Journal of Chromatography B (v.1007, #C)
Editorial Board (i).
Development of novel magnetic solid phase extraction materials based on Fe3O4/SiO2/poly(acrylamide-N,N′-methylene bisacrylamide)-Pluronic L64 composite microspheres and their application to the enrichment of natamycin by Miaomiao Tian; Yongcun Zou; Shaoyan Zhou; Tianpeng Wang; Xueju Lv; Qiong Jia (1-7).
Novel magnetic adsorbents based on Fe3O4/SiO2/poly(acrylamide-N,N'-methylene bisacrylamide) magnetic microspheres modified with non-ionic triblock copolymer surfactant were successfully prepared as a magnetic solid phase extraction adsorbent for the determination of trace natamycin in jam samples. The adsorbent was characterized by scanning electron microscopy, transmission electron microscopy, Fourier transformed infrared spectroscopy, vibrating sample magnetometer, and X-ray diffractometer analysis, confirming that Pluronic L64 was effectively functionalized on the magnetic materials. Various experimental parameters affecting the extraction capacity were investigated including adsorbent amount, extraction time, desorption time, sample pH, and ionic strength. For recovery evaluations, the jam samples were spiked at two concentration levels of 100 and 200 μg kg−1 of natamycin and the recovery values were in the range of 78.8–93.4%. The relative standard deviations (RSD) for the recoveries were less than 3.5%. The novel magnetic solid phase extraction method provided several advantages, such as simplicity, low environmental impact, convenient extraction procedure, and short analysis time when used for natamycin analysis.
Keywords: Magnetic solid phase extraction; Non-ionic triblock copolymers; Natamycin; Jam;
Enrichment and purification of total flavonoids from Cortex Juglandis Mandshuricae extracts and their suppressive effect on carbon tetrachloride-induced hepatic injury in Mice by Pan Zhao; Chao Qi; Gang Wang; Xinpeng Dai; Xiaohong Hou (8-17).
Display OmittedIn the present work, a simple and efficient chromatographic separation method was developed for preparative separation and enrichment of total flavonoids (TFs) from Cortex Juglandis Mandshuricae (CJM) extracts and then the protective effect of TFs against CCl4-induced acute liver injury in mice was investigated. Enrichment and purification of TFs from CJM extracts were studied using six macroporous resins and HPD-750 resin was selected as the best resin according to its adsorption and desorption properties. The operating parameters of resin column chromatography were optimized. Under the optimal conditions, TFs from CJM with purity larger than 50% were produced and their antioxidant activity was further evaluated in vitro. The mice were orally administrated with the purified TFs for seven days and then given CCl4 (0.3%, 10 mL/kg i.p.). The results showed that TFs of CJM significantly attenuated the activities of serum aspartate transaminase (AST) and alanine transaminase (ALT) compared with model group, as well as the relative liver weight. Histopathological observation also revealed that TFs reduced the incidence of liver lesions and improved hepatocyte abnormality. Moreover, oral administration of TFs significantly enhanced antioxidant enzyme activities (superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px)) and decreased the content of malondialdehyde (MDA). Histopathological and biochemical results elicited that TFs of CJM had significant hepatoprotective activity comparable to the standard silymarin. This is the first time to reveal the protective actions of the TFs from CJM against CCl4-induced liver damage in mice and this natural product should be developed as a new drug for treatment of live injury in future.
Keywords: Cortex Juglandis Mandshuricae; Total flavonoids; Enrichment; Cabon tetrachloride; Hepatoprotective effect;
Quantitative subcellular study of doxorubicin in MCF-7/Adr cells using liquid chromatography–tandem mass spectrometry by Wenzhuan Ma; Jinling Wang; Qiang Guo; Pengfei Tu (18-22).
A rapid, sensitive and selective high-performance liquid chromatography–tandem mass spectrometric (LC–MS/MS) method has been developed and validated for the determination of doxorubicin in intracellular compartments using glibenclamide as internal standard (IS). MCF-7/Adr cancer cells (1 × 106) were incubated with doxorubicin (8 μg/mL) for 0.5, 1, 2 and 4 h and then subjected to sequential extraction of cytosolic, membrane/organelle, nuclear and cytoskeleton soluble protein. Samples were extracted using protein precipitation with methanol. Chromatographic separation was carried out on a C18 column with acetonitrile and 0.1% formic acid water as mobile phase and with gradient elution at a flow rate of 0.2 mL/min. The method was linear over the range of 1–300 ng/mL with a lower limit of quantification (LLOQ) of 1 ng/mL. The distribution of doxorubicin in subcellular components of MCF-7/Adr cancer cells was mainly in nucleic protein fraction.
Keywords: Doxorubicin; LC–MS/MS; MCF-7/Adr cells; Intracellular distribution;
Separation of vitexin-4″-O-glucoside and vitexin-2″-O-rhamnoside from hawthorn leaves extracts using macroporous resins by Hongjuan Li; Ying Liu; Haizhu Jin; Sujing Liu; Shengtao Fang; Chunhua Wang; Chuanhai Xia (23-29).
Vitexin-4″-O-glucoside and vitexin-2″-O-rhamnoside are the major flavonoids of hawthorn leaves. In this work, the adsorption and desorption characteristics of vitexin-4″-O-glucoside and vitexin-2″-O-rhamnoside on seven macroporous resins were evaluated. Among the tested resins, the HPD-400 resin showed the best adsorption and desorption capacities. Adsorption isotherms were constructed for the HPD-400 resin and well fitted to Langmuir and Freundlich models. Dynamic adsorption and desorption tests were performed on column packed with the HPD-400 resin to optimize the chromatographic parameters. After one run treatment with the HPD-400 resin, the contents of vitexin-4″-O-glucoside and vitexin-2″-O-rhamnoside in the product were increased 8.44-fold and 8.43-fold from 0.720% and 2.63% to 6.08% and 22.2% with recovery yields of 79.1% and 81.2%, respectively. These results show that the developed method is a promising basis for the large-scale purification of vitexin-4″-O-glucoside and vitexin-2″-O-rhamnoside from hawthorn leaves and other plant materials.
Keywords: Macroporous resins; Vitexin-4″-O-glucoside; Vitexin-2″-O-rhamnoside; Purification; Hawthorn leaves;
Simultaneous quantification of acetaminophen and five acetaminophen metabolites in human plasma and urine by high-performance liquid chromatography–electrospray ionization–tandem mass spectrometry: Method validation and application to a neonatal pharmacokinetic study by Sarah F. Cook; Amber D. King; John N. van den Anker; Diana G. Wilkins (30-42).
Drug metabolism plays a key role in acetaminophen (paracetamol)-induced hepatotoxicity, and quantification of acetaminophen metabolites provides critical information about factors influencing susceptibility to acetaminophen-induced hepatotoxicity in clinical and experimental settings. The aims of this study were to develop, validate, and apply high-performance liquid chromatography–electrospray ionization–tandem mass spectrometry (HPLC–ESI–MS/MS) methods for simultaneous quantification of acetaminophen, acetaminophen-glucuronide, acetaminophen-sulfate, acetaminophen-glutathione, acetaminophen-cysteine, and acetaminophen-N-acetylcysteine in small volumes of human plasma and urine. In the reported procedures, acetaminophen-d4 and acetaminophen-d3-sulfate were utilized as internal standards (IS). Analytes and IS were recovered from human plasma (10 μL) by protein precipitation with acetonitrile. Human urine (10 μL) was prepared by fortification with IS followed only by sample dilution. Calibration concentration ranges were tailored to literature values for each analyte in each biological matrix. Prepared samples from plasma and urine were analyzed under the same HPLC–ESI–MS/MS conditions, and chromatographic separation was achieved through use of an Agilent Poroshell 120 EC-C18 column with a 20-min run time per injected sample. The analytes could be accurately and precisely quantified over 2.0–3.5 orders of magnitude. Across both matrices, mean intra- and inter-assay accuracies ranged from 85% to 112%, and intra- and inter-assay imprecision did not exceed 15%. Validation experiments included tests for specificity, recovery and ionization efficiency, inter-individual variability in matrix effects, stock solution stability, and sample stability under a variety of storage and handling conditions (room temperature, freezer, freeze-thaw, and post-preparative). The utility and suitability of the reported procedures were illustrated by analysis of pharmacokinetic samples collected from neonates receiving intravenous acetaminophen.
Keywords: Acetaminophen/Paracetamol; Metabolites; LC–MS/MS; Human plasma; Human urine;
Single-step microwave assisted headspace liquid-phase microextraction of trihalomethanes and haloketones in biological samples by Abdulnaser Alsharaa; Chanbasha Basheer; Muhammad Sajid (43-48).
A single-step microwave assisted headspace liquid-phase microextraction (MA-HS-LPME) method was developed for determination of trihalomethanes (THMs) and haloketones (HKs) in biological samples. In this method, a porous membrane envelope was filled with few microliters of extraction solvent and then placed inside the microwave extraction vial. A PTFE ring was designed to support the membrane envelope over a certain height inside the vial. An optimum amount of biological sample was placed in the vial equipped with magnetic stirrer. After that nitric acid was added to the vial for digestion of biological sample. The sample was digested and the volatile THMs and HKs were extracted at headspace in the solvent containing porous membrane. After simultaneous digestion and extraction, the extract was injected to gas chromatography/mass spectrometry for analysis. Factors affecting the extraction efficiency were optimized to achieve higher extraction performance. Quantification was carried out over a concentration range of 0.3–100 ng g−1 for brominated compounds while for the chlorinated ones linear range was between 0.5–100 ng g−1. Limit of detections (LODs) were ranged from 0.051 to 0.110 ng g−1 while limit of quantification (LOQ) were in the range of 0.175–0.351 ng g−1. The relative standard deviations (RSDs) of the calibrations were ranged between 1.1 and 6.8%. The MA-HS-LPME was applied for the determination of trace level THMs and HKs in fish tissue and green alga samples.
Keywords: Disinfection by-products; Microwave assisted headspace liquid-phase microextraction; Environmental applications; Gas chromatography–; mass spectrometry;
Determination of a novel phosphodiesterase-4 inhibitor chlorbipram in mouse plasma and brain by UFLC–MS/MS: Application in pharmacokinetic studies after intravenous administration by Yiwen Li; Yufang Cheng; Bingqing Zeng; Bo Niu; Haibiao Guo; Guoliang Li; Hongfang Feng; Jiangping Xu; Xuemei Yang (49-53).
In this study, we evaluated a simple and sensitive method for determination of a novel phosphodiesterase-4 (PDE4) inhibitor, chlorbipram, in mouse plasma and brain using ultra-fast liquid chromatography-tandem mass spectrometry (UFLC–MS/MS). Separation was achieved using an Acquity UPLC BEH C18 column (50 mm × 2.1 mm, particle size 1.7 μm) with a gradient mobile phase consisting of water and methanol at a flow rate of 0.25 ml/min. Detection was performed in the multiple reaction monitoring (MRM) mode using electrospray ionization (ESI) in the positive ion mode. The liquid–liquid extraction method with ethyl acetate was used for both pretreatment of plasma and brain homogenates. The calibration curves of chlorbipram showed good linearity over the concentration range of 0.5–200 ng/ml (R 2 > 0.994) for mouse plasma and over the range of 0.25–100 ng/ml (R 2 > 0.994) for mouse brain homogenate. The extraction recovery was in the range of 78.3–84.8% for chlorbipram and the internal standard (IS) ZXI14 in two different biological matrices. The intra- and inter-day precision values were less than 13.0% and the accuracy ranged from 97.8% to 106.0% for quality control samples. No noteworthy matrix effects and instability were observed for chlorbipram. This validated method was successfully applied to a pharmacokinetic study of chlorbipram in mice after intravenous administration. The results show that this novel drug crosses the blood-brain barrier and provides the basis for further studies on chlorbipram.
Keywords: Phosphodiesterase-4 inhibitor; Chlorbipram; UFLC–MS/MS; Pharmacokinetics;
Analysis of the chemical constituents and rats metabolites after oral administration of Nauclea officinalis by ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry by Fenxia Zhu; Jiaquan Chen; Hui Wang; Xiaobin Jia; Shuxia Wang; Zhiyuan Zhang; Xiaoting Zhai; Jindi Xu; Wei Tan; Qing Ning; Junfei Gu (54-66).
Nauclea officinalis has long been used in China for the treatment of cold, fever, swelling of throat, pink eyes, and so on; however, the in vivo integrated metabolism of its multiple bioactive components remains unknown. In this paper, an ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-QTOF/MS) method was established to identify chemical constituents in N. officinalis and metabolites in rat biological fluids after oral administration of N. officinalis. First, 40 chemical constituents in N. officinalis were detected within 19 min by UPLC-QTOF/MS. Among them, 18 alkaloids and 7 phenolic acids and iridoids were identified or tentatively characterized. Secondly, 22 metabolites were identified after oral administration of N. officinalis extract, including 3, 9, 6 and 4 metabolites in the plasma, feces, urine and bile samples, respectively. Finally, the metabolic pathway was proposed, which were the hydroxylation, the hydroxylation of deglycosyation product of parent compound, the hydroxylation and dehydrogenation product of parent compound, and acetylation. Among these, hydroxylation was considered as the main metabolic processes. This work suggests that the integrative metabolism approach makes a useful template for drug metabolism research of traditional Chinese medicine (TCM).
Keywords: UPLC-QTOF/MS; Chemical compounds; Metabolite identification;
Gas chromatographic separation of fatty acid esters of cholesterol and phytosterols on an ionic liquid capillary column by Simon Hammann; Walter Vetter (67-71).
Steryl esters are high molecular weight compounds (600–700 g/mol) regularly present as a minor lipid class in animal and plant lipids. Different sterol backbones (e.g., cholesterol, β-sitosterol and brassicasterol) which can be esterified with various fatty acids can result in highly complex steryl ester patterns in food samples. The gas chromatographic (GC) analysis of intact steryl esters is challenging, since high elution temperatures are required for their elution. On nonpolar GC phases, steryl esters with fatty acids with differing degree of unsaturation (e.g., oleate and linoleate) cannot be separated and there are only few polar columns available with sufficient temperature stability. In this study, we used gas chromatography with mass spectrometry (GC/MS) and analyzed intact steryl esters on a commercial room temperature ionic liquid (RTIL) column which was shortened to a length of 12 m. The column separated the steryl esters both by total carbon number and by degree of unsaturation of the fatty acid. For instance, cholesteryl esters with stearic acid (18:0), oleic acid (18:1n-9), linoleic acid (18:2n-6) and α-linolenic acid (18:3n-3) could be resolved (R ≥ 1.3) from each other. By analysis of synthesized standard substances, the elution orders for different steryl backbones and different fatty acids on a given sterol backbone could be determined. Analysis of spreads and plant oils allowed to determine retention times for 37 steryl esters, although a few co-elutions were observed. The ionic liquid column proved to be well-suited for the analysis of intact steryl esters.
Keywords: Steryl ester; Ionic liquid; Gas chromatography; Mass spectrometry;
Profile of bioactive compounds from grape pomace (Vitis vinifera and Vitis labrusca) by spectrophotometric, chromatographic and spectral analyses by L.F. Ribeiro; R.H. Ribani; T.M.G. Francisco; A.A. Soares; R. Pontarolo; C.W.I. Haminiuk (72-80).
The aim of this study was to characterize grape pomace (GP) from winemaking byproducts of different grape samples (Cabernet Sauvignon—CS; Merlot—ME; Mix composed of 65% Bordeaux, 25% Isabel and 10% BRS Violet—MI and Terci—TE) with a view to exploiting its potential as a source of bioactive compounds and an alternative to the reuse of waste. Bioactive compounds such as individual phenolic compounds and polyunsaturated fatty acids (PUFA) were identified and quantified by spectrophotometric, chromatographic and spectral analyses. The sample of MI had the highest concentrations for total phenolic compounds and total flavonoids, while TE had the highest content for total monomeric anthocyanins. For all samples it was possible to identify 13 different anthocyanins by high performance liquid chromatography (HPLC) and mass spectrometry (MS). Moreover, the GP samples showed phenolic acids; flavan-3-ols such as catechin; flavonols such as quercetin, rutin and kaempferol; and stilbenes such as trans-resveratrol. Therefore, grape pomace can be considered a source for the recovery of phenolic compounds having antioxidant activity as well as a rich source of PUFA. Thus it can be used as an ingredient in the development of new food products, since it is suitable for human consumption, and a viable alternative both to adding nutritional value to food and to reduce environmental contamination.
Keywords: Phenolic compounds; Anthocyanins; Flavonoids; Fatty acids; Infrared reflectance;
Highly sensitive method for simultaneous determination of nine alkaloids of Shuanghua Baihe tablets in human plasma by LC–MS/MS and its application by Yao Wu; Ruijuan Liu; Pan Gu; Minlu Cheng; Lu Zheng; Yujie Liu; Pengcheng Ma; Li Ding (81-92).
Shuanghua Baihe tablets (SBT) is a traditional Chinese medicinal formula which has been used to treat recurrent aphthous stomatitis for many years. To study the pharmacokinetic profiles of berberine, epiberberine, coptisine, palmatine, jatrorrhizine, magnoflorine, berberrubine, corynoline and acetylcorynoline in human after administration of SBT, a sensitive liquid chromatography–tandem mass spectrometry method was developed and fully validated for the simultaneous quantification of these nine alkaloids in human plasma. After protein precipitation, the nine alkaloids in human plasma sample was separated on a Hanbon C18 (150 mm × 2.1 mm, 5 μm) column with gradient elution using methanol and 0.5% formic acid water solution, and detected by a triple quadrupole mass spectrometer with an electrospray ionization source. It is a challenge to design different calibration ranges for different analytes in a bioanalytical method for simultaneous determination of multi-analytes in bio-samples. To ensure that each alkaloid in the plasma was determined accurately by the simultaneous quantitation method, the upper limits of quantification for the nine alkaloids were designed at 100, 300, 800, 1800 and 5000 pg/mL, respectively, according to the maximum plasma concentration value of each alkaloid obtained from the pilot pharmacokinetic study. The lower limit of quantification was 15 pg/mL for berberine, epiberberine, coptisine, magnoflorine, berberrubine, corynoline and acetylcorynoline, while for palmatine and jatrorrhizine it was 1.5 pg/mL. This method was successfully applied to investigate the pharmacokinetic profiles of the nine alkaloids in healthy Chinese volunteers after a single oral administration of SBT.
Keywords: Shuanghua Baihe tablets; LC–MS/MS; Berberine; Palmatine; Coptisine; Corynoline;
Separation of E. coli chaperonin groEL from β-galactosidase without denaturation by Sudheer K. Molugu; Jihui Li; Ricardo A. Bernal (93-99).
Chaperonins are a class of ubiquitous proteins that assist and accelerate protein folding in the cell. The Escherichia coli groEL is the best known and forms a complex with its co-chaperonin groES in the presence of ATP and assists in the folding of nascent and misfolded substrate proteins. The purification of recombinant groEL results in a nearly homogeneous sample that consistently co-purifies with the major contaminant E. coli β-galactosidase. Removal of β-galactosidase using column chromatography alone is exceedingly difficult. This is due to the fact that the overall size, surface charge, isoelectric point and hydrophobicity of groEL and β-galactosidase are very similar. Therefore purification of groEL chaperonin to homogeneity requires denaturation of the complex into monomers with urea for separating the groEL from contaminating β-galactosidase followed by reassembly of the chaperonin complex.Here, we present a simple procedure for separating β-galactosidase along with many other impurities from groEL preparations under non-denaturing conditions. The groEL is first salted out with 50% ammonium sulfate. This step also precipitates β-galactosidase but this is then salted out by the addition of magnesium chloride which leaves groEL in solution. All remaining contaminants are removed by column chromatography.
Keywords: GroEL; β-Galactosidase; Chaperonin purification; Protein salting out;
One-pot preparation of a novel monolith for high performance liquid chromatography applications by Xiaoyan Jiao; Shigang Shen; Tiesheng Shi (100-109).
Various novel porous organic-based monoliths with the mode of hydrophobicity were synthesized by in situ free-radical crosslinking copolymerization and optimized for the separations of small molecules and high-performance reversed-phase chromatography (RP-chromatography). These monoliths contained co-polymers based on glycidyl methacrylate (GMA)/ethylene glycol dimethacrylate (EDMA)/tripropylene glycol diacrylate (TPGDA) or EDMA/TPGDA. A mixture of cetanol, methanol and poly (ethylene glycol) (PEG) was used as the porogen, with the ratio of these solvents being varied along with the polymerization temperature to generate a library of monoliths. The conditions were optimized and the resulting poly (GMA-co-TPGDA-co-EDMA) monolith was investigated by infrared spectrometer (IR), field emission scanning electron microscope (FESEM), and mercury intrusion porosimetry (MIP), respectively. The column performance was assessed by the separation of a series of neutral solutes of benzene derivatives. The result demonstrated that the prepared monolith exhibited an RP-chromatographic behavior and relatively homogeneous structure, good permeability and separation performance. Moreover, the relative standard deviations (RSDs) of the retention factor values for benzene derivatives were less than 1.5% (n = 7, column-to-column). The approach used in this study was extended to the separation of anilines.
Keywords: Organic polymeric monolith; Tripropylene glycol diacrylate; Mixed-mode polymer monolith; High performance liquid chromatography (HPLC); Small molecules;
Effects of berberine and pomegranate seed oil on plasma phospholipid metabolites associated with risks of type 2 diabetes mellitus by U-HPLC/Q-TOF-MS by Xia Wu; Yan Li; Qiu Wang; Weimin Li; Yifan Feng (110-120).
A rapid and reliable ultra-performance liquid chromatography coupled with electrospray ionization/quadrupole-time-of-flight mass spectrometry (U-HPLC/Q-TOF-MS) has been firstly used to analyze the changes of plasma phospholipids, in type 2 diabetes mellitus (T2DM) mice after administration of berberine and pomegranate seed oil (PSO). The separation of plasma phospholipids was carried out on an Acquity U-HPLC BEH C18 column (2.1 mm × 50 mm, 1.7 μm, Waters) by linear gradient elution using a mobile phase consisting of 10 mM ammonium formate in water and acetonitrile: isopropanol (1:1, v/v) mixed solution added by 0.25% water and 10 mM ammonium formate. The method demonstrated a good precision and reproducibility. Linear regression analysis showed a good linearity. And potential biomarkers were discovered based on their mass spectra and chemometrics methods. The results demonstrated that the proposed U-HPLC/Q-TOF-MS method was successfully applied to analyze the dynamic changes of phospholipids components in plasma of T2DM mice after drug treatment and could provide a useful data base for meriting further study in humans and investigating pharmacological actions of drugs.
Keywords: Berberine; Biomarkers; Plasma phospholipids; Pomegranate seed oil; U-HPLC/Q-TOF-MS;
Combined analysis of N′-nitrosonornicotine and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol in the urine of cigarette smokers and e-cigarette users by Delshanee Kotandeniya; Steven G. Carmella; Makenzie E. Pillsbury; Stephen S. Hecht (121-126).
A liquid chromatography–electrospray ionization–tandem mass spectrometry (HPLC–ESI+–MS/MS) method for the analysis of the tobacco-specific carcinogens N′-nitrosonornicotine (NNN) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL) and their glucuronides (total NNN and total NNAL) in human urine was developed. The method has excellent accuracy and intra-day and inter-day precision, and limits of quantitation of 0.015 and 0.075 pmol/mL urine, respectively, for total NNN and total NNAL. A unique aspect of this method is internal assessment of possible artifactual formation of NNN by inclusion of the monitor amine [pyridine-D4]nornicotine. We found that artifactual formation of NNN comprised only 2.5% of the measured amounts of total NNN in urine of cigarette smokers, under our conditions using ammonium sulfamate as an inhibitor of nitrosation. The method was applied to urine samples from cigarette smokers and e-cigarette users. Levels of total NNN and total NNAL in the urine of cigarette smokers averaged 0.060 ± 0.035 pmol/mL and 2.41 ± 1.41 pmol/mL urine, (N = 38), respectively, which were both significantly greater than in the urine of 27 e-cigarette users.
Keywords: Combined analysis; N′-Nitrosonornicotine; NNAL; 4-(Methylnitrosamino)-1-(3-pyridyl)-1-butanol; Smokers’ urine; e-cigarette users’ urine; Liquid chromatography–electrospray ionization–tandem mass spectrometry;
Synthesis of monodisperse molecularly imprinted microspheres with multi-recognition ability via precipitation polymerization for the selective extraction of cyromazine, melamine, triamterene and trimethoprim by Xian-Hua Wang; Li-Fu Xie; Qian Dong; Hao-Long Liu; Yan-Ping Huang; Zhao-Sheng Liu (127-131).
Through precipitation polymerization, three monodisperse molecularly imprinted polymers (MIPs) containing imprints of 2,4-diamino-6-methyl-1,3,5-triazine (DM), cyromazine (CY) or trimethoprim (TM), were synthesized using methacrylic acid as functional monomer, divinylbenzene as cross-linker, and a mixture of acetonitrile–toluene (90/10, v/v) as porogen. The morphology and selectivity of the MIPs were characterized and compared systematically. The MIPs had the best specific binding in pure acetonitrile, and the data of adsorption experiment were fitted well with Langmuir and Freundlich model. In addition, DM-MIPs showed the excellent binding and multi-recognition capability for CY, melamine (ME), triamterene (TA) and TM, and the binding capacity were 7.18, 7.56, 5.66 and 5.45 μmol/g, respectively. Due to the pseudo template and the ability of multi-recognition, DM-MIPs as sorbent material could avoid the effect of template leakage on quantitative analysis. Therefore, DM-MIPs were used as a solid-phase extraction material to enrich ME, CY, TA and TM from different bio-matrix samples for high performance liquid chromatography analysis. Under the optimized conditions, the recoveries of three spiked levels in different bio-matrix samples were ranged from 80.9% to 91.5% with RSD ≤ 4.2 (n = 3).
Keywords: Molecularly imprinted polymers; Precipitation polymerization; Pseudo template; Multi-recognition;
Bioanalytical validation for simultaneous quantification of non-aromatic steroids in follicular fluid from cattle via ESI-LC–MS/MS by Martin Kunze; Elisa Wirthgen; Christina Walz; Marion Spitschak; Julia Brenmoehl; Jens Vanselow; Manfred Schwerin; Klaus Wimmers; Andreas Hoeflich (132-139).
The family of steroid hormones is quite attractive for the approach of phenotype monitoring in farm animals. Therefore, we developed a new protocol for the quantitative analysis of natural steroids in follicular fluid from dairy cows. The corresponding steroid profile, which consists of progesterone, corticosterone, hydrocortisone, testosterone, and androstenedione covering three distinct steroid classes, was determined by LC/MS. Quantification is achieved by use of steroid standards diluted in steroid-free follicular fluid as calibrators. Thus, the new protocol does not require deuterated standards. In order to correct for conditional performance of the analytical system we have used dexamethasone as an internal standard. The method was validated according to EMA guidelines. Within- and between-day variations were below 20% for most parameters assessed. All steroids assessed had lower limits of quantification in the range of 2.1 to 4.4 ng/ml. We have established a simple and sensitive analytical system in order to step towards a broader and cost-efficient phenotyping analysis in follicular fluid from dairy cows.
Keywords: Steroid hormones; ESI-LC–MS/MS; Quantification; Validation; Cattle; Follicular fluid; Phenotyping;
Metabolomic study on the faecal extracts of atherosclerosis mice and its application in a Traditional Chinese Medicine by Feng Tian; Lei Gu; Aiyong Si; Quanbao Yao; Xianwei Zhang; Jihui Zhao; Daode Hu (140-148).
The intestinal microbiota and their metabolites are closely related to the formation of atherosclerosis (AS). In this study, a metabolomic approach based on the reversed-phase liquid chromatography/quadrupole time-of-flight mass spectrometry (LC-Q-TOF-MS) platform was established to analyze the metabolic profiling of fecal extracts from AS mice model. The established metabolomic platform was also used for clearing the effective mechanism of a Traditional Chinese Medicine (TCM) named Sishen granule (SSKL). Totally, sixteen potential biomarkers in faeces of AS mice were identified and 5 of them could be reversed by SSKL. Through functional analysis of these biomarkers and the established network, lipid metabolism, cholesterol metabolism, energy cycle, and inflammation reaction were considered as the most relevant pathological changes in gastrointestinal tract of AS mice. The metabolomic study not only revealed the potential biomarkers in AS mice’ faeces but also supplied a systematic view of the pathological changes in gastrointestinal metabolite in AS mice. This metabolomic study also demonstrated that SSKL had the therapeutic effectiveness on AS through partly reversing the lipid metabolism, inflammation and energy metabolism.
Keywords: Metabolomic; Faecal extracts; Atherosclerosis; Traditional Chinese Medicine;