpensive procedure for the spectrofluorometric quantitation of ascorbic acid was proposed using acriflavine as a fluorescence quenching reagent. The procedure was based on the determined quenching effect of ascorbic acid on the natural fluorescence signal of acriflavine and the reaction between ascorbic acid and acriflavine in Britton–Robinson buffer solution (pH 6) to produce an ion‐associated complex. The reduction in acriflavine fluorescence intensity was detected at 505 nm, while excitation occurred at 265 nm. The relationship between quenching fluorescence intensity (?F) and concentration of ascorbic acid was linear (R² = 0.9967) within the range 2–10 μg/ml and with a detection limit of 0.08 μg/ml. No significant interference was detected from other materials often found in pharmaceutical nutritional tablets. The obtained results were compared with those from high‐performance liquid chromatography and appeared in good agreement, with no important differences in precision or accuracy. The proposed spectrofluorimetric method was used to determine the amount of ascorbic acid in a number of commercial pharmaceutical nutritional supplement tablets with a 95% confidence performance.     

真眼点藻类色素的提取与测定方法

分别采用甲醇、乙醇和丙酮3种有机溶剂提取7种真眼点藻的色素,比较3种有机溶剂提取色素的效果,测定3种有机溶剂色素提取液的吸收光谱,利用分光光度法计算藻的叶绿素a和类胡萝卜素的含量,并比较甲醇和乙醇色素提取液在A₄₇₀和A₆₆₆的最大吸收峰。结果表明:使用乙醇比甲醇和90%丙酮操作更简便、快捷并且毒害低。3种有机溶剂色素提取液的叶绿素a和类胡萝卜素的含量均无显著性差异(P>0.05),提取率基本一致。色素在3种有机溶剂中的吸收光谱相似,甲醇和乙醇的色素提取液在A₄₇₀和A₆₆₆的最大吸收峰并无显著性差异(P>0.05)。乙醇色素提取液可使用Lichtenthaler的公式计算色素含量。     

Chemometric-assisted kinetic-spectrophotometric method for simultaneous determination of ascorbic acid, uric acid, and dopamine

A chemometric-assisted kinetic spectrophotometric method has been developed for simultaneous determination of ascorbic acid (AA), uric acid (UA), and dopamine (DA). This method relies on the difference in the kinetic rates of the reactions of analytes with a common oxidizing agent, tris(1,10-phenanthroline) and iron(III) complex (ferritin, [Fe(phen)₃]³⁺) at pH 4.4. The changes in absorbance were monitored spectrophotometrically. The data obtained from the experiments were processed by chemometric methods of artificial neural network (ANN) and partial least squares (PLS). Acceptable techniques of prediction set, randomization t test, cross-validation, and Y randomization were applied for the selection of the best chemometric method. The results showed that feedforward artificial neural network (FFANN) is more efficient than the other chemometric methods. The parameters affecting the experimental conditions were optimized, and it was found that under optimal conditions Beer’s law is followed in the concentration ranges of 4.3–74.1, 4.3–78.3, and 2.0–33.0 μM for AA, UA, and DA, respectively. The proposed method was successfully applied to the determination of analytes in serum and urine samples.     

党参类多糖比色法含量测定

党参的主要成分是糖类,传统经验认为,党参“味甜者佳”,其品质好坏,多以含糖量来衡量。药理研究表明,党参的升血糖作用是其所含糖分所致。刘中煜等曾对5种党参中单糖含量进行了测定,但其多糖含量测定尚无报道。本文采用苯酚一硫酸比色法对党参类多糖含量进行了测定。     

Two methods for determination of transketolase activity

Two new optical methods for transketolase activity assay using only one substrate, xylulose 5-phosphate or glycol aldehyde, have been developed. For transketolase activity assay in the first method, it is necessary to add auxiliary enzyme, glyceraldehyde phosphate dehydrogenase. It is not needed in the second method. The range of transketolase concentration in the activity assay is 0.036–0.144 U/ml for the first method and 1.8–6.8 U/ml for the second one. Published in Russian in Biokhimiya, 2006, Vol. 71, No. 5, pp. 693–696.     

Colorimetric assay method for determination of the tannin acyl hydrolase (EC 3.1.1.20) activity

A new colorimetric method of tannase (tannin acyl hydrolase, EC 3.1.1.20) assay has been developed using its specific substrate tannic acid. It is based on the changes in optical density of substrate tannic acid after enzymatic reaction at 530 nm. The residual tannic acid was measured by a modified BSA precipitation method. This assay is very simple, reproducible, and very convenient, and with it tannase activity can be measured in relation to the growth of the organism.     

Quantitative determination of Coomassie R bound to proteins in polyacrylamide gel. A facilitated method for multi-spot analysis of electrograms

Dimethylsulfoxide (DMSO) was found to be an efficient solvent for extraction of Coomassie Blue R 250 (Coomassie R) from stained proteins on polyacrylamide gels. Kinetic measurements show that the extraction of the dye from a 2-D gel reached equilibrium in 48 h. Staining of E. coli ribosomal proteins by Coomassie R dissolved in trichloroacetic acid exhibited two types of dye-protein complexes, the majority of them yield a blue-purple colour, while the rest are stained with a light-blue tone and fluorescent appearance as well. The absorbance spectra of the complexes in the gel matrix differ significantly from each other. However, the DMSO-extracted Coomassie show identical absorbance profiles with lambda max at 602 nm, thus the amount of the bound dye can easily be measured spectrophotometrically.     

A rapid spectrophotometric method for the determination of esterase activity

We have developed a spectrophotometric assay method which continuously records esterase activity at 510 nm by monitoring absorbance changes due to the formation of a diazo dye complex. In our method, α-naphthyl ester substrates are hydrolyzed by enzymatic action to α-naphthol which couples to Fast Blue RR salt (a diazonium salt) forming a diazo dye complex. Our method is unique in directly monitoring the formation of the diazo dye complex without extracting the color of the complex as in other methods that use naphthyl esters and diazo coupling of reaction products. The method appears to be limited to α-naphthyl ester substrates, however, since β-naphthyl esters did not give a linear change in absorbance in the enzymatic reactions tested. With this assay method, one can use a single substrate both to determine esterase units quantitatively in solution and to detect esterase staining activity on gel electrophoresis.     

Rapid determination of amino acid concentrations in microbiological media: Evaluation of Borchers' cuprizone method

A micro-method adaptation of Borchers' cuprizone amino acid assay was ideal for the rapid determination of the concentrations and, thus the utilization, of sole amino acids in microbiological cultures. While the method has been in the literature for ≈ 30 yr, it does not seem to have been widely used. This communication report on the usefulness of the method in routine microbiological analyses and shows that interfering compounds were generally not a problem. Standard medium components were shown either not to interfere at all, or to be easily corrected for using a medium blank. A wide range of amino acids and a number of other N-containing compounds could be measured. The accuracy of the method was estimated at ±4.5%. a Thermus strain was grown on -glutamate and substrate utilization was followed using this method.     

Simultaneous determination of dopamine, ascorbic acid, and uric acid using carbon ionic liquid electrode

A recently constructed carbon composite electrode using room temperature ionic liquid as pasting binder was employed as a novel electrode for sensitive, simultaneous determination of dopamine (DA), ascorbic acid (AA), and uric acid (UA). The apparent reversibility and kinetics of the electrochemical reaction for DA, AA, and UA found were improved significantly compared to those obtained using a conventional carbon paste electrode. The results show that carbon ionic liquid electrode (CILE) reduces the overpotential of DA, AA, and UA oxidation, without showing any fouling effect due to the deposition of their oxidized products. In the case of DA, the oxidation and reduction peak potentials appear at 210 and 135mV (vs Ag/AgCl, KCl, 3.0M), respectively, and the CILE shows a significantly better reversibility for dopamine. The oxidation peak due to the oxidation of AA occurs at about 60mV. For UA, a sharp oxidation peak at 340mV and a small reduction peak at 250mV are obtained at CILE. Differential pulse voltammetry was used for the simultaneous determination of ternary mixtures of DA, AA, and UA. Relative standard deviation for DA, AA, and UA determinations were less than 3.0% and DA, AA, and UA can be determined in the ranges of 2.0x10(-6)-1.5x10(-3), 5.0x10(-5)-7.4x10(-3), and 2.0x10(-6)-2.2x10(-4)M, respectively. The method was applied to the determination of DA, AA, and UA in human blood serum and urine samples.     

Chemiluminescence determination of sulphite using a cyclometalated iridium complex as chemiluminescence reagent

A simple, fast and accurate chemiluminescence (CL) method for the determination of sulphite has been developed, based on its sensitizing effect on the CL reaction between a novel water‐soluble iridium complex, [(dpci)₂Ir(bvbbi)](PF₆) (dpci = 3,4‐diphenylcinnoline; bvbbi = N,N′‐bivinylester‐¹H,^1′H‐[2,2′] bibenzimidazole) and cerium(IV). Under the optimal experimental conditions, the increased CL response was linear, with the concentration of sulphite over the range 5.0 × 10^–7–5.0 × 10^–4 mol/L. The detection limit of the method was 1.6 × 10^–7 mol/L, with a relative standard deviation (RSD) of 2.7% for nine repetitive determination of 1.0 × 10^–4 mol/L sulphite. The method was successfully applied to the quantitative analysis of sulphite in sugar samples. The possible reaction mechanism of sulphite on the [(dpci)₂Ir(bvbbi)](PF₆)–cerium(IV) system is also briefly discussed. Copyright © 2012 John Wiley & Sons, Ltd.     

Determination of manganese (II) in some medicinal plants and their decoctions by a kinetic spectrophotometric method

The catalytic effect of manganese (II) on the oxidation of the azo dye tropaeolin 00 with potassium periodate in the presence of 1,10-phenanthroline was investigated. The reaction was followed spectrophotometrically by measuring the decrease in the absorbance of the azo dye at 445 nm. Under the optimum analytical conditions, manganese (II) in the range 0.05-2.5 ng/mL could be determined by the fixed-time method with a detection limit of 0.02 ng/mL. The developed method is highly sensitive, selective and simple, and was applied successfully to analyse decoctions of some medicinal plants for trace amounts of total manganese (II) and free manganese (II) ions without separation.     

Preparation of 2-nitro-5-thiobenzoate for the routine determination of reagent hypochlorous acid concentrations

Solutions of hypochlorous acid (HOCl) decay over time. This decay indicates the necessity for methods and reagents for the routine measurement of this oxidant. 2-Nitro-5-thiobenzoate is commonly used to measure HOCl concentrations. This article describes a method for the preparation of 2-nitro-5-thiobenzoate that is stable for at least 3 months. This method relies on the partial rather than full reduction of 5,5′-dithiobis-(2-nitrobenzoic acid) and the resulting equilibrium between the substrate and the product.     

Fluorescence derivatizing procedure for 5-hydroxytryptamine and 5-hydroxyindoleacetic acid using 1,2-diphenylethylenediamine reagent and their sensitive liquid chromatographic determination

A pre-column derivatization method using a fluorogenic reagent, 1,2-diphenylethylenediamine (DPE) was studied for the sensitive HPLC determination of 5-hydroxytryptamine (5-HT) and 5-hydroxyindoleacetic acid (5-HIAA), which are biosubstances used in the diagnosis of several diseases. For the quantitative determination, the biogenic indole compounds were converted to their corresponding fluorescent derivatives with DPE in the presence of potassium hexacyanoferrate (III) at room temperature, and then the derivatives were separated by reversed-phase liquid chromatography with fluorescence detection. The chromatographic detection limits of the fluorescent peaks at a signal-to-noise ratio of 3 were 0.3 fmol for 5-HT and 0.2 fmol for 5-HIAA. The proposed method permits the simultaneous quantification of 5-HT and 5-HIAA at concentrations higher than 2.4 nM in human urine without a clean-up procedure.     

A method for visual determination of sex, using the human hip bone

A new visual method for the determination of sex using the human hip bone (os coxae) is proposed, based on a revision of several previous approaches which scored isolated characters of this bone. The efficacy of the methodology is tested on a sample of 402 adults of known sex and age of French and Portuguese origins. With the simultaneous use of five characters of the hip bone, it is possible to provide a correct sexual diagnosis in 95% of all cases, with an error of 2% and an inability to identify sex in only 3%. The advantage of this new method is a reduction in observer subjectivity, since the evaluation procedure cannot involve any anticipation of the result. In addition, this method of sex determination increases the probability of a correct diagnosis with isolated fragments of the hip bone, provided that a combination of elements of one character is found to be typically male or female.     

Colorimetric method for the determination of ethanol by Flow Injection Analysis

An automated Flow Injection Analysis system using stop-flow technique for quantifying ethanol based in a colorimetric detection method was developed. The system permitted analysis in a linear range of 0.05–1 g ethanol l^–1 without external dilution, a sampling frequency of 15 analyses per hour, and a relative standard deviation of 3.5%. A dilution line was implemented in the FIA system permitting the extension of the linear range to 0.5–5.2 g ethanol l^–1 maintaining the same sampling frequency and standard deviation. The system was applied to measure ethanol concentrations present in samples of an alcoholic fermentation and the results showed no significant difference with other analytical procedures (GC).