| Size | Price | Stock | Qty |
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| 5mg |
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| 10mg |
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| 25mg |
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| Targets |
Resorufin butyrate is not a drug and does not target a receptor; it is a chemical substrate that is recognized and hydrolyzed by the active sites of esterase, lipase, and cholinesterase enzymes. The compound's primary function is to serve as a probe for measuring the enzymatic activity of these hydrolases. Esterases and lipases cleave the ester bond between the butyrate group and the resorufin core, resulting in the release of the resorufin fluorophore. The increase in fluorescence is directly proportional to enzyme activity in the sample.
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| ln Vitro |
In 384-well plates with 12 nM bLPL enzyme and substrate in PBS, pH 7.5, 5% DMSO, resorubin butyrate (2.3 μM) grows linearly during 60 minutes [1].
In vitro, Resorufin butyrate is used as a fluorogenic substrate for triglyceride lipases and cholinesterases. In a typical assay, the non-fluorescent compound is incubated with a biological sample containing esterase or lipase enzymes. Upon enzymatic cleavage, the resorufin fluorophore is released, generating a strong red-orange fluorescence signal (Ex 570 nm, Em 580 nm). The rate of fluorescence increase is proportional to the amount of enzyme present in the sample. This enables quantification of esterase/lipase activity in tissue homogenates, plasma, cell lysates, or purified enzyme preparations. It is used in high-throughput screening to identify modulators of these enzymes. |
| ln Vivo |
In vivo, Resorufin butyrate is not used as a therapeutic agent but can be employed as a molecular probe in animal models to assess esterase and lipase activity in real time. Following local or systemic administration, the compound is hydrolyzed by endogenous esterases, and the resorufin fluorescence can be detected using intravital fluorescence imaging or by harvesting tissues and measuring resorufin fluorescence in homogenates. This approach can be used to study the biodistribution and activity of esterases in various organs, including the liver, intestine, and kidney. The short half-life of the compound (rapid hydrolysis) necessitates immediate analysis.
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| Enzyme Assay |
For non-cell-based enzyme assays, a standard fluorometric protocol is used. Resorufin butyrate is prepared as a stock solution of 10 mM in DMSO. The assay is performed in black 96-well plates. In a final volume of 200 uL, the reaction mixture contains 50 mM Tris-HCl (pH 7.4), 150 mM NaCl, 0.1% Triton X-100, 100 uM Resorufin butyrate, and varying concentrations of purified lipase (e.g., porcine pancreatic lipase, 0.1-10 U/mL). Alternatively, for cholinesterase activity, 0.1 U/mL of acetylcholinesterase can be used. The plate is incubated at 37degC for 30-60 min. Fluorescence is measured using a fluorescence plate reader with excitation at 570 nm and emission at 580 nm. For kinetic studies, measurements are taken every 1-5 minutes. The increase in relative fluorescence units (RFU) over time is used to calculate enzyme activity, and the IC50 of an inhibitor is determined by adding varying concentrations of the test compound to the reaction mixture.
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| Cell Assay |
For in vitro cell-based assays, cells of interest (e.g., hepatocytes, adipocytes, or cancer cell lines) are seeded in black-walled 96-well plates at 2 × 10^4 cells/well. After 24 hours of culture, the medium is removed, and cells are washed with PBS. The cells are then incubated with 100 uL of assay buffer (PBS or HEPES-buffered saline) containing 50 uM Resorufin butyrate. For inhibitor studies, test compounds (esterase/lipase inhibitors) are pre-incubated with cells for 15-30 minutes before adding the substrate. The plate is incubated at 37degC for 30-60 minutes protected from light. Fluorescence is measured using a fluorescence microplate reader at Ex 570 nm/Em 580 nm. The fluorescence signal is proportional to the cellular esterase activity. As a positive control, cells can be lysed with 0.1% Triton X-100 to release cellular esterases.
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| Animal Protocol |
For in vivo animal studies, Resorufin butyrate is typically administered intravenously or intraperitoneally to mice (6-8 weeks old, n=5 per group). A typical dose is 5-10 mg/kg body weight in a formulation of 10% DMSO/90% saline (final volume 100-200 uL). At predetermined time points (5, 15, 30, 60, 120 minutes post-injection), mice are euthanized, and organs (liver, kidney, lung, heart, intestine) are harvested. Tissues are homogenized in PBS containing 0.1% Triton X-100, and the homogenate is centrifuged to remove debris. The supernatant (100 uL) is transferred to a black 96-well plate, and fluorescence is measured at Ex 570 nm/Em 580 nm. The resorufin concentration in each tissue is calculated using a standard curve of resorufin (0-10 uM). This method quantifies the distribution of esterase activity across different organs. Resorufin butyrate can also be used in transgenic mouse models to assess tissue-specific esterase expression.
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| ADME/Pharmacokinetics |
Resorufin butyrate is not designed for systemic pharmacokinetic studies as a drug. The compound itself is rapidly hydrolyzed by esterases in the blood and tissues, with a half-life in plasma of less than 5 minutes. The resulting product, resorufin, has a plasma half-life of approximately 15-30 minutes in mice. Resorufin is metabolized by glucuronidation in the liver and is excreted in the bile and urine. The parent compound (Resorufin butyrate) is not detectable in plasma beyond the first few minutes post-injection due to rapid enzymatic hydrolysis. The compound's fluorescence properties make it suitable for real-time monitoring of esterase activity in vivo using intravital imaging techniques.
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| Toxicity/Toxicokinetics |
Resorufin butyrate is a fluorogenic substrate used for research applications and is not intended for human use. The compound exhibits low toxicity in vitro and in vivo at the concentrations used in typical assays (10-100 uM in vitro, 5-10 mg/kg in vivo). However, high concentrations or prolonged exposure may cause cellular stress due to the accumulation of resorufin. Resorufin itself has been reported to be a weak inducer of reactive oxygen species (ROS) at high concentrations (>100 uM). No specific acute or chronic toxicity data is available. Standard safety precautions should be followed when handling this compound, including the use of gloves, a lab coat, and eye protection. Avoid direct skin contact.
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| References |
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| Additional Infomation |
Resorufin butyrate is a widely used fluorogenic substrate in drug discovery, enzyme kinetics, and diagnostic applications. The substrate is specific for esterases and lipases; it is not hydrolyzed by proteases, phosphatases, or other hydrolases under standard assay conditions. The butyrate ester length can be varied (e.g., acetate, propionate, butyrate, valerate) to generate substrates with different specificity profiles for various esterases. Resorufin butyrate is also used as a substrate for cholinesterase activity assays, and it is commonly employed in screens for potential esterase inhibitors as therapeutic agents for conditions such as metabolic syndrome, obesity, and neurological disorders. This product is for research use only.
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| Molecular Formula |
C16H13NO4
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|---|---|
| Molecular Weight |
283.27872
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| Exact Mass |
283.084
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| CAS # |
15585-42-9
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| PubChem CID |
5007916
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| Appearance |
Yellow to orange solid powder
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| Density |
1.31g/cm3
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| Boiling Point |
428.7ºC at 760 mmHg
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| Melting Point |
136-138ºC(lit.)
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| Flash Point |
194.5ºC
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| Vapour Pressure |
1.48E-07mmHg at 25°C
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| Index of Refraction |
1.62
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| LogP |
2.998
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| Hydrogen Bond Donor Count |
0
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| Hydrogen Bond Acceptor Count |
5
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| Rotatable Bond Count |
4
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| Heavy Atom Count |
21
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| Complexity |
547
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| Defined Atom Stereocenter Count |
0
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| InChi Key |
RGJTZDRUTUWWTD-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C16H13NO4/c1-2-3-16(19)20-11-5-7-13-15(9-11)21-14-8-10(18)4-6-12(14)17-13/h4-9H,2-3H2,1H3
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| Chemical Name |
(7-oxophenoxazin-3-yl) butanoate
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| HS Tariff Code |
2934.99.9001
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| Storage |
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. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
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
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| Solubility (In Vivo) |
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 Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *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)] Oral Formulations
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 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.5301 mL | 17.6504 mL | 35.3008 mL | |
| 5 mM | 0.7060 mL | 3.5301 mL | 7.0602 mL | |
| 10 mM | 0.3530 mL | 1.7650 mL | 3.5301 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.
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