SciCombinator

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δ-Valerobetaine (δVB), a constitutive metabolite of ruminant milk, is produced in the rumen from free dietary Nε-trimethyllysine occurring ubiquitously in vegetable kingdom. Biological role of δVB is poorly known. Here, the antioxidant and anti-inflammatory potential of buffalo milk δVB was tested in vitro during high-glucose (HG)-induced endothelial damage. Results indicated that δVB (0.5 mM) ameliorated the HG cytotoxicity (0.57±0.02 vs. 0.41±0.018 O.D. (P<0.01). Buffalo milk extracts enriched with δVB showed significant efficacy in decreasing reactive oxygen species, lipid peroxidation, and cytokine release during HG treatment compared to milk extracts alone (P<0.05). δVB reduced the HG-activated inflammatory signal by modulating SIRT1 (0.96±0.05 vs. 0.85±0.04 AU), SIRT6 (0.82±0.04 vs. 0.61±0.03 AU), and NF-κB (0.85±0.03 vs. 1.23±0.03 AU) (P<0.05). On the whole, our data show the first evidence of δVB efficacy in reducing endothelial oxidative stress and inflammation, suggesting a potential role of this betaine as a novel dietary compound with health-promoting properties.

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The synthesis of antibody-drug conjugates (ADCs) using the interchain cysteines of the antibody inherently gives a mixture of proteins with varying drug-to-antibody ratio. The drug distribution profiles of ADCs are routinely characterized by hydrophobic interaction chromatography (HIC). Since HIC is not in-line compatible with mass spectrometry (MS) due to the high salt levels, it is laborious to identify the constituents of HIC peaks. An MS-compatible alternative to HIC is reported here: native reversed phase liquid chromatography (nRPLC). This employs a mobile phase 50 mM ammonium acetate for high sensitivity in MS, and elution with a gradient of water/isopropanol. The key is a bonded phase giving weaker drug-surface interactions compared to the noncovalent interactions holding the antibody-drug conjugates together. The hydrophobicity of the bonded phase is varied and the least hydrophobic bonded phase in the series, polymethylmethacrylate, is found to resolve the intact constituents of a model ADC (Ab095-PZ) and a commercial ADC (brentuximab vedotin) under these MS-compatible conditions. The nRPLC-MS data show that all species, ranging from drug-to-antibody ratios of 1 to 8, remained intact in the column. This supports the premise that lower hydrophobicity of the bonded phase is the key to enabling nRPLC-MS of antibody-drug conjugates.

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Herein, we describe an alkyl thiolate-ligated iron complex that reacts with dioxygen to form an unprecedented example of an iron superoxo interme-diate, [FeIII(S2Me2N3(Pr,Pr))(O2)] (4), which is capable of cleaving strong C-H bonds. A cysteinate-ligated iron superoxo intermediate is proposed to play a key role in the biosynthesis of β-lactam antibiotics by isopenicllin N-synthase (IPNS). Superoxo 4 converts to a metastable putative Fe(III)-OOH intermediate, at rates that are de-pendent on the C-H bond strength of the H-atom donor, with a kinetic isotope effect (kH/kD= 4.8) comparable to that of IPNS (kH/kD= 5.6). The bond dissociation energy of the C-H bonds cleaved by 4 (92 kcal/mol) is compa-rable to C-H bonds cleaved by IPNS (93 kcal/mol). Both the calculated and experimental electronic absorption spectrum of 4 are comparable to that of the putative IPNS superoxo intermediate, and are shown to involve RS-→Fe-O2•- and O2•-→Fe charge transfer transitions. -back-donation by the electron-rich alkyl thiolate pre-sumably facilitates this reactivity by increasing the basicity of the distal oxygen. The frontier orbitals of 4 are shown to consist of two strongly coupled unpaired elec-trons of opposite spin, one in a superoxo *(O-O) orbital, and the other in an Fe(dxy) orbital.

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Glycan moiety of glycoproteins play key roles in various biological processes. However, there are few versatile methods for releasing, separating, and recovering monomeric reducing N-glycans for further functional analysis. In this study, we developed a new method to achieve the release, separation, and recovery of monomeric reducing N-glycans using enzyme E (Pronase E) combined with 9-chloromethyl chloroformate (Fmoc-Cl) and glycosylasparaginase (GA). Ovalbumin, Ribonuclease B (Ribo B), ginkgo, and pine nuts glycoproteins were used as materials and were sequentially enzymatically hydrolyzed with Pronase E, derivatized with Fmoc-Cl, and enzymatically hydrolyzed with GA. The products produced by this method were then detected by electrospray ionization mass spectrometry (ESI-MS), high-performance liquid chromatography (HPLC), and online hydrophilic interaction chromatography (HILIC-MS )separation. The results showed that all N-glycans with essentially one amino acid obtained by Pronase E were labeled with Fmoc-Cl and could be efficiently separated and detected via HPLC and HILIC-MS). Finally, the isolated Asn-glycan derivatives were digested with GA, enabling the recovery of all monomeric reducing N-glycans modified by core α-1,3 fucose. This method was simple, low cost, and broadly applicable and could therefore have great importance for analysis of the structure-function relationships of glycans.

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A copper-catalyzed iodination of carbazoles has been developed. Barluenga’s reagent IPy2BF4 is used to generate a soft electrophilic halonium species for the iodination of the carbazoles. This report represents the first concept of copper-catalyst promoted electrophilic halogenation of carbazoles. We demonstrated numerous applications of this methodology synthesizing diverse carbazole derivatives, i.e. both electron rich and electron deficient systems.

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A gradient system was developed for the separation of proteins at femtoliter scale utilizing nanofluidic channels. In the history of chromatography, miniaturization of the separation column is important for efficient separation and downsizing of instruments. Previously, our group developed a small and highly efficient chromatography system utilizing nanofluidic channels, although flexible design of the gradient was difficult and separation of proteins was not achieved. Here, we propose a flexible gradient system using standard HPLC pumps and an auxiliary mixer with a simple sample injection system. In contrast to our previous sample injection system using pressure balance, the system enables a femtoliter-scale sample injection which is compatible with gradient elution using HPLC pumps. The system was carefully designed, verified for sample injection and gradient elution, and finally applied to the separation of proteins from model and real samples. This femtoliter-scale, efficient separation system will contribute to omics studies at the single cell level.

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Although representativeness is often a prerequisite when sampling odors, the methods used have never been assessed from the analytical and the sensory perspective simultaneously. We validate several critical innovations in the methods used to sample odors, starting with a previously developed static-and-trapped headspace (S&T-HS) cell, to minimize sorptive biases and allow thermodesorption of trapped odors. The addition of a desorption oven allows restoration and testing of odors sampled not only by S&T-HS, but also by other techniques (solid-phase microextraction, HS sorptive extract, purge-and-trap HS). The S&T-HS cell exhibits satisfactory representativeness, much higher than the other three techniques. This allows, for the first time, a proposal to use this technology as an olfactive camera to capture and restore an odor. The method was tested on a sample of a complex fresh ashtray odor.

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The Toronto Alexithymia Scale-20 is arguably the most utilized measure of alexithymia. Although a three-factor solution has been found by numerous studies, these findings are not universal. This article examined and compared 18 competing factor structures for the Toronto Alexithymia Scale-20, which included between one and four correlated latent factor structures, common methods models that accounts for negatively worded items, and bifactor models. Although the two-factor bifactor model with a common methods factor had the better model fit compared with the other 17 models examined, it still did not achieve the requisites of a good model fit across all model fit indices. Issues stemmed primarily from the externally oriented thinking factor and the negatively worded items. Post hoc analyses indicated that a two-factor bifactor model with the negatively worded items dropped achieved the requisites of a good model fit and can be treated as a unidimensional measure despite the presence of multidimensionality. Multiple-group analysis indicated that the factor loadings were invariant across U.S. and Philippines samples. After controlling for noninvariance at the item intercept level, the Philippines sample had a higher alexithymia general score compared with the U.S. sample.

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The decisive factor in successful intensive insulin therapy is the ability to deliver need-based-adjusted nutrition-independent insulin dosages at the closest possible approximation to the physiological insulin level. Because this basal insulin requirement is strongly influenced by the patient’s lifestyle, its subtlety is of great importance. This challenge is very different between patients with type 1 diabetes and those with insulin-dependent type 2 diabetes. Furthermore, it is more difficult to finetune a basal insulin dosage with intensified conventional insulin therapy (ICT), due to delayed insulin delivery, compared to insulin pump therapy, which provides continuous delivery of small doses of exclusively short-acting insulin. In all cases, the goal is to achieve an optimal basal delivery rate.