건강과 대안 » MON71800

[GMO] ‘콩 100% 국내제조 콩기름’… 낚이셨습니다

건강과대안 — Thu, 20 Jun 2013 20:19:02 +0000

‘콩 100% 국내제조 콩기름’… 낚이셨습니다

한겨레21 등록 : 2013.06.20 15:12 수정 : 2013.06.20 16:11
http://www.hani.co.kr/arti/economy/economy_general/592583.html

유전자조작작물(GMO)을 식품 원료로 사용했더라도 단백질로 최종 제품에 남아 있지 않으면 GMO로 표시할 필요가 없다. 콩기름의 경우 100% GM 콩으로 만들어도 GMO 표시 대상이 아니다. 100% 수입콩으로 제조됐음에도 GMO 표시가 없는 국내 제품 모습.탁기형

미국산 GM 콩으로 만들어도 표시 의무 강제 않는 현행 식품법
“GMO 원재료 들어갔으면 반드시 표시” 홍종학 의원 개정안 발의

지난 5월29일 저녁 9시 농림축산식품부 검역정책과장에게 주한 미국대사관한테서 전화가 왔다. “미국의 오리건주에서 재배가 금 지된 유전자조작(GM) 밀이 발견됐다는 사실을 30일 새벽 3시(한국시각)에 발표한다.” 그게 통보의 전부였다. 미국 정부의 공식 보고서나 성명서는 없었다고 농림축산식품부는 박주선 의원(무소속)에게 밝혔다. 우리나라는 미국에서 가장 많은 밀을 수입하며 오 리건주산 밀은 국내에 들어오는 미국산 밀의 3분의 1을 차지하는데도 말이다. 2010년 이후 오리건주에서 우리나라로 수입된 밀은 171만t에 이른다. 이번에 발견된 GM 밀 (MON71800)은 미국의 거대 농업기업 몬샌토가 1998~2005년 미국 16개 주에서 100여 건에 걸쳐 시험재배해 승인을 추진하다가 부정적 여론에 밀려 완전 폐기한 상품이다.

수입 중단 않고 제 돈 들여 전수조사한 식약처

우리나라처럼 미국산 밀을 많이 수입하는 일본은 즉각 오리건주산 밀 수입을 잠정 중단했다. 하지만 우리 식품의약품안전처는 오 리건주에서 들어오는 밀과 밀가루의 수입 단계 검사를 강화하기로 했을 뿐이다. 미승인 GM 밀이 나오면 즉시 반송 조처하겠다고 했다. 앞서 2012년 4월 미국에서 광우병(소 해면상뇌증·BSE) 사례가 발생했을 때도 정부는 통상마찰 소지가 있다며 검역 중단이 나 수입 중단 조처를 취하지 않았다.

지난 6월5일 식약처는 국내 주요 제분업체 7곳과 식품수입업체 2곳에서 보관한 오리건 주산 밀과 밀가루 샘플 40건과 5건을 수거 해 분석한 결과를 발표했다. “미승인 GM 밀이 검출되지 않았다.” 식약처는 “미국산 밀에 대해 GMO 분석을 한 나라는 한국이 유일하다”고 강조했다. “일본도 수입 중단에 준하는 조처를 했지만 GMO 검사는 하지 않았다.”

일반 콩이 안전하니 GMO 콩도 안전?

허술한 GM 식품 승인 절차 유전자조작작물(GMO)이 식품으로 승인받으려면 식품의약품안전처(옛 식품의약품안전청)의 ‘유전자 재조합식품 등 안전성 평가자료 심사위원회’ 단계를 거쳐야 한다. 이 위원회의 의무적인 심사 승인 절차를 통과한 것만이 식품이고, 이를 통과하지 못한 것은 식품이 아니다. 첫 GM 식품 승인은 2000 년 6월에 이뤄졌다.

1999년 11월12일 몬샌토코리아는 식품의약품안전청장에게 ‘제초제 내성 유전자조작 콩(GTS 40- 3-2)’의 안전성 심사를 요청했다. 심사 대상은 ‘서류’였다. 몬샌토가 제출한 서류를 토대로 유전자조 작식품의 안전성을 회사가 잘 평가했는지 심사하는 것이다. 별도의 실험은 없다. 개발자가 몇 년에 걸쳐 안전성 검사를 실시한다는 점을 생각해보면, 심사위원회가 안전성 실험을 수행하기란 현실적 으로 불가능할 수도 있다. GMO 개발자가 작성한 서류의 진정성을 믿고 심사할 수밖에 없다. 그렇다 면 개발자인 몬샌토는 유전자조작 콩의 투여 독성 실험을 했을까? 그렇지 않았다. ‘유전자재조합 콩 안정성 평가자료 심사 결과’ 보고서 15쪽을 읽어보자.

“현재의 자료 이외의 단회 투여 독성, 반복 투여 독성, 생식·발생 독성, 유전 독성 발암성, 기타 필요 한 독성(소화기계 독성 등)의 in vivo(생체 내) 독성 자료의 필요성은 없을 것으로 생각됨.”

그렇다면 무슨 근거로 GM 콩이 안전하다고 결론을 내린 것일까? 답은 ‘실질적 동등성’이란 원리다. ‘사람들이 오랜 세월 먹어온 보통의 콩은 안전하다. 심사 대상 GM 콩은 보통 콩과 실질적으로 다르 지 않다. 그러므로 GM 콩은 안전하다.’ 1992년 5월29일 미국 조지 부시 행정부가 미국 연방 관보 57 권 22984쪽에 이러한 정책 성명을 냈고 우리나라 식품법이 이를 반영했다.

GM 식품을 승인하는 심사위원회의 구성은 어떻게 될까? 통상법 전문가 송기호 변호사가 <맛있는 식 품법 혁명>(2010)에서 국내 심사위원 명단(2001~2010년)을 처음 공개했다. 위원회는 매년 20명 안팎으로 구성되는데 그중 2명은 10년간 계속 위원을 맡았다. 김해영·김형진 교수다. 김해영 교수는 경희 대 GMO개발연구단 단장이며, 이 연구단은 GM 식품 기술을 개발하는 곳이다. 김형진 교수는 한국생 명공학연구원에서 일하는데, 이 연구원은 생명과학 기술의 연구·개발을 목적으로 정부가 세운 곳이 다. 또 2001~2008년 계속 심사위원이었던 신영철 교수는 같은 기간에 ‘주식회사 아미코젠’의 대표이 사도 맡았다. 이 회사의 법인등기부를 보면, 회사의 목적은 ‘생명공학 기술을 적용한 효소 및 신소재’ 의 개발·생산 및 판매다. 또 회사는 유전자에 의해 형질전환된 미생물을 이용한 특허권을 보유하고 있다. 만일 GM 식품이 사회적으로 수용되지 않으면 이 특허 기술의 가치가 떨어질 수밖에 없다.

“유전자조작식품을 개발하거나 특허를 받은 사람들, 그러니까 유전자조작식품을 승인하고 수용하 는 데 직업적 이해관계가 있는 사람들이 유전자조작식품의 수용 여부를 결정하는 위원직을 그토록 장기간 맡는 것이 과연 정의로운가?” 송 변호사가 반문한다.

박상표 건강과대안 연구위원은 이렇게 반 박했다. “미국에서 미승인 GM 밀이 발견됐는데, 우리나라 정부가 국민의 세금으로 조 사하는 게 말이 되느냐? 미국이 자국 예산으로 조사해 유전자조작 밀을 걸러낸 뒤 수출 하는 게 당연하다. 그때까지 우리나라는 미국산 밀 수입을 잠정 중단하면 그만이다.”

식약처의 검사법도 문제다. 보통 GMO가 농산물이나 가공식품에 섞여 있는지 확인하려면 ‘공인검사법’이 필요하다. 검사법은 GMO 품목별로 제각각이다. GMO에 대한 정보를 토대로 검사법을 개발하는데, 이 정보는 정부의 재배 승인을 받은 품목에 한해 서만 제공된다. 하지만 이제껏 GM 밀은 재배 승인을 받은 것이 없다. 재배 승인을 받은 적이 없으니 검사법을 개발할 자료 자체가 없다. 미국 농무부가 검사 결과를 아직 발표 하지 못한 이유다.

그런데 식약처는 어떻게 검사를 했다는 것일까? 답은 GMO 유전자와 단백질을 확인하는 식약처의 자체 검사법이다. 그러고는 앞으로 검사를 계속하겠다고 덧붙였다. 미 승인 GM 밀의 샘플을 이제야 미국에서 건네받았고 공인검사법도 통보받을 예정이라서다. 공인검사법도 없는 상태에서 오리건주산 밀을 전수조사하는 ‘쇼’를 감행한 이유는 분명하다. GMO에 대한 부정적 여론이 확산 되는 걸 차단하기 위해서다. 하지만 GMO는 ‘뜨거운 감자’로 떠오르고 있다.

변형? 재조합? 용어부터 혼선

GMO를 얘기하려면 먼저 용어를 정리해야 한다. 널리 통용되는 GMO의 영문 이름 은 ‘Genetically Modified Organism’이다. 우리 법률이 다르게 번역해 문제다. 식품위생법은 ‘유전자재조합식품’이라고 부르며, 농산물품질관리법은 ‘유전자변형농산물’이라는 용어를 사용한다. ‘유전자변형생물체의 국가 간 이동에 관한 법률’이라는 긴 이름의 법에서는 ‘유전자변형생물체’라고 한다. 언론 에서는 ‘유전자조작’이라고 쓰는 게 일반적이 다. 재조합보다 변형이, 변형보다는 조작이 부정적으로 느껴진다. GM 식품 안전성을 승 인하는 식약처가 재조합을 고집한다. 어떤 용어를 쓰든지 사실은 모두 같은 것을 말한다. 다른 생물체의 유전자를 동식물 유전자 에 삽입해 종래 자연적 수정을 통해서는 발생하지 않던 새로운 유전자를 갖는 동식물 을 창조하는 작업이다.

90일 대신 2년간 관찰해보니…

GMO 안전성 논란

유전자조작작물(GMO)의 안전성은 논란거리다. 안전성을 부정할 수도, 그렇다고 확신할 수도 없는 상태다. 다만 최근에 GMO의 안전성에 심각한 우려를 표명한 연구 결과가 발표됐다.

2012년 9월 프랑스 캉대학교 질에리크 셀라리니 교수가 이끄는 연구진은 몬샌토의 제초제 ‘라운드 업’에 내성을 갖도록 만든 GM 옥수수 NK603과 라운드업을 쥐에게 먹이면서 신체 기능의 변화를 관 찰했다. NK603은 우리나라도 2002년 식용(2004년 사료용)으로 수입을 승인한 품목이다.

연구진은 실험 결과 NK603과 라운드업을 먹지 않은 대조군에 비해 이를 먹은 쥐에서 유선 종양과 간·신장 손상이 크게 늘었다는 점을 발견했다. 실험 대상은 암수 각 100마리씩 총 200마리였다. 보통 NK603을 비롯한 GMO의 동물실험은 최대 90일을 넘지 않는다. 하지만 프랑스 연구진은 쥐의 평균수 명 기간인 2년에 걸쳐 관찰했고, 암컷이 수컷보다 이상 증세가 심각하다고 밝혔다. 실험군에서 암컷 쥐가 조기 사망한 비율이 최고 70%에 달했기 때문이다. 대조군의 조기 사망률은 20%에 그쳤다. 연구 결과가 미국의 전문 학술지 <식품과 화학독성학> 온라인판에 공개되자 즉각 반박이 제기됐다. 실험 자체가 과학적으로 문제가 있기 때문에 그 결과를 신뢰할 수 없다는 내용이었다. 그동안 많은 경우 이렇게 찬반 공방을 지속하면서 GMO의 안전성은 미궁에 빠졌다.

앞서 2005년 5월 영국 일간지 <인디펜던트>가 몬샌토의 비공개 실험 자료를 입수해 GM 옥수수 MON863을 먹인 쥐의 면역 기능과 신장 크기 등에서 이상 현상이 관찰됐다고 보도했다. MON863 도 이미 우리나라에서 안정성 승인을 받은 품목이다. 프랑스 셀라리니 교수팀이 몬샌토의 자료를 검 토한 결과 ‘위해성이 있다’고 판정했다. 하지만 과학계의 반박이 뒤를 이었고 우리나라도 나름의 검토 과정을 거쳐 ‘안전하다’고 밝혔다.

1983년 5월19일 세계적 과학저널 <네이처>에는 새로 만들어진 꽃에 대한 논문이 실렸다. 벨기에와 독일 연구진이 항생제에 저항하는 물질을 일부러 꽃의 유전자에 심어 항생제 저항성을 가진 꽃을 개발했다고 발표했다. 이전까지는 식물 종을 개량하려 면 여러 대에 걸쳐 식물을 교배하는 ‘육종’ 이 유일한 수단이었다. 육종은 생물분류학 에서 비슷한 종류, 즉 종(species)이나 속 (genus)에 속하는 식물끼리 인위적으로 교 배하는 방법이다. 방울토마토와 씨 없는 수박이 대표적인 사례다. 하지만 이같은 절차 없이 새로운 식물을 단시일에 만들 수 있는 다른 차원이 탄생했다. 종과 속을 뛰어넘어 생물분류학상으로 훨씬 멀리 떨어져 있는 종류의 유전자는 물론이고, 동물과 미생물 의 유전자도 인위적으로 삽입되는 GMO다.

30년이 지난 현재 GMO는 무섭게 컸다. 몬 샌토·듀폰·신젠타 등 다국적 종자·농약 회 사와 카길 등 곡물회사가 세계 농업을 쥐락펴 락할 정도다. 전세계 GMO 경작지는 1996년 170만ha에서 지난해 1억7030만ha로 16년 만에 100배가 늘었다. 우리나라는 GMO 승인 건수로 볼 때 세계 5위다. GMO 재배국인 미 국·캐나다·멕시코를 제외하고 수입국만 따지면 일본에 이어 2위다. 2012년 6월까지 국내에서 승인을 거쳐 공식 유통되는 GMO의 종류는 일곱 가지다. 콩(대두)·옥수수·면화 (목화)·유채(카놀라)·사탕무·감자·알팔파 다. 이 GMO에 포함된 유전자는 모두 미생물 에서 왔고 기능은 두 가지다. 제초제를 뿌려도 잘 견디는 기능(제초제 내성 또는 저항성), 그리고 작물을 해치는 병해충을 없애는 기능 (살충성 또는 해충 저항성)이다.

GMO 원료 포함해도 함량 순위 높아야 표기

2011년 우리나라는 식용 GMO를 187만5 천t 수입했다. 모두 옥수수와 콩이다. 수입한 식용 옥수수 가운데 49%가 GMO였다. 식용 콩은 4분의 3(75%)으로 GMO 비중이 더 많았다. 하지만 GMO는 옥수수나 콩 원래의 모습이 유지된 채 팔리지 않는다. 다양한 과정을 거쳐 가공된다. GM 옥수수는 대부분 전분(녹말), 그리고 전분으로 만든 감미료를 뜻하는 전분당(과당·물엿·올리고당 등)으로 바뀐다. 반면 GM 콩은 대부분(99% 이상) 콩기름 제조에 쓰인다. 국내 식용유 시장 점유율 상위 4개사가 GM 농산물을 싹쓸이하는 이유다. CJ제일제당·사조해표·대상·삼양제넥스가 지난 3년간 우리나라 GMO 수입량(565만7천t)의 86%(486만8천t)를 들여온 것으로 확인됐다. 이뿐만 아니다. GM 식품이 분명하지만 농산물이 아니라서 아예 GMO 수입량 통계에 잡히지 않는 경우도 있다.

김훈기 서울대 기초교육원 교수는 카놀라유를 사례로 들었다. “카놀라유는 유채의 한 종류에서 씨앗을 원료로 삼아 만든 식용유다. 이때 사용되는 유채는 1970년대 캐나다에서 식용으로 품종이 개량된 것인데, 80% 이상이 GMO다. 우리나라는 카놀라유를 씨앗이 아닌 기름 상태로 수입한다. 농산물이 아니고 가공식품이라서 정부는 GMO 통계에 명시하지 않는다.”

승인되지 않은 유전자조작(GM) 밀이 미국 오리건주에서 발견됐지만 우리 정부가 미국산 밀 수입을 중단하지 않자 소비자생활협동조합(iCOOP) 회원들이 지난 6월12일 서울 광화문광장에서 수입 중단을 요구하는 퍼포먼스를 벌이고 있다.한겨레 박종식

경제정의실천시민연합(경실련) 박지호 간사는 6월12일 서울의 한 대형마트를 방문했다. 콩과 옥수수가 포함된 제품에 GMO라는 표시가 돼 있는지 확인하기 위해서다. 지난 4월에도 100여 개를 조사했지만 “국내 제품에서 GMO 표시가 된 경우를 본 적이 없다”고 박 간사는 말했다. 가끔 수입 제품에서만 ‘유전자재조합 옥수수 포함 가능성 있음’이라는 문구가 발견됐다.

GMO는 표시하지 않으면 일반 농산물과 마구 뒤섞여도 소비자는 알 길이 없다. 그래서 GMO 표시제는 소비자의 ‘알 권리’와 ‘선택할 권리’를 보장하는 최소한의 필요조건이다. 우리나라에서도 표시제가 시행되고 있다. GM 식품 자체에는 ‘유전자재조합(변형)식품’, GM 콩이 포함돼 있으면 ‘유전자재조합 콩 포함 식품’, 정확하진 않지만 포함될 가능성이 있으면 ‘유전자재조합 콩 포함 가능성 있음’ 등으로 표기한다. 이러한 GMO 표시를 왜 발견할 수 없을까? 두 가지 이유 때문이다.

“소비자 주권 지키는 게 경제민주화”

첫째, 불가피하게 섞여 들어간 경우다. 워낙 많은 GMO를 생산하고 있어서 유통 과정에서 일반 농산물에 GMO가 혼합될 수 있다. 이러한 현실적 여건을 고려해 국가별로 ‘비의도적 혼입률’을 채택한다. 의도하지 않게 GMO가 섞인 경우 표시를 면제해주는 것이다. 우리나라는 혼입률을 3% 이하로 정했다. 2007년 이뤄진 GM 식품 조사 결과 전체 4521건 중 1057건은 GMO 성분이 3% 이내로 검출돼 GMO 표시 없이 유통됐다. 2011년 3월에는 서울환경운동연합이 국내에서 GMO 표시가 없는 햄과 소시지 24개 제품 가운데 6개에서 GM 콩 성분이 나왔다고 밝혔지만, 비의도적 혼입률로 피해갔다. 유럽연합(EU)은 0.9%, 일본은 5%로 한다.

둘째, GMO를 식품 원료로 사용했더라도 가공식품은 표시 대상이 아니다. 현행 표시제를 보면, GM 농산물에 삽입한 외래 유전자 또는 그 유전자가 만든 단백질이 최종 제품에 남아 있지 않거나 검출할 수 없는 경우에는 GMO 표시를 할 필요가 없다고 돼 있다. 예를 들어 콩기름의 경우 콩에서 지방 성분만 뽑았기 때문에 유전자나 단백질이 포함되지 않는다. 따라서 100% GM 콩으로 만들었어도 GM 식품이라고 표시하지 않아도 된다. 게다가 식품에 쓰인 원료 가운데 GM 농산물이 전체의 5순위에 들어 있지 않으면 역시 표시가 면제된다. 그 결과 GM 옥수수 전분으로 만드는 빵·과자·음료·스낵·소스 등에 GMO 표시가 없을 수밖에 없다. 옥수수차·팝콘·시리얼도 마찬가지다. 김은진 원광대 교수(법학전문대학원)의 지적이다. “옛 식품법은 주요 원재료를 함량이 많은 순으로 5가지 이상 표시하도록 했고, GMO 표시 기준도 이를 반영했다. 하지만 현재는 모든 원재료를 표시하도록 식품법이 바뀌었고 GMO 표시도 당연히 달라져야 한다.” GMO 재배국이라 표시제에 소극적이던 미국에서도 주별로 의무 표시제 법안이 통과되기 시작했다. 미국 버몬트주 의회가 GMO 표시법을 107 대 37로 최근 통과시켰다.

홍종학 의원(민주당)이 식품법 개정안을 발의했다. 제품의 주요 원재료 함량 순위와 잔류 여부에 상관없이 GMO가 첨가됐으면 모두 표시하도록 했다. EU와 동일한 방식이다. 또 정부 기관별로 서로 다르게 사용하는 GMO 용어를 ‘유전자변형식품’으로 통일하고 토종 먹을거리를 찾는 소비자를 위해 ‘무유전자변형식품’(GMO Free)도 명시할 수 있도록 했다. 홍 의원은 “소비자 주권을 지키는 게 경제민주화”라고 설명했다.

정은주 기자 ejung@hani.co.kr

참고 문헌 김훈기 <생명공학 소비시대 알 권리 선택할 권리>(2013), 송기호 <맛있는 식품법 혁명>(2010)

[GMO] 유전자조작 밀 MON 71800

건강과대안 — Sun, 02 Jun 2013 10:22:39 +0000

미 농무부에 따르면, 몬산토사의 GM 밀은 이번에 발견된 Oregon주를 포함하여
미국 내 15개 주의 100군데 이상에서 시험재배를 실시했다고 합니다.
“The Agriculture Department said that during that seven-year period,
it authorized more than 100 field tests with the same
glyphosate-resistant wheat variety. Tests were conducted in in 15
states, including Oregon.”
(http://phys.org/news/2013-05-usda-non-approved-wheat-oregon-field.html)

로이터통신 보도에도 몬산토가 최소한 16개주 279군데에서 GM 밀 야외시험을
실시했으며, 그 면적이 4천 에이커가 넘는다고 합니다.
Government records show Monsanto conducted at least 279 field tests of
herbicide-resistant wheat on over 4,000 acres in at least 16 states
from 1994 until the company abandoned its field testing of wheat in
2004.
(http://www.reuters.com/article/2013/05/31/us-wheat-control-idUSBRE94U06H20130531)

이번에 몬산토사의 GM밀이 발견된 Oregon주의 농장은 GM 밀을 시험재배한 적이
없기 때문에 해당 농장주가 몬산토사를 상대로 소송을 제기했다고 하구요… 그리고
AP통신에 따르면… 몬산토사의 GM 밀이 시험재배된 16개주의 명단은 아래와 같습니다.
Arizona, California, Colorado, Florida, Hawaii, Idaho, Illinois,
Kansas, Minnesota, Montana, Nebraska, North Dakota, Oregon, South
Dakota, Washington and Wyoming.
(http://www.spokesman.com/stories/2013/jun/04/kansas-farmer-sues-monsanto-over-gmo-wheat/)

[PDF]

Evaluation of Broiler Performance When Fed Roundup-Ready …
www.poultryscience.org/ps/paperpdfs/04/p0481325.pdf
(첨부파일)
=================
GM Crop Database

http://www.cera-gmc.org/?action=gm_crop_database&mode=ShowProd&data=MON71800&frmat=SHORT

Database Product Description

MON71800

Host Organism

Triticum aestivum L. (Wheat)

Trait

Glyphosate herbicide tolerance.

Trait Introduction

Agrobacterium tumefaciens-mediated plant transformation.

Proposed Use

Production of T. aestivum for human food and livestock feed. This material will not be grown outside the normal production area for wheat.

GM Crop Database

http://cera-gmc.org/index.php?action=gm_crop_database&mode=ShowProd&data=MON71800&frmat=LONG

Database Product Description

MON71800

Host Organism

Triticum aestivum L. (Wheat)

Trait

Glyphosate herbicide tolerance.

Trait Introduction

Agrobacterium tumefaciens-mediated plant transformation.

Proposed Use

Production of T. aestivum for human food and livestock feed. This material will not be grown outside the normal production area for wheat.

Company Information

Monsanto Company
Chesterfield Village Research Center (MO)
700 Chesterfield Parkway North
St. Louis
MO USA

Summary of Regulatory Approvals

Country	Environment	Food and/or Feed	Food	Feed	Marketing
Colombia			2004
United States		2004

Click on the country name for country-specific contact and regulatory information.

Introduction

The spring wheat variety MON 71800 (Roundup Ready wheat) was developed to allow the use of glyphosate, the active ingredient in the herbicide Roundup? as a weed control option in spring wheat. This genetically engineered wheat variety contains a glyphosate-tolerant form of the plant enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS), isolated from the soil bacterium Agrobacterium tumefaciens strain CP4. The novel form of this enzyme is termed hereafter CP4 EPSPS.

The EPSPS enzyme is part of the shikimate pathway that is involved in the production of aromatic amino acids and other aromatic compounds in plants (Steinrucken and Amrhein, 1980). When conventional plants are treated with glyphosate, the herbicide binds to EPSPS, thereby preventing the synthesis of aromatic amino acids needed for plant growth. The CP4 EPSPS enzyme in MON 71800 spring wheat has a reduced affinity for glyphosate; its enzymatic activity is therefore not hindered by the herbicide.

EPSPS is present in all plants, bacteria, fungi, but not in animals, which do not synthesize their own aromatic amino acids. Because the aromatic amino acid biosynthetic pathway is not present in mammalian, avian or aquatic life forms, glyphosate has little if any toxicity for these organisms (U.S. EPA, 1993; WHO, 1994; Williams et al. 2000). The EPSPS enzyme is normally present in food derived from plant and microbial sources.

MON 71800 was developed by introducing the CP4 EPSPS coding sequences into the spring wheat variety ?Bobwhite? using Agrobacterium-mediated transformation.

General Description

Monsanto used Bobwhite spring wheat (designated as MON 71900) as the parental variety to produce Roundup Ready® wheat event MON 71800. Event MON 71800 was developed through Agrobacterium-mediated transformation using the double border, binary vector PVTXGT10. The vector consists of the T-DNA segment intended for transformation, as well as the backbone DNA which is not expected to be incorporated into the transformed wheat.

The T-DNA contains two cp4 epsps gene cassettes and the left and right T-DNA border sequences. The segment is approximately 7.0 Kb in size. The cp4 epsps coding sequence in both cassettes is derived from Agrobacterium sp. strain CP4 and begins with a leader chloroplast transit peptide coding sequence (ArabTP) derived from the Arabidopsis thaliana epsps gene. The ArabTP transit peptide directs the CP4 EPSPS protein expressed in event MON 71800 to chloroplasts. The ArabTP-cp4 epsps fusion coding sequence in each cassette is followed by the 3′ non-translated region of the nopaline synthase gene (nos 3′) from Agrobacterium tumefaciens that provides the transcriptional termination signal. The upstream regulatory sequence of the first cassette consists of the 5′ region of the rice actin1 gene (Pract1- ract1I) which contains the promoter, transcription start site and the first intron. The upstream regulatory sequence of the second cassette consists of an enhanced 35S promoter from the cauliflower mosaic virus (CMV) (P-e35S) and the intron of the corn heat shock protein gene (hsp70I). The two different promoters were used to drive expression of the gene product in both vegetative and reproductive tissues.

Reference: U.S. Food and Drug Administration

Summary of Introduced Genetic Elements

Code	Name	Type	Promoter, other	Terminator	Copies	Form
CP4 epsps	5-enolpyruvyl shikimate-3-phosphate synthase (Agrobacterium tumefaciens CP4)	HT	enhanced CaMV 35S chloroplast transit peptide from A. thaliana EPSPS gene (CTP2)	A. tumefaciens nopaline synthase (nos) 3′-untranslated region
CP4 epsps	5-enolpyruvyl shikimate-3-phosphate synthase (Agrobacterium tumefaciens CP4)	HT	rice actin I promoter and intron sequences chloroplast transit peptide from A. thaliana EPSPS gene (CTP2)	A. tumefaciens nopaline synthase (nos) 3′-untranslated region

Characteristics of Triticum aestivum (Wheat)

Center of Origin	Reproduction	Toxins	Allergenicity
Asia Minor, Tigris-Euphrates drainage basin of the Middle East, as well as the regions of southern Caucasus and Crimea.	Primarily self-pollinated (autogamous). Some outcrossing by wind-pollination of less than 10%. Seed does not display dormancy.	Phytic acid, trypsin inhibitor, lectins. Gliadins responsible for celiac enteropathy.	Glutenins and gliadins (e.g., the IgE-inducing alpha-gliadin).

Donor Organism Characteristics

Latin Name	Gene	Pathogenicity
Agrobacterium tumefaciens strain CP4	CP4 EPSPS	A. tumefaciens is a common soil bacterium that is responsible for causing crown gall disease in susceptible plants. There have been no reports of adverse affects on humans or animals.

Modification Method

The wheat line MON 71800 was produced by Agrobacterium-mediated transformation of plant cells from ?Bobwhite? spring wheat. The plasmid vector PV-TXGT10 used for the transformation contained two cp4 epsps gene cassettes coding for glyphosate tolerance. Each gene cassette consisted of chloroplast transit peptide coding sequences from the Arabidopsis thaliana epsps gene (ArabTP) associated with the sequences of the cp4 epsps gene. Two different promoters were used to regulate the expression of each cp4 epsps gene: 1) the enhanced 35S promoter from the cauliflower mosaic virus and 2), the promoter, transcription start site, and first intron of the 5? region of the rice actin1 gene. Terminator sequences in each gene casette consisted of the 3? non-translated region of the nopaline synthase gene (nos 3?). The PV-TXGT10 vector backbone contained the origin of replication sequences ori-V and ori-322/rop. The vector backbone also contained the aad gene, which codes for streptomycin adenyltransferase, to allow the selection of bacteria containing the PV-TXGT10 vector.

Characteristics of the Modification

The Introduced DNA
Southern blot analysis and Polymerase Chain Reaction (PCR) amplification of the genomic DNA of MON 71800 wheat demonstrated one site of integration of a single copy of the T-DNA insert of PV-TXGT10. Southern blot analysis also confirmed the insertion of one intact copy of each cp4 epsps gene cassette, including the promoter, terminator and chlorophyll transit peptide sequences. None of the vector backbone sequences were integrated into the genome of MON 71800 wheat.

Genetic Stability of the Introduced Trait
The stability of the inserted DNA was evaluated, across several generations of wheat plants, using Southern blot analysis. The plants tested were progeny from several generations of self-fertilization, as well as from crosses with commercial varieties. The results of the genomic DNA blot analysis confirmed the stable inheritance of the inserted cp4 epsps gene cassettes. The stability of the introduced trait was also demonstrated after 18 generations of selfing of the original homozygous glyphosate-resistant plants. These generations showed no decrease in tolerance to glyphosate.

Mendelian segregation studies for the inheritance of the glyphosate tolerant trait were conducted with heterozygous first generation plants: these were selfed and the progeny were sprayed with glyphosate. The resulting 3:1 ratio of tolerant to sensitive plants was statistically significant and confirmed the inheritance of a single insertion site of the glyphosate tolerant trait.

Expressed material
In plant cells, the EPSPS enzyme is transported to the chloroplast by a transit peptide, which then cleaves from the enzyme. The introduced gene sequences in MON 71800 included a gene for a chloroplast transit peptide from Arabidopsis thaliana. Thorough analyses were conducted to investigate whether the same mechanism of binding, transporting, and cleaving of the chloroplast transit peptide to the CP4 EPSPS enzyme exists in MON 71800. The analyses revealed two forms of CP4 EPSPS expressed in MON 71800: one in which the chloroplast transit peptide has completely cleaved from the full length enzyme, and the other in which only part of the transit peptide is bound to the enzyme. Further analysis of Western blots showed that of the total amount of CP4 EPSPS expressed in MON 71800, 80% is in the form where the transit peptide is fully cleaved and 20%, where part of the transit peptide is still bound to the enzyme.

An enzyme-linked immunosorbent assay (ELISA) analysis was used to quantify the levels of the CP4 EPSPS proteins in forage and grain from MON 71800. The mean levels of both CP4 EPSPS proteins, on a fresh weight basis, were 106 μg/g in forage and 13 μg/g in grain. Assuming 14.5% moisture in stored grain, the concentration of both CP4 EPSPS proteins in the grain is approximately 0.0015%, on a dry matter basis.

Food and/or Feed Safety Considerations

Dietary exposure
The genetic modification of MON 71800 spring wheat will not result in any change in the consumption pattern of wheat and wheat-based products. MON 71800 is expected to be used in similar applications as other spring wheat cultivars by the food industry. MON 71800 did not express any novel compositional characteristics, as confirmed by the similarity in composition of the modified line to its parental counterpart, and other conventional spring wheat varieties. Furthermore, the availability of many spring wheat cultivars for cultivation, and the normal variation in wheat composition due to differences in grade and growing conditions, result in a wide variation in the composition of conventional wheat grain. Consequently, the dietary exposure of consumers in the United States to MON 71800 is anticipated to be the same as for other varieties of commercially available spring wheat.

Nutritional and Compositional Data
The nutritional components of MON 71800 grain and forage were determined analytically and compared to those of the parental line ‘Bobwhite’ and several commercial varieties grown at five locations in the United States and Canada. For the grain, these components included proximates (crude protein, crude fat, ash, moisture, total carbohydrates), total dietary fibre, sugars, starch, amino acids, fatty acids, B vitamins, vitamin E and minerals. Forage samples were analyzed for proximates, acid detergent fibre, neutral detergent fibre, calcium and phosphorus. At some of the locations, for both the grain and the forage, there were statistically significant differences in the levels of certain components between MON 71800 and its parental line. These differences within locations could be attributed to environmental effects rather than to any unintended effect of the genetic modification in MON 71800. However, the combined data from all test locations demonstrated that the nutritional composition of MON 71800 grain and forage was comparable to that of the parental line ‘Bobwhite,’ and other commercial spring wheat varieties.

Phytic acid occurs naturally in wheat and other cereals. It is indigestible by humans and non-ruminant livestock, and inhibits the absorption of iron and other minerals. Grain samples of MON 71800, the parental line ‘Bobwhite,’ other commercial wheat varieties were analyzed to determine levels of phytic acid. The concentration of phytic acid in MON 71800 was comparable to that in the parental line, and was within the range of values determined for the commercial varieties and those found in the literature.

Toxicity and Allergenicity
The potential for toxicity and allergenicity of MON 71800 wheat was investigated using the following data and information: results from the determination of amino acid sequence similarity between the CP4 EPSPS proteins and known toxins and allergens; analysis for possible glycosylation of the CP4 EPSPS proteins; analysis of the stability the novel proteins in simulated gastric fluids; results from an acute oral toxicity study in mice using the CP4 EPSPS proteins; and, information on the safety of the cp4 epsps gene donor, A. tumefaciens strain CP4. The potential for increased allergenicity of the grain from MON 71800 was investigated, specifically with regard to endogenous wheat allergens that induce an IgE reaction in susceptible humans. Various immunilogical assays, using sera from humans with an IgE-mediated wheat allergy, were performed with extracts from MON 71800, the parental line, and several other commercial wheat varieties. The possibility that the genetic modification would have also altered the levels of gliadin, proteins that cause celiac enteropathy in susceptible persons, was also investigated. Gliadin levels were measured, and gluten levels calculated in MON 71800, its parental line, and other commercial varieties.

The CP4 EPSPS proteins in MON 71800 showed no amino acid sequence similarity with known toxins and allergens and neither of the two forms of the protein are glycosylated. Both forms of the CP4 EPSPS protein were rapidly digested under simulated gastric fluid conditions, and the enzyme activity of each protein was substantially diminished within the same period as for the digestion. The results of the acute oral toxicity study on mice, at the highest administered dose, showed no adverse effects of the CP4 EPSPS proteins. Gluten levels were not significantly altered by the genetic modification of MON 71800, neither were the endogenous allergens, as demonstrated by the results of the immunological studies.

Both forms of the CP4 EPSPS protein were expressed at very low levels in MON 71800 grain and forage. This fact, along with negative results of the various safety studies, sequence homology investigations, and immunological assays led to the conclusion that MON 71800 spring wheat did not demonstrate any potential for toxicity and novel allergenicity, nor any altered endogenous allergenicity, compared to conventional spring wheat varieties.

Links to Further Information

Food Standards Australia New Zealand[PDF Size: 296073 bytes]

Initial Assessment Report: Application A524 – Food derived from herbicide – tolerant wheat MON 71800

U.S. Food and Drug Administration[PDF Size: 164547 bytes]

Biotechnology Consultation Note to the File BNF No. 000080

References

Steinrucken, H.C. & Amrhein, N. (1980). The herbicide glyphosate is a potent inhibitor of 5-enolpyruvylshikimic acid-3-phosphate synthase. Biochemical and Biophysical Research Communications, 94, 1207-1212.

U.S. EPA. (1993). Reregistration Eligibility Decision (RED): Glyphosate. Office of Prevention, Pesticides and Toxic Substances, U.S. Environmental Protection Agency, Washington, D.C.

WHO. (1994). Glyphosate. World Health Organization (WHO), International Programme of Chemical Safety (IPCS), Geneva. Environmental Health Criteria No. 159.

Williams, G.M., Kroes, R. & Munro, I.C. (2000). Safety evaluation and risk assessment of the herbicide Roundup and its active ingredient, glyphosate, for humans. Regulatory Toxicology and Pharmacology 31, 117-165.

================

Biotechnology Consultation Note to the File BNF No. 000080

Date: July 22, 2004

http://www.fda.gov/Food/FoodScienceResearch/Biotechnology/Submissions/ucm155777.htm

Subject: Monsanto Roundup Ready® Wheat Event MON 71800

Keywords: Wheat, Roundup Ready®, Triticum aestivum, Glyphosate (N-phosphonomethyl-glycine), EPSPS (5-enolpyruvylshikimate-3-phosphate synthase), cp4 epsps gene, Agrobacterium sp. CP4 strain, Herbicide-Tolerant, Glyphosate-Tolerant, event MON 71800, Bobwhite

1. Introduction

In a submission dated June 28, 2002, Monsanto provided information to support the safety and nutritional assessment of their glyphosate-tolerant (Roundup Ready®) wheat (Triticum aestivum) containing a transformation event designated MON 71800. The company provided additional information in a submission dated April 25, 2003. Monsanto concluded that their Roundup Ready® wheat event MON 71800 and the foods and feed derived from it are as safe and nutritious as current commercial varieties of wheat and the comparable foods and feed derived from them. Monsanto has previously completed consultations for other Roundup Ready® crops which are also tolerant to glyphosate. These other crops include bioengineered soybean, canola, corn, and cotton (BNFs 01, 20, 71, and 86, respectively). In a letter to FDA dated June 9, 2004, Monsanto states that the firm is deferring all further commercial development efforts to introduce Roundup Ready® wheat until such time that other wheat biotechnology traits are introduced. In its letter, Monsanto requests that FDA complete the consultation process for Roundup Ready® wheat event MON 71800.

2. Intended Effect

The intended effect of the genetic modification is to confer tolerance to the herbicidal compound glyphosate (N-phosphonomethyl-glycine) which is the active ingredient in Roundup® agricultural herbicides. In glyphosate-sensitive plants, glyphosate binds to the plant 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) and prevents the synthesis of aromatic amino acids that are necessary for plant growth. Roundup Ready® wheat MON 71800 contains the epsps gene from Agrobacterium sp. strain CP4 (cp4 epsps gene) encoding the CP4 EPSPS enzyme. This enzyme has a reduced affinity for glyphosate when compared to the native plant EPSPS enzyme. As a result, wheat plants expressing the CP4 EPSPS enzyme are tolerant to glyphosate and survive spraying with Roundup®

3. Development of Roundup Ready® Wheat Event MON 71800

3.1. The Parent Plant Wheat

Monsanto describes the history and biology of wheat. The scientific name for common bread wheat is Triticum aestivum L. Wheat belongs to the order Poales (Glumiflorae), family Poaceae (Graminae), tribe Triticeae, genus Triticum. T. aestivum is hexaploid with a total of 42 chromosomes. Modern wheat cultivars are either tetraploid (durum) or hexaploid (common and club types). Wheat is predominantly self-pollinating.

3.2. Genetic Modifications and Characterization of the Introduced DNA

Monsanto used Bobwhite spring wheat (designated as MON 71900) as the parental variety to produce Roundup Ready® wheat event MON 71800. Event MON 71800 was developed through Agrobacterium-mediated transformation using the double border, binary vector PV-TXGT10. The vector consists of the T-DNA segment intended for transformation, as well as the backbone DNA which is not expected to be incorporated into the transformed wheat.

The T-DNA contains two cp4 epsps gene cassettes and the left and right T-DNA border sequences. The segment is approximately 7.0 Kb in size. The cp4 epsps coding sequence in both cassettes is derived from Agrobacterium sp. strain CP4 and begins with a leader chloroplast transit peptide coding sequence (ArabTP) derived from the Arabidopsis thaliana epsps gene. The ArabTP transit peptide directs the CP4 EPSPS protein expressed in event MON 71800 to chloroplasts. The ArabTP-cp4 epsps fusion coding sequence in each cassette is followed by the 3′ non-translated region of the nopaline synthase gene (nos 3′) from Agrobacterium tumefaciens that provides the transcriptional termination signal. The upstream regulatory sequence of the first cassette consists of the 5′ region of the rice actin1 gene (P-ract1-ract1I) which contains the promoter, transcription start site and the first intron. The upstream regulatory sequence of the second cassette consists of an enhanced 35S promoter from the cauliflower mosaic virus (CMV) (P-e35S) and the intron of the corn heat shock protein gene (hsp70I). The two different promoters were used to drive expression of the gene product in both vegetative and reproductive tissues.

The vector backbone contains the origin of DNA replication ori-V that allows maintenance of PV-TXGT10 in Agrobacterium as well as the origin of replication ori-322/rop that allows the replication of PV-TXGT10 in the intermediate host E. coli. The vector backbone also contains the aad gene encoding the selectable marker enzyme streptomycin adenyltransferase that allows selection of bacteria containing PV-TXGT10.

Monsanto used Southern blot analysis to characterize the DNA introduced in the transformation event MON 71800. Genomic DNA isolated from event MON 71800 and the parent line MON 71900 was digested with restriction enzymes and subjected to Southern blot analysis using radiolabeled DNA probes corresponding to different segments of the T-DNA and plasmid backbone. Based on this analysis, Monsanto concludes that: 1) event MON 71800 contains a single T-DNA insert of the expected size (approximately 7.0 Kb) comprising one intact copy of each cp4 epsps expression cassette; 2) event MON 71800 does not contain the plasmid backbone sequences, including ori-V, ori-322/rop, and aad coding sequence; and 3) all genetic elements present in the expression cassettes before transformation are also present in event MON 71800.

Monsanto performed polymerase chain reaction (PCR) on event MON 71800 genomic DNA to verify the presence of the unique T-DNA insert-to-plant junction sequences. This analysis yielded PCR products of the expected size, thereby confirming the presence of these unique sequences in event MON 71800.

3.3. Stability of the Introduced DNA

Monsanto describes the experiments it conducted to evaluate the stability of the DNA insert. Monsanto used Southern blot analysis to test the insert stability across several generations of wheat plants containing event MON 71800. The tested plants included R2, R3, R4, and R5 progeny derived from the initial MON 71800 transformant by self-fertilization and three pre-commercial wheat varieties (Westbred 926 BC4F1, HJ-98 BC4F1, and BW251 BC5F4) containing event MON 71800 introduced by traditional breeding. These seed materials represent the 2nd, 3rd, 4th, 5th, 8th, 8th, and 12th generations, respectively. The non-transgenic wheat lines MON 71900, Westbred 926, HJ-98, and BW251 were used as control lines. Genomic DNA isolated from the seeds of all of these lines was digested with the restriction enzyme Bam HI and probed with the cp4 epsps coding sequence. The DNA from the non-transgenic control lines did not contain detectable hybridization bands. The digested transformation plasmid used as a positive control produced two bands of the expected length (3.7 and 8.6 Kb). The DNA from the test lines containing event MON 71800 produced two DNA bands – the expected 3.7 Kb band (which represents the internal segment of the insert) and a 9.0 Kb fragment (which represents a border segment containing the 3′ end of the insert and a portion of the flanking wheat DNA sequence). Monsanto concludes that these experiments demonstrate the stability of the inserted DNA spanning six different seed generations.

3.4 Inheritance of the Roundup Ready® Trait

Monsanto tested the inheritance of the Roundup Ready® trait as follows: the original event MON 71800 R0 plant was selected on a glyphosate medium. Initial R1 plants were sprayed with Roundup® herbicide and evaluated for their resistance and sensitivity. Homozygous glyphosate-resistant R1 plants were identified by a PCR-based homozygosity assay and confirmed by spray testing their R2 progeny with Roundup® herbicide. One of the homozygous glyphosate-resistant plants from the R1 generation was used to increase the number of generations to R18 by self-pollination. These generations showed no decrease in vegetative or reproductive tolerance and no sensitivity to Roundup® confirming the stability of the Roundup Ready®trait.

Monsanto also described a Mendelian segregation study for the Roundup Ready® trait in event MON 71800. Monsanto selected heterozygous R1 plants and self pollinated them to produce progeny. The R2 plants were sprayed with Roundup® herbicide and scored for their resistance or sensitivity to the herbicide. The ratio of resistant to sensitive plants was found to be in conformance with the expected 3:1 ratio by the chi square test. Monsanto concludes that this result is consistent with a single insertion site of the Roundup Ready® trait in the wheat genome.

4. Expressed Proteins: the CP4 EPSPS Enzymes

4.1. Identity, Function and Characterization

Monsanto notes that the CP4 EPSPS enzyme is structurally and functionally similar to native plant EPSPS enzymes, which are involved in the biosynthesis of aromatic amino acids that are necessary for growth and development of the plant.

In order for EPSPS to function in plants, it must be transported to the chloroplast. In event MON 71800, the A. thaliana chloroplast transit peptide coding sequence was joined to the cp4 epsps coding sequence so that a fusion protein of CP4 EPSPS (47.6 kilodaltons (kDa), 455 amino acids) and the A. thaliana chloroplast transit peptide (1.6 kDa, 76 amino acids) would be expressed. The transit peptide directs the protein to the chloroplast. Typically, transit peptides are completely cleaved from the protein following delivery to the chloroplast. However, there are examples in the literature of alternatively processed forms where the transit peptide is only partially cleaved.

Monsanto isolated and purified CP4 EPSPS expressed in the grain of event MON 71800 and used the following techniques for characterization: sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE); western blot analysis; glycosylation analysis; enzymatic assay; N-terminal amino acid sequence analysis; and matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry analysis. Monsanto identified two forms of the CP4 EPSPS protein in the grain of event MON 71800, i.e., a “mature form” and an “alternatively processed form”. The mature form is the full length CP4 EPSPS, where the transit peptide was fully cleaved; the alternatively processed form is the full length CP4 EPSPS protein plus seven amino acids from the transit peptide. Monsanto used the mature form of CP4 EPSPS produced in E. coli for comparison in the characterization analyses.

Based on quantitative image analysis of western blots, Monsanto reports that the relative percentages of the CP4 EPSPS proteins produced in event MON 71800 are 80% mature form and 20% alternatively processed form. Glycosylation analysis (ECL glycoprotein detection system) indicated that neither form of the protein is glycosylated. In addition, Monsanto concludes from enzyme activity assays that the average specific activity of both plant-produced CP4 EPSPS proteins is comparable to the specific activity of the protein produced in E. coli.

4.2. Expression Level and Human Exposure

Monsanto used a direct double antibody sandwich enzyme-linked immunosorbent assay (ELISA) analysis to estimate the levels of the CP4 EPSPS proteins in forage and grain tissues collected from event MON 71800. The forage and grain tissues from the parental line MON 71900 were used as control samples. Monsanto provides the results of the assay which show that the average level of CP4 EPSPS proteins are 106 μg/g in forage and 13 μg/g in grain on a fresh weight basis. Monsanto estimates a per capita exposure to the CP4 EPSPS proteins to be 0.039 mg/kg body weight/day assuming no loss due to food processing.

4.3. Presence in Food Crops

Monsanto discusses the similarity of the CP4 EPSPS proteins present in event MON 71800 to native EPSPS proteins that occur in plants and microorganisms and to CP4 EPSPS proteins present in bioengineered crops. Monsanto states that the CP4 EPSPS proteins expressed in event MON 71800 are similar to naturally occurring EPSPS proteins present in a variety of food and feed sources, such as, soybean, corn, and Baker’s yeast. Monsanto also states that the mature CP4 EPSPS protein expressed in event MON 71800 grain has been consumed by humans and animals since 1996, through the consumption of Roundup Ready® crops, such as soybean, corn and canola. The mature protein is identical to or shares greater than 99% amino acid sequence identity with the CP4 EPSPS proteins produced in these food crops. The alternatively processed form of CP4 EPSPS shares greater than 98% amino acid sequence identity with the CP4 EPSPS protein produced in other Roundup Ready® food crops.

4.4. Assessment of Potential Allergenicity

4.4.1. Donor

Agrobacterium species, the source of the CP4 EPSPS gene, are not known to be allergenic.

4.4.2. Amino Acid Sequence Homology

Monsanto searched the ALLPEPTIDES database for amino acid sequence homology to the CP4 EPSPS proteins found in event MON 71800 using the FASTA algorithm. The ALLPEPTIDES database is comprised of publically available protein sequences in SwissProt version 39+, TrEMBL (updated weekly), and GenPept version 124. Monsanto concluded that the CP4 EPSPS proteins do not demonstrate sequence similarity to proteins known to pose human health risks.

In addition, Monsanto screened all overlapping peptides of eight or more contiguous amino acids of the CP4 EPSPS proteins against the ALLERGEN3 database using a pairwise comparison algorithm. ALLERGEN3 is an allergen and gliadin protein sequence database compiled by Monsanto. Neither of the two forms of CP4 EPSPS contained sequences of eight or more contiguous amino acids identical to those from proteins in the allergen database.

4.4.3. Stability in Simulated Digestive Fluid

Monsanto examined the in vitro stability of both forms of CP4 EPSPS in simulated gastric fluids (SGF) prepared according to U.S. Pharmacopeia (1990). Stability was assessed by colloidal blue staining of SDS-PAGE gels, western blot analysis, and EPSPS enzyme activity assay.

In experiments performed with the purified E. coli-produced mature form of CP4 EPSPS, 95-98% of the protein was digested in SGF within 15 seconds, with no detectable degradation products present. The enzymatic activity of this form of CP4 EPSPS decreased by greater than 90% following SGF treatment for 15 seconds. Monsanto also refers to previous experiments which show that purified E. coli-produced mature form of CP4 EPSPS was digested in less than ten minutes in simulated intestinal fluid (SIF) (Harrison et al., 1996).

In experiments performed with the purified E. coli-produced alternatively processed form of CP4 EPSPS, 98% of the protein was digested in SGF within 15 seconds. The enzymatic activity of this form of CP4 EPSPS was lost following SGF treatment for 15 seconds. In addition, purified E. coli-produced alternatively processed form of CP4 EPSPS was degraded within four to eight hours of SIF treatment.

Monsanto performed additional experiments examining the in vitro stability of the CP4 EPSPS proteins produced in event MON 71800 grain in SGF. These experiments were performed to assess the stability of the CP4 EPSPS proteins within the matrix of other wheat proteins. The results indicated that these proteins were rapidly digested within a matrix of wheat grain proteins, with greater than 95% of CP4 EPSPS digested within 15 seconds.

Finally, Monsanto calculated that the amount of CP4 EPSPS proteins present in event MON 71800 grain is very low and represents only a small portion of the total protein. From these data and information, Monsanto concluded that the CP4 EPSPS proteins in event MON 71800 do not pose a significant allergenic risk.

4.5. Assessment of Potential Toxicity

Monsanto provided information about Agrobacterium sp. strain CP4, the donor of the cp4 epsps gene. Agrobacterium species are non-pathogenic and non-toxigenic. The safety of the Agrobacterium sp. strain CP4 has been previously evaluated during Monsanto’s consultations with FDA on other Roundup Ready&174; crops.

In two separate studies, Monsanto assessed the acute oral toxicity of the CP4 EPSPS proteins in mice. They used both the mature and alternatively processed CP4 EPSPS proteins, each expressed in E. coli. Each protein was administered by oral gavage. The highest doses tested were 572 mg/kg body weight for the mature protein and 1028 mg/kg body weight for the alternatively processed protein. Monsanto reports that no acute toxicity resulted from the oral administration of either protein in male or female mice at any of the doses administered in the experiments.

Monsanto compared the amino acid sequences of the CP4 EPSPS proteins expressed in the grain of event MON 71800 to the amino acid sequences of proteins (including toxins) available in the database ALLPEPTIDES using FASTA algorithm. Monsanto concluded that its search showed no relevant similarities between the CP4 EPSPS proteins present in event MON 71800 and proteins that are known to cause adverse health effects in humans or animals.

5. Food and Feed Uses of Wheat

Monsanto describes historical and current uses of wheat in food and animal feed. Wheat has been cultivated for use in food for several millennia. In the U.S., wheat is one of the most abundant crops in terms of planted acreage. Wheat grain contains approximately 60% carbohydrate, 10-16% protein, 2% fat, and 13% water. It is mainly used for the production of flour which is used in baked goods or other flour-based foods. U.S. consumers currently consume approximately 143 pounds of wheat flour per capita per year.

The use of wheat as animal feed is minor when compared to its use as food. Wheat grain is used as feed for poultry, swine, and cattle. It is estimated that on average 13 percent of total U.S. wheat grain has been utilized for animal feed in recent years. Wheat forage is also used as an animal feed source and is used for grazing cattle over the winter months in certain regions of the U.S.

Monsanto is not aware of any food or feed uses of standard spring wheat varieties that are not also applicable to spring wheat varieties containing event MON 71800.

6. Compositional Analysis of Roundup Ready® Wheat

Monsanto compared MON 71800 wheat to the parent line (Bobwhite MON 71900) and to commercially available non-transgenic spring wheat varieties.

6.1 Overview of the Approach to Compositional Analysis

To assess whether Roundup Ready® wheat event MON 71800 is as safe and nutritious as wheat varieties currently consumed, Monsanto conducted compositional analyses of grain and forage from event MON 71800, the non-transgenic parent line MON 71900 (Bobwhite), and several commercial wheat varieties. To conduct these compositional analyses, Monsanto collected both grain and forage samples from field trials and purchased commercially available grain produced from certified seed.

Monsanto planted event MON 71800, the parental line MON 71900, and 18 different commercial lines at five North American sites (three sites in the U.S. and two sites in Canada). Monsanto considered that these five sites provided a variety of environmental conditions representative of regions where Roundup Ready® wheat can be grown as a commercial product. Monsanto analyzed 76 components in grain collected from event MON 71800, the parental line MON 71900, the 18 different commercial spring wheat lines that Monsanto grew as part of its field trials, and grain from 4 spring wheat varieties purchased from certified seed producers. Monsanto also analyzed 9 components in forage tissues collected from event MON 71800, the parental line MON 71900, and the 18 different commercial spring wheat lines that Monsanto grew as part of its field trials.

Monsanto used standard analytical methods or other suitable methods for compositional analyses and provides references and descriptions for all analytical methods used. Monsanto provided the analytical results for the five test sites individually, and for all the sites combined. Monsanto reported the level of each analyzed component (mean and range) in the transgenic line MON 71800 and non-transgenic control line MON 71900; the difference between the level of each component (mean and range) obtained for line MON 71800 and line MON 71900; the range of levels obtained for 22 commercial lines; and the historical range based on values in the literature.

Monsanto subjected all of the compositional data obtained for Roundup Ready® wheat event MON 71800 and the parental control line MON 71900 to statistical analysis using a mixed model analysis of variance. Monsanto conducted the statistical analysis on data from each of the five replicated trials, as well as on data combined from all five trials.

6.2 Grain

Monsanto determined the levels of the following components of wheat grain:

Proximates: protein, fat, carbohydrate (by calculation), ash and moisture
Sugars: arabinose, fructose, galactose, glucose, maltose, raffinose, sucrose and xylose
Total dietary fiber (TDF)
Amino acid composition
Fatty acid composition (16:0 palmitic, 16:1 palmitoleic, 18:0 stearic, 18:1 oleic, 18:2 linoleic, 18:3 linolenic, 20:0 arachidic, and 20:1 eicosenoic)
B vitamins: niacin, riboflavin (vitamin B₂), thiamin (B₁), and vitamin B₆
Vitamin E
Minerals: cadmium, calcium, copper, iron, magnesium, manganese, phosphorus, potassium, selenium, and zinc
Starch

Monsanto also analyzed wheat grain for 14 other fatty acids, sodium, and the sugars mannose and stachyose; however, these substances were not detected in any samples tested (i.e., either from line 71800, line 71900, or commercial varieties).

Based on the combined statistical analysis of data from all test sites, Monsanto did not identify any statistically significant differences in grain composition between the event MON 71800 line and the parental MON 71900 line. Monsanto did identify some statistically significant differences (p<0.05) in composition between MON 71800 and MON 71900 at individual sites. In total, Monsanto found small differences in the levels of 16 components. However, these differences were found at only one or two test sites, and were not consistent over all trials. Monsanto also reported that for grain, all measured parameters fell within the range detected by Monsanto for the commercial varieties. The values also fell within the range found in the literature, except for the two amino acids, serine and valine; however, there were no differences in the levels of these amino acids when comparing MON 71800, MON 71900, or the commercial wheat varieties. Monsanto concludes that grain from event MON 71800 is compositionally equivalent to that of the non-transgenic parental control wheat and other wheat varieties grown commercially.

6.3 Forage

Monsanto determined the levels of the following components of wheat forage:

Proximates: protein, fat, carbohydrate (by calculation), ash and moisture
Fiber: acid detergent fiber, neutral detergent fiber
Minerals: calcium and phosphorus

Based on the combined statistical analyses from individual test sites, Monsanto identified no statistically significant differences (p<0.05) in composition between forage samples from MON 71800 and MON 71900. In the individual tests, Monsanto found significant (p<0.05) differences in the levels of five components from two out of five test sites. However, the differences were small and were not consistent over all locations as indicated by the combined analysis. In addition, the range of values for those components fell within the interval of values determined for the commercial varieties and within values reported in the literature.

Monsanto concludes that the forage from event MON 71800 is compositionally equivalent to that of the non-transgenic parental control wheat and other wheat varieties grown commercially.

6.4 Antinutrients

Phytic acid is an antinutrient that occurs naturally in wheat. Monsanto measured phytic acid in the grain of all test and control lines. The mean level of phytic acid in event MON 71800 was comparable to that of the parental line MON 71900 and fell within the range established for the 22 commercial lines and within the historical range for wheat based on values in the literature.

6.5 Endogenous Allergens

Because wheat is known to cause allergic reactions in sensitive individuals, Monsanto performed an evaluation to assess whether the transformation process may have increased the overall allergenicity of wheat grain from event MON 71800. Monsanto conducted IgE-inhibition ELISAs and IgE-immunoblot analysis with extracts from event MON 71800, parental MON 71900, and seven other commercial varieties of wheat. Sera used in these assays were obtained from ten human subjects with IgE-mediated allergic response to wheat consumption as evidenced by clinical history and response in single or double blind oral food challenge. From these experiments, Monsanto concluded that the transformation process did not significantly alter endogenous allergens in event MON 71800, because IgE binding properties were similar to those of other commercial wheat varieties.

6.6 Gluten

Celiac disease (gluten-sensitive enteropathy) is caused by a specific immune response to antigens present in gluten in susceptible individuals. Gluten refers to a mixture of glutenin and gliadin proteins present in wheat, barley and rye. Monsanto measured gliadin levels and calculated gluten levels in event MON 71800, parental MON 71900, and 22 other commercial wheat varieties. Gliadin and gluten levels in event MON 71800 are comparable to those of the parental and other commercial varieties tested.

6.7 Summary of Compositional Analysis

Monsanto concludes that the results of its compositional analysis established that the levels of nutrients and other components of the grain and forage derived from event MON 71800 fall within the ranges found for commercially available wheat varieties. Monsanto considered that the few minor differences in the levels of certain components seen at individual test sites are unlikely to be meaningful, and that grain and forage from event MON 71800 are compositionally equivalent to that of the non-transgenic parental control wheat and other wheat varieties grown commercially.

7. Conclusions

Monsanto has concluded that Roundup Ready® wheat event MON 71800 is not materially different in composition, safety, or any other relevant parameter from wheat now grown, marketed, and consumed. At this time, based on Monsanto’s data and information, the agency considers Monsanto’s consultation on Roundup Ready® event MON 71800 wheat to be complete.

Jason Dietz