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Introduction Mycotoxins are toxic metabolites produced by fungi that commonly contaminate foods. As recommended by the World Health Organization, total diet study (TDS) is the most efficient and effective way to estimate the dietary intakes of certain chemical substances for general populations. It requires sensitive and reliable analytical methods applicable to a wide range of complex food matrices and ready-to-eat dishes. Objectives A novel strategy with high selectivity and sensitivity, incorporating three methods based on ultra-high-performance liquid chromatography coupled with tandem mass spectrometry (UHPLC-MS/MS), was designed for measuring 43 mycotoxins in dietary samples in a China TDS. Methods The 43 mycotoxins were divided into 3 groups for analysis to achieve better performance. For each group, an UHPLC-MS/MS method was developed to determine the target compounds after clean-up by solid phase extraction. A total of 21 isotope internal standards were employed for accurate quantitation. Method validation in terms of linearity, selectivity, sensitivity, accuracy, and precision was performed for all the 43 mycotoxins in 12 complex food matrices. Results The limits of detection (LODs) and limits of quantitation (LOQs) were 0.002–1 ng mL−1 and 0.006–3 ng mL−1, respectively. The method recoveries of the 43 mycotoxins spiked in 12 food categories were in the range of 60.3%–175.9% after internal standard correction, with relative standard deviations (RSDs) below 13.9%. For practical application, this method was utilized for 72 dietary samples collected from 6 provinces in the 6th China TDS. More than 80% of the samples were found contaminated by mycotoxins. DON, SMC, FB1, ZEN, BEA, ENNB1, and ENNB were most detected. Conclusions The proposed methods with high sensitivity, accuracy, and robustness provide powerful tools for multi-mycotoxin monitoring and dietary exposure assessment, allowing 43 mycotoxins, including some emerging mycotoxins, to be accurately investigated in a total diet study for the first time.

Conventionally used spectrometric techniques of inductively coupled plasma optical emission spectrometry (ICP-OES) and inductively coupled plasma optical emission spectrometry (ICP-MS) usually involve time-consuming sample preparation procedure of a sample dissolution which requires the usage of aggressive and toxic chemicals. The need for suitable and sustainable analytical methods for direct multi-elemental analysis of plant samples has been increased in recent years. Spectrometric techniques for direct sample analysis, instrumental neutron activation analysis (INAA) and X-ray fluorescence (XRF) have been applied in environmental studies and various fields of screening tests. Nevertheless, these techniques are not commonly used for plant sample analysis and their performances need to be evaluated. This research aimed to assess how reliable non-destructive techniques are in the determination of elements in plants compared to conventionally used spectrometric techniques. A total of 49 plant samples of four conifer species (Pinus nigra, Abies alba, Taxus baccata and Larix decidua) were measured using two conventionally applied (ICP-MS, ICP-OES) and two non-destructive techniques (wavelength dispersive XRF (WD-XRF), INAA). The comparison was performed by investigation of relative ratios of concentrations and by correlation analysis. Moreover, precision of the techniques was examined and compared. The quality control included analysis of NIST pine needles certified reference material (1575a) using all examined techniques. Our results suggest that additional analytical and quality control steps are necessary for reaching the highest accuracy of multi-elemental analysis. Конвенционално коришћене спектрометријске технике (ICP-OES, ICP-MS) обично подразумевају дуготрајну процедуру припреме узорка која захтева коришћење агресивних и токсичних хемикалија за минерализацију. У последњих неколико година расте потреба за погодним и одрживим аналитичким методама за директну мулти-елементарну анализу биљних узорака. Спектрометријске технике за директну анализу (INAA, XRF) се већ примењују у еколошким истраживањима и у различитим пољима скрининг испитивања. Ипак, ове технике нису уобичајено коришћене за анализу биљака и њихове перформансе морају бити процењене. Циљ овог истраживања је био да се процени колико су недеструктивне технике поуздане код одређивања елемената у биљном материјалу, у односу на рутински коришћене спектрометријске технике. Укупно 49 узорака четири врсте четинара (Pinus nigra, Abies alba, Taxus baccata и Larix decidua) су анализиране помоћу две рутински коришћене (ICP-MS и ICP-OES) и две недеструктивне технике (WD-XRF и INAA). Технике су упоређене испитивањем релативних односа концентрација и помоћу корелационе анализе. Поред тога, испитана је и упоређена прецизност техника. Програм контроле квалитета је обухватао анализу сертификованог референтног материјала, иглица бора (NIST 1575a) помоћу свих коришћених техника. Резултати нашег истраживања сугеришу да су неопходни додатни аналитички и кораци контроле квалитета како би се постигла максимална тачност мулти-елементарне анализе.

Human biological fluids are quite varied and complex. Some examples include blood, cerebrospinal fluid, synovial fluid, peritoneal fluid, intraocular fluids, and fluids that do not require an invasive sample uptake, such as lacrimal fluid, seminal fluid, sweat, urine and saliva. They are present in the body in very different volumes, from the low microliter level to the liter range. For those requiring invasive sampling, just a small percentage can be sampled in order not to disrupt the living body. Furthermore, quite often, the extracted samples must be divided into several aliquots required for different independent (bio)chemical assays. In addition, volumes of biofluids are much smaller in new-borns and infants. Therefore, just a microvolume of the biofluid is quite often available for analysis. Inductively coupled plasma-mass spectrometry (ICP-MS) is a suitable technique for elemental and isotopic determination of biological fluids at the trace and ultra-trace levels. Such determinations contributes to the interpretation of biochemical processes within our bodies and in many cases serve for the diagnosis of diseases. A research trend along the years has been to increase the capabilities of ICP-MS for the analysis of microsamples. This review focusses on strategies for microsample collection of fluids from the human body and their introduction into an ICP-MS. In addition, representative examples of applications in the clinical field are briefly described.

Objective To figure out the source of uncertainty in the determination of 14C activity concentration in urine samples, and to establish a complete expression of analysis results, in order to improve the accuracy of 14C monitoring method. Methods Four urine samples of normal people were collected, and wet oxidation method was used in the pretreatment of the samples, then the activity concentration of 14C was determined by liquid scintillation counting (LSC). After the mathematical model was established, the uncertainty components were determined, the standard uncertainty was combined, and the extended uncertainty was calculated. Results The relative uncertainty of 4 samples was 22.5%, 17.0%, 17.3% and 18.2%. The activity concentration of urine was (0.32 ​± ​0.14), (0.60 ​± ​0.20), (0.86 ​± ​0.30) and (0.74 ​± ​0.27) Bq/L, respectively. In the relative uncertainty of 22.5%, 17.7% was from sample counting, 13.6% from detection efficiency, 2.1% from recovery rate, 0.58% from sample volume, and 0.01% from sample weight. Conclusions The main factors affecting the uncertainty include the error or deviation of sample counting, detection efficiency and recovery rate. The error of volume and weight also contributes to the measurement uncertainty.

Aflatoxin B1 (AFB1), as the main metabolite, found in aflatoxin contaminated food is highly toxic and strictly controlled in many countries. Aiming to construct a simplified, highly sensitive, and accurate quantitative detection platform of AFB1 in ordinary conditions, a one-step competitive quantum dot-labeled immunosorbent assay (cQLISA) was developed using biocompatible CdSe/ZnS core/shell quantum dots (QDs). The quantitative detection of AFB1 can be achieved by adding food sample and QD-AFB1 antibody simultaneously, with an analytical range between 1 and 40 ng/mL. The half maximal inhibitory concentration (IC50) of this method is 542 pg/mL and the limit of detection (LOD) is 56 pg/mL for standard samples, demonstrating a twofold improvement in sensitivity compared with that of the commercial enzyme-linked immunosorbent assay (ELISA) test kit. The recovery rates of negative corn spike-in samples range from 87.01 to114.7% (CV

In this study, a sensitive high-performance liquid chromatography method for zearalenone determination in feeds and grains was established based on the automatic immunomagnetic bead purifier analyzer (KY-AMI-I) for the one-step sample pre-treatment. Various parameters that may affect the analytical performance were investigated. Immunomagnetic bead separation system based on a monoclonal antibody (anti-ZEN antibody-immunomagnetic beads conjugation) was individually prepared for the ZEN purification. Besides, the average recoveries in spiked samples ranged from 75.23 to 102.62%, and the relative standard deviations were between 3.55 and 8.07% (

The Griess assay is widely used by regulation agencies as an official method for nitrite quantification in water and food samples. In Brazil, the official method, which has been used to determine nitrite in food, was described by Instituto Adolfo Lutz (283/IV) in 1984. It uses 8 mL of reactants and provides 50 mL (reactants plus sample) of waste per sample analyzed. Here, students scaled down the official method 50 times and quantified nitrite in water and food samples by using just 0.2 mL of the Griess reactant and 1 mL of sample. Quantitative analysis was carried out using absorbance measured at 540 nm (standard method) and 96-well-plate images (proposed method) obtained with a desktop scanner. The nitrite was extracted from solid food samples by heating it in a water bath. After heating, sample color and turbidity were eliminated by addition of K4[Fe(CN)6] and ZnSO4 solutions and filtering. During samples preparation, students evaluate the heating time effect in nitrite extraction from sausage samples using hypothesis tests. Students did a series of matrix matched samples to observe the matrix effect. Students also calculated the detection limit (DL) and the quantification limit (QL) for the proposed method (1.35 and 4.1 μmol/L nitrite, respectively) and for the standard method (1.1 and 3.4 μmol/L nitrite, respectively). DL and QL were determined using the standard deviation of the lowest concentration point on the standard curve. The major pedagogical value of this laboratory class was to scale down the official method and use it to prepare and analyze a solid food sample. As a learning model, finding real water samples, which have nitrite concentrations larger than method QL, was a hard task, but all sausage samples analyzed had larger nitrate concentrations than the method QL and nitrite quantification in sausages was a good learning model.

A continuous-flow microchip enabling high-accuracy DNA analysis was developed. Serial consecutive analysis for multiple amplified DNA samples was demonstrated. The sample segments were continuously introduced to the microchip from the PCR device which was interfaced to the microchip through capillary tubing. Electrokinetic co-injection of the DNA samples with size marker enabled reproducible and reliable injection of the DNAs into the gel-filled separation channel providing accurate size determination of the DNA samples. Cross-contamination between serially introduced DNA samples was minimized by plugging a washing solution segment following the previous sample segment between two sample plugs. Using this microchip, continuous separation of multiple samples was performed without any inconvenient and labor-intensive sample preparation steps such as sample mixing, staining, and gel loading which are necessary for conventional gel electrophoresis. It has taken about 4 min to separate single DNA sample and taken 37 min for three serially injected samples which implies that this microchip can be a platform device for fast as well as highly accurate DNA analysis.

In Japan, the use of mothproofing agents [dieldrin and 4,6-dichloro-7-(2,4,5-trichlorophenoxy)-2-trifluoromethylbenzimidazole; DTTB] in textiles is regulated by the Act on the Control of Household Products Containing Harmful Substances. Since official analytical methods for these agents have been in place for approximately 40 years, we developed an improved method in a previous study. In the present study, we validated this method. Accordingly, six laboratories analyzed the sample prepared at 3 μg/g (1/10 of the regulation value) and 30 μg/g (the regulation value). The high accuracy of the results for these samples in almost all the cases (accuracy: 70-120%, repeatability

Objectives: To evaluate four sample treatments in a safe and straightforward procedure to detect SARS-CoV-2 in saliva. Methods: Four sample treatments were evaluated in a 3-step procedure to detect SARS-CoV-2 in saliva: 1) heating at 95 °C for 5 min for sample inactivation; 2) sample treatment; 3) analysis by reverse-transcription loop-mediated isothermal amplification (LAMP). Saliva samples used were from infected individuals or were spiked with known quantities of viral particles. Results: Three treatments had a limit of detection (LOD) of 500.000 viral particles per ml of saliva and could be used to detect individuals with potential to transmit the disease. The treatment of phosphate buffer, dithiothreitol, ethylenediaminetetraacetic acid and proteinase K, with an additional 95 °C heating step, yielded a lower LOD of 95; its sensitivity ranged from 100% in patients with nasopharyngeal swab reverse-transcriptase polymerase chain reaction cycle threshold values 30. Conclusions: This report highlights the importance of an adequate sample treatment for saliva to detect SARS-CoV-2 and describes a flexible procedure that can be adapted to point-of-care. Although its sensitivity when LAMP is used is lower than reverse-transcriptase polymerase chain reaction, this procedure can contribute to COVID-19 control by detecting individuals able to transmit the disease.

In this study, the idea of sample-specific metabolites library with retention neighbor was presented for better identification and quantitation of metabolites. Compared with universal libraries, the established library was found to have significantly improved match indices (similarity, reverse, and probability), indicating better identification of metabolites. For quantitative metabolic profiling of temperature treated tobacco samples, the established method integrated and aligned 175 metabolites with no missing value, better than the widely used metabolomics software including XCMS Online, MetaboliteDetector, SpectConnect, and ChromaTOF. Partial least squares discriminant analysis models were also used for quantitative evaluation and the established method produced better values of model parameters than the compared software, indicating a better model was established using the present method.

Introduction Urine collection from incontinent individuals can be challenging. Various methods have been devised to collect the sample without catheterization. Recently the PureWick external catheter was developed to draw the sample gently away from external female genitalia. While the primary purpose of the device is to prevent moisture and maintain skin integrity, the urine that is collected may be sent for laboratory analysis. We sought to validate the use of this collection method for common urine chemistry assays and urinalysis. Methods Twenty pools of residual urine samples were separated into “control” and “PureWick” treated samples. The control samples were maintained at room temperature while 15 mL of urine was added to the PureWick device which was connected to a vacuum line through a collection canister. The urine collected in the canister and the controls samples were all subject to urine chemistry strip, microscopic, and automated urine chemistry analysis. Results were compared between pairs of treated and control samples. Results No clear affect was noted on urine strip semi-quantitative or automated chemistry analysis from the PureWick collection. There was a statistically significant decrease in microscopic measurements of white blood cells and crystals in the PureWick urine samples. Discussion This study supports the use of the PureWick external catheter for collection of samples for most urinalysis and urine chemistry tests.

Objectives In the absence of sample validity testing, a healthcare provider may fail to identify a patient’s adulteration of their urine sample. This study compared different methods for specific gravity (SG), pH, TECO™ Drug Adulteration Test Strip (dipstick) and oxidant assay to explain the differences and also make an informative decision on method selection. Methods Creatinine, SG and pH measurements are essential in sample validity testing. SG and pH automated chemical methods are compared against pH meter method, SG refractometer and dipstick method. Also, oxidant assay was compared against dipstick method. Results SG chemical method agreement with refractometer is 81.9% and with dipstick method is 64.7%. The refractometer method agreement with dipstick method is 66.1%. pH chemical method agreement with pH Meter method is 74.3% and with dipstick method is 81.4%. pH meter method agreement is 85.7% with dipstick method. Results were analysed using Deming regression analysis and F-test. SG chemical method correlated better with refractometer than the dipstick method. Oxidant assay correlated well with dipstick method in detecting adulterants such as pyridinium chlorochromate, nitrite and bleach. Conclusions Varying degrees of differences were seen in the SG and pH measurements. These differences were both method and instrument dependent. The automated chemical methods are recommended alongside oxidant assay for consistency, accuracy and faster turn-around time as part of sample validity testing for drugs of abuse.

Bacterial contamination of donated blood is defined as the presence of bacteria in the blood components which are collected and/or processed for transfusion. It is the second cause of death beyond ABO-mismatch. The aims were to determine the frequency of bacterial contaminations in stored packed RBC and platelet concentrate units. This cross-sectional study was conducted in Baghdad between 2nd of September to 27th of December 2019. Two hundred samples; 100 samples from packed RBC units and 100 samples from platelet concentrate units were randomly selected. There were 38/100 of platelet concentrate units found to be contaminated, while 28/100 samples studied of packed RBC units were contaminated by bacteria. The high rate of contamination of samples presented. Gram-positive bacteria were the most predominant, and this attributed to poor skin cleansing and antiseptic techniques used prior to donor blood collection.

Isobaric labeling via tandem mass tag (TMT) reagents enables sample multiplexing prior to LC-MS/MS, facilitating high-throughput large-scale quantitative proteomics. Consistent and efficient labeling reactions are essential to achieve robust quantification; therefore, embedded in our clinical proteomic protocol is a quality control (QC) sample that contains a small aliquot from each sample within a TMT set, referred to as "Mixing QC." This Mixing QC enables the detection of TMT labeling issues by LC-MS/MS before combining the full samples to allow for salvaging of poor TMT labeling reactions. While TMT labeling is a valuable tool, factors leading to poor reactions are not fully studied. We observed that relabeling does not necessarily rescue TMT reactions and that peptide samples sometimes remained acidic after resuspending in 50 mM HEPES buffer (pH 8.5), which coincided with low labeling efficiency (LE) and relatively low median reporter ion intensities (MRIIs). To obtain a more resilient TMT labeling procedure, we investigated LE, reporter ion missingness, the ratio of mean TMT set MRII to individual channel MRII, and the distribution of log 2 reporter ion ratios of Mixing QC samples. We discovered that sample pH is a critical factor in LE, and increasing the buffer concentration in poorly labeled samples before relabeling resulted in the successful rescue of TMT labeling reactions. Moreover, resuspending peptides in 500 mM HEPES buffer for TMT labeling resulted in consistently higher LE and lower missing data. By better controlling the sample pH for labeling and implementing multiple methods for assessing labeling quality before combining samples, we demonstrate that robust TMT labeling for large-scale quantitative studies is achievable.

Background: There are different methods of Vatsanabha shodhana but it is not mentioned in literature that which method should be used in preparation of Tribhuvankirti rasa which will make the drug safer. Objectives: To acquire knowledge regarding toxic principle in self prepared and market sample of Tribhuvankirti Rasa with respect to different shodhana methods of Vatsanabha and to develop a method of testing toxic principle in the formulation. Material and methods: Three samples were prepared out of which two samples were prepared using different shodhana media and one sample was prepared using impure ingredients and three samples of Tribhuvankirti Rasa were purchased from market. Physicochemical and phytochemical analysis of the final product was conducted and the observations were compared among six samples. Results: HPLC graphs indicate that the numbers of peaks are increased in the sample prepared with Godugdda shodhita Vatsanabha and all the market samples in comparison with Ashodhita and Gomutra shodhita samples of Tribhuvankirti Rasa. All samples contain alkaloids, however after purification there was decrease in the concentration of detected alkaloid. Conclusion: The toxic principle (Aconitine) in market and self prepared sample of Tribhuvankirti Rasa has been reduced by shodhana. In addition, flavonoids, glycosides and anti-oxidants are found which are beneficial for health.

Hemolysis, a common occurrence in blood collected for chemical analysis, has been reported to affect analytical test results for some analytes depending upon the material tested and the analytical technique employed. The potential for hemolysis to impact blood ethanol determinations using headspace gas chromatography of samples diluted with an internal standard was investigated. A sample of non-hemolyzed blood and a matched sample of hemolyzed blood were both analyzed thirty times for ethanol concentration using headspace gas chromatography. The mean ethanol concentration measured for the non-hemolyzed samples was 0.0639 g/dl. The mean ethanol concentration measured for the hemolyzed samples was 0.0642 g/dl. The calculated t value, 1.897, was less than the critical t value, 2.002, at a 0.05 level of significance. There was no measured statistical difference detected between the mean blood ethanol concentration determined for a hemolyzed whole blood sample and a non-hemolyzed whole blood sample.

Evaluating the short tandem repeat (STR) in the field is important for the timely identification of a suspect. Several lines showed that the RapidHIT® ID system is reliable for DNA genotyping with buccal swabs and naked DNA. However, the application of this approach with blood samples has been poorly investigated. Because blood samples are among the most common forensic samples in our laboratory, further studies should be conducted. Here, we assessed the analytical performance of 19 STR loci with a newly developed RapidINTEL (RI) Sample Cartridge Kit by using the blood samples with known genotypes. Several commonly used substrates were included in the sensitivity study, and FTA cards proved to be the most promising sample carrier for blood storage and later identification. There was superior sensitivity and specificity with a 100% concordance rate for 0.5 μL of blood or 7 ng of genomic DNA. The performance for blood samples was comparable with that for the standard protocol. High success rate (90.57%) and high-concordance (100%) genotyping were automatically achieved over a wide range of operating conditions except for TH01. No contamination was observed throughout the study. Hematin, indigo, and humic acid had limited influence on the instrument system, while urea and melanin dramatically affected the genotyping results. Generally, the newly developed RI sample cartridge provided an alternative method for the STR genotyping of single-source blood samples over a wide range of operating conditions.

In the present study, we developed a novel digital coding combination analysis (DCCA) to analyze the gene mutation based on the sample combination principle. The principle is that any numerically named sample is divided into two groups, any two samples are not grouped in the same two groups, and any sample can be tested within the detection limit. Therefore, we proposed a specific combination that N samples were divided into M groups. Then N samples were analyzed, which could obtain the mutation results of M mixed groups. If only two groups showed positive (mutant type) signals, the same sample number from two positive signal groups would be the positive sample, and the remaining samples were negative (wild type). If three groups or more exhibited positive results, the same sample number from three positive signal groups would be the positive sample. If some samples remained uncertain, individual samples could be analyzed on a small scale. In the present study, we used the two genotypes of a mutation site (A5301G) to verify whether it was a useful and promising method. The results showed that we could quantitatively detect mutations and demonstrate 100% consistent results against a panel of defined mixtures with the detection limit using pyrosequencing. This method was suitable, sensitive, and reproducible for screening and analyzing low-frequency mutation samples, which could reduce reagent consumption and cost by approximately 70-80% compared with conventional clinical methods.

Most quantitative research methods are based on measuring either the total or the free concentration of an analyte in a sample. However, this is often insufficient for the study of complex biological systems. The main objective of this research was to develop new methods for providing more information from samples: the free concentration (C(f)), the total concentration (C(t)), and the plasma binding capacity (PBC). Samples were processed using microextraction and ultrafiltration. For each of these techniques, two quantification procedures were used: addition of isotopically labeled standard and repeated analysis of the same sample. The new methods were validated by analyzing clinical samples and samples with known concentrations. Methods based on addition of labeled compound were found to be the fastest, and most reproducible. For analysis of clinical samples, methods based on microextraction were more sensitive and more accurate than those based on ultrafiltration. For analysis of pooled plasma samples, the overall accuracy of all approaches to determine PBC, testosterone C(f), and testosterone C(t) was between 94–109%, 87–113%, and 94–122% respectively. The new approach goes beyond a simple concentration measurement, giving more information from clinical samples, with great potential for personalizing drug dosage and therapy to the needs of individual patients.

This article describes the development of a suitable Gel Permeation Chromatography method for cellulose nitrate plasticised with camphor (celluloid) found in cultural heritage. Current sample preparation and dissolution methods, apart from focusing on native, nonderivatised cellulose, require long preparation times, and often employ solvents that induce degradation. This study aims to develop a systematic method for sample preparation of cellulose nitrate that uses the least sample amount possible, is nondegrading, and can be applied on differently aged samples. This is investigated through identification of a suitable solvent system and a statistically designed experiment testing the critical variables affecting the analysis, namely sample condition, sample, and salt concentration (lithium chloride) in N,N-dimethylacetamide. The use of 0.1% sample was inadequate for analysis because it did not fully dissolve in any salt concentration, while the 0.3% negatively impacted the analysis with its high molecular weight distributions. The 0.2% cellulose nitrate in a solution of 0.5% lithium chloride in N,N-dimethylacetamide offered the most consistent and repeatable molecular weight data. This method miniaturised the sample as much as possible and is suitable for museum objects in various ageing conditions.

In-depth LC–MS-based proteomic profiling of limited biological and clinical samples, such as rare cells or tissue sections from laser capture microdissection or microneedle biopsies, has been problematic due, in large, to the inefficiency of sample preparation and attendant sample losses. To address this issue, we developed on-microsolid-phase extraction tip (OmSET)-based sample preparation for limited biological samples. OmSET is simple, efficient, reproducible, and scalable and is a widely accessible method for processing ~200 to 10,000 cells. The developed method benefits from minimal sample processing volumes (1–3 μL) and conducting all sample processing steps on-membrane within a single microreactor. We first assessed the feasibility of using micro-SPE tips for nanogram-level amounts of tryptic peptides, minimized the number of required sample handling steps, and reduced the hands-on time. We then evaluated the capability of OmSET for quantitative analysis of low numbers of human monocytes. Reliable and reproducible label-free quantitation results were obtained with excellent correlations between protein abundances and the amounts of starting material (R(2) = 0.93) and pairwise correlations between sample processing replicates (R(2) = 0.95) along with the identification of approximately 300, 1800, and 2000 protein groups from injected ~10, 100, and 500 cell equivalents, resulting from processing approximately 200, 2000, and 10,000 cells, respectively.

A method for quantitative determination of the main active ingredient in traditional Chinese medicine was developed and validated, using liquid-liquid chromatography combined with high performance ...

Chromatography is used to separate analytes in a mixture. In the absence of a detector providing analyte-specific information, the retention time is used to identify the analyte, typically by comparing the retention time of the unknown with that of a standard. This is the simplest of identification routines, but it remains the most widely used. The challenge is that analyte retention times can and do shift; sole reliance on retention time for identification is not prudent. Retention time is influenced by the total sample concentration, elution of components near the peak of interest, aging of a column, and so on, complicating time window-based identification. We propose a sameness threshold (ST) that is based on numerical comparison of shapes of chromatographic peaks for identification confirmation. We demonstrate shape-based identification confirmation or lack thereof of an unconfirmed peak by comparison with a shape library of standards. In numerous cases we look at, only one analyte in the shape library fits the putative identity. ST can be based on r2, the commonly used coefficient of determination used in regression analysis, or on an "index of width mismatch" that compares the widths of the peaks throughout the available height range.

Centralized high-throughput purification laboratories routinely produce large numbers of test tubes with fractions containing the purified compounds of interest interspersed with test tubes containing fractions collected from undesired peaks. Because the next step after purification entails the removal of the solvent in a centrifugal evaporator with multiple sample positions per rotor, select test tubes must be labeled prior to dry-down to track the identity of each compound. The diversity of test tube sizes and tray configurations from different chromatography system vendors complicates this labeling task. Therefore, the development of an automated tube labeler that can accommodate a multitude of test tube and tray sizes can reduce the chances of error as well as reduce the hands-on labor required to complete this tedious but essential task. Custom hardware and software have been implemented to inform and to enable the Pick-n-Place arm of a commercially available Tecan EVO robotic system to pick up and present select tubes, filled with purified chromatography fractions from a multitude of vendor trays, to a custom label application station integrated with a commercially available Zebra label printer. Particular challenges existed with accurately positioning tubes in Agilent G1364-84544 trays onto the deck of the instrument. The resulting instrument reduces hands-on time for labeling fractions by approximately 60%.

This paper reports a new method for the determination of sludge water content by a multiple headspace extraction gas chromatographic (MHE-GC) method. It is based on measuring the GC signals for the water vapor in a sample vial from the first five headspace extractions, from which the water content in the sludge sample can be extrapolated according to the established calculation equation. The results show that the method has a good repeatability (the relative standard deviation is less than 1%) and accuracy. The maximum relative difference is less than 16% compared to the reference method. The present method is very simple and efficient, and suitable for rapid sample testing in related applications.

A portable self-flow filtration and separation platform was designed using soft lithography to create a polydimethylsiloxane (PDMS) microfluidic channel cover combined with a matching acrylic substrate. The separation zone was filled with microbeads of appropriate sizes to achieve universal filtration and separation. This simple structure requires only 20 μl of the sample for filtration separation. A vacuum of 760 torr is applied to the porous PDMS cover to drive the sample during testing. The average time required for a 20 μl sample of blood to pass through the separation zone is about 56 s, while the filling time for the detection zone of volume 6 μl is about 319 s. When the hematocrit of the blood sample is about 20-25%, the separation efficiency is 99.98%. Further, the separation efficiency of fat globules from raw milk is close to 100%, whereas almost all impurities are filtered out from juice and stool samples. It is also observed that E. coli in the stool can pass from the separation to detection zone at a maximum rate of about 81.21%, with an average of about 68.18%.

While meeting the pandemic demand of SARS-CoV-2 testing, clinical laboratories worldwide tend to adopt new test systems offering cost-effective and faster test outcomes. However, the reliability of SARS-CoV-2 test results has paramount importance in the management of such a health crisis. Therefore, this study sought to determine the accuracy of the test results from a novel duplex Microchip RT-PCR test system using patient saliva samples and nasal swabs stabilized in Viral Transport Medium (VTM) with reference threshold Cycle Values (Ct). The VTM used to stabilize these samples during transport was found to be inhibitory to the RT-PCR. Therefore, all the samples were subjected to spin column purification of total RNA to remove the influence of VTM. A total of 70 patient samples, including 24 positive- and 31 negative-saliva in VTM samples and 15 positive nasal swab samples, were tested. Results obtained from both the sample types were compared to their reference values and no false positive or false negatives were observed. From this data, accuracy, specificity, and sensitivity were determined to be 100% applying the corresponding formulae. The limit of detection with 95% confidence probability was determined to be 2.5 copies/μl in the original sample.

The bottom-up strategy for proteome analysis typically employs a multistep sample preparation workflow that suffers from being time-consuming and sample loss or contamination caused by the off-line manual operation. Herein, we developed a hollow fibre membrane (HFM)-aided fully automated sample treatment (FAST) method. Due to the confinement effects of HFMs and the immobilized enzymatic reactor, the proteome samples could be denatured, reduced, desalted and digested within 8–20 min via the one-stop service. This method also showed superiority in trace sample analysis. In one and half hours, we could identify about 1,600 protein groups for 500 HeLa cells as the starting materials, 1.5–8 times more than those obtained by previously reported methods. Through the on-line combination of FAST with nano-liquid chromatography-electrospray ionization tandem mass spectrometry (nanoLC-ESI-MS/MS), we further established a fully integrated platform for label-free quantification of proteome with high reproducibility and precision. Collectively, FAST presented here represents a major advance in the high throughput sample treatment and quantitative analysis of proteomes.

Environmental DNA (eDNA) is increasingly popular as a useful non-invasive method to monitor and study biodiversity and community structure in freshwater and marine environments. To effectively extract eDNA from the filter surface is a fundamental factor determining the representativeness of the samples. We improved the eDNA extraction efficiency of an established Sterivex method by 12- to 16-fold using a larger volume of lysis buffer mix coupled with backflushing the cartridges. The DNeasy extraction column could be overloaded when the environmental sample input is high, possibly due to a higher nonspecific binding present in environmental samples, thus resulting in a relatively lower quantity measured. Therefore, we included an internal control DNA in the extraction to monitor the extraction and purification efficiencies in field samples, which is crucial for quantification of original eDNA concentration. The use of Takara Probe qPCR Mix supplemented with protein-based additives improved the robustness of the real time PCR assay on inhibitor-rich environmental samples, but prior purification by Qiagen PowerClean Pro Cleanup kit could be essential for inhibitor-rich water samples, even though the recovery rate was unexpectedly low (average 33.0%). The improved extraction and quantification complement the qualitative analyses including metabarcoding and metagenomics in field application.

Contamination from the polymeric material released by vial caps used for sample introduction in liquid chromatography can significantly affect the signal of the analyte of interest. In particular, repeated injections from the same sample vial can enhance this suppressing effect. Multiple injections of the same sample are often used in metabolomics and lipidomics during routine analyses. Here we demonstrate how the presence of contaminant polymers, originating from the vial closures, significantly influences the estimation of the relative amount of endogenous lipids in human plasma. Furthermore, this can negatively impact other operations in mass spectrometric analysis, such as instrument equilibration and tuning or the common use of technical replicates to improve confidence in data interpretation. Our observations provide critical information on how to improve future analyses through the use of appropriate vial caps, solvents, chromatographic separations and equipment.

There is a need for biosensing systems that can be operated at the point-of-care (POC) for disease screening and diagnostics and health monitoring. In spite of this, simple to operate systems with the required analytical sensitivity and specificity in clinical samples, using a sample-in-answer-out approach, remain elusive. Reported here is an electrochemical bio-barcode assay (e-biobarcode assay) that integrates biorecognition with signal transduction using molecular (DNA/protein) machines and signal readout using nanostructured electrodes. The e-biobarcode assay eliminates multistep processing and uses a single step for analysis following sample collection into the reagent tube. A clinically relevant performance for the analysis of prostate specific antigen (PSA) in undiluted and unprocessed human plasma: a log-linear range of 1 ng mL-1 -200 ng mL-1 and a LOD of 0.4 ng mL-1 , was achieved. The e-biobarcode assay offers a realistic solution for biomarker analysis at the POC.

Accurate clotting time assay results are vital, as the test is employed to indicate the amount of oral anticoagulant to be prescribed, while it is also used for screening the hemorrhagic and thrombotic diseases. The procedure chosen for preparation of a patient blood sample including centrifugation can contribute to significant differences in the results obtained. Thus, for the purpose of proposing a standardized method to appropriately prepare blood samples prior to assay, the Japanese Society of Laboratory Hematology organized the Working Group for Standardization of Sample Preparation for Clotting Time Assays (WG). Following reviews of previously announced guidelines and original experimental results, consensus was obtained by the WG, with the main findings as follows. (1) The recommended anticoagulant in the blood collection tube is sodium citrate solution at 0.105–0.109 M (3.13–3.2%). (2) Whole blood samples should be stored at room temperature (18–25 ˚C) within 1 h of collection from the patient. (3) For plasma preparation, centrifugation at 1500 × g should be performed for at least 15 min or at 2000 × g for at least 10 min at room temperature. (4) After the plasma sample is prepared, it should be stored at room temperature and assayed within 4 h.

N-acetylaspartate is a human endogenous compound synthesized by neurons, which is involved in neuronal metabolism. It is used as a marker in brain magnetic resonance spectroscopy to investigate several neurological and metabolic disorders, that can be related to a variation of its concentration with respect to reference values. N-acetylaspartate is present also in biological fluids, such as plasma, urine, and cerebrospinal fluid, where it can be quantified. Here we describe the development and validation, in compliance with the EMA guidelines, of a novel assay method for the quantification of N-acetylaspartate in plasma based on tandem mass spectrometry coupled to liquid chromatography. Its peculiarity lies in the fact that sample preparation includes an esterification step, which significantly improves the chromatographic performances and, consequently, also the method sensitivity, reproducibility and accuracy. Instrumental LLOQ is 0.06 ng/mL, i.e. at least 300 times lower than the medium N-acetylaspartate concentration in samples, accuracy is in the range 98–103%, while precision lies between 1 and 3%. The method robustness was tested in about 1000 injections of plasma samples, 96 of which were used also to assess the reference ranges in control subjects (16.46–63.40 ng/mL). Controls were then compared to plasma samples from type 2 diabetic patients. Contrary to brain magnetic resonance spectroscopy, which demonstrated a decrease in the N-acetylaspartate levels in right frontal and parieto-temporal region of type 2 diabetic patients, plasma analysis showed no statistical difference with respect to controls. However, the method here described can be profitably used in studies concerning different disorders with CNS involvement, as confirmed by reports available in the literature.

Potential sources of a specific and considerable contamination with bisphenol A (BPA) via biological sample testing were investigated. Determination of BPA in animal plasma, collected in a commerci...

Objectives A matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) method (Mass-Fix) as a replacement for gel-based immunofixation (IFE) has been recently described. To utilize Mass-Fix clinically, a validated automated method was required. Our aim was to automate the pre-analytical processing, improve positive specimen identification and ergonomics, reduce paper data storage and increase resource utilization without increasing turnaround time. Methods Serum samples were batched and loaded onto a liquid handler along with reagents and a barcoded sample plate. The pre-analytical steps included: (1) Plating immunopurification beads. (2) Adding 10 μl of serum. (3) Bead washing. (4) Eluting the immunoglobulins (Igs), and reducing to separate the heavy and light Ig chains. The resulting plate was transferred to a second low-volume liquid handler for MALDI plate spotting. MALDI-TOF mass spectra were collected. Integrated in-house developed software was utilized for sample tracking, driving data acquisition, data analysis, history tracking, and result reporting. A total of 1,029 residual serum samples were run using the automated system and results were compared to prior electrophoretic results. Results The automated Mass-Fix method was capable of meeting the validation requirements of concordance with IFE, limit of detection (LOD), sample stability and reproducibility with a low repeat rate. Automation and integrated software allowed a single user to process 320 samples in an 8 h shift. Software display facilitated identification of monoclonal proteins. Additionally, the process maintains positive specimen identification, reduces manual pipetting, allows for paper free tracking, and does not significantly impact turnaround time (TAT). Conclusions Mass-Fix is ready for implementation in a high-throughput clinical laboratory.

Tissue samples are widely used for the study of metabolomics, lipidomics and proteomics due to their abundant useful biomolecular information. Modern analytical mass spectrometry has the ability to directly obtain the molecular information contained in various tissue samples (e.g., plant tissues, animal tissues, and clinical tissues, etc.) in real-time, with minimal or no sample pretreatment. Such features of analytical mass spectrometry promote the throughput of tissue analysis and extend the application of tissue analysis in multiple fields. This review summarizes the applications of mass spectrometry in direct analysis of plant tissues, animal tissues and clinical tissues, and the challenges and perspectives of mass spectrometry in direct analysis of tissue samples are discussed as well.

In forensic sciences, Blood and saliva are two of the most examined biological samples used in forensic toxicology as they essentially contains all the information (in the form of DNA as well as certain other chemical compounds like salivary amylase etc.) that is needed to narrow down possible suspects and also to determine the identity of the true criminal. There is a time gap between collection of blood from the criminal site and its analysis in laboratory so it is very important to store and handle the biological specimen appropriately in order to maintain its stability. This paper presents a review on storage of biological sample mainly blood and saliva in different temperature.

WOS:000560662700007 Doxylamine, which is an active drug ingredient used for insomnia, is an antihistaminic with sedative effect and can be received without a prescription. Although doxylamine succinate intoxication is encountered quite frequently, there is no data that shows the drug is used for intentional injury. This study was conducted to unravel a suspected case of intentional poisoning facilitated with the use of doxylamine, using a fast, accurate and reliable chromatographic method. The complainant realized that she had fallen asleep after drinking the tea given to her by her colleague, who has been treating her badly for some time. After experiencing a second similar situation, she asked for toxicological analysis. The analysis revealed the presence of 69.3 mu g/mL (13.9 mg/cup) of doxylamine in the sample, and it was concluded that this amount was sufficient to produce the symptoms the complainant mentioned. However, this study reveals that the complainant's claim is quite difficult to prove, and toxicological analysis alone are not enough to prove her claim. Considering the possibility of crimes of intentional injury, aspersion and torment on a legal basis, this study thought to be remarkable as it presents a case of suspected crime facilitated with the use of an active drug ingredient. Scientific Research Projects Unit of Istanbul UniversityIstanbul University [TSA-2017-24125] The Scientific Research Projects Unit of Istanbul University -Cerrahpasa with the project numbered TSA-2017-24125, supported this study.

This work illustrates four different spectrophotometric methods for metformin (MET) and canagliflozin (CANA) quantification in their recently approved pharmaceutical with trade name Invokamet®. The proposed methods are simple, definitive and accurate. Certain ensuing steps implementing zero spectra and/or ratio and/or derivative spectra were applied. Via implementing spectrum subtraction method accompanied with constant multiplication, zero order spectra were obtained for these drugs followed by their determination at their correlated λmax at 237 and 290 nm for MET and CANA, respectively. Meanwhile, through employing method of derivative subtraction the two drugs were obtained in their first derivative spectra and quantified at 242.7 and 319.0 nm for MET and CANA, respectively. The suggested methods were inspected via laboratory prepared (diverse ratios) mixtures and they were excellently applied for analysis of Invokamet® tablets. Moreover, sample enrichment via spiking technique was elected for a pharmaceutical formulation analysis containing CANA as a minor component. Accuracy, precision and specificity were between the valid limits. Validation steps were done in accordance with the ICH guidelines. Also, statistical compression was carried out between the obtained and reported results and no crucial divergence appeared.

The method of capillary electrophoresis can be used to assess the quality, safety, falsification and identification of food and beverages. It is characterized by high accuracy (up to 97%) and efficiency. Using it, you can determine the content of various cations and anions, organic acids, carbohydrates, vitamins, amino acids, preservatives, antioxidants, flavonoids, synthetic dyes, which are of great importance for the technological control of production at the enterprise. The method of capillary electrophoresis allows you to determine the naturalness and quality of soft drinks and alcoholic drinks by detecting the presence or absence of organic acids in the sample, as well as analyzing their quantitative content and ratio, makes it possible to control enzymatic processes and correlate with the taste of the final product. In addition, this method can be used to assess the quality of water used as the main raw material for the production of beverages. It allows you to determine the content of pesticides and fungicides in the raw materials used for the production of food and beverages. The method is based on the electromigration of ions and electroosmosis and, as a consequence, the separation of a complex multicomponent system in a quartz capillary. The bottom line is that after applying voltage up to 30 kV to the prepared capillaries for analysis, the components of the mixture begin to move along the capillary at different speeds, thereby reaching the detection zone at different times. Thus, separation and subsequently detection of the test substances in a particular product, drink or raw material is achieved.

The objective of this study is to evaluate in vitro stability of cocaine compounds, cocaine (COC), benzoylecgonine (BE), ecgonine methyl ester (EME) and benzoylecgonine ethyl ester (EBE), in blood and urine, during post-analysis custody. Stability was evaluated by measuring percent recovery. Parameters evaluated were time of custody (1 year), storage temperature (−20°C and 4°C), influence of preservative (only for blood samples) and pH (only for urine samples). The impact of the temperature is very important in blood samples. At −20°C all compounds demonstrated to be stable, with recoveries higher than 80% after 1 year. In contrast, degradation was observed in the concentration for all four compounds when the samples were maintained at 4°C. In these same conditions, the influence of the preservative was also noticeable and a higher stability was found in samples preserved with NaF. COC and EBE had similar profiles, and both compounds disappeared after 30 days in samples without NaF and after 150 days in samples with NaF added. EME disappeared after 185 days and after 215 days in samples without and with preservative, respectively. BE recoveries, after 365 days of storage, were 68.5% (in samples with NaF) and 3.7% (in samples without NaF). In urine samples, the four compounds were stable in all the studied conditions except when samples were at pH 8 and stored at 4°C where the compounds disappeared (COC and EBE after 75 days of storage and EME after 15 days). The exception was BE, with a recovery of 23% after 1 year of storage. Of the temperatures evaluated, −20°C seems to be optimal for storage to maintain the stability of cocaine and metabolites in biological samples. This can be further enhanced by maintaining a pH of 4 in urine samples and adding a NaF preservative to blood.

Because of the inherent affinity of proteins for bare, fused silica capillaries, the analysis of protein-containing samples has proven a challenging task for capillary electrophoresis. The adsorption of proteins to the capillary walls effectively changes the zeta potential and thus affects the electro-osmotic flow leading to significant shifts in migration time, peak broadening, and poor reproducibility. While there are several well-known methods to remove proteins from samples prior to the analysis (including precipitation) or to prevent their adsorption to the capillary (semi-permanent coatings), those approaches are often expensive, time consuming, or simply unreliable. Aiming to address these needs, this manuscript reports on the use of pyrolyzed cotton balls, as a simple and widely accessible hydrophobic material to remove proteins from serum samples. The material retains enough flexibility so it can be placed directly into the sample vials and has enough capacity to capture more than 75% of the proteins in the sample (1% dilution of 1 mL of serum). The advantages of the material are demonstrated by performing the analysis of five representative drugs (in serum) by capillary electrophoresis obtaining a change in migration time of only 5 ± 1%, after 10 consecutive runs.

There are so many ways to lose weight, such as consuming fat burner supplements to burn fat faster. A fat burner supplement consists of various ingredients, such as caffeine. Fat burner supplements are usually not registered in BPOM RI. This study aims to evaluate caffeine in fat burner supplements qualitatively and quantitatively. These supplements were analyzed in the pharmaceutical laboratory in Universitas Muhammadiyah Yogyakarta using Thin Layer Chromatography (TLC)–Densitometry and UV–Vis Spectrophotometer. Sample preparation was conducted by separating caffeine from supplements by a separatory funnel with chloroform as an organic solvent. Qualitative analysis was carried out by TLC and analyzing standard spectrum, and sampling technique was carried out with UV–Vis Spectrophotometer. The first quantitative analysis used densitometry by measuring the spot on the TLC plate. Meanwhile, the second quantitative analysis used UV–Vis Spectrophotometer by observing absorbency value on samples with λ 273.5 nm. The result of the qualitative test using TLC was analyzed by comparing the Rf value of the standard and the sample. The Rf value of caffeine standard was 0.63, and every sample showed similar value with caffeine standard, indicating that all samples contain caffeine. The result of quantitative test with TLC - Densitometry method revealed 1 st sample 5.68 mg/ml, 2 nd sample 5.74 mg/ml, 3 rd sample 3.43 mg/ml, 4 th sample 8.90 mg/ml, and 5 th sample 1.88 mg/ml. The qualitative test result was analyzed using the UV–Vis Spectrophotometer method, and all of the caffeine standard spectra can be read at wavelength 273.5 nm, which means all samples contain caffeine. The second quantitative test result analyzed by using UV–Vis Spectrophotometer method showed 1 st sample 3.22 mg/g, 2 nd sample 4.56 mg/g, 3 rd sample 2.23 mg/g, 4 th sample 11.22 mg/g, and 5 th sample 0.26 mg/g. It can be concluded that all samples (fat burners) contained caffeine.

The advantages of analysis of drugs in hair samples are recognized for the long window of detection, alongside easy sampling and long stability after sample collection. Alcohol markers, ethyl glucuronide (EtG) and total fatty acid ethyl esters (FAEEs) in hair, are widely used for monitoring alcohol consumption for clinical and forensic purposes. Although stability of drugs and EtG in hair samples is documented to a certain extent, stability of FAEEs in hair samples after collection has not been reported. This study covered hair samples that had been tested for FAEEs on the day of arrival at the laboratory and retested between 4 and 80 months later. The statistical analysis of the data set reveals significant lower FAEEs levels including ethyl palmitate (EtPa) ester levels when samples were retested for the second time after 6 days of storage under ideal conditions. Specifically, the results suggest that when measuring total FAEEs or solely EtPa in hair samples, the elapsed time between sample collection and analysis of the sample needs to be considered when interpreting the results. The recommendation is that whenever hair samples need to be tested for total FAEEs or EtPa, the analytical procedure needs to be performed within 1 week after collection in order to obtain meaningful results. The study results substantiate the case for the use of hair samples solely for the analysis of EtG, in conjunction with other measurements such as full blood count, carbohydrate-deficient transferrin test, liver function test or phosphatidylethanol alongside clinical assessment for a more effective evaluation of alcohol consumption.

Among the various strategies to enhance performance by the use of doping substances or methods, sample manipulation to cheat common detection strategies is prohibited by the World Antidoping Agency. Two cases of urine sample adulteration with surfactants are presented in this article, as well as an easy strategy for the detection of this sort of sample manipulation.