An analytical approach for on-site analysis of breath samples for Δ9-tetrahydrocannabinol (THC).

Increased acceptance of cannabis containing the psychoactive component, Δ9-tetrahydrocannabinol (THC), raises concerns about the potential for impaired drivers and increased highway accidents. In contrast to the "breathalyzer" test, which is generally accepted for determining the alcohol level in a driver, there is no currently accepted roadside test for THC in a motorist. There is a need for an easily collectible biological sample from a potentially impaired driver coupled with an accurate on-site test to measure the presence and quantity of THC in a driver. A novel breath collection device is described, which includes three separate sample collectors for collecting identical A, B, and C breath samples from a subject. A simple one-step ethanol extraction of the "A" breath collector sample can be analyzed by UHPLC/selected ion monitoring (SIM) liquid chromatography/mass spectrometry (LC/MS) to provide qualitative and quantitative determination of THC in breath sample in less than 4 min for samples collected up to 6 h after smoking a cannabis cigarette. SIM LC/MS bioanalyses employed d3-THC as the stable isotope internal standard fortified in negative control breath samples for quantitation including replicates of six calibrator standards and three quality control (QC) samples. Subsequent confirmation of the same breath sample in the B collectors was then confirmed by a reference lab by LC/MS/MS analysis. Fit-for-purpose bioanalytical validation consistent with pharmaceutical regulated bioanalyses produced pharmacokinetic (PK) curves for the two volunteer cannabis smokers. These results produced PK curves, which showed a rapid increase of THC in the breath of the subjects in the first hour followed by reduced THC levels in the later time points. A simpler single-point calibration curve procedure with calibrators and QC prepared in ethanol provided similar results. Limitations to this approach include the higher cost and operator skill sets for the instrumentation employed and the inability to actually determine driver impairment.

Key quality control aspects about cannabinoid-rich hemp products that a veterinarian needs to know: a practitioner's guide.

There is considerable confusion in the veterinary profession surrounding the rise in hemp cannabidiol-based animal products and what veterinarians should know before discussing these products with clients. There is emerging evidence for the potential use in case management across many veterinary indications; however, the cannabinoid concentrations and whether these are isolated cannabinoids or whole hemp extracts is difficult to elucidate, even from the published papers. Much like any extract from a plant, there are multiple considerations including quality control, pharmacokinetics in the intended species, microbiological and chemical contamination, and product consistency-all aspects that should be considered before a conversation can begin with a client. The aim of this review is to help practitioners make informed decisions and better facilitate discussions with clients for companion animals kept as pets. This review will not address food animal issues, as established withholding times have yet to be fully researched.

Intact Protein Mass Spectrometry for Therapeutic Protein Quantitation, Pharmacokinetics, and Biotransformation in Preclinical and Clinical Studies: An Industry Perspective.

Recent advancements in immunocapture methods and mass spectrometer technology have enabled intact protein mass spectrometry to be applied for the characterization of antibodies and other large biotherapeutics from in-life studies. Protein molecules have not been traditionally studied by intact mass or screened for catabolites in the same manner as small molecules, but the landscape has changed. Researchers have presented methods that can be applied to the drug discovery and development stages, and others are exploring the possibilities of the new approaches. However, a wide variety of options for assay development exists without clear recommendation on best practice, and data processing workflows may have limitations depending on the vendor. In this perspective, we share experiences and recommendations for current and future application of mass spectrometry for biotherapeutic molecule monitoring from preclinical and clinical studies.

The Role of Mass Spectrometry in the Cannabis Industry.

The focus of this critical insight article is a brief overview of analytical challenges the cannabis industry faces and how analytical chemists have new opportunities to demonstrate the merits of employing mass spectrometry for the chemical analysis of cannabis and its products. The current range of cannabis products extends from recreational use to medicines, edibles, beverages, and beyond. The standards employed to assure product quality, integrity, and safety are lacking compared to those currently used by the pharmaceutical, food, and beverage industries. This manuscript overviews some of the important analytical issues that exist for the growth and harvest of the cannabis plant to the production of a wide variety of its products. Currently, the topics of interest for safety in cannabis testing where mass spectrometry can play an important role include what are currently referred to as potency, pesticides, terpenes, heavy metals, and mycotoxins from bacteria. Since each state in the USA as well as several countries has their own regulations, the analytical opportunities and challenges vary depending upon which jurisdiction a laboratory is supporting. This Critical Insight report will suggest where mass spectrometry can play an important role and provide valuable input on these topics. Graphical Abstract.

Top-down LC-MS quantitation of intact denatured and native monoclonal antibodies in biological samples.

AIM: The requirements for developing antibody biotherapeutics benefit from understanding the nature and relevant aspects of the entire molecule. The method presented herein employs on-line multidimensional LC-quadrupole time-of-flight (QTOF)-MS for the quantitative determination of an antibody isolated from biological samples while maintaining the intact native biologically active conformation of the antibody. RESULTS: Following method optimization for a model antibody, an incurred biotherapeutic in cynomologus monkey was quantified in its intact top-down native conformation. A partial method validation demonstrated acceptable precision and accuracy although improved sensitivity requires further studies. CONCLUSION: An on-line multidimensional LC-MS approach presents a proof-of-principle example for quantifying an intact, native antibody isolated from an incurred biological sample via immunoaffinity techniques coupled with top-down QTOF LC-MS bioanalysis.

A Book-Type Dried Plasma Spot Card for Automated Flow-Through Elution Coupled with Online SPE-LC-MS/MS Bioanalysis of Opioids and Stimulants in blood.

Despite many benefits including simple point-of-care sample collection, reduced costs, and simplified shipping and storage, dried blood spot (DBS) techniques have faced adoption resistance due to factors such as the hematocrit (Hct) effects and the established preference for bioanalysis of plasma rather than whole blood. One way to potentially circumvent these challenges is to adopt the concept of dried plasma spot (DPS) techniques. One approach to accomplishing this is through an on-card red blood cell (RBC) filtration to generate plasma from whole blood without the need for centrifugation. In this report, a book-type DPS card has been developed and validated by employing fully automated flow-through elution coupled with online SPE-LC-ESI-MS/MS for the quantitative determination of four representative opioids (morphine, codeine, oxycodone, hydrocodone) and five stimulants (amphetamine, methamphetamine, 3,4-methylenedioxymethamphetamine (MDMA), phentermine, and mephedrone) in one method using their corresponding deuterium-labeled analogues as internal standards. Method validation results showed good linearity (R2 ≥ 0.9963) ranging from 5 to 1000 ng/mL. Intraday and interday precision and accuracy were within the acceptable limits at four quality control (QC) levels. Extraction recovery was ≥87.9% at both the lower limit of quantitation (LLOQ) and the upper limit of quantitation (ULOQ) along with acceptable selectivity and sensitivity. DPS on-card short-term stability was compound-dependent and storage-dependent. The additional benefits of the validated book-type DPS card include a wider applicability range of Hct (30% to 60%), automated online analysis compatibility, and a higher plasma volume yield.

Hematocrit-Independent Quantitation of Stimulants in Dried Blood Spots: Pipet versus Microfluidic-Based Volumetric Sampling Coupled with Automated Flow-Through Desorption and Online Solid Phase Extraction-LC-MS/MS Bioanalysis.

A workflow overcoming microsample collection issues and hematocrit (HCT)-related bias would facilitate more widespread use of dried blood spots (DBS). This report describes comparative results between the use of a pipet and a microfluidic-based sampling device for the creation of volumetric DBS. Both approaches were successfully coupled to HCT-independent, fully automated sample preparation and online liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) analysis allowing detection of five stimulants in finger prick blood. Reproducible, selective, accurate, and precise responses meeting generally accepted regulated bioanalysis guidelines were observed over the range of 5-1000 ng/mL whole blood. The applied heated flow-through solvent desorption of the entire spot and online solid phase extraction (SPE) procedure were unaffected by the blood's HCT value within the tested range of 28.0-61.5% HCT. Enhanced stability for mephedrone on DBS compared to liquid whole blood was observed. Finger prick blood samples were collected using both volumetric sampling approaches over a time course of 25 h after intake of a single oral dose of phentermine. A pharmacokinetic curve for the incurred phentermine was successfully produced using the described validated method. These results suggest that either volumetric sample collection method may be amenable to field-use followed by fully automated, HCT-independent DBS-SPE-LC-MS/MS bioanalysis for the quantitation of these representative controlled substances. Analytical data from DBS prepared with a pipet and microfluidic-based sampling devices were comparable, but the latter is easier to operate, making this approach more suitable for sample collection by unskilled persons.

Determination of emerging nitrogenous economic adulterants in milk proteins by high-performance liquid chromatography/compact mass spectrometry.

RATIONALE: Milk-derived ingredients are widely used around the world in the manufacturing of nutritional products. They are prone to economically motivated adulteration with nitrogenous compounds such as melamine and its analogs in order to increase the nitrogen content of these ingredients. The need to rapidly screen milk-derived ingredients to detect adulteration is of paramount public health concern. A liquid chromatography/mass spectrometry (LC/MS)-based method using a single quadrupole mass spectrometer has been developed for the rapid frontline analysis of six nitrogenous protein adulterants, i.e. melamine, ammeline, ammelide, amidinourea, cyromazine and cyanuric acid, in three key milk-derived ingredients, i.e. whole milk powder, nonfat milk powder and whey protein concentrate. METHODS: The sample preparation scheme involves both 'dilute and shoot' as well as solid-phase extraction (SPE)-based methods. The 'dilute and shoot' scheme uses a tenfold dilution of sample with water followed by protein precipitation using 2% formic acid in acetonitrile. The SPE scheme involves tenfold dilution of sample with water, followed by protein precipitation using acetonitrile, and further cleanup through Strata Melamine SPE cartridges. Sample extracts were analyzed by hydrophilic interaction chromatography/single quadrupole mass spectrometry (HILIC/MS) in both positive and negative electrospray ionization mode. Accurate quantitation was achieved using stable isotope labeled internal standards. RESULTS: A multi-day method validation study was conducted using three different milk-derived ingredients. Average accuracies, relative standard deviations (RSD) and method detection limits (MDL) for all analytes in whole milk powder were 65-118%, 7-11% and 0.9-30 mg/kg, using the 'dilute and shoot' extraction procedure. The SPE procedure results were 102-111%, 5-13%, and 0.4-2.5 mg/kg, respectively, for melamine, ammeline, ammelide and cyromazine only. CONCLUSIONS: A rugged and simple to use analytical method to screen for the presence of nitrogenous economic adulterants in milk-derived ingredients has been developed for routine frontline use. Copyright © 2016 John Wiley & Sons, Ltd.

Quantitative determination of opioids in whole blood using fully automated dried blood spot desorption coupled to on-line SPE-LC-MS/MS.

Opioids are well known, widely used painkillers. Increased stability of opioids in the dried blood spot (DBS) matrix compared to blood/plasma has been described. Other benefits provided by DBS techniques include point-of-care collection, less invasive micro sampling, more economical shipment, and convenient storage. Current methodology for analysis of micro whole blood samples for opioids is limited to the classical DBS workflow, including tedious manual punching of the DBS cards followed by extraction and liquid chromatography-tandem mass spectrometry (LC-MS/MS) bioanalysis. The goal of this study was to develop and validate a fully automated on-line sample preparation procedure for the analysis of DBS micro samples relevant to the detection of opioids in finger prick blood. To this end, automated flow-through elution of DBS cards was followed by on-line solid-phase extraction (SPE) and analysis by LC-MS/MS. Selective, sensitive, accurate, and reproducible quantitation of five representative opioids in human blood at sub-therapeutic, therapeutic, and toxic levels was achieved. The range of reliable response (R(2) ≥0.997) was 1 to 500 ng/mL whole blood for morphine, codeine, oxycodone, hydrocodone; and 0.1 to 50 ng/mL for fentanyl. Inter-day, intra-day, and matrix inter-lot accuracy and precision was less than 15% (even at lower limits of quantitation (LLOQ) level). The method was successfully used to measure hydrocodone and its major metabolite norhydrocodone in incurred human samples. Our data support the enormous potential of DBS sampling and automated analysis for monitoring opioids as well as other pharmaceuticals in both anti-doping and pain management regimens.

Novel membrane devices and their potential utility in blood sample collection prior to analysis of dried plasma spots.

BACKGROUND: Construction and application of a novel membrane substrate dried plasma spot (DPS) card is described. Results/methodology: The online automation compatible prototype DPS card employs a membrane filter to remove red blood cells from whole blood for the collection of plasma. The described autoDPS card provides acceptable quantitative precision and accuracy data from plasma filtered from 45% hematocrit whole blood. When significantly lower or higher hematocrit (30 and 60%) whole blood fortified with guanfacine is applied, the quantitative precision and accuracy data fall outside regulated 15/20 bioanalytical acceptance criteria. CONCLUSION: The described prototype DPS card works well for normal hematocrit whole blood, but further development is needed for samples of much lower or higher hematocrit.

Automated high-capacity on-line extraction and bioanalysis of dried blood spot samples using liquid chromatography/high-resolution accurate mass spectrometry.

RATIONALE: Pharmacokinetic data to support clinical development of pharmaceuticals are routinely obtained from liquid plasma samples. The plasma samples require frozen shipment and storage and are extracted off-line from the liquid chromatography/tandem mass spectrometry (LC/MS/MS) systems. In contrast, the use of dried blood spot (DBS) sampling is an attractive alternative in part due to its benefits in microsampling as well as simpler sample storage and transport. However, from a practical aspect, sample extraction from DBS cards can be challenging as currently performed. The goal of this report was to integrate automated serial extraction of large numbers of DBS cards with on-line liquid chromatography/high-resolution accurate mass spectrometry (LC/HRAMS) bioanalysis. METHODS: An automated system for direct DBS extraction coupled to a LC/HRAMS was employed for the quantification of midazolam (MDZ) and α-hydroxymidazolam (α-OHMDZ) in human blood. The target analytes were directly extracted from the DBS cards onto an on-line chromatographic guard column followed by HRAMS detection. No additional sample treatment was required. The automated DBS LC/HRAMS method was developed and validated, based on the measurement at the accurate mass-to-charge ratio of the target analytes to ensure specificity for the assay. RESULTS: The automated DBS LC/HRAMS method analyzed a DBS sample within 2 min without the need for punching or additional off-line sample treatment. The fully automated analytical method was shown to be sensitive and selective over the concentration range of 5 to 2000 ng/mL. Intra- and inter-day precision and accuracy was less than 15% (less than 20% at the LLOQ). The validated method was successfully applied to measure MDZ and α-OHMDZ in an incurred human sample after a single 7.5 mg dose of MDZ. CONCLUSIONS: The direct DBS LC/HRAMS method demonstrated successful implementation of automated DBS extraction and bioanalysis for MDZ and α-OHMDZ. This approach has the potential to promote workload reduction and sample throughput increase.

Automated direct extraction and analysis of dried blood spots employing on-line SPE high-resolution accurate mass bioanalysis.

BACKGROUND: Online automated extraction of dried blood spots (DBS) via direct extraction to a solid-phase extraction (SPE) cartridge and bioanalysis by high-resolution accurate mass spectrometry was examined. The methodology was validated and used to investigate the effect of hematocrit on assay bias using partial and whole spot extractions from accurately dispensed blood samples. RESULTS: The completed analysis of a DBS sample was accomplished within 2 to 3 min using the online DBS-SPE platform. Hematocrit related bias was observed (>15%) for the partial DBS extractions, but not when the whole DBS was eluted. CONCLUSION: RESULTS demonstrate successful implementation of automated online DBS-SPE high-resolution accurate mass spectrometry analysis and the remediation of hematocrit bias using a capillary micro dispenser for accurate spotting of blood samples.

Fully-automated approach for online dried blood spot extraction and bioanalysis by two-dimensional-liquid chromatography coupled with high-resolution quadrupole time-of-flight mass spectrometry.

An integrated automated approach has been developed for the direct determination of drug concentrations using a SCAP DBS system for online extraction and analysis of dried blood spots (DBS) from DBS paper cards to a multidimensional liquid chromatography system coupled to a high-resolution QTOF mass spectrometry (LC-HRMS). An accurate, precise, selective, and sensitive two-dimensional liquid chromatography-high-resolution mass spectrometry (2D LC-HRMS) assay was developed and validated using small volumes of rat blood (approximately 1.25 µL) from a 2 mm DBS punch. The methodology was validated according to internationally accepted regulated bioanalysis acceptance criteria in order to establish the validity of the combination of online DBS assay and use of HRMS for quantitative bioanalysis. The fully automated procedure exhibited acceptable linearity (r(2) > 0.997) over the concentration range of 5 to 1000 ng/mL. Intra- and interday precision and accuracy runs indicated relative errors less than 20% at the LLOQ level and less than 15% at all other levels. The direct extraction and analysis of DBS samples resulted in a 5-fold improvement in assay sensitivity compared to conventional off-line extraction of punched DBS samples. In addition, the impact of blood hematocrit (Hct) on accurate quantification of the studied drugs also was evaluated, comparing Hct values of 30% and 60% against standards prepared at 45%. Hematocrit experiments show that Hct can influence the accuracy of drugs quantified by DBS and needs to be thoroughly evaluated prior to committing to validating a DBS assay. The online DBS system coupled to the LC-HRMS was then successfully applied to a pharmacokinetic study performed on male Sprague-Dawley rats after administration of a single dose of 5 and 10 mg/kg for midazolam and desipramine, respectively.

Microsample analyses via DBS: challenges and opportunities.

The use of DBS is an appealing approach to employing microsampling techniques for the bioanalysis of samples, as has been demonstrated for the past 50 years in the metabolic screening of metabolites and diseases. In addition to its minimally invasive sample collection procedures and its economical merits, DBS microsampling benefits from the very high sensitivity, selectivity and multianalyte capabilities of LC-MS, which has been especially well demonstrated in newborn screening applications. Only a few microliters of a biological fluid are required for analysis, which also translates to significantly reduced demands on clinical samples from patients or from animals. Recently, the pharmaceutical industry and other arenas have begun to explore the utility and practicality of DBS microsampling. This review discusses the basis for why DBS techniques are likely to be part of the future, as well as offering insights into where these benefits may be realized.

Semi-automated direct elution of dried blood spots for the quantitative determination of guanfacine in human blood.

BACKGROUND: Direct analysis of dried blood spot (DBS) samples was investigated using a prototype semi-automated robotic device that allows the direct elution of sample spots from a DBS paper card to an online SPE cartridge. The eluted SPE samples were analyzed with high-performance LC-MS/MS. RESULTS: A LLOQ of 0.01 ng/ml was achieved with a linear calibration range from 0.01 to 25 ng/ml. Optimal performance data were obtained from spotting the internal standard solution on the card before blood spotting. Internal standard addition from the system injector loop produced intra-assay inaccuracy of -9.0-7.3% and precision of 1.3-8.2%, and inter-assay inaccuracy of -3.5-3.9% and precision of 4.4-8.7%. CONCLUSION: Results demonstrated the feasibility of a semi-automated online rapid direct elution method that avoids manual extraction for DBS sample analysis using the online DBS-SPE system coupled to LC-MS/MS.

The use of a membrane filtration device to form dried plasma spots for the quantitative determination of guanfacine in whole blood.

RATIONALE: A two-layered polymeric membrane is employed for the formation of separated dried plasma spots from whole blood as an alternative to the direct analysis of whole dried blood spots (DBS). This dried plasma spot (DPS) analysis procedure precludes potential issues of hematocrit differences in whole blood samples while providing pharmokinetic data from plasma rather than whole blood. The described procedure is also semi-automated thus providing a simpler work flow for LC/MS/MS bioanalysis procedures. METHODS: Molecular filtration of red blood cells (RBC) from applied microsamples of whole blood fortified with guanfacine and its stable isotope internal standard was accomplished with a two-layer polymeric membrane substrate. The lower membrane surface containing the separated plasma spot was physically separated from the upper membrane followed by semi-automated direct elution of the sample to an online solid-phase extraction (SPE) cartridge followed by liquid chromatography coupled to tandem mass spectrometry (LC/MS/MS). RESULTS: A two-layer membrane sample preparation substrate produced plasma from whole blood without centrifugation which could be directly eluted for semi-automated LC/MS/MS bioanalysis. Standard curves were constructed by plotting peak area ratios between the analyte and the stable isotope labeled internal standard (SIL-IS) versus the nominal concentration in whole blood. A weighted 1/x(2) linear regression was applied to the data from DPS samples. Standard curves were linear over the range 0.25-250 ng/mL human whole blood. The representative regression equation was y = 0.0142x + 0.00248 (R(2) = 0.995) for the described DPS assay. CONCLUSIONS: The described work demonstrates proof-of-principle using membrane sample preparation techniques to form DPS samples from whole blood for subsequent bioanalysis by LC/MS/MS. This approach has the potential to eliminate the hematocrit issues from the current controversy surrounding validation of DBS assays.

Dried blood spots as a sampling technique for the quantitative determination of guanfacine in clinical studies.

BACKGROUND: Dried blood spot (DBS) technology was evaluated for the quantitative determination of guanfacine in human blood in clinical studies. A very sensitive DBS assay has been developed using HPLC coupled with an AB Sciex 5500 QTRAP® (Applied Biosystems/MDS Sciex, ON, Canada) MS system (LC-MS/MS) with a linear calibration range of 0.05 to 25 ng/ml. High-resolution MS using an Exactive Orbitrap® (ThermoFisher, LLC., CA, USA) was compared with the QTRAP using extracted exact mass ion current profiles for guanfacine and its stable-isotope-incorporated internal standard. The sample preparation employed liquid-liquid extraction with methyl t-butyl ether of 5 mm punched DBS card disks, followed by reversed-phase HPLC separation coupled with either MS/MS or high-resolution MS. Routine experiments were performed to establish the robustness of the DBS assay, including precision, accuracy, linearity, selectivity, sensitivity and long-term stability of up to 76 days. In addition, several factors that potentially affect quantitation were investigated, including blood volume for DBS spotting, punch size and punch location. RESULTS: A sensitive research assay with a LLOQ of 0.05 ng/ml was developed and subjected to several components of a method validation common to a regulated bioanalysis procedure employing DBS. This method development and partial validation study determined that spot volume, punch size or punch location do not affect assay accuracy and precision. The DBS approach was successfully applied to a clinical study (a Phase I, randomized, double-blind, placebo-controlled, crossover study to assess the effect of varying multiple oral doses of guanfacine on the pharmacokinetic, pharmacodynamic, safety, and tolerability profiles in healthy adult subjects). The pharmacokinetic profiles for 12 volunteers generated from the DBS assay and from a previously validated plasma assay were compared and were found to be comparable. DBS incurred samples collected from finger prick blood and directly applied to the DBS cards were also analyzed for comparison. CONCLUSION: From a bioanalytical perspective, DBS sample collection and analysis is a potentially viable alternative for guanfacine determination in clinical studies, utilizing approximately 100 µl of blood per subject profile compared with a few millilitres of blood drawn for conventional plasma bioanalysis.

Liquid extraction surface analysis mass spectrometry (LESA-MS) as a novel profiling tool for drug distribution and metabolism analysis: the terfenadine example.

Liquid extraction surface analysis mass spectrometry (LESA-MS) is a novel surface profiling technique that combines micro-liquid extraction from a solid surface with nano-electrospray mass spectrometry. One potential application is the examination of the distribution of drugs and their metabolites by analyzing ex vivo tissue sections, an area where quantitative whole body autoradiography (QWBA) is traditionally employed. However, QWBA relies on the use of radiolabeled drugs and is limited to total radioactivity measured whereas LESA-MS can provide drug- and metabolite-specific distribution information. Here, we evaluate LESA-MS, examining the distribution and biotransformation of unlabeled terfenadine in mice and compare our findings to QWBA, whole tissue LC/MS/MS and MALDI-MSI. The spatial resolution of LESA-MS can be optimized to ca. 1 mm on tissues such as brain, liver and kidney, also enabling drug profiling within a single organ. LESA-MS can readily identify the biotransformation of terfenadine to its major, active metabolite fexofenadine. Relative quantification can confirm the rapid absorption of terfendine after oral dosage, its extensive first pass metabolism and the distribution of both compounds into systemic tissues such as muscle, spleen and kidney. The elimination appears to be consistent with biliary excretion and only trace levels of fexofenadine could be confirmed in brain. We found LESA-MS to be more informative in terms of drug distribution than a comparable MALDI-MS imaging study, likely due to its favorable overall sensitivity due to the larger surface area sampled. LESA-MS appears to be a useful new profiling tool for examining the distribution of drugs and their metabolites in tissue sections.

Liquid extraction surface analysis (LESA) of food surfaces employing chip-based nano-electrospray mass spectrometry.

An automated surface-sampling technique called liquid extraction surface analysis (LESA), coupled with infusion nano-electrospray high-resolution mass spectrometry and tandem mass spectrometry (MS/MS), is described and applied to the qualitative determination of surface chemical residues resulting from the artificial spraying of selected fresh fruits and vegetables with representative pesticides. Each of the targeted pesticides was readily detected with both high-resolution and full-scan collision-induced dissociation (CID) mass spectra. In the case of simazine and sevin, a mass resolution of 100,000 was insufficient to distinguish the isobaric protonated molecules for these compounds. When the surface of a spinach leaf was analyzed by LESA, trace levels of diazinon were readily detected on the spinach purchased directly from a supermarket before they were sprayed with the five-pesticide mixture. A 30 s rinse under hot running tap water appeared to quantitatively remove all remaining residues of this pesticide. Diazinon was readily detected by LESA analysis on the skin of the artificially sprayed spinach. Finally, incurred pyrimethanil at a level of 169 ppb in a batch slurry of homogenized apples was analyzed by LESA and this pesticide was readily detected by both high-resolution mass spectrometry and full-scan CID mass spectrometry, thus showing that pesticides may also be detected in whole fruit homogenized samples. This report shows that representative pesticides on fruit and vegetable surfaces present at levels 20-fold below generally allowed EPA tolerance levels are readily detected and confirmed by the title technologies making LESA-MS as interesting screening method for food safety purposes.