A Fluorescence Polarization Assay of Erythrocyte Membrane Fluidity on the Abbott TDx Analyzer

Four fluorescein dye derivatives (attached to cholic acid, deoxycholic acid, lithocholic acid, and 5-cholenic acid-3β-ol) were studied as a means of detecting alterations in erythrocyte membrane fluidity. In addition, membrane fluidity using these novel dye derivatives was correlated to nonenzymatic glycation of erythrocyte membrane ghosts. These dyes have excitation and emission wavelengths suitable for analysis on an Abbott TDx analyzer. Both the lithocholic acid and the 5-cholenic acid-3β-ol fluorescein derivatives were capable of detecting erythrocyte membrane fluidity with an increase in polarization of approximately 34% over the range of glycation studied, and the fluidity was strongly correlated with the degree of glycation. The cholic acid and deoxycholic acid fluorescein derivatives were unable to enter the erythrocyte membrane since no alteration in fluorescence intensity or polarization was observed upon incubation with the membrane preparations. The results obtained indicate that the lithocholic acid and 5-cholenic acid-3β-oI fluorescein derivatives are potential new reagents for the development of routine assays for membrane fluidity on the Abbott TDx analyzer.     

Kinetic micro determination of mercury in natural waters and biological materials

Summary A wet digestion has been developed to prepare water and biological samples for a kinetic determination of mercury using an iodide-catalyzed reaction between cerium(IV) and arsenite(III). A mercury-free control, prepared using ion-exchange with a selective chelating resin, was used by adding mercury standards to make a calibration curve. Both inorganic and organic mercury can be determined by the method described either in water or biological samples containing mercury in the range of 0.05 to 2.0g per ml. The procedure can be used satisfactorily down to the 0.05-ppm level for fresh water and urine with an overall error of less than 5 %. The method can also be employed for the determination of mercury in sea water or blood serum with an error of 10 % or less and gives results which compare favourably with other procedures.

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Zusammenfassung Die nasse Veraschung zur Aufbereitung von Wasserproben und von biologischem Material für die kinetische Quecksilberbestimmung mit Hilfe der jodkatalysierten Cer-Arsenit-Reaktion wurde beschrieben. Durch Ionenaustausch an einem selektiven Harz wurde eine quecksilberfreie Vergleichsprobe hergestellt. Diese diente nach Zusatz bekannter Quecksilbermengen zur Ermittlung einer Eichkurve. Anorganisches und organisch gebundenes Quecksilber kann nach diesem Verfahren in Wasser oder biologischem Material im Bereich zwischen 0,05 und 2,0g/ml bestimmt werden. Für die Größenordnung von 0,05 ppm kann das Verfahren für frisches Wasser und für Harn mit einem durchschnittlichen Fehler unter 5% verwendet werden. Ebenso dient es zur Untersuchung von Meerwasser und Blutserum mit einem Fehler bis zu 10%. Die Ergebnisse stehen mit denen anderer Methoden in gutem Einklang.

     

Titrimetric determination of α-substituted cystines and α-substituted cysteines with N-Bromosuccinimide

Summary N-Bromosuccinimide has been employed as a titrant for the determination of -substituted DL-cystines and -substituted DL-cysteines. Using NBS and Bordeaux Red indicator, quantitative recoveries in the 1–5 mg range were obtained for L-cystine, L-cysteine, L-methionine, and the -substituted DL-cystines. With NBS and the iodine displa'cement procedure, quantitative recoveries in the 10--20 mg range were obtained for L-cysteine and the s-substituted DL-cysteines.

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Zusammenfassung N-Bromsuccinimid (NBS) wurde zur Titration von -substituierten DL-Cystinen sowie DL-Cysteinen herangezogen. Mit Bordeaux-Rot als Indikator erhält man quantitative Ergebnisse im Bereich von 1–5 mgL-Cystin,L-Cystein,L-Methionin bzw. -substituierten DL-Cystinen. Mit Hilfe des Jodverdrängungsverfahrens führt NBS zu quantitativen Ergebnissen im Bereich von 10–20 mgL-Cysteine bzw. -substituierten DL-Cysteinen.

     

Spectrophotometric study of bilirubin and hemoglobin interactions in several hydrogen peroxide generating procedures

The interference effects of bilirubin and hemoglobin have been described for the peroxidase-hydrogen peroxide oxidation of a hydrogen donor and the catalase-hydrogen peroxide oxidation of methanol to formaldehyde. A competition between bilirubin and the intended hydrogen donor is shown for the substitute analyte, hydrogen peroxide, with a resultant diminution of color due to the loss of intended reaction. No inhibition of peroxidase action appears to take place; its action when complexed with hydrogen peroxide is directed toward the competing hydrogen donor, bilirubin. The final color measured appeared to be partially compensatory, that is the sum of intended color plus the color of residual bilirubin. The subtraction of a serum blank representing a static system will result in a lowered value and a larger error. Hemoglobin, with its strong Soret band can, if its concentration is excessive, cause a major interference in reactions such as the Hantzsch reaction which result in overlapping bands at the reaction wavelength. Samples which are both hemolyzed and jaundiced would present as formidable blanking problems. Further studies on bilirubin and its glucuronide and their individual effect on the peroxidase-peroxide reaction are presently in progress.     

Polarographic reduction of D-glucurono-γ-lactone in nonaqueous media

Summary The nonaqueous polarographic reduction of D-glucurono--lactone in dimethylformamide using tetrabutylammonium iodide as the supporting electrolyte (apparent E_1/2=– 1.94 V versus mercury pool) is analytically applicable over the concentration range of 50–300g/ml.

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Zusammenfassung Die polarographische Reduktion von D-Glucuronsäure--lacton in Dimethylformamid mit Tetrabutylammoniumjodid als Trägerelektrolyt (E_1/2=–1,94 V gegen Quecksilber) ist im Konzentrationsbereich 50 bis 300g/ml analytisch anwendbar.

     

The Zimmermann reaction and a correction for irrelevant absorption

Two similar modifications of a previously described aqueous Zimmermann reaction for 17-ketosteroids were studied to see whether the Allen correction for irrelevant background absorption could be accurate when pure steroids were compared against a single steroid as a calibrator. Dehydroisoandrosterone was selected as the standard because most procedures use it for that purpose. In one procedure in which varied spectral shapes, wide-ranging peak maximum values, and molar absorptivities were evident, accuracy of recovery was poor. In a second procedure, in which the primary variation was in band width rather than in either fine structures or molar absorptivities, the results were more accurate though still inaccurate as compared to peak measurement calculations. It would seem that this correction could only be valid for those procedures in which calibrator and analyte yield spectra which show better uniformity than the two procedures described here.     

Determination of polychlorinated dibenzo-p-dioxin and dibenzofuran congeners in air particulate and marine sediment standard reference materials (SRMs)

Due to the limited number of environmental matrix certified reference materials (CRMs) with assigned values for natural levels of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs), an interlaboratory study was undertaken by the National Institute of Standards and Technology (NIST) and Environment Canada to establish reference concentration values for selected PCDD/Fs in two well-characterized NIST Standard Reference Materials (SRMs): SRM 1649a (Urban Dust) and SRM 1944 (New York/New Jersey Waterway Sediment). Results from 14 laboratories were used to provide reference values for the seventeen 2, 3, 7, 8-substituted PCDD/F congeners, the totals for individual tetra- through hepta-substituted PCDD/F homologues, and the total amount of tetra- through hepta-substituted PCDD/Fs. The mass fractions for the individual 2, 3, 7, 8-substituted congeners range from approximately 0.01 microg/kg to 7 microg/kg dry mass.     

Titrimetric determination of glucose,galactose, and xylose with n-bromosuccinimide

Summary N-Bromosuccinimide has been employed as a reagent for the determination of glucose, galactose, and xylose. The reactions are carried out at room temperature for 15 minutes. The errors involved are legs than ± 6% at the mg level. The time required for a complete determination is 45 minutes and the only standard solution required is sodium thiosulfate.

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Zusammenfassung N-Bromsuccinimid wurde als Reagens für die Bestimmung von Glukose, Galaktose und Xylose verwendet. Die Umsetzungen wurden in 15 Minuten bei Zimmertemperatur durchgeführt. Der Fehler beträgt im Milligrammbereich weniger als ± 6%. Für eine vollständige Bestimmung benötigt man 45 Minuten. Die einzig erforderliche Standardlösung ist Natriumthiosulfat.