动态显色法鲎试验用于果汁细菌内毒素活性检测及干扰分析
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摘要
目的建立一种定量检测果汁细菌内毒素活性的方法,准确评估果汁潜在的内毒素污染风险。方法采用动态显色法东方鲎(Tachypleus amebocyte lysate, TAL)试验对常见的4种果汁(橙汁、桃汁、苹果汁和葡萄汁)细菌内毒素活性进行检测,对果汁样品中的干扰因素进行识别与排除,最后使用抗增液排除β-葡聚糖等类似物质带来的假阳性结果。结果市售果汁(n=36)的细菌内毒素活性范围为<0.2～209.5 EU/mL,自榨果汁(n=12)的细菌内毒素活性范围为<0.4～4.2EU/mL,加标回收率范围均在50%～200%之间,符合药典要求。结论动态显色法TAL试验适用于常见的4种果汁(橙汁、桃汁、苹果汁和葡萄汁)细菌内毒素活性的检测,为果汁的健康风险评价和卫生监管提供参考。
Objective To establish a quantitative detection method for endotoxin activities in juice, in order to provide technical support for assessment of potential endotoxin contamination risk in juice. Methods The endotoxin activities of juice were detected by Tachypleus amebocyte lysate(TAL) assay of kinetic chromogenic analysis, the interference factors were identified and eliminated, and the juice samples with positive results of TAL assay were verified by endotoxin-specific buffer in order to avoid the false positive caused by β-glucan. Results The endotoxin activities of 4 kinds of juice products(n=36) and self-squeezed juice(n=12) were <0.2-209.5 EU/mL and <0.4-4.2 EU/mL, respectively. The recovery rates met requirements of Chinese Pharmacopoeia(50%-200%).Conclusion The TAL assay of kinetic chromogenic analysis is suitable for the quantitative determination of endotoxin activities in 4 kinds of common fruit juice(orange juice, peach juice, apple juice and grape juice), which provides reference for health risk assessment and health supervision of fruit juice.
Objective To establish a quantitative detection method for endotoxin activities in juice, in order to provide technical support for assessment of potential endotoxin contamination risk in juice. Methods The endotoxin activities of juice were detected by Tachypleus amebocyte lysate(TAL) assay of kinetic chromogenic analysis, the interference factors were identified and eliminated, and the juice samples with positive results of TAL assay were verified by endotoxin-specific buffer in order to avoid the false positive caused by β-glucan. Results The endotoxin activities of 4 kinds of juice products(n=36) and self-squeezed juice(n=12) were <0.2-209.5 EU/mL and <0.4-4.2 EU/mL, respectively. The recovery rates met requirements of Chinese Pharmacopoeia(50%-200%).Conclusion The TAL assay of kinetic chromogenic analysis is suitable for the quantitative determination of endotoxin activities in 4 kinds of common fruit juice(orange juice, peach juice, apple juice and grape juice), which provides reference for health risk assessment and health supervision of fruit juice.
引文
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[4]张灿,刘文君,张明露,等.生活饮用水和瓶装饮用水的细菌内毒素活性调査[J].中国给水排水,2013,29(13):47-52.Zhang C, Liu WJ, Zhang ML, et al. Investigation on bacterial endotoxinactivities in municipal drinking water and bottled drinking water[J]. China Water Wastewater, 2013, 29(13):47-52.
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[7] Guizani M, Yusuke N, Dhahbi M, et al. Characterization of endotoxic indicative organic matter(2-keto-3deoxyoctulosonic acid)in raw and biologically treated domestic wastewater[J]. Water Res, 2011, 45(1):155-162.
[8] Man HD, Heederik DDJ, Leenen EJTM, et al. Human exposure to endotoxins and fecal indicators originating from water features[J]. Water Res, 2014,51:198-205.
[9] O'Toole J, Sinclair M, Jeavons T, et al. Alternative water sources and endotoxin[J]. Water Sci Technol, 2008, 58(3):603-607.
[10]张灿,刘文君,张明露,等.水体环境内毒素活性的鲎法定量检测及影响因素研究[J].环境科学, 2013, 34(9):3381-3385.Zhang C, Liu WJ, Zhang ML, et al. Detection of endotoxin activity in water environment and analysis of influence factors for TAL assay[J].Environ Sci, 2013, 34(9):3381-3385.
[11] Sipka S, Béres A, Bertók L, et al. Comparison of endotoxin levels in cow's milk samples derived from farms and shops[J]. Innate Immun, 2015,21(5):531-536.
[12] Health Council of the Netherlands, 2010. Endotoxins:Health-based recommended occupational exposure limit. No. 2010/040SH[Z].
[13]中国国家药典委员会.中国人民共和国药典[M].北京:中国医药科技出版社,2015.Chinese Pharmacopoeia Commission. Chinese pharmacopoeia of the People's Republic of China[M]. Beijing:Chemical Industry Press, 2015.
[14] GB 4789.2-2016食品安全国家标准食品微生物学检验菌落总数测定[S].GB 4789.2-2016 National food safety standard-Food microbiological examination-Detection of aerobicplate count[S].
[15] GB 4789.3-2016食品安全国家标准食品微生物学检验大肠菌群计数[S].GB 4789.3-2016 National food safety standard-Food microbiological examination-Enumeration of coliforms[S].
[2] Elin RJ, Wolff SM, Mcadam KP, et al. Properties of reference Escherichia coli endotoxin and its phthalylated derivative in humans[J]. J Infect Dis,1981,144(4):329-336.
[3] Levin J, Bang FB. Clottable protein in Limulus; its localization and kinetics of its coagulation by endotoxin[J]. Thromb Haemost, 1968, 19(1):186-197.
[4]张灿,刘文君,张明露,等.生活饮用水和瓶装饮用水的细菌内毒素活性调査[J].中国给水排水,2013,29(13):47-52.Zhang C, Liu WJ, Zhang ML, et al. Investigation on bacterial endotoxinactivities in municipal drinking water and bottled drinking water[J]. China Water Wastewater, 2013, 29(13):47-52.
[5] Dai S, Masaaki H, Hisashi H, et al. Occurrence and fate of endotoxin activity at drinking water purification plants and healthcare facilities in Japan[J]. Water Res, 2018, 145:1-11.
[6] Zhang C, Liu WJ, Sun W, et al. Endotoxin contamination and control in surface water sources and a drinking water treatment plant in Beijing,China[J]. Water Res, 2013, 47(11):3591-3599.
[7] Guizani M, Yusuke N, Dhahbi M, et al. Characterization of endotoxic indicative organic matter(2-keto-3deoxyoctulosonic acid)in raw and biologically treated domestic wastewater[J]. Water Res, 2011, 45(1):155-162.
[8] Man HD, Heederik DDJ, Leenen EJTM, et al. Human exposure to endotoxins and fecal indicators originating from water features[J]. Water Res, 2014,51:198-205.
[9] O'Toole J, Sinclair M, Jeavons T, et al. Alternative water sources and endotoxin[J]. Water Sci Technol, 2008, 58(3):603-607.
[10]张灿,刘文君,张明露,等.水体环境内毒素活性的鲎法定量检测及影响因素研究[J].环境科学, 2013, 34(9):3381-3385.Zhang C, Liu WJ, Zhang ML, et al. Detection of endotoxin activity in water environment and analysis of influence factors for TAL assay[J].Environ Sci, 2013, 34(9):3381-3385.
[11] Sipka S, Béres A, Bertók L, et al. Comparison of endotoxin levels in cow's milk samples derived from farms and shops[J]. Innate Immun, 2015,21(5):531-536.
[12] Health Council of the Netherlands, 2010. Endotoxins:Health-based recommended occupational exposure limit. No. 2010/040SH[Z].
[13]中国国家药典委员会.中国人民共和国药典[M].北京:中国医药科技出版社,2015.Chinese Pharmacopoeia Commission. Chinese pharmacopoeia of the People's Republic of China[M]. Beijing:Chemical Industry Press, 2015.
[14] GB 4789.2-2016食品安全国家标准食品微生物学检验菌落总数测定[S].GB 4789.2-2016 National food safety standard-Food microbiological examination-Detection of aerobicplate count[S].
[15] GB 4789.3-2016食品安全国家标准食品微生物学检验大肠菌群计数[S].GB 4789.3-2016 National food safety standard-Food microbiological examination-Enumeration of coliforms[S].