Chromatography-isotope Dilution Tandem Mass Spectrometry Research Articles

Association of bone mineral density with nine urinary personal care and consumer product chemicals and metabolites: A national-representative, population-based study

Personal care and consumer products are extensively used and contain various chemicals that are recognized as endocrine disrupting chemicals. Few epidemiological studies measured the associations of these chemicals and metabolites, especially phenols, with bone health. We measured the associations between nine urinary phenol biomarkers and BMD and the subsequent prevalence of osteopenia/osteoporosis. National representative data from US adults aged over 20 years who participated in the National Health and Nutrition Examination Survey from 2005 to 2010 were used. 2,267 men, 1,145 premenopausal women and 1,033 postmenopausal women were included in the final analyses. Bisphenol A, benzophenone-3, triclosan, butylparaben, ethylparaben, methylparaben, propylparaben, 2,5-dichlorophenol and 2,4-dichlorophenol were measured by high-performance liquid chromatography-isotope dilution tandem mass spectrometry. Paraben concentrations, especially ethyl-, methyl- and propylparabens, were related to greater BMD in the femoral neck and intertrochanter and lumbar spine in men and premenopausal women. 2,4-dichlorophenol concentrations was associated with lower BMD and a higher prevalence of osteopenia + osteoporosis in men. Bisphenol A was associated with a higher prevalence of osteopenia + osteoporosis in the lumbar spine in postmenopausal women. Benzophenone-3 tended to be associated with a higher BMD of femur in men and premenopausal women. In conclusion, certain urinary personal care and consumer product chemicals and metabolites were significantly associated with BMD or osteopenia + osteoporosis. These results further indicate that, in future research, determination of the causal relationships and detection of undelying mechanism could be next important directions.

Simple and accurate candidate reference measurement procedure for total testosterone in human serum by one-step liquid-liquid extraction coupled with isotope dilution mass spectrometry.

Testosterone in human serum is commonly tested in clinical laboratories using immunoassay methods as well as liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods. To standardize and ensure the accuracy of the measurement results, reference procedures with higher metrological order are required. A simple measurement procedure based on one-step liquid-liquid extraction (LLE) and liquid chromatography-isotope dilution tandem mass spectrometry (LC-IDMS/MS) was developed for total testosterone in human serum. The procedure involved serum spiked with 13C3-testosterone, equilibration for 2h, and extraction with an organic solvent. Testosterone certified reference material (CRM) was used as the calibration standard to ensure the traceability to the International System of Units (SI). Testosterone in serum CRMs from the National Institute for Standards and Technology (NIST) and LGC were used to validate the accuracy of the newly developed method. The deviations of the obtained values from the NIST and LGC certified values ranged from -0.55% to 0.45%. Similarly, the coefficient of variations (CVs) of the replicate measurements were in the range of 0.55% and 0.78%, respectively. The relative expanded uncertainties were comparable with those of the certified materials. The newly developed LC-IDMS/MS procedure demonstrated adequate trueness and precision, and was simple to perform. The method can be used for value assignment of testosterone in external quality assessment (EQA) materials as well as certification of CRMs in the future. Graphical abstract.

Commutable whole blood reference materials for hemoglobin A1c validated on multiple clinical analyzers.

Background The measurement of hemoglobin A1c (HbA1c) is important for diagnosing diabetes mellitus as well as assessing glycemic control in diabetic patients. Commutable whole blood certified reference materials (CRMs) are needed in the measurement of HbA1c for method validation and/or as quality controls. Methods We developed three levels of hemolyzed whole blood CRMs for HbA1c. The certified values were determined using liquid chromatography-isotope dilution tandem mass spectrometry method (LC-IDMS/MS) where two "signature" hexapeptides of HbA1c and hemoglobin A0 (HbA0) were used as the calibration standards. The concentrations of the hexapeptide solutions were determined by amino acid analysis by the LC-IDMS/MS method using amino acid CRMs as the calibration standards. The commutability study was conducted by measuring 25 patient specimens and the whole blood CRMs by both LC-IDMS/MS method and various routine methods using six different clinical analyzers. Results The certified values were determined to be 35.1±2.0, 50.3±1.9 and 65.8±2.6 mmol/mol, respectively. These CRMs showed good commutability on five of the six clinical analyzers but showed poor commutability on one of the clinical analyzers that used similar method as two other analyzers where good commutability was observed. Conclusions With certified target values based on metrological traceability and good commutability on most of the clinical analyzers, the developed whole blood CRMs can be used for method validation or as quality control materials in the measurement of HbA1c. The commutability study results also underscored the need of commutability testing of clinical CRMs using various clinical analyzers.

Identification and quantification of DNA repair protein poly(ADP ribose) polymerase 1 (PARP1) in human tissues and cultured cells by liquid chromatography/isotope-dilution tandem mass spectrometry

Poly(ADP ribose) polymerase 1 (PARP1) is a multifunctional DNA repair protein of the base excision repair pathway and plays a major role in the repair of DNA strand breaks and in replication and transcriptional regulation among other functions. Mounting evidence points to the predictive and prognostic value of PARP1 expression in human cancers. Thus, PARP1 has become an important target in cancer therapy, leading to the development of inhibitors as anticancer drugs. In the past, PARP1 expression levels in tissue samples have generally been estimated by indirect and semi-quantitative immunohistochemical methods. Accurate measurement of PARP1 in normal tissues and malignant tumors of patients will be essential for evaluating PARP1 as a predictive and prognostic biomarker in cancer and other diseases, and for the development and use of its inhibitors in cancer therapy. In this work, we present an approach involving liquid chromatography–isotope-dilution tandem mass spectrometry to positively identify and accurately quantify PARP1 in human tissues and cultured cells. We identified and quantified PARP1 in human normal ovarian tissues and malignant ovarian tumors, and in three pairs of human cell lines, each pair consisting of a normal cell line and its cancerous counterpart. Significantly greater expression of PARP1 was observed in malignant ovarian tissues than in normal ovarian tissues. In the case of one pair of cell lines, the cancerous cell line also exhibited greater expression of PARP1 than in normal cell line. We also show the simultaneous measurement of PARP1 and apurinic/apyrimidinic endonuclease 1 (APE1) in a given protein extract. The approach presented in this work is expected to contribute to the accurate quantitative assessment of PARP1 levels in basic research and clinical studies.

Ultra-High-Performance Liquid Chromatography-Isotope Dilution Tandem Mass Spectrometry for the Determination of Phthalate Secondary Metabolites in Human Serum Based on Solid-Phase Extraction.

>Background: A multitude of biomonitoring analytical methods are applied in urinary detection, but few are applicable to phthalate metabolites in human serum, and those that are invariably involve complex operation. Objective: To develop a novel method for the determination of phthalate monoesters in human serum using solid-phase extraction (SPE) and ultra-HPLC (UHPLC)-tandem mass spectrometry for multiple human serum sample determination. Method: Analytes in serum samples were extracted and purified with a novel SPE cartridge named Prime HLB and then directly injected and analyzed without eluent redissolution. After sample extraction and cleanup, 11 phthalate monoesters were separated on a C18 chromatographic column in UHPLC. Among them, mono-isobutyl phthalate and mono-n-butyl phthalate, a pair of isomers, were successfully separated. Results: Lower LODs of the 11 monoesters in 0.5 mL human serum were in the range of 0.03-3 ng/mL, and lower LOQs accordingly ranged from 0.1 to 10.0 ng/mL. Recoveries from spiked bovine serum samples were in the range of 95.3-109.9%, and RSDs were 8.0-12.0%. Intra- and inter-day recoveries were in the range of 94.1-115.7%, with RSDs <13.7%. Conclusions: The proposed method has been proved to be rapid, simple, accurate, and sensitive, so that it may potentially be used for high-throughput biological monitoring with high efficiency and precision. Highlights: Sample pretreatment does not require eluent drying and redissolution. Mass losses of analytes are avoided. Operational procedures are easy and rapid. The method has proved applicable for 11 phthalate metabolites in multiple human serum sample determinations.

Quantification of DEET and neonicotinoid pesticide biomarkers in human urine by online solid-phase extraction high-performance liquid chromatography-tandem mass spectrometry.

Neonicotinoid insecticides are widely used replacements for organophosphate and carbamate insecticides, but the extent of human exposure is largely unknown. On the other hand, based on urinary concentrations of DEET metabolites, human exposure to N,N-diethyl-m-toluamide (DEET) appears to be widespread. We developed a fast online solid-phase extraction high-performance liquid chromatography-isotope dilution tandem mass spectrometry (HPLC-MS/MS) method to measure in 200μL of human urine the concentrations of six neonicotinoid biomarkers (acetamiprid, N-desmethyl-acetamiprid, clothianidin, imidacloprid, 5-hydroxy-imidacloprid, thiacloprid), and two DEET biomarkers (3-diethyl-carbamoyl benzoic acid, 3-ethyl-carbamoyl benzoic acid). Limits of detection ranged from 0.01 to 0.1μg/L, depending on the biomarker. Accuracy ranged from 91 to 116% and precision ranged from 3.7 to 10 %RSD. The presented method can be used to increase our understanding of exposure to neonicotinoid insecticides and DEET, and to evaluate the potential health effects from such exposures.

Impact of enzymatic hydrolysis on the quantification of total urinary concentrations of chemical biomarkers

Human exposure to consumer and personal care products chemicals such as phenols, including parabens and other antimicrobial agents, can be assessed through biomonitoring by quantifying urinary concentrations of the parent chemical or its metabolites, often after hydrolysis of phase II conjugates. Developing suitable analytical methods for the concurrent quantification of multiple exposure biomarkers is challenging because optimal conditions for the hydrolysis of such conjugates (e.g., O-glucuronides, N-glucuronides, sulfates) may differ depending on the biomarker. We evaluated the effectiveness of seven commercial hydrolytic enzymes to simultaneously hydrolyze N-glucuronides (using the antibacterial triclocarban as example compound) and other conjugates (using select phenols and parabens as examples) by using on-line solid phase extraction–high performance liquid chromatography-isotope dilution-tandem mass spectrometry. Incubation (30 min, 55 °C) with a genetically engineered β-glucuronidase (IMCS, ≥15 units/μL urine) hydrolyzed N-glucuronide triclocarban, but did not fully hydrolyze the conjugates of phenols and parabens. By contrast, incubation (4 h, 37 °C) with solid β-glucuronidase (Helix pomatia, Type H-1, ≥30 units/μL urine) or liquid β-glucuronidase/arylsulfatase (Helix pomatia, 30 units/μL urine [i.e., 30 μL/100 μL urine]) in the presence of 100 μL methanol for 100 μL urine completely hydrolyzed N-glucuronide triclocarban and the conjugates of several phenols and parabens, without cleaving the ester bond of the parabens to form p-hydroxybenzoic acid. These results highlight the relevance of method validation procedures that include optimizing the hydrolysis of phase II urinary conjugates (e.g., enzyme type and amount used, reaction time, temperature) to quantify accurately and concurrently multiple exposure biomarkers for biomonitoring purposes.

Rapid qualitative and quantitative analysis of aflatoxin B1 in Pu-erh tea by liquid chromatography-isotope dilution tandem mass spectrometry coupled with the QuEChERS purification method

An efficient method for the determination of aflatoxin B1 in Pu-erh tea by isotope dilution mass spectrometry coupled with QuEChERs was established.

Urinary triclosan concentrations and diminished ovarian reserve among women undergoing treatment in a fertility clinic

To investigate the association between urinary triclosan concentrations and antral follicle count (AFC), a well-accepted marker of ovarian reserve, among women from a fertility center. Prospective cohort study. Hospital fertility center. A total of 109 women. Urinary triclosan concentrations quantified by online solid phase extraction-high performance liquid chromatography-isotope dilution tandem mass spectrometry. AFC through transvaginal ultrasonography on the third day of an unstimulated menstrual cycle or on the third day of a progesterone withdrawal bleed. The geometric mean of the specific gravity-adjusted urinary triclosan concentrations for the 225 samples provided by the 109 women was 13.0μg/L (95% confidence interval [CI], 8.9, 19.1). Women had median (with interquartile range) AFC of 13 (8, 18). The specific gravity-adjusted urinary triclosan concentrations were inversely associated with AFC (-4%; 95% CI, -7%, -1%). Women with triclosan concentrations above the median had lower AFC compared with those whose triclosan concentrations were equal to or below the median, with an adjusted difference of -3.2 (95% CI, -3.9, -1.6) among those with a body mass index <25kg/m2 and -1.8 (95% CI, -3.2, -0.3) among those who were <35years old. Specific gravity-adjusted urinary triclosan concentrations were inversely associated with AFC in women seeking care at a fertility center. This association was modified by age and body mass index, with the younger and leaner women showing larger decreases in AFC.

Liquid chromatography–isotope dilution tandem mass spectrometry method for the measurement of urea in human serum and assignment of reference values to external quality assessment samples

Reference measurement procedures are essential for the accurate measurement of urea in human serum. The widely used gas chromatography-isotope dilution mass spectrometry (GC–IDMS) involves tedious derivatization processes. We developed and validated a simple liquid chromatography-isotope dilution tandem mass spectrometry (LC-IDMS/MS) method for the measurement of urea in human serum.In this method, 13C,15N2-urea was spiked into serum samples as the internal standard, and pure urea certified reference material (CRM) was used as the calibration standard to ensure the traceability of the measurements to the International System of Units (SI). The sample cleanup procedure involved a simple protein precipitation step, followed by centrifugation and filtration. The developed LC-IDMS/MS method was found to be accurate and highly reproducible. The coefficient of variation (CV) was generally below 1%. Good accuracy was demonstrated by measuring serum CRMs from two National Metrology Institutes (NMIs) with relative deviations from the certified values being close to or below 1%, and measurement uncertainties of 2.0%–2.8%. The method was used to assign the reference values of serum urea in ten cycles of an external quality assessment (EQA) program for clinical laboratories over a period of four years. The robust mean of the reported results from clinical laboratories using automated analyzers were close to the assigned values from LC-IDMS/MS method.With good accuracy and precision, as well as simple sample cleanup procedure, the developed LC-IDMS/MS method can be used as a reference measurement procedure for the measurement of urea in human serum.

Elevated urinary levels of 8-oxo-2′-deoxyguanosine, (5′R)- and (5′S)-8,5′-cyclo-2′-deoxyadenosines, and 8-iso-prostaglandin F2α as potential biomarkers of oxidative stress in patients with prediabetes

Prediabetes is the preclinical stage of type 2 diabetes mellitus (T2DM) with intermediate state of hyperglycemia. Hyperglycemia results in a state of oxidative stress, which may contribute to the production of insulin resistance, β-cell dysfunction and long-term complications of diabetes. Novel approaches are required for prevention and treatment of diabetes. New biomarkers that can be used in risk stratification and therapy control as supplementary to current parameters are needed. These biomarkers may facilitate a more individualized and sufficient treatment of diabetes. Therefore, the aim of this study was to investigate the levels of oxidatively induced DNA damage products, 8-oxo-2′-deoxyguanosine (8-oxo-dG) (also known as 8-OH-dG), (5′R)- and (5′S)-8,5′-cyclo-2′-deoxyadenosines (R-cdA and S-cdA), and the lipid peroxidation product 8-iso-prostaglandin F2α (8-iso-PGF2α) as reliable oxidative stress markers in patients with prediabetes or T2DM in comparison with healthy volunteers. Urine samples were collected from these subjects. Absolute quantification of 8-oxo-dG, R-cdA, S-cdA and 8-iso-PGF2α was achieved by liquid chromatography-isotope dilution tandem mass spectrometry. The levels of 8-oxo-dG, S-cdA and 8-iso-PGF2α were significantly greater in prediabetes patients than those in healthy volunteers. T2DM patients also had higher levels of 8-oxo-dG than healthy volunteers. No statistically significant difference was observed for R-cdA levels. 8-Oxo-dG levels positively correlated with R-cdA and S-cdA levels for prediabetes and newly diagnosed T2DM. S-cdA levels and HbA1c were found negatively correlated in prediabetes patients. Also 8-iso-PGF2α levels and HbA1c were found negatively correlated in prediabetes patients. These results indicate that oxidatively induced macromolecular damage appears before the establishment of T2DM. Thus, our data suggest that oxidatively induced DNA damage and lipid peroxidation products that were found to be elevated in prediabetic stage may be used as early disease markers in patients at risk for T2DM.

Lewisite Metabolites in Urine by Solid Phase Extraction-Dual Column Reversed-Phase Liquid Chromatography-Isotope Dilution Tandem Mass Spectrometry.

Lewisite (2-chlorovinyldichloroarsine) is a chemical warfare agent developed during World War I. A quantitative method using solid phase extraction (SPE) followed by dual column liquid chromatography (LC)-isotope dilution tandem mass spectrometry(MS-MS) was developed for the determination of (2-chlorovinyl)arsonic acid (CVAOA), a metabolite of Lewisite, in human urine. The sample was treated with hydrogen peroxide to oxidize any (2-chlorovinyl)arsonous acid (CVAA) that remained in the trivalent arsenic oxidation state. There was 1.19% (arsenic purity) of bis-(2-chlorovinyl)arsinic acid (BCVAOA), a minor Lewisite metabolite, in the stock CVAA material. The high-throughput method qualitatively assessed BCVAOA simultaneously utilizing normal-phase silica SPE followed by reversed-phase C18 LC for an orthogonal separation. The chromatographic method results in a5.8-min cycle time with adequate retention (k'=2.4) of CVAOA. The mass spectrometer was operated in positive electrospray ionization mode with quantitativem/z 186.9→61.0 and confirmation 186.9→91.0 mass transitions. This selective method demonstrated linearity, accuracy and reproducibility for the clinically relevant calibration range (25-3,200 µg/L as CVAA). The method detection limit was 3.3 µg/L as CVAA from a 10µL injection. This LC-MS-MS emergency response method has a throughput of >240 samples(2.5 extracted 96-well plates) per day.

Simple and accurate measurement of carbamazepine in surface water by use of porous membrane-protected micro-solid-phase extraction coupled with isotope dilution mass spectrometry

To achieve fast and accurate analysis of carbamazepine in surface water, we developed a novel porous membrane-protected micro-solid-phase extraction (μ-SPE) method, followed by liquid chromatography-isotope dilution tandem mass spectrometry (LC-IDMS/MS) analysis. The μ-SPE device (∼0.8 × 1 cm) was fabricated by heat-sealing edges of a polypropylene membrane sheet to devise a bag enclosing the sorbent. The analytes (both carbamazepine and isotope-labelled carbamazepine) were first extracted by μ-SPE device in the sample (10 mL) via agitation, then desorbed in an organic solvent (1 mL) via ultrasonication. Several parameters such as organic solvent for pre-conditioning of μ-SPE device, amount of sorbent, adsorption time, and desorption solvent and time were investigated to optimize the μ-SPE efficiency. The optimized method has limits of detection and quantitation estimated to be 0.5 ng L−1 and 1.6 ng L−1, respectively. Surface water samples spiked with different amounts of carbamazepine (close to 20, 500, and 1600 ng L−1, respectively) were analysed for the validation of method precision and accuracy. Good precision was obtained as demonstrated by relative standard deviations of 0.7% for the samples with concentrations of 500 and 1600 ng kg−1, and 5.8% for the sample with concentration of 20 ng kg−1. Good accuracy was also demonstrated by the relative recoveries in the range of 96.7%–103.5% for all samples with uncertainties of 1.1%–5.4%. Owing to the same chemical properties of carbamazepine and isotope-labelled carbamazepine, the isotope ratio in the μ-SPE procedure was accurately controlled. The use of μ-SPE coupled with IDMS analysis significantly facilitated the fast and accurate measurement of carbamazepine in surface water.

Monitoring exposure to polycyclic aromatic hydrocarbons in an Australian population using pooled urine samples

Integrated exposure to polycyclic aromatic hydrocarbons (PAHs) can be assessed through monitoring of urinary mono-hydroxylated PAHs (OH-PAHs). The aim of this study was to provide the first assessment of exposure to PAHs in a large sample of the population in Queensland, Australia including exposure to infant (0–4years). De-identified urine specimens, obtained from a pathology laboratory, were stratified by age and sex, and pooled (n=24 pools of 100) and OH-PAHs were measured by gas chromatography–isotope dilution–tandem mass spectrometry. Geometric mean (GM) concentrations ranged from 30ng/L (4-hydroxyphenanthrene) to 9221ng/L (1-naphthol). GM of 1-hydroxypyrene, the most commonly used PAH exposure biomarker, was 142ng/L. The concentrations of OH-PAHs found in this study are consistent with those in developed countries and lower than those in developing countries. We observed no association between sex and OH-PAH concentrations. However, we observed lower urinary concentrations of all OH-PAHs in samples from infants (0–4years), children (5–14years) and the elderly (>60year old) compared with samples from other age groups (15–29, 30–44 and 45–59years) which may be attributed to age-dependent behaviour-specific exposure sources.

Simplified Method for Quantifying Sulfur Mustard Adducts to Blood Proteins by Ultrahigh Pressure Liquid Chromatography−Isotope Dilution Tandem Mass Spectrometry.

Sulfur mustard binds to reactive cysteine residues, forming a stable sulfur-hydroxyethylthioethyl [SHETE]adduct that can be used as a long-term biomarker of sulfur mustard exposure in humans. The digestion of sulfur mustard-exposed blood samples with proteinase K following total protein precipitation with acetone produces the tripeptide biomarker [S-HETE]-Cys-Pro-Phe. The adducted tripeptide is purified by solid phase extraction, separated by ultra high pressure liquid chromatography, and detected by isotope dilution tandem mass spectrometry. This approach was thoroughly validated and characterized in our laboratory. The average interday relative standard deviation was ≤ 9.49%, and the range of accuracy was between 96.1 and 109% over a concentration range of 3.00 to 250. ng/mL with a calculated limit of detection of1.74 ng/mL. A full 96-well plate can be processed and analyzed in 8 h, which is 5 times faster than our previous 96-well plate method and only requires 50 μL of serum, plasma, or whole blood. Extensive ruggedness and stability studies and matrix comparisons were conducted to create a robust, easily transferrable method. As a result, a simple and high-throughput method has been developed and validated for the quantitation of sulfur mustard blood protein adducts in low volume blood specimens which should be readily adaptable for quantifying human exposures to other alkylating agents.

Report on the two‐phase public consultation on the draft EFSA scientific opinion on bisphenol A (BPA)

Report on the two‐phase public consultation on the draft EFSA scientific opinion on bisphenol A (BPA)

T-2 and HT-2 toxins in oat flakes: development of a certified reference material.

Trichothecene mycotoxins, with T-2 and HT-2 toxins being the main representatives of the type A subgroup, are naturally and worldwide occurring contaminants frequently found in grain-based food and feed. Due to the high consumption of these products and the potential health risk associated herewith, concerns about the safety and quality of food and feed have increased over the last decades at both governmental and consumer levels. Since it is not possible to avoid their occurrence, tremendous efforts have been performed to identify and monitor mycotoxins in food and feed to make their consumption safe. However, suitable certified reference materials (CRMs) intended for quality assurance and quality control purposes are still lacking for many mycotoxin-matrix combinations. Therefore, in the framework of a European Reference Material (ERM®) project, the first CRM for T-2 and HT-2 toxin in ground oat flakes (ERM®-BC720) was developed according to the requirements of ISO Guide 35. The whole process of ERM®-BC720 development, including sample preparation, homogeneity and stability studies and value assignment, is presented. The assignment of the certified mass fractionswas based upon an in-house study using high-performance liquid chromatography isotope-dilution tandem mass spectrometry. Simultaneously, an interlaboratory comparison study involving 24 expert laboratories was conducted in order to support the in-house certification study. The certified values and their corresponding expanded uncertainties (k = 2) for both T-2 and HT-2 toxin in ERM®-BC720, traceable to the international system of units, are (82 ± 4)μgkg(-1) and (81 ± 4)μgkg(-1), respectively.

Significant disparity in base and sugar damage in DNA resulting from neutron and electron irradiation.

In this study, a comparison of the effects of neutron and electron irradiation of aqueous DNA solutions was investigated to characterize potential neutron signatures in DNA damage induction. Ionizing radiation generates numerous lesions in DNA, including base and sugar lesions, lesions involving base–sugar combinations (e.g. 8,5′-cyclopurine-2′-deoxynucleosides) and DNA–protein cross-links, as well as single- and double-strand breaks and clustered damage. The characteristics of damage depend on the linear energy transfer (LET) of the incident radiation. Here we investigated DNA damage using aqueous DNA solutions in 10 mmol/l phosphate buffer from 0–80 Gy by low-LET electrons (10 Gy/min) and the specific high-LET (∼0.16 Gy/h) neutrons formed by spontaneous 252Cf decay fissions. 8-hydroxy-2′-deoxyguanosine (8-OH-dG), (5′R)-8,5′-cyclo-2′-deoxyadenosine (R-cdA) and (5′S)-8,5′-cyclo-2′-deoxyadenosine (S-cdA) were quantified using liquid chromatography–isotope-dilution tandem mass spectrometry to demonstrate a linear dose dependence for induction of 8-OH-dG by both types of radiation, although neutron irradiation was ∼50% less effective at a given dose compared with electron irradiation. Electron irradiation resulted in an exponential increase in S-cdA and R-cdA with dose, whereas neutron irradiation induced substantially less damage and the amount of damage increased only gradually with dose. Addition of 30 mmol/l 2-amino-2-(hydroxymethyl)-1,3-propanediol (TRIS), a free radical scavenger, to the DNA solution before irradiation reduced lesion induction to background levels for both types of radiation. These results provide insight into the mechanisms of DNA damage by high-LET 252Cf decay neutrons and low-LET electrons, leading to enhanced understanding of the potential biological effects of these types of irradiation.

Measurement procedure for glucose in serum using liquid chromatography isotope dilution--tandem mass spectrometry (LC-ID-MS/MS).

We developed and validated a measurement procedure for glucose using liquid chromatography-isotope dilution tandem mass spectrometry. The proposed method is intended to be used for setting target values in EQAS samples and for certification of glucose reference materials, including those in biological matrices. Serum samples were spiked with internal standard 13C6 D-glucose. Protein precipitation was performed with ethanol. The samples were vortexed and centrifuged. An aliquot of the supernatant was evaporated to dryness, the residue dissolved in elution buffer and injected into the LC-MS/MS system. Measurements were performed in the positive ion mode monitoring the Cs+ adducts for glucose at m/z 313 --> 132.9 and m/z 319 --> 132.9 for the internal standard. The coefficient of variation (CV) of the measurement procedure for lyophilized, liquid, and fresh serum samples was between 0.27 and 1.77%. The bias from certified target values for NIST reference materials was < or = 0.62%. A Deming regression comparison demonstrated a good correlation of results obtained with the proposed LC-ID-MS/MS method and target values obtained in the internationally accepted IFCC-RELA ring trial using JCTLM-recognized reference measurement procedures. The proposed LC-ID-MS/MS measurement procedure with traceability to SI units shows excellent accuracy and precision and is suitable for use as reference measurement procedure for certification of target values in lyophilized and fresh serum samples.

Automated on-line column-switching high performance liquid chromatography isotope dilution tandem mass spectrometry method for the quantification of bisphenol A, bisphenol F, bisphenol S, and 11 other phenols in urine

Human exposure to bisphenol A (BPA) is widespread. However, in recent years, bisphenol analogs such as bisphenol S (BPS) and bisphenol F (BPF) are replacing BPA in the production of some consumer products. Because human exposure to these alternative bisphenols may occur, biomonitoring of these bisphenol analogs is warranted. In the present study, we developed and validated a sensitive and selective method that uses on-line solid phase extraction coupled to high performance liquid chromatography-isotope dilution tandem mass spectrometry with peak focusing to measure BPA, BPF, BPS, and 11 other environmental phenols in urine. The method required a small amount of sample (100μL) and minimal sample pretreatment. The limits of detection were 0.03ng/mL (BPS), 0.06ng/mL (BPF), 0.10ng/mL (BPA), and ranged from 0.1ng/mL to 1.0ng/mL for the other 11 phenols. In 100 urine samples collected in 2009-2012 from a convenience group of anonymous adults in the United States, of the three bisphenols, we detected BPA at the highest frequency and median concentrations (95%, 0.72ng/mL), followed by BPS (78%, 0.13ng/mL) and BPF (55%, 0.08ng/mL). This sensitive, rugged, and labor and cost-effective method could be used for the analysis of large number of samples for epidemiologic studies.