The fact that butylated hydroxytoluene (BHT) effectively promotes tumor-induced tumors is widely recognized. In tumors that are 7 weeks old, butylated hydroxytoluene (facial; 400 mg/kg; weekly) formulation boosts the efficacy of rasH2 tumorigenesis. Sensibility [3]

Absorption, Distribution and Excretion
The biliary metabolism of BHT, BHT-COOH, BHT-OH, and BHT-aldehyde was compared after i.p. or intravenous administration to male Wistar rats. For all four test compounds, the major metabolites in enterohepatic circulation were BHT-COOH and its ester glucuronide. ... Total biliary excretion after treatment with BHT or BHT-aldehyde was less after i.v. dosing than after i.p. dosing. ...
Rats and humans received single doses of BHT. Male Wistar rats (n=2-10) were treated with 20 to 200 mg/kg BHT. Human subjects were treated with 0.5 mg/kg BHT (seven non-smoking males). In rats, kinetic parameters increased dose-dependently. Rats excreted ~10% of the high dose as unchanged BHT in the feces, mostly on day 1. Urinary excretion of BHT-COOH was little more than 1%, in decreasing amounts, on days 1 to 4. In humans, the mean plasma concentration-time profile was decreased as compared to that of rats. Unchanged BHT was not detected in the feces, and urinary excretion of BHT-COOH was 0 to 5.5%.
After a single dose of [(14)C]BHT /to human males/ at least two-thirds of the radioactivity is excreted in urine, with the major portion appearing within 24 hr of dosing. As occasional assays demonstrated a fecal excretion of about one-half of that in urine, it is probable that the remainder of the radioactivity was excreted by this route. The bulk of the radioactivity appears on the first day after dosing, and thereafter a progressively diminishing slight excretion continues for a considerable period. ...
Solutions of BHT (10 mg/L) were prepared in polyethylene glycol 400-normal saline (1:1). The solutions were infused in rabbits for <2 minutes at a constant rate of 2 mL/min to provide a total dose of 10 mg/kg. Blood samples were collected at 0, 0.085, 0.173, 0.33, 0.50, 0.75, 1, 1.15, 2.0, 3.0, 4.0, 6.0 and 12 hours and thereafter twice per day for 2 days and daily for 3 days. Blood samples were analyzed for their concentration of BHT using a highly sensitive and specific GLC method. The fast disposition phase half-life of BHT was approximately 1 hour and the slow disposition phase decayed with a half-life of >11 days. These data suggest rapid accumulation and slow clearance from the body. BHT tends to be stored in body tissues upon multiple dosing and more than a 16-fold accumulation of BHT is possible on daily exposure.
For more Absorption, Distribution and Excretion (Complete) data for 2,6-DI-T-BUTYL-P-CRESOL (7 total), please visit the HSDB record page.

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Metabolism / Metabolites
The metabolism of BHT has been investigated extensively in rabbits, rat, mice and man. The principle routes of metabolism of BHT in all species involve oxidation of the para-methyl and of one, or both, of the tert-butyl substituents. Neither mechanism is mutually exclusive. Oxidation of the methyl-group is catalyzed by the microsomal enzyme, BHT-oxidase and several derivatives including the quinone-methide, 2,6-di-tert-butyl-4-methylene-2,5-cyclohexadienone and 4-hydroxy-4-methyl-2,6-di-tert-butyl-cyclahexe-2,5-dienone have been identified in rat liver. Whereas oxidation of the para-methyl substituent is the major route of metabolism in the rat and rabbit, where BHT-acid accounts for approximately 30% of the dose, some 30-40% of the dose in male and female mice and in man is excreted as metabolites involving oxidation of one or both of the tert-butyl groups. BHT is excreted principally in the urine in man whereas in rodents 50-80% is eliminated in the feces. This is presumed to be due to species differences in the molecular weight threshold for biliary excretion.
The biliary metabolism of BHT, BHT-COOH, BHT-OH, and BHT-aldehyde was compared after i.p. or intravenous administration to male Wistar rats. For all four test compounds, the major metabolites in enterohepatic circulation were BHT-COOH and its ester glucuronide. ... Total biliary excretion after treatment with BHT or BHT-aldehyde was less after i.v. dosing than after i.p. dosing. ...
A comparative metabolism study of BHT was conducted in mice and rats. In male and female DDY/Slc mice given single oral doses (20 or 500 mg/kg body weight) of BHT labelled with (14)C at the p-methyl group, (14)C was distributed mainly in the stomach, intestines, liver and kidney, and then excreted in the urine, feces and expired air. During the 7 days after treatment, 41-65, 26-50 and 69% of the (14)C dose was excreted in feces, urine and expired air, respectively, and the total recovery was 96-98%. Levels of (14)C in 21 male and 22 female tissues 7 days after treatment were less than 1 ug BHT equivalents/g tissue (ppm) in mice given 20 mg/kg and less than 11 ppm in mice given 500 mg/kg. When [(14)C]BHT was given orally to male mice at 20 mg/kg/day for 10 days, (14)C was rapidly excreted and did not exhibit any tendency to accumulate in any tissues. Thin-layer chromatography and high-performance liquid chromatography analyses showed that more than 43 metabolites were present in the urine and feces of both species, and all of these were identified to determine metabolic pathways for BHT in mice and rats. Major metabolic reactions of [(14)C]BHT in mice were the oxidation of the p-methyl group attached to the benzene nng and of the tert-butyl groups. The products from the latter reaction were cyclized to some extent by reacting with the adjacent phenolic OH group to give hemiacetals or lactones. The carboxyl derivatives from the p-methyl oxidation were conjugated with glucuronic acid. When single oral doses of 20 or 500 mg [(14)C]BHT/kg were given to male Sprague-Dawley rats, metabolites similar to those in mice were found. However, the major biotransformation was oxidation of the p-methyl group, and oxidation of the tert-butyl groups was a minor reaction in rats.
The principal metabolites of 2,6-di-tert-butyl-p-cresol (BC) in mouse bronchiolar Clara cells were 6-tert-butyl-2-(hydroxy-tert-butyl)p-cresol (BC-butOH; 4.4 +/- 1.1 pmol/10-6 cells/minute) and 2,6-di-tert-butyl-p-hydroxymethyl-phenol (BC-OH; 1.0 +/- 0.2 pmol/10-6 cells/minute). This metabolite pattern is nearly identical with that obtained with microsomes prepared from whole lungs. Quinone methide production occurred more readily from BC-butOH than from BC (0.52 +/- 0.14 compared to 0.41 +/- 0.06 pmol/10-6 cells/minute). The maximum concentration of the intermediate BC-butOH was very low relative to that of BC; similar quantities of the quinone methides were generated. Furthermore, two glutathion conjugates, expected from attack of BC-quinone methide and BC-butOH-quinone methide, were found. Incubation time was 15 minutes (Clara cells) or 10 minutes (microsomes).
Metabolite of BHT isolated from liver of orally treated male rats. Identified as 2,6-di-tert-butyl-4-methylene-2,5-cyclohexadienone.
Oxidative metabolism (phase 1 reactions) mediated by the microsomal monooxygenase system is the major route for BHT degradation. Oxidation of the tert-butyl groups is most common in man. Gallates and 2-tert-butylhydroquinone are mainly metabolized by non-oxidative pathways (methylation or conjugation with sulphate and glucuronic acid). (A15352). In particular BHT is frequently metabolized to quinone methides (QMs) which are thought to be responsible for promoting tumor formation. One example of a QM is 2,6-di-tert-butyl-4-methylenecyclohexa-2,5-dienone (BHT-QM). QMs are strongly electrophilic and readily form adducts with proteins.

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Biological Half-Life
Solutions of BHT (10 mg/L) were prepared in polyethylene glycol 400-normal saline (1:1). The solutions were infused in rabbits for <2 minutes at a constant rate of 2 mL/min to provide a total dose of 10 mg/kg. ... The fast disposition phase half-life of BHT was approximately 1 hour and the slow disposition phase decayed with a half-life of >11 days. ...

Toxicity Summary
IDENTIFICATION AND USE: Butylated hydroxytoluene( BHT) is a white, crystalline, odorless solid. It is used as an antioxidant for fats and oils or in packaging material for fat containing foods. HUMAN EXPOSURE AND TOXICITY: Potential symptoms of overexposure are irritation of eyes and skin. ANIMAL STUDIES: Rats fed high doses of BHT, showed increases in serum cholesterol in both sexes. Groups of weanling rats fed BHT in conjunction with lard supplementation had a reduction in growth rate, especially in males. BHT also increased absolute liver weight and the ratio of liver weight to body weight in both sexes. BHT increased the ratio of left adrenal weight to body weight in male rats but had no consistent effect in female rats. BHT administered to rats for 68-82 days caused reduction in rate of increase in weight and fatty infiltration of the liver. BHT was given in feed of rats and mice of both sex at 3000 or 6000 ppm; in rats 105 wk and 107 or 108 wk in mice. No tumors occurred in either sex of rats and mice. When tested for teratogenic properties BHT produced anophthalmia in offspring in rats, but not in mice. BHT administered to pregnant mice for 18 days along with another group fed BHT for 50 to 64 days including 18 das of pregnancy. No fetal abnormalities were observed. In a study using 144 mice, no blindness was observed in any of the 1162 litters representing 7765 offspring born throughout the reproductive life span of the mothers. BHT was tested for mutagenicity in the Salmonella/microsome preincubation assay in 5 Salmonella typhimurium strains (TA1535, TA1537, TA97, TA98, and TA100) in the presence and absence of metabolic activation. BHT was negative in these tests and the highest ineffective dose tested in any Salmonella typhimurium strain was 10 mg/plate. ECOTOXICITY STUDIES: In salmon fed graded levels of BHT during a 12-week feeding followed by a 2-week depuration period, BHT selectively modulated toxicological responses in the xenobiotic biotransformation pathways during the feeding period.
BHT is metabolized to quinone methides (QMs) which are responsible for promoting tumor formation in many animal models. One example of a QM is 2,6-di-tert-butyl-4-methylenecyclohexa-2,5-dienone (BHT-QM). QMs are strongly electrophilic and readily form adducts with proteins. Some of the QM targets include redox proteins such as glutathione S-transferase P1 (GST-P1), peroxiredoxin 6 (Prx6), Cu,Zn-superoxide dismutase (SOD1), carbonyl reductase, and selenium-binding protein 1, which have direct or indirect antioxidant functions. (A15087, A15355). The modification of these proteins leads to decreased cellular protection from electrophiles and oxidants. Alkylation also may interfere with GSTP1 regulation of stress kinases, thereby influencing phosphorylation and cell growth. BHT also binds to the retinoic acid receptor which can lead to changes in cell development.

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Interactions
During the DPPH scavenging assay carried out in non polar and non protic solvents, such as toluene, BHT regenerates alpha-tocopherol from tocopheryl radical, whereas in polar and protic solvents, like methanol, no regeneration is observed due to a fast electron transfer reaction from the tocopheryl radical to the reactive DPPH radical. ... In the presence of a small amount of alcohol, the synergy is exalted and BHT regenerates twice as much alpha-tocopherol due to a nucleophilic addition of short alcohols on the BHT oxidation product, giving a new phenolic co-antioxidant.
The natural retinoid, retinyl acetate (RA), and the phenolic antioxidant, butylated hydroxytoluene (BHT), are both effective inhibitors of mammary carcinogenesis in rats. The present study was designed to determine if an increased inhibition of mammary carcinogenesis is obtained when RA and BHT are administered in combination. At age 50 days (time 0), virgin, female Sprague-Dawley rats received a single intragastric instillation of 16 mg of 7,12-dimethylbenz(a)anthracene dissolved in 1 mL sesame oil. Groups of 30 carcinogen-treated rats received Wayne Lab Chow supplemented with (per kg diet) 250 mg RA, 5000 mg BHT, or 250 mg RA plus 5000 mg BHT by the following schedule: -2 to +1 week; +1 week until the end of the experiment; -2 weeks to end; or none. Combined administration of RA plus BHT by the -2 weeks to end schedule was more effective in mammary cancer chemoprevention than was RA alone or BHT alone; the interaction of RA and BHT was additive. Similarly, administration of RA plus BHT by the -2 weeks to end protocol was more active in chemoprevention than was RA plus BHT administered either from weeks -2 to +1 or +1 week to end. Chronic exposure to RA plus BHT induced a high incidence of hepatic fibrosis and bile duct hyperplasia; these changes were not observed in controls and were seen in low incidence in animals exposed to RA only or BHT only. These data indicate that enhanced anticarcinogenic activity can be obtained through the use of "combination chemoprevention" regimens; however, chemopreventive compounds may interact not only to inhibit carcinogenesis but also to induce toxicity.
The individual and combined (binary mixtures) (anti)androgenic effect of butylparaben (BuPB), butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and propyl gallate (PG) was evaluated using the MDA-kb2 cell line. Exposing these cells to AR agonists results in the expression of the reporter gene (encoding for luciferase) and luminescence can be measured in order to monitor the activity of the reporter protein. In case of the evaluation of the anti-androgenic effect, the individual test compounds or binary mixtures were tested in the presence of a fixed concentration of a strong AR agonist (1000 pM 5-alpha-dihydrotestosterone; DHT). Cell viability was assessed using a resazurin based assay. For PG, this is the first report in the literature concerning its (anti)androgenic activity. In case of both individual and mixture testing none of the compounds or binary combinations showed androgenic activity. When tested in the presence of DHT, BuPB, BHA and BHT proved to be weak anti-androgens and this was confirmed during the evaluation of binary mixtures (BuPB+BHA, BuPB+BHT and BHA+BHT). Besides performing the in vitro testing of the binary combinations, two mathematical models (dose addition and response addition) were evaluated in terms of accuracy of prediction of the anti-androgenic effect of the selected binary mixtures. The dose addition model guaranteed a good correlation between the experimental and predicted data. However, no estimation was possible in case of mixtures containing PG, due to the lack of effect of the compound in case of the individual testing.
The aim of this study was to compare the effects of L-arginine (L-arg) and food-antioxidant butylated hydroxytoluene (BHT) against oxidative stress of Escherichia coli endotoxin (LPS) in the liver. Ninety Wistar albino rats were assigned in three groups. Rats received one of the following pre-treatment previous to 5 mg/kg LPS intraperitoneally: saline, L-arg (NO donor, 100 mg/kg), or BHT (250 mg/kg/day), for 3 days. At second, fourth and sixth hours, plasma nitrite-plus-nitrate, circulating liver enzymes, glutathione levels, superoxide dismutase, glutathione peroxidase activities were measured. The most remarkable liver injury was evident in BHT pre-treated animals at all time points compared to L-arg pre-treated rats. While BHT enhanced superoxide dismutase activities following LPS, glutathione decreased simultaneously compared to L-arg group. ...
For more Interactions (Complete) data for 2,6-DI-T-BUTYL-P-CRESOL (15 total), please visit the HSDB record page.

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Non-Human Toxicity Values
LD50 Rat oral 890 mg/kg
LD50 Mouse oral 650 mg/kg
LD50 Mouse ip 138 mg/kg
LD50 Mouse iv 180 mg/kg
For more Non-Human Toxicity Values (Complete) data for 2,6-DI-T-BUTYL-P-CRESOL (11 total), please visit the HSDB record page.

[1]. Butylated hydroxytoluene (BHT): a review. Environ Res. 1982 Oct;29(1):1-29.

[2]. Ferroptosis: A Regulated Cell Death Nexus Linking Metabolism, Redox Biology, and Disease. ell. 2017 Oct 5;171(2):273-285.

[3]. Butylhydroxytoluene (BHT) increases susceptibility of transgenic rasH2 mice to lung carcinogenesis. J Cancer Res Clin Oncol. 2001 Oct;127(10):583-90.

Therapeutic Uses
/EXPL THER/ /The objective was to/ evaluate the efficacy of potential therapeutics in Rdh8(-/-)Abca4(-/-) mice, a rodent model of human age-related macular degeneration (AMD). Therapeutic efficacy of several antioxidant agents (ascorbic acid, alpha-lipoic acid, alpha-tocopherol, Mn(III)-tetrakis(4-benzoic acid)-porphyrin, and butylated hydroxytoluene), an immunosuppressive agent with antivascular endothelial growth factor (VEGF) activity (sirolimus, also known as rapamycin), a retinoid cycle inhibitor (retinylamine), and an artificial chromophore (9-cis-retinyl acetate) were evaluated side by side in a recently described murine model of AMD, the Rdh8(-/-)Abca4(-/-) mouse. This animal exhibits a retinopathy caused by delayed all-trans-retinal clearance resulting from the absence of both ATP-binding cassette transporter 4 (Abca4) and retinol dehydrogenase 8 (Rdh8) activities. Drug efficacy was evaluated by retinal histologic analyses and electroretinograms (ERGs). All tested agents partially prevented atrophic changes in the Rdh8(-/-)Abca4(-/-) retina with retinylamine demonstrating the greatest efficacy. A significant reduction of complement deposition on Bruch's membrane was observed in sirolimus-treated mice, although the severity of retinal degeneration was similar to that observed in antioxidant- and 9-cis-retinyl acetate-treated mice. Sirolimus treatment of 6-month-old Rdh8(-/-)Abca4(-/-) mice for 4 months prevented choroidal neovascularization without changing retinal VEGF levels. Mechanism-based therapy with retinylamine markedly attenuated degenerative retinopathy in Rdh8(-/-)Abca4(-/-) mice. ...
/EXPL THER/ The present study was undertaken to evaluate the possible ameliorating effect of butylated hydroxyl toluene (BHT), associated with ferric nitrilotriacetate (Fe-NTA)-induced oxidative stress and liver injury in mice. The treatment of mice with Fe-NTA alone enhances ornithine decarboxylase activity to 4.6 folds, protein carbonyl formation increased up to 2.9 folds and DNA synthesis expressed in terms of [(3)H] thymidine incorporation increased to 3.2 folds, and antioxidants and antioxidant enzymes decreased to 1.8-2.5 folds, compared with the corresponding saline-treated controls. These changes were reversed significantly (p < 0.001) in animals receiving a pretreatment of BHT. Our data show that BHT can reciprocate the toxic effects of Fe-NTA and can serve as a potent chemopreventive agent.

C15H24O

220.36

220.182

128-37-0

Butylated hydroxytoluene-d21;64502-99-4;Butylated hydroxytoluene-d24;1219805-92-1;Butylated hydroxytoluene-d3;86819-59-2

31404

White to off-white solid powder

1.048

265 ºC

69-71 ºC

127 ºC

0.0±0.6 mmHg at 25°C

1.499

5.32

1

1

2

16

207

0

NLZUEZXRPGMBCV-UHFFFAOYSA-N

InChI=1S/C15H24O/c1-10-8-11(14(2,3)4)13(16)12(9-10)15(5,6)7/h8-9,16H,1-7H3

Phenol, 2,6-bis(1,1-dimethylethyl)-4-methyl-

Butylated hydroxytoluene NSC-6347 NSC6347NSC 6347

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~100 mg/mL (~453.82 mM)
H2O : ~1 mg/mL (~4.54 mM)

Solubility in Formulation 1: 2.5 mg/mL (11.35 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (11.35 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (11.35 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 100 mg/mL (453.82 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.

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 (Please use freshly prepared in vivo formulations for optimal results.)