Metabolic Glycoengineering with Azide- and Alkene-Modified Hexosamines: Quantification of Sialic Acid Levels

Metabolic glycoengineering (MGE) is an established method to incorporate chemical reporter groups into cellular glycans for subsequent bioorthogonal labeling. The method has found broad application for the visualization and isolation of glycans allowing their biological roles to be probed. Furthermore, targeting of drugs to cancer cells that present high concentrations of sialic acids on their surface is an attractive approach. We report the application of a labeling reaction using 1,2-diamino-4,5-methylenedioxybenzene for the quantification of sialic acid derivates after MGE with various azide- and alkene-modified ManNAc, GlcNAc, and GalNAc derivatives. We followed the time course of sialic acid production and were able to detect sialic acids modified with the chemical reporter group – not only after addition of ManNAc derivatives to the cell culture. A cyclopropane-modified ManNAc derivative, being a model for the corresponding cyclopropene analog, which undergoes fast inverse-electron-demand Diels-Alder reactions with 1,2,4,5-tetrazines, resulted in the highest incorporation efficiency. Furthermore, we investigated whether feeding the cells with natural and unnatural ManNAc derivative results in increased levels of sialic acids and found that this is strongly dependent on the investigated cell type and cell fraction. For HEK 293T cells, a strong increase in free sialic acids in the cell interior was found, whereas cell-surface sialic acid levels are only moderately increased.     

快速准确的硅乙氧基容量分析法——高氯酸乙酰化法

论述了高氯酸乙酰化法分析乙氧基硅单体及中间的乙氧基的含量，该法具有精密（相对偏差小于０．５％）、准确、快速、经济的特点，是蔡塞尔法和重铬酸钾法不可比的，有推广价值。     

高效液相色谱法定量分析C.I.酸性蓝9隐色体中苯甲醛邻磺酸的含量

本文阐述了C．I．酸性蓝9隐色体中苯甲醛邻磺酸的高效液相色谱定量分析方法,采用Lichrospher C18色谱柱,以甲醇／醋酸／水的体积比率为20：0．2：80为流动相,检测波长为254nm,线性范围量0～0．52mg／mL（r=0.9991）,相对标准偏差为0．28％,回收率为99．01％-100．44％。     

紫外导数光谱法定量分析2-硝基氯苯-4-磺酸

本文介绍了一阶导数紫外光谱法定量分析2-硝基氯苯-4-磺酸的方法,并与常用的化学还原-重氮法,高效液相色谱法进行了对比,结果表明,一阶导数紫外光谱法测定2-硝基氯苯-4-磺酸的含量简便,快速,准确,是一种非常实用的定量分析方法。     

对氯化聚乙烯和硬脂酸产品标准的研究和分析

按对氯化聚乙烯标准中氯含量的测试和硬脂酸标准中0.001%硫酸标准溶液的配制方法进行试验后,发现按标准中要求的试验方法检测出来的数据与实际要求不一致。经试验研究,找出标准中存在的问题原因,为正确应用标准提供了技术资料。     

高效液相色谱定量分析2,6-二硝基氯苯-4-磺酸

本文报导了高效液相色谱法定量分析2，6-二硝基氯苯-4-磺酸的方法。采用Diamonsil C18色谱枉，甲醇／水为流动相(60／40)(每500ml流动相中舍有0．30g磷酸二氢钾)，检测波长为215nm，线性范围19.83mg／L～99.16nm／L，r=0.9999；标准偏差为0．16，回收率为99．50％～100．10％。     

高效液相色谱法定量分析4-氯-3-硝基苯磺酸的方法研究

本文建立了高效液相色谱（HPLC）定量分析4-氯-3-硝基苯磺酸及其有机杂质含量的方法。通过对检测波长、流动相等色谱条件的优化,实现4-氯3-硝基苯磺酸及其杂质良好分离。采用外标法对4-氯-3-硝基苯磺酸及其杂质进行了定量。     

赤霉素和6-苄氨基嘌呤混合物的色谱分析方法

采用C18不锈钢柱,以甲醇-水-磷酸为流动相,对赤霉素和6-苄氨基嘌呤的混合物能同时进行定量分析。该分析方法的标准偏差分别为0．0229、0．0235;变异系数分别为2．1604％、2．1963％;平均回收率分别为99．57％、99．28％。     

Common FABP4 Genetic Variants and Plasma Levels of Fatty Acid Binding Protein 4 in Older Adults

We examined common variants in the fatty acid binding protein 4 gene (FABP4) and plasma levels of FABP4 in adults aged 65 and older from the Cardiovascular Health Study. We genotyped rs16909187, rs1054135, rs16909192, rs10808846, rs7018409, rs2290201, and rs6992708 and measured circulating FABP4 levels among 3190 European Americans and 660 African Americans. Among European Americans, the minor alleles of six single nucleotide polymorphisms (SNP) were associated with lower FABP4 levels (all p ≤ 0.01). Among African Americans, the SNP with the lowest minor allele frequency was associated with lower FABP4 levels (p = 0.015). The C-A haplotype of rs16909192 and rs2290201 was associated with lower FABP4 levels in both European Americans (frequency = 16 %; p = 0.001) and African Americans (frequency = 8 %; p = 0.04). The haplotype combined a SNP in the first intron with one in the 3′untranslated region. However, the alleles associated with lower FABP4 levels were associated with higher fasting glucose in meta-analyses from the MAGIC consortium. These results demonstrate associations of common SNP and haplotypes in the FABP4 gene with lower plasma FABP4 but higher fasting glucose levels.     

CMAS and ST3GAL4 Play an Important Role in the Adsorption of Influenza Virus by Affecting the Synthesis of Sialic Acid Receptors

Influenza A viruses (IAVs) initiate infection by attaching Hemagglutinin (HA) on the viral envelope to sialic acid (SA) receptors on the cell surface. Importantly, HA of human IAVs has a higher affinity for α-2,6-linked SA receptors, and avian strains prefer α-2,3-linked SA receptors, whereas swine strains have a strong affinity for both SA receptors. Host gene CMAS and ST3GAL4 were found to be essential for IAV attachment and entry. Loss of CMAS and ST3GAL4 hindered the synthesis of sialic acid receptors, which in turn prevented the adsorption of IAV. Further, the knockout of CMAS had an effect on the adsorption of swine, avian and human IAVs. However, ST3GAL4 knockout prevented the adsorption of swine and avian IAV and the impact on avian IAV was more distinct, whereas it had no effect on the adsorption of human IAV. Collectively, our findings demonstrate that knocking out CMAS and ST3GAL4 negatively regulated IAV replication by inhibiting the synthesis of SA receptors, which also provides new insights into the production of gene-edited animals in the future.     

脂肪酸甘油酯的色谱分析

应用高效液相色谱法和高分辨凝胶渗透色谱法建立了脂肪酸甘油酯中单酯、双酯和三酯的族分离方法 ,采用乙醇作流动相的非水反相色谱法可以将饱和脂肪酸甘油酯按族进行分离 ,不饱和脂肪酸甘油酯在反相色谱条件下族分离效果不理想 ;相比于HPLC方法 ,采用四氢呋喃作流动相的GPC法可以迅速地将各种多元醇酯按族进行分离 ,而且具有快速、简便等特点     

牛血清蛋白与肉桂酸和阿魏酸的相互作用研究

采用亲和毛细管电泳法,研究了牛血清蛋白(BSA)与两种结构相似的羧酸类化合物肉桂酸(CA)和阿魏酸(FA)间的相互作用。采用淌度比计算得到25℃时CA与FA的结合常数分别为3.5318×104L/mol、8.0158×104L/mol,并根据不同温度的结合常数计算了ΔS、ΔH、ΔG等热力学参数。热力学参数表明,CA、FA与BSA体系间的相互作用力以氢键作用力和范德华作用力为主。     

嗪草酮中间体——三甲基丙酮酸高效液相色谱定量分析

建立一种采用高效液相色谱法定量分析除草剂嗪草酮的重要中间体三甲基丙酮酸的方法,为嗪草酮的高效合成提供有效保证。选用Agilent ZORBAX SB-Aq色谱柱、紫外检测器,以乙腈和水(0.01 mol/L KH_2PO_4)(体积比15:85)为流动相,流速1.0 mL/min,柱温为室温,波长215 nm定量分析三甲基丙酮酸。三甲基丙酮酸的色谱保留时间为4.8 min,线性相关系数为0.999 9,方法的标准偏差为0.081,变异系数为0.17%,平均回收率为99.75%。此方法分析快速、简便、分离完全、结果重现性好,所得结果可信度高,是一种实用的中间控制分析方法。     

一种制备唾液酸磁性表面分子印迹聚合物的新方法

一锅法合成氨基化磁性纳米颗粒Fe₃O₄@NH₂,与对甲酰基苯硼酸(FPBA)反应嫁接硼酸官能团,通过硼酸基与模板唾液酸Neu5Ac分子上的顺式二醇共价反应,将Neu5Ac定向固定于磁性纳米颗粒。以多巴胺(DA)及3-氨基苯硼酸(3-APBA)为功能单体,自聚合反应形成共聚壳层包覆在磁性纳米颗粒的表面,制备得Neu5Ac磁性分子印迹聚合物(Neu5Ac-MMIPs)。通过透射电镜、红外光谱对其形态及结构进行表征,并评价其吸附性能。结果表明,Neu5Ac磁性分子印迹聚合物对Neu5Ac具有较好的吸附量、印迹效率、特异性等优点。通过对Neu5Ac至少5次吸附-洗脱的循环实验表明,MMIPs具有较好的重复再利用能力。     

Structure and Stability of Carbohydrate–Lipid Interactions. Methylmannose Polysaccharide–Fatty Acid Complexes

We report a detailed study of the structure and stability of carbohydrate–lipid interactions. Complexes of a methylmannose polysaccharide (MMP) derivative and fatty acids (FAs) served as model systems. The dependence of solution affinities and gas‐phase dissociation activation energies (E_a) on FA length indicates a dominant role of carbohydrate–lipid interactions in stabilizing (MMP+FA) complexes. Solution ¹H NMR results reveal weak interactions between MMP methyl groups and FA acyl chain; MD simulations suggest the complexes are disordered. The contribution of FA methylene groups to the E_a is similar to that of heats of transfer of n‐alkanes from the gas phase to polar solvents, thus suggesting that MMP binds lipids through dipole‐induced dipole interactions. The MD results point to hydrophobic interactions and H‐bonds with the FA carboxyl group. Comparison of collision cross sections of deprotonated (MMP+FA) ions with MD structures suggests that the gaseous complexes are disordered.     

Synthesis and Biological Activity of the Lipoteichoic Acid Anchor from Streptococcus sp. DSM 8747

In this study, the role of lipoteichoic acid (LTA) anchors in the activation of the innate immune response was investigated through the chemical synthesis of a series of LTA derivatives and the determination of their ability to induce NO production in bone marrow‐derived macrophages (BMM). To this end, an efficient synthesis of the sn‐3‐O‐(α‐D ‐galactofuranosyl)‐1,2‐di‐O‐acylglycerol LTA core was developed, which was then used as a key structure to produce both phosphate and glycerylphosphate‐funtionalised LTA anchors, as well as galactofuranosyldiglycerides with different fatty acid chain lengths. With a series of LTA anchors in hand, we then determined the effect of these glycolipids on the innate immune response by exploring their capacity to activate macrophages. Here, we report that several of the LTA‐derivatives were able to induce NO production by BMMs. In general, the unnatural (sn‐1) core glycolipid anchors showed lower levels of activity than the corresponding natural (sn‐3) analogues, and the activity of the glycolipids also appears to be dependent on the length of lipid present, with an optimum lipid length of C20 for the sn‐3 derivatives. Interestingly, a triacylated anchor and the 6‐O‐phosphorylated anchor, showed only modest activity, while the 6‐O‐glycerophosphorylated derivative was unable to induce NO production. Taken as a whole, our results highlight the subtle effects that glycolipid length can have on the ability to activate BMMs.     

定量分析法测定核酸的技术进展

介绍核酸的定量分析方法，包括化学发光法、光度分析法、荧光分析法、共振光散射分析法及电化学分析法等方法，评述各方法的特点及研究进展情况。     

X射线荧光光谱法定量测定湿法磷酸中主次成分

研究了湿法磷酸样品与助熔剂四硼酸锂在高温下熔融制成玻璃片后，在X射线荧光光谱仪上测定其Fe2O3、Al2O3、MgO、P2O5含量的方法。方法测定结果与化学分析测定结果基本一致。Fe2O3、Al2O3、MgO、P2O5的变异系数（CV）分别为：0．88％、0．60％、1．07％、0．16％。     

Exploring the Potential of Norbornene‐Modified Mannosamine Derivatives for Metabolic Glycoengineering

Metabolic glycoengineering (MGE) allows the introduction of unnaturally modified carbohydrates into cellular glycans and their visualization through bioorthogonal ligation. Alkenes, for example, have been used as reporters that can react through inverse‐electron‐demand Diels–Alder cycloaddition with tetrazines. Earlier, norbornenes were shown to be suitable dienophiles; however, they had not previously been applied for MGE. We synthesized two norbornene‐modified mannosamine derivatives that differ in the stereochemistry at the norbornene (exo/endo linkage). Kinetic investigations revealed that the exo derivative reacts more than twice as rapidly as the endo derivative. Through derivatization with 1,2‐diamino‐4,5‐methylenedioxybenzene (DMB) we confirmed that both derivatives are accepted by cells and incorporated after conversion to a sialic acid. In further MGE experiments the incorporated sugars were ligated to a fluorophore and visualized through confocal fluorescence microscopy and flow cytometry.