Inside Cover: Artificial Ribozyme Switches Containing Natural Riboswitch Aptamer Domains (Angew. Chem. Int. Ed. 15/2009)

Fusions of a ribozyme and an aptamer of a natural riboswitch (thiamine pyrophosphate, TPP) are used to construct artificial thiamine‐dependent switches of gene expression. As J. S. Hartig et al. describe in their Communication on page 2715 ff. , insertion of these RNA elements into bacterial mRNAs allows translation of the message to be switched on or off. TPP triggers changes to the ribozyme‐mediated mRNA cleavage, resulting in liberation of the message for translational initiation.

     

高效液相色谱/荧光法测定血浆、红细胞与尿液中的硫胺素及其磷酸酯

建立了同时测定血浆、红细胞、尿液中硫胺素(T)、一磷酸硫胺素(TMP)、二磷酸硫胺素(TDP)的柱前衍生/反相高效液相色谱-荧光检测法。样品经高氯酸除蛋白,铁氰化钾衍生后,采用反相色谱柱分离,荧光检测器测定,外标法定量。色谱柱为C18柱(250 mm×4.6 mm,5μm i.d.),流动相为0.2 mol/L KH2PO4溶液(pH 7.0,含0.000 3 mol/L四丁基氢氧化铵)-甲醇和甲醇-水,采用梯度程序进行洗脱,流速为0.8mL/min。荧光检测器激发波长为365 nm,发射波长为435 nm。结果表明:T,TMP在0.5~20.0μg/L,TDP在5.0~200.0μg/L质量浓度范围内呈良好的线性关系,相关系数分别为0.99,0.98,0.99。T,TMP,TDP的平均回收率为86.8%~110.2%,相对标准偏差(RSDs)为2.5%~10.9%。3种化合物在血浆及红细胞中的检出限(LODs)为0.03~0.35μg/L,定量下限(LOQs)为0.09~1.18μg/L。T在尿液中的检出限(LODs)为1.58μg/L,定量下限(LOQs)为5.26μg/L。该方法的准确度和精密度均较高,能够满足临床检测和科研需要。     

The Mechanism of N-Heterocyclic Carbene Organocatalysis through a Magnifying Glass

The term “N-Heterocyclic carbene organocatalysis” is often invoked in organic synthesis for reactions that are catalyzed by different azolium salts in the presence of bases. Although the mechanism of these reactions is considered today evident, a closer look into the details that have been collected throughout the last century reveals that there are many open questions and even contradictions in the field. Emerging new theoretical and experimental results offer solutions to these problems, because they show that through considering alternative reaction mechanisms a more consistent picture on the catalytic process can be obtained. These novel perspectives will be able to extend the scope of the reactions that we call today N-heterocyclic carbene organocatalysis.     

铜在硫胺燐光分析中的作用机制

硫胺在碱中加入硫酸铜可促进硫胺转化为燐光体。提高反应温度或剧烈振荡及延长反应时间均可提高硫胺燐光，加入Ｎａ２ＳＯ２Ｏ４则燐光下降。从加铜后燐光光谱不变，活化能下降，说明铜是催化硫胺氧化。氧化的硫胺有强的燐光，再加入铜则燐光下降，且不受反应温度、时间的影响，说明燐光下降是铜的猝灭作用。在一定浓度范围内，无论催化氧化与猝灭，铜量与燐光变化呈线性相关。     

Development and validation of a dried blood spot test for thiamine deficiency among infants by HPLC–fluorimetry

Thiamine deficiency, if detected early in infancy, can be treated with thiamine supplementation and can prevent seizures, other disabilities and death. The dried blood spot (DBS) sampling technique is an attractive sample collection technique for infants. The present study reports the development and validation of a highly sensitive and precise method for quantification of thiamine diphosphate from DBS. The method utilizes full‐spot analysis of a volumetrically deposited 40 μl DBS. The analyte was extracted from the DBS using 50% methanol and then derivatized using potassium ferricyanide to thiochrome. Separation was achieved with the help of an Inertsil ODS C₁₈ column (5.0 μm, 250 × 4.6 mm) using 150 mm phosphate buffer pH 7–acetonitrile (90:10, % v/v) as the mobile phase. The use of a fluorimetric detector gave a good response to the thiochrome derivative offering good sensitivity for the method. The excitation and emission wavelengths were 367 and 435 nm, respectively. The limit of detection and lower limit of quantification were 5 and 10 ng/ml, respectively. Linearity was demonstrated from 10 to 1000 ng/ml, and precision (CV) was <12.08%, at all tested quality control levels. The method accuracy was 89.34–118.89% with recoveries >80%. Bland–Altman analysis of DBS sampling vs. whole blood demonstrated a mean bias of only 1.16 ng/ml, with a majority of the 60 investigated patient samples lying within 7.2% of the corresponding concentration measured in blood, thereby meeting the clinical desirable biological specification criterion and showing that the two methods are comparable.     

732强酸性阳离子树脂催化合成盐酸维生素B1

以硫羟硫胺为原料,经双氧水氧化得硫酸维生素B1(VB1·H2SO4);VB1·H2SO4经纯化水稀释后,流经732强酸型阳离子交换树脂,用稀盐酸洗脱,浓缩后用85%乙醇重结晶得盐酸VB1,其结构经1H NMR,13C NMR,MS和元素分析确证。     

Synthesis of new antidiabetic agent by complexity between vanadyl (II) sulfate and vitamin B1: Structural,characterization, anti‐DNA damage,structural alterations and antioxidative damage studies

Complexity between thiamine (vitamin B₁) and VSO4.xH₂O salt with the suggest formula, [VO (vitB₁)₂] has been synthesized by the chemical reaction in neutralization media pH = 7.5 at 70 °C. The assignments of the elemental analysis, conductivity measurements, FT‐IR, UV–Vis, ESR spectroscopy, thermal analyses (TGA‐DTA) and magnetic moment data visualize the stoichiometry, formula and chelation of the vanadyl (II) complex. The spectroscopic analyses revealed that vitamin B₁ reacted with vanadyl (II) ions as a bidentate ligand via hydroxyl ethyl‐oxygen and sulfur of the thiazole group. New vanadyl (II) complex has the protective effect against pancreatic toxicity induced by STZ. The target of this investigation was to assess the enhancement effect of new vanadyl (II)complex in two doses on pancreatic toxicity, oxidative stress, DNA damage and hyperglycemia persuade by STZ. The rats were divided into 7 groups; control group, STZ (diabetic untreated group) (50 mg/kg), STZ plus thiamine (10 mg/Kg) (Low dose), STZ plus thiamine (50 mg/Kg) (High dose), STZ plus VSO4. xH₂O (15 mg/kg), STZ + vanadyl (II) complex (10 mg/Kg) (Low dose), STZ+ vanadyl (II) complex (50 mg/Kg) (High dose). Vanadyl (II) complex in high dose afforded a significant decline in MDA level parallel to significant elevation in antioxidant enzymes (SOD, CAT, MPO and XO) in pancreas homogenates. It may be due to the capturing activities of reactive oxygen species by the new complex, which reduces oxidative damage and enhance antioxidant capacities. The novel complex succeeded in the restoration of lipid parameters to its normal levels beside lowering TNF‐α and CRP levels. The new complex also reduces hyperglycemia induced by STZ greatly and improve histological and ultrastructure of pancreas and has a high potency in reducing DNA damage in pancreatic tissues.     

Oxalyl‐CoA Decarboxylase Enables Nucleophilic One‐Carbon Extension of Aldehydes to Chiral α‐Hydroxy Acids

The synthesis of complex molecules from simple, renewable carbon units is the goal of a sustainable economy. Here we explored the biocatalytic potential of the thiamine‐diphosphate‐dependent (ThDP) oxalyl‐CoA decarboxylase (OXC)/2‐hydroxyacyl‐CoA lyase (HACL) superfamily that naturally catalyzes the shortening of acyl‐CoA thioester substrates through the release of the C₁‐unit formyl‐CoA. We show that the OXC/HACL superfamily contains promiscuous members that can be reversed to perform nucleophilic C₁‐extensions of various aldehydes to yield the corresponding 2‐hydroxyacyl‐CoA thioesters. We improved the catalytic properties of Methylorubrum extorquens OXC by rational enzyme engineering and combined it with two newly described enzymes—a specific oxalyl‐CoA synthetase and a 2‐hydroxyacyl‐CoA thioesterase. This enzymatic cascade enabled continuous conversion of oxalate and aromatic aldehydes into valuable (S)‐α‐hydroxy acids with enantiomeric excess up to 99 %.     

Expeditious synthesis of 2,3-dihydroquinazolin-4(1H)-ones in aqueous medium using thiamine hydrochloride (VB1) as a mild,efficient, and reusable organocatalyst

A simple and straightforward synthesis of 2,3-dihydroquinazolin-4(1H)-ones is developed by reacting anthranilamide with various aldehydes or ketones under mild reaction conditions, using thiamine hydrochloride as a cost-effective, readily available, and green catalyst in water. Simple purification process, high yields within short reaction time, wide substrate scope, operational simplicity, and reusability of the catalyst up to three cycles enrich the applicability of the protocol.