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PubMed1990

Analysis of the porphyrin content of fluorescent pus by absorption spectrophotometry and high performance liquid chromatography.

Extracts of 19 samples of pus which showed red fluorescence with ultraviolet light were screened for the presence of porphyrins by absorption spectrophotometry. All those which showed spectra typical of metal-free porphyrins were analysed by high performance liquid chromatography to identify the porphyrins present. These were predominantly the di-carboxylic porphyrins, deuteroporphyrin and mesoporphyrin, and another which was thought to be pemptoporphyrin. This combination matched those reported previously in normal stools. Protoporphyrin IX was shown not to be the most common fluorescent pigment in pus and was never present alone. However, the di-carboxylic porphyrins may be produced by bacterial metabolism of its labile vinyl side-chains. Black-pigmented bacteroides (the melaninogenicus group of Bacteroides spp. and Porphyromonas spp.) were isolated from 12 (63%) of the 19 pus samples; these may produce protoporphyrin IX by the demetallation of haem.

Bacterial Infections↗
PubMed1992

Determination of dissociation constants of some pharmaceutical compounds using derivative spectrophotometry.

Derivative spectrophotometry is applied to the determination of the dissociation constants (pKa) of some weakly dissociating compounds. The pKa value(s) can be calculated by measuring the amplitude of derivative maxima (D) at the selected wavelengths for the dissociated, undissociated and partially dissociated forms of the compound. A graphical technique, based on plotting the derivative spectrophotometric titration curves and interpolating pKa at D1/2 is also presented. Both methods are applied to pharmaceutical compounds with single and double dissociation constants. The results are in good agreement with the reported pKa values of the investigated compounds.

Chemistry, Pharmaceutical↗
PubMed2005

The measurement of tooth whiteness by image analysis and spectrophotometry: a comparison.

Digital image capturing and analysis techniques have been used to measure the colour of teeth and to compare with spectrophotometric results and visual observations. A non-linear image analysis approach was developed and, for the colour range of human teeth, allows device-dependant digital camera colour data to be quantitatively transformed to Commission Internationale de l'Eclairage (CIE) colorimetric values. With reference to a CIE standard illuminant, two different lighting arrays have been used. For flat and non-translucent white and yellow surfaces, spectrophotometric results showed that this transformation achieves required accuracy. It was found, in all of the present studies, which included measurements on the VITA Lumin Vacuum shade guide and extracted teeth, that spectrophotometry invariably underestimated values of the CIE whiteness index. However, the results from these two types of measurement correlated well. There was also a reasonably good correlation between earlier data obtained by visual assessment and the present data by the two instrumental methods. For extracted teeth, both instrumental methods used in this work did not confirm a whitening effect for 2-min brushing with toothpaste, but did show significant whitening results for bleaching with 15% hydrogen peroxide.

Algorithms↗
PubMed1975

Interactions of colipase with bile salt micelles. 2. Study by dialysis and spectrophotometry.

The finding reported in the preceding paper that colipase is able to bind one sodium taurodeoxycholate micelle per molecule was confirmed by dialysis and spectrophotometry. Dialysis in the presence of labelled sodium taurodeoxycholate provided a direct qualitative proof of taurodeoxycholate binding to colipase. This binding was found to occur only above the critical micelle concentration. But, dialysis did not give any information about the composition of the associations, because equilibrium was not attained at the end of the assays. Addition of sodium taurodeoxycholate above the critical micelle concentration was also observed to induce a strong perturbation of the ultraviolet spectrum of one or several of the three tyrosines of colipase. The variation of the perturbation as a function of sodium taurodeoxycholate concentration was consistent with the binding of a single micelle to colipase. The dissociation constant calculated in "micelle molarity" was approximately 1 X 10(-4) M. The colipase-bile salt micelle association can fix one molecule of lipase to form a ternary complex which represents an interesting model of a protein-protein interaction mediated by an organized lipid structure. The ternary complex is probably also a model for lipase-substrate interactions in the presence of an amphipath.

Binding Sites↗
PubMed1977

The reduction by dithionite of Fe(III) myoglobin derivatives with different ligands attached to the iron atom. A study by rapid-wavelength-scanning stopped-flow spectrophotometry.

1. The reductions of a number of sperm whale Fe(III) myoglobin-ligand complexes by sodium dithionite in a phosphate buffer pH 6.4, were investigated by using rapid-wavelength-scanning stopped-flow spectrophotometry. The ligands were azide, cyanide, fluoride, imidazole, thiocyanate and water. 2. The reduction of Fe(III) myoglobin cyanide led to the transient formation of Fe(II) myoglobin cyanide but no intermediate species were observable during the reductions of the other derivatives. The final product of the reaction in all cases was unliganded Fe(II)myoglobin. 3. Invesigation of the effect of dithionite concentration on the rate of reduction indicated that the SO2- radical ion was the active species in reducing the azide, cyanide, fluoride and thiocyanate derivatives. 4. Comparison of the observed rates of reduction at different ligand concentrations with those predicted for a pathway of reduction involving prior dissociation of the ligand, allowed us to estimate the rate of reduction with the ligand in position (outer-sphere reduction). There was a large variation in the relative rates of outer-sphere reduction in the order imidazole greater than CN- greater than SCN- greater than N3- greater than F-. The fluoride derivative was so resistant to outer-sphere reduction that the reaction with SO2- proceeded only by a pathway involving dissociation of F- before reduction. It was calculated that any direct reduction of this complex was at least 100 times slower than that of the azide derivative. 5. The results are discussed in terms of the possible rôle of the axial ligands in haem proteins and it is suggested that the pathway of the electron to the Fe(III) centre may be via the ppi orbitals of these ligands.

Animals↗
PubMed1982

The state of tyrosine and phenylalanine residues in proteins analyzed by fourth-derivative spectrophotometry. Histone H1 and ribonuclease A.

The analysis of the absorption spectra of model compounds of tyrosine and phenylalanine residues by means of fourth-derivative spectrophotometry is able to separate the contribution of the two chromophores, thus allowing the study of each one. Fourth-derivative analysis resolves the two main vibrational bands of tyrosine, giving rise to two peaks which are sensitive to changes in the environment of the phenolic ring. The parameters obtained from the fourth-derivative spectra were found to depend on the strength of the hydrogen bonds formed by the OH group of tyrosine, as well as on the heterogeneity of tyrosine environments. It is also shown that the fourth-derivative tyrosine peaks are not perturbed by broad bands, such as that arising from ionized tyrosine chromophores. The peaks arising from the phenylalanine model, although less sensitive than those of tyrosine, were found to depend on the polarity of the environment. As a check of the method, it is applied to the study of tyrosine and phenylalanine residues of calf thymus histone H1 and bovine pancreatic ribonuclease A.

Animals↗
PubMed1983

Fourth-derivative spectrophotometry analysis of tryptophan environment in proteins. Application to melittin, cytochrome c and bacteriorhodopsin.

Fourth-derivative spectrophotometry is applied to the analysis of solvent effects on the spectral transitions of N-acetyl-L-tryptophan amide, as well as of its mixtures with N-acetyl-L-tyrosine ethyl ester. These compounds were analyzed in different media. It was found that the position of the longest-wavelength minimum of the fourth-derivative spectrum is mainly determined by the nature of the Trp environment, with a minor contribution from that of Tyr. A geometrical parameter is also defined, which depends on both Trp and Tyr environments. The simultaneous consideration of both parameters allows an estimation to be made of the Trp environment. The method is applied to the study of conformational changes of three well-characterized proteins: melittin from bee venom, cytochrome c from horse heart and bacteriorhodopsin from Halobacterium halobium. The results obtained are found to be in accordance with the known conformational properties of these proteins. In addition, the fourth-derivative operation completely eliminates the interference from the near-ultraviolet bands of the prosthetic groups.

Animals↗
PubMed1989

Measurement of carboxyhaemoglobin by spectrophotometry and gas chromatography.

The purpose of this study was to evaluate state-of-the-art spectrophotometry for measurement of carboxyhaemoglobin (COHb). We measured the fractional concentration of COHb in 109 blood samples from patients under investigation of anaemia or with exposure to carbon monoxide (smokers) with the OSM3 Hemoximeter and by gas chromatography. Duplicate measurements were made with both methods in 42 samples. We found only a trivial systematic difference between the two methods. There was, however, a considerable scatter of the measurements, the limits of agreement (95% confidence limits for the difference between the two methods) being -0.98 and 0.86% COHb. The poor agreement between the methods was largely explained by a large random scatter in duplicate spectrophotometric measurements, whereas the method based on gas chromatography was highly reproducible. We conclude that the low accuracy of spectrophotometric measurements of COHb precludes its use for assessment of the endogenous production of CO, but that it may be useful for assessment of exposure to exogenous CO.

Anemia↗
PubMed2002

Reflectance spectrophotometry in the gastrointestinal tract: limitations and new applications.

OBJECTIVES: Reflectance spectrophotometry (RS) assesses blood flow changes by measuring an index of Hb concentration (IHB) and index of Hb oxygen saturation (ISO2). We tested the following hypotheses: 1) endoscopic RS measurements obtained by two observers and with the aid of fiber optic and video endoscopes are similar, and 2) the method is suitable for documenting mesenteric venoconstriction associated with systemic hypoxia and blood flow autoregulation associated with hemorrhagic hypotension. METHODS: Study 1: two investigators obtained baseline gastric mucosal RS measurements in anesthetized rats (n = 3) before and after stepwise reduction of blood pressure induced by arterial hemorrhage. Study 2: subjects were examined by both fiber optic and video endoscopes. Endoscopic RS measurements were obtained at 20 cm from the anal verge. Study 3: video endoscope was used to obtain RS measurements in oxygen-dependent patients on and off oxygen treatment. Study 4: the procedures in study 1 were repeated in five additional rats by one of the investigators. RESULTS: Study 1: there was good agreement between the measurements of IHB and ISO2 between the two investigators. Study 2: video endoscope-assisted measurements were consistently lower. Study 3: cessation of oxygen treatment produced a significant drop in oxygen saturation (pulse oximetry), decline in ISO2, and rise in IHB. Study 4: when blood pressure varied between 90% and 40% of baseline, gastric mucosal blood flow (IHB) was maintained at approximately 70% of baseline level. CONCLUSIONS: We confirmed that reproducible measurement can be obtained by different investigators using standardized techniques. Standardization of endoscopic equipment is also necessary to overcome the significant limitation of endoscopic equipment on RS measurements. RS measurements can document mesenteric venoconstriction associated with systemic hypoxia and blood flow autoregulation associated with hemorrhagic hypotension.

Animals↗
PubMed1997

Measurement of lip and skin pigmentation using reflectance spectrophotometry.

Previous methods for evaluating the color of oral epithelial tissues have relied primarily on subjective indices and interpretation. In this study, a narrow-band reflectance spectrophotometer was used to measure melanin and hemoglobin pigmentation in the lips and skin of individuals who self-reported their skin type from a choice of white, olive, or black. Significant differences in melanin pigmentation were observed among skin types and at most anatomical sites evaluated between skin types. Hemoglobin levels were not significantly different among skin types at most anatomical sites, but differences were observed among anatomical sites within each skin color group. Within each skin type, melanin (except in blacks) and hemoglobin were significantly higher for both the upper and lower lips compared with most other anatomical sites. Results suggest that reflectance spectrophotometry is a useful tool for quantifying differences in melanin and hemoglobin pigmentation in oral tissues.

Analysis of Variance↗
PubMed1989

Determination of Na and K in the rat lens by atomic absorption spectrophotometry.

A new procedure is described for determination of sodium and potassium in rat lenses with atomic absorption spectrophotometry. Samples were prepared by soaking of the ground or the intact lens in deionized water. For potassium determinations HNO3 dissolution was also tried. Deionized water or CsCl aqueous solution was adopted for dilution for measurement. At absorbances up to 1.0 absorbance unit only measurements on HNO3 dissolved lenses diluted in deionized water were compatible with a linear absorbance function. At absorbances below 0.2-0.3 absorbance units all combinations were in agreement with a linear absorbance function. Grinding of the lens was found to increase the variability but did not change the level. HNO3 dissolution did not increase the level. Dilution with CsCl increased the sensitivity and decreased the variability. A schedule is proposed for determination of sodium and potassium in which lens samples are obtained by soaking in deionized water. Aqueous solution of CsCl is suggested for dilution.

Animals↗
PubMed1979

Precise spectrophotometry of small quantities of drugs in the presence of irreproducible quantities of impurities.

A variety of methods of spectrophotometric assay of compounds in the presence of one or more irrelevant impurities have been recently described, employing, for example, the use of Legendre polynomials and trigonometric functions. The use of least-squares solutions of overdetermined systems (Scheid 1968) in spectrophotometry is outlined, with conditions under which simplified overdetermined systems have been used for precise spectrophotometric assay of small quantities of drug substances in the presence of largely irreproducible quantities of irrelevant impurities.

Chemistry, Pharmaceutical↗
PubMed1995

Acid-base equilibria of 1,4-benzodiazepine 4-oxides by spectrophotometry.

Chlordiazepoxide (a 1,4-benzodiazepine 4-oxide) is an anxiolytic/hypnotic drug in clinical use. It was reported to be predominantly protonated at the N-oxide oxygen in acidic aqueous solutions at pH << 4.6 (pKa). We have studied the acid-base equilibria of three 1,4-benzodiazepine 4-oxides (chlordiazepoxide, diazepam 4-oxide, and nordiazepam 4-oxide) by ultraviolet-visible spectrophotometry. The results indicate that chlordiazepoxide is not protonated at the N-oxide oxygen, but rather at the nitrogen of an imine bond between C2 carbon and its nitrogen substituent in acidic media.

Chemical Phenomena↗
PubMed1967

Estimation of calcium and magnesium in serum and urine by atomic absorption spectrophotometry.

A method has been described for the estimation of calcium and magnesium in serum and urine using atomic absorption spectrophotometry. The precision and accuracy of the techniques have been determined and were found to be acceptable. THE RANGE OF VALUES FOR CALCIUM AND MAGNESIUM IN THE SERA OF NORMAL ADULTS WAS FOUND TO BE: serum calcium (corrected to a specific gravity of 1.026) 8.38-10.08 mg. per 100 ml.; serum magnesium 1.83-2.43 mg. per 100 ml.

Absorption↗
PubMed1969

Micro determination of plasma and erythrocyte copper by atomic absorption spectrophotometry.

The free and total plasma copper and total erythrocyte copper levels have been determined by simple, yet sensitive and highly specific methods, using atomic absorption spectrophotometry. For total copper determination, the copper was split from its protein combination in plasma or red cells by the action of hydrochloric acid at room temperature. The liberated copper was chelated by ammonium pyrrolidine dithiocarbamate and extracted into n-butyl acetate by shaking and the organic extract was aspirated into the atomic absorption spectrophotometer flame. The entire procedure was carried out in polypropylene centrifuge tubes, capped during shaking. For the free plasma copper measurement the hydrochloric acid step was omitted. Removal of the plasma or erythrocyte proteins was found to be unnecessary, and, in addition, the presence of trichloracetic acid caused an appreciable lowering of absorption. Using a double-beam atomic absorption spectrophotometer and scale expansion x 10, micro methods have been derived for determining the total copper of plasma or erythrocytes with 0.1 ml of sample, and the free copper of plasma with 0.5 ml. The macro plasma copper method requires 2 ml of plasma and is suitable for use with single-beam atomic absorption spectrophotometers. With blood from 50 blood donors, normal ranges of plasma and erythrocyte copper have been determined.

Blood Proteins↗
PubMed1988

Demonstration of aluminium in iliac bone: correlation between aluminon and solochrome azurine staining techniques with data on flameless absorption spectrophotometry.

The presence of excess aluminium was investigated in 204 samples of iliac bone from 197 patients with chronic renal failure by using the aluminon and solochrome azurine staining techniques. The results were compared with values obtained by atomic absorption spectrophotometry (AAS). Overall, the staining reactions correlated with the AAS data, but the solochrome azurine stain was positive more often than was the aluminon stain (in 90.6% and 62.3%, respectively, of bone samples with greater than the control group mean +3SD (that is, in the range 17.8 to 113.4 micrograms aluminium/g Solochrome azurine staining was consistently positive in all cases, with greater than 23.1 micrograms aluminium/g bone but the corresponding aluminon stain was occasionally inexplicably negative in this range. In some samples solochrome azurine was positive in parts of old unresorbed cement lines when the aluminon stain was negative and the bone aluminium concentration was within the normal range or slightly increased. The stains, particularly solochrome azurine, have the advantage over AAS in that they indicate the location as well as the presence of excess bone aluminium.

Aluminum↗
PubMed1978

Increased plasma levels of lead in patients with amyotrophic lateral sclerosis compared with control subjects as determined by flameless atomic absorption spectrophotometry.

The levels of lead in plasma were determined in 16 cases of amyotrophic lateral sclerosis (ALS) and 18 control subjects, using flameless atomic absorption spectrophotometry. The mean values were 0.52+/-0.22 microgram/100ml (ALS) and 0.37+/-0.13 microgram/100ml (controls), the difference is statistically significant (5% level). The values in both groups are lower than reported earlier for normal subjects. The findings are discussed against the background of the possible pathogenetic significance of retrograde axoplasmic flow in ALS.

Adolescent↗
PubMed1987

Reflectance spectrophotometry for the assessment of gastroduodenal mucosal perfusion.

Reflectance spectrophotometry in assessing gastroduodenal mucosal perfusion was evaluated. Ischemia without congestion, e.g., during hemorrhagic hypotension or celiac artery occlusion, was associated with a reduction in the indexes of mucosal hemoglobin concentration and of oxygen saturation. Ischemia with congestion, e.g., during portal vein occlusion, or in absolute ethanol or suction-induced mucosal lesions, was associated with an increase in the index of mucosal hemoglobin concentration but a reduction in the index of oxygen saturation. An increase in the index of mucosal hemoglobin concentration associated with a normal index of oxygen saturation was found in the postischemic hyperemia after release of celiac artery occlusion and during the sustained increase in corpus mucosal blood flow induced by vagus nerve stimulation. Thus reflectance spectrophotometric measurements reflected ischemia, without or with congestion, and hyperemia. Additionally, although regional differences in reflectance spectrophotometric measurements were demonstrated in the duodenal, antral, and corpus mucosa, such differences bore no consistent relationship to regional differences in blood flow demonstrated in previous studies.

Animals↗