Achieving the Lowest Limits of Identification – GC-MS with Cold EI versus Standard EI with High Efficiency Source

Aviv Amirav, Tel Aviv University and Aviv Analytical Ltd.

Executive Summary  

GC-MS sensitivities are specified with octafluoronaphthalene (OFN). However, for many GC-MS users the most important operational parameter is the sample limits of identification. We compared the Aviv Analytical GC-MS with Cold EI with the Agilent 5977B GC-MS with high efficiency ion source (HES) in sample identification. We found that Cold EI far outperforms the 5977B-HES in both detection and identification limits. Cold EI detected and identified thirteen impurity compounds in a given test mixture while the HES standard EI failed to detect most of these impurities and failed to identify any of them. In this article we demonstrate and discuss several Cold EI benefits of superior sensitivity, better identification capability, greater range of compounds amenable for analysis and faster speed of analysis. The graphical abstract figure above demonstrates the absence of impurity peaks in the TIC with HES versus at least thirteen detected peaks in Cold EI in which each peak exhibits a molecular ion and is amenable for trustworthy identification.   

Permethrin Drug Impurity Analysis with GC-MS with Cold EI and the Road to Failure in Such Analysis by GC-MS with Standard EI

Aviv Amirav, Tel Aviv University and Aviv Analytical

Abstract

A Permethrin impurity was successfully analyzed by GC-MS with Cold EI after it failed to be analyzed by GC-MS with standard EI. In this application note we demonstrate and discuss the ways GC-MS with standard EI analysis of relatively large compounds gradually becomes more difficult as the sample compound size is increased due to reduced total ion count signal, reduced molecular ion relative abundance and increased noise. Accordingly, as the analyzed sample compound becomes larger its GC-MS analysis becomes harder in a gradual fashion until it fails. In contrast, GC-MS with Cold EI can analyze twice larger compounds and thus significantly extends the range of compounds amenable for GC-MS analysis.    

Cold EI Versus Low Electron Energy EI

Aviv Amirav, Tel Aviv University and Aviv Analytical

Executive Summary

Recently, the use of low electron energy electron ionization is claimed to serve as a soft ionization method and it is sometimes referred to as "Soft EI". In this application note, we show and discuss that low-eV EI is not a universal soft ionization method, its applicability is limited to small molecules that exhibit molecular ions in 70 eV EI, and its enhancement of molecular ion abundances is small or non-existent for many compounds. Furthermore, the addition of a 2nd analysis and loss of signal with low eV EI rarely justify its use in real-world applications.

In contrast, Cold EI (electron ionization of cold molecules in supersonic molecular beams) is a far superior "Soft EI" ion source with close to universal applicability. We demonstrate that for squalane (C30H62), the molecular ion is the base peak in the 70 eV Cold EI mass spectrum, which is 10,000-fold higher than its 0.01% relative ion abundance in 14 eV low electron energy EI. Furthermore, Cold EI is the best ion source in all other main performance aspects, and most importantly, it significantly increases the range of compounds and applications amenable for analysis. Thus, Cold EI bridges the GC-MS gap with LC-MS and can increase the total GC-MS market.

Lipids in Human Serum Analysis by the 5975-SMB GC-MS with Cold EI

Aviv Amirav and Svetlana Tsizin, Tel Aviv University and Aviv Analytical, Tel Aviv Israel. 

Gabi Shefer, Ichilov hospital Tel Aviv Israel 

Introduction 

Cholesterol and triglycerides analysis in human blood is among the most widely used chemical medical diagnostics tests. Some estimate the number of such analysis at 1 Billion/year with cost of about $30 for each analysis (three analyses of cholesterol LDL, HDL and triglycerides at $10 each). Thus, the total lipids in blood analysis market is about $30 Billion/year. However, current analysis brings limited information and mass spectrometry can provide far better and more detailed lipids in blood information if an appropriate instrument for such analysis will be available. We used our GC-MS with Cold EI and demonstrated as below the analysis of large range of lipids in blood. Each analysis provided information on the amount of several free fatty acids, cholestadiene, cholesterol, vitamins E and 25 Hydroxy vitamin D3 and many diglycerides, cholesteryl esters and triglycerides. Notably, we can clearly distinguish differences among samples from different people. Each analysis took only 10 min via the use of column flow programing. Our results yielded extensive lipidomics information that may include new diagnostic tools. We feel that this new and advantageous assay for lipids profiling in blood is worthy of further investigation and evaluation.  

Triglycerides in Oils Analysis by the 5975-SMB GC-MS with Cold EI

Aviv Amirav, Tel Aviv University and Aviv Analytical, Tel Aviv Israel.
Hans-Gerd Janssen, Unilever R&D Vlaardingen and University of Amsterdam, Amsterdam The Netherland

Introduction 

Triglycerides analysis is challenging since these large compounds do not elute from standard GC columns under the conditions used in GC-MS and furthermore their standard EI mass spectra do not exhibit any molecular ion, as shown in the NIST library. In addition, their analysis by LC-MS is also challenging in view of their poor proton affinity. Thus, and in view of the importance of triglycerides analysis we decided to analyze three oil samples that were previously analyzed by GC-FID in order to evaluate these oils analysis by GC-MS with Cold EI. The challenge was to properly elute these large compounds and demonstrate the availability of abundant molecular ions plus useful and informative high mass fragments. Triglycerides analysis is very important for the food industry and such analysis can also be potentially important for human medical diagnostics in the form of cholesterol and triglycerides analysis in blood. Currently triglycerides are hydrolyzed and methylated to form FAMEs (fatty acid methyl esters) that are analyzed by GC-MS. However, in such analysis vital information is lost on the actual structure and concentration of the various parent triglycerides and GC-MS with Cold EI is challenged to exhibit such information.     

Linearity, Sensitivity and Response Uniformity Comparison of the Aviv Analytical 5975-SMB with Cold EI and the Agilent 5977A GC-MS with Standard EI

Aviv Amirav (1,2), Uri Keshet (1) and Bogdan Belgorodsky (1) 
1.   Tel Aviv University 
2.   Aviv Analytical Ltd 

Introduction 

Standard GC-MS analysis of low volatility and polar compounds is known to be plagued by ion source related peak tailing and degradation (see our post on peak tailing here). We found that these intra ion source tailing and sample degradation effects strongly depend on the sample concentration. Consequently, when these compounds are analyzed, the response of the standard EI ion source of GCMS is both non-uniform and non-linear. In this article we evaluate and compare the Aviv Analytical 5975-SMB GC-MS with Cold EI with the new Agilent 5977 Extractor GC-MS in the analysis of a test mixture containing five components, using the same on-column amounts. In contrast to the Agilent 5977 the Aviv Analytical 5975-SMB GC-MS with Cold EI exhibits uniform, linear, compound independent response. We concluded that the harder the compound analysis the greater is the sensitivity gain of the 5975-SMB over 5977 extractor with standard EI, up to and over a factor of 100.

Keep reading to find out how the Aviv Analytical 5975-SMB GC-MS with Cold EI uniquely enables uniform and linear ion source response with a substantial improvement of sensitivity in comparison with a state of the art GC-MS with standard EI.

Explosives Analysis with the 5975-SMB GC-MS with Cold EI

Aviv Amirav, Professor of Chemistry at Tel Aviv University and Director - Aviv Analytical Ltd, December 4, 2013.

Introduction

Effective explosives analysis is very important for homeland security and forensic applications. However, explosives analysis is challenging and the number of explosive compounds that require monitoring is growing with time. Furthermore, the growing use by terrorists of peroxide explosive compounds such TATP (triacetone triperoxide, C9H18O6) and HMTD (Hexamethylene triperoxide diamine, C6H12N2O6) implies that modern analytical techniques of explosive analysis must be as effective with peroxide explosive as with the traditional poly nitro explosives. In fact, there is no limit to the desirable LOD and selectivity of explosives analysis but these features must be combined with an analytical system that enables the analysis of the entire explosives family used by terrorists. We note that the actual system sensitivity and selectivity should be measured by its performance with the most difficult to analyze explosives such as the peroxide explosives and not with TNT and alike. 

Keep reading to find out how the Aviv Analytical 5975-SMB GC-MS with Cold EI uniquely enables the sensitive analysis of the full range of explosive compounds including TATP, HMTD, R-Salt, TNT, ETN, PETN, RDX, Tetril, HMX and Urea Nitrate while providing abundant molecular ions or high mass fragments that improve the detection selectivity and identification capability of these labile compounds.

Extending the Range of Compounds Amenable for GC-MS Analysis

Aviv Amirav, Alexander B. Fialkov and Tal Alon 

Tel Aviv University and Aviv Analytical. July 22, 2013.  

Introduction

GC-MS is a powerful technique yet it suffers from a major Achilles Heel of limited range of low volatility and thermally labile compounds amenable for analysis. However, it was discovered and investigated by A. B. Fialkov, A. Gordin and A. Amirav, J. Chromatog. A.  991, 217-240 (2003) that this range can be significantly extended via two major changes in GC-MS:

1. The use of short columns with high column flow rates and preferably with a temperature programmable injector which lowers the elution temperatures from the injector liner to the column and from the column itself (slower temperature programming rate and thinner films also help) and; 
2. The use of a fly-through ion source in combination with sample compounds in supersonic molecular beams (Cold EI) eliminates any ion source peak tailing and/or sample degradation on the hot metallic ion source surfaces. 

The use of GC-MS with Cold EI opens the door for significantly increased range of compounds and applications that are amenable for GC-MS analysis. Consequently, GC-MS with Cold EI bridges the gap with LC-MS, helps to analyze the bottleneck hard to analyze compounds in current methods and opens the door for new and untapped opportunities in GC-MS analysis.                

Keep reading to find out how the Aviv Analytical 5975-SMB GC-MS with Cold EI uniquely enables significant extension of the range of compounds amenable for GC-MS analysis and as a result enables new types of GC-MS applications.

Organo-Iodine Compounds Analysis by the 5975-SMB GC-MS with Cold EI

Aviv Amirav, Professor of Chemistry at Tel Aviv University and Director - Aviv Analytical

Larisa Panz Ph.D., Ksenia Kulbitski and Professor Mark Gendelman, Schulich Faculty of Chemistry at the Technion Haifa Israel. 

Introduction

Organo-iodine compounds are used in several applications, mostly as intermediates in organic synthesis because of the easy formation and cleavage of the C–I bond. The analysis of organo-iodine compounds is highly challenging since most of them are not compatible with LC-MS and electrospray ionization while in GC-MS their analysis is hampered via their possible decomposition at the GC injector, column and ion source since the iodine-carbon bond is by far the weakest among all halogen-carbon bonds. Furthermore, even if the organo-iodine compounds elute from the GC column their standard EI mass spectra are often void of any molecular ions and as a result their identification becomes impossible. Motivated by the above challenges we explored the analysis of novel synthetic organo-iodine compounds with the Aviv Analytical 5975-SMB GC-MS with Cold EI, developed a method for its successful analysis and compared the results with that obtained with GC-MS with standard EI.

Keep reading to find out how the Aviv Analytical 5975-SMB GC-MS with Cold EI uniquely enables the effective analysis of synthetic organo-iodine compounds and provides unambiguous identification of all the synthetic organo-iodine reaction mixture compounds and as a result helps optimizing the synthesis reaction conditions and yields.

What Can be Improved in GC-MS – When Multi Benefits are Transformed into a GC-MS Revolution

Aviv Amirav, Alexander B. Fialkov and Tal Alon, Tel Aviv University and Aviv Analytical

GC-MS vendors typically describe and characterize their systems using a small set of specifications that include octafluoronaphthalene (OFN) signal to noise ratio (SNR), mass range and scan speed. As a result, improvements in GC-MS are often focused on the incremental increase of these few specifications. However, GC-MS is characterized by many additional features and operational parameters that contribute to its performance, and their improvements can make a big impact on the GC-MS analytical capabilities. Many such GC-MS aspects are improved by incorporating the new Cold-EI GC and MS interface and ion source technology and by using "out of the box" thinking. In LC-MS, the biggest revolution was brought not by LC or MS improvements but rather by the development of a new interface and ionization method namely Electrospray. Similarly, Cold EI with its supersonic molecular beams interface and fly-through ion source brings multiple benefits and improvements into GC-MS which can initiate a new GC-MS revolution. This blog post lists 62 GC-MS improvements brought forth by the Cold EI interface and ion source, encompassing any and every important aspect of GC-MS, and explains how the unique features of GC-MS with Cold EI enable these benefits. 

Keep reading to find out how the 5975-SMB GC-MS with Cold EI improves GC-MS not by one or few but by 62 different aspects. When these multiple benefits are combined they are destined to be transformed into the next GC-MS revolution.  

Free Fatty Acids Analysis by GC-MS - Cold EI Versus Standard EI

Aviv Amirav (1)  and David Benanou (2) 

1. Professor of Chemistry at Tel Aviv University and Director – Aviv Analytical 

2. Veolia Environment Recherche and Innovation 

Gas Chromatography Mass Spectrometry (GC-MS) is known to be limited in the analysis of polar organic compounds such as free acids and bases which require derivatization for their proper analysis. The problem in free acids analysis emerges from extended peak tailing at the GC separation column (and liner) as well as peak tailing and sample compound degradation at the metallic surfaces of the ion source. Recently, a few of the major column vendors introduced new brands of inert columns and liners that alleviates and significantly reduces column related peak tailing by polar compounds. However, the problem of ion source activity remained unsolved and it is currently the main reason why free fatty acids and other polar organic compounds cannot be properly analyzed by standard GC-MS and require derivatization. While derivatization enables the GC-MS analysis of polar compounds such as fatty acids, it requires another step in the sample preparation. Furthermore, derivatization is not always practical since for a large variety of sample types the presence of compounds that require derivatization is not known hence they are analyze without derivatization and in such cases important sample information on such polar compounds is lost.

In this Advanced GC-MS Blog post we describe the analysis of an anti foam agent which in this case is hydrocarbon based oil that also contains free fatty acids. However, the presence of these free fatty acids was not initially known.            

                  

Keep reading to find out how the 5975-SMB GC-MS with Cold EI uniquely enables the analysis of free underivatized fatty acids which could not be properly analyzed by standard GC-MS.  

Drug Impurities Analysis by the Aviv Analytical 5975-SMB GC-MS with Cold EI

Aviv Amirav, School of Chemistry, Tel Aviv University and Director – Aviv Analytical 

Drugs are typically analyzed by LC-MS. When active pharmaceutical ingredients (API) are analyzed for the presence of impurities, the general requirements are that every impurity found should be at concentration below 0.1% of the API or it must be either cleaned or identified and toxicologically characterized which is a time consuming and expensive procedure. However, the ionization yields of Electrospray and/or APCI in LC-MS are highly non-uniform and compound dependent, and as a result when an impurity is found in a given LC-MS mass chromatogram its actual concentration is not known. Consequently, regardless of its actual concentration that can be well below 0.1% every impurity that is found must be isolated, fully identified, synthesized (or obtained) and serve for the provision of a calibration curve in order to measure the actual concentration of that small impurity peak. In addition, several types of impurities such as non-polar compounds are not properly ionized and are not detected by LC-MS. Thus, clearly drug impurity analysis represents an unmet analytical challenge.  

Alternatively, GC-MS with its standard electron ionization (EI) can be used and has, for volatile compounds, approximately uniform, semi-quantitative ionization yield. However, GC-MS with standard EI is compatible only with a limited range of thermally stable drugs and it provides useful molecular ions only for a portion of the analyzed compounds. Furthermore, ion source degradation and peak tailing severely erode the standard EI response uniformity for semi-volatile compounds. 

We used the Aviv Analytical 5975-SMB GC-MS with Cold EI for the analysis of drug impurities. Cold EI is the electron ionization of cold molecules in supersonic molecular beams (SMB). The technology of GC-MS with Cold EI is reviewed by A. Amirav, A. Gordin, M. Poliak, and A. B. Fialkov, J. Mass Spectrom., 43, 141-163. (2008). The later is available upon request. 

GC-MS with Cold EI as implemented with the Aviv Analytical 5975-SMB uniquely extends the range of drugs that are amenable for GC-MS analysis, including thermally labile drugs that are not compatible with standard GC-MS analysis. Furthermore, it is characterized by having uniform response to all compounds due to its contact-free fly-through ion source that eliminates any ion source peak tailing or losses. In addition, it provides trustworthy enhanced molecular ions that can be converted with the Tal-Aviv Molecule Identifier software into elemental formulae. Thus, the high sensitivity and uniform response of Cold EI uniquely enables the detection, identification and quantitation of low level impurities in drugs without lengthy calibration procedures. 

In order to evaluated the above attributes of the 5975-SMB we analyzed impurities in two Active Pharmaceutical Ingredient (API) samples (powder form) of Haloperidol and Ramelteon (Structures are given below).   

      

 

Keep reading to find our how the Aviv Analytical 5975-SMB GC-MS with Cold EI uniquely enables quantitative drug impurities analysis and helps in their identification. 

A Universal Method for Pesticide Analysis with the 5975-SMB GC-MS with Cold EI

 Aviv Amirav, Professor of Chemistry at Tel Aviv University and Director – Aviv Analytical

Introduction 

An estimated 2000 chemicals are being used as pesticides worldwide, including banned pesticides and other hazardous chemicals. In view of international food trade this large number implies that pesticide analysis should not be treated as target compounds analysis. Thus, current mass spectrometry instrument development is challenged to provide a one system and method that will be capable of analyzing as many pesticides as possible in full scan mode, in the needed instrumental sensitivity and selectivity in complex agricultural matrices, and in a short amount of time for effective (and lower cost) pesticides screening.   

In order to advance towards meeting the above challenges, a GC-MS with Cold EI (EI of Cold Molecules in Supersonic Molecular Beams) was evaluated in the past with pesticide analysis in agricultural products as described in M. Kochman, A. Gordin, P. Goldshlag, S. J. Lehotay and A. Amirav "Fast, High Sensitivity, Multi-Pesticide Analysis of Complex Mixtures with the Supersonic GC-MS" J. Chromatogr. A. 974, 185-212 (2002). Recently, we further evaluated pesticide analysis with the Aviv Analytical 5975-SMB GC-MS with Cold EI (GC-MS with supersonic molecular beam (SMB) interface and its fly through ion source) and developed a method to meet the above challenges. 

Keep reading to find out how the Aviv Analytical 5975-SMB GC-MS with Cold EI enables a truly universal method of pesticide analysis in agricultural matrices with improved range of compounds amenable for analysis, reduced matrix interference, lower LOD particularly for difficult to analyze pesticides, improved pesticide identification and shorter analysis time.   

Measurement and Optimization of Organic Chemical Reaction Yields by GC-MS with Cold EI

Aviv Amirav, School of Chemistry, Tel Aviv University and Director – Aviv Analytical 

Alexander Gordin, Bogdan Belgorodsky, Boaz Seemann, Michael Gozin and Alexander B. Fialkov, School of Chemistry, Tel Aviv University.  

Introduction

Chemical reactions are typically involved with mixing the reactants together in an appropriate solvent, often with addition of a catalyst, and allowing them to undergo a reaction, which may take from couple of minutes to few days. The progress of the reaction is either assumed or monitored most-commonly by thin layer chromatography. However, little or no information is obtained on-line on the reaction actual progress and its yield. At the perceived end of the reaction, the products are separated and purified by preparative chromatography, distillation, sublimation, selective precipitation or crystallization, processes which may take several hours or even several days. Subsequently, the purified products are analyzed by 1H and 13C NMR and by high resolution mass spectrometry (often Quadrupole Time of Flight (QTOF) mass spectrometer), typically via flow injection electrospray ionization (FI-ESI). Yet, while FI-ESI-QTOF provides elemental formulas information, it does not provide information about the synthesis yield, products purity, availability of isomers and on the reaction mechanism, since the already-purified compounds are analyzed and since ESI has non-uniform, highly compound dependent ionization yields. 

In order to alleviate the above mentioned shortcomings of organic synthesis, we used the Aviv Analytical 5975-SMB GC-MS with Cold EI for semi-on-line monitoring of organic chemical reactions for obtaining information on the reaction products identity and purity and for mechanism elucidation and reaction yield optimization. The 5975-SMB was used with reaction mixtures without their prior separation and purification as required for NMR analysis, thereby saving time and effort. Our unique 5975-SMB GC-MS with Cold EI is based on GC interface with the MS with supersonic molecular beams (SMB) and on the ionization of the sample molecules during their axial flight through an open electron ionization ion source as vibrationally cold molecules (hence the name Cold EI). GC-MS with Cold EI was demonstrated to significantly extend the range of compounds amenable for analysis, practically always giving molecular ions, enabling effective fast GC-MS analysis and providing elemental formulas via isotope abundance analysis with unit mass resolution quadrupole MS. In addition, it uniquely provides uniform response to all compounds, a feature which is vital for the measurement of chemical reaction yields. 

Keep reading to find out how the Aviv Analytical 5975-SMB GC-MS with Cold EI can change the way organic synthesis is performed via enabling semi on-line monitoring of the progress of organic reactions and via the provision of information on the reaction mechanism, products identity and purity and the chemical reaction yield for its optimization. 

Jojoba Oil Analysis using the Aviv Analytical 5975-SMB GC-MS with Cold EI

Aviv Amirav, Professor of Chemistry at Tel Aviv University and Director – Aviv Analytical

N. Gabriel Lemcoff, Associate Professor of Chemistry at Ben-Gurion University

Introduction 

Jojoba oil is produced from the seeds of the Jojoba plant, a shrub native to the South West USA and now also grown in the Middle East. The oil makes up approximately 50% of the Jojoba seeds by weight. Jojoba oil appears as a clear golden liquid wax and among oils it is unique because it does not contain triglycerides unlike most other vegetable oils. 

Jojoba oil is used as an additive in many cosmetic products, especially those marketed as being made from natural ingredients. In particular, such products commonly containing jojoba are lotions and moisturizers, hair shampoos and conditioners, or the pure oil itself may be used on skin or hair.

Recently Jojoba is explored as a source of bio-fuel since it has an attraction – the Jojoba plant can be grown in saline soils, and in desert lands. In fact, Jojoba is considered as one of the most practical and profitable solutions for desert plantation. Hot summers, warm winters, desert soil, minimal water, great salinity tolerance, lesser possibilities for infection, lesser need for fertilizers and generous financial income are all valuable characteristics of the plant.  

Jojoba oil has a unique composition of almost pure heavy esters with two double bonds. Professor Gabriel Lemcoff from the Ben Gurion University in Beer Sheva Israel is exploring its unique features while performing various novel organic synthesis experiments with this oil. 

Professor Lemcoff provided a pure Jojoba oil sample and asked Prof. Amirav to help with its analysis since it can not be analyzed by standard GC-MS (being too heavy and without expected molecular ions) or LC-MS.

We were able to analyze the pure oil and its chemical reaction products (which are not shown in this report) using the Aviv Analytical 5975-SMB GC-MS with Cold-EI, and in this post we present the Jojoba oil analysis results and conditions.

Beeswax in Fruit and Vegetables Analysis and the Emerging Challenge of Material Identification

Aviv Amirav, Professor of Chemistry at Tel Aviv University and Director – Aviv Analytical 

Igal Bar Ilan Ph.D.  Migal Laboratories, Kiryat Shmona Israel    

Introduction 

Beeswax serves to treat growing numbers of fruits and vegetables in order to increase its shelf time, reduce product dehydration, suppress mold growth and improve the appearance (shiny) of the treated fruits or vegetables. However, while wax is generally considered as safe, some say that such treatment serves as an evidence for extended (long) time between harvest and consumption hence reduction in certain nutrients. A few types of waxes are used but beeswax is considered as an "organic" wax hence of high quality. Recently beeswax analysis is required in certain fruits and vegetables (by the EU) and standard GC-MS or LC-MS failed to perform such challenging analysis. Beeswax is used in combination with minor addition of olive oil and the challenge is to develop a method to analyze such wax in the 1-100 ppm range in certain fruits and vegetables. Note that wax is a mixture of several heavy compounds, most notably hydrocarbons and heavy esters plus some triglycerides, thus 1 ppm means about 40 ppb each of its major heavy ingredients. In addition, unlike in pesticide analysis in this case the emerging challenging need is to identify a material as opposed to compound. 

After method development we successfully analyzed the wax in 16 fruit and vegetable samples including avocado, cucumber, tomato and apple using the Aviv Analytical 5975-SMB GC-MS with Cold-EI.

Keep reading for the analysis details, results and conclusions.