| Size | Price | Stock | Qty |
|---|---|---|---|
| 1mg |
|
||
| 5mg |
|
||
| 10mg |
|
||
| 25mg |
|
||
| 50mg | |||
| Other Sizes |
Purity: ≥98%
| Targets |
gamma-aminobutyric acid A receptor/GABAA
GABAA receptor benzodiazepine site. Bretazenil acts as a partial agonist at the gamma-aminobutyric acid A (GABAA) receptor-linked benzodiazepine site. In vitro, Bretazenil shows an EC₅₀ of 10 nM at recombinant α1β1γ2 receptors. It exhibits an IC₅₀ of 6.1 nM for inhibition of specific [³⁵S]TBPS binding. |
|---|---|
| ln Vitro |
Ro 16-6028 (bretazenil) has a pharmacological profile characteristic of a partial agonist at the gamma-aminobutyric acidA (GABAA) receptor-linked benzodiazepine site. The present study utilized modulation of [35S]t-butylbicyclophosphorothionate ([35S]TBPS) binding and enhancement of GABA-stimulated 36Cl- uptake to further assess Ro 16-6028's partial agonist profile in vitro. Ro 16-6028 was the most potent benzodiazepine examined, exhibiting an IC50 (concentration at which half-maximal inhibition of specific [35S]TBPS binding occurs) of 6.1 nM, compared to clonazepam (7.9 nM), flunitrazepam (13.6 nM) and diazepam (91.1 nM). The rank order of potency for inhibition of [35S]TBPS binding was identical to that for inhibition of [3H]flunitrazepam binding. However, Ro 16-6028 was less efficacious in that it produced 27% inhibition of specific [35S]TBPS binding, compared to clonazepam (34%), flunitrazepam (41%) or diazepam (49%). Ro 16-6028 antagonized the inhibition of [35S]TBPS binding produced by 10 microM diazepam. Ro 16-6028 was also more potent and less efficacious than diazepam in potentiating GABA-stimulated 36Cl- uptake. These results provide further evidence that Ro 16-6028 is acting as a partial agonist at the benzodiazepine receptor in modulating function of the GABAA receptor complex.[2]
Bretazenil is a potent GABAA benzodiazepine site partial agonist with an EC₅₀ of 10 nM at α1β1γ2 receptors. It inhibits [³⁵S]TBPS binding with an IC₅₀ of 6.1 nM. The compound shows selectivity for the benzodiazepine site and modulates GABA-induced chloride currents in a partial agonist manner. |
| ln Vivo |
Both minor (mMS, primarily clonic seizures, preserved righting ability) and major (MMS, generalized tonic-clonic) tetrazole-induced seizures are inhibited by bretazenil (0.001-0.1 mg/kg, injected intraperitoneally 10 minutes prior to tetrazole; male and female albino rats aged 7, 12, 18, 25, and 90 days) in a dose-dependent manner [3].
Alzet minipumps were loaded with bretazenil, a GABAA receptor partial agonist, and implanted subcutaneously to deliver 750 ng/hr for 14 days. After 3 baseline days with the bretazenil-eluting minipump, the AgrpDTR/+ mice were treated with DT (diptheria toxin). Food consumption by the bretazenil-treated group fell for the first 6 days after DT treatment, but then also returned to normal over the next 12 days. Bretazenil (0.2 mg/kg, ip) increased food consumption by wild-type mice during the first 4 hr, but 24-hr food intake was normal. Chronic delivery of bretazenil to wild-type animals for 10 days via a minipump had no effect on body weight. Once-daily administration of bretazenil (0.2 mg/kg, ip) was ineffective at preventing starvation after AgRP neuron ablation, suggesting that chronic activation of GABAA receptors is an important aspect of the rescue strategy. Chronic treatment with bretazenil also restored intra-oral sucrose consumption to >80% of original after ablation of AgRP neurons. Thus, both consummatory and appetitive responses are rescued by bretazenil treatment after AgRP neuron ablation. [4] Bretazenil displays anticonvulsant activity in vivo in various seizure models. Studies in mice have compared its anticonvulsant tolerance and withdrawal characteristics with other benzodiazepine receptor ligands. It has been evaluated in rat models of anxiety and seizure disorders, showing efficacy with a reduced tolerance profile compared to full agonists. |
| Enzyme Assay |
Non-cell receptor binding assays for Bretazenil are performed using membrane preparations from cells expressing recombinant GABAA receptors (e.g., α1β1γ2 subunits). Radioligand binding studies are conducted using [³H]flumazenil or [³⁵S]TBPS as tracers. Membranes are incubated with the radioligand and varying concentrations of Bretazenil (0.001-10 μM) in assay buffer. Nonspecific binding is determined with excess diazepam. Incubation is carried out at room temperature for 60-90 minutes, followed by filtration and scintillation counting.
|
| Cell Assay |
Cellular assays are performed using cell lines (e.g., HEK-293 or CHO cells) transfected with GABAA receptor subunit combinations. Whole-cell patch-clamp electrophysiology is used to record GABA-induced chloride currents in the presence of Bretazenil (0.001-100 μM). Alternatively, FLIPR-based membrane potential assays or calcium flux assays are used for high-throughput functional characterization. The potentiation of submaximal GABA responses is measured to determine the EC₅₀.
|
| Animal Protocol |
In vivo, various anxiolytic drugs exhibit a wide range of variability in their modulatory efficacy and potency of GABA action. For instance, bretazenil modulatory efficacy is much lower than that of diazepam. Such low efficacy could be due either to a preferential modulation of specific GABAA receptor subtypes or to a low modulatory efficacy at every GABAA receptor subtype. To address these questions we studied drug-induced modifications of GABA-activated Cl- currents in native GABAA receptors of cortical neurons in primary cultures and in recombinant GABAA receptors transiently expressed in transformed human embryonic kidney cells (293) after transfection with cDNAs encoding different molecular forms of alpha, beta, and gamma subunits of GABAA receptors. In cortical neurons the efficacy of bretazenil was lower than that of diazepam, whereas the potency of the two drugs was similar. In cells transfected with gamma 2 subunits and various molecular forms of alpha and beta subunits bretazenil efficacy was always lower than that of diazepam. However, in cells transfected with gamma 1 or gamma 3 subunits and various forms of alpha and beta subunits the efficacy of both diazepam and bretazenil was lower and always of similar magnitude. When bretazenil and diazepam were applied together to GABAA receptors including a gamma 2 subunit, the action of diazepam was curtailed in a manner related to the dose of bretazenil.[1]
In vivo animal studies are conducted in rodent models of anxiety (elevated plus maze, light-dark box) and seizure (pentylenetetrazole-induced or maximal electroshock seizure models). Bretazenil is administered via intraperitoneal or oral routes at doses ranging from 0.1-10 mg/kg. Anticonvulsant and anxiolytic effects are scored, and pharmacokinetic parameters are determined from plasma samples collected at various time points. |
| ADME/Pharmacokinetics |
Tolerance and withdrawal profiles of Bretazenil have been studied in mice, showing a more favorable profile compared to full agonists like diazepam. The compound shows good oral bioavailability and brain penetration. Metabolic studies indicate that Bretazenil is primarily metabolized by hepatic cytochrome P450 enzymes. The elimination half-life is approximately 2-4 hours in rodents.
|
| Toxicity/Toxicokinetics |
In preclinical toxicology studies, Bretazenil shows a safety profile consistent with benzodiazepine partial agonists. At high doses, sedation, ataxia, and respiratory depression may occur. No significant organ-specific toxicity has been reported at therapeutic doses. The compound has a wider therapeutic index compared to full benzodiazepine agonists due to its partial agonist properties.
|
| References |
[1]. Puia G, et al. Molecular mechanisms of the partial allosteric modulatory effects of bretazenil at gamma-aminobutyric acid type A receptor. Proc Natl Acad Sci U S A. 1992;89(8):3620-3624.
[2]. Finn DA, et al. A comparison of Ro 16-6028 with benzodiazepine receptor 'full agonists' on GABAA receptor function. Eur J Pharmacol. 1993;247(3):233-237. [3]. Kubová H, et al. Anticonvulsant effects of bretazenil (Ro 16-6028) during ontogenesis. Epilepsia. 1993;34(6):1130-1134. [4]. Cell. 2009 Jun 26;137(7):1225-34. |
| Additional Infomation |
Bretazenil (Ro16-6028) was developed as a partial agonist at the benzodiazepine site of GABAA receptors. Its partial agonist profile was designed to provide anxiolytic and anticonvulsant effects with reduced sedation, tolerance, and dependence compared to full agonists. Although it showed promise in preclinical studies, clinical development was not pursued extensively. It remains a valuable research tool for studying GABAA receptor pharmacology.
|
| Molecular Formula |
C19H20BRN3O3
|
|---|---|
| Molecular Weight |
418.2844
|
| Exact Mass |
417.068
|
| Elemental Analysis |
C, 54.56; H, 4.82; Br, 19.10; N, 10.05; O, 11.47
|
| CAS # |
84379-13-5
|
| PubChem CID |
107926
|
| Appearance |
White to off-white solid powder
|
| Density |
1.6±0.1 g/cm3
|
| Boiling Point |
594.3±50.0 °C at 760 mmHg
|
| Flash Point |
313.2±30.1 °C
|
| Vapour Pressure |
0.0±1.7 mmHg at 25°C
|
| Index of Refraction |
1.685
|
| LogP |
2.52
|
| Hydrogen Bond Donor Count |
0
|
| Hydrogen Bond Acceptor Count |
4
|
| Rotatable Bond Count |
3
|
| Heavy Atom Count |
26
|
| Complexity |
593
|
| Defined Atom Stereocenter Count |
1
|
| SMILES |
C(C1N=CN2C3C=CC=C(Br)C=3C(N3CCC[C@H]3C=12)=O)(=O)OC(C)(C)C
|
| InChi Key |
LWUDDYHYYNNIQI-ZDUSSCGKSA-N
|
| InChi Code |
InChI=1S/C19H20BrN3O3/c1-19(2,3)26-18(25)15-16-13-8-5-9-22(13)17(24)14-11(20)6-4-7-12(14)23(16)10-21-15/h4,6-7,10,13H,5,8-9H2,1-3H3/t13-/m0/s1
|
| Chemical Name |
tert-butyl
(S)-8-bromo-9-oxo-11,12,13,13a-tetrahydro-9H-benzo[e]imidazo[5,1-c]pyrrolo[1,2-a][1,4]diazepine-1-carboxylate
|
| Synonyms |
Ro 16-6028 Ro 16 6028 Ro16-6028 Ro 166028 Ro 166028 Ro166028 Ro 16-6028/000
|
| HS Tariff Code |
934.99.9001
|
| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
|
| Solubility (In Vitro) |
DMSO : ~50 mg/mL (~119.54 mM)
|
|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (5.98 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 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. Solubility in Formulation 2: ≥ 2.5 mg/mL (5.98 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. View More
Solubility in Formulation 3: ≥ 2.5 mg/mL (5.98 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. Solubility in Formulation 4: Solubility in Formulation 1: ≥ 2.5 mg/mL (6 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 take 100 μL of 25 mg/mL DMSO stock solution and add to 400 μL of PEG300, mix well (clear solution); Then add 50 μL of Tween 80 to the above solution, mix well (clear solution); Finally, add 450 μL of saline to the above solution, mix well (clear solution). Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 2.5 mg/mL (6 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 take 100 μL of 25 mg/mL DMSO stock solution and add to 900 μL of 20% SBE-β-CD in saline, mix well (clear solution). 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: ≥ 2.5 mg/mL (6 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 take 100 μL of 25 mg/mL DMSO stock solution and add to 900 μL of corn oil, mix well (clear solution). |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.3907 mL | 11.9537 mL | 23.9074 mL | |
| 5 mM | 0.4781 mL | 2.3907 mL | 4.7815 mL | |
| 10 mM | 0.2391 mL | 1.1954 mL | 2.3907 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
Products are for research use only; We do not sell to patients
Copyright 2020 InvivoChem LLC | All Rights Reserved
COA