城镇污泥中神经酰胺的分离纯化
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摘要
以A~2O-MBR工艺的冷冻干燥污泥作为原料,以索氏提取法,获得污泥中的粗脂;再经硅胶柱层析,以石油醚-乙酸乙酯100:1,10:1和7:3(V/V)的顺序梯度洗脱,利用薄层色谱法,将洗脱液与神经酰胺标准品对比,V(石油醚):V(乙酸乙酯)=7:3为合适的洗脱梯度.经Q-Exactive液相色谱-高分辨质谱检测各组成成分,就峰面积比,经柱层析洗脱前后神经酰胺从60.88%提高到98.34%.最后,用另一种A~2O工艺的污泥进行尝试,柱层析前后神经酰胺从79.06%提高至94.15%.
Using the freeze-dried sludge of A~2O-MBR process as raw material, we firstly obtained the crude lipid in the sludge by Soxhlet extraction. Then the crude lipid was eluted in a sequence of petroleum ether-ethyl acetate 100:1, 10:1 and 7:3(V/V) by silica gel column chromatography. Compared the eluent with ceramide standard by thin layer chromatography, it was proved that the appropriate elution gradient was petroleum ether-ethyl acetate 7:3(V/V). Through Q Exactive bench quadrupole orbitrap high-resolution mass spectrometry, the peak area ratio of ceramide was increased from 60.88% to 98.349% in the process of column chromatography. When the sludge of A~2 O process was used as raw material in the same experiment, the peak area ratio of ceramide was increased from 79.06% to 94.15%.
Using the freeze-dried sludge of A~2O-MBR process as raw material, we firstly obtained the crude lipid in the sludge by Soxhlet extraction. Then the crude lipid was eluted in a sequence of petroleum ether-ethyl acetate 100:1, 10:1 and 7:3(V/V) by silica gel column chromatography. Compared the eluent with ceramide standard by thin layer chromatography, it was proved that the appropriate elution gradient was petroleum ether-ethyl acetate 7:3(V/V). Through Q Exactive bench quadrupole orbitrap high-resolution mass spectrometry, the peak area ratio of ceramide was increased from 60.88% to 98.349% in the process of column chromatography. When the sludge of A~2 O process was used as raw material in the same experiment, the peak area ratio of ceramide was increased from 79.06% to 94.15%.
引文
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[21]李海池,马晋芳,史庆龙,等.超临界CO2萃取-分子蒸馏提取分离磨芋神经酰胺[J].中药材,2017,40(10):2401-2405.Li H C, Ma J F, Shi Q L, et al. Supercritical CO2 extraction-molecular distillation extraction for ceramide of koniac[J]. Journal of Chinese Medicinal Materials, 2017, 40(10):2401-2405.
[22] Qi J J, Zhu F F, Wei X, et al. Comparison of biodiesel production from sewage sludge obtained from the A(2)/O and MBR processes by in situ transesterification[J]. Waste Management, 2016,49(4):212-220.
[2]崔建鑫,赵海霞.城镇污水处理设施空间优化配置研究[J].中国环境科学,2016,36(3):943-952.Cui J X, Zhao H X. Method for spatial optimal allocation of urban sewage treatment facilities[J]. China Environmental Science, 2016,36(3):943-952.
[3] Dufreche S, Hernandez R., French T, et al. Extraction of lipids from municipal wastewater plan microorganisms for production of biodiesel[J]. Journal of the American Oil Chemists Society, 2007,84(2):181-187.
[4] Eurostat. Sewage sludge production and disposal from urban wastewater(in dry substance(d.s))[EB/OL]. https://ec.europ-a.eu/eurostat/web/products-datasets/-/ten00030.
[5] Zhu F F, Takaoka M, Wang H C, et al. Sewage sludge disposal and recycling trends in japan[J]. China Water&Wastewater, 2012,28(11):102-104.
[6]戴晓虎.我国城镇污泥处理处置现状及思考[J].给水排水,2012,48(2):1-5.Dai X H. The current state and consideration on urban sewage sludge disposal in China[J]. Water&Wastewater Engineering, 2012,38(2):1-5.
[7] Zhu F F, Wu X M, Zhao L Y, et al. Lipid Profiling in Sewage Sludge[J]. Water Research, 2017,116:149-158.
[8] Urubauer U, Feingold K R., Elias P M. Relationship of epidermal lipogenesis to cutaneous barrier function[J]. Journal of Lipid Research,1987,28(6):746-752.
[9] Hannun Y A, Bell R M. Functions of sphingolipids and sphingolipid breakdown products in cellular regulation[J]. Science, 1989,234(4890):500-507.
[10] Merrill J A, Schmelz E M, Wang E, et al. Role of dietary sphingolipids and inhibitors of sphingolipid metabolism in cancer and other diseases[J]. Journal of Nutrition,1995,125(6Suppl):1 677S-1682S.
[11] Zhang L,Hellgren L I, Xu X. Kinetic study of sphingomyelin hydrolysis for ceramide production[J]. Journal of Molecular Catalysis B Enzymatic,2008,51(3):93-99.
[12] Kwun K H., Lee J H, Rho K H, et al. Production of ceramide with saccharomyces cerevisiae[J]. Applied biochemistry and biotechnology,2006,133(3):203-210.
[13]刘希望.神经酰胺类似化合物的合成及其生物活性研究[D].杨凌:西北农林科技大学,2010.Liu X W. Synthesis of ceramide analogs and study of their bioactivities[D]. Yangling:Northwest A&F University, 2010.
[14]刘静雯,李健,马亚瑞,等.一种从海洋球石藻中提取神经酰胺的方法:中国,CN108017554A[P]. 2018.Liu J W, Li J, Ma Y R, et al. A method for extracting ceramide from marine coccidian, China, CN108017554A[P]. 2018.
[15] Sahoo M R,Varma S R, Patel N K,et al. In vitro anti-inflammatory and skin protective properties of extract comprising ceramides from de-oiled rice bran[J]. Journal of Food Biochemistry, 2017,41.
[16] Cui S H, Kun M A, Zhao Y N, et al. Extraction and structure analysis of a species of ceramide from wheat powder[J]. Science&Technology of Food Industry, 2011,32(2):79-78.
[17]李玮滋,李献.一种从魔芋中高效提取神经酰胺的方法:中国,CN108623492A[P]. 2018.Li W Z, Li X. A highly efficient method for extracting ceramide from konjac:China, CN108623492A[P]. 2018.
[18]王兴林,侯秋红.魔芋飞粉中神经酰胺的提取分离方法:中国,CN103755583A[P]. 2014-04-30.Wang X L, Hou Q H. Extraction and separation of ceramide from konjac flying powder:China, CN103755583A[P]. 2014-04-30.
[19] Coderch L,Fonollosa J,Marti M,et al. Extraction and analysis of ceramides from internal wool lipids[J]. Journal of the American Oil Chemists Society, 2002,79(12):1215-1220.
[20]周秀秀,丁亚平,徐乐乐,等.花魔芋中三种功能性成分联产分离工艺[J].天然产物研究与开发,2017,29(7):1224-1229.Zhou X X, Ding Y P, Xu L L, et al. Joint producing technology of three function ingredients from konjac[J]. Natural Product Research and Development. 2017,29(7):1224-1229.
[21]李海池,马晋芳,史庆龙,等.超临界CO2萃取-分子蒸馏提取分离磨芋神经酰胺[J].中药材,2017,40(10):2401-2405.Li H C, Ma J F, Shi Q L, et al. Supercritical CO2 extraction-molecular distillation extraction for ceramide of koniac[J]. Journal of Chinese Medicinal Materials, 2017, 40(10):2401-2405.
[22] Qi J J, Zhu F F, Wei X, et al. Comparison of biodiesel production from sewage sludge obtained from the A(2)/O and MBR processes by in situ transesterification[J]. Waste Management, 2016,49(4):212-220.