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| Targets |
DL-Propargylglycine (PAG) specifically and irreversibly targets cystathionine gamma-lyase (CTH or CSE), a pyridoxal-5-phosphate (PLP)-dependent enzyme. It acts as a suicide inhibitor, meaning it becomes covalently bound to the active site of the enzyme, leading to its permanent inactivation. By inhibiting CTH, PAG blocks the endogenous production of hydrogen sulfide (H2S) from L-cysteine in the transsulfuration pathway. This makes it a primary tool for studying H2S biology. At higher concentrations, it may also affect other PLP-dependent enzymes, but it is generally considered selective for CSE over CBS.
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| ln Vitro |
In vitro, DL-Propargylglycine is used to inhibit H2S production in cell cultures. Studies using primary mouse hepatocytes showed that treatment with PPG dramatically decreased both taurine and hypotaurine levels compared to untreated cells, confirming the role of CSE in the transsulfuration pathway. In cancer cell lines, inhibition of endogenous CSE by DL-PAG led to spontaneous epithelial-mesenchymal transition (EMT), characterized by decreased E-cadherin expression and increased vimentin expression. It is also used to study the role of H2S in cell proliferation, inflammation, and metabolic regulation across various cell types.
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| ln Vivo |
In vivo, DL-Propargylglycine is used to reduce H2S levels in animal models. In rats, inactivation of cystathionine gamma-lyase by D,L-propargylglycine was found to be less profound in the kidney than in the liver. It is used to study the role of endogenous H2S in disease models. For example, PAG treatment has been shown to alleviate symptoms of diseases that impair the formation or action of H2S. In cancer studies, DL-PAG is administered to investigate the contribution of CSE-derived H2S to tumor growth. It is also used in gastric mucosa research to study the upregulation of mRNA expressions of sodium bicarbonate cotransporter 1.
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| Enzyme Assay |
The in vitro enzyme inhibition assay for cystathionine gamma-lyase (CTH) using DL-Propargylglycine is typically performed in a 96-well plate format. The reaction mixture contains Tris-HCl buffer (pH 8.0), L-cysteine (substrate), pyridoxal phosphate (cofactor), and the purified CTH enzyme. DL-PAG is added at varying concentrations and pre-incubated with the enzyme for 15 minutes. The reaction is initiated by adding L-cysteine. After incubation at 37degC for 30-60 minutes, the production of H2S is detected using N,N-dimethyl-p-phenylenediamine and FeCl3 to form methylene blue, which is measured spectrophotometrically at 670 nm. The decrease in H2S production compared to control wells indicates the percentage of CTH inhibition.
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| Cell Assay |
For cell-based activity assays, primary hepatocytes or cancer cell lines are cultured in appropriate media. Cells are seeded in 6-well plates and allowed to reach 70-80% confluence. DL-Propargylglycine is then added to the culture medium at concentrations ranging from 0.5 mM to 5 mM. Cells are incubated for 24 to 48 hours. After treatment, cell lysates are collected, and the H2S production capacity is measured using a specific fluorescent H2S probe or the methylene blue method. Additionally, the levels of taurine and hypotaurine in the cell lysates or culture medium can be quantified by LC-MS/MS. The reduction in these metabolites compared to untreated control cells serves as a functional readout of CTH inhibition.
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| Animal Protocol |
A typical in vivo animal protocol for DL-Propargylglycine involves male Sprague-Dawley rats (200-250g) or C57BL/6 mice. Animals are randomized into treatment and control groups (n=6-8 per group). DL-PAG is dissolved in sterile saline and administered via intraperitoneal (IP) injection. A common dosing regimen is 50 mg/kg once daily for 7 to 14 consecutive days. For acute studies, a single IP injection may be sufficient. After the treatment period, blood is collected via cardiac puncture, and tissues of interest (liver, kidney, or tumor) are harvested. Tissue samples are homogenized to measure H2S content using a sulfide-sensitive electrode or biochemical assays. This protocol helps determine the extent of CTH enzyme inhibition in vivo and its downstream pathophysiological effects.
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| ADME/Pharmacokinetics |
DL-Propargylglycine is a synthetic amino acid analog, and its pharmacokinetics are characterized by rapid distribution into tissues, particularly the liver and kidneys. Following intraperitoneal administration, maximum tissue concentrations are achieved within 30-60 minutes. The half-life is relatively short (approximately 1-2 hours) due to renal clearance and potential metabolic degradation. The compound is not extensively protein-bound in plasma. Because it acts as an irreversible suicide inhibitor of CTH, its pharmacodynamic effect (H2S reduction) lasts significantly longer than the presence of the compound in the circulation, often persisting for 24-48 hours. Tissue-specific inhibition profiles have been observed, with less profound inactivation in the kidney compared to the liver.
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| Toxicity/Toxicokinetics |
Based on safety data from its parent compound (PAG, CAS 64165-64-6), DL-Propargylglycine is classified with GHS hazard statements H302 (Harmful if swallowed), H315 (Causes skin irritation), and H319 (Causes serious eye irritation). The acute oral LD50 is estimated to be around 1000-2000 mg/kg in rodents. In chronic animal studies, high doses of PAG have been associated with hepatotoxicity and nephrotoxicity due to the accumulation of its substrate cystathionine and disruption of the transsulfuration pathway. It should be handled with care in the laboratory, using personal protective equipment such as gloves and safety goggles to avoid exposure. As an irreversible enzyme inhibitor, off-target effects on other PLP-dependent enzymes are possible at high concentrations.
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| References | |
| Additional Infomation |
DL-Propargylglycine sodium is also commonly referred to as PAG, PPG, or 2-Amino-4-pentynoic acid. Its molecular formula for the free base is C5H7NO2. The compound is used not only as an H2S inhibitor but also in metabolic labeling strategies. For instance, it has been utilized in metabolic labeling of peptidoglycan with NIR-II dyes for in vivo imaging of gut microbiota. Additionally, it has applications in photothermal and photodynamic synergistic bacteria-infected wound therapy. The compound is typically stored as a powder at -20degC to maintain stability. It is soluble in DMSO and water, making it versatile for both in vitro and in vivo research applications.
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| Molecular Formula |
C5H6NNAO2
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| Molecular Weight |
135.10
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| Exact Mass |
135.03
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| CAS # |
73823-66-2
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| Related CAS # |
DL-Propargylglycine; DL-Propargylglycine hydrochloride
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| PubChem CID |
178201544
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| Appearance |
Light brown to brown solid powder
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| Hydrogen Bond Donor Count |
1
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| Rotatable Bond Count |
2
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| Heavy Atom Count |
9
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| Complexity |
137
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| Defined Atom Stereocenter Count |
0
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| SMILES |
C#CCC(C(=O)[O-])N.[Na+]
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| InChi Key |
WDRHMUGGOUVCLC-UHFFFAOYSA-M
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| InChi Code |
InChI=1S/C5H7NO2.Na/c1-2-3-4(6)5(7)8;/h1,4H,3,6H2,(H,7,8);/q;+1/p-1
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| Chemical Name |
sodium 2-aminopent-4-ynoate
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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 | 7.4019 mL | 37.0096 mL | 74.0192 mL | |
| 5 mM | 1.4804 mL | 7.4019 mL | 14.8038 mL | |
| 10 mM | 0.7402 mL | 3.7010 mL | 7.4019 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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