CYP2

Bufuralol 1'-hydroxylation is a prototypical reaction catalyzed by cytochrome P450 (P450) 2D6, an enzyme known to show debrisoquine/sparteine-type genetic polymorphism in humans. In the present study we further examined the roles of several human P450 enzymes, as well as P450 2D6, in the hydroxylation of (+/-)-bufuralol, using liver microsomes from several human samples and human P450 enzymes expressed in human lymphoblastoid cell lines or Escherichia coli. Kinetic analysis of bufuralol 1'-hydroxylation by liver microsomes showed that there were different Km and Vmax values in seven human samples examined; low Km values (approximately 0.05 mM) were observed in four samples (including sample HL-18), high Km values (approximately 0.25 mM) in two samples (including sample HL-67), and an intermediate Km value (approximately 0.1 mM) in one sample. Quinidine and anti-rat P450 2D1 antibody almost completely inhibited bufuralol 1'-hydroxylation in human sample HL-18 at a substrate concentration of 0.4 mM, whereas these effects were not so drastic when liver microsomes from human sample HL-67 were used. In contrast, a very low concentration (< 10 microM) of alpha-naphthoflavone or anti-human P450 1A2 antibody significantly inhibited bufuralol 1'-hydroxylation catalyzed by human sample HL-67, but not HL-18, with 0.4 mM bufuralol. When the relative contents of P450 2D6 and P450 1A2 in 20 human samples were determined, bufuralol 1'-hydroxylation in samples containing large amounts of P450 2D6 tended to be more sensitive to quinidine, whereas the P450 1A2-rich samples were highly susceptible to alpha-naphthoflavone. However, at low substrate concentrations bufuralol 1'-hydroxylation was shown to be catalyzed principally by P450 2D6, based on the inhibitory effects of anti-rat P450 2D1 antibody and quinidine, in both human samples HL-18 and HL-67. At least five other, minor, bufuralol products were formed by human liver microsomes, in addition to 1'-hydroxybufuralol. Two of them were identified as 4- and 6-hydroxybufuralol by 1H NMR spectroscopy and mass spectrometry. The formation of the 4- and 6-hydroxylated products was suggested to be catalyzed by P450 1A2, based on the results of correlation with P450 1A2 contents in 60 human samples and inhibition by anti-P450 1A2 and alpha-naphthoflavone. Purified recombinant P450 1A2 (expressed in E. coli) produced 1'-, 4-, and 6-hydroxybufuralol in a reconstituted system, although P450 2D6 (expressed in human lymphoblast cell lines) was found to catalyze only bufuralol 1'-hydroxylation[2].

[1]. D₂-dopaminergic receptor-linked pathways: critical regulators of CYP3A, CYP2C, and CYP2D. Mol Pharmacol. 2012 Oct;82(4):668-78.

[2]. Bufuralol hydroxylation by cytochrome P450 2D6 and 1A2 enzymes in human liver microsomes. Mol Pharmacol. 1994 Sep;46(3):568-77.

Various hormonal and monoaminergic systems play determinant roles in the regulation of several cytochromes P450 (P450s) in the liver. Growth hormone (GH), prolactin, and insulin are involved in P450 regulation, and their release is under dopaminergic control. This study focused on the role of D₂-dopaminergic systems in the regulation of the major drug-metabolizing P450s, i.e., CYP3A, CYP2C, and CYP2D. Blockade of D₂-dopaminergic receptors with either sulpiride (SULP) or 4-(4-chlorophenyl)-1-(1H-indol-3-ylmethyl)piperidin-4-ol (L-741,626) markedly down-regulated CYP3A1/2, CYP2C11, and CYP2D1 expression in rat liver. This suppressive effect appeared to be mediated by the insulin/phosphatidylinositol 3-kinase/Akt/FOXO1 signaling pathway. Furthermore, inactivation of the GH/STAT5b signaling pathway appeared to play a role in D₂-dopaminergic receptor-mediated down-regulating effects on these P450s. SULP suppressed plasma GH levels, with subsequently reduced activation of STAT5b, which is the major GH pulse-activated transcription factor and has up-regulating effects on various P450s in hepatic tissue. Levels of prolactin, which exerts down-regulating control on P450s, were increased by SULP, which may contribute to SULP-mediated effects. Finally, it appears that SULP-induced inactivation of the cAMP/protein kinase A/cAMP-response element-binding protein signaling pathway, which is a critical regulator of pregnane X receptor and hepatocyte nuclear factor 1α, and inactivation of the c-Jun N-terminal kinase contribute to SULP-induced down-regulation of the aforementioned P450s. Taken together, the present data provide evidence that drugs acting as D₂-dopaminergic receptor antagonists might interfere with several major signaling pathways involved in the regulation of CYP3A, CYP2C, and CYP2D, which are critical enzymes in drug metabolism, thus affecting the effectiveness of the majority of prescribed drugs and the toxicity and carcinogenic potency of a plethora of toxicants and carcinogens.[1]

C16H23NO3

277.36

277.168

57704-16-2

1'-Hydroxy bufuralol-d9;1185069-74-2

162836

Off-white to light yellow solid powder

1.139g/cm3

433ºC at 760mmHg

41-49ºC

215.7ºC

1.577

3.298

3

4

5

20

315

0

GTYMTYBCXVOBBB-UHFFFAOYSA-N

InChI=1S/C16H23NO3/c1-10(18)12-7-5-6-11-8-14(20-15(11)12)13(19)9-17-16(2,3)4/h5-8,10,13,17-19H,9H2,1-4H3

2-(tert-butylamino)-1-[7-(1-hydroxyethyl)-1-benzofuran-2-yl]ethanol

1'-Hydroxybufuralol; 57704-16-2; 1'-Hydroxy bufuralol; 2-(tert-butylamino)-1-[7-(1-hydroxyethyl)-1-benzofuran-2-yl]ethanol; 5,6-Dimethoxyindane-1,3-dione; Ro 037410; Ro 03-7410; 1&prime-Hydroxybufuralol;

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, 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)

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.

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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).

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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)

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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).

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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

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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.

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