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A spectrophotometric method is described for the determination of tilidine in its dosage forms (injection, drops, suppositories). The method is based on ion-pair extraction with chloroform at pH 3.5 using bromocresol green or bromophenol blue as the ion-pairing reagents. The spectrophotometric measurements are carried out at the absorption maxima at 415 and 411 nm, respectively.
In erythroblasts of varying maturational stages, cytoplasm stained bright yellow with bromochlorophenol blue. Since this staining reaction appeared to be distinctive among various cytological types of marrow cells, it may constitute a rapid method for visualization of cells of erythroid origin.
The stability of porcine insulin in biodegradable polymers, i.e., poly(DL-lactide-co-glycolide) 50:50 (50:50 DL-PLGA) and poly(L-lactide) (L-PLA) was investigated. Insulin encapsulated microspheres were fabricated from both polymers using double-emulsion-solvent evaporation and emulsion-solvent evaporation techniques and subjected to accelerated stability studies at 40 degrees C and 75% relative humidity. Porcine insulin was found to degrade in all microsphere formulations with an average of < 50% of the initial loading amount remaining intact at the end of 4 weeks. The two major degradation products observed in these formulations were determined to be A-21 desamido insulin and covalent insulin dimer with trace amounts of high molecular weight transformation products. In vitro release studies in phosphate buffered saline at 37 degrees C resulted in very slow and incomplete (< 30% in 30 days) release kinetics for all microsphere formulations. Extraction and analyses of the unreleased insulin within the microspheres revealed that an average of approximately 11% of the encapsulated insulin remained intact. The degradation products observed consisted of approximately 15% of three distinct deamidated hydrolysis products including A-21 desamido insulin, approximately 22% covalent insulin dimer, and trace amounts of high molecular weight transformation products. The degradation of porcine insulin within biodegradable polyester microspheres during stability and release studies can be attributed to the gradual decrease in the pH within the microspheres due to progressive polymer hydrolysis resulting in the production of DL-lactic and glycolic acids. The encapsulation of an acid-base indicator, bromophenol blue, in 50:50 PLGA microspheres (as a probe to estimate pH within the microspheres during accelerated stability studies) indicated that the pH decreased to approximately 3.8 after 3 weeks.
In this study, we have examined the acid-base characteristics of various citrate buffer systems alone and in the presence of the pH indicator dye, bromophenol blue, in aqueous solution, and after lyophilization to produce amorphous material. Fourier transform Raman and solid-state nuclear magnetic resonance spectroscopy have been used to monitor the ratio of ionized to un-ionized citric acid under various conditions, as a function of initial pH in the range of 2.65-4.28. Ultraviolet-visible spectrophotometry was used to probe the extent of proton transfer of bromophenol blue in the citrate buffer systems in solution and the amorphous state. Spectroscopic studies indicated greater ionization of citric acid and bromophenol blue in solution and the solid state with increasing initial solution pH, as expected. Fourier transform Raman measurements indicated the same ratio of ionized to un-ionized citrate species in solution, frozen solution, and the amorphous state. It is shown that the ratio of species at any particular initial pH is primarily determined by the amount of sodium ion present so as to maintain electroneutrality and not necessarily to the fact that pH and pK(a) remain unchanged during freezing and freeze drying. Indeed, for bromophenol blue, the relative ultraviolet-visible intensities for ionized and un-ionized species in the amorphous sample were different from those in solution indicating that the extent of protonation of bromophenol blue was significantly lower in the solid samples. It is concluded that under certain conditions there can be significant differences in the apparent hydrogen activity of molecules in amorphous systems.
The study of a number of diphenhydramine-dye systems was carried out in order to determine the most suitable precipitate for the turbidimetric determination of diphenhydramine using flow injection (FI). The reagent selected was Bromophenol Blue. The chemical and FI variables were optimised. The calibration graph was linear over the concentration range 50-230 p.p.m. of diphenhydramine hydrochloride. A number of interfering substances were also investigated.
The forward rate constant for the binding of Bromophenol blue to lysozyme is estimated to be 10(5)M-1 X S-1 at room temperature and ionic strength 0.01, from the line broadening of the 1H-NMR spectrum of Bromophenol blue at 270 MHz. The broadening of the 1H-NMR line corresponding to histidine-15 of lysozyme, in the presence of Bromophenol blue in solution at pH 4.5, suggests that the binding site of Bromophenol blue is not far from this protein residue. The extent of this broadening is pH-dependent and can be interpreted in terms of the average coulombic interaction between the dye and His-15. Since Bromophenol blue is known to inhibit lytic activity of lysozyme towards cell wall substrates, we can say that the region near His-15 may also be important for the efficient catalytic activity of lysozyme.
Nineteen fungi were tested for their ability to degrade aflatoxin B1 (AFB1). An extracellular enzyme from the edible mushroom Pleurotus ostreatus showed afaltoxin-degradation activity detected by thin-layer chromatography (TLC). An enzyme with this activity was purified by two chromatographies on DEAE-Sepharose and Phenyl-Sepharose. The apparent molecular mass of the purified enzyme was estimated to be 90 kDa by SDS-PAGE. Optimum activities were found in the pH range between 4.0 and 5.0 and at 25 degrees C. Also, degradation activity of several dyes in the presence of H2O2 was tested, resulting in the detection of bromophenol blue-decolorizing activity. Based on these data, we suggest this enzyme is a novel enzyme with aflatoxin-degradation activity. Fluorescence measurements suggest that the enzyme cleaves the lactone ring of aflatoxin.
The assay is based on the property of proteins to form a complex with bromophenol blue. In alkaline solution this complex shows an intensive blue colour which is well measurable at 598 nm. The method is especially useful for small volumes (5-50 microliter) with a content of 5-30 micrograms of protein. By modification the test can be also used for the measurement of higher amounts of protein.
A simple, economic, and available method for measuring protein in the urine is proposed. It is based on the capacity of bromophenol blue stain to form a complex with proteins in acid medium, which is characterized by absorption maximum at a wave-length of 597 nm. The proposed method is compared with the sulfosalicylic acid test and the Biuret method.
Protein chip technology provides a new and useful tool for high-throughput screening of drugs because of its high performance and low sample consumption. In order to screen elastase inhibitors on a large scale, we designed a composite microarray integrating enzyme chip containing chemical arrays on glass slides to screen for enzymatic inhibitors. The composite microarray includes an active proteinase film, screened chemical arrays distributed on the film, and substrate microarrays to demonstrate change of color. The detection principle is that elastase hydrolyzes synthetic colorless substrates and turns them into yellow products. Because yellow is difficult to detect, bromochlorophenol blue (BPB) was added into substrate solutions to facilitate the detection process. After the enzyme had catalyzed reactions for 2 h, effects of samples on enzymatic activity could be determined by detecting color change of the spots. When chemical samples inhibited enzymatic activity, substrates were blue instead of yellow products. If the enzyme retained its activity, the yellow color of the products combined with blue of BPB to make the spots green. Chromogenic differences demonstrated whether chemicals inhibited enzymatic activity or not. In this assay, 11,680 compounds were screened, and two valuable chemical hits were identified, which demonstrates that this assay is effective, sensitive and applicable for high-throughput screening (HTS).
An albumin selective urine strip based on bis (3',3''-diiodo-4', 4''-dihydroxy-5',5''-dinitrophenyl)-3,4,5,6-tetrabromo sulfonphthalein dye (DIDNTB) dye was examined in populations with clinical proteinuria. The relationship of albumin to the sum concentration of all protein in urine was found to vary widely even though the albumin concentration generally increased with the total protein concentration. The albumin reagent strips correlated well with immuno-nephrometric assays for albumin on specimens from hypertensives, diabetics, and renal disease which tended to have albumin contents of >/= 50.0%. High proteinuria concentrations of > 250 mg/l, with low albumin contents of </= 30%, occurred more frequently in cases of cancer, infection, and myeloma. The albumin strip read higher than the immuno assay in samples with high proteinuria and low albuminuria. The albumin strip was also less affected by albumin fragmentation than by the immunological assay. Overall, the albumin strip gave a lower risk of false negatives than a protein strip based on tetrabromophenol blue (TBPB) dye and was more sensitive to disease condition. The protein strip was not sensitive to low levels of albumin and the agreement between TBPB dye strip and the quantitative analysis was not as affected by the albumin content.
A test for demonstrating ringed sideroblasts was developed, using the dye bromchlorphenol blue. The test is based upon the ability of the dye to form a colored water-insoluble precipitate with iron. Because it is rapid and permits localization of ferritin-containing cytoplasmic inclusions, the reaction may be useful in the assessment of sideroblastic anemias.
Bromophenol blue and tetrabromophenol blue are two triphenylmethane dyes. Triphenylmethane derivatives and their structurally related compounds, such as fluoresceins and xathenes, are widely used as industrial dyes for foods, drugs, cosmetics, textiles, printing inks or laboratory indicators. Since a number of these types of dyes have been reported to be genotoxic, safety concerns on these two dyes of interest have been raised. Consequently, a battery of genetic toxicology assays, including the Ames Salmonella/microsome assay, L5178Y TK+/- mouse lymphoma assay, mouse micronucleus test and mitotic recombination assay with yeast Saccharomyces cerevisiae strain D5, has been performed on each of the two dyes. The results of the evaluations indicate that both bromophenol blue and tetrabromophenol blue were not active and can be considered non-genotoxic for the three genetic endpoints assessed (gene mutation, chromosome aberrations and primary DNA damage). Genetic activities in some structurally related compounds of these dyes have been reported but may be attributed to the presence of mutagenic impurities rather than the compound itself.
Three simple, accurate and sensitive spectrophotometric methods were developed for determination of enoxacin. The methods based on extraction of this drug into chloroform as ion pairs with sulphonphthalein dyes as bromophenol blue and bromocresol purple. The optimum conditions of the reactions were studied and optimized. The absorbance of yellow products was measured at 412 nm for enoxacin-bromophenol blue and 410 nm for enoxacin-bromocresol purple. Linearity ranges were found to be 2.0-20.0 microg ml(-1) for enoxacin-bromophenol blue and 0.77-17.62 microg ml(-1) for enoxacin-bromocresol purple. The detection limits were found to be 0.084 microg ml(-1) and 0.193 microg ml(-1) for enoxacin-bromophenol blue and enoxacin-bromocresol purple, respectively. The composition of the ion pairs was found 1:1 by Job's method. The developed methods were applied successfully for the determination of this drug in pharmaceutical preparation. The data obtained by developed methods were compared with the spectrophotometric method in literature. No differences were found statistically.
The spectrophotometric determination of trace amounts of chlorhexidine was carried out by liquid-liquid extraction using bromophenol blue with a flow system. The determination of chlorhexidine in the range of 1 x 10(-4) to 1 x 10(-5) M was possible with a sampling frequency of 40 samples per hour. The method was satisfactorily applied to the determination of chlorhexidine in pharmaceutical preparations.
In this paper an isochromatic dye ion-pairs extraction-catalytic spectrophotometric method for the determination of trace copper is studied. The method is based on the copper catalyzed oxidation of isochromatic dye ion-pairs formed by buthylrhodamine B and bromophenol blue by hydrogen peroxide in ammoniac medium. The reaction time, concentration of isochromatic dye ion-pairs in aqueous phase and degree of reaction are controlled by extraction equilibrium. The detection limit of this method is 0.02 microgram.L-1. The linear range is 0.1-10 micrograms.L-1. The method has been applied to the determination of trace copper in tumor tissue with satisfactory results.
Biocompatibility and diffusional characteristics of recently synthesized unique amphiphilic networks, i.e., copolymers of methacryloyl-capped polyisobutylene (MA-PIB-MA) with 2-(dimethylamino)-ethyl methacrylate (DMAEMA) have been examined. Excellent biocompatibility and biostability were obtained after 7 weeks with films containing 53-58% MA-PIB-MA implanted dorsally or abdominally in rats. In contrast, networks with higher or lower MA-PIB-MA contents showed decreased biocompatibility. These amphiphilic networks were also studied as potential implantable drug release systems. Bromophenol blue and folic acid were used as model compounds for drug release. Diffusion of these chemicals from loaded networks into water showed a marked pH dependence. Under specific well-defined conditions (MA-PIB-MA/DMAEMA content, pH, time range) release was observed to be independent of time (zero-order release).
The authors have modified R. Flores' method of protein measurements in solutions with bromophenol blue and suggest their modification for measurements of the blood serum total proteins. Experimental and clinical data have confirmed the significant effect of elevated blood levels of glucose and dextrans on the values of blood serum total proteins measured by the common biuret and refractometric methods. The bromophenol blue method is free from these disadvantages. The technique is recommended, first of all, for the infusion therapy departments. The method is highly specific, unsophisticated, and rapid, and is recommended for wide practical use in both inpatient and outpatient clinics. It can be used to measure proteins in other biologic fluids and solutions, provided its levels are not lower than 100 micrograms/ml.