Reporter

Volume 29.3

Improved Detection and Analysis of Airborne Contaminants

Liquid Chromatography Sample Preparation Gas Chromatography

Detection and identification of airborne pollutants leads to better quality of life.

Standards Accessories Chiral Chromatography

Reporter
Volume 29.3

Fully Supported Air Monitoring Solutions

Visit us on the web at sigma-aldrich.com/thereporter

Kristen Schultz Product Manager Sample Preparation and Purification Dear Colleague: Air monitoring has evolved from the first regulations on reduction of smoke emissions from industrial and transportation operations as early as 1890s; to the Clean Air Act of 1970, to the numerous ambient, indoor and occupational air monitoring programs that exist today for a wide range of pollutants. The formation of regulatory agencies such as the Environmental Protection Agency (EPA), Occupational Health & Safety Administration (OSHA) and National Institute for Occupational Safety & Health (NIOSH) has paved the way for occupational, consumer and environmental protection. These agencies provide the guidance and the required or recommended sampling media and methods to sample, collect, and accurately measure the concentration levels of these pollutants in the air. As a result, there is a wide variety of sampling media and analytical hardware and techniques required to accomplish these measurements. The most widely sampled chemical pollutants are volatile organic compounds (VOCs); benzene, toluene, ethylene, and xylenes (BTEX), semi-volatile compounds (SV); carbonyls (such as formaldehyde); ammonia, and sulfur compounds (hydrogen sulfide, sulfur dioxide) as well as hundreds of other compounds. For these pollutants there is more than one type of acceptable sampling media. Most methods specify a carbon, silica, or resin based adsorbent packed into a specific size glass or stainless steel tube that will collect these pollutants (compounds) when air flows through or analytes diffuse into the adsorbent bed(s) via active or passive means. These compounds are then extracted from the adsorbent bed via solvent desorption or thermal desorption and injected into the analytical instrument mostly by GC or HPLC. We have a comprehensive Air Monitoring website sigma-aldrich.com/air-monitoring which features a wide range of air monitoring products for active, passive (featuring radiello), and whole air sampling. If you are looking for specific industrial applications such as Vapor Intrusion or application in the Petrochemical Industry to name a few, we have featured Product Guides to assist you in selection of the sampling media as well as the recommended analytical GC and HPLC columns and air monitoring standards. We are the only fully supported Air Monitoring provider to offer not only the sampling media products, but also expertise in carbon adsorbent technology, analytical columns, and standards. We invite you to visit our website and find what you need or let us know if we can assist you in your air sampling application.

Table of Contents
Liquid Chromatography Ion Exchange Chromatography for the Characterization of Biotherapeutics ............................................20 Novel Performance-Tested Solvents for UHPLC Applications.................................22 Sample Preparation Simultaneous Evaluation of Odor Episodes and Air Quality in Urban Areas by Multi-Sorbent Sampling and TD-GC/MS Analysis ...................................3 Radiello Passive Samplers and Vapor Intrusion......................................................6 Gas Chromatography Comparison of the Supelco 37-Component FAME Mix on the SP-2560 and SLB-IL111 .....................................8 Edible Oil FAMEs on the SLB-IL111 ........11 Standards Prescreened, In-Stock Chemicals ............15 TraceCERT Organic and Inorganic CRMs ..................................................16 Primary Standards for the Analysis of Herbal Medicinal Products....................17 Accessories Quality Syringes for Gas Sampling.........14 Vial Accessories ..................................................16 Autosampler Vial Convenience Packs ..17 Chiral Chromatography Accessing the Benefits of Polar Organic Mobile Phases on Cellulose-Based CSPs for Chiral HPLC........................................18 Kind regards,

Kristen Schultz Product Manager, Sample Preparation and Purification kristen.schultz@sial.com

Reporter is published 5 times a year by Supelco Marketing, 595 North Harrison Road, Bellefonte, PA 16823-0048.

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Reporter 29.3

Simultaneous Evaluation of Odor Episodes and Air Quality in Urban Areas by Multi-Sorbent Sampling and TD-GC/MS Analysis
Contributed Article
The following was generated with the assistance of an outside source using Sigma-Aldrich products. Technical content was generated and provided by: E. Gallego 1*, F. X. Roca1, J. F. Perales1, X. Guardino2
1 Laboratori del Centre de Medi Ambient. Universitat Politcnica de Catalunya (LCMA-UPC). Avda. Diagonal, 647. E 08028 Barcelona. Phone: 34934016683, Fax: 34934017150, e-mail: Lcma.info@upc.edu Centro Nacional de Condiciones de Trabajo. INSHT. C/Dulcet, 2-10. E 08034 Barcelona. Phone: 34932800102, Fax: 34932803642, e-mail: cnctinsht@mtin.es

Sample Preparation

Multi-Sorbent Bed Adsorbent Tubes

A validated analytical method, based on thermal desorption (TD) coupled to gas chromatography (GC) and mass spectrometry (MS), is used for air-quality measurements of volatile organic compounds in indoor and outdoor air, including alkanes, aromatic hydrocarbons, aldehydes, alcohols, chlorides, esters, ketones, terpenes, amides, carbon disulfide and isocyanates (Table 1, Figure 1). Sorbent materials with a wide range of properties (surface and pores structure) are commercially available both for industrial/occupational and environmental applications (1, 2, 3). For air quality determination and pollution control a good combination of different sorbents (2) allows us the qualitative and quantitative determinations of a wide range of C2-C14 VOCs, around 200-300 compounds present in a standard urban sample. A multi-sorbent tube filled with carbon blacks Carbotrap (20/40 mesh, weak sorption strength, 70 mg) and Carbopack X (40/60 mesh, medium sorption strength, 100 mg) as well as Carboxen 569 (spherical carbon molecular sieve, 20/45 mesh, high sorption strength, 90 mg) (Supelco, Bellefonte, PA, USA) has been selected (4). Adsorbents are placed in glass tubes arranged from weak to strong sorption strength (sampling direction, desorption in reverse). Air quality criteria can be applied to the chemical compounds concentrations obtained with the presented methodology.

klaus.buckendahl@sial.com

Odor Episodes and Air Quality

Odor episodes and environmental air quality are topics of worldwide concern, mainly due to industrial facilities in the vicinity of inhabited areas. Several atmospheric pollutants, mainly volatile organic compounds (VOC), are responsible for odor episodes of varying degrees of annoyance. Interest in determining VOC both in indoor and outdoor air has increased over the last several decades, as they are known precursors of photochemical smog formation as well as can cause shortand long-term adverse health effects (irritation of mucous membranes, psychological stress and long-term toxic reactions) and discomfort (bad odors). Generally, specific atmospheric pollutants are established and measured chemically in real-time outdoors and indoors. Indoor air quality (residential, commercial, office and public buildings) would depend mainly on the emissions from sources like building materials and appliances, consumer products, tobacco smoke and intake of outdoor air.

Sampling
VOC are dynamically sampled connecting custom packed glass multi-sorbent cartridge tubes (Carbotrap, Carbopack X and Carboxen 569) to an air collector pump sampler specially designed
(continued on page 4)

Figure 1. GC chromatogram for stock standard solution. Correspondent VOC reference numbers are listed in Table 1
56 57 50 36,37 51,52

27 20 28,29 11,12,13 2,3,4 1 1 5,6 78 10 9,10 14 18 17 19 16 15 21,22,23 25 24 26 30 31 20 Min 34,35 33 32 4344 42

46 48,49 47

54 55

39,40 38 41

45

53 30 Chromatogram2

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Table 1. Method target VOC analytes, retention time (RT, min) and quantification ions m/z 1 (low concentration range) and m/z 2 (high concentration range)
Target VOCs Ethanol Propanal Acetone Carbon disulfide Methyl acetate Isopropanol Tert-butylmethylether n-Hexane Butanal Ethyl acetate Chloroform Methylethylketone Tetrahydrofuran 1,1,1-Trichloroethane Cyclohexane Carbon tetrachloride Isobutanol Benzene 1-Butanol Trichloroethylene Methylcyclohexane Pentanal Methyl methacrylate Methylisobutylketone Toluene 1,1,2-Trichloroethane Tetrachloroethylene Butyl acetate

VOC Ref. No. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28

RT (min) 7.4 7.5 7.56 7.84 8.14 8.16 8.81 9.11 10.16 10.48 11.02 11.12 11.12 11.42 11.54 11.74 11.9 12.12 13.11 13.13 13.77 13.78 13.83 15.53 15.97 16.83 17.22 17.55

Quantification Ions m/z 1 m/z 2 45 46 58 58 43 44 76 76 74 74 45 59 73 57 57 86 44 72 61 88 83 87 72 57 42 72 97 117 56 84 117 121 43 74 78 51 56 31 130 134 55 98 44 86 100 100 43 100 92 65 166 168 166 168 73 73

Compound with only one characteristic quantification ion

Target VOC VOCs Ref. No. Hexanal 29 N,N-Dimethylformamide 30 N-Methylformamide 31 Ethylbenzene 32 n-Nonane 33 m-Xylene 34 p-Xylene 35 o-Xylene 36 Styrene 37 Heptanal 38 2-Butoxyethanol 39 -Pinene 40 Cyclohexanone 41 Propylbenzene 42 n-Decane 43 1,3,5-Trimethylbenzene 44 -Pinene 45 1,2,4-Trimethylbenzene 46 Benzaldehyde 47 Isocyanatocyclohexane 48 Limonene 49 p-Dichlorobenzene 50 n-Undecane 51 Phenol 52 1-Octanol 53 Naphthalene 54 Isothiocyanatocyclohexane 55 2-Methylnaphthalene 56 1-Methylnaphthalene 57

RT (min) 17.59 18.56 19.06 19.45 19.63 19.73 19.73 20.75 20.75 21.2 21.64 21.65 21.97 22.71 23.04 23.15 23.4 24.16 24.25 24.88 24.88 25.24 26.22 26.32 26.88 31.4 32.9 35.34 35.9

Quantification Ions m/z 1 m/z 2 44 72 73 58 59 30 106 65 57 128 106 77 106 77 91 91 104 104 44 86 57 87 93 136 98 83 91 120 71 142 105 120 93 136 105 120 77 106 82 125 93 136 140 75 57 156 94 66 41 84 128 102 55 141 142 115 142 115

Sample Preparation

(continued from page 3)

in the LCMA-UPC laboratory (Figure 2). The operating flow ranges between 40 and 200 mL/min. Samples can be taken dynamically during 24-hour controls or during odor episodes. By means of remote control (both radio frequency and mobile phone), the air sampler can be activated during odor episodes when medium and high odor intensity and nuisance is percept (5).

Table 2. Instrument Settings and TD-GC/MS Operating Conditions

TD Desorption Temp.: Desorption Time: Transfer Line: Cold Trap Sorbent: 300 C 10 min 200 C TD Capillary Column: DB-624 (60 m x 0.25 mm x 1.4 m) Temperature Program: 40 C (1 min), 6 C min-1 until 230C (5 min) Carrier Gas: He (19.1 psi)

Analytical Instrumentation
The analysis of VOC is performed by Thermal Desorption GC/MS. Mass spectral data are acquired over a mass range of 20-300 amu (Table 2). Qualitative identification of target compounds is based on the match of the retention times and the ion ratios of the target quantification ions and the qualifier ions (Table 1). Quantification of field samples is conducted by the external standard method. Limits of detection (LOD), determined applying a signal-to-noise ratio of 3, range from 0.001 to 10 ng. The studied compounds show repeatabilities (% relative standard deviation values) 25%, accomplishing the EPA performance criteria (6). Extreme precautions are established for quality assurance, injecting periodically blank samples and a known concentration of toluene (4).

Tenax + Carbotrap Cold Trap Low: -30 C Cold Trap High: 300 C Desorption Flow Rate: He (50 mL min-1) Inlet Split: 4 mL min-1 Oultlet Split: 7 mL min-1 Split Ratio: 12%

MS Interface: 250 C Ionization Source: 200 C Ionization mode: Electron impact Electron Energy: 70 eV Mass Range: 20-300 amu

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Reporter 29.3

Figure 2. Collector pump sampler

Indoor Air Applications

Most indoor air pollution originates from sources inside the building, such as adhesives, carpeting, wood products or cleaning products. However, the outdoor air that enters the building through e.g. air intake vents and windows can also be a source of indoor air VOC pollution. Examples are pollutants from motor vehicle exhausts, plumbing vents, building exhausts (kitchen and bathroom exhausts) and combustion products from a nearby garage. It has to be noted that VOC have been generally less studied than other indoor air contaminants, even though their concentrations have been described to contribute to the Sick Building Syndrome (SBS). To exemplify the methodology, a characterisation of the indoor air quality of a dwelling where occupants experienced symptoms that resembled the SBS symptoms was done in May-July 2006 in Barcelona city. It was observed that there was indoor air pollution by several major compounds including ethanol, acetone, acetic acid and 1-metohoxy-2-propanol, which altered the VOC family distributions expected for a standard dwelling. The external source of VOC was found to be a not declared activity of storage and manipulation of solvents located at the bottom of a contiguous building (7).

Sample Preparation

Collector Pump

Outdoor Air Applications

Outdoors, a sampling system can be used in situations involving permanent and discontinuous odor episodes in air, either in urban, industrial, waste treatment facilities or rural areas. A thorough analysis of the meteorological conditions in the area during the sampling period in combination with the chemical information obtained through the individual VOC determination and an inspection of the activities on the site allow an irrefutable tracking of the permanent odor sources and to generate a list of potential odor episodic sources. To exemplify the results obtained from chemical control (VOC concentrations, both 24-hour average and maximum episodic values), interpolated concentration maps are represented in a study area (Figure 3). Those maps are important tools for determining the origin of VOC emissions in a concrete study area, showing focuses with higher concentrations. Figure 3. Concentration maps of three families of compounds and TVOC in Banyoles city (Catalonia, Spain).
Carboxylic Acids
2500 2000 1500 1000 500 0 40 2 30 20 0 1000 2000 0 1000 2000 1 g Mn-3 50 3

Conclusions
The presented methodology can identify in outdoor air the compounds that cause odor episodes as well as their sources. A real inventory of all potential emitting activities can be obtained, including not only nearby emitters but also sources that are several kilometres away. Air quality criteria can be applied to the chemical compound concentrations obtained with the presented methodology. In indoor air, the methodology has been demonstrated to be useful in an urban dwelling. It has been observed that people repetitively exposed long-term to relatively high levels of VOC can modulate their physiological response to a given compound. Legislated guideline recommendations should be established. In the interim period, VOC indoor concentrations should be maintained as low as reasonably achievable (ALARA) (8).

Trimethylbenzenes

g Mn-3

References
1. 2. 3. 4. 5. Shirey, R., 2005. The Reporter Europe 16, 13-14. Buchanan, M., Betz, B., Buckenhdahl, K, 2006. The Reporter Europe 22, 15-16. Brown, J.L., 2007. The Reporter Europe 28, 11-12. Ribes, A., Carrera, G., Gallego, E., Roca, X., Berenguer, M.J., Guardino, X., 2007. J Chromatogr A 1140, 44-55. Gallego, E., Roca, F.J., Perales, J.F., Guardino, X., 2008. In: Romano, G.C., Conti A.G. (Eds.), Air Quality in the XXI Century, Nova Science Publishers, New York. U.S. EPA, 1999. Compendium of Methods for the Determination of Toxic Organic Compounds in Ambient Air, Method TO-17. Gallego, E., Roca, X., Perales, J.F., Guardino, X., 2009. J Environ Sci 21, 333- 339. ECA-IAQ, 1997. Indoor air quality and its impact on man. Total volatile organic compounds in indoor air quality investigations. EUR 17675 EN, Report No. 19, Luxemburg, pp. 48.

Carboxilic acids

TMBs

Organonitrogenated Compounds
2500 2000 1500 1000 500 0

6. 7. 8.

g Mn-3

TVOC's

g Mn-3 140 120 100

20

10

80 60 40

1000

2000

1000

2000

Organonitro...

TOTAL_COVs

Reporter 29.3

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Radiello Passive Samplers and Vapor Intrusion

Kristen Schultz kristen.schultz@sial.com

Table 1. Select VOC Sampling Rates for Passive Sampling Devices

Passive Sampling Rates (mL/min) VOCs by VOCs by Typical Solvent Thermal Axial Desorption Desorption Sampling RAD130 RAD145 Rates 80 27.8 10-16 59 25.4 10-13 62 65 74 69 65 68 25 20.0 24.6 30.0 27.1 30.0 25.7 NA NA 8-12 8-15 11-15 9-14 9-13 NA

Sample Preparation

Introduction
Vapor intrusion assessments necessitate sampling of soil gas and in some cases indoor air. The traditional method for sample collection relies on Summa canisters for vapor collection and storage as required by EPA Compendium Method TO-15. Other approved active sampling methods include EPA TO-13 and TO-17 which utilize sorbent tubes such as thermal desorption tubes as the sample collection medium (1). Other types of devices such as gas sampling bags are used in addition to, or in lieu of, Summa canisters depending on the compounds of interest. Passive sampling devices such as Radiello have been deployed in numerous studies in the US for sampling vapor intrusion compounds because of their low maintenance, ease of use, and low detection limits. The Radiello VOC samplers such as the RAD130 VOCs/BTEX by Solvent Desorption (Figure 1) and RAD145 VOCs/BTEX by Thermal Desorption are presented. Figure 1. Radiello Passive Sampler

Compound Benzene Perchloroethylene (Tetrachloroethylene) 1,1,1-Trichloroethane o-Xylene Toluene Trichloroethylene Methyl tert-butyl ether Ethyl benzene Naphthalene

Compared to SKC Ultra Samplers for Indoor Air, Publication 1720 Revision 1103 and Ultra III Publication 1804 Issue 1104

NA = Not available

Figure 2. Comparison of Radial Design to Axial/Planar Design

Radial design allows air adsorption from all directions Diffusive surface Axial/Planar design limits air adsorption from only one direction

Adsorbing surface
P001114a P001111

P001113a

Radiello Q=80 mL/min

Axial/Planar Q=<10 mL/min

Sampling soil gas with any passive sampling device has its limitations. Because passive sampling relies on air flow, the device may become starved for air at lower face velocities and affect the sampling rate. Radiello diffusive samplers provide reliable results when airflow is 0.1 10 m/s and relative humidity is in the range of 15-90%. Radiello sampling rates (Q) are 2-5 times higher than axial (badge) type samplers (Table 1), due to the radial design of the radiello sampler (Figure 2).

Diffusion rates for benzene at 25 C

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Reporter 29.3

Table 2. Passive Sampling Exposure Time & Limits of Detection

Exposure Time RAD130 VOCs by Solvent Desorption Indoor Outdoor* 15 min-8 hr 7-30 days 15 min-8 hr 7-30 days 15 min-8 hr 15 min-8 hr 15 min-8 hr 15 min-8 hr 15 min-8 hr 15 min-8 hr 15 min-8 hr 7-30 days 7-30 days 7-30 days 7-30 days 7-30 days 7-30 days 7-30 days Limits of Detection (g/m) LOD Typical Range: 0.05-1.0 g/m3 Varies by laboratory, please consult your lab. Exposure Time RAD145 VOCs by Thermal Desorption Indoor Outdoor 8 hr 7-14 days 8 hr 7-14 days 8 hr 8 hr 8 hr 8 hr 8 hr 8 hr NA 7 days 7-14 days 7-14 days 7 days 7 days 7-14 days NA Limits of Detection (g/m) LOD 0.05 0.02 0.10 0.01 0.01 0.02 0.20 0.01 NA

Compound Benzene Perchloroethylene (Tetrachloroethylene) 1,1,1-Trichloroethane o-Xylene Toluene Trichloroethylene Methyl tert-butyl ether Ethyl benzene Naphthalene

Sample Preparation

*7 days is ideal but the device can be sampled for 30 days in low concentration environments. NA = Not available

Exposure Limits
Depending on the concentration of compounds in the sampling environment, radiello devices can be deployed from 8 hours to 7-14 days for thermal desorption and 8 hours to 7-30 days for solvent desorption (Table 2). Radiello has a significantly higher capacity compared to active sampling devices and other passive sampling devices. The Radiello 130 (RAD130) cartridge has a high loading capacity of about 80 mg corresponding to an overall VOC concentration of 3,000-3500 g/m3 sampled for 8 hours or 70,000-80,000 mg/m3 for 14 days. The thermal desorption sampler is more sensitive and applicable for lower concentration VOC environment (<2,000 g/m3) but can sample from 8 hours to 7-14 days. Radiello is suitable for both time-weighted average (TWA) and short term exposure limit (STEL) concentrations.

Featured Products
Qty. 20 20 2 1 1 20 20 2 1 20 20 198 20 Cat. No. RAD130 RAD120 RAD130S RAD405 RAD406 RAD145 RAD1202 RAD145S RAD407 RAD121 RAD195 RAD190 RAD122

Description VOCs/BTEX by Solvent Desorption (SD) Cartridge Adsorbent Diffusive Body White VOC/BTEX Starter Kit (SD) BTEX Calibration Kit (SD) VOC Calibration Kit (IH environment) VOCs/BTEX by Thermal Desorption (TD) Cartridge Adsorbent Diffusive Body Yellow VOC/BTEX Starter Kit (TD) BTEX Calibration Kit (TD) Radiello Accessories Triangular Support Plate Radiello Clips Bar Code Labels Radiello Vertical Adapter Threaded for Standard Use

Summary
Radiello is the ideal passive sampler for indoor vapor intrusion environments because of its high sampling rates, high capacity, and low detection limits. It is versatile for sampling from 15 min for STEL sampling and 8 hrs for TWA sampling to 7-30 days depending on the sampling environment. Like other passive samplers, there are limitations to sample vapor intrusion or soil gases such as in micro-wells and like environments. Radiello is suitable for sampling in crawl spaces and indoor air environments to the surrounding outdoor environment (1). In addition to samplers for VOCs, radiello samplers are available for sampling a wide range of compounds such as: aldehydes, hydrogen sulfide, ammonia, nitrogen and sulfur dioxides, phenols, ozone, hydrochloric acid and anesthetic gases. Radiello analytical testing services are available in several US locations, Italy, Canada, Australia, and New Zealand.

Each starter kit contains 1 triangular support plate, 1 diffusive body, 1 vertical adapter, and 2 cartridge adsorbents.

Did you know . . .

Radiello passive sampling devices are suitable for a wide range of compounds. Download a copy of the Radiello Manual (IYP) from our website - sigma-aldrich.com/radiello - for a complete list of compounds by sampler type.

References
1. Shamory, Brett; Hayes, Heidi; Mahfood, John; Schultz, Kristen, Radiello Passive Sampling Method for Evaluating Vapor Intrusion, Business of the Brownfields Conference 2009, Pittsburgh, PA SKC Technical Note, Measuring Sub-ppb Levels of VOCs in Indoor Air, Publication 1720 Rev 1103. Ultra III: The Creative Evolution of Indoor Air and Ambient Sampling, SKC Publication 1804, Issue 1104

2. 3.

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Comparison of the Supelco 37-Component FAME Mix on the SP-2560 and SLB-IL111
Gas Chromatography
Katherine K. Stenerson, Leonard M. Sidisky, and Michael D. Buchanan mike.buchanan@sial.com

Table 2. Supelco 37-Component FAME Mix Composition

1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22. 23. 24. 25. 26. 27. 28. 29. 30. 31. 32. 33. 34. 35. 36. 37. Butyric Acid Methyl Ester (C4:0) at 4 wt % Caproic Acid Methyl Ester (C6:0) at 4 wt % Caprylic Acid Methyl Ester (C8:0) at 4 wt % Capric Acid Methyl Ester (C10:0) at 4 wt % Undecanoic Acid Methyl Ester (C11:0) at 2 wt % Lauric Acid Methyl Ester (C12:0) at 4 wt % Tridecanoic Acid Methyl Ester (C13:0) at 2 wt % Myristic Acid Methyl Ester (C14:0) at 4 wt % Myristoleic Acid Methyl Ester (C14:1) at 2 wt % Pentadecanoic Acid Methyl Ester (C15:0) at 2 wt % cis-10-Pentadecenoic Acid Methyl Ester (C15:1) at 2 wt % Palmitic Acid Methyl Ester (C16:0) at 6 wt % Palmitoleic Acid Methyl Ester (C16:1) at 2 wt % Heptadecanoic Acid Methyl Ester (C17:0) at 2 wt % cis-10-Heptadecenoic Acid Methyl Ester (C17:1) at 2 wt % Stearic Acid Methyl Ester (C18:0) at 4 wt % Oleic Acid Methyl Ester (C18:1n9c) at 4 wt % Elaidic Acid Methyl Ester (C18:1n9t) at 2 wt % Linoleic Acid Methyl Ester (C18:2n6c) at 2 wt % Linolelaidic Acid Methyl Ester (C18:2n6t) at 2 wt % -Linolenic Acid Methyl Ester (C18:3n6) at 2 wt % -Linolenic Acid Methyl Ester (C18:3n3) at 2 wt % Arachidic Acid Methyl Ester (C20:0) at 4 wt % cis-11-Eicosenoic Acid Methyl Ester (C20:1n9) at 2 wt % cis-11,14-Eicosadienoic Acid Methyl Ester (C20:2) at 2 wt % cis-8,11,14-Eicosatrienoic Acid Methyl Ester (C20:3n6) at 2 wt % cis-11,14,17-Eicosatrienoic Acid Methyl Ester (C20:3n3) at 2 wt % Arachidonic Acid Methyl Ester (C20:4n6) at 2 wt % cis-5,8,11,14,17-Eicosapentaenoic Acid Methyl Ester (C20:5n3) at 2 wt % Heneicosanoic Acid Methyl Ester (C21:0) at 2 wt % Behenic Acid Methyl Ester (C22:0) at 4 wt % Erucic Acid Methyl Ester (C22:1n9) at 2 wt % cis-13,16-Docosadienoic Acid Methyl Ester (C22:2) at 2 wt % cis-4,7,10,13,16,19-Docosahexaenoic Acid Methyl Ester (C22:6n3) at 2 wt % Tricosanoic Acid Methyl Ester (C23:0) at 2 wt % Lignoceric Acid Methyl Ester (C24:0) at 4 wt % Nervonic Acid Methyl Ester (C24:1n9) at 2 wt %

Ionic Liquids

Introduction
For the food chemist, developing a single method to determine the fat composition of different products may be difficult because foods contain a complex mixture of fatty acids, each with a variety of carbon chain lengths.

Milk and butter contain saturated C4 to C20, monounsaturated C16 and C18, and polyunsaturated C18 fatty acids Vegetable oils contain saturated C6 to C24, monounsaturated C16, and monounsaturated cis C18, C20, and C22 fatty acids Margarines contain the same fatty acids as vegetable oils plus monounsaturated trans C18, C20, and C22, and polyunsaturated C18 fatty acids Fish and meat typically contain saturated and monounsaturated C12 to C24+ fatty acids, plus polyunsaturated omega 3 C18, C20, and C22, and polyunsaturated omega 6 C18 and C20 fatty acids Fish tends to be richer in the polyunsaturated omega 3 fatty acids, whereas meats are richer in the polyunsaturated omega 6 fatty acids

Materials and Experimental

Supelco 37-Component FAME Mix. To properly identify fatty acid composition, standards of known reference must be used. One such standard is the Supelco 37-Component FAME Mix (47885-U). This standard contains methyl esters of fatty acids ranging from C4 to C24, including key monounsaturated and polyunsaturated fatty acids, making this standard very useful to food analysts since it can be used to identify fatty acids in many

Saturated, monounsaturated, polyunsaturated, cis, trans, and omega all refer to the structure of fatty acid moieties. Descriptions and their effects on human health are summarized in Table 1.

Table 1. Fatty Acid Types and Health Effects

Type Saturated fatty acids Mono- and polyunsaturated cis fatty acids Mono- and polyunsaturated trans fatty acids Omega 3 fatty acids Description No double bonds >1 cis double bond >1 trans double bond Initial double bond located directly after the third carbon atom as measured from the methyl end Initial double bond located directly after the sixth carbon atom as measured from the methyl end Health Effects Raise LDL cholesterol (increases risk of cardiovascular disease) Lower LDL cholesterol (reduces risk of cardiovascular disease) Raise LDL cholesterol (increases risk of cardiovascular disease) and also lower HDL (increases risk of type II diabetes) Linked with reducing coronary heart disease

Omega 6 fatty acids

Can interfere with the health benefits of omega 3 fatty acids, and also linked with several detrimental health conditions

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Reporter 29.3

different types of foods. The composition of this mixture is shown in Table 2. It is diluted to 10 mg/mL in methylene chloride. SP-2560 Column. The highly polar SP-2560 column was specifically designed for the separation of geometric-positional (cis/trans) isomers of FAMEs, and is extremely effective for Figure 1. 37-Component FAME Mix Chromatograms

special FAME applications including the separation of FAMEs in hydrogenated vegetable oil samples. It is offered in a 100 m x 0.25 mm I.D., 0.20 m dimension (24056), providing the necessary plates for the detailed separation of cis/trans FAME isomers. This well-established column is specified in several methods. (1-2)
(continued on page 10)

Gas Chromatography

SP-2560 Conditions column: SP-2560, 100 m x 0.25 mm I.D., 0.20 m (24056) oven: 140 C (5 min.), 8 C/min. to 180 C, 4 C/min. to 210 C, 20 C/min. to 250 C (7 min.) inj.: 250 C det.: FID, 250 C carrier gas: hydrogen, 40 cm/sec injection: 1 L, 100:1 split liner: 4 mm I.D., split type, cup design sample: Supelco 37-Component FAME Mix (47885-U) SLB-IL111 Conditions column: SLB-IL111, 100 m x 0.25 mm I.D., 0.20 m (29647-U) oven: 140 C (5 min.), 8 C/min. to 180 C, 5 C/min. to 260 C det.: FID, 260 C All other conditions the same as those used for the SP-2560. See Table 2 for Peak IDs

SP-2560 Column
12 23 2 1 3 4 6 8 16 17 24 30 25 5 7 9 10 11 13 14 15 18 20 19 21 22 32 35 26 33 28 29 34 27 37 31 36

10 Min

20

G005366

12

SLB-IL111 Column
3 2 1

16

17

23 31 36 27 28 3235 25 26 33 29 34 37

10 9

11

14 13

15

18 20 19

24 30 21 22

10 Min

20 G005367

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(continued from page 9)

Conclusion
The SLB-IL111 column proved to be a highly efficient column for the analysis of the Supelco 37-Component FAME Mix. Based on the data presented here and the work reported by Delmonte et al. (regarding the resolution of C18:2 9c,11t from C18:2 7t,9c the two most abundant conjugated linoleic acid (CLA) isomers found in ruminant fats), the SLB-IL111 should be considered when selecting a column for FAME analysis.

Gas Chromatography

SLB-IL111 Column. The extremely polar SLB-IL111 column exhibits the highest polarity of any GC phase, providing an alternative selectivity for FAME applications typically performed on highly polar cyanopropyl siloxane columns. It is able to provide resolution of some key isomers that cannot be resolved on other columns. (3-4) Its maximum temperature of 270 C is very impressive for such an extremely polar column. The 100 m x 0.25 mm I.D., 0.20 m version of this column (29647-U) was used to compare directly to the SP-2560. Experimental. The standard was analyzed on both columns, using conditions optimized for each. The resulting chromatograms and operating conditions are shown in Figure 1.

References
1. AOAC Method 996.06, Fat (Total, Saturated, and Unsaturated) in Foods Official Methods of Analysis, 18th Edition (on-line) Association of Official Analytical Chemists, Inc. AOCS Method Ce 1h-05, Determination of cis-, trans-, Saturated, Monounsaturated and Polyunsaturated Fatty Acids in Vegetable or Nonruminant Animal Oils and Fats by Capillary GLC AOCS Official Methods (2005) American Oil Chemists Society. P. Delmonte, A-R.F. Kia, J.K.G. Kramer, M.M. Mossoba, L. Sidisky, and J.I. Rader, Separation Characteristics of Fatty Acid Methyl Esters Using SLB-IL111, A New Ionic Liquid Coated Capillary Gas Chromatographic Column J. Chromatogr. A 1218 (2011) p. 545. L.M. Sidisky and M.D. Buchanan, Detailed Analysis of C18:1 cis/trans FAME Isomers Using the New Supelco SLB-IL111 Reporter 29.1 (2011) p. 3.

2.

Results and Discussion

Baseline and Peak Shape. Both columns produced chromatograms with minimal bleed and sharp peaks. Signal-to-noise ratios for the later eluding peaks are slightly better with the SLB-IL111 column. Run Time. Compared to the SP-2560 phase, the SLB-IL111 phase is not able to undergo as much dispersive interaction with analytes. Therefore, all analytes elute at lower oven temperatures on the SLB-IL111, resulting in a slightly quicker analysis (22 minutes compared to 26 minutes). Selectivity. The SLB-IL111 phase is able to undergo more dipoleinduced dipole interaction with analytes, as compared to the SP-2560 phase. These interactions cause the increased retention of polarizable analytes (those with double bonds) relative to nonpolarizable analytes (those that are saturated). In fact, the higher the degree of unsaturation (the greater the number of double bonds), the stronger the interaction is. This results in alternative selectivity. A comparison of elution orders and elution locations are summarized in Table 3 and Table 4, respectively.
3.

4.

Introductory Offer:

40% off SLB-IL111 Columns

Use Promo Code 962 when placing your order. Offer expires September 30, 2011. Offer not valid in Argentina, Brazil, China, India, and Japan.

Featured Products
Cat. No. 29647-U 24056 47885-U

Description SLB-IL111, 100 m x 0.25 mm I.D., 0.20 m SP-2560, 100 m x 0.25 mm I.D., 0.20 m Supelco 37-Component FAME Mix 10 mg/mL (total wt.) in methylene chloride, 1 mL Analytes and wt% listed in Table 2

Table 3. Comparison of Elution Orders

Peak ID 19 21 22 25 26 27 33 Isomer C18:2n6c C18:3n6 C18:3n3 C20:2 C20:3n6 C20:3n3 C22:2 SP-2560* C18:0 C20:0 C20:0 C21:0 C22:0 C22:0 C23:0 SLB-IL111* C20:0 C21:0 C21:0 C22:0 C23:0 C23:0 C24:0

Related Products
Cat. No. 28925-U 28927-U 23348-U 23362-U 18919-1AMP

Description SLB-IL111, 15 m x 0.10 mm I.D., 0.08 m SLB-IL111, 30 m x 0.25 mm I.D., 0.20 m SP-2560, 75 m x 0.18 mm I.D., 0.14 m SP-2560, 100 m x 0.25 mm I.D., 0.20 m Wound onto a 5 inch cage to fit the Agilent 6850 C4-C24 FAME Mix
Neat mixture of 37 analytes, 100 mg total wt.

* On this column, this isomer elutes after this saturated FAME.

Table 4. Comparison of Elution Locations

Peak ID 28 29 32 34 Isomer C20:4n6 C20:5n3 C22:1n9 C22:6n3 SP-2560 Right after C23:0 Right after C24:0 Between C22:0 and C23:0 ~2.5 min. after C24:0 SLB-IL111 Right before C24:0 >1 min. after C24:0 Right before C23:0 ~3.2 min. after C24:0

Analytes and wt % listed in Table 2

Related Information Additional information regarding the SLB-IL111, or any of our ionic liquid GC columns, can be found on our website: sigma-aldrich.com/il-gc

10

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Reporter 29.3

Edible Oil FAMEs on the SLB-IL111

Katherine K. Stenerson, Leonard M. Sidisky, and Michael D. Buchanan mike.buchanan@sial.com

A typical esterification procedure (using BCl3-methanol) follows:

1. Weigh 1-25 mg of sample into a 5-10 mL micro reaction vessel. 2. Add 2 mL BCl3-methanol, 12% w/w. A water scavenger (such as 2,2-dimethoxypropane) can be added at this point. 3. Heat at 60 C for 5-10 minutes. Derivatization times may vary, depending on the specific compound(s) being derivatized. 4. Cool, then add 1 mL water and 1 mL hexane. 5. Shake the reaction vessel (it is critical to get the esters into the non-polar solvent). 6. After allowing the layers to settle, carefully transfer the upper (organic) layer to a clean vial. Dry the organic layer by either passing through a bed of anhydrous sodium sulfate during the transfer step to the clean vial, or by adding anhydrous sodium sulfate to the clean vial then shaking. 7. Proceed to the appropriate GC analysis procedure.

Introduction
The edible oil industry boasts revenues measured in the tens of billions of dollars. As such, it can be subject to criminal acts of fraud aimed at increasing profits. A fingerprinting technique using gas chromatography (GC) can be used to monitor for adulteration (adding a cheaper, inferior oil to boost the volume of a premium, higher priced oil), and also identify the source of oils in unknown samples. This article will discuss the use of high quality reference standards paired with the SLB-IL111 column to perform this application.

Gas Chromatography

Characterized Reference Oils

To properly identify an edible oil through pattern recognition, it is necessary to have standards of known reference (such as our Characterized Reference Oils). These can be used as part of a quality control program, providing an excellent means of standardizing procedures and comparing results between facilities. Each of these standards includes a Certificate of Analysis (C of A) showing the % composition of each isomer based on peak areas. Prior to analysis, these standards must undergo derivatization.

This procedure is intended as a guideline and may need to be altered to meet the needs of a specific application. Samples can be derivatized neat or after dissolving in solvent. If appropriate, dissolve sample in a non-polar solvent (such as hexane, heptane, or toluene). If the sample is in an aqueous solvent, first evaporate to dryness then use neat or dissolved in an organic, non-polar solvent. It is important to prepare a reagent blank along with the samples to help identify any issues that may arise.

Derivatization Procedure
GC can be used to analyze fatty acids either as free fatty acids or as fatty acid methyl esters (FAMEs). The primary reasons to analyze fatty acids as fatty acid methyl esters include:

Derivatization Tips
Use only high-quality reagents, to ensure that no artifacts are introduced during the processing of samples. Additionally, only derivatization reagents with low moisture should be used, as the esterification reaction will be hindered by the presence of water. The storage conditions of all reagents should be strictly adhered to, as some are susceptible to degradation during long-term storage. (2) Perform derivatization reactions in micro reaction vessels. These heavy-walled borosilicate glass vessels are cone-shaped inside, providing an ideal geometry for performing derivatization reactions. Use screw caps with PTFE-faced septa, to ensure an inert environment. To determine the proper derivatization time, analyze aliquots of a representative sample using different derivatization times. Plot peak area (y-axis) vs. derivatization time (x-axis). The minimum time to use is when no further increase in peak area is observed with increasing derivatization time (where the curve becomes flat). If it is suspected that complete derivatization is never achieved, use additional reagent or re-evaluate temperature.

In their free, underivatized form, fatty acids may be difficult to analyze because these highly polar compounds tend to form hydrogen bonds, leading to adsorption issues. Reducing their polarity may make them more amenable for analysis. To distinguish between the very slight differences exhibited by unsaturated fatty acids, the polar carboxyl functional groups must first be neutralized. This then allows column chemistry to perform separations by degree of unsaturation, position of unsaturation, and even the cis vs. trans configuration of unsaturation.

The esterification of fatty acids to FAMEs is performed using an alkylation derivatization reagent. Methyl esters offer excellent stability, and provide quick and quantitative samples for GC analysis. The esterification reaction involves the condensation of the carboxyl group of an acid and the hydroxyl group of an alcohol. Esterification is best done in the presence of a catalyst (such as boron trichloride). The catalyst protonates an oxygen atom of the carboxyl group, making the acid much more reactive. An alcohol then combines with the protonated acid to yield an ester with the loss of water. The catalyst is removed with the water. The alcohol that is used determines the alkyl chain length of the resulting esters (the use of methanol will result in the formation of methyl esters whereas the use of ethanol will result in ethyl esters). (1)

Analysis Using the SLB-IL111

The extremely polar SLB-IL111 column exhibits the highest polarity of any GC phase, providing an alternative selectivity for FAME applications typically performed on highly polar cyanopropyl siloxane columns. Its maximum temperature of 270 C is very impressive for such an extremely polar column.
(continued on page 12)

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11

Figure 1. Characterized Reference Oils on the SLB-IL111

column: oven: inj.: det.: carrier gas: injection: liner: samples: 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. SLB-IL111, 30 m x 0.25 mm I.D., 0.20 m (28927-U) 180 C 250 C FID, 260 C helium, 25 cm/sec 1 L, 50:1 split 4 mm I.D., split type, cup design Characterized reference oils, methylated using BF3-methanol prior to analysis Lauric acid methyl ester (C12:0) Myristic acid methyl ester (C14:0) Palmitic acid methyl ester (C16:0) Palmitoleic acid methyl ester (C16:1) Stearic acid methyl ester (C18:0) Oleic acid methyl ester (C18:1n9c) Linoleic acid methyl ester (C18:2n6c) Linolenic acid methyl ester (C18:3n3) Arachidic acid methyl ester (C20:0) cis-11-Eicosenoic acid methyl ester (C20:1) 6

(continued from page 11)

Gas Chromatography

Following derivatization, five of our Characterized Reference Oils were analyzed on the SLB-IL111. The resulting chromatograms are shown in Figure 1. A 30 m column was selected because the higher resolution provided by the 100 m column was not necessary for this application. Additionally, the shorter column resulted in shorter analysis times. The % compositions, based on peak areas, agree with those listed on the Certificate of Analysis for each mix.

Conclusion
The SLB-IL111 was able to produce good resolution, peak shape, and signal-to-noise ratios, all with a short analysis time. Food analysts wishing to perform quality control of edible oils should consider employing Characterized Reference Oils and SLB-IL111 columns for this application.

Reference(s)
7 3 5 9 10 8

Canola Oil

1. 2.

W. W. Christie, Gas Chromatography and Lipids The Lipid Library, http:// www.lipidlibrary.co.uk/GC_lipid/gc_lip.html (accessed Jun 26, 2008). W. W. Christie, Why I Dislike Boron Trifluoride-Methanol Lipid Technology (1994) 6, 66-68.

6 3 7 4 2 5 8

G005361

Introductory Offer:

Lard Oil

40% off SLB-IL111 Columns

Use Promo Code 962 when placing your order. Offer expires September 30, 2011. Offer not valid in Argentina, Brazil, China, India, and Japan.

3 6

G005362

Featured Products
Cat. No. 28927-U 46961 47115-U 46962 47122 47123

Description Palm Oil

7

SLB-IL111 Capillary GC Column 30 m x 0.25 mm I.D., 0.20 m Characterized Reference Oils Canola Oil, 1 g Lard Oil, 1 g Palm Oil, 1 g Soybean Oil, 1 g Sunflower Seed Oil, 1 g

1 2

G005363

Soybean Oil
3 5 6 8

Did you know . . .

Fatty Acid/FAME Application Guide
Analysis of Foods for Nutritional Needs

G005364

Sunflower Seed Oil

6 3 5
Free Fatty Acid Analysis FAME Preparation FAME Analysis

4 Min

6 G005365

Fatty Acid/FAME Application Guide: Analysis of Foods for Nutritional Needs (T408126 KUK) includes sections on free fatty acids, derivatization to FAMEs, SPE fractionation, FAMEs by boiling point elution, FAMEs by degree of unsaturation, omega 3 and omega 6 FAMEs, and cis/trans FAME isomers. Request your no-charge copy on the Business Reply Card.

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Reporter 29.3

Related Products
Cat. No. 28925-U 29647-U 46949 47112-U 47113 47559-U 47116 47118 47119 47120-U 33356 33020-U 33040-U 33021 33353 33089-U 33033 Description Micro Reaction Vessels and Caps 5 mL Clear, with Hole Caps, 12 ea 5 mL Clear, with Solid Caps, 12 ea 5 mL Amber, with Hole Caps, 12 ea 10 mL Clear, with Hole Caps, 12 ea Water Scavenger 2,2-Dimethoxypropane, 98%, 25 mL Sodium Sulfate, Anhydrous, >=99.0% Granular, 500 g Granular, 1 Kg Granular, 2.5 Kg Cat. No. 33299 27039 27478-U 27479 D136808-25ML 239313-500G 239313-1KG 239313-2.5KG

Description SLB-IL111 Capillary GC Columns 15 m x 0.10 mm I.D., 0.08 m 100 m x 0.25 mm I.D., 0.20 m Characterized Reference Oils Coconut Oil, 1 g Corn Oil, 1 g Cottonseed Oil, 1 g Linseed (Flaxseed) Oil, 1 g Menhaden Fish Oil, 1 g Olive Oil, 1 g Peanut Oil, 1 g Safflower Oil, 1 g Derivatization Reagents BF3-Methanol, 10% w/w, 20 1 mL BF3-Methanol, 10% w/w, 19 2 mL BF3-Methanol, 10% w/w, 10 5 mL BF3-Methanol, 10% w/w, 400 mL BCl3-Methanol, 12% w/w, 20 x 1 mL BCl3-Methanol, 12% w/w, 20 x 2 mL BCl3-Methanol, 12% w/w, 400 mL

Gas Chromatography

Related Information Additional information regarding the SLB-IL111, or any of our ionic liquid GC columns, can be found on our website: sigma-aldrich.com/il-gc

Supelco GC Web Node The Supelco GC Web Node

Detailed information available on the following types of GC columns

Looking for a particular column?

Capillary GC columns can be located by industry/application, and also by phase polarity.
There are also links to several detailed application topics (such as biofuels and fatty acids/FAMEs). To start your experience, visit sigma-aldrich.com/gc

Ionic liquid columns MS-grade columns Fast GC columns GCxGC columns Chiral columns PLOT columns SCOT columns Guard columns/retention gaps

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Quality Syringes for Gas Sampling

Sigma-Aldrich inventories a broad selection of quality Hamilton, SGE and VICI Precision Sampling gastight syringes for air sampling applications and for use withdrawing samples from transportable gas cylinders. Syringe volumes range in size from 10 L to 500 mL.

Accessories

Below is a sampling of our many gastight syringe products. To view the complete line, visit sigma-aldrich.com/syringes.

Hamilton Gastight SampleLock Syringes

P000894_KO

Autoclavable SGE Gas Tight Syringes

SGE Gas Tight syringes are constructed of borosilicate glass, PTFE plunger tips, and KEL-F Luer cones where applicable. An interference fit between the plunger tip and the barrel enhances the gas sealing properties of the syringe. Plungers are interchangeable. Syringes can be autoclaved, but the plunger should be removed. Needles and valves are sold separately.

The Hamilton 1000 and 1700 syringe series are available in a SampleLock style for use with gas samples. The twist valve feature locks the sample in the syringe and allows you to collect, transport and test your sample. The positive rear plunger stock prevents sample loss and plunger blowout at high pressures on 250 mL to 10 mL syringes. Male and female luer lock adapters are available to thread onto the SampleLock valve, making the syringe compatible with many connectors and fittings. All adapters have a Kel-F (CTFE) fluid path, with a port diameter of 0.039.
Description 1705SL 1710SL 1725SL 1750SL 1001SL 1002SL 1005SL 1010SL Replacement Needles Description 1705SL-1725SL 1750SL-1010SL Size 22 ga, bevel tip 22s ga, bevel tip Needle Length (mm) 51 (2 in.) 51 (2 in.) Qty. 6 6 Cat. No. 19136-U 19137-U Volume 50 L 100 L 250 L 500 L 1 mL 2.5 mL 5 mL 10 mL Qty. 1 1 1 1 1 1 1 1 Cat. No. 26280-U 26281 26282 26283 26284 26285 26286 26287

Maximum inertness for liquid or gas samples. Suitable for manual dispenser/diluter applications. High accuracy of dispensed volume. Wide variety of terminations.
Description 10R-GT 25R-GT 50R-GT 100R-GT 250R-GT 500R-GT 1MR-GT 2.5MDR-GT 5MDR-GT 10MDR-GT Volume 10 L 25 L 50 L 100 L 250 L 500 L 1 mL 2.5 mL 5 mL 10 mL Qty. 1 1 1 1 1 1 1 1 1 1 Cat. No. 26237 26262 26263 26264 26265 26266 509388 509396 509418 509426

SGE Syringe Valves and Needles 509345

P000431_KO

509337 Description Push-Pull Valve with needle Push-Pull Valve with Luer Lock Push Button Valve with Luer Lock Replacement Needles Description For 10 L For 25-500 L For 1-2.5 mL For 5-10 mL Size 26 ga, bevel tip 25 ga, bevel tip 23 ga, bevel tip 23 ga, bevel tip

509353 Volume 25 L - 2.5 mL 25 L - 2.5 mL 5-10 mL Qty. 1 1 1 Cat. No. 509337 509345 509353

Needle Length (mm) 50 (1.97 in.) 50 (1.97 in.) 50 (1.97 in.) 50 (1.97 in.)

Qty. 5 5 5 --

Cat. No. 24437 24447 29864-U Inquire

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Reporter 29.3

VICI Precision Sampling Pressure-Lok Syringes Pressure Lok Syringes u g

Series
P000433

Volume 25 L 50 L 100 L 250 L 500 L 1 mL 2 mL 5 mL 10 mL

Qty. 1 1 1 1 1 1 1 1 1

Cat. No. 22268 22269-U 22270-U 22271 22272 22273 22274 22275 22276

Precision Sampling Series A-2 syringes are an economically priced general purpose syringe for use in air monitoring applications. The syringes are leak tight to 250 psi and are suitable for both gas and liquids. These syringes include a push-button valve that allows instantaneous injection and has all the standard Pressure-Lok features such as a PTFE plunger tip, PTFE-sealed needle, and ultrasmooth bore sample. The smaller volume syringes are excellent for small volatile samples. The positive rear stop prevents plunger blowout at elevated pressures.

A-2 A-2 A-2 A-2 A-2 A-2 A-2 A-2 A-2 Replacement Needles Volume 25-100 L 250 L - 10 mL Point Style bevel tip bevel tip

Standards

Needle L x OD x ID (mm) 51 x 0.29 x .005 51 x 0.29 x .012

Qty. 3 3

Cat. No. 22298-U 22299

Prescreened, In-Stock Chemicals

Pesticides, PAHs, PCBs, VOCs, and More
What do these compounds have in common?
They are examples of the thousands of prescreened, in-stock chemicals available through the Sigma-Aldrich custom standards group. We can formulate, test, and package custom standard solutions to meet your needs for all your chromatographic applications. Our custom standard chemists will gladly discuss stability and solubility concerns with you, and make suggestions where needed to improve the quality of your purchase.

You can rely on Sigma-Aldrich custom standard solutions to include:

raw materials and solvents screened for identity and purity your choice of gravimetric, qualitative, and quantitative testing packaging choices from ampuls to bottles manufacturing processes following the guidelines of ISO 9001/2000 proper handling of light- and/or oxygen-sensitive chemicals documentation and Material Safety Data Sheets free technical support strict adherence to all shipping regulations

If you are interested in a customized standard, please email us at customstandards@sial.com, or complete the on-line custom standards quote request form, available 24 hours a day 7 days a week, at our website sigma-aldrich.com/standards

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Standards

TraceCERT Organic and Inorganic CRMs

CRMs for: AAS, ICP, IC, qNMR and Chromatography

Production in accordance with ISO/IEC 17025 and ISO Guide 34 Superior level of accuracy, calculated uncertainties and lot-specific values Traceability to internationally accepted reference materials (i.e. NIST SRM)

For more information and product listings, please visit our website sigma-aldrich.com/inorganiccrm or sigma-aldrich.com/organiccrm. To obtain a copy of our TraceCERT Inorganic CRMs brochure, click the Request Literature link at sigma-aldrich.com/standards, or request a copy on the attached reply card.

Vial Accessories
Glass Magnet Products
Our glass magnet sheet prevents your glass apparatus from being knocked over accidentally. This 2 x 2 sheet (61 cm x 61 cm), with its tacky surface, effectively secures the vessels in place. It can be cut with a razor blade or utility knife to fit strategically on counter tops or in drawers containing fragile glassware. Add it to carts when transporting glass to another lab or into the field.

Space Saver Stack Racks

Economical and convenient storage of samples in your screw cap and crimp cap vials. Can be stacked when empty or full. Autoclavable, 120 C. Each case contains 5 racks.
P000396 P000392

Vial Diam (mm) 6-8 11-12 15-16 17 22 29

No. Vials Per Rack 96 50 48 90 36 50

Rack Dimensions L x W x H (mm) 183 x 125 x 23 192 x 100 x 23 268 x 95 x 30 320 x 170 x 30 320 x 90 x 30 336 x 175 x 30

Cat. No. 23204-U 23207 23205-U 23202 23201 23206

The glass magnet vial holder keeps individual vials, ampuls, and other vessels in place.
Description Glass magnet sheet, 2 x 2 (61 x 61 cm) Glass magnet vial holder, 4 (10 cm) dia., pk. of 2 Cat. No. 57269 57270

Will accommodate 40 mL EPA vial (Cat. No. 23189)

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Autosampler Vial Convenience Packs

Supelco now offers our best selling line of 2 mL autosampler vials in convenience packs. Each convenience pack contains 100 vials of Type 1 borosilicate glass and a bag of 100 pre-assembled caps and septa supplied on a clear lid tray. Purchasing vials in convenience packs provides several benefits for the laboratory. The clear lid tray keeps the products visible and free of particulate contamination. Analysts are assured the components are compatible with each other and there are an equal number of vials and caps. Ordering replacement vials, caps, and septa requires the use of one part number, not three. If you have additional questions, or require help in choosing the correct product, please contact Supelco Technical Service at techservice@sial.com or visit us at sigma-aldrich. com/vials
P001181_KO

Accessories

Featured Products
Cat. No. 29056-U 29057-U 29058-U 29064-U 29065-U 29066-U 29061-U 29062-U 29063-U 29067-U 29068-U Description Cat. No. Screw Top Glass Vials with Large Opening (6.0 mm), pk of 100 Clear glass with PTFE/red rubber septa 29116-U Clear glass with PTFE/silicone septa 29118-U Amber glass with PTFE/red rubber septa 29117-U Amber glass with PTFE/silicone septa 29119-U Crimp Top Glass Vials with Large Opening (6.0 mm), pk of 100 Clear glass with PTFE/red rubber septa 29124-U Clear glass with PTFE/silicone septa 29125-U Clear glass with PTFE/silicone/PTFE septa 29126-U Amber glass with PTFE/red rubber septa 29127-U Amber glass with PTFE/silicone septa 29128-U Amber glass with PTFE/silicone/PTFE septa 29129-U Snap Seal Glass Vials with Large Opening (6.0 mm), pk of 100 Clear glass with PTFE/red rubber septa 29141-U Clear glass with PTFE/silicone septa 29142-U Clear glass with PTFE/silicone/PTFE septa 29143-U Amber glass with PTFE/red rubber septa 29144-U Amber glass with PTFE/silicone septa 29145-U Amber glass with PTFE/silicone/PTFE septa 29146-U

Description Screw Top Glass Vials with 9 mm Thread, pk of 100 Clear glass with PTFE/red rubber septa Clear glass with PTFE/silicone septa Clear glass with PTFE/silicone/PTFE septa Clear glass with graduation spot, PTFE/red rubber septa Clear glass with graduation spot, PTFE/silicone septa Clear glass with graduation spot, PTFE/silicone/PTFE septa Amber glass with PTFE/red rubber septa Amber glass with PTFE/silicone septa Amber glass with PTFE/silicone/PTFE septa Amber glass with graduation spot, PTFE/red rubber septa Amber glass with graduation spot, PTFE/silicone septa

Screw Top Glass Vials with Standard Opening (4.6 mm), pk of 100 Clear glass with PTFE/silicone septa 29104-U Clear glass with graduation spot, PTFE/silicone/PTFE septa 29107-U Amber glass with PTFE/silicone septa 29106-U Amber glass with graduation spot, PTFE/silicone/PTFE septa 29108-U

Primary Standards for the Analysis of Herbal Medicinal Products

For use in quality control, in-process control and stability testing of herbal medicinal products Content assignment by quantitative NMR

For more information and a list of products, please visit our website at sigma-aldrich.com/phytopharma

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Accessing the Benefits of Polar Organic Mobile Phases on Cellulose-Based CSPs for Chiral HPLC
Chiral Chromatography
J.T. Lee jt.lee@sial.com

Column Screening in NP, POM, and PIM Systems

Successful separation of several test compounds on Astec Cellulose DMP using a simple, LC-MS compatible mobile phase of 0.1% ammonium formate in methanol is shown in Figure 1. We then compared the selectivity of this column to Astec CHIROBIOTIC V2 (vancomycin-based) using the same LC-MS mobile phases, as well as normal phase on the cellulose column. Their performance was found to be complementary. Table 1 summarizes the screening results of twenty-six basic racemic drugs. The best results for each compound are highlighted in red. In just three screening runs (one NP on Astec Cellulose DMP and one POM/PIM on both columns), twenty-two baseline separations were achieved, mostly in POM/PIM operation. The example of mianserin on the two columns and three mobile phases is shown in Figure 2. Table 1. Screen Results on 26 Basic Molecules Using Astec Cellulose DMP and CHIROBIOTIC V2 Columns
Conditions same as Figure 1 except: column: Astec Cellulose DMP, 15 cm x 4.6 mm I.D., 5 m particles (51098AST) or Astec CHIROBIOTIC V2, 25 cm x 4.6 mm I.D., 5 m particles (15023AST) mobile phases: normal phase (NP): 10:90:0.1, IPA:Heptane:DEA polar organic mode (POM): 0.1% w/v ammonium formate in methanol polar ionic mode (PIM): 0.1% w/v ammonium formate in methanol flow rate: 0.5 mL/min. temp.: 25 C det.: UV at 230 nm

Although the majority of chiral HPLC separations are done in normal phase mode (including SFC), mobile phases comprising polar solvents have advantages. They may offer different selectivity because of a conformational change of the CSP, better UV sensitivity, better solubility of the analyte, or LC-MS compatibility. The latter is a distinct advantage for bioanalytical applications. You can also use these mobile phases on instruments plumbed for reversed-phase, much more widely used than normal phase. Two mobile phase systems used in chiral HPLC are the POM (polar organic mode) and PIM (polar ionic mode). (In PIM, salts are added to facilitate ionization. In POM, if salts are used it is to suppress ionization.)

Column: Mode: Compound Atropine Bupivacaine Citalopram Clenbuterol Diperodon Disopyramide Esmolol Fluoxetine Homatropine Hydroxyzine Indapamide Ketamine Ketoconazole Mefloglumide Methocarbamol Methoxypheamine Metoprolol Mianserin Ofloxacin Ondasetron Promethazine Propranolol Ritalin Thalidomide Tolperison Trger's base

Mobile Phase Limitations of Cellulose-Based CSPs

Historically, the benefits of POM and PIM have been out of reach for the polysaccharide-based CSPs (cellulose, amylose) that dominate in chiral HPLC since they are often restricted to normal phase or SFC operation. In this short article, we will demonstrate high resolving power of the new Astec Cellulose DMP phase in polar organic mobile phases toward a set of basic pharmaceutical compounds. This unique column can operate in both normal phase and polar organic phase without detrimental effects, even with repeated cycles on the same column (1). We will also show its complementary character to the popular macrocyclic glycopeptide CSPs (Astec CHIROBIOTIC).

Astec Cellulose DMP normal phase (NP) k1/selectivity 0.06/1.33 0.86/1.00 2.75/1.14 1.34/1.00 No elution 1.65/1.07 3.36/1.57 1.09/1.08 2.40/1.62 1.16/1.23 No elution 0.80/1.14 No elution 1.59/1.19 No elution 0.86/1.21 1.25/2.66 0.79/1.23 No elution No elution 0.58/1.05 2.36/2.22 0.66/1.09 No elution 0.41/1.00 0.78/1.22

Astec Cellulose DMP polar organic mode (POM) k1/selectivity 0.18/1.00 0.23/1.00 0.26/1.00 0.03/1.00 0.73/3.89 0.11/1.02 0.09/1.25 0.07/1.02 0.08/2.04 0.40/1.10 0.37/2.27 0.48/1.00 4.31/1.06 0.07/1.00 0.30/1/35 0.07/1.00 0.08/1.38 0.96/1.26 1.91/1.13 1.62/1.07 0.47/1.00 0.16/1.24 0.16/1.00 1.20/1.00 0.27/1.00 1.33/1.28

Astec CHIROBIOTIC V2 polar ionic mode (PIM) k1/selectivity 3.54/1.00 0.31/1.34 2.37/1.12 1.02/1.22 0.66/1.00 1.08/1.14 1.34/1.12 2.00/1.24 0.13/1.00 0.71/1.00 0.26/1.00 0.27/1.00 0.31/1.00 2.86/1.36 1.08/1.00 1.52/1.16 1.22/1.12 0.65/1.98 No Elution 1.02/1.00 1.76/1.68 1.60/1.16 1.32/1.45 0.47/2.97 1.14/1.24 0.18/1.00

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Reporter 29.3

Figure 1. Separation of Basic Drugs Using Polar Organic Mobile Phase on Astec Cellulose DMP
column: Astec Cellulose DMP, 15 cm x 4.6 mm I.D., 5 m particles (51098AST) mobile phase: 0.1% w/v ammonium formate in methanol flow rate: 0.5 mL/min. temp.: 25 C det.: UV at 230 nm injection: 5 L sample: 1 mg/mL in methanol

Figure 2. Example of Complementary Selectivity of Astec Cellulose DMP and Astec CHIROBIOTIC V2 in Various Mobile Phases (Mianserin Enantiomers)
Conditions (same as Figure 1 except): columns: Astec Cellulose DMP, 15 cm x 4.6 mm I.D., 5 m particles (51098AST) or Astec CHIROBIOTIC V2, 25 cm x 4.6 mm I.D., 5 m particles (15023AST) mobile phases: normal phase, NP (a): 10:90:0.1, IPA:Heptane:DEA polar organic mode, POM (b): 0.1% w/v ammonium formate in methanol polar ionic mode, PIM (c): 0.1% w/v ammonium formate in methanol flow rate: 0.5 mL/min. (a,b); 0.8 mL/min. (c) sample: mianserin

Chiral Chromatography

Diperodon

G005183

2a. NP on Astec Cellulose DMP, 10:90:0.1, IPA:Heptane:DEA R()

G004327
*

8 Min

10

12

14
G005355

S(+)

Indapamide

G005217

4 Min

G005358

10

2b. POM on Astec Cellulose DMP, 0.1% w/v ammonium formate in methanol R()
0 2 4 Min 6 8
G005356

Trgers Base

S(+)
G005180

4 Min

10
G005359

4 Min

10
G005357

2c. PIM on Astec CHIROBIOTIC V2, 0.1% w/v ammonium formate in methanol R()

In conclusion, LC-MS compatible polar organic mobile phases (PIM/POM) can be applied to macrocyclic glycopeptide-based Astec CHIROBIOTIC CSPs and polysaccharide-based Astec Cellulose DMP columns for complementary results. The Astec Cellulose DMP is rugged enough to permit operation in both types of mobile phases. These findings demonstrate that polysaccharide-based CSPs can now extend their application areas to pharmacological/clinical trial studies where biological samples are routinely analyzed. To learn more, request our brochures on Astec Cellulose DMP and Astec CHIROBIOTIC on the attached reply card. For complete information on all of our chiral HPLC and GC columns, visit our website sigma-aldrich.com/chiral.

S(+)

4 Min

10
G005360

Featured Products
Cat. No. 51098AST 15023AST

Description Astec Cellulose DMP, 15 cm x 4.6 mm I.D., 5 m particles Astec CHIROBIOTIC V2, 25 cm x 4.6 mm I.D., 5 m particles

Reference
1. Lee, J. T.; Campbell, W., HPLC Enantiomeric Separations of Pharmaceuticals Using Polar Organic Mobile Phases. Presented at The Pittsburgh Conference, Atlanta, GA, March 17, 2011.

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Ion Exchange Chromatography for the Characterization of Biotherapeutics

Liquid Chromatography

Contributed Article
The following was generated with the assistance of an outside source using Sigma-Aldrich products. Technical content was generated and provided by: Regina Rmling Tosoh Bioscience GmbH
tracy.ascah@sial.com

Figure 1. Separation of Charge Variants of mAbs on a TSKgel CM-STAT Column

column: TSKgel CM-STAT, 10 cm x 4.6 mm I.D., 7 m particles (821966) mobile phase: A: 20 mM MES buffer, pH 6.0 B: 0.5 M NaCl in buffer A, pH 6.0 gradient: 10% B (0 min.), 30% B (15 min.), 100% B (15 min.), 0% B (17 min.), 10% B (17 min.), 10% B (21 min.) flow rate: 1 mL/min. temp.: ambient det.: UV at 280 nm injection: 20 L sample: monoclonal antibodies (mAb A through E) 40

Development and production of biopharmaceuticals is a growing segment of the pharmaceutical industry. Therapeutic proteins include blood factors, thrombolytic agents, growth factors, interferons, interleukins, therapeutic enzymes, and last but not least, monoclonal antibodies. A thorough characterization of therapeutic biomolecules is a key task for the successful submission of research and production data for regulatory approvals of new drugs. Due to its high degree of automation, robustness, and reproducibility, liquid chromatography is the workhorse of the pharmaceutical laboratory. Therefore, HPLC is also frequently used when establishing analysis methods for biotherapeutics. The typical chromatographic modes for separation of proteins in native form, such as size exclusion chromatography (SEC) and ion exchange chromatography (IEC), are routinely used for the characterization of biotherapeutics. Detection is usually performed by UV, fluorescence, or light scattering. This article features the use of IEC for the characterization of biopharmaceuticals, specifically the analysis of charge variants of monoclonal antibodies and PEGylated proteins. Besides isoelectric focusing, ion exchange chromatography is the method of choice to analyze charge heterogeneity of proteins. Charge isoforms of proteins result from deamidation of asparagine or glutamine residues or from incomplete removal of C-terminal lysine residues (1). TSKgel STAT ion exchange columns from Tosoh Bioscience can help to increase throughput in quality control (QC) of biopharmaceuticals. TSKgel STAT columns are non-porous polymer columns with a high surface density of functional groups: quaternary ammonium for anion exchange (TSKgel Q- and DNA-STAT), carboxymethyl (TSKgel CM-STAT) and sulphopropyl (TSKgel SP-STAT) for cation exchange. Particle sizes and dimensions of these IEC columns are optimized either for highest throughput or for highest efficiency. Applications for the TSKgel STAT cation exchange columns include the separation of peptides and proteins, PEGylated proteins and charge isomers of monoclonal antibodies. A TSKgel CM-STAT weak cation exchange (WCX) column was applied to separate charge variants of several monoclonal antibodies (Figure 1). The typical analysis time on conventional 25 cm long WCX columns of about eighty minutes could be significantly reduced when separation was performed on a 10 cm TSKgel CM-STAT column, filled with 7 m particles. The analysis profiles for five antibodies show that high resolution analysis can be obtained in about 20 minutes analysis time.

30

mV

20

E D C B

10

0 0 10

A
20 Min 30 40
G005353

PEGylation, the process by which polyethylene glycol (PEG) chains are attached to protein and peptide drugs, is a common practice in the development of biopharmaceuticals to prolong serum half-life and improve pharmacokinetics of a drug (2). Lysine residues of the protein typically serve as PEGylation sites. There is increasing demand for chromatographic methods to separate the modified isoforms from the native protein. The degree of PEGylation can be monitored by analyzing the increase in molecular weight by size exclusion chromatography, as every PEG group remarkably increases the apparent size of the protein. As every PEG residue also shields parts of the protein and, therefore, alters the overall surface charge, ion exchange separation can be used as well to analyze PEGylated proteins. SEC separates the native protein, mono- and di-PEGylated forms but because of the specific mode of interaction, free separation, it requires a certain analysis time and is, therefore, not well suited for high throughput applications. IEC, however, can be accelerated by using shorter columns and the latest particle technology. In addition IEC has the ability to resolve isoforms differing in the position of PEGylated lysine residues, as the surface charge of the protein varies depending on the position of the modification (3).

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Reporter 29.3

Figure 2. Monitoring PEGylation Reaction Using TSKgel SP-STAT Column

column: TSKgel SP-STAT, 3.5 cm x 3 mm I.D., 10 m particles (821963) mobile phase: A: 20 mM sodium acetate buffer, pH 5.0 B: 1 M NaCl in mobile phase A, pH 5.0 gradient: 0% B (0 min.), 100% B (2 min.) flow rate: 2 mL/min. det.: UV at 280 nm sample: PEGylated -lactoglobulin 50

Liquid Chromatography

Native

100

UV @280nm (1Abs/1000mV)

40

80

Native

30

Peak %

60

20

Mono-PEG Di-PEG Tri-PEG

40

Mono-PEG
20

Di-PEG Tri-PEG
0 10 20 30 40 Reaction Time (Min) 50 60 70

10 0 0 1 Min 2
G005354

Figure 2 shows the monitoring of a PEGylation reaction of -lactoglobulin in intervals of 5 minutes on a high throughput TSKgel SP-STAT, 10 m, strong cation exchange (SCX) column. A fast gradient of sodium chloride in sodium acetate buffer at a high flow rate of 2 mL/min. was applied to achieve fast separation. An efficient reaction monitoring has to follow the progress of the reaction in short intervals. Therefore, a high throughput column was applied here. For a more detailed characterization of the final product, a better separation of the different Mono-PEG isomers can be achieved by using a high resolution SCX column (10 cm length, smaller particles) and a more shallow gradient profile (data not shown).

TSKgel STAT ion exchange non-porous resin columns contain large particles (5, 7 and 10 m). This relatively large particle size, combined with an innovative bonding chemistry, results in columns that enable fast equilibration and analysis of complex biomolecular samples, as demonstrated in this article.

References
1. 2. 3. Ahrer, K.; Jungbauer, A. J. Chromatogr. B, 2006, 841, 110-122. Harris, J. M.; Chess, R. B. Nature Reviews, 2003, 2, 214-221. Yamamoto, S. et al. J. Biotechnol., 2007, 132, 196-201.

Featured Products
Functional Group quaternary ammonium quaternary ammonium quaternary ammonium sulfopropyl sulfopropyl carboxymethyl carboxymethyl Counter Ion Cl Cl Cl Na Na Na Na pH Range 3.0 - 10.0 3.0 - 10.0 3.0 - 10.0 3.0 - 10.0 3.0 - 10.0 3.0 - 10.0 3.0 - 10.0 pKa 10.5 10.5 10.5 2.6 2.6 4.9 4.9 Cat. No. 821960 821961 821962 821963 821964 821965 821966

Description* TSKgel Q-STAT, 3.5 cm x 3 mm I.D., 10 m particles TSKgel Q-STAT, 10 cm x 4.6 mm I.D., 7 m particles TSKgel DNA-STAT, 10 cm x 4.6 mm I.D., 5 m particles TSKgel SP-STAT, 3.5 cm x 3 mm I.D., 10 m particles TSKgel SP-STAT, 10 cm x 4.6 mm I.D., 7 m particles TSKgel CM-STAT, 3.5 cm x 3 mm I.D., 10 m particles TSKgel CM-STAT, 10 cm x 4.6 mm I.D., 7 m particles

*matrix, hydrophilic polymer; porosity, non-porous; hardware, stainless steel tubing and fittings, PEEK frits

Supelco is pleased to carry the full line of TSKgel columns from Tosoh Bioscience. Visit sigma-aldrich.com/tsk for the complete listing of TSKgel columns.

TRADEMARKS: Agilent Agilent Technologies; Ascentis, Carbopack, Carbotrap, Carboxen, CHIROBIOTIC, CHROMASOLV, Fluka, SLB, SP, Supelco, TraceCERT Sigma-Aldrich Biotechnology LP; Fused-Core Advanced Materials Technology, Inc.; Gastight, Hamilton, SampleLock Hamilton Co.; Pressure-Lok Valco; Kel-F 3M Corp; Radiello Fondazione Salvatore Maugeri; STAT, TSKgel Tosoh Bioscience

Reporter 29.3

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Novel Performance-Tested Solvents for UHPLC Applications

Reagents

New LC-MS Ultra CHROMASOLV Grade Solvents and Additives

Rudolf Khling rudolf.koehling@sial.com

Introduction
Recent innovations in HPLC and mass spectrometry (MS), including Fused-Core particles and ultra high performance/ pressure liquid chromatography (UHPLC) systems, have pushed the limits of speed (throughput), efficiency, and sensitivity. They have greatly increased the amount and quality of data obtained from HPLC and LC/MS experiments. While the technical buzz is usually around columns and instruments, the solvents used for mobile phases, sample preparation, and sample dissolution are also critical components of the system. Their influence on reducing background noise and baseline instability, extending column lifetime, and maintaining system integrity cannot be underestimated nor overlooked. These solvent-related issues as they apply to UHPLC are discussed in this article; the same arguments apply to traditional HPLC as well.

Figure 1. UV baseline UV drift of the solvent baseline resulting from a gradient application of LC-MS Ultra CHROMASOLV Acetonitrile/water (red) compared with LC-MS grade Acetonitrile/ water (blue)

10.0 7.5 5.0 2.5 Intensity (mAu) 0.0 -2.5 8 6 4 2

The Problem of Solvent-Derived Impurities

Background noise is typically caused by impurities that enter the system from the sample, by leaching from system components (bottles, tubes, valves, vials, etc.), and from solvents and additives. To reduce the latter contribution, Sigma-Aldrich developed and recently introduced Fluka-brand LC-MS Ultra CHROMASOLV grade solvents and additives. This new product line is tested for UHPLC performance, including gradient applications, under both UV and MS detection. Experimental problems resulting from these can be significantly reduced, if not eliminated entirely, by using the new LC-MS Ultra CHROMASOLV grade solvents. The solvents are tested for performance in UHPLC gradient separations, as well as UV, and positive and negative ion mode MS detection. The tests and the corresponding solvent performance are discussed below.

0 -2 0 1 2 Min 3 4 5 CHR_1

Blank runs of standard reversed-phase gradients of 30% to 80% acetonitrile in water generally result in low and stable baselines with HPLC columns packed with particles >3 m. However, this is not the case in UHPLC separations because of the typically higher flow rates and short columns. Figures 2 and 3 (positive and negative ion mode ESI, respectively) show that UHPLC gradients using standard solvents have many more impurity peaks and rising baselines compared to LC-MS Ultra CHROMASOLV solvents.

Measurement of Solvents Performance Under Gradient Conditions

The UHPLC/UV/MS testing is performed with state-of-the art LC and MS instruments. Columns are Ascentis Express Fused-Core C18 particles (2.7 m) and totally porous sub-2 m particle C18. Using a gradient of acetonitrile in water, the new LC-MS Ultra CHROMASOLV grade solvents have extremely low total UV baseline drift compared to classical LC-MS grade and gradient grade acetonitrile. Drift is below 8 mAU at 210 nm for acetonitrile (Figure 1). Compared to standard LC-MS grade solvent, this new grade has a significantly lower baseline and fewer minor peaks.

Conclusion
The advent of UHPLC and other high-efficiency HPLC and LC-MS techniques have set new standards for sensitivity, efficiency, and throughput. In order for the UHPLC system to provide this reliable data and high performance and eliminate system downtime, it is critical to use solvents that are as carefully developed, prepared, and tested as other components of the system. The solvents must be tested in situ under demanding UHPLC conditions, under various detection modes. Flukas new LC-MS Ultra CHROMASOLV grade solvents fulfill these requirements.

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Reporter 29.3

Figure 2. Baseline in Positive Ion Mode MS Detection ESI(+) LC-MS chromatograms of a standard blank gradient using LCMS Ultra CHROMASOLV Acetonitrile/water (red) compared to standard HPLC solvents (blue)

Figure 3. Baseline in Negative Ion Mode MS Detection Baseline of LC-MS Ultra CHROMASOLV Acetonitrile/water gradient in negative ion ESI mode (red). The upper blue trace is from standard HPLC grade solvent under the same conditions as Figure 2
1200

Reagents

800 600 400 Intensity (mAu) 200 0 800 600 400 200 0 0

Low UV Drift

1000 800 600 Intensity (mAu) 3 4 Min 5 6 7 8 CHR_2 400 200 1200

High UV Drift

1000 800 600 400 200

4 Min

8 CHR_3

Featured Products
Description LC-MS Ultra CHROMASOLV, ** LC-MS Ultra eluent additive, 99.0% LC-MS Ultra eluent additive, 98% + LC-MS Ultra eluent additive, + LC-MS Ultra eluent additive, + Package Size 1 L, 2 L 1 mL, 2 mL 1 mL, 2 mL 25 g 25 g Cat. No. 14263 14264 14265 14266 14267

Name Water Trifluoroacetic acid Formic acid Ammonium formate Ammonium acetate
** gradient tested for UHPLC + suitable for UHPLC-MS

Seminars about Innovative Technologies for HPLC, GC, & Sample Prep
Supelco and Fluka Sigma-Aldrichs Analytical & Chromatography brands invite you to our complimentary half day, open seminars in your region. Delivered by our technical experts and special guest speakers, the seminars show how our practical, innovative products and technologies can help you today.
Date July 19 July 20 July 21 July 25 July 27 Location Rochester, NY Syracuse, NY Albany, NY Vancouver, BC Calgary, AB Date July 28 August 16 August 18 September 13 September 14 Location Edmonton, AB Denver, CO Phoenix, AZ Rahway, NJ Princeton, NJ Date September 15 October 18 October 19 October 20 November 29 Location Collegeville, PA Boston, MA Wallingford, CT Long Island, NY Birmingham, AL Date November 30 December 1 December 13 December 14 December 15 Location Memphis, TN Little Rock, AR Knoxville, TN Greenville, SC Atlanta, GA

For registration and details, visit sigma-aldrich.com/analytical-seminars, e-mail ac_seminars@sial.com, or call 814-359-5910.

Reporter 29.3

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