Exposure to aflatoxin B1 attenuates cell viability and induces endoplasmic reticulum-mediated cell death in a bovine mammary epithelial cell line (MAC-T)

Park, Wonhyoung,Park, Min Young,Song, Gwonhwa,Lim, Whasun Elsevier 2019 Toxicology in vitro Vol.61 No.-

<P><B>Abstract</B></P> <P>Mammary gland epithelial cells play a key role in milk production in dairy cattle, which are prone to feed contamination from microorganisms, especially pathogenic fungi and their mycotoxin products. When mycotoxins enter the body, they cause a reduction in feed intake and milk production, leading to the retardation of growth and productivity in dairy cattle. Among them, aflatoxin B1 is a ubiquitous food contaminant frequently found in stored crops, which are used as animal feed, and causes many adverse health effects in domestic animals. Unfortunately, there is a lack of knowledge on the mechanisms underlying the detrimental effects of aflatoxin B1. Therefore, in this study, we found that aflatoxin B1 reduced cellular proliferation, induced apoptosis by DNA fragmentation, and disrupted intracellular homeostasis, including mitochondrial membrane potential and calcium concentration, in an immortalized bovine mammary epithelial cell line (MAC-T). Additionally, we identified aflatoxin B1-mediated cell signaling pathways associated with cell survival and the endoplasmic reticulum stress response. These results revealed that exposure to aflatoxin B1 attenuates cell viability and induces endoplasmic reticulum-mediated cell death in MAC-T cells.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Mechanisms underlying the detrimental effects of aflatoxin B1 were studied. </LI> <LI> Aflatoxin B1 reduced cellular proliferation and induced apoptosis. </LI> <LI> It disrupted intracellular homeostasis. </LI> <LI> Aflatoxin B1 regulated cell survival related signaling pathways. </LI> <LI> It also regulated ER stress response related signaling pathways. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>Schematic illustration of the cell signaling pathways induced by aflatoxin B1 treatment in bovine mammary epithelial cells. Aflatoxin B1 exposure increases ER stress by activation of GRP78, ATF6α, IRE1α, PERK, and eIF2α proteins. Aflatoxin B1 also suppresses cell viability through inhibition of the PI3K and ERK1/2 cell signaling pathways, whereas it activates the mitochondrial-mediated apoptotic pathway. Taken together, exposure to aflatoxin B1 induces cell death in bovine mammary gland cells.</P> <P>[DISPLAY OMISSION]</P>

Aflatoxin B₁에 대한 유산균의 결합력에 영향을 미치는 배양조건과 물리화학적 인자

임성미,안동현,Lim, Sung-Mee,Ahn, Dong-Hyun 한국미생물학회 2013 미생물학회지 Vol.49 No.3

본 연구에서는 숙성된 된장으로부터 분리된 유산균에 의한 aflatoxin $B_1$의 결합 정도를 배양조건에 따라 측정하였고, 물리화학적 처리조건이 aflatoxin $B_1$에 대한 유산균 세포의 결합력에 미치는 영향을 살펴보았다. Enterococcus faecium DJ22, Lactobacillus fermentum DJ35, Lactobacillus rhamnosus DJ42 및 Lactobacillus pentosus DJ47는 19.3-52.1% 정도의 aflatoxin $B_1$ 결합 효과를 나타내어 균종에 따라 결합력에 차이가 있었다. 하지만 E. faecalis DJ14, Lactobacillus panis DJ29 및 Pediococcus halophilus DJ50 균주는 aflatoxin $B_1$에 대한 결합력을 나타내지 않았다. Aflatoxin $B_1$에 대한 유산균의 결합력과 결합속도는 독소의 농도, 반응시간 및 온도와 초기 세포수 등의 배양 조건에 따라 유의한 차이가 있었다. Aflatoxin $B_1$의 결합력은 세척 횟수에 따라 현저하게 감소하였고, 감소율은 살아있는 세포와 가열 처리한 세포에서 비슷하게 나타났다. 가열, 산성 pH, ${\alpha}$-amylase, protease, lysozyme 혹은 sodium metaperiodate의 처리에 의해 결합력이 유의하게 감소된 것으로 보아 주로 세포벽에 존재하는 당이나 단백질에 aflatoxin $B_1$이 결합되며, urea의 처리에 의해 결합력에 낮아지는 것은 이들 사이에는 소수성 결합이 작용하는 것으로 추정되었다. The purpose of this study was to investigate the aflatoxin $B_1$ binding of lactic acid bacteria (LAB) isolated from Korean traditional soybean paste and to evaluate the effect of incubation conditions and physico-chemical factors on the binding ability of LAB to this mutagen. The amount of aflatoxin $B_1$ bound by Enterococcus faecium DJ22, Lactobacillus fermentum DJ35, Lactobacillus rhamnosus DJ42, and Lactobacillus pentosus DJ47 was strain specific with the percent bound ranging from 19.3% to 52.1%. However, Enterococcus faecalis DJ14, Lactobacillus panis DJ29, and Pediococcus halophilus DJ50 strains did not exhibit any of the binding ability to aflatoxin $B_1$. For most strains, the binding ability was significantly affected by the environmental conditions such as the aflatoxin $B_1$ level, incubation time and temperature, and the initial cell count of LAB. The stability of the aflatoxin $B_1$-bacteria complexes was significantly more unstable after washing. In addition, the binding stability between viable and nonviable cells was not statistically significant. Treatment with heating, acidic pH, ${\alpha}$-amylase, protease, lysozyme, or sodium metaperiodate caused a significant (P<0.05) decrease in aflatoxin $B_1$ binding for the tested strains, suggesting that carbohydrates or proteins in the cell walls may be involved in aflatoxin $B_1$ binding ability. Since the aflatoxin $B_1$ binding of LAB was significantly reduced (P<0.05) by the pretreatment of the urea, the binding force observed in this study may have resulted from hydrophobic interaction.

생약추출물이 Aspergillus parasiticus Aflatoxin B1 생성에 미치는 영향

정상진 한국약용작물학회 2003 한국약용작물학회지 Vol.11 No.5

본 연구에서 8종의 생약 추출물이 A. parasiticus의 배양시 aflatoxin B1 생성에 미치는 효과를 조사하였다. 배지의 pH는 배양 3일 후에 모든 생약 추출물이 pH 4 이하를 나타냈으며 구기자, 오매, 계피, 두충은 배양 6일에 다시 pH 4 이상으로 상승하였고 이중 대추가 배양 기간 중 가장 낮은 pH를 나타냈다. 균체 생성량은 모든 실험군이 대조군보다 높았으며 갈근, 두충, 오미자, 대추, 오매,구기자, 목과의 순으로 나타났다. 이중 갈근이 최대 생성량을 나타냈으며 목과가 가장 낮은 생성량을 나타냈다. Aflatoxin B1은 갈근과 대추 추출물을 제외한 모든 실험군에서 생성이 억제되었다. 특히 계피, 오매, 두충 구기자, 오미자 추출물 에서 aflatoxin B1 생성이 현저히 저하되었으며 계피가 가장 큰 억제 효과를 나타냈다. 균체량이 많이 생성되면 aflatoxin B1 생성이 적어지고 균체량이 적게 생성되면 aflatoxin B1의 생성이 많아졌다. Aspergillus parasiticus에 의해 aflatoxin B1을 가장 적게 생성하는 계피와 가장 많이 생성하는 갈근 추출물의 총단백질 생성량은 계피는 3일째 (34.5%), 갈근 ext ract는 4일째 (36.4%) 총단백질의 함량이 최대가 되어 균체 내의 총단백질의 함량은 계피와 갈근 추출물을 첨가한 시험군이 게조군의 총단백질 함량 (32.7%)보다 약간 많았으며 aflatoxin B1 생성 및 축적이 최대가 되는 시기는 총단백질량이 최대가 되는 시기보다 대체로 1일 정도 늦게 나타났다. The influences of the extracts from Cinnamomi Cortex, Eucommiae Cortex, Puerariae Radix, Lycii Fructus, Zizyphi Fructus, Schisandrae Fructus, Mume Fructus, Chaenomelis Fructus on mycelial growth and aflatoxin B1 production from Aspergillus parasiticus were analyzed. The pH of the culture media were reduced to below pH 4 by all the herbal extracts after 3 days incubation. However, the pH of the culture media increased above pH 6 after 6 days incubation using the extracts from Cinnamomi Cortex, Eucommiae Cortex, Puerariae Radix and Lycii Fructus. The mycelial growth of A. parasiticus was increased over the amount of the control. Puerariae Radix produced the largest amount of mycelial growth and Chaenomelis Fructus produced the smallest amount of mycelial growth. The productions of aflatoxin B1 from A. parasticus culture were increased by the extracts of Puerariae Radix and Zizyphi Fructus, while inhibited by the extracts of Cinnamomi Cortex, Eucommiae Cortex, Lycii Fructus, Schisandrae Fructus, Mume Fructus and Chaenomelis Fructus. In particular, the extracts of Cinnamomi Cortex, Lycii Fructus and Schisandrae Fructus almost inhibited the production of aflatoxin B1. The production of the total protein from Cinnamomi Cortex, which produced much less aflatoxin B1, and Puerariae Radix, which produced a great deal of aflatoxin B1 from A parasticus were slightly higher than the production of the total protein of the control medium.

곡류 중 아플라톡신 B1의 위해성평가

이효민,윤은경,최윤호,이근영,조연숙,임철주,김종욱,양지선,양기화 식품의약품안전청 2001 식품의약품안전청 연보 Vol.5 No.-

Aflatoxins은 fspergijfus fovur,4.forofj'cus,4.uonlius의 B차 대사산물로써, 유전독성을 지닌 강력한 발암물질로 알려지고 있으며, 간장, 신장, 신경계, 내분비계, 면역계 독성을 지니고있다. Aflatoxin은 열에 대한 저항성o'S 매우 커서 일반적인 열처리에 의해서 쉽게 파괴되지 않고, 식품섭친패턴이나 기후언1 따라 인체노출량이 상이하여 나라별 1일인체노출량이 3.5-Tln토Ag/day 정도인 것으로 보곤되고 있다. 본 연구에서는 aflatoxin중 가장 대표적인 Bl을 대상으로 한 위해성평가방땁론 연구를 통해 국내 곡류중 aflatoxin Bl 오염도자료와 식품섭취 패턴을 고려한 위해성평가를 수행하고 우리나라 국민들의 식품섭취와 관.된 aflatoxin 위해정도를 고려핫으로 그 관리대안을 제시코자 하였다. Aflatoxin의 인체노출평가를 위하여 우리나라 일반성인들을 대상으로 국민건강영양조사결과보고서에 제시된 식품소비량자료와 국내에써 조사된 바 있는 곡류, 두류, 견과류의 식붐오염도자료(1989-2000)를 활총하였다. 식품오염도자료는 결과의 대표성을 부여하기 위하여 bootstrap방법을 활용하여 정확도 검정을 실시함으로 얻어진 평근값을 활용하였다. 위해도결정에 쓰여진 a(latoxin 31의 발암력은 Yeh 등 t1989)이 제안한 자료로 B형간염비보균자의 경우 9(mg/kg/day)-'. B형간염보균자의 경우 230(mgAg/daH)-'을 사응하였다. 계산죈=ftatoxin Bl의 총인체노출량은 6.05×10-img/kg/'day였고, 인체노출량에 발암력을 고려하므로 계산된추골나라 일반성인에게 aflatoxin Bl식이노출로 나타날 수 있는 간암발생확률은 B형간암fT보균자의 경우차 보균f·『의 경우 각각 5.45×10-"와 1.39r10-'로 산출되었다. 머린이나 노약자, 그궈고 B형간염보균자 등을 포함한 민감그룹과 aflatorin B2, Cl, G2 등의 동시노출을 고려한다딴 식픔섭취를 통한 a긴atoxiri의 인체위해수준은 더 커질것으로 예측된다. 실제로 B형간염보균자의 경우, 식이섭취를 통한 만성 a친atoxin Bl 노출로 인해 간암이 발생할 확률이 비보균자에 비해 26배 정도 높은 것으로 나타나 철저한 씩이 관리가 필요하다. 일반성인(B형간염비보군자)의 경우, aflatoxin 31곡 대글 관리목표치를 십만명당 1명으로 할 경우 인체노출수준은 1.Ix10-srng/kg/day 이하로 조절되어야하고, 우리나라 일반성인의 1일 곡류, 두류, 견과류 섭취량 393.46g./da!·(보건복지부, 1999)과 체중(60kg)을 고려하고, 조리시 감소콘는 오염수준, aflatoxin에 오염된 식픔을 동시에 섭취할 가능성의 배제, 현실적 인 분석기술 수준의 미흡, 그리고 현기준과 비교된 비용-편익적인 면엔서의 문제점을 고려한다만, 곡류, 두류, 견과류 등을 포함한 식품들의 aflatorin Bl 최대오염수준은 0.2ppb이하로 조정되어야 할 것을 제아할 수 있다 Aflatoxins are unaviodable contaminats of corn, peanuts, rice, barley, soybean and other agricultural products that are stored under warm and humid conditions for sometime. The prominent aflatoxins found on food is aflatoxin B1, Which has most potent toxicological and carcinogenic properties of the aflatoxins. The range of human daily intake has been reported as 3.5-77ng/kg/day from various countries and human daily intake was mainly affected by food consumption pattern and climate of each other country. This study was conducted to identify excess cancer risk on aflatoxin B1 induced by domestic cereals ingestion and to suggest regulatory option on aflatoxin B1 for management based on acceptable risk value. For the quantification of human exposure dose to general adult, aflatoxin pollution data in foods were collected(1989-2000)and more accurate mean±data were obtained by bootstrapping, using the S-plus^()(Mathsoft) and also, daily food consumption data from National Health and Nutrition Survey Report(Minstry of Health and Welfare, 1999) and body weight(60kg, Korea Reserch institute of Standard and Science, 1997) were regarede. The used cancer potency of aflatoxin B1 was 9(mg/kg/day)^(-1) for individuals negative for hepatitis B and 230(mg/kg/day)^(-1) for individuals positive for hepatitis B suggested from epidemiological study by Yeh et al(1989). Estimated human exposure dose of aflatoxin B1 by cereals ingestion were 5.89×10^(-7)mg/kg/day and excess cancer risk value to liver cancer incidence were 5.30×10^(-6) for individuals negative for hepatitis B and 1.4×10^(-4) for individuals positive for hepatitis B. Individuals positive for hepatitis B revealed 26 fold higher excess liver cancer risk than individuals negative and therefore, they should be managed having special consideration. And, if the excess cancer risk for aflatoxins B1 is 10^(-5) as managing goal, human exposure level should be controlled as below 1.1×10^(-6)mg/kg/day. Thus, this study has suggested regulatory option as below 0.2ppb for aflatoxin B1 in cereals, legumes. and nuts.

아플라톡신 B1 섭취가 위암 발생에 미치는 영향

윤정국,강종원,김헌 충북대학교 의과대학 충북대학교 의학연구소 2003 忠北醫大學術誌 Vol.13 No.2

연구목적: 콩과류 식물에서 많이 생성되는 아플라톡신 B1은 강력한 발암물질로서 간암 등을 유발하는 것으로 알려져 있다. 최근 여러 연구에서 아플라톡신 B1이 폐암이나 위암을 일으킬 가능성이 있다는 보고가 있으나 음식물을 통한 아플라톡신 B1의 섭취와 위암의 관련성에 대한 연구가 위암발생률이 높은 우리 나라에서도 아직까지 거의 이루어지지 않고 있다. 또한, 아플라톡신 B1 섭취가 간조직의 p53과 Ki-ras 유전자의 특정 돌연변이를 일으킨다는 연구가 있으나, 현재 우리나라에서는 이러한 연구가 아직까지 이루어지지 않고 있다. 따라서, 본 연구는 음식물을 통한 아플라톡신 B1 섭취와 한국인 위암과의 관련성과 아플라톡신 B1 섭취와 p53 유전자 및 Ki-ras 유전자 돌연변이와의 관련성을 알아보고자 하였다. 대상 및 방법: 위암 환자 214명과, 이들과 성별과 나이를 짝지은 대조군 214명을 이 연구의 대상으로 하였다. 반정량적 설문을 이용하여 89개 식이항목에 해당하는 음식 섭취량을 직접면접을 통해 측정하였고, 이들 식이항목을 수집하여 경쟁적 효소면역반응법(competitive ELISA)으로 음식물에 함유된 아플라톡신 B1의 양을 정량화 하였다. 이 결과를 이용하여 대상자들의 아플라톡신 B1 섭취량을 추정하고 위암 환자군과 대조군 사이에 아플라톡신 B1 섭취량에 차이가 있는지를 검정하였다. 또한, 위암환자의 종양조직에서 추출한 RNA를 이용하여 RT-PCR을 시행하여 염기서열을 분석하는 방법으로 p53와 Ki-ras 돌연변이를 검출하고, 아플라톡신 B1 섭취량에 따른 이들 유전자 돌연변이의 차이 여부도 조사하였다. 결과: 칼로리를 보정한 주당 총 아플라톡신 B1 평균 섭취량은 위암 환자군과 대조군에서 각각 6,714.08ng과 6,312.65ng으로 유의한 차이를 나타냈다(p=0.009). 주당 아플라톡신 B1 섭취량을 저섭취군과 고섭취군의 2단계로 나누어 OR을 산출한 결과 아플라톡신 B1 고섭취량군의 OR(95% CI)이 1.54(1.05-2.58)로 유의하게 증가하였다. 환자군을 대상으로 p53과 Ki-ras 유전자 돌연변이와 아플라톡신 B1 섭취량의 관계를 검정한 결과 아플라톡신 B1 섭취정도에 따라 두 유전자의 돌연변이 빈도는 유의한 차이가 나타나지 않았다. 결론: 본 연구 결과는 아플라톡신 B1의 섭취가 위암의 위험인자임을 시사하는 것으로 사료된다. 그러나, 아플라톡신 B1 섭취가 p53이나 Ki-ras 돌연변이를 유발하는 것은 아닌 것으로 보인다. Puorpose: Aflatoxin B1, which is toxic, mutagenic and carcinogenic has been suspected of contributing to human hepatocellular carcinoma. In recent researches aflatoxin B1 intake have been reported to be related with lung cancer and stomach cancer. However, studies on the associations between dietary intake of aflatoxin B1 and stomach cancer risk are scarce. We have assessed the relationship between dietary intake of aflatoxin B1 and the risk of stomach cancer in Koreans. Materials and Methods: Two hundred fourteen pathologically-proven stomach cancer patients and 214 age-matched controls were included as the study subjects. Information about food item and dietary intake was collected by direct interview. Level of aflatoxin B1 was determined in 89 Korean food items using competitive ELISA. Intake amount of aflatoxin B1 was estimated by multiplying intake amount of food items and average concentration of aflatoxin B1 in the food items. Detection of p53 and Ki-ras gene mutation using RT-PCR direct sequencing. Results: The mean aflatoxin B1 intake amount was significantly higher in stomach cancer patients than in controls(OR:1.54 95% CI: 1.05, 2.28). p53 and Ki-ras gene mutations were not significantly related with calorie-adjusted intake amount of aflatoxin B1. Conclusion: It is suggest that dietary intake of aflatoxin B1 would be a risk factor for stomach cancer in Koreans and low level aflatoxin intake would not induce p53 or Ki-ras mutation in stomach tissue.

유통 건고추 및 고춧가루의 아플라톡신 B1과 오크라톡신 A 오염도 조사

제갈승,김지형,주광식,정세진,나현주,조남규,이제만,김용희 한국식품위생안전성학회 2013 한국식품위생안전성학회지 Vol.28 No.3

2012년 2월부터 11월까지 인천 지역에서 유통된 건고추및 고춧가루 193건을 대상으로 아플라톡신 B1과 오크라톡신 A의 오염도를 조사하였다. Immunoaffinity column 및HPLC를 이용한 시험법은 모두 80% 이상의 회수율을 보였고, 아플라톡신 B1 및 오크라톡신 A의 검출한계는 각각0.13 μg/kg, 0.30 μg/kg였다. 오염도 조사를 한 결과 아플라톡신 B1은 17.1%의 검출율을 보였고 오크라톡신 A는 20.7%의 검출율을 보였으며, 아플라톡신 B1의 검출농도는 0.14~9.67 μg/kg였고, 오크라톡신 A의 검출 농도는 0.31~3.31 μg/kg였다. 이는 우리나라 식품공전 상의 기준인 10 μg/kg(아플라톡신 B1), 7 μg/kg(오크라톡신 A)보다는 낮은 수치로비교적 안전한 수준이었다. A survey of aflatoxin B1 and ochratoxin A was conducted on dried red pepper and red pepper powder. Total number of 193 samples were collected from local markets in Incheon. The presence of aflatoxin B1 and ochratoxin A was determined by high performance liquid chromatography (HPLC) with fluorescence detector using immunoaffinity column clean-up. The recovery rate of aflatoxin B1 and ochratoxin A were more than 80% and the limits of quantification were 0.13 μg/kg for aflatoxin B1 and 0.30 μg/kg for ochratoxin A. Aflatoxin B1 was detected in 33 samples (17.1%) with a range of 0.14~9.67 μg/kg and ochratoxin A was detected in 40 samples (20.7%) with a range of 0.31~3.31 μg/kg. These results show that the occurrence of aflatoxin B1 and ochratoxin A in dried red pepper and red pepper powder tested in this study is low compared with the standard in Korea Food Code (10 μg/kg as aflatoxin B1 and 7 μg/kg as ochratoxin A).

유통 건고추 및 고춧가루의 아플라톡신 B₁과 오크라톡신 A 오염도 조사

제갈승,김지형,주광식,정세진,나현주,조남규,이제만,김용희,Jegal, Seung,Kim, Ji-Hyeung,Joo, Gwang-Sig,Jung, Se-Jin,Na, Hyeon-Ju,Jo, Nam-Gyu,Lee, Jea-Man,Kim, Yong-Hee 한국식품위생안전성학회 2013 한국식품위생안전성학회지 Vol.28 No.3

2012년 2월부터 11월까지 인천 지역에서 유통된 건고추 및 고춧가루 193건을 대상으로 아플라톡신 $B_1$과 오크라톡신 A의 오염도를 조사하였다. Immunoaffinity column 및 HPLC를 이용한 시험법은 모두 80% 이상의 회수율을 보였고, 아플라톡신 $B_1$ 및 오크라톡신 A의 검출한계는 각각 0.13 ${\mu}g/kg$, $0.30{\mu}g/kg$였다. 오염도 조사를 한 결과 아플라톡신 $B_1$은 17.1%의 검출율을 보였고 오크라톡신 A는 20.7%의 검출율을 보였으며, 아플라톡신 $B_1$의 검출농도는 0.14~9.67 ${\mu}g/kg$였고, 오크라톡신 A의 검출 농도는 0.31~3.31 ${\mu}g/kg$였다. 이는 우리나라 식품공전 상의 기준인 10 ${\mu}g/kg$(아플라톡신 $B_1$), 7 ${\mu}g/kg$(오크라톡신 A)보다는 낮은 수치로 비교적 안전한 수준이었다. A survey of aflatoxin $B_1$ and ochratoxin A was conducted on dried red pepper and red pepper powder. Total number of 193 samples were collected from local markets in Incheon. The presence of aflatoxin $B_1$ and ochratoxin A was determined by high performance liquid chromatography (HPLC) with fluorescence detector using immunoaffinity column clean-up. The recovery rate of aflatoxin $B_1$ and ochratoxin A were more than 80% and the limits of quantification were 0.13 ${\mu}g/kg$ for aflatoxin $B_1$ and 0.30 ${\mu}g/kg$ for ochratoxin A. Aflatoxin $B_1$ was detected in 33 samples (17.1%) with a range of 0.14~9.67 ${\mu}g/kg$ and ochratoxin A was detected in 40 samples (20.7%) with a range of 0.31~3.31 ${\mu}g/kg$. These results show that the occurrence of aflatoxin $B_1$ and ochratoxin A in dried red pepper and red pepper powder tested in this study is low compared with the standard in Korea Food Code (10 ${\mu}g/kg$ as aflatoxin $B_1$ and 7 ${\mu}g/kg$ as ochratoxin A).

영남지방 농상물에 대한 위생학적 연구(제2보) ELISA 법에 의한 Aflatoxin B1 검색

정덕화,하기수,정혜경,김동술,James J. Pestka 한국식품위생안전성학회 1989 한국식품위생안전성학회지 Vol.4 No.3

영남지방 곡류에 오염되어 있는 Aflatoxin B1을 측정하기 위해 식속하고 정확한 Enzyme-Linked Immunosorbent Assay(ELISA) 법을 응용하였다. 생산된 Antibody 는 aflatoxin B2와 강한 corss reaction을 보였고, 정량적으로 표준곡선을 보인 aflatoxin B1 범위는 1-100ppb이었으며, methanol-PBS-dimethylformamide로 spike시킨 쌀 추출물이 ELISA 반응에 약간의 간섭반응을 보였다. ELISA법에 의한 쌀 65종, 보리 116종의 aflatoxin 분석결과는 다음과 같다. 즉, 쌀시료 65개 중 3개의 시료가 positive 반응을 보였으며, 그중 R-18이 $7.5\;\mu\textrm{g}/kg$, R-30이 $3.6\;\mu\textrm{g}/kg$, R-59가 $3.3\;\mu\textrm{g}/kg$ 순으로 aflatoxin B1 함량을 나타내었고 시료의 4.6%가 aflatoxin B1에 오염되었음을 보였다. 보리 116종 중 겉보리에서 4개의 시료가 positive 반응을 나타내었으며, 그 중 겉보리 UB-37이 $9.6\;\mu\textrm{g}/kg$의 함량으로 가장 높은 함량을 나타내었고, 그의 UB-35, UB-54순이었으며 전체 시료의 3.4%가 aflatoxin B1에 오염되었음을 보였다. A rapid, simple method of ELISA was applied for the determination of aflatoxin BI in cereals from Y oungnam districts. Antibodies obtained cross reacted with aflatoxin B2 and to a less extent with other aflatoxin BI analogs. Response range for a typical standard curve was between I and 100 ppb. Fewer interference by spiked methanol-PBSdimethylformamide extracts ofrice was evidenced. Contents of aflatoxin BI from rice (65) and barley (116) were determined by competitive direct enzyme- linked immunosorbent assay as follows. Three out of 65 rices samples were positive. Rice samples of R-IS, R-30, and R-59 represent the aflatoxin B1 levels of $7.5\;\mu\textrm{g}.kg,\;6.0\;\mu\textrm{g}/kg,\;3.5\;\mu\textrm{g}/kg,\;3.3\;\mu\textrm{g}/kg$, respectively, and showed 4.6% aflatoxin BI contamination in rice samples. Meanwhile, four out of 116 barley samples were positive. VB-37 showed the highest aflatoxin Bllevels of $9.6\;\mu\textrm{g}/kg$ and VB-35, VB-15 and VB-54 represent $7.5\;\mu\textrm{g}.kg,\;6.0\;\mu\textrm{g}/kg\;and\;3.6\;\mu\textrm{g}/kg$, respectively, and showed 3.4% aflatoxin B1 contamination in barley samples.

Development of aflatoxin B₁ aptasensor based on wide-range fluorescence detection using graphene oxide quencher

Joo, Minyoung,Baek, Seung Hoon,Cheon, Seon Ah,Chun, Hyang Sook,Choi, Sung-Wook,Park, Tae Jung Elsevier 2017 Colloids and surfaces. B, Biointerfaces Vol.154 No.-

<P><B>Abstract</B></P> <P>Aflatoxin B<SUB>1</SUB> (AFB<SUB>1</SUB>) is a carcinogenic substance produced by fungi of genus <I>Aspergillus</I>, especially <I>Aspergillus flavus</I>. Few nanograms of AFB<SUB>1</SUB> that permeated through the skin is sufficient to cause liver cancer and stunted growth. In this study, a rapid aptamer-based assay for AFB<SUB>1</SUB> was developed using the fluorescence quenching property of graphene oxide (GO) and a fluorescein amidite (FAM)-modified aptamer specific to AFB<SUB>1</SUB>. The aptamer, modified with the fluorescence dye FAM on its 5′-end, was used as a probe. Once bound by AFB<SUB>1</SUB>, a conformational change of the aptamer was caused that led to its interaction with the well-known fluorescence quencher GO, resulting in a decrease of the fluorescence intensity of the system. In the absence of AFB<SUB>1</SUB>, the fluorescence intensity remained unchanged. The aptamer-based AFB<SUB>1</SUB> assay process was conducted through 3 steps within 40min. The aptamer was incubated with AFB<SUB>1</SUB> before the addition of GO. The amount of AFB<SUB>1</SUB> present was measured by the change in fluorescence intensity. The detection system was evaluated with standard solutions of AFB<SUB>1</SUB> of various concentrations. The results showed that the fluorescence intensity decreased linearly as the concentration of AFB<SUB>1</SUB> gradually increased. Although the assay was specific to AFB<SUB>1</SUB>, there was slight interference by other types of aflatoxin. When the assay was applied to a real sample, the limit of detection was 4.5 ppb, which was within the wide detection range of up to 300ppb with good linearity. Thus, this biosensor is considered to be competitive with the conventional detection methods in the field owing to its wide detection range and assay rapidity.</P> <P><B>Highlights</B></P> <P> <UL> <LI> A simple and rapid detection for AFB1 mycotoxin produced by <I>Aspergillus flavus</I>. </LI> <LI> Fluorescence spectroscopy using organic fluorescent dye and graphene oxide quencher. </LI> <LI> Wide detection range of AFB1 with linear scale from 4.5 to 300ppb concentration. </LI> <LI> The limit of detection with 4.5ppb on real samples. </LI> </UL> </P> <P><B>Graphical abstract</B></P> <P>Graphene oxides were applied for the excellent signal-to-background ratio enhancer based on fluorescence quenching effect and interaction between FAM-labeled aptamers and aflatoxin B1 targets.</P> <P>[DISPLAY OMISSION]</P>

Aflatoxin $B_1$이 계배 간조직의 미세구조에 미치는 영향

조자향,전향미,서숙재,Cho, Ja-Hyang,Cheon, Hyang-Mi,Seo, Sook-Jae 한국현미경학회 1996 Applied microscopy Vol.26 No.2

To investigate the effect of aflatoxin $B_1$, on survival rate and ultrastructure of liver during chick embryogenesis electron microscopic methods were used. After injection of aflatoxin $B_1$ into the yolk, ultrastructural changes in the liver of chicken embryo were observed. The results were as followed. 1. 12-day old chicken embryos were treated with single injection of aflatoxin $B_1$ with the dose of $0.0005{\mu}g,\;0.005{\mu}g,\;0.05{\mu}g,\;0.5{\mu}g,\;2.5{\mu}g,\;5.0{\mu}g$ each. Chicken embryos treated with the dose of $0.5{\mu}g$ of aflatoxin $B_1$ had survival rate of 22%. The embryos treated with $2.5{\mu}g$ of aflatoxin $B_1$ hardly survived. 2. Chicken embryos treated with $0.05{\mu}g$ of afatoxin $B_1$ had hatched in 30%, but once hatched, they all survived. 3. After administration of $0.05{\mu}g$ of aflatoxin $B_1$ into the 12-day old chicken embryo, the electron microscopic studies were examined during development stages. The nuclei of hapatocytes became irregularly shaped and the structures of endoplasmic reticulum were changed to spherical types at 20-day old chicken embryo. Also, mitochondria became to be dilated and severe fibrosis was induced in the cytoplasm. However, the hepatocytes became almost normal in 30-day old young chicken.