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Soybean-derived phosphatidylethanolamine (PE) is a valuable phospholipid component yet its high-purity form is costly and its molecular structure is poorly understood. The present study combined solvent extraction and cryopurification to purify PE. The optimal extraction conditions were as follows: material-liquid ratio 1:15 (g/mL), ethanol base concentration 100:4 (Vanhydrous ethanol /V25% ammonia), extraction temperature 40 °C, time 60 min, extraction twice. The cryopurification conditions were: material-liquid ratio 1:60, ethanol base concentration 100:6 (Vanhydrous ethanol/V25% ammonia), freezing temperature - 20 °C, time 20 h. UPLC-QTOF-MS/MS analysis revealed phospholipid composition of raw material, crude product, and purified product. The results showed that the purity of PE in the purified products was 76.74%, and the yield was 72.43% under optimal conditions. 181 phospholipid molecules were quantified. The study successfully explored high-purity PE preparation method and the composition of PE product. It provides a basis for the subsequent exploration of its biofunction and potential applications.
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It is known that phospholipase C (PLC) enzymatic degumming can hydrolyze phospholipids into diacylglycerol (DAG), which improves the efficiency of oil processing. However, it is unclear whether the presence of DAG and the use of enzymes affect the performance of the oil. This paper evaluated the frying performance of PLC-degummed refined soybean oil. Following the chicken wings and potato chips frying trials, results revealed that after 30 cycles of frying, free fatty acid (FFA) levels were 0.22% and 0.21%, with total polar compounds (TPC) at 23.75% and 24.00%, and peroxide value (PV) levels were 5.90 meq/kg and 6.45 meq/kg, respectively. Overall, PLC-degummed refined soybean oil showed almost the same frying properties as traditional water-degummed refined oil in terms of FFA, PV, TPC, polymer content, viscosity, color, foaming of frying oils, and appearance of foods. Moreover, FFA, TPC, polymer content, foaming, and color showed significant positive correlations with each other (p < 0.05) in soybean oil intermittent frying processing.
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Accurate oxylipin annotation is crucial for advancing our understanding of physiological processes in health and disease and identifying biomarkers. However, a full view of oxylipins for early diagnosis needs further attention due to the lack of proper analytical methods, which may be attributed to the wide dynamic range, poor sensitivity, extreme molecular complexity, and limited commercially available standards of oxylipins. Here, we devised a novel method by combining a chemical derivatization (CD)-based retention index (RI) algorithm and feature tandem mass spectrometric fragmentation annotation (CD-RI-LC-MS/MS) for identification and quantification of oxylipins. To this end, N,N-diethyl-1,3-diaminopropane (DEPA) was used for fast labeling of oxylipin (within 0.5 min at room temperature) to improve separation resolution and detection sensitivity. The RI algorithm was established to calibrate the retention time variances and assist the identification of oxylipins during liquid chromatography-tandem mass spectrometry (LC-MS) analysis. MS/MS analysis of in total 58 DEPA derivatives of authentic oxylipin standards was subsequently employed to obtain the tandem mass spectrometric feature fragmentation rules for further structure elucidation of the unknown regio-isomers. Finally, a method based upon CD-RI-LC-MS/MS was established for profiling oxylipins from Standard Reference Material (SRM) 1950 human plasma and nonalcoholic fatty liver disease (NAFLD) mouse liver tissue samples. A total of 87 and 96 potential oxylipins including 12 and 14 unreported oxylipins were detected and identified from human plasma and mouse liver tissues, respectively. The results showed that compared to the control group, in the liver samples of the NAFLD mouse, the content levels of prostaglandin (PG) E2, PGF2a, 8-hydroxy-eicosatrienoic acid (8-HETrE), and the newly discovered 2-hydroxy-octadecatrienoic acid (2-HOTrE) were remarkably increased, while the oxidation product of n-3 PUFA (p < 0.05) and all hydroperoxy oxylipins significantly decreased. On balance, this method contributes to future studies on oxylipin screening and application in other biological samples for facilitating the understanding of oxylipin roles in metabolic regulation of numerous diseases.
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Enfermedad del Hígado Graso no Alcohólico , Oxilipinas , Humanos , Ratones , Animales , Oxilipinas/análisis , Espectrometría de Masas en Tándem/métodos , Cromatografía Liquida , Enfermedad del Hígado Graso no Alcohólico/diagnóstico , BiomarcadoresRESUMEN
Collective systems self-organize to form globally ordered spatiotemporal patterns. Finding appropriate measures to characterize the order in these patterns will contribute to our understanding of the principles of self-organization in all collective systems. Here we examine a new measure based on the entropy of the neighbor distance distributions in the characterization of collective patterns. We study three types of systems: a simulated self-propelled boid system, two active colloidal systems, and one centimeter-scale robotic swarm system. In all these systems, the new measure proves sensitive in revealing active phase transitions and in distinguishing steady states. We envision that the entropy by neighbor distance could be useful for characterizing biological swarms such as bird flocks and for designing robotic swarms.
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BACKGROUND: Volume replacement is one of the vital techniques of oncoplastic surgery (OPS) when applying breast-conserving surgery. The clinical application of peri-mammary artery perforator flaps for this indication is uneven in China. Here, we describe the results of our clinical experience with peri-mammary artery flaps for partial breast reconstruction. METHODS: In this study, 30 patients underwent partial breast resection for quadrant breast cancer followed by partial breast reconstruction with peri-mammary artery perforator flaps, which included the thoracodorsal artery perforator flap (TDAP), anterior intercostal artery perforator flap (AICAP), lateral intercostal artery perforator flap (LICAP), and lateral thoracic artery perforator flap (LTAP). All the patients' operation plans were discussed comprehensively and were performed by sticking to every step. The satisfaction outcome was assessed with the extracted version of the BREAST-Q version 2.0, Breast Conserving Therapy Module Preoperative and Postoperative Scales both preoperatively and postoperatively. RESULTS: According to the study outcomes, the mean flap size was 5.3*4.2*2.8 cm (3.0-7.0*3.0-5.0*1.0-3.5 cm). The mean surgical time was 142 min (100-250 min). No partial flap failure was detected, and no severe complications were observed. Most patients were satisfied with the outcomes regarding the dressing, sexual life, and breast shape postoperation. Furthermore, the sensation of the surgical area, scar satisfaction, and recovery state gradually improved. Overall, LICAP and AICAP had higher scores when different flaps were compared. CONCLUSIONS: Based on this study, we found that peri-mammary artery flaps had significant value in breast-conserving surgery, especially in patients with small or medium-sized breasts. Perforators could be detected by vascular ultrasound before the operation. More than one perforator could be found most of the time. No severe complications occurred when performing a suitable plan, including discussing and recording the operation procedure; the focus of care, the choice for precise and proper perforators, and the mechanism for hiding the scars were all considered and recorded in a specific chart. Patients were satisfied with the reconstruction technique of peri-mammary artery perforator flaps after breast-conserving, and the satisfaction of AICAP and LICAP was higher. In general, this technique is suitable for partial breast reconstruction and has no negative impact on patient satisfaction.
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Neoplasias de la Mama , Mamoplastia , Arterias Mamarias , Colgajo Perforante , Humanos , Femenino , Colgajo Perforante/irrigación sanguínea , Arterias Mamarias/cirugía , Mamoplastia/métodos , Mama/cirugía , Neoplasias de la Mama/cirugía , CicatrizRESUMEN
In this study, a two-step extraction strategy (TSES) and targeted metabolomics combined with chemometrics was successfully applied for profiling of phenolic compounds in different colored rapeseeds. To this end, organic solvent extraction followed by deep eutectic solvent extraction made up the TSES with improved extraction coverage of free phenolics and enhanced extraction yield of conjugated phenolics, which combined with liquid chromatography tandem mass spectrometry (LC-MS/MS) for further profiling of phenolics. TSES-LC-MS/MS method was established with determination coefficients for phenolic compounds greater than 0.9989. Finally, the relationship between color differences and phenolic compounds in rapeseeds was investigated upon TSES-LC-MS/MS method combined with chemometrics. Syringin, kaempferol, isorhamnetin, and sinapic acid were found to be the differential phenolics for the six different colored rapeseeds and their spatial distribution in rapeseeds were presented. Consequently, our method showed great potential for future studies based on comprehensive extraction and profiling of phenolics from complex matrices.
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Brassica napus , Brassica rapa , Brassica napus/química , Cromatografía Liquida , Quempferoles/análisis , Espectrometría de Masas en Tándem/métodos , Quimiometría , Disolventes Eutécticos Profundos , Fenoles/análisis , Solventes , Cromatografía Líquida de Alta Presión/métodosRESUMEN
The application of the Shannon entropy to study the relationship between information and structures has yielded insights into molecular and material systems. However, the difficulty in directly observing and manipulating atoms and molecules hampers the ability of these systems to serve as model systems for further exploring the links between information and structures. Here, we use, as a model experimental system, hundreds of spinning magnetic micro-disks self-organizing at the air-water interface to generate various spatiotemporal patterns with varying degrees of order. Using the neighbor distance as the information-bearing variable, we demonstrate the links among information, structure, and interactions. We establish a direct link between information and structure without using explicit knowledge of interactions. Last, we show that the Shannon entropy by neighbor distances is a powerful observable in characterizing structural changes. Our findings are relevant for analyzing natural self-organizing systems and for designing collective robots.
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Lipidomics refers to the large-scale study of pathways and networks of cellular lipids in biological systems. A lipidomic analysis often involves the identification and quantification of the thousands of cellular lipid molecular species within a complex biological sample and therefore requires a well optimized method for lipid profiling. In this chapter, the methods for lipidomic analysis, including sample collection and preparation, lipid derivatization and separation, mass spectrometric identification of lipids, data processing and interpretation, and quality control, are overviewed.
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Lipidómica , Lípidos , Espectrometría de Masas , Control de CalidadRESUMEN
Echium oil has great nutritional value as a result of its high content of α-linolenic acid (ALA, 18:3ω-3) and stearidonic acid (SDA, 18:4ω-3). However, the comprehensive lipid profiling and exact structural characterization of bioactive polyunsaturated lipids in echium oil have not yet been obtained. In this study, we developed a novel pseudotargeted lipidomics strategy for comprehensive profiling and lipid structural elucidation of polyunsaturated lipid-rich echium oil. Our approach integrated untargeted lipidomics analysis with a targeted lipidomics strategy based on Paternò-Büchi (PB)-tandem mass spectrometry (MS/MS) using 2-acetylpyridine (2-AP) as the reaction reagent, allowing for high-coverage lipid profiling and simultaneous determination of CâC locations in triacylglycerols (TGs), diacylglycerols (DGs), free fatty acids (FFAs), and sterol esters (SEs) in echium oil. A total of 209 lipid species were profiled, among which 162 unsaturated lipids were identified with CâC location assignment and 42 groups of ω-3 and ω-6 CâC location isomers were discovered. In addition, relative isomer ratios of certain groups of lipid CâC location isomers were revealed. This pseudotargeted lipidomics strategy described in this study is expected to provide new insight into structural characterization of distinctive bioactive lipids in food.
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Echium , Ácidos Grasos Omega-3 , Isomerismo , Lipidómica , Lípidos , Espectrometría de Masas en TándemRESUMEN
Macadamia nuts have high nutritional value and positive health attributes. Changes to the composition and availability of these compounds during roasting contribute to product quality. In this study, changes to the chemical composition of lipids (fatty acids, triglycerides, and free fatty acids) and other phytochemicals were analyzed, and a sensory evaluation was carried out of two major varieties of macadamia nuts planted in China, after roasting. Only small changes in fatty acid (FA) content and a slight decrease in total triglycerides (TAGs) were observed after roasting. The free fatty acid (FFA) content and the peroxide value were increased by roasting. The total available polyphenol content increased by 25.6% and the oxidative stability index of kernels increased by 21.6%. The sensory scores for taste and aroma were doubled by roasting. Overall, the sensory, nutritional quality, and oxidative stability of roasted macadamia nuts were greatly improved, compared with raw nuts.
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Flax lignans (SDG) and sinapic acid (SA) both have the function of antioxidation and anti-inflammation. However, previous studies have focused mainly on biochemical measurements, gene expression analysis, and clinical assessments. There are limited studies that systematically reveal the underlying mechanism of the anti-inflammation effect of SDG or SA from the lipidomic point of view. Herein, the integrated lipidomic profiling platform was used for the analysis of free fatty acids (FFAs), phospholipids (PLs), triacylglycerols (TAGs), and oxylipins in high-fat (HF)-diet-fed mice after SDG or SA administration. Dietary supplementation of SDG or SA downregulated the levels of total TAGs and FFAs in the ApoE-/- mice model. Furthermore, 28 potential lipids were screened out and considered as key evaluation factors to understand the anti-inflammation function and mechanism of SDG and SA. The results indicated that the anti-inflammatory effect of SDG and SA was principally exerted via regulation of lipid homeostasis.
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Lino , Lignanos , Animales , Antiinflamatorios , Butileno Glicoles , Ácidos Cumáricos , Dieta Alta en Grasa/efectos adversos , Glucósidos , Lipidómica , RatonesRESUMEN
Free fatty acids (FFAs) are key intermediates of lipid metabolism that have a crucial role in many critical biological processes. The specific locations of carbon-carbon double bonds (CâC) in FFAs are often associated with distinct biological functions. Despite the rapid development of analytical techniques, identification of CâC locations in FFAs with more than three CâC bonds in complex biological matrices remains a challenge. Herein, we describe a double derivatization strategy, coupled with shotgun-mass spectrometry (MS), for unambiguous and sensitive determination of a high-coverage CâC bond (from 1 to 6) locations of FFAs. Our approach is based on combination of acetone labeling of CâC bonds and N,N-diethyl-1,2-ethanediamine (DEEA) labeling of carboxyl groups within FFAs. Acetone labeling of CâC bonds via photochemical reaction provides diagnostic ions, specific to CâC locations, and DEEA labeling of carboxyl groups significantly enhances MS response of diagnostic ions, by invoking a readily protonated tertiary amine group on FFA analytes. By exploiting this double derivatization strategy, the assignment of CâC locations of FFAs with more than three CâC bonds was achieved with high sensitivity (limit of quantitation (LOQ) 0.1-1.5 nmol/L). In contrast, such assignments were not possible by acetone labeling alone, because of the low sensitivity of diagnostic ions in negative ionization mode of MS. The applicability of our method was demonstrated by profiling of FFAs, including unsaturated FFA CâC positional isomers, in liver samples from mice with nonalcoholic fatty liver disease (NAFLD) and their lean controls. The study showed that the high-specificity and high-sensitivity method developed here is promising for accurate identification and quantitation of a wide array of FFAs in biological samples.
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In this research, a novel ultrasound-assisted extraction (USAE) method using a one-phase solvent (isopropanol, IPA) combined with liquid chromatography-electrospray ionization quadrupole time-of-flight mass spectrometry (LC-ESI-QToF-MS) was developed for exhaustive profiling of egg yolk lipids, which are considered to be essential nutrients that have important functions for human health. Compared with IPA-vortex extraction method, the optimized IPA-USAE method shows better performance in terms of extraction time, lipid losses, lipid recoveries and the number of lipids identified. With this established simple, high throughput and reliable analytical approach, untargeted comprehensive lipid profiling of four kinds of egg yolks was achieved, thus providing a powerful tool for nutritional research of egg lipids. Considering that the lipids contained in egg yolk are quite abundant in both quantity and species, the proposed IPA-USAE combined with LC-ESI-QToF-MS is also potentially applicable to comprehensive extraction and profiling of lipids in other biological samples.
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Yema de Huevo/química , Lípidos/química , 2-Propanol/química , Cromatografía Líquida de Alta Presión , Solventes/química , Espectrometría de Masa por Ionización de Electrospray , UltrasonidoRESUMEN
Microwave pretreatment of rapeseeds prior to cold-pressing is a simple and desirable method for producing high quality oils. In this study, a rapid and sensitive lipid profiling platform employing an accurate quantification strategy was established based on direct infusion electrospray ionization tandem mass spectrometry. Using this developed platform, we further investigated the effect of microwave pretreatment of rapeseeds on the contents of important lipids such as triglycerides (TAGs), phospholipids (PLs), and free fatty acids (FFAs) in 15 different rapeseed oils. Our results demonstrated that no significant changes of total FAs and TAGs contents were observed after microwave pretreatment, while FFA contents increased and PLs contents significantly increased up to 40 folds. The potential mechanism of lipid changes was also discussed. The established lipidomics profiling platform can provide reliable lipids profiling data and help to better understand the potential mechanism of microwave pretreatment in oil processing.
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Lípidos/análisis , Aceite de Brassica napus/análisis , Espectrometría de Masa por Ionización de Electrospray/métodos , Brassica rapa/química , Brassica rapa/metabolismo , Cromatografía de Gases , Ácidos Grasos no Esterificados/análisis , Microondas , Fosfolípidos/análisis , Aceites de Plantas/análisis , Aceites de Plantas/química , Aceite de Brassica napus/química , Temperatura , Triglicéridos/análisisRESUMEN
In this study, a novel strategy based on acetone stable-isotope derivatization coupled with HPLC-MS for profiling and accurate quantification of aminophospholipids (phosphatidylethanolamine and phosphatidylserine) in biological samples was developed. Acetone derivatization leads to alkylation of the primary amino groups of aminophospholipids with an isopropyl moiety; the use of deuterium-labeled acetone (d6-acetone) introduced a 6 Da mass shift that was ideally suited for profiling and quantification analysis with high selectivity and accuracy. After derivatization, significantly increased column efficiency for chromatographic separation and detection sensitivity for MS analysis of aminophospholipids was observed. Furthermore, an accuracy quantification method was developed. Aminophospholipids in biological samples were derivatized with d0-acetone; while more than two aminophospholipid standards were selected for each class of aminophospholipid and derivatized with d6-acetone, which were then used as the internal standards to typically construct a calibration curve for each class to normalize the nonuniformity response caused by the differential fragmentation kinetics resulting from the distinct chemical constitution of individual aminophospholipid species in the biological samples. The excellent applicability of the developed method was validated by profiling and quantification of aminophospholipids presented in liver samples from rats fed with different diets.
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Métodos Analíticos de la Preparación de la Muestra/métodos , Cromatografía Líquida de Alta Presión/métodos , Espectrometría de Masas/métodos , Fosfolípidos/análisis , Fosfolípidos/química , Acetona/química , Animales , Límite de Detección , Hígado/química , Masculino , Fosfolípidos/aislamiento & purificación , Ratas , Ratas Sprague-DawleyRESUMEN
Atherosclerosis is a chronic disease associated with oxidative stress and inflammatory activation and is the main underlying trigger for cardiovascular disease. There are many cardiovascular health products in the market; in order to evaluate the effect of these products, in this paper, a novel lipid profiling platform was established using the shot-gun mass spectrum method for the analysis of free fatty acid and phospholipids, and the high performance liquid chromatography coupled with mass spectroscopy method for the analysis of lipid mediators and triacylglycerol, respectively, in serum from male apolipoprotein E-knock out mice after different diet interventions. Changes in the four lipids above, and pathways and regulation of lipid metabolism in mice from different groups were further investigated. The result showed that all cardiovascular health products showed some certain potential to alleviate atherogenesis and ameliorate inflammation; among them, lemon apple cider vinegar drink and seal oil could significantly decrease triacylglycerol in mouse serum. The establishment of this lipidome profiling platform helps to better understand the metabolism regulation and intervention mechanism of different cardiovascular health products in chronic diseases, such as atherogenesis. This platform could be applied to other cardiovascular health products and provide reliable lipid profiling data for their potential effect.
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Apolipoproteínas E/deficiencia , Aterosclerosis/dietoterapia , Metabolismo de los Lípidos , Lípidos/sangre , Animales , Apolipoproteínas E/genética , Aterosclerosis/sangre , Aterosclerosis/genética , Aterosclerosis/metabolismo , Dieta , Modelos Animales de Enfermedad , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Estrés OxidativoRESUMEN
A novel composite material for prednisone molecular capture (PS-MC) was prepared by surface imprinting technique in combination with a polyethylene filter plate coated with multi-walled carbon nanotubes for the first time. PS-MC was achieved by using prednisone as the template molecule, 3-aminopropyltriethoxysilane as the monomer, and tetraethoxysilane as the cross-linker. The structure, morphology, and thermal stability of the prepared PS-MC were studied by fourier-transform infrared spectrometry, field emission scanning electron microscopy, energy-dispersive X-ray spectroscopy, and thermogravimetric analysis. PS-MC was assessed by re-binding experiments such as adsorption kinetics, adsorption isotherms, molecular identification, and applied to the separation and enrichment of prednisone in cosmetics. The results indicated that PS-MC has rapid binding kinetic, high adsorption capacity, and favorable reusability. The imprinted materials were coupled with HPLC to selectively separation, purification, and detection of prednisone from spiked cosmetic samples. The recoveries of spiked cosmetic samples were in the range of 83.0-106.0%, with relative standard deviations of less than 2.10%, and the limit of detection of 5â¯ng/mL (S/Nâ¯=â¯3).
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Cosméticos/análisis , Impresión Molecular/métodos , Nanotubos de Carbono/química , Prednisona/química , Adsorción , Cosméticos/química , Cinética , Límite de Detección , Polietileno/química , Propilaminas/química , Silanos/química , Extracción en Fase Sólida/métodos , Propiedades de SuperficieRESUMEN
Molecularly imprinting polymers (MIPs) are typically prepared using a single template molecule, which allows selective separation and enrichment of only one target analyte. It is not suitable for determination of complex real samples containing multiple analytes. In order to expand the practical application of imprinted polymers, novel dual-template magnetic molecularly imprinted polymers (MMIPs) were synthesized by surface polymerization using hydrocortisone and dexamethasone as the dual-template molecules in this study. The dual-template MMIPs were prepared by copolymerization on the surface of Fe3O4@ SiO2-NH2, the template molecules, the functional monomer acrylamide (AM), the cross-linking agent ethylene glycol dimethacrylate (EGDMA), and the initiator 2,2-azobisisobutyronitrile. The morphology, magnetic properties and adsorption characteristics of the obtained dual-template MMIPs were studied by field emission scanning electron microscopy, dynamic light scattering, Fourier transform infrared spectroscopy, thermal gravimetric analysis, and vibrating sample magnetometry, and re-binding experiments. The results indicated that dual-template MMIPs had uniform particle size, strong magnetic properties, high thermal stability, and good mass transfer rate. To investigate the selectivity of dual-template MMIPs, the template molecules were mixed along with their structural analogs. The dual-template MMIPs revealed a significantly higher adsorption amount for the template molecule than its structure analog. The dual-template MMIPs can be used for the enrichment and determination of hydrocortisone and dexamethasone in cosmetic products with the recoveries of spiked cosmetic samples ranging from 86.8-107.5% and 91.2-104.3%, respectively. The relative standard deviation (RSD) for hydrocortisone was <2.89%, and RSD for dexamethasone was <2.62%.
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Cromatografía Líquida de Alta Presión/métodos , Cosméticos/análisis , Glucocorticoides/análisis , Imanes/química , Impresión Molecular/métodos , Polímeros/química , Adsorción , Dexametasona/análisis , Hidrocortisona/análisis , Polimerizacion , Dióxido de Silicio/químicaRESUMEN
In this study, novel magnetic molecularly imprinted polymers (MMIPs) were developed as a sorbent for solid-phase extraction (SPE) and used for the selective separation of metronidazole (MNZ) in cosmetics; MNZ was detected by high-performance liquid chromatography (HPLC). First, magnetic Fe3O4 nanoparticles (NPs) were prepared by the co-precipitation of Fe(2+)and Fe(3+) ions in an ammonia solution; then oleic acid (OA) was modified onto the surface of Fe3O4NPs. Finally, the MMIP was prepared by aqueous suspension polymerization, involving the copolymerization of Fe3O4NPs@OA with MNZ as the template molecule, methacrylic acid (MAA) as the functional monomer, ethylene glycol maleic rosinate acrylate (EGMRA) as the cross-linking agent, and 2,2-azobisisobutyronitrile (AIBN) as the initiator. The MMIP materials showed high selective adsorption capacity and fast binding kinetics for MNZ; the maximum adsorption amount of the MMIP to MNZ was 46.7 mg/g. The assay showed a linear range from 0.1 to 20.0 µg/mL for MNZ with the correlation coefficient 0.999. The relative standard deviations (RSD) of intra- and inter-day ranging from 0.71 to 2.45% and from 1.06 to 5.20% were obtained. The MMIP can be applied to the enrichment and determination of MNZ in cosmetic products with the recoveries of spiked toner, powder, and cream cosmetic samples ranging from 90.6 to 104.2, 84.1 to 91.4, and 90.3 to 100.4%, respectively, and the RSD was <3.54%.
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Cromatografía Líquida de Alta Presión/métodos , Cosméticos/análisis , Nanopartículas de Magnetita/química , Metronidazol/análisis , Metronidazol/química , Impresión Molecular/métodos , Cosméticos/química , Contaminación de Medicamentos/prevención & control , Nanopartículas de Magnetita/ultraestructura , Polímeros/químicaRESUMEN
A cDNA encoding a bifunctional acetylxylan esterase/xylanase, XynS20E, was cloned from the ruminal fungus Neocallimastix patriciarum. A putative conserved domain of carbohydrate esterase family 1 was observed at the N-terminus and a putative conserved domain of glycosyl hydrolase family 11 was detected at the C-terminus of XynS20E. To examine the enzyme activities, XynS20E was expressed in Escherichia coli as a recombinant His(6) fusion protein and purified by immobilized metal ion-affinity chromatography. Response surface modeling combined with central composite design and regression analysis was then applied to determine the optimal temperature and pH conditions of the recombinant XynS20E. The optimal conditions for the highest xylanase activity of the recombinant XynS20E were observed at a temperature of 49 degrees C and a pH of 5.8, while those for the highest carbohydrate esterase activity were observed at a temperature of 58 degrees C and a pH of 8.2. Under the optimal conditions for the enzyme activity, the xylanase and acetylxylan esterase specific activities of the recombinant XynS20E toward birchwood xylan were 128.7 and 873.1 U mg(-1), respectively. To our knowledge, this is the first report of a bifunctional xylanolytic enzyme with acetylxylan esterase and xylanase activities from rumen fungus.