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This research was supported entirely by the European Ramazzini Foundation
of Oncology and Environmental Sciences.
The authors declare that they have no competing financial interests.
Short running head: Carcinogenicity of Aspartame
Key words: artificial sweeteners, aspartame, carcinogenicity, lymphomas/leukemias, mammary cancers, prenatal exposure, Sprague-Dawley
ADI acceptable daily intake
BW body weight
CMCRC Cesare Maltoni Cancer Research Center
EFSA European Food Safety Authority
ERF European Ramazzini Foundation
EU European Union
FDA Food and Drug Administration
Outline of section headers
Materials and Methods
Total paper word count: 4,960
Aspartame (APM) is one of the most widely used artificial sweeteners in the world.
First approved by the US Food and Drug Administration (FDA) for limited use in solid food in 1981, its
authorization was extended to soft drinks in 1983 and then approved as a general sweetener in 1996
(FDA 1981; FDA 1983; FDA 1996).
Likewise, the sweetener was approved for general use in the European Union (EU) in 1994 (EC Directive 1994).
Today APM is present in over 6000 consumer packaged goods and in nearly 500 pharmaceutical products, including children’s medicines
(Aspartame Information Center 2005).
In the United States, over 70% of aspartame sales are attributed to soft-drinks (American Dietetic Association 2004).
The acceptable daily intake (ADI) of aspartame is currently 50 mg/kg body weight (bw) in the USA and 40 mg/kg bw in the EU for both
children and adults.
Daily assumption of artificial sweeteners by women of childbearing age and children has been estimated between 2.5 - 5.0 mg/kg bw (Butchko et al. 2002).
In a study of Swedish diabetics, although the general APM intake was lower than the ADI, the worst-case calculation of intake in the children’s group was reported to be 114% of ADI (Ilbäck et al 2003).
APM is metabolized in the gastric tract of rodents, non-human primates and humans to its three constituents: aspartic acid, phenylalanine and methanol.
When absorbed, aspartic acid is transformed into alanine plus oxaloacetate (Stegink 1984);
phenylalanine is transformed mainly into tyrosine and, to a lesser extent, phenylethylamine and phenylpyruvate (Harper 1984);
and methanol is transformed into formaldehyde and then to formic acid (Opperman 1984).
In vitro and in vivo tests demonstrate that APM is not genotoxic.
Likewise, long term carcinogenicity studies conducted by the manufacturers of aspartame using rats and mice in the ‘70s and ‘80s did not demonstrate any carcinogenic effects.
We have reported a detailed review of the genotoxicity and carcinogenicity studies available to date on APM in previous publications. (Belpoggi et al. 2006; Soffritti et al. 2005; Soffritti et al. 2006).
In our opinion, the small number of animals used per sex/per group and the termination of these experiments after 110 weeks of age rather than observing animals for the lifespan, represent limiting factors when evaluating the
carcinogenic risk or safety of artificial sweeteners such as aspartame.
It was for this reason, together with the growing use of APM in industrialized countries, that we designed and performed a mega-experiment using 7 groups of Sprague-Dawley rats (100-150/sex/group), treated with APM in feed at various dose levels (including one very close to the ADI for humans), from 8 weeks of age until natural death.
The study demonstrated for the first time that APM is a multipotential carcinogenic agent, capable of inducing, in our experimental conditions:
a) a significant, doserelated increased incidence of malignant tumor-bearing animals in males (p 0.05) and in females (p 0.01), in particular in females treated at 50,000 ppm (p 0.01);
b) a significant dose-related increase in lymphomas/leukaemias in both males (p 0.05) and females (p 0.01), in particular in females treated at doses of 100,000 (p 0.01), 50,000 (p 0.01), 10,000 (p 0.05), 2,000 (p 0.05), or 400 ppm (p 0.01);
c) a significant, dose-related increased incidence (p 0.01), of transitional cell carcinomas of the renal pelvis and ureter and their precursors (dysplasias) in females treated at
100,000 (p 0.01), 50,000 (p 0.01), 10,000 (p 0.01), 2,000 (p 0.05), or 400 ppm (p 0.05);
d) a significant, dose-related increased incidence of malignant schwannomas of peripheral nerves (p 0.05) in males (Belpoggi et al. 2006; Soffritti et al. 2005; Soffritti et al. 2006).
Given the consolidated experience of the European Ramazzini Foundation (ERF) in the conduct of long term bioassays, and the large number of rodents used in the study, the results attracted the attention of the scientific community, consumer and industry associations, and the national and international agencies responsible for food safety, including the Italian Superior Council of Health, the European Food Safety Authority (EFSA), the US FDA, Health Canada, and others.
Per their request, each of these agencies was provided with all available raw data related to the study.
To our knowledge, only EFSA has issued an official opinion on our study, releasing on May 5, 2006 a 40-page report in which they concluded that it is not necessary to revise their previous
opinion on the absolute safety of APM (EFSA 2006).
Subsequent to our findings of hematopoietic cancers in rats, and in light of persistent concerns among the scientific community of an association between APM and brain cancers, Lim et al (2006)
published the results of a study which assessed the correlation between the consumption of aspartame-containing beverages and the incidence of these types of cancers.
The findings were based on data derived from a prospective study conducted by the US National Institutes of Health and the American Association of Retired Persons, using a cohort of over 285,000 men and over 188,000 women between the ages of 50-71, who had satisfactorily responded to a self-administered food frequency questionnaire.
Among the survey questions was the consumption of beverages (soda, fruit drinks, sweetened iced tea) potentially containing APM during the previous year.
The questionnaires were mailed from 1995 to 1996 and the follow-up lasted until 2000.
The conclusions of the study did not support the hypothesis that APM increases hematopoietic or brain cancer risks.
Recently a group of Italian authors (Gallus et al. 2007) published the results of an integrated network of case-control studies conducted in Italy between 1991-2004 on the potential correlation between artificial sweeteners (including APM) and cancer.
The authors interviewed patients with histologically confirmed cancers of the oral cavity and pharynx (598), esophagus (304), colon (1225), rectum (728), larynx (460), breast (2569), ovary (1031), prostate (1294) and kidney (renal cell carcinoma 767).
Controls were 7028 patients (3301 men and 3727 women) admitted to the same hospitals for acute, non-neoplastic disorders.
Cases and controls were interviewed during their hospital stay, using a questionnaire on subjects' usual diet in the 2 years before diagnosis.
The results reported a lack of association between artificial sweeteners and the risk of the aforementioned cancers.
As soon as we perceived the carcinogenic effects of APM during the elaboration of the data in our first mega-experiment, we planned an integrated program of long-term bioassays, beginning treatment from prenatal life, on a total of more than 4000 rats and mice in order to better quantify the carcinogenic risks of aspartame.
In this report we present the results of a second study on APM in which male and female Sprague-Dawley rats were exposed to very low doses of APM in feed (100 or 20 mg/kg bw) from fetal life until natural death.
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