Keywords: Capillary Gas Chromatography, MultiPurpose Sampler, Headspace, GC/MS, Volatiles, Urine
A MultiPurpose sampler (GERSTEL MPS), designed for liquid large volume, gaseous and headspace samples was tested for its suitability in the GC/MS analysis of organic volatiles in human urine. Headspace sampling with a volume, temperature and speed controlled gas tight syringe was combined with a temperature controlled cold injection system for cold trapping, enrichment and focusing of analyte. Regular 2 mL GC vials filled with 1 mL urine were used as headspace sampling vials. A 100 vial autosampler tray was equipped with an additional temperature and heating time controlled pre-heating module for 5 vials. Preliminary experiments were done with urine samples from our clinical laboratory with a ketone or glucose positive test result. In addition urine samples from healthy volunteers were taken and analysis were done with and without acidifying the urine. The promising results in regard to sensitivity and practicality in a day to day routine together with the cost cutting philosophy of a MultiPurpose sampler makes this highly automated system very attractive for clinical routine use.
Keywords: Blood Alcohol, Headspace Analysis, Ethanol
Forensic laboratories face the need to analyze many samples of human blood and body fluids for alcohol content. The large number of samples that require quantification of ethanol in these facilities creates a challenge for the methodology employed. Factors that need to be considered are sample throughput, resolution, and carryover. A successful method for these analyses should be fast, precise, and accurate. Current methods used in these analyses use a gas chromatograph coupled to a static headspace sampler and flame ionization detector (FID). The x, y, z robotic autosampler used in this study has a capacity of up to 128 headspace samples, which is a distinct advantage compared to other samplers commercially available. Results obtained with the instrument and methodology described in this report meet the specifications set by the California Department of Justice Blood Alcohol Operating Procedures (Title 17). A dual-column, dual-FID blood alcohol analysis system that can be used for confirmation of ethanol peaks was also tested and produced results with good precision (below 5 % RSD).
Keywords: Stir Bar Sorptive Extraction, SBSE, THC, GC/MS
This study demonstrates the feasibility of extracting THC from an aqueous matrix using Stir Bar Sorptive Extraction (SBSE) followed by thermal desorption into the injection port of a standard single quadrupole electron impact GC/MS to achieve detection limits less than 1.0 ng/mL. The specimen is diluted with water + methanol and extracted with a stir bar coated with a non-polar polydimethylsiloxane phase.
Keywords: Stir Bar Sorptive Extraction, SBSE, Basic Drugs, Forensic Analysis
Stir bar sorptive extraction (SBSE) is an innovative sample extraction technique that can be used to process blood, urine, and tissue samples for routine drug screening in the forensic toxicology laboratory. Case specimens containing drugs typically encountered in forensic samples were evaluated using SBSE and compared with a liquid/liquid extraction method followed by GC/MS analysis. This evaluation demonstrated that SBSE method worked equally as well as the routine extraction method for the detection of basic drugs in screening forensic samples.
Keywords: Drugs, Urine, Blood, Dean‘s Switch, Multidimensional GC, Low Thermal Mass, LTM, Fast GC/MS
Reduction in analysis time is an important goal for easing the burden of large sample sets associated with routine screening of blood samples and sample sets produced from drug metabolism studies. This study focuses on reduction in analysis time for simultaneous detection of delta-9-THC, 11-OH-THC, and THC-COOH in whole blood and urine extracts. This was achieved using the GERSTEL fast GC system combined with an Agilent GC-MSD. Addition of the fast GC system allows three independently heated temperature zones for multidimensional chromatography using Agilent Capillary Flow Technology Dean’s Switches along with fast heating/cooling rates. A novel pre-column approach protects the analytical columns in independent temperature zones and adds a high level of robustness. The Agilent Capillary Flow Technology Dean’s Switch allows a combination of heartcutting multidimensional GC and back flushing to reduce the amount of unwanted background components. Standard analysis time was reduced from 15 minutes to less than 11 minutes for blood samples and from 15 to less than 8 minutes for urine samples. Two Agilent Capillary Flow Technology Deans Switches in tandem were used for this analysis. Three independently programmed pressure zones were used in conjunction with three independent heated zones. The MS was operated in the EI mode.
Keywords: Sample Preparation, DPX, Opiates, Automation
The analysis of opiates in blood and urine was accomplished using automated Disposable Pipette Extraction (DPX) followed by LC/MS analysis. The automated extractions were performed in about 5 minutes using a GERSTEL MultiPurpose Sampler (MPS). The eluents were subsequently dried and reconstituted in solvent and injected into a HPLC/MS/MS instrument. Recoveries ranged from 60 to 85% for the opiates in blood, and recoveries ranged from 78 to 85% for the opiates in hydrolyzed urine. The %RSDs were lower than 6% for all analytes.
Keywords: DPX, LC/MS/MS, Sample Preparation, Lab Automation
This study focuses on the automated extraction of small sample volumes coupled to LC/MS/MS in order to provide high throughput analysis of an extended list of benzodiazepines. Using a GERSTEL MPS autosampler, DPX extractions of a variety of biological matrices were performed, using a mixed-mode cation exchange (DPX-CX) sorbent. The resulting eluents from the automated DPX extractions were then introduced into an Agilent 6410 LC/MS/MS instrument. Coupling DPX to LC/MS/MS provides rapid, just-in-time sample preparation for high throughput analysis. Data show the use of an Agilent 6410 LC/MS/MS instrument to be a highly sensitive procedure for the analysis of benzodiazepines with limits of quantitation of 0.5 ng/mL, and good linearity. The DPX extraction removes potential matrix interferences and ion suppression, and high sensitivity is therefore achieved.
Keywords: Drugs of abuse, Pharmaceuticals, Tissue samples, Stir Bar Sorptive Extraction, SBSE, Thermal Desorption
Stir Bar Sorptive Extraction (SBSE) is an innovative and efficient method for the extraction of drugs and pharmaceuticals from blood, urine and tissue samples in a forensic toxicology laboratory. As shown in this application note and earlier publications, SBSE is an effective screening tool for drugs and pharmaceuticals in biological fluids and tissue. The SBSE technique is easy to use and the method described in this publication is performed without additional use of organic solvents, centrifugation, or the liquid transfer steps that are normally necessary when performing liquid-liquid or solid-phase extraction.
Keywords: DPX, LC-MS/MS, Sample Preparation, Laboratory Automation
This study focuses on the automated extraction of small sample volumes combined with LC-MS/MS analysis providing high throughput analysis of common pain management drugs. Using a GERSTEL MultiPurpose Sampler (MPS), DPX extractions of hydrolyzed urine were performed, using a reversed phase sorbent with a proprietary salt additive (DPX-RP-S). The resulting eluents from the automated DPX extractions were introduced into an Agilent 6460 LC-MS/MS instrument.
Keywords: DPX, LC-MS/MS, Sample Preparation, High Throughput Laboratory Automation
This study focuses on the automated extraction of small volumes of urine samples (< 500 uL) using disposable pipette extraction (DPX) for the comprehensive screening for pain management drugs by LC-MS/MS. Using a GERSTEL MPS autosampler, DPX extractions of hydrolyzed urine were performed, using a reversed phase (DPX-RP-S) sorbent. The resulting eluents from the DPX extractions were automatically diluted and injected into an Agilent® Technologies LC-MS/MS system. Sample preparation was performed just-in-time enabling high throughput screenings, averaging a cycle time of 7 min/sample. Validation results show that the automated DPX-LC-MS/MS screening method provides adequate sensitivity for over 65 analytes and internal standards. Lower limits of quantitation (LLOQ) ranged between 0.5 – 50 ng/mL and % RSDs were below 10% in most cases.
Keywords: DPX, LC/MS/MS, Sample Preparation, High Throughput Laboratory Automation
This study focuses on the rapid cleanup of urine samples (< 500 uL) using disposable pipette extraction (DPX) for high throughput LC/MS/MS screening of buprenorphine (Bup) and main active metabolite norbuprenorphine (Nbup). Using a GERSTEL MultiPurpose autosampler (MPS), DPX extractions of hydrolyzed urine were performed, and analysis results were directly compared to results obtained using the "dilute-and-shoot" (D&S) approach for the same samples. The automated DPX cleanup process significantly reduced matrix effects without compromising the required minimum reportable limits (MRLs), whereas when using the D&S approach, samples had to be diluted by up to a factor 100 to remove such effects, which directly affected the ability to determine Bup and Nbup at the MRLs. Moreover, the addition of valve switching control from MAESTRO software further increased the throughput potential of the solution. The resulting eluents from the automated DPX extractions were injected into an Agilent 6460 LC/MS/MS instrument configured with 2 LC pumps (gradient and re-generative) allowing rapid, just-in-time sample preparation for high throughput screening, averaging a cycle time of 4 min/sample.
Keywords: Doping, Equine Urine, Automated Sample Preparation, Disposable Pipette Extraction, Triple Quadrapole Mass Spectrometry, Gas Chromatography
This application note describes a fully automated method for the determination of selected doping compounds in equine urine. A GERSTEL MultiPurpose autosampler (MPS) with Disposable Pipette Extraction (DPX) option is employed for extraction and cleanup. After gas chromatographic separation the analytes are detected by a triple-quadrupole mass spectrometer (QqQ-MS). The method is rugged, provides an excellent cleanup of the complex sample matrix and shows very good limits of detection, from below 0.1 to just under 10 ng/mL, when compared to labor intensive manual manual SPE methods.
Keywords: DPX, LC/MS/MS, Sample Preparation, High Throughput Laboratory Automation
This work demonstrates the use of disposable pipette extraction (DPX) as a fast and automated sample preparation technique for the determination of barbiturates and 11-nor-9-carboxy- 9-THC (COOH-THC) in urine. Using a GERSTEL MultiPurpose autosampler (MPS) with DPX option coupled to an Agilent 6460 LC/MS/MS instrument, 8 barbiturates and COOH-THC were extracted and their concentrations determined. The resulting average cycle time of 7 min/sample, including just-in-time sample preparation, enabled high throughput screening. Validation results show that the automated DPX-LC/MS/MS screening method provides adequate sensitivity for all analytes and corresponding internal standards that were monitored. Lower limits of quantitation (LLOQ) were found to be 100 ng/mL for the barbiturates and 10 ng/mL for COOH-THC and % CVs were below 10 % in most cases.
Keywords: DPX, LC/MS/MS, Sample Preparation, Drugs of Abuse, High Throughput Lab Automation
This application demonstrates the use of Disposable Pipette Extraction (DPX) for rapid, automated sample preparation of urine samples for comprehensive LC/MS/MS screening. The combination of automated sample cleanup and introduction with mass spectrometric detection using a Scheduled MRM™ (AB SCIEX) algorithm and fast MS/MS spectral acquisition allowed high confidence compound identification based on mass spectral library matching. The automated workflow enabled monitoring of large panels of analytes (100+ drugs); detecting and quantifying these compounds in a single run. The new automated DPX-LC/MS/MS workflow provides rapid extractions, high recoveries, and minimized matrix interferences with complete automation capabilities towards high throughput chromatographic analysis.
Keywords: Opiates, Opioids, Cocaine, Solid Phase Extraction, SPE, Automation, GC/MS
Analyzing blood serum for opioids, cocaine and metabolites is a routine task in forensic laboratories. The most commonly used methods involve several manual or partly-automated sample preparation steps such as protein precipitation, solid phase extraction, evaporation and derivatization followed by GC/MS or LC/MS determination. In this study a comprehensively automated method is compared with a validated, partly-automated routine method. Following manual protein precipitation, the automated method relies on a MultiPurpose Sampler (MPS) to perform all remaining sample preparation steps. These include solid phase extraction (SPE), evaporation of the eluate, derivatization and introduction to the GC/MS. Quantitative analysis of close to 170 serum samples, as well as more than 50 samples of other matrices like urine, different tissues and heart blood, was performed using both methods. Cocaine, benzoylecgonine, methadone, morphine, codeine, 6-monoacetylmorphine, dihydrocodeine and 7-aminofl unitrazepam were determined quantitatively and the methods were found to produce equivalent analytical results even near the limits of quantification
Keywords: Mycotoxins, LC/MS/MS, Sample Preparation, Lab Automation, Food Safety
In this report, we describe a completely automated sample preparation work multi-mycotoxin residues in different food matrices (corn, wheat) by LC/MS/MS. The extraction and cleanup was performed using a GERSTEL MultiPurpose Sampler (MPS) followed by LC/MS/MS determination using an AB SCIEX QTRAP® 4500. The automated sample preparation work flow involved centrifugation, dispersive solid phase extraction (dSPE) and evaporative concentration, providing extraction efficiencies greater than 70 % with RSDs less than 15 % for most analytes. The LC/MS/MS method was developed for screening for a panel of 14 mycotoxins (aflatoxins, trichotecenes and fuminosins) using the Scheduled MRM™ algorithm in combination with fast polarity switching, achieving excellent linearity (R2 values of 0.98 or greater) , average accuracies greater than 88 % and limits of quantitation lower than the action levels established by the EC and FDA.
Keywords: Sample Preparation, LC/MS/MS, High Throughput Laboratory Automation, DPX, Urine, Glucuronides
A major mechanism of the metabolism of many pain management drugs involves conjugation of the analyte with glucuronic acid. To ensure accurate results when drugs are determined from urine matrices, the analytes must be deconjugated which is typically performed by hydrolysis using enzymes such as beta-glucuronidase. Typical hydrolysis procedures involve long incubation periods and specified temperatures and have traditionally been performed manually. This study shows how a typical enzymatic hydrolysis procedure can be easily automated using a GERSTEL MultiPurpose Sampler (MPS), combining an automated extraction and clean-up procedure with introduction to the LC/MS/MS, in order to provide high throughput analysis of common pain management drugs.
Keywords: THC, THC-OH, THC-COOH, Serum, Solid Phase Extraction, SPE, Automation, GC/MS
This note presents a fully automated analysis system for the determination of THC and its metabolites in blood serum. Automation is based on the GERSTEL MultiPurpose Sampler (MPS) equipped for solid phase extraction and a module for automated eluate evaporation (GERSTEL mVAP). A validated, semi-automated analysis method used for routine analysis was transferred and automated using the described system. Improvements were realized such as a reduction of sample volume and the use of a smaller SPE cartridge format. The method was validated according to Society for Toxicology and Forensic Chemistry (GTFCh) guidelines. Limits of quantification below 1 ng/mL for THC and THC-OH, extraction efficiencies between 70 and 93% and RSDs between 3 and 10% were achieved.The SPE system performs sample preparation in parallel with the chromatographic run, enabling the GC/MS system to operate at maximum capacity.
Keywords: Urine Hydrolysis, β-Glucuronidase, LC/MS/MS
In this report, a completely automated, 96 well plate format “Prep-and-Shoot” workflow including enzymatic hydrolysis, dilution and injection is described. A GERSTEL MultiPurpose autosampler (MPS) coupled to an AB SCIEX QTRAP® 4500 LC/MS/MS system was used for a fast enzymatic hydrolysis process (15 minutes), dilution and injection of urine samples. The procedure was applied to the analysis of multiple drug classes (e.g., opiates, opioids, benzodiazepines, muscle relaxants, hallucinogens) in urine. This automated workflow employed an ultra-pure β-Glucuronidase enzyme yielding hydrolysis efficiencies of glucuronide conjugates above 80 % for the analytes tested. The methodology developed allowed the reproducible injection and analysis of over 960 samples on the same analytical column, with % RSDs ≤ 10 %. Moreover, the combined automation of urine hydrolysis, injection and analysis allowed the system to process more than 200 samples in a day.
Keywords: Forensic, Biological Fluids, Clinical Research
Below is a list of publications for your reference. These cannot be downloaded due to Copyright protection.
Ethanol analysis from biological samples by dual rail robotic autosampler C. L. Morris-Kukoski, E. Jagerdeo, J. E. Schaff, M. A. LeBeau (Federal Bureau of Investigation Laboratory, Quantico, VA) J. Chromatography B. 850 (2007) 230-235
Stir bar sorptive extraction-thermal desorption-capillary GC-MS for profiling and target component analysis of pharmaceutical drugs in urine. Tienpont, B. et al. J. Pharm. Biomed. Anal. 2003, 32, 569-579.
Identification of in Vivo Diethylhexylphthalate Metabolites in Breath, Plasma and Urine of Dialysis Patients H.G. Wahl, A. Chrzanowski, D. Luft, H.U. Häring, H. Liebich 20th International Symposium on Capillary Chromatography-Riva del Garda, Italy (1998)
Enantiomer Separation of Desflurane in Human Blood and Urine Samples by Headspace GC/MS R. Schmidt, M. Hadjidimos, H.G. Wahl, V. Schurig 23rd International Symposium on Capillary Chromatography-Rivadel Garda, Italy (2000)
Quantification of Volatiles in Mammalian Urine by Stir Bar Sorptive Extraction (SBSE) Techniques and Gas Chromatography H.A. Soini, K.E. Bruce, D. Wiesler, M.V. Novotny Symposium on Capillary Chromatography and Electrophoresis-Riva del Garda, Italy (2002)
Comprehensive Profiling of Drugs of Abuse in Biological Fluids by SBSE-TD-CGC-MS B. Tienpont, F. David, P. Sandra Symposium on Capillary Chromatography and Electrophoresis-Riva del Garda, Italy (2002)
Analysis of Polycyclic Aromatic Hydrocarbon Metabolites in Urine Using in-situ Derivatisation Stir Bar Sorptive Extraction-CGC/MS K. Desmet, P. Kiss, M. DeHertogh, P. Sandra Symposium on Capillary Chromatography and Electrophoresis-Riva del Garda, Italy (2002)
Stir bar sorptive extraction-thermal desorption-capillary gas chromatography-mass spectrometry applied to the analysis of polychlorinated biphenyls in human sperm Tom Benjits, Joeri Vercammen, Riet Dams, Hai Pham Tuan, Willy Lambert, Pat Sandra Journal of Chromatography B, 755 (2001) 137-142
Breath Analysis in Mechanically Ventilated Patients: Comparison of Different MethodsW. Miekisch , J.K. Schubert, W.P.E. Mueller, K. Geiger 20th International Symposium on Capillary Chromatography-Rivadel Garda, Italy (1998)
Stir bar sorptive extraction-thermal desorption-capillary GC-MS applied to biological fluids B. Tienpont, F. David, K. Desmet, P. Sandra Anal Bioanal Chem (2002) 373:46-55
Keywords: Forensics ,Veterinary, Sample Preparation, LC/MS/MS, High Throughput Lab Automation
The extraction of dried blood spots (DBS) typically involves manual intervention. First, a small disc is punched out of the center of a dried blood spot placed on a DBS card. Following solvent extraction of the sample, it is also common to include further cleanup steps, using solid phase extraction (SPE) to improve detection limits or exchanging solvents for compatibility with subsequent chromatographic separations. Modern analytical labs are looking to automate the process to help reduce solvent usage and to increase sample throughput while ensuring the high quality of the resulting data.
In this report, the complete automation of dried blood spot analysis is demonstrated and the results evaluated. A novel, automated DBS Autosampler (DBS A) automatically inserts DBS cards into a Flow Through Desorption (FTD™) cell in which individual blood spots are rapidly and effectively desorbed. The DBS A is integrated into a complete cleanup and analysis system using online SPE with replaceable cartridges combined with automated injection to an LC/MS/ MS system. Automated DBS extraction methods for a variety of analytes from different matrices are examined along with the use of different SPE cartridge sorbents. The resulting precision and accuracy data are provided.