Aflatoxin B1

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Aflatoxin B1
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Executive Summary

Compound Aflatoxin B1
Toxicities Apoptosis
Mechanisms Aflatoxin is metabolized to a reactive epoxide that forms adducts with DNA and protein lysine groups. The epoxide is hydrolyzed to a dialdehyde, which also forms adducts with protein lysine groups. Because mitochondrial energy production is not significantly disrupted by this toxin, ATP levels are maintained and cellular damage triggers apoptosis rather than necrosis. It is not clear, however, which specific biological adduct(s) trigger apoptosis.
Comments This compound was selected as standard to exemplify the chemistry of reactive epoxides and lysine reagents. Carcinogenic effects that are observed on long-term, low-dose exposure are of less interest in these studies.

CAUTION: Aflatoxin is extremely toxic. Long-term exposure to sub-microgram levels can cause cancer. Handle the solid compound with extreme care, and handle the compound in solution when possible. Not recommended for high-throughput screening assays.

Feedback Contact Gold Compound Working Group (GCWG)
Aflatoxin B1
Aflatoxin B1.png


Identifiers
Leadscope Id LS-584
CAS 1162-65-8
ChemSpider 162470
ChEBI 2504
Pathway DBs
KEGG C06800
Assay DBs
PubChem CID 186907
Omics DBs
Properties
ToxCast Accepted no
Toxic Effect Apoptosis
ToxBank Accepted yes
Approved on 2011-06-28
Target possibly DNA binding
Toxicities Apoptosis


In Vivo Data ? Compound Assessment
Adverse Events ? The primary target of aflatoxins is the hepatic system. Acute effects include hemorrhagic necrosis of the liver and bile duct proliferation while chronic effects include hepatocellular carcinoma. Acute exposure can result in aflatoxicosis, which manifests as severe, acute hepatotoxicity with a case fatality rate of approximately 25%.Hepatocellular carcinoma as a result of chronic exposure has been well documented, generally in association with hepatitis B virus or other risk factors.

References:

-Public Health Strategies for Preventing Aflatoxin Exposure workgroup report for the International Mycotoxin Workshop, sponsored by WHO and CDC, held in Geneva in July 2005.
-I.Dvorackova 1990 Aflatoxins and human health CRC Press 1990
-Mohamed IF Shariff et al. Current Trends in Hepatocellular Carcinoma: Aflatoxin B1, Expert Rev Gastroenterol Hepatol. 2009;3(4):353-367.
-Current Trends in Hepatocellular Carcinoma: Aflatoxin B1
-C.P.Wild et al. The toxicology of aflatoxins as a basis for public health decisions, Mutagenesis vol.17 no.6 pp.471-481, 2002
-Strosnider H et al. 2006 Workgroup Report: Public Health Strategies for Reducing Aflatoxin Exposure in Developing Countries. Environ Health Perspect 114(12): 1898-1903

Toxicity is dose dependent.

References:

-Sara Hale Henry et al. "Reducing Liver Cancer-- Global Control of Aflatoxin", Science 1999: Vol. 286 no. 5449 pp. 2453-2454
Toxicity Mechanisms ? AFB1 is activated by cytochromes P450 CYP3A4 and 1A2 to AFB1-8,9-exoepoxide and AFB1-8,9-endo-epoxide, both of which can covalently bind to nucleic acids or proteins, provoking cell membrane damage, necrosis, and mutagenesis in the affected cells. The exo-epoxide binds to DNA to form the predominant 8,9-dihydro-8-(N7-guanyl)-9-hydroxy-AFB1 (AFB1–N7-Gua) adduct that confers the mutagenic properties of the compound.

References:

-Michelle McLean and Michael f. Dutton, "Cellular Interactions and Metabolism of Aflatoxin: An Update", Pharmac. Ther. 65: 163-192 (1995).
-Bedard, L. L. and Massey, T. E. (2006) "Aflatoxin B1-induced DNA damage and its repair", Cancer Lett., 241, 174–183.

Adducts are formed with mitochondrial DNA with 3-4-fold preference over nuclear DNA. Because repair of mDNA is also slower, there is a preferential reduction in mitochondrial protein synthesis compared to cytosolic synthesis.

References:

-Niranjan BG, Bhat NK, Avadhani NG (1982) “Preferential attack of mitochondrial DNA by aflatoxin B1 during hepatocarcinogenesis”, Science, 215: 73-75.

AFB1 induces cell death via apoptosis in HepG2 cells, without depletion of ATP.

-Christopher McKean, Lili Tang, Madhavi Billam, Meng Tang, Christopher W. Theodorakis, Ronald J. Kendall and Jia-Sheng Wang, “Comparative acute and combinative toxicity of aflatoxin B1 and T-2 toxin in animals and immortalized human cell lines”, J. Appl. Toxicol. 2006; 26: 139–147.
-Travis O’Brien, George Babcock, James Cornelius, Mike Dingeldein, Glenn Talaska, David Warshawsky, and Kent Mitchell, “A Comparison of Apoptosis and Necrosis Induced by Hepatotoxins in HepG2 Cells”,Toxicology and Applied Pharmacology 164, 280–290 (2000).

AFB1-8,9-epoxides are conjugated to GSH. This appears to be primarily a detoxification mechanism, and depletion of GSH is not required for toxicity.

-Kevin D. Raney, David J. Meyer, Brian Ketterer, Thomas M. Harris, and F. Peter Guengerich, “Glutathione Conjugation of Aflatoxin B1 exo- and endo-Epoxides by Rat and Human Glutathione S-Transferases”, Chem. Res. Toxicol. 1992,5, 470-478.

The exo-AFB1-8,9-epoxide is the DNA-alkylating metabolite. Both endo- and exo-epoxides have a very short half-life of 1 s and are hydrolyzed to the dialdehyde. The dialdehyde is the species that modifies proteins and reacts with lysine groups. This chemistry gives aflatoxin B1 a different reactivity profile than the common quinone imine metabolites for inducing toxicity.

-Guengerich FP, Arneson KO, Williams KM, Deng Z, and Harris TM, “Reaction of aflatoxin B(1) oxidation products with lysine”, Chem Res Toxicol (2002) 15:780-792.

AFB1 also induces formation of reactive oxygen species, lipid peroxidation and formation of 8-hydroxydeoxyguanosine in vivo and in vitro. The ability of AFB1 to induce oxidative damage to cells and DNA may add to its toxicity. The quantitative importance of these reactions relative to protein and DNA adduct formation is not clear.

-R. J. Verma, "Aflatoxin Causes DNA Damage" , Int J Hum Genet, 4(4): 231-236 (2004).
-R. Liu et al. "In vitro toxicity of aflatoxin B1 and its photodegradation products in HepG2 cells", J. Appl. Toxicol. 2011, DOI 10.1002/jat.1669.
-Shen H.M et al. "Detection of elevated reactive oxygen species level in cultures rat hepatocytes treated with aflatoxin B1". Free Rad. Biol. Med. 1996, 21, 139–146.
Therapeutic target ?

PubMed references

The following resource link will perform a PubMed query for the terms "Aflatoxin B1" and "liver toxicity".
Aflatoxin B1 Search

EPA Summary

EPA summary

The table listed below contains a summarized listing of toxic effect information leveraged from the 6th European Framework Programme project LIINTOP. For a complete listing of the Gold Compound evaluation criteria please see the Gold Compound Evaluation and Comments immediately following the summary table below.

SMILES COC1=C2C3=C(C(=O)CC3)C(=O)OC2=C4C5C=COC5OC4=C1
InChI

InChI=1S/C17H12O6/c1-20-10-6-11-14(8-4-5-21-17(8)22-11)15-13(10)7-2-3-9(18)12(7)16(19)23-15/h4-6,8,17H,2-3H2,1H3

InChI-Key

OQIQSTLJSLGHID-UHFFFAOYSA-N

Summary Hepatotoxic Effects from the LIINTOP FP6 Program
Hepatocellular necrosis.gif Apoptosis.gif Transporter inhibition.gif Cholestatic.gif Steatotic.gif Phospholipidosis.gif Hepatocyte function.gif Mithochondria impairment.gif Oxidative stress.gif DNA synthesis genotoxicity.gif Covalent binding.gif Idiosyncrasia metabolic.gif Idiosyncrasia immune.gif Bioactivation required.gif LIINTOP severity.gif References
+ + + + +

[1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15] [16]

References

  1. Li, A.P., 2002. A review of the common properties of drugs with idiosyncratic hepatotoxicity and the ‘‘multiple determinant hypothesis” for the manifestation of idiosyncratic drug toxicity. Chem. Biol. Interact. 142, 7–23. doi:10.1016/S0009-2797(02)00051-0
  2. Feldmann, G., 2006. Liver apoptosis. Gastroenterol. Clin. Biol. 30, 533–545.
  3. Gomez-Lechon, M.J., O’Connor, E., Castell, J.V., Jover, R., 2002. Sensitive markers used to identify compounds that trigger apoptosis in cultured hepatocytes. Toxicol. Sci. 65, 299–308. doi:10.1093/toxsci/65.2.299
  4. Gomez-Lechon, M.J., O’Connor, J.E., Lahoz, A., Castell, J.V., Donato, M.T., 2008. Identification of apoptotic drugs: multiparametric evaluation in cultured hepatocytes. Curr. Med. Chem. 15, 2071–2085. pmid:18691057
  5. Gomez-Lechon, M.J., Ponsoda, X., O’Connor, E., Donato, T., Jover, R., Castell, J.V., 2003. Diclofenac induces apoptosis in hepatocytes. Toxicol. in Vitro 17, 675– 680. doi:10.1016/S0887-2333(03)00105-X
  6. Kass, G.E., Macanas-Pirard, P., Lee, P.C., Hinton, R.H., 2003. The role of apoptosis in acetaminophen-induced injury. Ann. NY Acad. Sci. 1010, 557–559. doi:10.1196/annals.1299.103
  7. Ioannides, C., Lewis, D.F., 2004. Cytochromes P450 in the bioactivation of chemicals. Curr. Top. Med. Chem. 4, 1767–1788. doi:10.2174/1568026043387188
  8. Kang, P., Dalvie, D., Smith, E., Zhou, S., Deese, A., Nieman, J.A., 2008. Bioactivation of flutamide metabolites by human liver microsomes. Drug Metab. Dispos. 36, 1425–1437. doi:10.1124/dmd.108.020370
  9. Li, A.P., 2002. A review of the common properties of drugs with idiosyncratic hepatotoxicity and the ‘‘multiple determinant hypothesis” for the manifestation of idiosyncratic drug toxicity. Chem. Biol. Interact. 142, 7–23.doi:10.1016/S0009-2797(02)00051-0
  10. Park, K., Williams, D.P., Naisbitt, D.J., Kitteringham, N.R., Pirmohamed, M., 2005b. Investigation of toxic metabolites during drug development. Toxicol. Appl. Pharmacol. 207, 425–434. {{doi|10.1016/j.taap.2005.02.029
  11. Reddy, M.V., Storer, R.D., Laws, G.M., Armstrong, M.J., Barnum, J.E., Gara, J.P., McKnight, C.G., Skopek, T.R., Sina, J.F., DeLuca, J.G., Galloway, S.M., 2002. Genotoxicity of naturally occurring indole compounds: correlation between covalent DNA binding and other genotoxicity tests. Environ. Mol. Mutagen. 40, 1–17. doi:10.1002/em.10088
  12. Hynes, J., Marroquin, L.D., Ogurtsov, V.I., Christiansen, K.N., Stevens, G.J., Papkovsky, D.B., Will, Y., 2006. Investigation of drug-induced mitochondrial toxicity using fluorescence-based oxygen-sensitive probes. Toxicol. Sci. 92, 186–200. doi:10.1093/toxsci/kfj208
  13. Johannsen, D.L., Ravussin, E., 2009. The role of mitochondria in health and disease. Curr. Opin. Pharmacol. 9, 780–786. doi:10.1016/j.coph.2009.09.002
  14. Jones, D.P., Lemasters, J.J., Han, D., Boelsterli, U.A., Kaplowitz, N., 2010. Mechanisms of pathogenesis in drug hepatotoxicity putting the stress on mitochondria. Mol. Interv. 10, 98–111. doi:10.1124/mi.10.2.7
  15. Labbe, G., Pessayre, D., Fromenty, B., 2008. Drug-induced liver injury through mitochondrial dysfunction: mechanisms and detection during preclinical safety studies. Fundam. Clin. Pharmacol. 22, 335–353. doi:10.1111/j.1472-8206.2008.00608.x
  16. Masubuchi, Y., 2006. Metabolic and non-metabolic factors determining troglitazone hepatotoxicity: a review. Drug Metab. Pharmacokinet. 21, 347–356. doi:10.2133/dmpk.21.347

PK-ADME ? Compound Assessment
PK parameters ? Pharmacokinetics in humans:
  • Volume of distribution = 1 L/kg
  • Cmax = 0.9 pg/mL for 500 pg/kg dose
  • Half-life = 3 hr (alpha phase, free species), 64 h (beta phase, protein-bound species) (parent + metabolites)

References:

-Carole Jubert, John Mata, Graham Bench, Roderick Dashwood, Cliff Pereira, William Tracewell, Kenneth Turteltaub, David Williams, and George Bailey, “Effects of Chlorophyll and Chlorophyllin on Low-Dose Aflatoxin B1 Pharmacokinetics in Human Volunteers“, Cancer Prev Res 2009;2:1015-1022.

Human skin penetration = 2.1 x 10-4 cm/h. At this penetration rate, it is calculated that farm workers can be exposed to a hepatocarcinogenic dose of AFB1 from contaminated liquids, e.g. fruit juices.

References:

-Boonen J, Malysheva SV, Taevernier L, Di Mavungu JD, De Saeger S, De Spiegeleer B (2012) "Human skin penetration of selected mycotoxins", Toxicology 301, 21-32
Therapeutic window ? Maximum exposure levels for cereals a maximum level of 2 μg/kg (0.007 umol/kg) and for maize and rice a maximum level of 5 μg/kg for aflatoxin B1 has been established.

References:

-COMMISSION REGULATION (EU) No 165/2010 setting maximum levels for certain contaminants in foodstuffs as regards aflatoxins.
Metabolically activated ? AFB1 8, 9-epoxide is generally accepted as the active electrophilic form of AFB1 that may attack nucleophilic nitrogen, oxygen and sulphur heteroatoms in cellular constituents. This highly reactive substance may combine with DNA bases such as guanine to produce alterations in DNA. This may be the most important product from the carcinogenic point of view.

Other major metabolites in the human include AFM1, aflatoxicol, AFLH1, AFP1, AFB2á and AFB1-2, 2-dihydrodiol (detoxification products).

References:

-R. J. Verma, Aflatoxin Cause DNA Damage, Int J Hum Genet, 4(4): 231-236 (2004)
-Massey TE, Stewart RK, Daniels JM, Liu L. 1995. “Biochemical and molecular aspects of mammalian susceptibility to aflatoxin B1 carcinogenicity”. Proc. Soc. Exp. Biol. Med. 208: 213–227.

Omics and IC50 Data ? Compound Assessment
Gene expression profiles known. ? References:
-Jossé R et al. Early target genes of aflatoxin B1 in human hepatic cells, Public on Dec 07, 2010
-C. Magkoufopoulou, S.M.H. Claessen, D.G.J. Jennen, J.C.S. Kleinjans, and J.H.M. van Delft, “Comparison of phenotypic and transcriptomic effects of false-positive genotoxins, true genotoxins and non-genotoxins using HepG2 cells”, Mutagenesis vol. 26 no. 5 pp. 593–604, 2011.
Proteomics profiles known. ? References:
-Yuan Li et al. Proteome analysis of aflatoxin B1-induced hepatocarcinogenesis in tree shrew (Tupaia belangerichinensis) and functional identification of candidate protein peroxiredoxin II, Proteomics 2008, 8, 1490–1501
Metabonomics profiles known. ? References:
-Limin Zhang et al. Systems Responses of Rats to Aflatoxin B1 Exposure Revealed with Metabonomic Changes in Multiple Biological Matrices, J. Proteome Res., 2011, 10 (2), pp 614-623
Fluxomics profiles known. ?
Epigenomics profiles known. ? References:
-Z. Herceg Epigenetics and cancer: towards an evaluation of the impact of environmental and dietary factors, Mutagenesis (2007) 22 (2): 91-103 (page 96)
-Zhang YJ et al. High frequency of promoter hypermethylation of RASSF1A and p16 and its relationship to aflatoxin B1-DNA adduct levels in human hepatocellular carcinoma. Mol. Carcinog. 2002;35:85-92
-Zhang YJ et al. Inactivation of the DNA repair gene O6-methylguanine-DNA methyltransferase by promoter hypermethylation and its relationship to aflatoxin B1-DNA adducts and p53 mutation in hepatocellular carcinoma. Int. J. Cancer 2003;103:440-444
Observed IC50 for in vitro cellular efficacy. ?
Observed IC50 for in vitro cellular toxicity studies. ? Acute toxicity:

Rat LD50 at 7 days: 2-4 mg/kg (7-1.3 umol/kg), single dose HepG2 cell LC50 at 24h: 0.9-7 uM (tetrazolium dye-based WST-1 assay)

References:

- Christopher McKean, Lili Tang, Madhavi Billam, Meng Tang, Christopher W. Theodorakis, Ronald J. Kendall and Jia-Sheng Wang, "Comparative acute and combinative toxicity of aflatoxin B1 and T-2 toxin in animals and immortalized human cell lines", J. Appl. Toxicol. 2006; 26: 139–147.

Apoptosis in HepG2 cells at 24 h. Note that ATP is not depleted at these concentrations. DNA fragmentation MEC 80 uM LDH release MEC 100 uM p53 elevation 3 uM

References:

-Travis O’Brien, George Babcock, James Cornelius, Mike Dingeldein, Glenn Talaska, David Warshawsky, and Kent Mitchell, "A Comparison of Apoptosis and Necrosis Induced by Hepatotoxins in HepG2 Cells",Toxicology and Applied Pharmacology 164, 280–290 (2000).

Doses of 1 uM have been used for gene expression studies in HepG2 studies.

References:

-C. Magkoufopoulou, S.M.H. Claessen, D.G.J. Jennen, J.C.S. Kleinjans, and J.H.M. van Delft, "Comparison of phenotypic and transcriptomic effects of false-positive genotoxins, true genotoxins and non-genotoxins using HepG2 cells", Mutagenesis vol. 26 no. 5 pp. 593–604, 2011.

Physical Properties ? Compound Assessment
Accepted and listed within the ToxCast/Tox21 program. ? No - Not included in ToxCast Phase I and II Chemicals List


soluble in DMSO
(-1 < log P < 6, i.e., log of the octanol/water partition coefficient; 97.5% meet this criteria)
Log P 2.04 (predicted ACD/Labs from ChemSpider) meets criteria
molecular weight range 250-1000 (90% meet this criteria)
Aflatoxin B1 312.3 g/mol meets criteria

Substance stability. ? Sensitivity to light


The amount of aflatoxin in water did not vary after 100 days of storage at 10°C. Little changes in concentration of AFB 1in deionized water solution after 140 days at 25°C

References:

-Garcia ME, Blanco JL, Suarez G. Aflatoxins B, and G1 solubility in standard solutions and stability during cold storage. Mycotoxin Research 1994 Vol. 10 (1994)

Soluble in buffer solution at 30 times the in vitro IC50 for toxicity. ? 0.01 mg/ml (25°C after 24 hours mixing); 0.05-0.09 mg/ml soluble with lower stability and probable formation of crystals.

References:

-Garcia ME, Blanco JL, Suarez G. Aflatoxins B, and G1 solubility in standard solutions and stability during cold storage. Mycotoxin Research 1994 Vol. 10 (1994)


estimated intrinsic solubility: 0.414 mg/ml
estimated solubility in pure water at pH 7.0: 0.414 mg/ml
estimated solubility in water at pH 7.4: 0.414 mg/ml
Calculations performed using ACD/PhysChem v 9.14
Solubility in DMSO 100 times buffer solubility. ? Soluble Sigma Aldrich Product Information Sheet

20mg/ml Enzo Lifesciences Aflatoxin B1 Product Informaton

Vessel binding properties. ? Adsorbtion to glass.

References:

-Rodricks IV. Note on adsorption of aflatoxin standards to glass. I Assoc Offic Anal Chern 1969; 52:979-980

Vapor pressure. (Non-volatile) ? estimated vapor pressure: 1.74E-09 mmHg (Calculation performed using EPI Suite v4.10)

Calculated/Predicted Properties

Water Solubility Results
pH Sol,mg/ml Flags  % Graph
2 0.41 N 100 Aflatoxin b1 solubility.png
5.5 0.41 N 100
6.5 0.41 N 100
7.4 0.41 N 100
10 0.41 N 100
Summary Solubility Data
Intrinsic Solubility,mg/ml 0.414
Intrinsic Solubility,log(S,mol/l) -2.8775
Solubility in Pure Water at pH = 7,mg/ml 0.414
Calculations performed using ACD/PhysChem v 9.14
Single-valued Properties
Property Value Units Error
Molar Refractivity 76.01 cm3 0.4
Molar Volume 199.58 cm3 5
Parachor 574.64 cm3 6
Index of Refraction 1.69 3.33E-2
Surface Tension 68.72 dyne/cm 5
Density 1.56 g/cm3 0.1
Polarizability 30.13 10E-24 cm3 0.5
Calculations performed using ACD/PhysChem v 9.14
Property Name Value Units Source
pKa - SPARC v4.5
Estimated VP 1.74E-09 mm Hg EPI Suite v4.10
Estimated VP 2.32E-07 Pa EPI Suite v4.10
Estimated Water Solubility 918.3 mg/L EPI Suite v4.10
WATERNT Frag Water Solubility Estimate 6.619 mg/L EPI Suite v4.10

Authors of this ToxBank wiki page

David Bower, Egon Willighagen, Matthew Clark
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