Acetyl-CoA targets a wide range of enzymes and transporters that utilize it as a substrate or ligand. Its targets include: (1) Acetyl-CoA Carboxylase (ACC): Acetyl-CoA serves as the substrate for this enzyme to synthesize malonyl-CoA ; (2) Citrate Synthase: In the TCA cycle, acetyl-CoA condenses with oxaloacetate catalyzed by this enzyme to form citrate ; (3) SLC33A1 (AT-1): This is the only identified acetyl-CoA transmembrane transporter, located on the ER membrane, responsible for translocating acetyl-CoA from the cytosol into the ER lumen ; (4) NLRX1: A mitophagy receptor. Cytosolic acetyl-CoA acts as a signaling metabolite that directly binds to NLRX1 to regulate nutrient starvation-induced mitophagy. In vitro binding assays showed that unlabeled acetyl-CoA competes with biotin-acetyl-CoA for binding to recombinant NLRX1 protein with a saturation concentration of approximately 25 μM . ; Endogenous Metabolite

In famine-starved U2OS cells, acetyl coenzyme A trisodium promotes cytoplasmic protein acetylation while decreasing starvation-induced autophagic fluxes. (U2OS cells that express GFP-LC3 steadily are microinjected with Acetyl Coenzyme A Trisodium; they are then cultured in the absence of nutrients with 100 nM BafA1 and fixed after three hours)[2].

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- In starved U2OS cells, microinjection of acetyl-CoA reduced starvation-induced autophagic flux while increasing cytoplasmic protein acetylation .
- In U2OS cells stably expressing GFP-LC3, treatment with mild starvation medium (containing 5 mM glucose and 2 mM glutamine) reduced cytosolic acetyl-CoA levels, significantly decreased the levels of mitochondrial proteins TIM23, MT-CO2, and HSP60, and increased the recruitment of LC3 to mitochondria, indicating that the reduction in mitochondrial mass resulted from autophagic degradation .
- In KRAS-mutant cancer cells (e.g., KPC and AsPC-1 cells), treatment with KRAS inhibitors (MRTX1133 and RMC-6236) significantly reduced ACLY expression and cytosolic acetyl-CoA levels and induced mitophagy, an effect that was blocked by acetate supplementation .

- In vivo, acetyl-CoA protects mice from cardiomyopathy induced by pressure overload .
- In mouse studies, one day of fasting led to significantly lower total acetyl-CoA levels in organs including the heart and muscle, which correlated with lower protein acetylation levels; however, under the same experimental conditions, liver protein acetylation and acetyl-CoA levels increased, with no apparent effect on brain acetyl-CoA concentration .
- In a study on KRAS inhibitor resistance mechanisms, using an in vivo xenograft tumor model, it was found that NLRX1 knockout significantly enhanced the anti-tumor effect of the KRAS inhibitor MRTX1133 on KPC tumors (manifested as reduced tumor volume and weight), while reactive oxygen species levels increased and mitophagy was impaired in the tumor tissue, indicating that KRAS inhibitors induce NLRX1-dependent mitophagy to help tumor cells resist oxidative stress and develop drug resistance .

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In a mouse cardiac pressure overload model, acetyl coenzyme A trisodium attenuates pressure overload-induced cardiomyopathy via inhibiting maladaptive autophagy[2][3]. Mice denied food (but allowed unlimited access to water) for a whole day have markedly lower levels of total Acetyl coenzyme A trisodium in the heart and muscles, which correlates with lower levels of protein acetylation. Nevertheless, the identical experimental setups actually raise hepatic levels of protein acetylation and Acetyl coenzyme A trisodium while having no discernible impact on brain concentrations of the enzyme[4].

- Acetyl-CoA Carboxylase (ACC) Activity Assay (NADH Rate Method): This principle is based on ACC catalyzing the conversion of acetyl-CoA, NaHCO3, and ATP to malonyl-CoA, ADP, and inorganic phosphate. The generated ADP is converted to ATP and pyruvate from phosphoenolpyruvate via pyruvate kinase, and pyruvate is further converted to lactate with the concomitant oxidation of NADH to NAD+. Since NADH has a characteristic absorption peak at 340 nm, ACC activity is calculated by monitoring the decrease in absorbance at 340 nm .
- Acetyl-CoA Carboxylase Activity Assay (Non-radioactive Spectrophotometric Method): Permeabilized Corynebacterium glutamicum cells are added to an assay mixture containing acetyl-CoA, bicarbonate, magnesium, and ATP. Aliquots are removed at set time points, and the reaction is stopped by adding trifluoroacetic acid. The level of acetyl-CoA remaining in each aliquot is determined via a citrate synthase assay, in which the formation of the yellow compound dithiobisnitrobenzoic acid-thiophenolate is followed spectrophotometrically at 412 nm .
- NLRX1 and Acetyl-CoA Direct Binding Assay (Pull-down Assay): Recombinant NLRX1 protein with an N-terminal MBP tag is purified using an insect cell expression system. In the in vitro pull-down assay, the recombinant MBP-NLRX1 protein is incubated with biotin-labeled acetyl-CoA, and the complex is captured using streptavidin magnetic beads. After elution, MBP-NLRX1 is detected by Western blot. Unlabeled acetyl-CoA competes with biotin-acetyl-CoA for binding to recombinant NLRX1 protein in a dose-dependent manner, with a saturation concentration of approximately 25 μM for competition .

- Cell Culture and Starvation Treatment: U2OS cells (stably expressing GFP-LC3) are cultured in regular medium. To induce autophagy, the medium is replaced with mild starvation medium (containing 5 mM glucose and 2 mM glutamine) and incubated for a specified time (e.g., 3 hours). To block autophagic flux, BafA1 (100 nM) can be added before treatment. After treatment, cells are washed with PBS, lysed, and mitochondrial protein levels (e.g., TIM23, MT-CO2, HSP60) are detected by Western blot, or GFP-LC3 puncta formation is observed by fluorescence microscopy .
- Acetyl-CoA Microinjection: In starved U2OS cells, acetyl-CoA is directly introduced into the cytoplasm via microinjection. After injection, cells are treated with BafA1 (100 nM) under nutrient-free conditions for 3 hours and then fixed. Autophagic flux is assessed by detecting GFP-LC3 puncta, while cytoplasmic protein acetylation levels are also measured .
- KRAS Inhibitor Treatment: KRAS-mutant cancer cells (e.g., KPC or AsPC-1 cells) are seeded in culture plates. Cells are treated with KRAS inhibitors MRTX1133 or RMC-6236. Control groups (DMSO) and acetate supplementation groups (to verify the specific effect of acetyl-CoA) are also set up. After treatment, cells are harvested, and ACLY expression levels, mitochondrial protein levels, and acetyl-CoA levels are detected by Western blot .

- Cardiomyopathy Model: A mouse model of cardiomyopathy is induced by pressure overload. Acetyl-CoA is administered to evaluate its preventive effect on cardiomyopathy .
- Fasting Experiment: Mice are divided into an ad libitum feeding group and a fasting group (fasted for 24 hours with free access to water). After fasting, mice are sacrificed, and heart, muscle, liver, and brain tissues are collected. Total acetyl-CoA levels in each tissue are measured by LC-MS, and protein acetylation levels are detected by Western blot .
- Tumor Xenograft Model: KPC cells (KRAS-mutant pancreatic cancer cells) are subcutaneously inoculated into immunodeficient mice to establish a xenograft tumor model. After tumors reach a certain volume, mice are randomly divided into a control group and a treatment group (e.g., KRAS inhibitor MRTX1133). In some experiments, NLRX1-knockout KPC cells are used. KRAS inhibitors are administered by oral gavage or intraperitoneal injection. Tumor volume and body weight are measured regularly during the administration period. At the end of the experiment, mice are sacrificed, tumor tissues are excised and weighed, and reactive oxygen species levels and mitophagy-related indicators are measured in the tumor tissue .

Traditional pharmacokinetic parameters for acetyl-CoA itself are not described in the literature. Acetyl-CoA is a large, charged molecule (MW ~800) that is unable to freely diffuse across cellular membranes and requires specific transporter proteins (such as SLC33A1) to facilitate its transmembrane movement . Under laboratory conditions, aqueous solutions of acetyl-CoA can be stored at -20°C for up to 2 weeks .

Direct toxicity information for acetyl-CoA itself is not described in the literature. As an endogenous metabolite, it does not exhibit toxicity at physiological concentrations. According to supplier information, Acetyl Coenzyme A Trilithium Salt is not regulated for transport and is considered non-hazardous . It is important to note that disruption of acetyl-CoA homeostasis is associated with various diseases, including neurodegenerative disorders, aging-related conditions, and cancer . For example, either insufficient or excessive activity of the SLC33A1 transporter can lead to diseases, especially those affecting the nervous system .

[1]. The growing landscape of lysine acetylation links metabolism and cell signalling. Nat Rev Mol Cell Biol. 2014 Aug;15(8):536-50.

[2]. Regulation of autophagy by cytosolic acetyl-coenzyme A. Mol Cell. 2014 Mar 6;53(5):710-25.

[3]. Cardiac autophagy is a maladaptive response to hemodynamic stress. J Clin Invest. 2007 Jul;117(7):1782-93.

[4]. Acetyl coenzyme A: a central metabolite and second messenger. Cell Metab. 2015 Jun 2;21(6):805-21.

Acetyl-CoA is involved in the biosynthesis of fatty acids and sterols, fatty acid oxidation, and the metabolism of various amino acids. It also functions as a biological acetylation agent. Lysine acetylation is a conserved post-translational modification of proteins that links acetyl-CoA metabolism to cellular signaling. In recent years, mass spectrometry has made significant progress in the identification and quantification of lysine acetylation, deepening our understanding of its role in various biological processes by regulating protein-protein interactions, activity, and localization. Furthermore, proteins frequently undergo other types of acylation modifications, such as formylation, butyrylation, propionylation, succinylation, malonylation, myristylation, glutarylation, and crotonylation. The intricate link between lysine acylation and cellular metabolism has been elucidated through various metabolite-sensitive acylation reactions and their selective removal by sirtuin deacylases. These new findings reveal novel functions of different lysine acylations and deacylases, while also highlighting the mechanisms by which acetylation regulates various cellular processes. [1] Acetyl-CoA (AcCoA) is an important integrator of nutritional status at the intersection of fat, sugar and protein catabolism. This study shows that nutritional deprivation leads to rapid consumption of AcCoA. Consumption of AcCoA results in a corresponding decrease in the overall acetylation level of cytoplasmic proteins and induces autophagy—a self-digestive process that maintains cellular homeostasis. Various manipulations aimed at increasing or decreasing cytoplasmic acetyl-CoA (AcCoA) levels resulted in inhibition or induction of autophagy in cultured human cells and mice, respectively. Furthermore, in a cardiac stress overload model, maintaining high levels of AcCoA inhibited maladaptive autophagy. AcCoA depletion reduced the activity of acetyltransferase EP300, which is essential for the inhibition of autophagy by high levels of AcCoA. In summary, our results demonstrate that cytoplasmic AcCoA plays a role as a core metabolic regulator of autophagy, thus elucidating an AcCoA-centered pharmacological strategy for the therapeutic regulation of autophagy. [2] Cardiac hypertrophy is a major predictor of heart failure and a common disease with high mortality. However, little is known about the mechanisms of transition from stable cardiac hypertrophy to decompensated heart failure. This study investigated the role of autophagy, a conserved pathway that mediates the massive degradation of long-lived proteins and organelles, ultimately leading to cell death. To quantify autophagy activity, we constructed an "autophagy reporter gene" mouse strain and demonstrated that short-term nutrient deprivation can induce cardiomyocyte autophagy in vivo. Pressure load induced by aortic constriction can lead to heart failure and significantly enhance cardiac autophagy. Load-induced autophagy activity peaked at 48 hours and remained significantly elevated for at least 3 weeks. Furthermore, autophagy activity was not spatially uniformly distributed but was particularly pronounced in the basal septa. Heterozygous loss of the gene encoding Beclin 1 (a protein essential for early autophagosome formation) reduced cardiomyocyte autophagy and attenuated pathological remodeling induced by severe stress. Conversely, overexpression of Beclin 1 enhanced autophagy activity and exacerbated pathological remodeling. In summary, these results suggest that autophagy is involved in the pathogenesis of load-induced heart failure and hints at its potential as a target for novel therapeutic interventions. [3] Acetyl-CoA (acetyl-CoA) is an important metabolic intermediate. The abundance of acetyl-CoA in different subcellular compartments reflects the overall energy state of the cell. In addition, the concentration of acetyl-CoA affects the activity or specificity of a variety of enzymes by allosteric regulation or altering substrate availability. Finally, acetyl-CoA controls key cellular processes, including energy metabolism, mitosis, and autophagy, by influencing the acetylation profiles of a variety of proteins, including histones, directly or through epigenetic regulation of gene expression. Thus, acetyl-CoA determines the balance between catabolism and anabolism by acting as both a metabolic intermediate and a second messenger. [4]

C23H35N7NA3O17P3S

875.52

809.125

102029-73-2

Acetyl coenzyme A lithium;32140-51-5;Acetyl coenzyme A;72-89-9

6302

White to off-white solid powder

1.9±0.1 g/cm3

1.718

-3.89

9

22

20

51

1380

4

O[C@@H]1[C@@H]([C@@H](COP(O)(=O)OP(O)(=O)OCC(C)(C)[C@@H](O)C(=O)NCCC(=O)NCCSC(=O)C)O[C@H]1N1C=NC2C(=NC=NC1=2)N)OP(O)(O)=O.[Na+].[Na+].[Na+]

ZSLZBFCDCINBPY-KMYLAXNMSA-N

InChI=1S/C23H38N7O17P3S/c1-12(31)51-7-6-25-14(32)4-5-26-21(35)18(34)23(2,3)9-44-50(41,42)47-49(39,40)43-8-13-17(46-48(36,37)38)16(33)22(45-13)30-11-29-15-19(24)27-10-28-20(15)30/h10-11,13,16-18,22,33-34H,4-9H2,1-3H3,(H,25,32)(H,26,35)(H,39,40)(H,41,42)(H2,24,27,28)(H2,36,37,38)/t13-,16-,17-,18?,22-/m1/s1

S-[2-[3-[[4-[[[(2R,3S,4R,5R)-5-(6-aminopurin-9-yl)-4-hydroxy-3-phosphonooxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-hydroxyphosphoryl]oxy-2-hydroxy-3,3-dimethylbutanoyl]amino]propanoylamino]ethyl] ethanethioate

Acetyl coenzyme A sodium salt; 102029-73-2; Coenzyme A, S-acetate, trisodium salt (9CI); Acetyl Coenzyme A Trisodium Salt; Acetyl-CoA sodium; XGCDESRMLMCTPI-UHFFFAOYSA-N;

2934.99.9001

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.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

H2O : 83.33 mg/mL (95.18 mM)
DMSO : < 1 mg/mL

Solubility in Formulation 1: 100 mg/mL (114.22 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.

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 (Please use freshly prepared in vivo formulations for optimal results.)