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锅碗瓢盆儿

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虫号: 1251336
注册: 2011-03-31
性别: GG
专业: 果树学

[求助] 离子色谱，新手，准备摸索植酸的测定，不懂，求解，谢谢各位

0.5  g  each  of  the legume  samples  powder  were  extracted  under  mechanical  agitation  with  20 ml  0.5  M HCl  for  2  h  at  room  temperature  (Graf  & Dintzis,  1982).  The  extract  was  centrifuged  (700  g,  10 min)  and  the  supematant  decanted,  frozen  overnight, centrifuged  and  filtered  through  MF-Millipore  (0.22 pm).  The  inositol  phosphates  were  separated  from  the filtrate  and  concentrated  by  the  ion-exchange  procedure  of  Sandberg  and  Ahderinne  (1986)  with  some modifications.  The  filtrate  was  evaporated  to  dryness under  vacuum  (35℃,  dissolved  in  15 ml,  25 mM HCI, and  passed  through  a strong  anion-exchange (SAX)  column  that  was  connected  to  a  vacuum  manifold  (Visiprep; Supelco,  Bellefonte,  PA,  USA). The  loaded  SAX  column  was  washed  successively  with  10  and  5 ml  of  25 mM  HCI.  The  resin-bound  inositol  polyphosphates (IP3,  IP4,  IP5  and  IP6)  were  then  eluted  with  ten  1 ml portions  of  2  M HCl.  The  eluent  was  evaporated  to dryness  and  diluted  with  0.5 ml  of  mobile  phase.
自己大致翻译了下，求助各位大神，该段话有很多不理解的，
取0.5g上述粉末用20ml的0.5M的盐酸室温下机械搅拌浸提2h (Graf  & Dintzis,  1982)，离心700g×10min后取上清，冷冻过夜，并过0.22μ滤膜(MF-Millipore)，肌醇磷酸盐通过滤膜过滤至滤液中，并经过Sandberg  and  Ahderinne  (1986)改进的离子交换过程进行浓缩。滤液在抽真空后35摄氏度干燥(真空干燥器 35℃)然后用15ml浓度为25mM盐酸将其溶解，上强阴离子交换柱(季铵键合)，该强阴离子交换柱与真空歧管装置上相连，依次用10m 25mM盐酸和5m l25mM盐酸洗脱，与树脂结合的肌醇磷酸(IP3，IP4，IP5和IP6)被ten 个1ml 部分2M的盐酸洗脱
有几个地方不是很明白，特求助大牛们给点自己的看法
1、上面说0.5g的粉末在0.5M的盐酸中机械搅拌，用磁力搅拌是否可以？
2、离心取上清后冷冻过夜的目的是什么？
3、该强阴离子交换柱与真空歧管装置上相连，想问下实验室没有真空歧管装置怎么办？该装置的作用是什么？可否用其他代替？
4、后面说用25mM盐酸连续洗脱，我想问下为什么写成The  loaded  SAX  column  was  washed  successively  with 10  and  5 ml  of  25 mM  HCI，是先拿10ml洗再拿5ml洗吗？为什么要这么做，洗的时候是10ml全部加进去之后再将5ml全部加进去吗？
5、还有最最最不理解的后面的一句，The  resin-bound  inositol  polyphosphates (IP3,  IP4,  IP5  and  IP6)  were  then  eluted  with  ten  1 ml portions  of  2  M HCl，说是与树脂结合的肌醇多磷酸被 ten  1 ml portions  of 2M盐酸洗脱，这个是第10个1ml吗？是什么意思？是加10次吗？每次加1ml，然后收集加第10个1ml时候的洗脱液？那是收集1ml吗？真心不懂，希望大牛能帮帮忙~~谢谢！！！

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