Simultaneous and dynamic measurement of Schisandrol A changes in rat blood and brain and its comparative pharmacokinetic study in control and Parkinson's disease rats by dual-probe in vivo microdialysis.

Schisandrol A (SchA) is the main active ingredient of Schisandra chinensis (Turcz.) Baill., which is a famous traditional Chinese herbal medicine. SchA can penetrate the blood-brain barrier and has a significant neuroprotective effect. A group of multiplexed stable isotope mass tags (MSIMTs, m/z 332, 338, 346, 349, 351, 354, 360, 363, 374 and 377) were synthesized to perform multiplexed stable isotope labeling derivatization (MSILD) of SchA in rat microdialysates and standards. A new magnetic molecularly imprinted polymer was prepared using MSIMT-375-SchA as dummy template. All the 10-plexed derivatives of MSIMTs-SchA can be efficiently and selectively enriched and purified using this adsorbent by magnetic dispersive solid phase extraction (MDSPE) before ultra high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) analysis. It should be pointed out that the MSIMT-346-SchA standard derivative was used as internal standard in the process of MDSPE and UHPLC-MS/MS. On these bases, 9 different rat microdialysate samples can be determined by UHPLC-MS/MS in a single run. The utilization of MSIMTs significantly increased the sensitivity, accuracy, selectivity and analysis throughput. Under the optimized conditions, satisfactory linearity (R2> 0.987), limit of detection (LODs, 0.15-0.26 pg/mL) and lower limit of quantitative (LLOQ, 0.8-2.0 pg/mL) were obtained. Intra- and inter-day precisions were in the range of 2.2% -12.5%, and recoveries 94.2% -106.2%. The matrix effects were very low, and the average derivatization efficiency of 10-plex MSIMTs to SchA was as high as 97.8%. Using the developed dual-probe in vivo microdialysis sampling technique, the proposed analytical method has been applied for comparative pharmacokinetics of SchA in the brain and blood of control and Parkinson's disease (PD) rats.

Ratiometric fluorescence assay for sulfide ions with fluorescent MOF-based nanozyme.

A novel ratiometric fluorescence strategy for sulfide ions (S2-) analysis has been developed using metal-organic framework (MOF)-based nanozyme. NH2-Cu-MOF displays blue fluorescence (λem = 435 nm) originating from 2-amino-1,4-benzenedicarboxylic acid ligand. Besides, it possesses oxidase-like activity due to Cu2+ node, which can trigger chromogenic reaction. o-Phenylenediamine (OPD), as a common enzyme substrate, can be oxidized by NH2-Cu-MOF to form luminescent products (oxOPD) (λem = 570 nm). Inner filter effect occurs between oxOPD and MOF. Upon exposure to S2-, oxidase-like activity of MOF is depressed significantly because of the generation of CuS. On one hand, the amount of free Cu2+ decreases, affecting the yielding of oxOPD. On the other hand, CuNPs with larger size are obtained during the oxidation-reduction reaction between Cu2+ and OPD, which show weaker autocatalytic ability for OPD oxidation. These result in the decrease and increase of intensities at 570 and 435 nm, respectively. This method exhibits sensitive and selective responses towards S2- with LOD of 0.1 µM. Furthermore, such ratiometric strategy has been applied to detect S2- in food samples.

Ratiometric fluorescence detection of artemisinin based on photoluminescent Zn-MOF combined with hemin as catalyst.

Artemisinin (ART) is a type of frontline drug to treat drug-resistant falciparum malaria. Simple, accurate and selective determination of ART is significant to monitor its clinical pharmaceutical efficacy. Herein, a new ratiometric fluorescence method has been designed for the determination of ART with Zn-MOF as fluorescence reference and hemin as catalyst, respectively. Zn-MOF possesses intrinsic fluorescence at 443 nm owing to 2-aminoterephthalic acid ligand. When o-phenylenediamine (OPD) is mixed with hemin, a weak fluorescent signal at 570 nm ascribed to oxidized product of OPD (oxOPD) is observed. In the presence of ART, hemin can catalyze ART to break its peroxide bridge and release a large number of reactive oxygen species, which effectively oxidize OPD into luminescent oxOPD. Therefore, the fluorescence at 570 nm is enhanced significantly while the fluorescence of Zn-MOF remains basically unchanged. Thus, a ratiometric fluorescence sensing platform has been constructed for the detection of ART. This method exhibits wider linear range (0.15 µM-150 µM) with detection limit of 50 nM. This novel and selective method has been used to detect ART in compound naphthoquinone phosphate tablets.

Ratiometric fluorescence sensing of formaldehyde in food samples based on bifunctional MOF.

A new fluorescence strategy was described for ratiometric sensing of formaldehyde (FA) with bifunctional MOF, which acted as a fluorescence reporter as well as biomimetic peroxidase. With the assistance of H2O2, NH2-MIL-101 (Fe) catalyzes the oxidation of non-luminescent substrate o-phenylenediamine (OPD) to produce fluorescent product (oxOPD) with the maximum emission at 570 nm. Besides, intrinsic fluorescence of MOF (λem = 445 nm) was quenched by oxOPD through inner filter effect (IFE). However, FA and OPD reacted to generate Schiff bases, which competitively consumed OPD inhibiting the generation of oxOPD. Under the excitation wavelength of 375 nm, a ratiometric strategy was designed to detect FA with the fluorescence intensity ratio at 445 nm and 570 nm (F445/F570) as readout signal. This strategy exhibited a wide linear range (0.1-50 µM) and low detection limit of 0.03 µM. This method was confirmed for FA detection in food samples. In addition to establishing a new method to detect FA, this work will open new applications of MOF in food safety.

CoS2@TiO2 nanoarray: a heterostructured electrocatalyst for high-efficiency nitrate reduction to ammonia.

Electrochemical nitrate (NO3-) reduction emerges as a promising strategy to maintain the balance of the global nitrogen cycle and an alternative to nitrogen electroreduction for ambient ammonia (NH3) synthesis. However, the complicated multiple-electron transfer process of NO3--to-NH3 conversion demands catalysts with high selectivity for NH3 production. Herein, CoS2 nanoparticle decorated TiO2 nanobelt array on a titanium plate (CoS2@TiO2/TP) is reported as a superb electrocatalyst for the NO3- reduction reaction. In 0.1 M NaOH containing 0.1 M NO3-, CoS2@TiO2/TP offers a large NH3 yield of 538.21 µmol h-1 cm-2 at -0.7 V vs. reversible hydrogen electrode (RHE) and a high faradaic efficiency of 92.80% at -0.5 V vs. RHE. Additionally, it also shows strong stability for the 20 h electrolysis test.

Visible-Light-Initiated Multicomponent Reactions of α-Diazoesters to Access Organophosphorus Compounds.

A simple visible-light-initiated strategy has been established for the construction of organophosphorus compounds via aerobic multicomponent reaction of α-diazoesters, cyclic ethers, and P(O)H compounds under air. A number of phosphonates and phosphinates could be efficiently isolated in moderate to good yields without the use of photosensitizers and metal reagents. This multicomponent reaction has advantages of mild condition, simple operation, eco-friendly energy, good functional-group tolerance, and gram-scale synthesis.

Carbon quantum dots-functionalized silica stationary phase for pharmaceutical analysis by a green liquid chromatography mode.

Facing the trends of green chemistry, this work tries to find a novel material for per aqueous liquid chromatography (PALC) aiming to reduce the consumption of hazardous reagents. As a kind of green nanomaterials, the chromatographic performance of carbon quantum dots (CQDs) in PALC was rarely studied. Here, hydrophilic CQDs were prepared by a simple hydrothermal method using citric acid and ethylenediamine as carbon sources. The synthesized CQDs with functional groups of amino, carboxyl, and hydroxyl were decorated on silica gels forming a novel Si-CQDs stationary phase. This Si-CQDs column possesses the typical retention feature of PALC. Compounds with different polarities including hydrophobic pesticides, polar sulfonamides, ß-adrenoceptor blockers and agonists, as well as hydrophilic nucleosides and bases obtained satisfactory separation on this Si-CQDs column under PALC mode, even better resolution than in hydrophilic interaction liquid chromatography (HILIC) mode. A mixture of four sulfonamides can be separated within 6 min using a mobile phase containing only 5% acetonitrile, and the resolution achieves 2.39, 2.13, and 1.83 with an average column efficiency of 1400. For certain compounds, this Si-CQDs column showed better separation performance than commercial SiO2 column, NH2 column, and C18 column. The retention mechanism includes hydrophobic and electrostatic interactions due to the multifunctional groups of CQDs. This Si-CQDs column achieved the rapid detection of residual sulfonamides in milk with simplified sample pretreatment process and the detection of atenolol in commercial atenolol tablets. The developed Si-CQDs column has great prospects in low-cost and environmentally friendly separation and analysis.

Cascade amplification strategy combined with analyte-triggered fluorescence switching of dual-quenching system for highly sensitive detection of isoniazide.

A sensitive fluorescence sensing platform consisting of manganese dioxide nanosheets (MnO2) and gold nanoparticles (AuNPs) as dual nanoquenchers has been constructed to detect isoniazid combined with analyte-triggered cascade reactions. The fluorescence of 2,3-diaminophenazine (DAP) is quenched simultaneously by MnO2 and AuNPs via inner filter effect. MnO2 is decomposed by isoniazid to generate Mn2+, which makes AuNPs aggregated. The quenching abilities of both the decomposed MnO2 and aggregated AuNPs are inhibited, causing remarkable fluorescence recovery. The usage of dual nanoquenchers enhances the quenching efficiency and reduces the fluorescence background. Moreover, the isoniazid-triggered cascade reaction further amplifies the readout signal. Thus, this strategy exhibits higher sensitivity towards the detection of isoniazid. Compared with MnO2-based fluorescence assay, this strategy possesses lower limit of detection. This strategy has been successfully used to detect isoniazid in pharmaceutical preparations, which is of great significance for drug analysis.

A T-rich nucleic acid-enhanced electrochemical platform based on electroactive silver nanoclusters for miRNA detection.

DNA-templated silver nanoclusters (DNA/AgNCs) serve as a useful electrochemical sensing nanomaterial characterized by excellent electroactivity and good stability, while the effect of surrounding nucleotides on their electroactivity has not been studied. Herein, we validated a nucleotide-assisted enhancement mechanism of the DNA/AgNCs electroactivity caused by T-rich nucleic acid sequences in the vicinity of AgNCs. Based on the T-rich nucleic acid-enhanced AgNCs (NAE-AgNCs) combined with hybrid chain reaction (HCR), a novel signal-enhanced electrochemical biosensing platform was established for the ultrasensitive detection of miRNA. In the presence of target miRNA-155, HCR could be triggered to generate duplex strands containing both numerous AgNC synthesis sites and T-rich overhang strands upon the electrode. With the electrodeposition of adjacent AgNCs on the electrode, the larger oxidation potential of T-rich nucleic acid leaded to stronger electron-accepting capacity, which could contribute to increased current responses. The T-rich NAE HCR electrochemical strategy resulted in a detection limit of 0.39 fM for miRNA-155 detection, one order of magnitude lower than conventional HCR-based electrochemical sensors. This T-rich nucleic acid-assisted enhancement mechanism provided a new direction to construct highly sensitive, label-free, low-cost, and simple sensing platforms for applications in biomarker assays and clinic diagnosis.

Terbium (III)-based Metallacrowns with aggregation-induced emission feature coupled with cu (II) for fluorescence detection of cysteine.

The Metallacrowns (MCs) composed of repeated [Metal-N-O] subunits are a type of new material, but the MCs have not been developed and utilized in analytical applications. This essay reports on a new kind of terbium(III)-based Metallacrowns (Tb-MCs) with aggregation-induced emission (AIE) feature to build a sensing platform. It is first time that Tb-MCs are able to aggregate to larger aggregates in water along with a bright green emission, so that the property makes it possible to apply in biosensing. Thereafter, the AIE of Tb-MCs can be quenched effectively by Cu2+. Based on the high affinity of thiol to Cu2+, cysteine (Cys) recovers the fluorescence of Tb-MCs in the presence of Cu2+. There is a good linear range varying from 0.02 to 20 µM with a low limit of detection (LOD) 9.67 nM of Cys. In the end, this novel probe is also successfully applied to the determination of Cys in human serum with satisfactory results.

Co-crystallization and structure determination: An effective direction for anti-SARS-CoV-2 drug discovery.

Safer and more-effective drugs are urgently needed to counter infections with the highly pathogenic SARS-CoV-2, cause of the COVID-19 pandemic. Identification of efficient inhibitors to treat and prevent SARS-CoV-2 infection is a predominant focus. Encouragingly, using X-ray crystal structures of therapeutically relevant drug targets (PLpro, Mpro, RdRp, and S glycoprotein) offers a valuable direction for anti-SARS-CoV-2 drug discovery and lead optimization through direct visualization of interactions. Computational analyses based primarily on MMPBSA calculations have also been proposed for assessing the binding stability of biomolecular structures involving the ligand and receptor. In this study, we focused on state-of-the-art X-ray co-crystal structures of the abovementioned targets complexed with newly identified small-molecule inhibitors (natural products, FDA-approved drugs, candidate drugs, and their analogues) with the assistance of computational analyses to support the precision design and screening of anti-SARS-CoV-2 drugs.

[Recent advances in chemical derivatization-based chromatography-mass spectrometry methods for analysis of aldehyde biomarkers].

Human exposure to chemical pollutants in the environment can cause a variety of diseases, including cancer, diabetes, cardiovascular disease, and neurodegenerative diseases (Alzheimer's disease, Parkinson's disease, etc.). Exogenous and environmental pollutant exposure-induced endogenous aldehydes are highly reactive electrophilic compounds that can form covalently modified products with a variety of important biological molecules in the human body, thus inducing toxic effects. Exposome research has become a hotspot since it was first proposed in 2005. Exposure studies can map the complex relationships between biomarkers and disease risk. Therefore, the measurable and characteristic changes of all biomarkers together constitute a key basis for exposome research. Aldehydes are among the main components of chemical exposure. Because of the physical and chemical properties of aldehydes and the existence of multiple matrix interferences in the samples, it is particularly difficult to analyze and characterize them. The analysis and detection methods for aldehydes mainly include sensing analysis, electrochemical methods, fluorescence imaging, chromatography, mass spectrometry (MS), and chromatography-MS. Analytical techniques based on gas chromatography-MS (GC-MS) and liquid chromatography-MS (LC-MS) have emerged as the main methods for chemical exposome research. Chemical derivatization, especially stable isotope labeling derivatization (also known as chemical isotope labeling) combined with LC-MS analytical techniques, can help circumvent the problems encountered in targeted and non-targeted metabolome and exposome analysis. The combination of chemical derivatization with chromatography-MS is one of the most important solutions for the accurate analysis of aldehydes in complex samples. Over the past five years, the development and application of chromatography-MS analytical methods based on chemical derivatization have become key topics in aldehyde analysis. This paper summarizes and reviews the latest progress in GC-MS and LC-MS methods based on chemical derivatization (2015-2020). The review focuses on analytical method development for aldehyde exposure biomarkers in bio-matrices (blood, urine, saliva, biological tissue, etc.). Various derivatization reagents for labeling small-molecule aldehydes, qualitative/quantitative analytical methods and their application value, advantages/disadvantages of different analytical methods for aldehyde exposure biomarkers, and future development trends are also included. The manuscript contents may aid the integrated development of exposome, metabolomics, and lipidomics, as well as research on the environment, ecology, and health. To clarify the complex actions of exogenous and endogenous aldehydes in physiological and pathological events, it is necessary to improve the analysis and characterization techniques and tools for studying the "aldehydome." With the development and application of sophisticated mass spectrometers, advances in high-performance chromatographic separation and bioinformatics, and advent of single-cell analysis and MS imaging, future aldehyde exposome analytical methods will have higher sensitivity and throughput. This in turn would be more useful for screening and identifying unknown aldehyde compounds and discovering new exposome biomarkers.

MOF@MnO2nanocomposites prepared usingin situmethod and recyclable cholesterol oxidase-inorganic hybrid nanoflowers for cholesterol determination.

In this work, through thein situgrowth of MnO2nanosheets on the surface of terbium metal-organic frameworks (Tb-MOFs), MOF@MnO2nanocomposites are prepared and the fluorescence of Tb-MOFs is quenched significantly by MnO2. Additionally, the hybrid nanoflowers are self-assembled by cholesterol oxidase (ChOx) and copper phosphate (Cu3(PO4)2·3H2O). Then a new strategy for cholesterol determination is developed based on MOF@MnO2nanocomposites and hybrid nanoflowers. Cholesterol is oxidized under the catalysis of hybrid nanoflowers to yield H2O2, which further reduces MnO2nanosheets into Mn2+. Hence, the fluorescence recovery of Tb-MOFs is positively correlated to the concentration of cholesterol in the range of 10 to 360µM. The limit of detection (LOD) of cholesterol is 1.57µM. On the other hand, the hierarchical and confined structure of ChOx-inorganic hybrid nanoflowers greatly improve the stability of the enzyme. The activity of hybrid nanoflowers remains at a high level for one week when stored at room temperature. Moreover, the hybrid nanoflowers can be collected by centrifugation and reused. The activity of hybrid nanoflowers can continue at a high level for five cycles of determination. Therefore, it can be concluded that the hybrid nanoflowers are more stable and more economic than free enzymes, and they show a similar sensitivity and specificity to cholesterol compared with free ChOx. Finally, this strategy has been further validated for the determination of cholesterol in serum samples with satisfactory recoveries.

pH-modulated aggregation-induced emission of Au/Cu nanoclusters and its application to the determination of urea and dissolved ammonia.

A fluorescence platform is designed based on aggregation-induced emission of Au/Cu nanoclusters (Au/Cu NCs) driven by pH value. When pH increases from 6.0 to 7.0, Au/Cu NCs change from aggregation to dispersion, accompanied by the oxidation of Cu cores. Under the catalysis of urease, urea is hydrolysed to release ammonia, which further undergoes a hydrolysis reaction to produce OH-, causing the pH to increase. The fluorescence of Au/Cu NCs quenches linearly at 590 nm with the excitation wavelength at 320 nm when the concentration of urea varies from 5.0 to 100 µM. The limit of detection (LOD) and limit of quantification (LOQ) of urea are 2.23 and 7.45 µM, respectively. Combined with headspace single-drop microextraction technology, Au/Cu NCs are employed to monitor dissolved ammonia with low-cost and simple operation. The linear range of dissolved ammonia is from 20 to 300 µM. The LOD and LOQ of dissolved ammonia are 7.04 and 23.4 µM, respectively. The relative standard deviation (RSD) values of the intra-day and inter-day precision of urea are 2.4-3.0% and 3.0-3.7%, respectively, and those of dissolved ammonia are in the range 3.4-5.1% (intra-day precision) and 4.2-5.8% (inter-day precision). No interferences have been indentified in the determination of urea and dissolved ammonia. Finally, the proposed method has been applied to determine urea in human urine samples and dissolved ammonia in water samples with satisfactory results.Graphical abstract The pH increase produces the dispersion and decomposition of Au/Cu NCs, leading to the fluorescence quenching. Both urea and dissolved ammonia are detected successfully because they cause the pH change to alkaline.

Lead halide perovskites with aggregation-induced emission feature coupled with gold nanoparticles for fluorescence detection of heparin.

Herein, a new kind of lead halide perovskite (LHP, (C12H25NH3)2PbI4) with aggregation-induced emission (AIE) feature is developed as a fluorescent probe for heparin (Hep). The LHPs exhibit high emission when they aggregate in water. Interestingly, a few picomoles of dispersed gold nanoparticles (AuNPs) can quench the emission of LHPs, but the aggregated AuNPs are invalid. When protamine (Pro) is mixed with AuNPs at first, the negatively charged AuNPs aggregate through electrostatic interaction, producing the AIE recovery. Nevertheless, Hep disturbs the interaction between AuNPs and Pro due to its strong electrostatic interaction with Pro. Therefore, the dispersed AuNPs quench the fluorescence of LHPs again. A response linear range of Hep of 0.8-4.2 ng ml-1is obtained, and the detection limit is 0.29 ng ml-1. Compared with other probes for determination of Hep with AuNPs, this strategy exhibits better sensitivity due to the small quantity of AuNPs used. Finally, it is also successfully applied to detect Hep in human serum samples with satisfactory recoveries.

A fluorescence strategy for monitoring α-glucosidase activity and screening its inhibitors from Chinese herbal medicines based on Cu nanoclusters with aggregation-induced emission.

Herein, the self-assembly of 1-dodecanethiol-capped Cu nanoclusters (DT-Cu NCs) is obtained by annealing of dibenzyl ether solution of nanoclusters. These aggregates are composed of small clusters and emit a high level of aggregation-induced emission (AIE) in water. Based on the quenching effect of 4-nitrophenol (4-NP) on DT-Cu NCs, a fluorescence strategy is developed to monitor α-glucosidase (α-Glu) activity and screen its inhibitors from Chinese herbal medicines. 4-Nitrophenyl-α-D-glucopyranoside (NGP) is selected as the substrate, which is further hydrolyzed to yield 4-NP through the catalysis of α-Glu. The quenching efficiency is positively correlated to the concentration of α-Glu. Furthermore, the inhibitory effects of the extracts from four Chinese herbal medicines (i.e., the rind of Punica granatum L., Momordica grosvenorii Swingle., Crataegus pinnatifida Bge., and Lycium barbarum L.) on the α-Glu activity have been studied. The IC50 values of extracts from the rind of Punica granatum L. and Momordica grosvenorii Swingle are 0.23 and 0.37 g/L, respectively, so they show obvious inhibitory effects on α-Glu. The extracts of Crataegus pinnatifida Bge. and Lycium barbarum L. exhibit relatively weak inhibitory effects. Hence, the proposed strategy can be applicable for screening α-Glu inhibitors from Chinese herbal medicines. Last but not the least, by immobilizing DT-Cu NCs into agarose hydrogels in polyethylene tubes, a visual device is fabricated to screen α-Glu inhibitors with high throughput and sensitivity.

A ratiometric fluorescence assay for bleomycin based on dual-emissive chameleon DNA-templated silver nanoclusters.

The authors design dual-emissive DNA-templated silver nanoclusters (DNA-AgNCs) for ratiometric fluorescence sensing bleomycin (BLM) for the first time. A hairpin probe containing two different C-rich DNA templates at two terminals is used to synthesize chameleon DNA-AgNCs, which possess two emission peaks when they are in close proximity. A strong emission is founded at 622 nm (λex = 570 nm) while a weak one is located at 572 nm (λex = 504 nm). Meanwhile, the loop of this probe contains the scission site (5'-GC-3') of BLM. The loop can be cleaved into two parts by BLM-Fe(II) complex, inducing the two DNA-AgNCs away from each other. The fluorescence intensity at 572 nm and 622 nm increases and decreases, respectively. Such chameleon DNA-AgNCs exhibit an obvious fluorescence discoloration from orange to yellow. Therefore, a sensitive ratiometric fluorescent strategy for BLM detection has been proposed with the detection limit of 67 pM. Finally, this ratiometric method is used to detect BLM in serum samples.

Ce4+-triggered cascade reaction for ratiometric fluorescence detection of alendronate.

A ratiometric fluorescence assay for alendronate (ALDS) has been designed with Ce4+-triggered cascade chromogenic reaction. This strategy involves three processes: (1) Ce4+ oxidizes ascorbic acid (AA) into dehydroascorbic acid (DHAA), which then condenses with o-phenlenediamine (OPD) to generate fluorescent 3-(dihydroxyethyl)furo[3,4-b] quinoxaline-1-one (DFQ), presenting the maximum emission at 434 nm; (2) As oxidase-mimics, Ce4+ can oxidize OPD into fluorescent 2,3-diaminophenazine (DAP) which shows a strong emission at 568 nm; (3) ALDS inhibits the oxidation ability of Ce4+ towards OPD, thus inhibiting the generation of DAP. Accordingly, a homogeneous ratiometric fluorescence system with dual emission comes into being and the presence of ALDS can change the fluorescence intensity ratio obviously. With F434/F568 as readout, ALDS can be detected sensitively with the detection limit of 30 nM. Moreover, this ratiometric method was used to analyze ALDS in both human serum and pharmaceutical samples.

12-Plex UHPLC-MS/MS analysis of sarcosine in human urine using integrated principle of multiplex tags chemical isotope labeling and selective imprint enriching.

Urinary sarcosine was considered to be a potential biomarker for prostate cancer (Pca). In this work, an integrated strategy of multiplex tags chemical isotope labeling (MTCIL) combined with magnetic dispersive solid phase extraction (MDSPE), was proposed for specific extraction and high-throughput determination of sarcosine by ultra high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). In the past three months, we have developed 8-plex MTCIL reagents with excellent qualitative and quantitative performance. In this work, the multiplexing capacity of MTCIL reagents (MTCIL360/361/362/363/364/365/366/375/376/378/379/381) was increased 1.5-fold from 8-plex to 12-plex. MTCIL359 was prepared and used to label sarcosine standard as internal standard (IS). The structural analogue derivative (MTCIL373-sarcosine) of all targeted MTCIL-sarcosine derivatives was synthesized and used as a novel dummy template to prepare dummy magnetic molecularly imprinted polymers (DMMIPs). The integration of MTCIL and DMMIPs procedures were extremely favorable to excellent chromatographic separation and efficient mass spectrometric detection. The labeling efficiency, chromatographic retention and mass spectrometry responses of MTCIL reagents were consistent for sarcosine. In a single UHPLC-MS/MS run (2.0 min), this method can simultaneously quantify sarcosine in 12-plex urine samples and achieve unbiased concentrations comparison between different urine samples. Analytical parameters including linearity (R2 0.989-0.997), detection limits (0.02 nM), precision (2.6-11.5%), accuracy (96.1-107.4%), matrix effect, labeling and extraction efficiency were carefully validated. The proposed method was successfully applied for urinary sarcosine determination of healthy male individuals and Pca patients. It was found that the sarcosine concentrations in these two groups were statistically extremely significantly different (P < 0.001). The developed method was a powerful analytical tool to substantially promote the analysis throughput and large-scale experiments about the potential biomarker research.

13-Plex UHPLC-MS/MS Analysis of Hexanal and Heptanal Using Multiplex Tags Chemical Isotope Labeling Technology.

In this work, a new series of chemical isotope labeling reagents, levofloxacin-hydrazide-based mass tags (LHMTs) named as LHMT359/360/361/362/363/364/365/366/373/375/376/378/379/381 were first designed and synthesized for the high-throughput analysis of potential biomarkers containing hexanal and heptanal of lung cancer. We exploited a new core structure of levofloxacin-d3, which significantly enhanced the multiplexing capability. Among them, LHMT359 was used for labeling standard compounds as internal standards for quantification. Using LHMT373-heptanal as dummy template, dummy magnetic molecularly imprinted polymers (DMMIPs) were prepared for magnetic dispersive solid-phase extraction after derivatization procedure. Other 12 LHMTs were established for high-throughput labeling hexanal and heptanal in human serum samples. The presynthesized DMMIPs can selectively extract LHMTs-derivatives of hexanal and heptanal from equally mixed derivatization solutions. The enriched derivatives of hexanal and heptanal were quantified by ultrahigh-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). A single UHPLC-MS/MS run enabled simultaneously quantifying hexanal and heptanal from 12 serum samples only within 2 min. The limits of detection were all 0.5 pM for hexanal and heptanal. The accuracies from human serum samples ranged from -10.2% to +11.0% with the intra- and interday precisions less than 11.3%. Meanwhile, this method was successfully applied for the analysis of hexanal and heptanal in serum samples from healthy people and lung cancer patients. The results show that this method has the significant advantages of high sensitivity, accuracy, selectivity, and analysis-throughput. The method application indicates that the developed method is promising in the screening of suspected lung cancer patients.