Fluorescent Contaminants Research Articles

Optical methods in oceanography with special reference to oil fluorometry

A single channel profiling oil in-situ fluorometer has been constructed and tested in the marine environment as well as during controlled conditions in the laboratory. Its performance range for oil was shown to lie between 5 ppb up to 100 ppm (5 × 10 −9 and 100 × 10 −6, respectively) assuming no so-named fluorescent contaminants such as dissolved organic, inorganic and planktonic material. The fluorescent contaminants are generally of importance in coastal waters like the Baltic, the North Sea and the Norwegian Coastal Current. Steps taken to correct the total fluorescent signal for these contaminants in order to obtain thefluorescent signal due to oil are described and discussed. It is suggested that single channel oil fluorometry in the sea preferably should be combined with auxilliary optical measurements in order to arrive at reliable quantifications of spatial distribution of oil spills in the sea especially in the vicinity of land, oceanic fronts and pycnoclines, respectively.

High-performance liquid chromatographic determination of urinary catecholamines by pre-column solid-phase dansylation on alumina

Sensitive and selective high-performance liquid chromatographic determination of catecholamines by pre-column solid-phase dansylation is described. After catecholamines are adsorbed on alumina, the amino groups not responsible for adsorption are dansylated by a solid-phase reaction. The excess reagent and fluorescent contaminants are washed out, and the dansylated catecholamines are eluted and separated by reversed-phase high-performance liquid chromatography. The four catecholamine derivatives can be separated within 10 min and no major interfering peak is observed on chromatograms. The response of each catecholamine is linear from 10 to 500 pmol per sample and the detection limit is 0.5 pmol. This method was applied to determination of catecholamines in human urine.

Ultraviolet detection of synthetic oil contamination of skin.

A study was conducted to evaluate skin contamination occurring on coal liquefaction process workers using quantitative (Spill Spotter) and qualitative (blacklight) ultraviolet fluorescence methods. Other objectives were to determine the effectiveness of showering in removing contamination, and to determine the quantitative relationship between Spill Spotter response and skin contamination concentration. Field measurements showed wide ranges of background skin fluorescence caused by differing skin types. Visible dirt on the skin partially quenched the fluorescence response of the Spill Spotter. Fluorescent skin contamination on selected workers was quantified relative to a calibration curve developed on pigskin using a reference synfuel oil. One group of workers was shown to exhibit a two-to three-fold increase in fluorescence during the work shift. Showering removed skin contamination although residual contamination sometimes remained on workers that exhibited the greatest degree of skin contamination during the workshift. Further refinement of the ultraviolet method for absolute quantification of fluorescent skin contamination is needed.

Ultraviolet Detection of Synthetic Oil Contamination of Skin

A study was conducted to evaluate skin contamination occurring on coal liquefaction process workers using quantitative (Spill Spotter) and qualitative (blacklight) ultraviolet fluorescence methods. Other objectives were to determine the effectiveness of showering in removing contamination, and to determine the quantitative relationship between Spill Spotter response and skin contamination concentration. Field measurements showed wide ranges of background skin fluorescence caused by differing skin types. Visible dirt on the skin partially quenched the fluorescence response of the Spill Spotter. Fluorescent skin contamination on selected workers was quantified relative to a calibration curve developed on pigskin using a reference synfuel oil. One group of workers was shown to exhibit a two-to three-fold increase in fluorescence during the work shift. Showering removed skin contamination although residual contamination sometimes remained on workers that exhibited the greatest degree of skin contamination during the workshift. Further refinement of the ultraviolet method for absolute quantification of fluorescent skin contamination is needed.

A fluorometric microassay procedure for monitoring the enzymatic activity of GM 1-ganglioside β-galactosidase by use of high-performance liquid chromatography

For the measurement of the enzymatic activity of GM 1-ganglioside (II 3 NeuAcGgOse 4Cer, galactosyl- N-acetylgalactosaminyl-( N-acetylneuraminosyl)-galactosyl-glucosylceramide) β-galactosidase in crude enzyme samples, a microassay using nonradioisotopic GM 1-ganglioside was devised. To reduce the volume of the reaction mixture and eliminate the interferences due to the fluorescent contaminants in the reaction mixture, NADH, a product after the oxidation of the released galactose with NAD and β-galactose dehydrogenase, was fluorometrically estimated by use of high-performance liquid chromatography. By this method, as little as 10 pmol of galactose can be detected. Using rat brain homogenates as an enzyme sample, the several parameters were reexamined to define the optimal conditions for the assay. This assay method was also applied to human cultured skin fibroblast homogenates, and it was found that this method can be used for the diagnosis of GM 1-gangliosidosis, instead of the usual method using the radioisotopelabeled natural substrate.

Precision and accuracy of indole‐3‐acetic acid analyses performed with the 2‐methylindolo‐α‐pyrone fluorescence assay and with a high performance liquid chromatography technique with spectrofluorimetric detection, exemplified on pine tissue (Pinus sylvestris L.)

The precision and accuracy of two versions of the 2‐methylindolo‐α‐pyrone (2‐MIP) assay and of a high performance liquid chromatography (HPLC) technique was assessed when applied to analysis of indole‐3‐acetic acid (IAA) in pine (Pinus sylvestris) tissue. In one version of the 2‐MIP assay total fluorescence at wavelengths > 450 nm was recorded, whereas in the other version fluorescence spectra were recorded and analyzed. Such analysis revealed the occurrence in some samples of fluorescent contaminants that impaired both precision and accuracy. The precision and accuracy of the 2‐MIP assay could be improved by additional sample purification, and the over‐all accuracy of the assay was finally verified, at a high level of sample purity, by successive approximation. The precision of the assay was then 14%. Successive approximation also verified that the HPLC technique was accurate at a less advanced stage of sample purity than the 2‐MIP assay, and had a precision of 14%.

Separation of fluorescent impurities from Amphotericin B

Reverse phase high pressure liquid chromatography of Amphotericin B samples provides one means of cleanly separating Amphotericin B from a strongly fluorescent contaminant. Purified Amphotericin B does not exhibit any fluorescence.

Fluorescence immunoassay based on long time correlations of number fluctuations.

We report the development of a fluorescence-based immunoassay technique relying on the physical phenomena of random number fluctuations and diffusion, which we review. By determining the autocorrelation of the fluctuations in the fluorescent intensity, this methid is able to measure the amount of labeled antigen or antibody that is bound to micrometer-sized carrier particles in solution. The principal advantage of this technique is its insensitivity to small, fast-diffusing sources. It also discriminates against weakly fluorescent contaminants of size comparable to the carrier particles. We demonstrate these attributes by using two model systems: a human IgG assay and an idealized system consisting of polystyrene fluorescent spheres and rhodamine dye.

A modified method for the isolation and determination of brain histamine using the Bio-Rex 70.

Bio-Rex 70, a weak cation exchange resin, has been used to specifically isolate histamine from rat brain tissue. This method compares favourably to other extraction procedures with respect to selectivity, reproducibility and time taken to perform the procedure. However, because it combines the optimum of these properties, it appears more adaptable for routine laboratory use. Following isolation, histamine is quantified fluorometrically after condensation with o-phthaldialdehyde (OPT). The sensitivity of the procedure allows for the chromatographic isolation, using Bio-Rex 70, of 12.5 ng histamine to give fluorescence twice that of blank. In addition, the use of Bio-Rex 70 enables the selective separation of histamine from fluorescent contaminants such as spermidine. The stability and the reproducibility of the adsorption and elution characteristics of Bio-Rex 70 enables the determination of 30 brain samples in a working day. This method has been applied to determine whole brain and regional brain levels of histamine in control and L-histidine-treated rats. The whole brain level of histamine, which was 50 ng/g, was increased by L-histidine and the highest concentration of histamine was found in the hypothalamus. Since the reliability of existing histamine extraction procedures is questionable, under certain conditions, it is suggested that the use of Bio-Rex 70 is a valuable addition in evaluating the possible physiological role of brain histamine.

A sensitive fluorometric assay for avidin and biotin

The quenching of the tryptophan fluorescence of avidin by biotin forms the basis for a rapid, sensitive assay for biotin and avidin in solutions free of fluorescent contaminants. Avidin solutions are assayed by titration with a standard biotin solution until no further decrease in fluorescence at 350 nm, with excitation at 290 nm, is observed. Unknown quantities of biotin in solution may be determined by titration with aliquots of standard avidin until the change in fluorescence observed following each addition undergoes a sharp increase. As little as 20 ng of biotin may be measured in this way.

Plasma Retinol Assay by Elution from Silicic Acid with Cymene (p-Isopropyl Toluene)

Abstract We developed a microcolumn adsorption chromatographic technique for assay of plasma retinol. A petroleum ether extract of plasma was fractionated on silicic acid, to isolate retinol. The first eluent, petroleum ether, removed "unidentified fluorescent contaminant" (UFC); the second eluent, cyclohexene, removed traces of UFC and retinyl palmitate. Retinol was eluted by the third eluent, cymeme (p-isopropyl toluene). No recognized plasma vitamin A derivatives, other than retinol, were detectable in the cymene eluate. With the procedure, we could detect a minimum concentration of 7 µg of retinol per deciliter of plasma. Sixty-six plasmas from preschool children were assayed by our "cymene assay," and by the Neeld—Pearson modification of the Carr—Price procedure, the method of Thompson et al., and the column assay of Garry et al. Results by the latter three procedures were similar (P >0.025): the mean plasma retinol ranged from 29.4 to 31.2 (SD, ± 5.8 to 7.4) µg/dl. By the present assay the same samples contained 24.5 (SD, ± 6.8) µg/dl; significantly more was detected by the other procedures (P <0.001). This difference (16-22%) between the cymene assay for retinol and other assays for total vitamin A may reflect, in part, the concentration of other vitamin A derivatives in plasma.

Fluorometric Determination of Plasma 11-Hydroxycorticosteroids. II. Studies on the Specificity of the Method

Abstract We have investigated the specificity of the fluorometric method we proposed [Clin. Chem. 19, 710 (1973)] for plasma or serum 11-hydroxycorticosteroid determinations. The principal specific fluorogenic contaminants in the cortisol-containing extract of plasma, as detected by thin-layer chromatography and chemically, were triglycerides and total cholesterol. These contaminants contributed an average of 1.6 µg/dl to the total normal cortisol value. Fatty acids were also found, but did not fluoresce. Nonspecific serum fluorogens were quantitated at 1.3 ± 1.2 µg/dl. Cortisol, corticosterone, and fluorogenic contaminants represented an average of 62.0, 29.1, and 9.9%, respectively, of the total fluorometric plasma cortisol value obtained (expressed in terms of cortisol standard). A ratio of 8.3 ± 2.4 was found for cortisol/corticosterone when each component was determined in terms of its respective standard. Fluorescence scans of the plasma cortisol extract indicated cortisol to be the main component present, accompanied by minor fluorescent contaminants.

THE RAPID DETECTION OF BREWERY CONTAMINANTS BELONGING TO THE GENUS SACCHAROMYCES BY A SEROLOGICAL TECHNIQUE

The cells of contaminant Saccharomyces spp. are rendered fluorescent by means of fluorescein chemically bound to antibody protein. The fluorescent contaminants are observed microscopically, and an estimate of levels of contamination likely to be encountered in the brewery can be obtained within 3 hr. A serum of suitable specificity is obtained by careful cross-absorption with a brewing yeast, and by combining two sera, all brewery contaminants of the genus Saccharomyces so far encountered are detected. Careful selection of the brewing strain used for cross-absorption, and control of the cross-absorption procedure itself, allows the recognition of certain wild-type Sacch. cerevisiae strains.

Fluorescent contaminants from plastic and rubber laboratory equipment.

Plastic and rubber laboratory apparatus contains materials that are extractable with water, aqueous solutions, and nonaqueous solvents. These extractable substances fluoresce under ultraviolet light and were found as contaminants in nutrient solution used for growing excised lemon fruit tissue in vitro. The source of these fluorescent contaminants has been removed by using all-glass laboratory equipment.