Standard to Meet
| Compound Assessment
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Human Toxicity
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Compound causes cholestasis, lipid-accumulating disorders (steatosis/phopholipidosis) or cell death in humans.
| Salicylates are weak acids that may produce metabolic acidosis through various mechanisms. In toxic concentrations, salicylates interfere with energy production by uncoupling oxidative phosphorylation and may produce renal insufficiency that causes accumulation of phosphoric and sulfuric acids. The metabolism of fatty acids is likewise increased in patients with salicylate toxicity, generating ketone body formation. These processes all contribute to the development of an elevated anion gap metabolic acidosis in patients with salicylate poisoning. Ref. Medscape Article 1009987
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Toxicity is concentration dependent (non-idiosyncratic).[1]
| Matindale indicates that hepatoxicity appears to be correlated with serum salicylate levels greater than 150mgs/ml. Although generally manifests as mild to moderate elevation in Aminotransferase activity.
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Therapeutic target.
| COX inhibitor
The analgesic, antipyretic, and anti-inflammatory effects of acetylsalicylic acid are due to actions by both the acetyl and the salicylate portions of the intact molecule as well as by the active salicylate metabolite. Acetylsalicylic acid directly and irreversibly inhibits the activity of both types of cyclooxygenase (COX-1 and COX-2) to decrease the formation of precursors of prostaglandins and thromboxanes from arachidonic acid.
The platelet aggregation-inhibiting effect of acetylsalicylic acid specifically involves the compound's ability to act as an acetyl donor to cyclooxygenase; the nonacetylated salicylates have no clinically significant effect on platelet aggregation. Irreversible acetylation renders cyclooxygenase inactive, thereby preventing the formation of the aggregating agent thromboxane A2 in platelets (drugbank)
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Biochemical mechanism of toxicity.
| The mechanism of salicylic acid-induced liver injury has not been sufficiently clarified. It is known that salicylic acid acts as an uncoupler against mitochondria. Mitochondrial dysfunction is speculated to be closely related to Reye's syndrome, which is an acute encephalopathy and is characterized by liver damage. Results suggest that salicylic acid-induced lipid peroxidation was related to oxidative metabolism mediated by CYP2E1 and CYP2C11. On the other hand, 5 mM salicylic acid induced a drastic decrease of ATP contents in rat isolated hepatocytes. Furthermore, mitochondrial respiration control ratio (RC ratio), calculated by State 3/State 4 also decreased with the increase of salicylic acid concentration. These findings suggest that salicylic acid would trigger mitochondrial dysfunction and cause ATP decrease, leading to lethal liver cell injury by lipid peroxidation, although this hypothesis remains to be elucidated in vivo.
Doi et al.(2010) Chemico-Biological Interactions Volume 183, Issue 3, 12 February 2010, Pages 363-368
Oral, mouse: LD50 = 250 mg/kg;
Oral, rabbit: LD50 = 1010 mg/kg;
Oral, rat: LD50 = 200 mg/kg
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PK-ADME
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PK parameters: recommended dose, Cmax, Vd, and half-life.[2]
| The plasma half-life is approximately 15 minutes; that for salicylate lengthens as the dose increases: doses of 300 to 650 mg have a half-life of 3.1 to 3.2 hours; with doses of 1 gram, the half-life is increased to 5 hours and with 2 grams it is increased to about 9 hours.
Table from Zeitoun et al The JOURNAL of APPLIED RESEARCH In Clinical and Experimental Therapeutics based on single dose 100mg oral tablet.
| AUC0-t (µg/mL)
| Cmax (µg/mL)
| Tmax (h)
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Test Formulation
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Mean ± SD
| 66.2 ± 42.9
| 23.6 ± 16.3
| 4.9 ± 2.4
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Minimum
| 21.6
| 8.1
| 1
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Maximum
| 176.5
| 59.7
| 10
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95% Cl
| 24.3
| 9.19
| 1.33
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Reference Formulation
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Mean ± SD
| 64.8 ± 38.0
| 21.7 ± 11.3
| 4.9 ± 1.5
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Minimum
| 24.6
| 9.8
| 2
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Maximum
| 160.2
| 44.3
| 8
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95% Cl
| 21.5
| 6.41
| 0.85
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Therapeutic window.[3]
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Metabolically activated (optional), active metabolite known and available for testing.[3]
| High binding (99.5%) to albumin. Decreases as plasma salicylate concentration increases, with reduced plasma albumin concentration or renal dysfunction, and during pregnancy.
Acetylsalicylic acid is rapidly hydrolyzed primarily in the liver to salicylic acid, which is conjugated with glycine (forming salicyluric acid) and glucuronic acid and excreted largely in the urine
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Omics and IC50 Data
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Gene expression profiles known.[4]
| GDS332 record:
Colon carcinoma response to butyrate and aspirin [Rattus norvegicus]
Summary: Temporal analysis of colon carcinoma cell line CC531 response to 4.5 mM butyrate or 3 mM aspirin. Samples taken at 2, 6, 12, 16 and 24 hours.
Toxicology (2008) 250, Issue: 1, Pages: 15-26
Uehara et al. A toxicogenomics approach for early assessment of potential non-genotoxic hepatocarcinogenicity of chemicals in ratsGene expression profiling in rat liver treated with compounds inducing elevation of bilirubin
E-GEOD-5258
Transcription profiling of human cell lines freely cycling treated with 164 distinct small molecules - Broad Connectivity Map CMAP Set 1
E-GEOD-424
Transcription profiling time series of the response of Rattus norvegicus colon carcinoma cell line CC531 to treatment with butyrate or aspirin
E-GEOD-16394
Rat liver. Control vs. Chemical treated, 28 days
E-GEOD-8251
Non-genotoxic Hepatocarcinogens Iconix dataset
A gene expression biomarker provides early prediction and mechanistic assessment of hepatic tumor induction by nongenotoxic chemicals. Fielden MR, Brennan R, Gollub J. , PubMed 17557906
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Proteomics profiles known.[5]
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Metabonomics profiles known.[5]
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Fluxomics profiles known.[5]
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Epigenomics profiles known.[5]
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Observed IC50 for in vitro cellular efficacy.[5]
| PGE2 synthesis was investigated in primary rat microglial cells IC50 = 1.85uM (Fiebich et al.) J.of Neurochemistry Volume 86, Issue 1, pages 173–178, July 2003.
Human chondriocyte cells COX-1 (IC50 = 3.57 uM) COX-2 (IC50 = 29.3 uM)
Blanco et al. J Rheumatol. 1999 Jun;26(6):1366-73.
Ovine test kit pirin (COX-1 IC50 = 0.35 μM; COX-2 IC50 = 2.4 μM)
Bioorganic & Medicinal Chemistry Volume 13, Issue 15, 1 August 2005, Pages 4694-4703
Cox-2 IC50
Aspirin A549 cells, A23187 gastric cell line 2.9 (1.4–5.6)
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Observed IC50 for in vitro cellular toxicity studies.
| Hela cell line
P E > 15 mM n = 36
BM 7.8 mM n = 45
NRU 8.3 mM n = 49
Neoplasma. 2000;47(3):172-6. Comparison of three in vitro assays at evaluation of IC50 of acetylsalicylic acid, ferrous sulfate, amitriptyline, methanol, isopropanol and ethylene glycol in human cancer cells HeLa.
cell viability
IC50 rat hepatocyte 3955 uM, 3T3 cells 2386 uM, Hela 8523 uM, HepG2 7454 uM
Wang et al. J of Toxicological Sciences Vol. 27 (2002) , No. 3 August 229-237
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Physical Properties
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Accepted and listed within the ToxCast/Tox21 program.[6]
| Included in phase II ToxCast List U.S EPA/ORD/NCCT. ToxCast Phase I and II Chemicals. December 14, 2010.
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Defined and confirmed structure and isomeric form(s).
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Substance stability.
| Half life in distilled water, unbuffered (22.5°C): 153.30 ± 3.70
Bakar SK and Niazi S. Stability of aspirin in different media. J Pharm Sci 1981; 72(9): 1024-1027
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Soluble in buffer solution at 30 times the in vitro IC50 for toxicity.[7]
| Water solubility: 4.6 mg/ml (25°C)
YALKOWSKY,SH & DANNENFELSER,RM (1992) from SRC PhysProp Database
Water solubility: 3 mg/ml (25°C) Sigma A5376 Product details.
estimated intrinsic solubility : 2.03 mg/ml
estimated solubility in pure water at pH 2.71: 2.38 mg/ml
estimated solubility in water at pH 7.4: 1000 mg/ml
(Calculations performed using ACD/PhysChem v 9.14)
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Solubility in DMSO 100 times buffer solubility.
| 41 mg/ml Cayman Chemical Aspirin Product Information
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Vessel binding properties.[8]
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Commercial availability at > 95% (> 99% is preferred).
| Sigma Aldrich (A5376) 100g/17.50€ purity >99% Sigma A5376 Product details
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Vapor pressure. (Non-volatile)
| Estimated vapor pressure: 6 .56E-05 mmHg
(Calculation performed using EPI Suite v4.10)
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Criteria Notes
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1. | The in vivo therapeutic window is used to estimate an appropriate concentration for in vitro toxicity assays. This in vitro concentration should also be consistent with the exposure implied by pharmacokinetics parameters. |
2. | We prefer compounds that require metabolic activation, although standards that are active in themselves will be accepted if they have otherwise valuable properties. We require knowing the active metabolite, and we prefer compounds where the metabolite is stable and can be independently tested in order to verify the mechanism of toxicity as well as of metabolic activation in the test cell line. |
3. | Literature data for at least one, but not necessarily all, of the ‘omics datasets is desired. This requirement can be waived in special cases. |
4. | The IC50’s for in vitro efficacy and toxicity should be consistent with the therapeutic ratio observed in the clinic. These parameters will be dependent on specific cell type and culture conditions, but differences of more than 30-fold in the in vitro vs. in vivo therapeutic ratios should be considered suspect and carefully justified. |
5. | This is not a requirement, but compounds utilized in the EPA testing program can be assumed to have physical properties verified to be suitable for in vitro cellular assays. |
6. | Sparing soluble compounds may be assayed for solubility in serum and the percent serum used specified here. |
7. | This property will be measured when a sample of compound becomes available. |
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