Elemental Analyzer/isotope Ratio Mass Spectrometry Research Articles

Simultaneous observation of 2H and 13C enrichment of methyl phosphonic acid via Orbitrap-IRMS with applications to nerve agent forensics

Quantification of the stable isotopes within a compound aids forensic investigations as it provides a fingerprint which can determine that compound's source substrates, synthetic route, and possible mechanisms of degradation. Previous stable isotope studies have explored 13C and 2H measurements of the sarin precursors methylphosphonic dichloride (DC) and methylphosphonic difluoride (DF) as forensic signatures. However, these measurements required different sample preparations and measurement techniques. Orbitrap isotope ratio mass spectrometry (Orbitrap-IRMS) is a developing technique which can characterize multiple stable isotopes simultaneously. Here, we apply Orbitrap-IRMS to simultaneously observe the 13C and 2H content of methylphosphonic acid (MPA), the hydrolysis product of DC and DF, which can be used as a proxy for the isotopic content of DC and DF. Our method requires 20 min analyses and consumes ≈60 nmol of sample, with precisions of ≈0.9 ‰ (13C) and ≈3.6 ‰ (2H). We apply our method to both commercially acquired MPA and MPA obtained from the hydrolysis of commercially acquired DC. We validate our methods via comparison to elemental-analyzer isotope ratio mass spectrometry (EA-IRMS). The combined 13C and 2H measurement creates a more robust forensic tool than either isotope individually. Our results demonstrate the viability of Orbitrap-IRMS for chemical forensic measurements.

Pottery use across the Neolithic transition in northern Belgium: evidence from isotopic, molecular and microscopic analysis

This study represents the first extensive residue analysis of prehistoric pottery from northern Belgium. It examines pottery use and culinary practices across the Mesolithic-Neolithic transition, from the late 6th to the early 4th millennium cal BC. Residue analyses were performed on more than 200 samples from nine archaeological sites, representing different cultural groups from this transitional phase. This includes the analysis of charred food residues encrusted on the vessel surfaces by elemental analysis-isotope ratio mass spectrometry (EA-IRMS), gas chromatography-mass spectrometry (GC-MS), stereomicroscopic analysis and Scanning Electron Microscopy (SEM), as well as the analysis of absorbed lipids by gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). This study provides the first evidence of ruminant dairy fats in Early Neolithic Limburg pottery, supporting the hypothesis that this pottery was produced and used by LBK farmers rather than hunter-gatherer populations. The first indigenous pottery of the Swifterbant culture was frequently used to process freshwater fish (often together with plant foods) and ruminant meat, although several of the studied vessels likely contained mixtures of resources which could also include porcine products. Ruminant dairy is nearly absent from this pottery. Similar results were obtained for pottery of the subsequent Michelsberg culture/Group of Spiere of the late 5th and early 4th millennium cal BC. The limited presence of ruminant dairy fats in this pottery contrasts with the findings for Middle Neolithic pottery from neighbouring regions, providing further evidence for the existence of regional variations in pottery use or culinary practices throughout prehistoric NW Europe. However, our current view of pottery use during the Mesolithic-Neolithic transition in northern Belgium might be biased by the difficulties in distinguishing between wild and domesticated ruminant adipose fats as well as in detecting plant foods through lipid residue analysis.

Sulfur isotope analyses using 3× elemental analysis/isotope ratio mass spectrometry: Saving helium and energy while reducing analytical time and costs.

Helium (He) and energy shortages have caused price increases and reduced their availability. Using three combustion reactions per acquisition of carbon and nitrogen isotope ratios saves 50% He and energy during the elemental analysis/isotope ratio mass spectrometry (EA/IRMS). This approach needs to be tested for sulfur isotope (δ34S) analyses. A new method to measure δ34S in three sequential combustion reactions within one EA/IRMS acquisition was developed. The same material or blank samples could be used in the three reactions. After SO2 was used, a N2 purging method was employed to prolong the lifetime of the valves in the EA/IRMS interface. The 3×EA/IRMS was applied to measure δ34S in precious samples, such as Ag2S from acid-volatile and chromium-reducible sulfur extracted with a multiple-port setup. The 3×EA/IRMS-δ34S method was validated with replicate analyses of international reference materials and laboratory standards with a wide range of mineralogical compositions and δ34S values. The method provided a strategic advantage for the δ34S measurements of small precious samples (measured between blanks). The accuracy and precision of the 3×EA/IRMS values effectively matched those obtained using conventional EA/IRMS, with good agreement between the mean ± SD values and the recommended values with their uncertainties. Compared with the conventional EA/IRMS, the proposed method provides accurate and precise δ34S measurements of the sulfate and sulfide samples while saving approximately 50% of He, energy, SO2 reference gas, O2, analysis time, and cost. Notably, 3×EA/IRMS can provide up to three δ34S values unaffected by memory effects.

Experimental assessment of elemental analyzer isotope ratio mass spectrometry normalization methodologies for environmental stable isotopes.

In stable isotope mass spectrometry, isotope delta values are normalized to internationally recognized reference scales using a combination of certified and in-house isotope reference materials. Numerous techniques exist for performing this normalization, but these methodologies need to be experimentally assessed to compare their effect on reproducibility of isotope results. We tested normalization methods by the number of reference materials used, their matrix, their isotope range, and whether normalization required extrapolating beyond the isotope range. We analyzed eight commercially available isotope reference materials on a ThermoFinnigan Delta-V isotope ratio mass spectrometer (IRMS) and an Elementar VisION IRMS for nitrogen and carbon isotope composition via solid combustion with an elemental analyzer and computed every possible isotope normalization (n = 612). Additionally, we assessed how sample matrix affected linearity effects on both instruments using five in-house reference materials. Normalizations exhibited the best performance when the reference materials spanning an isotope range of at least 20‰ were matrix matched with the samples and did not require extrapolation beyond the calibration curve. When these conditions were not met, the number of reference materials used had a significant effect on accuracy, with normalizations composed of two reference materials exhibiting particularly inconsistent performance at isotope ranges below 20‰. Linearity effects were found to exceed instrument precision by two orders of magnitude irrespective of matrix type and were not predicted by working gas diagnostics. Interlaboratory comparability of isotope results is improved when operators of elemental analyzer isotope ratio mass spectrometry (EAIRMS) systems select reference materials spanning an isotope range of at least 20‰. Additionally, using three or more isotopic reference materials, avoiding extrapolation beyond the range of the normalization curve, and matching the matrix of the reference materials to the samples improve normalizations.

Determining sugar and molasses origin by non‐exchangeable hydrogen stable isotope of ethanol and carbon isotope ratio mass spectrometry

AbstractThis study explores the differentiation of sugar and molasses produced from sugar beet and cane, which are susceptible to fraudulent labeling due to differing production costs. The research aimed to authenticate these products by botanical origin using novel analytical techniques. Utilizing ethanol isotopic measurement–isotope ratio mass spectrometry (IRMS) for non‐exchangeable hydrogen stable isotopes alongside carbon stable isotopes analysis through elemental analyzer–IRMS, the study accurately identified the origin of various sugar and molasses samples, pinpointed mislabeled goods, and determined the source of products with previously unknown provenance. These methods were also effective in revealing sugar and molasses adulteration and quantifying the extent of such fraud. The combined isotope analyses demonstrated their potential as robust tools for combating misrepresentation and adulteration in the sugar industry.

Influence of microbially fermented 2´-fucosyllactose on neuronal-like cell activity in an in vitro co-culture system.

2´-Fucosyllactose (2´-FL), the most abundant oligosaccharide in human milk, plays an important role in numerous biological functions, including improved learning. It is not clear, however, whether 2´-FL or a cleavage product could influence neuronal cell activity. Thus, we investigated the effects of 2´-FL, its monosaccharide fucose (Fuc), and microbial fermented 2´-FL and Fuc on the parameters of neuronal cell activity in an intestinal-neuronal transwell co-culture system in vitro. Native 13C-labeled 2´-FL and 13C-Fuc or their metabolites, fermented with Bifidobacterium (B.) longum ssp. infantis and B. breve, which were taken from the lag-, log- and stationary (stat-) growth phases of batch cultures, were applied to the apical compartment of the co-culture system with Caco-2 cells representing the intestinal layer and all-trans-retinoic acid-differentiated SH-SY5Y (SH-SY5YATRA) cells mimicking neuronal-like cells. After 3 h of incubation, the culture medium in the basal compartment was monitored for 13C enrichment by using elemental analysis isotope-ratio mass spectrometry (EA-IRMS) and effects on cell viability, plasma, and mitochondrial membrane potential. The neurotransmitter activation (BDNF, GABA, choline, and glutamate) of SH-SY5YATRA cells was also determined. Furthermore, these effects were also measured by the direct application of 13C-2´-FL and 13C-Fuc to SH-SY5YATRA cells. While no effects on neuronal-like cell activities were observed after intact 2´-FL or Fuc was incubated with SH-SY5YATRA cells, supernatants from the stat-growth phase of 2´-FL, fermented by B. longum ssp. infantis alone and together with B. breve, significantly induced BDNF release from SH-SY5YATRA cells. No such effects were found for 2´-FL, Fuc, or their fermentation products from B. breve. The BDNF release occurred from an enhanced vesicular release, which was confirmed by the use of the Ca2+-channel blocker verapamil. Concomitant with this event, 13C enrichment was also observed in the basal compartment when supernatants from the stat-growth phase of fermentation by B. longum ssp. infantis alone or together with B. breve were used. The results obtained in this study suggest that microbial products of 2´-FL rather than the oligosaccharide itself may influence neuronal cell activities.

Elemental (CNS) and isotopic (δ13C and δ15N) characterization of local sources to atmospheric dust at Trombay, Mumbai, using elemental analyzer–isotope ratio mass spectrometry

Particulate matter (PM) in the air has significant implications for human health and the environment, particularly in urban areas with high concentrations. This study focuses on the isotopic (δ13C and δ15N), elemental (CNS) as well as the total carbon to total nitrogen ratio (TC/TN) characterization of PM in Trombay, Mumbai. Monthly atmospheric dust samples were collected from 2016 to 2020 during the premonsoon and postmonsoon seasons. The average elemental concentrations for premonsoon samples were N (1.73% ± 0.25%), C (7.38% ± 0.72%), S (0.84% ± 0.13%), and TC/TN ratio (4.38 ± 0.86). For postmonsoon samples, the average elemental concentrations were N (1.47% ± 0.185%), C (6.95% ± 1.04%), S (0.84% ± 0.13%), and TC/TN ratio (4.80 ± 0.94). The average δ13C values for premonsoon and postmonsoon seasons were −24.31 ± 1.15‰ and −24.56 ± 1.20‰, respectively. The average δ15N values for premonsoon and postmonsoon seasons were 5.22 ± 1.92‰ and 4.57 ± 1.32‰, respectively. The results showed variations in elemental concentrations between the seasons, with higher values observed in premonsoon samples which can be attributed to increased dust generation from land and sea. Overall, this study provides insights into the local sources and characteristics of PM in the Trombay area and highlights the importance of long-term monitoring to understand the seasonal variations in PM composition.

Using Liquid Chromatography-Isotope Ratio Mass Spectrometry for Detection of Economically Motivated Adulteration of Maple Syrup.

Maple syrup is a sought-after commodity, and used as a condiment and a sweetener. Also, it is an active target of economically motivated adulteration (EMA), similar to other foods such as lemon juice and honey. This study is aimed to detect low cost sugar adulteration in maple syrup via an internal standard method using malic acid through solid-phase extraction (SPE) and LC with isotope ratio mass spectrometric detection (LC-IRMS). In this work, an optimized SPE sample preparation procedure was used for the isolation of organic acids from maple syrup. Using LC-IRMS, malic acid was separated from other organic acids and the δ13C value of malic acid was determined. Eleven maple syrup samples, domestic or imported from Canada, were evaluated for 13C/12C ratios (δ13C values) using combustion module-cavity ring down spectrometry (CM-CRDS) and compared to the δ13C values obtained from well-established elemental analyzer-isotope ratio mass spectrometry (EA-IRMS) methods. The δ13C values of isolated malic acid analyzed by SPE-LC-IRMS were used as internal standards and compared to the δ13C values of bulk maple syrup; difference (δ13Csugars - δ13Cmalic acid) values greater than 3.6‰ are indicative of low-cost sugar adulteration. Overall, the results obtained from SPE-LC-IRMS provided a faster, novel analysis approach for determining low-cost sugar adulteration in maple syrup for regulatory purposes. This method also provided lower detectable limits of adulteration versus current literature reports using bulk analysis and comparable detection limits to Tremblay and co-workers who utilized an internal standard method. SPE-LC-IRMS is a robust method that can be used for detecting adulteration in maple syrup samples for regulatory purposes. SPE-LC-IRMS is a faster, novel analysis approach for determining C4 adulteration in maple syrup with lower detection limits.

Using stable carbon isotope ratio analysis to detect adulteration in red yeast rice dietary supplements

Red yeast rice (RYR) is marketed as a dietary supplement because it contains natural 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins), including monacolin K. However, there is concern that some RYR supplements may be adulterated with the pharmaceutical drug lovastatin to enhance health claims. We have developed an optimized method to isolate monacolin K/lovastatin from complex RYR dietary supplement matrices to then test for adulteration in RYR supplements using stable carbon isotope (δ13C) analysis. Samples were initially screened for monacolin K/lovastatin using liquid chromatography with mass spectrometric detection (LC-MS). To ensure the extraction process did not affect the measured isotopic values (i.e., isotopic fractionation effects), neat lovastatin standards were spiked into two types of blank RYR matrices (powder and gel). The monacolin K/lovastatin peaks were detected using high performance liquid chromatography with ultraviolet detection (HPLC-UV) and isolated using fraction collection. Residual matrix components were removed from targeted fractions by solid phase extraction (SPE) using graphitized carbon black cartridges. The resulting isolates were then analyzed using elemental analyzer-isotope ratio mass spectrometry (EA-IRMS) to measure δ13C values. The δ13C values of the extracted lovastatin standards were compared to their respective neat lovastatin δ13C values and demonstrated negligible isotopic fractionation effects. Using this optimized clean up method and carbon isotope analysis, thirty-one samples were screened. Eight RYR dietary supplement samples had >0.8 mg/g of monacolin K/lovastatin, our minimum threshold for analyzing samples using this method. Four of these eight samples had δ13C values greater than −28.3‰, a previously proposed cutoff value for natural monacolin K, indicating likely adulteration. Additionally, five RYR powder samples were analyzed as part of a collaborative study using in-house methods from two laboratories and the data shows acceptable agreement in the δ13C values of monacolin K/lovastatin (differences ranging from ±0.02‰ to ±0.76‰). This optimized method represents a robust, reproducible procedure for detecting lovastatin adulteration in dietary supplements with minimal isotopic fractionation.

Application of a porous graphitic carbon column to carbon and nitrogen isotope analysis of underivatized individual amino acids using high-performance liquid chromatography coupled with elemental analyzer/isotope ratio mass spectrometry.

Isolation of underivatized amino acids (AAs) using high-performance liquid chromatography (HPLC) is becoming a popular method for carbon (δ13 C) and nitrogen isotope (δ15 N) analyses of AAs because of the high analytical precision and for performing dual-isotope analysis. However, some AAs in natural samples, especially small, hydrophilic AAs, are not suitably separated using reversed-phase columns (e.g., C18) and ion-exchange columns (e.g., Primesep A). We developed a new method for HPLC using a porous graphitic carbon column for the separation of nine hydrophilic AAs. After purification, δ13 C and δ15 N values of AAs were determined using elemental analyzer/isotope ratio mass spectrometry (EA/IRMS). We demonstrated the application of this method by determining δ13 C and δ15 N values of individual hydrophilic AAs in a biological sample, the muscle of blue mackerel (Scomber australasicus). Chromatographically, the baseline separation of hydrophilic AAs was achieved in both the standard mixture and the biological sample. We confirmed that δ13 C and δ15 N values of AA standards remained unchanged during the whole experimental procedure. The δ13 C values of AAs in mackerel muscle are also in good agreement with the values obtained using another verified method for δ13 C analysis. The good separation performance of hydrophilic AAs and the reliability of δ13 C and δ15 N analyses of individual AAs using the porous graphite column offer a significant advantage over conventional settings. We suggest that, in the future, the HPLC × EA/IRMS method can be used for reliable δ13 C and δ15 N analyses of AAs in natural samples.

Bias Estimation in the Certification of Steroid Reference Materials for Carbon Isotope Delta Measurements via EA- and GC-C-IRMS.

Two new CRMs have been prepared providing three steroids certified for stable carbon isotope delta values, δ(13 C) ‰. These materials have been designed to assist anti-doping laboratories validate their calibration method or employed as calibrant for stable carbon isotope measurements of Boldenone, Boldenone Metabolite 1 and Formestane. These CRMs will allow for accurate and traceable analysis in compliance with WADA Technical Document TD2021IRMS. Certification was performed using an EA-IRMS primary reference method on the bulk carbon isotope ratios of nominally pure steroid starting materials. EA-IRMS analyses were carried out on a Flash EA Isolink CN coupled via a Conflo IV to a Delta V plus Mass Spectrometer. Confirmation analysis was performed by GC-C-IRMS using a Trace 1310 GC coupled to a Delta V plus Mass Spectrometer via GC Isolink II. Based on the EA-IRMS analysis the materials were certified with δ(13 C) values of -30.38 ‰ (Boldenone), -29.71 ‰ (Boldenone Metabolite 1) and 30.71 ‰ (Formestane). Noting that the assumption of 100% purity in the starting materials has the potential to introduce bias this was investigated by GC-C-IRMS analysis and theoretical modelling based on purity assessment data. Careful application of this theoretical model was shown to provide reasonable estimates of uncertainty while avoiding the introduction of errors associated with analyte specific fractionation during GC-C-IRMS analysis.

Evaluation of honey authenticity in Thailand by analysis of carbon stable isotope ratio using elemental analyser coupled to isotope ratio mass spectrometry and cavity ring‐down spectrometry

SummaryIn Thailand, honey is used in a variety of commercial products that are distributed in domestic markets and abroad. As honey in Thailand comes from many locations, it is challenging to control the production quality of commercial products. The main problem commonly found in the honey industry is adulteration with undeclared added sugars. Identification of adulterated honey is crucial for authentication, which can increase consumer confidence on honey purity. In this study, 49 honey samples from beekeepers (natural) and 15 commercial honey producers were collected from different cities across Thailand. They were analysed for stable carbon isotope ratios (expressed as 13C/12C or δ13C and reported in relation to the international reference standard unit Vienna Pee Dee Belemnite, ‰ VPDB) using Elemental Analysis Isotope Ratio Mass Spectrometry (EA‐IRMS) for developing a δ13C database of natural honey and for checking the adulteration of honey. The δ13C values detected in 49 natural honey samples varied from −23.1‰ to −29.1‰ VPDB with an average of −26.8 ± 1.2‰ VPDB. According to the AOAC standard (No. 998.12, 2019), natural honey should have δ13C values lower than −23.5‰ VPDB, with an acceptable difference in δ13C values between honey and the protein extracted from honey up to 1‰. Compared to the standard guideline, 60% of commercial honey samples collected for this study (9 out of 15 samples) were found to contain undeclared added sugars.

Identification of biodegradable plastics using differential scanning calorimetry and carbon composition with chemometrics

Biodegradable plastics have become an important commodity to replace non-degradable plastics and alleviate environment pollution. It is important to identify instances of counterfeit and mislabeled biodegradable plastics in the marketplace to avoid sustainability fraud and improve plastics waste recycling. In this study, differential scanning calorimetry (DSC) and elemental analyzer-isotope ratio mass spectrometry (EA-IRMS) were used to analyze biodegradable plastics (polylactic acid (PLA), poly butylene adipate-co-terephthalate (PBAT), PBAT/PLA blend (PBAT+PLA)) and non-degradable plastics (polyethylene terephthalate (PET), high density polyethylene (HDPE), low density polyethylene (LDPE), polypropylene (PP)). A verification model was built using orthogonal partial least squares-discriminant analysis (OPLS-DA). Results showed that the melting point of biodegradable and non-degradable plastics ranged from 120 °C to 176 °C and 105 °C to 253 °C, respectively. The melting points of biodegradable PLA and non-degradable PP overlapped. The δ13C values of biodegradable and non-degradable plastics ranged from -25.8 ‰ to -11.6 ‰ and -32.3 ‰ to -25.4 ‰, respectively. The % C of biodegradable and non-degradable plastics ranged from 36.0% to 61.0% and 56.4% to 85.3%, respectively. Three OPLS-DA models were established by combining DSC variables, δ13C values and % C. The PLA discriminant accuracy of these models was 100%, suggesting this approach holds promise for the regulation and identification of biodegradable plastics in the marketplace.

Dynamics of the heavy metals content (Zn, Cd, Pb, Cu) during the decomposition of sphagnum moss at the drained and post-pyrogenic sites of oligotrophic bogs

The aim of the study. Assessment of heavy metals (Zn, Cd, Pb, Cu) dynamic during the decomposition of Sphagnum fuscum L. in native, drained and post-pyrogenic oligotrophic peatlands. Location and time of the study. The study was carried out in 2019–2021 in two oligotrophic bogs: “Bakcharskoye” (field station “Vasyuganye” (IMCES SB RAS)) and “Iksinskoye”, which are the north-eastern spurs of the Great Vasyugan mire and located in the Bakcharsky district of the Tomsk region. Methods. Decomposition rate of Sph. fuscum L. was determined by the method of partially isolated samples, which is widely used to study the processes of transformation of plant material and peat. In the original and decomposed samples of sphagnum moss and peat soil (in layers 0-10, 10-20, 20-30), the ash content of peat was determined according to GOST-11306-83. The percentage of total carbon and nitrogen in samples of peat (up to 30 cm from the surface) and plant residues was determined together with the determination of isotopic composition using EA-IRMS (Elemental Analyzer/Isotope Ratio Mass Spectrometry). The concentrations of heavy metals (Zn, Cd, Pb, Cu) in samples of peat and plant residues were determined by stripping voltammetry. Results. Three-year experiment on decomposition of Sph. fuscum in four ecosystems (natural – VASnat, drained – VASdry, post-pyrogenic – Iksa 1 and Iksa2) showed that fire affected the decomposition rate: on average, mass loss in post-pyrogenic bogs was 1.4–1.6 times lower compared to the natural site. For almost all sites, a relationship was found between the mass loss and the concentration of heavy metals, with the exception of Cd during the first year of the experiment. The concentration of all elements in moss residues decreased during the decomposition process at all study sites. During moss decomposition the similarity in the release of biogenic elements Cu and Zn was recorded in the drained and post-pyrogenic sites (VASdry, Iksa1, Iksa2), while for Cd and Pb, which are not biogenic elements, such similarity was not revealed. Conclusions. Peat fires affect plant residue decomposition by decreasing its rate of decomposition. The study revealed incremental dynamics of HMs content in plant residues: at the initial stage, HMs are released, whereas during the second and third years, in some cases, the accumulation of HMs is observed. However, by the end of the experiment, the release of heavy metals during decomposition was observed for all elements.

Stable Isotope Analysis of Residual Pesticides via High Performance Liquid Chromatography and Elemental Analyzer-Isotope Ratio Mass Spectrometry.

To broaden the range of measurable pesticides for stable isotope analysis (SIA), we tested whether SIA of the anthranilic diamides cyantraniliprole (CYN) and chlorantraniliprole (CHL) can be achieved under elemental analyzer/isotope ratio mass spectrometry with compound purification in high-performance liquid chromatography (HPLC). Using this method, carbon isotope compositions were measured in pesticide residues extracted from plants (lettuce) grown indoors in potting soil that were treated with 500 mg/kg CHL and 250 mg/kg CYN and were followed up for 45 days. Our results show that the CYN and CHL standard materials did not have significant isotope differences before and after clean-up processing in HPLC. Further, when applied to the CYN product and CHL product in soil, stable isotope differences between the soil and plant were observed at <1.0‱ throughout the incubation period. There was a slight increase in the variability of pesticide isotope ratio detected with longer-term incubation (CHL, on average 1.5‱). Overall, we measured the carbon isotope ratio of target pesticides from HPLC fraction as the purification and pre-concentration step for environmental and biological samples. Such negligible isotopic differences in pesticide residues in soils and plants 45 days after application confirmed the potential of CSIA to quantify pesticide behavior in environments.

Rapid discrimination of the authenticity and geographical origin of bear bile powder using stable isotope ratio and elemental analysis.

Bear bile powder (BBP) is one of the most famous traditional Chinese medicines derived from animals. It has a long history of medicinal use and is widely used in the treatment of hepatobiliary and ophthalmic diseases. Due to its similar morphological characterizations and chemical composition compared with other bile powders, it is difficult to accurately identify its authenticity. In addition, there are very few methods that could analyze the geographical origins of BBP. In this study, elemental analysis isotope ratio mass spectrometry (EA-IRMS) and inductively coupled plasma mass spectrometry (ICP-MS) were used to determine stable isotope ratios and elemental contents, respectively. Combined these variables with chemometrics, the discrimination models were established successfully for identifying the authenticity and geographical origins of BBP. Meanwhile, the discrimination markers were identified by calculating the variable importance for the projection (VIP) value of each variable. A total of 13 discrimination markers (δ13C, δ15N, C, Li, Mg, K, Ca, Cr, Ni, Zn, As, Se, and Sr) were used to further establish the fingerprint of BBP. According to similarity analysis, the authenticity and geographical origins of BBP could be identified without chemometrics. In conclusion, the present study established a reliable method for authenticity identification and origin traceability of BBP, which will provide references for the quality control of bile medicines.

Application of stable isotopes with machine learning techniques for identifying Aconiti Lateralis Radix Praeparata (Fuzi) geographical origins

Authentication of geographical traceability is essential to the safety of traditional Chinese medicines (TCMs). Aconiti Lateralis Radix Praeparata (Chinese: Fuzi), a well-known TCMs, the therapeutic efficacy and toxicity are clearly correlated with its geographical origin, and a rapid and effective method of origin tracing must be established. Herein, an elemental analysis-isotope ratio mass spectrometry (EA-IRMS) combined with a machine learning model is constructed for the origin traceability of Fuzi. We analyzed stable isotope information (δ13C‰, δ15N‰, δ2H‰, δ18O‰, C% and N%) for a total of 243 samples from eight major producing areas. Classification models were built using seven machine learning techniques, including random forest (RF), support vector machine (SVM), AdaBoost, etc., to investigate the feasibility of stable isotopes in Fuzi geographic identification. Results show that stable isotopes exhibit an overwhelming origin discrimination advantage. Among them, the RF model showed the best recognition performance with 94.2 % accuracy, 95.45 % sensitivity, and 99.15 % specificity in the independent test set, respectively. The results show that stable isotopes combined with machine learning techniques have great potential for the origin traceability of TCMs.

Stable carbon isotopic characterization of rice vinegar protein as an intrinsic reference for discriminating the authenticity of brewed rice vinegar.

Rice vinegar plays an important role in daily life. However, some unscrupulous manufacturers may deliberately add synthetic acetic acid in vinegar products to reduce fermentation time and save production costs. To protect the rights and health of consumers, vinegar authenticity must be controlled. The rice vinegar protein was used as an intrinsic reference and its stable carbon isotope ratio (δ13Cprotein) was analyzed by elemental analyzer-isotope ratio mass spectrometry. The stable carbon isotope ratio difference between the acetic acid and the rice vinegar protein (Δδ13Cacetic acid-protein) was calculated to evaluate vinegar authenticity. Sixteen rice vinegar samples were analyzed and a stable carbon isotopic pattern of rice vinegar was established by the 95% confidence interval for Δδ13Cacetic acid-protein (0.27‰-2.10‰). An acetic acid adulteration curve of Δδ13Cacetic acid-protein was also assumed according to the data from rice vinegar samples, and its validity was confirmed by rice vinegar deliberately blended with acetic acid at different ratios (25, 50, and 75%). The Δδ13Cacetic acid-protein values of the adulterated vinegars decreased with increasing amounts blended acetic acid, but the δ13Cprotein values did not, showing that rice vinegar protein could be used as an intrinsic reference for identifying the adulterated rice vinegar. The rice vinegar adulterated with acetic acid at higher than approximately 10% could be detected.

New method for simultaneous determination of dissolved organic carbon and its stable carbon isotope ratio in liquid samples: environmental applications

ABSTRACT This study reports the development of an all-in-one elemental analyser isotope ratio mass spectrometry (EA-IRMS) system modified for simultaneous analysis of dissolved organic carbon (DOC) concentration and its stable carbon isotope footprint (δ 13CDOC) in aqueous samples. The method involves a quantitative oxidation of DOC in a 200 µL liquid sample to CO2, after sample acidification and stripping by nitrogen. The detection limit of the method for DOC quantification was 0.2 mg C/L with an analytical precision of 12 %. Uncertainty of stable isotope determinations was 2 % at 0.2 mg DOC/L, while decreasing to 0.3 % at 20 mg DOC/L. Quantitative oxidation of DOC in aqueous samples was validated by using ring test water samples and Deep Sea reference seawater. The method performances of isotope analysis were evaluated by analysing different isotopic standard solutions. The applicability of the method was tested through the analysis of different environmental types of water, showing that δ 13CDOC ranged from – 23.30 to –31.85 ‰, allowing to characterize samples of different environmental origin. The developed method offers several advantages including rapidity, use of small sample volumes and minimal sample pre-treatment, making it a valuable tool for routine DOC concentration measurements paired with isotopic characterization.

A Case Study to Establish a Basis for Evaluating Geographic Origin Claims of Timber From the Solomon Islands Using Stable Isotope Ratio Analysis

Global demand for low-cost forest products is leading manufacturers and traders to source timber and wood products from vulnerable nations and delicate ecosystems. One small island nation, the Solomon Islands, is seeing exploitation of natural resources accelerating to such a point that its natural forests may be exhausted by 2036. The main causes of natural forest loss on the archipelago are unsustainable or illegal logging practices. Various laws in consumer countries require that members of industry ensure that only legally sourced timber is placed onto their respective national markets. Those that break these laws or fail to act in a way that is compliant may be subject to harsh penalties. This study aims to establish scientific data to evaluate claims that timber has originated from the Solomon Islands. This will enable Operators to carry out due diligence analysis and permit members of Law Enforcement to conduct forensic investigations. Eighty timber core samples comprising 13 different genera of tropical trees were obtained from mature trees in two sites in the Solomon Islands (Guadalcanal and Kolombangara islands) during the period August 2019 to November 2019 using a Pickering Punch sampling device. Homogenised core samples were subject to δ18O, δ2H, δ13C, and δ34S stable isotope analysis using elemental analysis-isotope ratio mass spectrometry. Additional stable isotope data from relevant taxa and geographic origins (elevation, geographic co-ordinates) were also included in this research as an initial assessment of differences in stable isotope ratios between countries. Results show that significant differences are evident in the stable isotope ratios of the sampled taxa within the Solomon Islands (Guadalcanal and Kolombangara Islands) and between other countries. These data can be used as a basis of evaluation to evaluate origin claims of timber or wood products from the Solomon Islands, particularly Kolombangara Island. Furthermore, in the right context, these data can also be used to establish whether timber or wood products declared to be from origins other than the Solomon Islands have stable isotope ratios that are consistent with data from the Solomon Islands. If not, this would suggest foreign timber/forest products are from elsewhere and are being passed-off as originating from the Solomon Islands.