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1 36 6 Vol.36, No JOURNAL OF FISHERIES OF CHINA June, 2012 文章编号 : (2012) DOI: /SP.J 饲料中黄曲霉毒素 B 1 对凡纳滨对虾生长肝胰腺和血淋巴生化指标及肝胰腺显微结构的影响, *,,, (, ) : B 1 (AFB 1 ) , μg/kg 6, ( ) g, 8 AFB 1 : AFB 1,, μg/kg μg/kg AFB μg/kg (304.67%), μg/kg, (58.33%)AFB μg/kg (T-AOC) (P<0.05), μg/kgs(gst) ATP(SOD), GST (P<0.05), AFB 1, AFB 1, AFB 1, μg/kg AFB 1 (95.49 μg/kg), AFB 1 : ; B 1 ; ; : S 917; S : A ( ),,,, tk-ur, B 1 B 2 G 1 G 2 4 B 1 (AFB 1 ), [1],, [2-3],,,,, El-Banna [4] 0.1 mg AFB 1 /kg 10, 0.2 mg AFB 1 /kg 16.7% Cagauan [5] μg AFB 1 /kg 90 d, 67%, Tuan [6] 250 μg AFB 1 /kg, 2.5 mg AFB 1 /kg Deng [7] 245 μg AFB 1 /kg 收稿日期 : 修回日期 : 资助项目 : B 通讯作者 : toguoran@163.com

2 6, : B 1 953, AFB μg/kg AFB 1 24 h AFB mg/l, AFB ~ ~ mg/kg,, AFB 1, AFB 1 400~2 000 μg/kg, 0 μg/kg AFB 1,, AFB 1, AFB (Litopanaeus vannamei),, 1.5 cm, ( ) g,, AFB 1 Sigma 1 1, AFB μg/kg, 1. 5 mm, 60, 4 h 20, 40 cm 50 cm 60 cm,, 30 6, 3 18, 30 6%~7%, 7:00 12:00 17:00, 1/3,,,,, ph ( ), ( ) mg/l, ph 8.0~8.5, ( ) mg/l 表 1 实验饲料配方及营养组成 Tab. 1 Formulation and composition of experimental diets % raw materials content proximate analysis content casein 37.0 moisture 9.70 gelatin 9.2 crude protein cellulose 20.7 crude lipid 6.33 others 33.1 total carbohydrate 7.38 crude ash 8.50 : 20%, 3.5%, 3.5%, 0.5%, 0.5%, 2.0%, (50%) 0.5%, Vc 0.5%, 0.1%, CMC 2.0% Notes: Others include corn starch 20%, fish oil 3.5%, corn oil 3.5%, soybean lecithin 0.5%, mineral mixture 0.5%, vitamin mixture 2.0%, choline(50%) 0.5%, ascorbic phosphate ester 0.5%, betaine 0.1%, CMC 2.0%. 1.3 AOAC [8], 24 h, 5,, 105 4, 105 [9] - ( ),, H.E [10] AFB 1 (ELISA), 1.4 (mean SE), Duncan, P<0.05,

3 SPSS AFB 1 3, 4 6 8, (P<0.05)( 2) 6, (P<0.05) AFB 1,, 5 6 (P<0.05) 4 6 表 2 AFB 1 对凡纳滨对虾生长及机体营养成分组成的影响 Tab. 2 Growth performance and body composition of L. vannamei fed with aflatoxin B 1 % AFB 1 /μg group aflatoxin B 1 survival rate weight gain crude lipid crude protein hepatopancrease ratio b cd a a b bc a c b ab a bc b a a ab b ab b ab a d b ab : (n=3); (P<0.05); =(W t W 0 )/W 0 100%; = /100%; = /100%; W 0, W t Notes: Values are means SE (n=3); Means with different superscript in each line are significant different (P<0.05); Weight gain= (W t W 0 )/W 0 100%; Survival rate = final shrimp/initial shrimp 100%; Hepatosomatic index (HSI) = liver weight/ body weight 100%; W 0 is initial average wet weight, W t is final average wet weight. 表 3 AFB 1 对凡纳滨对虾免疫机能的影响 Tab. 3 Immune function of L. vannamei fed with aflatoxin B 1 group hepatopancreas biochemical indicators /(U/mg prot) T-AOC c b b b b a S /(U/mg prot) GST d c b b a c ATP /(U/mg prot) ATPase e d c b a d (U/mg prot) SOD d c c b a b haemolymph biochemical indicators S /(U/mL) GST a a b ab ab c bc /(U/mL) SOD d d d c a b : (n=3)(p<0.05) 1U=1 Notes: Values are means SE (n=3). Means with different superscript in each row are significant different (P<0.05).1U=1 enzyme activity unit. 2.2 AFB 1 3, 2 5 S (GST) ATP (SOD) (P<0.05) 6 (T-AOC), AFB 1, GST SOD (P<0.05), GST, SOD, AFB 1, ( 1), 6 AFB 1 (P<0.05) AFB AFB 1 AFB 1,, 5 6, 3

4 6期王静, 等: 饲料中黄曲霉毒素 B1 对凡纳滨对虾生长肝胰腺和血淋巴生化指标及肝胰腺显微结构的影响 955 肝胰腺出现明显的破损图 2 显示实验进行 8 周后凡纳滨对虾肝胰腺组织切片与对照组相比, 实验组的肝细胞受到不同程度的损害, 可以明显看出, 第 3 组对虾肝胰腺的贮存细胞严重萎缩, 出现大片的空泡, 肝胰腺边缘的细胞尚完整, 第 5 组和第 6 组对虾肝胰腺出现细胞紧缩 3 讨论 Ostrowski-Meissner 等[11]的研究结果表明当饲料中 AFB1 为 μg/kg 时, 凡纳滨对虾成活率显著下降 μg/kg 时, 对虾开始厌食并伴随生长减缓, 体质量降低 μg/kg 时, 经 14 d 饲养对虾全部死亡本次实验饲料中 AFB1 在 0~1 600 μg/kg 时, 对虾的成活率未受到明显影响, μg/kg 时, 成活率明显降低当饲料中 AFB1 增加时, 对虾生长减慢增重率下降根据 Burgos-Hernandez 等[12] 的研究, AFB1 对凡纳滨对虾的毒性导致了消化能力降低和肝胰腺发育异常, 可能是由于霉菌毒素改变了胰蛋白酶及其他消化酶, 如酯酶和淀粉酶的活性, 从而导致对虾对营养素的消化吸收能力降低, 造成虾的摄食量减少虽第 6 组对虾增重率显著高于对照组, 但实验结果显示第 6 组 AFB1 在图 1 AFB1 在凡纳滨对虾肝胰腺和肌肉中的残留量 Fig. 1 Residues in hepatopancreas and muscle of L. vannamei fed with aflatoxin B1 肝胰腺中含量最高且观察到死亡对虾个体幼小, 夜间死亡对虾在次日清晨被发现时只剩下头部未被其他对虾摄食, 成活的对虾个体均较大, 从而使对虾增重率较高实验开始 4 周后, 第 4 至 6 组对虾的蜕皮次数多于其他组, 且实验期间死亡的对虾多为软壳虾产生这种现象的原因可能是由于对虾摄食较高浓度的 AFB1 使其应激蜕皮, 抵抗力减弱从而造成死亡, 接受高 AFB1 的虾表现为抗病力下降, 使对虾生产性能下降本实验结果显示对虾机体中粗蛋白含量并没有明显的变化, 虽粗脂肪含量有差异, 但各组之间的变化并不大且 AFB1 在肌肉中无残留, 因此 AFB1 对虾体营养成分的影响不显著 SOD 是重要的一种抗氧化酶, 具有催化超氧离子自由基进行歧化反应的功能, 可以消除氧自由基, 减少自由基对正常细胞的损伤, 使机体免受氧化伤害 Munoz 等 [13] 在凡纳滨对虾体内证实了超氧阴离子的存在 T-AOC 是机体抗氧化能力的总体反映根据 Bintvihok 等[14] 的研究, 实验 10 d 后, 饲料中低于 20 μg/kg 的 AFB1 已经导致对虾肝胰腺损伤和血淋巴的生化指标改变图 2 饲料中 AFB1 对凡纳滨对虾肝胰腺显微结构的影响 Fig. 2 Hepatopancreas microstructure of L. vannamei fed with AFB1 本研究的结果显示 AFB 1 会直接影响肝胰腺的总抗氧化能力, μg/kg 时影响力最大 AFB 1 凡纳滨对虾肝胰腺组织切片 100 第 4 组图片箭头所指为食道, 第通过影响可以清除自由基和抗氧化的酶, 以及 5 组图片箭头所指为空泡细胞, 第 6 组图片箭头所指为萎缩肝细胞肝脏的主要解毒系统, 从而破坏对虾正常的体 Digestive gland of shrimp biopsies 100. The arrow of the fourth group 内代谢, 使其抗病能力减弱, 对外界细菌及病 indicate esophagus, the fifth group arrows indicate vacuoles cells, and the sixth group arrows indicate atrophy of hepatopancreas cell. 毒的防御性降低对鱼类而言 AFB 1 最直接的毒

5 AFB 1 AFB 1-8,9- (AFBO) DNA DNA, S AFBO [15] GST,, GST,, AFB 1, GST GST AFB 1, GST, (Ictalurus punctatus), AFB 1,, T-AOC, 6, 6 Ostrowski-Meissner [11],,, AFB 1 Boonyaratpalin [16] (Penaeus monodon) AFB 1 100~2 500 μg/kg 8,,, AFB 1,, AFB 1, AFB 1,, 4,,, μg/kg, AFB 1 AFB 1, 6,,,,, AFB 1, AFB 1,,, AFB 1 : [1] IARC. Aflatoxins, some naturally occurring substances: food items and constituents, heterocyclic aromatic amines and mycotoxins[r]. Lyon: IARC Monograph on the Evaluation of Carcinogenic Risks to Humans, 1994, 56: [2] Bailey G S, Williams D E, Wilcox J S, et al. Aflatoxin B1 carcinogenesis and its relation to DNA adduct formation and adduct persistence in sensitive and resistant salmonid fish[j].carcinogenesis, 1988, 9: [3] Hussain M, Gabal M A, Wilson T, et al. Effect of aflatoxin-contaminated feed on morbidity and residues in walleye fish [J]. Veterinary and Human Toxicology, 1993, 35: [4] El-Banna R, Teleb H M, Hadi M M, et al. Performance and tissue residue of tilapia fed dietary aflatoxin[j]. Veterinary Medical Journal Giza, 1992, 40: [5] Cagauan A G, Tayaban R H, Somga J R, et al. Effect of aflatoxin-contaminated feeds in Nile tilapia (Oreochromis niloticus)[m]//remedios R B, Mair G C. Proceedings of the Sixth International Symposium on Tilapia in Aquaculture, 2004: [6] Tuan N A, Grizzle J M, Lovell R T, et al. Growth and hepatic lesions of Nile tilapia (Oreochromis niloticus) fed diets containing aflatoxin B1[J]. Aquaculture, 2002, 212: [7] Deng S X, Tian L X, Liu F J, et al. Toxic effects and residue of aflatoxin B1 in tilapia (Oreochromis niloticus O. aureus) during long-term dietary exposure[j]. Aquaculture, 2010, 307: [8] Horwitz W, Latimer G W. Official Methods of Analysis of AOAC international [M]// 14th edn. Gaithersburg: AOAC International, [9] Folch J, Lees M, Sloane-Stanley G H. A simple method of the isolation and purification of total lipid from animal tissues [J]. Journal of Biological Chemistry, 1957, 226(1): [10],,. H.E [J]., 1997, 20(5): 508.

6 6, : B [11] Ostrowski-Meissner H, LeaMaster B, Duerr E, et al. Sensitivity of the Pacific white shrimp, Penaeus vannamei, to aflatoxin B1 [J].Aquaculture, 1995, 131: [12] Burgos-Hernandez A, Farias S I, Torres-Arreola W, et al. In vitro studies of the effects of aflatoxin B1 and fumonisin B1 on trypsinlike and collagenase-like activity from the hepatopancreas of white shrimp (Litopanaeus vannamei)[j]. Aquaculture, 2005, 250: [13] Munoz M, Rodnguez J, Vandei K, et al. Measurement of reactive oxygen intermediate production in haemocyres of the penaeid shrimp, Penaeus vannamei [J]. Aquaculture, 2000, 191: [14] Bintvihok A, Ponpornpisit A, Tangtrongpiros J, et al. Aflatoxin contamination in shrimp feed and effects of aflatoxin addition to feed on shrimp production[j]. Journal of Food Protection, 2003, 66(5): [15] Gallagher E P, Eaton D. In vitro biotransformation of aflatoxin B1 (AFB1) in channel catfish liver[j]. Toxicology and Applied Pharmacology, 1995, 132: [16] Boonyaratpalin M, Supamattaya K, et al. Effects of aflatoxin B1 on growth performance, blood components, immune function and histopathological changes in black tiger shrimp (Penaeus monodon)[j]. Aquaculture, 2001, 32: Toxic effects of aflatoxin B 1 on growth performance, biochemical and hepatopancreas microstructure of Litopanaeus vannamei WANG Jing, GUO Ran *, SU Li, XIA Hui, CUI Min (Ocean College, Agricultural University of Hebei, Qinhuangdao , China) Abstract: The toxic effects of aflatoxin B 1 (AFB 1 ) on Litopanaeus vannamei were examined in terms of growth indices, body composition, hepatopancreas microstructure and biochemical indexes of haemolymph and hepatopancreas. Six isonitrogenous purified diets were fed to shrimp [initial mean weight (0.3±0.02) g] for 8 weeks in triplicate tanks connected to recirculating system. Diets contained different levels of AFB 1 (0, 400, 800, 1 200, 1 600, μg/kg). Crude protein of whole shrimp had no significant differences (P > 0.05). Body lipid tended to be higher within shrimp fed diets with higher AFB 1 level (1 600 and μg/kg). The lowest weight gain (304.67%) was found in shrimp fed with μg/kg diet. However the shrimp fed with μg AFB 1 /kg got the lowest survival rate (58.33%) and the highest weight gain. Enzyme activities generally indicated a downward trend. Total antioxidant capacity of hepatopancreas decreased significantly (P<0.05) in μg/kg diet. The activity of GST, Na +, K + -ATPase and SOD in hepatopancreas reached minimum in shrimp fed with μg/kg AFB 1 diet, but GST of haemolymph was significantly higher than others (P<0.05). Histological study on shrimp fed more AFB 1 exhibited histological changes. Besides, higher AFB 1 residues in hepatopancreas were observed in shrimp fed with higher level AFB 1. The most AFB 1 residues in hepatopancreas were detected at μg/kg in shrimp fed with μg/kg. However, no AFB 1 residue in muscle was detected in all groups. Key words: Litopanaeus vannamei; aflatoxin B 1 ; biochemical; hepatopancreas microstructure Corresponding author: GUO Ran. toguoran@163.com

中草药 Chinese Traditional and Herbal Drugs 第 46 卷第 9 期 2015 年 5 月 1339 非酒精性脂肪性肝病 (nonalcoholic fatty liver disease,nafld) 是一种与胰岛素抵抗遗

中草药 Chinese Traditional and Herbal Drugs 第 46 卷第 9 期 2015 年 5 月 1339 非酒精性脂肪性肝病 (nonalcoholic fatty liver disease,nafld) 是一种与胰岛素抵抗遗 1338 中草药 Chinese Traditional and Herbal Drugs 第 46 卷第 9 期 2015 年 5 月九味肝泰胶囊对高脂饮食诱导大鼠非酒精性脂肪肝的治疗作用陈菲 1 2,, 艾国 3, 盛柳青 1, 麻佳蕾 1, 黄正明 1. 金华职业技术学院医学院, 浙江金华