Nano- and microbiodevices were developed for the high-performance separation of DNA, RNA, protein, sugar chain, and other biomolecules. We describe numerous advantages of nano- and microbiodevices as the separation technologies, including HPLC and capillary electrophoretic separation of DNA, nanopillar devices for the ultra-fast separation of DNA and proteins, nanoball materials for the fast separation of wide range of DNA fragments, nanowire devices for ultra-fast separation of DNA, RNA, and proteins. Because of the high efficiency of nano- and microbiodevices for the separation of biomolecules in the clinical blood and urea samples, numerous types of nano- and microbiodevices were developed and applied to the diagnosis of diseases, including cancer, diabetes, hypertension, infectious diseases, and other noncommunicable diseases.
Recent analytical advancements for sedative drug monitoring are a major topic in modern clinical stages. The liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) technique has been used for conventional therapeutic drug monitoring in the clinical stages. From the middle and late-2000s, LC-MS/MS with an electrospray ionization (ESI) interface has played an important role in pharmacokinetics, drug monitoring and metabolism studies since its introduction to the pharmaceutical industry and laboratory. Obviously, clinical researchers have needed highly sensitive, selective and accurate techniques for the sedative drug monitoring in various clinical stages. We review the current developments and reports regarding the application of LC-MS/MS with an ESI interface for sedative drug monitoring in the clinical stage.
In this work, quaternary ammonium group-modified celluloses (QCs) were homogeneously synthesized by reacting cellulose with 3-chloro-2-hydroxypropyltrimethylammonium chloride in NaOH/urea solutions. The structure and solution properties of the QCs were characterized by elemental analysis, FT/IR, 1H-NMR, and size exclusion chromatography. The results show that water-soluble QCs, with degree of substitution values, defined as the substitution of free hydroxyl groups of cellulose, 0.49–0.72 and molecular weight 21–66 kDa could be obtained by optimizing the reaction time. The synthesized QCs were tested for protein separation as physically adsorbed coatings in capillary electrophoresis. Among the derivatives studied, quaternary ammonium cellulose showed rapid electroosmotic mobility and effective suppression of protein adsorption. The EOF can be manipulated for various applications by using QCs with different molecular weight. Because the reversed EOF can be obtained over a broad pH range, it is possible to separate basic, neutral, and acidic proteins under physiological conditions. Eight proteins; lysozyme, ribonuclease A, cytochrome C, α-chymotrypsinogen, α-lactalbumin, ovalbumin, transferrin, and myoglobin were baseline separated within 25 min by 25 mM sodium phosphate buffers (pH 7.0) containing 100 µg/mL QC.
Flavor compounds in beverages were determined by a purge-and-trap (PT) method with a needle-type extraction device in gas chromatography-mass spectrometry. Optimum extraction medium was quantitatively evaluated for extraction/desorption of target analytes. A graphite carbon adsorbent of Carbopack X showed excellent extraction and desorption efficiencies for all the investigated flavor compounds in this PT method. On the basis of a systematic optimization of the several experimental conditions, a simple and quantitative analytical method was developed for a wide range of flavors including esters, alcohols, and aromatic compounds. The limit of quantification of the analytes were in the range from 0.5 to 10 μg/L. The proposed PT method was successfully applied to the determination of flavor compounds in commercially available beverages and fruit juices.
A simple preparation method for C60-fullerene bonded silica monolithic capillary by thermal coupling with a specific agent is reported. A silica monolithic capillary was modified with aminopropyl silane by a typical modification method, and then a thermal coupling agent, perfluorophenyl azide (PFPA) was covalently reacted with the amino groups. After C60-fullerene was covered over the PFPA-monolith at high density, thermal reaction was carried out at 170 C for 5 h, resulting C60-fullerene bonded silica monolith. We qualitatively confirmed all the modification process by elemental analysis and FT-IR, which showed the accomplishment of the all the reactions. According to liquid chromatographic evaluations, C60-fullerene bonded silica monolithic capillary provided the specific separation of polycyclic aromatic hydrocarbons (PAHs) by an effective π−πinteraction based on the bonded C60-fullerene. Even if we used non-polar solvent, n-hexane, as a mobile phase, the effective base-line separation of PAHs was achieved. Additionally, the capillary allowed the separation of aromatic couples, including benzene/anisole, dibenzosuberone/dibenzosuberenone, and chrysene/triphenylene, which have slightly structural differences, depending on the density of π electrons in these compounds.
To facilitate development of the androgen receptor inhibitor, enzalutamide, we developed and validated methods for simultaneous determination of enzalutamide, its carboxylic acid metabolite (M1), and N-desmethyl enzalutamide (M2) in plasma from mice, rats, and dogs, and brain from mice. Following addition of stable isotope-labeled internal standards, plasma and brain samples (0.05 and 0.3 mL, respectively) were mixed with 0.1% formic acid and extracted with tert-butyl methyl ether, and then injected into a liquid chromatography-tandem mass spectrometry system. The eluent was monitored in positive electrospray ionization mode. To bracket the concentrations of the drug and metabolites in study samples, calibration curves were constructed from 20 to 50000 ng/mL for plasma and 10 to 25000 ng/g for brain. Validation data demonstrated that these methods were selective, reproducible, and accurate. Using these methods, brain-to-plasma concentration ratios in mice were determined to be 0.72 for enzalutamide, 0.048 for M1, and 1.4 for M2.
Hydrophilic interaction liquid chromatography (HILIC) is advantageous for separation of polar compounds and sensitive detection in mass spectrometry. Sample dilution can negatively affect separation. We previously reported that separation efficiency in HILIC is affected by sample water content and perhaps the wash solution used in the injector sample loop. In this study, the effects of sample composition, sample volume, and wash solutions, on separation in HILIC was investigated using a mixture of catechol compounds including basic, acidic, and zwitterionic analogs. A ZIC-cHILIC column with a phosphorylcholine stationary phase was used for analysis. In the investigation of the effect of sample composition and volume, it was shown that separation efficiency was dependent on the amount of water injected into the column. Furthermore, we found that the most distorted peaks had similar elution times to water. We also investigated the effect of the sample loop wash solution, with separation efficiency reduced by increasing water content in the wash solutions. These results indicated that the water content of both sample and washing solutions should be kept to a minimum in HILIC separation.
To support therapeutic drug monitoring (TDM) a quantitative method for eight HIV protease inhibitors (PIs): nelfinavir, amprenavir, indinavir, saquinavir, atazanavir, darunavir, ritonavir and lopinavir in human plasma was developed. Separation of the analytes was achieved using core-shell column for UHPLC separation and mass spectrometric detection in fast multiple reaction monitoring (MRM) mode. Human plasma samples were collected on filter paper in the tube-based dried blood spot device and the analytes were extracted with methanol directly. Core-shell column and fast MRM allowed achieving an analysis time of six minutes. Using a plasma aliquot of 15 µL a dynamic linear range of 25-5000 ng/mL (ritonavir), 25-10000 ng/mL (nelfinavir and amprenavir) and 25-20000 ng/mL (indinavir, saquinavir, atazanavir, darunavir and lopinavir) could be achieved with precision and accuracies better than 15 %.
LC/MS analysis of triacylglycerols was carried out by cationization using Frit-FAB interface. Conventional triacylglycerol analysis utilizes m-NBA as the FAB matrix. However, by using methanol as the mobile phase for separating triacylglycerols, a glycerol matrix could also be applied. Through cationization in glycerol matrix containing 3 mM NaI, the molecular weight of triacylglycerol could be determined from [M+Na]+ ions, similar to using a m-NBA matrix. Furthermore, the fatty acid compositions of triacylglycerols could be determined from [M-(RCOO)]+ fragment ions. Additionally, dipalmitoyl-oleoyl-glycerol stereoisomers were separated with a C30 column by the slot model effect, the different mass spectra of 1,3-dipalmitoyl-2-oleoyl-sn-glycerol and 1,2-dipalmitoyl-3-oleoyl-rac-glycerol were determined.