The Developmental and Reproductive Risks Associated with Glyphosate Exposure

Aaron Lobsenz

Glyphosate is the primary active ingredient in many glyphosate-based herbicides (GBHs) including RoundUp, Touchdown, Glifloglex, and Glyphogan. When GBHs were first introduced to agriculture in the 1970s, they were regarded as harmless to humans. However, recent studies suggest that human exposure to GBHs may be associated with reproductive inhibition. This paper describes glyphosate and GBHs and the risks that they potentially pose to human reproduction and development.

Plants use the shikitame pathway to synthesize the amino acids tryptophan, phenylalanine, and tyrosine (see Figure 1)1 . Glyphosate’s ability to actively inhibit 3phosphoshikimate 1-carboxyvinyltransferase (EPSP synthase) prevents this mechanism.

Figure 1: This diagram depicts the mechanism of the shikimate pathway, which glyphosate and GBHs act upon to inhibit the synthesis of three essential amino acids. Researchers initially believed that GBHs would pose no significant health risks to humans, who do not use this pathway. Because of the increasing usage of GBH resistant crops – to remove unwanted weeds and plants while protecting crops – GBHs’ utilization has significantly increased. Between 1974 and 2014, overall glyphosate use in the United States (US) rose approximately 300-fold in agriculture. Globally, the use of glyphosate increased from 12 to 20 million liters from 2008 to 20122 . With its increasing presence in the environment and food supplies, some studies that examined GBHs’ impact on human and animal health are finding concerning results. Varying regulatory bodies have made different determinations regarding its safety. In 2017, the International Agency for Research on Cancer classified glyphosate as “probably carcinogenic” to humans. However, currently, the European Food Safety Authority and the US Environmental Protection Agency both consider glyphosate to pose no significant risks2 .

Glyphosate is frequently found within human urine, blood, and maternal milk, which primarily accumulates in the kidneys, liver, colon, and small intestine. Glyphosate in urine is detected at a rate of 0.16-7.6 µg/L in the general population, 0.26-73.5 µg/L in exposed workers, and up to 292 µg/L in workers in China involved in glyphosate production. One of the few studies that directly tested glyphosate on humans discovered that it could remain in the body for at least 106 hours after initial consumption. The same study also found that, in comparison to prior animal studies, less glyphosate is removed from urine in humans than in animals, raising possible concerns over bodily absorption rate and lingering presence of glyphosate in humans3 .

A 2005 study examined how cell viability changed depending on the amount of glyphosate concentration exposure at different durations of time3 . Researchers used an MTT assay to test the effects of glyphosate and Roundup on JEG3 placental cells by incubating the cells with 250 µL MTT and varying concentrations of glyphosate, then using a spectrophotometer to measure cell viability. The study measured aromatase activity – which is involved in estrogen synthesis – by measuring the conversion of androstenedione to estrone. The study found that the toxicity of glyphosate and Roundup on JEG3 cells increased when exposed to higher glyphosate and Roundup concentrations for longer durations, particularly 24 to 48 hours. It also found a decrease in cell viability among cells treated with low glyphosate concentrations -- less than 10 times of what is typically used in agriculture (a 1-2% concentration). It further concluded that cells exposed to Roundup for only an hour had increased in aromatase activity, but the same cells had decreased in aromatase activity when exposed for 18 hours (see Figure 2).

Figure 2 The effects of Roundup and glyphosate on JEG3 aromatase activity at time lengths of 1 and 18 hours at varying concentrations in a serum-free medium. These results make the researchers consider if Roundup should be labeled an endocrine disruptor, particularly when cells are exposed to glyphosate at the higher concentrations typically used in agriculture4 .

Multiple studies suggested that glyphosate may alter testicular morphology and testosterone levels. A 2010 study5 divided 68 weaned male Wistar rats into four groups, where each group received a different concentration of diluted glyphosate-Roundup Transorb (GRT) from the twenty-third-day post-natal (PND23) until PND53. The researchers monitored their weights and balanopreputial separation -- which is used as an index to determine the onset of male puberty. At PND53, researchers performed a cardiac puncture of the rats to collect serum, which was later used in a radioimmunoassay analysis to determine hormone concentrations. -There was little difference in overall body weight for rats when exposed to different GRT concentrations. However, the rats’ testicular weights were significantly greater in those exposed to higher concentrations of GRT. In addition, exposure to higher concentrations of GRT correlated with a delay in preputial separation and lower testosterone levels, but no change in estradiol presence (see Figure 3). The researchers found this particularly interesting as testosterone is a precursor of estradiol. This finding suggests that glyphosate consumption may pose a potential risk to male testosterone levels5 .

Figure 3: Testosterone levels in mice treated with varying amounts of Roundup (RU) from PND 23 to PND53. 5 *P<0.005, ***P<0.001

Multiple toxicology studies suggest the potential for morphological, functional, and biochemical alterations associated with increased GBH exposure. One research group found that mice (F1 generation) exposed prenatally to GBHs – where their mothers received a solution of 0.5% of the active ingredient glyphosate in their drinking water – had lower bodyweights at weaning. Then, these F2 offspring, whose parents and F1 mice were both exposed perinatally to GBH formulation in doses of 200 mg/kg/bodyweight/day, often developed

1 Peillex, Cindy, and Martin Pellletier. "The Impact and Toxicity of Glyphosate and Glyphosate-based Herbicides on Health and Immunity." Journal of Immunotoxicology, vol. 17, no. 1, 2020, pp. 163-74, www.tandfonline.com/doi/full/10.1080/1547691X.2020.1 804492. Accessed 25 Apr. 2022. 2 Richmond, Martha E. "Glyphosate: A Review of Its Global Use, Environmental Impact, and Potential Health Effects on Humans and Other Species." Journal of Environmental Studies and Sciences, 28 Sept. 2018, https://www.sciencedirect.com/science/. article/pii/S2667010021001281. Accessed 27 Apr. 2022. 3 Richard, S., Moslemi, S., Sipahutar, H., Benachour, N., & Seralini, G.-E. (2005). Differential effects of glyphosate and roundup on human placental cells and aromatase. Environmental Health Perspectives, 113(6), 716–720. https://doi.org/10.1289/ehp.7728

4 Faniband, Moosa, et al. "Human Experimental Exposure to Glyphosate and Biomonitoring of Young Swedish Adults." International Journal of Hygiene and Environmental Health, Jan. 2021, 6 bodily anomalies including conjoined fetuses and maldeveloped limbs. There are also concerns over glyphosate’s role in human neurodegenerative disorders. One population-based case-control study found an increase in the likelihood of developing autism spectrum disorder in people prenatally exposed to glyphosate.6 Other studies performed on rats found a decrease in locomotor activity and anxiety, exacerbated emotionality, and impairment in recognition memory in prenatally exposed offspring. The authors suggest that this may occur because GBHs can induce gene deregulation, which causes changes in the concentrations of stress-related metabolites, including lysophosphatidylcholine and phosphatidylcholine, which are commonly associated with neurodegenerative diseases7 .

There are concerns over the human health risks resulting from glyphosate exposure. As GBH use increases, further research is needed to better understand the multiple health effects associated with glyphosate exposure. Epidemiological studies are particularly needed to gain a more comprehensive understanding of the risks that glyphosate and GBHs pose to humans.

file:///Users/a22lobse/Downloads/Humanexperimentale xposuretoglyphosateandbiomonitoringofyoungSwedishadul ts.pdf. Accessed 28 Apr. 2022. 5 Romano, R. M., et al. "Prepubertal Exposure to Commercial Formulation of the Herbicide Glyphosate Alters Testosterone Levels and Testicular Morphology." Reproductive Toxicology, no. 84, 12 Dec. 2009, pp. 309-17, hhra.org/wp-content/uploads/Romano-Prepubertalexposure-to-commercial-form.pdf. Accessed 30 Apr. 2022. 6 von Ehrenstein, O. S., Ling, C., Cui, X., Cockburn, M., Park, A. S., Yu, F., Wu, J., & Ritz, B. (2019). Prenatal and infant exposure to ambient pesticides and autism spectrum disorder in children: Population based case-control study. BMJ, l962. https://doi.org/10.1136/bmj.l962 7 Milesi, Maria Mercedes, et al. "Glyphosate Herbicides: Reproductive Outcomes and Multigenerational Effects." Frontiers in Endocrinology, vol. 12, 7 July 2021, pp. 1-22, www.frontiersin.org/articles/10.3389/fendo.2021.672532/ full. Accessed 30 Apr. 2022.