News from LabRulezGCMS Library - Week 13, 2025

Our Library never stops expanding. What are the most recent contributions to LabRulezGCMS Library in the week of 24th March 2025? Check out new documents from the field of the gas phase, especially GC and GC/MS techniques!

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This week we bring you application notes by Agilent Technologies, Shimadzu and Thermo Fisher Scientific!

1. Agilent Technologies: Fast Analysis of Human Malodor Compounds, Volatile Organic Acids by SPME and Column Backflushing

• Application note
• Full PDF for download

Human body odor is caused by the metabolism of sweat by a host of microorganisms in the human underarm. Human sweat is odorless and certain microbial species metabolize secreted sweat components into volatile, malodorous compounds. These bacterial species exist as a microbiome community. Some members of this community can enzymatically convert nonodorous components of human sweat into malodorous compounds, while others are not responsible for creating noxious smells. The human axillary hosts a wide array of bacterial species, including Corynebacterium jeikeium and Corynebacterium tuberculostearicum along with Staphylococcus aureus, Staphylococcus epidermidis and Staphylococcus hominis.¹ The relative population of each species within the microbiome community influences the number of malodorous compounds liberated from digested human sweat. Typically, a variety of volatile organic acids (VOAs) are produced and are comprised of long-chain fatty acids, fatty acids bound to amino acids, and sulfur-containing amino acids. However, several studies suggest that only a few, key odorants are predominately responsible for the negative human perception of body odor, which include E-3-methyl-2-hexenoic acid (E-3M2H), isovaleric acid, 3-methyl-3-sulfanylhexan-1-ol (3M3SH), and 3-hydroxy-3-methylhexanoic acid (HMHA)^.2

SPME is a technique invented in the early 1990s, where a fiber coated in an absorbent polymer phase is exposed to a sample, either by direct immersion or in the headspace above the sample (Figures 1 and 2).4 The target analytes are adsorbed from the sample to the phase, depending on affinity to the phase, and once reaching equilibrium, will be removed from the sample and desorbed onto the inlet of the GC. These methods of sample preparation and sample introduction have been especially useful in the analysis of low concentration compounds in volatile compounds and in decreasing the effects of heavy matrices.

Determining which GC column phase to select can be difficult when analyzing trace level aromatic compounds. Spectral deconvolution can be helpful in removing ions of coeluting compounds and in the identification of all peaks. The Agilent MassHunter Quantitative Analysis software with the Unknowns Analysis program is a tool that can be used to analyze multiple samples, deconvolute spectra, and compare retention times or retention index (RI) commercially available libraries such as NIST or user-created libraries.9 To demonstrate the benefit of combining these techniques, the application of analyzing malodor compounds was optimized to improve overall data quality and productivity.

Materials and methods

Method 

A Gerstel MPS Robotic Pro was installed on an Agilent 8890 GC system with an Agilent 5975 MSD. The SPME headspace parameters and MS conditions are listed in Tables 1 through 4. Data were analyzed using Agilent MassHunter Unknowns Analysis.

Conclusion 

The use of solid phase microextraction (SPME) is a valuable sampling technique for measuring volatile organic compounds produced by human skin commensals, especially when considering the complex sample matrix of multispecies in vitro bacterial communities. The use of a polar column phase such as an Agilent J&W DB-Wax UI GC column will provide better peak shape for organic acids than a nonpolar column. When switching to a more polar phase, the implementation of midcolumn backflush will help to remove unwanted high-boiling matrix contamination in the VOA sample at lower oven temperatures. This method allows for faster run times, extended column lifetimes, and reduced contamination of the MS source. The Agilent MassHunter Quantitative Analysis software with Unknowns Analysis can be used to easily deconvolute samples by searching against a commercial library or a user-created library to aid in faster sample analysis. Finally, combining SPME with midcolumn backflushing provides a major improvement in method runtimes compared to other methodologies.

2. Shimadzu: Extractable Study of Pharmaceutical Packaging and Delivery System Used for Ophthalmic Drug Product

• Application note
• Full PDF for download

User Benefits:

• ASSP™ and simultaneous Scan/MRM analysis facilitates accurate qualitative identification and quantitation at trace levels
• UFsweeper™ allows quicker ion transmission in the collision cellsuppressing crosstalk & enables faster MRM analysis
• Thermal Desorption (TD) System offers a direct sampling feature that significantly reduces extraction time and prevents the loss of VOCs

Extractable and leachable (E & L) studies are becoming increasingly important in the pharmaceutical industry as it is mandatory requirement from FDA during filing of the drug product. The purpose of E & L studies is to identify and evaluate possible toxicological risks. E & L studies in the regulatory references aim to identify traces of potential chemical substances. These substances may be harmful to patients due to their toxicity or may impact the activity of the drug product. Hence, to ensure the safety and efficacy of the drug throughout its shelf-life period, E & L studies play an important role. Impact of Extractables on efficacy and safety of the drug product is presented in Figure 1.

What are Extractables and Leachables? 

Extractables are organic and inorganic chemical entities that are released from a pharmaceutical packaging / delivery system, packaging component, or packaging material of construction into an extraction solvent under laboratory conditions. [1] 

Leachables are foreign organic and inorganic chemical entities that are present in a packaged drug product because they have leached into the packaged drug product from a packaging/delivery system, packaging component, or packaging material of construction under normal conditions of storage and use or during accelerated drug product stability studies [1].

Experimental design

There are many ways by which extractable study can be designed like generating extract by maceration (solvent soaking), by reflux/Soxhlet/sealed vessel/sonication or solvent extraction (manually and automated) etc. [1] However, one can perform this study by direct heating the CCP using the technique such as TD System. 

Physical as well as chemical properties of the sample should be considered while designing the extraction experiment. Polarity, pH and chemical components of the drug product plays an important role in the extraction process. Extractable study is performed at accelerated temperature conditions considering worst case scenario for storage of the drug product. 

Here, we conducted extractable study at variable pH and by refluxing the CCP with solvents of different polarities. Also, TD System was utilized for the assessment of extractable study. Empty CCP designed for the storage of Ophthalmic drug product were purchased from the local resource. Using these CCP’s, multiple experiments were performed and acquired data were evaluated to identify the potential extractables. Below are the detailed explanation for Experiment A, B, C, D, E etc. Experiment A is performed by incubating CCP in aqueous solutions having different pH values and experiment B, C, D & E were performed by refluxing the CCP with organic solvents of different polarities.

Mathods of analysis

Method-I: 

In this method, seventeen standards were analyzed by GCMS-TQ8040 NX with AOC-20i autosampler. For the method suitability, few important parameters like system suitability, linearity and LOQ precision were performed. Instrument Parameters are as mentioned below.

Conclusion

• The chromatographic profile of solvent reflux experiment shows higher number of extractables than aqueous incubation. Furthermore, in aqueous incubation, the content of extractables is well below the AET level. Since, placebo of the ophthalmic drug product is completely aqueous, CCP may be used for the storage. 
• Chromatographic profile from different experiments conclude that, the compounds detected in solvent extraction are different than the compounds obtained in TD analysis. Thus, additional information obtained from TD profile will further facilitate material characterization. 
• Shimadzu’s GCMS-TQ8040 NX triple quadrupole mass spectrometer provides high scan speed of 20000 amu/sec with ASSP technology which enables simultaneous SCAN/MRM analysis which is critical in E&L study. The UFsweeper technology enables 800 MRMs/sec which minimizes cross-talk, enhances selectivity & sensitivity.

3. Thermo Fisher Scientific: Quantitation of volatile PFAS in environmental samples using SPME Arrow and Orbitrap Exploris GC

• Application note
• Full PDF for download

Application benefits

• A simple and robust sample preparation for volatile PFAS based on SPME Arrow for reducing manual handling and potential contamination issues.
• Sensitive and quantitative analysis of volatile PFAS in complex environmental samples using high resolution accurate mass (HRAM) that meets challenging reporting limits.
• Flexible data processing using full scan HRAM to include additional points of confirmation and quickly increase scope of analysis without reanalysis.

Per- and polyfluoroalkyl substances (PFAS) contain one or more alkyl radicals with all the hydrogens replaced by fluorine atoms. Traditionally, two groups of PFAS have been of the most concern and subject to control and monitoring. The first group includes ionic (or acidic) PFAS—the perfluorocarboxylic acids (perfluorooctanoic acid (PFOA)) and perfluoroalkylsulfonates (perfluorooctanesulfonic acid (PFOS))—where LC-MS-MS is the most common analytical technique. The second group includes neutral (or volatile) PFAS—the fluorotelomer alcohols1 (FTOHs) and N-substituted fluoroalkylsulfonamides (FOSAs). For this group, due to the volatility, GC-MS is the analytical method of choice and is the focus of this study.

The accurate analysis of PFAS by testing laboratories requires robust and streamlined analytical workflows. These methods must overcome the challenges of an ever-growing list of PFAS compounds, diversity of sample matrices, and demanding analytical performance requirements. Typically, a gas chromatography instrument coupled to low-resolution, nominal mass triple quadruple mass spectrometer (GC-MS/MS) has been the system of choice for the sensitive and selective detection of a wide range of target PFAS compounds. However, the development of high-resolution accurate mass (HRAM) Orbitrap mass spectrometry coupled to GC has proved to be a valuable alternative to triple quadrupole GC-MS. With HRAM mass spectrometry, the default acquisition mode is untargeted (fullscan) meaning that all the ions are acquired with high selectivity at the same time across a specified mass range, making the method setup and data acquisition simple to manage and giving the analyst the flexibility to decide on which compounds to focus. This can extend into retrospective analysis of data to evaluate for the presence/absence of other contaminants not necessarily of interest at the time of acquisition. For example, this could allow the search for other PFAS compounds beyond the target list. 

In this study, the performance of the Orbitrap Exploris GC high-resolution accurate mass (HRAM) spectrometer together with the headspace solid phase micro extraction (SPME) Arrow for the quantitative analysis of volatile PFAS including FTOH, fluorotelomer acrylates (FTACs), fluorotelomer methacrylates (FTMACs), fluorotelomer iodides (FTIs), Me/Et-FOSAs, and Me/Et-FOSEs is demonstrated.

Experimental

Instrument and method setup Headspace extraction and injection of samples were performed using the Thermo Scientific™ TriPlus™ RSH SMART autosampler equipped with the Thermo Scientific™ SMART SPME Arrow 1.1 mm PDMS 100 µm fiber (P/N 36SA10P1-SM). Incubation and extraction were performed online followed by sample injection/ desorption. After sample injection, the SPME Arrow fiber was re-conditioned at high temperature under a nitrogen flow using an SPME conditioning station to avoid sample carryover between injections. Further details surrounding the SPME Arrow operating parameters can be found in Table 1.

A Thermo Scientific™ TRACE™ 1610 GC equipped with a Thermo Scientific™ TraceGOLD™ TG-5SilMS (30 m × 0.25 mm I.D. × 0.25 µm film) capillary column (P/N 26096-1420) was used to perform the chromatographic separation. Oven program conditions can be found in Table 1. Data acquisition was carried out in full scan analysis using an Orbitrap Exploris GC mass spectrometer. Additional MS method parameters are summarized in Table 2. External mass calibration was performed prior to analysis, while characteristic ions representing column bleed were used as lock masses when scanning in EI to perform internal mass calibration. Sample acquisition and qualitative processing was performed using the Thermo Scientific™ Chromeleon™ version 7.3.2 Chromatography Data System (CDS) software. Additional screening data processing was done using Thermo Scientific™ Compound Discoverer™ software with the Thermo Scientific™ Orbitrap GC-MS HRAM Contaminants Library.

Conclusion 

Volatile perfluoroalkyl substances (PFAS) have become a significant focus in environmental analysis due to their potential health and environmental risks. The inherent complexity of environmental samples and the need for minimal sample handing of volatile PFAS is critical in accurate quantitation and detection. The method presented utilizes a combination of HS-SPME Arrow with an Orbitrap Exploris GC to provide an efficient workflow for the analysis of PFAS compounds giving analytical advantages including: 

• Minimal sample preparation and online extraction using the TriPlus RSH SMART robotic autosampler increase the sample throughput and minimize the risk of contamination. Using SPME Arrow extraction technique, good recovery of all target volatile PFAS compounds was demonstrated. 
• Full scan acquisition at high mass resolution provides targeted quantitative analysis together with non-target analysis to quickly increase the scope of analysis and screen for other compounds outside the target list. 
• Full scan acquisition facilitates versatile data processing and allows for the incorporation of additional confirmation points, including supplementary confirmation ions, spectral matching, and isotope pattern. 
• Limits of detection ranging from 0.1 to 1.4 ng/L are well below the target 5 ng/L level of PFAS compounds in complex environmental matrices. 
• Application of the method to real samples showed PFAS compounds present in all samples with some very high concentrations in crude sewage and landfill leachate.