| Size | Price | Stock | Qty |
|---|---|---|---|
| 10g |
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| Other Sizes |
Purity: ≥98%
| Targets |
Triacylglycerol (TG) lipases hydrolyze triacylglycerol to diacylglycerol and free fatty acids. The major TG lipases in murine white adipose tissue (WAT) are adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL). The HSL-specific inhibitor 76-0079 (NNC 0076-0000-0079) was used to differentiate between these activities. [1]
In rat liver, alkaline triacylglycerol lipase activity (pH 7.5) is associated with the plasma membrane and released by heparin. [2] In yeast, Tgl4 is a major TG lipase and a functional ortholog of murine ATGL, activated by cyclin-dependent kinase 1 (Cdk1/Cdc28)-dependent phosphorylation at threonine 675 and serine 890. [3] |
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| ln Vitro |
In cytosolic preparations of murine WAT, the HSL-specific inhibitor 76-0079 (5 μM) reduced TG hydrolase activity in wild-type and ATGL-knockout (ATGL-ko) WAT but had no effect on HSL-knockout (HSL-ko) WAT, demonstrating its specificity for HSL. Combined inhibition of ATGL and HSL (using ATGL-ko cytosol plus 76-0079) resulted in >95% reduction of TG hydrolase activity. [1]
CGI-58, a coactivator of ATGL, stimulated TG hydrolase activity in wild-type and HSL-ko WAT cytosolic preparations by 1.7- and 2.1-fold, respectively, but had no effect in ATGL-ko WAT, indicating ATGL is the sole target for CGI-58. [1] In rat liver homogenates, alkaline TG lipase activity was optimal at pH 7.5, with linearity maintained for approximately 20 minutes at substrate concentrations >3 mM. No significant accumulation of di- or mono-[1-¹⁴C]oleoylglycerol was detected, suggesting the rate of degradation of triacylglycerol is limited by the first ester bond hydrolysis. [2] In yeast, Tgl4 lipase activity was stimulated by Cdk1/Cdc28-dependent phosphorylation. Mutation of phosphorylation sites (threonine 675 and serine 890 to alanine) reduced lipolytic activity by up to 90% in vivo, while mutation to glutamic acid (mimicking constitutive phosphorylation) resulted in high lipolytic activity resistant to Cdk1 inhibition. [3] |
| ln Vivo |
In murine WAT organ cultures, forskolin-stimulated FFA release was reduced by 38% in HSL-ko and 70% in ATGL-ko WAT compared to wild-type. The HSL inhibitor 76-0079 (25 μM) reduced forskolin-stimulated FFA release by 68% in wild-type and 94% in ATGL-ko WAT. In HSL-ko WAT, 76-0079 had no effect. [1]
In rats, streptozotocin-induced diabetes reduced hepatic alkaline TG lipase specific activity to 39% of normal at 24 hours, remaining low at 72 hours and 7 days. Chronic insulin treatment (4 subcutaneous injections of 10 U/kg protamine zinc insulin at 16-hour intervals) increased total lipase activity in normal rats (by 40%) and diabetic rats (to 155% of normal). Withholding insulin for 32 hours reduced activity to 65% of normal. Starvation for 24 hours also significantly reduced lipase activity (35% of control). Acute insulin (35 U/kg, 90 min) or glucagon (1 mg/kg, 30 min) had no effect on hepatic alkaline lipase activity. [2] In yeast, Tgl4 phosphorylation and lipolysis were most active during late G1 phase of the cell cycle, coinciding with bud formation. tgl3 tgl4 double mutants lacking TG lipolysis showed delayed bud formation (~30 min delay) and delayed entry into S phase. In the presence of cerulenin (which inhibits de novo fatty acid synthesis), bud formation was blocked in tgl3 tgl4 mutants but not in wild-type cells. [3] |
| Enzyme Assay |
TG Hydrolase Assay (Murine): The substrate containing triolein and [9,10-³H]triolein was emulsified with phosphatidylcholine/phosphatidylinositol. Cytosolic fractions (0.1 mL) were incubated with 0.1 mL substrate (167 nmol triolein/assay, 8000 cpm/nmol) at 37°C for 60 minutes. The reaction was terminated with methanol/chloroform/heptane (10:9:7) and potassium carbonate/boric acid buffer (pH 10.5). Radioactivity in the upper phase was determined by liquid scintillation counting. [1]
Lipase Assay (Rat Liver): Tri[1-¹⁴C]oleoylglycerol was emulsified with 5% gum arabic. The assay mixture (1 mL) contained 10 mg fatty acid-poor bovine serum albumin, 3.2 μmol trioleoylglycerol with 0.08 μCi tri[1-¹⁴C]oleoylglycerol, and liver protein in 0.10 M potassium phosphate, 1 mM EDTA, pH 7.5. Incubations were at 37°C for 10-40 minutes. [¹⁴C]oleate liberation was determined by liquid scintillation spectrometry. [2] |
| Cell Assay |
Organ Culture of Murine WAT: Gonadal fat pads (~20 mg) were incubated in DMEM containing 2% fatty acid-free BSA with or without 10 μM forskolin and/or the HSL inhibitor 76-0079 for 1 hour at 37°C. Tissues were then transferred to fresh medium and incubated for another 60 minutes. Aliquots of medium were analyzed for FFA and glycerol content using commercial kits. [1]
Yeast Cell Culture and Analysis: Yeast cells (wild-type, tgl3, tgl4, tgl3 tgl4 mutants) were grown to stationary phase and synchronized by RediGrad gradient centrifugation or α factor arrest. For lipolysis analysis, cells were incubated in fresh medium with 40 μg/mL cerulenin to inhibit de novo fatty acid synthesis. TG content was analyzed by lipid extraction and TLC. For cell cycle analysis, bud formation was assessed by microscopy, and DNA content by FACS. Tgl4 phosphorylation was assessed by immunoblotting on 6% SDS-PAGE gels after protein precipitation. [3] |
| Animal Protocol |
Rat Diabetes Study: Adult male Sprague-Dawley rats (250-300 g) were injected intracardially with 100 mg/kg streptozotocin. For chronic insulin treatment, rats received four subcutaneous injections of 10 U/kg protamine zinc insulin at 16-hour intervals and were killed 16-18 hours after the final injection. For acute insulin treatment, regular insulin (35 U/kg) was injected intraperitoneally 90 minutes before sacrifice. For glucagon treatment, 1 mg/kg was injected intracardially 30 minutes before sacrifice. Livers were removed, homogenized, and assayed for alkaline TG lipase activity. Plasma glucose, fatty acids, and triglycerides were measured. [2]
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| References |
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| Additional Infomation |
ATGL and HSL are the major TG lipases in murine white adipose tissue, together accounting for >95% of TG hydrolase activity. ATGL preferentially performs the initial hydrolysis of TG to generate diacylglycerol (DG), while HSL efficiently degrades DG to monoacylglycerol (MG) and FFA. [1]
In rat liver, alkaline TG lipase activity is decreased in diabetes and starvation, and increased by chronic insulin treatment, but is not acutely responsive to insulin or glucagon. The activity appears to be reciprocally related to plasma free fatty acid concentrations. [2] In yeast, Tgl4 lipase activity is regulated by Cdk1/Cdc28-dependent phosphorylation during the cell cycle, providing fatty acids and lipid precursors for membrane synthesis during bud formation. This represents a direct link between cell-cycle regulatory kinases and TG degradation. [3] |
| CAS # |
9001-62-1
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|---|---|
| Appearance |
White to off-white solid powder
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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) |
H2O : ~3.33 mg/mL
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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.) |
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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