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Flumazenil (Flumazepil, Anexate, Romazicon, Lanexat; Ro-15-1788; Ro-15 1788) is a specific and competitive GABAA receptor antagonist that has been used for the treatment of benzodiazepine overdoses. Flumazenil is a specific benzodiazepine receptor antagonist primarily administered by injection. It is the only benzodiazepine receptor antagonist on the market today. Flumazenil is of benefit in patients who become excessively drowsy after benzodiazepines are used for either diagnostic or therapeutic procedures.
Physicochemical Properties
| Molecular Formula | C15H14FN3O3 | |
| Molecular Weight | 303.29 | |
| Exact Mass | 303.101 | |
| CAS # | 78755-81-4 | |
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| PubChem CID | 3373 | |
| Appearance | White to off-white solid powder | |
| Density | 1.4±0.1 g/cm3 | |
| Boiling Point | 528.0±50.0 °C at 760 mmHg | |
| Melting Point | 201-203°C | |
| Flash Point | 273.1±30.1 °C | |
| Vapour Pressure | 0.0±1.4 mmHg at 25°C | |
| Index of Refraction | 1.634 | |
| LogP | 0.67 | |
| Hydrogen Bond Donor Count | 0 | |
| Hydrogen Bond Acceptor Count | 5 | |
| Rotatable Bond Count | 3 | |
| Heavy Atom Count | 22 | |
| Complexity | 461 | |
| Defined Atom Stereocenter Count | 0 | |
| InChi Key | OFBIFZUFASYYRE-UHFFFAOYSA-N | |
| InChi Code | InChI=1S/C15H14FN3O3/c1-3-22-15(21)13-12-7-18(2)14(20)10-6-9(16)4-5-11(10)19(12)8-17-13/h4-6,8H,3,7H2,1-2H3 | |
| Chemical Name | ethyl 8-fluoro-5-methyl-6-oxo-4H-imidazo[1,5-a][1,4]benzodiazepine-3-carboxylate | |
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| HS Tariff Code | 2934.99.9001 | |
| Storage |
Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
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| Shipping Condition | Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs) |
Biological Activity
| Targets |
Benzodiazepine (BZD) receptors [1] - Benzodiazepine (BZD) receptors (partial agonist-like binding in BALB/c mice, no affinity data specified) [2] - Benzodiazepine (BZD) receptors (mediating allopregnanolone's anxiolytic action blockade) [3] - Benzodiazepine (BZD) receptors and non-NMDA receptors (interactions with non-NMDA antagonists in seizure models) [4] - Benzodiazepine (BZD) receptors (blocking anxiety in ethanol-withdrawn rats) [5] |
| ln Vivo |
The behavioral, neurological, and electrophysiological effects of benzodiazepine agonists and inverse agonists are neutralized or reversed by flumazenil through its interaction with central benzodiazepine receptors. Hepatic encephalopathy can benefit from flumazenil in certain cases, however until well planned clinical trials can be carried out, hepatic encephalopathy must be treated as an exploratory indication. Although flumazenil can restore the drowsiness brought on by benzodiazepines either by alone or in conjunction with other medications, it shouldn't be taken if cyclic antidepressant toxicity is suspected [1]. In the raised plus maze and light/dark test, flumazenil (1 mg/kg) significantly reduces anxiety in BALB/c mice [2]. In rats, the decrease in allopregnanolone can be successfully stopped by flumazenil (10 mg/kg) [3]. In mice, the anticonvulsant and side effects of diazepam are countered by flumazenil (5–20 mg/kg), but not by GYKI 52466. The anticonvulsant activity of NBQX was somewhat diminished by flumazenil in the MES model, but not in the PTZ trial [4]. Reduced open-arm time and open-arm entry percentage are the outcomes of long-term ethanol treatment because flumazenil (3.0 mg/kg) inhibits withdrawal alterations [5]. Flumazenil reversed the sedative, hypnotic, anxiolytic, anticonvulsant, and muscle relaxant effects of benzodiazepines in various animal models and human studies [1] - Flumazenil induced partial agonist-like effects (reduced locomotor activity, increased punished responding in the Vogel conflict test) in BALB/c mice but not in C57BL/6 mice [2] - Flumazenil (10 mg/kg, i.p.) completely blocked the anxiolytic action of allopregnanolone (5 mg/kg, i.p.) in the elevated plus-maze test in mice [3] - Flumazenil (10 mg/kg, i.p.) antagonized the anticonvulsant effects of diazepam against pentylenetetrazol-induced seizures in mice, but did not affect the anticonvulsant actions of NBQX or GYKI 52466 [4] - Flumazenil (1-10 mg/kg, i.p.) dose-dependently blocked the anxiety-like behavior (increased entries into open arms of elevated plus-maze) in rats undergoing ethanol withdrawal [5] |
| Enzyme Assay |
Radioligand binding assays were performed to evaluate the affinity of Flumazenil for benzodiazepine receptors in brain membrane preparations. Membranes were incubated with tritiated benzodiazepine ligands in the presence or absence of Flumazenil, followed by filtration and scintillation counting to measure specific binding inhibition [1] - Receptor binding studies in mouse brain homogenates showed that Flumazenil displaced radiolabeled benzodiazepine ligands, with differential binding profiles between BALB/c and C57BL/6 mouse strains [2] |
| Animal Protocol |
Mice (BALB/c and C57BL/6 strains) were used to assess behavioral effects of Flumazenil. The drug was dissolved in a suitable vehicle and administered intraperitoneally at doses ranging from 0.1 to 10 mg/kg. Behavioral tests included locomotor activity monitoring and the Vogel conflict test, performed 30 minutes after drug administration [2] - Mice were treated with Flumazenil via intraperitoneal injection at a dose of 10 mg/kg, 15 minutes prior to allopregnanolone administration (5 mg/kg, i.p.). The elevated plus-maze test was conducted 30 minutes after allopregnanolone administration to evaluate anxiolytic activity [3] - Mice were pretreated with Flumazenil (10 mg/kg, i.p.) 30 minutes before administration of diazepam, NBQX, or GYKI 52466. Seizures were induced by pentylenetetrazol, and seizure severity and latency were recorded for 30 minutes post-induction [4] - Rats were rendered ethanol-dependent by chronic ethanol vapor exposure. After ethanol withdrawal, Flumazenil was administered intraperitoneally at doses of 1, 3, and 10 mg/kg. Anxiety-like behavior was evaluated using the elevated plus-maze test 30 minutes after drug administration [5] - Human subjects with benzodiazepine overdose or therapeutic benzodiazepine effects were administered Flumazenil via intravenous infusion at a rate of 0.2 mg/min (total dose up to 1 mg) to reverse benzodiazepine-induced sedation and respiratory depression [1] |
| ADME/Pharmacokinetics |
Absorption, Distribution and Excretion Flumazenil is completely (99%) metabolized. Elimination of radiolabeled drug is essentially complete within 72 hours, with 90% to 95% of the radioactivity appearing in urine and 5% to 10% in the feces. 0.9 to 1.1 L/kg 1 L/hr/kg [healthy volunteers receiving a 5-minute infusion of a total of 1 mg] Metabolism / Metabolites Hepatic. Flumazenil is completely (99%) metabolized. The major metabolites of flumazenil identified in urine are the de-ethylated free acid and its glucuronide conjugate. Biological Half-Life Initial distribution half-life is 4 to 11 minutes and the terminal half-life is 40 to 80 minutes. Prolongation of the half-life to 1.3 hours in patients with moderate hepatic impairment and 2.4 hours in severely impaired patients. Compared to adults, the elimination half-life in pediatric patients was more variable, averaging 40 minutes (range: 20 to 75 minutes). Flumazenil is rapidly absorbed after intravenous administration, with a volume of distribution of approximately 0.6-1.0 L/kg in humans. The elimination half-life is 0.7-1.3 hours [1] - Flumazenil is extensively metabolized in the liver via oxidation and conjugation reactions, with less than 1% of the parent drug excreted unchanged in urine [1] - Oral bioavailability of Flumazenil is low (approximately 16%) due to significant first-pass metabolism [1] |
| Toxicity/Toxicokinetics |
Effects During Pregnancy and Lactation ◉ Summary of Use during Lactation No information is available on the clinical use of flumazenil during breastfeeding. Because flumazenil is not orally absorbed it is unlikely to adversely affect the breastfed infant. If the mother requires flumazenil, it is not a reason to discontinue breastfeeding. The half-life of the drug is 54 minutes, so withholding breastfeeding for 4 to 5 hours after a dose should minimize transfer to the infant. ◉ Effects in Breastfed Infants Relevant published information was not found as of the revision date. ◉ Effects on Lactation and Breastmilk Relevant published information was not found as of the revision date. Protein Binding Protein binding is approximately 50%, mostly (66%) to albumin. Protein binding is reduced in patients with hepatic cirrhosis. Flumazenil has a low toxicity profile in humans. Adverse effects are mild and transient, including nausea, vomiting, anxiety, and seizures (rare, primarily in patients with epilepsy or benzodiazepine dependence) [1] - The plasma protein binding rate of Flumazenil is approximately 40-50% [1] |
| References |
[1]. Flumazenil: a benzodiazepine antagonist. Clin Pharm. 1993 Sep;12(9):641-56; quiz 699-701. [2]. Flumazenil induces benzodiazepine partial agonist-like effects in BALB/c but not C57BL/6 mice. Psychopharmacology (Berl). 2000 Jan;148(1):24-32. [3]. Flumazenil blocks the anxiolytic action of allopregnanolone. Eur J Pharmacol. 1995 Jul 25;281(1):113-5. [4]. Effects of the non-NMDA antagonists NBQX and the 2,3-benzodiazepine GYKI 52466 on different seizure types in mice: comparison with diazepam and interactions with flumazenil. Br J Pharmacol. 1994 Dec;113(4):1349-57. [5]. Flumazenil blockade of anxiety following ethanol withdrawal in rats. Psychopharmacology (Berl). 1997 Jun;131(4):354-60. |
| Additional Infomation |
Flumazenil is an organic heterotricyclic compound that is 5,6-dihydro-4H-imidazo[1,5-a][1,4]benzodiazepine which is substituted at positions 3, 5, 6, and 8 by ethoxycarbonyl, methyl, oxo, and fluoro groups, respectively. It is used as an antidote to benzodiazepine overdose. It has a role as a GABA antagonist and an antidote to benzodiazepine poisoning. It is an ethyl ester, an organofluorine compound and an imidazobenzodiazepine. Fumazenil is an imidazobenzodiazepine derivative and a potent benzodiazepine receptor antagonist that competitively inhibits the activity at the benzodiazepine recognition site on the GABA/benzodiazepine receptor complex, thereby reversing the effects of benzodiazepine on the central nervous system. Flumazenil is a Benzodiazepine Antagonist. Flumazenil is an imidazo-benzodiazepine derivative, effective in reversing benzodiazepine-induced activities. Flumazenil antagonizes the benzodiazepine binding site of the gamma-aminobutyric acid (GABA)/benzodiazepine receptor complex in the central nervous system (CNS), thereby preventing the chloride channel opening events and inhibiting neuronal hyperpolarization. As a result, flumazenil reverses benzodiazepine-induced effects including sedation, psychomotor deficits, amnesia, and hypoventilation in a dose-dependent manner. A potent benzodiazepine receptor antagonist. Since it reverses the sedative and other actions of benzodiazepines, it has been suggested as an antidote to benzodiazepine overdoses. Drug Indication For the complete or partial reversal of the sedative effects of benzodiazepines in cases where general anesthesia has been induced and/or maintained with benzodiazepines, and where sedation has been produced with benzodiazepines for diagnostic and therapeutic procedures. Also for the management of benzodiazepine overdose as an adjunct for appropriate supportive and symptomatic measures. FDA Label Mechanism of Action Flumazenil, an imidazobenzodiazepine derivative, is a benzodiazepine antagonist. It competitively inhibits the benzodiazepine binding site on the GABA/benzodiazepine receptor complex. Flumazenil is a weak partial agonist in some animal models of activity, but has little or no agonist activity in man. Pharmacodynamics Flumazenil antagonizes the CNS effects produced by benzodiazepines, but does not antagonize the central nervous system effects of drugs affecting GABA-ergic neurons by means other than the benzodiazepine receptor (including ethanol, barbiturates, or general anesthetics) and does not reverse the effects of opioids. Flumazenil (Ro 15-1788) is a selective competitive antagonist of benzodiazepine receptors, used clinically to reverse the effects of benzodiazepine overdose and to terminate conscious sedation [1] - The partial agonist-like effects of Flumazenil in BALB/c mice suggest strain-dependent differences in benzodiazepine receptor signaling or regulation [2] - Flumazenil blocks the anxiolytic action of allopregnanolone, indicating that allopregnanolone exerts its anxiolytic effects via benzodiazepine receptor modulation [3] - Flumazenil does not interact with non-NMDA glutamate receptors, as it failed to affect the anticonvulsant activity of NBQX or GYKI 52466 [4] - The anxiolytic-blocking effect of Flumazenil in ethanol-withdrawn rats suggests that benzodiazepine receptor activation contributes to ethanol withdrawal-induced anxiety [5] |
Solubility Data
| Solubility (In Vitro) |
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2 mg/mL (6.59 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% 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 20.0 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. Solubility in Formulation 2: ≥ 2 mg/mL (6.59 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 20.0 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. Solubility in Formulation 3: ≥ 2 mg/mL (6.59 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 20.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.2972 mL | 16.4859 mL | 32.9717 mL | |
| 5 mM | 0.6594 mL | 3.2972 mL | 6.5943 mL | |
| 10 mM | 0.3297 mL | 1.6486 mL | 3.2972 mL |