GC-MS) And Gas Chromatography-combustion-isotope Ratio Mass Spectrometry Research Articles

Tracing culinary practices in the western provinces of the Roman Empire using Organic Residue Analysis

This study aims at reconstructing foodways in the north-east (NE) of the Iberian Peninsula, focusing on lipid residue analysis of utilitarian vessels and using as case studies the sites of Puig Castellar of Biosca (180–120 BCE) and Guissona (120 BCE-third century CE). In total, fifty vessel fragments of different types and origins were analysed with techniques such as gas chromatography-mass spectrometry (GC-MS) and gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). As some vessel fragments were recovered from waterlogged contexts, they had excellent lipid preservation, which enabled the detection of various compounds that are rarely reported in archaeological contexts in Iberia. Analyses revealed both animal and plant products in the vessels, suggested that a variety of food resources was consumed, and that vessels had multiple uses. The detection of levoglucosan in some extracts, along with other heat markers, further suggested the presence of cellulose or starchy products in contact with fire as well as the heating of animal fats. Combined with available bioarchaeological and palaeoenvironmental studies from both sites, the analyses indicated a continuity in diet across the time periods during which the sites were occupied. Some differences were also observed, including the possible use of tubers until the first century CE. The results are then contextualised and compared with other available organic residue studies from the Roman Iberian Peninsula. The study demonstrates how the combination of multiple bioarchaeological proxies and biomolecular approaches can provide a holistic means to approach Roman foodways.

Technological variability in foragers’ pottery productions at the early-mid Holocene site of Sphinx, western part of Jebel Sabaloka, Sudan

The site of Sphinx (SBK.W-60) is located about 3.5 km from the present Nile in the western part of Jebel Sabaloka, upstream of the Sixth Nile Cataract, in Sudan. This site uniquely includes Early Khartoum (Mesolithic) artifacts with no intrusive elements and has been dated from the ninth to the end of the sixth millennium cal BC. Excavations at Trench 7, in particular, brought to light a 1.2-m thick deposit with the quantitatively and qualitatively richest artifactual materials. Analysis and classification of the pottery assemblage from this site have been conducted with the aim of observing manufacturing techniques and examining the correlation between pottery production, cultural change and chronological variability. We undertook visual examinations of the manufacturing techniques combined with petrographic (optical microscopy, OM) and chemical analyses (instrumental Neutron Activation Analysis, iNAA), observations of manufacturing and decorative techniques, gas chromatography-mass spectrometry (GC-MS) and gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) on absorbed organic residues. The vertical distribution of the ceramic assemblage in Trench 7 reveals the existence of a relative sequence suggesting consistent technological variability throughout the site's occupation.

Pastoralist Foodways Recorded in Organic Residues from Pottery Vessels of Modern Communities in Samburu, Kenya

Organic residue analyses of archaeological ceramics can provide important insights into ancient foodways. To date, however, there has been little critical reflection on how lipid residues might (or might not) reflect dietary practices or subsistence strategies more generally. A combination of ethnoarchaeological research and chemical and isotopic analyses of lipid residues from pottery made and used by modern Samburu pastoralists in northern Kenya was undertaken to supplement the interpretive framework used in archaeological investigations. A total of 63 potsherds were collected from various contexts, including settlement sites and rockshelters, and analysed using gas chromatography (GC), gas chromatography-mass spectrometry (GC-MS) and gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). The results showed that the free fatty acids, palmitic (C16:0) and stearic (C18:0), dominated the lipid profiles, with extremely broad ranges of δ13C values. The majority of the Δ13C values from Samburu pots suggest that vessels were intensively used to process ruminant carcass products, yet the Samburu economy is not, in fact, meat-based at all. Despite an overall reliance on dairy products, milk is rarely processed in ceramic vessels, largely due to cultural prohibitions. Surprisingly, a number of vessels from one site, Naiborkeju Hill, were used to process dairy products. Compound-specific radiocarbon dating of lipids from these sherds suggests that this pottery originated from an earlier period, demonstrating a possible shift in ceramic use by pastoralist communities in this region over time. The overall conclusion is that lipid residues may not necessarily reflect, in a simple way, the day-to-day consumption or the perceived relative importance of different foodstuffs. In the Samburu case, lipid residues reflect the functional and ideological suitability of ceramics for processing only certain types of food (meat/fat/bones), despite an overall reliance on milk. These conclusions are important when considering the origins and development of African pastoralism, for example, as interpreted from the archaeological record.

MALDI-FT-ICR-MS for archaeological lipid residue analysis.

Soft-ionization methods are currently at the forefront of developing novel methods for analysing degraded archaeological organic residues. Here, we present little-used soft ionization method of matrix assisted laser desorption/ionization-Fourier transform-ion cyclotron resonance-mass spectrometry (MALDI-FT-ICR-MS) for the identification of archaeological lipid residues. It is a high-resolution and sensitive method with low limits of detection capable of identifying lipid compounds in small concentrations, thus providing a highly potential new technique for the analysis of degraded lipid components. A thorough methodology development for analysing cooked and degraded food remains from ceramic vessels was carried out, and the most efficient sample preparation protocol is described. The identified components, also controlled by independent parallel analysis by gas chromatography-mass spectrometry (GC-MS) and gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS), demonstrate its capability of identifying very different food residues including dairy, adipose fats as well as lipids of aquatic origin. The results obtained from experimentally cooked and original archaeological samples prove the suitability of MALDI-FT-ICR-MS for analysing archaeological organic residues. Sample preparation protocol and identification of compounds provide future reference for analysing various aged and degraded lipid residues in different organic and mineral matrices.

Archaeological bone lipids as palaeodietary markers.

Stable isotope analysis of archaeological and fossil bone samples can provide important insights into past environments, ecologies and diets. Previous studies have focused on stable carbon and nitrogen isotopes in bone collagen, or carbon isotopes in bone mineral (bioapatite). Carbon isotope analysis of lipids from archaeological bone has received much less attention, partly due to the lack of suitable methodologies allowing sufficient recovery of compounds for structural and isotopic characterisation. Here we show that lipids can be easily and reliably recovered from archaeological bone using a modified protocol, and that these provide complementary dietary information to other bone components. Human and animal bones were obtained from a variety of archaeological contexts. Lipids were sequentially extracted using solvent extraction (dichloromethane/methanol), followed by acidified methanol extraction (methanol/H2SO4). The lipids were then analysed by gas chromatography/mass spectrometry (GC/MS) and gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS). Appreciable amounts of endogenous lipid were recovered from archaeological bone. Importantly, a comparison between compound-specific and bulk collagen isotopic data shows that archaeological bone lipids reflect dietary input and can be used to distinguish between marine and terrestrial consumers, as well as between C3 and C4 plant consumers. Furthermore, the presence of essential fatty acids directly incorporated from diet to bone may provide additional palaeodietary information. Our findings suggest that archaeological bone lipids are a hitherto untapped resource of dietary information that offer additional insights to those gained from other isotopic analyses of bone.

Lipids, pots and food processing at Hočevarica, Ljubljansko barje, Slovenia

The paper presents the results of lipid analyses of pottery samples from Ho≠evarica (Ljub- ljansko barje, Slovenia). Total lipid extracts were subjected to high temperature gas chromatography (HT-GC), gas chromatography-mass spectrometry (GC-MS) and gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). The results show that some vessels were used for prepar- ing ruminant meat and vegetable, but also the remains of aquatic food were identified. The processing of non-ruminant meat was detected in a few samples. A high number of pottery samples yielded the presence of beeswax lipids. The charred residual on pottery was AMS 14C dated.

Complementary stable carbon isotope ratio and amount of substance measurements in sports anti‐doping

The detection of steroids originating from synthetic precursors against a background of their chemically identical natural analogues has proven to be a significant challenge for doping control laboratories accredited by the World Anti-Doping Agency (WADA). The complementary application of gas chromatography-mass spectrometry (GC-MS) and gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS) has been demonstrated to provide specific detection of endogenous steroid misuse for improved anti-doping analysis. Markers of synthetically derived steroids are reviewed on the basis of abnormal urinary excretions and low (13)C content. A combinatorial approach is presented for the interpretation of GC-MS and GC-C-IRMS data in the anti-doping context. This methodology can allow all relevant information concerning an individual's metabolism to be assessed in order to make an informed decision with respect to a doping violation.

13C tracer recovery in human stools after digestion of a fat‐rich meal labelled with [1,1,1‐13C3]tripalmitin and [1,1,1‐13C3]triolein

Lipid metabolism studies focus mainly on oxidation and storage but rarely on faecal elimination, which is needed to assess total lipid distribution during the postprandial period. The purpose of the present work was to set up and validate the analysis of lipid tracers in stools, with an aim of later using this methodology in studies of postprandial lipid tracer metabolism. Eight subjects received a mixture of [1,1,1-(13)C3]tripalmitin and [1,1,1-(13)C3]triolein with a fat-rich meal. The nature and amounts of (13)C lipids excreted in stools during 3 days post-dose were determined by gas chromatography/mass spectrometry (GC/MS) and gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) analysis of fatty acid methyl esters (FAMEs) from total fatty acid (TFA), free fatty acid (FFA) and triacylglycerol (TAG) fractions. The results were expressed as the Cumulative Tracer Recovery of the administered dose (CTR%). The quantities and labelling of FAMEs were higher in FFA than in TAG, indicating that label loss was not due to a lack of digestive lipase activity. The labelling was higher for C16:0 than for C18:1. The CTRs were 7.03 ± 0.77% and 6.87 ± 0.91%, respectively, in TFA and FFA for [1-(13)C] C16:0, while they were 0.60 ± 0.15% and 0.51 ± 0.11% for [1-(13)C] C18:1 (mean ± sem). By studying the kinetics of lipid excretion from subjects, two groups emerged. The first one showed rapid excretion in stool #1, whereas the second showed slower excretion in stools #2-#3. A significant difference was found in the FFA in stool #1 for C16:0 (p < 0.01) and C18:1 (p < 0.05). Individual excretion kinetics showed marked variability. Nevertheless, the CTR over the 3-day study period was substantial and homogenous for all subjects. These results confirm that the assessment of faecal elimination is of great importance when establishing total lipid distribution during the postprandial period and validate the analysis of cumulative tracer loss during 72 h post-tracer ingestion.

Validation of pentaacetylaldononitrile derivative for dual 2H gas chromatography/mass spectrometry and 13C gas chromatography/combustion/isotope ratio mass spectrometry analysis of glucose

A reference method to accurately define kinetics in response to the ingestion of glucose in terms of total, exogenous and endogenous glucose is to use stable-isotope-labelled compounds such as 2H and 13C glucose followed by gas chromatography/mass spectrometry (GC/MS) and gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) analysis. The use of the usual pentaacetyl (5Ac) derivative generates difficulties in obtaining accurate and reproducible results due to the two chromatographic peaks for the syn and anti isomers, and to the isotopic effect occurring during acetylation. Therefore, the pentaacetylaldononitrile derivative (Aldo) was validated for both isotopes, and compared with the 5Ac derivative. A correction factor including carbon atom dilution (stoichiometric equation) and the kinetic isotopic effect (KIE) was determined. Analytical validation results for the 2H GC/MS and 13C GC/C/IRMS measurements produced acceptable results with both derivatives. When 2H enrichments of plasma samples were < or = 1 mol % excess (MPE), the repeatability (RSD(Aldo Intra assay and Intra day) <0.94%, RSD(5Ac Intra assay and Intra day) <3.29%), accuracy (Aldo <3.4%, 5Ac <29.0%), and stability of the derivatized samples were significantly better when the Aldo derivatives of the plasma samples were used (p < 0.05). When the glucose kinetics were assessed in nine human subjects, after glucose ingestion, the plasma glucose 2H enrichments were identical with both derivatives, whereas the 13C enrichments needed a correction factor to fit together. Due to KIE variation, this correction factor was not constant and had to be calculated for each batch of analyses, to obtain satisfactory results. Mean quantities of exogenous glucose exhibit marked difference (20.9 +/- 1.3g (5Ac) vs. 26.7 +/- 2.5g (Aldo)) when calculated with stoichiometric correction, but fit perfectly when calculated after application of the correction factor (22.1 +/- 1.3g (5Ac) vs. 22.9 +/- 1.9g (Aldo)). Finally, the pentaacetylaldononitrile derivative, used here in GC/C/IRMS for the first time, enables measurement of 2H and 13C enrichments in plasma glucose with a single sample preparation.

Development of criteria for the detection of adrenosterone administration by gas chromatography‐mass spectrometry and gas chromatography‐combustion‐isotope ratio mass spectrometry for doping control

Adrenosterone (androst-4-ene-3,11,17-trione, 11-oxoandrostenedione) is an endogenous steroid hormone that has been promoted as a dietary supplement capable of reducing body fat and increasing muscle mass. It is proposed that adrenosterone may function as an inhibitor of the 11beta-hydroxysteroid dehydrogenase type 1 enzyme (11beta-HSD1), which is primarily responsible for reactivation of cortisol from cortisone. The urinary metabolism of adrenosterone was investigated, after a single oral administration in two male subjects, by gas chromatography-mass spectrometry (GC-MS) and gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). Substantially increased excretion of 11beta-hydroxyandrosterone, 11beta-hydroxyetiocholanolone, 11-oxoandrosterone and 11-oxoetiocholanolone was observed. Minor metabolites such as 3alpha,17beta-dihydroxy-5beta-androstan-11-one, 3alpha-hydroxyandrost-4-ene-11,17-dione and 3alpha,11beta-dihydroxyandrost-4-en-17-one were also identified. The exogenous origin of the most abundant adrenosterone metabolites was confirmed by GC-C-IRMS according to World Anti-Doping Agency criteria. Through analysis of a reference population data set obtained from urine samples provided by elite athlete volunteers (n = 85), GC-MS doping control screening criteria are proposed: 11beta-hydroxyandrosterone concentration greater than 10 000 ng/mL (specific gravity adjusted to 1.020) or 11beta-hydroxyandrosterone/11beta-hydroxyetiocholanolone ratio greater than 20.Urine samples fulfilling these screening criteria may be subjected to GC-C-IRMS analysis for confirmation of adrenosterone administration.

Determination of13C- and15N-enrichment of glutamine by gas chromatography/mass spectrometry and gas chromatography/combustion/isotope ratio mass spectrometry after N(O,S)-ethoxycarbonyl ethyl ester derivatisation

To measure 13C- and 15N-Gln enrichments in plasma samples by gas chromatography/mass spectrometry (GC/MS) and gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) we investigated the use of the N(O,S)-ethoxycarbonyl ethyl ester derivative. We found that this derivative is very stable at room temperature, even after 5 days storage. It allows reproducible and accurate 13C- and 15N-isotopic enrichment determinations with values of RSD below 0.8 and 3.2% for GC/MS and GC/C/IRMS, respectively. This derivative enables high sample throughput analyses when automated GC/MS and GC/C/IRMS are used. The methodology was applied to measure isotopic enrichments in rat plasma after oral force-feeding with [2,5-15N2]-Gln and in human plasma samples obtained after intravenous infusion of [1-13C]-Gln. In rats after oral force-feeding, we found that the maximal appearance of [2,5-15N2]-Gln occurred between 40 and 80 min in the two compartments studied (portal and peripheral blood). In humans infused intravenously with [1-13C]-Gln for 4 h, the plateau in 13C-Gln enrichments in plasma was reached after 3 h of infusion and coefficients of variation between samples were 1.7 and 12.4% for the two subjects. Using the N(O,S)-ethoxycarbonyl ethyl ester derivative, 13C- and 15N-isotopic enrichment can be determined in the range 2–100 ± 0.3 MPE by GC/MS and 0.3–2 ± 0.02 MPE by GC/C/IRMS. Copyright © 2001 John Wiley & Sons, Ltd.

Characterization of tetramethylammonium hydroxide thermochemolysis products of near-shore marine sediments using gas chromatography/mass spectrometry and gas chromatography/combustion/isotope ratio mass spectrometry

The study describes the application of gas chromatography/mass spectrometry (GC/MS) and gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) to characterize the molecular and compound-specific stable carbon isotope composition, respectively, of tetramethylammonium hydroxide (TMAH) thermochemolysis products of organic matter present in marine sediments. The objective of the study was to examine the usefulness of TMAH thermochemolysis products in identifying organic markers of terrestrial-derived sources in marine sediments. ‘Batch-wise’ thermochemolysis conditions (excess TMAH. 250 °C, sealed glass tube, 30 min) were used on sediments and standards. Compound-specific isotope analysis (CSIA) of standards showed no evidence of isotopic fractionation at methyl carbon during methylation of a number of phenolic acid and fatty acid compounds. It was also shown that no appreciable isotopic affect occurs during bond cleavage and methylation of fatty acid groups of the standard triglyceride, tricaprin. These preliminary tests indicate that the initial δ 13C composition of chemical markers containing acidic functional groups can be obtained by measuring the δ 13C of the corresponding methyl esters and/or ethers after TMAH methylation. In order to confirm the number of methoxy groups in lignin-derived markers, tetraethylammonium hydroxide was employed. Useful chemical markers found in near-shore sediments in reasonable abundance were a range of fatty acids, vanillin, vanillic acid and P-coumaric acid. Preliminary results of CSIA of these phenolic markers and selected saturated fatty acids in three different marine sediment cores are reported.

Isotope Ratio Mass Spectrometry, Compared with Conventional Mass Spectrometry in Kinetic Studies at Low and High Enrichment Levels: Application to Lipoprotein Kinetics

The aim of the present study was to compare the performances of gas chromatography/mass spectrometry (GC/MS) and gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) in stable isotope kinetic studies. In the analysis of cholesterol and leucine, GC/C/IRMS gave precise and linear results over a large scale of13C enrichment (−22 to +760 δ‰ for cholesterol, −26 to +600 δ‰ for leucine). Compared with GC/MS, GC/C/IRMS was much more accurate and reproducible, especially at low [13C]cholesterol enrichment (−12 δ‰), with cholesterol samples ranging from 0.11 to 17 ng. Cholesterol ester kinetics in rabbit plasma low-density lipoproteins was studied after injection of 3 mg [3,4-13C]cholesterol. A smooth and regular kinetic curve was obtained with GC/C/IRMS; results were much less reproducible with GC/MS. Finally, the performances of GC/C/IRMS were demonstrated in the simultaneous kinetic study of three human plasma apolipoproteins during a primed constant infusion of 0.7 mg·kg−1·h−1l-[1-13C]leucine. Kinetic curves were obtained in very-low-density lipoproteins and low-density lipoproteins for apolipoprotein B100, and in high-density lipoproteins for apolipoproteins AI and AII.