IPEM code of practice for high-energy photon therapy dosimetry based on the NPL absorbed dose calibration service.

The 1990 code of practice (COP), produced by the IPSM (now the Institute of Physics and Engineering in Medicine, IPEM) and the UK National Physical Laboratory (NPL), gave instructions for determining absorbed dose to water for megavoltage photon (MV) radiotherapy beams (Lillicrap et al 1990). The simplicity and clarity of the 1990 COP led to widespread uptake and high levels of consistency in external dosimetry audits. An addendum was published in 2014 to include the non-conventional conditions in Tomotherapy units. However, the 1990 COP lacked detailed recommendations for calibration conditions, and the corresponding nomenclature, to account for modern treatment units with different reference fields, including small fields as described in IAEA TRS483 (International Atomic Energy Agency (IAEA) 2017, Vienna). This updated COP recommends the irradiation geometries, the choice of ionisation chambers, appropriate correction factors and the derivation of absorbed dose to water calibration coefficients, for carrying out reference dosimetry measurements on MV external beam radiotherapy machines. It also includes worked examples of application to different conditions. The strengths of the 1990 COP are retained: recommending the NPL2611 chamber type as secondary standard; the use of tissue phantom ratio (TPR) as the beam quality specifier; and NPL-provided direct calibration coefficients for the user's chamber in a range of beam qualities similar to those in clinical use. In addition, the formalism is now extended to units that cannot achieve the standard reference field size of 10 cm × 10 cm, and recommendations are given for measuring dose in non-reference conditions. This COP is designed around the service that NPL provides and thus it does not require the range of different options presented in TRS483, such as generic correction factors for beam quality. This approach results in a significantly simpler, more concise and easier to follow protocol.

Solid Sampling in Analysis of Various Plants Using Two-Jet Plasma Atomic Emission Spectrometry.

The possibility of two-jet plasma atomic emission spectrometry for analysis of different plants using solid sample preparation and unified calibration samples was investigated. The certified reference materials of wheat, maize, rice, potato, grass mix, birch leaves, and Elodea canadensis were used for analysis. On the basis of the behavior of these plants in the plasma, they were divided into two groups: starch-containing materials (cereal and root crops) and leaves/grass. It was found that the previous sample carbonization should be used for analysis of starch-containing plants while leaves and grass could be analyzed by the direct technique. Carbonization was only applied for determining low concentrations of trace elements in leaves and grass. The calibration samples based on graphite powder and simple sample preparation, dilution of powdered sample with a spectroscopic buffer, were used for both direct analysis and analysis after carbonization. Such an approach allowed estimation of B, Ba, Be, Cd, Co, Cr, Cu, Ga, Fe, Mn, Ni, Pb, Si, Sr, V, and Zn in different plants. The limits of detection (LODs) provided by the direct technique were at the level of (µg·g-1): n × 0.1 for Cd, Cu, and Mn; n for B, Ba, Co, Cr, Fe, Ga, Ni, Pb, Sr, V, and Zn; n × 10 for Si. Carbonization allowed improving LODs of elements several times depending on the thermal stability and mineral composition of plants. The LODs of elements in plants obtained after carbonization are the following (µg·g-1): n × 0.01 for Be, Cd, Cu, and Mn; n × 0.1 for Co, Cr, Fe, Ga, Ni, Pb, Sr, V, Zn; and n for Si. The techniques suggested are fast, easily workable, and do not require harmful chemical reagents. In some cases, the influence of variable matrices and different element species on analytical signal of elements was not completely suppressed; the deviation of element concentrations from the true values was discussed.

New simple and low-cost methods for periodic checks of Cyclone® Plus Storage Phosphor System.

The recent large use of the Cyclone® Plus Storage Phosphor System, especially in European countries, as imaging system for quantification of radiochemical purity of radiopharmaceuticals raised the problem of setting the periodic controls as required by European Legislation. We described simple, low-cost methods for Cyclone® Plus quality controls, which can be useful to evaluate the performance measurement of this imaging system.

Progress in standardizing and harmonizing thyroid function tests.

Iodine is an essential component of thyroid hormone. Because thyroid hormone synthesis is affected by iodine deficiency on the one hand and by excess iodine intake on the other, thyroid function biomarkers may be useful for assessing iodine status and studying the effects of iodine supplementation. However, reference intervals for some of the most useful thyroid function biomarkers, including serum concentrations of thyroid-stimulating hormone (TSH), free thyroxine (FT4), and thyroglobulin, vary widely due to variability in the commercially available immunoassays for these tests. Recognizing the need for standardization of thyroid function testing, the International Federation of Clinical Chemistry and Laboratory Medicine established a working group, later restructured as the Committee for Standardization of Thyroid Function Tests, to examine its feasibility. The committee has established a conventional reference measurement procedure for FT4 and an approach to harmonization of results for TSH. Panels of single-donation human blood specimens that span the measuring interval of the immunoassays were used to assess the performance of commercially available immunoassays and form the basis for their recalibration. Recalibration of the manufacturers' methods for both FT4 and TSH has shown that the variability among immunoassays can be successfully eliminated for euthyroid individuals as well as for patients with thyroid disease. The committee is not investigating the standardization of thyroglobulin at the present time.

A round-robin gamma stereotactic radiosurgery dosimetry interinstitution comparison of calibration protocols.

PURPOSE: Absorbed dose calibration for gamma stereotactic radiosurgery is challenging due to the unique geometric conditions, dosimetry characteristics, and nonstandard field size of these devices. Members of the American Association of Physicists in Medicine (AAPM) Task Group 178 on Gamma Stereotactic Radiosurgery Dosimetry and Quality Assurance have participated in a round-robin exchange of calibrated measurement instrumentation and phantoms exploring two approved and two proposed calibration protocols or formalisms on ten gamma radiosurgery units. The objectives of this study were to benchmark and compare new formalisms to existing calibration methods, while maintaining traceability to U.S. primary dosimetry calibration laboratory standards. METHODS: Nine institutions made measurements using ten gamma stereotactic radiosurgery units in three different 160 mm diameter spherical phantoms [acrylonitrile butadiene styrene (ABS) plastic, Solid Water, and liquid water] and in air using a positioning jig. Two calibrated miniature ionization chambers and one calibrated electrometer were circulated for all measurements. Reference dose-rates at the phantom center were determined using the well-established AAPM TG-21 or TG-51 dose calibration protocols and using two proposed dose calibration protocols/formalisms: an in-air protocol and a formalism proposed by the International Atomic Energy Agency (IAEA) working group for small and nonstandard radiation fields. Each institution's results were normalized to the dose-rate determined at that institution using the TG-21 protocol in the ABS phantom. RESULTS: Percentages of dose-rates within 1.5% of the reference dose-rate (TG-21+ABS phantom) for the eight chamber-protocol-phantom combinations were the following: 88% for TG-21, 70% for TG-51, 93% for the new IAEA nonstandard-field formalism, and 65% for the new in-air protocol. Averages and standard deviations for dose-rates over all measurements relative to the TG-21+ABS dose-rate were 0.999±0.009 (TG-21), 0.991±0.013 (TG-51), 1.000±0.009 (IAEA), and 1.009±0.012 (in-air). There were no statistically significant differences (i.e., p>0.05) between the two ionization chambers for the TG-21 protocol applied to all dosimetry phantoms. The mean results using the TG-51 protocol were notably lower than those for the other dosimetry protocols, with a standard deviation 2-3 times larger. The in-air protocol was not statistically different from TG-21 for the A16 chamber in the liquid water or ABS phantoms (p=0.300 and p=0.135) but was statistically different from TG-21 for the PTW chamber in all phantoms (p=0.006 for Solid Water, 0.014 for liquid water, and 0.020 for ABS). Results of IAEA formalism were statistically different from TG-21 results only for the combination of the A16 chamber with the liquid water phantom (p=0.017). In the latter case, dose-rates measured with the two protocols differed by only 0.4%. For other phantom-ionization-chamber combinations, the new IAEA formalism was not statistically different from TG-21. CONCLUSIONS: Although further investigation is needed to validate the new protocols for other ionization chambers, these results can serve as a reference to quantitatively compare different calibration protocols and ionization chambers if a particular method is chosen by a professional society to serve as a standardized calibration protocol.

[Measurement of peak correction factor of Farmer chamber for calibration of flattening filter free (FFF) clinical photon beams]. / Kiegyenlítõ szûrõ nélküli mezõk Farmer-kamrával történõ kalibrációjánál alkalmazandó csúcshatás-korrekció mérése.

Farmer-type ionization chambers are considered the most reliable detectors and for this reason they are most frequently used for the calibration of photon beams of medical linear accelerators. Flattening filter free (FFF) photon beams of linear accelerators have recently started to be used in radiotherapy. The dose profile of FFF beams is peaked in the center of the field and the dose distribution will be inhomogeneous along the axis of the 2.3 cm long measuring volume of the Farmer chamber. The peaked radiation field will result in volume averaging effects in the large Farmer chamber, therefore this chamber will underestimate the true central axis dose. Our objective was to determine the value of the peak correction factor (Kp) of Farmer-type chamber with measurements to avoid the underestimation of the central axis dose during the calibration of FFF radiation fields. Measurements were made with 6 MV and 10 MV flattened (6X and 10X) and FFF beams (6XFFF and 10XFFF) of a Varian TrueBeam medical linear accelerator in a solid water phantom at 10 cm depth. The source surface distance (SSD) was 100 cm, the field size was 10×10 cm and the dose rate was always 400 MU/min during the measurements. We delivered 100 MU in each measurement and the absorbed dose to water was calculated according to the IAEA TRS-398 dosimetry protocol. The measured signals of the ionization chambers were always corrected for the ion recombination loss. The ion recombination correction factors (Kr) were determined with the two-voltage method separately for the used ion chambers and for flattened and unflattened beams. First, we measured the dose to water with PTW TM30012 Farmer chamber in 6XFFF and 6X beams, then calculated the ratio of doses of 6XFFF and 6X beams (R6,Farmer). Immediately after this we repeated the above measurements with PTW TM31010 Semiflex chamber and determined the ratio of doses of 6XFFF and 6X beams again (R6,Semiflex). The length of the sensitive volume of the Semiflex chamber is only 6.5 mm. According to our dose profile measurements the peak correction factor of this chamber equals to unity for both photon energies. As a consequence R6,Semiflex is larger than R6,Farmer and Kp6XFFF = R6,Semiflex / R6,Farmer, where Kp6XFFF is the peak correction factor of the Farmer chamber in 6XFFF beam. The advantage of this method is that we have to calculate ratio of doses, so it is not necessary to know the calibration factors of the chambers. Repeating the above measurements with 10X and 10XFFF beams we determined the peak correction factor of Farmer chamber for 10XFFF beam, too (Kp10XFFF). According to our measurements Kp6XFFF = 1.0025 and Kp10XFFF = 1.009. The bigger peak correction factor for 10XFFF beam is in accordance with the fact that the peak of dose profile is steeper for higher photon energy. The above described method for the determination of Kp can be used for other photon energies and other large volume ionization chambers.

Standardization of autoantibody testing: a paradigm for serology in rheumatic diseases.

Autoantibody measurement is an excellent tool to confirm the diagnosis of rheumatic autoimmune diseases. Hence, reliability and harmonization of autoantibody testing are essential, but these issues are still a matter of debate. Intrinsic variability in analytes and reagents as well as heterogeneity of the techniques are the main reasons for discrepancies in inter-laboratory variations and reporting of test results. This lack of reliability might be responsible for wrong or missed diagnoses, as well as additional costs due to assay repetition, unnecessary use of confirmatory tests and/or consequent diagnostic investigations. To overcome such issues, the standardization of autoantibody testing requires efforts on all aspects of the assays, including the definition of the analyte, the pre-analytical stages, the calibration method and the reporting of results. As part of such efforts, the availability of suitable reference materials for calibration and quality control would enable the development of a reliable reference system. Strong-positive sera from patients have been used as reference materials in most of the autoantibody assays for rheumatic diseases; however, antigen-affinity-purified immunoglobulin fractions or in some cases reliable monoclonal antibody preparations offer more adequate tools for standardization. Systematic assessments of reference materials are currently underway, and preliminary results appear to be encouraging.

Effect of calibration method on distortion-product otoacoustic emission measurements at and around 4 kHz.

OBJECTIVES: Distortion-product otoacoustic emissions (DPOAEs) collected after sound pressure level (SPL) calibration are susceptible to standing waves that affect measurements at the plane of the probe microphone due to overlap of incident and reflected waves. These standing-wave effects can be as large as 20 dB, and may affect frequencies both above and below 4 kHz. It has been shown that forward pressure level (FPL) calibration minimizes standing-wave effects by isolating the forward-propagating component of the stimulus. Yet, previous work has failed to demonstrate more than a small difference in test performance and behavioral-threshold prediction with DPOAEs after SPL and FPL calibration. One potential limitation in prior studies is that measurements were restricted to octave and interoctave frequencies; as a consequence, data were not necessarily collected at the standing-wave null frequency. In the present study, DPOAE responses were measured with f2 set to each participant's standing-wave frequency in an effort to increase the possibility that differences in test performance and threshold prediction would be observed for SPL and FPL calibration methods. DESIGN: Data were collected from 42 normal-hearing participants and 93 participants with hearing loss. DPOAEs were measured with f2 set to 4 kHz and at each participant's notch frequency after SPL and FPL calibration. DPOAE input/output functions were obtained from -10 to 80 dB in 5 dB steps for each calibration/stimulus condition. Test performance was evaluated using clinical decision theory. Both area under receiver operating characteristic curves for all stimulus levels and cumulative distributions when L2 = 50 dB (a level at which the best performance was observed regardless of calibration method) were used to evaluate the accuracy with which auditory status was determined. A bootstrap procedure was used to evaluate the significance of the differences in test performance between SPL and FPL calibrations. DPOAE predictions of behavioral threshold were evaluated by correlating actual behavioral thresholds and predicted thresholds using a multiple linear regression model. RESULTS: First, larger DPOAE levels were measured after SPL calibration than after FPL calibration, which demonstrated the expected impact of standing waves. Second, for both FPL and SPL calibration, test performance was best for moderate stimulus levels. Third, differences in test performance between calibration methods were evident at low- and high-stimulus levels. Fourth, there were small but statistically significant improvements in test performance after FPL calibration for clinically relevant conditions. Fifth, calibration method had no effect on threshold prediction. CONCLUSIONS: Standing waves after SPL calibration have an impact on DPOAE levels. Although the effect of calibration method on test performance was small, test performance was better after FPL calibration than after SPL calibration. There was no effect of calibration method on predictions of behavioral threshold.

[Development of the 60Co gamma-ray standard field for therapy-level dosimeter calibration in terms of absorbed dose to water (N(D,w))].

A primary standard for the absorbed dose rate to water in a 60Co gamma-ray field was established at National Metrology Institute of Japan (NMIJ) in fiscal year 2011. Then, a 60Co gamma-ray standard field for therapy-level dosimeter calibration in terms of absorbed dose to water was developed at National Institute of Radiological Sciences (NIRS) as a secondary standard dosimetry laboratory (SSDL). The results of an IAEA/WHO TLD SSDL audit demonstrated that there was good agreement between NIRS stated absorbed dose to water and IAEA measurements. The IAEA guide based on the ISO standard was used to estimate the relative expanded uncertainty of the calibration factor for a therapy-level Farmer type ionization chamber in terms of absorbed dose to water (N(D,w)) with the new field. The uncertainty of N(D,w) was estimated to be 1.1% (k = 2), which corresponds to approximately one third of the value determined in the existing air kerma field. The dissemination of traceability of the calibration factor determined in the new field is expected to diminish the uncertainty of dose delivered to patients significantly.

Shrink-induced single-cell plastic microwell array.

The ability to interrogate and track single cells over time in a high-throughput format would provide critical information for fundamental biological understanding of processes and for various applications, including drug screening and toxicology. We have developed an ultrarapid and simple method to create single-cell wells of controllable diameter and depth with commodity shrink-wrap film and tape. Using a programmable CO(2) laser, we cut hole arrays into the tape. The tape then serves as a shadow mask to selectively etch wells into commodity shrink-wrap film by O(2) plasma. When the shrink-wrap film retracts upon briefly heating, high-aspect plastic microwell arrays with diameters down to 20 µm are readily achieved. We calibrated the loading procedure with fluorescent microbeads. Finally, we demonstrate the utility of the wells by loading fluorescently labeled single human embryonic stem cells into the wells.

Correlation between chromatographic fingerprint and antioxidant activity of Turnera diffusa (Damiana).

In the present contribution, the partial least squares (PLS) method was used to establish a correlation between the antioxidant activity (obtained by DPPH assay) and chromatographic profiles of TURNERA DIFFUSA extracts. Chromatograms were obtained using HPLC-DAD. A model was constructed using 40 samples with 2550 X variables corresponding to the responses obtained at different times; the Y variables consisted of experimental values of antioxidant activity of each extract (measured as EC50). Prior to this analysis, alignment of chromatograms was performed based on consideration of seven high-intensity signals present in all samples. The PLS1 model was validated by cross-validations; its capacity was evaluated using correlation parameters R², root mean square error of calibration (RMSEC), and root mean square error of prediction (RMSEP). The best results were achieved with zero order chromatograms using five-point smoothing (R² = 0.96, RMSEC = 3.31, and RMSEP = 7.86). Under these conditions, the optimal number of components was five. The model was applied to the prediction of antioxidant activity of commercial products; no significant differences were found between the experimental and predicted antioxidant activities for 83 % of them.

Separation and determination of four ganoderic acids from dried fermentation mycelia powder of Ganoderma lucidum by capillary zone electrophoresis.

Ganoderic acids (GAs) were bioactive secondary metabolites produced by a traditional mushroom Ganoderma lucidum. We describe a simple and efficient method for the separation and quantitative determination of four GAs, namely Ganoderic acid T (GA-T), Ganoderic acid Mk (GA-Mk), Ganoderic acid Me (GA-Me) and Ganoderic acid S (GA-S) from dried triterpene-enriched extracts of G. lucidum mycelia powder by capillary zone electrophoresis (CZE). Under the optimum conditions, the four GAs reached the baseline separation in 9 min with Glycyrrhetinic acid (GTA) as internal standard. The four GAs and internal standard (GTA) were detected at a wavelength 245 nm. All calibration curves showed good linearity (r(2)>0.9958) within test ranges. Limit of detection (LOD) and limit of quantification (LOQ) were less than 0.6 and 1.8 microg/mL, respectively. The relative standard deviation (R.S.D.) values of precision and recoveries were less than 5% and recoveries ranged from 91.4% to 103.6%. This was the first report on simultaneous determination of the four GAs and the results provided a firm basis for the trace analysis of GAs in dried fermentation mycelia powder of G. lucidum with high accuracy.

Long-term fade study of the DT-702 LiF: Mg,Cu,P TLD.

LiF thermoluminescent dosemeters (TLDs) are used by the US Navy to record radiation exposure of personnel. The Model DT-648 LiF:Mg,Ti TLD has been replaced by a new Model DT-702 LiF:Mg,Cu,P TLD. The DT-648 was used for many years and has undergone extensive testing to identify its pre- and post-irradiation fade operating characteristics. Studies have shown that the addition of copper increases the thermoluminesence sensitivity of the TLD for improved low-level radiation monitoring. This study evaluates various fading characteristics of the new copper-doped dosemeter using current equipment for processing of TLDs and calibrating to a National Institute of Standards and Technology standard source. The 57-week study took place at the Naval Dosimetry Center, Bethesda, MD, USA. TLDs were stored for various lengths of time before and after being exposed to a National Institute of Standards and Technology calibrated radiation sources. TLDs were then processed using current US Navy instructions and the resulting dose compared with the calibrated exposure. Both loss of signal and loss of sensitivity were evaluated. The results of this study have shown that the DT-702 TLD has no statistically significant change in sensitivity or change in signal with up to 57 weeks of pre- or post-irradiation time. The results of this study will increase the accuracy of exposure record keeping for the Navy and will allow longer issue periods. This will increase flexibility with international and domestic shipping procedures, as well as reduce workload requirements for dosimetry processing.

Orthogonal field calibration analysis for myocardial electrode arrays used in defibrillation studies.

Mapping the myocardial electric field during a defibrillation pulse requires the recording of potential differences between electrodes. The field is then calculated from these quantities and the corresponding calibration matrix. One straightforward calibration technique involves alignment of a known electric field along each of the orthogonal axes of an electrode array and recording the resulting potential differences. However, no results have been reported to support the efficacy of this technique. This study performs a detailed error analysis including a one-to-one comparison to a precision calibration technique, and quantitatively establishes the efficacy of the orthogonal field technique.

Detection and quantification of soy allergens in food: study of two commercial enzyme-linked immunosorbent assays.

The objective of this study was to evaluate the efficacy of 2 commercially available soy enzyme-linked immunosorbent assays (ELISA) and use them in detecting soy proteins in selected food commodities. Both ELISA kits exhibited high sensitivity. The determined limits of detection (LOD) (approximately 2 and <1 microg/mL for Tepnel Biosystems and Elisa Systems kits, respectively) were lower than those claimed by the manufacturer. Quantification range for both kits was, however, narrower and in a lower concentration range than defined by the kit providers. Our examination revealed a positive cross-reactivity with chickpea proteins and matrix interferences for both kits. The immunoreactivity of soy proteins, when tested by the Tepnel Biosystems kit, was partially reduced by papain and bromelain hydrolysis; it was significantly decreased by protein glycation (>47%). Nondenatured and nonheated soy protein isolate (SPI) samples were also significantly less antigenic than the treated ones.

Call for calibration standard for newborn screening using auditory brainstem responses.

The mode of stimulation employed in newborn screening of the auditory brainstem response has evolved from the clinically standardized supraaural earphone to the tubal insert earphone, to most recently a circumaural earphone developed for this test. Considered here is the need to develop a standard for calibration of such devices for newborn screening applications, in particular. At risk is the prospect of missing the milder degrees of hearing loss, assuming a goal of detecting all clinically-significant congenital hearing losses. Two commercially manufactured test instruments for automated newborn screening were scrutinized via bench testing of sound output from their respective transducers, using a variety of measurements. By convention or design, none of the measurement approaches involved a model of the newborn ear, per se. While it was concluded that the manufacturers' method shows promise, namely as a relatively simple and potentially reliable method of calibration, concerns arose regarding output levels when measured according to both the manufacturers' and the authors' methods. Further work is needed to critically assess calibration methods and to establish, to the extent possible, appropriate norms and validation studies in newborns to provide a better understanding of the actual sound pressure level of the screening stimulus.

HPLC determination of safflor yellow A and three active isoflavones from TCM Naodesheng in rat plasma and tissues and its application to pharmacokinetic studies.

A high-performance liquid chromatographic method was developed for the simultaneous determination and pharmacokinetic studies of safflor yellow A, puerarin, 3'-methoxyl-puerarin, and puerarinapioside in the plasma and tissues of rats that had been administered with the traditional Chinese medicine (TCM) preparation Naodesheng via the caudal vein. Samples taken from rats were subjected to protein precipitation with acetone. Separation of these four compounds was accomplished on a Kromisil C18 stationary phase using a mobile phase of acetonitrile-0.1% phosphoric acid-tetrahydrofuran (8:92:2, v/v/v) at a flow-rate of 1.0 mL/min. The detection wavelength was set at 250 nm. The calibration curves of the four components were linear in the given concentration ranges. The intra- and inter-day precisions in plasma and tissues were less than 15% and the extraction recoveries were higher than 60%. The lower limits of quantitation of four components were low enough to determine the four components. These four components all exhibited kinetics that fitted a two-compartment model in rats. The elimination half-life was 1.19 h for safflor yellow A, 2.69 h for puerarin, 2.94 h for 3'-methoxyl-puerarin, and 0.87 h for puerarinapioside, respectively. Following administration of a single injection of Naodesheng, the concentration (C) of the four components in the tissues showed C(kidney) > C(lung), C(liver) > C(spleen), C(stomach), C(heart), approximately. The method is a reliable tool for performing studies of safflor yellow A and three puerarin isoflavones in different biological material.

Capillary electrophoretic method for the determination of inorganic and organic anions in real samples. Strategies for improving repeatability and reproducibility.

Selectivity and robustness of the pyromellitic acid (PMA) based background electrolyte was improved in order to increase its applicability for routine analysis of inorganic and organic anions in real samples. An electrolyte composed of 6.75 mM PMA, 0.5 mM hexamethonium hydroxide as electroosmotic flow (EOF) modifier, Ca(2+) 0.05 mM as complexation agent and pH adjusted to 7.6 with TEA 1M allows for the separation of 22 inorganic and organic anions in less than 17 min. Good RSDs for within-day migration time reproducibility (0.03-0.9%) and day-to-day analyses (0.04-1.4%) were obtained by the use of two internal standards, allowing for an accurate compound identification. The detection limits ranged from 0.1 to 0.4 mgL(-1) (S/N=3) for hydrodynamic injection (1250 mbars). The applicability of the proposed method was demonstrated by the analysis of inorganic and organic anions in diverse real samples. The recoveries obtained ranged from 93 to 106%.

Ultra-low flow nanospray for the normalization of conventional liquid chromatography/mass spectrometry through equimolar response: standard-free quantitative estimation of metabolite levels in drug discovery.

Nanospray experiments were performed on an ensemble of drug molecules and their commonly known metabolites to compare performance with conventional electrospray ionization (ESI) and to evaluate equimolar response capabilities. Codeine, dextromethorphan, tolbutamide, phenobarbital, cocaine, and morphine were analyzed along with their well-known metabolites that were formed via hydroxylation, dealkylation, hydrolysis, and glucuronidation. Nanospray exhibited a distinct trend toward equimolar response when flow rate was reduced from 25 nL/min to less than 10 nL/min. A more uniform response between the parent drug and the corresponding metabolites was obtained at flow rates of 10 nL/min or lower. The largest discrepancy was within +/-50% for plasma samples. Nanospray was used as a calibrator for conventional ESI liquid chromatography/tandem mass spectrometry (LC/MS/MS) and normalization factors were applied to the quantitation of an acyl-glucuronide metabolite of a proprietary compound in rat plasma. A nanospray calibration method was developed with the standard curve of the parent drug to generate quantitative results for drug metabolites within +/-20% of that obtained with reference standards and conventional ESI. The nanospray method provides a practical solution for the quantitative estimation of drug metabolites in drug discovery when reference standards are not available.