Calculation and verification of blood ethanol measurement uncertainty for headspace gas chromatography

An estimate was made of the measurement uncertainty for blood ethanol testing by headspace gas chromatography. While uncertainty often focuses on compliance to a single threshold level (0.08 g/100 mL), the existence of multiple thresholds, related to enhanced sentencing, subject age, or commercial vehicle licensure, necessitate the use of an estimate with validity across multiple specification levels. The uncertainty sources, in order of decreasing magnitude, were method reproducibility, linear calibration, recovery, calibrator preparation, reference material, and sample preparation. A large set of reproducibility data was evaluated (n = 15,433) in order to encompass measurement variability across multiple conditions, operators, instruments, concentrations and timeframes. The relative, combined standard uncertainty was calculated as ±2.7%, with an expanded uncertainty of ±8.2% (99.7% level of confidence, k = 3). Bias was separately evaluated through a recovery study using standard reference material from a national metrology institute. The uncertainty estimate was verified through the use of proficiency test (PT) results. Assigned values for PT results and their associated uncertainties were calculated as robust means (x*) and standard deviations (s*) of participant values. Performance scores demonstrated that the uncertainty estimate was appropriate across the full range of PT concentrations (0.010-0.370 g/100 mL). The use of PT data as an empirical estimate of uncertainty was not examined. Until providers of blood ethanol PT samples include details on how an assigned value is obtained along with its uncertainty and traceability, the use of PT data should be restricted to the role of verification of uncertainty estimates.     

HPLC法测定血浆中万古霉素浓度的不确定度评定

目的：对人血浆万古霉素浓度高效液相色谱法（HPLC）测定的不确定度进行评定。方法：对HPLC法测定血浆中万古霉素浓度的全过程进行分析,对于分析过程中称量过程、溶液配制、萃取过程、仪器及标准曲线拟合等因素带来的不确定度进行逐一分析,用A类评定程序评定了分析过程中随机效应引起的不确定度．用B类评定程序评定了分析过程中其他因素引起的不确定度,根据合成不确定度的公式计算出合成不确定度并进行了扩展。结果：置信概率P为95％时．血浆中低（8．44μg／mL）、中（54．22μg／mL）、高（171．63μg／mL）浓度万古霉素的扩展不确定度分别是2．57μg／mL,7．66μg／mL．24．03μg／mL。结论：线性拟合过程是本实验中低浓度时不确定度的主要来源．而中、高浓度时标准曲线配制是不确定度的主要来源。本方法可用于HPLC法测定血浆中万古霉素浓度的不确定度评定．为药物分析过程中的不确定度评定提供了参考。     

HPLC-MS/MS 法测定人血浆中伏立康唑浓度的不确定度评价

目的 评价高效液相色谱 - 串联质谱法 (HPLC-MS/MS) 测定人血浆中伏立康唑浓度的不确定度。方法 分析 HPLCMS/MS 法测定人血浆中伏立康唑浓度过程中的不确定度来源,计算其大小并合成。结果 HPLC-MS/MS 法测定伏立康唑浓度 的不确定度在低浓度时主要由标准曲线拟合、样品提取和重复性引入,在高浓度时主要由仪器允差,重复性和样品提取引入。 人血浆中伏立康唑在低浓度(0.025μg/mL)、中浓度(1.5μg/mL)和高浓度(8.0μg/mL)的扩展不确定度分别为0.0018、0.2057和0.8723 ( 置信概率 P=95.45%, k=2)。结论 为有效减小测量结果的不确定度,建议在今后工作中应尽量避免过宽的标准曲线范围,增加 测定点的个数和重复测定次数,同时提高向处理后的基质中添加内标或质控溶液的加样技巧。     

UPLC-MS/MS测定大鼠尿样中26-OH-PD浓度的不确定度评定

目的 评定超高液相色谱-串联质谱法（UPLC-MS/MS）测定大鼠尿样中26-OH-PD浓度的不确定度。方法 分析UPLC-MS/MS法测定大鼠尿样中26-OH-PD浓度的不确定度来源,根据各分量计算出合成不确定度并进行了扩展。结果 大鼠尿样中低浓度（6.16 ng/mL）、中浓度（97.94 ng/mL）和高浓度（398.02ng/mL）26-OH-PD的扩展不确定度分别为1.44、6.97、22.05 ng/mL（P=95%,k=2）。结论 UPLC-MS/MS法测定大鼠尿样中26-OH-PD低浓度样品的不确定度主要分别由线性拟合引入,中、高浓度样品的不确定度主要由仪器允差引入。该法适用于评定UPLC-MS/MS法测定尿样中26-OH-PD的不确定度研究,能为复杂生物样品分析过程的不确定度评定提供一定参考。     

Antioxidant properties, total phenols and pollen analysis of propolis samples from Portugal

Pollen analysis, total phenols content and antioxidant activity were studied for the first time in Portuguese propolis samples from Bornes and Fundão regions. Total phenols content was determined by colorimetric assay and their amount was of 329 mg/g of GAE in Bornes sample and 151 mg/g of GAE in Fundão propolis. The antioxidant capacity of propolis extracts was assessed through the scavenging effects on DPPH (2,2-diphenyl-1-picrylhydrazyl) and reducing power of iron (III)/ ferricyanide complex assays. A concentration-dependent antioxidative capacity was verified in DPPH and reducing power assays. Low values of EC₅₀ on DPPH scavenging assay were obtained for Bornes and Fundão propolis (of 6.22 μg/mL and 52.00 μg/mL, respectively). For reducing power the values were 9.00 μg/mL, for Bornes propolis, and 55.00 μg/mL, for Fundão propolis. The high activity of propolis from Bornes could be related with their different pollen composition. The results obtained indicate that Portuguese propolis is an important source of total phenols showing antioxidant properties that could be beneficial for human health.     

气相色谱法测定非吸收性外科缝线中环氧乙烷残留量的不确定度分析

目的：对气相色谱法测定非吸收性外科缝线中环氧乙烷残留量的不确定度进行分析,以找出影响不确定度的因素。方法：建立气相色谱法测定非吸收性外科缝线中环氧乙烷残留量的数学模型,并根据《测量不确定度评定与表示》（JJF1059-1999）中有关规定,量化各不确定度分量,计算合成不确定度从而得出测定结果的扩展不确定度。结果：测定结果的合成不确定度为0.7μg·g^-1,扩展不确定度为1.4μg·g^-1,非吸收性外科缝线中环氧乙烷残留量为（27.5±1.4）μg·g^-1（k=2）。结论：测定结果的不确定度主要来源于拟合曲线的影响,故气相色谱仪的稳定性是测量准确与否的关键所在。建立的不确定度评估方法适用于气相色谱法测定非吸收性外科缝线中环氧乙烷残留量的不确定度分析。     

Inhibitory activity of α-amylase and α-glucosidase by plant extracts from the Brazilian cerrado

Diabetes mellitus is the most common disease in the world. One therapeutic approach for treating diabetes is inhibition of α-amylase and α-glucosidase activities to reduce postprandial blood glucose levels. In vitro tests showed that several plant extracts from Brazilian cerrado species can inhibit the activity of α-amylase and α-glucosidase. The extracts of Eugenia dysenterica, Stryphnodendron adstringens, Pouteria caimito, Pouteria ramiflora, and Pouteria torta showed strong α-amylase and α-glucosidase inhibitory activity. Eugenia dysenterica, P. caimito, P. ramiflora, and P. torta aqueous extracts exerted the highest activity against α-amylase (IC??) values of 14.93, 13.6, 7.08, and 5.67 μg/mL, respectively) and α-glucosidase (IC?? values of 0.46, 2.58, 0.35, and 0.22 μg/mL, respectively). Stryphnodendron adstringens ethanol extract also exhibited inhibitory activity against both enzymes (IC??) 1.86 μg/mL against α-amylase and 0.61 μg/mL against α-glucosidase). The results suggest that the activity of these cerrado plants on α-amylase and α-glucosidase represents a potential tool for development of new strategies for treatment of diabetes.     

Simultaneous determination of efavirenz, rifampicin and its metabolite desacetyl rifampicin levels in human plasma

A simple and rapid isocratic, high performance liquid chromatography (HPLC) assay employing solid phase extraction (SPE) for the simultaneous determination of the anti HIV drug, efavirenz, the anti-tuberculosis drug, rifampicin and the desacetyl metabolite of rifampicin in plasma from HIV/tuberculosis infected patients has been developed. Using a Zorbax SB-Phenyl reverse-phase analytical column with UV detection, good separation and detection of the drugs was attained within a 10 min run time. Intra- and inter-assay precision RSD values were found to be less than 15% at the concentrations examined (0.1-20 μg/mL). The LOQ was found to be 0.1 μg/mL for each agent and the assay was found to generate a linear response up to 20 μg/mL.This low cost assay can accurately detect efavirenz and rifampicin concentrations within a clinically relevant concentration range using standard chromatography equipment, making it particularly applicable to resource-limited settings.     

Biomonitoring equivalents for di-isononyl phthalate (DINP)

Biomonitoring Equivalents (BEs) are defined as the concentration or range of concentrations of a chemical or its metabolite in a biological medium (blood, urine, or other medium) that is consistent with an existing health-based exposure guideline such as a reference dose (RfD) or tolerable daily intake (TDI). BE values can be used as a screening tool for the evaluation of population-based biomonitoring data in the context of existing risk assessments. This study reviews available health based risk assessments and exposure guidance values for di-isononyl phthalate (DINP) from Health Canada, the United States Consumer Product Safety Commission (US CPSC), and the European Food Safety Authority (EFSA). Controlled dosing data reporting the urinary excretion fractions of major DINP metabolites following administration of labeled DINP are reviewed, and BE values corresponding to the available exposure guidance values are derived assuming chronic, steady-state intake and excretion at those exposure values. The BE values range from 1500 to 3600μg/L (1900-4600μg/g creatinine) based on the sum of three oxidative metabolites. Sources of uncertainty relating to both the basis for the BE values and their use in evaluation of biomonitoring data, including the transience of the biomarkers relative to exposure frequency, are discussed. The BE values derived here can be used as screening tools for evaluation of population biomonitoring data for DINP in the context of existing risk assessments and can assist in prioritization of the potential need for additional risk assessment efforts for DINP relative to other chemicals.     

GC-MS-MS determination of gamma-hydroxybutyrate in blood and urine

A gas chromatographic-tandem mass spectrometric (GC-MS-MS) method for determining trace concentrations of gamma-hydroxybutyrate (GHB) in blood and urine has been developed. Multiple reaction monitoring was used to detect parent and daughter ions of GHB, 233 and 147, respectively, following liquid-liquid extraction with acetonitrile and derivatisation with N,O-bis[trimethylsilyl]trifluoroacetamide (BSTFA). Deuterated GHB was used as an internal standard. The assay produced excellent linearity and sensitivity without conversion to gamma butyrolactone. The lower limit of quantitation (LLOQ) in 50 microL of sample was 2.5 microg/mL. The expanded uncertainty values for intra- and interassay results were +/- 0.097 and +/- 0.123 ng/mL at a confidence level of 95% for blood and urine, respectively. Endogenous concentrations of GHB were found to be in the range of 0.3 to 6 microg/mL in urine and 0.5 to 2.3 microg/mL in blood, confirming previously suggested cut-off values for forensic analysis.     

Free radical scavenging and antiacetylcholinesterase activities of Origanum majorana L. essential oil

In the present study, Origanum majorana L. essential oil (EO) was analyzed by gas chromatography-mass spectrometry (GC-MS) and evaluated for free radical scavenging and anticholinesterase activities. GC-MS analysis revealed the presence of 4-terpineol (29.97%), γ-terpinene (15.40%), trans-sabinene hydrate (10.93), α-terpinene (6.86%), 3-cycolohexene-1-1 methanal,a,a4-trimethyl-,(S)-(CAS) (6.54%), and sabinene (3.91%) as main constituents. Origanum majorana L. EO exhibited concentration-dependent inhibitory effects on 2,2'-diphenylpicrylhydrazyl (DPPH(?)), hydroxyl radical, hydrogen peroxide, reducing power, and lipid peroxidation with IC(50) values of 58.67, 67.11, 91.25, 78.67, and 68.75 μg/mL, respectively; while the IC(50) values for the standard trolox were noted to be 23.95, 44.97, 51.30, 42.22, and 52.72 μg/mL, respectively. Interestingly, cholinesterase inhibitory activity was also found with IC(50) values of 36.40 μg/mL. We can conclude that the marjoram EO has a significant potential to be used as a natural antioxidant and anti-AChE.     

HPLC法测定血浆中法罗培南浓度的不确定度分析

目的:对HPLC法测定血浆中法罗培南浓度的测量不确定度进行分析,以找到影响不确定度的因素,为评价检测报告提供科学依据。方法:根据《测量不确定度评定与表示》,并参考《化学分析中不确定度的评估指南》,对血浆中法罗培南含量测定进行测量不确定度的分析与评定。结果:标准不确定度为1.28μg·ml^-1,扩展不确定度为2.56μg·ml^-1,含量测定结果可表示为（10.14±2.56）μg·ml^-1。结论:建立的不确定度评估方法可用于HPLC法测定血浆中法罗培南含量的不确定度评估。测量不确定度的评定方法的确立对于血浆中法罗培南含量测定方法标准的研究具有重要意义。     

In vitro assessment of selected Korean plants for antioxidant and antiacetylcholinesterase activities

Context: Antiacetylcholinesterase (AChE) drugs have been a main therapeutic treatment for Alzheimer’s disease because increased AChE levels play a key role in reducing neurotransmission.

Objectives: Extracts from 35 Korean plants were selected and screened for antioxidant and anti-cholinesterase activity to explore new sources derived from Korean natural resources that could be used as AD therapeutic agents.

Materials and methods: The antioxidant effect of extracts from 35 selected Korean plants was determined using two most common free radical scavenging assays using 1,1-diphenyl-2-picrylhydrazyl (DPPH) and 2,2′-azino-bis-3-ethylbenzthiazoline-6-sulphonic acid (ABTS). Additionally, the effect of extracts, identified as antioxidants, on acetylcholinesterase inhibition was assessed by an acetylcholinesterase assay kit.

Results: Out of 36 extracts of 35 plants tested, Oenothera biennis L. (9.09?μg/mL), Saururus chinensis (Lour.) Baill. (9.52?μg/mL) and Betula platyphylla var. japonica (9.85?μg/mL) showed strong DPPH scavenging activity. Twelve other extracts also exerted moderate free radical scavenging activities with IC₅₀ values ranging from 10 to 50?μg/mL. Antioxidant capacity detected by ABTS assay was only significant in O. biennis (23.40?μg/mL), while the other extracts were weak or unable to reduce the production of ABTS. Based on the antioxidant activities of these plant extracts, 19 extracts with IC₅₀ values less than 100?μg/mL in DPPH assay were selected for further AChE inhibition assay. Among the extracts tested, the IC₅₀ value for Prunella vulgaris var. lilacina NAKAI (18.83?μg/mL) in AChE inhibitory activity was the lowest, followed by O. biennis (20.09?μg/mL) and Pharbitis nil Chosy (22.79?μg/mL).

Conclusions: Considering complex multifactorial etiology of AD, the extracts of P. vulgaris var. lilacina (aerial part), O. biennis (seed) and P. nil (seed) may be safe and ideal candidates for future AD modifying therapies.     

Pharmacokinetics of ceftriaxone in liver-transplant recipients

The disposition of ceftriaxone was studied after a single 2 g intravenous dose in seven patients 3 to 5 days after liver transplantation. Ceftriaxone concentrations in plasma, urine, and bile were measured by HPLC, and plasma protein binding was determined by equilibrium dialysis. Plasma protein binding was nonlinear, and the unbound fraction varied between 0.05 and 0.56. Both capacity and affinity were markedly different from reported values for normal subjects. The pharmacokinetic parameters obtained were: total body clearance (TBC), 11.2 +/- 7.8 mL/hr/kg total and 44.8 +/- 29.1 mL/hr/kg unbound; volume of distribution (V(area)), 224 +/- 76 mL/kg total and 767 +/- 432 mL/kg unbound; steady-state volume of distribution (Vss), 212 +/- 68 mL/kg total and 651 +/- 368 mL/kg unbound; terminal disposition half-life (t1/2), 21.6 +/- 14.3 hour total and 16.3 +/- 11.1 hour unbound. TBC for both total and free drug was considerably lower than literature values for normal subjects. V(area) for total drug was greater than normal, whereas the corresponding value for free drug was smaller than normal. The plasma ceftriaxone concentrations at 12 and 24 hours were above the reported minimum inhibitory concentration (MIC). The fraction of the administered dose excreted in urine over 24 hours was 38 +/- 29% and did not differ markedly from that reported for normal subjects. Less than 2% of the administered dose was excreted in 24-hour bile; however, biliary concentrations were always above MIC. Ceftriaxone can be administered once or twice daily at a dose of 2 g/day for prophylaxis in liver transplant recipients.     

Science-policy in environmental and health risk assessment: if we cannot do without, can we do better?

How can empirical evidence of adverse effects from exposure to noxious agents, which is often incomplete and uncertain, be used most appropriately to protect human health? We examine several important questions on the best uses of empirical evidence in regulatory risk management decision-making raised by the US Environmental Protection Agency (EPA)'s science-policy concerning uncertainty and variability in human health risk assessment. In our view, the US EPA (and other agencies that have adopted similar views of risk management) can often improve decision-making by decreasing reliance on default values and assumptions, particularly when causation is uncertain. This can be achieved by more fully exploiting decision-theoretic methods and criteria that explicitly account for uncertain, possibly conflicting scientific beliefs and that can be fully studied by advocates and adversaries of a policy choice, in administrative decision-making involving risk assessment. The substitution of decision-theoretic frameworks for default assumption-driven policies also allows stakeholder attitudes toward risk to be incorporated into policy debates, so that the public and risk managers can more explicitly identify the roles of risk-aversion or other attitudes toward risk and uncertainty in policy recommendations. Decision theory provides a sound scientific way explicitly to account for new knowledge and its effects on eventual policy choices. Although these improvements can complicate regulatory analyses, simplifying default assumptions can create substantial costs to society and can prematurely cut off consideration of new scientific insights (e.g., possible beneficial health effects from exposure to sufficiently low 'hormetic' doses of some agents). In many cases, the administrative burden of applying decision-analytic methods is likely to be more than offset by improved effectiveness of regulations in achieving desired goals. Because many foreign jurisdictions adopt US EPA reasoning and methods of risk analysis, it may be especially valuable to incorporate decision-theoretic principles that transcend local differences among jurisdictions.     

New issues in cancer risk assessment

When a nonlinear dose-response at low doses can be justified, an acceptable daily intake for a carcinogen can be obtained by dividing a benchmark dose, associated with a low incidence of tumors in animals, by uncertainty factors to account for animal-to-human extrapolation, human variability, and risk reduction from a low observed adverse-effect level. This approach can utilize mechanistic information to justify smaller uncertainty factors than typical default values of 10. If a nonlinear dose-response cannot be justified, traditional linear extrapolation from the benchmark dose to zero sometimes gives similar results. This suggests a unified risk-assessment procedure based on uncertainty factors. The issue of cross-species extrapolation based on the risk relative to background risks, rather than excess risk, is examined. The relative risk approach reduces the estimates of cancer risk in humans based on common rodent tumors, such as the liver in some strains of mice.     

Antimicrobial, antimalarial, and antileishmanial activities of mono- and bis-quaternary pyridinium compounds

Pyridinium-based oxime compounds have been utilized worldwide as antidotes following exposure to anticholinesterase agents. In the event of combined chemical and biological incident, it is of vital importance to know the ability of antidotes to provide additional protection against biological threats. This paper reports results of in vitro antimicrobial and antiprotozoal activities of a series of quaternary pyridinium oximes against a number of lower pathogenicity BSL-1 and 2 agents. In general, our compound panel had little to no antimicrobial action except for thiophene- and benzothiophene-substituted monoquaternary pyridinium compounds 21 and 24 that showed moderate antibacterial activity against Staphylococcus aureus and methicillin-resistant S. aureus with IC(50) values ranging from 12.2 to 17.7 μg/mL. Compounds 21 and 24 also exhibited antileishmanial activity against Leishmania donovani with IC(50) values of 19 and 18 μg/mL, respectively. Another monoquaternary pyridinium compound with a bromobutyl side chain 17 showed antimalarial activity against both a chloroquine sensitive and resistant strains of Plasmodium falciparum with IC(50) values of 3.7 and 4.0 μg/mL, respectively. None of the bisquaternary pyridinium compounds showed antimicrobial or antiprotozoal activity. None of the compounds showed cytotoxic effects toward mammalian kidney fibroblasts. Results of this study indicate that the pyridinium compounds, some of which are already in use as antidotes, do not have significant antimicrobial and antiprotozoal activities and cannot be relied upon for additional protection in the event of combined chemical-biological incident.     

Isolation,characterization and antioxidant activities of the endophytic fungi from Hosta. Ventricosa

A total of 13 types of endophytic fungi, classified into six genera, were isolated from Hosta. ventricosa based on 18S rDNA sequencing of ITS region and microscopic examination. The antioxidant activities of the crude extracts were investigated by T-AOC, DPPH and ABTS⁺ methods. The results showed that the crude extracts from Fusarium oxysporum (HoV1) and Epicoccum (HoV4), which presented higher phenol levels (1.90±0.03 mg/g, and 2.41±0.01 mg/g, respectively), exhibited stronger comprehensive antioxidant activities, with higher T-AOC values (63.46±0.26 U/g, and 66.05±0.71 U/g, respectively), lower EC₅₀ values against ABTS⁺ (30.25±0.05 μg/mL, and 24.66±0.65 μg/mL, respectively)and DPPH radicals (83.42±0.24 μg/mL, and 59.136 μg/mL, respectively). Moreover, Fusarium oxysporum (HoV8) showed excellent scavenging activity against DPPH radicals (EC₅₀ = 16.02 μg/mL), which was better than BHT. All of them could be useful in the developing of new natural antioxidant agents.     

Application of qualitative and quantitative uncertainty assessment tools in developing ranges of plausible toxicity values for 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin

Increasing interest in characterizing risk assessment uncertainty is highlighted by recent recommendations from the National Academy of Sciences. In this paper we demonstrate the utility of applying qualitative and quantitative methods for assessing uncertainty to enhance risk‐based decision‐making for 2,3,7,8‐tetrachlorodibenzo‐p‐dioxin. The approach involved deconstructing the reference dose (RfD) via evaluation of the different assumptions, options, models and methods associated with derivation of the value, culminating in the development of a plausible range of potential values based on such areas of uncertainty. The results demonstrate that overall RfD uncertainty was high based on limitations in the process for selection (e.g., compliance with inclusion criteria related to internal validity of the co‐critical studies, consistency with other studies), external validity (e.g., generalizing findings of acute, high‐dose exposure scenarios to the general population), and selection and classification of the point of departure using data from the individual studies (e.g., lack of statistical and clinical significance). Building on sensitivity analyses conducted by the US Environmental Protection Agency in 2012, the resulting estimates of RfD values that account for the uncertainties ranged from ~1.5 to 179 pg/kg/day. It is anticipated that the range of RfDs presented herein, along with the characterization of uncertainties, will improve risk assessments of dioxins and provide important information to risk managers, because reliance on a single toxicity value limits the information needed for making decisions and gives a false sense of precision and accuracy.     

Analysis of ritonavir in plasma/serum and tissues by high-performance liquid chromatography

A method has been developed to quantify ritonavir concentrations in human plasma and in mouse serum, liver, and brain using high-performance liquid chromatography. Extraction recoveries for ritonavir and its internal standard averaged greater than 95%. Within-day variability, expressed as a coefficient of variation, averaged 6% over the concentration range 0.5 μg/mL to 15 μg/mL ritonavir, and between-day variability averaged 5.6% over 5 μg/mL to 15 μg/mL ritonavir. The method was applied to quantitation of ritonavir in mouse serum and tissue. Measured values deviated less than 5% from the actual values in mouse serum, liver, and brain samples containing 5 μg/mL ritonavir. The slopes of calibration curves for extracted calf serum, mouse serum, mouse liver and mouse brain standards were nearly identical to the calibration slope of standards which were not extracted. All curves were linear through zero, and r² was no less than 0.998 for any form of calibration. In addition, there was no chromatographic interference from commonly prescribed medications.