bluuebunny/crossref_metadata_2025 huggingface dataset:
A single‐step FACS sorting strategy in conjunction with fluorescent vital dye imaging efficiently assures clonality of biopharmaceutical production cell lines
Authors: Fieder Jürgen , Schulz Patrick , Gorr Ingo , Bradl Harald , Wenger Till
Published: April 2017

The development of biopharmaceutical production cell lines typically starts with generation of heterogeneous populations of cells, from which then single cell clones are established. Several regulatory guidelines require that production cell lines are clonal, and the actual demonstration of clonality has been increasingly demanded by regulatory authorities over the last years. Here, the authors describe the relative contribution of flow cytometry mediated deposition of single cells in multiwell plates and subsequent imaging to assurance of clonality in a state of the art approach to single cell generation. Within the flow cytometry step, two unit operations are evaluated separately, doublet discrimination during event selection for deposition and droplet deposition accuracy. The imaging procedure is evaluated for the accuracy of detection of non‐clonal populations. By employing mixing experiments of cell populations, the authors demonstrate that doublet discrimination is highly efficient, and that an appropriately set up flow cytometry system already can generate >99.5% true single cell clones. The efficiency of the described imaging process depends on several factors, reaching an optimal detection rate of non‐clonal wells of about 99.8%. Our results demonstrate that one well characterized cloning step generate biopharmaceutical production cell lines with a probability of clonality of >99.99%.
Non-technical skills of surgeons and anaesthetists in simulated operating theatre crises
Authors: Doumouras A G , Hamidi M , Lung K , Tarola C L , Tsao M W , Scott J W , Smink D S , Yule S
Published: April 2017

Background

Deficiencies in non-technical skills (NTS) have been increasingly implicated in avoidable operating theatre errors. Accordingly, this study sought to characterize the impact of surgeon and anaesthetist non-technical skills on time to crisis resolution in a simulated operating theatre.

Methods

Non-technical skills were assessed during 26 simulated crises (haemorrhage and airway emergency) performed by surgical teams. Teams consisted of surgeons, anaesthetists and nurses. Behaviour was assessed by four trained raters using the Non-Technical Skills for Surgeons (NOTSS) and Anaesthetists' Non-Technical Skills (ANTS) rating scales before and during the crisis phase of each scenario. The primary endpoint was time to crisis resolution; secondary endpoints included NTS scores before and during the crisis. A cross-classified linear mixed-effects model was used for the final analysis.

Results

Thirteen different surgical teams were assessed. Higher NTS ratings resulted in significantly faster crisis resolution. For anaesthetists, every 1-point increase in ANTS score was associated with a decrease of 53·50 (95 per cent c.i. 31·13 to 75·87) s in time to crisis resolution (P < 0·001). Similarly, for surgeons, every 1-point increase in NOTSS score was associated with a decrease of 64·81 (26·01 to 103·60) s in time to crisis resolution in the haemorrhage scenario (P = 0·001); however, this did not apply to the difficult airway scenario. Non-technical skills scores were lower during the crisis phase of the scenarios than those measured before the crisis for both surgeons and anaesthetists.

Conclusion

A higher level of NTS of surgeons and anaesthetists led to quicker crisis resolution in a simulated operating theatre environment.
Enrichment of apoplastic fluid with therapeutic recombinant protein for efficient biofarming
Authors: Chatterjee Aparajita , Das Narayan C. , Raha Sumita , Maiti Indu B. , Shrestha Ankita , Khan Ahamed , Acharya Sefali , Dey Nrisingha
Published: April 2017

Objective

For efficient biofarming we attempted to enrich plant interstitial fluid (IF)/apoplastic fluid with targeted recombinant therapeutic protein. We employed a synthetic human Glucocerebrosidase (GCB), a model biopharmaceutical protein gene in this study.

Results

Twenty one Nicotiana varieties, species and hybrids were initially screened for individual IF recovery and based on the findings, we selected Nicotiana tabacum NN (S‐9‐6), Nicotiana tabacum nn (S‐9‐7) and Nicotiana benthamiana (S‐6‐6) as model plants for raising transgenic expressing GCB via Agrobacterium mediated transformation under the control of M24 promoter; GCB specific activity in each transgenic lines were analyzed and we observed higher concentration of recombinant GCB in IF of these transgenic lines (S‐9‐6, S‐9‐7, and S‐6‐6) in comparison to their concentration in crude leaf extracts.

Conclusion

Recovery of valuable therapeutics in plant IF as shown in the present study holds great promise for promoting plant based biofarming. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:726–736, 2017
A strategy to accelerate protein production from a pool of clones in Chinese hamster ovary cells for toxicology studies
Authors: Hu Zhilan , Hsu Wendy , Pynn Abby , Ng Domingos , Quicho Donna , Adem Yilma , Kwong Zephie , Mauger Brad , Joly John , Snedecor Bradley , Laird Michael W. , Andersen Dana C. , Shen Amy
Published: April 2017

In the biopharmaceutical industry, a clonally derived cell line is typically used to generate material for investigational new drug (IND)‐enabling toxicology studies. The same cell line is then used to generate material for clinical studies. If a pool of clones can be used to produce material for IND‐enabling toxicology studies (Pool for Tox (PFT) strategy) during the time a lead clone is being selected for clinical material production, the toxicology studies can be accelerated significantly (approximately 4 months at Genentech), leading to a potential acceleration of 4 months for the IND submission. We explored the feasibility of the PFT strategy with three antibodies—mAb1, mAb2, and mAb3—at the 2 L scale. For each antibody, two lead cell lines were identified that generated material with similar product quality to the material generated from the associated pool. For two antibody molecules, mAb1 and mAb2, the material generated by the lead cell lines from 2 L bioreactors was tested in an accelerated stability study and was shown to have stability comparable to the material generated by the associated pool. Additionally, we used this approach for two antibody molecules, mAb4 and mAb5, at Tox and GMP production. The materials from the Tox batch at 400 L scale and three GMP batches at 2000 L scale have comparable product quality attributes for both molecules. Our results demonstrate the feasibility of using a pool of clonally derived cell lines to generate material of similar product quality and stability for use in IND‐enabling toxicology studies as was derived from the final production clone, which enabled significant acceleration of timelines into clinical development. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1449–1455, 2017
Microbial cell disruption for improving lipid recovery using pressurized CO2: Role of CO2 solubility in cell suspension, sugar broth, and spent media
Authors: Howlader Md Shamim , French William Todd , Shields‐Menard Sara A. , Amirsadeghi Marta , Green Magan , Rai Neeraj
Published: April 2017

The study of in situ gas explosion to lyse the triglyceride‐rich cells involves the solubilization of gas (e.g., carbon dioxide, CO2) in lipid‐rich cells under pressure followed by a rapid decompression, which allows the gas inside the cell to rapidly expand and rupture the cell from inside out. The aim of this study was to perform the cell disruption using pressurized CO2 as well as to determine the solubility of CO2 in Rhodotorula glutinis cell suspension, sugar broth media, and spent media. Cell disruption of R. glutinis was performed at two pressures of 2,000 and 3,500 kPa, respectively, at 295.2 K, and it was found from both scanning electron microscopy (SEM) and plate count that a substantial amount of R. glutinis was disrupted due to the pressurized CO2. We also found a considerable portion of lipid present in the aqueous phase after the disruption at P = 3,500 kPa compared to control (no pressure) and P = 2,000 kPa, which implied that more intracellular lipid was released due to the pressurized CO2. Solubility of CO2 in R. glutinis cell suspension was found to be higher than the solubility of CO2 in both sugar broth media and spent media. Experimental solubility was correlated using the extended Henry's law, which showed a good agreement with the experimental data. Enthalpy and entropy of dissolution of CO2 were found to be −14.22 kJ mol−1 and 48.10 kJ mol−1 K−1, 9.64 kJ mol−1 and 32.52 kJ mol−1 K−1, and 7.50 kJ mol−1 and 25.22 kJ mol−1 K−1 in R. glutinis, spent media, and sugar broth media, respectively. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:737–748, 2017
miR‐143 targets MAPK7 in CHO cells and induces a hyperproductive phenotype to enhance production of difficult‐to‐express proteins
Authors: Schoellhorn Melanie , Fischer Simon , Wagner Andreas , Handrick René , Otte Kerstin
Published: April 2017

In recent years, the number of complex but clinically effective biologicals such as multi‐specific antibody formats and fusion proteins has increased dramatically. However, compared to classical monoclonal antibodies (mAbs), these rather artificially designed therapeutic proteins have never undergone millions of years of evolution and thus often turn out to be difficult‐to‐express using mammalian expression systems such as Chinese hamster ovary (CHO) cells. To provide access to these sophisticated but effective drugs, host cell engineering of CHO production cell lines represents a promising approach to overcome low production yields. MicroRNAs (miRNAs) have recently gained much attention as next‐generation cell engineering tools. However, only very little is known about the capability of miRNAs to specifically increase production of difficult‐to‐express proteins. In a previous study we identified miR‐143 amongst others to improve protein production in CHO cells. Thus, the aim of the present study was to examine if miR‐143 might be suitable to improve production of low yield protein candidates. Both transient and stable overexpression of miR‐143 significantly improved protein production without negatively affecting cell growth and viability of different recombinant CHO cells. In addition, mitogen‐activated protein kinase 7 (MAPK7) was identified as a putative target gene of miR‐143‐3p in CHO cells. Finally, siRNA‐mediated knock‐down of MAPK7 could be demonstrated to phenocopy pro‐productive effects of miR‐143. In summary, our data suggest that miR‐143 might represent a novel genetic element to enhance production of difficult‐to‐express proteins in CHO cells which may be partly mediated by down‐regulation of MAPK7. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1046–1058, 2017
Superbanone, A New 2‐Aryl‐3‐benzofuranone and Other Bioactive Constituents from the Tube Roots of Butea superba
Authors: Boonsombat Jutatip , Prachyawarakorn Vilailak , Pansanit Acharavadee , Mahidol Chulabhorn , Ruchirawat Somsak , Thongnest Sanit
Published: April 2017

Phytochemical investigation from the tube roots of Butea superba, led to the isolation and identification of a new 2‐aryl‐3‐benzofuranone named superbanone (1), one benzoin, 2‐hydroxy‐1‐(2‐hydroxy‐4‐methoxyphenyl)‐2‐(4‐methoxyphenyl)ethanone (2), eight pterocarpans (3 – 10), and eleven isoflavonoids (11 – 21). Compound 2 was identified for the first time as a natural product. The structure of the isolated compounds was elucidated using spectroscopic methods, mainly 1D‐ and 2D‐NMR. The isolated compounds and their derivatives were evaluated for α‐glucosidase inhibitory and antimalarial activities. Compounds 3, 7, 8, and 11 showed promising α‐glucosidase inhibitory activity (IC50 = 13.71 ± 0.54, 23.54 ± 0.75, 28.83 ± 1.02, and 12.35 ± 0.36 μm, respectively). Compounds 3 and 11 were twofold less active than the standard drug acarbose (IC50 = 6.54 ± 0.04 μm). None of the tested compounds was found to be active against Plasmodium falciparum strain 94. On the basis of biological activity results, structure–activity relationships are discussed.
Chemical Synthesis and Evaluation of a Disialic Acid‐Containing Dextran Polymer as an Inhibitor for the Interaction between Siglec 7 and Its Ligand
Authors: Yamaguchi Sho , Yoshimura Atsushi , Yasuda Yu , Mori Airi , Tanaka Hiroshi , Takahashi Takashi , Kitajima Ken , Sato Chihiro
Published: April 2017

A new sialic acid (Sia)‐containing glycopolymer—a fluorescent probe with high‐density disialic acid (diSia) on the surface of polysaccharide dextran (diSia‐Dex)—was synthesized as a key molecule to regulate the Sia recognition lectins, Siglecs, that are involved in the immune system. According to our original methods, diSia was synthesized by α‐selective sialylation, and a dextran template possessing terminal acetylenes and amino groups was prepared. A diSia and a fluorescent molecule were subsequently introduced to surface‐modified dextran by Hüisgen reaction and amidation, respectively. The modulatory activity of Siglec7 was evaluated by using synthetic probes. DiSia‐Dex showed high binding avidity toward Siglec7, with a KD value of 5.87×10−10 m, and a high inhibitory activity for the interaction between Siglec7 and a ligand (GD3), with a IC50 value of 1.0 nm. Notably, diSia‐Dex was able to release Siglec7 from the pre‐existing Siglec7–GD3 complex, possibly due to its unique properties of a slow dissociation rate and a high association rate. Together, these data show that diSia‐Dex can be widely applicable as a modulator of Siglec7 functions.
New Human CD22/Siglec‐2 Ligands with a Triazole Glycoside
Authors: Prescher Horst , Schweizer Astrid , Kuhfeldt Elena , Nitschke Lars , Brossmer Reinhard
Published: April 2017

CD22 is a member of the Siglec family. Considerable attention has been drawn to the design and synthesis of new Siglec ligands to explore target biology and innovative therapies. In particular, CD22‐ligand‐targeted nanoparticles with therapeutic functions have proved successful in preclinical settings for blood cancers, autoimmune diseases, and tolerance induction. Here we report the design, synthesis and affinity evaluation of a new class of Siglec ligands: namely sialic acid derivatives with a triazole moiety replacing the natural glycoside oxygen atom. In addition, we describe important and surprising differences in binding to CD22 expressed at the cell surface for compounds with distinct valences. The new class of compounds might serve as a template for the design of ligands for other members of the Siglec family and next‐generation CD22‐ligand‐based targeted therapies.
The Lipids of the Early Endosomes: Making Multimodality Work
Authors: Arumugam Senthil , Kaur Amandeep
Published: April 2017

Early endosomes are dynamic intracellular compartments that fuse with incoming endocytic carrier vesicles and associated cargoes from the plasma membrane. It has been long known that the chemical structures of lipids confer striking properties and rich biochemistry on bilayers. Although the organisational principles of the plasma membrane are relatively better understood, understanding endosomal membranes has been challenging. It has become increasingly apparent that endosomal membranes, because of their lipid compositions and interactions, use distinct lipid chemistries. We discuss the biochemical and biophysical phenomena in play at the early endosomal membrane. We focus on cholesterol, phosphoinositides, and phosphatidylserine and their clear roles in endosome functions. We discuss the various principles and mechanisms underpinning how these lipids are implicated at the functional level in the working of endosomes, and we summarise early endosomes as a multimodal organelle employing distinct lipid‐specific mechanisms.
Sialylation Is Dispensable for Early Murine Embryonic Development in Vitro
Authors: Abeln Markus , Borst Kristina M. , Cajic Samanta , Thiesler Hauke , Kats Elina , Albers Iris , Kuhn Maike , Kaever Volkhard , Rapp Erdmann , Münster‐Kühnel Anja , Weinhold Birgit
Published: April 2017

The negatively charged nonulose sialic acid (Sia) is essential for murine development in vivo. In order to elucidate the impact of sialylation on differentiation processes in the absence of maternal influences, we generated mouse embryonic stem cell (mESC) lines that lack CMP‐Sia synthetase (CMAS) and thereby the ability to activate Sia to CMP‐Sia. Loss of CMAS activity resulted in an asialo cell surface accompanied by an increase in glycoconjugates with terminal galactosyl and oligo‐LacNAc residues, as well as intracellular accumulation of free Sia. Remarkably, these changes did not impact intracellular metabolites or the morphology and transcriptome of pluripotent mESC lines. Moreover, the capacity of Cmas−/− mESCs for undirected differentiation into embryoid bodies, germ layer formation and even the generation of beating cardiomyocytes provides first and conclusive evidence that pluripotency and differentiation of mESC in vitro can proceed in the absence of (poly)sialoglycans.
Complications caused by adenosine during catheter ablation of atrial fibrillation
Authors: Liang Jackson J. , Kumareswaran Ramanan , Hyman Matthew C. , Moore Roger A. , Lin David , Frankel David S.
Published: April 2017

Key Clinical Message

Adenosine is increasingly used to assess for dormant conduction following pulmonary vein isolation during atrial fibrillation ablation. While the half‐life of adenosine is typically short and side effects transient, operators should be aware of more serious, lasting adverse reactions including anaphylaxis and bronchospasm.
Less Frustration, More Activity—Theoretical Insights into Frustrated Lewis Pairs for Hydrogenation Catalysis
Authors: Mane Manoj V. , Vanka Kumar
Published: April 2017

The field of frustrated Lewis pair (FLP) chemistry has seen rapid development in only a few years. FLPs have performed most spectacularly in hydrogenation catalysis: a wide variety of FLP‐based systems can catalyze the hydrogenation of a range of different substrates, including imines, enamines, ketones, alkynes, and alkenes. However, FLP‐based hydrogenation catalysts are yet to match the efficiency of their transition‐metal counterparts. The current investigation reveals an important aspect of FLPs that can be exploited to improve their efficiency, that is, the more sterically hindered the FLP catalyst is, the lower is its turnover frequency. Full quantum chemical calculations with DFT for a family of different, experimentally known hydrogenation FLP catalysts shows that superior FLP catalysts can be designed by reducing the frustration (by reducing the steric demand and acid/base strength) in the FLP. However, as lowering the steric demand without reduction in the frustration can result in unwanted side reactions, the design of the most efficient FLP catalysts depends on tuning the system so that both the steric demand and the frustration are reduced appropriately.
Copper–Porphyrin–Metal–Organic Frameworks as Oxidative Heterogeneous Catalysts
Authors: Castro Kelly A. D. F. , Figueira Flávio , Mendes Ricardo F. , Cavaleiro José A. S. , Neves Maria da Graça P. M. S. , Simões Mário M. Q. , Almeida Paz Filipe A. , Tomé João P. C. , Nakagaki Shirley
Published: April 2017

The synthesis and structural characterization of novel metal–organic‐framework‐type materials obtained from the reaction of 5,10,15,20‐tetrakis[2,3,5,6‐tetrafluoro‐4‐(4‐pyridylsulfanyl)phenyl]porphyrin (H2P1) and 5,10,15,20‐tetrakis(4‐pyridyl)porphyrin (H2P2) with copper(II) acetate is reported. The new material, CuP1S, shows higher heterogeneous catalytic activity than the homogeneous porphyrin–copper(II) complex CuP1 in the oxidation of catechol into ortho‐benzoquinone in the presence of air (O2) or H2O2 (30 %).
Identifying the Electrochemical Processes in LiFeSO4F Cathodes for Lithium Ion Batteries
Authors: Blidberg Andreas , Sobkowiak Adam , Tengstedt Carl , Valvo Mario , Gustafsson Torbjörn , Björefors Fredrik
Published: April 2017

The electrochemical performance of tavorite LiFeSO4F can be considerably improved by coating the material with a conducting polymer (poly(3,4‐ethylenedioxythiophene); PEDOT). Herein, the mechanisms behind the improved performance are studied systematically by careful electrochemical analysis. It is shown that the PEDOT coating improves the surface reaction kinetics for the Li‐ion insertion into LiFeSO4F. For such coated materials no kinetic limitations remain, and a transition from solid state to solution‐based diffusion control was observed at 0.6 mA cm−2 (circa C/2). Additionally, the quantity of PEDOT is optimized to balance the weight added by the polymer and the improved electrochemical function. Post mortem analysis shows excellent stability for the LiFeSO4F‐PEDOT composite, and maintaining the electronic wiring is the most important factor for stable electrochemical cycling of LiFeSO4F. The insights and the methodology used to determine the rate‐controlling steps are readily transferable to other ion‐insertion‐based electrodes, and the findings are important for the development of improved battery electrodes.
A Novel Electrophoretic Deposited Coordination Supramolecular Network Film for Detecting Phosphate and Biophosphate
Authors: Chu Tianshu , Zhang Feng , Wang Yi , Yang Yangyi , Ng Seik Weng
Published: April 2017

A series of new lanthanide coordination supramolecular networks (CSNs) based on nalidixic acid (HNA) ligand were designed and synthesized. Their crystal structures were determined by single‐crystal XRD techniques. Thin films of these compounds were first successfully deposited on indium tin oxide (ITO) glass through one‐step electrophoretic deposition under mild conditions. The europium CSN film exhibits intense red emission and can act as a highly sensitive luminescent sensor for H2PO4− anions and adenosine triphosphate (ATP) in aqueous solution. The limits of detection of H2PO4− and ATP can achieve 0.68 and 1.9 μm respectively. The sensing mechanism was further explored through fluorescence and UV techniques. The fabricated sensor also showed excellent selectivity towards these phosphates in the presence of other coexisting ions (F−, Cl−, Br−, I−, SO4−, CO32−, NO3−, and AcO−) without interference. Moreover, we demonstrated the feasibility of using this sensor to detect ATP in bovine serum samples, suggesting the potential value of application in real biosystems.
Phenothiazinyl Boranes: A New Class of AIE Luminogens with Mega Stokes Shift, Mechanochromism, and Mechanoluminescence
Authors: Arivazhagan C. , Maity Arunava , Bakthavachalam K. , Jana Arijit , Panigrahi Suraj Kumar , Suresh Eringathodi , Das Amitava , Ghosh Sundargopal
Published: April 2017

Phenothiazines with a dimesityl boron moiety, a new class of aminoboranes with B−N linkage, were synthesized. These aminoboranes exhibited interesting photophysical behavior including aggregation‐induced emission (AIE), mechanochromism (MC), mechanoluminescence (ML), and a mega Stokes shift (up to 312 nm in hexane). The solid‐state emission of the aminoboranes could be switched reversibly by grinding–fuming processes. Furthermore, the phenothiazine derivative with a bromo and an arylborane group at 3‐ and 7‐positions exhibited bright mechanoluminescence.
Probing Phosphorus Nitride (P≡N) and Other Elusive Species Formed upon Pyrolysis of Dimethyl Phosphoramidate
Authors: Liang Shuyu , Hemberger Patrick , Levalois‐Grützmacher Joëlle , Grützmacher Hansjörg , Gaan Sabyasachi
Published: April 2017

The thermal behavior of organophosphorus compounds is intricate and poorly understood but crucial for understanding gas‐phase flame inhibition, syntheses of thermally active phosphorus‐based reactive precursors, catalytic combustion, incineration of toxic nerve gases, and astrochemistry. In this work, the pyrolysis of dimethyl phosphoramidate was investigated using photoion photoelectron coincidence spectroscopy in combination with vacuum ultraviolet synchrotron radiation. This technique enables isomer‐selective detection of reactive intermediates, which are crucial in the understanding of the decomposition process. Combined with quantum chemical calculations, the experimental results permit the formulation of a comprehensive pyrolysis reaction pathway for dimethyl phosphoramidate, consisting of several reactive phosphorus species on four possible decomposition pathways. Compared to the decomposition of dimethyl methyl phosphonate, which leads exclusively to the formation of PO radicals, substitution of the methyl with an amino group most notably yields phosphorus nitride (P≡N). This mostly favored reaction pathway involves the subsequent loss of methanol and formaldehyde to yield three PONH2 tautomers, which eliminate water to generate P≡N. The thermally induced production of PN species and its possible role in flame inhibition has not previously been reported. In addition, the adiabatic ionization energy of O=P(OCH3)2NH2 was determined to be 9.79±0.02 eV.
Preparation and Characterization of Group 13 Cyanides
Authors: Haiges Ralf , Deokar Piyush , Vasiliu Monica , Stein Trent H. , Dixon David A. , Christe Karl O.
Published: April 2017

Binary Group 13 cyanides [PPh4][Ga(CN)4], [PPh4]2[In(CN)5], and [PPh4]2[Tl(CN)5] were obtained in quantitative yields from the corresponding metal trifluorides MF3 (M=Ga, In, Tl) by fluoride–cyanide exchange reactions with Me3SiCN in the presence of stoichiometric amounts of [PPh4]CN in acetonitrile solution. The 2,2′‐bipyridine (bipy) adducts [(bipy)2Ga(CN)2][Ga(CN)4], [(bipy)In(CN)3], and [(bipy)Tl(CN)3] were obtained from the reaction of MF3 with Me3SiCN and bipy in acetonitrile. While the reaction of the metal trifluorides with Me3SiCN in acetonitrile resulted in recovery of the starting materials, the reaction of MF3 with Me3SiCN in pyridine (py) solution resulted in the formation of the pyridine adducts [(py)2Ga(CN)3], [(py)3In(CN)3], and [(py)2Tl(CN)3]. The cyano compounds were characterized by their vibrational spectra and, in most cases, by their X‐ray crystal structures.
The Prebiotic C‐Terminal Elongation of Peptides Can Be Initiated by N‐Carbamoyl Amino Acids
Authors: Abou Mrad Ninette , Ajram Ghinwa , Rossi Jean‐Christophe , Boiteau Laurent , Duvernay Fabrice , Pascal Robert , Danger Grégoire
Published: April 2017

The formation of peptides upon 1‐ethyl‐3‐(3‐dimethylaminopropyl)carbodiimide (EDC)‐promoted activation of N‐carbamoylamino acids (CAA), was considered in the scope of our recent works on carbodiimide promoted C‐terminus elongation of peptides in a prebiotic context. Thus EDC promoted activation of CAA derivatives of Tyr(Me) or Ala in dilute aqueous medium pH 5.5–6.5 in the presence of excess of AA, resulted in peptide formation by C‐terminus activation/elongation. Kinetic results similar to those of EDC‐mediated activation of N‐acyl‐AA lead us to postulate the formation of a 2‐amino‐5(4H)‐oxazolone intermediate by cyclization of the activated CAA, in spite of the absence of epimerization occurred at CAA residues. Thus, in a prebiotic context, CAA may have played a similar role as N‐acyl‐AA in the initiation of C‐terminus peptide elongation.
High‐Throughput Optical Imaging and Spectroscopy of One‐Dimensional Materials
Authors: Yao Fengrui , Chen Cheng , Liu Can , Zhang Jin , Wang Feng , Liu Kaihui
Published: April 2017

Direct visualization of one‐dimensional (1D) materials under an optical microscope in ambient conditions is of great significance for their characterizations and applications. However, it is full of challenges to achieve such goal due to their relative small size (ca. 1 nm in diameter) in the optical‐diffraction‐limited laser spot (ca. 1 μm in diameter). In this Concept article, we introduce a polarization‐based optical homodyne detection method that can be used as a general strategy to obtain high‐throughput, real‐time, optical imaging and in situ spectroscopy of polarization‐inhomogeneous 1D materials. We will use carbon nanotubes (CNTs) as an example to demonstrate the applications of such characterization with respect to the absorption signal of individual nanotubes, real‐time imaging of individual nanotubes in devices, and statistical structure information of nanotube arrays.
Synthesis, Structure, and Properties of Clathrate Si30.3(8)P15.7(8)Se7.930(3)
Authors: Zhang Hui , Peng Wei , Mu Gang , Hu Tao , Huang Fuqiang , Xie Xiaoming
Published: April 2017

The new clathrate single crystal Si30.3(8)P15.7(8)Se7.930(3) was synthesized through a solid‐state reaction, and it was characterized to have a unit cell parameter a=19.7614(1) Å and belong to space group Fm
. In this clathrate structure, a Se1−Si3 bond forms in place of the tetrakaidecahedron structure, and pental dodecahedral cages composed of Si and P atoms encapsulate Se2, Se3, and Se4 guest atoms. Similar structures are observed in partial Te‐ and Ge‐substituted compounds. Theoretical calculations based on an ordered formula Si32P14Se7 showed a semiconducting electronic structure with a band gap of 0.711 eV. A photoelectric response was observed in this compound at room temperature. Across the whole temperature range, this material exhibited a weak temperature‐dependent diamagnetic signal, and no phase transition or thermal anomaly was observed in heat capacity measurements.
Silyl Chalconium Ions: Synthesis, Structure and Application in Hydrodefluorination Reactions
Authors: Kordts Natalie , Künzler Sandra , Rathjen Saskia , Sieling Thorben , Großekappenberg Henning , Schmidtmann Marc , Müller Thomas
Published: April 2017

The synthesis of two series of silylated chalconium borates,9and10, which are based on theperi‐naphthyl andperi‐acenaphthyl framework, is reported (chalcogen (Ch): O, S, Se, Te). NMR investigations of the selenium‐ and tellurium‐containing precursor silanes3 d–fand8 d, frevealed a significant through‐spaceJ‐coupling between the chalcogen nuclei and the Me2SiH group. Experimental and computational results typify the synthesized cations9and10as chalconium ions. The imposed ring strain weakens the Si−Ch linkage compared to acyclic chalconium ions. This attenuation of the Si−Ch bond strength is more pronounced in the acenaphthene series. Surprisingly, the Si−O bonds in oxonium ions9 aand10 aare the weakest Si−Ch linkage in both series. The synthesized silyl chalconium borates are active in hydrodefluorination reactions of alkyl fluorides with silanes. A cooperative activation of the silane by the Lewis acidic (silicon) and by the Lewis basic side (chalcogen) is suggested.
The Surprising Importance of Peptide Bond Contacts in Drug–Protein Interactions
Authors: Parrish Robert M. , Sitkoff Doree F. , Cheney Daniel L. , Sherrill C. David
Published: April 2017

The study of noncovalent interactions, notably including drug–protein binding, relies heavily on the language of localized functional group contacts: hydrogen bonding, π–π interactions, CH–π contacts, halogen bonding, etc. Applying the state‐of‐the‐art functional group symmetry‐adapted perturbation theory (F‐SAPT) to an important question of chloro versus methyl aryl substitution in factor Xa inhibitor drugs, we find that a localized contact model provides an incorrect picture for the origin of the enhancement of chloro‐containing ligands. Instead, the enhancement is found to originate from many intermediate‐range contacts distributed throughout the binding pocket, particularly including the peptide bonds in the protein backbone. The contributions from these contacts are primarily electrostatic in nature, but require ab initio computations involving nearly the full drug–protein pocket system to be accurately quantified.
Structural Studies of Nicotinoids: Cotinine versus Nicotine
Authors: Uriarte Iciar , Pérez Cristóbal , Caballero‐Mancebo Elena , Basterretxea Francisco J. , Lesarri Alberto , Fernández José A. , Cocinero Emilio J.
Published: April 2017

Invited for the cover of this issue is the group of Emilio J. Cocinero at the University of the Basque Country. The image depicts a fight between two biomolecules, cotinine and nicotine. Read the full text of the article at 10.1002/chem.201700023.
Generation of β‐Halo Vinylsulfones through a Multicomponent Reaction with Insertion of Sulfur Dioxide
Authors: Xiang Yuanchao , Kuang Yunyan , Wu Jie
Published: April 2017

A four‐component reaction of terminal alkynes, aryldiazonium tetrafluoroborates, sulfur dioxide surrogate of DABCO⋅(SO2)2, and potassium halide in the presence of copper(I) chloride (10 mol %) gives rise to β‐halo vinylsulfones with good stereoselectivity. The vicinal difunctionalization of alkynes through sulfonylation and halogenation with the insertion of sulfur dioxide works efficiently. A plausible mechanism is proposed, which includes a radical process.
A chiral, porous, organic cage‐based, enantioselective potentiometric sensor for 2‐aminobutanol
Authors: Duan Ai‐Hong , Wang Bang‐Jin , Xie Sheng‐Ming , Zhang Jun‐Hui , Yuan Li‐Ming
Published: April 2017

A new enantioselective potentiometric sensor containing R‐type chiral porous organic cage CC9 as the chiral selector was designed for the assay of 2‐aminobutanol. Optimized membrane electrodes displayed a linear dynamic range from 10−3 ~ 10−1 mol·L−1 with a detection limit of 2.5 × 10−4 mol·L−1 and a Nernstian response of 27 ± 0.5mV·decade−1 toward S‐2‐aminobutanol within the pH range 7.0–10.0. The potentiometric enantioselectivity coefficient (
) of this sensor was −1.333, indicating that the porous organic cage‐based electrode exhibited good discrimination toward S‐2‐aminobutanol over R‐2‐aminobutanol.
Anaerobic Digestion Model (AM2) for the Description of Biogas Processes at Dynamic Feedstock Loading Rates
Authors: Arzate Juan A. , Kirstein Max , Ertem Funda C. , Kielhorn Erich , Ramirez Malule Howard , Neubauer Peter , Cruz‐Bournazou M. Nicolas , Junne Stefan
Published: April 2017

The anaerobic digestion of biomass is a complex, dynamic, and highly nonlinear process. Hence, a mathematical model that is capable of describing the complete operability region with sufficient accuracy cannot be identifiable even with the most advanced process monitoring technology. The complexity of the anaerobic digestion model no. 1 (ADM1) leads to the application of either parameter‐reduced versions of the ADM1 or simpler models as, e.g., the anaerobic digestion model AM2. A comparative study for the simulation of biogas formation at dynamic feedstock loading using maize silage was performed with both models.
New Fluorescent Conjugates Displaying Solvatochromic Properties
Authors: Liu Huijing , Huang Rongrong , Fang Yu
Published: April 2017

Four new fluorescent dyes based on bis(phenyl‐ethynyl‐)‐2‐naphthyl (BPEN) were designed and synthesized. To improve the solvatochromic property and enhance the brightness of the fluorescent BPEN, an electron‐donating unit of azetidine and/or an electron deficient group of ‐NO2 was introduced. The corresponding fluorophores are denoted as T1, T2, T3 and T4, respectively. Moreover, to facilitate derivatization of the probes, two ethoxy carbonyl residues were grafted onto the side positions of BPEN. Spectroscopic studies demonstrated that introduction of azetidine leads to superior solvatochromic properties and largely enhanced fluorescence quantum yields as evidenced by the fact that T3 shows more than 150 nm shift in its maximum emission when dissolved in solvents of very different polarities and displays high fluorescence quantum yields in the solvents studied. However, T2, which is the one bearing a ‐NO2 group, is non‐fluorescent. Theoretical analysis and Lippert‐Mataga modeling revealed the intra‐molecular charge transfer (ICT) nature of the solvatochromic behavior of the compounds. Further test reveals that the fluorophores, in particular T3, are sensitive to the presence of trace water in less polar solvents, such as THF and 1,4‐dioxane. Moreover, it is believed that the new fluorophores may serve as building blocks for creating environment‐sensitive fluorescent sensors.
High‐dose chemotherapy with thiotepa, busulfan, and cyclophosphamide and autologous stem cell transplantation for patients with primary central nervous system lymphoma in first complete remission
Authors: DeFilipp Zachariah , Li Shuli , El‐Jawahri Areej , Armand Philippe , Nayak Lakshmi , Wang Nancy , Batchelor Tracy T. , Chen Yi‐Bin
Published: April 2017

BACKGROUND

High‐dose chemotherapy and autologous stem cell transplantation (HDC‐ASCT) is a therapeutic option for patients with primary central nervous system lymphoma (PCNSL). To the authors’ knowledge, data are limited regarding its use among patients in first complete remission (CR1) with the CNS‐directed conditioning regimen of thiotepa, busulfan, and cyclophosphamide (TBC).

METHODS

A retrospective analysis of patients with PCNSL in CR1 who underwent transplantation using a TBC‐based conditioning regimen at 2 academic institutions was performed.

RESULTS

Forty‐six consecutive patients who underwent HDC‐ASCT while in CR1 were identified. The most common induction regimen was high‐dose methotrexate plus temozolomide and rituximab (59%). No patients received whole‐brain radiotherapy. A total of 40 patients (87%) received cytarabine before undergoing ASCT as either induction intensification, early consolidation therapy, or mobilization. The median time from diagnosis to transplantation was 6 months (range, 4‐15 months). The median age of the patients at the time of transplantation was 59 years (range, 27‐69 years). With a median follow‐up of 2.7 years after ASCT (range, 6 months‐7.5 years), the Kaplan‐Meier estimates of 2‐year overall survival and progression‐free survival were 95% (95% confidence interval [95% CI], 80%‐99%) and 92% (95% CI, 77%‐97%), respectively. The most common toxicities were severe mucositis (35%) and bacterial infections occurring within 100 days of transplantation (35%). The estimated 2‐year nonrecurrence mortality rate was 2.9% (95% CI, 0.2%‐13.4%).

CONCLUSIONS

HDC‐ASCT with a CNS‐directed conditioning regimen such as TBC should be considered for patients with PCNSL who are in CR1 because this approach is associated with encouraging disease control and survival in this select patient population. Cancer 2017;123:3073–79. © 2017 American Cancer Society.
Method for DNA Ploidy Analysis Along with Immunophenotyping for Rare Populations in a Sample using FxCycle Violet
Authors: Tembhare Prashant , Badrinath Yajamanam , Ghogale Sitaram , Subramanian Papagudi Ganesan
Published: April 2017

The clinical use of flow cytometric DNA ploidy assay has been extended towards stratifying the risk of diseases, such as monoclonal gammopathies or B cell acute lymphoblastic leukemia, and to detect circulating tumor cells, both of which require detection of minute cell populations. This unit describes a protocol for determining DNA ploidy in fixed samples with simultaneous surface immunophenotyping. It is an easy method for simultaneous 6‐ to 8‐color immunophenotyping and DNA content analysis using FxCycle Violet (FCV; DAPI) dye. This protocol is a one‐step modification of routine multicolor immunophenotyping that includes surface staining followed by fixation and then DNA staining with FCV. It utilizes mature lymphocytes from the sample as an internal control for determination of DNA index. It is a sensitive method that allows DNA‐ploidy determination and cell cycle analysis in a rare tumor population as low as 100 events, as well as DNA ploidy determination in various subsets of hematopoietic cells in the same sample based on their immunophenotype. © 2017 by John Wiley & Sons, Inc.
Bioequivalence Study of Warfarin in Healthy Chinese Volunteers With a Validated High‐Performance Liquid Chromatography‐Mass Spectrometry Method
Authors: Li Wenlong , Bu Fanlong , Li Rong , Wang Benjie , Shaikh Abdul Sami , Zhang Yunyun , Guo Ruichen , Zhang Rui
Published: April 2017

This study was designed to investigate the pharmacokinetics of an innovative film‐coated warfarin sodium tablet and to compare it with the marketed sugar‐coated warfarin sodium tablet in humans. A single‐dose, open‐label, randomized, two‐way crossover study was performed in 24 healthy Chinese male volunteers. They were administered 2.5 mg of innovative film‐coated warfarin sodium tablets or the marketed sugar‐coated warfarin sodium tablets. Blood samples were collected at different time points after dosing for investigation of the pharmacokinetics of warfarin in human plasma. A sensitive liquid chromatography mass spectrometry method was established to determine warfarin in plasma. Drug and Statistics 2.1.1 was applied to calculate the pharmacokinetics parameters. The main pharmacokinetic parameters for film‐coated and sugar‐coated warfarin were the following: t½, 103.5 ± 18.8 and 105.8 ± 21.3 hours; Tmax, 0.7 ± 0.5 and 1.3 ± 0.8 hours; Cmax, 347.8 ± 74.8 and 322.9 ± 75.7 ng/mL; AUC0∼360, 16,024.2 ± 3713.9 and 15,586.6 ± 3477.0 ng·mL−1·h; AUC0∼∞, 17,335.7 ± 4089.1 and 16,912.0 ± 3911.2 ng·mL−1·h, respectively. The human pharmacokinetics of film‐coated and sugar‐coated warfarin were slightly different. The oral absorption and bioavailability of innovative film‐coated warfarin were slightly higher than those of the sugar‐coated warfarin. This study is vital to clinical usage of warfarin not only because of the pharmacokinetic parameters of the 2 pharmaceutical dosage forms of warfarin but also to obtain data on the prevalence of CYP2C9 and VKORC1 genes and their influence on the concentration of warfarin.
Effect of Solvent Choice on the Self‐Assembly Properties of a Diphenylalanine Amphiphile Stabilized by an Ion Pair
Authors: Mayans Enric , Ballano Gema , Sendros Javier , Font‐Bardia Merçè , Campos J. Lourdes , Puiggalí Jordi , Cativiela Carlos , Alemán Carlos
Published: April 2017

A diphenylalanine (FF) amphiphile blocked at the C terminus with a benzyl ester (OBzl) and stabilized at the N terminus with a trifluoroacetate (TFA) anion was synthetized and characterized. Aggregation of peptide molecules was studied by considering a peptide solution in an organic solvent and adding pure water, a KCl solution, or another organic solvent as co‐solvent. The choice of the organic solvent and co‐solvent and the solvent/co‐solvent ratio allowed the mixture to be tuned by modulating the polarity, the ionic strength, and the peptide concentration. Differences in the properties of the media used to dissolve the peptides resulted in the formation of different self‐assembled microstructures (e.g. fibers, branched‐like structures, plates, and spherulites). Furthermore, crystals of TFA⋅FF‐OBzl were obtained from the aqueous peptide solutions for X‐ray diffraction analysis. The results revealed a hydrophilic core constituted by carboxylate (from TFA), ester, and amide groups, and the core was found to be surrounded by a hydrophobic crown with ten aromatic rings. This segregated organization explains the assemblies observed in the different solvent mixtures as a function of the environmental polarity, ionic strength, and peptide concentration.
Transfection by Electroporation
Authors: Potter Huntington , Heller Richard
Published: April 2017

Electroporation—the use of high‐voltage electric shocks to introduce DNA into cells—can be used with most cell types, yields a high frequency of both stable transformation and transient gene expression, and, because it requires fewer steps, can be easier than alternative techniques. This unit describes electroporation of mammalian cells, including ES cells, for the preparation of knock‐out, knock‐in, and transgenic mice. Protocols are described for the use of electroporation in vivo to perform gene therapy for cancer, as well as for DNA vaccination. © 2017 by John Wiley & Sons, Inc.
Gut Microbiome Standardization in Control and Experimental Mice
Authors: McCoy Kathy D. , Geuking Markus B. , Ronchi Francesca
Published: April 2017

Mouse models are used extensively to study human health and to investigate the mechanisms underlying human disease. In the past, most animal studies were performed without taking into consideration the impact of the microbiota. However, the microbiota that colonizes all body surfaces, including the gastrointestinal tract, respiratory tract, genitourinary tract, and skin, heavily impacts nearly every aspect of host physiology. When performing studies utilizing mouse models it is critical to understand that the microbiome is heavily impacted by environmental factors, including (but not limited to) food, bedding, caging, and temperature. In addition, stochastic changes in the microbiota can occur over time that also play a role in shaping microbial composition. These factors lead to massive variability in the composition of the microbiota between animal facilities and research institutions, and even within a single facility. Lack of experimental reproducibility between research groups has highlighted the necessity for rigorously controlled experimental designs in order to standardize the microbiota between control and experimental animals. Well controlled experiments are mandatory in order to reduce variability and allow correct interpretation of experimental results, not just of host‐microbiome studies but of all mouse models of human disease. The protocols presented are aimed to design experiments that control the microbiota composition between different genetic strains of experimental mice within an animal unit. © 2017 by John Wiley & Sons, Inc.
Elucidation of Intramolecular Through‐Space Electronic Communication in a Propeller‐Shaped Molecule
Authors: Kodama Takuya , Hirao Yasukazu , Nishiuchi Tomohiko , Kubo Takashi
Published: April 2017

A novel π‐extended trinaphtho[3.3.3]propellane (TNP) derivative was prepared and through‐space electronic communication in its radical cation species was characterized by electrochemical and spectroscopic methods. Cyclic voltammetry and UV/Vis‐NIR titrations suggested weak electronic communication among the three aromatic rings. Variable‐temperature ESR measurements of the radical cation showed a drastic change in the spectra in the range of 230–180 K, indicating hopping‐type spin delocalization that would originate from migration of a counter anion.
Direct Isolation of Seamless Mutant Bacterial Artificial Chromosomes
Authors: Lyozin George T. , Kosaka Yasuhiro , Bhattacharje Gourab , Yost H. Joseph , Brunelli Luca
Published: April 2017

Seamless (i.e., without unwanted DNA sequences) mutant bacterial artificial chromosomes (BACs) generated via recombination‐mediated genetic engineering (recombineering) are better suited to study gene function compared to complementary DNA (cDNA) because they contain only the specific mutation and provide all the regulatory sequences required for in vivo gene expression. However, precisely mutated BACs are typically rare (∼1:1,000 to 1:100,000), making their isolation quite challenging. Although these BACs have been classically isolated by linking the mutation to additional genes, i.e., selectable markers, this approach is prone to false positives and is labor‐intensive because it requires the subsequent removal of the selectable marker. We created Founder Principle–driven Enrichment (FPE), a method based on the population genetics “founder principle,” to directly isolate rare mutant BACs, without any selectable marker, from liquid cultures via the polymerase chain reaction (PCR). Here, we provide a detailed description of FPE, including protocols for BAC recombineering and PCR screening. © 2017 by John Wiley & Sons, Inc.
Measuring Protein Interactions by Optical Biosensors
Authors: Zhao Huaying , Boyd Lisa F. , Schuck Peter
Published: April 2017

This unit gives an introduction to the basic techniques of optical biosensing for measuring equilibrium and kinetics of reversible protein interactions. Emphasis is placed on description of robust approaches that will provide reliable results with few assumptions. How to avoid the most commonly encountered problems and artifacts is also discussed. © 2017 by John Wiley & Sons, Inc.