Anesthetics

Tribromoethanol is a potent anesthetic agent used before surgery in animals, in particular, rodents.

Prior to surgery, experimental animals—particularly rodents—are put to sleep with tribromoethanol.
Mice (n = 68) were randomly assigned to 1 of 7 groups to receive tribromoethanol (500 mg/kg IP) on day 0 or days 0 and 8; vehicle (tert-amyl alcohol in sterile water) only on day 0 or days 0 and 8; sterile water injection on day 0 or days 0 and 8; or no treatment. A single dose of tribromoethanol failed to produce loss of pedal reflex and had no effect on median food and water consumption but altered median body weight on days 1 through 4 when compared with that in mice that received vehicle only or no treatment. Median body weight did not differ between mice that received a single dose of tribromoethanol and those that received an injection of water. Among mice given 2 doses of tribromoethanol, induction time, anesthetic duration, and recovery time varied widely. Repeated administration of tribromoethanol had no effect on median food and water consumption or body weight compared with those in controls. Median liver weight was significantly greater in mice that received 2 doses compared with a single dose of tribromoethanol. Median liver weight did not differ between untreated mice and those that received tribromoethanol. No significant organ or tissue pathology was observed in any study animal. Although tribromoethanol did not produce morbidity, mortality, or pathologic changes in treated animals, we urge caution in use of tribromoethanol in C57BL/6NHsd mice due to its variable anesthetic effectiveness [1].

Group allocation.[1]
Mice were acclimated for 5 d prior to experimental use. After acclimation, mice were randomly assigned to 1 of 7 groups to receive Tribromoethanol (500 mg/kg IP) on day 0 or days 0 and 8; vehicle (tert-amyl alcohol in sterile water) intraperitoneally on day 0 or days 0 and 8; sterile water intraperitoneally on day 0 or days 0 and 8; or no treatment. All groups contained 10 mice each, except for the no-treatment group (n = 8).

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Tribromoethanol.[1]
The Tribromoethanol dose (500 mg/kg IP) used in this study followed the preparation and dosing recommendations outlined by our IACUC.19 Briefly, a 1.61-g/mL stock solution was prepared by adding 6.2 mL tert-amyl alcohol to 10 g 2,2,2-tribromoethanol. A 25-mg/mL working solution was prepared by adding 0.78 mL of the Tribromoethanol stock solution to 49.2 mL tissue-grade double-distilled H20; the working solution was filtered through a 0.2-mm syringe filter prior to injection in mice. The stock solution was made 1 d prior to injection and allowed to stir overnight at room temperature; the working solution was made immediately prior to injection. The pH of the working solution was not determined. Tribromoethanol powder from the same bottle and lot number was used throughout this study. The vehicle-only solution contained the same ratio of tert-amyl alcohol and sterile water as that in the Tribromoethanol working solution.

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Tribromoethanol efficacy and safety.[1]
Intraperitoneal injections were performed according to group allocation and current body weight and by a single investigator (JT). All mice (except the untreated group) received injection volumes of 0.31 to 0.44 mL. After Tribromoethanol injection, mice were maintained on a circulating-water heating pad and monitored for anesthetic induction time, duration of anesthesia, and recovery time. Induction time was defined as the time from Tribromoethanol administration to loss of the pedal reflex. Anesthetic duration was defined as the time between the loss and return of pedal reflex. Recovery time was defined as the time between return of the pedal reflex to movement around the primary enclosure. According to a previously described procedure,8 we assessed the pedal reflex by using a Touch-Test Sensory Evaluator (North Coast Medical, Gilroy, CA) with a target force of 300 g (Figure 1); a single investigator (CC) performed all assessments. Presence of the pedal reflex was defined as withdrawal of the limb on contact by the sensory evaluator. Body weight and food and water intake were measured daily by using a digital laboratory scale. Daily intake was quantified by determining the remaining mass of offered food and water.

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Pathologic evaluation.[1]
Mice that received injections only on day 0 were euthanized on day 4; those injected on days 0 and 8 were euthanized on day 11. Untreated mice were euthanized on day 4 (n = 3) or 11 (n = 5). All mice were euthanized via cervical dislocation under CO2 anesthesia. After euthanasia, each mouse was necropsied, and gastrointestinal tract, spleen, and liver were harvested and immediately weighed. The stomach, small intestine, cecum, colon, liver, spleen, and a sample of the body wall were collected for histopathologic evaluation. Tissues were fixed in 10% formalin, stained with hematoxylin and eosin, and evaluated by a single veterinary pathologist (KN) blinded to treatment group. By using a previously published algorithm for evaluation of Tribromoethanol-induced changes,8 histopathologic lesions were scored on a scale of 0 to 4 reflecting the severity of inflammation and percentage of organ affected.

rat LDLo oral 1 gm/kg Journal of Pharmacology and Experimental Therapeutics., 63(183), 1938
rat LDLo intraperitoneal 300 mg/kg GASTROINTESTINAL: OTHER CHANGES; LIVER: HEPATITIS, FIBROUS (CIRRHOSIS, POST-NECROTIC SCARRING) Laboratory Animal Science., 49(665), 1999 [PMID:10638506]
rat LDLo subcutaneous 530 mg/kg Naunyn-Schmiedeberg's Archiv fuer Experimentelle Pathologie und Pharmakologie., 182(348), 1936
mouse LD50 oral 930 mg/kg BEHAVIORAL: CHANGES IN MOTOR ACTIVITY (SPECIFIC ASSAY) Journal of Pharmaceutical Sciences., 56(920), 1967 [PMID:6034844]
rabbit LDLo oral 1100 mg/kg BEHAVIORAL: GENERAL ANESTHETIC; LUNGS, THORAX, OR RESPIRATION: OTHER CHANGES Naunyn-Schmiedeberg's Archiv fuer Experimentelle Pathologie und Pharmakologie., 132(214), 1928

[1]. Repeated administration of tribromoethanol in C57BL/6NHsd mice. J Am Assoc Lab Anim Sci. 2013 Mar;52(2):176-179.

Tribromoethanol is an organobromine compound and an alcohol.
Tribromoethanol, a nonpharmaceutical-grade anesthetic, has been used extensively for various manipulations in laboratory rodents due to its ready availability, lack of state and federal drug regulations associated with its use, and rapid anesthetic induction and recovery times. Tribromoethanol is commercially available as a white crystalline powder that, when reconstituted and administered, produces generalized CNS depression, including depression of respiratory and cardiovascular centers.11 Despite routine use, tribromoethanol use in rodents remains controversial due to contradictory reports regarding the compound's efficacy and associated pathology and mortality. Morbidities reported with tribromoethanol use in mice include intestinal ileus, peritonitis, muscle necrosis, serositis of abdominal organs, and death. n an attempt to balance animal welfare concerns and investigator needs, many IACUC have developed guidelines for tribromoethanol use, including prohibiting its repeated use in individual animals. A single study published in 1979 described high mortality after repeated tribromoethanol injection, and although no experimental details were provided, the report likely has influenced institutional policies.
In the current study, we determined the effect of repeated tribromoethanol administration on induction time, anesthetic duration, recovery time, food and water consumption, body weight, morbidity, mortality, and pathology in C57BL/6NHsd mice. To our knowledge, this study is the first to thoroughly evaluate the safety and efficacy of repeated tribromoethanol administration.[1]

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The tribromoethanol preparation used in this study appeared to be safe for use in C57BL/6NHsd mice, given that no morbidity, mortality, or pathologic changes were observed at the administered dose and frequencies. In a previous study, 10 of 47 female ICR mice were found dead or moribund after the administration of 400 mg/kg of freshly prepared tribromoethanol.8 The lethal dose of tribromoethanol may vary by mouse strain. Moreover, tribromoethanol purity may vary by supplier, lot number, and bottle and thus affect lethality. We noted no organ or tissue pathology in any study animal. Prior reports have described abdominal muscle necrosis, fluid distension of the stomach and small intestine, peritonitis, splenic serositis, and fibrinous visceral adhesions and tribromoethanol administration in ICR, CD-1, OF-1, NMRI, and (NCR) nu/nu mice.2,8,20 The tribromoethanol concentration and dose administered in these cited studies ranged from 12 to 25 mg/mL and 240 to 500 mg/kg, respectively, and included stored and freshly prepared preparations. Endpoints in the cited studies ranged from 24 h to 6 wk after tribromoethanol administration. In our current study, mice were euthanized 3 and 4 d after tribromoethanol administration. As a result, histopathologic changes associated with an acute inflammatory response would not have been observed. The absence of morbidity and mortality in the C57BL/6NHsd mice used in our study suggests that tribromoethanol toxicity may be strain-related. On the basis of our current findings, tribromoethanol appears to be safe for repeated administration in C57BL/6NHsd mice. However we urge caution regarding the use of this anesthetic in this strain due to variable effectiveness.[1]

C2H3BR3O

282.76

279.773

C, 8.50; H, 1.07; Br, 84.78; O, 5.66

75-80-9

6400

White to yellow solid

2.9±0.1 g/cm3

199.0±0.0 °C at 760 mmHg

73-79 °C(lit.)

99.7±25.9 °C

0.1±0.8 mmHg at 25°C

1.651

2.3

1

1

0

6

38.5

0

BrC(Br)(Br)CO

YFDSDPIBEUFTMI-UHFFFAOYSA-N

InChI=1S/C2H3Br3O/c3-2(4,5)1-6/h6H,1H2

2,2,2-Tribromoethanol

NSC-2189; NSC 2189; 2,2,2-Tribromoethanol; 75-80-9; Tribromoethyl alcohol; Avertin; Bromethol; Ethobrome; Tribromethanol; Narcolan; Tribromoethanol

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

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

DMSO : ≥ 125 mg/mL (~442.07 mM)
Ethanol :≥ 100 mg/mL (~353.66 mM)
H2O : ≥ 25 mg/mL (~88.41 mM)

Solubility in Formulation 1: 40 mg/mL (141.46 mM) in 5% Ethanol 95% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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