Leuco Crystal Violet Research Articles

Sensitive Spectrophotometric Method for Determining Pentachlorophenol in Various Environmental Samples

A new, simple, and sensitive spectrophotometric method is described for determination of pentachlorophenol, a widely used insecticide and herbicide, in various environmental samples. The method is based on the reaction of pentachlorophenol with concentrated nitric acid to form chloranil, which liberates iodine from potassium iodide. The liberated iodine then selectively oxidizes leucocrystal violet to form crystal violet, which has an absorption maximum at 592 nm. Beer's law is obeyed over the concentration range of 0.1-1.6 micrograms pentachlorophenol/25 mL (0.004-0.064 ppm). The method was applied satisfactorily to determination of pentachlorophenol in air, water, plant material, textile effluent, and biological samples.

A sensitive spectrophotometric method for the determination of dithiocarbamate fungicide and its application in environmental samples

A sensitive spectrophotometric method based on the evolution of CS 2 and colour development by leuco crystal violet is described for the determination of dithiocarbamate fungicides, e.g. thiram, ziram and zineb. Dithiocarbamate fungicides release CS 2 on acid hydrolysis. This CS 2 is absorbed in ethanolic sodium hydroxide and forms xanthate. The xanthate formed is subsequently treated with potassium iodate and N-chlorosuccinimide, during which free iodine is liberated. Crystal violet dye was formed through selective oxidation of leuco crystal violet by liberated iodine, which has an absorbance maxima at 595 nm. The colour systems obey Beer's law in the range of 0.02–0.20, 0.02–0.24 and 0.04–0.32 ppm for thiram, ziram and zineb respectively. The molar absorptivity of the colour system were found to be 9.6×10 5, 1.1×10 6 and 6.8×10 5±100 l mol −1 cm −1 for thiram, ziram and zineb respectively. The method has been successfully applied to the determination of these dithiocarbamate fungicides in various environmental samples.

Mechanism of Dye Bleaching upon Laser Excitation of Crystal Violet Bound to Bovine Serum Albumin

Abstract The laser induced photobleaching of crystal violet non-covalently bound to bovine serum albumin yields leuco crystal violet and Michler’s ketone as reaction products. The first step of the bleaching process is postulated to be an electron or hydrogen atom transfer from the protein to the dye moiety.

Use of leuco crystal violet to enhance shoe prints in blood

Use of leuco crystal violet to enhance shoe prints in blood

Triplet-sensitized photobleaching of crystal violet

Crystal violet (CV +) and five related triarylmethane dyes quench triplet benzophenone ( 3K) in acetonitrile at diffusion-controlled rates ( k q ∼ 1 × 10 10 M −1 s −1). The interaction with CV + results in bleaching of CV + with a quantum yield, K Φ(−CV +), dependent on the concentration of CV + with a limiting value of 0.2 at high [CV +. Benzopinacol (K 2H 2) is also formed in this reaction but its yield decreases as [CV +] increases. In contrast, although both triplet anthracene ( 3An) and triplet naphthalene ( 3Nap) are also efficiently quenched by CV + at diffusion-controlled rates, there is only very inefficient fading, An Φ(−CV +) = 1.1 × 10 −5 . These results are interpreted in terms of a mechanism in which energy transfer to form triplet crystal violet, 3CV +, is the only major channel in the case of anthracene and naphthalene, i.e. 3CV + (like 1CV +) is unreactive. However, in the case of benzophenone, reduction of 3K by CV + with a rate constant k h = 1.4 × 10 9 M −1 s −1 to form ketyl radicals (KH .) can compete (about 14%) with this energy transfer. The ketyl radicals so formed can either reduce CV + to form leuco crystal violet (CVH) or form benzopinacol.

Glutathione Peroxidase-Like Activity of Simple Selenium Compounds. Peroxides and the Heterocyclic N-Oxide Resazurin Acting as O-Atom Donors.

Selenite and selenocystamine [(CyaSe)2] efficiently activate the decomposition of H2O2 y GSH and by other thiols, as demonstrated using a leuco crystal violet POD-based H2O2 assay which is applicable (unlike other assays) also in presence of thiols. The GPx-like activities were estimated to be 3.6 and 2.7 μmol H2O2/min per μmol SeO3 2- and (CyaSe)2, respectively. Both selenium compounds also activate reduction of the heterocyclic N-oxide resazurin (RN→O) to resorufin (RN) by GSH; H2O2 competes with reduction of this dye. GSSeH and CyaSeH, formed by interaction of GSH with SeO3 2- and (CyaSe)2, respectively, are likely to be the active reductants. CyaSeH, generated γ-radiolytically from (CyaSe)2, exhibits an absorption peak at 243 nm and is removed by H2O2 with a rate constant of 9.7x102 ᴍ-1 s-1, and slightly slower by hydroperoxides. We have no evidence for one-electron interactions between GSSeH or CyaSeH and H2O2, with formation of free radical intermediates, as previously proposed in the case of selenium-activated reduction of cytochrome c by GSH (Levander et al., Biochemistry 23, 4591-4595 (1973)). Our results can be explained by O-atom transfer from the substrate to the active selenol group. RSeH + H2O2 (RN→O)→RSeOH + H2O (RN), and recycling of RSeOH to RSeH (+ H2O) by GSH, analogous to the selenenic acid pathway of GPx. The substrate specificity appears to be different, however, in that GPx is unable to catalyse RN→O reduction, and GSSeH hardly catalyses the decomposition of cumene- or t-butyl-hydroperoxide; CyaSeH, on the other hand, is active also with the hydroperoxides. RN→O is reduced to RN also by certain oxidizing free radicals, e.g. by the thiyl CyaS·; O -atom transfer may in this case lead to the generation of reactive oxyl radicals.

Spectrophotometric determination of H 2O 2 in marine waters with leuco crystal violet

H 2O 2 in marine waters may be determined by measuring the absorbance of crystal violet (CV +), formed by the oxidation of leuco crystal violet (LCV) by H 2O 2 in the presence of the enzyme horseradish peroxidase, at 592 nm at pH 4. The detection limit and the precision of the method are about 0.02 μ M and ± 1% (at a concentration of 0.03 μ M), respectively. The results obtained by this method and by a widely used fluorimetric method agreed with each other well. After color development, samples together with the sub-samples for constructing the calibration curve may be stored for up to 5 days before their absorbances are measured without significantly changing the estimated concentration of H 2O 2.

Hydride transfer reactions of Michler's hydride with different π‐accetors

AbstractThe hydride transfer reactions of 4,4′‐bis(dimethylaminophenyl)methane (Michler's hydride) with p‐benzoquinones were studied. The rate of formation of Michler's Hydrol Blue was followed spectrophotometrically. The second‐order rate constants and the activation parameters were estimated. The formation of a charge‐transfer complex was observed at low temperatures. Stable and unstable paramagnetic species formed during the reactions were assigned by ESR spectroscopy. The observed kinetic behaviour and the stoichiometry were in line with those previously obtained for the systems involving Leuco Crystal Violet, Leuco Malachite Green and Leuco Bindschedler's Green. Hence the reaction is considered to proceed according to the so‐far accepted multi‐step mechanism. However, when tetracyano‐p‐quinodimethane or tetracyanoethylene was used as a π‐acceptor, a comparatively stable radical ion pair was formed as a result of a one‐electron transfer, followed by the gradual formation of Michler's Hydrol Blue. The kinetic behaviour and the stoichiometry of the reaction were examined, together with the enhanced kinetic effects of added triethylamine. A modified mechanism for these systems is proposed. The role of cyanomethylenes was found to be essentially different from that of p‐benzoquinones, and therefore π‐acceptors are divided into two groups of cyanomethylenes and p‐benzoquinones.

Mechanism of the hydride transfer reaction of leuco crystal violet with cyanomethylene acceptors

AbstractIn the hydride transfer of Leuco Crystal Violet to form the Crystal Violet, cation, the role of cyanomethylene acceptors was found to be essentially different from that of p‐benzoquinones, both previously believed to act as π‐acceptors in the same manner.

Extraction Spectrophotometric Determination of Sulfur(IV) in Atmosphere with Leuco Crystal Violet and N-Chlorosuccinimide

A new and sensitive method is described for the determination of sulfur(IV). Sulfur(IV) is passed through potassium iodate solution and liberated iodine is then reacted with N-chlorosuccinimide and leuco crystal violet. The crystal violet dye formed shows maximum absorbance at 605 nm and has apparent molar absorptivity of 5.2 × 104 mol liter−1 cm−1 Beer′s law is obeyed in the range of 0.006-0.09 μg ml−1 sulfur(IV). The dye is extractable in toluene and is stable for 2 days. The method is fairly reproducible with coefficient of variation ± 1.69% for five replicate analyses. The method is superior to the other reported methods in terms of manipulation, use of nontoxic and easily available reagents, and selectivity. The method has been applied for spectrophotometric determination of sulfur dioxide in polluted and ambient air samples.

The Effect of Changing Water Chemistry on Rates of Manganese Oxidation in Surface Sediments of a Temperate Saltmarsh and a Tropical Mangrove Estuary

The influence of changing water chemistry on manganese oxidation in surface sediments of a Sapelo Island, Georgia saltmarsh and an Andros Island, Bahamas mangrove swamp was examined. Surface sediment interstitial water salinities for sample stations in the mangrove swamp, and for saltmarsh creek bank sample sites on Sapelo Island were similar to that of the overlying waters. Intertidal sediments from high marsh sites on Sapelo Island were frequently exposed to the atmosphere on low tides, resulting in enhanced evaporation, and interstitial water salinities up to 105 were observed. Rates of microbial Mn oxide production were determined using the oxidation of leuco crystal violet in sediment/water suspensions. Sodium azide was added to some incubations to measure rates of abiotic Mn oxide formation. Overall, rates of Mn oxide production were much higher in saltmarsh sediments than in mangrove sediments. Within the mangrove swamp estuary, sediments approximately 15 km inland exhibited the highest rates of microbial Mn oxide production (station salinities of 0 and 8, 50 to 119 pmol mg dwt -1h -1, respectively), compared with sediments from the mouth of the estuary (station salinities of 24 and 34, 3 to 16 pmol mg dwt -1h -1, respectively). Suspension of mangrove sediments in water from different parts of the estuary, and in coastal seawater, had no affect on rates of Mn oxidation. In the saltmarsh rates of Mn oxidation were much higher for creek bank sediments than for sediments from high marsh sites (2·31 ± 0·28 SD and 0·45 ± 0·14 SD nmol mg dwt -1h -1, respectively). Suspension of creek bank sediments in coastal seawater diluted with distilled water had no affect on rates of Mn oxidation when compared with suspensions in natural saltmarsh creek water. Sediments from high marsh sites exhibited short-term enhancement of Mn oxidation when the coastal seawater used for suspensions was experimentally diluted to 24% of full strength with distilled water (final salinity 8). High marsh sediments experimentally exposed to hypersaline conditions, demonstrated decreasing rates of Mn oxidation as sediment suspension salinity reached 102. This inhibition of Mn oxidation appears to have been due to the effects of the concentrated bulk salts present in the water. Differences in organic or other trace components of the creek water appear to have had a negligible effect on rates of Mn oxidation.

Microbial Manganese Oxidation in Saltmarsh Surface Sediments Using a Leuco Crystal Violet Manganese Oxide Detection Technique

Microbial manganese (Mn) oxide production in surface sediments of a Georgia saltmarsh was directly measured using an assay involving the oxidation of 4,4′,4″-methylidynetris (N,N-dimethylaniline), leuco crystal violet (LCV), by Mn oxides to produce crystal violet. The assay exhibits high specificity for Mn oxides without interference by Mn(II) and is sufficiently sensitive to determine rates of Mn oxidation in surface sediment or saltmarsh creek water suspensions. Sample salinity affects crystal violet absorbance in the 0-25 salinity range and must be corrected for in Mn oxide determinations for estuarine samples of variable salinity. Other oxidants found to oxidize LCV slowly included Cl(I), Cr(III), I(V), Fe(III), and Mn(III), although the sensitivity of the assay for Mn(IV) oxides was found to be seven times greater than for Mn(III), and at least 100 times greater than for any of the other oxidants. Rates of abiotic Mn oxide production in sediment suspensions treated with either sodium azide or formalin, or autoclaved, were much slower than rates determined for untreated sediments. Sodium azide (7·7 mM) inhibited Mn oxide production in these sediment suspensions to rates between 5 and 10% of the rates of Mn oxidation determined for unamended suspensions. Manganese oxidation was highly temperature dependent, with maximal rates on a dry weight basis (8·9 nmol mg dwt-1 h-1), occurring at 60°C, and negligible activity at 100 and 0°C. Rates were also dependent on sample pH, with maximal rates at pH 6·7, decreasing to near 0 as the pH was lowered to approximately 3·0. For Mn(II) concentrations ranging from 9 to 91 μM, rates of Mn oxide production were independent of Mn(II) concentration, while Mn oxide production was inhibited at concentrations greater than 91 μM (e.g. by 25-40% at 450 μM). Rates of microbial Mn oxide production in surface sediment/saltmarsh creek water suspensions incubated under natural conditions of temperature, pH, and Mn(II) concentration ranged from 0·49 to 2·7 nmol mg dwt-1 h-1 and were nearly four times higher in surface sediments from creek banks than in sediments from the high marsh. The microorganisms associated with saltmarsh creek water particulate matter oxidized Mn(II) at rates intermediate to the values obtained for the two types of surface sediments, averaging 0·98 nmol mg dwt-1 h-1.

Spectrofluorimetric hydrodynamic voltammetry: the investigation of electrode reaction mechanisms

Spectrofluorimetric hydrodynamic voltammetry (SFV) is used to study the electroreduction of crystal violet in acetonitrile solution. The SFV technique is shown to have high sensitivity and to complement conventional electrochemical experiments through its identification of leucocrystal violet as an electrolysis product generated in low concentration in addition to the radical species formed as a one-electron adduct of the parent compound and previously thought to be produced exclusively. Mechanistic SFV studies show that the leucocrystal violet is formed via an EC process for which kinetic parameters are reported

Leaching of hydrogen peroxide from bleached bovine enamel

Accurately weighed bovine enamel slabs were individually immersed in 2 ml of 35% hydrogen peroxide for 1, 3, 5, 30, or 60 min. A control group was obtained by individual immersion of bovine enamel slabs in 2 ml of saline for 60 min. All samples were washed, dried, acid-etched with 37% phosphoric acid for 60 s, then washed and dried again. Two milliliters of double-distilled water were used for individual sample leaching. Leaching was done for 1, 5, 10, 20 min, or 7 days for the experimental groups and for 7 days for the control group. The samples of one of the experimental groups were leached for a second time for 1 min. A total of 112 samples was used in this study. Hydrogen peroxide was spectrophotometrically identified and quantified in all leaching solutions based on the oxidation reaction of leuco-crystal violet buffer solution by hydrogen peroxide, a reaction catalyzed by horseradish peroxidase. The results revealed a significant difference in the quantity of leached peroxide between bleached samples (irrespective of the duration of leaching) and control, saline-treated ones. No difference was observed in the quantity of leached peroxide between releached samples and control, saline-treated ones. However, these were small, random, and numerically insignificant. Statistically significant differences were also noted among some of the experimental groups. They were thought to hold no clinical significance. The results suggested that upon immersion, the complete leaching of peroxide from bleached enamel occurs rapidly.

Histamine and Histamine-Producing Bacteria in Retail Swiss and Low-Salt Cheeses

Seventeen low-sodium and low-salt cheeses and 50 Swiss cheeses were surveyed for histamine and histamine-producing organisms. Two of the low-salt cheeses and nine of the Swiss cheese samples contained greater than 45 mg histamine per 100 g of cheese, as determined by the AOAC method. Over 800 total colonies were randomly chosen and screened for histamine production by the leucocrystal violet detection method following their initial isolation from MRS media. However, none of the leucocrystal violet-positive isolates from the low-salt cheese and only five from the Swiss cheese were found to produce histamine in MRS broth supplemented with L-histidine. Proteolysis (determined by the trinitrobenzene-sulfonic acid assay) was also measured in the low-salt cheeses in an attempt to further understand the role of free histidine as a substrate with respect to histamine content. In general, the cheese samples with high histamine levels also had the high values for trichloroacteic acid-soluble nitrogen. However, the highest proteolysis values did not necessarily correlate with the highest histamine values. Two samples of low-salt Swiss cheese that had high trichloroacetic acid-soluble nitrogen (greater than 200 μmoles glycine equivalents per g of cheese) contained less than 15 mg histamine per 100 g cheese.

Reactions of Leuco Bindschedler’s Green with Some p-Benzoquinones in Acetone

Abstract The rate of reaction between Leuco Bindschedler’s Green (BGH) and some p-benzoquinone derivatives (Q’s) in acetone was followed spectrophotometrically by means of the stopped-flow technique. The reaction was found to proceed reversibly. The rate data and the activation parameters for the forward and backward reactions were estimated. The rates were susceptible to the oxidation–reduction potentials of the Q’s. The absorption spectra observed at low temperatures indicated that three kinds of intermediates were involved. Radicals formed during the reaction were detected by the ESR method. The results have been discussed in relation to those previously obtained for systems involving Leuco Crystal Violet (LCV) and Leuco Malachite Green (LMG).

Post-column reaction for simultaneous analysis of chromatic and leuco forms of malachite green and crystal violet by high-performance liquid chromatography with photometric detection

The chromatic and leuco forms of malachite green and crystal violet were readily separated and detected by a sensitive and selective high-performance liquid chromatographic procedure. The chromatic and leuco forms of the dyes were separated within 11 min on a C 18 column with a mobile phase of 0.05 M sodium acetate and 0.05 M acetic acid in water (19%) and methanol (81%). A reaction chamber, containing 10% PbO 2 in Celite 545, was placed between the column and the spectrophotometric detector to oxidize the leuco forms of the dyes to their chromatic forms. Chromatic and leuco malachite green were quantified by their absorbance at 618 nm; and chromatic and leuco Crystal Violet by their absorbance at 588 nm. Detection limits for chromatic and leuco forms of both dyes ranged from 0.12 to 0.28 ng. A linear range of 1 to 100 ng was established for both forms of the dyes.

Biodegradation of triiodophenol by cell-free extracts of a pentachlorophenol-degrading Flavobacterium sp

Pentachlorophenol (PCP) degrading Flavobacterium sp. ATCC 39723 was found to degrade other polyhalogenated phenolic compounds, including triiodophenol, tribromophenol, and trichlorophenol. Each Compound was able to induce the degradation of the other compounds. A PCP Flavobacterium sp. mutant, F-2, was unable to degrade any of the halogenated compounds. The results suggest that all of the polyhalogenated phenols were degraded by the same enzyme system. This observation led us to exploit the sensitive leuco crystal violet assay, which measures the iodide released from triiodophenol, Cell free extracts from PCP-induced cells were able to release lodide from triiodophenol. The reaction required NADPH and oxygen.

Liquid chromatographic assay of plasma hemoglobin based on oxidative quenching of serotonin by pseudoperoxidase effect of hemoglobin.

A high-performance liquid chromatographic method for the determination of hemoglobin is described. The method uses the pseudoperoxidase effect of hemoglobin in the presence of hydrogen peroxide and is based on the oxidative quenching of the native fluorescence of serotonin. The determination range for hemoglobin was between 3 and 100 mg/dl. The intra-assay coefficient of variation was 2.04% or less, with a mean recovery of 102.0% at hemoglobin concentrations of 17.28, 43.20, and 69.12 mg/dl. A close correlation was found between the results of this method and the leuco crystal violet method.

Coulombic effects on photoinduced electron-transfer reactions between benzophenone derivatives and leuco crystal violet

Photo-oxidation of leuco crystal violet (DH) to crystal violet dye (D +) was studied in the benzophenone (BP) derivatives—DH—acetonitrile systems. The BP derivatives with a tetraalkylammonium substituent (R 3N +; I, II and III exhibited much higher photo-oxidizing efficiencies than BP itself. Under deaerated conditions, nearly quantitative photo-oxidation of DH by the absorbed photons was attained by I whereas BP itself showed a poor ability to photo-oxidize DH. Addition of tetraalkylammonium salts (R 4NX), such as cetyltrimethylammonium chloride or tetra- n-butylammonium chloride to the BPDH system also enhanced the quantum yield by a factor of about 10. The enhancement of quantum yield by the use of I, II or III, or by the addition of R 4NX to BP was interpreted as due to stabilization of the BP anion radical or the DH cation radical by ion pairing with the tetraalkylammonium substituent on BP or with the added R 4NX. In the case of I, II and III, the electrostatic repulsion between the DH cation radical and the R 3N + substituent on BP, which enchanced the charge separation efficiency, was the main cause of the high photo-oxidation yields.