Arnon法是叶绿素提取和测定最经典、最常用的方法,此法虽经多次改进,但仍存在着检测波长误差大、计算公式有误、提取速度慢、测定结果不够准确以及操作步骤繁琐等缺陷。该文提出了二甲基亚砜(DMSO)高温提取、80%丙酮稀释的两步快速浸提...Arnon法是叶绿素提取和测定最经典、最常用的方法,此法虽经多次改进,但仍存在着检测波长误差大、计算公式有误、提取速度慢、测定结果不够准确以及操作步骤繁琐等缺陷。该文提出了二甲基亚砜(DMSO)高温提取、80%丙酮稀释的两步快速浸提法,使叶绿素提取和测定过程缩短至3小时以内。通过对提取温度、提取时间、稀释比例及吸收光谱等进行系统分析,筛选出了叶绿素的最佳提取条件和叶绿素含量的准确计算公式,并用多种类型的植物材料验证了改进后的提取方法,证明该方法具优越性和可靠性。具体测定方法是将植物材料切成1 mm宽的细丝或细段,取50-100 mg材料于10 m L具塞试管中;加入2 m L DMSO,使植物材料浸没其中,65°C高温避光提取至植物材料变白或透明;冷却后加入8 m L 80%丙酮,混匀,测定663.6和646.6 nm处的吸光度。用公式计算叶绿素浓度:Ca(mg·L^(-1))=12.27A663.6-2.52A646.6;Cb(mg·L^(-1))=20.10A646.6-4.92A663.6;CT(mg·L^(-1))=Ca+Cb=7.35A663.6+17.58A646.6。展开更多
Feasible and effective cell models for hepatitis B virus(HBV) infection are required for investigating the complete lifecycle of this virus, including the early steps of viral entry. Resistance to dimethyl sulfoxide/p...Feasible and effective cell models for hepatitis B virus(HBV) infection are required for investigating the complete lifecycle of this virus, including the early steps of viral entry. Resistance to dimethyl sulfoxide/polyethylene glycol(DMSO/PEG), h NTCP expression, and a differentiated state are the limiting factors for successful HBV infection models. In the present study, we used a hepatoma cell line(Hu7^(hDNTCPh)) to overcome these limiting factors so that it exhibits excellent susceptibility to HBV infection. To achieve this goal, different hepatoma cell lines were tested with 2.5% DMSO/4%PEG8000, and one resistant cell line(Huh7 D) was used to construct a stable h NTCP-expressing cell line(Hu7^(hDNTCPh)) using a recombinant lentivirus system. Then, the morphological characteristics and differentiation molecular markers of Hu7^(hDNTCPh) cells with or without DMSO treatment were characterized. Finally, the susceptibility of Hu7^(hDNTCPh) cells to HBV infection was assessed. Our results showed that Huh7 D cells were resistant to 2.5% DMSO/4% PEG8000, whereas the others were not. Hu7^(hDNTCPh) cells were established to express a high level of h NTCP compared to liver extracts, and Hu7^(hDNTCPh) cells rapidly transformed into a non-dividing, well-differentiated polarized phenotype under DMSO treatment. Hu7^(hDNTCPh) cells fully supported the entire lifecycle of HBV infection. This cell culture system will be useful for the analysis of host-virus interactions, which should facilitate the discovery of antiviral drugs and vaccines.展开更多
采用汽液双循环相平衡仪测定了101.33 k Pa下苯-噻吩、苯-二甲基亚砜及苯-噻吩-二甲基亚砜的等压汽液相平衡数据。两组二元体系实验数据均通过了Herington面积法的热力学一致性检验,分别采用NRTL和UNIQUAC活度系数模型对二元体系实验数...采用汽液双循环相平衡仪测定了101.33 k Pa下苯-噻吩、苯-二甲基亚砜及苯-噻吩-二甲基亚砜的等压汽液相平衡数据。两组二元体系实验数据均通过了Herington面积法的热力学一致性检验,分别采用NRTL和UNIQUAC活度系数模型对二元体系实验数据进行热力学关联,得到了相应的模型参数,温度偏差和汽相组成偏差,关联结果表明两种模型均能较好的关联实验数据,其中UNIQUAC模型的关联结果稍优于NRTL。利用二元体系UNIQUAC模型参数预测三元体系汽液平衡数据,预测值与实验值相比,平均温度偏差1.37 K,汽相各组分平均绝对偏差均在3%以内,获得了较优的预测结果,从而为工业上苯-噻吩萃取精馏分离过程提供可靠的热力学基础数据。展开更多
文摘Arnon法是叶绿素提取和测定最经典、最常用的方法,此法虽经多次改进,但仍存在着检测波长误差大、计算公式有误、提取速度慢、测定结果不够准确以及操作步骤繁琐等缺陷。该文提出了二甲基亚砜(DMSO)高温提取、80%丙酮稀释的两步快速浸提法,使叶绿素提取和测定过程缩短至3小时以内。通过对提取温度、提取时间、稀释比例及吸收光谱等进行系统分析,筛选出了叶绿素的最佳提取条件和叶绿素含量的准确计算公式,并用多种类型的植物材料验证了改进后的提取方法,证明该方法具优越性和可靠性。具体测定方法是将植物材料切成1 mm宽的细丝或细段,取50-100 mg材料于10 m L具塞试管中;加入2 m L DMSO,使植物材料浸没其中,65°C高温避光提取至植物材料变白或透明;冷却后加入8 m L 80%丙酮,混匀,测定663.6和646.6 nm处的吸光度。用公式计算叶绿素浓度:Ca(mg·L^(-1))=12.27A663.6-2.52A646.6;Cb(mg·L^(-1))=20.10A646.6-4.92A663.6;CT(mg·L^(-1))=Ca+Cb=7.35A663.6+17.58A646.6。
基金supported by the National Natural Science Foundation of China (Grant number: 81601760, 31621061 and 81461130019)General Financial Grant from the China Postdoctoral Science Foundation (Grant number: 2016M602587)+1 种基金the Shenzhen Foundation of Science and Technology (Grant number: JCYJ20160425 104534335)supported by the Youth Innovation Promotion Association CAS (No.201603)
文摘Feasible and effective cell models for hepatitis B virus(HBV) infection are required for investigating the complete lifecycle of this virus, including the early steps of viral entry. Resistance to dimethyl sulfoxide/polyethylene glycol(DMSO/PEG), h NTCP expression, and a differentiated state are the limiting factors for successful HBV infection models. In the present study, we used a hepatoma cell line(Hu7^(hDNTCPh)) to overcome these limiting factors so that it exhibits excellent susceptibility to HBV infection. To achieve this goal, different hepatoma cell lines were tested with 2.5% DMSO/4%PEG8000, and one resistant cell line(Huh7 D) was used to construct a stable h NTCP-expressing cell line(Hu7^(hDNTCPh)) using a recombinant lentivirus system. Then, the morphological characteristics and differentiation molecular markers of Hu7^(hDNTCPh) cells with or without DMSO treatment were characterized. Finally, the susceptibility of Hu7^(hDNTCPh) cells to HBV infection was assessed. Our results showed that Huh7 D cells were resistant to 2.5% DMSO/4% PEG8000, whereas the others were not. Hu7^(hDNTCPh) cells were established to express a high level of h NTCP compared to liver extracts, and Hu7^(hDNTCPh) cells rapidly transformed into a non-dividing, well-differentiated polarized phenotype under DMSO treatment. Hu7^(hDNTCPh) cells fully supported the entire lifecycle of HBV infection. This cell culture system will be useful for the analysis of host-virus interactions, which should facilitate the discovery of antiviral drugs and vaccines.
文摘采用汽液双循环相平衡仪测定了101.33 k Pa下苯-噻吩、苯-二甲基亚砜及苯-噻吩-二甲基亚砜的等压汽液相平衡数据。两组二元体系实验数据均通过了Herington面积法的热力学一致性检验,分别采用NRTL和UNIQUAC活度系数模型对二元体系实验数据进行热力学关联,得到了相应的模型参数,温度偏差和汽相组成偏差,关联结果表明两种模型均能较好的关联实验数据,其中UNIQUAC模型的关联结果稍优于NRTL。利用二元体系UNIQUAC模型参数预测三元体系汽液平衡数据,预测值与实验值相比,平均温度偏差1.37 K,汽相各组分平均绝对偏差均在3%以内,获得了较优的预测结果,从而为工业上苯-噻吩萃取精馏分离过程提供可靠的热力学基础数据。
