β-adrenoceptor

Bufuralol (Ro 3-4787), which possesses the aromatic ring and basic nitrogen properties of CYP2D6 substrates, is frequently used to assess CYP2D6 activity [3].

In line with findings in myocardium Consistent [4], bufuralol (Ro 3-4787) has biphasic kinetics mediated by NADPH and is less effective than that seen in the presence of cumene hydroperoxide (CuOOH) in monkey disruption.

Cytochromea P450 2D6 (CYP2D6) is a highly polymorphic enzyme that metabolizes a large number of therapeutic drugs. To date, more than 100 CYP2D6 allelic variants have been reported. Among these variants, we recently identified 22 novel variants in the Chinese population. The aim of this study was to functionally characterize the enzymatic activity of these variants in vitro. A baculovirus-mediated expression system was used to express wild-type CYP2D6.1 and other variants (CYP2D6.2, CYP2D6.10 and 22 novel CYP2D6 variants) at high levels. Then, the insect microsomes containing expressed CYP2D6 proteins were incubated with Bufuralol or dextromethorphan at 37°C for 20 or 25 min., respectively. After termination, the metabolites were extracted and used for the detection with high-performance liquid chromatography. Among the 24 CYP2D6 variants tested, two variants (CYP2D6.92 and CYP2D6.96) were found to be catalytically inactive. The remaining 22 variants exhibited significantly decreased intrinsic clearance values for Bufuralol 1'-hydroxylation and 20 variants showed significantly lower intrinsic clearance values for dextromethorphan O-demethylation than those of the wild-type CYP2D6.1. Our in vitro results suggest that most of the variants exhibit significantly reduced catalytic activities compared with the wild-type, and these data provide valuable information for personalized medicine in Chinese and other Asian populations.[2]

---

Metabolic phenotype can be affected by multiple factors, including allelic variation and interactions with inhibitors. Human CYP2D6 is responsible for approximately 20% of cytochrome P450-mediated drug metabolism but consists of more than 100 known variants; several variants are commonly found in the population, whereas others are quite rare. Four CYP2D6 allelic variants-three with a series of mutations distal to the active site (*34, *17-2, *17-3) and one ultra-metabolizer with mutations near the active site (*53), along with reference *1 and an active site mutant of *1 (Thr309Ala)-were expressed, purified, and studied for interactions with the typical substrates dextromethorphan and Bufuralol and the inactivator SCH 66712. We found that *34, *17-2, and *17-3 displayed reduced enzyme activity and NADPH coupling while producing the same metabolites as *1, suggesting a possible role for Arg296 in NADPH coupling. A higher-activity variant, *53, displayed similar NADPH coupling to *1 but was less susceptible to inactivation by SCH 66712. The Thr309Ala mutant showed similar activity to that of *1 but with greatly reduced NADPH coupling. Overall, these results suggest that kinetic and metabolic analysis of individual CYP2D6 variants is required to understand their possible contributions to variable drug response and the complexity of personalized medicine.[3]

(+)-Bufuralol 1'-hydroxylation, a commonly used marker of hepatic CYP2D6 activity, was investigated in human and rhesus monkey intestinal microsomes and compared with that in hepatic microsomes. The cumene hydroperoxide (CuOOH)-mediated metabolism of (+)-bufuralol suggested that at least two enzymes were responsible for bufuralol 1'-hydroxylation in both human and monkey intestinal microsomes. In contrast, the kinetics of the CuOOH-mediated metabolism in human and monkey livers were monophasic. The Km values for the higher affinity component of the intestinal enzyme(s) of both species were similar to, while the corresponding Vmax values were much lower than, those obtained with the livers. Bufuralol metabolism mediated by NADPH exhibited biphasic kinetics and was less efficient than that observed in the presence of CuOOH in both human and monkey intestines, in agreement with the observations in the livers. Inhibition of bufuralol hydroxylase activity in the intestine and liver preparations from the same species by known CYP2D6 inhibitors/substrates was qualitatively similar. Quinidine was the most potent inhibitor of (+)-bufuralol 1'-hydroxylation in all tissues studied. Western immunoblots using anti-CYP2D6 peptide antibody revealed a protein band in human and monkey intestinal microsomes of the same molecular weight as that observed in the liver preparations. The intestinal CYP2D protein content appeared to be much less than that of liver, and correlated with the (+)-bufuralol hydroxylase activity. Immunoinhibition studies indicated significant (up to 50%) inhibition of the CuOOH-mediated (+)-bufuralol metabolism in human and monkey intestines only by anti-CYP2D6, and not by anti-CYP2A6, or anti-CYP2E1. Inhibition of the bufuralol 1'-hydroxylase activity by anti-rat CYP3A1 was only slight (20%) in human, but marked (60-65%) in monkey intestinal microsomes. The hepatic metabolism of (+)-bufuralol in humans and monkeys was only inhibited (75%) by anti-CYP2D6, but not by anti-CYP3A1. Overall, the results suggest that (1) tissue and species differences exist in the catalysis of (+)-bufuralol 1'-hydroxylation, and (2) CYP2D6-related enzymes are partially or primarily responsible for the bufuralol hydroxylase activity in human and monkey intestines or monkey liver[4].

Metabolism / Metabolites: Bufuralol has known human metabolites that include 1'2'-Ethenylbufuralol, 4-Hydroxybufuralol, and 6-Hydroxubufuralol.

---

Observations were made in eight subjects who exercised before and at 1, 2, 4, 6, 8 and 24 h after the double-blind oral administration of placebo, bufuralol 7.5, 15, 30, 60 and 120 mg and propranolol 40 and 160 mg. The exercise heart rate remained constant after placebo. Bufuralol 7.5 mg and propranolol 40 mg reduced exercise heart rate up to 6 and 8 h respectively after dosing but bufuralol 15, 30, 60 and 120 mg and propranolol 160 mg were still active at 24 h. The lowest exercise heart rate occurred at 2 h after all active treatments. Bufuralol 60 and 120 mg produced similar reduction in exercise tachycardia as propranolol 40 mg but less than propranolol 160 mg. Plasma levels of bufuralol and its two major metabolites were measured. The peak plasma concentrations of bufuralol occurred at 1.5 h after 7.5 mg and at 2 h after the other doses of bufuralol. In six subjects the plasma elimination half-life of bufuralol was 2.61 +/- 0.18 h and in the other three subjects 4.85 +/- 0.35 h. There was a corresponding longer time to peak concentration and plasma elimination half-life of the two metabolites in these three subjects. These findings show that bufuralol is a potent beta-adrenoceptor antagonist with partial agonist activity. It has a long duration of action and there is bimodal metabolism of the drug in man.[1]

rat LD50 oral 750 mg/kg BEHAVIORAL: CONVULSIONS OR EFFECT ON SEIZURE THRESHOLD; BEHAVIORAL: ATAXIA; LUNGS, THORAX, OR RESPIRATION: RESPIRATORY DEPRESSION Arzneimittel-Forschung. Drug Research., 27(1410), 1977 [PMID:20114]
rat LD50 subcutaneous 1400 mg/kg BEHAVIORAL: CONVULSIONS OR EFFECT ON SEIZURE THRESHOLD; BEHAVIORAL: ATAXIA; LUNGS, THORAX, OR RESPIRATION: RESPIRATORY DEPRESSION Arzneimittel-Forschung. Drug Research., 27(1410), 1977 [PMID:20114]
mouse LD50 oral 177 mg/kg BEHAVIORAL: CONVULSIONS OR EFFECT ON SEIZURE THRESHOLD; BEHAVIORAL: ATAXIA; LUNGS, THORAX, OR RESPIRATION: RESPIRATORY DEPRESSION Arzneimittel-Forschung. Drug Research., 27(1410), 1977 [PMID:20114]
mouse LD50 intraperitoneal 88 mg/kg BEHAVIORAL: CONVULSIONS OR EFFECT ON SEIZURE THRESHOLD; BEHAVIORAL: ATAXIA; LUNGS, THORAX, OR RESPIRATION: RESPIRATORY DEPRESSION Arzneimittel-Forschung. Drug Research., 27(1410), 1977 [PMID:20114]
mouse LD50 intravenous 29700 ug/kg BEHAVIORAL: CONVULSIONS OR EFFECT ON SEIZURE THRESHOLD; BEHAVIORAL: ATAXIA; LUNGS, THORAX, OR RESPIRATION: RESPIRATORY DEPRESSION Arzneimittel-Forschung. Drug Research., 27(1410), 1977 [PMID:20114]

[1]. T H Pringle, et al. Pharmacodynamic and pharmacokinetic studies on bufuralol in man. Br J Clin Pharmacol. 1986 Nov;22(5):527-34.
[2]. Jie Cai, et al. Effects of 22 Novel CYP2D6 Variants Found in the Chinese Population on the Bufuralol and Dextromethorphan Metabolisms In Vitro. Basic Clin Pharmacol Toxicol. 2016 Mar;118(3):190-9.
[3]. Sarah M Glass, et al. CYP2D6 Allelic Variants *34, *17-2, *17-3, and *53 and a Thr309Ala Mutant Display Altered Kinetics and NADPH Coupling in Metabolism of Bufuralol and Dextromethorphan and Altered Susceptibility to Inactivation by SCH 66712. Drug Metab Dispos. 2018 Aug;46(8):1106-1117.
[4]. T Prueksaritanont, et al. (+)-bufuralol 1'-hydroxylation activity in human and rhesus monkey intestine and liver. Biochem Pharmacol. 1995 Oct 26;50(9):1521-5.

C16H23NO2.HCL

297.82026

297.15

C, 64.53; H, 8.12; Cl, 11.90; N, 4.70; O, 10.74

59652-29-8

59652-29-8 (HCl); 54340-62-4; 60398-91-6 (racemic HCl)

151573

Typically exists as solid at room temperature

143-146ºC

4.609

3

3

5

20

287

0

CCC1=CC=CC2=C1OC(=C2)C(CNC(C)(C)C)O.Cl

KJBONRGCLLBWCJ-UHFFFAOYSA-N

InChI=1S/C16H23NO2.ClH/c1-5-11-7-6-8-12-9-14(19-15(11)12)13(18)10-17-16(2,3)4;/h6-9,13,17-18H,5,10H2,1-4H3;1H

2-(tert-butylamino)-1-(7-ethyl-1-benzofuran-2-yl)ethanol;hydrochloride

Bufuralol hydrochloride; 60398-91-6; Bufuralol HCl; 59652-29-8; Angium; bufuralol, hydrochloride; Bufuralol (hydrochloride); Ro-34787; Ro 34787; Ro34787; Ro3-4787; Ro 3-4787; Ro3-4787; Bufurolol hydrochloride;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

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

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

---

Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

View More

Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

---

Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

View More

Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

---

Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

---

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