WHO FOOD ADDITIVES SERIES: 69 Prepared by the Seventy-eighth meeting of the Joint FAO/WHO Expert Committee on Food Additives (JECFA) GENTIAN VIOLET page 3-34 Toxicological evaluation of certain veterinary drug residues in food , 2 7 The summaries and evaluations contained in this book are, in most cases, based on o. N unpublished proprietary data submitted for the purpose of the JECFA assessment. A registration es authority should not grant a registration on the basis of an evaluation unless it has first received i r e authorization for such use from the owner who submitted the data for JECFA review or has received the data on which the summaries are based, either from the owner of the data or from es es S a second party that has obtained permission from the owner of the data for this purpose. v i t i Add d World Health Organization, Geneva, 2014 oo F O O 6 1 H 0 W 2 GENTIAN VIOLET First draft prepared by Mr John Reeve 1 and Dr Susan Barlow 2 1 Science and Risk Assessment Branch, Ministry for Primary Industries, Wellington, New Zealand 2 Consultant, Brighton, East Sussex, England, United Kingdom 1. Explanation ........................................................................................... 4 2. Biological data ...................................................................................... 4 2.1 Biochemical aspects ....................................................................... 4 2.1.1 Absorption, distribution and excretion ................................... 4 (a) Mice ................................................................................. 4 (b) Rats ................................................................................. 5 2.1.2 Biotransformation .................................................................. 7 (a) Bacteria ............................................................................ 7 (b) In vitro .............................................................................. 7 (c) Mice ................................................................................. 8 (d) Rats ................................................................................. 8 2.2 Toxicological studies ....................................................................... 8 2.2.1 Acute toxicity ......................................................................... 8 2.2.2 Short-term studies of toxicity ................................................. 9 (a) Rats ................................................................................. 9 (b) Dogs ................................................................................ 9 2.2.3 Long-term studies of toxicity and carcinogenicity.................. 10 (a) Mice ................................................................................. 10 (b) Rats ................................................................................. 13 2.2.4 Genotoxicity .......................................................................... 17 2.2.5 Reproductive and developmental toxicity.............................. 22 (a) Multigeneration reproductive toxicity ............................... 22 (b) Developmental toxicity ..................................................... 23 2.2.6 Special studies ...................................................................... 25 2.3 Observations in humans ................................................................. 25 3. Comments ............................................................................................ 26 3.1 Biochemical data ............................................................................ 26 3.2 Toxicological data ........................................................................... 26 4. Evaluation ............................................................................................. 30 5. References ........................................................................................... 31 - 3 - 4 GENTIAN VIOLET 1. EXPLANATION Gentian violet (Chemical Abstracts Service No. 548-62-9) has many common names, including CI Basic Violet 3, crystal violet and methyl violet 10B. It is a triphenylmethane dye with antibacterial, antifungal and anthelminthic properties. Gentian violet has been used for the treatment of fungal and parasitic infections in fish and topically for skin and eye infections in livestock. It was previously used in poultry feeds to inhibit the growth of mould and fungus; however, several countries have withdrawn approval or registration of this use. In humans, gentian violet has been used as a hair dye, to treat gut parasites and for topical fungal treatment. It has also been used in human medicine to treat blood held for transfusions in order to prevent the transmission of Chagas disease caused by Trypanosoma cruzi . It also has activity as a topical antiviral agent. Gentian violet is used in industrial processes for wood, leather, silk, nylon, paper and ribbon tapes and also as a biological stain. Contamination of the environment can occur, as about 1015% of all dyes are lost directly to wastewater in the dyeing process. Gentian violet in water originating from contamination as a result of its industrial applications or from its illegal use in aquaculture is efficiently taken up from the water by fish. Gentian violet has not previously been evaluated by the Joint FAO/WHO Expert Committee on Food Additives (JECFA). It was evaluated by the Committee at the current meeting at the request of the Twentieth Session of the Codex Committee on Residues of Veterinary Drugs in Foods (FAO/WHO, 2012), which asked for advice as to whether an acceptable daily intake (ADI) can be established and whether the continued use of gentian violet in food-producing animals is safe for humans. 2. BIOLOGICAL DATA 2.1 Biochemical aspects 2.1.1 Absorption, distribution and excretion Docampo & Moreno (1990) reviewed the metabolism and mode of action of gentian violet. No data were available on skin absorption of gentian violet in food-producing animals when the drug is applied topically, although Diamante et al. (2009) considered gentian violet to be poorly absorbed through human skin. (a) Mice Groups of 12 male and 12 female B6C3F1 mice were housed three per metabolism cage and given 14 doses of 14 C-labelled gentian violet (94.8% gentian violet and 5.2% pentamethylpararosaniline) at 12-hour intervals by gavage. The total gentian violet doses were 5.6 mg/kg body weight (bw) (0.72 MBq/animal) and 7.1 mg/kg bw (0.72 MBq/animal) for males and females, respectively. The mice were killed 2 hours after receiving the final dose. Urine, faeces, liver, kidney, muscle, testes or ovaries and a fat sample were collected during the study, and radioactivity was measured ( Table 1 ). The data show that gentian violet residues GENTIAN VIOLET 5 Table 1. Disposition and excretion of multiple oral doses of 14 C-labelled gentian violet administered to mice Sample Gentian violet residues a ( g/g or g/mL) Males Females Liver 17.8 ± 2.6** 10.7 ± 3.4** Kidney 1.6 ± 0.1** 2.7 ± 0.8** Muscle 0.6 ± 0.4** 1.3 ± 0.7** Gonad 0.49 ± 0.08 3.66 ± 1.08 b Fat 14.3 ± 3.0** 24.1 ± 7.0** Urine 1.16 (5.9%) 1.58 (8.1%) Faeces 12.89 (65.9%) 13.17 (67.4%) **: P < 0.01 (student t-test for a signi cant sex difference) a Values are means ± 1 standard deviation for 12 male and 12 female mice. Numbers in parentheses indicate the percentage of the total dose. b Mean of eight mice. Source : McDonald et al. (1984a); McDonald (1989) were highest in adipose tissue (particularly in females), although a major portion (6667%) was excreted in faeces (McDonald et al., 1984a; McDonald, 1989). (b) Rats Groups of three male and three female F344 rats were housed individually in metabolism cages and given a single dose by gavage of 14 C-labelled gentian violet (94.8% gentian violet and 5.2% pentamethylpararosaniline). Gentian violet doses were 4.8 mg/kg bw (0.11 MBq/animal) and 5.2 mg/kg bw (0.34 MBq/animal) for males and females, respectively. Rats were killed 2, 4, 14 or 24 hours after dosing. Urine, faeces, liver, kidney, muscle, testes or ovaries and a fat sample were collected, and radioactivity was measured ( Table 2 ). Half-lives of 14.5 and 14.4 hours were calculated following a single dose for the liver and kidney, respectively, for males and 17.0 and 18.3 hours, respectively, for females (McDonald et al., 1984a). Groups of eight male and eight female F344 rats were housed individually in metabolism cages and given 14 doses of 14 C-labelled gentian violet (94.8% gentian violet and 5.2% pentamethylpararosaniline) at 12-hour intervals by gavage. The total gentian violet doses were 3.5 mg/kg bw (5.2 MBq/animal) and 5.69 mg/kg bw (2.9 MBq/animal) for males and females, respectively. The rats were killed 2 hours after receiving the final dose. Urine, faeces, liver, kidney, muscle, testes or ovaries and a fat sample were collected, and radioactivity was measured ( Table 3 ). As for mice, gentian violet residues concentrated in the adipose tissue of females; in males, the levels in liver and fat were similar. The percentages of administered gentian violet radioactivity excreted in the faeces of rats and mice were very similar, whereas considerably more was excreted in the urine of mice than in that of rats (McDonald et al., 1984a). Two female bile ductcannulated rats were administered a single dose of 14 C-labelled gentian violet (94.8% gentian violet and 5.2% pentamethylpara- rosaniline) by gavage at 300 g (0.12 MBq) or 840 g (0.34 MBq), and bile was 6 GENTIAN VIOLET Table 2. Deposition and excretion