CN101596381A - 用于去除过程液体中生物污染物的吸收性过滤介质 - Google Patents

用于去除过程液体中生物污染物的吸收性过滤介质 Download PDF

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CN101596381A
CN101596381A CNA2009101417603A CN200910141760A CN101596381A CN 101596381 A CN101596381 A CN 101596381A CN A2009101417603 A CNA2009101417603 A CN A2009101417603A CN 200910141760 A CN200910141760 A CN 200910141760A CN 101596381 A CN101596381 A CN 101596381A
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D·P·亚沃尔斯基
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Abstract

本发明涉及用于去除过程液体中生物污染物的吸收性过滤介质。吸附性过滤介质，尤其适于去除过程液体中生物污染物。仅采用颗粒吸附剂和水不溶性热塑粘合剂形成吸附材料的多孔固定床。所得的复合过滤器和常规深度过滤器相比，可以允许具有吸附颗粒更小的、更大量的吸附剂。消除纤维素纤维以及消除热固性粘合剂，使得过程液体的污染物减少。

Description

用于去除过程液体中生物污染物的吸收性过滤介质
本申请是申请日为2007年2月17日提交的200710078971.8，发明名称为“用于去除过程液体中生物污染物的吸收性过滤介质”的专利申请的分案申请。
本申请要求2006年2月17日提交的临时申请序列No.60/774773的优先权，其公开内容在此通过引用结合进来。
技术领域
本发明涉及用于去除过程液体中生物污染物的吸收性过滤介质。
背景技术
纤维素深度过滤器(depth filter)，比如来自Millipore Corporation的
Figure A20091014176000031
+可商购过滤器，通常在生物药物的制备中用于澄清各种粗产物流体，所述生物药物比如源于哺乳动物细胞培养物。这些复合过滤器包括结构紧密的纤维素深度介质层，可以针对具体应用而进行最优化，比如保留胶体颗粒和细胞碎片或者保留全细胞和较大的碎片。它们在单一滤筒中结合了连续等级的介质。这些过滤器最常用于抛光或者二次澄清过程中，以从含水产物(蛋白质)流中去除少量悬浮物质。这些过滤器的主要功能是保护或者延长更昂贵的下游分离过程，比如无菌过滤和亲和色谱仪的使用寿命。也就是说，这些过滤器的共同应用是作为“预过滤器”，即保护下游装置和介质免受胶态污染物和其它细胞碎片的影响。另外，所述深度过滤器也用于通过去除痕量的黏结蛋白质(agglomerated protein)来保护病毒清除型过滤器。
在本行业中同样公知的是，复合深度过滤器也能够以不同程度保留在哺乳动物细胞培养物中常见的一些可溶性污染物，比如核酸、宿主细胞蛋白质、脂质、表面活性剂等。这种对于特定可溶性污染物的保留能力是基于深度过滤介质的吸附性能。
在这些深度过滤器中通常采用的过滤介质包括精制纤维素纤维(源于木浆和/或棉花)、硅藻土和水溶性热固树脂粘合剂。这些复合体中的硅藻土(二氧化硅的天然形式，含有痕量的各种硅酸盐)通常为40-60重量％，被认为是必要组分，吸收胶体大小的生物物质，比如细胞断片、细胞器和黏结蛋白质，以及各种可溶性的生物化学物质，比如蛋白质、脂质和核酸。
但是，采用这些纤维素深度过滤器制备肠道外药和其它药物的主要缺点之一是它们释放到体系中的水溶性污染物的含量相对较高。实际上，在使用前为了将这些有机和无机污染物含量降低到可接受的水平，需要大量的预冲洗。而且，硅藻土吸附剂在深度过滤介质中的最大加载量被限制为大约60重量％，而且所述吸附剂保留在纤维基后中所需的最小颗粒尺寸为大约10微米。
所以，本发明的目标是减少或者消除从吸附性过滤器中释放污染物。
本发明的另一目标是提高吸附剂在过滤介质中的含量或加载量。
本发明的又一目标是提供具有更小吸附剂颗粒的过滤器，以使用于吸附的可用表面积最大化。
通过下面的详述和附图，本发明的其它目标和优点将变得显而易见。
发明内容
通过本发明解决了现有技术的问题，本发明提供了特别适于去除过程液体中生物污染物的吸附性过滤介质。采用仅仅颗粒吸附剂和水不溶性热塑粘合剂形成了吸附性材料的多孔固定床。所得到的复合过滤器和常规深度过滤器相比，可以具有更高量的、吸附性颗粒更小的吸附剂。消除纤维素纤维以及消除水溶性热固粘合剂，使得过程液体的污染程度下降。结果，不再需要大量的预冲洗来减少外部污染物。通过提高吸附性材料的含量和/或通过采用更小的吸附性介质来最大化可用于吸附的表面积，实现了介质性能的改进。
所得到的复合过滤材料基本上或者完全没有纤维素和热固性粘合剂，可以置于生物反应器下游和无菌过滤器上游的澄清系统中。其它应用包括在病毒清除型过滤和色谱分离之前对细胞培养物流体进行预处理。
附图说明
图1是比较本发明复合体和常规复合体的透水性的图；
图2是预过滤压差和通过量的关系图；
图3是无菌过滤压差和通过量的关系图；和
图4是通过各种介质的流出物的传导率图表。
具体实施方式
本发明的过滤介质包括吸附性材料和水不溶性热塑粘合剂。这种组合可用于形成吸附性材料的多孔固定床，具有对所述应用而言合适的机械性质，包括渗透性、拉伸强度和弯曲强度。过滤介质特别适于用于深度过滤器。
合适的吸附性材料包括硅藻土、二氧化硅、多孔玻璃、沸石和活性碳。特别优选硅藻土。另外，任何形式(珠子、研磨的粉末、等)的、任何表面化学性质的(离子交换、疏水性等)的色谱分析介质可用作所述多孔介质中的吸附剂。合适的粘合剂包括热塑粘合剂，比如聚烯烃，优选聚乙烯、聚丙烯或者其混合物。粘合剂优选以珠子、粉末或者纤维形式使用。通过正确选择粘合剂(从具有足够高的熔点或软化点方面考虑)，介质可以经过热压处理或者以其它方式蒸汽消毒或者经过γ照射来减少或者消除其中的任何生物污染物。
介质制备方法可能取决于所用的粘合剂形式。可以通过将粘合剂和吸收性材料掺混，然后通过比如部分熔融或者软化粘合剂将吸附性颗粒熔合在一起，来制备介质。例如，聚乙烯粉可以和吸附性颗粒，比如硅藻土或者二氧化硅珠子，以大约1∶1-大约1∶3的粘合剂∶吸附性颗粒的重量比干混(比如通过摇晃/翻转数分钟)。所得的掺合材料可以放置于模具中，加热(例如，在加热的水压机中)到合适的温度以熔融所述吸附性颗粒，比如130℃-160℃。通过本方法已经采用MitsuiChemical市售的超高分子量聚乙烯粉(MipelonTM)将硅藻土和二氧化硅珠子结合到2-4mm厚的垫子中，其中平均珠子直径为20-30微米。随着材料在水压机中加热和软化，压力应该进行周期性调节，以在加热循环中(大约5-10分钟)维持恒定的力。
或者，可以采用湿铺法(wet-laid)制备介质，尤其在粘合剂是纤维形式时。例如，可以将细的聚乙烯纤维(FybrelTM，来自MitsuiChemical)分散在异丙醇和水中，然后和硅藻土粉以大约1∶1-大约1∶3的粘合剂∶吸附剂的重量比例掺混成浆料。将浆料转移到布氏漏斗中，所述漏斗在基部容纳有标称为1微米的非织造支持材料，以防纤维和吸附剂直接通过漏斗的开孔。然后，通过真空瓶将液体主体吸走。所形成的饼被转移到烘箱中进行干燥，饼由于部分熔融或者软化聚乙烯纤维将吸附性颗粒热结合在一起。
如上构建的复合材料和常规纤维素深度过滤器相比，具有非常高的渗透性(高孔隙率)和低的颗粒保留性质。硅藻土颗粒和聚乙烯纤维或粉的重力沉降获得了具有较大孔隙空间的低密度复合结构。
为了提高本发明硅藻土复合材料的分离性质，所述材料在加热之前和/或之中应该致密化或压实。例如，发现采用气动压力机在100-325psi压缩过滤介质试样大约30秒是有效的。介质试样在压缩后基本不会松弛；在经过加热以将所述结构熔合成单块后，介质保持了其压缩后的厚度尺寸。复合物可以被压缩到其原始体积的大约50-70％，具体取决于组成和压缩力。
也可以在复合材料上施加机械压缩来调节透水性，所述透水性可以接近标准纤维素深度过滤垫的透水性。图1举例说明了和常规
Figure A20091014176000061
+AlHC深度过滤器试样相比，由CelpureTM(AdvancedMinerals)硅藻土和FybrelTM(Mitsui Chemical)聚乙烯微纤维制成的复合物的透水性测量结果(流速和压差的关系)。该图表明本发明复合物的透水性范围和市售纤维素深度过滤介质的相一致。
可以在吸附剂/粘合剂混合物中加入第三种颗粒状或者纤维状组分作为操控制品渗透性的手段，从而可以更有效地使用所含的吸附剂。所述添加物可以是功能吸附性或者惰性材料，但是其尺寸和量必须对介质渗透性产生可测量的影响。测试结果表明以这种方式影响介质渗透性并不会对介质的吸附能力有明显的损坏(下降)。在介质中有选择地形成更大的流道使得过程流体可以在多孔基质中渗透得更深/更宽，从而可以有效补偿介质的整体较低的吸附容量。
除了捕捉颗粒以外，本发明的吸附性多孔单块和常规纤维素吸附性深度过滤器相比具有明显的优点，尤其在水可提取物方面。在肠道外药物的制备中，任何可提取的材料都是很严重的问题。已知常规纤维素深度介质具有较高的可提取量，所述可提取量要求在使用之前进行大量冲洗。本发明人已经证实对传导率(无机物)有贡献的可提取物并不仅仅或者主要来自硅藻土。实际上，本发明的复合材料不含纤维素和热固化粘合剂，和常规纤维素介质相比，其使得流出物传导率下降75-90％。消除纤维素以及用水不溶性粘合剂比如聚乙烯代替热固性水溶性树脂粘合剂，获得了无机可提取物的量显著下降的DE基介质。对于所述材料的使用者而言，在降低冲洗要求但同时降低制品污染风险方面，具有相当的益处。
一般而言，制备本发明的优选复合过滤材料的一种优选方法如下。在包括内部高速搅拌器的旋转V型掺混机中，分批干混平均颗粒尺寸为25微米的超高分子量聚乙烯粉和颗粒尺寸为0.2-25微米的天然硅藻土。混合物从掺混机转移到粉体分配器(或者施加器)中。施加器将混合的粉体于室温分配到移动的多孔非织造聚酯材料的卷幅(web)上，可控厚度为不超过0.5英寸。可以以相似方式多次施加不同的粉体混合物，从而形成具有梯度组成的吸附性介质。
然后，通过和顶置滚筒接触将松散的混合粉体层轻微致密化和平整，然后通过电热板从下部并同时通过IR灯从上部加热，使聚乙烯粉体软化。受热的板的温度沿着制备途径逐渐升高到大约340的最终温度。粉体混合物温度保持在最大约340数分钟，然后在顶部施加另外的非织造聚醚卷幅。
然后，将复合材料通过温度同样设置在大约340的两个加热的压延机滚筒，以大约100psi连续压缩至0.10-0.20英寸的厚度。然后，使最终材料在和空气连通的金属板上冷却。
实施例1
在和从哺乳动物细胞培养物回收蛋白质制品相关的标准澄清化工艺步骤中，测试熔合成厚度大约2-4mm的垫的硅藻土和聚乙烯微纤维的各种掺混物试样。所述测试是采用E.coli溶胞产物悬浮液(在缓冲液)在恒流条件下对吸附性介质进行测试，同时监控介质试样上的升压(堵塞速率)和流出物品质与已处理的流体体积的关系。通过使流体直接通过0.2微米灭菌级膜式过滤器(在本例中是
Figure A20091014176000071
GV)过滤，测量流出物或者滤出液的品质。这些试验复合物试样被再次和
Figure A20091014176000072
+纤维素深度过滤介质进行了比较。
图2和3给出了吸附性介质试样和
Figure A20091014176000073
+垫的压力分布以及相关的灭菌过滤器压力分布。通过量定义为处理的流体体积和采用的深度过滤介质体积(床体积)的关系。
如图所示，DE/PE纤维复合体在通过量和保留方面都能够和最密级或者保留能力最强级别的
Figure A20091014176000081
+DE介质(75级)相配。各种复合体试样的升压率都等于或者低于75DE
Figure A20091014176000082
+介质的升压率。另外，对于本发明的复合体而言，和75DE+介质相比，下游灭菌过滤器的升压率都几乎相等，这表明颗粒保留水平具有可比性。
实施例2
对采用聚乙烯粉(MipelonTM，Mitsui Chemical)将硅藻土熔合到固定床垫中的试样进行测试，测试其保护病毒保留型膜，NFP Viresolve180的能力。在本测试中，用浓度为0.5gm/L的多克隆人体IgG蛋白质溶液测试DE/PE复合体(大约3mm厚)。对于这种原料而言，
Figure A20091014176000084
膜通常的能力是大约150L/m2。采用
Figure A20091014176000085
+AlHC深度过滤器对这种原料进行预处理以去除蛋白质黏结体，可以使Viresolve膜的能力提高到750-1500L/m2。将硅藻土试样(Celpure 25和Celpure 300(Advanced Minerals))与Mipelon PE粉掺混，在加热后形成2-3mm的垫(无需压制)。然后，将DE/PE复合试样用于对IgG原料进行预处理，滤出液再次通过Viresolve 180进行处理以确定对膜能力的影响。
Celpure 25(精细级DE)使得Viresolve能力为440L/m2，Celpure300(较大的粗颗粒DE)使得Viresolve能力＞1000L/m2。这些测试表明通过PE粉体粘合剂形成的硅藻土单块为病毒保留型膜提供的保护水平(通过去除蛋白质黏结物)，和通过纤维素深度过滤器比如
Figure A20091014176000086
+深度过滤器目前提供的相同。
实施例3
为了测量本发明复合材料的洁度，用干净的去离子水冲洗材料试样，在预定的冲洗量之后测量流出物的传导率。传导率值被用来表示在过滤介质中可溶金属的含量。图4给出了各种DE/PE复合试样的传导率值和市售
Figure A20091014176000087
+深度过滤器试验的关系。
Figure A20091014176000088
+DE介质是纤维素、硅藻土和水溶性热固性树脂粘合剂的复合体。CE介质仅仅含有纤维素纤维和粘合剂。从这些测量中可以证明，对传导率(无机物)有贡献的这些可提取物并不主要来自硅藻土。将这些值和所测试的DE/PE复合体相比，流出物传导率下降75-90％。
实施例4
当在粉体聚乙烯(20-30微米)和硅藻土(0.5-10微米)的2∶1混合物中加入10-20％的75-100微米多孔玻璃珠子时，可以使最终介质的透水性下降10-30％，但通过颗粒捕捉以及过程体积测量发现吸附能力没有可测量的损失。

Claims (8)

1.一种复合过滤材料，其包括吸附性材料和将所述吸附性材料粘合在一起的水不溶性粘合剂，所述材料基本不含纤维素。
2.权利要求1的复合过滤材料，其中所述吸附性材料选自硅藻土、二氧化硅、玻璃、沸石和活性炭。
3.权利要求1的复合材料，其中所述吸附性材料是硅藻土。
4.权利要求1的复合材料，其中所述粘合剂包括聚烯烃。
5.权利要求1的复合材料，其中所述粘合剂包括聚乙烯和聚丙烯。
6.权利要求1的复合材料，其中通过将所述吸附性材料和所述粘合剂加热到其中所述粘合剂至少部分熔融或者软化的温度以将所述材料热结合在一起，形成所述复合材料。
7.权利要求1的复合材料，其中所述粘合剂和吸附性材料的重量比是1∶1-1∶3。
8.权利要求1的复合材料，其基本不含树脂粘合剂。
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CN110194784A (zh) * 2019-05-07 2019-09-03 上海药明生物技术有限公司 运用硅藻土澄清细胞发酵液的方法

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US8672144B2 (en) 2014-03-18
ATE515308T1 (de) 2011-07-15
CN101053717A (zh) 2007-10-17
CN101596381B (zh) 2013-01-23
US8403153B2 (en) 2013-03-26
US20070193938A1 (en) 2007-08-23
JP2007216224A (ja) 2007-08-30
US8562875B2 (en) 2013-10-22
CN101053717B (zh) 2011-09-28
EP1820555B1 (en) 2011-07-06
JP5117067B2 (ja) 2013-01-09
US20130062280A1 (en) 2013-03-14
US20100159581A1 (en) 2010-06-24
US20120175805A1 (en) 2012-07-12
ES2367850T3 (es) 2011-11-10
SG135097A1 (en) 2007-09-28
US7673757B2 (en) 2010-03-09

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