Isotope Dilution Research Articles

A potential primary method for peptide purity analysis by gas chromatography-isotope dilution infrared spectrometry.

Here we proposed a method for peptide purity analysis using gas chromatography-isotope dilution infrared spectroscopy. The principle and feasibility of the proposed measurement method were investigated. The derivatization, separation, and infrared detection conditions for amino acids were optimized, and the performance of the method was investigated. Then, the proposed method was used for assessment of [Glu1]-fibrinopeptide B purity, and the results were compared with those obtained by high performance liquid chromatography-isotope dilution mass spectrometry. The average purity of six sub-samples using the proposed method was (0.755 ± 0.017)g/g, which agreed well with that obtained by isotope dilution mass spectrometry (0.754 ± 0.012) g/g. The repeatability of the proposed method was 2.2%, which was similar to that of isotope dilution mass spectrometry (1.7%). The proposed method has a similar principle and had similar accuracy, precision, and linearity to isotope dilution mass spectrometry; however, the developed method had higher limit of detection (LOD) and limit of quantitation (LOQ) values because of the low sensitivity of infrared detection. The results were also Système International d'Unités (SI) traceable. The developed method has the advantage of lower cost compared with isotope dilution mass spectrometry because only one isotope-labeled atom in an analog is required, and several infrared spectra can be extracted, averaged, and used for an amino acid calculation during one run, potentially leading to higher accuracy. This method could be easily expanded to the accurate quantitation of other organic compounds, including proteins. It is expected that the proposed method will be widely used in chemical and biological measurements as a new primary method.

Development of a potential primary method for protein quantification via electrospray differential mobility analysis

Accurate protein quantification is the basis for establishing the metrological traceability of in vitro diagnostics or drug products. In this study, we established and validated a potential primary method for protein quantification based on electrospray-differential mobility analysis coupled with a condensation particle counter (ES-DMA-CPC). The analytical performance of this method was assessed using the certified reference material NIMCmAb, and the uncertainty of measurement was evaluated. The method was applied to the quantification of three other protein reference materials and one highly purified protein, including myoglobin, bovine serum albumin, IgG monoclonal antibody, and one highly purified fibrinogen, with a molecular weight range between 17 kDa and 340 kDa. In addition, when compared with isotope dilution mass spectrometry (IDMS) and UV‒VIS spectrophotometry approaches, the ES-DMA-CPC method showed good agreement with IDMS method for the quantification of these protein reference materials. Our proposed method provided an accurate quantification of proteins, especially those with large molecular weights. Moreover, our method could be a potential primary method for protein quantification and serve as a complement to IDMS method.

Associations of Maternal Urinary Concentrations of Phenols, Individually and as a Mixture, with Serum Biomarkers of Thyroid Function and Autoimmunity: Results from the EARTH Study.

The associations between urinary phenol concentrations and markers of thyroid function and autoimmunity among potentially susceptible subgroups, such as subfertile women, have been understudied, especially when considering chemical mixtures. We evaluated cross-sectional associations of urinary phenol concentrations, individually and as a mixture, with serum markers of thyroid function and autoimmunity. We included 339 women attending a fertility center who provided one spot urine and one blood sample at enrollment (2009-2015). We quantified four phenols in urine using isotope dilution high-performance liquid chromatography-tandem mass spectrometry, and biomarkers of thyroid function (thyroid-stimulating hormone (TSH), free and total thyroxine (fT4, TT4), and triiodothyronine (fT3, TT3)), and autoimmunity (thyroid peroxidase (TPO) and thyroglobulin (Tg) antibodies (Ab)) in serum using electrochemoluminescence assays. We fit linear and additive models to investigate the association between urinary phenols-both individually and as a mixture-and serum thyroid function and autoimmunity, adjusted for confounders. As a sensitivity analysis, we also applied Bayesian Kernel Machine Regression (BKMR) to investigate non-linear and non-additive interactions. Urinary bisphenol A was associated with thyroid function, in particular, fT3 (mean difference for a 1 log unit increase in concentration: -0.088; 95% CI [-0.151, -0.025]) and TT3 (-0.066; 95% CI [-0.112, -0.020]). Urinary methylparaben and triclosan were also associated with several thyroid hormones. The overall mixture was negatively associated with serum fT3 concentrations (mean difference comparing all four mixture components at their 75th vs. 25th percentiles: -0.19, 95% CI [-0.35, -0.03]). We found no evidence of non-linearity or interactions. These results add to the current literature on phenol exposures and thyroid function in women, suggesting that some phenols may alter the thyroid system.

Determination of 1,3-Diphenylguanidine, 1,3-Di-o-tolylguanidine, and 1,2,3-Triphenylguanidine in Human Urine Using Liquid Chromatography-Tandem Mass Spectrometry.

1,3-Diphenylguanidine (DPG), 1,3-di-o-tolylguanidine (DTG), and 1,2,3-triphenylguanidine (TPG) are rubber additives widely present in the indoor environment. Nevertheless, little is known about their human exposure. We developed a method for the quantification of DPG, DTG, and TPG in human urine, using high-performance liquid chromatography-tandem mass spectrometry. The quantitative analysis of target analytes at parts-per-trillion levels in urine was optimized using hydrophilic-lipophilic balanced solid-phase extraction and isotopic dilution. The method limits of detection and quantification were in the range of 0.002-0.02 and 0.005-0.05 ng/mL, respectively. The recoveries of all analytes in human urine fortified at 1, 5, 10, and 20 ng/mL concentrations were in the range of 75.3-111%, with standard deviations of 0.7-4%. The repeated measurement of similarly fortified human urine yielded intra-day and inter-day variations of 0.47-3.90 and 0.66-3.76%, respectively. The validated method was applied in the measurement of DPG, DTG, and TPG in real human urine samples, which revealed the occurrence of DPG in children's urine samples (n = 15) with a detection frequency of 73% and at a median concentration of 0.05 ng/mL. DPG was found in 20% of adults' urine samples (n = 20).

Association of plasma aflatoxin with persistent detection of oncogenic human papillomaviruses in cervical samples from Kenyan women enrolled in a longitudinal study

BackgroundCervical cancer is caused by oncogenic human papillomaviruses (HR-HPV) and is common among Kenyan women. Identification of factors that increase HR-HPV persistence is critically important. Kenyan women exposed to aflatoxin have an increased risk of HR-HPV detection in cervical specimens. This analysis was performed to examine associations between aflatoxin and HR-HPV persistence.MethodsKenyan women were enrolled in a prospective study. The analytical cohort for this analysis included 67 HIV-uninfected women (mean age 34 years) who completed at least two of three annual study visits and had an available blood sample. Plasma aflatoxin was detected using ultra-high pressure liquid chromatography (UHPLC)-isotope dilution mass spectrometry. Annual cervical swabs were tested for HPV (Roche Linear Array). Ordinal logistic regression models were fitted to examine associations of aflatoxin and HPV persistence.ResultsAflatoxin was detected in 59.7% of women and was associated with higher risk of persistent detection of any HPV type (OR = 3.03, 95%CI = 1.08–8.55, P = 0.036), HR-HPV types (OR = 3.63, 95%CI = 1.30-10.13, P = 0.014), and HR-HPV types not included in the 9-valent HPV vaccine (OR = 4.46, 95%CI = 1.13–17.58, P = 0.032).ConclusionsAflatoxin detection was associated with increased risk of HR-HPV persistence in Kenyan women. Further studies, including mechanistic studies are needed to determine if aflatoxin synergistically interacts with HR-HPV to increase cervical cancer risk.

Plasma and cerebrospinal fluid nonenzymatic protein damage is sustained in Alzheimer's disease

BackgroundOxidative stress is considered to play an important role in the pathogenesis of Alzheimer's disease (AD). It has been observed that oxidative damage to specific protein targets affecting particular functional networks is one of the mechanisms by which oxidative stress contributes to neuronal failure and consequently loss of cognition and AD progression. Studies are lacking in which oxidative damage is measured at both systemic and central fluid levels and in the same cohort of patients. We aimed to determine the levels of both plasma and cerebrospinal fluid (CSF) nonenzymatic protein damage in patients in the continuum of AD and to evaluate the relation of this damage with clinical progression from mild cognitive impairment (MCI) to AD. MethodsDifferent markers of nonenzymatic post-translational protein modification, mostly from oxidative processes, were detected and quantified in plasma and CSF by isotope dilution gas chromatography‒mass spectrometry using selected ion monitoring (SIM-GC/MS) for 289 subjects: 103 AD, 92 MCI, and 94 control subjects. Characteristics of the study population such as age, sex, Mini-mental state examination, CSF AD biomarkers, and APOE ϵ4, were also considered. ResultsForty-seven (52.8%) MCI patients progressed to AD during follow-up (58 ± 12.5 months). After controlling for age, sex, and APOE ϵ4 allele, plasma and CSF concentrations of protein damage markers were not associated with either diagnosis of AD or MCI. The CSF levels of nonenzymatic protein damage markers were associated with none of the CSF AD biomarkers. In addition, neither in CSF nor in plasma were the levels of protein damage associated with the MCI to AD progression. ConclusionThe lack of association between both CSF and plasma concentrations of nonenzymatic protein damage markers and AD diagnosis and progression suggests that oxidative damage in AD is a pathogenic mechanism specifically expressed at the cell-tissue level, not in extracellular fluids.

Polychlorinated dibenzo-p-dioxins, dibenzofurans and dioxin-like polychlorinated biphenyls in bivalve molluscs. Risk to Polish consumers?

Concentrations of polychlorinated dibenzo-p-dioxins and polychlorinated dibenzofurans (PCDD/Fs) and dioxin-like polychlorinated biphenyls (DL-PCBs) were investigated in six species of bivalve mollusc purchased on the Polish retail market. The risk to consumers was calculated as a percentage of the tolerable weekly intake (TWI) (2 pg World Health Organization toxic equivalent (WHO-TEQ) kg-1 body weight (b.w.)). Altogether 32 samples were analysed using an isotope dilution technique with high resolution gas chromatography coupled with high resolution mass spectrometry. Low levels of all analysed compounds were found. The range of PCDD/Fs was 0.08-0.37 pg WHO-TEQ g-1 of wet weight (w.w.) and 0.04-0.41 pg WHO-TEQ g-1 w.w. for DL-PCBs. The highest concentrations of all analysed compounds were found in Pacific oysters, at 0.30 pg WHO-TEQ g-1 w.w. for the sum of PCDD/Fs and 0.19 pg WHO-TEQ g-1 for the sum of DL-PCBs. These concentrations were 2-4 times higher than those detected in the other analysed mollusc species. Different species-dependent congener profiles were observed for PCDD/F concentrations, while PCB congener concentration profiles were species independent. The risk to consumers was assessed relating theoretical intakes of PCDD/Fs and DL-PCBs per 25, 50 and 100 g of consumption of molluscs per week to the TWI. Taking into account the low consumption of molluscs in Poland and low concentrations of analysed compounds, neither adults nor children are likely to exceed the TWI by ingestion of food in this category.

Determination of four aflatoxins in feeds by high throughput automated immunoaffinity magnetic beads purification-ultra performance liquid chromatography

Aflatoxin (AFT) is an extremely toxic and highly toxic carcinogenic substance. This is particularly problematic due to the risk of aflatoxin contamination in raw feed materials and products during production, transportation, and storage. In this study, immunoaffinity magnetic beads (IMBs) were prepared for the purification of four aflatoxins (aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1) and aflatoxin G2 (AFG2)). The aflatoxin contents were then determined rapidly and accurately using ultra performance liquid chromatography (UPLC). More specifically, the coupling ratio of magnetic beads (MBs) to the aflatoxin monoclonal antibody was initially optimized, wherein an MB volume of 1 mL and an antibody content of 2.0 mg was found to meet the purification requirements of this method. The magnetic properties of the MBs and the IMBs were then investigated using a vibrating sample magnetometer (VSM) at room temperature. As a result, the maximum saturation super magnetizations of the MBs and the IMBs were determined to be 28.61 and 23.22 emu/g, respectively, indicating that the saturation magnetization intensity of the IMBs was reduced by coupling with a non-magnetic antibody. However, the saturation magnetization intensity remained sufficiently high to permit magnetic separation from the solution. In addition, the appearance of the IMBs was examined using a biomicroscope, and it was clear that the magnetic cores were wrapped in agarose gel. Furthermore, the reaction time between the IMBs and the aflatoxins was investigated, and the optimal reaction time for meeting the purification requirements was determined to be 2 min. The stability of the IMBs was then evaluated under refrigerated storage conditions at 4 ℃. It was found that the prepared IMBs maintained a high aflatoxin enrichment capacity for at least eight months. Through the examination of three different extraction solutions, a mixture of acetonitrile and water (70∶30, v/v) was found to be optimal for the extraction of aflatoxins from the feed samples. Moreover, five sample dilutions and purification effects were also examined, and phosphate-buffered saline (containing 0.5% Tween-20) was selected as the preferred sample dilutant. With the optimized conditions, the effectiveness of using IMB for the purification of different feed samples was investigated. The resulting UPLC chromatogram showed no spurious peaks close to the target peaks, demonstrating a good purification performance. Following matrix spiking (5, 20, and 40 μg/kg, calculated based on AFB1) of the four feed samples (i. e., soybean meal, distillers dried grains with solubles, pig feed, and chicken feed), the spiked recoveries of the four aflatoxins ranged from 91.1% to 119.4% with a relative standard deviation (RSD) of <6.9%. In addition, the inter-day precision was 4.5% to 7.5%, and the method exhibited a good reproducibility. Subsequently, the developed method was used to detect AFB1 using reference materials. The test value was 18.6 μg/kg with an accuracy of 110.3%, thereby constituting satisfactory results. Upon testing 21 randomly purchased feed samples using this method, four of these samples contained AFB1, and the test results obtained using the developed method and stable isotope dilution LC-MS/MS were comparable. It was therefore apparent that the IMB purification method combined with UPLC analysis exhibited a good accuracy for aflatoxin determination. Thus, an automatic purification system was established to facilitate the operation and use of IMBs. This system was able to purify 24 samples simultaneously in 30 min. An IMB purification kit for was also designed and produced for aflatoxin detection in feed samples. The kit contained the sample dilutant, IMBs, the washing solution, and the eluent. After extraction of the feed sample, the extraction solution was added to the sample wells provided in the kit, and the purification system automatically completed the steps of aflatoxin enrichment, impurity washing, and elution of the target toxin. It should be noted that the purification process does not require the operator to manually add the solution, thereby simplifying operation. Overall, the purification method established in this study achieved the high-throughput and automatic purification of the four aflatoxins in feed samples.

Determination of 15 halogenated polycyclic aromatic hydrocarbons in aquatic products by stable isotope dilution coupled with gas chromatography-triple quadrupole mass spectrometry

Halogenated polycyclic aromatic hydrocarbons (H-PAHs), including chlorinated polycyclic aromatic hydrocarbons (Cl-PAHs) and brominated polycyclic aromatic hydrocarbons (Br-PAHs), are compounds in which one or more hydrogen atoms replaced by chlorine or bromine atoms. These compounds are not only difficult to degrade but also highly fat soluble and toxic. They are a new type of high-risk organic pollutants with structures similar to those of dioxins, and their toxicity is even higher than that of the parent polycyclic aromatic hydrocarbons (PAHs). The bioaccumulation of H-PAHs can be predicted by their octanol-water partition coefficient (Kow); in general, higher bioaccumulation capacity and Kow values indicate greater fat solubility. Therefore, animal-derived foods with higher fat contents, such as animal meat, milk, aquatic products, and their processed forms, are more likely to be contaminated with higher contents of H-PAHs than those with lower fat contents. In this work, a gas chromatography-triple quadrupole mass spectrometry (GC-MS/MS) method coupled with stable isotope dilution was established to determine 15 H-PAHs in aquatic products. The instrument and pretreatment methods were systematically optimized. The GC-MS/MS used in this method can effectively eliminate matrix interferences and features high sensitivity and low analytical cost; thus, it has good application prospects. The samples were added with an isotope internal standard before extraction to calibrate the loss of the tested substance during the pretreatment process, extracted by accelerated solvent extraction, purified using gel permeation chromatography and PRiME HLB columns, and then analyzed by GC-MS/MS. The use of two DB-5MS chromatographic columns (30 m×0.25 mm×0.25 μm) and microplate fluidics technology to connect chromatographic columns 1 and 2 in series led to better separation effects, good peak shapes, and high target compound responses. The 15 H-PAHs demonstrated good linearities in the range of 1-50 μg/L, with correlation coefficients (r) greater than or equal to 0.993. The relative standard deviation (RSD) values of the relative response factor (RRF) of the H-PAHs were less than 9%, the method detection limit (MDL) was 0.009-0.072 μg/kg, and the method quantification limit (MQL) was 0.031-0.240 μg/kg. Three spiked levels of 0.25, 1.0, 2.5 μg/kg were added to the blank samples to determine the recovery and precision. The recoveries for these spiked levels were 74.6%-116.8%, 77.8%-123.2%, and 71.9%-124.8%, respectively, and the corresponding RSDs were 0.6%-8.2%, 0.6%-9.0%, and 0.4%-10.6%, respectively. The total actual content of H-PAHs in aquatic product samples was 0.60-3.54 μg/kg. Among the H-PAHs investigated, 9-chlorophenanthrene (9-ClPhe) showed the greatest detection rate (100%) and highest content (1.15 μg/kg), indicating that H-PAHs widely exist in aquatic products. Thus, further assessment of the dietary exposure risk of these compounds is necessary. The developed method simplifies the pretreatment step, and has the advantages of simplicity, rapid analysis, high recoveries, and good stability. It is suitable for the qualitative and quantitative analysis of H-PAHs in actual aquatic product samples and provides reliable technical support for the residue status and risk assessment of H-PAHs in aquatic products.

Impact of LC-MS/MS quantification methods on analytical results by example of nationwide U.S. environmental samples

Although different quantification methods are extensively used in environmental chemistry, the impact of the choice of method on the quality and range of analytical results is understudied. This two-part study consists of (a) in-lab evaluation and (b) a traditional meta-analysis (n = 66) of commonly used quantification methods): (i) external calibration; (ii) isotope dilution method with authentic target analogs; (iii) isotope dilution with non-target standards; and (iv) standard addition prior to LC-MS/MS in liquid chromatography tandem mass spectrometry (LC-MS/MS) by example of antibiotics in sewage sludge from across the U.S. Using method (i) as the benchmark quantification method for the antibiotic erythromycin in biosolids, other quantification methods resulted in an overestimation (110–450 %) or an underestimation (10–60 %). Using the method (iv) as the benchmark for other compounds resulted in an overestimation (101–14,700 %) or an underestimation (6–98 %). Matrix effects were also observed and were dependent on the matrix and analyte type. For example, in the case of erythromycin, all sample matrices showed signal suppression. This study showed that in the absence of isotopically labeled analogs, the most accurate alternate quantification method may need to be experimentally determined depending on the analyte. Analysis of published literature on pharmaceuticals in sewage sludge indicated that isotope dilution with authentic target analog is most commonly used, followed by non-target isotope standards, standard addition, and finally external calibration.

Loss of body weight and lean mass in long-stay, hospitalized canine patients.

A high prevalence of malnutrition occurs in human hospitals and has been associated with detrimental consequences. By comparison, much less is known in hospitalized veterinary patients. The aims of this study were to evaluate the prevalence of malnutrition and body composition changes in long-stay hospitalised patients using an isotopic dilution technique. An additional objective was to compare the changes in composition with commonly used methods measuring body fat and lean mass. The dogs consumed on average 77.5% of their estimated resting energy requirements during their stay. The majority (78.3%) of dogs lost body weight, of which a greater proportion was lean mass (61.8%) than fat mass (FM) (38.2%). There was a moderate correlation between body condition score and percentage FM measured at admission (Kendall's τ = 0.51; p = 0.002), and at discharge (Kendall's τ = 0.55; p = 0.001). However, there was no correlation between muscle condition score and fat-free mass at either admission or discharge (p > 0.1). Duration of stay was positively associated with loss of body weight (p < 0.001), but was not associated with changes in either lean or FM expressed as a percentage of body weight or in absolute terms (p > 0.1), which was presumed to be explained by small sample size and variation. Food intake was not found to a significant factor for lean or FM loss (p > 0.1). These findings indicate that weight loss is common in hospitalized canine patients, which is not explained by simple under-eating. Other factors such as inflammation and inactivity should be evaluated in future studies to determine their role in influencing muscle and FM changes in hospitalized canine patients.

High-Throughput Analysis of Underivatized Amino Acids and Acylcarnitines in Infant Serum: A Micromethod Based on Stable Isotope Dilution Targeted HILIC-ESI-MS/MS.

Amino acids and acylcarnitines are important biomarkers of the body's energy state and can be used as diagnostic markers of certain inborn errors of metabolism. Few multianalyte methods for high-throughput analysis in serum exist for these compounds, but micromethods suitable for use in young children and infants are lacking. Therefore, we developed a quantitative high-throughput multianalyte hydrophilic interaction liquid chromatography-tandem mass spectrometry method preceded by a derivatization-free sample preparation using minimum amounts of serum (25 μL). Isotopically labeled standards were utilized for quantification. Forty amino acids and amino acid derivatives and 22 acylcarnitines were detected by applying a multiple reaction monitoring mode within a 20 min run. The method was comprehensively validated, comprising linearity, accuracy, (intraday/interday) precision, and quantitation limits, of which the latter ranged from 0.25 to 50 nM for acylcarnitines and from 0.005 to 1 μM for amino acids and their derivatives. Application of the method to 145 serum samples of three- to four-month-old healthy infants showed excellent reproducibility for multiday analyses and enabled simultaneous amino acid and acylcarnitine profiling in this age group.

Toward High-Throughput Analysis of Aroma Compounds Using Ultrahigh-Performance Liquid Chromatography-Tandem Mass Spectrometry: Screening of Key Food Odorants in Various Foods.

Recent studies show the immense capacities of the unified quantitation of aroma and taste compounds using liquid chromatography-mass spectrometry (LC-MS). The goal of this study was to highlight the broad application of this unified method. Thus, a stable isotope dilution analysis quantification method of the most important key food odorants in various food categories by LC-MS was developed. Using the well-known derivatization agent 3-nitrophenylhydrazine for carbonyl derivatization and a newly developed approach for alcohol and thiol derivatization, a method for the quantitation of 20 key food odorants was established. Intraday precision was determined to be ≤26%, and interday precision was between 24 and 31%. Limits of quantitation were determined between 0.014 and 283 μg/kg. The work shows that a wide array of aroma compounds can be analyzed accurately by LC-MS.

Ultra-high throughput dual channel liquid chromatography with tandem mass spectrometry for quantification of four immunosuppressants in whole blood for therapeutic drug monitoring

Liquid chromatography with tandem mass spectrometry (LC-MS/MS) is the golden standard for immunosuppressants analyses, where optimising throughput by parallel chromatography can reduce costs and turnaround time. We aimed to double our system throughput using a dual LC-MS/MS setup. Therefore, two independent UPLC systems were hyphenated to one triple quadrupole MS, with staggered injections from one autosampler on alternating columns. The method simultaneously measured the analytes tacrolimus, sirolimus, everolimus, and cyclosporin A in whole blood using isotope dilution, with a run time of 1.5 min. Using the dual LC-MS/MS system, net run-to-run time improved from 2.3 to 0.98 min per injection, where throughput increased from 26 to 61 injections per hour. For Performance Qualification, 1101 clinical samples were measured on the dual LC-MS/MS system in addition to the standard system, during a period of one month, and the results were compared using Passing Bablok regression and Bland Altman analysis. There was excellent agreement for all four analytes, with regression slopes of 0.98–1.02x and intercepts of -0.11–0.88 µg/L. Minor bias was demonstrated between the systems with mean differences from -0.93 to 1.43%. In conclusion, the throughput was doubled and idle MS time was reduced with good agreement to the standard system. Currently, the method is applied for clinical routine with frequent peak intensities of >180 injections per day.

Characterization of the Major Odor-Active Off-Flavor Compounds in Normal and Lipoxygenase-Lacking Soy Protein Isolates by Sensory-Directed Flavor Analysis.

The major off-flavor odorants of normal (NOR) and lipoxygenase-lacking (LOX-lack) soy protein isolates (SPIs) were discriminated by sensory-directed flavor analysis. A total of 32 odor-active off-flavor compounds were detected in SPIs, and 19 of them with flavor dilution (FD) factors ranging from 3 to 2187 were quantified by external standard curves. It was found that hexanal and nonanal dominated in the off-flavor profile of SPIs regarding their odor activity values (OAVs) and FD contributions, followed by octanal, 1-hexanol, 1-octen-3-ol, 2-heptone, and benzaldehyde. To further improve the precision of quantification, the above 7 major odor-active off-flavor compounds were requantified by stable isotope dilution assays (SIDA) for the first time. The results suggested that the sensory differences between NOR and LOX-lack SPIs could be mainly due to the reduced C6/C9 aldehydes and alcohols rather than 1-octen-3-ol and benzaldehyde. Finally, these differential compounds were further verified by the spiking experiment.

Association between Organochlorine Pesticides and Vitamin D in Female Subjects.

In human population studies, organochlorine pesticides (OCPs) have been linked to vitamin D deficiency. Therefore, this study examined the association between OCPs, vitamin D3 (cholecalciferol, 25(OH)D3), and the active metabolite 1,25-dihydrovitamin D3 (1,25(OH)2D3) in a cohort of non-obese women. The serum samples of 58 female participants (age-31.9 ± 4.6 years; body mass index (BMI)-25.7 ± 3.7 kg/m2) were screened for 10 indicator OCPs. 25(OH)D3 and 1,25(OH)2D3 levels were determined via isotope dilution liquid chromatography tandem mass spectrometry. In this cohort, the 25(OH)D3 and 1,25(OH)2D3 levels were 22.9 ± 11.2 ng/mL and 0.05 ± 0.02 ng/mL, respectively, with 28 participants classified as 25(OH)D3-deficient (<50 nmol/L). In the study cohort, no correlations were found between individual or total OCPs (ƩOCPs) and 25(OH)D3. p,p'-dichlorodiphenyldichloroethylene (DDE) and ƩOCPs correlated positively with 1,25(OH)2D3, with the latter being negatively correlated with estimated glomerular filtration rate (eGFR). In women with sufficient 25(OH)D3 levels, p,p'-dichlorodiphenyltrichloroethan (DDT) was positively correlated with 1,25(OH)2D3, whilst in the deficient group, hexachlorobenzene (HCB) and p,p'-(DDE) were positively correlated with 1,25(OH)2D3, β-Hexachlorocyclohexane (HCH) was positively correlated with 25(OH)D3, and none of the OCPs were associated with measures of renal function. Overall, OCPs and ƩOCPs were not associated with 25(OH)D3, suggesting that they are unrelated to vitamin D deficiency, but p,p'-DDE and ƩOCPs correlated positively with active 1,25(OH)2D3, while ƩOCPs correlated negatively with eGFR, suggesting a possible renal effect. Analysis of vitamin D deficiency revealed an association between β-HCH and 25(OH)D3, and between HCB and p,p'-DDE and 1,25(OH)2D3, suggesting that OCP effects may be enhanced in cases of vitamin D deficiency.

Various seasonal patterns and potential mechanisms of soil nitrogen transformation along an elevational gradient in the Hengduan Mountains

Abstract Soil nitrogen (N) transformation, a key microbial process in global N cycling, is thought to alter soil N availability and subsequently regulate ecosystem functioning. In particular, the gross rates of N transformation can provide a deeper understanding of internal N dynamics and mechanisms, but questions of whether gross N transformation processes and their underlying drivers change with seasons and elevations remain large uncertain. Based on field collection along an elevational gradient and laboratory incubation experiments, we investigated the seasonal processes of soil N mineralization and nitrification with 15N isotope dilution technique, and also explored the potential mechanisms involved in the Hengduan Mountains. Unimodal soil gross/net mineralization rates were higher in the wet season (61.32 mg kg−1 day−1; 1.01 mg kg−1 day−1 respectively) than in the dry season (10.88 mg kg−1 day−1; 0.55 mg kg−1 day−1 respectively) (p &lt; 0.001), with a peak at medium elevation. Soil gross/net nitrification rates were lower in the wet season (1.43 mg kg−1 day−1; −0.017 mg kg−1 day−1) than in the dry season (2.49 mg kg−1 day−1; 0.004 mg kg−1 day−1 respectively) (p &lt; 0.001), which increased with increasing elevation. The results detected the dominant drivers of the gross transformation rates, specifically microbial attributes occupied crucial roles in controlling the gross mineralization/nitrification in the wet season, and soil physicochemical properties were dominant controllers on the gross mineralization/nitrification during the dry season. This study revealed the divergent patterns and drivers of N transformation, suggesting that seasonal N cycling should no longer be overlooked if we are to predict N biogeochemical cycles in response to environmental change accurately. Read the free Plain Language Summary for this article on the Journal blog.

Features of steroidogenesis and arterial hypertension in men in different types of "physiological" male hyperandrogenism

To reveal the peculiarities of steroidogenesis and arterial hypertension in «physiological» hyperandrogenism in men. One-stage simultaneous study. The groups of men with hyperandrogenism caused by increased total testosterone (n=34) and those with hyperandrogenism caused by increased dihydrotestosterone (DHT) (n=66) were compared. In determining the type of hyperandrogenism and allocating patients to groups, DHT and total testosterone levels were determined by enhanced chemiluminescence. Subgroups of men with and without arterial hypertension were compared in the group of patients with hyperandrogenism due to an increase in total testosterone. Body mass index, waist circumference, systolic and diastolic blood pressure, pulse, and LH, SBHG, estradiol, blood multisteroid levels by isotope dilution liquid chromatography/tandem mass spectrometry, glucose, blood lipid spectrum, uric acid, creatinine, renin, potassium, sodium, and blood chloride were assessed in all patients. Patients with arterial hypertension additionally underwent daily BP monitoring, albuminuria assessment, electrocardiography, ocular fundus examination. The baseline threshold level of significance was p&lt;0.05. For multiple comparisons, the p significance level was calculated using the Bonferroni correction. Statistically significant differences were found in the levels of 17-hydroxypregnenolone, 17-hydroxyprogesterone, and androstenedione, which were higher in men with elevated levels of total testosterone. No statistically significant differences in other laboratory parameters were found. No cases of increased blood pressure were detected in the group of men with elevated DHT. In the group of men with elevated total testosterone, 23,5% of men with arterial hypertension without targetorgan lesions were identified, while hyperandrogenism was associated with 17,6% of cases. Arterial hypertension associated with hyperandrogenism was characterized by a rise in blood pressure in the early morning hours. Estradiol levels, while remaining within normal limits, were statistically significantly lower in patients with arterial hypertension compared with men with elevated testosterone but without hypertension. No cases of arterial hypertension were observed in «physiological» hyperandrogenism due to elevated DHT levels, whereas its incidence in «physiological» hyperandrogenism due to elevated total testosterone was 23,5%. The features of steroidogenesis were increased production of 17-hydroxypregnenolone, 17-hydroxyprogesterone, and androstenedione in men with testosterone hyperandrogenism and decreased estradiol production in patients with arterial hypertension compared with patients without testosterone hyperandrogenism.

Calibrating the Early Cretaceous Urho Pterosaur Fauna in Junggar Basin and implications for the evolution of the Jehol Biota

Over the past decades, abundant and well-preserved vertebrate fossils, known as the Urho Pterosaur Fauna, have been recovered from the Lower Cretaceous Tugulu Group in Junggar Basin, Xinjiang, NW China. Excavated materials belong to pterosaur, plesiosaur, dinosaur, crocodylomorph, and turtle taxa. As such, they provide key insights into the evolutionary history of several critical vertebrate groups in the Early Cretaceous. The Junggar assemblages have been interpreted as belonging to the Jehol Biota sensu lato, representing its northwesternmost known geographic extent. This research presents a new chemical abrasion−isotope dilution−thermal ionization mass spectrometry U-Pb age of 135.2 ± 0.9 Ma (2σ internal error) from a tuffaceous bed stratigraphically below the fauna-bearing layers, indicating a Valanginian maximum age for the Urho fauna. Combined with available biostratigraphic data, the results bear several important paleobiologic implications for the Early Cretaceous vertebrates. First, the Dsungaripterus pterosaur and Psittacosaurus ornithischian fauna appear to have emerged earlier than previously believed. Second, the data suggest that the oldest carcharodontosaurids in Asia appeared during the Valanginian Stage and extend the age range of basal coelurosaurs and basal crocodyliforms. Our results do not support the notion of the Jehol Biota sensu lato migrating as far west as the Junggar Basin in their later stages. The new information calls into question the temporal and spatial bases for the conventional, three-stage evolutionary theory of the Jehol Biota.

Determination of 24,25-dihydroxyvitamin D3 in Vitamin D External Quality Assessment Scheme samples using a reference measurement procedure

Ninety archived human serum samples from the Vitamin D External Quality Assessment Scheme (DEQAS) were analyzed using a reference measurement procedure (RMP) based on isotope dilution liquid chromatography – tandem mass spectrometry (ID LC-MS/MS) for the determination of 24,25-dihydroxyvitamin D3 [24,25(OH)2D3]. These 24,25(OH)2D3 results, in conjunction with concentration values assigned using RMPs for 25-hydroxyvitamin D2 [25(OH)D2] and 25-hydroxyvitamin D3 [25(OH)D3], provide a valuable resource for assessing the accuracy of measurements for 24,25(OH)2D3 and for investigating the relationship between 24,25(OH)2D3 and 25(OH)D3. Results for 24,25(OH)2D3 using the RMP were compared to DEQAS consensus values demonstrating that the consensus values were not sufficient to assess the accuracy of measurements among different laboratories and methods. A multivariable regression analysis approach using historical DEQAS consensus values for various total 25(OH)D assays was used to assess the contribution of 24,25(OH)2D3 concentration on the assay response. The response of several ligand binding assays for total 25(OH)D was shown to be impacted by the presence of 24,25(OH)2D3.