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The metabolism and mode of action of gentian violet.

Gentian violet has been used in medicine for almost 100 years: as an antiseptic for external use, as an antihelminthic agent by oral administration, and more recently, as a blood additive to prevent transmission of Chagas' disease. To date, no serious side effects have been reported when used externally. However, oral administration can cause gastrointestinal irritation, and intravenous injection can cause depression in the white blood cell count. Surprisingly, no acute toxic side effects were reported after administration of large amounts of gentian violet-treated blood. No studies have been done on long-term effects (chronic toxicity, carcinogenicity) of gentian violet-treated blood either in humans or in laboratory animals. Gentian violet is a mutagen, a mitotic poison, and a clastogen. The carcinogenic effects of gentian violet in rodents have been reported recently. In addition, a number of triphenylmethane-classed dyes, of which gentian violet is a member, have been recognized as animal and human carcinogens. A photodynamic action of gentian violet, apparently mediated by a free-radical mechanism, has been described in bacteria and in T. cruzi. However, the main target of gentian violet toxicity in the dark is the mitochondrion. Gentian violet is actively demethylated by liver microsomes from different animals and is reduced to leucogentian violet by intestinal microflora. Although the first process may represent a detoxication reaction, the second pathway may have toxicological significance because the completely demethylated derivative leucopararosaniline has been demonstrated to be carcinogenic in rats. A free-radical derivative of gentian violet is also formed by the action of rat liver microsomes, but whether this radical is involved in the cytotoxic effects of gentian violet in mammalian cells remains to be elucidated. Other pathways of gentian violet metabolism have recently been investigated that involve its oxidative N-demethylation by peroxidases. The N-demethylation of gentian violet by prostaglandin synthetase deserves further study. In this regard, the PGS system is being studied as an alternative activating pathway in xenobiotic metabolism because some carcinogenic intermediates can be formed during this cooxidation reaction.

Animals↗

In vitro antifungal activity of gentian violet.

Gentian violet, a compound that gained wide acceptance in the poultry industry as a mold inhibitor in feed, was tested for its activity against eight aflatoxigenic strains of Aspergillus flavus and A. Parasiticus. In a simple nutrient medium, it completely inhibited growth of all strains at 8 micrograms/ml. When chicken feed was added to the medium, the inhibition by the same concentration ranged from 12 to 44% depending on the strain. Observations suggested that both the feed and the fungi were involved in the resistance conferred by chicken feed. Growth and aflatoxin production on moist rice was inhibited in one strain but not in a second strain. Inoculating autoclaved chicken feed of varying moisture contents with pure cultures revealed that the activity of gentian violet was better at high moisture contents than at low moisture contents approximating those found in poultry feed. It was not possible to assess the activity of gentian violet in unaltered chicken feed. Attempts failed because the formation of reproducible levels of aflatoxin could not be attained.

Animal Feed↗

Determination of leucogentian violet and gentian violet in catfish tissue by high-performance liquid chromatography with visible detection.

A sensitive analytical procedure for the determination of residues of leucogentian violet (LGV) and gentian violet (GV) in catfish tissue is presented. Frozen (-20 degrees C) catfish fillets were cut into chunks and then blended in a Waring blender. A 10-g amount of catfish muscle tissue was homogenized and extracted with acetonitrile-buffer, partitioned against methylene chloride, and cleaned up on tandem neutral alumina and propylsulfonic acid cation-exchange solid-phase extraction cartridges. Samples of 100 microliters (0.5 g equiv.) were chromatographed isocratically in 15 min using an acetonitrile-buffer mobile phase on a cyano phase column in-line with a post-column PbO2 oxidation reactor. The PbO2 post-column reactor efficiently oxidized the LGV to the chromatic GV permitting visible detection at 588 nm for both LGV and GV. Linearity was demonstrated with standards over the range 0.5-50 ng per injection. Recoveries of LGV and GV from catfish tissues fortified at 20, 10, and 1 ng/g were 83.1 +/- 1.2, 78.4 +/- 4.0, 84 +/- 8 and 92.7 +/- 1.8, 95.0 +/- 2.2, 93 +/- 2 (mean +/- S.D., n = 4), respectively.

Animals↗

[Experimental study of parotid gland injected with gentian violet].

OBJECTIVE: The Gentian Violet was used for treating the chronic parotitis since 1960's, but the dosage and the histologic changes of the tissues were undefined until now. We try to study the histologic changes of the salivary glands, heart, liver and kidneys after large volume of gentian violet was injected into the parotid gland through the duct. METHODS: 5 ml of Gentian Violet was injected to each side of 6 dogs. The macropathologic and histologic examination of the salivary glands, heart, liver and kidneys were performed at 30 min, 60 min, 120 min, 21 days and 9 months separately after the perfusion. RESULTS: Local swelling was obvious in two or three days after infusion and subsided in one week in most of the dogs. Degeneration of the gland took place after 20 days. After 9 months, the parotid gland tissues were replaced by the connective tissues. On the other hand, no histologic change was observed in heart, liver and kidney in the investigation. CONCLUSION: The Gentian Violet can cause degeneration of the parotid gland including the duct and replaced by connective tissues later. There is no obvious changes were noted in other organs.

Animals↗

Chronic toxicity and carcinogenicity studies of gentian violet in mice.

Gentian violet is a dye belonging to a chemical class known as the di- and triaminophenylmethanes. Although it has been used for many years for the control of fungal and intestinal parasites, for various uses in veterinary medicine, and as an additive to the feed of chickens to inhibit propagation of mold and fungus, very few long-term toxicity data are available. A life span dosing study of gentian violet in the diet of 720 males and 720 females of B6C3F1 mice (C57BL/6 X C3H) at dose levels of 0, 100, 300, and 600 ppm was done to determine its toxicity and carcinogenicity. Sacrifices were conducted after 12, 18, and 24 months of continuous dosing. There was no effect on food consumption or body weight gain; however, a dose effect was noted for mortality rates. Mortality (adjusted for sacrifices) in the controls of both sexes was less than 15% at 24 months, but was approximately 64% in the females and 23% in the males given the high dose. Females appeared to be more susceptible than males. A positive dose response for hepatocellular carcinoma was noted in males at 24 months and in females at 18 and 24 months. Statistical tests for dose-related trends with respect to mortality due to liver neoplasms, prevalence of liver neoplasms, and time to onset of liver neoplasms showed positive trends in both males and females. Other dose-related toxicological responses, particularly in the female mice, included erythropoiesis in the spleen, atrophy of the ovaries, adenoma of the Harderian gland, and the presence of type A reticulum cell sarcomas in the urinary bladder, uterus, ovaries, and vagina. The estimation of risk of 10(-6) over background for malignant liver neoplasms using linear extrapolations showed a lower bound on the virtually safe dose (VSD) to be 2 ppb for the female mice and 1 ppb for the male mice. For benign and malignant liver tumors together, the lower bound on the VSD was essentially the same as for malignant liver neoplasm alone. Under the conditions of the experiment described above, gentian violet appears to be a carcinogen in mice at several different organ sites.

Animals↗

Further study of the genetic toxicity of gentian violet.

The genetic toxicity of gentian violet was studied with the Ames and the Rosenkranz bacterial assays as well as the cytogenetic assays (Chinese hamster ovary cells in vitro in the presence of rat-liver S-9 fractions, the chicken-embryo and mouse-bone-marrow cells in vivo). Gentian violet was found to be toxic but not mutagenic in the Ames assay. However, it was active in the Rosenkranz assay causing reparable DNA damage. The presence of S-9 in the in vitro cytogenetic assay and in the bacterial assays showed that the activity of gentian violet could be reduced or eliminated. In the in vivo assays, gentian violet was not clastogenic and failed to induce sister-chromatid exchanges. However, gentian violet proved to be highly toxic to growing chick embryos at high dosage and depressed mitotic activities in mouse bone marrow after prolonged treatment. Our study suggested that gentian violet can be inactivated by the liver detoxification system. However, it is potentially hazardous to cells that are exposed to the dye directly (e.g. skin epithelium and cell lining of the gastrointestinal tract).

Animals↗

Simultaneous determination of malachite green, gentian violet and their leuco metabolites in catfish or trout tissue by high-performance liquid chromatography with visible detection.

A sensitive analytical procedure for the determination of residues of leucomalachite green (LMG)-malachite green (MG) and leucogentian violet (LGV)-gentian violet (GV) in catfish or trout tissue is presented. Frozen (-20 degrees C) fish fillets were cut into small pieces and blended in a Waring blender. A 20-g amount of homogenized fish tissue was extracted with acetonitrile-buffer, partitioned against methylene chloride, and cleaned up on tandem neutral alumina and propylsulfonic acid cation-exchange solid-phase extraction cartridges. Samples of 100 microliters (0.8 g equiv.) were chromatographed isocratically in 10 min using an acetonitrile-buffer mobile phase on a short-chain deactivated (SCD) reversed-phase column (250 x 4.6 mm I.D.) in-line with a post-column PbO2 oxidation reactor. The PbO2 post-column reactor efficiently oxidized LMG to the chromatic MG, and LGV to the chromatic GV permitting visible detection at 588 nm for all four compounds. Linearity was demonstrated with standards over the range of 0.5-50 ng per injection. Recoveries of LMG, MG, LGV and GV from catfish tissues fortified at 10 ng/g were 75.4 +/- 3.0, 61.3 +/- 4.1, 72.6 +/- 3.7 and 87.9 +/- 2.5, respectively, while trout tissues fortified at 10 ng/g yielded recoveries of 82.6 +/- 2.3, 48.6 +/- 1.8, 72.1 +/- 2.1 and 83.8 +/- 4.6 (mean +/- S.D., n = 4), respectively.

Aniline Compounds↗

Interactions during inhibition of growth of Aspergillus parasiticus by gentian violet.

The interrelationships of gentian violet and the growth of Aspergillus parasiticus NRRL 2999 were investigated with static cultures in a yeast extract-sucrose liquid medium. The dye was fungistatic but not fungicidal at concentrations up to 32 microgram/ml. Growth inhibition was dependent on the concentration of the added dye and on the age of the culture. At 8 microgram/ml with a spore inoculum, growth was inhibited for 6 days while 32 microgram/ml added to a 4-day-old culture inhibited for only 24 hr. Initiation of a normal rate of growth coincided with disappearance of the dye from the medium. The disappearance followed first order kinetics. The dye appeared to bind to the mycelium prior to its apparent disappearance from the medium. Gentian violet appeared to inhibit spore germination by inhibiting a critical point in the spore swelling phenomenon. In addition, growth subsequent to germination was retarded.

Animals↗

Physiological effects of gentian violet on broiler chickens.

The effects of dietary gentian violet upon certain physiological parameters of broiler chickens were studied. Gentian violet exhibited no effect upon growth rates or feed conversion ratios at dietary levels of 16, 32, or 64 micrograms/g. In vitro intestinal absorption of methionine and glucose was also unaltered. Dietary gentian violet significantly increased hemoglobin concentration without an effect on packed cell volume. Furthermore, commercial gentian violet containing preparations, when incorporated into the diet, resulted in increased intestinal absorption of Fe59 but this increase was dependent upon type of inert carrier used. Dietary gentian violet alleviated some of the growth suppression caused by dietary aflatoxin; however, no effect was observed on plasma pigmentation. These data suggest that dietary gentian violet possesses effects other than those for which it has been traditionally employed.

Animals↗

A study of the feasibility of the use of gentian violet as a fungistat for poultry feed.

A 96% formulation of gentian violet was fungistatic to Aspergillus flavus, Alternaria sp., Candida albicans, Fusarium moniliforme, and Penicillum citrinum when incorporated into corn meal agar at 6.5, 39.0, and 156.0 p.p.m. A. flavus, Alternaria sp., F. roseeum, and P. citrinum were inhibited on corn meal agar which had been amended with 12.8, 26.6, and 38.4 p.p.m. of gentian violet in a 2.54% commercial liquid concentrate of gentian violet employed as the active ingredient in a commercial poultry feed additive. Sporulation of these fungi was not inhibited by either of the formulations of gentian violet at any concentration. An anhydrous gentian violet poultry feed additive (containing 2.05% gentian violet) significantly reduced the numbers of A. flavus and F. moniliforme propagules recovered from artificially contaminated feed samples treated with 1,000, 2,000, and 4,000 g./1000 kg. of the additive (20, 40, and 80 p.p.m. of gentian violet, respectively) and stored for 12 weeks at 8.5% moisture and compared with the numbers of propagules recovered from contaminated-unamended samples. The numbers of propagules of F. moniliforme and C. albicans recovered from the pre-mix amended samples held for 12 weeks at 19% moisture also were fewer (significantly fewer for F. moniliforme) than those recovered from the contaminated, unamended feed. Amending the A. flavus or F. moniliforme contaminated feed samples did not reduce the numbers of propagules recovered below the numbers present in the original feed samples. GV was fungistatic to the test fungi under the conditions in these studies, since it inhibited, but did not prevent development of the test fungi. A. flavus and F. moniliforme were the fungi isolated in greatest numbers from the original sample.

Animal Feed↗

Corneal vital staining with gentian violet.

We have used 0.5% gentian violet solution as a corneal vital stain in 112 patients with variable degrees of corneal involvements and in 40 normal eyes as control. Gentian violet stained the epithelial defects and degenerated epithelial cells of cornea. The stain persisted 3-5 minutes and disappeared by 8-10 minutes. There was no cross infection from dye use. The dye did not hamper usual process of repair of corneal lesions. It is found highly feasible, with a sensitivity about 100% and specificity about 95%, with some minor side effects. Thus it could be an excellent method in community ophthalmology for early diagnosis of corneal affections, thereby commencing prompt and appropriate treatment in early stage.

Corneal Diseases↗

High-performance liquid chromatography of gentian violet, its demethylated metabolites, leucogentian violet and methylene blue with electrochemical detection.

High-performance liquid chromatographic conditions are reported for the electrochemical detection (ED) of Gentian Violet, its demethylated metabolites, Leucogentian Violet and Methylene Blue. Gentian Violet, its demethylated metabolites and Leucogentian Violet were separated within 14 min on a cyano column eluted isocratically with methanol-buffer (60:40) as the mobile phase. ED responses for Gentian Violet, Leucogentian Violet and Methylene Blue were linear over the ranges 0.54-6.75, 0.50-25.2, and 5.7-285 ng, respectively. Under these conditions, the compounds were eluted in the following order: Leucogentian Violet, N"-2-tetra-methylparaosaniline chloride, N'-1-tetramethylpararosaniline chloride, pentamethylpararosaniline chloride and Gentian Violet. Methylene Blue and Gentian Violet had essentially the same retention time under these parameters. The detection limit for Gentian Violet, its demethylated metabolites and Leucogentian Violet was determined to be 0.1 pmol. A detection limit of 3 pmol was established for Methylene Blue. Detector response, elution, separation, linearity and sensitivity of detection are discussed.

Chemical Phenomena↗

Prevention of Chagas' disease resulting from blood transfusion by treatment of blood: toxicity and mode of action of gentian violet.

Blood transfusion is the second most important mechanism of transmission of Chagas' disease. Gentian violet, a cationic dye, is currently used in blood banks in endemic areas in attempts to eliminate such transmission. A photodynamic action of gentian violet has been demonstrated in Trypanosoma cruzi. Visible light causes photoreduction of gentian violet to a carbon-centered radical. Under aerobic conditions this free radical autooxidizes generating superoxide anion whose dismutation yields hydrogen peroxide. This photodynamic action of gentian violet is thus probably mediated by the oxygen reduction products. Since irradiation with visible light in the presence of sodium ascorbate reduces the effective dose and time of contact of the dye with T. cruzi-infected blood, a possible application of these findings can be envisaged. In addition to this photodynamic action, an uncoupling effect of gentian violet on mitochondrial oxidative phosphorylation has been described in rat liver and T. cruzi mitochondria. Gentian violet released respiratory control, hindered ATP synthesis, enhanced ATPase activity, released the inhibition of State 3 respiration by oligomycin, and produced swelling of isolated rat liver mitochondria or T. cruzi mitochondria in situ. Taken together, these results indicate that the T. cruzi mitochondrion is the main target of gentian violet toxicity in the dark.

Animals↗

Outer penetration barrier of Escherichia coli K-12: kinetics of the uptake of gentian violet by wild type and envelope mutants.

Wild-type strains of Escherichia coli K-12 adsorb gentian violet to the cell surface, but the dye is not transported into the cytoplasm. However, when some mutants that have an altered outer membrane are exposed to gentian violet, the dye is also found in the ribosomal fraction. The transport into the cytoplasm is inhibited at 0 C and requires that the concentration of gentian violet exceeds a threshold value. The initial rate of uptake as well as the amount of gentian violet found in the cytoplasm increases with the concentration of the dye in the medium. The rate of transport of the dye into the cytoplasm is much lower for stationary mutant cells than for exponentially growing cells. The rate of uptake into the cytoplasm increases with increasing deficiency of carbohydrate in the lipopolysaccharide (carbohydrate content lpsB > lpsA > galU). However, other components are also responsible for the barrier since an envA mutant which is not altered in the lipopolysaccharide carbohydrates show an extremely rapid uptake of the dye. The rate of uptake for the envA mutant was the highest found and the same as that of spheroplasts. Growth in the presence of agents affecting the murein sacculus, e.g., lysozyme and sublethal concentrations of penicillin, increased the rate of uptake of gentian violet. Brief treatments with tris(hydroxymethyl)aminomethane-ethylenediaminetetraacetic acid drastically impaired the barrier function. Inhibition of protein synthesis by chloramphenicol also opened the barrier to gentian violet. In conclusion, the outer part of the bacterial envelope is a penetration barrier for gentian violet and probably also for other substances. The lipopolysaccharide, the murein and also other components are important for the function of this barrier. Resistance to gentian violet was found to be inversely correlated to the rate of penetration of the dye into the cytoplasm.

Bacterial Proteins↗

Gentian violet solution for staining the anterior capsule.

PURPOSE: To evaluate the histopathological changes after injecting gentian violet solution into the anterior chamber of rats and to describe a technique that uses gentian violet to allow a clear view of the anterior capsule during continuous curvilinear capsulorhexis (CCC) in human eyes with white mature cataract. SETTING: Department of Ophthalmology, University of Dicle, Diyarbakir, Turkey. METHODS: In this masked, experimental study (first stage), 0.05 mL of gentian violet 0.01% or 0.001% solution or balanced salt solution (BSS) (control group) was injected into the anterior chamber of 30 eyes of 30 Wistar albino rats. One, 24, and 48 hours after injection, 4 eyes in each group and 2 eyes in the control group were enucleated, and histopathological examination was performed. In the second stage, these solutions were used for staining the anterior capsule in the 18 human eyes with white mature cataract. The success rate of CCC and intraoperative and postoperative complications were evaluated. RESULTS: Histopathological examination revealed no pathology in any group. A CCC was completed in all cases. No intraoperative or postoperative complications were observed in human eyes except mild corneal edema and mild inflammatory reaction in the anterior chamber that improved within 1 week. Mean follow-up was 3.4 months. Visualization of the anterior capsule was better with gentian violet 0.01% solution. CONCLUSIONS: Gentian violet solutions at 0.01% and 0.001% concentrations had no evident toxic effect that caused significant histopathological changes. The staining technique was practical and helped the surgeon visualize the anterior capsule. However, gentian violet may have adverse effects that lead to corneal edema.

Adult↗

Factors influencing antifungal activity of gentian violet in poultry feed and ingredients.

The inhibition of fungal activity in poultry feed and ingredients by gentian violet was investigated by measuring respiratory CO2 liberated into the headspace gas above samples of feed and ingredients. Inhibition by gentian violet depended on 1) the lot of corn meal, 2) the concentration of gentian violet, 3) the moisture of the substrate, 4) time of incubation, 5) particle sizes of corn meal and gentian violet, and 6) temperature of a short heating episode mimicking the feed-pelleting process. These same factors were reported earlier to control and limit the activity of organic acid mold inhibitors. These results imply that differences between gentian violet and other mold inhibitors used in poultry feed are quantitative rather than qualitative.

Animal Feed↗

Cytogenetic toxicity of gentian violet and crystal violet on mammalian cells in vitro.

The cytogenetic toxicity of gentian violet in Chinese hamster CHO cells in vitro has been studied by analyzing (1) squash preparations from direct fixation for recording mitotic anomalies and (2) air-dried preparations (with colcemid--hypotonic pretreatments) for recording metaphase chromosome aberrations. It was concluded that this compound is a mitotic poison as well as a clastogen in vitro. Its clastogenic property was confirmed in 5 other different mammalian cel types. 10 samples of different gentian violet and crystal violet were surveyed and all were shown to be clastogenic. Unless in vivo studies prove otherwise, gentian violet and crystal violet should be regarded as biohazardous substances.

Animals↗