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| Other Sizes |
| Targets |
As a leucine derivative with a tert-butyl ester protecting group, H-Leu-OtBu·HCl does not have a specific biological target. Its primary utility is as a chemical building block in peptide synthesis. However, leucine itself is a branched-chain amino acid that plays important roles in protein synthesis and metabolic regulation. In vitro studies have shown that leucine esters can increase phosphorylation of the mTORC1 substrate p70 ribosomal S6 kinase (p70S6K). The compound may also inhibit L-[¹⁴C]leucine transport, suggesting interactions with amino acid transporters. These effects are related to leucine's biological functions rather than specific pharmacological targeting of a receptor. In its protected ester form, the compound is primarily used as a synthetic intermediate.
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| ln Vitro |
Commercial ergot supplements have been made from amino acids and their derivatives. They affect the release of anabolic hormones, the availability of fuel for activity, the ability to think clearly under pressure, and the prevention of muscular damage brought on by exertion. They are regarded as advantageous synergistic food ingredients [1].
H-Leu-OtBu·HCl exhibits limited pharmacological activity in vitro beyond its role as a leucine precursor. In cellular studies, leucine and its derivatives are known to activate the mTOR signaling pathway, which regulates protein synthesis and cell growth. The compound may be used in cell culture to study the effects of leucine on mTORC1 activation or as a source of leucine for amino acid supplementation experiments. However, the compound's primary in vitro application is as a reagent in peptide synthesis rather than as a bioactive molecule. It does not possess intrinsic receptor binding, enzyme inhibition, or cytotoxic activities at concentrations typically used for synthesis. Its utility in cell-based assays is limited to providing leucine in a membrane-permeable form. |
| ln Vivo |
H-Leu-OtBu·HCl is not a pharmacologically active drug and does not have defined in vivo activity as a therapeutic agent. When administered to animals, the compound is expected to be hydrolyzed by esterases to release leucine, which then enters normal metabolic pathways. Leucine is an essential amino acid that plays critical roles in protein synthesis, muscle metabolism, and regulation of the mTOR signaling pathway. The compound may be used in nutritional studies to investigate leucine metabolism, amino acid transport, or the effects of leucine supplementation on muscle protein synthesis. These studies typically involve oral or intraperitoneal administration to rodents, followed by measurement of plasma leucine levels and assessment of metabolic or physiological responses.
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| Enzyme Assay |
In vitro enzyme assays for H-Leu-OtBu·HCl typically focus on esterase activity or amino acid transport. A standard protocol for esterase assays involves incubating the compound with an enzyme preparation, such as plasma, tissue homogenates, or purified esterases, in a suitable buffer at physiological pH. The hydrolysis of the tert-butyl ester releases leucine and tert-butanol, which can be quantified by HPLC, GC, or mass spectrometry. Alternatively, the decrease in substrate concentration can be monitored. For amino acid transport studies, the compound can be used as a substrate to investigate the specificity and kinetics of amino acid transporters. Radiolabeled or fluorescently labeled derivatives may be employed to measure uptake into cells or tissues. These assays help characterize the biochemical properties of the compound and its interactions with biological systems.
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| Cell Assay |
In vitro cellular assays using H-Leu-OtBu·HCl are typically conducted to study leucine metabolism or mTOR signaling. Cells are cultured in media supplemented with the compound, which is taken up by cells and hydrolyzed to release leucine. The effects of leucine on cellular processes such as protein synthesis, autophagy, or cell proliferation can be assessed. A common endpoint is the phosphorylation of p70S6K, a downstream target of mTORC1, which can be measured by Western blotting. These experiments are typically performed in cell lines such as C2C12 myotubes, HepG2 hepatocytes, or primary muscle cells. The compound's ability to penetrate cell membranes may be enhanced by the lipophilic tert-butyl ester group, making it useful for studying leucine's intracellular functions.
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| Animal Protocol |
In vivo animal studies with H-Leu-OtBu·HCl are primarily conducted in the context of nutritional science or metabolic research. A typical protocol involves oral gavage or intraperitoneal injection of the compound to rodents at doses ranging from 10 to 500 mg/kg. Blood samples are collected at various time points to measure leucine and tert-butanol levels, allowing assessment of the compound's absorption, hydrolysis, and pharmacokinetics. The compound's ability to elevate plasma leucine concentrations and its effects on muscle protein synthesis, mTOR signaling, or whole-body nitrogen balance may be evaluated. These studies help to understand the bioavailability of amino acid esters and their utility as leucine delivery agents for nutritional supplementation or metabolic research.
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| ADME/Pharmacokinetics |
As a leucine tert-butyl ester, H-Leu-OtBu·HCl is expected to be absorbed after oral administration, although detailed pharmacokinetic data are not well-documented. The compound is likely hydrolyzed by esterases in the gastrointestinal tract, liver, and plasma to release leucine and tert-butanol. The tert-butyl ester may enhance lipophilicity and membrane permeability compared to free leucine, potentially improving oral bioavailability. Following hydrolysis, leucine enters the endogenous amino acid pool and is distributed throughout the body via the circulation. Leucine is metabolized primarily in muscle tissue through transamination and oxidative decarboxylation. The pharmacokinetic profile of the compound is primarily determined by the rate of ester hydrolysis and the subsequent metabolism of leucine.
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| Toxicity/Toxicokinetics |
The hydrochloride salt of leucine tert-butyl ester is generally considered to have low toxicity, consistent with its use as a chemical reagent and its metabolic conversion to the essential amino acid leucine. Acute toxicity is expected to be minimal, as leucine has a very low toxicity profile. However, the compound may cause irritation upon contact with skin, eyes, or mucous membranes due to its acidic nature. Inhalation of the powder may cause respiratory irritation. The compound is not classified as a carcinogen, mutagen, or reproductive toxicant. Standard laboratory safety precautions, including the use of personal protective equipment, are recommended. No significant systemic toxicity is anticipated at typical research doses, as the compound is rapidly metabolized to leucine and tert-butanol.
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| References | |
| Additional Infomation |
L-Leucine tert-butyl ester hydrochloride (H-Leu-OtBu·HCl, CAS 2748-02-9) is a protected amino acid derivative used as a building block in peptide synthesis. Its chemical formula is C₁₀H₂₁NO₂·HCl and molecular weight is 223.74. The compound appears as a white to off-white solid. It is designed for biological research and industrial applications and is not intended for clinical or medical use. The tert-butyl ester group provides acid-labile protection for the carboxyl function, allowing selective deprotection during peptide synthesis. The compound is typically stored at room temperature in a cool, dry place.
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| Molecular Formula |
C10H22CLNO2
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| Molecular Weight |
223.7402
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| Exact Mass |
223.133
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| CAS # |
2748-02-9
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| PubChem CID |
7022311
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| Appearance |
White to off-white solid powder
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| Density |
0.929g/cm3
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| Boiling Point |
222.4ºC at 760 mmHg
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| Melting Point |
167 °C
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| Flash Point |
90.3ºC
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| Vapour Pressure |
0.102mmHg at 25°C
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| Index of Refraction |
1.444
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| LogP |
3.203
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| Hydrogen Bond Donor Count |
1
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| Hydrogen Bond Acceptor Count |
3
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| Rotatable Bond Count |
5
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| Heavy Atom Count |
13
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| Complexity |
170
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| Defined Atom Stereocenter Count |
1
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| SMILES |
Cl[H].O(C([C@]([H])(C([H])([H])C([H])(C([H])([H])[H])C([H])([H])[H])N([H])[H])=O)C(C([H])([H])[H])(C([H])([H])[H])C([H])([H])[H]
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| InChi Key |
RFUWRXIYTQGFGA-QRPNPIFTSA-N
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| InChi Code |
InChI=1S/C10H21NO2.ClH/c1-7(2)6-8(11)9(12)13-10(3,4)5;/h7-8H,6,11H2,1-5H3;1H/t8-;/m0./s1
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| Chemical Name |
tert-butyl (2S)-2-amino-4-methylpentanoate;hydrochloride
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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) |
DMSO : ≥ 50 mg/mL (~223.47 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 | 4.4695 mL | 22.3474 mL | 44.6947 mL | |
| 5 mM | 0.8939 mL | 4.4695 mL | 8.9389 mL | |
| 10 mM | 0.4469 mL | 2.2347 mL | 4.4695 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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