Internal Standard Peak Research Articles

Transfer learning improves predictions in lignin content of Chinese fir based on Raman spectra

Lignin in biomass plays significant role in substitution of synthetic polymer and reduction of energy expenditure, and the lignin content was usually determined by wet chemical methods. However, the methods' heavy workload, low efficiency, huge consumption of chemicals and use of toxic reagents render them unsuitable for sustainable development and environmental protection. Chinese fir, a prevalent angiosperm tree, holds immense importance for various industries. Since our previous work found that Raman spectroscopy could accurately predict the lignin content in poplar, we propose that the lignin content of Chinese fir can be estimated by similar strategy. The results suggested that the peak at 2895 cm−1 is the optimal choice of internal standard peak and algorithm of XGBoost demonstrates the highest accuracy among all algorithms. Furthermore, transfer learning was successfully introduced to enhance the accuracy and robustness of the model. Ultimately, we report that a machine learning algorithm, combining transfer learning with XGBoost or LightGBM, offers an accurate, high-efficiency and environmental friendly method for predicting the lignin content of Chinese fir using Raman spectra.

Whiteness and greenness appraisal approaches for quantitative determination of recently approved combination of indacaterol and mometasone via utilization of nuclear magnetic resonance technique

Quantitative proton nuclear magnetic resonance (1H-qNMR) is currently being used in a variety of applications in many disciplines due to the great privileges. Economical and minimal solvent system volume is utilized; no sample labelling or pre-treatment is necessary. Furthermore, no sample destruction is occurred. The all aforementioned benefits are contributed to its suitability for employment in pharmaceutical companies and quality control laboratories. Starting from this significance, a simple, quick, and sustainable method was developed for the determination of indacaterol (IND) and mometasone (MOM) in the raw material and newly approved pharmaceutical dosage form Atectura Breezhaler®. The developed approach involved the utility of pyridoxine (PYR) as an internal standard (IS) and DMSO-d6 as the suitable deuterated solvent for the analysis. The quantitative peaks of IND (doublet at 8.247 and 8.254 ppm) and MOM (singlet at 1.014 ppm) were obviously disengaged from the IS peak; which was singlet peak at 8.082 ppm. The developed methodology was perfectly linear over concentration range of 0.05–5.00 and 0.001–1.00 mg/mL for IND and MOM, respectively. The developed protocol was applied successfully for the determination of IND and MOM mixtures, including capsule-based inhaler dosage form with % recovery of 100.15% and 97.98% for IND and MOM, respectively. Also, the greenness and whiteness (GAPI, AGREE and RGB-algorithm approaches) of the evolved method were evaluated in order to assess its impacts on the ecosystem. The method was proved to possess ecofriendly properties without any expected harmful effects on the environment.

In Situ Ratiometric Determination of Cerebral Ascorbic Acid after Ischemia Reperfusion.

Ascorbic acid (AA) is significant in protecting the brain from further damage and maintaining brain homeostasis after ischemia stroke (IS); however, the dynamic change of cerebral AA content after different degrees of ischemic stroke is still unclear. Herein, carboxylated single-walled carbon nanotube (CNT-COOH)- and polyethylenedioxythiophene (PEDOT)-modified carbon fiber microelectrodes (CFEs) were proposed to detect in situ cerebral AA with sensitivity, selectivity, and stability. Under differential pulse voltammetry scanning, the CFE/CNT-COOH/PEDOT gave a ratiometric, electrochemically responsive signal. The internal standard peak at -310 mV was from the reversible peak of O2 reduction and the deprotonation and protonation of quinone groups, while AA was oxidized at -70 mV. In vivo experimental results indicated that the cerebral AA level gradually increased with the ischemic time increasing in different middle cerebral artery occlusion (MCAO) model mice. This work implies that the increasing cerebral AA level may be highly related to the glutamate excitotoxicity and ROS-led cell apoptosis and paves a new way for further understanding the release and metabolic mechanisms of AA during ischemia reperfusion and IS.

Chemical QuantArray: A Quantitative Tool for Mass Spectrometry Imaging.

Matrix-assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI) is a powerful analytical technique that provides spatially preserved detection and quantification of analytes in tissue specimens. However, clinical translation still requires improved throughput, precision, and accuracy. To accomplish this, we created "Chemical QuantArray", a gelatin tissue microarray (TMA) mold filled with serial dilutions of isotopically labeled endogenous metabolite standards. The mold is then cryo-sectioned onto a tissue homogenate to produce calibration curves. To improve precision and accuracy, we automatically remove pixels outside of each TMA well and investigated several intensity normalizations, including the utilization of a second stable isotope internal standard (IS). Chemical QuantArray enables the quantification of several endogenous metabolites over a wide dynamic range and significantly improve over current approaches. The technique reduces the space needed on the MALDI slides for calibration standards by approximately 80%. Furthermore, removal of empty pixels and normalization to an internal standard or matrix peak provided precision (<20% RSD) and accuracy (<20% DEV). Finally, we demonstrate the applicability of Chemical QuantArray by quantifying multiple purine metabolites in 14 clinical tumor specimens using a single MALDI slide. Chemical QuantArray improves the analytical characteristics and practical feasibility of MALDI-MSI metabolite quantification in clinical and translational applications.

Simple and Sensitive RP-HPLC and UV Spectroscopic Methods for the Determination of Remogliflozin Etabonate in Pure and Pharmaceutical Formulations

Simple, novel and selective reverse phase-high performance liquid chromatography (RP-HPLC) and ultraviolet (UV) spectroscopic methods have been developed and optimized for the determination of remogliflozin etabonate (RMZ) in bulk and dosage forms. In the HPLC method, the principal peak and internal standard peak were eluted separately at different retention times (RT) with the chromatographic conditions such as, mobile phase consisting of 0.02 M ammonium acetate buffer (pH was adjusted to 4.0 by 1.0 M ortho phosphoric acid), acetonitrile and tetrahydrofuran in the ratio 50:45:05, respectively (v/v) and the stationary phase used was C18, 5 μm, 4.6 mm x 250 mm kromasil column. The flow rate was 2.0 mL min-1, sample injection volume was 10 μL, and the wavelength of detection was fixed at 228 nm. In case UV spectroscopic method, the RMZ was diluted with pure ethanol. The RMZ showed a maximum absorbance at 228 nm. Hence throughout analysis 228 nm was used for the determination of RMZ. The RT of RMZ and internal standard, atorvastatin (ATST) were 6.2 min and 7.0 min, respectively. The resolution between the peaks was found to be more than 2.0. The total run time was fixed at 10 min. The linearity range for RP-HPLC method was found to be 10 μg mL-1 to 50 μg mL-1, at a fixed concentration of ATST. The linearity range for the UV spectroscopic method was found to be in the range of 100 to 250 μg mL-1. Regression coefficients (R2) were found above 0.999 for both of the techniques. The limit of detection and quantification for RMZ were found to be 1.0 μg mL-1 and 3.5 μg mL-1 respectively, in RP-HPLC method and 10.0 μg mL-1 and 40 μg mL-1, respectively, in UV spectroscopic method. The developed methods were found to be simple, accurate, reproducible, and precise. The RMZ can be analyzed in dual techniques, i.e., chromatographic and UV spectroscopic methods for its routine analysis.

Reliable and realistic models for lignin content determination in poplar wood based on FT-Raman spectroscopy

Studying lignin content is critical for analyzing the inherent constitution, performance, and application of lignocellulosic materials. However, the traditional methods (wet chemical methods) for the determination of lignin content have several limitations such as labor intensive, time-consuming preparations, and the use of toxic reagents. To address these shortcomings, based on 140 groups of poplar samples, a strategy of 1064 nm FT-Raman spectroscopy combined with several algorithms was proposed in our study. Before modeling, smoothing algorithm (Savitzky-Golay), baseline correction algorithm (adaptive iteratively reweighted penalized least squares), and deconvolution algorithm (Gaussian and Lorentzian) were applied to extract the Raman information. Also, several peaks including 1095, 1378, and 2895 cm−1 were selected as the candidates of internal standard peak for normalizing the lignin-related peaks. Subsequently, lignin content predictive models were established based on data extracted from FT-Raman spectra combined with regression algorithms, including principal components regression (PCR), partial least square regression (PLSR), ridge regression (RR), lasso regression (LR), and elastic net regressions (ENR). Consequently, credible models were obtained based on data normalized by peak of 2895 cm−1 combined with RR, LR, and ENR algorithms (Pearson’s R > 0.88, RMSE < 0.62, MAE < 0.46, and MAPE < 1.92%). Thus, the lignin quantitative models based on Raman spectroscopy developed here can be used to predict the lignin content in poplar clones accurately and efficiently.

~1H-qNMR quantification of total ginsenosides in Shenmai Injection

A content determination method based on ~1H-qNMR was developed for the determination of total ginsenosides in Shenmai Injection. The parameters were optimized with CD_3OD as the solvent, dimethyl terephthalate as the internal standard, the peak at δ 8.11 as the internal standard peak, and the peaks at δ 1.68 and δ 0.79 as quantitative peaks of total ginsenosides. The developed ~1H-qNMR-based method was validated methodologically. The results showed that the method could achieve accurate measurement of total ginsenosides in Shenmai Injection in the range of 0.167 6-3.091 1 mmol·L~(-1). The developed ~1H-qNMR-based method for total ginsenosides is simple in operation, short in analysis time, strong in specificity, independent of accompanying standard curve, and small in sample volume, which can serve as a reliable mean for the quality control of Shenmai Injection. This study is expected to provide new ideas for the development of quantification methods of total ginsenosides.

Analytical validation of plasma amyloid β measuring system by immunoprecipitation‐mass spectrometry

AbstractBackgroundWith the increasing number of patients with Alzheimer's disease, construction of inspection system for blood biomarkers to replace or complement amyloid PET and CSF biomarkers has been required. Since our first report in 2014 (Kaneko et al., Proc Jpn Acad Ser B Phys Biol Sci. 2014), we have been focusing on three amyloid peptides, Aβ1‐40, Aβ1‐42, and APP669‐711 in human plasma, and found that the combination of peptide ratios APP669‐711/Aβ1‐42 and Aβ1‐40/Aβ1‐42 became a biomarker for amyloid peptide accumulation in human brain (Nakamura et al., Nature 2018). Here we describe the analytical validation of plasma amyloid β measuring system by immunoprecipitation‐matrix‐assisted laser desorption ionization time‐of‐flight mass spectrometry (IP‐MALDI‐MS).MethodCommercially available human plasma and artificial plasma were immunoprecipitated with anti‐Aβ antibody (clone 6E10) covalently bound to the magnetic beads, then measured by MALDI‐TOF‐MS. The peak intensity of the Aβ‐related peptides was normalized by the internal standard peak, and biomarker values were calculated from those intensities. A method validation including experiments of calibration range, sensitivity, specificity, reproducibility, interfering substances, dilution linearity, recovery, and stability was performed.ResultThe system showed good linearity over the range from 15 to 150 pM for Aβ1‐40, and 3 to 15 pM for Aβ1‐42 and APP669‐711. The trueness was within 100 ± 25% at the lower limit of quantification and within 100 ± 20% at all other concentrations. The precision of the intra‐ and inter‐assay was 1% to 13% as a CV value. Recovery from spiked human plasma ranged from 83% to 101% at low, middle, and high concentrations. Stability testing has shown that the antibody beads are stable for up to 6 months in the refrigerator so far. The addition of the interfering substances, such as anticoagulants, Alzheimer's disease drugs, etc., had only a slight effect on the biomarker values showing 10% or less difference relative to the control. Further results will be reported in the presentation.ConclusionThe results demonstrate that the plasma amyloid β measuring system by IP‐MALDI‐MS is an accurate, reliable, and robust assay that would be most suitable for screening subjects in clinical trials.

Determination of phosphite (HPO3−2) by a new IC/MS/MS method using an 18O-labeled HPO3−2 internal standard

A new method has been developed to determine trace amounts of phosphite (HPO3−2) in environmental samples using ion chromatography with electrospray tandem mass spectrometry (IC-ESI/MS/MS). The method includes the production and use of an 18O-labeled HPO3−2 internal standard (IS). This isotopically labeled IS significantly improved sensitivity and could account for matrix suppression. The method detection limit (MDL) was determined as 0.017 and 0.034 μg L−1 of HPO3−2 (6.5 and 13 ng P L−1) using a 500 and 25 μL injection loop, respectively. Precision (1–10%) and accuracy (recoveries = 96–106%) were established for a range of environmental samples using known (spiked) addition. The impact of ionic interferences was investigated by evaluating the response of the internal standard in the presence of common anions with respect to distilled deionized water. The most significant interference was due to nitrate (100 mg-NO3− L−1) with a 99.99% reduction in IS intensity. The method was successfully applied to wastewater effluent, surface water, tap water, and soil samples. Relatively low concentrations <0.25 μg HPO3−2 L−1 were measured in tap water, surface water and wastewater effluent, and ~1.6 μg kg−1 HPO3−2 in soil samples, using both injection loops. Limited suppression was observed for all matrices. The largest IS peak area suppression (~98%) was observed in WW effluent with 500 μL injection loop; however, this method was able to quantify HPO3−2 with good recoveries and precision despite the mentioned suppression, supporting the ability of the proposed method to quantify HPO3−2 in different environmental matrices.

Direct infusion nano-electrospray ionization mass spectrometry for therapeutic drug monitoring of ciprofloxacin and its metabolites in human saliva

A rapid and sensitive method based on direct infusion-nano-electrospray ionization mass spectrometry (DI-nESI-MS) has been developed for the detection and quantification of ciprofloxacin and its metabolites in human saliva. Saliva samples were collected after the oral administration of 500 mg ciprofloxacin tablets. Internal standard (IS), tamoxifen, was added to the collected samples, and then diluted with the ionization solvent, centrifuged and filtered. An aliquot of 4 μL of the filtrate was loaded into a nanospray (NS) capillary. The NS capillary was then fitted into an off-line ion source and the instrument was operated to acquire a two-minute run by applying a voltage of 1000 V (positive-ion detection mode). Quantification of ciprofloxacin relied on the ratio of its peak intensity to the IS peak intensity. The DI-nESI-MS method was validated and provided satisfactory precision with relative standard deviation ranging from 0.39 to 7.48 % and accuracy with relative error ranging from -2.12 to 9.72 %. The calibration curve showed good linearity (r2) > 0.999 over the concentration range of 10−4000 ng/mL. These results verify the effectiveness of the DI-nESI-MS method for monitoring of ciprofloxacin and its metabolites in human saliva samples.

Internal Standards for Quantitative Analysis of Chemical Warfare Agents by the GC/MS Method: Nerve Agents.

General conditions and requirements for an internal standard useful in the determination of chemical warfare agents (CWAs) by the method of gas chromatography coupled with mass detection (GC/MS) were defined. The determination is based on a GC/MS analysis of a mixture of a CWA with an internal standard, conversion of the TIC chromatogram to a chromatogram extracted at a particular m/z ratio, and calculation of the CWA concentration from the internal standard concentration, response factor, and chromatographic peak areas. Available internal standards were identified, and they were verified for seven organophosphorus nerve-paralysing agents. Corresponding response factors were determined as a ratio of gradients of the linear functions of the peak area and compound concentration. Linearity, repeatability, and accuracy of the measurements were evaluated. The determination can be performed on all GC/MS systems of the Fire Rescue Service of the Czech Republic (FRS), where no CWA standards are available.

Quantification and targeted detection of ciprofloxacin in dosage form and human urine by direct injection nano-electrospray ionization multi-stage mass spectrometry

A rapid, selective and sensitive direct infusion nano-electrospray ionization multi-stage mass spectrometry (DI-nano-ESI-MS3) method has been developed and validated for ciprofloxacin identification and quantification. An aliquot of 4 µl containing ciprofloxacin sample solution along with an internal standard (IS), in this case tamoxifen, was loaded into a nanospray (NS) capillary. The NS capillary was then fitted off-line into the ion source where the distance between NS capillary tip and MS aperture was set to 5 mm. The instrument was operated to acquire a two-minute run by applying a voltage of 1000 V (positive-ion detection mode). Ciprofloxacin quantification relied on the ratio of its parent peak (m/z 332) to the IS peak (m/z 372), while identification relied on MS3 of the parent peak and its fragments with m/z values of 314, 288, 268 and 245, respectively. Due to the observed ion suppression effect exerted by the IS over the analyte ion signal at lower concentration levels (< 500 ng/ml ciprofloxacin), the calibration curve was split into two sections. As a result, linearity correlation between the ratios of the analyte to the IS and the corresponding concentrations over the ranges 5–500 and 500–4000 ng/ml respectively was observed. The applicability of the developed method was confirmed by analyzing ciprofloxacin in a pharmaceutical dosage form as well as detection of the analyte in random human clinical urine samples after oral administration of the drug.

Development and validation of a high performance liquid chromatography method to determine nevirapine in plasma in a resource-limited setting.

BackgroundThere are several instances where nevirapine pharmacokinetic monitoring may be useful, such as in special populations or pharmacokinetic drug interaction studies that require the ascertainment of nevirapine pharmacokinetics in the sub-Saharan region.ObjectivesThe main aim of this study was to produce a validated, sustainable and relevant nevirapine assay method that meets bio-analytical regulatory requirements.MethodsThe developed method utilised a Waters 2795 Alliance high performance liquid chromatography system with a 2996 photo diode array detector, an Atlantis dC18 5 micron, 3.9 mm × 150 mm analytical column and a gradient flow rate of 1 mL/min. Ultraviolet detection data were collected from 210 nm to 400 nm, extracted at 260 nm, and processed for nevirapine and internal standard peak height responses.ResultsThe method proved to be linear (R2 0.995), precise (+1.92% – +9.69%) and accurate (-9.70% – 12.0%). Recovery for the analyte and internal standard was between 98.8% and 114%. The method showed good specificity as no interferences were caused by common African traditional medicines, anti-tuberculosis medications or other concomitant antiretrovirals nor endogenous components.ConclusionThe method is reproducible, relevant to our setting and uses considerably low plasma volumes with preservation of some consumables, a desirable key factor in a resource-limited setting.

Screening and quantitation of forty-six drugs of abuse and toxic compounds in human whole blood by capillary electrophoresis: Application to forensic cases

A novel capillary electrophoresis method, coupled with photo-diode array detection, was established for the analysis of forty-six drugs of abuse, including opiates, cocaine, amphetamines, benzodiazepines, and alkaloids, in human whole blood. Doxapram was used as the internal standard, and an Oasis HLB column was used for solid-phase extraction of drugs from human whole blood. This method was applied to analyze real blood samples in forensic cases. The separation was performed on a fused-silica capillary (75 μm I.D. × 50.2 cm, with 40 cm of effect length). In this study, the running buffer was 150 mM phosphate buffer (pH 2.4) containing 20% methanol. Samples were injected at a pressure of 0.5 psi for 10 s and separated at a running voltage of 16 kV. The precision, accuracy, sensitivity, linear range, limit of detection, and recovery of the method were evaluated. The time taken for the entire separation process was 18.5 min. All compounds were identified by relative retention times, and the peak height ratios (the ratio between the target analyte peak height and the internal standard peak height) were monitored at 200 nm and 210 nm in corresponding ultraviolet spectra. 210 nm was used as quantitative detection wavelength. Calibration curves were established for all compounds of interest. For human blood samples, the detection limits were 8–80 ng/mL, and the linear range was 0.03–10 μg/mL. The intraday and interday precisions were both less than 13%. This method offers simple and effective screening and quantitation of drugs of abuse and toxic compounds in human blood at toxicologically relevant concentrations, and was applied to analyze blood samples from forensic cases.

Quantitative Assessment of Poorly Soluble Anticoagulant Rivaroxaban by Microemulsion Electrokinetic Chromatography.

Microemulsion electrokinetic chromatography (MEEKC) is an electrophoretic methodology based on the separation of compounds by a microemulsionated electrolyte. There are few options for the evaluation of the stability and content of the oral anticoagulant rivaroxaban (RIV) in pharmaceutical formulations. RIV has low water solubility and undergoes ionization only under restricted pH conditions (pH < 1 or pH > 13), thus, hindering the application of free zone capillary electrophoresis as an analytical method. Therefore, the work aimed at developing and validating a stability-indicating MEEKC method for the analysis of RIV in pharmaceutical formulations. Separation was performed in a fused-silica capillary applying a voltage of 30 kV. The microemulsion system consisted of 13 mM tetraborate, pH 9.75 + 1.2% SDS + 1.0% ethyl acetate + 2.4% butanol. The linearity range was 25-150 μg mL-1, with r = 0.9982. Drug degradations were performed in acid and basic media (HCl 1 M and NaOH 0.1 M, respectively), oxidation with 3%H2O2, 60°C temperature and exposure to UV-C radiation. No interferences with RIV or internal standard peaks were detected. Method robustness was accessed through Plackett-Burman experimental design, after evaluation of model validity. Trueness values between 100.49 and 100.68% were obtained with repeatability. The method developed was found appropriate for quality control of RIV tablets, as a consistent analytical technique that is considered less damaging to the environment due to its low consumption of organic reagents.

Degree of conversion and depth of cure of Ivocerin containing photo-polymerized resin luting cement in comparison to conventional luting agents.

Objective:To evaluate the degree of conversion (DC) and depth (extent) of cure of four resin cements (Variolink E, Calibra, NX3 and Variolink N) using Fourier transform infrared (FTIR) and Vickers Micro hardness (MH).Methods:Ten disks (1mmx2mm) of each resin cement were light cured through a ceramic disk for 40 seconds prior to assessment. The ATR spectra of the uncured resin were collected in absorbance mode from 16 scans at 4 wave number resolutions. Degree of conversion was calculated by estimating the changes in peak height ratio of the absorbance intensities of aliphatic C=C peak at 1638 cm-1 and that of an internal standard peak of aromatic C=C at 1608 cm-1 during polymerization. For Vickers microhardness testing 10 disks of each cement specimen was exposed to 100 grams of load for 15 seconds. Three indentations were made 0.5mm apart and an average Vickers micro-hardness (MH) for each specimen. Two way ANOVA and multiple comparison tests were performed to assess data.Results:The highest degree of conversion by peak area was shown by Variolink-Esthetic [light-cure (87.18±2.90%)]; however the lowest was observed in samples of Variolink-N [Dual cure (44.55±4.33%)]. Similarly, Variolink-Esthetic and NX3 cement showed significantly higher MH as compared to other groups.Conclusion:Ivocerin containing Variolink–E cement showed high degree of conversion and extent of polymerization when compared to conventional light and dual cure luting cements.

Liquid Chromatographic-Chemometric Techniques for the Simultaneous HPLC Determination of Lansoprazole, Amoxicillin and Clarithromycin in Commercial Preparation.

Two multivariate calibration-prediction techniques, principal component regression (PCR) and partial least-squares regression (PLSR) were applied to the chromatographic multicomponent analysis of the drug containing lansoprazole (LAN), clarithromycin (CLA) and amoxicillin (AMO). Optimum chromatographic separation of LAN, CLA and AMO with atorvastatin as the internal standard (IS) was obtained by using Xterra® RP18 column 5 μm 4.6 × 250 mm2, and 25 mM ammonium chloride buffer prepared ammonium chloride, acetonitrile and bidistilled water (45:45:10 v/v) as the mobile phase at flow rate 1.0 mL/min. The high pressure liquid chromatography data sets consisting of the ratios of analyte peak areas to the IS peak area were obtained by using diode array detector detection at five wavelengths (205, 210, 215, 220 and 225 nm). LC-chemometric calibration for LAN, CLA and AMO were separately constructed by using the relationship between the peak-area ratio and training sets for each analyte. A series of synthetic solutions containing different concentrations of LAN, CLA and AMO were used to check the prediction ability of the PCR and PLS. Both of the two-chemometric methods in this study can be satisfactorily used for the quantitative analysis and for dissolutions tests of multicomponent commercial drug.

A rapid screening of histamine concentration in fish fillet by direct analysis in real time mass spectrometry (DART-MS)

Histamine is a biogenic amine causing scombroid foodborne poisoning. A rapid determination of the amine is effective in preventing the distribution of histamine-accumulated foods to market. Conventional methods for the determination of histamine concentration in foods include analyses by high performance liquid chromatography (HPLC). The present study tried to utilize a direct analysis in real-time ionization coupled to mass spectrometer (DART-MS) to estimate the histamine concentration in fish filets as an alternative to HPLC. The DART-MS responses to histamine and 1,8-diaminooctane as an internal standard were highly dependent on heater temperature; the highest responses were obtained at 400 °C. Histamine was spiked from 16.3 mg/kg to 407.0 mg/kg in tuna and yellowtail filets to obtain contaminated samples. The contaminated samples were homogenized with 0.1 mM EDTA containing 1,8-diaminooctane. The filtrates of the suspension were analyzed by DART-MS, and peak areas corresponding to the histamine and internal standard were obtained. The ratios of the histamine peaks to the internal standard peaks increased with higher histamine concentrations in the fish filets. The high linearity between the histamine concentrations and peak area ratios and the determination coefficient was more than 0.95. The results suggested that DART-MS is an attractive tool as a rapid screening method for determining histamine concentrations in fish filets.

Ultra-high performance liquid chromatographic determination of levofloxacin in human plasma and prostate tissue with use of experimental design optimization procedures.

Fluoroquinolones are considered as gold standard for the prevention of bacterial infections after transrectal ultrasound guided prostate biopsy. However, recent studies reported that fluoroquinolone- resistant bacterial strains are responsible for gradually increasing number of infections after transrectal prostate biopsy. In daily clinical practice, antibacterial efficacy is evaluated only in vitro, by measuring the reaction of bacteria with an antimicrobial agent in culture media (i.e. calculation of minimal inhibitory concentration). Such approach, however, has no relation to the treated tissue characteristics and might be highly misleading. Thus, the objective of this study was to develop, with the use of Design of Experiments approach, a reliable, specific and sensitive ultra-high performance liquid chromatography- diode array detection method for the quantitative analysis of levofloxacin in plasma and prostate tissue samples obtained from patients undergoing prostate biopsy. Moreover, correlation study between concentrations observed in plasma samples vs prostatic tissue samples was performed, resulting in better understanding, evaluation and optimization of the fluoroquinolone-based antimicrobial prophylaxis during transrectal ultrasound guided prostate biopsy. Box-Behnken design was employed to optimize chromatographic conditions of the isocratic elution program in order to obtain desirable retention time, peak symmetry and resolution of levofloxacine and ciprofloxacine (internal standard) peaks. Fractional Factorial design 2(4-1) with four center points was used for screening of significant factors affecting levofloxacin extraction from the prostatic tissue. Due to the limited number of tissue samples the prostatic sample preparation procedure was further optimized using Central Composite design. Design of Experiments approach was also utilized for evaluation of parameter robustness. The method was found linear over the range of 0.030-10μg/mL for human plasma and 0.300-30μg/g for human prostate tissue samples. The intra-day and inter-day variability for levofloxacine from both plasma and prostate samples were less than 10%, with accuracies between 93 and 108% of the nominal values. The limit of detection and the limit of quantification for human plasma were 0.01μg/mL and 0.03μg/mL, respectively. For the prostate tissue, the limit of detection and the limit of quantification were 0.1μg/g and 0.3μg/g, respectively. The average recoveries of levofloxacin were in the range from 99 to 106%. Also, the method fulfills requirements of robustness what was determined and proved by Design of Experiments. The developed method was successfully applied to examine prostate tissue and plasma samples from 140 hospitalized patients enrolled into the clinical study, 12h after oral administration of LVF at a dose of 500mg. The mean (±SD) LVF concentration in prostate was 6.22±3.52μg/g and in plasma 2.54±1.14μg/mL. Due to simplicity of the method and relative small amount of sample needed for the assay, the method can be applied in clinical practice for monitoring of LVF concentrations in plasma and prostate gland.

Surface enhanced Raman spectroscopy hyphenated with surface microextraction for in-situ detection of polycyclic aromatic hydrocarbons on food contact materials

Highly bioaccumulated polycyclic aromatic hydrocarbons (PAHs) have cause health concerns because of their carcinogenic properties. PAHs could migrate to food from contaminated food contact materials. In this study, a hyphenated technique combining surface enhanced Raman spectroscopy (SERS) with surface microextraction was developed for in-situ on site screening of PAHs on food contact materials. Methanol and 1-propanethiol-modified silver nanoparticles (PTH-Ag NPs) were used to perform the in-situ microextraction and detection of PAHs, respectively. The SERS spectra can be obtained by a portable Raman spectrometer. The vibration of the C–C bond of PTH at 1030cm−1 was chosen as an internal standard peak. The PTH-Ag NPs showed high uniformity with an RSD of 2.96%. A plot of the normalized SERS intensity against fluoranthene concentration showed a linear relationship (R2=0.98). The detection limit could reach 0.27ngcm−2. The in-situ microextraction-SERS hyphenated technique for the detection of three PAHs at five food contact materials was demonstrated. The method can be also applied to detect PAH mixtures. This in-situ microextraction-SERS hyphenated method demonstrated its ability to rapidly screen PAHs on contaminated food contact materials free from complex sample pretreatment.