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Pioneer Hi-Bred International, Inc.

High Oleic 305423 Soybean

OECD Unique Identifier:

DP-3Ø5423-1

Final Environmental Assessment

May 2010

Agency Contact

Cynthia Eck

Document Control Officer

Biotechnology Regulatory Services

USDA, APHIS

Riverdale, MD 20737

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(voice and TDD).

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Whitten Building, 1400 Independence Avenue, SW, Washington, DC 20250–9410 or call (202) 720–

5964 (voice and TDD). USDA is an equal opportunity provider and employer.

______________________________________________________

Mention of companies or commercial products in this report does not imply recommendation or 

endorsement by the U.S. Department of Agriculture over others not mentioned. USDA neither 

guarantees nor warrants the standard of any product mentioned. Product names are mentioned 

solely to report factually on available data and to provide specific information.

______________________________________________________

This publication reports research involving pesticides. All uses of pesticides must be registered by 

appropriate State and/or Federal agencies before they can be recommended. 

______________________________________________________

 
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Table of Contents

I. PURPOSE & NEED................................................................................................3

II. INTRODUCTION..................................................................................................5

III. AFFECTED ENVIRONMENT............................................................................7

A. Soybean.............................................................................................................................................................7

B. Agricultural Production of Soybean ............................................................................................................7

C. Soybean Composition.....................................................................................................................................9

IV. ALTERNATIVES...............................................................................................11

A. No Action: Continuation as a regulated article.......................................................................................11

B. Grant nonregulated status to Pioneer 305423 soybean, “in whole”-Preferred Alternative: 

Determination that Pioneer 305423 soybean is no longer a regulated article..........................................12

Alternatives Considered but Rejected from Further Consideration.........................................................12

V. ENVIRONMENTAL CONSEQUENCES............................................................13

A. No Action........................................................................................................................................................13

A. Soybean.......................................................................................................................................................13

B. Agricultural Production of Soybean..........................................................................................................14

C. Soybean Composition................................................................................................................................15

B. Preferred Alternative....................................................................................................................................16

A. Soybean.......................................................................................................................................................16

B. Agricultural Production of Soybean..........................................................................................................17

C. Soybean Composition................................................................................................................................20

Cumulative effects..............................................................................................................................................24

Potential impact on non-target organisms, including beneficial organisms and threatened or 

endangered species.............................................................................................................................................25

Consideration of Executive Orders, Standards and Treaties relating to environmental impacts........28

VI. REFERENCES...................................................................................................31

VII. PREPARERS AND REVIEWERS ...................................................................39

VIII. APPENDIX 1 - BIOTECHNOLOGY CONSULTATION NOTE TO THE 

FILE BNF NO. 000110 .............................................................................................40
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I. PURPOSE & NEED

"Protecting American agriculture" is the basic charge of the U.S. Department of 

Agriculture's (USDA) Animal and Plant Health Inspection Service (APHIS). APHIS 

provides leadership in ensuring the health and care of plants and animals. The agency 

improves agricultural productivity and competitiveness, and contributes to the national 

economy and the public health. USDA asserts that all methods of agricultural production 

(conventional, organic, or the use of genetically engineered varieties) can provide 

benefits to the environment, consumers, and farm income. 

In 1986, the Federal Government’s Office of Science and Technology Policy (OSTP) 

published a policy document known as the Coordinated Framework for the Regulation of 

Biotechnology. This document specifies three Federal agencies that are responsible for 

regulating biotechnology in the U.S.: USDA’s APHIS, the U.S. Department of Health 

and Human Services’ Food and Drug Administration (FDA), and the Environmental 

Protection Agency (EPA). APHIS regulates genetically engineered (GE) organisms 

under the Plant Protection Act of 2000. FDA regulates GE organisms under the authority 

of the Federal Food, Drug, and Cosmetic Act. The FDA policy statement concerning 

regulation of products derived from new plant varieties, including those genetically 

engineered, was published in the Federal Register on May 29, 1992 (57 FR 22984-

23005). Under this policy, FDA uses what is termed a consultation process to ensure that 

human food and animal feed safety issues or other regulatory issues (e.g., labeling) are 

resolved prior to commercial distribution of bioengineered food. The EPA regulates 

plant-incorporated protectants under the Federal Insecticide, Fungicide, and Rodenticide 

Act (FIFRA) and certain biological control organisms under the Toxic Substances 

Control Act (TSCA). Together, these agencies ensure that the products of modern 

biotechnology are safe to grow, safe to eat, and safe for the environment. USDA, EPA, 

and FDA enforce agency-specific regulations to products of biotechnology that are based 

on the specific nature of each GE organism. Products are regulated according to their 

intended use and some products are regulated by more than one agency. 

The APHIS Biotechnology Research Service’s (BRS) mission is to protect America’s 

agriculture and environment using a dynamic and science-based regulatory framework

that allows for the safe development and use of genetically engineered organisms. 

APHIS regulations at 7 Code of Federal Regulations (CFR) part 340, which were 

promulgated pursuant to authority granted by the Plant Protection Act, as amended (7 

United States Code (U.S.C.) 7701–7772), regulate the introduction (importation, 

interstate movement, or release into the environment) of certain GE organisms and 

products. A GE organism is no longer subject to the regulatory requirements of 7 CFR 

part 340 when APHIS determines that it is unlikely to pose a plant pest risk. A GE 

organism is considered a regulated article if the donor organism, recipient organism, 

vector, or vector agent used in engineering the organism belongs to one of the taxa listed 

in the regulation (7 CFR 340.2) and is also considered a plant pest. A GE organism is 

also regulated under Part 340 when APHIS has reason to believe that the GE organism 

may be a plant pest or APHIS does not have sufficient information to determine if the GE 

organism is unlikely to pose a plant pest risk.
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A person may petition the agency to evaluate submitted data and determine that a 

particular regulated article is unlikely to pose a plant pest risk, and, therefore, should no 

longer be regulated under 7 CFR 340.6 entitled “Petition for determination of 

nonregulated status.” The petitioner is required to provide information under § 

340.6(c)(4) related to plant pest risk that the agency uses to determine whether the 

regulated article is unlikely to present a greater plant pest risk than the unmodified 

organism. A GE organism is no longer subject to the regulatory requirements of 7 CFR 

part 340 when APHIS determines that it is unlikely to pose a plant pest risk. 

Pioneer Hi-Bred International, Inc. (Pioneer) of Johnston, IA submitted a petition to 

APHIS seeking a determination of non-regulated status for their transgenic high oleic 

acid soybean event DP-3Ø5423-1 (hereafter referred to as Pioneer 305423 soybean)

(Pavely, 2007). According to Pioneer, their 305423 soybean is engineered to produce 

increased amounts of monounsaturated fatty acid (oleic) and decreased amounts of 

polyunsaturated fatty acids (linoleic and linolenic) and to lesser extent, saturated fatty 

acid (palmitic acid). This soybean is also engineered to express a new protein, a 

modified soybean acetolactate synthase. The modified soybean acetolactate synthase was 

used as a selectable marker for transformation. The Pioneer 305423 soybean is currently 

regulated under 7 CFR part 340. Interstate movements and field trials of Pioneer 305423 

soybean have been conducted under permits issued or notifications acknowledged by 

APHIS. 

Under the authority of 7 CFR part 340, APHIS has the responsibility for the safe 

development and use of genetically engineered organisms under the provisions of the 

Plant Protection Act. APHIS must respond to petitioners that request a determination of 

the regulated status of genetically engineered organisms, including genetically 

engineered crop plants such as Pioneer 305423 soybean. If a petition for nonregulated 

status is submitted, APHIS must make a determination if the genetically engineered 

organism is unlikely to pose a plant pest risk. 

As a Federal agency subject to compliance with the National Environmental Policy Act 

(NEPA) (42 U.S.C. 4321 et seq.), APHIS has prepared this environmental assessment 

(EA) to consider the potential environmental effects of this proposed action (granting 

nonregulated status) and the reasonable alternatives to that action consistent with NEPA 

regulations (40 CFR parts 1500-1508, 7 CFR 1b, and 7 CFR part 372) and the USDA and 

APHIS NEPA implementing regulations and procedures. This EA has been prepared in 

order to specifically evaluate the effects on the quality of the human environment 1

The Pioneer 305423 soybean is designed for human and animal consumption and as such,

may also be subject to regulation by Food and Drug Administration (FDA). FDA policy 

uses what is termed a consultation process to ensure that human food and animal feed 

safety issues or other regulatory issues (e.g., labeling) are resolved prior to commercial 

distribution of biotechnology-derived food. Pioneer submitted a summary of its safety 

that 

may result from the deregulation of Pioneer 305423 soybean. 

 1 Under NEPA regulations, the “human environment” includes “the natural and physical environment and 

the relationship of people with that environment” (40 CFR §1508.14)
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and nutritional assessment to FDA for Pioneer 305423 soybean in 2007. Pioneer 

concluded that, with the exception of the intended change in fatty acid composition, the 

305423 soybean and the foods and feeds derived from it are not materially different in 

composition, safety, or any other relevant parameter from soybeans now grown, 

marketed, and consumed. In January 2009, FDA considered Pioneer's consultation on the 

305423 soybean to be completed regarding the safety and nutritional assessment for 

Pioneer 305423 soybean and had no further questions regarding the safety of Pioneer 

305423 soybean (FDA, 2009). The text of the FDA’s scientific and regulatory

assessment response for Pioneer 305423 is available at the FDA website (FDA, 2009). 

Because Pioneer 305423 soybean does not contain any GE pesticides or the genetic 

machinery necessary to produce them, or tolerance to herbicides, EPA consultation is not 

required.

Public Involvement

APHIS-BRS routinely seeks public comment on draft environmental assessments

prepared in response to petitions to deregulate GE organisms. APHIS-BRS does this 

through a notice published in the Federal Register. This EA, the petition submitted by 

Pioneer, and APHIS’s plant pest risk assessment, were made available for public 

comment for 120 days, from September 2, 2009 until December 28, 2009. Comments 

that were received within the comment period were fully analyzed and used by APHIS to 

determine if the petition to deregulate the Pioneer 305423 soybean should be granted. 

Decision to Be Made

APHIS will also use the information from this EA, and the comments received, to inform 

and to assist APHIS’ decisionmaker to determine whether to grant nonregulated status, or 

to continue to regulate Pioneer 305423 soybean under the regulations at 7 CFR part 340, 

or that an Environmental Impact Statement for Pioneer 305423 soybean is necessary prior 

to the decision of whether to grant nonregulated status to this soybean variety.

II. Introduction

Pioneer has developed a transgenic soybean line “Pioneer 305423 Soybean” that 

produces soybean seeds with increased levels of monounsaturated (oleic) fatty acid, 

decreased levels of polyunsaturated fatty acids (linoleic and linolenic) and decreased 

levels of palmitic acid (Pavely, 2007). In addition, Pioneer 305423 soybean also contains 

a slightly modified version of a soybean acetolactate synthase gene. The expression of 

the modified version of a soybean acetolactate synthase protein can increase the inherent 

tolerance level to the ALS-inhibiting class of herbicides. This trait is intended for 

selecting and identifying this high oleic soybean bioengineered event, rather than as a 

separate commercial trait as this version of the gene does not confer commercial levels of 

herbicide tolerance in Pioneer 305423 soybean. Pioneer indicated there is no plan to 

commercially promote Pioneer 305423 soybean as tolerant to sulfonylurea herbicides

(Pavely, 2007).
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Genetic modification of the fatty acid composition of soybean oil has been one of the 

major goals of many soybean breeders over the last 50 years. The levels of 

polyunsaturated fatty acids are one of the major factors influencing the quality of 

vegetable oils. Soybean oils rich in monounsaturated fatty acids could provide improved 

commercial value. Pioneer identifies users of the modified oil as both food 

manufacturers and industrial product manufacturers, especially those who are presently 

using soybean oil products.

Unmodified soybean oil has poor oxidative stability due to its unstable chemical structure 

and naturally occurring levels of polyunsaturated fatty acids. Polyunsaturated fatty acids

increase rancidity compared with saturated and monounsaturated fatty acids, especially 

after prolonged contact with oxygen, light or heat. This characteristic reduces product 

stability and shelf life. Hydrogenation is a chemical process that improves stability and 

shelf life necessary for food application; however, hydrogenation has the undesirable 

consequences of creating trans-fatty acids. 

In recent years, trans-fatty acid have come under considerable scrutiny because of their

negative affects on human health. On July 9, 2003, the FDA issued a regulation requiring 

manufacturers to list trans-fatty acids, or trans fat, on the Nutrition Facts panel of foods 

and some dietary supplements (FDA, 2003). With this rule, consumers have more 

information to make food choices that could lower their consumption of trans-fat as part 

of a heart-healthy diet. 

USDA APHIS has previously granted nonregulated status to a high oleic soybean variety

(USDA-APHIS, 1997) developed by DuPont in which the high oleic phenotype was 

conferred by introduction of the soybean omega-6 desaturase gene 1 (fad2-1 gene).

Those DuPont high oleic soybean varieties received regulatory approval in Australia, 

Canada, Japan, New Zealand, and the United States.

 

Pioneer 305423 soybean has been field tested in the United States since 2005 as 

authorized by APHIS. Associated notifications acknowledged and permits issued by 

APHIS are listed in Figure 23 (Pavely, 2007, p.67). The list compiles a total of 13 test 

sites in diverse regions of the U.S. and Canada including the major soybean growing area 

of the Midwest and winter nurseries in Hawaii. Field tests conducted under APHIS 

oversight allow for evaluation in agricultural settings under confinement measures 

designed to minimize the likelihood of persistence in the environment after completion of

the field trial. Under confined field trial conditions, data are gathered on multiple 

parameters and used by applicants to evaluate agronomic characteristics and product 

performance. These data are also valuable to APHIS as the agency assesses the potential 

for a new variety to pose a plant pest risk. The evaluated data may be found in the 

APHIS plant pest risk assessment (USDA-APHIS, 2009).
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III. Affected Environment

A. Soybean 

The soybean (Glycine max (L.) Merr.) is an economically important leguminous crop, 

providing oil and protein. Soybean plants are grown for their seed, which is further 

processed to yield oil and meal. Soybean is ranked number one in oil production (56%) 

among the major oil seed crops production in the world (Soy Stats, 2008). Other 

expanding uses for soybeans in the U.S. include soy biodiesel, animal agriculture, 

exports, and edible soybean oil (USB, 2007). Increased public focus on data supporting 

the human health benefits of soybeans could create more consumer demand, and will be 

examined further in the Environmental Consequences section of this EA.

The OECD Consensus Document (OECD, 2000) provides detailed information about the 

crop biology of soybean. The genus Glycine is divided into two subgenera, glycine and 

soja. The subgenus soja consists of three annual species: G. soja Sieb. and Zucc., the 

wild form of soybean, G. gracilis Skvortz., the weedy form of soybean and G. max,

which is the cultivated soybean. These species do not exist naturally in the United States 

(USDA-NRCS, 2008). Soybean lacks sexually compatible wild relatives in the United 

States and its territories. Therefore, there is no potential for gene flow from cultivated 

soybean plants to wild soybean relatives in the U.S.

Transgenes in crops have the potential to move within a population. The potential for 

outcrossing can be defined as the ability of gene escape to other soybean fields. Soybean 

is a highly self-pollinating species with a cross-pollination rate of less than one percent in 

plants grown in close proximity (OECD, 2000; Caviness, 1966). Cross-pollination 

greater than 4.6 m from a pollen source has been rarely observed although it has been 

reported that insects can sometimes transfer the pollen that distance or more (Caviness,

1996). Even if gene flow occurred, the nature of this trait (oil composition changes) 

would not confer a selective advantage. The only known propagation method for 

soybean is through seed germination (i.e., there are no reports of vegetative propagation 

under field conditions in the United States). Mature soybean seeds have no innate 

dormancy, are sensitive to cold, and are not expected to survive in freezing winter 

conditions (Raper et al., 1987). Volunteer plants that might grow under certain 

environmental conditions can be easily controlled mechanically or with herbicides

(Zollinger, 2005). Soybean is not weedy (Holm et al., 1977), is not found outside of 

cultivated areas, and does not compete well with other cultivated plants (Hymowitz et al.,

1987).

B. Agricultural Production of Soybean 

To determine the areas of soybean production, APHIS used data from the National 

Agricultural Statistics Service (NASS) 2002/2007 Census of Agriculture to determine 

where soybean is produced in the United States (USDA-NASS 2009). Only 17 states in 

2008 produced more than 1 million acres among the 31 that produced soybean according 

to the 2002 Census of Agriculture. These states include IA, IL, MN, IN, MO, NE, OH, 

SD, AR, ND, KS, MI, MS, WI, NC, KY, TN (USDA-NASS, 2008). In the U.S.,

soybeans were harvested on 72.1 million acres in 2008 (USDA-NASS, 2008). This 
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harvested soybean acreage was only one percent below the record high acreage in 2006

(USDA-NASS, 2008a).

Pioneer states that this variety will “offer growers another value-added specialty soybean 

option” and do not expect that soybean market dynamics will change with the 

availability of this product (Pavely, 2007). Properties of soybean oil from this variety 

will be preferred over those commodity soybeans with less stability, and over the inferior 

flavor properties that currently characterize hydrogenated commodity oils (Pavely, 

2007). Thus, products derived from Pioneer 305423 soybean will likely only replace 

commodity soybean products, and not create new demand. APHIS concludes that it is 

not likely that “new,” previously uncultivated land will be brought into soybean 

production beyond that currently or historically used for such production.

Processed soybeans are the largest source of protein feed and the second largest source 

of vegetable oil in the world. Soybeans represent about 90 percent of U.S. total oilseed 

production, while other oilseeds—such as cottonseed, sunflower seed, canola, and 

peanuts—account for the remainder (USDA-ERS, 2008).

Agricultural production of genetically engineered herbicide tolerantsoybean 

Based on USDA survey data, adoption of genetically engineered herbicide-tolerant (HT) 

soybeans increased from 17 percent of U.S. soybean acreage in 1997 to 68 percent in 

2001 and 92 percent in 2008 (Fernandez-Cornejo, 2008). Use of herbicide-resistant crops 

has been a major development in agriculture over the past 20 years. Weed control had 

been one of the biggest challenges for soybean growers. Infestation with weeds during 

an entire growing season have resulted in soybean yield losses ranging from 12 to 80% 

(Barrentine, 1989). By the early 1990’s, Gianessi et al. (2002) reported over 70 

individual herbicides or combination products registered for weed control in soybean. 

Along with the increased use of herbicides, biotypes of various plant species developed 

resistance to certain herbicide modes of action (Heap, 2007). With the 1996 commercial 

introduction of glyphosate tolerant soybean, a major shift occurred with an increased use 

of glyphosate accompanying the increased planting of glyphosate tolerant soybean (92%

of all soybean planted in the United States in 2008) and a decrease in use of other 

soybean herbicides(Gianessi et al., 2002). According to USDA’s Agricultural and 

Resource Management Surveys (ARMS) in 2001-03, growers who used glyphosatetolerant soybean technology cited the simplicity in weed control as the most common

reason for growing HT soybean varieties. The popularity of glyphosate-tolerant soybean 

is due to advantages of the technology over conventional weed control practice. 

Agricultural production of conventional soybean and genetically-engineered, non 

herbicide tolerant soybean

According to the report, Adoption of Genetically Engineered Crops in the U.S.,

“approximately 8% of total soybean acres in 2008 were planted with non-herbicide 

tolerant varieties (Fernandez-Cornejo, 2008).” A portion of this area (about 0.13% of 

U.S. soybean production in 2008) was devoted to the production of organic soybean 

(USDA-NASS, 2010). 
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Weed control is one of the biggest challenges for all soybean farmers. United States 

soybean farmers began switching from the use of tillage for weedcontrol to herbicides in 

the late1950s. Herbicides were estimated to be used on more than 97 percent of the total 

soybean acreage in 1997 in the U.S. (Fernandez-Cornejo et al., 2002). As a result, 

soybean is a major market for pesticides in general and for herbicides in particular.

For Pioneer 305423 soybean (a non-herbicide tolerant soybean), there are no anticipated 

changes in pesticide use for weed management compared to conventional soybean

varieties. The potential herbicide use in growing conventional soybean or Pioneer 

305423 soybean should be very similar. Currently, there are at least 70 registrations for 

herbicides for weed management in soybean, plus numerous mixtures (Crop Protection 

Reference, 2009).

Organic soybean production

The production of organic soybeans represents about 0.13% of U.S. soybean production 

(USDA-NASS, 2010). In 2008, 98,199 acres of certified or exempt organic soybean 

were harvested in United States. Under the USDA National Organic Program guideline, 

the use of synthetic pesticides, fertilizers, and genetically engineered crops is strictly 

limited. Pioneer 305423 soybean is not approved for use in organic systems because it is 

genetically engineered.

Maintaining the integrity of the organic production process is important to producers of 

organic soybeans. There are many practices organic producers use to prevent movement 

of GE soybean or the transfer of pollen from GE soybean into their organic production 

fields (Organic Farming Research Foundation (2002). Growers may choose to plant only 

organic seed; plant earlier or later than neighboring farmers who may be using GE crops, 

ensuring that the flowering times between GE and organically produced crops will differ,

thus minimizing the change of pollen movement between fields; and also employ 

adequate isolation distances between the organic field and the fields of neighbors to 

minimize the chance that pollen will be carried between the fields. Additionally, organic 

growers must maintain records to show that production and handling procedures comply 

with USDA organic standards.

C. Soybean Composition

Generally, soybean seed consists of oil (about 20%), protein (about 40%), carbohydrate 

(about 35%) and ash (about 5%) (Liu, 1997). Soybean oil is currently the predominant 

plant oil produced in the world, and is used in a wide variety of food applications (FAO, 

2009; Lusas, 2004). After the oil is extracted the remaining solid materials in the form of 

flakes are toasted and ground to produce soybean meal. 

Oil

For total world vegetable oil production, soybean is ranked number one among the major 

oilseed crops, and in the US supplies 71% of all fats and oils consumed (Soy Stats, 2008). 

Conventional soybean oil is composed of a mixture of several fatty acids. The major 

unsaturated fatty acids in conventional soybean oil are 7% linolenic acid; 51% linoleic 

acid; and 23% oleic acid. It also contains saturated fatty acids 4% stearic acid and 10% 
10

palmitic acid (Codex standard for edible fats and oil, 1996). Soybean oil has a relatively 

high proportion of oxidation prone linolenic acid, which reduces product stability and 

shelf life and which is an undesirable property for the food industry. The hydrogenation 

process (chemical addition of hydrogen) is used to enhance the oil’s stability by reducing 

its polyunsaturated fatty acid content. But this process has undesirable consequences 

including the formation of trans fatty acid isomers and a characteristic “hydrogenated 

flavor” (Fernandez, 1995). Partially hydrogenated oils are used by the food industry 

because they extend product shelf life and have a desirable taste and texture (The 

Pennsylvania State University, 2006). Although use of soybean oil in food is a large part 

of the market for soybean products, soy oil is also used for industrial products as Pioneer 

notes, and modified oils may be used for biodegradable products such as certain types of 

lubricants. 

Meal

Soybean meal, which contains about 50% protein by dry weight, remains the primary 

product from soybean, and of this more than 95% of domestic soybean meal is consumed 

by livestock (ABG, 2007). Only a small proportion of the soybean crop is consumed 

directly by humans. Soybeans are considered to be a source of complete protein. A 

complete protein is one that contains significant amounts of all the essential amino acids 

that must be provided to the human body because of the body’s inability to synthesize 

them. The ten essential amino acids are arginine, histidine, isoleucine, leucine, lysine, 

methionine, phenylalanine, threonine, tryptophan, and valine (Kuiken et al., 1949). 

Cystine is also an important amino acid as it can partially substitute for methionine.

Protein Digestibility Corrected Amino Acids Score (PDCAAS) is considered the gold 

standard for measuring protein quality for humans since 1990. The PDCAAS rating has 

been adopted by the FDA and FAO/WHO as the preferred method to determine protein 

quality (FAO/WHO, 1990). By this criterion, soybean protein was reported to be 

equivalent to animal protein with the highest rating of 1.0 (Hasler, 2002). Humans can 

produce 10 of the 20 amino acids. Soybean contains the other ten essential amino acids

(Kuiken, et al., 1949) that are necessary for human nutrition and are not produced 

naturally in the body. The essential amino acid composition of soybean is included in the 

USDA Nutrition database (USDA-ARS, 2006).

Isoflavones

Soybeans naturally contain isoflavone compounds which are reported to possess 

biochemical activity, including estrogenic and hypocholesterolemic effects. The major 

isoflavones in soybeans include genistein and daidzein. Isoflavones are polyphenol

compounds, produced primarily by beans and other legumes, including peanuts and 

chickpeas. Isoflavones are closely related to the antioxidant flavonoids found in other 

plants, vegetables and flowers (Manach et al., 2004). 

Antinutrients

Soybean grain contains several key antinutrients, such as oligosaccharides, lectins, phytic 

acid and protease inhibitors (OECD, 2001). The trypsin and chymotrypsin inhibitors can 

inhibit digestion of proteins (OECD, 2001). Others such as lectins are sugar-binding 

proteins, preventing digestion of proteins resulting in decreased animal growth (OECD, 
11

2001). The activity of these inhibitors is destroyed during the heat treatment processing 

of the soybean products. The low molecular weight carbohydrates (e.g. stachyose, 

raffinose) can cause flatulence when consumed (Rackis, 1974). Phytic acid binds most of 

the phosphorus in soybean and can chelate various minerals as well (OECD, 2001). 

Livestock producers commonly add phytic acid degrading enzymes to animal feed 

formulas (Vats and Banerjee, 2004).

 

IV. Alternatives

This EA analyzes the potential environmental consequences of a proposal to grant 

nonregulated status to Pioneer 305423 soybean. In order for Pioneer 305423 soybean to 

be granted nonregulated status, APHIS must determine that Pioneer 305423 soybean is 

unlikely to pose a plant pest risk. The analysis by APHIS in its plant pest risk 

assessment (USDA-APHIS, 2009) demonstrates that there was sufficient data to 

determine that Pioneer 305423 soybean is unlikely to pose a plant pest risk and therefore 

is eligible for nonregulated status. 

The regulations at 7 CFR 340.6(d)(3)(i) state that APHIS may "approve the petition in 

whole or in part." Because APHIS has found that Pioneer 305423 soybean is unlikely to 

pose a plant pest risk, the only action alternative considered in this EA is to grant 

nonregulated status “in whole” to Pioneer 305423 soybean. An “in part” deregulation 

can be given if there is a plant pest risk associated with some, but not all lines requested 

in a petition. The petition for Pioneer 305423 soybean only requested APHIS to grant 

nonregulated status to one soybean event, therefore, an “in part” determination is not an 

appropriate consideration. Thus, only two alternatives will be considered in this EA: (1) 

no action and (2) to grant nonregulated status to Pioneer 305423 soybean, “in whole.” 

APHIS has assessed the potential for environmental impacts for each alternative in the 

“Environmental Consequences” sections below. 

A. No Action: Continuation as a regulated article

Under the "no action" alternative, APHIS would deny the petition. Pioneer 305423 

soybeans and progeny derived from them would continue to be regulated articles under 

the regulations at 7 CFR part 340. Permits issued or notifications acknowledged by 

APHIS would still be required for introductions of Pioneer 305423 soybeans and 

measures to ensure physical and reproductive confinement would continue to be 

implemented. APHIS might choose this alternative if there were insufficient evidence to 

demonstrate the lack of plant pest risk from the unconfined cultivation of Pioneer 305423 

soybeans. 

Soybean breeders have achieved soybean oil compositional changes by both conventional 

breeding and genetic engineering (Fehr 2007). Under this no action alternative, growers 

and other parties who are involved in production, handling, processing or consumption of 

soybean would continue to have access to existing deregulated GE high oleic acid 
12

soybean products as well as conventional high or mid level oleic soybean varieties. 

However, growers would not have widespread access to soybean varieties based on 

Pioneer 305423 soybean since it would continue to be regulated under Part 340. There is 

no potential for human consumption of Pioneer 305423 soybean high oleic acid soybean 

under this alternative. 

This alternative is not the preferred alternative because APHIS has already determined 

through a plant pest risk assessment (USDA-APHIS, 2009) that Pioneer 305423 soybean 

is unlikely to pose a plant pest risk. Choosing this alternative would hinder the purpose 

and need of APHIS to allow for the safe development and use of GE organisms given that 

Pioneer 305423 soybean is unlikely to pose a plant pest risk.

B. Grant nonregulated status to Pioneer 305423 soybean, “in whole”- Preferred 

Alternative: Determination that Pioneer 305423 soybean is no longer a regulated 

article.

Under this alternative, Pioneer 305423 soybeans and progeny derived from them would 

no longer be regulated articles under the regulations at 7 CFR part 340. Pioneer 305423 

soybean is eligible for nonregulated status because APHIS has determined that this GE 

organism is unlikely to pose a plant pest risk (USDA-APHIS, 2009). Permits issued or 

notifications acknowledged by APHIS would no longer be required for introductions of 

high oleic acid soybeans derived from this event. APHIS might choose this alternative if 

there was sufficient evidence to demonstrate the lack of plant pest risk from the 

unconfined cultivation of high oleic acid soybeans derived from this event. 

Under this alternative, growers may have future access to Pioneer 305423 soybean and 

progeny derived from this variety if the developer decides to commercialize Pioneer 

305423. In addition, growers and other parties that are involved in production, handling, 

processing or consumption of soybean would continue to be able to use the current high 

or mid level oleic soybean products by conventional breeding as well as the genetically 

engineered soybean variety. Consumers may benefit by having access to a greater range 

of potentially healthful food products. By granting nonregulated status to Pioneer 

305423 soybean, the purpose and need to allow the safe development and use of GE 

organisms is met. 

APHIS has chosen Alternative B as the preferred alternative for the proposed action

because APHIS has determined that Pioneer 305423 soybean is unlikely to pose a plant 

pest risk (USDA-APHIS, 2009). 

Alternatives Considered but Rejected from Further Consideration

Geographic restrictions

APHIS considered geographic restrictions based upon geographic variation in plant pest 

risk. As presented in APHIS plant pest risk assessment for Pioneer 305423 soybean, 

there is no geographic differences in the plant pest risks for Pioneer 305423 soybean 
13

(USDA-APHIS 2009). This alternative was rejected and not analyzed in detail because 

Pioneer 305423 soybean is unlikely to pose a plant pest risk and therefore, APHIS will 

have no regulatory authority over Pioneer 305423 soybean and will be unable to impose 

regulatory restrictions on this GE soybean variety.

V. Environmental Consequences

According to APHIS regulations at 7 CFR part 340, an organism is no longer subject to 

regulatory requirements when it is demonstrated not to present a plant pest risk (USDAAPHIS, 2009). Under the regulations, APHIS is required to render a determination on a 

petition for nonregulated status. The analysis of potential environmental consequences in 

the following sections address the potential impacts to the human environment from the 

alternatives analyzed in this EA, namely taking no action and granting nonregulated 

status to Pioneer 305423 soybean, “in whole”.

SCOPE OF THE ENVIRONMENTAL ANALYSIS

Although the preferred alternative would allow for plantings of Pioneer 305423 soybean

to occur anywhere in the U.S., APHIS limited the environmental analysis to those areas 

that currently support soybean production. To determine areas of soybean production, 

APHIS used data from the National Agricultural Statistics Service (NASS) 2002 Census 

of Agriculture to determine where soybean is produced in the United States (USDANASS, 2009). Only 17 states in 2008 produced more than 1 million acres among the 31 

that produced soybean according to the 2002 Census of Agriculture. These states include 

IA, IL, MN, IN, MO, NE, OH, SD, AR, ND, KS, MI, MS, WI, NC, KY, TN (USDANASS, 2008). Because the oil derived from this Pioneer 305423 soybean will likely 

replace that from existing soybean varieties, it is not likely that new land will be planted 

to soybean by growers. The most recently harvested area of soybean production (72.1 

million acres) is already only one percent below the record high acreage of 2006 (USDANASS, 2008a).

A. No Action

Under the “no action” alternative, Pioneer 305423 soybean would continue to be a 

regulated article. APHIS’ assessment of environmental consequences under the no 

action alternative is described below.

A. Soybean 

Under the ‘no action’ alternative, flowering and reproduction in soybean production 

fields will remain unchanged. DuPont’s GE high oleic soybean variety remains eligible 

for commercial production and Pioneer 305423 will remain a regulated article.

The food/feed nutritional and safety assessment for Pioneer 305423 soybean has been 

reviewed by FDA. Under Federal Food, Drug, and Cosmetic Act (FFDCA), it is the 

responsibility of food and feed manufacturers to ensure that the products they market are 

safe and properly labeled. Food and feed derived from Pioneer 305423 soybean must be 
14

in compliance with all applicable legal and regulatory requirements. In January 2009, 

FDA completed the safety and nutritional assessment for this product and had no further 

questions regarding the safety of Pioneer 305423 soybean (FDA, 2009). 

APHIS assessment of the safety of this product focuses on its potential to pose a plant 

pest risk, and that analysis, is based on the comparison of the GE-soybean to its non-GE 

counterpart (USDA-APHIS, 2009). Based on the assessment of the laboratory evidence 

provided by Pioneer (Pavely, 2007) and accompanying scientific literature (Reference 

section), APHIS has concluded that Pioneer 305423 soybean would have no significant 

impacts on human or animal health

B. Agricultural Production of Soybean

Most of the soybean acreage in the U.S. is planted to GE varieties. Of the total

soybean acres planted in 2008, 92% were GE glyphosate tolerant soybean varieties 

(USDA-NASS, 2008). In 2008, the production of organic soybeans represented about 

0.13% of U.S. soybean production (USDA-NASS, 2010). Conventional production 

practices that use GE varieties (such as herbicide tolerant varieties and new GE traits 

planned by four companies) will likely still dominate in terms of acreage, or perhaps

increase in acreage (Bonny, 2009), even without granting nonregulated status to Pioneer 

305423 soybean under the “no action” alternative. Available seed for both GE and 

conventional varieties will remain the same under the “no action” alternative. Pioneer 

305423 soybean variety will remain unavailable for commercial production. Soybean is 

currently produced in more than 30 states according to the 2008 State Soy Crop Statistics

(USDA-NASS, 2008), and under the “no action” alternative, this area for production will 

likely be unchanged. 

Weed control is one of the biggest challenges for conventional soybean farmers because 

poorly controlled weeds drastically decrease crop yield and quality. Full season 

infestation of weeds can result in soybean yield losses ranging from 12 to 80% 

(Barrentine, 1989). Weeds growing with soybeans compete with the crop for light, 

moisture, and nutrients. Uncontrolled weeds reduce soybean yields and interfere with 

harvest. Soybeans are very competitive with weeds once they develop a canopy, but 

early emerging weeds can cause serious problems. Thus, early-season weed control is 

the key to providing the soybeans with a competitive advantage and minimizing the 

effect of weeds. Crop rotation is one of the most effective ways to manage certain weed 

problems specific to a crop. Herbicides, when used properly, are a safe and effective 

method to control certain weeds in soybeans. 

 

Under the “no action” alternative, herbicides will still be used alone or in combination 

and selected on the basis of their effectiveness on the different weed species in the 

soybean field. Different herbicides have different modes of action; the correct herbicide 

rate must be used for each in order to obtain good weed control results and to minimize 

soybean plant injury. APHIS has no authority under the Plant Protection Act to regulate

herbicide use. The use of herbicides is regulated by the Federal Insecticide,

Fungicide, and Rodenticide Act (FIFRA) restrictions administered by the EPA, which 

mandate registration and use of all pesticides. EPA includes instructions and restrictions 
15

on how herbicides can be applied, and has determined that there is no unreasonable 

environmental risk if the user adheres to the directions. Directions include application 

restrictions that minimize impacts on nearby environments. Violators of the

regulations are liable for all negative consequences of their actions; therefore, farmers 

who use herbicides are very likely to follow its label restrictions, and thereby limiting any 

potential adverse impacts.

The large scale commercial cultivation of GE glyphosate tolerant soybean crop acreage 

has steadily increased from 1996, accounting for 92 % of soybean acreage in 2008. The 

primary reason that farmers have switched to glyphosate-tolerant soybean is the 

simplicity of the weed management programs. Glyphosate is a highly effective, 

nonselective, broad-spectrum herbicide and in general, considered “environmentally 

friendly” when compared to other herbicides (Cerdeira and Duke, 2006). Under the ‘no 

action’ alternative, APHIS expects no change to the availability or use of GE glyphosate 

tolerant soybean varieties. These GE varieties will remain available for commercial use. 

Thus, it is likely that under the ‘no action’ alternative, greater than 90% of the soybean 

acres produced in the U.S. will continue to be GE varieties and production of organic 

varieties will remain unchanged (Bonny, 2009).

If APHIS chooses the no action alternative there would be no direct impact on organic or 

other non-transgenic soybean farmers. The current cultivation practices are unlikely to 

change and 92% of the soybeans produced would likely be planted with the current 

herbicide tolerant biotech soybean varieties. 

C. Soybean Composition

Soybean production for modified fatty acid composition 

The genetic changes in soybeans that resulted in soybean oil composition changes have 

been achieved by both conventional and genetic engineering techniques. Mutagenesis 

was used to develop the major genes for reduced palmitic and linolenic acids that are in 

the cultivars currently grown for commercial production (Fehr, 2007). Conventional 

soybean breeders have obtained varieties with greater than 70% oleic acid by 

intercrossing (Alt et al., 2005). As described in this EA, DuPont (USDA-APHIS, 1997)

has used genetic engineering techniques to increase oleic acid composition in its 

commercially available soybean to near 80%. Therefore, under the ‘no action’ 

alternative, soybean varieties that have modified oil content, either through conventional 

breeding techniques or through genetic engineering, will still be available and on the 

market. Additionally, other seed high in oleic oil (e.g. GE sunflower oil) will also remain 

in commerce. 

A variety of soybean oil choices are presented to food manufacturers, particularly in 

response to the requirement by FDA that those oils with trans-fats be labeled. These 

choices may be altered linolenic acid varieties or interesterified soybean oils (Eckel et al., 

2007). In response to any soybean oils with altered fatty acid ratios that are used in food 

products, substantial efforts in reformulating the entire product may be needed (Eckel et 

al., 2007). Although these specialty oils are marketed exclusively as identity preserved 
16

(see Summary and Section I.B, Taveley, 2007) and are not likely to be found in 

commodity soybean oils, food manufacturers need to be especially attentive to altered 

fatty acid ratios that may be present in commodity oils.

Pioneer proposes to use current industry standard mechanisms to maintain product 

identity, from seed production to delivery to customers after oil refining (Pioneer Public 

Comment APHIS Docket 2007-0156-0045; Regulations.gov (2009). These methods include 

sales of seed for planting to growers under contracts to deliver produced specialty oilseed 

at defined locations, and also to meet standards for oil quality. Other agreements by 

oilseed crushers and refiners could specify precisely the acceptable range of fatty acids 

that would meet established standards for the commodity oil. The Biotechnology 

Industry Organization in cooperation with its member companies is also developing 

stewardship standards for any specialty oilseed products that may differ substantially 

from commodity oils (Jeff Barach, Grocery Manufacturers Association, personal 

communication, 2009).

High oleic oils are found to be useful for industrial products, such as biodegradable 

lubricants (National Ag-Based Lubricants Center, 2008; Machinery Lubrication (2010). 

Such products are used in food processing applications or farm tractor transmissions. 

High oleic oils are used in the industrial sector for base oils that are produced for 

hydraulic or gear oils (National Ag-Based Lubricants Center, 2008). Because properties 

of oils are dependent upon properties of constituent fatty acids, the specific content 

deriving from differing soybean varieties is of interest to these manufacturers. Again, 

specialty soy oils (See Section I.B., Taveley, 2007) are marketed as identity preserved 

products, and are not likely to be found in commodity soybean oils. 

B. Preferred Alternative

Under this alternative, Pioneer 305423 soybean would no longer be a regulated article 

under 7 CFR part 340. Permits issued and/or notifications acknowledged by APHIS 

would no longer be required for introductions of Pioneer 305423 soybean. APHIS has 

chosen the preferred alternative for the proposed action because the Pioneer 305423 

soybean lacks plant pest characteristics, as determined in APHIS’ Plant Pest Risk 

Assessment (USDA-APHIS, 2009). APHIS’ assessment of environmental consequences 

under the preferred alternative is described below.

A. Soybean 

Under this alternative, Pioneer 305423 soybean would be available to growers. A 

potential environmental impact to consider as a result of planting this soybean variety is 

the potential for gene flow to sexually compatible plant species. Based on the plant pest 

risk assessment (USDA-APHIS, 2009), APHIS has determined that Pioneer 305423 

soybean is not a plant pest and that gene flow between this product and weedy and wild 

relatives will not occur in the United States. Weedy and wild relative species do not exist 

naturally in the United States (USDA-NRCS, 2008). Therefore, soybean lacks sexually 

compatible wild relatives in the United States and its territories. APHIS does note that 

gene flow can take place between a field planted with Pioneer 305423 soybean and a 

neighboring soybean crop. The potential for outcrossing can be defined as the ability of 
17

gene escape to other soybean fields. Soybean is a highly self-pollinating species with a 

cross-pollination rate of less than one percent in plants grown in close proximity (OECD, 

2000; Caviness, 1966). Cross-pollination greater than 4.6 m from a pollen source has 

been rarely observed although it has been reported that insects can sometimes transfer the 

pollen that distance or more (Caviness, 1996). Although the biology of the soybean crop 

limits the amount of gene flow that may occur between two soybean fields, soybean 

farmers may implement certain management practices to minimize gene flow between 

production fields (e.g., plant at different dates than neighboring soybean, to encourage 

differing flowering times, establish isolation distances to avoid mechanical mixing and 

short-distance pollen flow).

The food/feed nutritional and safety assessment for Pioneer 305423 soybean has been 

reviewed by FDA. Under Federal Food, Drug, and Cosmetic Act (FFDCA), it is the 

responsibility of food and feed manufacturers to ensure that the products they market are 

safe and properly labeled. Food and feed derived from Pioneer 305423 soybean must be 

in compliance with all applicable legal and regulatory requirements. In January 2009, 

FDA completed the safety and nutritional assessment for this product and had no further 

questions regarding the safety of Pioneer 305423 soybean (FDA, 2009). 

APHIS assessment of the safety of this product focuses on its potential to pose a plant 

pest risk, and that analysis, is based on the comparison of the GE-soybean to its non-GE 

counterpart (USDA-APHIS, 2009). Based on the assessment of the laboratory evidence 

provided by Pioneer (Pavely, 2007) and accompanying scientific literature (Reference 

section), APHIS has concluded that Pioneer 305423 soybean would have no significant 

impacts on human or animal health with overall impacts similar to the no action 

alternative. 

B. Agricultural Production of Soybean

In 2008, GE soybean was planted on 92% of all soybean acres in the US, and the use of 

GE soybean has been steadily increasing over the last 3 years (USDA-NASS 2006, 2007, 

2008). Conventional and GE soybean production occurs on land that is dedicated to crop 

production. Most soybean is planted in agricultural fields that have been in crop 

production for multiple years.

Pioneer 305423 soybean has been field tested under permits issued or notifications 

acknowledged by APHIS in the United States since 2002. Field test sites included the 

major soybean growing areas of the Midwest and the winter nursery areas in Hawaii. 

Agronomic and phenotypic data were collected by the applicant to assess agronomic 

comparability to conventional soybean. To evaluate the agronomic and phenotypic 

characteristics of Pioneer 305423 soybean, the data were collected to address specific 

characteristics that influence reproductive and survival biology. Based on these data, 

APHIS agronomists evaluated the potential for weediness as compared to conventional 

soybean. The evaluation concluded that there were no biologically significant differences 

in weediness potential between Pioneer 305423 soybean and other soybean varieties (see 

Section VIII-A-D and X-C in Pavely, 2007). No differences were found in dormancy or 

germination potential, in responses to diseases or insects, seedling vigor, plant maturity, 
18

plant height, and the lowered linolenic acid content would be correlated with reduced 

cold tolerance. These results show that the agronomic and phenotypic characteristics of 

Pioneer 305423 soybean were either not altered when compared to nontransgenic

(conventional) soybean or were characteristic of less fitness (cold tolerance potential).

Therefore, granting nonregulated status to Pioneer 305423 soybean under the “preferred” 

alternative is not expected to substantially alter the range of soybean cultivation as the 

new GE trait (high oleic acid) does not change the production needs or growth habits of 

GE soybean varieties compared to conventional varieties (USDA-APHIS, 2009).

Pioneer 305423 soybean oil will be marketed as “TREUSTM2

” high oleic soybean oil.

This soybean variety will likely be introduced to areas where soybean is currently grown 

for oil production as an alternative to other choices (such as conventional and GE

varieties producing altered fatty acids) already available in the market (see Summary, 

Pavely (2007)). Therefore, the current distribution of soybean production acreage will 

likely remain unchanged as a result of APHIS granting nonregulated status to Pioneer 

305423 soybean.

Pioneer 305423 soybean was genetically engineered to express a modified soybean 

acetolactate synthase. However, this modified soybean acetolactate synthase was used as 

a selectable marker for transformation and does not confer commercial levels of herbicide 

tolerance in this event. Also, Pioneer 305423 soybean is not genetically engineered for 

tolerance to glyphosate. Because of the herbicide usage similarity of Pioneer 305423 

soybean to conventional soybean, there is no expected change in herbicide use with 

Pioneer 305423 soybean under the preferred alternative. Therefore, potential impacts 

from the use of EPA registered herbicides are the same as those from the no action 

alternative.

Production of mid and high oleic soybean

In 2003, Dupont Protein Technologies provided grower contracts for production of G-94-

1, G-94-19 or G-168 GE high oleic soybean. Production over several thousand acres 

through 2005 revealed no environmental or agronomic issues. Beginning in 2007, 

growers in Iowa planted mid-oleic, low linolenic producing varieties (Stockhausen, 

2008). These varieties, available through the Iowa State University Research Foundation, 

such as IA3036 and IA3039 have oleic acid content of between 49 and 53% (Iowa State 

University, 2010). These lines were grown under contracts with commercial oilseed 

crushers (J. J.G. Minot, personal communication, IA State Univ. Res. Fndtn). No 

agronomic issues were identified, nor impacts on agricultural production noted.

Organic and Conventional Soybean Production

Organic farming operations as described by the National Organic Program, which is 

administered by USDA’s Agricultural Marketing Service, requires organic production 

operations to have distinct, defined boundaries and buffer zones to prevent unintended 

contact with excluded methods from adjoining land that is not under organic 

management. Organic production operations must also develop and maintain an organic 

production system plan approved by their accredited certifying agent. This plan enables 

 2 TREUSTM is a trademark of Pioneer Hi-Bred International, Inc.
19

the production operation to achieve and document compliance with the National Organic 

Standards, including the prohibition on the use of excluded methods. Excluded methods 

include a variety of methods used to genetically modify organisms or influence their 

growth and development by means that are not possible under natural conditions or 

processes. 

Organic certification involves oversight by an accredited certifying agent of the materials 

and practices used to produce or handle an organic agricultural product. This oversight 

includes an annual review of the certified operation’s organic system plan and on-site 

inspections of the certified operation and its records. Although the National Organic 

Standards prohibit the use of excluded methods, they do not require testing of inputs or 

products for the presence of excluded methods. The presence of a detectable residue of a 

product of excluded methods alone does not necessarily constitute a violation of the 

National Organic Standards (USDA-AMS, 2007). The unintentional presence of the 

products of excluded methods will not affect the status of an organic product or operation 

when the operation has not used excluded methods and has taken reasonable steps (such 

as isolation zones, use of buffer rows surrounding the organic crops or adjusting planting 

dates and appropriate cleaning of planting and harvesting equipment) to avoid contact 

with the products of excluded methods as detailed in their approved organic system plan. 

Organic certification of a production or handling operation is a process claim, not a 

product claim. 

It is not likely that farmers, including organic and conventional farmers, who choose not 

to plant transgenic soybean varieties or sell transgenic soybeans varieties, will be 

significantly impacted by the commercial use of Pioneer 305423 soybean. Nontransgenic soybean will likely still be sold and will be readily available to those who wish 

to plant it. An internet search of “Soybean Seed Company” identifies vendors that offer 

all types of soybean seeds for purchase including conventional and transgenic (Vendors, 

2009). 

If Pioneer receives regulatory approval from all appropriate agencies, Pioneer 305423 

soybean will likely be widely available to growers and breeders. Other farmers who 

choose not to plant or sell Pioneer 305423 soybean or other transgenic soybeans will not 

likely be significantly impacted by the expected commercial use of this product because:

(a) non-transgenic soybeans will likely still be sold and will be readily available to those 

who wish to plant these varieties; 

(b) soybean is a highly self-pollinated plant and therefore buffer requirements to isolate 

GE and organic varieties would be minimal; 

(c) 92% of the 2008 soybean acreage in the United States is already planted to transgenic 

herbicide tolerant varieties with organic soybean currently coexisting in the largely 

biotech agricultural environment; and 
20

(d) APHIS expects that Pioneer 305423 soybean may replace some of the presently 

available GE soybean varieties without significantly affecting the overall total soybean 

acreage so organic farmers will be able to coexist with biotech soybean producers as they 

do now. 

If Pioneer 305423 soybean is granted nonregulated status, similar to the no action 

alternative there would be no direct impact on organic or other non-transgenic soybean 

farmers as the market share of transgenic soybean are unlikely to change by the 

introduction of Pioneer 305423 soybean. 

C. Soybean Composition

Soybean is one of the leading agricultural products in the United States. Soybean is 

classified as an oilseed and is characterized by its high (38–45%) protein content as well 

as its high (20%) oil content. The molecular analysis data supplied in the Petition show 

that Pioneer 305423 soybean contains multiple copies of soybean fatty acid desaturase 

gene (Gm--fad2-1) and one modified version of the soybean acetolactate synthase gene 

(Gm-hra). Pioneer 305423 soybean is a transgenic soybean product that provides 

soybean seeds with increased levels of monounsaturated (oleic) fatty acids and decreased 

levels of polyunsaturated fatty acids (linoleic and linolenic). The compositional 

assessment was conducted in accordance with the Organization of Economic Cooperation and Development (OECD) consensus document on compositional 

considerations for new varieties of soybean (OECD, 2001) (Pavely, 2007, p 76). 

On July 9, 2003, the Food and Drug Administration (FDA) announced that it would 

require the labeling of all food products for their trans-fat content beginning January 1, 

2006. As a result of the federal regulation, the food industry in the United States has 

been actively pursuing alternatives to hydrogenated oils so that their products can be 

labeled as containing 0 g of trans-fat. The oil from Pioneer 305423 soybean with high 

oleic acid could be adopted as one of the alternatives to hydrogenated oil.

Soybean seeds both genetically engineered and conventionally breed for altered fatty acid 

composition of the soybean oil have been sold commercially and have a history of safe 

use. The commercialization of Pioneer 305423 soybean could be beneficial for the 

consumer’s health. Increased intake of oils high in monounsaturated fatty acids, such as 

oleic acid have been shown to have positive effects on total cholesterol levels when 

compared to equal intakes of hydrogenated oils (Lichtenstein et al., 2006). Likewise, 

increased intake of oils high in oleic acid can decrease LDL-cholesterol levels compared 

to equal intakes of saturated oils (Mensink et al., 1989) and increased HDL-cholesterol 

levels compared to an equal intake of polyunsaturated oil (Mata et al., 1992). Moderate 

consumption of oil high in oleic acid has also demonstrated decreases in systolic blood 

pressure (Bondia-Pons et al., 2007).

Similar to the no action alternative, a variety of soybean oil choices are presented to food 

manufacturers and for use in industral products. These specialty oils are marketed 

exclusively as identity preserved (see Summary and Section I.B, Taveley, 2007) and will 
21

not likely be found in commodity soybean oils. Pioneer 305423 soybaen would be an 

additional soybean variety available to this specility food oil market. 

Oil Composition

Pioneer 305423 soybean was generated by the insertion of a second copy of a soybean 

fatty acid desaturase gene (Gm-fad2-1) into a publicly available soybean cultivar “Jack”.

The fatty acid desaturase gene is responsible for the synthesis of linoleic acid, which is 

the major polyunsaturated fatty acid present in soybean oil. By silencing3

APHIS has previously granted nonregulated status to a GE high oleic soybean (APHIS 

Petition # 97-008-01p) (USDA-APHIS, 1997) developed by DuPont in which high oleic 

phenotype was conferred by introduction of the soybean omega-6 desaturase gene 1 

(fad2-1 gene). DuPont high oleic soybean varieties have received regulatory approval for 

food use in the Australia, Canada, Japan, New Zealand, and the United States (AgBios, 

2010). Australia New Zealand Food Authority (ANZFA) developed a draft “risk analysis 

report” (ANZFA, 2000) for the DuPont high oleic product. ANZFA concluded that high 

oleic acid soybeans do not raise any public health and safety concerns. Health Canada 

gave DuPont a “no objection to the food use of high oleic soybeans” in 2000 (AgBios, 

2010). 

the fatty acid

desaturase gene, it prevents linoleic acid from being synthesized and leads to the 

accumulation of oleic acid in the seeds. An overview of fatty acid biosynthesis in 

soybean is illustrated in Figure 11, page 41 of the Petition (Pavely, 2007). The intended 

change to Pioneer 305423 soybeans is to greatly increase oleic acid content in the seed 

(Heppard, 1996). Multiple copies of the Gm-fad2-1 gene appear to be necessary for 

effective co-suppression of the endogenous fatty acid desaturase gene 1 (El-Shemy et al.,

2004; Mishra et al., 2005). 

Data that support the safety of GmFad2-1 gene was reviewed by FDA resulting in their 

issuing a FDA Biotech Consultation BNF #000039 (FDA, 1996b). FDA has evaluated 

the food safety of the new proteins in bioengineered plants since 1992 and provided

recommendations concerning its food safety (FDA, 2004). In January, 2009, FDA 

completed the voluntary consultation of the Pioneer 305423 soybean for “Food and Feed 

Safety and Nutritional Assessment” and had no further questions concerning Pioneer 

305423 soybean based on the information provided by Pioneer (FDA, 2009).

Fatty acid compositional data was collected on Pioneer 305423 soybean and comparisons 

were made to conventional control lines and a set of reference soybean varieties (OECD,

2001). A total of 25 fatty acids were analyzed in Pioneer 305423 soybean and control 

lines. Eleven fatty acid concentrations were near or below the detection limits of the 

assay. These fatty acids are listed in Petition (Pavely, 2007, page 79). The analyses of 

other 14 fatty acids are presented in Table 5, Petition (Pavely, 2007, page 81-82). Six of 

the 14 fatty acids were statistically significantly different from control soybean lines: 

myristic acid, palmitoleic acid, stearic acid, arachidic acid, eicosenoic acid and lignoceric 

acid (Pavely, 2007, page 79-82). However, the levels of these fatty acids are not 

 3 Gene silencing refers to a technique for selectively turn off specific genes within a cell.
22

biologically significant; they are relatively minor fatty acids (together comprising less 

than 6% of fatty acid content) and are common fatty acids in vegetable oils at similar 

levels (USDA-ARS, 2006). Two of the 14 fatty acids were not statistically significantly 

different: behenic acid and linoleic acid isomer (9,15). The remaining six fatty acids

(oleic acid, linoleic acid, linolenic acid, palmitic acid, heptadecanoic acid, and 

heptadecenoic acid) are discussed below.

Fatty acids analyses confirmed that Pioneer 305423 soybean has significantly more oleic 

acid (from an average of 21% to 76% of total fatty acid content), less in linoleic acid 

(from an average of 52% down to 3.6%), less in linolenic (9.4% to 5.4%) and less in 

palmitic acid (from an average of 10.3% down to 6.28%) than conventional soybeans. A 

mid-oleic acid soybean variety is offered by Iowa State Research Foundation producing 

more than 50% oleic acid (Stockhausen, 2008). High oleic acid levels are found in other 

commonly consumed premium edible oils (e.g., olive oil 55-83% oleic, Vossen (2007) 

and high oleic sunflower (at least 75% oleic, CODEX, 2009) and high oleic canola oil

(70% oleic, Canolainfo.org, 2010). The consumption of high levels of oleic acid is not 

considered to pose any safety concerns (Lunn, 2007; FDA, 2009). Linolenic acid is 

produced directly from conversion of linoleic acid and therefore the decrease (from an 

average of 9.3% down to 5.4%) in the linolenic acid was directly related to decreased 

linoleic acid levels. Linoleic acid and alpha-linolenic acid are fatty acids the human body 

requires, and cannot be constructed from other components by known chemical 

pathways. They therefore must be obtained from the diet. These two fatty acids are 

widely distributed in plant oils at (e.g. safflower oil, poppy seed oil, walnut oil, olive oil)

and other food sources (USDA-ARS, 2006). The decrease in palmitic acid content in 

Pioneer 305423 soybean is one of the intended effects. Palmitic acid is one of the most 

common saturated fatty acids found in animals and plants. Palmitic acid constitutes 

between 22 and 30 percent of most animal fats and is also an important constituent of 

most vegetable fats (2-17 percent, but 35-45 percent of palm oil) (Table 1, Nishina et al., 

1993; Beare-Rogers et al., 2001).

Pioneer also noted increased levels of two minor fatty acids, heptadecanoic acid and 

heptadecenoic acid (Pavely, 2007, pp. 80-82). These increases likely result from changes

in the GM-HRA protein that shifts a metabolic pathway leading to production of the C174

fatty acids. Pioneer describes this complex oil biosynthesis in the Petition (Pavely, 2007, 

Appendix 7, pp. 171-172). The combined value of heptadecanoic and heptadecenoic 

fatty acids in control soybean line is less than 0.2% of total fatty acids. In Pioneer 

305423 soybean, the combined value of these two fatty acids is about 2% of total fatty 

acids. Heptadecanoic acid is commonly found in meat, tofu and butter (USDA-ARS,

2006), and heptadecenoic acid is commonly found in meat (Senaratne, 2009) and tofu

(USDA-ARS, 2006). The levels of these two fatty acids in Pioneer 305423 soybean are 

comparable to those already found in the diet and there is no evidence to indicate that 

exposure to either fatty acid through dietary sources would have adverse effects in 

humans.

 4 C17 fatty acids are fatty acids containing 17 carbon atoms (e.g. heptadecanoic acid and heptadecenoic 

acid). 
23

Soybean Meal Composition

Amino Acids

Soybean meal is fed to animals primarily as a source of protein. Amino acid content in 

Pioneer 305423 soybean was determined for 18 amino acids. APHIS reviewed and 

analyzed the data presented in the petition, and concludes that there are no significant 

differences in amino acid composition between Pioneer 305423 soybean and control

(non-GE) soybean lines (Pavely, 2007).

Novel protein

The inserted Gm-hra gene is a slightly modified version of the soybean acetolactate 

synthase gene (als gene) that is responsible for tolerance to ALS inhibiting herbicides. 

The Gm-hra gene encodes the GM-HRA protein with two amino acid residues modified 

from the endogenous enzyme. In Pioneer 305423 soybean, GM-HRA is the only novel 

protein expressed and was solely used as a selectable marker during the transformation 

procedure. The HRA fragment in Pioneer 305423 soybean does not confer commercial 

levels of sulfonylurea or other ALS (acetolactate synthase) inhibitor herbicide tolerance. 

Pioneer has no plan to commercially market Pioneer 305423 as a sulfonylurea herbicide 

tolerant variety.

ALS (acetolactate synthase) proteins are present in nature, as als genes have been isolated 

from bacteria, fungi, algae and plants (Friden et al., 1985; Falco et al., 1985; Reith et al.,

1995; Mazur et al., 1987). Several commercialized non-GE crops (Clearfield trade mark) 

with the herbicide tolerant als gene are available in the current market. Data that support 

the safety of modified soybean acetolactate synthase proteins have been reviewed by 

FDA in the FDA Biotech Consultation BNF #000110 (FDA, 2009) for GE Pioneer 

soybeans Petition #06-354-01p (Pavely, 2007). FDA Biotech Consultation BNF #000050 

(FDA, 1998) for GE flax, FDA Biotech Consultation BNF #000030 (FDA, 1996a) for GE 

cotton, FDA Biotech Consultation BNF #000108 (FDA, 2007) for GE Pioneer soybeans 

Petition #06-271-01p (USDA-APHIS, 2006).

Pioneer also assessed the acute toxicity in mice of a dose of 2000 mg purified GM-HRA 

protein per kilogram of body weight (Pavely 2007, pp 62-63). Even at this high dosage, 

no clear threshold of acute toxicity in mice was reached. Since GM-HRA protein is 

expressed at such a low level in Pioneer 305423 soybean, the amount of these soybeans 

that would need to be consumed to cause harm in mammals would be outside the limits 

of any realistic scenario for consumption. It is clear there is a wide margin of safety for 

GM-HRA.

Based on Pioneer's data and information in Petition, the recently published 

Environmental Assessment for APHIS Petition # 06-271-01p for GE Pioneer soybeans 

(USDA-APHIS 2006) and results of multiple previous FDA consultations, there is no 

indication of risk from GM-HRA protein due to exposure or other environmental safety 

concerns under the preferred alternative. GM-HRA protein was also assessed by the 

applicant for possible allergenicity and toxicity using internationally accepted guidance 

from the Codex Alimentarius Commission. APHIS reviewed this information (Pavely,
24

2007, p62) and concludes that GM-HRA proteins are unlikely to be either allergenic or 

toxic to humans or animals. 

Proximate, isoflavones and antinutrients 

Data on proximate and fiber in soybean grain was provided to APHIS in the petition 

(Pavely, 2007). Proximate analysis is a chemical method of assessing and expressing the 

nutritional value of a feed. Soybeans are occasionally used as an alternative forage 

source when alfalfa or clovers are in short supply. In 2001, an OECD consensus 

document on compositional considerations for new soybean varieties, suggested

parameters that soybean developers should measure. The proximate nutrient content, 

including crude protein, crude fat, fiber, and ash content of soybean meal is one of the 

parameters. APHIS reviewed this information and concludes that the ranges for crude 

protein, fat, ash, neutral detergent fiber (NDF) and acid detergent fiber (ADF) of Pioneer

305423 soybean are comparable to nontransgenic soybean lines or reference soybean 

lines. APHIS also concludes that proximate and fiber analysis of soybean forage and 

grain samples demonstrate that there is no unexpected difference between Pioneer 

305423 soybean and control (non-GE) soybean lines.

Antinutrients are compounds which decrease the nutritional value, usually by making an 

essential nutrient unavailable or indigestible when consumed by humans/animals. 

Soybean seeds are known to contain different compounds displaying antinutrient effects. 

Soybean grain contains several key antinutrients, such as oligosaccharides, trypsin 

inhibitors, lectins and phytic acid. APHIS reviewed the information on the key 

antinutrients stachyose, raffinose, lectins, phytic acid and trypsin inhibitor presented in 

the petition for Pioneer 305423 soybean and the control (non-GE) soybean lines and 

concludes there were no statistically significant differences were seen between Pioneer 

305423 soybean and control (non-GE) soybean lines. 

APHIS also reviewed the information presented on isoflavones and concludes there were 

no statistically significant differences observed between Pioneer 305423 soybean and 

control (non-GE) soybean lines. 

APHIS has concluded that Pioneer 305423 soybean, and the foods and feeds derived 

from it are not materially different in safety, composition, or any other relevant parameter 

from soybeans now grown, marketed, and consumed. Results of these comparisons 

indicate that Pioneer 305423 soybean is compositionally and nutritionally equivalent to 

conventional soybean varieties currently in commerce except for the intended changes in 

the fatty acid profiles.

Cumulative effects

APHIS considered whether the proposed action could lead to significant cumulative 

impacts, when considered in light of other past, present, and reasonably foreseeable 

future actions, regardless of what agency or person initiated such actions. As mentioned 

above, Pioneer 305423 soybean is not the first high oleic soybean product to be granted 
25

nonregulated status. APHIS has previously made determinations of nonregulated status 

for DuPont high oleic soybean in the market currently. 

APHIS has evaluated the potential cumulative impacts of granting nonregulated status to 

Pioneer 305423 soybean. In 2008, GE soybean was planted on 92% of all soybean acres 

in the US, and the use of GE soybean has been steadily increasing over the last 3 years 

(USDA-NASS, 2006, 2007, 2008). Conventional and GE soybean production occurs on 

land that is dedicated to crop production. Most soybean is planted on agricultural fields 

that have been in crop production for years. Pioneer 305423 soybean will not affect the 

amount of acreage devoted to GE varieties, nor will Pioneer 305423 soybean alter the 

growing regions available for soybean production. There would be no direct impact on 

organic or other non-transgenic soybean farmers as the market share of transgenic 

soybean are unlikely to change by the introduction of Pioneer 305423 soybean. Pioneer 

305423 is not herbicide tolerant at commercial herbicide application rates. Therefore, 

APHIS has no reason to believe that there will be any substantive change in current 

herbicide use rates as a result of granting nonregulated status to Pioneer 305423. 

In 2003, several thousand acres of DuPont high oleic soybeans were grown under 

contract in Iowa for DuPont Protein Technologies. There were no known or reported 

environmental impacts due to the production of this high oleic soybean variety. This 

DuPont high oleic soybean variety has not been planted since 2005 due to changes in 

market strategy for DuPont.

 

Based on this information, APHIS has determined that there are no past, present, or 

reasonably foreseeable actions that would aggregate with effects of the proposed action to 

create cumulative impacts or reduce the long-term productivity or sustainability of any of 

the resources associated with the ecosystem in which Pioneer 305423 soybean is planted. 

Potential impact on non-target organisms, including beneficial organisms 

and threatened or endangered species

Unintended effects on non-target organisms are one of the safety concerns considered for 

genetically engineered foods, including the potential for altered expression of plant 

produced toxicants and anti-nutrients or the formation of novel toxins. Generally, 

unintended effects can be predicted or explained through our current knowledge of plant 

biology and metabolic pathways. Molecular and biochemical analysis data can also help 

the risk assessors to determine the levels of transcriptional and translational changes. 

APHIS reviewed and analyzed the data presented for Pioneer 305423 soybean and 

evaluated the potential of this GE soybean variety to cause damage or have toxic effects 

directly or indirectly on non-target organisms.

Toxcity and Allergencity

The only novel protein expressed in Pioneer 305423 soybean is GM-HRA. The potential 

toxicity of the GM-HRA protein was assessed by comparing the amino acid sequence of 

the GM-HRA protein with known protein toxins in the bioinformative database (NIHNCBI, 2010) as well as in an acute toxicity feeding study in mice (Pavely, 2007, p 63).
26

The study found no evidence of acute toxicity in mice fed with high doses of purified 

GM-HRA protein (Pavely, 2007, p. 63). Since Pioneer 305423 soybean was proposed for 

use as a food/feed product, there is a wide safety margin taken into account (by the 

dosage range) in the acute toxicity feeding study. APHIS concludes, after reviewing the 

data presented, including FDA’s food safety and nutritional assessment, that there will 

not be any toxic effect at any feasible level of consumption.

The information on the possible allergenicity of Pioneer 305423 soybean was reviewed 

and analyzed by APHIS. GM-HRA protein is not known to be a protein toxin/allergen 

based on the information in the database of the Food Allergy Research and Resource 

Program (FARRP), University of Nebraska, Allergen Database (Version 6.0, January 

2006). Likewise, the rapid proteolytic degradation (less than 30 seconds) under 

simulated mammalian digestion conditions provides further evidence to confirm the 

safety of the protein (FAO, 1996). In addition, Pioneer provided a detailed assessment of 

human and animal safety assessment of the GM-HRA protein to FDA as part of the 

consultation on food and feed safety for Pioneer 305423 soybean (FDA, 2009). FDA had 

no further questions on the safety of Pioneer 305423 soybean.

Nutrition

Nutritional studies on targeted animals are performed in cases where the composition of 

the GE plant differs significantly from the non-GE counterpart. Pioneer conducted a 42-

day feeding study on broiler chickens comparing Pioneer 305423 soybean and nontransgenic soybean as the main diets. Broilers are the choice animal for feeding studies 

because they are fast growing and are particularly sensitive to the presence of toxic 

elements in their feed. APHIS evaluated the results of feeding tests on broilers (Pavely,

2007) and concluded that there is no difference between Pioneer 305423 soybean and 

traditional soybean variety counterparts, and Pioneer 305423 soybean can be considered 

as nutritionally equal to the non-GE counterparts (Kuiper et al., 1999). Therefore, when 

compared to conventional soybean varieties, Pioneer 305423 soybean is not expected to 

have any adverse effects on birds and animals feeding in the field.

Pest and Disease

Plant protease inhibitors have been determined to play a potent defensive role against 

certain pests and pathogens of soybeans (Boulter, 1993; Hilder et al., 1987; Williamson et 

al., 1996; Joshi et al. 1998) as described in the Plant Pest Risk Assessment (USDAAPHIS, 2009, p 3-5). The reduced protease activity could potentially result in Pioneer 

305423 soybean being more susceptible to insect damage or fungal disease than the nonGE control lines. Pioneer provided data (Pavely, 2007, pp. 158-160) to support the 

conclusion that Pioneer 305423 soybean is comparable to non-GE control lines in 

environmental fitness and defense against pests and diseases. The mean value for trypsin 

inhibitor was significantly different (lower; adjusted P-value < 0.05) in Pioneer 305423 

soybean as compared to the control line although remaining within the statistical 

tolerance interval and the “combined literature range.” APHIS reviewed the information 

and concludes that reducing the protease activities of the Pioneer 305423 soybean is not 

expected to have any effect on insects or threatened and endangered insect species 

feeding in a soybean field (Pavely, 2007, pp. 158-160). 
27

Soil Communities

APHIS also evaluated the effects of production of Pioneer 305423 soybean on soil 

communities in agricultural settings. In this case the genetic modification does not confer 

herbicide tolerance or insect resistance. Therefore, the potential pesticide (herbicide and 

insecticide) usage should be similar as that for conventional soybean plantings. 

Currently, there are over 70 registrations for herbicides for weed management in soybean 

(Gianessi et al., 2002), plus numerous mixtures (Crop Protection Reference, 2009). In 

planting Pioneer 305423 soybean, the pesticide or herbicide runoff into ground and 

surface water should have no more deleterious effects on non-target organisms, including 

aquatic animals or aquatic invertebrates in ponds and streams than conventional 

plantings. Reports issued by the OECD (OECD, 1986) and a working group coordinated 

by the Royal Society of London (Royal Society, 2000) have indicated that environmental 

risks of biotechnology-derived crops are not fundamentally different from risks of 

conventionally derived products. Therefore, cultivation of Pioneer 305423 soybean 

should have no effects on either the microbial or invertebrate communities of soil 

ecosystems when compared to current agricultural practices for soybean. 

Threatened and Endangered Species

Using the information pertaining to the potential impacts to non-target organisms 

described above, APHIS also considered the potential impact of deregulating Pioneer 

305423 soybean on federally listed threatened or endangered species (TES) and species 

proposed for listing [http://ecos.fws.gov/tess_public/SpeciesReport.do (accessed April 

10, 2009)], as well as designated critical habitat and habitat proposed for designation, as 

required under Section 7 of the Endangered Species Act. In this analysis, APHIS 

considered the biology of Pioneer 305423, and how its production would differ from 

typical agricultural practices associated with cultivation of soybean.

To identify negative effects or significant impacts on TES animal species, APHIS 

evaluated the risks to TES animals from consuming Pioneer 305423 soybean. Risk is a 

combination of hazard and exposure. APHIS first conducted hazard identification for 

Pioneer 305423 soybean. APHIS reviewed and analyzed the composition and nutritional 

quality of Pioneer 305423 soybean submitted in the petition, and compared the 

composition of Pioneer 305423 to the composition of a non-genetically engineered 

control soybean line and the natural variation found in commercial soybean varieties

(Pavely, 2007). If the composition of Pioneer 305423 soybean is similar to other 

commercial soybean plants, it is unlikely that Pioneer 305423 poses a hazard to TES 

animal species. If no hazards are identified, then the risk of Pioneer 305423 soybean 

harming TES animal species is also unlikely, regardless of exposure. 

As discussed in this EA, the proteins produced by the inserted genes and the changes in 

fatty acid composition do not raise safety issues. As noted above in this section, 

consumption of GM-HRA protein has shown no toxicity in lab testing with mice, and no 

measurable change in nutritional response on broiler chickens. The Pioneer 305423 

soybeans do not express additional proteins, natural toxicants, allelochemicals, 

pheromones, hormones, etc. that could directly or indirectly affect a listed TES or species 
28

proposed for listing. Data submitted on the composition of the Pioneer 305423 soybeans 

indicate that these soybeans are not significantly different from non-transgenic soybeans 

and would not be expected to have any effect on TES that would be different from nontransgenic soybeans. The Pioneer 305423 soybean is not sexually compatible with a 

federally listed TES or a species proposed for listing. APHIS has not identified any 

stressor caused by the production of Pioneer 305423 that could affect the reproduction, 

numbers, or distribution of a listed TES or species proposed for listing. Consequently, an 

exposure analysis for individual species is not necessary.

Soybeans do not grow and persist in unmanaged habitats and would not be expected to 

invade and/or persist in the natural environment. Soybean fields are typically highly 

managed agricultural areas that can be expected to be dedicated to crop production for 

many years and cultivation of Pioneer 305423 soybean is not expected to differ from 

typical soybean cultivation. After reviewing possible effects of deregulating Pioneer 

305423 soybean, APHIS expects Pioneer 305423 soybean to replace some of the 

presently available soybean varieties, but APHIS does not expect that Pioneer 305423 

soybean will cause new soybean acres to be planted in areas that are not already devoted 

to agriculture. APHIS has considered the effect of Pioneer 305423 soybean production 

on habitat that could be used by TES, including critical habitat, and could identify no 

difference from affects that would occur from the production of other soybean varieties. 

Based on the above information, APHIS has determined that the preferred alternative, 

deregulating Pioneer 305423 soybean would have no effect on federally listed threatened 

or endangered species and species proposed for listing, or on designated critical habitat or 

habitat proposed for designation. Consequently, a written concurrence or formal 

consultation with the USFWS is not required for this action.

Consideration of Executive Orders, Standards and Treaties relating to 

environmental impacts

Executive Order (EO) 12898 (US-NARA, 2008), “Federal Actions to Address 

Environmental Justice in Minority Populations and Low-Income Populations”, 

requires Federal agencies to conduct their programs, policies, and activities that 

substantially affect human health or the environment in a manner so as not to exclude 

persons and populations from participation in or benefiting from such programs. It also 

enforces existing statutes to prevent minority or low-income communities from being 

subjected to disproportionately high and adverse human health or environmental effects. 

EO 13045 (US-NARA, 2008), “Protection of Children from Environmental Health 

Risks and Safety Risks”, acknowledges that children may suffer disproportionately from 

environmental health and safety risks because of their developmental stage, greater 

metabolic activity levels, and behavior patterns, as compared to adults. The EO (to the 

extent permitted by law and consistent with the agency’s mission) required each Federal 

agency to identify, assess, and address environmental health risks and safety risks that 

may disproportionately affect children. 
29

Each alternative was analyzed with respect to EO 12898 and 13045. Based on the 

information submitted by the applicant and assessed by APHIS, Pioneer 305423 soybean 

is not significantly different than conventional soybean and has successfully completed 

the FDA voluntary consultation for food and feed use. Therefore, Pioneer 305423 

soybean is not expected to have a disproportionate adverse effect on minorities, lowincome populations, or children. 

EO 13112 (US-NARA, 2008), “Invasive Species”, states that Federal agencies take 

action to prevent the introduction of invasive species, to provide for their control, and to 

minimize the economic, ecological, and human health impacts that invasive species 

cause. Both non-GE and GE soybean varieties that have been granted nonregulated 

status are widely grown in the U.S. Based on historical experience with soybean and the 

data submitted by the applicant and assessed by APHIS, Pioneer 305423 soybean plants 

are very similar in fitness characteristics to other soybean varieties currently grown and 

are not expected to become weedy or invasive [see (USDA-APHIS, 2009) for the plant 

pest risk assessment of Pioneer 305423 soybean]. 

EO 13186 (US-NARA, 2008), “Responsibilities of Federal Agencies to Protect 

Migratory Birds”, states that Federal agencies taking actions that have, or are likely to 

have, a measurable negative effect on migratory bird populations are directed to develop 

and implement, within 2 years, a Memorandum of Understanding (MOU) with the Fish 

and Wildlife Service that shall promote the conservation of migratory bird populations. 

Data submitted by the applicant has shown no difference in compositional and nutritional 

quality of Pioneer 305423 soybean compared to conventionalsoybean, apart from the 

presence of Pioneer 305423 soybean. Pioneer also conducted feeding experiments on 

broiler chickens to evaluate the effects of Pioneer 305423 soybean on birds (page 15 of

EA). The applicant reported no harmful effects to chickens from Pioneer 305423 

soybean. The migratory birds that occasionally forage in soybean fields are unlikely to 

contain high amounts of Pioneer 305423 soybean as soybean availability is limited by 

seed germination and harvest. Based on APHIS’ assessment of Pioneer 305423 soybean 

it is unlikely that granting nonregulated status to this soybean variety will have a negative 

effect on migratory bird populations. 

INTERNATIONAL IMPLICATIONS 

EO 12114 (US-NARA, 2008), “Environmental Effects Abroad of Major Federal 

Actions”, requires Federal officials to take into consideration any potential 

environmental effects outside the U.S., its territories, and possessions that result from 

actions being taken. APHIS has given this due consideration and does not expect a 

significant environmental impact outside the U.S. should nonregulated status be 

granted to Pioneer 305423 soybean. It should be noted that all the considerable, 

existing national and international regulatory authorities and phytosanitary regimes that 

currently apply to introductions of new soybean cultivars internationally, apply equally 

to those covered by an APHIS determination of nonregulated status under 7 CFR part 

340. Any international trade of Pioneer 305423 soybean subsequent to a determination 

of nonregulated status for the product would be fully subject to national phytosanitary 

requirements and be in accordance with phytosanitary standards developed under the 
30

International Plant Protection Convention (IPPC, 2008). 

The purpose of the IPPC “is to secure a common and effective action to prevent the 

spread and introduction of pests of plants and plant products and to promote appropriate 

measures for their control” (IPPC, 2008); the protection it affords extends to natural flora 

and plant products and includes both direct and indirect damage by pests, including 

weeds. The IPPC set a standard for the reciprocal acceptance of phytosanitary 

certification among the nations that have signed or acceded to the Convention (169 

countries as of September 2008). In April 2004, a standard for pest risk analysis (PRA) 

of living modified organisms (LMOs) was adopted at a meeting of the governing body of 

the IPPC as a supplement to an existing standard, International Standard for 

Phytosanitary Measure No. 11 (ISPM-11, Pest Risk Analysis for Quarantine Pests). The 

standard acknowledges that all LMOs will not present a pest risk and that a determination 

needs to be made early in the PRA for importation as to whether the LMO poses a 

potential pest risk resulting from the genetic modification. APHIS pest risk assessment 

procedures for genetically engineered organisms are consistent with the guidance 

developed under the IPPC. In addition, issues that may relate to commercialization and 

transboundary movement of particular agricultural commodities produced through 

biotechnology are being addressed in other international forums and through national 

regulations. 

The Cartagena Protocol on Biosafety is a treaty under the United Nations Convention 

on Biological Diversity (CBD) that established a framework for the safe transboundary 

movement, with respect to the environment and biodiversity, of LMOs, which includes 

those modified through biotechnology. The Protocol came into force on September 11, 

2003, and 150 countries are Parties to it as of January, 2009 (CBD-UN, 2008). Although 

the U.S. is not a party to the CBD, and thus not a party to the Cartagena Protocol on 

Biosafety, U.S. exporters will still need to comply with domestic regulations that 

importing countries that are Parties to the Protocol have put in place to comply with their 

obligations. The first intentional transboundary movement of LMOs intended for 

environmental release (field trials or commercial planting) will require consent from the 

importing country under an advanced informed agreement (AIA) provision, which 

includes a requirement for a risk assessment consistent with Annex III of the Protocol, 

and the required documentation. 

LMOs imported for food, feed, or processing (FFP) are exempt from the AIA procedure, 

and are covered under Article 11 and Annex II of the Protocol. Under Article 11 Parties 

must post decisions to the Biosafety Clearinghouse database on domestic use of LMOs 

for FFP that may be subject to transboundary movement. To facilitate compliance with 

obligations to this protocol, the U.S. Government has developed a website that provides 

the status of all regulatory reviews completed for different uses of bioengineered products 

(NBII, 2008). These data will be available to the Biosafety Clearinghouse. APHIS 

continues to work toward harmonization of biosafety and biotechnology consensus 

documents, guidelines, and regulations, including within the North American Plant 

Protection Organization (NAPPO), which includes Mexico, Canada, and the U.S., and 

within the Organization for Economic Cooperation and Development. NAPPO has 

completed three modules of a standard for the Importation and Release into the 
31

Environment of Transgenic Plants in NAPPO Member Countries (NAPPO, 2008). 

APHIS also participates in the North American Biotechnology Initiative (NABI), a 

forum for information exchange and cooperation on agricultural biotechnology issues for 

the U.S., Mexico and Canada. In addition, bilateral discussions on biotechnology 

regulatory issues are held regularly with other countries including: Argentina, Brazil, 

Japan, China, and Korea. 

COMPLIANCE WITH CLEAN WATER ACT AND CLEAN AIR ACT 

This Environmental Assessment evaluated the changes in soybean production due to the 

unrestricted use of Pioneer 305423 soybean. Pioneer 305423 soybean will not lead to the 

increased production of soybean in U.S. agriculture. There is no expected change in water 

use due to the production of Pioneer 305423 soybean compared to current soybean 

production regimes, nor is it expected that air quality will change due to do the 

production of Pioneer 305423 soybean. 

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10/2007)

VII. PREPARERS AND REVIEWERS 

Michael Gregoire, Deputy Administrator 

Biotechnology Regulatory Services 

Beverly Simmons, Associate Deputy Administrator for Emerging and International 

Programs

Sidney Abel, Assistant Deputy Administrator 

BRS, Environmental Risk Analysis Division

Michael Watson, Ph.D., Division Director 

Margaret Jones, Ph.D., Senior Biotechnologist (Reviewer) 

Karen Green, Biotechnologist (Preparer of Environmental Assessment) 

Luc Hebou, Biotechnologist (Reviewer) 

BRS, Policy and Coordination Division 
40

John Turner, Ph.D., Director

Andrea Huberty, Ph. D., Branch Chief (Reviewer)

Richard Coker, Regulatory Analyst

Michael Blanchette, Environmental Protection Specialist (Reviewer)

AGENCY CONTACT 

Ms. Cynthia Eck, Document Control Officer 

USDA, APHIS, BRS 

4700 River Road, Unit 147 

Riverdale, MD 20737-1237 

Phone: (301) 734-0667 

Fax: (301) 734-8669 

Cynthia.A.Eck@aphis.usda.gov

VIII. Appendix 1 - Biotechnology Consultation Note to

the File BNF No. 000110

Date: January 14, 2009

Subject: High Oleic 305423 Soybean

Keywords: soybean, Glycine max, high oleic 305423 soybean, TREUSTM, OECD unique 

identifier DP-3Ø5423-1, omega-6 desaturase, FAD2-1, gm-fad2-1, seed-specific 

silencing, gm-hra, soybean acetolactate synthase

1. Introduction

In a submission dated December 27, 2006, Pioneer Hi-Bred International, Inc. (Pioneer), 

a DuPont Company, submitted to the Food and Drug Administration (FDA) a safety and 

nutritional assessment of the bioengineered high oleic 305423 soybean line containing 

the transformation event DP-3Ø5423-1. Pioneer provided additional information on 

August 31, 2007, January 30, February 19, March 18, November 21, December 12, and 

December 19, 2008. Pioneer concluded that food and feed derived from the 305423 

soybean are as safe and nutritious as food and feed derived from conventional soybean 

varieties currently being marketed.

2. Intended Effect

The intended effect of the modification in 305423 soybean is to produce soybean seeds 

with increased levels of monounsaturated fatty acid (oleic) and decreased levels of 

polyunsaturated fatty acids (linoleic and linolenic). To accomplish this objective, Pioneer 

inserted a fragment of the soybean microsomal omega-6 desaturase gene (FAD2-1) into 
41

soybean. The fragment of the FAD2-1 gene does not code for a protein. Transcription of 

the gene fragment under the control of a seed-preferred KTi3 promoter acts to silence the 

expression of the endogenous soybean omega-6 desaturase, which results in an increased 

level of oleic acid and decreased levels of linoleic and linolenic acids in the soybean seed. 

A gene (gm-hra) encoding a modified version of the soybean acetolactate synthase (als

gene) that confers tolerance to a sulfonylurea herbicide was used as a selectable marker 

for the transformation.

3. Genetic Modifications and Characterization

3.1. Parental Variety

The publicly available soybean variety "Jack" was used as the recipient in the DNA 

transformation to create 305423 soybean.

3.2. Transformation DNA and Method

Pioneer used microprojectile bombardment to co-transform secondary plant cell embryos 

with two purified linear DNA fragments: a 2924 base pair fragment containing the gmfad2-1 cassette, and the 4512 base pair fragment containing the gm-hra cassette. The gmfad2-1 cassette includes the promoter region from the soybean Kunitz trypsin inhibitor 

gene (KTi3), a fragment of the FAD2-1 gene that corresponds to approximately 40% of 

the middle portion of the FAD2-1 gene, and the 3' untranslated region of the KTi3 gene 

(KTi3 terminator).

The gm-hra cassette includes a promoter and an intron from the 5'regulatory region of the 

S-adenosyl-L-methionine synthetase (SAMS) gene from soybean, the gm-hra gene that 

encodes the GM-HRA protein, and the terminator from the endogenous als gene.

1

3.3. Characterization, Stability, and Inheritance of the Introduced DNA

In order to characterize the introduced DNA, Pioneer conducted Southern blot and 

sequencing analyses of the DNA inserted into the 305423 soybean. Pioneer reports that 

the 305423 soybean contains four genetically linked insertions. The 305423 soybean has 

multiple intact and partial copies of the gm-fad2-1 cassette that contain, in total, eight 

copies of the KTi3 promoter, seven copies of the gm-fad2-1 gene fragment, and five 

copies of the KTi3 terminator. Pioneer states that it appears that multiple copies of the 

gm-fad2-1 gene fragment are necessary for effective co-suppression of the endogenous 

gene. One copy of the KTi3 promoter is associated with a small non-functional fragment 

of plasmid backbone DNA. Pioneer determined that a single intact gm-hra cassette is 

inserted into the genome of 305423 soybean. Pioneer noted that further analysis of 

305423 soybean for plasmid backbone sequence using Southern hybridization showed 

that 305423 soybean contains neither the hygromycin gene nor the bacterial plasmid 

origin of replication present on the plasmids from which the gm-fad2-1 cassette and the 

gm-hra cassette are derived. Pioneer sequenced the inserted DNA and reported that the 

sequence confirmed the results of the Southern analysis.
42

Each of the four insertions in 305423 soybean was screened for the presence of open 

reading frames (ORFs) containing both a start and stop codon that spanned any novel 

junctions. Pioneer identified two such ORFs. Pioneer reports that neither ORF contains 

the necessary regulatory elements for transcription. Northern blot analysis detected no 

transcripts of the ORFs in developing seeds from either 305423 soybean or the control 

soybean. Pioneer concluded that it is very unlikely that a protein is expressed from either 

ORF. Pioneer also reports that screening of the ORFs against a database containing 

known protein toxins2 and to a database of known allergens3 showed no biologically 

significant identities to known protein toxins or allergens. Based upon their analyses, 

Pioneer concluded that there are no safety concerns resulting from these ORFs.

Pioneer reports that Southern blot analyses across three generations showed that the 

inserted DNA in 305423 soybean is stably integrated into the genome. The same eventspecific hybridization pattern was observed for all but one plant that apparently lost the 

gm-hra cassette due to a recombination event. Pioneer investigated the frequency of 

recombination in 305423 soybean by examination of 1000 additional segregating 305423 

soybean plants by PCR-based assays and found no other recombinants.

Pioneer conducted chi square analysis of trait inheritance data. They report that the 

expected segregation ratios were observed in crosses showing the Mendelian inheritance 

and stability of the introduced trait. Pioneer reports that when both traits were analyzed in 

the same plants, data confirmed co-segregation of the gm-fad2-1 gene fragment and the 

gm-hra gene.

4. Introduced Protein – GM-HRA

4.1. Identity, Function, and Characterization

Pioneer describes the GM-HRA protein as a modified version of the endogenous soybean 

acetolactate synthase (ALS) enzyme and that this modified ALS enzyme confers 

tolerance to ALS-inhibiting herbicides. ALS enzymes are widely distributed in nature and 

have been isolated from bacteria, fungi, algae, and plants. ALS-inhibiting herbicides 

inhibit plant growth by blocking the action of the ALS enzyme thereby inhibiting 

branched-chain amino acid biosynthesis. The mature GM-HRA protein differs from the 

endogenous ALS enzyme at two specific amino acids and is responsible for GM-HRA 

insensitivity to ALS-inhibiting herbicides. The gm-hra gene was used only as a selectable 

marker for the transformation.

Pioneer characterized 305423 soybean-produced GM-HRA protein using various 

methodologies4 and demonstrated its equivalence with Escherichia coli-produced GMHRA protein. The E. coli-derived GM-HRA protein was used for in vitro and in vivo

biochemical and toxicological studies.

The GM-HRA protein levels in 305423 soybean were measured in replicated samples of 

leaf, root, forage and grain tissues using a quantitative enzyme-linked immunosorbent 

assay (ELISA). Pioneer reports that the mean GM-HRA protein concentrations in 305423 

soybean leaf, root, forage, and grain were 4.0, 0.18, 5.7, and 2.5 nanograms per milligram 
43

(ng/mg) tissue dry weight respectively. Pioneer concluded that the above results confirm 

that the expression of the GM-HRA protein in 305423 soybean is constitutive.

4.2. Assessment of Potential Toxicity and Allergenicity

Pioneer reports the results of a global sequence similarity search of the GM-HRA amino 

acid sequence against the National Center for Biotechnology Information (NCBI) Protein 

dataset. The search was conducted using the BLASTP algorithm.5 GM-HRA showed 

sequence similarity to other ALS proteins. None of the proteins returned by the search 

was identified as a toxin. Pioneer concluded that the GM-HRA protein did not share 

relevant sequence similarities with known protein toxins.

Pioneer conducted an acute oral toxicity study in mice. A single dose of 582 mg per 

kilogram of body weight (mg/kg bw) of GM-HRA protein was administered by oral 

gavage to five male and five female mice. No clinical symptoms of toxicity, body weight 

loss, gross organ lesions or mortality were observed. Pioneer concluded that the result of 

this study shows that the GM-HRA protein does not cause acute toxicity.

Pioneer reports that while soybean is one of the major food allergens, none of the 

identified allergens is a member of the ALS family and the ALS protein from soybean 

has not been characterized as a soy allergen. Pioneer compared the amino acid sequence 

of the GM-HRA protein to the amino acid sequences of known allergens in the Food 

Allergy Research and Resource Program (FARRP, version 6.0) database using the 

FASTA34 sequence alignment program.6 None of the identified alignments met or 

exceeded the threshold of greater than or equal to 35% identity over 80 amino acids and 

no contiguous stretches of 8 or greater amino acids were shared between the GM-HRA 

protein and the proteins in the allergen database. Additionally, the GM-HRA protein is 

not glycosylated.

Pioneer also reports that the GM-HRA protein is rapidly (< 30 seconds) hydrolyzed in 

simulated gastric fluid (SGF) to fragments of < 3 kDa; and when subjected to simulated 

intestinal fluid (SIF), the GM-HRA protein, including the low weight molecular 

fragments seen in SGF, is completely and rapidly (< 2 minutes) hydrolyzed.

Pioneer concluded that the GM-HRA protein is unlikely to be a toxin and is not a 

potential allergen.

5. Evidence for Silencing of FAD2-1 Gene Expression

Pioneer determined that a 597 base pair gm-fad2-1 gene fragment, which is identical to a 

portion of the coding sequence of the endogenous soybean microsomal omega-6 

desaturase gene (FAD2-1), resulted in the down regulation of expression of the 

endogenous FAD2-1 gene. Pioneer examined the expression of the endogenous FAD2-1

gene in the leaf and developing seed of ten 305423 soybean plants and five control Jack 

soybean plants. Northern blots indicated that transcripts of the FAD2-1 gene in 

developing seeds of 305423 soybeans were greatly diminished when compared to the 
44

control soybean. This greatly decreased level of transcription confirms that the 

endogenous FAD2-1 gene is effectively silenced.

6. Food and Feed Uses of Soybean

Pioneer describes historical and current uses of soybean varieties in food and animal feed. 

The primary use of commodity soybeans is for soybean meal that is consumed by 

animals. Raw soybeans contain several antinutritional factors (trypsin inhibitors, urease, 

and hemagglutinins). Heat treatment is the most common processing method used to 

minimize the activity of such factors. Soybean oil is the major soybean fraction 

consumed by humans. Soybean oil accounts for 80% of total United States consumption 

of oils and fats.

Pioneer states that the 305423 soybean variety is intended to be used for the production 

of high oleic soybean oil. The oil is intended to be a highly stable vegetable oil suitable 

for frying applications without the need for hydrogenation which produces trans fatty 

acids and "hydrogenated flavor." Pioneer states that it is aware of no food or feed uses of 

current soybean varieties for which the 305423 soybean variety would not be equally 

suitable.

7. Overview of Compositional Analysis

Pioneer assessed the composition of grain and forage from the 305423 soybean and a null 

segregant (non-transgenic isoline) control. Pioneer states that the null segregant plants are 

an appropriate control because they are almost genetically identical to the corresponding 

305423 soybean plants with the exception that they do not carry the transgenic DNA. 

Both the transgenic and control soybeans were grown at six field locations in soybeangrowing areas of North America using a randomized complete block design with three 

replicates at each location. Pioneer also analyzed grain and forage from four different 

commercial soybean varieties.

Pioneer measured 52 components in grain and 5 in forage. Pioneer used the composition 

data obtained from the commercial varieties to calculate 99% tolerance intervals with 

95% confidence for all measured components. To interpret the composition results for 

305423 soybeans, Pioneer used the confidence intervals and established a combined 

literature range using data from published literature and databases on soybean 

composition.

Pioneer performed statistical analyses of composition data obtained for 305423 soybean 

and control soybean using mean values calculated from data aggregated from all tests 

sites. Pioneer used a linear mixed model analysis of variance (ANOVA). In order to hold 

the rate of false positive results to 5% or less, Pioneer employed the false discovery rate 

(FDR) approach (Benjamini and Hochberg (1995) and Westfall et al. (1999)). Pioneer 

reported the composition data by providing mean values, ranges, FDR-adjusted P-values, 

unadjusted P-values, tolerance intervals, and literature ranges, as available. Pioneer 

discussed analytical results in the context of FDR-adjusted P-values. Unless so indicated, 
45

statistical analyses using the unadjusted and adjusted P-values are in agreement. Pioneer 

used a P-value of 0.05 to denote a statistically significant difference between the control 

and the 305423 soybean.

Pioneer analyzed grain samples for proximates (protein, fat, and ash), acid detergent fiber 

(ADF), neutral detergent fiber (NDF), fatty acids, amino acids, isoflavones, and 

antinutrients. Compositional analysis of forage samples included proximates, ADF, and 

NDF. Table 1 contains the complete list of all measured components.

Table 1. Components measured in grain and forage

Proximates* & Fiber* Fatty Acids** Amino Acids Isoflavones+ Antinutrients

ash

fat

protein

acid detergent fiber (ADF)

neutral detergent fiber (NDF)

myristic (14:0)

palmitic (16:0)

palmitoleic (16:1)

heptadecanoic (17:0)

heptadecenoic (17:1)

stearic (18:0)

oleic (18:1)

linoleic (18:2)

linoleic (18:2)

 isomer (9,15)

linolenic (18:3)

arachidic (20:0)

eicosenoic (20:1)

behenic (22:0)

lignoceric (24:0)

methionine

cystine

lysine

tryptophan

threonine

isoleucine

histidine

valine

leucine

arginine

phenylalanine

glycine

alanine

aspartic acid

glutamic acid

proline

serine

tyrosine

genistin

genistein

malonylgenistin

daidzin

daidzein

malonyldaidzin

glycitin

glycitein

malonylglycitin

stachyose

raffinose

lectins

phytic acid

trypsin 

inhibitor

* Proximates and fiber were measured in both soybean grain and forage. All other 

components were measured in grain only.

** The levels of eleven additional fatty acids were near or below the lower limit of 

quantitation.

+ The levels of acetylgenistin, acetyldaidzin, and acetylglycitin were below the limit of 

quantitation.

7.1 Compositional Analysis of Soybean Grain

Intended Compositional Change – Fatty Acids

Pioneer analyzed the fatty acid composition of the oil extracted from the grain of the 

305423 and control soybeans. Pioneer provides the levels of 14 fatty acids (see Table 1) 

calculated as percentages of the total fatty acids. Pioneer states that the fatty acid analysis 

confirmed the expected high oleic acid phenotype as shown by a substantial increase in 

the mean level of oleic acid to 76.5% in 305423 soybean as compared to a mean level of 

21% in the control soybean. As expected, the increase in the level of oleic acid was 

accompanied by a decrease in the level of linoleic acid. The mean level of linoleic acid in 

the control soybean was 52.5% and that in the 305423 soybean was 3.6%. The level of 
46

linolenic acid also decreased since linolenic acid is formed in soybeans directly from 

linoleic acid. Pioneer also notes that the levels of two minor fatty acids, heptadecanoic 

and heptadecenoic, increased in the 305423 soybean to 0.8% and 1.2% of the total fatty 

acids, respectively. Pioneer explains that the increase in the levels of heptadecanoic and 

heptadecenoic fatty acids is not unexpected because the GM-HRA enzyme most likely 

results in the increased concentration of 2-ketobutyrate, which is the substrate in the 

biosynthesis of hepta fatty acids in soybeans. Pioneer also reported changes in the levels 

of all remaining fatty acids, which are not biologically significant.

Proximates and Amino Acids

Pioneer reports that the mean levels of protein and fiber in grain from the 305423 and 

control soybean are not statistically significantly different. While the mean level of fat 

was statistically significantly lower in the 305423 soybean than in the control using the 

unadjusted P-value, no statistical difference was detected using the FDR-adjusted Pvalue. The mean level of ash was statistically significantly lower in the 305423 soybean 

as compared to the control soybean. Mean levels of the proximates and fiber in grain 

from 305423 and control soybean lines were within the 99% tolerance intervals for the 

reference varieties and within the combined literature range.

Pioneer reports that there are no statistically significant differences observed in the mean 

levels of amino acids between 305423 and control soybean grain with the exception of 

threonine and glutamic acid. The mean levels of these two amino acids were statistically 

significantly increased in 305423 soybean when the unadjusted P-values were used, but 

not when the FDR-adjusted P-values were used. All these levels were within the 99% 

tolerance intervals.

Isoflavones

Pioneer analyzed grain from the 305423 and control soybeans for twelve isoflavones, of 

which nine were quantified (see Table 1). The mean levels for daidzin, malonylgenistin, 

and malonyldaidzin were statistically significantly increased in the 305423 versus the 

control soybean. Mean genestin levels were only shown to be statistically significantly 

increased when the unadjusted P-value was used. Mean values for all the measured 

isoflavones were within the 99% tolerance intervals.

In both the control and 305423 soybean the mean levels of genistin, glycitin, 

malonylgenistin, malonylglycitin, and malonyldaidzin were above the combined 

literature range. Daidzin levels were only above the combined literature range in the 

305423 soybean. No literature data were available for the level of glycitein. The values 

for the other two isoflavones (genistein and daidzein) were within the combined literature 

range.
47

Antinutrients

Pioneer measured several antinutrients in soybean grain including non-digestible 

oligosaccharides stachyose and raffinose, lectins, phytic acid, and trypsin inhibitor (see 

Table 1). No statistically significant differences were observed between the 305423 and 

control soybean in mean values for raffinose, lectins, and phytic acid. The mean value for 

trypsin inhibitor was statistically significantly lower for 305423 soybean as compared to 

the control soybean. The reduction in the mean value of trypsin inhibitor in 305423 

soybean was expected as Pioneer reported that the promoter used to drive expression of

the FAD2-1 gene, the Kunitz trypsin inhibitor promoter, has been shown to silence the 

KTi3 gene which encodes the Kunitz trypsin inhibitor. The mean value for stachyose was 

statistically significantly increased when the unadjusted P-value was used, but not when 

the FDR-adjusted P-value was used. Mean values for all the measured antinutrients were 

within the 99% tolerance intervals and within the combined literature ranges.

7.2 Compositional Analysis of Soybean Forage

Pioneer analyzed forage for protein, fat, ash, ADF, and NDF. Pioneer reports that no 

statistically significant differences were observed between the mean levels of these 

components in forage from the 305423 and control soybeans with the exception of the 

level of fat. The mean level of fat was statistically significantly decreased in 305423 

using the unadjusted P-value, but not when using the FDR-adjusted P-value. All mean 

levels were within the 99% tolerance intervals and combined literature ranges, with the 

exception of NDF. The mean level of NDF in both the control and 305423 soybean was 

above the combined literature range.

7.3 Endogenous Allergens

Pioneer conducted a study to assess whether the transformation process may have 

increased the overall allergenicity of 305423 soybean compared to conventional soybean. 

Using sera from clinically reactive soy allergic patients, Pioneer conducted IgE 

immunoblot and ELISA studies using protein extracts from 305423 soybean and 

conventional soybean. Pioneer reports that the SDS-PAGE Coomassie blue-stained 

protein profiles for 305423 and control soybean extracts appeared to be the same; they 

are similar in their IgE binding profile, and showed the same IgE binding capacity for 

305423 and control soy extracts. Pioneer concluded that the levels of endogenous 

allergens in and the allergic potential of 305423 soybean are comparable to those in 

nontransgenic control soybean.

8. Fatty Acids Intake

8.1 Human Diet

Pioneer generated estimates of dietary exposure to various fatty acids from the 

consumption of soy oil derived from conventional soybeans, as well as 305423 soybeans. 

Pioneer concluded that the intake of oleic acid would increase, while the intakes of 
48

linoleic acid and trans fatty acid would decrease. Pioneer states that based on very 

conservative intake estimates calculated on the assumption that high oleic soybean oil 

would replace all soybean oil in commercial applications, the dietary intake of linoleic 

acid would still fall within the current intake levels. Pioneer also notes that a variety of 

other oils used by the food industry would provide significant amounts of linoleic acid in 

the diet. Other noted changes in dietary intakes of fatty acids would result in small 

increases in the consumption of minor fatty acids, heptadacanoic acid (C17:0), 

heptadecenoic acid (C17:1), and the (9, 15) isomer of linoleic acid (cis-9, cis-15-

octadecadienoic acid).

Pioneer states that the 17-carbon fatty acids, heptadecanoic and heptadecenoic acids, 

occur at low levels in commonly consumed foods. For example, heptadecanoic acid is 

commonly found in meat (lamb, beef, pork), butter, and tofu, while heptadecenoic acid is 

found in foods such as tofu, beef, cheese, and olive oil. Pioneer stated that odd-chain fatty 

acids such as 17-carbon fatty acids are readily metabolized.

8.2 Animal Diets

When oil is removed from the soybean a defatted meal is generated that is used as a 

primary protein supplement for animal feed. Pioneer provided examples of swine and 

poultry diets to demonstrate that the reduced intake of linoleic acid, an essential fatty 

acid, would not lead to a nutritional deficit for animals consuming feeds containing meal 

derived from 305423 soybeans. Corn, the major ingredient of animal diets, is the primary 

source of linoleic acid in such diets. Even though soybean meal derived from 305423 

soybeans would have reduced amounts of linoleic acid, the quantity of linoleic acid 

provided by corn is several-fold above the animals' requirement.

9. Common or Usual Name of the Oil Product

Pioneer concluded that based on the intended increase in the level of oleic acid and 

decrease in the levels of linoleic and linolenic acids, a new common or usual name is 

appropriate for the oil from 305423 soybeans to distinguish this oil from the conventional 

soybean oil as defined in the Food Chemicals Codex (FCC). Pioneer proposed the name 

"high oleic soybean oil" for the oil that will be produced from 305423 soybeans.

10. Conclusion

Pioneer has concluded that, with the exception of the intended change in fatty acid 

composition, the 305423 soybean and the foods and feeds derived from it are not 

materially different in composition, safety, or any other relevant parameter from 

soybeans now grown, marketed, and consumed. At this time, based on Pioneer's data and 

information, the agency considers Pioneer's consultation on the 305423 soybean to be 

complete.

Mary D. Ditto, Ph.D.
49

1

The gm-hra cassette also contains three Flp recombinase target sequences; however, 

these sequences were not used in the development of the 305423 soybean. The presence 

of these sites does not cause recombination. Recombination requires the presence of the 

specific Flp recombinase enzyme that is not present in plants.

2

National Center for Biotechnology Information (NCBI) Protein Dataset release 156.0. 

The NCBI dataset incorporates all non-redundant entries from all GenBank nucleotide 

translations along with protein sequences from SWISS-PROT, PIR, PRF, and PDB 

databases.

3

Allergen database derived from the Food Allergy Research and Resource Program 

(FARRP, version 6.0).

4

Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), Western blot 

analysis, glycoprotein staining, mass determination of the tryptic peptides by matrix 

assisted laser desorption ionization mass spectrometry (MALDI-MS), and N-terminal 

amino acid sequence analysis.

5

The E-score was set at 1.0 to ensure that proteins even with limited similarity would not 

be overlooked.

6

University of Nebraska Allergen Database, version 6, January 2006; 

www.allergenonline.com.

From FDA website page: 

http://www.fda.gov/Food/Biotechnology/Submissions/ucm155595.htm