13 results on '"Long-Hui Liang"'
Search Results
2. Generic detection of organophosphorus nerve agent adducts to butyrylcholinesterase in plasma using liquid chromatography-tandem mass spectrometry combined with an improved procainamide-gel separation and pepsin digestion method
- Author
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Chang-Cai Liu, Long-Hui Liang, Long Yan, Bo Chen, Xiu-Jie Liu, Yang Yang, Shi-Lei Liu, and Hai-Ling Xi
- Subjects
Organic Chemistry ,General Medicine ,Biochemistry ,Analytical Chemistry - Published
- 2023
3. Quantitative detection of ricin in beverages using trypsin/Glu-C tandem digestion coupled with ultra-high-pressure liquid chromatography-tandem mass spectrometry
- Author
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Ji-Na Wu, Long Yan, Hui-Lan Yu, Xi-Hui Mu, Long-Hui Liang, Bo Chen, Shi-Lei Liu, Yang Yang, Chang-Cai Liu, Xi Cheng, and Xiao-Sen Li
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Quality Control ,Streptavidin ,Peptide ,Ricin ,02 engineering and technology ,Mass spectrometry ,Sensitivity and Specificity ,01 natural sciences ,Biochemistry ,Analytical Chemistry ,Beverages ,Magnetics ,chemistry.chemical_compound ,Limit of Detection ,Tandem Mass Spectrometry ,Liquid chromatography–mass spectrometry ,medicine ,Biotinylation ,Trypsin ,Chromatography, High Pressure Liquid ,Detection limit ,Orange juice ,chemistry.chemical_classification ,Chromatography ,Chemistry ,010401 analytical chemistry ,Reproducibility of Results ,021001 nanoscience & nanotechnology ,0104 chemical sciences ,Isotope Labeling ,Calibration ,Solvents ,Peptides ,0210 nano-technology ,medicine.drug - Abstract
The toxic protein of ricin has drawn wide attention in recent years as a potential bioterrorism agent due to its high toxicity and wide availability. For the verification of the potential anti-terrorism activities, it is urgent for the quantification of ricin in food-related matrices. Here, a novel strategy of trypsin/Glu-C tandem digestion was introduced for quantitative detection of ricin marker peptides in several beverage matrices using isotope-labeled internal standard (IS)-mass spectrometry. The ricin in beverages was captured and enriched by biotinylated anti-ricin polyclonal antibodies conjugated to streptavidin magnetic beads. The purified ricin was cleaved using the developed trypsin/Glu-C tandem digestion method and then quantitatively detected by ultra-high-pressure liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) with isotope-labeled T7A and TG11B selected as IS. The use of trypsin/Glu-C digestion allows shorter peptides, which are more suitable for MS detection, to be obtained than the use of single trypsin digestion. Under the optimized tandem digestion condition, except for T7A in the A-chain, two resulting specific peptides of TG13A, TG28A from the A-chain and two of TG11B, TG33B from the B-chain were chosen as novel marker peptides with high MS response. The uniqueness of the selected marker peptides allows for unambiguous identification of ricin among its homologous proteins in a single run. The MS response of the four novel marker peptides is increased by more than 10 times compared with that of individual corresponding tryptic peptides. Both the marker peptides of A-chain T7A and B-chain TG11B were selected as quantitative peptides based on the highest MS response among the marker peptides from their individual chains. The limit of detection (LOD) of ricin is 0.1 ng/mL in PBS and 0.5 ng/mL in either milk or orange juice. The linear range of calibration curves for ricin were 0.5-300 ng/mL in PBS, 1.0-400 ng/mL in milk, and 1.0-250 ng/mL in orange juice. The method accuracy ranged between 82.6 and 101.8% for PBS, 88.9-105.2% for milk, and 95.3-118.7% for orange juice. The intra-day and inter-day precision had relative standard deviations (%RSD) of 0.3-9.4%, 0.7-8.9%, and 0.2-6.9% in the three matrices respectively. Furthermore, whether T7A or TG11B is used as a quantitative peptide, the quantitative results of ricin are consistent. This study provides not only a practical method for the absolute quantification of ricin in beverage matrices but also a new strategy for the investigation of illegal use of ricin in chemical weapon verification tasks such as OPCW biotoxin sample analysis exercises.
- Published
- 2020
4. [Highly toxic type Ⅱ ribosome-inactivating proteins ricin and abrin and their detection methods: a review]
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Long-Hui Liang, Junmei Xia, Chang-Cai Liu, and Shi-Lei Liu
- Subjects
Chromatography ,Oligonucleotide ,Chemistry ,General Chemical Engineering ,Ribosome-inactivating protein ,Aptamer ,Organic Chemistry ,Ribosome Inactivating Proteins ,Ricin ,Biochemistry ,Analytical Chemistry ,chemistry.chemical_compound ,Protein structure ,Targeted mass spectrometry ,Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ,Electrochemistry ,Depurination ,Abrin ,Ribosomes ,Plant Proteins - Abstract
Type Ⅱ ribosome-inactivating proteins (RIPs) are an important class of protein toxins that consist of A and B chains linked by an interchain disulfide bond. The B-chain with lectin-like activity is responsible for binding to the galactose-containing receptors on eukaryotic cell surfaces, which is essential for A-chain internalization by endocytosis. The A-chain has N-glycosidase activity that irreversibly depurinates a specific adenine from 28S ribosomal RNA (28S rRNA) and terminates protein synthesis. The synergistic effect of the A-B chain inactivates the ribosome, inhibits protein synthesis, and exhibits high cytotoxicity. Ricin and abrin that are expressed by the plants Ricinus communis and Abrus precatorius, respectively, are typical type Ⅱ RIPs. The toxicity of ricin and abrin are 385 times and 2885 times, respectively, more that of the nerve agent VX. Owing to their ease of preparation, wide availability, and potential use as a bioterrorism agent, type Ⅱ RIPs have garnered increasing attention in recent years. Ricin is listed as a prohibited substance under schedule 1A of the Chemical Weapons Convention (CWC). The occurrence of ricin-related bioterrorism incidents in recent years has promoted the development of accurate, sensitive, and rapid detection and identification technology for type Ⅱ RIPs. Significant progress has been made in the study of toxicity mechanisms and detection methods of type Ⅱ RIPs, which primarily involve qualitative and quantitative analysis methods including immunological assays, mass spectrometry analysis methods, and toxin activity detection methods based on depurination and cytotoxicity. Immunoassays generally involve the specific recognition of antigens and antibodies, which is based on oligonucleotide molecular recognition elements called aptamers. These methods are fast and highly sensitive, but for highly homologous proteins in complex samples, they provide false positive results. With the rapid development of biological mass spectrometry detection technology, techniques such as electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI) are widely used in the identification of proteins. These methods not only provide accurate information on molecular weight and structure of proteins, but also demonstrate accurate quantification. Enzyme digestion combined with mass spectrometry is the predominantly used detection method. Accurate identification of protein toxins can be achieved by fingerprint analysis of enzymatically digested peptides. For analysis of protein toxins in complex samples, abundant peptide markers are obtained using a multi-enzyme digestion strategy. Targeted mass spectrometry analysis of peptide markers is used to obtain accurate qualitative and quantitative information, which effectively improves the accuracy and sensitivity of the identification of type Ⅱ RIP toxins. Although immunoassay and mass spectrometry detection methods can provide accurate identification of type Ⅱ RIPs, they cannot determine whether the toxins will retain potency. The widely used detection methods for activity analysis of type Ⅱ RIPs include depurination assay based on N-glycosidase activity and cytotoxicity assay. Both the methods provide simple, rapid, and sensitive analysis of type Ⅱ RIP toxicity, and complement other detection methods. Owing to the importance of type Ⅱ RIP toxins, the Organization for the Prohibition of Chemical Weapons (OPCW) has proposed clear technical requirements for the identification and analysis of relevant samples. We herein reviewed the structural characteristics, mechanism of action, and the development and application of type Ⅱ RIP detection methods; nearly 70 studies on type Ⅱ RIP toxins and their detection methods have been cited. In addition to the technical requirements of OPCW for the unambiguous identification of biotoxins, the trend of future development of type Ⅱ RIP-based detection technology has been explored.
- Published
- 2021
5. Rapid Differential Detection of Abrin Isoforms by an Acetonitrile- and Ultrasound-Assisted On-Bead Trypsin Digestion Coupled with LC-MS/MS Analysis
- Author
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Chang-Cai Liu, Yang Yang, Hui-Lan Yu, Long-Hui Liang, Shu Geng, Shi-Lei Liu, and Xi Cheng
- Subjects
Streptavidin ,RIP II protein toxins ,Health, Toxicology and Mutagenesis ,Size-exclusion chromatography ,Isotope dilution ,Toxicology ,medicine.disease_cause ,01 natural sciences ,Article ,03 medical and health sciences ,chemistry.chemical_compound ,Tandem Mass Spectrometry ,Abrus precatorius ,HPLC-ESI-MS/MS ,medicine ,Animals ,Protein Isoforms ,Computer Simulation ,Trypsin ,Ultrasonics ,Chromatography, High Pressure Liquid ,030304 developmental biology ,Toxins, Biological ,0303 health sciences ,Chromatography ,biology ,Toxin ,010401 analytical chemistry ,biology.organism_classification ,0104 chemical sciences ,Milk ,chemistry ,Polyclonal antibodies ,ultrasound-assisted trypsin digestion ,Biotinylation ,Abrus ,biology.protein ,Medicine ,marker peptides ,Abrin ,Rabbits ,detection of abrin ,Chromatography, Liquid - Abstract
The high toxic abrin from the plant Abrus precatorius is a type II ribosome-inactivating protein toxin with a human lethal dose of 0.1–1.0 µg/kg body weight. Due to its high toxicity and the potential misuse as a biothreat agent, it is of great importance to developing fast and reliable methods for the identification and quantification of abrin in complex matrices. Here, we report rapid and efficient acetonitrile (ACN)- and ultrasound-assisted on-bead trypsin digestion method combined with HPLC-MS/MS for the quantification of abrin isoforms in complex matrices. Specific peptides of abrin isoforms were generated by direct ACN-assisted trypsin digestion and analyzed by HPLC-HRMS. Combined with in silico digestion and BLASTp database search, fifteen marker peptides were selected for differential detection of abrin isoforms. The abrin in milk and plasma was enriched by immunomagnetic beads prepared by biotinylated anti-abrin polyclonal antibodies conjugated to streptavidin magnetic beads. The ultrasound-assisted on-bead trypsin digestion method was carried out under the condition of 10% ACN as denaturant solvent, the entire digestion time was further shortened from 90 min to 30 min. The four peptides of T3Aa,b,c,d, T12Aa, T15Ab, and T9Ac,d were chosen as quantification for total abrin, abrin-a, abrin-b, and abrin-c/d, respectively. The absolute quantification of abrin and its isoforms was accomplished by isotope dilution with labeled AQUA peptides and analyzed by HPLC-MS/MS (MRM). The developed method was fully validated in milk and plasma matrices with quantification limits in the range of 1.0-9.4 ng/mL for the isoforms of abrin. Furthermore, the developed approach was applied for the characterization of abrin isoforms from various fractions from gel filtration separation of the seeds, and measurement of abrin in the samples of biotoxin exercises organized by the Organization for the Prohibition of Chemical Weapons (OPCW). This study provided a recommended method for the differential identification of abrin isoforms, which are easily applied in international laboratories to improve the capabilities for the analysis of biotoxin samples.
- Published
- 2021
6. Direct Acetonitrile-Assisted Trypsin Digestion Method Combined with LC-MS/MS-Targeted Peptide Analysis for Unambiguous Identification of Intact Ricin
- Author
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Long Yan, Xiao-Sen Li, Hai-Ling Xi, Hui-Lan Yu, Yang Yang, Bo Chen, Chang-Cai Liu, Shi-Lei Liu, and Long-Hui Liang
- Subjects
0301 basic medicine ,Acetonitriles ,Ricin ,Cleavage (embryo) ,medicine.disease_cause ,Biochemistry ,Matrix (chemical analysis) ,03 medical and health sciences ,chemistry.chemical_compound ,Liquid chromatography–mass spectrometry ,Tandem Mass Spectrometry ,medicine ,Trypsin ,chemistry.chemical_classification ,Chromatography ,030102 biochemistry & molecular biology ,Toxin ,General Chemistry ,030104 developmental biology ,Enzyme ,chemistry ,Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ,Digestion ,Peptides ,medicine.drug ,Chromatography, Liquid - Abstract
Ricin is a type II ribosome-inactivating protein toxin consisting of A and B chains linked by one interchain disulfide bond. Because of its high toxicity depending on both chains together, confirming the presence of both A and B chains of intact ricin is required during the investigation of the illegal production and application. Here, we report a novel and sensitive acetonitrile (ACN)-assisted trypsin digestion method for unambiguous identification of intact ricin by simultaneous detection of its marker peptides from A and B chains. Marker peptides were generated with a simple procedure by direct cleaving the native ricin at 45 °C for 4 h using Promega modified sequencing grade trypsin under the assistance of 10% ACN, and then directly analyzed by ultrahigh performance liquid chromatography tandem mass spectrometry. The type of trypsin was found to be one critical factor for cleavage of intact ricin based on a significant difference in the yields of specific peptides generated while using various types of trypsin. A low content of ACN in enzymatic buffer significantly reduced the digestion time from overnight to 4 h. There was commonly a better MS response of marker peptides when using the developed ACN-assisted trypsin digestion method than methanol-assisted trypsin digestion within the same 4 h. Totally, seven specific peptides with high sensitivity and specificity including three in the A-chain (TA7, TA11, and TA10) and four in the B-chain (TB6, TB14-ss-TB16, TB20, and TB18) were obtained as good marker peptides for unambiguous identification of intact ricin. The lowest concentration of native ricin for unambiguous identification was 20 ng/mL, in which three marker peptides from both the A-chain and B-chain could be measured with a minimum of three ion transitions. Combined with affinity enrichment, the developed approach was successfully applied for the measurement of intact ricin from the complicated matrix samples of the second, third, and fourth biotoxin exercises organized by the Organisation for the Prohibition of Chemical Weapons (OPCW). This study has provided a recommended detection method combined with one novel ACN-assisted trypsin digestion with MS for forensic unambiguous confirmation of trace ricin intact with high confidence.
- Published
- 2020
7. Advances in sulfur mustard-induced DNA adducts: Characterization and detection
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Chang-Cai Liu, Yang Yang, Junmei Xia, Bo Chen, Hui-Lan Yu, Long-Hui Liang, Yihe Li, Xi Cheng, and Shi-Lei Liu
- Subjects
0301 basic medicine ,DNA damage ,Toxicology ,medicine.disease_cause ,Adduct purification ,Adduct ,03 medical and health sciences ,chemistry.chemical_compound ,DNA Adducts ,0302 clinical medicine ,DNA adduct ,Mustard Gas ,medicine ,Animals ,Humans ,Chemical Warfare Agents ,Sulfur mustard ,General Medicine ,DNA extraction ,030104 developmental biology ,chemistry ,Biochemistry ,030217 neurology & neurosurgery ,Genotoxicity ,DNA ,Biomarkers - Abstract
Sulfur mustard (SM) is a blister chemical warfare agent with severe cytotoxicity and genotoxicity. It can extensively alkylate important macromolecules in organisms, such as proteins, DNA, and lipids, and produce a series of metabolites, among which the characteristic ones can be used as biomarkers. The exact toxicological mechanisms of SM remain unclear but mainly involve the DNA lesions induced by alkylation and oxidative stress caused by glutathione depletion. Various methods have been used to analyze DNA damage caused by SM. Among these methods, liquid chromatography-tandem mass spectrometry (LC-MS/MS) technology stands out and makes it possible to observe damage in view of biomarkers induced by SM. Sample preparation is critical for detection by LC-MS/MS and mainly includes DNA isolation, adduct hydrolysis, and adduct purification. Moreover, optimization of chromatographic conditions, selection of MS transitions, and quantitative strategies are also essential. SM-DNA adducts are generally considered to be N7-HETEG, O6-HETEG, N7-BisG, and N3-HETEA. This article proposes some other possibilities of SM-DNA adducts for the identification of SM genotoxicity.
- Published
- 2020
8. A novel BODIPY-based fluorescent probe for sensitive and selective detection of nerve agent simulants through base-assisted photo-induced electron transfer process
- Author
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Sheng-Song Li, Xiao-Ming Zhu, Xu-Zhe Wang, Ling Yuan, Fa-Heng Zhang, Long-Hui Liang, He Zheng, Yong-Chao Zheng, Hong-Bo Wang, and Chong-Lin Zhao
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Detection limit ,Metals and Alloys ,Conjugated system ,Condensed Matter Physics ,Photochemistry ,Fluorescence ,Surfaces, Coatings and Films ,Electronic, Optical and Magnetic Materials ,chemistry.chemical_compound ,Electron transfer ,chemistry ,Intramolecular force ,Materials Chemistry ,medicine ,Electrical and Electronic Engineering ,BODIPY ,Instrumentation ,Tabun ,Nerve agent ,medicine.drug - Abstract
A fluorescent BODIPY probe (SO-BOD) conjugated with a 2-salicylaldoxime group via an ethynyl spacer was designed and synthesized for highly sensitive and selective detection of DCNP (a simulant of Tabun). The probe exhibited significant fluorescent quenching in the presence of Et3N, and 30-fold fluorescence enhancement was achieved in the presence of DCNP. Integration of experimental and computational methods well illustrated that these fluorescence variations were attributed to the intramolecular photo-induced electron transfer (PET) process. Except for the low limit of detection (34 nM) to DCNP, SO-BOD also showed a rapid response (t1/2 = 175 s) and high selectivity. Furthermore, naked-eye detection of DCNP was carried out in both the liquid and vapor phases by test strips immobilized with the probe.
- Published
- 2021
9. Simultaneous quantification of four metabolites of sulfur mustard in urine samples by ultra-high performance liquid chromatography-tandem mass spectrometry after solid phase extraction
- Author
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Chang-Cai Liu, Long-Hui Liang, Hai-Ling Xi, Huang Guilan, Shi-Lei Liu, Hui-Lan Yu, Jing-Quan Liu, and Shi-Kun Zhou
- Subjects
Analyte ,Metabolite ,Urine ,Acetates ,Thiodiglycol ,01 natural sciences ,Biochemistry ,Analytical Chemistry ,Matrix (chemical analysis) ,chemistry.chemical_compound ,Tandem Mass Spectrometry ,Liquid chromatography–mass spectrometry ,Mustard Gas ,Humans ,Chemical Warfare Agents ,Sulfhydryl Compounds ,Solid phase extraction ,Chromatography, High Pressure Liquid ,Detection limit ,Chromatography ,010405 organic chemistry ,Solid Phase Extraction ,010401 analytical chemistry ,Organic Chemistry ,Reproducibility of Results ,General Medicine ,0104 chemical sciences ,chemistry ,Biomarkers - Abstract
Four HD urinary metabolites including hydrolysis metabolite thiodiglycol (TDG), glutathione-derived metabolite 1,1'-sulfonylbis[2-S-(N-acetylcysteinyl)ethane] (SBSNAE), as well as the β-lyase metabolites 1,1'-sulfonylbis[2-(methylsulfinyl)ethane] (SBMSE) and 1-methylsulfinyl-2-[2-(methylthio) ethylsulfonyl]ethane (MSMTESE) are considered as important biomarkers for short-term retrospective detection of HD exposure. In this study, a single method for simultaneous quantification of the four HD metabolites in urine samples was developed using ultra high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS). The four urinary metabolites were simultaneously extracted from urinary samples using a solid phase extraction (SPE) method with high extraction recoveries for all four metabolites varied in the range of 71.1-103% followed by UHPLC-MS/MS analysis. The SPE is simple and high effective only requiring 0.1mL of urinary samples and 0.5h time consuming. The problem of previous co-elution of TDG and SBSNAE in UHPLC was well solved, and complete separation of TDG, SBSNAE, SBMSE and MSMTESE from SPE-processed urine matrix was obtained to increase specificity and sensitivity. A full method validation was performed for each analyte in urine matrix. The linear range of calibration curves for the four analytes were respectively from 0.50-500ngmL-1 for TDG and SBSNAE, 0.05-500ngmL-1 for SBMSE and MSMTESE with coefficient of determination value (R2) ≥0.990. The limit of detection was 0.25ngmL-1 for TDG and SBSNAE, 0.01ngmL-1 for SBMSE and MSMTESE spiked in normal urine. The intra/inter-day precision for each analyte at three QC levels had relative standard deviation (%RSD) of ≤10.3%, and the intra/inter-day accuracy ranged between 88.0-108%. This developed method allows for simultaneous and trace measurement of four HD urinary metabolites within one single determination with the lowest usage amount of urine samples over all previous methods This study provides a useful tool for early diagnosis and monitoring of HD poisoning for medical treatment with high confidence, avoiding the need for application of several analysis methods.
- Published
- 2017
10. Simultaneous quantification of soman and VX adducts to butyrylcholinesterase, their aged methylphosphonic acid adduct and butyrylcholinesterase in plasma using an off-column procainamide-gel separation method combined with UHPLC–MS/MS
- Author
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Chang-Cai Liu, Ling Yuan, Shi-Kun Zhou, Long-Hui Liang, Hui-Lan Yu, Hai-Ling Xi, Shi-Lei Liu, Jing-Quan Liu, and Huang Guilan
- Subjects
0301 basic medicine ,Soman ,Clinical Biochemistry ,Procainamide ,Immunomagnetic separation ,01 natural sciences ,Biochemistry ,Analytical Chemistry ,Adduct ,03 medical and health sciences ,chemistry.chemical_compound ,Organophosphorus Compounds ,Limit of Detection ,Tandem Mass Spectrometry ,medicine ,Humans ,Sample preparation ,Chemical Warfare Agents ,Methylphosphonic acid ,Chromatography, High Pressure Liquid ,Butyrylcholinesterase ,Nerve agent ,Detection limit ,Chromatography ,010401 analytical chemistry ,Organothiophosphorus Compounds ,Equipment Design ,Cell Biology ,General Medicine ,0104 chemical sciences ,030104 developmental biology ,chemistry ,Cholinesterase Inhibitors ,Gels ,medicine.drug - Abstract
This work describes a novel and sensitive non-isotope dilution method for simultaneous quantification of organophosphorus nerve agents (OPNAs) soman (GD) and VX adducts to butyrylcholinesterase (BChE), their aged methylphosphonic acid (MeP) adduct and unadducted BChE in plasma exposed to OPNA. OPNA-BChE adducts were isolated with an off-column procainamide-gel separation (PGS) from plasma, and then digested with pepsin into specific adducted FGES*AGAAS nonapeptide (NP) biomarkers. The resulting NPs were detected by UHPLC-MS/MS MRM. The off-column PGS method can capture over 90% of BChE, MeP-BChE, VX-BChE and GD-BChE from their respective plasma materials. One newly designed and easily synthesized phosphorylated BChE nonapeptide with one Gly-to-Ala mutation was successfully reported to serve as internal standard instead of traditional isotopically labeled BChE nonapeptide. The linear range of calibration curves were from 1.00-200ngmL-1 for VX-NP, 2.00-200ngmL-1 for GD-NP and MeP-NP (R2≥0.995), and 3.00-200ngmL-1 for BChE NP (R2≥0.990). The inter-day precision had relative standard deviation (%RSD) of
- Published
- 2016
11. LC-HRMS Screening and Identification of Novel Peptide Markers of Ricin Based on Multiple Protease Digestion Strategies
- Author
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Long Yan, Hui-Lan Yu, Long-Hui Liang, Xiao-Sen Li, Shi-Lei Liu, Ji-Na Wu, Bo Chen, Yang Yang, and Chang-Cai Liu
- Subjects
analytical_chemistry ,endocrine system ,Health, Toxicology and Mutagenesis ,lcsh:Medicine ,Peptide ,Toxicology ,Article ,chemistry.chemical_compound ,Pepsin ,medicine ,Chymotrypsin ,Trypsin ,Amino Acid Sequence ,mass spectrometry ,chemistry.chemical_classification ,biology ,lcsh:R ,Proteinase K ,ricin ,Pepsin A ,Amino acid ,unambiguous identification ,Ricin ,chemistry ,Biochemistry ,biology.protein ,Solvents ,marker peptides ,Endopeptidase K ,Digestion ,Peptides ,medicine.drug ,Chromatography, Liquid - Abstract
Both ricin and R. communis agglutinin (RCA120), belonging to the type II ribosome-inactivating proteins (RIPs-Ⅱ), are derived from the seeds of the castor bean plant. They share very similar amino acid sequences, but ricin is much more toxic than RCA120. It is urgently necessary to distinguish ricin and RCA120 in response to public safety. Currently, mass spectrometric assays are well established for unambiguous identification of ricin by accurate analysis of differentiated amino acid residues after trypsin digestion. However, diagnostic peptides are relatively limited for unambiguous identification of trace ricin, especially in complex matrices. Here, we demonstrate a digestion strategy of multiple proteinases to produce novel peptide markers for unambiguous identification of ricin. Liquid chromatography-high resolution MS (LC-HRMS) was used to verify the resulting peptides, among which only the peptides with uniqueness and good MS response were selected as peptide markers. Seven novel peptide markers were obtained from tandem digestion of trypsin and endoproteinase Glu-C in PBS buffer. From the chymotrypsin digestion under reduction and non-reduction conditions, eight and seven novel peptides were selected respectively. Using pepsin under pH 1~2 and proteinase K digestion, six and five peptides were selected as novel peptide markers. In conclusion, the obtained novel peptides from the established digestion methods can be recommended for the unambiguous identification of ricin during the investigation of illegal use of the toxin.
- Published
- 2019
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12. A sensitive quantification approach for detection of HETE-CP adduct after benzyl chloroformate derivatization using ultra-high-pressure liquid chromatography tandem mass spectrometry
- Author
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Xinhai Li, Bo Chen, Hui-Lan Yu, Chang-Cai Liu, Shi-Lei Liu, Ji-Na Wu, Yang Yang, Long-Hui Liang, and Xiao-Sen Li
- Subjects
Alkylation ,Formates ,Serum Albumin, Human ,02 engineering and technology ,Pronase ,01 natural sciences ,Biochemistry ,Analytical Chemistry ,Adduct ,chemistry.chemical_compound ,Liquid chromatography–mass spectrometry ,Limit of Detection ,Tandem Mass Spectrometry ,Mustard Gas ,medicine ,Chemical Precipitation ,Humans ,Solid phase extraction ,Chemical Warfare Agents ,Derivatization ,Chromatography, High Pressure Liquid ,Benzyl chloroformate ,Detection limit ,Chromatography ,010401 analytical chemistry ,Solid Phase Extraction ,Dipeptides ,021001 nanoscience & nanotechnology ,Human serum albumin ,0104 chemical sciences ,chemistry ,Proteolysis ,0210 nano-technology ,Biomarkers ,medicine.drug - Abstract
Sulfur mustard (HD) reacts with human serum albumin (HSA) at Cys34 and produces a long-term biomarker of HD exposure. Here, we present a novel, sensitive, and convenient method for quantification of HD exposure by detection of HD-HSA adducts using pronase digestion, benzyl chloroformate (Cbz-Cl) derivatization, and ultra-high-pressure liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). The HSA in HD-exposed plasma in vitro was precipitated with acetone and digested (2 h, 50 °C) with pronase to form the alkylated dipeptide, S-hydroxyethylthioethyl-CysPro (HETE-CP). The HETE-CP adduct was derivatized with Cbz-Cl to generate N-carbobenzoxy HETE-CP (HETE-C(Cbz)P). The derivatized product was analyzed by UHPLC-MS/MS. HD surrogate, 2-chloroethyl ethyl sulfide (2-CEES), was introduced as a non-isotope internal standard (ISTD) instead of traditional d8-HD for quantification. The method was found to be linear between 1.00 and 200 ng/mL HD exposure (R2 > 0.998) with precision of ≤ 9.0% relative standard deviation (RSD) and accuracy ranged between 97.1 and 111%. The limit of detection (LOD) is 0.500 ng/mL (S/N~5), over 15 times lower than that of the previous method (7.95 ng/mL). Time-consuming affinity purification or solid phase extraction (SPE) is not needed in the experiment and the operation takes less than 5 h. This study provides a new strategy and useful tool for retrospective analysis of HD exposure by HETE-CP biomarker detection.
- Published
- 2018
13. An improved method for retrospective quantification of sulfur mustard exposure by detection of its albumin adduct using ultra-high pressure liquid chromatography-tandem mass spectrometry
- Author
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Long-Hui Liang, Shi-Lei Liu, Hai-Ling Xi, Yu Xiang, Chang-Cai Liu, Jing-Quan Liu, Shi-Kun Zhou, and Hui-Lan Yu
- Subjects
Analytical chemistry ,Poison control ,Tripeptide ,Mass spectrometry ,Sensitivity and Specificity ,Biochemistry ,Analytical Chemistry ,Adduct ,Tandem Mass Spectrometry ,Liquid chromatography–mass spectrometry ,Mustard Gas ,Protein Interaction Mapping ,Humans ,Sample preparation ,Chemical Warfare Agents ,Solid phase extraction ,Chromatography, High Pressure Liquid ,Serum Albumin ,Retrospective Studies ,Detection limit ,Chromatography ,Chemistry ,Reproducibility of Results ,Environmental Exposure ,Biological Assay ,Blood Chemical Analysis ,Protein Binding - Abstract
Sulfur mustard (HD) adduct to human serum albumin (ALB) at Cys-34 residue has become an important and long-term retrospective biomarker of HD exposure. Here, a novel, sensitive, and convenient approach for retrospective quantification of HD concentration exposed to plasma was established by detection of the HD-ALB adduct using ultra-high pressure liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) with a novel non-isotope internal standard (IS). The HD-ALB adduct was isolated from HD-exposed plasma with blue Sepharose. The adduct was digested with proteinase K to form sulfur-hydroxyethylthioethyl ([S-HETE])-Cys-Pro-Phe tripeptide biomarker. The tripeptide adduct could be directly analyzed by UHPLC-MS/MS without an additional solid phase extraction (SPE), which was considered as a critical procedure in previous methods. The easily available 2-chloroethyl ethylsulfide (2-CEES) as HD surrogate was first reported to be used as IS in place of traditional d8-HD for quantification of HD exposure. Furthermore, 2-CEES was also confirmed to be a good IS alternative for quantification of HD exposure by investigation of product ion spectra for their corresponding tripeptide adducts which exhibited identical MS/MS fragmentation behaviors. The method was found to be linear between 1.00 and 250 ng•mL(-1) HD exposure (R(2)>0.9989) with precision of
- Published
- 2015
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