GM Free Cymru

Glyphosate and Roundup toxicity in humans confirmed

Glyphosate Toxic to Mouth Cells & Damages DNA, Roundup Much Worse

Further evidence of genotoxic and cytotoxic effects – a prelude to cancer, birth defects and reproductive problems by Dr Eva Sirinathsinghji

ISIS Report 28/03/12

A fully referenced version of this articles is posted on ISIS members website

New research finds that glyphosate causes cell and DNA damage to epithelial cells derived from the inside of the mouth and throat [1]. It raises concerns over the safety of inhaling glyphosate, one of the most common ways in which people are exposed to the herbicide.

Siegfried Knasmueller and his colleagues the Medical University of Vienna, Austria, found that Monsanto’s formulated version of glyphosate called Roundup Ultra Max caused cellular damage and DNA damage including chromosomal abnormalities and ultimately killed the cells at higher concentrations. Importantly, DNA damage occurred at concentrations below those required to induce cell damage, suggesting that the DNA damage was caused directly by glyphosate instead of being an indirect result of cell toxicity.

These are not the first findings of glyphosate-based herbicides’ cytotoxic and genotoxic effects. Numerous independent research teams have been documenting the hazards of glyphosate exposure over the last few years with in vivo, in vitro and clinical studies.

DNA damage was observed in blood samples from exposed residents in Argentina and Ecuador [2, 3]. Lab mice were found to harbour chromosomal and DNA damage in bone marrow, liver and kidney cells as well as lymphoid cells [4]. Similar effects were found in non-mammalian species, including sea urchins [5], goldfish [6, 7], eels [8], tilapia fish [9] as well as the fruitfly [10]. These experiments show that glyphosate herbicides are dangerous for humans and many other animals. Glyphosate is highly soluble in water, so impacts on aquatic wildlife may be of particular concern, especially following the recent report on the presence of glyphosate in rain water, groundwater, rivers and air [11, 12]. Its extreme toxic effects on amphibians such as frogs has already been shown (see [13] Roundup Kills Frogs, SiS 26). Cell damage has been documented in many cell types including those derived from the rat testis (see [14] Glyphosate Kills Rat Testes Cells, SiS 54), human placenta, umbilical cord, and embryo (see [15] Death by Multiple Poisoning, Glyphosate and Roundup, SiS 42), rat and carp neurones [16, 17], and liver [18, 19].

Multiple tests all show cellular damage in response to Roundup

To reflect occupational exposure, human buccal epithelial cells were exposed to glyphosate and Roundup for 20 minutes only at concentrations from 10 mg/L to 200 mg/L. The Roundup formulation used for the experiments contains 450 g/L of glyphosate and should be diluted according to the manufacturer’s instructions to 1–3 % before use (final concentration 4 500–13 500 mg/l). The researchers found some significant effects with 10-20 mg/l, equivalent to a 225–1 350-fold dilution of the spraying solution.

Cell damage was assessed by the release of the membrane-bound enzyme lactose dehydrogenase into the culture medium. The integrity and viability of cells was indicated by their staining with neutral red as only healthy cells retain the dye. Mitochondrial function was assessed by measuring the activity of the enzyme mitochondrial dehydrogenase with the substrate XXT that gives a yellow colour product. And cell proliferation was measured by the total protein content of the cell cultures.

The results showed that the cells were much more sensitive to the Roundup formulation than glyphosate. With Roundup, a significant effect was seen at a dose level of 40 mg/L with the XXT assay, while a clear increase of the lactose dehydrogenase levels was seen already with 10 mg/L. The cell proliferation and the neutral red assays were less responsive, with significant effects detected at 80 and 100 mg/L, respectively (still well below agricultural use levels). All effects were dose-dependent.

With glyphosate, no significant effects were seen in 3 of the 4 assays, with only lactose dehydrogenase showing significant effects at over 80 mg/l.

Multiple tests show Roundup causes DNA damage

DNA damage was analysed by two methods. The first is the Single Cell Gel Electrophoresis (SCGE) assay, which reveals single or double-stranded breaks in DNA. The second is a special comprehensive assay of chromosome instability that picks up many DNA aberrations including chromosome breakage, DNA misrepair, chromosome loss, as well as cell death by either necrosis (cell death that results from external stressors such as toxins), apoptosis (programmed cell-death) and cell growth. Different nuclear anomalies were measured including micronuclei, a biomarker of chromosomal damage, breakage or loss; nuclear buds, a biomarker of elimination of amplified DNA and/or DNA repair complexes; and nucleoplasmic bridges reflecting the formation of dicentric chromosomes (chromosomes with 2 instead of 1 centromere) , a marker of DNA misrepair and/or end-fusions of the chromosomes.

Significant effects on DNA integrity as assessed by the SCGE assay were seen at 20 mg/l of both Roundup and glyphosate, increasing in a dose-dependent manner.

Exposure of the cells for 20 minutes also led to a significant and dose-dependent increase of nuclear anomalies including increases in the total number of micronuclei beginning at 10 mg/L of Roundup, and 15 mg/L of glyphosate. The number of nuclear buds increased with exposure concentrations, starting at 10 mg/L with both glyphosate and Roundup. In the case of the nucleoplasmic bridges, the only significant effect was obtained with the highest dose of Roundup used (20 mg/L). Apoptotic cells were observed following 20mg/L of Roundup but not glyphosate, while necrosis occurred in response to 20mg/L of both Roundup and glyphosate.

In summary, Roundup was cytotoxic at concentrations as low as 20 mg/L, while its active ingredient was not generally cytotoxic to buccal epithelial cells. Both glyphosate and Roundup elicited genotoxic effects at concentrations below the level required to induce cell damage. The different effects between the active ingredient and its commercial formulation is consistent with previous work, including experiments done on testicular, placental, embryonic and umbilical cord cells (see above). These results may explain some of the ailments observed in people who work with this herbicide and adds yet more weight to an outright ban of the herbicide [20] Ban Glyphosate Herbicides Now, SiS 43).

Death by Multiple Poisoning, Glyphosate and Roundup

Scientists pinpoint how very low concentrations of the herbicide and other chemicals in Roundup formulations kill human cells, strengthening the case for phasing them out, and banning all further releases of Roundup-tolerant GM crops

by Dr. Mae-Wan Ho and Brett Cherry

This article was submitted to the USDA on behalf of ISIS ISIS Press Release 11/02/09

Four different Roundup formulations of the herbicide glyphyosate manufactured by Monsanto are highly toxic to human cells, and at concentrations far below the recommended agricultural use levels. Researchers at the Institute of Biology in Caen , France published their latest results in the current issue of Chemical Research in Toxicology [1] .

Roundup formulations are lethal cocktails

The four Roundup formulations are mixtures of glyphosate with various adjuvants. (An adjuvant is ‘helper' substance added to aid the effect of the active ingredient.) The Roundup formulations are currently the top non-selective herbicides worldwide and increasing, as more than 75 percent of genetically modified (GM) crops are Roundup tolerant. Glyphosate and its major metabolite, aminomethylphosphonic acid (AMPA) are main contaminants in rivers. The adjuvants, not often measured in the environment, are usually considered ‘inert' and protected as trade secret in manufacturing. Among them, the predominant one is polyethoxylated tallow amine (POEA) . POEA is used as a surfactant in Roundup formulations to improve solubility and penetration into plants.

Three human cell lines were tested: primary cell line HUVEC from umbilical cord vein epithelium, embryonic cell line 293 derived from kidney, and placenta cell line JEG3. All cells died within 24 hours of exposure to the Roundup formulations.

The Roundup formulations (Rs) contain different amounts of the active ingredient glyphosate: Roundup Express, 7.2 g/L (R7.2) ; Roundup Bioforce, 360 g/L (R360) ; Roundup Grand Travaux, 400 g/L (R400); and Roundup Grand Travaux Plus, 450 g/L (R450). They were compared with glyphosate (G), AMPA, and POEA . All Roundup formulations in the study, along with individual chemical ingredients, were tested at concentrations from10 ppm (parts per million) to 2 percent (the recommended agricultural usage level), which means that the Roundup formulations were diluted up to 100 000 times or more.

The researchers found that the presence of the other chemical ingredients in Roundup formulations, such as POEA, actually amplified glyphosate's toxic effects. The toxicities of the Roundup formulations were not proportional to the amount of glyphosate they contained, and are most likely due to POEA and other as yet undisclosed ingredient(s) present in all the formulations. POEA by itself is much more toxic than the Roundup formulations, while AMPA is more toxic than glyphosate.

Multiple targets in toxicity

The researchers tested Rs, G, AMPA, and POEA for effects on three targets that could kill the cell: damage to the cell membrane, poisoning of the mitochrondria (site of energy metabolism), and programmed cell death that results in fragmentation of the DNA in the cell nucleus. They measured specific enzyme markers at different concentrations for each damage at 24 h of exposure, and also obtained images of the cell cultures under the microscope.

All Rs, as well as G, caused cells to die; the results are the same for all human cell types, but at different concentrations. Thus, R400, the most toxic formulation, killed all cells at 20 ppm, which is equivalent to 8ppm in G. However, 4-10 ppm G alone is non-toxic, its toxicity begins around 1 percent (10 000 ppm), and is not connected with the cell membrane. The R formulations damage the cell membrane, and also poison the mitochondria. In contrast, G poisons the mitochondria without damaging the cell membrane

Unexpectedly, R400 is more toxic than R450, the latter in turn more harmful than R360, R7.2 and G. However, the toxicities are not proportional to the concentration of G present. The cell killing power of R7.2 was almost the same as that of R360, and these results are consistent across all cell lines. This suggests other unknown substances are involved in the toxic effects.

Thus AMPA and POEA also kill cells by poisoning the mitochrondia and damaging the cell membrane. POEA is so potent that it begins to damage the cell membrane in HUVEC and poison the mitochondria in 2 93 and JEG3 at 1 ppm. Roundup formulations are more toxic than either G or AMPA. AMPA itself destroys the cell membrane, however, which G does not do, though it is 3-8 times more toxic for the mitochondria than AMPA. But as cell membrane damage is more serious for the cell, AMPA is more toxic than G, while POEA is the most toxic of all.

What happens when these ingredients are combined? The researchers found that for HUVEC and 293 cells, combinations of G and POEA, G and AMPA, AMPA and POEA were all more toxic than the same concentration of the single ingredients

For programmed cell death, the action is quicker. The marker enzymes are activated from 6 h of exposure, with a maximum at 12 h in all cases. HUVEC was 60-160 times more sensitive than the other cell lines; G and R360 were effective at exactly the same concentration, from 50 ppm. The adjuvants do not seem necessary. G alone is 30 percent more potent here than Rs; it acted rapidly at concentrations 500 –1 000 times lower than agricultural use

Ban Roundup tolerant GM crops

These latest studies confirm a wealth of evidence on the toxicities of glyphosate and Roundup formulations [2] ( Glyphosate Toxic & Roundup Worse , SiS 26), and pinpoint the different sites of action, all of which result in cell death. Epidemiological studies have previously linked glyphosate to spontaneous abortions, non-Hodgkin lymphoma, and multiple myeloma. Laboratory studies showed that glyphosate inhibits transcription in sea urchin eggs and delays development. Brief exposures to glyphosate in rats caused liver damage, and adding the surfactant in Roundup had a synergistic effect, causing greater liver damage. Roundup was also found to be much more lethal to frogs than to weeds, and could have contributed to the global demise of amphibians within the past decades [3] ( Roundup Kills Frogs , SiS 26).

We have called for a new regulatory review on glyphosate and Roundup in 2005 [2]. There is a now a strong case for restricting, if not phasing out glyphosate and Roundup; in the first instance, by banning the release of Roundup tolerant GM crops worldwide. For the same reason, no further Roundup tolerant GM crops should be approved for commercial release.


• Benachour N and Séralini G-E.. Glyphosate formulations Induce Apoptosis and Necrosis in Human Umbilical, Embryonic, and Placental Cells Chem. Res. Toxicol. , 2009, 22 (1), pp 97–105

• Ho MW and Cummins J. Glyphosate toxic and Roundup worse Science in Society 26, 12, 2005.

• Ho MW. Roundup kills grogs Science in Society 26. 13, 2005.