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4-Nitrophenyl phosphate disodium hexahydrate is a substrate for alkaline phosphatase (ALP) enzymes. Upon hydrolysis by ALP, the compound is converted into 4-nitrophenolate, which can be monitored using UV-Vis spectrophotometry at 405 nm to quantify ALP activity. The compound does not have a pharmacological target but serves as a research tool for measuring enzyme activity.
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| ln Vitro |
4-Nitrophenyl phosphate (PNPP) disodium hexahydrate is a substrate that alkaline phosphatases (ALPs) frequently employ. 4-ALP hydrolyzes nitrophenyl phosphate disodium hexahydrate to PNP (p-nitrophenol), which uses the inner filter effect (IFE) to dim the fluorescence of new gold nanoclusters (AuNCs)[2].
In vitro, 4-nitrophenyl phosphate disodium hexahydrate is used as a chromogenic substrate for alkaline phosphatase. Upon hydrolysis, it is converted into 4-nitrophenolate, which can be monitored using UV-Vis spectrophotometry at 405 nm to quantify ALP activity. It is widely used in enzyme activity assays, ELISA, and other biochemical applications. |
| ln Vivo |
In vivo data for 4-nitrophenyl phosphate disodium hexahydrate as a therapeutic agent are not available, as the compound is a chromogenic substrate used exclusively for in vitro enzymatic assays. It is not intended for in vivo administration and has no established in vivo pharmacokinetic or pharmacodynamic profile.
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| Enzyme Assay |
For in vitro enzyme activity assays, 4-nitrophenyl phosphate disodium hexahydrate is used as a substrate for alkaline phosphatase. Standard protocols involve dissolving the compound in an appropriate buffer (e.g., 1 M diethanolamine buffer, pH 9.8) and adding it to the enzyme solution. The release of 4-nitrophenol is monitored at 405 nm over time using a spectrophotometer or plate reader. The initial rate of absorbance increase is proportional to ALP activity.
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| Cell Assay |
For in vitro cell-based experiments, 4-nitrophenyl phosphate disodium hexahydrate is not typically used directly in cell culture. It is a chromogenic substrate used for measuring ALP activity in cell lysates or biological fluids. Cells are lysed in appropriate buffer, and the substrate is added to the lysate. After incubation, absorbance is measured at 405 nm to quantify ALP activity.
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| Animal Protocol |
In vivo animal studies for 4-nitrophenyl phosphate disodium hexahydrate are not applicable, as the compound is a chromogenic substrate used exclusively for in vitro enzymatic assays. No animal studies have been conducted for this compound as a therapeutic agent.
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| ADME/Pharmacokinetics |
Pharmacokinetic data for 4-nitrophenyl phosphate disodium hexahydrate are not available, as the compound is not a drug and is not administered to living organisms. The compound is a hexahydrate salt and should be stored under appropriate conditions to maintain its stability.
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| Toxicity/Toxicokinetics |
4-Nitrophenyl phosphate disodium hexahydrate is a research chemical and should be handled with appropriate laboratory safety precautions. As a nitrophenyl compound, it may cause skin and eye irritation. The compound is for research use only and not for human therapeutic or diagnostic applications.
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| References |
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| Additional Infomation |
A type of hydrolase that can catalyze the hydrolysis of monophosphate esters to produce one mole of orthophosphate.
4-Nitrophenyl phosphate disodium hexahydrate (p-Nitrophenyl phosphate disodium hexahydrate) (CAS 333338-18-4) is primarily a research-grade chromogenic substrate, not an FDA-approved pharmaceutical drug. Its primary application is as a substrate for alkaline phosphatase in enzyme activity assays, ELISA, and other biochemical applications. |
| Molecular Formula |
C6H16NNA2O12P
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|---|---|
| Molecular Weight |
371.14
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| Exact Mass |
371.021
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| CAS # |
333338-18-4
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| Related CAS # |
4-Nitrophenyl phosphate ditromethamine;68189-42-4
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| PubChem CID |
2723710
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| Appearance |
Off-white to light yellow solid powder
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| Boiling Point |
457.8ºC at 760mmHg
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| Melting Point |
>300ºC(lit.)
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| Flash Point |
230.7ºC
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| LogP |
2.08
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| Hydrogen Bond Donor Count |
6
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| Hydrogen Bond Acceptor Count |
12
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| Rotatable Bond Count |
1
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| Heavy Atom Count |
22
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| Complexity |
238
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| Defined Atom Stereocenter Count |
0
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| SMILES |
C1=CC(=CC=C1[N+](=O)[O-])OP(=O)([O-])[O-].O.O.O.O.O.O.[Na+].[Na+]
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| InChi Key |
KAKKHKRHCKCAGH-UHFFFAOYSA-L
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| InChi Code |
InChI=1S/C6H6NO6P.2Na.6H2O/c8-7(9)5-1-3-6(4-2-5)13-14(10,11)12;;;;;;;;/h1-4H,(H2,10,11,12);;;6*1H2/q;2*+1;;;;;;/p-2
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| Chemical Name |
disodium;(4-nitrophenyl) phosphate;hexahydrate
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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) |
H2O: 100 mg/mL (269.44 mM)
DMSO: 10 mg/mL (26.94 mM) |
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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 | 2.6944 mL | 13.4720 mL | 26.9440 mL | |
| 5 mM | 0.5389 mL | 2.6944 mL | 5.3888 mL | |
| 10 mM | 0.2694 mL | 1.3472 mL | 2.6944 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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