Continuous-flow enzymatic determination of creatinine with improved on-line removal of endogenous ammonia

A new continuous-flow automated enzymatic method suitable for the direct determination of creatinine in physiological samples is described. The proposed system utilizes an on-line gas predialysis unit in conjuction with a flow-through enzyme reactor coil and a potentiometric ammonia detector. The enzyme reactor contains immobilized creatinine iminohydrolase (EC 3.5.4.21) which converts creatinine to ammonia and N-methylhydantoin. Ammonia liberated from this reaction is detected downstream with the membrane electrode-based detector. The novel gas predialysis unit effectively removes >99.8% of endogenous ammonia (up to 1 mM) present in the sample. Thus, final peak potentials recorded by the electrode detector are directly proportional to the logarithm of creatinine concentrations present. The method is shown to be precise (<3%), selective, and capable of accurately determining creatinine in serum and urine samples containing abnormally high endogenous ammonia levels. Determinations of creatinine in serum samples (n = 30) using this new method correlate well with an existing Technicon AutoAnalyzer colorimetric method (r = 0.996).     

Simultaneous determination of creatine and creatinine using amperometric biosensors

In order to determine creatine and creatinine amperometric biosensors were proposed. A bienzymatic biosensor based on creatinase (CI) and sarcosine oxidase (SO) was used for the assay of creatine and a trienzymatic biosensor based on CI, SO and creatininase (CA) for the assay of creatinine. The linear concentration ranges are of pmol l⁻¹ to nmol l⁻¹ magnitude order, with very low limits of detection. The biosensors proved high reliability for determination of creatine and creatinine as raw material, and in the pharmaceutical formulation.     

An improved model for the kinetic description of the thermal degradation of cellulose

In spite of the large amount of work performed by many investigators during last decade, the actual understanding of the kinetics of thermal degradation of cellulose is still largely unexplained. In this paper, recent findings suggesting a nucleation and growth of nuclei mechanism as the main step of cellulose degradation have been reassessed and a more appropriate model involving chain scission and volatilization of fragments has been proposed instead. The kinetics of cellulose pyrolysis have been revisited by making use of a novel kinetic method that, without any previous assumptions regarding the kinetic model, allows performing the kinetic analysis of a set of experimental curves recorded under different heating schedules. The kinetic parameters and kinetic model obtained allows for the reconstruction of the whole set of experimental TG curves.     

An improved method for the potentiometric determination of plutonium using cerium/IV/ as an oxidant

An improved method for the determination of plutonium in an aliquot using cerium/IV/ as an oxidant is reported. Plutonium is oxidized quantitatively to plutonium/VI/ in nitric acid medium by cerium/IV/, the excess of which is chemically destroyed in a single step by hydrochloric acid. Plutonium/VI/ is then reduced to plutonium/IV/ with a known amount of Fe/II/, the excess of which is back titrated potentiometrically with standard dichromate. Results of analysis of 3–5 mg amounts of plutonium in aliquots containing standard plutonium nitrate solution are reliable within 0.2%. Effect of the presence of some relevant foreign ions has been studied. The application of the method for the analysis of mixtures containing various amounts of uranium and plutonium has been examined.     

An improved method for determination of air-borne polycyclic hydrocarbons

Summary In a search for a more effective solvent system than those discovered up to 1971 for separating the polycyclic hydrocarbon mixtures occurring in high-temperature tars, modifications were made to one found in 1957. Whatman No. 4 paper impregnated with 10% liquid paraffin solution in petrol ether was used as stationary phase, and methanol saturated with liquid paraffin as ascending mobile phase. Chromatography at 18° for 6 hr on 45×45 cm sheets of paper gave a separation considerably better than that achieved originally. Special attention was paid to the fraction containing carcinogenic hydrocarbons. No column chromatography or other pretreatment was necessary. Owing to the better separation it is possible to determine further polycyclic hydrocarbons such as 1,2, 5, 6-dibenzoanthracene, 3, 4, 9, 10-dibenzopyrene, and 3, 4, 8, 9-di-benzopyrene. A spot-evaluating procedure and a pretreatment of air samples have been developed. This procedure permits analysis of imponderable air samples.

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Zusammenfassung Auf der Suche nach einem besseren Lösungsmittelsystem zur Auftrennung polycyklischer Kohlenwasserstoffgemische aus Hochtemperaturteer wurde ein Verfahren aus dem Jahr 1957 modifiziert. Whatmanpapier Nr. 4 wurde mit einer 10% igen Lösung von flüssigem Paraffin in Petroläther als stationäre Phase imprägniert und Methanol, gesättigt mit flüssigem Paraffin, diente als aufsteigende Phase. Chromatographiert wurde bei 18° C durch 6 Stunden auf Papierblättern der Größe 45×45 cm. Die Trennung verlief erheblich besser als nach der Originalvorschrift. Besondere Aufmerksamkeit galt der Fraktion carcinogener Kohlenwasserstoffe. Säulenchromatographie oder eine andere Vorbehandlung ist nicht nötig. Infolge der besseren Auftrennung lassen sich polycyklische Kohlenwasserstoffe wie 1, 2, 5, 6-Dibenzoanthracen, 3, 4, 9,10-Dibenzopyren und 3, 4, 8, 9-Dibenzopyren bestimmen. Ein Verfahren zur Auswertung der Chromatogramm-flecken sowie die Vorbehandlung der Luftproben wurden ausgearbeitet. Die Analyse unwägbarer Luftproben wurde so ermöglicht.

     

An improved method for technetium determination in environmental samples

Summary The recent developments of extraction chromatography and ICP-MS made easier the determination of ⁹⁹Tc in environmental samples. However, in the non-contaminated area, a pre-concentration procedure is necessary, because usually a large amount of sample is used for analysis. In this study, the ferrous ion (Fe²⁺) added as a reductant can make chemical yield from 50 to 80%, when larger than 100 liter water samples or 500 g soil samples are analyzed. The extraction chromatography with TEVA resin (EIChrom) and the measurement by ICP-MS have been developed using ^95mTc and ¹⁰³Ru as yield tracers. Detection limits of 3σ are 0.054 mBq/kg for 500 g soil and 0.032 µBq/lfor 500 l water. A pond named Hinotani, Mie Prefecture in the central part of Japan, was selected to be investigated as a natural system in a non-contaminated area. Surface soil near this pond, pond water and sediment were collected and analyzed for ⁹⁹Tc. In a high fall-out area, Okuetsu, Fukui Prefecture forest soil was collected and analyzed. The ⁹⁹Tc in the surface (0-5 cm) was 10.5±0.8 mBq/kg. The ⁹⁹Tc in Hinotani surface (0-5 cm) soil were 0.77±0.06 mBq/kg less than in Okuetsu. Technetium-99 has been determined in pond water, sediment (0-5 cm) and shrimps in the Hinotani pond, 0.25±0.02 mBq/l, 3.3±0.3 mBq/kg, 1.5±0.2 mBq/kg, respectively.     

An improved method for the determination of free sphingosine in serum

Summary A rapid, sensitive and selective high-performance liquid chromatographic method has been developed for the determination of sphingosine in human serum. After precipitation with methanol, the samples were extracted using Carbopack B disposable columns; the sphingosine was eluted with 0.05 M hydrochloric acid in methanol-dichloromethane (20∶80, v/v) and the extract evaporated to dryness at 40°C. The sample residue was then reconstituted with methanol and reacted with o-phthaldialdehyde reagent to produce a fluorescent compound. Separation was performed using an LC-18 column with 0.05 M phosphate buffer (pH 7)-methanol-acetonitrile (15∶80∶5, v/v) as mobile phase. Fluorescence detection was performed with excitation and emission wavelengths of 340 and 455 nm, respectively. The serum extract was re-analyzed with a cyano LC column to minimize the possibility of false positive results. The possible interference of compounds having a structure similar to that of sphingosine was evaluated. The mean recovery of sphingosine was >94.5%. The limit of detection of the assay was 1 ng mL⁻¹. The between-run and within-run coefficients of variation for replicate analyses were <4.0% and <3.4%, respectively. The levels of free sphingosine in the serum of 40 normal subjects (20 male and 20 female) was investigated; the average level was 81.6±41.1 ng mL⁻¹ (mean ±S.D.) for males and 85.5±33.7 ng mL⁻¹ for females.     

An improved rapid method for the determination of actinides in water

A new rapid method has been developed for the determination of Th, Pu, Np, U, Am and Cm isotopes in water samples of about 1 L. Actinides are pre-concentrated by co-precipitation with Ca phosphate, sequentially separated on stacked TEVA and TK221 cartridges and measured by alpha spectrometry. The TK221 extraction chromatographic resin contains i.e. CMPO and DGA extractants. It has been characterized by measuring the weight distribution ratios (D_w) of actinides which are higher than 1000 for all actinides in 3 M HNO₃. The method has been optimized, applied for the analysis of tap and seawater samples and validated by participating in an IAEA proficiency test. Chemical recoveries for all actinides are better than 50%. The method can be performed within one day.

     

An improved method for the determination of glycosyltransferases usingpara-nitrophenyl-glycosides as substrates

Due to their easy purification nitrophenyl-glycosides have become of increasing interest as acceptor substrates for the assay of glycosyltransferase activity. In this communication reversed-phase chromatography is shown to provide a convenient means for the isolation of the radioactively labelled nitrophenylglycoside(s) formed in this reaction, superior to the methods hitherto used with regard to speed and specifity.     

An improved flow injection determination of nitrite in waters by using intermittent flows

An injector—commutator with a 0:1:2 section is employed to achieve intermittent flows without intermittent pumping. The resulting flow injection system is very simple, manually operated, and requires only one peristaltic pump. When applied to the spectrophotometric determination of nitrite in waters, the device leads to an enhancement of the sensitivity and sampling rate without affecting other analytical characteristics. The merging zones approach is employed to decrease consumption of reagents.     

Application and evaluation of a new cold-stable kinetic Jaffé reagent to the Hitachi 747 for the determination of serum creatinine

One of the most commonly requested tests of the clinical laboratory is that of serum creatinine. Since the late 19th century the most commonly used reagents have been those based upon the work of Jaffé [1], which are composed of alkali and picric acid. Upon addition of creatinine to this reagent a red color, referred to as a Janovsky complex [2 and 3] is rapidly formed. Almost all of the currently available approaches to the automated determination of creatinine are based upon this reaction. Furthermore, the rate of the reaction may be controlled so that either equilibrium [4] or kinetic [5] measurements can be made. Although various enzymic approaches have been proposed [6, 7, 8, 9 and 10] they, as yet, have not gained the same popular support of the modified Jaffé reagents.     

An improved technique for the spectrophotometric determination of nitrate with 2,4-xylenol

An improved method is proposed for the spectrophotometric determination of nitrate with 2,4-xylenol. The sample in aqueous (1.7 + 1 ) sulfuric acid is treated with 2,4-xylenol to produce 6-nitro-2,4-xylenol which is distilled into an ammoniacal water—isopropanol mixture. The intense yellow color of the ammonium salt of 6-nitro-2,4-xylenol is measured at 455 nm. The distillation is done in a Parnas—Wagner Kjeldahl Semimicro distillation apparatus. The isopropanol keeps the excess of 2,4-xylenol in solution. Two procedures are described. In the first (applicable to samples containing alkali nitrates but no chloride, alkaline earth, or ammonium salts), the solution is evaporated to dryness, and (1.7 + 1) sulfuric acid and 2,4-xylenol in acetone are added. In the second (applicable to samples containing chloride, alkaline earth, or ammonium salts), concentrated sulfuric acid is added dropwise to a cooled aliquot and the 2,4-xylenol reagent is then added; if chloride is present, it must be removed by prior precipitation with silver sulfate. Nitrite shows a slight interference which depends on the amount of nitrate and nitrite present.     

An improved iodometric method for the successive determination of copper and zinc

The method described by haider and khundkar for the succesive iodometric dctcrmination of copper and zinc gives results that arc too low both for copper and for zinc. The errors may amount to —20% for copper and to —5% for zinc. The causes of these errors arc traced and eliminated and an improved method for the succesive iodometric determination of copper and zinc is described.