Research on the Upper Limit of Accuracy for Predicting Theoretical Tandem Mass Spectrometry

In recent years, numerous theoretical tandem mass spectrometry prediction methods have been proposed, yet a systematic study and evaluation of their theoretical accuracy limits have not been conducted. If the accuracy of current methods approaches this limit, further exploration of new prediction techniques may become redundant. Conversely, a need for more precise prediction methods or models may be indicated. In this study, we have experimentally analyzed the limits of accuracy at different numbers of ions and parameters using repeated spectral pairs and integrating various similarity metrics. Results show significant achievements in accuracy for backbone ion methods with room for improvement. In contrast, full-spectrum prediction methods exhibit greater potential relative to the theoretical accuracy limit. Additionally, findings highlight the significant impact of normalized collision energy and instrument type on prediction accuracy, underscoring the importance of considering these factors in future theoretical tandem mass spectrometry predictions.

Rapid analysis of fifteen sulfonamide residues in pork and fish samples by automated on-line solid phase extraction coupled to liquid chromatography–tandem mass spectrometry

The aim of this work was to develop an automated on-line solid phase extraction(SPE)with liquid chromatography-tandem mass spectrometry method for the detection of fifteen sulfonamides in pork and fish samples.Samples were extracted with 0.2%formic acid acetonitrile solution,purified by on-line SPE device with HLB column,then separated by XBridge C18 column,using 0.1%formic acid solution and acetonitrile as the mobile phase.Mass spectrometric data was acquired under multiple reaction monitoring(MRM)mode using positive ionization electrospray.Internal standard method was used in the quantification,good linear relationship was got in range of 0.1–100 ng/mL and correlation coefficient was higher than 0.9990.The limits of detection were in the range of 0.125–2.00g/kg and the limits of quantitation were in the range of 0.250–5.00g/kg.Recoveries of the method were in range of 78.3%–99.3%,relative standard deviation were lower than 10%.The method was simple,sensitivity,and could be used for routine supervision and analysis of fifteen sulfonamides in pork and fish.

Liquid Chromatography-tandem Mass Spectrometry for Analysis of Acylcarnitines in Dried Blood Specimens Collected at Autopsy from Neonatal Intensive Care Unit

Objective To investigate the feasibility of analyzing acylcarnitine in dry filter-paper blood spots by liquid chromatography-tandem mass spectrometry(LC-MS/MS) which could be applied to detect inborn errors of metabolism in neonates.Methods We obtained filter-paper blood from 26 dead infants from a neonatal intensive care unit(NICU) between October 1,2008 and September 30,2009.Acylcarnitine and amino acid profiles were obtained with LC-MS/MS.Four infants underwent routine autopsy.The postmortem blood specimens were compared with newborn blood specimens,and with specimens obtained from older infants with metabolic disorders.Results Of all the 26 patients,5(19.2%) were diagnosed as having different kinds of diseases:3 with methylmalonic acidemia(the concentration of C3,and the ratio of C3/C16,C3/C2 increased),1 with maple syrup urine disease(the concentration of leucine and isoleucine increased),and 1 with isovaleric aci-demia(the concentration of C5 increased).Conclusions Postmortem metabolic test can explain infant deaths and provide estimates of deaths attributable to inborn errors of metabolism in NICU.LC-MS/MS is suitable for analysis of postmortem specimens and can be considered for routine application in NICU autopsy.

Liquid Chromatography-electrospray Ionization Tandem Mass Spectrometry for Simultaneous Determination of Metformin and Glimepiride in Beagle Dog Plasma and Bioequivalence Study

A sensitive and selective liquid chromatography-electrospray ionization tandem mass spectrometry（LC- ES1-MS/MS） was used for the simultaneous determination of metformin and glimepiride in beagle dog plasma with glipizide as internal standard（IS）. After simplified protein precipitation with methanol, both the analytes and IS were chromatographed on a Zorbax CN column via gradient elution with methanol（containing 5 mmol/L ammonium ace- tate） and 5 mmol/L aqueous ammonium acetate as the mobile phase. Detection was performed by multiple reaction monitoring（MRM） scanning via ESI source operated in positive ionization mode. Specificity, linearity, accuracy, pre- cision, recovery, matrix effect and stability were validated for metformin and glimepiride in beagle dog plasma. The calibration curves were linear in a concentration range of 10-10000 ng/mL for metformin and 4-4000 ng/mL for glimepiride with both correlation coefficients higher than 0.99. The recoveries obtained for the analytes and IS were all between 82.7% and 101.2%. The method exhibited excellent performance in terms of selectivity, robustness, short analytical time and simplicity of sample preparation. Finally, the proposed method was applied to a bioequivalence study of self-made bilayer tablet and commercial formulation containing 500 mg of metformin and 1 mg of glimepi- ride in beagle dogs.

Stereoselective separation and determination of quizalofopethyl in tobacco by ultra-performance convergence chromatography with tandem mass spectrometry

The purpose of this study was to develop and validate a novel and sensitive method for simultaneous enantiomeric analysis of quizalofop-ethyl in tobacco using ultra-performance converge nee chromatography with tandem mass spectrometry(UPC-MS/MS).Sample preparation prior to UPC^2-MS/MS analysis followed a QuEChERS(quick,easy,cheap,effective,rugged and safe)method.The UPC^2-MS/MS method used an Acquity UPC2 Trefoil CEL2 column with a mobile phase of CO2 and methanol.The injection volume was 2 μL and run-time was 6 min with the mobile phase running at 2.0 mL/min flow rate.Column temperature,auto back pressure regulator pressure(ABPR),and modifier solvent were optimized for separation efficiency.Under the optimal conditio ns,the recoveries for all the ena ntiomers were 81.3%-94.8% with relative standard deviations(RSD)less than 5.0% at 0.1,1.0 and 2.0 mg/kg levels,respectively.Good coefficients of determination(R^2≥0.993 7)were achieved for all the studied enantiomers in the tobacco over the concentration range of 10-250 ng/mL.The limits of detection(LODs)varied from 0.02 mg/kg to 0.04 mg/kg,and the limits of quantification(LOQs)were ≤ 0.13 mg/kg.The results confirm that the proposed method is green,convenient and reliable for the enantioselective determination of the enantiomers of quizalofop-ethyl in tobacco.

Determination of urine catecholamines and metanephrines by reversed-phase liquid chromatography-tandem mass spectrometry

The measurement of urine catecholamine and metanephrine concentrations is important for biochemical screening and diagnosis of pheochromocytoma.The goal of this work was to develop a simple liquid chromatography-tandem mass spectrometry(LC-MS/MS)method for determining catecholamines and metanephrines in urine to replace an existing liquid chromatographic method using electrochemical detection.Urine samples were prepared using Oasis weak-cation-exchange cartridges.The eluate was analyzed on an Agilent ZORBAX Eclipse Plus Phenyl-Hexyl column in 3 min.Adrenaline,noradrenaline,dopamine,metanephrine,normetanephrine,and their deuterated internal standards were monitored in positive electrospray ionization mode by multiple reaction monitoring(MRM).No evidence of ion suppression was observed.The assay was linear up to 5μmol/L for adrenaline,5μmol/L for noradrenaline,6.1μmol/L for dopamine,5.6μmol/L for metanephrine,and 34.6μmol/L for normetanephrine,with lower limits of quantification of 5,5,12,6 and 7nmol/L,respectively.The intra-day and inter-day precisions for all analytes ranged from 0.59%to 4.64%and1.98%to 4.80%,respectively.External quality assurance samples were assayed and showed excellent agreement with the target values.This simple method provides an improved assay for determining urine catecholamines and metanephrines.

Uncertainty Evaluation of Determination of Microcystin MC-LR in Environmental Samples by Solid Phase Extraction-Ultra Performance Liquid Chromatography-Tandem Mass Spectrometry

To assess uncertainty of determination of MC-LR in environmental samples by solid phase extraction- ultra performance liquid chromatography- tandem mass spectrometry,the sources of the uncertainty were evaluated firstly,and the expanded uncertainty was calculated finally.The results show that when MC-LR concentration in the water samples was 0.50 μg/L,the expanded uncertainty was 0.00628 μg/L(k=2).

Development of an analytical method for multi-residue quantification of 18 anthelmintics in various animal-based food products using liquid chromatography-tandem mass spectrometry

In this study,we developed a simple screening procedure for the determination of 18 anthelmintics(including benzimidazoles,macrocyclic lactones,salicylanilides,substituted phenols,tetrahydropyrimidines,and imidazothiazoles)in five animal-derived food matrices(chicken muscle,pork,beef,milk,and egg)using liquid chromatography-tandem mass spectrometry.Analytes were extracted using acetonitrile/1% acetic acid(milk and egg)and acetonitrile/1% acetic acid with 0.5 mL of distilled water(chicken muscle,pork,and beef),and purified using saturated n-hexane/acetonitrile.A reversed-phase analytical column and a mobile phase consisting of(A)10 mM ammonium formate in distilled water and(B)methanol were used to achieve optimal chromatographic separation.Matrix-matched standard calibration curves(R^(2)≥0.9752)were obtained for concentration equivalent to ×1/2,×1,×2,×3,×4,and×5 fold the maximum residue limit(MRL)stipulated by the Korean Ministry of Food and Drug Safety.Recoveries of 61.2e118.4%,with relative standard deviations(RSDs)of ≤19.9%(intraday and interday),were obtained for each sample at three spiking concentrations(×1/2,×1,and ×2 the MRL values).Limits of detection,limits of quantification,and matrix effects were 0.02e5.5 mg/kg,0.06e10 mg/kg,and -98.8 to 13.9%(at 20 μg/kg),respectively.In five samples of each food matrix(chicken muscle,pork,beef,milk,and egg)purchased from large retailers in Seoul that were tested,none of the target analytes were detected.It has therefore been shown that this protocol is adaptable,accurate,and precise for the quantification of anthelmintic residues in foods of animal origin.

Trace-Level Analysis of Hexavalent Chromium in Lake Sediment Samples Using Ion Chromatography Tandem Mass Spectrometry

The analysis of hexavalent chromium, Cr(VI), in soil and sediment samples has been predominantly carried out in materials containing elevated levels. Reliable analysis of trace-level of Cr(VI) in sediment samples remains challenging. Cr(VI) analyses with multipoint calibration and speciated isotope dilution (SID) adapted from U.S. EPA method 6800 were used to measure lower-level Cr(VI) on an ion chromatograph coupled with a tandem mass spectrometer (IC-MS/MS). Lake sediment samples were collected from various locations in Northern Ontario and Cr(VI) was extracted using both alkaline digestion and ethylene diaminetetraacetic acid (EDTA) extraction. Certified reference materials were extracted and analyzed by IC-MS/MS and UV-VIS detection. The SID-MS approach allowed for the quantification of Cr(VI) in samples with concentration levels below 0.5 μg.g-1 wet weight.

Rapid Quantitative Determination of Isoprene Monomer in Living Taraxacum kok-saghyz by Ultra-High Performance Liquid Chromatography Tandem Mass Spectrometry

Taraxacum kok-saghyz(TKS)is rich in natural rubber(NR),a natural organic macromolecular compound composed of cis-1,4-polyisoprene,and may become the second NR-bearing plant for biochemical engineering development.In this paper,a rapid and quantitative ultra-high performance liquid chromatography tandem mass spectrometry(UHPLCMS/MS)method was established for determination of macromolecular biosynthesis substrate(dimethylallyl pyrophosphate,DMAPP)and initiator(farnesyl pyrophosphate,FPP)contained in TKS.A Kromasil C18 chromatographic column was used for separation,and the multi-reaction monitoring mode(MRM)of triple quadrupole mass spectrometry was used for detection.Quantification was performed by external calibration method.The results showed that the limit of detection(LOD)and the limit of quantitation(LOQ)of DMAPP were 2.42μg/L and 7.26μg/L,respectively,and the LOQ and the LOD of FPP were 1.02μg/L and 3.05μg/L,respectively.At a concentration of 1—1000μg/L,both analytes had good determination coefficients(>0.999)of calibration curve.The recoveries of DMAPP and FPP were between 99.0%and 117.1%.In real samples detection,the contents of DMAPP and FPP in TKS samples were between 23.32—82.77μg/L and 12.03—85.67μg/L,respectively.Thus,this approach is a reliable method to quantify DMAPP and FPP in TKS.

Development of a Liquid Chromatography-tandem Mass Spectrometry Method for Determination of Butoconazole Nitrate in Human Plasma and Its Application to a Pharmacokinetic Study

Summary： A liquid chromatography-tandem mass spectrometry （LC-MS/MS） method was developed and validated for the determination of butoconazole in human plasma. Human plasma samples of 0.2 μL were pretreated by a single step protein precipitation procedure and analyzed using a high performance liquid chromatography （HPLC） electrospray tandem mass spectrometer system. The compounds were eluted isocratically on an Inertsil ODS-SP column （100 min×2.1 mm, 3 μm）, ionized using a positive ion atmospheric pressure electrospray ionization source and analyzed using multiple reaction monitoring （MRM） mode. The ion transitions monitored were m/z 412.8→q65.1 for butoconazole and m/z 453.4→230.3 for the internal standard. The chromatographic run time was 3.5 min per injection, with retention time of 2.47 rain and 2.15 min for butoconazole and repaglinide, respectively. The method was validated to be linear over the range of 20 to 8000 pg/mL （r〉0.999） by using a weighted （1/x2） quadratic regression. The mean recovery rate was more than 86.7%, and the intra- and inter-day precision of the quality control samples （QCs） was less than 8.3% and the accuracy ranged from 96.0% to 110.2%, which indicated that the quantitative method was reliable and accurate. The method is simple, rapid, and has been applied successfully to a pharmacokinetics study of butoconazole nitrate suppositories in healthy Chinese females.

Application of Liquid Chromatography-Tandem Mass Spectrometry in Determination of Veterinary Drug Residues

With the long -term and irrational use of veterinary drugs and drug additives in the process of animal feeding, the veterinary drug resi- dues in animals and their additives can be ingested into the human body and pose potential threat to human health. Detection of veterinary drug resi- dues are extremely important practical significance to protect the ecological environment and human health. In recent years, liquid chromatography -tandem mass spectrometry has been widely used in vetednary drug residue detection because of its applicability, qualitative and quantitative fea- tures as well as high sensitivity. This article summarized the application of liquid chromatography -tandem mass spectrometry in detection veterinary drug residues from feed, animal products, arid udne as well as illicit drugs and harmful additives.

Stable Isotope Dilution Analysis of Gibberellin Residues in Tomato Paste by Liquid Chromatography-Tandem Mass Spectrometry

An accurate and sensitive method for the simultaneous determination of gibberellic acid（GA3）, gibberellin A4（GA4） and gibberellin A7（GA7） residues in tomato paste was developed by coupling solid phase extraction to high performance liquid chromatography-tandem mass spectrometry（LC-MS/MS） with electrospray ionization based stable isotope dilution analysis（SIDA）. The isotope labeled internal standard can compensate for the losses during the extraction and cleanup steps and for discrimination due to ion suppression. After extraction from methanol, hydrophile lipophilic balance（HLB） solid phase extraction（SPE） column was tested for the capacity of the cleanup of the tomato paste in compared with C18 SPE column which is the common way to the detection of GAs, and the former gained better result. Spiked experiments were performed in the non-contaminated tomato pastes and the recoveries of GA3, GA4 and GA7 were 42.6%―75.0% in external standard method（ESM） and 91.1%―103.8% in internal standard method（ISM） respectively. The validities of this method were investigated and good analytical performance for the three GAs was obtained, including low limits of method detection（2 ng/g for GA3 and GA4, 0.3 ng/g for GA7）, excellent linear dynamic ranges（5―500 ng/g for GA3 and GA4, 1―100 ng/g for GA7） and good relative standard deviation ranges（4.8%―9.4% for the intra-day test and 3.5%―11.9% for the inter-day test）.

Liquid Chromatography-Tandem Mass Spectrometry Assay to Detect Ethyl Glucuronide in Human Fingernail: Comparison to Hair and Gender Differences

Over the past decade, the use of hair specimens for the long-term detection of the alcohol biomarker ethyl glucuronide has been increasing in popularity and usage. We evaluated the usefulness of fingernail clippings as a suitable alterna-tive to hair for ethyl glucuronide detection. A liquid chromatography-tandem mass spectrometry method for the detection of ethyl glucuronide in fingernail clippings was fully validated and used to analyze the hair and/or fingernail specimens of 606 college-aged study participants. The limit of detection was 2 pg/mg, the limit of quantitation was 8 pg/mg and the method was linear from 8 to 2000 pg/mg. Intra- and inter-assay imprecision studies at three different concentrations (20, 40, 200 pg/mg) were all within 7.8% and all intra- and inter-assay bias studies at these levels were within 115.1% of target concentration. Ethyl glucuronide levels in fingernail (mean = 29.1 ± 55.6 pg/mg) were higher than ethyl glucuronide levels in hair (mean = 9.48 ± 22.3 pg/mg) and a correlation of the matched pairs was observed (r = 0.552, P < 0.01, n = 529). Evaluating each gender separately revealed that the correlation of male fingernail to male hair was large and significant (r = 0.782, P < 0.01, n = 195) while female hair to female fingernail was small yet sig-nificant (r = 0.249, P < 0.01, n = 334). The study results demonstrated that fingernail may be a suitable alternative to hair for ethyl glucuronide detection and may be the preferred sample type due to the lack of a gender bias.

Improved preparation and identification of aristolochic acid-DNA adducts by solid-phase extraction with liquid chromatography-tandem mass spectrometry

Aristolochic acid （AA） is a known nephrotoxin and potential carcinogen, which can form covalent DNA adducts after metabolic activation in vivo and in vitro. A simple method for preparation and characterization of aristolochic acid-DNA adducts was developed. Four AA-adducts were synthesized by a direct reaction of AAI/AAII with 2′-deoxynucleosides. The reaction mixture was first cleaned-up and pre-concentrated using solid phase extraction （SPE）, and further purified by a reversed-phase high performance liquid chromatography （HPLC）. By the application of developed SPE procedure, matrices and byproducts in reaction mixture could be greatly reduced and adducts of high purity （more than 94% as indicated by HPLC） were obtained. The purified AA-DNA adducts were identified and characterized with liquid-electrospray ionization-quadrupole-time of flight-mass spectrometry （LC-ESI-Q-TOF-MS/MS） and LC-Diode array detector-fluorescence （LC-DAD-FL） analysis. This work provides a robust tool for possible large-scale preparation of AA-DNA adduct standards, which can promote the further studies on carcinogenic and mutagenic mechanism of aristolochic acids.

Modernization of Chinese herbal compound and the high performance liquid chromatography tandem mass spectrometry (HPLC-MS)

Chinese herbal compound is playing an important role on curing human diseases.And it has been a trend that Chinese herbal compound is being used all over the world in 21 century.However,our Chinese herbal compound is facing serious challenge for the lack of canonical system of quality criterion for Chinese herbal compound so it has been a urgent problem to set up the quality control standards and reveal therapeutic basis of Chinese herbal compound.In order to give full play to the advantages of Chinese herbal compound,modern scientific and technological is used to research of Chinese herbal compound,especially the high performance liquid chromatography tandem mass spectrometry(HPLC-MS),because it is high sensitive,rapid,and obtain more information.It is very necessary that HPLC-MS is uesed to elucidate the effective components of basic substances of Chinese Herbal Compound,and endow traditional Chinese medicine with modern scientific connotation.

Determination of sterols in tobacco leaves by gas chromatography-tandem mass spectrometry

The purpose of the study was to establish an analytical method to simultaneously determine multiple sterols in tobacco leaves rapidly and accurately.In this gas chromatography-triple quadruple tandem mass spectrometry(GC-MS/MS)based method,various conditions for sterols extraction were assessed and the instrumental operation parameters were optimized with 5α-cholane as an internal standard.Using this method,eight plant sterols,namely lanosterol,campesterol,chenodeoxycholic acid,cholesterol,β-sitosterol,stigmasterol,dihydrotachysterol and ergosterol,were separated in less than 30 min.The linear correlation coefficients were over 0.998 9,the low detection limits were in the range of 0.010 3-0.141 4 μg/g.The recoveries were from 87.30% to 115.60%,with low standard deviations.In conclusion,this method demonstrates good repeatability,accuracy,and high sensitivity,and is best suited for rapid analysis of multiple sterols in tobacco leaves.

Studies on Fragmentation Pathways of N-Ethoxy(phenyl) phosphoryl Amino Acids by Electrospray Ionization Tandem Mass Spectrometry

The positive and negative ESI-MS/MS spectra of N-ethoxy（phenyl） phosphoryl amino acids（EPP-AA） were investigated by electrospray ionization（ESI） ion trap mass spectrometry. The fragmentation pathways of [ M ＋ Na]^＋ and [ M - H]^- ions are proposed and rationalized. The observation may have some potential applications in the interpretation of the MS/MS spectra of novel N-phosphoryl compounds. The complexity of MS/MS spectra of EPP-AA [ M ＋ Na]^＋ ions is decreased compared with that of N-dialkyloxyphosphoryl amino acid. Therefore, the new phosphonamidate method may be considered one of the superior methods that can be used in sequencing peptides and proteins extensively.

Studies on Fragmentation of Peptide by Nano-electrospray Ionization Fourier Transform Ion Cyclotron Resonance Multi-stage Tandem Mass Spectrometry

The sequence analysis of peptides was performed by nano-electrospray ionization Fourier transform ion cyclotron resonance tandem mass spectrometry（Nano-ESI-FT-ICR-MSn） and several peptides were chosen as examples. With the aid of the collision induced dissociation（CID）, FT-ICR provides not only precise mass/charge ratio, but also structure information of the selected peptides. The fragment ions were identified according to the observed molecular weights and peptide sequence was determined successfully. So Nano-ESI-FT-ICR-MSn is a useful tool for identification of the amino acid sequence of peptides with high confidence. Besides, a pathway for the dehydration of y ions without amino acids containing carboxylic acid under sustained off-resonance irradiation collision-induced dissociation（SORI-CID） condition was proposed.

Determination of Impurities and Degradation Products/Causes for <i>m</i>-Iodobenzylguanidine Using HPLC-Tandem Mass Spectrometry

m-Iodobenzylguanidinium hemisulfate (MIBGHS) is a precursor in the preparation of radioiodine-labeled m-iodobenzylguanidine (MIBG), which is used as a radio-imaging and therapy agent for neuroendocrine tumors and myocardial sympathetic nerve function. To ensure the quality and efficacy of the medicine and prevent side effects, the precursor purity, source of impurity, and derivatives have to be determined. In this study, the purity of synthesized MIBGHS and the amount of contaminants therein were determined by high-performance liquid chromatography and ultraviolet detection, gradient eluted by ammonium formate aqueous solution and acetonitrile mobile phase on both C8 and phenyl type column. The impurities were identified on the basis of molecular and fragmented ion mass spectra using of electrospray ionization triple quadrupole tandem mass spectrometry. The results revealed the presence of process-related impurities including m-bromobenzylguanidine (MBrBG) and overreacted byproducts. Stress test results indicated that MIBGHS is stable under acidic and dry thermal conditions for at least 72 h but MIBG aqueous solution was deteriorated slowly when exposed to UV light, thermal, oxidative and alkaline environments. Thus, m-iodobenzylamine, the starting material intended for the synthesis of MIBGHS should be analyzed to ensure that it is free from m-bromobenzylamine impurity. Stored under normal condition (-18°C), MIBGHS is stable for at least 12 months. The chemically labile guanidine and amine groups in MIBGHS are the major causes of its instability, while iodide loss from the phenyl group is a minor cause.