Direct Spectrophotometric Method Research Articles

Carotene-bleaching activities of lipoxygenase and heme proteins as studied by a direct spectrophotometric method

A procedure is described for the direct spectrophotometric study of carotene-bleaching in aqueous solution. Beta-carotene, with or without added linoleate, is dissolved in suitable buffers with Tween 80, the decrease in absorbance at 460 nm is recorded, and the initial rate of bleaching is computed from the bleaching curves. The technique was applied to a comparative study of the catalytic effects of lipoxygenase, cytochrome c and peroxidase. The latter enzyme, which displayed much less activity in the presence of linoleate, compared to the other two catalysts, was the only one to cause significant carotene bleaching in the absence of added fatty acids. Activity was linearly related to the concentrations of lipoxygenase and cytochrome c, but not to the amount of peroxidase. The inhibitory effects of some antioxidants on carotene bleaching in the presence of the three catalysts were studied.

Přímé spektrofotometrické stanovení dusíku.

Přímé spektrofotometrické stanovení dusíku.

A direct spectrophotometric method for the assay of tannase.

A direct spectrophotometric method for the assay of tannase.

Kinetochromic spectrophotometry—II: Determination of sulphate by catalysis of the zirconium-methylthymol blue reaction

A direct spectrophotometric method is proposed for the determination of sulphate, based on its ability to catalyse the slow reaction between Methylthymol Blue and zirconium in slightly aged solution. The procedure is operated in the same way as a normal spectrophotometric method but with stricter control of timing. The interference of 100-fold excesses of 40 other ions was studied. Of these, Ce(III), Sr, Fe(III), Th, Sn(II), U(V), Mn(II), Sb(III), Se(IV), Bi(III), Te(IV), SO 3 2−, F −, PO 4 3−, AsO 4 3−, S 2−, tartrate, oxalate and citrate interfered. Cationic interferences may be removed by cation-exchange. The interference of equimolar ratios of arsenate, fluoride and phosphate may be removed by preliminary treatment with magnesium oxide. Sulphate may be determined in the range 0·1–2·4 ppm by the recommended procedure with a net molar absorptivity of 2·0 × 10 4 at 586 nm. The colour development time is 60 min.

Direct spectrophotometric method for determination of traces of boron in iron, steels and other compounds

For the determination of traces of boron in iron, low- and high-alloy steels and other materials a direct spectrophotometric method was developed. In this method the absorbance of the boron-Curcumin complex formed in the sample solution is compared with the absorbance of another aliquot of sample solution wherein the boron is rendered inactive with fluoride before the Curcumin is added.

The reduction of tetrazolium salts by plant mitochondria.

Data has been obtained concerning the reduction of tetrazolium salts by mitochondria isolated from Jerusalem artichoke tubers with succinate as the substrate using a direct recording spectrophotometric method of assay. ATP was found to increase the rate of reduction of the tetrazolium salts, this being independent of the effect ATP had on the rate of oxygen uptake. The magnitude of the stimulation by ATP depended on the concentration of tetrazolium salts present and under certain circumstances was suppressed by the addition of azide and cyanide. The sites at which the tetrazolium salts were reduced along the electron transport chain were investigated. The role of ATP has been discussed in relation to the mechanism of tetrazolium reduction.

Nicotinamide Adenine Dinucleotide Kinase from Azotobacter vinelandii: I. PURIFICATION AND PROPERTIES OF THE ENZYME

Abstract A method is described for the purification of NAD+ kinase from extracts of Azotobacter vinelandii (ATCC 9104) cells. The specific activity of the purified enzyme is the highest so far reported. The enzyme requires a metal ion for activity. There is a lack of specificity for the nature of the phosphate donor in the kinase reaction. The Michaelis constants for the enzyme substrates have been determined by a direct spectrophotometric assay method which is described. The enzyme is protected to a remarkable extent from heat inactivation by NAD+. The enzyme appears to be a sulfhydryl enzyme; the sulfhydryl groups are protected by NAD+.

Mechanism of color fading in the direct spectrophotometric method for Ca using glyoxal bis(2-hydroxyanil)

Mechanism of color fading in the direct spectrophotometric method for Ca using glyoxal bis(2-hydroxyanil)

Automatic Method for the Determination of Calcium in the Presence of Magnesium and Phosphate Ions

Abstract A direct spectrophotometric method for manual analysis of calcium using the indicator dye Eriochrome Blue S. E. was selected for automation. Most indicator dyes used have been somewhat unstable in solutions sufficiently alkaline to minimize the interference of magnesium. Eriochrome Blue S. E., when kept in an ice bath in the absence of light, permitted the use of sufficient alkali to remove interfering magnesium ions. Magnesium did not interfere in concentrations up to 12 mg./100 ml. when added to standards. The addition of 0.5% (w/v) citric acid removed interference of phosphate in concentrations up to 300 mg./100 ml. The automated method has the advantage of speed, yet retains the accuracy and precision of the conventional methods used for determining calcium.

Improved micro-method for determination of serum bilirubin

A simple, rapid, and precise diazo method for determining serum bilirubin has been described. The method requires 0.1 ml of serum. Interferences have been largely resolved by improved kinetics and stoichiometry, and by the use of a direct spectrophotometric technic as a reference method. Reliability of the diazo method has been established by comparison with a well-known diazo method as well as with the direct spectrophotometric reference method, by demonstrating the effect of hemolysis and lipemia, and by replicate analyses.

Simple chromatogaphic method for the separation and estimation of gibberellic acid in crude products

The estimation of gibberellic acid by a direct spectrophotometric method is described. For the separation a paper-chromatographic technique is used with a modified solvent and with a colour reaction of alkaline permanganate. Comparative data proved that the new method is superior to other techniques and can be applied to pure gibberellic acid or crude preparations as no interference occurs after chromatographic separation.

Purification and properties of succinyl-CoA synthetase from Jerusalem artichoke mitochondria.

Summary A direct spectrophotometric method for measurement of the activity of succinyl-CoA synthetase (succinate:CoA ligase (ADP), EC 6.2.1.5) having wider applications than those previously used has been developed. Succinyl-CoA synthetase was isolated from mitochondria and the resulting enzyme preparation had a specific activity more than 15-fold greater than that previously reported with the enzyme isolated from plant tissue. The apparent Km values for the substrates necessary for both the forward and reverse reactions were determined. The enzyme was shown to be highly specific for succinic acid and adenine nucleotide phosphates; in the reverse direction malonic acid was shown to act as a competitive inhibitor with succinic acid. The enzyme was less specific in its requirement for the metal-ion cofactor. Mn2+ and Co2+ could replace Mg2+, Zn2+ and Ca2+ could not.

A spectrophotometric method for the assay of threonine dehydratase

The enzymic conversion of β-hydroxy α-amino acids to α-keto acids by ThrDH (sheep liver) was followed by a direct spectrophotometric method at 230 mμ. The assay was more sensitive to changes in keto acid concentrations than the 2,4-dinitrophenylhydrazone method because of the absence of the high and variable blanks of the latter method. However, in the applicable range of concentrations both methods gave the same results. This procedure is not handicapped by the physical properties inherent in the coupled reactions.

Factors Affecting the Activity of Pentose Phosphatemetabolizing Enzymes in Rat Liver

A direct spectrophotometric method for the assay of pentose phosphate-metabolizing enzyme activity, also referred to as transketolase activity, is described. This method was used to study the effect of diets and hormones on the liver activity of this enzyme system. High protein, high fructose, and a starvation-refed regimen all produced significant increases in pentose phosphate-metabolizing enzyme activity. Administration of hydrocortisone or cortisone also caused increases and feeding iodinated casein or thyroxine administration had a negligible effect on the activity of this system. A 4-day fast, diabetes, adrenalectomy, and hypophysectomy all caused significant decreases in liver pentose phosphate-metabolizing enzyme activity. Both vitamin-free and thiamine-free diets produced large decreases in the activity of this system, whereas a pyridoxine-free diet causd only a slight decrease in activity. The addition of oxythiamine to the thiamine-free diet caused a further decrease in activity of the pentose phosphate-metabolizing enzymes.

METHANOL AS THE ABSORBING REAGENT IN THE DETERMINATION OF BENZENE, TOLUENE, XYLENE AND THEIR MIXTURES IN AIR.

Abstract A direct quantitative spectrophotometric method is presented for the determination of benzene, toluene, xylene and their mixtures in air. Methanol is used as the absorbing reagent and dry ice as the chilling agent to prevent evaporation. Vapor concentration in air is determined by direct ultraviolet spectrophotometry.

The Direct Spectrophotometric Determination of Protein in Cerebrospinal Fluid

Abstract A rapid and sensitive direct spectrophotometric method for the determination of cerebrospinal fluid protein which uses as little as 0.1 ml. of sample has been described. The result is not affected by the ratio of albumin to globulin in the sample. Interfering substances are removed prior to spectrophotometric measurement with an automatic separation chamber employing the principle of gel filtration. Slight alterations of the procedure described for the measurement of protein enable one to obtain the nonprotein moiety for study or to measure proteins in other biologic fluids.

Bilirubin Studies on the Serum of Newborn Infants

Abstract Total bilirubin concentration was determined on the sera of a group of newborn infants. Results obtained by a direct spectrophotometric method were compared simultaneously with those of a diazo procedure, using a simple spectrophotometer. The results showed close agreement provided that the sera were unhemolyzed. The effect of hemolysis on both methods is discussed.

Direct Spectrophotometry of Total Serum Bilirubin in the Newborn

Abstract A simple, rapid direct spectrophotometric method for the determination of total serum bilirubin on a micro basis, with a 1:21 dilution of serum in phosphate buffer, is presented for use with the Junior Coleman spectrophotometer. The formula, mg./100 ml. bilirubin = 50 A455 - 119 A575, is applied, where A represents the absorbancy at the wave length specified. The method for obtaining this or a similar equation is presented. The spectrophotometric method, when applied to sera obtained from newborns, gives apparently reliable results in the presence or absence of hemolysis. An evaluation of the method and its limitations is presented.

Investigation of the protective ultraviolet absorbers in a space environment. I. Rate of evaporation and vapor pressure studies

AbstractCompounds which are effective in protecting materials from terrestial ultraviolet radiation may, in a high vacuum environment, suffer loss both by evaporation and by photochemical degradation from the shorter wavelength ultraviolet radiation. The rates of evaporation from a free surface in a high vacuum of the commercially available protective ultraviolet absorbers have been measured as a function of temperature. Vapor pressures of the solid absorbers were calculated from the rate of evaporation data. The equilibrium vapor pressures of the liquid absorbers were determined by a direct vapor spectrophotometric method. Absorbers containing more than one hydroxy group were found to be considerably less volatile than the monohydroxy compounds, probably because of intermolecular hydrogen bonding of the former.

A direct spectrophotometric determination of butylated hydroxyanisole in lard and in hardened lard

SummaryA direct spectrophotometric method for the determination of butylated hydroxyanisole has been presented. This procedure allows good precision in the analysis of lard and hardened lard containing from 0 to 100 p.p.m. of BHA and is more reliable than existing colorimetric techniques, especially in the lower range of antioxidant concentrations.