Solid-phase microextraction of endogenous metabolites from intact tissue validated using a Biocrates standard reference method kit

Improved analytical methods for the metabolomic profiling of tissue samples are constantly needed.Currently,conventional sample preparation methods often involve tissue biopsy and/or homogenization,which disrupts the endogenous metabolome.In this study,solid-phase microextraction(SPME)fibers were used to monitor changes in endogenous compounds in homogenized and intact ovine lung tissue.Following SPME,a Biocrates AbsoluteIDQ assay was applied to make a downstream targeted metabolomics analysis and confirm the advantages of in vivo SPME metabolomics.The AbsoluteIDQ kit enabled the targeted analysis of over 100 metabolites via solid-liquid extraction and SPME.Statistical analysis revealed significant differences between conventional liquid extractions from homogenized tissue and SPME results for both homogenized and intact tissue samples.In addition,principal component analysis revealed separated clustering among all the three sample groups,indicating changes in the metabolome due to tissue homogenization and the chosen sample preparation method.Furthermore,clear differences in free metabolites were observed when extractions were performed on the intact and homogenized tissue using identical SPME procedures.Specifically,a direct comparison showed that 47 statistically distinct metabolites were detected between the homogenized and intact lung tissue samples(P<0.05)using mixed-mode SPME fibers.These changes were probably due to the disruptive homogenization of the tissue.This study's findings highlight both the importance of sample preparation in tissue-based metabolomics studies and SPME's unique ability to perform minimally invasive extractions without tissue biopsy or homogenization while providing broad metabolite coverage.

Comparison of different approaches for direct coupling of solid-phase microextraction to mass spectrometry for drugs of abuse analysis in plasma

The direct coupling of solid-phase microextraction(SPME)to mass spectrometry(MS)(SPME-MS)has proven to be an effective method for the fast screening and quantitative analysis of compounds in complex matrices such as blood and plasma.In recent years,our lab has developed three novel SPME-MS techniques:SPME-microfluidic open interface-MS(SPME-MOI-MS),coated blade spray-MS(CBS-MS),and SPME-probe electrospray ionization-MS(SPME-PESI-MS).The fast and high-throughput nature of these SPME-MS technologies makes them attractive options for point-of-care analysis and anti-doping testing.However,all these three techniques utilize different SPME geometries and were tested with different MS instruments.Lack of comparative data makes it difficult to determine which of these methodologies is the best option for any given application.This work fills this gap by making a comprehensive comparison of these three technologies with different SPME devices including SPME fibers,CBS blades,and SPME-PESI probes and SPME-liquid chromatography-MS(SPME-LC-MS)for the analysis of drugs of abuse using the same MS instrument.Furthermore,for the first time,we developed different desorption chambers for MOI-MS for coupling with SPME fibers,CBS blades,and SPME-PESI probes,thus illustrating the universality of this approach.In total,eight analytical methods were developed,with the experimental data showing that all the SPME-based methods provided good analytical performance with R^(2)of linearities larger than 0.9925,accuracies between 81%and 118%,and good precision with an RSD%≤13%.

Comparison of Headspace Solid-Phase Microextraction with Simultaneous Steam Distillation Extraction for the Analysis of the Volatile Constituents in Chinese Apricot

Volatile constituents in fully mature fruits of apricot （Prunus armeniaca L.） cultivar Xinshiji were extracted using headspace solid-phase microextraction （HS-SPME） and simultaneous steam distillation extraction （SSDE） and then analyzed using capillary gas chromatography and gas chromatography-mass spectrometry. A total of 70 components were identified by HSSPME, including 20 esters, 19 hydrocarbons, 5 alcohols, 5 ketones, 4 acids, 4 lactones, 3 aldehydes, and 10 miscellaneous components, with the esters being the dominant constituent. On the basis of the odor unit values, it is believed that the following compounds probably contributed to the fresh apricot odor： hexyl acetate, β-ionone, butyl acetate, （E）-2-hexenal, linalool, limonene, γ-decalactone, and hexanal. A total of 49 components were also detected by SSDE, including 13 hydrocarbons, 9 alcohols, 7 aldehydes, 9 esters, 4 ketones, 4 lactones, 2 acids, and 1 miscellaneous component, of which the monoterpene alcohols were the dominant constituents. It could be judged from the odor unit values that the following compounds were the major contributors to boiled apricot aroma： β-ionone, linalool, hexyl acetate, γ-dodecalactone, γ- decalactone, （E）-2-hexenal, hexanal, γ-octalactone, phenylacetaldehyde, butyl acetate, limonene, α-terpineol, and δ-decalactone. The results show that HS-SPME is a simple, rapid, and solvent-free method, which is an alternative to the classical SSDE.

Preparation of polypyrrole/nanosilica composite for solid-phase microextraction of bisphenol and phthalates migrated from containers to eye drops and injection solutions

This paper describes the electrodeposition of polyphosphate-doped polypyrrole/nanosilica nano-composite coating on steel wire for direct solid-phase microextraction of bisphenol A and five phthalates. We optimized influencing parameters on the extraction efficiency and morphology of the nanocomposite such as deposition potential, concentration of pyrrole and polyphosphate, deposition time and the nanosilica amount. Under the optimized conditions, characterization of the nanocomposite was inves-tigated by scanning electron microscopy and Fourier transform infra-red spectroscopy. Also, the factors related to the solid-phase microextraction method including desorption temperature and time, extrac-tion temperature and time, ionic strength and pH were studied in detail. Subsequently, the proposed method was validated by gas chromatography-mass spectrometry by thermal desorption and acceptable figures of merit were obtained. The linearity of the calibration curves was between 0.01 and 50 ng/mL with acceptable correlation coefficients (0.9956-0.9987) and limits of detection were in the range 0.002-0.01 ng/mL. Relative standard deviations in terms of intra-day and inter-day by five replicate analyses from aqueous solutions containing 0.1 ng/mL of target analytes were in the range 3.3%-5.4% and 5%-7.1%, respectively. Fiber-to-fiber reproducibilities were measured for three different fibers prepared in the same conditions and the results were between 7.3% and 9.8%. Also, extraction recoveries at two different concentrations were ≥96%. Finally, the suitability of the proposed method was demonstrated through its application to the analysis of some eye drops and injection solutions.

Covalent organic nanospheres as a fiber coating for solid-phase microextraction of genotoxic impurities followed by analysis using GC-MS

Covalent organic nanospheres(CONs)were explored as a fiber coating for solid-phase microextraction of genotoxic impurities(GTIs)from active ingredients(AIs).CONs were synthesized by an easy solutionphase procedure at 25℃.The obtained nanospheres exhibited a high specific surface area,good thermostability,high acid and alkali resistance,and favorable crystallinity and porosity.Two types of GTIs,alkyl halides(1-iodooctane,1-chlorobenzene,1-bromododecane,1,2-dichlorobenzene,1-bromooctane,1-chlorohexane,and 1,8-dibromooctane)and sulfonate esters(methyl p-toluenesulfonate and ethyl ptoluenesulfonate),were chosen as target molecules for assessing the performance of the coating.The prepared coating achieved high enhancement factors(5097-9799)for the selected GTIs.The strong affinity between CONs and GTIs was tentatively attributed to π-π and hydrophobicity interactions,large surface area of the CONs,and size-matching of the materials.Combined with gas chromatography-mass spectrometry(GC-MS),the established analytical method detected the GTIs in capecitabine and imatinib mesylate samples over a wide linear range(0.2-200 ng/g)with a low detection limit(0.04-2.0 ng/g),satisfactory recovery(80.03%-109.5%),and high repeatability(6.20%-14.8%)and reproducibility(6.20%-14.1%).Therefore,the CON-coated fibers are promising alternatives for the sensitive detection of GTIs in AI samples.

Flash Evaporation and Headspace Solid-phase Microextraction for the Analysis of the Essential Oils in Traditional Chinese Medicine,Houttuynia Cordata Thunb

We have investigated the use of flash evaporation, headspace solid-phase microextraction (HS-SPME) and steam distillation (SD) as sample concentration and preparation techniques for the analysis of volatile constituents present in Houttuynia cordata Thunb. The samples were analyzed by gas chromatography (GC) and identified by mass spectrometry (MS). Comparison studies were performed. It was found that the results obtained between Headspace solid-phase microextraction HS-SPME and SD techniques were in good agreement. Seventy-nine compounds in Houttuynia cordata Thunb were identified by MS. In flash evaporation, thirty-nine compounds were identified. Discrimination in the response for many constituents studied was not observed, which can be clearly observed in SD and HS-SPME techniques. As a conclusion, HS-SPME is a powerful tool for determining the volatile constitutes present in the Houttuynia cordata.

Solid-phase Microextraction with Benzoxy-calix[6]arene Fiber Coupled to Gas Chromatography for the Analysis of Polycyclic Aromatic Hydrocarbons in Water

Headspace solid-phase microextraction（HS-SPME） with sol-gel calix[6]arene-containing fiber followed by gas chromatography with a flame ionization detector was used to examine the composition and distribution of seven polycyclic aromatic hydrocarbons（PAHs） in water. The novel SPME fiber exhibited higher extraction efficiency to PAHs compared with poly（dimethylsiloxane） and other calixarene-containing fibers. Extraction/retention mechanism based on the interactions between calixarenes and PAHs was discussed. Owing to the good selectivity and high extraction capability of this calixarene fiber, low detection limits were obtained in a range of 0.34―6.50 ng/L and the relative standard deviation values were ≤12.3% for all of the analytes. The linear ranges of the proposed method were five orders of magnitude for the tested compounds, with linear correlation coefficients（r） greater than 0.998. The method was applied to the determination of polycyclic aromatic hydrocarbons in nine water sources in Wuhan City, China. Standard addition method was selected for the quantification and the recovery values were in a satisfactory range. Total PAHs concentrations in the nine surface water samples were found to vary between undetectable and 8.840 μg/L with two- and three-ring PAHs predominating.

Sol-Gel-Coated Calix[4]arene Fiber for Solid-Phase Microextraction

5, 11, 17, 23-tetra-tert-butyl-25, 27-diethoxy-26, 28-dihydroxycalix \ arene /hydroxy-terminated silicone oil coated fiber was first prepared and applied for solid-phase microextraction with sol-gel technology. The properties of the new coating were investigated by analysis of benzene derivatives, polycyclic aromatic hydrocarbons and aromatic amines. The fiber is characterized by good sensitivity and selectivity to these aromatics. The fiber has stable performance at a high temperature of 380℃ and after a long solvent immersing, thus its lifetime is very long. It shows good fiber-to-fiber and batch-to-batch reproducibility. Furthermore, broad linear ranges and low detection limits are also its characteristics.

Preparation of A New Fiber by Sol-gel Technology in Solid-phase Microextraction (SPME)

The sol-gel technology is applied for the preparation of solid-phase microextraction (SPME) fiber. The fiber demonstrates high thermal stability, efficient extraction rate and the selectivity for non-polar or low-polar analytes. Efficient SPME-GC-FID analyses of benzene- toluene-ethylbenzene-xylenes (BTEXs) and low-polar halocarbon were achieved by the sol-gel coated DSDA-DDBT-TiO2 fiber. Some parameters of the SPME fiber for the determination of halocarbon in aqueous sample were investigated.

Time-course monitoring of in vitro biotransformation reaction via solid-phase microextraction-ambient mass spectrometry approaches

The solid-phase microextraction technique quantifies analytes without considerably affecting the sample composition.Herein,a proof-of-concept study was conducted to demonstrate the use of coated probe electrospray ionization(coated-PESI)and coated blade spray(CBS)as ambient mass spectrometry approaches for monitoring drug biotransformation.The ability of these methods was investigated for monitoring the dephosphorylation of a prodrug,combretastatin A4 phosphate(CA4P),into its active form,combretastatin A4(CA4),in a cell culture medium supplemented with fetal bovine serum.The CBS spot analysis was modified to achieve the same extraction efficiency as protein precipitation and obtained results in 7 min.Because coated-PESI performs extraction without consuming any samples,it is the preferred technique in the case of a limited sample volume.Although coated-PESI only extracts small quantities of analytes,it uses the desorption solvent volume of 5-10 pL,resulting in high sensitivity,thus allowing the detection of compounds after only 1 min of extraction.The biotransformation of CA4P into CA4 via phosphatases occurs within the simple matrix,and the proposed sample preparation techniques are suitable for monitoring the biotransformation.

Application of Phenyl Bonded Mesoporous Silica as A Novel Coating Layer of Solid-phase Microextraction for Determination of Benzo[a]pyrene in Water Samples

Phenyl bonded mesoporous silica (C6H5-MCM-41) was applied as the fiber coating of solid-phase microextraction (SPME). The performance of the fiber coating was discussed coupling to HPLC. Applicability of mesoporous fiber coating was examined for the determination of benzo[a]pyrene (B[a]P) in water samples. The limit of detection (LOD) is 0.28μg·L-1. Good recovery and relative standard deviation (RSD) were obtained.

Solid-phase microextraction for flavor analysis in Harari Khat （Catha edulis） stimulant

This research examined the typical flavor compounds in the commonest type of Khat called Harari Khat grown in the region of Ethiopia.Twenty-eight compounds, which includes 1,2-Propanedione,1-Phenyl,Hexanol,Hexanal compounds,Limonene, Benzaldehyde with other flavors, were extracted by polydimethylsiloxane at room temperature for 30min from Khat samples,and identified by solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS).This method needs no organic solvents and required minimal sample.

Determination of Trihalomethanes in Water Samples Using Headspace Solid-Phase Microextraction Gas Chromatography

The chlorination process is one of the water treatment method used for the disinfection of water. The disinfection by products are trihalomethanes such as chloroform, dichloromethane, dibromochloromethane and bromoform. A headspace solid-phase microextraction method has been developed for determination oftrihalomethanes in water samples. The experimental parameters such as the stirring rate, extraction time, extraction temperature and desorption time were investigated. The linearity, detection limits and percentage recovery were evaluated. The optimum conditions were stirring rate 800 rpm/min, extraction time 6 min, extraction temperature 20 ~C, desorption time 2.5 min and desorption temperature 220 ~C. The detection limits were 0.01 ~g/L and the recoveries were in the range of 86-110 %, The proposed method was successfully applied to determination of THM4 in tap water samples. The THM4 contents were varied depending on the sample sites and the season. The total THM4 contents in cool, summer and rainy season were in the range of 27.58-41.89, 32.06-60.73 and 46.26-69.87 p.g/L, respectively. Confirmation of the detected compounds in water samples were performed by gas chromatograph-mass spectrometer. The mass spectra of the target compounds in water samples is in good agreement with trihalomethanes standard spectra.

Determination of Aroma Composition of Santalum album Linn by Solid-phase Microextraction-Gas Chromatography-Mass Spectrometry

[Objectives]This study aimed to determine the volatile components in Santalum album Linn and gradually clarify the aroma composition of S.album Linn.[Methods]Solid-phase microextraction method was used to obtain the volatile components of S.album Linn.The aroma components were analyzed by gas chromatography-mass spectrometry and their relative contents were calculated using the area normalization method.[Results]In a dry state at room temperature,39 chemical components were identified from S.album Linn,mainly olefins(91.15%),alkanes(3.00%),alcohols(2.56%),esters(2.19%),ketones(0.55%),aldehydes(0.41%)and heterocyclics(0.14%).[Conclusions]This method has the advantages of low sample consumption,easy operation,rapid identification of aroma components and high sensitivity,and can effectively separate and determine volatile components in S.album Linn,realizing the rapid identification of different S.album Linn varieties and providing technical support for further research on Chinese medicinal materials.

In vivo solid phase microextraction for therapeutic monitoring and pharmacometabolomic fingerprinting of lung during in vivo lung perfusion of FOLFOX

In vivo lung perfusion(IVLP)is a novel isolated lung technique developed to enable the local,in situ administration of high-dose chemotherapy to treat metastatic lung cancer.Combination therapy using folinic acid(FOL),5-fluorouracil(F),and oxaliplatin(OX)(FOLFOX)is routinely employed to treat several types of solid tumours in various tissues.However,F is characterized by large interpatient variability with respect to plasma concentration,which necessitates close monitoring during treatments using of this compound.Since plasma drug concentrations often do not reflect tissue drug concentrations,it is essential to utilize sample-preparation methods specifically suited to monitoring drug levels in target organs.In this work,in vivo solid-phase microextraction(in vivo SPME)is proposed as an effective tool for quantitative therapeutic drug monitoring of FOLFOX in porcine lungs during pre-clinical IVLP and intravenous(IV)trials.The concomitant extraction of other endogenous and exogenous small molecules from the lung and their detection via liquid chromatography coupled to high resolution mass spectrometry(LC-HRMS)enabled an assessment of FOLFOX's impact on the metabolomic profile of the lung and revealed the metabolic pathways associated with the route of administration(IVLP vs.IV)and the therapy itself.This study also shows that the immediate instrumental analysis of metabolomic samples is ideal,as long-term storage at80℃ results in changes in the metabolite content in the sample extracts.

Pre-concentration and determination of amitriptyline residues in waste water by ionic liquid based immersed droplet microextraction and HPLC

This paper describes a new approach for the determination of amitriptyline in wastewater by ionic liquid based immersed droplet microextraction （IL-IDME） prior to highperformance liquid chromatography with ultraviolet detection. 1-Hexyl-3-methylimidazolium hexafluorophosphate （[C6MIM][PF6]） was used as an ionic liquid. Various factors that affect extraction, such as volume of ionic liquid, stirring rate, extraction time, pH of the aqueous solution and salting effect, were optimized. The optimal conditions were as follows： microextraction time, 10 min; stirring rate, 720 rpm; pH, 11; ionic drop volume, 100 uL; and no sodium chloride addition. In quantitative experiments the method showed linearity in a range from 0.01 to 10 ug/mL, a limit of detection of 0.004 ug/mL and an excellent pre-concentration factor （PF） of 1100. Finally, the method was successfully applied to the determination of amitriptyline in the hospital wastewater samples.

Integration of GC-MS Based Non-Targeted Metabolic Profiling with Headspace Solid Phase Microextraction Enhances the Understanding of Volatile Differentiation in Tobacco Leaves from North Carolina, India and Brazil

In this report, gas chromatography-mass spectrometry (GC-MS) based non-targeted metabolomics is used to develop appropriate headspace solid phase microextractions (HS-SPME) to enhance the understanding of volatile complexity of flue-cured tobacco leaves. Non-targeted metabolic profiling of GC-MS shows that the extraction condition of HS-SPME at 100?C for 30 min provides a better metabolite profile than other extraction conditions tested. GC-MS and principal component analyses (PCA) show that among five types of fibers tested, 100 μm polydimethylsiloxane (PMDS), 65 μm polydimethylsiloxane/divinylbenzene (PMDS/DVB) and 75 μm carboxen/polydimethylsiloxane (CAR/ PMS) provide a better reproducible metabolite profile. Based on an appropriate PDMS extraction condition optimized, we use GC-MS analysis and PCA to compare metabolite profiles in flue-cured leaves of tobacco plants grown in North Carolina, India and Brazil, respectively. The resulting data of PCA show that the global metabolic profiles in North Carolina samples are separated from those in Brazil and India samples, two groups of which are characterized by a partially overlapped pattern. Several peaks that were differentially accumulated in samples were annotated to known metabolites by deconvolution analysis, such as norsolanadione, solavetivone and rishtin. Norsolanadione is detected only in Brazil samples. Solavetivone is detected in samples of India and Brazil but not in those of North Carolina. Rishtin is detected in samples of North Carolina and India but not in Brazil samples. These data indicate that not only can a non-targeted metabolic profiling approach enhance the understanding of volatile complexity, but also can identify marker volatile metabolites in tobacco leaves produced in different growth regions.

Modifying current thin-film microextraction(TFME)solutions for analyzing prohibited substances:Evaluating new coatings using liquid chromatography

For identifying and quantifying prohibited substances,solid-phase microextraction(SPME)continues to arouse interest as a sample preparation method.However,the practical implementation of this method in routine laboratory testing is currently hindered by the limited number of coatings compatible with the ubiquitous high-performance liquid chromatography(HPLC)systems.Only octadecyl(C18)and polydimethylsiloxane/divinylbenzene ligands are currently marketed for this purpose.To address this situation,the present study evaluated 12 HPLC-compatible coatings,including several chemistries not currently used in this application.The stationary phases of SPME devices in the geometry of thin filmcoated blades were prepared by applying silica particles bonded with various functional ligands(C18,octyl,phenyl-hexyl,3-cyanopropyl,benzenesulfonic acid,and selected combinations of these),as well as unbonded silica,to a metal support.Most of these chemistries have not been previously used as microextraction coatings.The 48 most commonly misused substances were selected to assess the extraction efficacy of each coating,and eight desorption solvent compositions were used to optimize the desorption conditions.All samples were analyzed using an HPLC system coupled with triple quadrupole tandem mass spectrometry.This evaluation enables selection of the best-performing coatings for quantifying prohibited substances and investigates the relationship between extraction efficacy and the physicochemical characteristics of the analytes.Ultimately,using the most suitable coatings is essential for trace-level analysis of chemically diverse prohibited substances.

直接浸入式固相微萃取-气相色谱法同时测定调味品中的山梨酸、苯甲酸和脱氢乙酸含量

目的建立直接浸入固相微萃取结合气相色谱法同时测定调味品中3种常见防腐剂(山梨酸、苯甲酸、脱氢乙酸)含量的分析方法。方法本研究对直接浸入固相微萃取的多个关键参数,包括萃取温度、萃取时间和盐酸添加量,进行了系统优化,以寻求最佳的萃取效率。采用气相色谱仪进行测定,并利用外标法进行定量分析。对建立的直接浸入固相微萃取结合气相色谱法进行了全面的方法学评价,并实际应用于调味品样品的检测。结果3种防腐剂在标准曲线范围内均呈现出优异的线性关系,相关系数(r)≥0.9960,方法的检出限为0.08~0.96 mg/L,定量限为0.25~3.20 mg/L,回收率为92.8%~106.3%。结论该方法操作简便、灵敏度高、结果准确可靠,为同时检测调味品中山梨酸、苯甲酸、脱氢乙酸含量提供了有效技术手段,在食品安全检测领域具有一定的应用价值和推广前景。

Mapping the metabolic responses to oxaliplatin-based chemotherapy with in vivo spatiotemporal metabolomics

Adjuvant chemotherapy improves the survival outlook for patients undergoing operations for lung metastases caused by colorectal cancer (CRC). However, a multidisciplinary approach that evaluates several factors related to patient and tumor characteristics is necessary for managing chemotherapy treatment in metastatic CRC patients with lung disease, as such factors dictate the timing and drug regimen, which may affect treatment response and prognosis. In this study, we explore the potential of spatial metabolomics for evaluating metabolic phenotypes and therapy outcomes during the local delivery of the anticancer drug, oxaliplatin, to the lung. 12 male Yorkshire pigs underwent a 3 h left lung in vivo lung perfusion (IVLP) with various doses of oxaliplatin (7.5, 10, 20, 40, and 80 mg/L), which were administered to the perfusion circuit reservoir as a bolus. Biocompatible solid-phase microextraction (SPME) microprobes were combined with global metabolite profiling to obtain spatiotemporal information about the activity of the drug, determine toxic doses that exceed therapeutic efficacy, and conduct a mechanistic exploration of associated lung injury. Mild and subclinical lung injury was observed at 40 mg/L of oxaliplatin, and significant compromise of the hemodynamic lung function was found at 80 mg/L. This result was associated with massive alterations in metabolic patterns of lung tissue and perfusate, resulting in a total of 139 discriminant compounds. Uncontrolled inflammatory response, abnormalities in energy metabolism, and mitochondrial dysfunction next to accelerated kynurenine and aldosterone production were recognized as distinct features of dysregulated metabolipidome. Spatial pharmacometabolomics may be a promising tool for identifying pathological responses to chemotherapy.