| Size | Price | Stock | Qty |
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| 1mg |
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| Other Sizes |
| Targets |
(Rac)-Timolol-d5 maleate is a stable isotope-labeled internal standard that targets the same beta-adrenergic receptors (beta-ARs) as its non-labeled counterpart. Timolol acts as a non-selective beta-adrenergic receptor antagonist, blocking both beta1 and beta2 receptors with equal potency. In the cardiovascular system, blocking beta1 receptors reduces heart rate, contractility, and cardiac output. In the eye, the mechanism of action for lowering intraocular pressure (IOP) is not fully understood, but it is thought to work primarily by reducing the production of aqueous humor by the ciliary body via beta2 receptor blockade. This leads to decreased outflow resistance and lower IOP. The deuterated version, (Rac)-Timolol-d5 maleate, is not used for its pharmacological effect but rather as an analytical standard to accurately quantify the concentration of Timolol in research samples.
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| ln Vitro |
Drug compounds have included stable heavy isotopes of carbon, hydrogen, and other elements, mostly as quantitative tracers while the drugs were being developed. Because deuteration may have an effect on a drug's pharmacokinetics and metabolic properties, it is a cause for concern [1].
The in vitro biological activity of (Rac)-Timolol-d5 maleate is expected to be the same as its unlabeled parent, Timolol maleate. Timolol is a potent, non-selective beta-adrenergic receptor antagonist. In vitro, it competitively inhibits the binding of isoproterenol (a beta-agonist) to beta1 and beta2 receptors in membrane preparations with a Ki (inhibition constant) in the low nanomolar range. In isolated rabbit iris-ciliary body preparations, Timolol (0.1-10 uM) has been shown to effectively inhibit the production of aqueous humor in a concentration-dependent manner. This effect is mediated by the blockade of beta2 adrenergic receptors on the non-pigmented epithelial cells of the ciliary body, which are responsible for fluid secretion. The labeled (Rac)-Timolol-d5 maleate is not typically used in these efficacy assays but serves as an internal standard for LC-MS analysis to validate drug concentrations in the organ bath or culture medium. |
| ln Vivo |
(Rac)-Timolol-d5 maleate is a stable isotope-labeled compound used as an analytical internal standard, not for direct pharmacological evaluation. The in vivo activity of its non-labeled counterpart, Timolol maleate, is well-characterized. In animal models (e.g., normotensive or hypertensive rabbits, dogs, monkeys), topical ophthalmic administration of Timolol maleate (0.25-0.5% solution) significantly reduces intraocular pressure (IOP) within 30-60 minutes, with the maximal effect observed at 2 hours and duration of action lasting up to 24 hours. Systemically, when administered intravenously (0.1-1 mg/kg), it reduces heart rate and blood pressure in normotensive rats and dogs. The drug has no intrinsic sympathomimetic (ISA) or membrane-stabilizing activity. (Rac)-Timolol-d5 maleate is used as an internal standard for the precise quantification of Timolol in these animal model studies, allowing for accurate assessment of its ocular absorption and systemic exposure.
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| Enzyme Assay |
A generic non-cell-based assay for (Rac)-Timolol-d5 maleate involves the establishment of a calibration curve for Timolol. Prepare a standard stock solution of unlabeled Timolol maleate in methanol (1 mg/mL). Prepare a separate stock solution of the internal standard ((Rac)-Timolol-d5 maleate) at the same concentration. Prepare calibration standards by spiking the unlabeled analyte into a blank matrix (e.g., artificial aqueous humor or plasma) to achieve concentrations ranging from 0.1 ng/mL to 1000 ng/mL. To each calibration standard, add a fixed concentration of the internal standard (e.g., 50 ng/mL). Also prepare blank and double-blank samples. For sample preparation, add 200 uL of acetonitrile to 100 uL of standard to precipitate proteins. Centrifuge at 10,000g for 5 minutes. Transfer the supernatant to autosampler vials. Analyze samples using LC-MS/MS in positive ion mode. Monitor mass transitions: m/z 317.2 → 261.1 for Timolol, and m/z 322.2 → 266.1 for Timolol-d5. Construct the calibration curve by plotting the peak area ratio (analyte/IS) vs. the nominal concentration. This curve is used to calculate the concentration of Timolol in unknown samples.
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| Cell Assay |
(Rac)-Timolol-d5 maleate is not typically used in cell-based assays for efficacy, as its non-labeled parent compound is used. A standard protocol for testing the pharmacological activity of Timolol involves the use of cells overexpressing the human beta-2 adrenergic receptor (beta2-AR). Culture CHO-K1 cells stably expressing the human beta2-AR in Ham‘s F12 medium supplemented with 10% fetal bovine serum and antibiotics. Seed the cells in 96-well plates (20,000 cells/well) and grow them for 24 hours. Then, measure the inhibition of cAMP production using a homogeneous time-resolved fluorescence (HTRF) assay. First, treat the cells with the phosphodiesterase inhibitor IBMX to prevent cAMP breakdown. Then, add increasing concentrations of unlabeled Timolol maleate (0.1 nM to 10 uM) to the wells and incubate for 15 minutes. Subsequently, stimulate the cells with 1 uM isoproterenol (a beta-agonist) for 30 minutes to induce cAMP production. Lyse the cells and add the HTRF detection reagents (cAMP-d2 and anti-cAMP-Eu cryptate). Read the fluorescence at 620 nm and 665 nm. Calculate the ratio (665/620) and generate a dose-response curve. Determine the IC₅0 for the inhibition of cAMP production. Use the deuterated Timolol as an internal standard to measure drug concentration in the medium.
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| Animal Protocol |
A standard in vivo animal experiment protocol for (Rac)-Timolol-d5 maleate involves a pharmacokinetic (PK) study in rabbits, which are a common model for ocular drug delivery. Use male New Zealand White rabbits (2-3 kg, n=4-5). Administer a single topical ocular dose (e.g., 35 uL of a 0.5% solution) of unlabeled Timolol maleate to the lower conjunctival sac of one eye. Collect blood samples from the marginal ear vein at various time points (0, 0.25, 0.5, 1, 2, 4, 6, 8, 12, 24 h). Additionally, collect aqueous humor samples by paracentesis at selected time points (0.5, 1, 2, 4 h). Immediately process the blood samples to obtain plasma. Store all samples at -80degC until analysis. For bioanalysis, spike the plasma and aqueous humor samples with a fixed concentration of (Rac)-Timolol-d5 maleate as the internal standard. Precipitate proteins with acetonitrile and analyze by LC-MS/MS. Calculate the PK parameters (Cmax, Tmax, AUC, t½) in both plasma and aqueous humor using WinNonlin software.
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| ADME/Pharmacokinetics |
(Rac)-Timolol-d5 maleate is not a therapeutic drug but an internal standard. The pharmacokinetics of its non-labeled parent, Timolol maleate, are well established. Following topical ocular administration, Timolol is absorbed systemically through the conjunctiva and nasal mucosa. The plasma elimination half-life of Timolol in humans is approximately 2-5 hours. It has a moderate volume of distribution (approx. 2 L/kg) and low plasma protein binding (approx. 10-20%). Timolol is extensively metabolized in the liver by CYP2D6 to several metabolites, including an inactive hydroxy derivative. Approximately 20% of a systemic dose is excreted unchanged in the urine. In patients with renal impairment, the half-life can be prolonged. The labeled (Rac)-Timolol-d5 maleate allows for highly accurate quantification of Timolol in PK studies, especially in ocular tissues where matrix effects are prominent, enabling precise determination of these parameters.
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| Toxicity/Toxicokinetics |
(Rac)-Timolol-d5 maleate is a research-grade compound used as an analytical internal standard, not as a therapeutic. The toxicity profile of its non-labeled counterpart, Timolol maleate, is well known from clinical use. In humans, ophthalmic Timolol is generally well-tolerated locally, but systemic absorption can cause beta-blockade side effects including bradycardia, hypotension, bronchospasm (due to beta2 blockade in the lungs), dizziness, and fatigue. In animal toxicity studies, the oral LD₅0 of Timolol maleate is approximately 1,200 mg/kg in rats. At high doses, it produces signs of exaggerated pharmacology such as bradycardia, reduced blood pressure, and respiratory depression. It is not teratogenic in rats and rabbits at clinically relevant doses. For laboratory handling of (Rac)-Timolol-d5 maleate, standard safety precautions should be followed, including the use of gloves and a lab coat. It should be stored as a powder at -20degC.
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| References |
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| Additional Infomation |
(Rac)-Timolol-d5 maleate is a stable isotope-labeled version of the non-selective beta-adrenergic receptor antagonist Timolol. It is exclusively intended for research use and serves as an internal standard for the accurate quantification of Timolol in biological samples by LC-MS/MS. Timolol maleate is a standard medication used to lower intraocular pressure (IOP) in the treatment of glaucoma and ocular hypertension. It is also used off-label for other conditions like infantile hemangiomas. The drug works by reducing the production of aqueous humor. The racemic mixture (Rac) refers to the presence of both S-enantiomer (which is the active pharmaceutical ingredient) and the R-enantiomer. This labeled version corrects for matrix effects and extraction losses, allowing precise measurement of Timolol levels in research settings, including pharmacokinetic and bioequivalence studies.
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| Molecular Formula |
C17H28N4O7S
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| Molecular Weight |
437.522631645203
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| Exact Mass |
437.199
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| CAS # |
1217260-21-3
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| Related CAS # |
(S)-Timolol Maleate;26921-17-5;Timolol;26839-75-8
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| PubChem CID |
46783095
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| Appearance |
White to off-white solid powder
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| Hydrogen Bond Donor Count |
4
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| Hydrogen Bond Acceptor Count |
12
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| Rotatable Bond Count |
9
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| Heavy Atom Count |
29
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| Complexity |
429
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| Defined Atom Stereocenter Count |
0
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| SMILES |
S1N=C(C(=N1)N1C([H])([H])C([H])([H])OC([H])([H])C1([H])[H])OC([2H])([2H])C([2H])(C([2H])([2H])N([H])C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H])O[H].O([H])C(/C(/[H])=C(/[H])\C(=O)O[H])=O
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| InChi Key |
WLRMANUAADYWEA-RJAXMONVSA-N
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| InChi Code |
InChI=1S/C13H24N4O3S.C4H4O4/c1-13(2,3)14-8-10(18)9-20-12-11(15-21-16-12)17-4-6-19-7-5-17;5-3(6)1-2-4(7)8/h10,14,18H,4-9H2,1-3H3;1-2H,(H,5,6)(H,7,8)/b;2-1-/i8D2,9D2,10D;
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| Chemical Name |
(Z)-but-2-enedioic acid;1-(tert-butylamino)-1,1,2,3,3-pentadeuterio-3-[(4-morpholin-4-yl-1,2,5-thiadiazol-3-yl)oxy]propan-2-ol
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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 |
| 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 | 2.2856 mL | 11.4280 mL | 22.8561 mL | |
| 5 mM | 0.4571 mL | 2.2856 mL | 4.5712 mL | |
| 10 mM | 0.2286 mL | 1.1428 mL | 2.2856 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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