Improved phosphometabolome profiling applying isotope dilution strategy and capillary ion chromatography-tandem mass spectrometry

Publication date: Available online 6 February 2018
Source:Journal of Chromatography B
Author(s): Marit H. Stafsnes, Lisa M. Røst, Per Bruheim
The phosphometabolome is comprised of all phosphorylated metabolites including the major metabolite classes sugar phosphates and nucleoside phosphates. Phosphometabolites are invaluable in any cell as a part of primary- and energy- metabolism, and as building blocks in the biosynthesis of macromolecules. Here, we report quantitative profiling of the phosphometabolome by applying capillary ion chromatography-tandem mass spectrometry (capIC-MS/MS), ensuring improved chromatographic separation, robustness and quantitative precision. Baseline separation was achieved for six out of eight tested hexose phosphates. Quantitative precision and reproducibility was improved by introducing a fully uniformly (U) 13C–labeled biological extract and applying an isotope dilution (ID) correction strategy. A 13C–labeled biological extract does in principle contain internal standards (IS) for all metabolites, but low abundant metabolites pose a challenge, and solutions to this are discussed. The extreme reproducibility and reliability of this capIC-MS/MS method was demonstrated by running the instrumentation continuously for ten days.

Publication date: Available online 6 February 2018
Source:Journal of Chromatography B

Author(s): Marit H. Stafsnes, Lisa M. Røst, Per Bruheim

The phosphometabolome is comprised of all phosphorylated metabolites including the major metabolite classes sugar phosphates and nucleoside phosphates. Phosphometabolites are invaluable in any cell as a part of primary- and energy- metabolism, and as building blocks in the biosynthesis of macromolecules. Here, we report quantitative profiling of the phosphometabolome by applying capillary ion chromatography-tandem mass spectrometry (capIC-MS/MS), ensuring improved chromatographic separation, robustness and quantitative precision. Baseline separation was achieved for six out of eight tested hexose phosphates. Quantitative precision and reproducibility was improved by introducing a fully uniformly (U) 13C–labeled biological extract and applying an isotope dilution (ID) correction strategy. A 13C–labeled biological extract does in principle contain internal standards (IS) for all metabolites, but low abundant metabolites pose a challenge, and solutions to this are discussed. The extreme reproducibility and reliability of this capIC-MS/MS method was demonstrated by running the instrumentation continuously for ten days.





A modified multiscale peak alignment method combined with trilinear decomposition to study the volatile/heat-labile components in Ligusticum chuanxiong Hort – Cyperus rotundus rhizomes by HS-SPME-GC/MS

Publication date: 15 March 2018 Source:Journal of Chromatography B, Volume 1079 Author(s): Min He, Pan Yan, Zhi-Yu Yang, Zhi-Min Zhang, Tian-Biao Yang, Liang Hong Head Space/Solid Phase Micro-Extraction (HS-SPME) coupled wi…

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079

Author(s): Min He, Pan Yan, Zhi-Yu Yang, Zhi-Min Zhang, Tian-Biao Yang, Liang Hong

Head Space/Solid Phase Micro-Extraction (HS-SPME) coupled with Gas Chromatography/Mass Spectrometer (GC/MS) was used to determine the volatile/heat-labile components in Ligusticum chuanxiong Hort - Cyperus rotundus rhizomes. Facing co-eluting peaks in k samples, a trilinear structure was reconstructed to obtain the second-order advantage. The retention time (RT) shift with multi-channel detection signals for different samples has been vital in maintaining the trilinear structure, thus a modified multiscale peak alignment (mMSPA) method was proposed in this paper. The peak position and peak width of representative ion profile were firstly detected by mMSPA using Continuous Wavelet Transform with Haar wavelet as the mother wavelet (Haar CWT). Then, the raw shift was confirmed by Fast Fourier Transform (FFT) cross correlation calculation. To obtain the optimal shift, Haar CWT was again used to detect the subtle deviations and be amalgamated in calculation. Here, to ensure there is no peaks shape alternation, the alignment was performed in local domains of data matrices, and all data points in the peak zone were moved via linear interpolation in non-peak parts. Finally, chemical components of interest in Ligusticum chuanxiong Hort - Cyperus rotundus rhizomes were analyzed by HS-SPME-GCMS and mMSPA-alternating trilinear decomposition (ATLD) resolution. As a result, the concentration variation between herbs and their pharmaceutical products can provide a scientific basic for the quality standard establishment of traditional Chinese medicines.





Simultaneous determination of Vitamin A, 25-hydroxyl vitamin D3 α-tocopherol in small biological fluids by liquid chromatography-tandem mass spectrometry

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079
Author(s): Haifeng Zhang, Li Quan, Pei Pei, Ye Lin, Chao Feng, Hongyan Guan, Fang Wang, Ting Zhang, Jianxin Wu, Junsheng Huo
A sensitive method for the simultaneous analysis of vitamin A (VA), 25-hydroxyl vitamin D3 (25-OH VD3) and α-tocopherol (VE) in children plasma by liquid chromatography-tandem mass spectrometry (LC–MS/MS) has been developed and validated. Sample preparation chose the solid phase extraction. 100 μL of plasma was mixed with 300 μL ethanol contained 4 μL isotope-labelled analytes. After a series operation, the supernatant was applied to the solid phase extraction (SPE) plate (HLB μElution plate). The eluate was evaporated, and reconstituted in 100 μL methanol. And then, 6 μL reconstituted sample was injected into LC–MS/MS. Quantitative analysis was carried out by multiple reaction monitoring (MRM) with a positive mode electrospray (ESi + ). Separations of VA, 25-OH VD3 and VE were performed on an Acquity UPLC reversed-phase Phenyl-Hexyl analytical column (CSH, 2.1 × 50 mm, 1.7 μm). Gradient elution was used at a flow rate of 0.4 mL/min with a mobile phase consisting of 0.1% formic acid solution and 0.1% formic acid, 5 mM ammonium formate in acetonitrile. The total time of analysis was 10 min. The method had a limit of quantification (LOQ) of 10.03, 1.20, and 0.04 ng/mL for VA, 25-OH VD3 and VE in methanol, respectively. The linear calibration curves were fitted over the range of 0.14–14.32 μg/mL, 1.80–180.29 ng/mL, and 6.03–602.99 ng/mL for VA, 25-OH VD3 and VE in methanol. The correlation coefficients were greater than 0.998 for all analytes. The recoveries for all analytes were between 80 and 120% with the inter- and intra-day precisions (presented as relative standard deviation, RSD%) less than 10.0%. Analysis of VA, 25-OH VD3 and VE in recurrent respiratory tract infection children plasma and anemic infants’ fingertip blood was then carried out using this method and statistical analysis of the data with statistic package for social science 20.0 (SPSS 20.0). Using this method, multiple fat-soluble vitamins could be detected at the same time. Solid phase extraction was used to simplify sample pretreatment. μElution plate used here could reduce the sample volume, only 100 μL sample was used in this method, and 6 μL reconstituted sample was injected into LC–MS/MS. This makes the method appropriate for larger sample pretreatment, and suitable for children, especially infants and newborns’ sample detection, in whom the circulation blood was low.

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079

Author(s): Haifeng Zhang, Li Quan, Pei Pei, Ye Lin, Chao Feng, Hongyan Guan, Fang Wang, Ting Zhang, Jianxin Wu, Junsheng Huo

A sensitive method for the simultaneous analysis of vitamin A (VA), 25-hydroxyl vitamin D3 (25-OH VD3) and α-tocopherol (VE) in children plasma by liquid chromatography-tandem mass spectrometry (LC–MS/MS) has been developed and validated. Sample preparation chose the solid phase extraction. 100 μL of plasma was mixed with 300 μL ethanol contained 4 μL isotope-labelled analytes. After a series operation, the supernatant was applied to the solid phase extraction (SPE) plate (HLB μElution plate). The eluate was evaporated, and reconstituted in 100 μL methanol. And then, 6 μL reconstituted sample was injected into LC–MS/MS. Quantitative analysis was carried out by multiple reaction monitoring (MRM) with a positive mode electrospray (ESi + ). Separations of VA, 25-OH VD3 and VE were performed on an Acquity UPLC reversed-phase Phenyl-Hexyl analytical column (CSH, 2.1 × 50 mm, 1.7 μm). Gradient elution was used at a flow rate of 0.4 mL/min with a mobile phase consisting of 0.1% formic acid solution and 0.1% formic acid, 5 mM ammonium formate in acetonitrile. The total time of analysis was 10 min. The method had a limit of quantification (LOQ) of 10.03, 1.20, and 0.04 ng/mL for VA, 25-OH VD3 and VE in methanol, respectively. The linear calibration curves were fitted over the range of 0.14–14.32 μg/mL, 1.80–180.29 ng/mL, and 6.03–602.99 ng/mL for VA, 25-OH VD3 and VE in methanol. The correlation coefficients were greater than 0.998 for all analytes. The recoveries for all analytes were between 80 and 120% with the inter- and intra-day precisions (presented as relative standard deviation, RSD%) less than 10.0%. Analysis of VA, 25-OH VD3 and VE in recurrent respiratory tract infection children plasma and anemic infants’ fingertip blood was then carried out using this method and statistical analysis of the data with statistic package for social science 20.0 (SPSS 20.0). Using this method, multiple fat-soluble vitamins could be detected at the same time. Solid phase extraction was used to simplify sample pretreatment. μElution plate used here could reduce the sample volume, only 100 μL sample was used in this method, and 6 μL reconstituted sample was injected into LC–MS/MS. This makes the method appropriate for larger sample pretreatment, and suitable for children, especially infants and newborns’ sample detection, in whom the circulation blood was low.





The metabolism of a natural product mogroside V, in healthy and type 2 diabetic rats

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079
Author(s): Guisheng Zhou, Yulong Zhang, Yang Li, Mengyue Wang, Xiaobo Li
Mogroside V, a natural compound isolated from the fruits of Siraitia grosvenorii (Swingle), is a promising candidate for anti-diabetic activity. The present study aims to develop a simple and practical strategy for comparing the in vivo metabolite profiling of mogroside V in healthy and type 2 diabetic (T2D) model rats. In this paper, a highly sensitive and rapid ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) with MetaboLynx™ software combined with mass defect filtering (MDF) method was established and successfully applied to detect and identify the metabolites in plasma, urine, bile and feces samples of healthy and model rats administrated with mogroside V. The distribution of metabolites in plasma, bile, urine and feces of healthy and model rats had obvious differences. A total of 23 metabolites were observed in healthy rats while 26 metabolites were detected in model rats. The results indicated that dehydrogenation, deoxidation, oxidation and isomerization were the major metabolic transformations of mogroside V. Additionally, it was noticed that the peak areas of metabolites in T2D rat plasma samples were much larger than those of metabolites in healthy rat plasma sample, whereas in T2D rat urine samples they were remarkably smaller compared with healthy rat urine sample. These high blood concentrations of metabolites might be beneficial for the treatment of T2D. The results of this study are valuable and important in understanding the metabolic process and therapeutic mechanism of mogroside V.

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079

Author(s): Guisheng Zhou, Yulong Zhang, Yang Li, Mengyue Wang, Xiaobo Li

Mogroside V, a natural compound isolated from the fruits of Siraitia grosvenorii (Swingle), is a promising candidate for anti-diabetic activity. The present study aims to develop a simple and practical strategy for comparing the in vivo metabolite profiling of mogroside V in healthy and type 2 diabetic (T2D) model rats. In this paper, a highly sensitive and rapid ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) with MetaboLynx™ software combined with mass defect filtering (MDF) method was established and successfully applied to detect and identify the metabolites in plasma, urine, bile and feces samples of healthy and model rats administrated with mogroside V. The distribution of metabolites in plasma, bile, urine and feces of healthy and model rats had obvious differences. A total of 23 metabolites were observed in healthy rats while 26 metabolites were detected in model rats. The results indicated that dehydrogenation, deoxidation, oxidation and isomerization were the major metabolic transformations of mogroside V. Additionally, it was noticed that the peak areas of metabolites in T2D rat plasma samples were much larger than those of metabolites in healthy rat plasma sample, whereas in T2D rat urine samples they were remarkably smaller compared with healthy rat urine sample. These high blood concentrations of metabolites might be beneficial for the treatment of T2D. The results of this study are valuable and important in understanding the metabolic process and therapeutic mechanism of mogroside V.





Pharmacokinetic comparisons of six components from raw and vinegar-processed Daphne genkwa aqueous extracts following oral administration in rats by employing UHPLC–MS/MS approaches

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079
Author(s): Yi Tao, Dandan Su, Weidong Li, Baochang Cai
A sensitive UHPLC–MS/MS approach was developed and validated for the quantification of genkwanin, 3′-hydroxygenkwanin, apigenin, luteolin, yuanhuacine and genkwadaphnin in biological samples after oral administration of raw and vinegar-processed Daphne genkwa. Liquiritin and glycyrrhetinic acid were employed as internal standards. Six components were extracted by using protein precipitation with acetonitrile. Chromatographic separation was achieved on a Waters BEH C18 column (50 mm × 2.1 mm, 1.7 μm) by using a mobile phase composed of water (containing 0.1% formic acid) and acetonitrile. Mass spectrometric detection was conducted using multiple reaction monitoring (MRM). The intra- and inter-day precisions of the six analytes were below 4.87%, and the accuracies were within ±5.0%. The extraction recoveries of the six constituents were determined between 97.5% and 105.4% and the matrix effects ranged from 97.3% to 103.7%. All the samples showed satisfactory precision and accuracy after various stability tests. The established approach was successfully applied to the comparative pharmacokinetic study. Compared to the raw group, the parameters of Cmax and AUC0−t of genkwanin, 3′-hydroxygenkwanin, apigenin and luteolin elevated remarkably (p < 0.05) after oral delivery of vinegar-processed Daphne genkwa while the parameters of Cmax and AUC0−t of yuanhuacine and genkwadaphnin decreased significantly (p < 0.05). The results revealed that vinegar-processing could enhance bioavailability of genkwanin, 3′-hydroxygenkwanin, apigenin and luteolin but reduce the bioavailability of yuanhuacine and genkwadaphnin.

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079

Author(s): Yi Tao, Dandan Su, Weidong Li, Baochang Cai

A sensitive UHPLC–MS/MS approach was developed and validated for the quantification of genkwanin, 3′-hydroxygenkwanin, apigenin, luteolin, yuanhuacine and genkwadaphnin in biological samples after oral administration of raw and vinegar-processed Daphne genkwa. Liquiritin and glycyrrhetinic acid were employed as internal standards. Six components were extracted by using protein precipitation with acetonitrile. Chromatographic separation was achieved on a Waters BEH C18 column (50 mm × 2.1 mm, 1.7 μm) by using a mobile phase composed of water (containing 0.1% formic acid) and acetonitrile. Mass spectrometric detection was conducted using multiple reaction monitoring (MRM). The intra- and inter-day precisions of the six analytes were below 4.87%, and the accuracies were within ±5.0%. The extraction recoveries of the six constituents were determined between 97.5% and 105.4% and the matrix effects ranged from 97.3% to 103.7%. All the samples showed satisfactory precision and accuracy after various stability tests. The established approach was successfully applied to the comparative pharmacokinetic study. Compared to the raw group, the parameters of C max and AUC0−t of genkwanin, 3′-hydroxygenkwanin, apigenin and luteolin elevated remarkably (p<0.05) after oral delivery of vinegar-processed Daphne genkwa while the parameters of C max and AUC0−t of yuanhuacine and genkwadaphnin decreased significantly (p < 0.05). The results revealed that vinegar-processing could enhance bioavailability of genkwanin, 3′-hydroxygenkwanin, apigenin and luteolin but reduce the bioavailability of yuanhuacine and genkwadaphnin.





Systematic evaluation of matrix effects in supercritical fluid chromatography versus liquid chromatography coupled to mass spectrometry for biological samples

Publication date: 15 March 2018 Source:Journal of Chromatography B, Volume 1079 Author(s): Vincent Desfontaine, Francesca Capetti, Raul Nicoli, Tiia Kuuranne, Jean-Luc Veuthey, Davy Guillarme Matrix effects (ME) is acknowle…

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079

Author(s): Vincent Desfontaine, Francesca Capetti, Raul Nicoli, Tiia Kuuranne, Jean-Luc Veuthey, Davy Guillarme

Matrix effects (ME) is acknowledged as being one of the major drawbacks of quantitative bioanalytical methods, involving the use of liquid chromatography coupled to mass spectrometry (LC-MS). In the present study, the incidence of ME in SFC-MS/MS and LC-MS/MS in the positive mode electrospray ionization (ESI+) was systematically compared for the analysis of urine and plasma samples using two representative sets of 40 doping agents and 38 pharmaceutical compounds, respectively. Three different SFC stationary phase chemistries were employed, to highlight the importance of the column in terms of selectivity. Biological samples were prepared using two different sample treatments, including a non-selective sample clean-up procedure (dilute and shoot (DS) and protein precipitation (PP) for urine and plasma samples, respectively) and a selective sample preparation, namely solid phase extraction (SPE) for both matrices. The lower susceptibility to ME in SFC vs. reversed phase LC (RPLC) was verified in all the experiments performed on urine, and especially when a simple DS procedure was applied. Also, with the latter, the performance strongly varied according to the selected SFC stationary phase, whereas the results were quite similar with the three SFC columns, in the case of SPE clean-up. The same trend was observed with plasma samples. Indeed, with the PP procedure, the occurrence of ME was different on the three SFC columns, and only the 2-picolylamine stationary phase chemistry displayed lower incidence of ME compared to LC-MS/MS. On the contrary, when a SPE clean-up was carried out, the results were similar to the urine samples, with higher performance of SFC vs. LC and limited discrepancies between the three SFC columns. The type of ME observed in LC-MS/MS was generally a signal enhancement and an ion suppression for urine and plasma samples, respectively. In the case of SFC-MS/MS, the type of ME randomly varied according to the analyzed matrix, selected column and sample treatment.





Headspace solid-phase microextraction (HS-SPME) combined with GC–MS as a process analytical technology (PAT) tool for monitoring the cultivation of C. tetani

Publication date: Available online 6 February 2018
Source:Journal of Chromatography B
Author(s): Masoud Ghader, Nader Shokoufi, Ali Es-haghi, Kazem Kargosha
Vaccine production is a biological process in which variation in time and output is inevitable. Thus, the application of Process Analytical Technologies (PAT) will be important in this regard. Headspace solid – phase microextraction (HS-SPME) coupled with GC–MS can be used as a PAT for process monitoring. This method is suitable to chemical profiling of volatile organic compounds (VOCs) emitted from microorganisms. Tetanus is a lethal disease caused by Clostridium tetani (C. tetani) bacterium and vaccination is an ultimate way to prevent this disease. In this paper, SPME fiber was used for the investigation of VOCs emerging from C. tetani during cultivation. Different types of VOCs such as sulfur-containing compounds were identified and some of them were selected as biomarkers for bioreactor monitoring during vaccine production. In the second step, the portable dynamic air sampling (PDAS) device was used as an interface for sampling VOCs by SPME fibers. The sampling procedure was optimized by face-centered central composite design (FC-CCD). The optimized sampling time and inlet gas flow rates were 10 min and 2 m L s−1, respectively. PDAS was mounted in exhausted gas line of bioreactor and 42 samples of VOCs were prepared by SPME fibers in 7 days during incubation. Simultaneously, pH and optical density (OD) were evaluated to cultivation process which showed good correlations with the identified VOCs (>80%). This method could be used for VOCs sampling from off-gas of a bioreactor to monitoring of the cultivation process.

Publication date: Available online 6 February 2018
Source:Journal of Chromatography B

Author(s): Masoud Ghader, Nader Shokoufi, Ali Es-haghi, Kazem Kargosha

Vaccine production is a biological process in which variation in time and output is inevitable. Thus, the application of Process Analytical Technologies (PAT) will be important in this regard. Headspace solid - phase microextraction (HS-SPME) coupled with GC–MS can be used as a PAT for process monitoring. This method is suitable to chemical profiling of volatile organic compounds (VOCs) emitted from microorganisms. Tetanus is a lethal disease caused by Clostridium tetani (C. tetani) bacterium and vaccination is an ultimate way to prevent this disease. In this paper, SPME fiber was used for the investigation of VOCs emerging from C. tetani during cultivation. Different types of VOCs such as sulfur-containing compounds were identified and some of them were selected as biomarkers for bioreactor monitoring during vaccine production. In the second step, the portable dynamic air sampling (PDAS) device was used as an interface for sampling VOCs by SPME fibers. The sampling procedure was optimized by face-centered central composite design (FC-CCD). The optimized sampling time and inlet gas flow rates were 10 min and 2 m L s−1, respectively. PDAS was mounted in exhausted gas line of bioreactor and 42 samples of VOCs were prepared by SPME fibers in 7 days during incubation. Simultaneously, pH and optical density (OD) were evaluated to cultivation process which showed good correlations with the identified VOCs (>80%). This method could be used for VOCs sampling from off-gas of a bioreactor to monitoring of the cultivation process.





Metabolic profiling of Gegenqinlian decoction in rat plasma, urine, bile and feces after oral administration by ultra high performance liquid chromatography coupled with Fourier transform ion cyclotron resonance mass spectrometry

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079
Author(s): Ting Liu, Xiumin Tian, Zhaoqin Li, Fei Han, Bin Ji, Yunli Zhao, Zhiguo Yu
Gegenqinlian decoction (GQLD) as a traditional Chinese medical (TCM) prescription is made from four Chinese herbs and has been used for centuries to treat pyrexia and diarrhea. In this work, an ultra-high performance liquid chromatography coupled with Fourier transform ion cyclotron resonance mass spectrometry (UHPLC-FT-ICR-MS) method was established to profile the metabolic fate of GQLD in rat plasma, urine, bile and feces. The analysis was performed on a Waters Acquity UPLC HSS T3 column (2.1 mm × 100 mm, 1.8 μm) at 35 °C within 25 min. A total of 174 compounds mainly including flavonoids, alkaloids and triterpenoids in rat plasma, urine, bile and feces after oral administration of GQLD were identified or tentatively characterized by comparison with retention times, accurate mass within 5 ppm error and their characteristic MS fragmentation ions. Among compounds in rats, 107 compounds are prototypes and 67 compounds are metabolites. Results demonstrated that metabolic pathways of GQLD in rats included methylation, demethylation, hydrogenation, hydrolysis, hydroxylation, methoxylation, sulfate conjugation and glucuronide conjugation. A clear understanding of the absorption and metabolism of GQLD is very important in its rational clinical use and pharmacological research. These findings enhance our understanding of the metabolism and effective forms of GQLD.

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079

Author(s): Ting Liu, Xiumin Tian, Zhaoqin Li, Fei Han, Bin Ji, Yunli Zhao, Zhiguo Yu

Gegenqinlian decoction (GQLD) as a traditional Chinese medical (TCM) prescription is made from four Chinese herbs and has been used for centuries to treat pyrexia and diarrhea. In this work, an ultra-high performance liquid chromatography coupled with Fourier transform ion cyclotron resonance mass spectrometry (UHPLC-FT-ICR-MS) method was established to profile the metabolic fate of GQLD in rat plasma, urine, bile and feces. The analysis was performed on a Waters Acquity UPLC HSS T3 column (2.1 mm × 100 mm, 1.8 μm) at 35 °C within 25 min. A total of 174 compounds mainly including flavonoids, alkaloids and triterpenoids in rat plasma, urine, bile and feces after oral administration of GQLD were identified or tentatively characterized by comparison with retention times, accurate mass within 5 ppm error and their characteristic MS fragmentation ions. Among compounds in rats, 107 compounds are prototypes and 67 compounds are metabolites. Results demonstrated that metabolic pathways of GQLD in rats included methylation, demethylation, hydrogenation, hydrolysis, hydroxylation, methoxylation, sulfate conjugation and glucuronide conjugation. A clear understanding of the absorption and metabolism of GQLD is very important in its rational clinical use and pharmacological research. These findings enhance our understanding of the metabolism and effective forms of GQLD.





Metal-organic framework UiO-66 for rapid dispersive solid phase extraction of neonicotinoid insecticides in water samples

Publication date: 1 March 2018
Source:Journal of Chromatography B, Volumes 1077–1078
Author(s): Xiaolin Cao, Zejun Jiang, Shanshan Wang, Sihui Hong, Hui Li, Chao Zhang, Yong Shao, Yongxin She, Fen Jin, Maojun Jin, Jing Wang
UIO-66 crystals were explored for the first time to adsorb neonicotinoid insecticides in environmental water samples. HPLC coupled with tandem MS was used for quantification and determination of neonicotinoid insecticides. UiO-66 crystals was successfully synthesized by a simple constant-temperature bath method. Synthesized UiO-66 was characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM), thermogravimetry and nitrogen adsorption porosimetry (NAP), which demonstrated a uniform particle size, a large Brunauer–Emmett–Teller (BET) surface area and high thermostability. The adsorbing results showed that UIO-66 crystals could be used as a promising adsorbents for rapid extraction of neonicotinoid insecticides and be reused at least 10 times. Under the optimized conditions, the limits of detection (LODs, S/N = 3) and limits of quantification (LOQs, S/N = 10) for the five insecticides were found to be 0.02–0.4 ng/mL and 0.05–1.0 ng/mL, respectively. This developed approach not only provided more simple and sensitive method, as well as possessing satisfactory recovery for neonicotinoid insecticides, but also for other traces in environmental samples.

Publication date: 1 March 2018
Source:Journal of Chromatography B, Volumes 1077–1078

Author(s): Xiaolin Cao, Zejun Jiang, Shanshan Wang, Sihui Hong, Hui Li, Chao Zhang, Yong Shao, Yongxin She, Fen Jin, Maojun Jin, Jing Wang

UIO-66 crystals were explored for the first time to adsorb neonicotinoid insecticides in environmental water samples. HPLC coupled with tandem MS was used for quantification and determination of neonicotinoid insecticides. UiO-66 crystals was successfully synthesized by a simple constant-temperature bath method. Synthesized UiO-66 was characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM), thermogravimetry and nitrogen adsorption porosimetry (NAP), which demonstrated a uniform particle size, a large Brunauer–Emmett–Teller (BET) surface area and high thermostability. The adsorbing results showed that UIO-66 crystals could be used as a promising adsorbents for rapid extraction of neonicotinoid insecticides and be reused at least 10 times. Under the optimized conditions, the limits of detection (LODs, S/N = 3) and limits of quantification (LOQs, S/N = 10) for the five insecticides were found to be 0.02–0.4 ng/mL and 0.05–1.0 ng/mL, respectively. This developed approach not only provided more simple and sensitive method, as well as possessing satisfactory recovery for neonicotinoid insecticides, but also for other traces in environmental samples.





Chinese patent medicine Xin-Ke-Shu inhibits Ca2+ overload and dysfunction of fatty acid β-oxidation in rats with myocardial infarction induced by LAD ligation

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079
Author(s): Yong Yang, Hongmei Jia, Meng Yu, Chao Zhou, Lili Sun, Yang Zhao, Hongwu Zhang, Zhongmei Zou
Myocardial infarction (MI) occurs during a sustained insufficient blood supply to the heart, eventually leading to myocardial necrosis. Xin-Ke-Shu tablet (XKS) is a prescription herbal compound and a patented medicine extensively used in the clinical treatment of coronary heart disease (CHD). To understand the molecular mechanism of the XKS action against MI in detail, it is necessary to investigate the altered metabolome and related pathways coincident with clinical features. In this study, tissue-targeted metabonomics based on ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) were developed to explore the metabolic changes associated with XKS treatment in the heart tissue of rats with MI induced by a left anterior descending coronary artery ligation (LAD). The metabolic disorder induced by LAD was alleviated after low-dose XKS (LD) and intermediate-dose XKS (MD) treatment. XKS modulated six perturbed metabolic pathways. Among them, inhibition of Ca2+ overload and dysfunction of fatty acid β-oxidation-related metabolic pathways likely underlie the therapeutic effects of XKS against MI. In agreement with its observed effect on metabolite perturbation, XKS reversed the over-expression of the four key proteins, long-chain acyl-CoA synthetase 1 (ACSL1), carnitine palmitoyl transferase-1 (CPT1B), calcium/calmodulin-dependent kinase II (CaMKII), and phospholipase A2IIA (PLA2IIA). Both metabolite and protein changes suggested that XKS exerts its therapeutic effect on metabolic perturbations in LAD-induced MI mainly by inhibiting the Ca2+ overload and fatty acid β-oxidation dysfunction.

Publication date: 15 March 2018
Source:Journal of Chromatography B, Volume 1079

Author(s): Yong Yang, Hongmei Jia, Meng Yu, Chao Zhou, Lili Sun, Yang Zhao, Hongwu Zhang, Zhongmei Zou

Myocardial infarction (MI) occurs during a sustained insufficient blood supply to the heart, eventually leading to myocardial necrosis. Xin-Ke-Shu tablet (XKS) is a prescription herbal compound and a patented medicine extensively used in the clinical treatment of coronary heart disease (CHD). To understand the molecular mechanism of the XKS action against MI in detail, it is necessary to investigate the altered metabolome and related pathways coincident with clinical features. In this study, tissue-targeted metabonomics based on ultra-high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF/MS) were developed to explore the metabolic changes associated with XKS treatment in the heart tissue of rats with MI induced by a left anterior descending coronary artery ligation (LAD). The metabolic disorder induced by LAD was alleviated after low-dose XKS (LD) and intermediate-dose XKS (MD) treatment. XKS modulated six perturbed metabolic pathways. Among them, inhibition of Ca2+ overload and dysfunction of fatty acid β-oxidation-related metabolic pathways likely underlie the therapeutic effects of XKS against MI. In agreement with its observed effect on metabolite perturbation, XKS reversed the over-expression of the four key proteins, long-chain acyl-CoA synthetase 1 (ACSL1), carnitine palmitoyl transferase-1 (CPT1B), calcium/calmodulin-dependent kinase II (CaMKII), and phospholipase A2IIA (PLA2IIA). Both metabolite and protein changes suggested that XKS exerts its therapeutic effect on metabolic perturbations in LAD-induced MI mainly by inhibiting the Ca2+ overload and fatty acid β-oxidation dysfunction.





Enantiomeric separation and quantification of R/S-amphetamine in urine by ultra-high performance supercritical fluid chromatography tandem mass spectrometry

Publication date: 1 March 2018
Source:Journal of Chromatography B, Volumes 1077–1078
Author(s): S. Hegstad, H. Havnen, A. Helland, O. Spigset, J. Frost
To distinguish between legal and illegal consumption of amphetamine reliable analytical methods for chiral separation of the R- and S-enantiomers of amphetamine in biological specimens are required. In this regard, supercritical fluid chromatography (SFC) has several potential advantages over liquid chromatography, including rapid separation of enantiomers due to low viscosity and high diffusivity of supercritical carbon dioxide, the main component in the SFC mobile phase. A method for enantiomeric separation and quantification of R- and S-amphetamine in urine was developed and validated using ultra-high performance supercritical fluid chromatography-tandem mass spectrometry (UHPSFC-MS/MS). Sample preparation prior to UHPSFC-MS/MS analysis was a semi-automatic solid phase extraction method. The UHPSFC-MS/MS method used a Chiralpak AD-3 column with a mobile phase consisting of CO2 and 0.2% cyclohexylamine in 2-propanol. The injection volume was 2 μL and run-time was 6 min. MS/MS detection was performed with positive electrospray ionization and two multiple reaction monitoring transitions (m/z 136.1 > 119.0 and m/z 136.1 > 91.0). The calibration range was 50–10,000 ng/mL for each enantiomer. The between-assay relative standard deviations were in the range of 3.7–7.6%. Recovery was 92–93% and matrix effects ranged from 100 to 104% corrected with internal standard. After development and validation, the method has been successfully implemented in routine use at our laboratory for both separation and quantification of R/S-amphetamine, and has proved to be a reliable and useful tool for distinguishing intake of R- and S-amphetamine in authentic patient samples.

Publication date: 1 March 2018
Source:Journal of Chromatography B, Volumes 1077–1078

Author(s): S. Hegstad, H. Havnen, A. Helland, O. Spigset, J. Frost

To distinguish between legal and illegal consumption of amphetamine reliable analytical methods for chiral separation of the R- and S-enantiomers of amphetamine in biological specimens are required. In this regard, supercritical fluid chromatography (SFC) has several potential advantages over liquid chromatography, including rapid separation of enantiomers due to low viscosity and high diffusivity of supercritical carbon dioxide, the main component in the SFC mobile phase. A method for enantiomeric separation and quantification of R- and S-amphetamine in urine was developed and validated using ultra-high performance supercritical fluid chromatography-tandem mass spectrometry (UHPSFC-MS/MS). Sample preparation prior to UHPSFC-MS/MS analysis was a semi-automatic solid phase extraction method. The UHPSFC-MS/MS method used a Chiralpak AD-3 column with a mobile phase consisting of CO2 and 0.2% cyclohexylamine in 2-propanol. The injection volume was 2 μL and run-time was 6 min. MS/MS detection was performed with positive electrospray ionization and two multiple reaction monitoring transitions (m/z 136.1 > 119.0 and m/z 136.1 > 91.0). The calibration range was 50–10,000 ng/mL for each enantiomer. The between-assay relative standard deviations were in the range of 3.7–7.6%. Recovery was 92–93% and matrix effects ranged from 100 to 104% corrected with internal standard. After development and validation, the method has been successfully implemented in routine use at our laboratory for both separation and quantification of R/S-amphetamine, and has proved to be a reliable and useful tool for distinguishing intake of R- and S-amphetamine in authentic patient samples.





Simultaneous determination of eight cyclopolypeptide antibiotics in feed by high performance liquid chromatography coupled with evaporation light scattering detection

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076
Author(s): Xuqin Song, Jingmeng Xie, Meiyu Zhang, Yingxia Zhang, Jiufeng Li, Qiwen Huang, Limin He
A high throughput, reliable and reproducible analysis strategy based on high performance liquid chromatography combined to evaporative light scattering detector (HPLC-ELSD) was developed for simultaneous determination of eight cyclopolypeptide antibiotics including vancomycin, polymyxin B (polymyxin B1 and polymyxin B2), polymyxin E (colistin A and colistin B), teicoplanin, bacitracin A, daptomycin and virginiamycin M1 in animal Feed. Feed samples were extracted with methanol-2% formic acid aqueous solution, followed by a solid-phase extraction step using a HLB cartridge. Under the optimum chromatographic conditions and ELSD parameters, target compounds were separated well on a short column filled with biphenyl stationary phase. The method was developed in accordance with pig complete feed and then extended to detect polypeptide antibiotics in piglet premix, pig feed additive, poultry complete feed and fattening pig premix. The results showed that logarithmic calibration curves of eight analytes were linear (r2 > 0.99) within the concentration range of 5–200 mg mL−1. The developed method provided good accuracy and precision for quantification of eight polypeptides in five kinds of feeds with recoveries ranging from 72.0% to 105.4% with relative standard deviations <9.5%. The limits of detection ranged from 2 to 5 mg kg−1. Finally, the method was successfully applied to analyze polypeptide antibiotics in commercial feed.

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076

Author(s): Xuqin Song, Jingmeng Xie, Meiyu Zhang, Yingxia Zhang, Jiufeng Li, Qiwen Huang, Limin He

A high throughput, reliable and reproducible analysis strategy based on high performance liquid chromatography combined to evaporative light scattering detector (HPLC-ELSD) was developed for simultaneous determination of eight cyclopolypeptide antibiotics including vancomycin, polymyxin B (polymyxin B1 and polymyxin B2), polymyxin E (colistin A and colistin B), teicoplanin, bacitracin A, daptomycin and virginiamycin M1 in animal Feed. Feed samples were extracted with methanol-2% formic acid aqueous solution, followed by a solid-phase extraction step using a HLB cartridge. Under the optimum chromatographic conditions and ELSD parameters, target compounds were separated well on a short column filled with biphenyl stationary phase. The method was developed in accordance with pig complete feed and then extended to detect polypeptide antibiotics in piglet premix, pig feed additive, poultry complete feed and fattening pig premix. The results showed that logarithmic calibration curves of eight analytes were linear (r 2 > 0.99) within the concentration range of 5–200 mg mL−1. The developed method provided good accuracy and precision for quantification of eight polypeptides in five kinds of feeds with recoveries ranging from 72.0% to 105.4% with relative standard deviations <9.5%. The limits of detection ranged from 2 to 5 mg kg−1. Finally, the method was successfully applied to analyze polypeptide antibiotics in commercial feed.





Determination of the low-abundant protein biomarker hCG from dried matrix spots using immunocapture and nano liquid chromatography mass spectrometry

Publication date: 1 March 2018
Source:Journal of Chromatography B, Volumes 1077–1078
Author(s): Cecilie Rosting, Elin Vyvy Tran, Astrid Gjelstad, Trine Grønhaug Halvorsen
Using LC-MS/MS for determination of low-abundance protein biomarkers from dried blood spots is challenging due to the combination of low biomarker levels (low pM-level) and small sample volumes (typically <50 μL). In the present paper it is demonstrated that use of state-of-the-art nano liquid chromatography triple quadrupole mass spectrometry in combination with immunoaffinity sample clean-up enable determination of the low abundance biomarker human chorionic gonadotropin (hCG) from four different biological matrices (whole blood, serum, plasma and urine) at its upper reference level (low pM). Detection limits for hCG was determined for all matrices from both commercially available non-soluble DBS sampling material (DMPK-C) and the water-soluble material carboxymethyl cellulose (CMC). The detection limits (S/N = 3) were ranging from 5.0 IU/L (14.5 pM; whole blood) to 10.5 IU/L (30.5 pM; urine) for DMPK-C and from 2.1 IU/L (6.1 pM; urine) to 6.4 IU/L (18.6 pM; plasma) for CMC. A brief evaluation was performed for both sampling materials using serum as matrix resulting in sufficient linearity (r2 ≥ 0.93, range 20–1000 IU/mL (58–2900 pM) for DMPK-C and 10–1000 IU/mL (29–2900 pM) for CMC), repeatability (RSD% = 13–31%) and accuracy (95–106%). To demonstrate the applicability of the method to real samples, a serum sample from a patient previously diagnosed with cancer was also analyzed using both sampling materials. The concentration levels found using the two materials were similar (5280± 595 IU/L (15,312 ± 1726 pM, n = 3) in the DMPK-C spot and 5060 ± 430 IU/L (14,674 ± 1247 pM, n = 3) in the CMC spot). All in all this demonstrated that the tools for determination of low abundance biomarkers at upper reference level from dried matrix spots now is available through a combination of immunoaffinity enrichment and state-of-the-art LC-MS/MS.

Publication date: 1 March 2018
Source:Journal of Chromatography B, Volumes 1077–1078

Author(s): Cecilie Rosting, Elin Vyvy Tran, Astrid Gjelstad, Trine Grønhaug Halvorsen

Using LC-MS/MS for determination of low-abundance protein biomarkers from dried blood spots is challenging due to the combination of low biomarker levels (low pM-level) and small sample volumes (typically <50 μL). In the present paper it is demonstrated that use of state-of-the-art nano liquid chromatography triple quadrupole mass spectrometry in combination with immunoaffinity sample clean-up enable determination of the low abundance biomarker human chorionic gonadotropin (hCG) from four different biological matrices (whole blood, serum, plasma and urine) at its upper reference level (low pM). Detection limits for hCG was determined for all matrices from both commercially available non-soluble DBS sampling material (DMPK-C) and the water-soluble material carboxymethyl cellulose (CMC). The detection limits (S/N = 3) were ranging from 5.0 IU/L (14.5 pM; whole blood) to 10.5 IU/L (30.5 pM; urine) for DMPK-C and from 2.1 IU/L (6.1 pM; urine) to 6.4 IU/L (18.6 pM; plasma) for CMC. A brief evaluation was performed for both sampling materials using serum as matrix resulting in sufficient linearity (r2 ≥ 0.93, range 20–1000 IU/mL (58–2900 pM) for DMPK-C and 10–1000 IU/mL (29–2900 pM) for CMC), repeatability (RSD% = 13–31%) and accuracy (95–106%). To demonstrate the applicability of the method to real samples, a serum sample from a patient previously diagnosed with cancer was also analyzed using both sampling materials. The concentration levels found using the two materials were similar (5280± 595 IU/L (15,312 ± 1726 pM, n = 3) in the DMPK-C spot and 5060 ± 430 IU/L (14,674 ± 1247 pM, n = 3) in the CMC spot). All in all this demonstrated that the tools for determination of low abundance biomarkers at upper reference level from dried matrix spots now is available through a combination of immunoaffinity enrichment and state-of-the-art LC-MS/MS.





Determination of pyrethrin and pyrethroid residues in animal fat using liquid chromatography coupled to tandem mass spectrometry

Publication date: 1 March 2018
Source:Journal of Chromatography B, Volumes 1077–1078
Author(s): M. Moloney, S. Tuck, A. Ramkumar, A. Furey, M. Danaher
A method was developed for the confirmatory and quantitative analysis of one pyrethrin and 18 pyrethroid residues in animal fat. Fat was extracted was collected from adipose tissue melted in an oven at 65 °C for 2 h. Fat samples (1 g) were dispersed with deactivated Florisil® sorbent and extracted with MeCN. Sample extracts were purified by cold temperature precipitation at −30 °C for 4 h and further purified using dispersive solid-phase extraction (d-SPE) clean-up in tubes containing 500 mg of Z-SEP+ and 125 mg of PSA bonded silica. Purified samples were analysed by ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC–MS/MS) detection. Chromatographic separation was carried out on a Acquity C8 BEH column, using a binary gradient separation comprising of mobile phase A, 5 mM ammonium formate in water:MeOH (80:20, v/v,) and mobile phase B, 5 mM ammonium formate in MeOH. The mass spectrometer was operated in the positive electrospray ionisation mode (ESI(+)). Validation was performed following the 2002/657/EC guidelines. Trueness ranged between 84% and 143% and precision ranged between 3.9% and 29%. The developed method is particularly advantageous because the sample preparation procedure does not require complex sample extraction equipment and uses less solvent compared to other sample preparation protocols.

Publication date: 1 March 2018
Source:Journal of Chromatography B, Volumes 1077–1078

Author(s): M. Moloney, S. Tuck, A. Ramkumar, A. Furey, M. Danaher

A method was developed for the confirmatory and quantitative analysis of one pyrethrin and 18 pyrethroid residues in animal fat. Fat was extracted was collected from adipose tissue melted in an oven at 65 °C for 2 h. Fat samples (1 g) were dispersed with deactivated Florisil® sorbent and extracted with MeCN. Sample extracts were purified by cold temperature precipitation at −30 °C for 4 h and further purified using dispersive solid-phase extraction (d-SPE) clean-up in tubes containing 500 mg of Z-SEP+ and 125 mg of PSA bonded silica. Purified samples were analysed by ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC–MS/MS) detection. Chromatographic separation was carried out on a Acquity C8 BEH column, using a binary gradient separation comprising of mobile phase A, 5 mM ammonium formate in water:MeOH (80:20, v/v,) and mobile phase B, 5 mM ammonium formate in MeOH. The mass spectrometer was operated in the positive electrospray ionisation mode (ESI(+)). Validation was performed following the 2002/657/EC guidelines. Trueness ranged between 84% and 143% and precision ranged between 3.9% and 29%. The developed method is particularly advantageous because the sample preparation procedure does not require complex sample extraction equipment and uses less solvent compared to other sample preparation protocols.





Determination and long-term stability of twenty-nine cathinones and amphetamine-type stimulants (ATS) in urine using gas chromatography–mass spectrometry

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076
Author(s): Khalid A. Alsenedi, Calum Morrison
A method was developed for the screening and quantification of seven amphetamine-type stimulants (ATS) and 22 cathinones, including three metabolites, in urine with Gas Chromatography–Mass Spectrometry. This method allowed the detection and quantification of ATS and cathinones group molecules using one procedure. A study of the stability of the drug mixtures for a period of 201 days in human urine samples under three different conditions has been carried. The ATS and cathinones include amphetamine, methamphetamine, MDA, MDEA, MDMA, PMA, PMMA, cathinone, methcathinone, 3′-position-substituted, ring-substituted, methylenedioxy-substituted, N-alkyl-substituted and pyrrolidinyl-substituted. Twenty drugs out of twenty-nine were validated with a quantitative method. This method can be applied to the nine remaining drugs as a screening method. The linearity of the assay was from 50 to 2000 ng/ml, with limits of detection of 0.5 to 10 ng/ml. In terms of accuracy, between-run and within-run precision were ≤20% for 20 compounds with good selectivity. No carryover was seen, and the recovery was between 80 and 120% for most drugs tested. ATS and pyrrolidinyl-substituted groups were conducted to be stable compounds under all conditions. All compounds tested were stable at −20 °C. Some cathinones were primarily degraded after 21 days at 4 °C. They were detectable but unstable after 201 days at 4 °C. Most cathinones were unstable after a day and completely lost after 28 days at RT.

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076

Author(s): Khalid A. Alsenedi, Calum Morrison

A method was developed for the screening and quantification of seven amphetamine-type stimulants (ATS) and 22 cathinones, including three metabolites, in urine with Gas Chromatography–Mass Spectrometry. This method allowed the detection and quantification of ATS and cathinones group molecules using one procedure. A study of the stability of the drug mixtures for a period of 201 days in human urine samples under three different conditions has been carried. The ATS and cathinones include amphetamine, methamphetamine, MDA, MDEA, MDMA, PMA, PMMA, cathinone, methcathinone, 3′-position-substituted, ring-substituted, methylenedioxy-substituted, N-alkyl-substituted and pyrrolidinyl-substituted. Twenty drugs out of twenty-nine were validated with a quantitative method. This method can be applied to the nine remaining drugs as a screening method. The linearity of the assay was from 50 to 2000 ng/ml, with limits of detection of 0.5 to 10 ng/ml. In terms of accuracy, between-run and within-run precision were ≤20% for 20 compounds with good selectivity. No carryover was seen, and the recovery was between 80 and 120% for most drugs tested. ATS and pyrrolidinyl-substituted groups were conducted to be stable compounds under all conditions. All compounds tested were stable at −20 °C. Some cathinones were primarily degraded after 21 days at 4 °C. They were detectable but unstable after 201 days at 4 °C. Most cathinones were unstable after a day and completely lost after 28 days at RT.





High efficiency and fast separation of active proteins by HIC chromatographic pie with sub-2 μm polymer packings

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076
Author(s): Cong-Yu Ke, Guo-Min Lu, Wu-Juan Sun, Xun-Li Zhang
This paper reports the development of hydrophobic interaction chromatography (HIC) by synthesizing sub-2 μm polymer packings which was packed into a chromatographic pie for fast separation of native proteins at low pressures demonstrating high efficiency. Using styrene as monomer and ethylene dimethacrylate (EDMA)as swelling agent, the polystyrene seeds with an average particle size of 0.8 μm and monodisperse polymeric microspheres with a particle size of 1.5–5.0 μm were synthesized through dispersion polymerization and one-step swelling method, respectively. In order to separate active proteins, the microspheres were modified to hydrophobic chromatographic packings through covalent bonding with benzene methanol. Compared with the traditional column chromatography, the sub-2 μm polymer packings in chromatographic pie exhibited higher column efficiency for protein separation at lower column pressures, even at higher flow rates. The van Deemter curve showed that the flow rate had insignificant effect on column efficiency of chromatographic pie. Seven example proteins were clearly separated within 3 min at a flow rate of 10 mL/min. The applicability of this method was further demonstrated by the separation of human serum samples. The results indicated that this chromatographic mode can be potentially applied for the fast separation of complex active proteins, such as protein drugs from natural products.

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076

Author(s): Cong-Yu Ke, Guo-Min Lu, Wu-Juan Sun, Xun-Li Zhang

This paper reports the development of hydrophobic interaction chromatography (HIC) by synthesizing sub-2 μm polymer packings which was packed into a chromatographic pie for fast separation of native proteins at low pressures demonstrating high efficiency. Using styrene as monomer and ethylene dimethacrylate (EDMA)as swelling agent, the polystyrene seeds with an average particle size of 0.8 μm and monodisperse polymeric microspheres with a particle size of 1.5–5.0 μm were synthesized through dispersion polymerization and one-step swelling method, respectively. In order to separate active proteins, the microspheres were modified to hydrophobic chromatographic packings through covalent bonding with benzene methanol. Compared with the traditional column chromatography, the sub-2 μm polymer packings in chromatographic pie exhibited higher column efficiency for protein separation at lower column pressures, even at higher flow rates. The van Deemter curve showed that the flow rate had insignificant effect on column efficiency of chromatographic pie. Seven example proteins were clearly separated within 3 min at a flow rate of 10 mL/min. The applicability of this method was further demonstrated by the separation of human serum samples. The results indicated that this chromatographic mode can be potentially applied for the fast separation of complex active proteins, such as protein drugs from natural products.





Method development, matrix effect, and risk assessment of 49 multiclass pesticides in kiwifruit using liquid chromatography coupled to tandem mass spectrometry

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076
Author(s): Yool-Ah Kim, A.M. Abd El-Aty, Md. Musfiqur Rahman, Ji Hoon Jeong, Ho-Chul Shin, Jing Wang, SungShik Shin, Jae-Han Shim
In the present study, a liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method with a minimal matrix effect (ME) was developed and validated for simultaneous determination of a diverse range of pesticides (49) in kiwifruit. Samples extracted by the quick, easy, cheap, effective, rugged, and safe (QuEChERS) citrate-buffered method were analyzed either without purification or following purification (with primary secondary amine (PSA) or PSA + graphitized carbon black (GCB)). With the addition of a clean-up step, the suppression of the ME decreased, with a higher number of pesticides determined by the application of PSA + GCB. The method exhibited good linearity with coefficients of determination (R2) ≥ 0.9972 and satisfactory recoveries (70–120%) with a relative standard deviations (RSDs) <10%. The limits of quantification (LOQ) were lower than the maximum residue limits (MRLs) set by the Korean Ministry of Food and Drug Safety (MFDS) and the CODEX Alimentarius. The developed method was applied to the real samples and the results indicated that the quantitated levels of all pesticides, except for pyraclostrobin and carbendazim, are lower than the MRLs set by the regulatory authorities. The percentage of the acceptable daily intake was <20%, suggesting that there is no risk associated with the intake of residual pesticides through kiwifruit.

Graphical abstract

image

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076

Author(s): Yool-Ah Kim, A.M. Abd El-Aty, Md. Musfiqur Rahman, Ji Hoon Jeong, Ho-Chul Shin, Jing Wang, SungShik Shin, Jae-Han Shim

In the present study, a liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) method with a minimal matrix effect (ME) was developed and validated for simultaneous determination of a diverse range of pesticides (49) in kiwifruit. Samples extracted by the quick, easy, cheap, effective, rugged, and safe (QuEChERS) citrate-buffered method were analyzed either without purification or following purification (with primary secondary amine (PSA) or PSA + graphitized carbon black (GCB)). With the addition of a clean-up step, the suppression of the ME decreased, with a higher number of pesticides determined by the application of PSA + GCB. The method exhibited good linearity with coefficients of determination (R 2 ) ≥ 0.9972 and satisfactory recoveries (70–120%) with a relative standard deviations (RSDs) <10%. The limits of quantification (LOQ) were lower than the maximum residue limits (MRLs) set by the Korean Ministry of Food and Drug Safety (MFDS) and the CODEX Alimentarius. The developed method was applied to the real samples and the results indicated that the quantitated levels of all pesticides, except for pyraclostrobin and carbendazim, are lower than the MRLs set by the regulatory authorities. The percentage of the acceptable daily intake was <20%, suggesting that there is no risk associated with the intake of residual pesticides through kiwifruit.

Graphical abstract

image




Rapid and sensitive determination of formamidines and metabolites with HPLC-MS/MS using core-shell columns

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076
Author(s): Yuhong Wu, Hao Guo, Juan Bu, Yanglan Tan, Jian Zhong, Qingbiao Zhao
A number of poisoning and suicide cases involving formamidine pesticides have been reported, thus developing a rapid and low cost determination method is crucial. In this work, a rapid, sensitive and low-cost method for the simultaneous determination of formamidine pesticides (amitraz, chlordimeform, formetanate) and their main metabolites, N-(2,4-dimethylphenyl)-N-methyl-formamidine, 2,4-dimethylformamidine, 2,4-dimethylaniline, 4-chloro-2-methylaniline and 3-hydroxyacetanilide in human blood by high-performance liquid chromatography with tandem mass spectrometry is developed. The application of columns with core-shell particles significantly reduced the analysis time. Very low LODs (0.01–0.04 μg L−1) were obtained for formamidine pesticides and their metabolites. The method was successfully applied to the analysis of human blood samples from a real forensic case. The significantly reduced analysis time, high sensitivity and low cost are the primary advantages of the developed method. This methodology provides important value for sensitive and rapid determination of residue pesticides and metabolites, study of residue pesticides behavior in human body, as well as application in real forensic cases.

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076

Author(s): Yuhong Wu, Hao Guo, Juan Bu, Yanglan Tan, Jian Zhong, Qingbiao Zhao

A number of poisoning and suicide cases involving formamidine pesticides have been reported, thus developing a rapid and low cost determination method is crucial. In this work, a rapid, sensitive and low-cost method for the simultaneous determination of formamidine pesticides (amitraz, chlordimeform, formetanate) and their main metabolites, N-(2,4-dimethylphenyl)-N-methyl-formamidine, 2,4-dimethylformamidine, 2,4-dimethylaniline, 4-chloro-2-methylaniline and 3-hydroxyacetanilide in human blood by high-performance liquid chromatography with tandem mass spectrometry is developed. The application of columns with core-shell particles significantly reduced the analysis time. Very low LODs (0.01–0.04 μg L−1) were obtained for formamidine pesticides and their metabolites. The method was successfully applied to the analysis of human blood samples from a real forensic case. The significantly reduced analysis time, high sensitivity and low cost are the primary advantages of the developed method. This methodology provides important value for sensitive and rapid determination of residue pesticides and metabolites, study of residue pesticides behavior in human body, as well as application in real forensic cases.





Supercritical fluid extraction (SFE) of ketamine metabolites from dried urine and on-line quantification by supercritical fluid chromatography and single mass detection (on-line SFE–SFC–MS)

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076
Author(s): Robert Hofstetter, Georg M. Fassauer, Andreas Link
On-line solid-phase supercritical fluid extraction (SFE) and chromatography (SFC) coupled to mass spectrometry (MS) has been evaluated for its usefulness with respect to metabolic profiling and pharmacological investigations of ketamine in humans. The aim of this study was to develop and validate a rapid, highly selective and sensitive SFE–SFC–MS method for the quantification of ketamine and its metabolites in miniature amounts in human urine excluding liquid-liquid extraction (LLE). Several conditions were optimized systematically following the requirements of the European Medicines Agency: selectivity, carry-over, calibration curve parameters (LLOQ, range and linearity), within- and between-run accuracy and precision, dilution integrity, matrix effect, and stability. The method, which required a relatively small volume of human urine (20 μL per sample), was validated for pharmacologically and toxicologically relevant concentrations ranging from 25.0 to 1000 ng/mL (r2 > 0.995). The lower limit of quantification (LLOQ) for all compounds was found to be as low as 0.5 ng. In addition, stability of analytes during removal of water from the urine samples using different conditions (filter paper or ISOLUTE® HM-N) was studied. In conclusion, the method developed in this study can be successfully applied to studies of ketamine metabolites in humans, and may pave the way for routine application of on-line SFE–SFC–MS in clinical investigations.

Publication date: 15 February 2018
Source:Journal of Chromatography B, Volume 1076

Author(s): Robert Hofstetter, Georg M. Fassauer, Andreas Link

On-line solid-phase supercritical fluid extraction (SFE) and chromatography (SFC) coupled to mass spectrometry (MS) has been evaluated for its usefulness with respect to metabolic profiling and pharmacological investigations of ketamine in humans. The aim of this study was to develop and validate a rapid, highly selective and sensitive SFE–SFC–MS method for the quantification of ketamine and its metabolites in miniature amounts in human urine excluding liquid-liquid extraction (LLE). Several conditions were optimized systematically following the requirements of the European Medicines Agency: selectivity, carry-over, calibration curve parameters (LLOQ, range and linearity), within- and between-run accuracy and precision, dilution integrity, matrix effect, and stability. The method, which required a relatively small volume of human urine (20 μL per sample), was validated for pharmacologically and toxicologically relevant concentrations ranging from 25.0 to 1000 ng/mL (r2 > 0.995). The lower limit of quantification (LLOQ) for all compounds was found to be as low as 0.5 ng. In addition, stability of analytes during removal of water from the urine samples using different conditions (filter paper or ISOLUTE® HM-N) was studied. In conclusion, the method developed in this study can be successfully applied to studies of ketamine metabolites in humans, and may pave the way for routine application of on-line SFE–SFC–MS in clinical investigations.