no. 9 march 2009...known as omega-6-fatty acids) among these linoleic acid and its derivatives, and...

Focus onFood Analysis

Fats, FAMEs, Fatty Acid Profiles, Rancidity ...

Archaeology meets chemistry

Peeking intoPharaoh’s wine glass

Polymer analysis

EfficientAutomated Pyrolysis GC

Automated DPX

Drugs of Abuse:Extraction in seconds

News from GERSTEL GmbH & Co. KG · Eberhard-Gerstel-Platz 1 · D-45473 Mülheim an der Ruhr · Germany · Phone +49 (0) 2 08 - 7 65 03-0 · [email protected]

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No. 9 March 2009

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G L O B A L A N A L Y T I C A L S O L U T I O N S

In this issue

GERSTEL Solutions Worldwide Editorial

News■Cooperation: New

Solution for Fat

Determination 3

Application■Determining

markers for fatty

acid decomposition 4■FAMEs: Automated,

Accurate and Reproducible Fatty Acid

Derivatization and Determination 8

Innovation■Polymer analysis:

Efficient Automated Pyrolysis GC 12■Drugs of abuse:

Extraction in seconds 14 ■MAESTRO-Software:

Perfectly Synchronized

Sample Prep and Analysis 24

Interview■Robert J. Collins, Ph.D., President

GERSTEL, Inc.: Looking

forward to shaping the future

of the business 18

Report■Archaeology meets chemistry:

Peeking into Pharaoh’s wine glass 21

International Sales under new leadership

As of July 2008, Ralf Löscher, Ph.D. has joined GERSTEL as International Sales Manager. A passionate marathon runner with a Ph.D. degree in biology, Dr. Löscher brings new energy to the International Sales team, coupled with extensive expe-rience in the international analytical instrumentation business. Before joining GERSTEL, the frequent flyer was International Sales Manager for a reputable supplier of GC and LC Time of Flight (TOF) Mass Spectrometry systems and had before that worked for a major international corporation, responsible for sales of LC and LC/MS instrumentation in Germany. Among Ralf Löscher’s many activities in his first year in office have been participation in the Analytica China and ArabLab exhibitions, reinvigorating and re-focusing GERSTEL activities in growth markets with a stronger presence and forming new contacts and alliances.

GERSTEL introduces Automated Disposable Pipette Extraction (DPX)

Disposable Pipette Extraction (DPX) is a fast and efficient SPE technique used for a wide range of applications such as drugs of abuse and therapeutic drug monitoring. Only 200 - 250 µL of sample is needed to reach the required limits of detection using a fully automated process. The extraction step is performed in 30 - 60 seconds and the complete process including elution and rinse steps takes 3 - 6 minutes depending on the application. Addition-

ally, automated DPX is performed during the GC or LC run of the preceding sample en-suring maximum throughput and best possible GC/MS or LC/MS system utilization. Elu-tion requires only a small amount of solvent, which means that DPX effectively provides a concentration step. For many applications, such as pesticides in fruit and vegetables, solvent evaporation is not required. For more information on the DPX system, please see article on page 14.

Keeping VOCs and SVOCs out of car interiors

All leading worldwide car manufacturers and a large number of their suppliers rely on GERSTEL technology. Analyses are regularly performed to ensure that materials used in or near the vehicle interior do not emit dangerous levels of Volatile and Semi-Volatile Organic Compounds (VOCs and SVOCs). A method that is often used world-wide is the VDA Method 278 from the German Automobile Producers Association (VDA). The VDA 278 method spec-ifies thermal desorption/ thermal extraction of materials that are placed directly in thermal desorption tubes followed by GC/MS determination. Some manufacturers have their own methods based on GERSTEL equipment. Recently, two new methods were established by a renowned leading world-wide manufacturer. One meth-od complements the VDA 278 method in the determination of emissions from materials used in car interiors; the other meth-od specifies direct air monitor-ing inside the passenger cab-ins of vehicles. As far as GER-STEL Solutions Magazine is in-formed, the methods are to be used by all world-wide opera-tions including suppliers. Both methods specify the GERSTEL Thermal Desorption System (TDS 3) with Thermal Desorption Autosampler (TDSA) in combi-nation with a GERSTEL Cooled Injection System (CIS 4).

Published by GERSTEL GmbH & Co. KG Eberhard-Gerstel-Platz 1 45473 Mülheim an der Ruhr Germany

Editorial Director Guido Deußing ScienceCommunicationNeuss, Germany [email protected]

Translation and editingKaj Petersen [email protected]

Scientific advisory boardEike Kleine-Benne, Ph.D. [email protected]

Oliver Lerch, Ph.D. [email protected]

Malte Reimold, [email protected]

Contact [email protected]

Design Paura Design, Hagen, Germany www.paura.com

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Imprint

GERSTEL online

You can find more information on products, applications and services on the GERSTEL home page at www.gerstel.com.

News

2 GERSTEL Solutions Worldwide – March 2009

GERSTEL Solution Worldwide News

GERSTEL and Büchi in cooperation

New Solution for Fat Determination

In order to further improve the patented Caviezel® rapid analysis method for determination of fat in food and feed, the Swiss company Büchi has entered into cooperation with GERSTEL.

W hen it comes to the determination of fats in food and feed, the Swiss company Büchi plays a substantial

role. This is due to the patented Caviezel method, which is increasingly specified in standardized methods. Examples are the determination of fat contents that must be listed on food products as well as the de-termination of milk fat in foods.

Following the Caviezel method, an in-ternal standard is added to the sample and the fat is extracted before undergoing alka-

line digestion. The resulting salts are con-verted to free acids and these are subse-quently determined using a gas chromato-graph (GC) with flame ionization detection (FID). Leading Büchi managers looked to GERSTEL to provide technical and applica-tion know-how in order to fully unlock the potential of the Caviezel method.

GERSTEL has provided improved au-tomation and simplified, accelerated GC analysis; in short: a single system that per-forms three different analyses in one short run, providing the analyst in the laboratory with a simpler, more efficient method.

Büchi was faced with the challenge of improving the automation of their fat anal-ysis solution, and they seized the opportu-nity to team up with outside expertise. The goal of the GERSTEL-Büchi cooperation was to create a comprehensive integrated solution, comprised of the Büchi extraction unit B 815, an Agilent 7890 GC with a split/splitless inlet and a FID, a GERSTEL Mul-tiPurpose Sampler (MPS), integrated soft-ware control as well as a customized report. GERSTEL has exclusive sales and distribu-tion rights world-wide for this solution.

MPS provides high throughputAutomated sample preparation and sam-ple introduction are among the core com-petencies of the GERSTEL MPS, making it the ideal candidate for automating all the required steps in the GERSTEL-Büchi fat analysis solution. „The MPS combines high sample capacity with the flexibility to adapt to any sample preparation challenge”, says Ralf Bremer, Managing Director in charge of R&D and Production at GERSTEL. Us-ing the PrepAhead functionality of the GERSTEL MAESTRO software, the MPS can even prepare the next sample or mul-tiple samples while the current analysis is ongoing. This means that the next sample is always prepared and ready for injection as soon as the GC finishes running the pre-vious sample. The GC system never has to be idle; it is always utilized to its fullest ca-pacity providing best possible return on in-

vestment. MAESTRO additionally offers an unsurpassed degree of flexibility, which is often sorely needed in a production envi-ronment. Priority samples that need to be analyzed as soon as possible, (for example, to release a batch of product or to accept an incoming raw material shipment), can simply be inserted into the running analy-sis sequence table.

3-in-1 chromatography systemBüchi’s pre-GERSTEL solution was based on a GC method using packed columns, an older technology that has limited sep-aration power. Because of this limitation, three different GC runs, each based on a different column was required per sample. The total analysis time needed was of course quite long and the user had to change and condition the columns prior to performing the next analysis. Last, but not least, lack of separation power resulted in wider peaks and less accurate results. “Implementing a method that uses capillary column tech-nology along with the 7890 GC from Agi-lent Technologies enabled us to improve the separation and provide more accurate peak integration and thus more accurate results”, says Ralf Bremer, “and all three analyses are now performed in one run in just 9 min-utes”. There is no longer a need to change columns and this of course improves the stability, performance and productivity of the system. The system provides values for the following: Total fat content, fat profile and butter fat (milk fat) content. According to Jochen Knecht, Ph.D., Managing Direc-tor of Büchi Germany and initiator of the cooperation, “This is a big step forward”.

GERSTEL delivers the complete system pre-loaded with required analysis method parameters and a reporting tool that de-livers all relevant results nicely organized on one page. “To use our system, you don’t have to be a GC expert”, says Ralf Bremer, “the system produces first class results in both production and R&D environments – and it does so much faster than it’s pre-decessor”.

Complete solution from Büchi and GERSTEL for the determination of fat levels in food and feed products. The system is based on the Büchi Extraction unit B 815, a 7890 GC from Agilent Technologies with a split/splitless inlet and a FID, a GERSTEL MultiPurpose Sampler (MPS) and MAESTRO software.

3GERSTEL Solutions Worldwide – March 2009

GERSTEL Solutions Worldwide Application

Determining markers for fatty acid decomposition

Fighting rancidity

Assessing the quality of food oils, fats and products that contain fatty acids may be cumbersome, but it is necessary in order to safeguard product quality and consumer safety. Scientists from GERSTEL have developed a new and more efficient method for monitoring quality markers in oily ma-trices: Oxidation products such as aldehydes and ketones are determined using automated Dynamic Headspace (DHS) coupled with GC/MS.

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Structure of the determined fatty acid decomposition products

S almon, bluefish, trout, walnuts, rape-seed oil, sesame seeds, sunflower seeds, soybeans, corn, vegetable oil-

based spread... this is not a shopping list, but these items are recommended as part of a healthy and nutritious diet. The rea-son for such a recommendation is that these foods contain long-chain polyunsaturated fatty acids (LCPs). LCPs can be divided in-to two categories: n-6-fatty acids (formerly known as omega-6-fatty acids) among these linoleic acid and its derivatives, and n-3-fat-ty acids (formerly known as omega-3-fatty acids) to which group the α-linolenic acid and its derivatives belong.

LCPs are said to have many beneficial properties. Provided to us via our mother’s milk, they provide an essential contribu-tion to the development of the brain, ner-vous system and vision in the child. At any age, LCPs are known to benefit our heart and circulatory system and they prevent or reduce arteriosclerosis and related illness-es. A deficiency of essential fatty acids can have dire negative consequences starting with skin conditions, such as calloused skin, increased susceptibility for infections, re-duced growth, hair loss, and reduced blood plate-count.

It is hardly surprising that LCPs are ex-tracted from natural products and added to foods in order to provide health bene-fits via pre-natal and post-natal baby nutri-tion. These extracts are also used to enrich foods that are low in LCPs, such as stan-dard bread products, and turn these into more valuable so-called functional foods that have health benefits.

Strength and WeaknessLCPs strengthen us, but are themselves of weak constitution. Getting too much “fresh air” doesn’t help them either. Oxygen mole-cules attack and destroy their double bonds and degradation products are formed. And this doesn’t exactly go unnoticed, since com-pounds with unpleasant odors and very low odor thresholds are formed. Degradation products such as aldehydes and ketones, 4-heptenal for one, are among the oxidative degradation products formed from the fat-ty acids. A concentration of less than 10 ng per gram is typically sufficient to give an oil or food a distinct rancid smell or flavor.

For most foods, the contact with oxy-gen cannot be prevented, only reduced, but degradation reactions can at least be slowed by storing foods at low temperatures. In the end, freshness of food oils and fats should be monitored. How then to monitor these reactions, when odor thresholds are so very low? Instrumental techniques are required that can extract and concentrate analytes without accelerating the very process they are meant to monitor, i.e. without heating

The GERSTEL scientists mainly fo-cused on eleven compounds that are known degradation products of LCPs: 1-pentene-3-one, 2-(E)-pentenal, hexanal, 2-(E)-hexenal, 4-(Z)-heptenal, 2-pentylfurane, 1-octene-3-one, 2,4-(E,E)-heptadienal, 2,6-(E,Z)-nonadienal, 2,4-(E,E)-nonadie-nal and 2,4-(E,E)-decadienal. One gram of each oil sample was stored in a 20 mL screw cap headspace vial. “Weighing in the sam-ples was the only manual preparation step we had to perform”, says Oliver Lerch, “and if we had used the automated weighing op-tion for the MPS, even this step could have been automated” (cf. GERSTEL Solutions Worldwide No. 8). All further sample prep-aration steps, including adding an internal standard, performing Dynamic Headspace extraction, and introducing the concentrat-ed analytes to the GC/MS were performed automatically.

Some details: Standards containing from 5 to 500 ng/µL of the target com-pounds were prepared from a 1 µg/µL stock solution by dilution with hexane. 1 µL of standard solution was added to each of the vials containing 1 g of oil sample. The vials were placed in the MPS sample trays and successively transferred into the DHS sta-tion agitator, where they were kept at 70 °C for 4 minutes for equilibration. DHS extrac-tion was then performed for 10 minutes us-ing a 50 mL/min flow of nitrogen to purge and transfer analytes onto a replaceable Te-nax TA adsorbent trap for concentration.

the sample too much in the process. Ac-cording to Oliver Lerch, Ph.D., applica-tion scientist at GERSTEL, Static Head-space (HS) coupled with GC/MS is a use-ful technique for the determination of vola-tile compounds. However, for the determi-nation of oxidation products in oily foods, detection limits reached using Headspace GC/MS are much too high; it is simply not possible to monitor these compounds at the required concentration levels using Static Headspace. More sensitive techniques are needed, such as Headspace Solid Phase Mi-cro-Extraction (HS-SPME), which relies on a fiber coated with sorbent to concentrate analytes from the sample headspace or the Dynamic Headspace (DHS) technique, which purges and concentrates analytes on-to an adsorbent trap.“Of these more sensi-tive techniques, we achieved the best re-sults”, says Lerch, “when we used the Dy-namic Headspace (DHS) technique for the determination of oxidation markers such as aldehydes and ketones”.

Theory guides, experiment decidesBased on a customer request, Dr. Lerch and his colleagues in the Analytical Services De-partment of GERSTEL investigated ten dif-ferent oil samples using automated DHS coupled with GC/MS. Most of the samples were vegetable oils, among them olive oils and rapeseed oils from different producers. Two of the samples were fish oils.


Analyte RT [min] m/z rapseed oil No.1 rapseed oil No.2 (fresh) in ng/g (fresh) in ng/g1-Penten-3-one 7.530 55 1.1 17.52-(E)-Pentenal 10.432 83 1.5 13.0Hexanal 11.892 56 26.1 > 5002-(E)-Hexenal 14.532 83 0.4 17.94-(Z)-Heptenal 16.010 94 0.2 nd2-Pentylfurane 18.858 81 0.5 35.61-Octen-3-one 19.304 70 nd 16.82,4-(E,E)-Heptadienal 21.484 81 12 14.72,6-(E,Z)-Nonadienal 26.900 70 nd 4.32,4-(E,E)-Nonadienal 29.364 81 0.9 63.72,4-(E,E)-Decadienal 32.964 81 5.9 9.2 nd = not detected

Table 2: Comparison of two fresh rapeseed oils from different producers.

Analyte RT [min] m/z rapseed oil No.1 rapseed oil No.1 (fresh) in ng/g (aged) in ng/g1-Pentene-3-one 7.530 55 1.1 52-(E)-Pentenal 10.432 83 1.5 15.7Hexanal 11.892 56 26.1 244.72-(E)-Hexenal 14.532 83 0.4 19.94-(Z)-Heptenal 16.010 94 0.2 4.92-Pentylfurane 18.858 81 0.5 0.31-Octen-3-one 19.304 70 nd 2.32,4-(E,E)-Heptadienal 21.484 81 12 902,6-(E,Z)-Nonadienal 26.900 70 nd nd2,4-(E,E)-Nonadienal 29.364 81 0.9 6.92,4-(E,E)-Decadienal 32.964 81 5.9 52.6 nd = not detected

Table 1: Comparison of fresh and aged rapeseed oil, which had been stored for 6 months under normal household conditions.

Oliver Lerch, Ph.D., Application Specialist,GERSTELGmbH & Co. KG

The standard addition calibration curves for the different compounds in an oil sample are linear up to 500 ng/g. The correlation coefficients were around 0.999 for almost all compounds.


The user can specify whether a new trap is used for each sample and of course which adsorbent is used, the tubes are avail-able with any stan-

dard adsorbent. Following dynamic head-space extraction and analyte concentra-tion, the adsorbent tube is transferred to the GERSTEL Thermal Desorption Unit (TDU). Analytes are desorbed in the TDU, transferred to the Cooled Injection System (CIS), where they are again focused, and fi-nally transferred in a narrow band to the GC/MS system for identification and quan-tification.

Fatty acid decomposition provides a clear picture of product quality Many samples were analyzed using the GERSTEL DHS in order to prove its use-fulness over the entire concentration range of decomposition products (generally from 1 to 100 ng/g) that represents fresh, aged or slightly rancid product.

As an example, Lerch mentions rape-seed oil products, which were analyzed as described above and categorized: Fresh oils mostly had a very low concentration of de-composition products. When such oil had been stored under normal household con-ditions for six months, however, levels had increased significantly (cf. table 1). Apart from the aspect of aging, large differenc-es in freshly purchased oils from different producers were demonstrated (cf. table 2). “The statistics supported our findings nice-ly”, says Lerch, “the standard addition cali-bration curves ranged up to 500 ng/g with excellent linearity; most correlation coef-ficients were at 0.999”. Standard deviation


Repeatability was tested with fresh rapeseed oil which had been spiked with 5 ng/g of each analyte. The RSD for five runs was under 6 % for most compounds, this was at least as good as the results obtained when performing the analysis with HS-SPME.

Analytical conditions

Adsorbent: Tenax TA

DHS: Trap temperature: 30 °C; Incubation temperature: 70 °C; extraction/purge volume: 500 mL N2

TDU: Splitless mode; Temperature program: 40 °C (0 min) – 720 °C/min to 280 °C (5 min)

CIS: TDU desorption flow: 70 mL/min Analyte transfer: Split 2.5:1 Temperature program: -150 °C (0 min); 12 °C/s to 270 °C (7 min).

Column: DB-624 (Agilent Technologies); Length: 30 m; I.D. = 0.25 mm; df = 1.4 µm

Carrier gas: He, constant flow: 1.5 mL/min

GC oven 40 °C (1 min); 4 °C/min to 170 °C;program: 30 °C/min to 240 °C (5 min)

MSD mode: Selected Ion Monitoring (SIM)

For the determination of quality markers from fatty acid decomposition, the GERSTEL scientists relied on an automated DHS-GC/MS system. The instrument set-up has been implemented successfully for this application in customer laboratories.


and repeatability was tested with fresh rape-seed oil that had been spiked with 5 ng/g of each target analyte. The RSD for five runs was under 6 % for most compounds. The method proved to be robust and run-to-run carry-over was below 0.01% for almost all compounds.

When using the DHS-GC/MS tech-nique, the analyst is able to get a clear and unequivocal picture of LCP decomposition and thereby of the quality and freshness of oils, fats, and foods that contain fat. The concentration of aldehydes and ketones that are “marker compounds” for fat de-composition will rise over time as the prod-uct ages and this makes it easy to determine product freshness. The limits of determina-tion for the marker compounds are in the range from 0.05 to 5 ng/g. The excellent cor-relation is also significant, “it is proof that the method is well suited for quality con-trol of foods”, says Oliver Lerch.



Every laboratory is under pressure to quickly and consi-stently deliver accurate results and to provide clear an-swers faster than ever before while reducing the cost per analysis. Analysis systems that meet these criteria are al-ways welcome. Application scientists from Bespak Euro-pe Ltd. and from Anatune Ltd., both based in the U.K., ha-ve provided such a solution for automated derivatization and GC/MS determination of fatty acids. The approach ta-ken to the challenge: Automated sample preparation com-bined with discrimination free introduction of the deriva-tized analytes to the GC/MS system. Automated sample preparation in this case includes adding an internal stan-dard and derivatizing the fatty acids.

Determining FAMEs with just-in-time

sample preparation

Automated, Accurate and Reproducible

Fatty Acid Derivatization and Determination



T he more complex the method, the higher the demands on the labora-tory robotics used. A system that is

able to perform two independent robot-ic functions is by default the more flexible and can frequently provide better produc-tivity and throughput. The GERSTEL Prep-Station happens to be such a system; it was successfully used to automate the determi-nation of fatty acid profiles and concentra-tions in dried extracts of polymer materials used in the pharmaceutical industry. Ex-traction was first performed using acceler-ated solvent extraction (ASE). The work was performed in collaboration between John Colwell from Bespak Europe Ltd. and Ray Perkins, Keith Summerhill and Jona-than Angove from Anatune Ltd. in Cam-bridge, U.K., and it was reported in Chro-matography Today (Vol. 1, Issue 4, Sept./Oct. 2008, p. 17-19) as well as in Anatune Appli-cation note AS54 (www.anatune.co.uk).

Derivatizing fatty acids for GC/MS determinationAnimal and vegetable fats are key compo-nents in our nutrition, but lipids are also used in various industrial applications such as polymers used for packaging. This means that there is a significant analytical market for determination of fat content in foods and fatty acid profiles in both foods and polymers used for packaging. Fats and fat oils are mainly triglycerides, glycerol esters of monocarboxylic fatty acids (glycerol is also known as propane-1,2,3-triol). Most triglycerides are made up of three differ-ent, linear, saturated fatty acids, each with an even number of carbon atoms. Triglycer-ides of fatty acids cannot be analyzed direct-ly by gas chromatography (GC), they must first be hydrolyzed and derivatized. The es-ter bonds are hydrolyzed and the free fat-ty acids that are formed in the process are converted to the corresponding fatty acid methyl esters (FAMEs). FAMEs are moder-ately apolar and sufficiently volatile to be determined by GC or GC/MS. The derivati-zation step is typically quite labor intensive, which makes the work by Colwell, Perkins, Summerhill and Angove even more inter-esting. Using the PrepStation, the scientists implemented and automated a widely used manual derivatization method that is based on boron trifluoride and methanol (Jour-nal of Liquid Research, 1965. 5: p. 600-608). Using an established method as a base en-abled the authors to compare their results with those from existing methods. First the dried polymer extracts containing fatty ac-ids were placed in 10mL vials, and deuterat-

ed fatty acids were added as recovery stan-dards. All further steps were performed ful-ly automated by the PrepStation. The quan-tification was performed using 1-bromo-tetradecane as internal standard and cali-bration curves were prepared from FAME standards.

Technical DetailsAll steps of the derivatization process for the fatty acids were performed using the GERSTEL PrepStation, which has two in-dependent parallel rails, each fitted with in-dependent robotic towers capable of per-forming liquid handling steps.

The upper robot of the GERSTEL Prep-Station covers the entire spectrum of liq-uid handling. This includes liquid sample

introduction, the addition of an internal standard, dilution and/or derivatization. The lower robot complements the upper robot, enabling other types of analyte en-richment and sample introduction such as Headspace (HS), Solid Phase Micro-Ex-traction (SPME) or automated Solid Phase Extraction (SPE). “The MPS PrepStation enables efficient automation of complex tasks”, says Ray Perkins, owner and Gener-al Manager of Anatune: “Sample prepara-tion is performed during GC or LC analy-sis of the preceding sample, there is no loss of productivity, samples are prepared just-in-time for introduction to the GC or LC exactly when it is ready for the next run. This means that the analysis system is nev-er waiting idly for the next sample. Equally,

MPS PrepStation as it was used by Colwell, Perkins, Summerhill and Angove for derivatization of fatty acids.

Analytical conditions

Injection volume: 1 µL (10 µL syringe)

Column: Phenomenex Zebron ZB1, 30 m x 240 µm (0.1 µm Film)

Pre-column / Approximately 1 m x 0.53 µm I.D., deactivatedretention gap: fused silica

Carrier gas: Helium (1mL/min), constant flow, vacuum compensated.

Oven temperature program: 40 °C (1 min), 10 °C/min to 300 °C (5 min)

Detection mode: Selected Ion Monitoring (SIM)

Interface temperature: 280 °C

MSD setting: Standard Auto Tune (ATUNE)

MSD Solvent delay: 5 min


Acetone was used as a syringe rinsing sol-vent in the work reported here. Acetone is miscible with water and with most organic solvents in any ratio. This means that ace-tone is especially well suited for condition-ing and rinsing solvent syringes to avoid sample carry over and surface adhesion problems when changing from aqueous to oily phases and vice versa.

prepared samples are never kept waiting in the autosampler. The risk of decomposition of labile analytes or labile derivatized ana-lytes is thereby greatly reduced, all samples are treated exactly the same, which reduc-es the risk of variations in results.

The PrepStation used in the work re-ported here was equipped with a 1 mL sy-ringe and a 10 µL syringe respectively in the lower and upper robotic towers. Two heat-ed agitators and a Solvent Filling Station (SFS) configured with four solvent reser-voirs were mounted on the system as well. One solvent reservoir was filled with HPLC-grade water, one with acetone and one with internal standard in hexane. The derivati-zation reagent (BF3 in methanol) was kept in a separate 100 mL vial.

Intelligent scheduling of sample preparation and analysisThe GERSTEL MAESTRO software was used to control all sample preparation steps. The MAESTRO Scheduler provides a complete at-a-glance overview of sam-ple preparation and sample introduction timing including total sample preparation

and analysis time for all samples. This func-tionality facilitates planning and schedul-ing of the laboratory work load. The analy-sis was performed on a 6890 GC/5973 MSD GC/MS system from Agilent Technologies. The PrepStation can be used as a bench top WorkStation, independent of the GC/MS system or it can be mounted on top of the GC/MS system enabling it to perform syn-chronized, overlapping sample preparation and sample introduction in one automat-ed analysis system.

Automated derivatization and addition of an internal standard10 mL sample vials were manually placed on the MPS PrepStation. The vials con-tained ASE extracts that had been concen-trated by solvent evaporation. All further steps were fully automated: 1 mL of the BF3/methanol mixture was aspirated from the 100 mL storage vial and added to the sam-ple. The sample was then transferred to the agitator, where it was kept at 70 °C for 5 minutes under agitation before being re-turned to the sample tray. The 1 mL syringe was subsequently used to add 1 mL internal

Table 1 GC/MS Peak Areas for Methyl Esters from Manually Derivatized Extracts.

Table 2GC/MS Peak Areas for Methyl Esters from Prepstation Derivatized Extracts.

Table 3Comparison of Mean Peak Areas.

Acetone


In the work reported here, deuterated fatty acids were used as recovery stan-dards to demonstrate that the derivati-zation process had been completed sat-isfactorily.

As an aside, the widely used technique of adding deuterated internal standards to your sample brings a number of benefits: 1) The analyst can be almost certain that he or she is using internal standards that do not occur naturally; there can be little doubt that the concentration of the stan-dard compound is equal to what was add-ed to the sample. 2) The properties of the internal standard compounds closely re-semble the target analytes, which means that deviations in response factors, reten-tion times and recoveries will be minimal. 3) The isotopes effectively provide a quali-ty check on the analysis: Less check stan-dards will need to be run; productivity and sample throughput per instrument can be increased. 4) In general, using isotopical-ly labelled standards can serve as a cal-ibration or a calibration check: Less cali-bration standards will need to be run; pro-ductivity and sample throughput per in-strument can be increased.

Using isotopically labelled standards


son for this can be found in the intelligent MAESTRO PrepAhead function that en-ables the user to perform sample prepa-ration of one or more samples in parallel with ongoing GC/MS analysis. Using the PrepAhead function, samples can be pre-pared well ahead of the time when they must be ready for injection. This is a win-win situation: The chromatography system wins in terms of productivity; it never has to wait idly for the next sample. The sam-ples win in terms of uniform treatment: Ev-ery sample is introduced immediately after it has been prepared; this means that there is less risk of sample to sample variations in terms of, for example, analyte degradation. Of course, the laboratory gains in terms of productivity and quality of results.

The MAESTRO Scheduler even pro-vides a complete, at-a-glance overview of sample preparation and sample introduc-tion timing including total sample prep-aration and analysis time for all samples. This functionality facilitates planning and scheduling of the laboratory work load.

Higher recovery and improved accuracy through automation“This work has shown”, the scientists said, “that our proven manual derivatization method for methylation of free fatty acids can easily and successfully be automated”. Furthermore, a comparison between the re-sults obtained from manual and automat-ed derivatization procedures clearly showed the advantages of automation: “The results we got from the automated system with the MPS PrepStation showed better recovery and much lower RSDs for all compounds”, Ray Perkins stated, while noting that a part of the already low RSDs could even be at-tributed to the Accelerated Solvent Extrac-tion (ASE) procedure performed prior to the derivatization process.

Fatty acid methyl esters (FAMEs) are formed by esterification of fatty acids with methanol. The fatty acids are ini-tially formed when oils (triglycerides) are hydrolyzed. A triglyceride normally con-tains different fatty acids and a mixture of different FAMEs is therefore formed in the reaction.

Did you know? Bio diesel is a vegetable oil based fuel that is comparable to diesel fu-el even though it is not produced from crude oil, but rather from vegetable oils, most often rapeseed oil, or from animal fats. Bio diesel is considered a renewable source of energy; chemically it is based on FAMEs.

GC/MS Chromatogram (SIM-Mode) of a sample to which deuterated fatty acids have been added as recovery standards. Peaks in the order of elution: Methylmyristate (deuterated), Methylmyristate, 1-bromotetradecane (internal standard), Methylpalmitate (deuterated), Methylpalmitate, Methylstearate (deuterated), Methylstearate.

Bio diesel

Fatty acid methyl esters (FAMEs)


standard (1-Bromotetradecane in hexane) to the sample. The deri vatization reaction was stopped by adding 3 mL HPLC-grade water to the sample.

Partitioning of FAMEs followed by sample introduction The FAMEs were partitioned into the or-ganic hexane phase, the process was accel-erated by agitating the vial in the second ag-itator at room temperature for 35 minutes. Following an equilibration time of 1 min-ute in the sample tray, the organic phase

that contained the FAMEs had separated out and settled. Using the 10 µL syringe, the PrepStation aspirated 1 µL of the or-ganic phase and introduced it to the GC/MS system.

High sample throughput and accurate results „Since the sample preparation steps for this analysis require much more time than the GC run, the PrepStation gives us signifi-cant time savings and it greatly improves productivity”, says Ray Perkins. The rea-


GERSTEL Solutions Worldwide Innovation

M onomers are generally low molecular weight compounds with spe-

cial functional groups that enable po-lymerization. Depending on the structure

and properties of monomers, and on the conditions chosen, polym-erization can lead to the forma-tion of linear, branched, or cross-linked polymers, which have dif-ferent chemical and physical prop-erties. In order to determine the structure of a polymer, pyrolysis GC is often used as the technique of choice; it is a powerful tool in the characterization of complex poly-mers whether they are in solid or liquid form or in emulsion.

Curie-Point pyrolyzers are widely used for polymer analysis. Some are based on re-sistive heating, some on microwave tech-nology. Technical aspects aside, the use of special pyrolyzers can be labor intensive, sometimes requiring additional cumber-some sample preparation steps as well as significant added investment volume. A different, simpler, and more cost-effective way has been described by scientists from

Polymer analysis

Efficient Automated Pyrolysis GC

Scientists from Dow, a leading producer and supplier of chemicals and polymer products, have collaborated with GERSTEL scientists in devel-oping a novel method for determining the structure and composition of polymers. The approach: Following a high-temperature liquid sam-ple introduction, pyrolysis is performed in the high-temperature version Cooled Injection System 6 (CIS 6) GC inlet. Pyrolysis break-down prod-ucts are subsequently determined using GC/FID or GC/MS.

DOW and GERSTEL who cooperated on developing an attractive alternative to stan-dard methods.

“Instead of a dedicated pyrolyzer, we used the CIS 6, the high-temperature ver-

sion of the GERSTEL Cooled In-jection System (CIS)”, says Pat-ric Eckerle, Dow Germany. The GERSTEL CIS is the most widely used PTV-type inlet in the world. Liquid polymers and polymer mixtures were pyrolyzed directly in the GC inlet in an oxygen-free carrier gas atmosphere. Pyrolysis break-down products were then transferred to the GC column,

separated and determined using a Flame Ionization Detector (FID).

To prove the validity of the method, the scientists analyzed different polymer mix-tures: 1. An emulsion based on a 1:100 styrene-

butadiene polymer mixture diluted with water was examined in order to determine recovery rates. In this con-text, Eckerle examined the influence of the GERSTEL CryoTrap System (CTS) on the quality of the separation.

2. A styrene-butadiene polymer mixture containing varying amounts of emul-gated copolymer (butylacrylate-sty-rene) was analyzed in order to verify the quantitation.

3. Polyethylene (PE), dissolved in hot xy-lene, was analyzed using two-dimen-sional GC (2D GC and GCxGC).

For the polymer analysis, the experts used a GC system with a GERSTEL Multi-Purpose Sampler (MPS), which was used for automated sample preparation and sam-ple introduction. The MPS was equipped with a headspace syringe adapter, a heated 10 µL syringe and a heated agitator. The GC 6890 from Agilent Technologies was fitted with a GERSTEL CIS 6 programmed tem-perature vaporizer as well as an FID.

And this is how Eckerle and his col-leagues approached the task: Following sample preparation, 0.5 to 2 µL of the dis-solved polymer was introduced into the cool CIS liner. Solvents were then purged from the inlet through the split vent, leav-ing the polymer material condensed on the CIS liner walls. The temperature of the CIS was kept at 90 °C for 3.5 minutes during the

Improved peak shape: The GERSTEL CryoTrap System (CTS) significantly improved both separation capacity and accuracy in the determination of volatile pyrolysis fragments by focusing these and sharpening the peaks.

First class results: Repeatability of 10 CIS pyrolysis runs of an S/B copolymer. The CIS was not cleaned or replaced over the course of these runs.


Polymer analysis and the instrument set-up used: Agilent Technologies 6890 GC with GERSTEL CIS 6 inlet and FID as well as a MultiPurpose Sampler used for automated sample preparation and sample introduction. A GERSTEL CryoTrap System (CTS) was used to focus and improve the determination of volatile pyrolysis breakdown fragments.

Success all around: The repeatability experiments gave good results and even after ten CIS Pyrolysis runs the liner needed neither replacing nor cleaning. “No trace of carry over between samples” said Dirk Bremer, GERSTEL R&D Manager, shown to the right in the picture along with Patric Eckerle from DOW.


solvent purge step. It was then programmed at 10 °C/min to 600 °C, a temperature suf-ficiently high for complete pyrolysis of the sample. The temperature was lowered after a one minute hold time.

The separation was performed using GC with and without column switching based on the following columns: HP 5 ms, 30 m x 0.25 mm I.D. x 0.25 µm film thick-ness from Agilent Technologies; in the 2D GC (GC x GC) setup, the following column was also used: Zebron ZB 50 ms, 30 m x 0.25 mm ID x 0.25 µm film thickness from Phenomenex. The oven temperature was kept at 50 °C for 6 minutes and then pro-grammed at 15 °C/min to 325 °C (8 min). The split flow was 20 mL/min and the FID temperature was set to 330 °C (standard gas flow conditions). Quantitation was per-formed based on standard addition.

The purpose of the exercise„Using the CIS 6 as a pyrolysis module was a complete success“, Patric Eckerle said. „A butylacrylate/styrene copolymer in a sty-rene/butadiene copolymer was determined qualitatively and quantitatively with min-imal method development time. In addi-

tion, pyrolysis patterns of standards con-taining different amounts of butylacry-late/styrene copolymer in a styrene/buta-diene copolymer were successfully repro-duced. Key fragments such as butanol and butylacrylate were identified by GC/MS and when we added a GERSTEL CryoTrap System (CTS), peaks were sharpened sig-nificantly, enabling us to improve separa-tion and to focus and accurately determine volatile pyrolysis products”.

Reproducibility and repeatability were excellent, and even after 10 pyrolysis runs, it was not necessary to clean or replace the CIS liner. “There was no sign of memory effects”, said Dirk Bremer, GERSTEL R&D Manager. “The CIS 6 - Pyrolysis - GCxGC system we used for polymer analysis gave a high peak yield and the correlation be-tween the peak areas and control standards was outstanding”. J. Sep. Sci. 2008, 31, 3416-3422.

The conclusion drawn by Mr. Eckerle is that CIS 6 pyrolysis is well suited for sever-al things: The determination of monomers in polymer mixtures; to gain information on micro-structures; and to identify addi-tives in polymers. “Our CIS 6 based pyroly-

sis method is efficient, fast and inexpensive - and it requires much less manual sample preparation than standard methods”, says Eckerle, while adding: “It is also very prom-ising that the method is faster than many spectroscopy-based methods we have oth-erwise used and the quality of the data is outstanding. You could literally place py-rograms on top of each other with a perfect match and we were able to get both quali-tative and quantitative results”.

Successful quantitation: Peak pattern obtained following pyrolysis of a synthetic standard. Standards with various concentrations of butylacrylate and styrene in a styrene/butadiene copolymer were pyrolyzed in the project. Results were obtained faster than when using spectroscopic methods. The key fragments butanol and butylacrylate were simultaneously identified by GC/MS.

Sty

rene

But

ylac

ryla

te

But

anol

Pyrolysis of polyethylene using the GERSTEL CIS 6 inlet. Overlay of two successive pyrolysis GC runs. The sample was dissolved in hot toluene. Agitator temperature: 125 °C; Syringe temperature: 125 °C.

Pyrolysis GCxGC enables high performance polymer analysis: 2D chromatogram of polyethylene fragments shown in a 3D presentation. The identification of micro-structures, information on monomers used, as well as the identification of additives, is significantly improved compared with standard GC chromatograms.


Forensic Sciences and Toxicology

Drugs of abuse: Extraction in seconds

In Forensic Science and in Toxicology, body fluids are regularly analyzed for residues of drugs of abuse, therapeutic drugs and of their metabolites. In general this type of analysis requires extensive sample prep-aration. At PittCon 2009, GERSTEL is presenting automated Disposable Pipette Extraction (DPX). DPX is a fast and efficient SPE technique used for a wide range of applications such as drugs of abuse, therapeu-tic drug monitoring, comprehensive screening, pharmacology studies (NNK), as well as pesticides in fruit and vegetables. DPX is based on unique and patented SPE devices: Pipette tips that incorporate loose-ly contained sorbent material, which is mixed with the sample solution. Turbulent air bubble mixing cre-ates a suspension of sorbent in the sample ensuring optimal contact and highly efficient extraction. The extraction is performed much faster than with traditional SPE techniques.

B ody fluids represent a complex and heterogeneous matrix. Accurate de-termination of drugs, pharmaceu-

ticals and metabolites in blood and urine requires both a suitable chromatographic system and adequate sample preparation. Sometimes more than one extraction tech-nique is needed for a successful result. Solid Phase Extraction (SPE) is among the most widely used sample clean-up and analyte extraction techniques in forensic and toxi-cology laboratories.


Traditionally, SPE requires the use of significant quantities of solvent, some of which are toxic. Following several labor in-tensive steps, many methods require that the solvent be evaporated in order to con-centrate the analytes of interest and achieve the necessary detection limits. Depending on the chemical properties of the analytes, a chromatographic determination may al-so require further sample preparation steps such as derivatization. The sum total of sample preparation steps can amount to a

significant bottleneck for laboratory pro-ductivity and a risk to occupational health unless adequate and costly safety precau-tions are taken.

If the tedious and labor intensive steps can be eliminated, the overall task can be performed more efficiently and faster while producing accurate results and using on-ly a fraction of the amount of solvent nor-mally used. All this is possible thanks to the novel Disposable Pipette Extraction (DPX) technique developed by Professor William

All steps are performed automatically by the MPS.If needed, the sorbent is conditioned with solvent prior to the extraction process.

1 Sample is drawn into the pipette tip for direct contact with the solid phase sorbent. There is no contact between the sample and the syringe used to aspirate the sample and therefore no risk of cross contamination.

2 Air is drawn into the pipette tip from below through the frit. Turbulent air bubble mixing creates a suspension of sorbent in the sample, ensuring optimal contact, highly efficient extraction, and high recovery.

3 The extracted sample is discharged, typically after 30 seconds.

If needed, the sorbent can be washed to remove unwanted residue.

4 Extracted analytes are eluted using a suitable solvent, which is added from above for most efficient elution. The eluate is collected in a vial for subsequent sample introduction to LC/MS or GC/MS.

The total time required for extraction in the examples shown in this article was always less than 6 minutes. Sample preparation and GC/MS or LC/MS determination can be performed in parallel for best possible throughput and system utilization.

Automated DPX process



In contrast to conventional SPE, DPX does not rely on standard cartridg-es with a packed sorbent bed. DPX is based on unique and patented SPE devices: Pipette tips that incorporate loosely contained sorbent material, which is mixed with the sample solution. DPX is a dispersive SPE tech-nique, turbulent air bubble mixing creates a suspension of sorbent in the sample ensuring optimal contact and highly efficient extraction. Extractions are performed much faster than with traditional SPE techniques. Elution requires only a small amount of solvent, which means that DPX effectively provides a concentration step: For many applications, such as pesticides

in fruit and vegetables, solvent evaporation is not required. DPX methods are readily automated using the GERSTEL MPS, which can

introduce the extract into a GC/MS or LC/MS system. Additional sample prepa-ration steps can be performed, including derivatization or adding an internal stan-dard. The analyst only needs to place the samples in the MPS autosampler and activate the sequence table from the MAESTRO software. Everything else is performed automatically including GC/MS and LC/MS analysis.

The patented DPX tips comprise loose solid phase sorbent contained inside a pipette tip fitted with a frit at the bottom and a polymer barrier at the top. The barrier mounted in the upper opening of the DPX tip has some additional functions: It is used as a transport adaptor, enabling the GERSTEL MPS to move the tip around and fully automate the DPX process. An orifice in the transport adapter enables the autosampler syringe to deposit liquids and to aspirate air through the sam-ple/sorbent suspension for highly efficient mixing and extraction. „DPX is a unique and patented extraction technique that provides the user a previously unheard of level of automation, efficiency and throughput“, says Ralf Bremer, Managing Director for R&D and production. GERSTEL has exclusive rights to sell automat-ed DPX instrumentation world-wide.

The DPX process is wonderfully simple: DPX tips are placed on the sample tray of the MPS. Aspirating the sample; adding liquids for conditioning, rinsing or elution; and aspirating air through the sample/sorbent suspension for efficient mixing – all this is performed by the microliter syringe in the autosampler. Depend-ing on the application, the sorbent is conditioned using a suitable solvent: This is aspirated into the DPX tip out of a vial or added from above. The defined amount of sample is aspirated into the DPX tip without making contact with the syringe or syringe needle. “There is no cross-contamination between samples”, says Ralf Bremer and notes: “Furthermore, since DPX is a dispersive SPE technique, it is not affected by flow rates or by changes in the flow path (i.e. channelling) through the sorbent. Such things have no impact on the extraction efficiency.“

When the DPX tip is safely retracted from the sample vial, air is aspirated through it, through the polymer frit and through the sample inside. “Fine air bub-bles quickly rise through the sample / sorbent suspension resulting in extreme-ly efficient, turbulent mixing. This significantly accelerates the extraction of ana-lytes onto the sorbent”, says Bremer. The extraction is performed under optimal conditions while requiring much less sorbent material than other SPE techniques. The extracted sample is now discarded into a waste vial. A rinse step follows and the extracted analytes are then eluted into an empty vial using a suitable solvent. The DPX tip is discarded and the MPS is ready for the next sample.

When performing SPE extractions manually, it is often necessary to concen-trate the extract by evaporating the solvent under an inert atmosphere and then adding a keeper solvent to take up the residue. Such a labor intensive and time consuming step is not required when performing fully automated DPX: “The ex-tract is simply introduced into the GC/MS system using a Large Volume Injection (LVI). The solvent is evaporated in the GC inlet and the analytes are concentrat-ed inside the inlet liner, provided you have a temperature programmable GC inlet such as the GERSTEL Cooled Injection System (CIS)”, says Ralf Bremer.

Disposable PipetteExtraction (DPX)

(Bill) E. Brewer, Ph.D. from the University of Southern Carolina. Professor Brewer is the Owner-President of DPX Labs (www.dpxlabs.com).

The DPX technique has now been auto-mated by GERSTEL, a leader in automation of sample preparation and sample intro-duction for GC/MS and LC/MS. GERSTEL is presenting automated DPX at PittCon 2009 in Chicago after it was initially intro-duced at the annual meeting of the Society of Forensic Toxicologists (SOFT) in Phoe-nix, Arizona (USA) in October 2008.

„The reactions we got from the foren-sic scientists at the SOFT meeting“, says Ro bert J. Collins, Ph.D., President of GER-STEL, Inc. in Baltimore, MD, “lead us to be-lieve that automated DPX is seen by experts as a very promising alternative to standard extraction techniques”.

In order to provide an efficient solution for the determination of drugs and metab-olites in blood in a routine laboratory en-vironment, GERSTEL and DPX Labs LLC collaborated on automating the DPX tech-nique. “DPX immediately struck us as the right solution”, says Bob Collins. Further-more, for this application, samples should be prepared and derivatized just prior to analysis. Just in time sample preparation eliminates analyte degradation and under-reporting of concentration levels since no sample is waiting for an extended period of time in the autosampler before being ana-lyzed. Also, in order to optimize GC/MS system utilization and sample throughput, a sample should be ready for introduc-tion every time the GC/MS finishes its run and becomes ready for the next sample. In summary, performing GC/MS analysis and sample preparation carefully synchronized and in parallel benefits both the quality of results and the throughput. The MAESTRO software with its PrepAhead and Schedul-er functions makes it extremely easy for the

Ralf Bremer,GERSTEL Managing Director for R&D and production



analyst to plan, optimize, and set up the whole process.

From theory to practiceIn order to test the MPS-DPX-CIS-

GC/MS system in practice, the scien-tists analyzed blood and urine samples that had been spiked with different drugs and pharmaceuticals. Compounds deter-mined included amphetamines, benzodi-azepines, cocaine and methadone as well as tetrahydrocannabinol (THC) and me-tabolites. For details, please see the graph-ic representations on this page. The analy-sis was performed using deuterated inter-nal standards: For blood samples, d5-PCP (0.2 ppm) was used. For the determination of benzodiazepines, d5-Nordiazepam (0.2 ppm) and d5-OH-Alprazolam (0.2 ppm) were used. For the opiates, equivalent deu-terated compounds were used (each at a level of 0.1 ppm).

Required manual sample preparation steps Preparing blood samples: 0.5 mL of ace-tonitrile was added to a 0.25 mL sample of whole blood followed by mixing to precip-itate proteins in the sample. The mixture was centrifuged and the supernatant trans-ferred to a clean labelled test tube contain-ing 0.1 mL of 0.1 M HCl.

Preparing urine samples for the deter-mination of benzodiazepines: In order to determine the total level of free, bound and metabolized residues of drugs and pharma-ceuticals in urine, hydrolysis must be per-formed of the respective conjugates, such as for example glucuronides of benzodiaz-epines, which are metabolites of the drugs that are formed to facilitate excretion of the substances from the body.

The hydrolysis reaction is started by add ing 10 µL of a solution of the enzyme b-glucuronidase and 50 µL of a 0.1 M sodi-

um phosphate buffer with pH 4 to a 0.2 mL sample of urine. The mixture is kept at 55 °C for two hours and is then allowed to cool to room temperature. Acetonitrile (0.25 mL) is then added in order to preci pitate the enzyme. Following centrifugation, the su-pernatant is transferred to a clean, labelled test tube and 200 µL of 0.1M HCl is added. The prepared samples are then placed in the MPS sample tray.

Automated Disposable Pipette Extrac-tion (DPX) is subsequently performed on the prepared samples using the MultiPur-pose Sampler (MPS) equipped with 1 mL CX tips from DPX Labs (www.dpxlabs.com). As the name suggests, CX tips con-tain a novel and unique cation exchange ma-terial with additional slightly apolar cha-racteristics. The DPX process is complete-ly automated: 250 µL of a 30 % solution of acetonitrile in water is dispensed onto the DPX sorbent inside the tip for condi-

Drugs of abuse in blood: Total ion chromatogram of a DPX extract of 250 µL whole blood spiked at 0.5 ppm with drugs of abuse using d5-PCP as internal standard. Chromatograms from the 5th and 20th injections are shown alongside each other to demonstrate the ruggedness of the analysis. The insert shows the extracted ion chromatogram (EIC) from the 5th injection. The sample was protein precipitated with 0.5 mL acetonitrile, and the supernatant was transferred to clean tubes. After adding 0.1 mL of 0.1 M HCl, automated DPX was performed. 1) Meperidine, 2) d5-PCP (ion 205), 3) PCP, 4) Methadone, 5) Methaqualone, 6) Amitriptyline, 7) Cocaine, 8) cis-Doxepin, 9) Imipramine, 10) trans-Doxepin, 11) Desipramine, 12) Pentazocine, 13) Codeine (Septum bleed from vial cap after repeat injections from the same vial).

EIC

TICDrug Average RSDAmphetamine 13.9 %Methamphetamine 14.4 %Meperidine 5.8 %PCP 2.2 %Methadone 3.3 %Methaqualone 3.6 %Amitriptyline 3.1 %Cocaine 3.8 %Cis Doxepin 2.8 %Imipramine 3.3 %Trans Doxepin 3.2 %Pentazocine 5.3 %Codeine 4.2 %Desipramine 6.4 %

CIS: 1 min solvent purge (150 mL/min); splitless; temperature program: 80 °C (1 min) – 12 °C/min – 300 °C (3 min)

Column: 30 m HP5 (Agilent Technologies); di = 0.25 mm; df = 0.25 µm

Pneumatics: He, constant flow, 1.5 mL/min

GC oven: 100 °C (0.5 min) – 20 °C/min – 300 °C (12.5 min)

MSD: Selectable Full Scan or SIM mode

Analysis conditions



tioning. The conditioning solvent is subse-quently discarded to waste. The DPX tip is then immersed into the sample and a de-fined volume is aspirated into the tip. Air is then aspirated into the tip, causing turbu-lent mixing and efficient extraction of an-alytes into the sorbent. Following a 30 sec-ond equilibration time, in which the sorbent is allowed to settle, the extracted sample is discarded to waste. The sorbent is rinsed twice, first with 0.5 mL of a 30 % solution of Acetonitrile in water and then with 0.5 mL acetonitrile. The extracted analytes are elut-ed using 0.7 mL of a solution consisting of 2 % concentrated ammonia, 78 % CH2Cl2 and 20 % isopropanol.

The eluate was dispensed directly into an autosampler vial. The total amount of time required for extraction and liquid han-dling was less than 6 minutes per sample.

InstrumentationThe analysis was performed on a 6890N/5975 (inert XL) GC/MS system from Agilent Technologies. The GC was fitted with a Cooled Injection System (CIS 4) PTV-type inlet. An MPS PrepStation with DPX option and MAESTRO Software control was used for automated sample preparation and sam-ple introduction. The complete system in-cluding GC/MS was operated using one in-tegrated method and one sequence table di-rectly from the Agilent Technologies Chem-Station Software, operated integrated with the MAESTRO software.

DerivatizationSome analytes must be derivatized to enable GC/MS determination. „The Cooled Injec-tion System (CIS) inlet offers an inert and temperature programmable environment“, says Prof. Brewer, „which is highly suited

Benzodiazepines in urine: Total ion chromatogram of 0.2 ppm benzodiazepines in 0.2 mL urine following enzymatic hydrolysis and DPX. Derivatization was performed in the CIS inlet by injecting 50 µL of DPX eluent together with 20 µL of 50/50 MTBSTFA/acetonitrile. No separate solvent evaporation step was performed. Increasing the sample volume to 0.5 mL and performing multiple DPX extractions would increase the sensitivity. 1) Diazepam, 2) Nordiazepam-d5-TBDMS, 3) Nordiazepam-TBDMS, 4) Flunitrazepam, 5) 7-aminoflunitrazepam, 6) Oxazepam-2TBDMS, 7) Temaze-pam-TBDMS, 8) Nitrazepam, 9) Lorazepam-2TBDMS, 10) Clonazepam-TBDMS, 11) Alprazolam, 12) a-OH-Alprazolam-d5-TBDMS, 13) a-OH-Alprazolam-TBDMS

THC and metabolites in blood: Total ion chromatogram of 10 ng/mL THC and metabolites extracted from 0.5 mL whole blood following protein precipitation, centrifugation and DPX-RP. Derivatization was performed in the CIS inlet by injecting 50 µL of DPX eluent together with 20 µL of 50/50 BSTFA/acetonitrile. No additional solvent evaporation or derivatization step was performed. Analytes: 1) THC-TMS, 2) OH-THC-TMS, 3) COOH-THC-2TMS

to evaporating and purging excess solvent while simultaneously, or at least sequential-ly, performing derivatization of analytes.“

For the derivatization of benzodiaze-pines, 20 µL of 50 % N-(t-butyldimethyl-silyl)-N-methyl-trifluoracetamide (MTB-STFA) in acetonitrile was aspirated into the autosampler syringe followed by 20 µL of air and 50 µL of the DPX eluate. „The resulting „Sandwich“ injection was performed slow-ly, using a programmed stop flow method to ensure that the solvent was completely removed through the split vent prior to the derivatization step”, the application special-ist explains. The CIS temperature quickly ramped to 300 °C, which started the deriva-tization process and helped transfer the de-rivatized analytes to the GC column in split-less mode for highest possible recovery and lowest limits of determination.

“Automated analyte derivatization in the GC inlet proved to be both simple and highly practical”, said Prof. Brewer. “The method was successfully applied to the de-termination of benzodiazepines in blood. Compounds that were not successfully de-rivatized in this way were derivatized direct-ly in the sample vial”. For this approach, the DPX eluate was evaporated to dryness un-der a flow of nitrogen in the sample vial. 50 µL of MTBSTFA and 50 µL of ethyl acetate were added and the mixture kept at 70 °C for 20 minutes. When the extract had cooled off, 50 µL of the solution was introduced to the CIS inlet using the Large Volume Injec-tion (LVI) technique.

The conclusion reached by the experts“As we had expected, the analysis based on automated DPX delivers excellent results”, said Bob Collins. Even though all analyses

were performed on very small sample vol-umes (250 µL blood or 200 µL urine), the resulting peak intensities were highly satis-factory – even in full scan mode. The Mul-tiPurpose Sampler (MPS) with automated DPX enables fast sample preparation of dif-ficult samples while delivering high sensi-tivity and accurate results. The additional liquid handling capabilities of the dual rail MPS PrepStation enabled full automation of all the required liquid handling steps such as derivatization and addition of an inter-nal standard. This added level of automa-tion provided best possible productivity and throughput. The instrument combination used proved to be especially useful for the determination of basic drugs such as co-caine, methadone, PCP, TCAs and Mepe-ridine.

Most benzodiazepines were easy to de-termine using MTBSTFA derivatization in the GC inlet. The following were deter-mined: Diazepam, Nordiazepam, Oxaze-pam, Temazepam, Alprazolam and a-OH-alprazolam. “DPX combined with GC/MS determination provided excellent results for the 11 listed benzodiazepines in urine“, Bob Collins notes, ”plus we got good recov-ery and great sensitivity for the opiates. For most opiates, we achieved limits of deter-mination under 1 ng/mL in whole blood”. Ralf Bremer, General Manager for produc-tion and R&D, is thrilled about the use of the CIS 4, PTV-type inlet for evaporative con-centration and analyte derivatization: “This year, we are celebrating the 25th anniversa-ry of the introduction of the first GERSTEL CIS. It is very reassuring to see that the con-stant improvements we have engineered in-to the CIS over the years enable us to stay well ahead of the competition”.


GSW: You have been with GERSTEL Inc. for 15 years, you are so to speak a man of the first hour, and you have been in charge of steering the compa-ny for a number of years. Please describe to us how the company has developed? Bob Collins: In the beginning it was a very small operation of just two people. The challenge was covering the entire country instantly. This had to be done even though we were so small, since U.S. companies typically have a country-wide presence and need to replicate their so-lutions in other branches as well. So you can’t just sell in Baltimore, you also need to be able to support customers in Los Angeles, which is a 6-7 hour flight away. In the beginning, I was the sales person, the service engineer and the technical support person all in one. With such a small staff it was a real challenge initially, but as sales grew, we added people and things became easier. We have had organic, steady growth, adding people as sales grew while doing our best to provide world-class support along with the world-class prod-ucts and solutions GERSTEL offers.

What is the distinguishing feature of GERSTEL, Inc.? What are the strengths of your compa-ny and what is the added value you offer your customers? The unique feature of GERSTEL is of course the products themselves; they do things that no other products can. They provide un-equalled sample preparation capabilities that are easy to use due to our advanced software. They allow the analyst to achieve very low detection limits in even the most difficult sample matrices; within the field of chemical analysis our products and so-

lutions are truly unique. Additionally, we offer comprehensive application support,

technical training and service support. Our customers con-sistently tell us that we do this better than anyone else, in the US alone, we have five applica-tion scientists. Our service en-gineers are excellent, and our sales people are very experi-enced and knowledgeable. This can be a great help to customers when they are trying to deter-mine which GERSTEL product best suits their needs. All system purchases from GERSTEL in the U.S. include a training course.

We have just moved to new headquarters with a wonderful new training facility that has greatly improved our capacity. Also, ev-ery time you call GERSTEL, you get an actu-al person on the line. You can speak directly to an application scientist or a service engi-neer; you are not just asked to leave a mes-sage or just given a reference number with the message that you will be called back. We try to give personalized high-level support in all aspects of the business.

In order to be innovative and to offer custom-ers added value, a company needs to invest in research and development. What is GERSTEL doing in that field? We are doing a lot! GERSTEL, Inc. is a sub-sidiary, product development is performed in Germany, but we are very active in method development and provide ideas for product development based on input from our U.S. customers and our sales and support staff. This combined with input from Japan and Germany allows GERSTEL to constantly re-fine existing products and develop new ones. If you look at GERSTEL products and solu-

tions today, you can see we’ve come a long way from just GC inlets. We now offer a wide range of sample preparation and analyte en-richment capabilities that can be used in al-most all areas of GC and LC analysis.

Speaking of innovations, you are a long time PittCon exhibitor, which products are you high-lighting this year?The main thing we will be highlighting is the new automated DPX technology, which we are extremely excited about. DPX is an ab-breviation of Disposable Pipette Extraction, it is essentially a miniaturized, fast SPE tech-nique. We are also showing a host of other new solutions: Automated SPE; Automated SPME Fiber Exchange, a multi-fiber system that will be a useful tool especially for the food, flavor and beverage industry. It enables the use of different SPME phases in one au-tomated sequence for flexible and efficient screening and analysis.

Will any other innovations from the past year be shown? A lot of automated sample preparation tech-niques have been developed over the past few years that we will present such as an automated blood analyzer with centrifuga-tion, sonication, sample heating and bar-code reading. We have also integrated sam-ple weighing. We have a great tool kit for au-tomated sample preparation.

Better than the competition?I would say yes, since it is much more mod-ular, easier to customize to the exact needs of each laboratory and surprisingly inexpen-sive compared with other robotic solutions. And since everything is controlled through our powerful, yet easy to use, MAESTRO software, it is really amazing how quickly we can put a solution together.

GERSTEL Solutions Worldwide Interview

Interview with Robert J. Collins, GERSTEL Inc., USA

Looking forward to shaping the future of the business

GERSTEL is recognized as one of the leading providers of automated sample preparation and sample intro-duction solutions for GC/MS and increasingly for LC/MS. After building a reputation as a leader in special-ized solutions for GC in Germany and Western Europe, management decided to expand GERSTEL’s pres-ence to the United States. In 1994, the company took the plunge and GERSTEL, Inc. was founded. Today, almost 15 years later, GERSTEL, Inc. has moved into new larger headquarters near the Baltimore Wash-ington International Airport in order to provide the best possible support for its large and growing family of customers. GERSTEL Solutions worldwide spoke with long-time GERSTEL, Inc. President Robert (Bob) Col-lins, Ph.D., about how GERSTEL, Inc. developed over the past 15 years.

GERSTEL, Inc. President Robert J. Collins, Ph.D.


Why do you think that DPX will be a success?DPX is really a faster, more efficient form of the SPE technique. And it is much less sensitive to changes in method parameters such as flow rates, with which you have to be really careful when you do SPE. DPX is just so much quicker than SPE and that will be a key factor.

You introduced automated DPX at the Society of Forensic Toxicology (SOFT) 2008 annual meet-ing in the U.S. – what was the reaction?DPX was extremely well received; we had a great deal of interest from people visit-ing our booth, and our presentations. Since then, DPX Labs has developed even more phase materials and we have been working closely with them to provide automated so-lutions for the entire range of tips available. We have already sold some systems in the U.S. and are about to place a system into one of the largest drug testing laboratories in the United States.

Is DPX useful outside the field of Forensic Sci-ence? Oh yes, DPX is useful for anything where you need sample clean-up, extraction and con-centration. Areas that come to mind imme-diately are clinical analysis, foods, including the QuEChERS method, and beverages.

Looking back: Which were the most success-ful products from GERSTEL over the years? Where do the particular strengths of the com-pany lie?A big success has been what we call „bundled systems“, for example the Thermal Desorp-tion Unit (TDU) bundle, which is extreme-ly flexible. I’ve always said that a typical day has the analyst come to work in the morn-ing; and while having his or her first cup of coffee and thinking of what the plans are for the day, the first emergency comes through the door. The original plan is set aside, and the emergency must be dealt with. The TDU bundle enables the analyst to quickly set up and use any of eight different analytical tech-niques and allows them to quickly meet any challenge. I would have loved to have had one when I was active in the laboratory.

The U.S. is a geographically very large coun-try. How do you support customers through-out such a vast territory? What is your strate-gy for reaching every part of the map both in terms of sales and support?We use a hybrid system with GERSTEL em-ployed regional sales managers that work with independent manufacturers’ repre-sentatives to cover their territory. We have four regional managers and about 35 man-ufacturers’ representatives covering the U.S.

and Canada. Every region has direct service support as well as third party service engi-neers that support our customers through-out this huge nation.

Is it important for U.S. customers that GERSTEL products are made in Germany? I don’t think U.S. customers place a lot of emphasis on where a product comes from, as long as the quality is good. Of course, Ger-man engineering and German products, es-pecially cars, have a great reputation, but ev-ery company has to build its own reputation, and more importantly, maintain this reputa-tion by delivering quality products and ser-vice. If you look at our products, they defi-nitely live up to the expectations associated with “Made in Germany”.

GERSTEL develops, produces and markets ad-vanced instruments and technologies. These are used in many different industries, univer-sities, state and public laboratories, and on top of it all, they are used both for routine analysis and for research. How do you go about support-ing so many different types of users and keep-ing them satisfied? Before we sell a system we make sure that it fits the customer’s needs, and allows them to successfully do whatever they need to do. This eliminates a lot of problems up front.


Impressions of the new company building



We do not try to put product out there just to rack up big sales numbers, this doesn’t help the customer or us. Secondly, as I mentioned before, all system purchases from GERSTEL in the U.S. include a training course at our Baltimore facilities. This covers theory, hands-on instrument operation and main-tenance. This ensures that the user is fully trained to perform the analysis. Addition-ally, an extensive set of maintenance videos and user manuals help a lot. Thirdly, our users can always reach us over the phone or through e-mail. Most problems are lit-tle things that are quickly resolved. Fast as-sistance means that productivity is quickly restored. This saves a lot of everyone’s time and makes the customer very happy. Large companies simply cannot or just do not of-fer that level of support.

How well is GERSTEL accepted in the market place?Our recognition and acceptance has grown enormously over the past 10 years. In the food, flavor and fragrance markets, for ex-ample, almost everyone has our products.

In State Health labs across the nation and in some homeland security applications we are also well accepted. In a U.S.-wide project headed up by the Centers for Disease Con-trol, we equipped almost every State Public Health laboratory in the U.S. with a unified custom solution that included sample prep-aration, sample introduction and integrat-ed custom software for homeland security. There are about 100 of these units out there now. Apart from that, we have steadily built a presence in the U.S. and now have a loy-al following. Word of mouth also helps us grow – and we still have tremendous room to grow, which is a good thing.

What was your reason to move to new and larg-er headquarters? As we grew at our previous location, we add-ed more and more space as adjacent suites became available. The disadvantage of this approach is that the layout of the facility is always inefficient because you have to use the previous tenant’s design and it can only be changed by expensive and disruptive de-molition. In the end, the laboratory, train-

ing, and warehouse facilities were no lon-ger adequate for the size of the business, and before our lease was about to expire, I decided to look into options to allevi-ate the problems we were having. We were very fortunate to find an excellent proper-ty close to the Baltimore airport. We were able to design the interior from scratch and it has greatly improved our work-flow. We can now do a lot of things simultaneous-ly, which gives us more flexibility and bet-ter efficiency. In the end we are now able to provide even better support for our grow-ing number of customers.

Do the new headquarters mean that you’ll pursue a different corporate strategy? Which plans and goals do you have and what is the outlook? A big part of our plans with the new facil-ity is that we want to increase the amount of customer training and bring more cus-tomers in to see our instrumentation first hand. We also recently had an Agilent 6410 Triple Quad LC/MS/MS system installed in our applications laboratory in order to sup-port our expansion into the LC/MS market. We now have the facilities and the people we need to take the business to the next level – I am very much looking forward to shap-ing the future of the business ... ... and GERSTEL is the leader in automated sample preparation? Yes, I think we are; especially if you are talking about combined automated sam-ple preparation and sample introduction. When you combine our technologies and hardware with MAESTRO software con-trol and extensive applications support, it is fair to say that GERSTEL is a recognized leader.

Is it important in this context that GERSTEL is a family owned company? I think it is extremely important. When we make a decision to pursue an opportunity, we can take a much longer view than pub-licly listed companies and that has made a huge difference. Customer support and satisfaction is the overriding priority at GERSTEL, and as a family owned company, we can stick to this priority without pres-sure from share holders that are looking for short term gains. Our customers know and see this and it makes a big difference for em-ployees too; they are more involved, better informed and feel more secure.

When other companies are laying off employ-ees, do you see this as an opportunity?Yes, absolutely, and not just because it makes it easier for us to compete. It is a great opportunity to get excellent and expe-rienced new people on board to help GER-STEL continue the steady growth that we have seen for over ten years now.


GERSTEL Solutions Worldwide Report

W ine from ancient Egypt is thought to have been honey-sweet – though now it is just bone-dry. What was

once refreshing, stimulating and thirst-quenching has mostly evaporated; only dust and residues remain in the 3-5 millennia old amphorae that were found in the tombs of those ancient rulers and demigods, the Pha-raohs. On their way to the netherworld, they were given gold and ample riches along with food and amphorae filled with precious wine. One amphora was marked: “Year 5. Wine of the House of Tutankhamen, Ruler of-the Southern On, the Western River. By the chief Vintner Khaa.’’ (Source BBC).

Some tombs are embellished with wall paintings depicting scenes from ancient Egyptian vineyards (cf. picture on p. 22). From such graphical renderings, as well as from a separate hieroglyph for the word “wine”, archaeologists were able to deter-

Archaeology meets chemistry

Details on viticulture in ancient Egypt are quite well understood by modern-day archaeologists. But what exactly was in Pharaoh’s glass when he savored the gift of the wine gods – and was it just imbibed for relaxation and merriment or was it taken as a stimulating aphrodisiac or maybe prescribed by his physician to cure or alleviate pharaohnic ailments? Answers to these questions have eluded us for ages. When archaeologists recently consulted analytical chemists armed with thermal desorption GC/MS systems, information began to trickle out, offering insight into ingredients used in ancient Egyptian wine.

Peeking into Pharaoh’s glass*

* Ancient Egyptian sign for „Wine“

mine that grapes were being grown and wine produced as early as 3,000 B.C. in the Nile Delta (Lower Egypt). At that point a thriv-ing wine-producing industry controlled by the rulers had already taken root. Vines were planted in pits filled with fertile Nile river silt. Given sufficient irrigation, vines could be grown successfully in oases.

Sacrifices to the godsArchaeologists have found evidence that wine was well appreciated for festive occa-sions in ancient Egypt. The only drop of bit-terness in the chalice was that many an out-standing droplet was reserved for the gods and donated as sacrifice. We have until now relied only on speculation as to what Tut-ankhamen and his contemporaries imbibed when “communicating” with the wine gods. The uncovered amphorae have been com-pletely dry and empty; the wine evaporat-

ed an eternity ago. Not until chemists were called upon to inspect the grave goods more closely did hard facts begin to emerge. In the amphorae found in the grave of Tut-ankhamen, malvidine-3-glucoside was iden-tified among the remains (Armen Mirzoian et al., „Analytical Chemistry“, Vol. 76, No. 6, March 15, 2004).

This compound is one of the more stable anthocyanins, the group of compounds that lends a warm red hue to the class of wines known as red wines. The 18 year old Pha-raoh, in other words, had been given ampho-rae of red wine to accompany him, possibly wine that he had favored during his short life. As an aside, anthocyanins form the main group of flavonoids that, along with phenols, make up the class polyphenols, which are thought to have positive health effects.

Equally scientifically intriguing was the search for wine residues in 700 wine jugs found in Abydos, Egypt. The jugs had been dated to 3,150 B.C., around 1,800 years pri-or to the birth of Tutankhamen. They were found in what was probably the tomb of the first Egyptian Pharaoh, Scorpion, from the first dynasty. Initial research had revealed that the Abydos jugs had contained around 4,000 Liters (1,000 Gallons) of wine from the Valley of Jordan, about 600 km (400 miles) away.

The project described here was per-formed by scientists from the Museum of the University of Pennsylvania (MASCA) in Philadelphia, PA, and from the Beverage Al-cohol Laboratory in Beltsville, MD, part of the U.S. Alcohol and Tobacco, Tax and Trade

To look into the soul of a wine that no longer has a body, the scientists had no other option but to grind clay from the inside of the amphorae and jugs and extract wine-related substances from the granulate.

Polyphenols are universally praised for their positive health effects, in large part ascribed to antioxidant and radi-cal scavenging properties. One might also turn to grapes, raisins, black cur-rants, cranberries or elderberries for a less stimulating source. Most chemical research on polyphenols is reportedly performed on wine.



Mural in the tomb of Nebamun, soldier in the army of Pharaoh Thutmose IV (Photo: archive).

Section of „Papyrus Ebers“ (1,500 B.C.) a 20 meter long list of medicinal recipes, and thereby the most comprehensive documentation of medical knowledge in ancient Egypt known to man. The content mainly deals with internal diseases and their treatment. The papyrus was acquired for the University of Leipzig, Germany in 1872 by Georg Ebers (1837-1898), Professor of Egyptology.

Bureau (TTB). The scope of the project was to determine volatile and semi-volatile com-pounds in the wine residues, but not neces-sarily to determine the origins of the wine. In order to have a historically differentiated reference, a wine amphora from the Nubi-an town of Djebel Adda, dating back to the year 400 A.D., was analyzed as well.

Wine tasting based on pottery shardsTo look into the soul of a wine that no lon-ger has a body, the scientists had no choice but to grind the ancient pottery and extract the oenological residues from the resulting powder using acidic or alkaline solutions. The extracts were filtered and analyzed us-ing chromatographic techniques. The fol-lowing provides an overview of the meth-ods used for analysis.

Traces of resin and herbsThe results: In both samples, the scientists identified a range of terpenoids, esters and alcohols as well as various volatile com-pounds and L-tartaric acid. This was defin-itive proof that the amphorae and jugs had contained wine. Further, the identified com-pounds indicated that resin and herbs had been added to the wines, making them a kind of ancient day Retsina wine, possibly simi-lar to what is produced, and mainly served to tourists, in Greece today.

The project provided facts that sup-port the theory of a preference for wines enriched with resin and herbs at the court of the Pharaohs, covering the entire period from the beginning of the ancient Egyptian High Culture (Abydos find) until the latter parts (Djebel Adda find). The herbs may have been added mainly to produce a sought after taste or they could have been added for medicinal purposes. Herbs and tree res-

SPME Total Ion Chromatogram (top) of the Abydos sample and enlarged (12.00-13.60 min) Selected Ion Chromatograms (Middle), shown along with mass spectra and library mass spectra of select compounds (below).



RT (min) Compound Abydos Djebel Adda Possible origin

3.75 1-Hexanol x Wine5.81 Benzaldehyde x x Wine5.62 Camphene x Pine5.99 Heptanol x Wine6.27 Phenol x Mint6.48 Menthene x Mint7.68 p-Cymol x x Pine, Rosemary7.82 Limonene x Mint, Pine7.97 Benzyl alcohol x Wine9.27 1-Octanol x Mint, Wine9.96 Fenchone x Rosemary, Fennel, Sage10.09 2-Nonanone x 10.91 Phenethyl alcohol x Wine10.92 Fenchol x x Pine12.17 Camphene x x Pine, Mint, Wild Fennel, Sage, Mugwort, Rosemary12.53 g-Heptalactone x 13.05 Borneol x x Pine, Rosemary, Mint, Oregano13.31 1-Nonanol x Mint13.42 L-Menthol x Mint14.12 a-Terpineol x Pine, Mint, Wine14.46 Ethyloctanoate x Wine15.63 Cuminaldehyde x Rosemary16.41 Carvone x Mint, Yarrow, Wild Fennel, Sage, Mugwort17.57 Ethyl Salicylate x Wine17.67 Decanol x Mint18.55 Thymol x Mint, Wild Fennel, Sage, Basil, Thyme23.11 Ethyl Decanoate x Wine23.13 Vanillin x Rosemary, Thyme25.53 Geranyl Acetone x Rosemary35.34 Farnesol x Pine37.76 Benzyl Benzoate x Pine38.85 Ethyl Palmitate x x Wine45.76 Ethyl Stearate x x Wine46.28 Manoyl Oxide x Pine47.65 Biformene x Pine56.24 Methyl Dehydroabietate x Pine

Compounds in the jugs from Abydos and amphorae from Djebel Adda identified using SPME-GC/MS and Thermal Desorption GC/MS.

Solid Phase Micro-Extraction (SPME)A 50/30 µm DVB/CAR/PDMS fiber was used. The fiber was immersed in a sodi-um chloride solution containing the sam-ple powder inside the sample vial for 40 min. at a temperature of 80 °C. The con-centrated analytes were desorbed from the SPME fiber in the GC inlet for 3 min-utes at 250°C. The SPME process was automated using the GERSTEL Multi-Purpose Sampler (MPS).

Gas Chromatography / Mass Spectrometry (GC/MS)A GC/MS system consisting of a 6890 GC and a 5973 MSD, both from Agilent Technologies, was used. Separation was achieved using a HP 5MS column, 30 m x 0.25 mm ID x 0.25 µm film thick-ness. Analyte transfer was performed in splitless mode, the MSD was set to scan mode from m/z = 40 to m/z = 400. GC oven program was started at 60 °C and programmed to 240°C at 3 °C/min. Car-rier: Helium at 1.2 ml/min constant flow. Compounds were identified using mass spectral libraries and Kovats Retention Indices, calculated from a series of n-al-kanes from C5 to C22.

Thermal DesorptionResidues from amphorae and jugs were also desorbed, or thermally extracted, using a Thermal Desorption System (TDS) from GERSTEL. The desorption temperature was programmed from a 50 °C starting temperature to 250 °C at a rate of 50 °C/min.

Liquid Chromatography – Tandem Mass Spectrometry (LC/MS/MS)A Waters Acquity UPLC and a Micro-Mass Quattro Premier XE Triple Quadro-pole mass spectrometer were used. LC parameters: UPLC BEH C18 column. Isocratic flow at 0.20 mL/min, 98 % H2O : 2 % ACN, 0.1 % Formic acid. MS/MS: Electron Spray Ionization (ESI), Cap. 4.50 KV, CV 20 V, CE 16 V.

in were part of the ancient Egyptian phar-macopoeia as we have learned from 13 an-cient papyri with information on medicine and various recipes. Among these are the “Papyrus Smith” (2,500 B.C.), the “Papyrus Ebers” (1,500 B.C.) or the “Papyrus Hearst” (1,500 B.C.), all named after the people by whom they were later purchased. In “Papy-rus Smith”, diseases were clearly divided into incurable and curable afflictions; for the lat-ter group, systematic instructions for treat-ment were listed. Knowledge about anato-my and physiology (e.g. functions of organs) was, however, very limited, which means that physicians at the time were quickly out of options for effective treatment. In many cases patients were diagnosed as being pos-sessed by demons; prayers or redemptive magic was prescribed. Empirical work, reli-gion and authorized magic often went hand

in hand. A word on alcohol content of the wine in

ancient Egypt: Alcohol plays a useful role as an extraction solvent for, and carrier of, ac-tive compounds in herbal medicine. The in-toxicating role is of course equally well rec-ognized and this seems to have been a cher-ished side-effect to what the doctor ordered. Beer, not wine, was the national beverage in ancient Egypt, often used in religious cere-monies and as a meal-time beverage. Legend has it that Osiris, the god of the underworld, taught humans how to brew beer. Prepared

from malted barley, a type of wheat called emmer and date juice, beer was counted as a staple food on the same level as bread. Brew-ing beer was of course also a way of preserv-ing drinking water and keeping it from being infested with undesirable microorganisms. Those ancient Egyptians who could afford it often preferred to drink wine when they wanted to have a good old time. Almost four thousand years ago, an Egyptian teacher la-mented that one of his students was leading a debauched and alcoholized life. “Oh if only you would recognize that wine is a horror, if only you would forget the chalice”.

Multiple reaction monitoring (MRM) LC/MS/MS chromatogram traces of an L-tartaric acid standard (top) based on the m/z 149 and 87 molecular fragments. The middle and bottom traces are from the aqueous extracts of the samples from Abydos and Djebel Adda respectively.

Selected Ion Chromatogram of the peak at 23.13 min; retention time and mass spectrum match those of vanillin.


GERSTEL GmbH & Co. KGEberhard-Gerstel-Platz 145473 Mülheim an der RuhrGermany

+49 208 - 7 65 03-0+49 208 - 7 65 03 33

[email protected]

GERSTEL, Inc.701 Digital DriveSuite JLinthicum, MD 21090USA

+1 410 - 247 5885+1 410 - 247 5887

[email protected]


Subject to change. GERSTEL®, GRAPHPACK® and TWISTER® are registered trademarks of GERSTEL GmbH & Co. KG.Printed in Germany · 0309 · © Copyright by GERSTEL GmbH & Co. KG

GERSTEL K.K.2-13-18 Nakane, Meguro-ku152-0031 TokyoDai-Hyaku Seimei ToritsudaiEkimae Bldg 2FJapan

+81 3 57 31 53 21+81 3 57 31 53 22

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GERSTEL AGEnterpriseSurentalstrasse 106210 SurseeSwitzerland

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S ample preparation for GC/MS or LC/MS analysis can involve sample clean-up and

liquid handling processes, such as additions or dilutions, as well as analyte concentration steps to reach the required limits of detec-tion (LOD, see box).

Using the MAESTRO Software, sample preparation is performed while chromato-graphy of the preceding sample is in prog-ress to optimize through-put. Urgent samples can be inserted into the running sequence without slowing or halting the workflow. If the analy-sis has been stopped before comple-tion of the entire batch, an e-mail notification can be sent to one or more addresses enabling us-ers to quickly restore produc-tivity. Context-sensitive help is available for all functions and entry fields in the software and a remote support software tool is included. The user can enable the GERSTEL support team to view the soft-ware screen via internet for fast and effi-cient advice on trou-ble-shooting or for training as needed.

GERSTEL MAESTRO Software

Perfect synchronization is essential, when performing sample preparation and chromatographic analysis in parallel for optimized productivity. Automated sample preparation requires reliable and compatible hardware combined with integrated software control of the whole analysis process, including LC/MS or GC/MS analysis. The GERSTEL MAESTRO software helps analysts find the fastest route to the results. The complete process can be viewed using the graphical Scheduler to aid laboratory workflow planning and allows the analyst to monitor important details while viewing the whole picture.

Perfectly SynchronizedSample Preparation and Analysis

MAESTRO supports a range of automated sample preparation techniques for LC/MS and GC/MS:

• Liquidhandling(e.g.,addinganinternal standard, derivatizing, extracting or generating a series of calibration standards)

• Centrifugation,Sonication,Weighing and Bar Code Reading

• DisposablePipetteEXtraction(DPX)

• SolidPhaseExtraction(SPE)

• DynamicHeadspace(DHS)

• ThermalDesorption(TD)andThermal Extraction from µ-vials

• Twister/StirBarSorptiveExtraction (SBSE)

• HeadspaceandSolidPhaseMicro-Extraction (SPME)

MAESTRO operates independently, or fully integrated with the Agilent ChemSta-tion software, or coupled with the Agilent MassHunter software. Only one method and one sequence table is required to operate the complete system from sample prep to GC/MS or LC/MS analysis. Further information: www.gerstel.com/maestro_eng.htm

GERSTEL GmbH & Co. KGEberhard-Gerstel-Platz 145473 Mülheim an der RuhrGermany

+49 208 - 7 65 03-0+49 208 - 7 65 03 33

[email protected]

GERSTEL, Inc.Caton Research Center1510 Caton Center Drive,Suite HBaltimore, MD 21227USA

+1 410 - 247 5885+1 410 - 247 5887

[email protected]


Subject to change. GERSTEL®, GRAPHPACK® and TWISTER® are registered trademarks of GERSTEL GmbH & Co. KG.Printed in Germany · 0208b · © Copyright by GERSTEL GmbH & Co. KG

GERSTEL K.K.2-13-18 Nakane, Meguro-ku152-0031 TokyoDai-Hyaku Seimei ToritsudaiEkimae Bldg 2FJapan

+81 3 57 31 53 21+81 3 57 31 53 22

[email protected]

GERSTEL AGEnterpriseSurentalstrasse 106210 SurseeSwitzerland

+41 41 - 9 21 97 23+41 41 - 9 21 97 25

[email protected]

no. 9 march 2009...known as omega-6-fatty acids) among these linoleic acid and its derivatives, and...

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