Federal Food, Drug, and Cosmetic Act (FFDCA) Considerations for Lipochitooligosaccharide (LCO) SP104
                                       
                    Docket ID Number: EPA-HQ-OPP-2017-0080
                            Date: January 22, 2018
                                       
Section 408(c)(2)(A)(i) of FFDCA allows the U.S. Environmental Protection Agency (EPA) to establish an exemption from the requirement for a tolerance (the legal limit for a pesticide chemical residue in or on a food) only if EPA determines that the exemption is "safe." Section 408(c)(2)(A)(ii) of FFDCA defines "safe" to mean that "there is a reasonable certainty that no harm will result from aggregate exposure to the pesticide chemical residue, including all anticipated dietary exposures and all other exposures for which there is reliable information." This includes exposure through drinking water and in residential settings but does not include occupational exposure. Pursuant to FFDCA section 408(c)(2)(B), in establishing or maintaining in effect an exemption from the requirement of a tolerance, EPA must take into account the factors set forth in FFDCA section 408(b)(2)(C), which require EPA to give special consideration to exposure of infants and children to the pesticide chemical residue in establishing a tolerance or tolerance exemption and to "ensure that there is a reasonable certainty that no harm will result to infants and children from aggregate exposure to the pesticide chemical residue . . . ." Additionally, FFDCA section 408(b)(2)(D) requires that EPA consider "available information concerning the cumulative effects of [a particular pesticide's] . . . residues and other substances that have a common mechanism of toxicity."
EPA performs a number of analyses to determine the risks from aggregate exposure to pesticide residues. First, EPA assesses the toxicity of the pesticide and identifies endpoints of concern, if any. Second, EPA examines aggregate exposure to the pesticide and other related substances through food, drinking water, and other non-occupational exposures. The anticipated aggregate exposure is then compared to any relevant endpoints to assess safety of the tolerance or exemption.
I. Summary of Petitioned-for Establishment of an Exemption from the Requirement of a Tolerance 
In the Federal Register of June 8, 2017 (82 FR 26641), EPA issued a document pursuant to FFDCA section 408(d)(3), 21 U.S.C. 346a(d)(3), announcing the filing of a pesticide tolerance petition (PP 6F8520) by Monsanto Company, 1300 I (Eye) St. NW., Suite 450 East, Washington, DC 20005. The petition requested that 40 CFR 180 be amended to establish an exemption from the requirement of a tolerance for residues of the plant growth regulator Lipochitooligosaccharide (LCO) SP104 in or on raw agricultural commodities and processed foods. That document referenced a summary of the petition prepared by the petitioner, Monsanto Company, LLC, which is available in Docket ID Number EPA-HQ-OPP-2017-0080 via http://www.regulations.gov. 
II. Toxicological Profile
Consistent with FFDCA section 408(b)(2)(D), EPA reviewed the available scientific data and other relevant information on LCO SP104 and considered their validity, completeness, and reliability, as well as the relationship of this information to human risk. EPA also considered available information concerning the variability of the sensitivities of major identifiable subgroups of consumers, including infants and children.
A. Overview of LOCs and LCO SP104

Lipochitooligosaccharides (LCOs) are signaling molecules involved in the initiation of plant-microbe endosymbiosis (the scenario when a microbe colonizes a plant) in an estimated 70-80% of land plants. LCOs are produced by bacteria on the plant roots. LCOs act as nodulation (Nod) factors or mycorrhization (Myc) factors inducing plant root formation. Since LCOs evolved tens to hundreds of millions of years ago and today are active signal transducers in the majority of land plants, there is a history of safe use/exposure to LCOs through the consumption of plants (Gough and Cullimore, 2011). 

LCO SP104 is a synthetically derived member of the LCO chemical class. The complete chemical name for LCO SP104 is D-Glucose, O - 2-deoxy-2- [[(11Z)-1-oxo-11-octadecen-1-yl] amino]- b-D-glucopyranosyl-(1-->4)-O - 2- (acetylamino)-2-deoxy-b-Dglucopyranosyl-(1-->4)-O - 2- (acetylamino)-2-deoxy-b-Dglucopyranosyl-(1-->4)-O - 2- (acetylamino)- 2-deoxy-b-Dglucopyranosyl-(1-->4)-2-(acetylamino)- 2-deoxy-. As a pesticide, LCO SP104 acts as a plant growth regulator (PGR) and is intended for use, specifically, to increase growth and decrease stress in growing crops. Typical of a PGR, LCO SP104 should be applied at low concentrations because use at high concentrations can result in detrimental effects to the plant. Humans are already exposed to LCOs as they are present in the roots of food crops and in the bacteria and fungi that are associated with the roots of these crops. Molecules identical to LCO breakdown products (such as chitin, vaccenic acid, and N-acetyl-D-glucosamine) are also present in insects, crustaceans, fungi, bacteria, and humans, and are regularly consumed by humans as part of a normal diet. 

Based on the data submitted in support of this petition (summarized in Unit II. B., below) and the comprehensive risk assessment conducted by the Agency (References 1 and 2, below), EPA concludes that there is a reasonable certainty of no harm from aggregate exposures to residues of LCO SP104, including the consumption of food treated with this active ingredient in accordance with label directions and good agricultural practices. EPA has made this determination because available toxicology data indicate that the active ingredient is of low acute toxicity and is not a developmental toxicant, a mutagen, or toxic via repeat oral exposure.

1. Acute Toxicity

LCO SP104 is of low acute toxicity based on the available data. To satisfy the Tier I mammalian acute toxicology data requirements for biochemical pesticides (listed at 40 CFR 158.2050, OCSPP Guideline numbers 870.1100, 870.1200, 870.1300, 870.2400, 870.2500, and 870.2600), toxicity studies on LCO SP104 were conducted according to the EPA/OCSPP guidelines and are presented in Table 1., below. In summary, based on the available data, LCO SP104 is of low toxicity for all routes of exposure. The chemical is classified into toxicity category IV for acute oral, dermal and inhalation toxicity and dermal and eye irritation. The substance is not a dermal sensitizer. 

Table 1. Acute Toxicology Data for LCO SP104 
                           Study/OCSPP Guideline No.
                                    Results
                         Toxicity Category/Description
                                     MRID
Acute oral toxicity 
(870.1100)
                         LD50  > 5,000 mg/kg (rat)
                                       
                                      IV
                                   49819912
Acute dermal toxicity 
(870.1200)
                          LD50 > 5,000 mg/kg (rat)
                                      IV
                                   49819913
Acute inhalation toxicity	
(870.1300)
                                       
                           LC50 > 5.14 mg/L (rat)
                                       
                                      IV
                                   49819914
Primary eye irritation 
(870.2400)
Minimally irritating (rabbit): minimal irritation observed at one-hour post instillation of the test substance with clearance by 24 hours 
                                      IV
                                   49819915
Primary dermal irritation 
(870.2500)
Nonirritating (rabbit): no irritation noted at any point throughout the 72-hour study.
                                      IV
                                   49819916
Dermal sensitization (Buehler)
(870.2600)
                         Not a sensitizer (guinea pig)
                                      N/A
                                   49819917


2. Subchronic Toxicity

90-Day Oral
	
Based on a weight of the evidence (WOE) approach, considering all the available LCO SP104 hazard and other data on similar compounds, EPA has determined that a 90-day oral study on LCO SP104 is not required at this time, thus the data requirement has been waived. This approach included the following considerations:
(1) several oral toxicity studies conducted on similar compounds to LCO SP104 showed no adverse effects at the highest dose tested (EPA, 2017a); (2) the acute toxicology studies performed on a manufacturing-use product containing LCO SP104 demonstrate Toxicology Category IV, suggesting that the compound has low toxicity on an acute basis; (3) LCO SP104 is structurally similar to naturally occurring LCOs that have long been part of the normal diet (seafood and plants); (4) prior data waivers for all subchronic toxicity requirements were granted by the agency for use of chitin and chitosan as food additives and plant growth regulators, respectively; (5) the LCO SP104 mode of action is not relevant outside the plant kingdom; (6) there is an existing U.S. EPA tolerance exemption for chitin and chitosan; (7) similar LCO compounds are ubiquitous in the environment; (8) due to low toxicity, the EPA will be performing only a qualitative assessment; 9) LCO SP104 is proposed to be used only a very low application rates; and (10) worst-case exposure estimates for LCO SP104 fall well below generally accepted Thresholds of Toxicological Concern (TTCs) for corn and canola (the proposed LCO SP104 use sites), which is 1800 micrograms (μg)/chemical/person/day for Cramer Class I, indicating that expected dietary exposure of LCO SP104 to the general population is well below the level of concern using this metric (Dewhurst and Renwick, 2013).

90-Day Dermal

EPA has determined that a 90-day dermal study on LCO SP104 is not required at this time (the data requirement has been waived) based upon the following considerations: (1) the percent of LCO SP104 proposed to be used ranges from 0.000001% to 0.000025% in end-use formulations and (2) dermal, inhalation, and combined Margins of Exposure (MOEs) were calculated (from a structurally similar chemical) and all are well above the Agency's level of concern of 100 (ranging from 8.1 x 10[9]-7.8 x 10[14]) (EPA, 2017a).


90-Day Inhalation

EPA has determined that a 90-day inhalation study on LCO SP104 is not required at this time (the data requirement has been waived) based upon the following considerations: (1) the percent of LCO SP104 proposed to be used ranges from 0.000001% to 0.000025% in end-use formulation and (2) dermal, inhalation, and combined Margins of Exposure (MOEs) were calculated and all are well above the Agency's level of concern of 100 (ranging from 8.1 x 10[9]-7.8 x 10[14]) (EPA, 2017a).

3. Mutagenicity 

Mutagenicity data are required to support food uses, but are unavailable for LCO SP104. However, EPA has determined that genetic toxicity testing on LCO SP104 is not required at this time, waiving these data requirements. This approach included the following considerations: (1) available literature studies demonstrated a lack of genotoxicity on compounds structurally similar to LCOs (EPA, 2017a); (2) structure-activity relationships of LCO SP104 demonstrate a lack of genotoxic potential; (3) prior genotoxicity data waivers were granted by the agency for use of (structurally similar) chitin and chitosan as food additives and plant growth regulators, respectively; (4) U.S. EPA has established a tolerance exemption for chitin and chitosan; (5) the ubiquitous nature of LCOs in the environment; (6) a History of Safe Use (HOSU) of LCO's; and (7) the maximum predicted exposure levels of LCO SP104 are orders of magnitude lower than generally recognized Thresholds of Toxicological Concern (TTCs).

4.  Developmental Toxicity

Based on a WOE approach, considering all the available hazard and exposure data on
LCO SP104, the HASPOC recommends that a Prenatal Developmental Study be not
required at this time. This approach included the following considerations: (1) LCO SP104 is structurally similar to naturally occurring LCOs that have long been part of the normal diet (seafood and plants); (2) the acute toxicology studies are classified as Toxicity Category IV, indicating low acute toxicity for LCO SP104; 90-day oral toxicity studies of similarly-structured compounds (chitin-glucan, chitin, N-acetylglucosamine, and oligo-N-acetylglucosamine) have found NOAEL's greater than 2000 mg/kg in rats (3) prior data waivers for all subchronic toxicity requirements were granted by the agency for use of chitin and chitosan as food additives and plant growth regulators, respectively; (4) mode of action is not relevant outside the plant kingdom; (5) there is an existing U.S. EPA tolerance exemption for the related compounds, chitin and chitosan; (6) ubiquity in the environment; (7) due to the low toxicity for this chemical, a qualitative assessment will be performed; (8) the low application rates of LCO SP104; and (9) worst-case exposures that fall well below the rabbit developmental Threshold of Toxicological Concern (TTCs) for corn and canola of 280 μg/chemical/person/day (van Ravenzwaay et al., 2012).

III. Aggregate Exposure

In examining aggregate exposure, FFDCA section 408 directs EPA to consider available information concerning exposures from the pesticide residue in food and all other non-occupational exposures, including drinking water from ground water or surface water and exposure through pesticide use in gardens, lawns, or buildings (residential and other indoor uses).

Food and Drinking Water Exposure:  An aggregate risk assessment for LCO (food and drinking water) was not conducted, as no toxicological endpoints have been identified in the toxicity database. Furthermore, residues of LCO SP104 in drinking water are not expected when products are used according to label instructions. The active ingredient is applied at low concentrations, is very soluble in water, and will dissociate within minutes once applied (EPA, 2017b). 

Other Non-Occupational Exposure: As there are no residential uses for LC SP104, there are no residential exposure contributions to aggregate exposure. Therefore, aggregate risk is equivalent to the dietary (food and drinking water) risk and is not of concern, as noted above.

IV. Cumulative Effects from Substances with a Common Mechanism of Toxicity

Section 408(b)(2)(D)(v) of FFDCA requires that, when considering whether to establish, modify, or revoke a tolerance, EPA consider "available information concerning the cumulative effects of [a particular pesticide's] . . . residues and other substances that have a common mechanism of toxicity."

LC SP104 is of low toxicity and does not have a common mechanism of toxicity with other substances and cumulative effects are not expected.  

V. Determination of Safety for the U.S. Population, Infants and Children

A. U.S. Population

For all of the reasons discussed previously, EPA concludes that there is reasonable certainty that no harm will result to the U.S. population, including infants and children, from aggregate exposure to residues of LCO SP104. This includes all anticipated dietary exposures and all other exposures for which there is reliable information.

B. Infants and Children

FFDCA section 408(b)(2)(C) provides that EPA shall apply an additional tenfold (10X) margin of safety for infants and children in the case of threshold effects to account for prenatal and postnatal toxicity and the completeness of the database on toxicity and exposure, unless EPA determines based on reliable data that a different margin of safety will be safe for infants and children. This additional margin of safety is commonly referred to as the Food Quality Protection Act Safety Factor. In applying this provision, EPA either retains the default value of 10X or uses a different additional safety factor when reliable data available to EPA support the choice of a different factor. As discussed above, EPA has concluded that LCO SP104 is not toxic to mammals, including infants and children. Because there are no threshold levels of concern to infants, children, and adults when LCO SP104 is used according to label directions and good agricultural practices, EPA concludes that no additional margin of safety is necessary to protect infants and children.

VI. Conclusions
EPA concludes that there is a reasonable certainty that no harm will result to the U.S. population, including infants and children, from aggregate exposure to residues of LCO SP104. Therefore, an exemption from the requirement of a tolerance is established for LCO SP104 in or on all food commodities when applied/used pre-harvest and in accordance with label directions and good agricultural practices. 

VII. References

EPA, 2017a. U.S. Environmental Protection Agency (EPA). October 2, 2017. LCO (lipochitooligosaccharide) SP104: Summary of Hazard and Science Policy Council (HASPOC) Meeting on September 7, 201 7: Recommendations on the Need for Waivers for the Following Subchronic Studies: 90-Day Oral Study (OCSPP 870.3000), 90-Day Dermal Study (OCSPP 870.3250), 90-Day Inhalation Study (OCSPP 870.3465) Prenatal Developmental Study (OCSPP 870.3700), and Genetic Toxicity Testing (OCSPP 870.5100, 870.5300, 870.5375). Memorandum from Austin Wray, through Kelly Lowe, to Sadaf Shaukat. 

EPA 2017b. U.S. Environmental Protection Agency (EPA). August 1, 2017. Risk Assessment in Support of the Registration of LCO SP104 MP, Containing 0.013% Lipochitooligosaccharide (LCO) SP104 as its Active Ingredient. Memorandum from Sadaf Shaukat, through Miachel Rexrode, to Gina Burnett. 

Gough, C. and J. Cullimore. 2011. Lipo-chitooligosaccharide Signaling in Endosymbiotic Plant-
Microbe Interactions. Molecular Plant-Microbe Interactions 24:867-878

Dewhurst, I. and A.G. Renwick. 2013. Evaluation of the Threshold of Toxicological Concern
(TTC)  -  Challenges and approaches. Regulatory Toxicology and Pharmacology 65:168-177.

van Ravenzwaay, B., M. Dammann, R. Buesen, B. Flick and S. Schneider. 2012. The threshold of
toxicological concern for prenatal developmental toxicity in rabbits and a comparison to TTC
values in rats. Regulatory Toxicology and Pharmacology 64:1-8.
