Cytidine 5'-monophosphate(5'-CMP)is an essential nucleotide for additives.In this study,enhanced production of 5'-CMP was realized by the transformation of cytidine using co-immobilized di-enzymes,uridine-...Cytidine 5'-monophosphate(5'-CMP)is an essential nucleotide for additives.In this study,enhanced production of 5'-CMP was realized by the transformation of cytidine using co-immobilized di-enzymes,uridine-cytidine kinase(UCK)and acetate kinase(AcK).The immobilization yield of the enzyme had a clear correlation with the surface charges as zeta potential(ξ).Among them,ε-polylysinefunctionalized sepharose(SA-EPL,ξ=9.31 m V)showed high immobilization yield(78.8%),which was4.9-fold than that of nitrilotriacetic acid functionalized sepharose(SA-NTA,ξ=-12.6 m V).The residual activity of affinity co-immobilized enzyme(EPL-Ni/EPL@Ac K-UCK)was higher than 70.6%after recycled 10 times.Thus,this study provides an effective approach for the production of 5'-CMP with the advantages of low adenosine 5'-triphosphate(ATP)consumption,reduced side reactions,and improved reusability by co-immobilized UCK and Ac K on the functionalized Sepharose.展开更多
[Objective] This study aimed to establish an efficient process for separation of phycoerythrin by using Q Sepharose Fast Flow resin and verity its feasibility for scale-up. [Method] Elution gradient, sample volume and...[Objective] This study aimed to establish an efficient process for separation of phycoerythrin by using Q Sepharose Fast Flow resin and verity its feasibility for scale-up. [Method] Elution gradient, sample volume and flow rate were optimized to determine the optimal separation condition, under which the scale-up process was verified. [Result] The optimal condition for separation of phycoerythrin by using Q Sepharose FF resin was investigated： 30 ml of laver extract was loaded to the Q Sepharose FF column with a bed volume of 8 ml; subsequently, the column was stepwise eluted with 0-0.10-0.35-1.00 mol/L NaCI solution （pH 6.0） at a constant flow rate of 1 ml/min; the elution peak under 0.35 mol/L NaCI solution was collected, and the recovery rate and purity coefficient （A565/A280） of phycoerythrin were determined as 44.3 and 1.15, respectively. Based on the established process, 75 ml of phycoerythrin extract was loaded to the Q Sepharose FF column with a bed volume of 20 ml for separation, while no significant variation was observed in the separation result. [Conclusion] Phycoerythrin can be well separated from laver extract by using Q Sepharose FF resin and the process is feasible for scale-up.展开更多
Q Sepharose Fast Flow(Q Sepharose F.F.) was adopted to purify the suspensions of type Ⅰ,Ⅱ,Ⅲ oral poliovaccine (OPV), which is prepared from Vero cells. After clarification and concentration by ultrafiltration,th...Q Sepharose Fast Flow(Q Sepharose F.F.) was adopted to purify the suspensions of type Ⅰ,Ⅱ,Ⅲ oral poliovaccine (OPV), which is prepared from Vero cells. After clarification and concentration by ultrafiltration,the recovery of virus infected titre may attain above 85%.Using column chromatography on Q Sepharose F.F. to purify the concentrated virus suspensions,the purified viruses attain 100% recovery of virus infectivity. Dot membrane hybrization was used to detect DNA with the probe of Vero cell genome DNA which was labeled with α P 32 dATP,the contents of residual substrate DNA was less than 100 Pg/dose. The process of downstream had no significant influence on some biological characters of purified viruses,such as virus morphology ,tumorigenicity, rct/40 character and capsid protein.This downstream reseach indicates that Q Sepharose F.F. is an ideal chromatography material for purifying OPV prepared from Vero cells.展开更多
The separations of single-wall carbon nanotubes on length by sepharose gel were investigated in this work. The solutions of sodium dodecyl sulfate and sodium deoxycholate were applied as the eluent in sequence. SEM an...The separations of single-wall carbon nanotubes on length by sepharose gel were investigated in this work. The solutions of sodium dodecyl sulfate and sodium deoxycholate were applied as the eluent in sequence. SEM and Raman were used to characterize the length of nanotube bundles. The results show that the longer nanotubes were eluted out first, and then the shorter tubes were followed by the sodium dodecyl sulfate. However, the separated order was totally reversed by the sodium deoxycholate. By this method, the process generated nanotube fractions not only were narrower in length distributions, but also could control the separation orders by changing the eluents. Moreover, the separation principle was also discussed.展开更多
双特异性抗体(BsAbs),简称双抗,是一种能同时结合两个不同靶点或表位的抗体,BsAbs在表达过程中通常伴随产生多种副产物,在纯化工艺中很难被去除。本研究运用高通量筛选技术,通过对SP Sepharose Fast Flow填料的纯化条件进行探索,建立基...双特异性抗体(BsAbs),简称双抗,是一种能同时结合两个不同靶点或表位的抗体,BsAbs在表达过程中通常伴随产生多种副产物,在纯化工艺中很难被去除。本研究运用高通量筛选技术,通过对SP Sepharose Fast Flow填料的纯化条件进行探索,建立基于Sepharose Fast Flow填料的双特异性抗体纯化方法开发的通用流程,每个筛选条件只需要0.4 mg双抗样品,可同时筛选多达32个条件,筛选过程总耗时2 h,而传统柱层析需耗时64 h。通过高通量迅速筛选方法可以有效去除双抗分子副产物,为解决纯化工艺难题提供了一种新的工艺路线。展开更多
目的分离、纯化具有降血糖作用的仙人掌多糖组分。方法经热水提取、乙醇沉淀、DEAE Sepharose fast flow离子交换色谱和Sephadex G系列凝胶滤过色谱纯化得到5种仙人掌多糖组分。醋酸纤维薄膜电泳检测多糖纯度,凝胶色谱测定相对分子质量...目的分离、纯化具有降血糖作用的仙人掌多糖组分。方法经热水提取、乙醇沉淀、DEAE Sepharose fast flow离子交换色谱和Sephadex G系列凝胶滤过色谱纯化得到5种仙人掌多糖组分。醋酸纤维薄膜电泳检测多糖纯度,凝胶色谱测定相对分子质量,高效液相色谱测定糖的组成。结果5种多糖都已达到电泳纯。它们的相对分子质量依次为2.0×103、1.0×106、4.0×103、9.2×105、5.0×103。ODP1由鼠李糖组成,ODP2可能由鼠李糖和葡萄糖组成,ODP4可能由鼠李糖和D-半乳糖组成。ODP3和ODP5分别由鼠李糖和一未知糖组分组成。结论仙人掌多糖的提取没有必要采用脱蛋白、脱色素步骤,本实验所提取的仙人掌多糖是均一的。展开更多
基金supported by grants from the National Key Research and Development Program of China(2021YFC2102805,2019YFD1101204)the National Natural Science Foundation of China(21878142,21776132)+3 种基金Key Research and Development Plan of Jiangsu Province(BE2020712)Key Research and Development Plan of Jiangsu Province(BE2019001)Jiangsu Natural Science Fund for Distinguished Young Scholars(BK20190035)the Priority Academic Program Development of Jiangsu Higher Education Institutions(PAPD)。
文摘Cytidine 5'-monophosphate(5'-CMP)is an essential nucleotide for additives.In this study,enhanced production of 5'-CMP was realized by the transformation of cytidine using co-immobilized di-enzymes,uridine-cytidine kinase(UCK)and acetate kinase(AcK).The immobilization yield of the enzyme had a clear correlation with the surface charges as zeta potential(ξ).Among them,ε-polylysinefunctionalized sepharose(SA-EPL,ξ=9.31 m V)showed high immobilization yield(78.8%),which was4.9-fold than that of nitrilotriacetic acid functionalized sepharose(SA-NTA,ξ=-12.6 m V).The residual activity of affinity co-immobilized enzyme(EPL-Ni/EPL@Ac K-UCK)was higher than 70.6%after recycled 10 times.Thus,this study provides an effective approach for the production of 5'-CMP with the advantages of low adenosine 5'-triphosphate(ATP)consumption,reduced side reactions,and improved reusability by co-immobilized UCK and Ac K on the functionalized Sepharose.
基金Supported by National Natural Science Foundation of China(51143012)Natural Science Foundation of Shandong Province(ZR2009BM006)~~
文摘[Objective] This study aimed to establish an efficient process for separation of phycoerythrin by using Q Sepharose Fast Flow resin and verity its feasibility for scale-up. [Method] Elution gradient, sample volume and flow rate were optimized to determine the optimal separation condition, under which the scale-up process was verified. [Result] The optimal condition for separation of phycoerythrin by using Q Sepharose FF resin was investigated： 30 ml of laver extract was loaded to the Q Sepharose FF column with a bed volume of 8 ml; subsequently, the column was stepwise eluted with 0-0.10-0.35-1.00 mol/L NaCI solution （pH 6.0） at a constant flow rate of 1 ml/min; the elution peak under 0.35 mol/L NaCI solution was collected, and the recovery rate and purity coefficient （A565/A280） of phycoerythrin were determined as 44.3 and 1.15, respectively. Based on the established process, 75 ml of phycoerythrin extract was loaded to the Q Sepharose FF column with a bed volume of 20 ml for separation, while no significant variation was observed in the separation result. [Conclusion] Phycoerythrin can be well separated from laver extract by using Q Sepharose FF resin and the process is feasible for scale-up.
文摘Q Sepharose Fast Flow(Q Sepharose F.F.) was adopted to purify the suspensions of type Ⅰ,Ⅱ,Ⅲ oral poliovaccine (OPV), which is prepared from Vero cells. After clarification and concentration by ultrafiltration,the recovery of virus infected titre may attain above 85%.Using column chromatography on Q Sepharose F.F. to purify the concentrated virus suspensions,the purified viruses attain 100% recovery of virus infectivity. Dot membrane hybrization was used to detect DNA with the probe of Vero cell genome DNA which was labeled with α P 32 dATP,the contents of residual substrate DNA was less than 100 Pg/dose. The process of downstream had no significant influence on some biological characters of purified viruses,such as virus morphology ,tumorigenicity, rct/40 character and capsid protein.This downstream reseach indicates that Q Sepharose F.F. is an ideal chromatography material for purifying OPV prepared from Vero cells.
基金supported by the National Science & Technology Pillar Program of MOST (Grant No. 2011BAK15B04)
文摘The separations of single-wall carbon nanotubes on length by sepharose gel were investigated in this work. The solutions of sodium dodecyl sulfate and sodium deoxycholate were applied as the eluent in sequence. SEM and Raman were used to characterize the length of nanotube bundles. The results show that the longer nanotubes were eluted out first, and then the shorter tubes were followed by the sodium dodecyl sulfate. However, the separated order was totally reversed by the sodium deoxycholate. By this method, the process generated nanotube fractions not only were narrower in length distributions, but also could control the separation orders by changing the eluents. Moreover, the separation principle was also discussed.
文摘双特异性抗体(BsAbs),简称双抗,是一种能同时结合两个不同靶点或表位的抗体,BsAbs在表达过程中通常伴随产生多种副产物,在纯化工艺中很难被去除。本研究运用高通量筛选技术,通过对SP Sepharose Fast Flow填料的纯化条件进行探索,建立基于Sepharose Fast Flow填料的双特异性抗体纯化方法开发的通用流程,每个筛选条件只需要0.4 mg双抗样品,可同时筛选多达32个条件,筛选过程总耗时2 h,而传统柱层析需耗时64 h。通过高通量迅速筛选方法可以有效去除双抗分子副产物,为解决纯化工艺难题提供了一种新的工艺路线。
文摘目的分离、纯化具有降血糖作用的仙人掌多糖组分。方法经热水提取、乙醇沉淀、DEAE Sepharose fast flow离子交换色谱和Sephadex G系列凝胶滤过色谱纯化得到5种仙人掌多糖组分。醋酸纤维薄膜电泳检测多糖纯度,凝胶色谱测定相对分子质量,高效液相色谱测定糖的组成。结果5种多糖都已达到电泳纯。它们的相对分子质量依次为2.0×103、1.0×106、4.0×103、9.2×105、5.0×103。ODP1由鼠李糖组成,ODP2可能由鼠李糖和葡萄糖组成,ODP4可能由鼠李糖和D-半乳糖组成。ODP3和ODP5分别由鼠李糖和一未知糖组分组成。结论仙人掌多糖的提取没有必要采用脱蛋白、脱色素步骤,本实验所提取的仙人掌多糖是均一的。
