Mean Recovery Research Articles

Simultaneous determination of protoporphyrin IX and magnesium protoporphyrin IX in Arabidopsis thaliana and Camellia sinensis using UPLC-MS/MS

BackgroundsInsertion of Mg2+ into protoporphyrin IX (PPIX) to produce magnesium-protoporphyrin IX (Mg-PPIX) was the first step toward chlorophyll biosynthesis, which not only imparts plants green pigmentation but underpins photosynthesis. Plants that blocked the conversion of PPIX to Mg-PPIX displayed yellowish or albino-lethal phenotypes. However, the lack of systematic study of the detection method and the metabolic difference between species have caused the research on chloroplast retrograde signaling controversial for a long time.ResultsAn advanced and sensitive UPLC-MS/MS strategy for determining PPIX and Mg-PPIX was established in two metabolic different plants, Arabidopsis thaliana (Columbia-0) and Camellia sinensis var. sinensis. Two metabolites could be extracted by 80% acetone (v/v) and 20% 0.1 M NH4OH (v/v) without hexane washing. Since the Mg-PPIX could be substantially de-metalized into PPIX in acidic conditions, analysis was carried out by UPLC-MS/MS with 0.1% ammonia (v/v) and 0.1% ammonium acetonitrile (v/v) as mobile phases using negative ion multiple reaction monitoring modes. Interestingly, it could be easier to monitor these two compounds in dehydrated samples rather than in fresh samples. Validation was performed in spiked samples and mean recoveries ranged from 70.5 to 916%, and the intra-day and inter-day variations were less than 7.5 and 10.9%, respectively. The limit of detection was 0.01 mg·kg− 1 and the limit of quantification was 0.05 mg·kg− 1. The contents of PPIX (1.67 ± 0.12 mg·kg− 1) and Mg-PPIX (3.37 ± 0.10 mg·kg− 1) in tea were significantly higher than in Arabidopsis (PPIX: 0.05 ± 0.02 mg·kg− 1; Mg-PPIX: 0.08 ± 0.01 mg·kg− 1) and they were only detected in the leaf.ConclusionsOur study establishes a universal and reliable method for determining PPIX and Mg-PPIX in two plants using UPLC-MS/MS. This procedure will facilitate studying chlorophyll metabolism and natural chlorophyll production.

Synthesis and Characterization of Magnetic Molecularly Imprinted Polymer for the Monitoring of Amoxicillin in Real Samples Using the Chromatographic Method

Amoxicillin (AMX) is an antibiotic frequently used for the treatment of bacterial disorders and respiratory problems in both humans and animals. This work aims to synthesize a molecularly imprinted superparamagnetic polymer (SP-MIP) with a core-shell structure for the selective detection of AMX in real samples. Magnetite superparamagnetic nanoparticles (SNP) were prepared by the polyol method, coated with silica, and functionalized with silane groups. The polymerization process was executed using the free-radical precipitation method. Thermogravimetric analysis (TGA) was used to evaluate the thermal stability of the synthesized materials. The results obtained from N2 adsorption and desorption analyses showed that the surface area of SP-MIP (19.8 m2/g) was higher than that of the non-molecularly imprinted superparamagnetic polymer (SP-NIP—9.24 m2/g). The optimized adsorption analysis showed that both SP-MIP and SP-NIP followed SIP-type behavior, with adsorption constant KS 0.01176, 1/n 1.73. The selectivity tests showed that SP-MIP is highly selective for AMX in the presence of other molecules. Finally, for the recovery analysis, the application of SP-MIP for determining AMX in samples of tap water, river water, and drugs using HPLC yielded a mean recovery value of 94.3%.

Spinal Versus General Anesthesia for Spine Surgery During the COVID-19 Pandemic: A Case Series.

Hospitals are one of the primary resources for disease transmission, so many guidelines were published, and neurosurgeons were advised to postpone elective spine surgeries during the COVID-19 pandemic. To avoid pulmonary complications and reduce the risk of spreading the virus and contracting the disease during the COVID-19 era, we operated a group of our patients under spinal anesthesia rather than general anesthesia. We retrospectively analyzed all patients who underwent discectomy surgery for lumbar spinal disc herniation under SA between September 2020 and 2021. Sixty-four patients diagnosed with lumbar disc herniation underwent lumbar discectomy with SA. All patients except three were male. The mean age was 44.52 ± 7.95 years (28 to 64 years). The mean procedure time for SA was 10 minutes. The duration of the surgery was 40 to 90 minutes per each level of disc herniation. The mean blood loss was 350 cc (200 to 600 cc). The most common involved level was L4/L5 intervertebral disc (n = 40 patients; 63.5%). The mean recovery time was 20 minutes. Only three patients requested more analgesics for relief of their pain postoperatively. All patients with discectomy were discharged a day after surgery, and in the case of fusion, two days after surgery. All the patients were followed up for six months, showing no recurrence symptoms, good pain relief, satisfaction with the surgery, and no bad memory of the surgery. Spinal anesthesia is a good alternative or even the main anesthesia route for patients with lumbar disc herniation. More studies are needed to elucidate the best candidate for SA in patients with lumbar pathology.

Determination of Cations and Anions in Desulfurization Waste Water of Coal-Fired Power Plant by Ion Chromatography.

The reliable and accurate determination of corrosive anions at sub- to low-μg/L levels is a challenging analytical problem. In this manuscript, a simultaneous determination method of cations and anions in power plant water samples was established by large volume injection ion chromatography. The analytical parameters including separation column, the suppressor current and the elute concentration were optimized. Results showed good separation under the optimum conditions, and the calibration curves of all analytes were linear with good coefficient of determination (r2) >0.997, and the mean recoveries for all analytes ranged from 75.62 to 118.58% with RSD of 0.07-4.83%. The established method was successfully applied to determine the cations and anions in realwaste water samples from coal-fired power, and was verified by inductively coupled plasma optical emission spectrometry and electrometric titration. The relative deviation between methods was all below 6.72%, which indicated good accuracy of the established ion chromatography method. The results could also provide reference for the precise and rapid detection of cations and anions in environmental water samples.

Comparing Recovery Methods for Wastewater Surveillance of Arthropod-Borne and Enveloped Viruses

The potential public health utility of arthropod-borne virus (arbovirus) wastewater surveillance (WWS) warrants the effort of establishing an effective sample preparation workflow. Here, we investigated different sample clarification, concentration, and extraction methods to determine the best workflow for recovery of spiked arboviral and murine hepatitis virus RNA signals in wastewater. We determined that the sample clarification method with the least loss is filtration through a 0.22 μm filter (9.06 ± 3.68%) when compared with centrifugation. Out of four different concentration methods (centrifugal ultrafiltration, polyethylene glycol precipitation, charged membrane capture and elution, and hollow-fiber ultrafiltration), centrifugal ultrafiltration, specifically using Macrosep Advance column (30 kDa), has the highest mean recovery of 84.76 ± 17.45%. With this method, we can reliably recover seeded arboviruses at concentrations corresponding to 10 infections in 400 to 180,000 people, depending on the shedding rate. As arboviruses are expected to be on the rise due to the expansion of their vectors’ geographic distribution, this research will lay the foundation for managing and reducing the impact of arboviral outbreaks through WWS-informed public health measures.

Simultaneous Estimation of Ampicillin and Sulbactam in Human Plasma by Liquid Chromatography Tandem Mass Spectrometry

A LCMS/MS method for the simultaneous determination of ampicillin and sulbactam in human plasma was described. After protein precipitation using 2mL of acetonitrile, 250µL of supernatant was mixed with 1.000 mL of 0.1% Acetic Acid in Milli-Q-water. 10µL was injected to a Biobasic AX column and eluted with 10mM Ammonium acetate and Acetonitrile: 60:40, v/v at a flow rate of 0.5mL/min. MRM transitions were monitored in negative mode as m/z 348.1 → 206.8 (AMP), 231.9 → 187.8 (SUL) and m/z 353.0 → 211.9 (AMP D5). Sample concentrations were calculated by linear regression analysis using the analyst software1.6.3. An excellent linear response was obtained over the concentration ranges 0.1040µg/mL to 10.1562µg/mL for Ampicillin and 0.0510µg/mL to 6.1552µg/mL for Sulbactam. The intra-day and inter-day precision were within 3.50% for all analytes. The assay accuracy was 96.27 –103.59 %. Mean recoveries were 84.51% and 98.54% for ampicillin and sulbactam, respectively. The limits of detections were 0.026µg/mL and 0.013µg/mL for ampicillin and sulbactam. This method was successfully used for a bioequivalence study.

Analytical methods for determination of closantel enantiomers by HPLC and their residual elimination profiles in ten tissues of black goats

Closantel is a halogenated derivative compound with a salicylanilide-base, broad-spectrum, potent anthelmintic against several types of trematodes, nematodes, and other external parasites. It is a racemate of an equimolar mixture of two enantiomers, mainly used in livestock farming. It is significant to develop the analysis methods of closantel enantiomers in animal tissues. In this study, a reliable, precise, and selective HPLC method was established to separate and determine closantel enantiomers in ten tissues of black goats. The mean recoveries for closantel enantiomers were between 70.0% and 93.8%, and the limits of quantification were 0.5 µg/g. The proposed method was successfully applied to investigate the distribution and residual elimination of closantel enantiomers in ten edible tissues of black goats following an intramuscular injection of 5 mg/kg body weight of closantel sodium. The results revealed that closantel was stereoselective, and the two closantel enantiomers were eliminated slowly, with half-lives ranging from 9 to 14 days in the tissues of black goats. These findings would support further evaluation of the enantioselective pharmacokinetics, pharmacodynamics, and toxicity of individual closantel enantiomers in food-producing animals.

Surgical Management of Cervical Ossification of Posterior Longitudinal Ligament: The Treatment Algorithm and Outcome.

Introduction Managing patients who presentwith symptoms of cervical myelopathy secondary to cervical ossification of the posterior longitudinal ligament (OPLL) is challenging. Various factors such as the number of levels involved with OPLL, types of OPLL, canal occupying ratio, K-line characteristics, and C2-C7 lordosis angle were found to guide decision-making and surgical approaches in managing this condition. However, no clear treatment algorithm has been published. This study aims to investigate the outcome of the management of cervical OPLL using a treatment algorithm used in a tertiary university hospital. Methods This is a retrospective cross-sectional study. Patients with cervical myelopathy secondary to cervical OPLL who were treated surgically in our center from 2014 to 2020 were included in this study. Demographic data and preoperative parameters that determined the treatment given according to our treatment algorithm were analyzed. Result A total of 24 patients fit the inclusion and exclusion criteria of the study. The mean recovery rate for all groups is 61.8[Formula: see text]21.9%and the mean postoperative neck disability index (NDI) is 17.83[Formula: see text]16.67%. There was a statistically significant difference between preoperative and postoperative Japanese Orthopaedic Association (JOA) scores for both anterior and posterior surgery subgroups. Conclusion We believe that the treatment algorithm used in our center could benefit other surgeons as a guide in managing patients who suffer from cervical myelopathy secondary to cervical OPLL. Further study including newer techniques would increase the surgeon's arsenal in providing the best outcome in managing this condition.

Establishment of Residual Methods for Matrine in Quinoa Plants and Soil and the Effect on Soil Bacterial Community and Composition

A method was developed for the determination of matrine residues in quinoa (Chenopodium quinoa Willd.) plants and soil by liquid chromatography triple quadrupole tandem mass spectrometry (LC-MS/MS) with QuEChERS clean-up. Matrine from soil, quinoa roots, stems, leaves and seeds was extracted with 25% ammonia, 20 mL acetonitrile/methanol, salted with sodium chloride (NaCl) and purified with anhydrous magnesium sulfate (MgSO4), N-propyl ethylenediamine (PSA) and graphitized carbon black (GCB). Then a chromatographic separation was performed on a Shim-pack XR-ODS II (75 mm × 2.0 mm, i. d., 2.2 µm) column with a gradient elution of 5 mmol/L ammonium formate-methanol as the mobile phase and monitored in multiple reaction monitoring modes (MRM) in electrospray positive ionization mode. The results showed that in the range of 0.005~1 mg/L, the linear correlation coefficients of matrine in the five matrices were all above 0.999. The LOQs for soil, quinoa roots, stems, leaves and seeds were 0.005, 0.005, 0.01, 0.01 and 0.005 mg/kg, respectively. The mean recoveries ranged from 74.42% to 98.37%, with RSDs of 1.25-6.84% at the three concentration addition levels. The average intra-day and inter-day recoveries were 73.92-92.36% and 78.56-90.18%, respectively, with RSDs below 8.72% and 9.43%. The recoveries and reproducibility of the method were superior. The method was used to determine the actual samples, which indicated that the half-lives of matrine in quinoa seeds, leaves, stems and soil were 1.28-1.32, 1.03-1.21, 0.81-0.92 and 0.93-0.97 d. It has a half-life below 30 d, which is an easily dissipated pesticide. The method is simple, sensitive, accurate, reliable and applicable to a wide range of applications, and it can achieve the rapid multi-residue determination of matrine to a certain extent. Next Generation Sequencing was used to explore the effects of exposure to high and low doses of matrine on soil bacterial communities and the composition of the three soils in the Qinghai Province (Haixi, Haidong and Haibei). The results showed that the number of ASVs increased significantly after treatment with matrine at an effective dose of 0.1 mg/kg than after treatment with matrine at an effective dose of 5.0 mg/kg. Similarly, bacterial abundance was higher after 0.1 mg/kg of matrine treatment than after 5.0 mg/kg of matrine treatment. The inhibitory effect on some bacterial flora was enhanced with an increase in matrine application, while the inhibitory effect on bacterial flora was weakened with time. Applying a certain dose of matrine e changed the relative abundance of the dominant bacterial genera of the soil bacteria.

DEVELOPMENT OF METHOD FOR QUANTITATIVE DETERMINATION OF CEFQUINOME SULFATE IN PIGLETS BLOOD PLASMA USING HPLC/DMD

This manuscript presents a developed method for determining cefquinome sulfate in piglets blood plasma intended for clinical and pharmaceutical research of veterinary drugs based on it. Blood plasma proteins were precipitated twice with a solution of trichloroacetic acid. The supernatant was further purified by a series of solid-phase extraction. Separation was performed on an inverted phase Kinetex EVO С18 column using acetonitrile and 0,1 % trifluoroacetic acide solusion as the mobile phase. The gradient mode of eluents was used during 10 min at a flow rate of 1,4 ml/min. The peak retention time of cefquinome sulfate is 4,2 min. The specificity of the analytical method was checked by comparing the chromatographic separation of a sample of blood plasma enriched with a standard solution of cefquinome sulfate and a sample of blood plasma placebo. The preparing loaded blood plasma samples procedure for building a calibration graph is described in the article. The validation parameters of the method “recovery” and “coefficient of variation” were considered in accordance with the criteria of Council Directive 2002/657/EC. The procedure of sample preparation of fortified blood plasma to construct calibration graph is described in the manuscript. The mean recovery from fortified blood plasma samples in the range of 0.1-2.0 μg/ml cefquinom sulfate was 102.3 %. The method is linear in the concentration range of 0.1 – 4.0 μg/ml of cefquinome sulfate. The correlation coefficient for the determination method is 0.9998. The results obtained in the study of the linearity of this technique were used to estimate the correctness and convergence. The accuracy of the measurements was evaluated by examining the known amounts of analyte added to the control blood plasma. Recovery data are acceptable because they are within ± 10% of the target value. The method has sufficient convergence (accuracy). The evaluation of the intermediate precision of cefquinome sulfate determination was assessed on three different days of analysis. The limit of detection for cefquinom sulfate is 0.05 μg/ml and limit of quantification - 0.10 μg/ml. The average CV for each compound was <10%. The procedure was confirmed and then applied to determination cefquinome sulfate in the pig blood plasma obtained during the study of the pharmacokinetics of the veterinary drug. The HPLC/DMD method can be used for study of the pharmacokinetics of the veterinary drug.

A NOVEL RP-HPLC GRADIENT ELUTION TECHNIQUE FOR BIOANALYTICAL METHOD DEVELOPMENT AND VALIDATION FOR ESTIMATING GALLIC ACID IN WISTAR RAT PLASMA

Objective: Present study aimed to develop and validate a novel, unique, simple, quick, cost-effective, sensitive, specific, accurate, precise, rugged, and robust bioanalytical method for the quantification of gallic acid in rat plasma by reverse phase high-performance liquid chromatography (RP-HPLC) using gradient elution technique. Methods: The stationary phase was a Zorbax SB C18 5 µ (4.6*150) mm column, with the mobile phase being water with 0.1 percent formic acid (A): acetonitrile (ACN) with 0.08 percent formic acid (B). Gradient chromatographic method was used throughout this experiment from the point of view of the estimation of gallic acid from herbal formulations when present along with other phytoconstituents. So at the gradient method, all the present phytoconstituents has cleared off from the column and no any strongly adsorption of phytoconstituents occurred. The experiment was carried out at a flow rate of 1.0 ml/min at 30 °C utilising PDA detectors at 271 nm. The proposed method was validated for different parameters. Results: The approach was found to be linear in the concentration range of 0.5-100 µg/ml, with a r2 of 0.9998. There was not observed any interference of co-eluting peaks of endogenous compounds from the biological matrix at the same retention time (Rt) of gallic acid. The RSD (%) of intra and interday precision was found to be within acceptable limit. The overall % mean recovery was found to be 99.97%. LOD and LOQ were found to be 0.1 and 0.5 μg/ml, respectively. In terms of fluctuation in essential parameters and operating settings, the devised bioanalytical approach was shown to be rugged and resilient. Short-term, long-term, autosampler, bench-top, and freeze-thaw stability experiments revealed that gallic acid is stable. Conclusion: The developed method described in this report was found to be well within an acceptable range. Hence, in the future, this method can be used successfully for the estimation of gallic acid alone or in combination with another analyte or marker present in bulk or an extract containing various phytoconstituents in pharmacokinetic, bioequivalence, and therapeutic drug monitoring studies in clinical laboratories.

In Vitro Viral Recovery Yields under Different Re-Suspension Buffers in Iron Flocculation to Concentrate Viral Hemorrhagic Septicemia Virus Genotype IVa in Seawater.

Iron flocculation is widely used to concentrate viruses in water, followed by Fe-virus flocculate formation, collection, and elution. In the elution stage, an oxalic or ascorbic acid re-suspension buffer dissolved iron hydroxide. After the concentration of viral hemorrhagic septicemia virus (VHSV) in seawater (1 × 101 to 1 × 105 viral genome copies or plaque-forming unit (PFU)/mL), the recovery yield of the viral genome using quantitative real-time PCR (qRT-PCR) and viral infectivity using the plaque assay were investigated to evaluate the validity of the two re-suspension buffers to concentrate VHSV. The mean viral genome recovery yield with oxalic and ascorbic acid was 71.2 ± 12.3% and 81.4 ± 9.5%, respectively. The mean viral infective recovery yields based on the PFU were significantly different between the two buffers at 23.8 ± 22.7% (oxalic acid) and 4.4 ± 2.7% (ascorbic acid). Notably, although oxalic acid maintains viral infectivity over 60% at a viral concentration above 105 PFU/mL, the infective VHSVs were not sufficiently recovered at a low viral concentration (102 PFU/mL, <10%). To support this result, concentrated VHSV was inoculated in Epithelioma papulosum cyprini (EPC) cells to confirm cell viability, viral gene expression, and extracellular viral titer. All results demonstrated that oxalic acid buffer was superior to ascorbic acid buffer in preserving viral infectivity.

Occurrence of aflatoxins B1, B2, G1, and G2 in beers produced in Brazil and their carcinogenic risk evaluation

The aflatoxins (B1, B2, G1, and G2) were investigated in 60 beer samples of the leading commercial beer brands, with and without adjuncts, produced in Brazil. An analytical method was standardized and validated by high-performance liquid chromatography (HPLC) with fluorescence detection. The extraction was carried out using immunoaffinity columns. The limit of quantification (LOQ) ranged from 10 to 30 ng/L−1, and the mean recoveries were 88.5%, 98.0%, 98.3%, and 110.1% for AFB1 and AFB2 AFG1, and AFG2, respectively. The percentage of samples positive for total aflatoxins (AFT) was 100% for beers with adjuncts and 80% for beers without adjuncts (“pure malt”). Of the whole beer samples, 88.3% showed the occurrence of at least one of the aflatoxins. AFB2 had the highest incidence in the beer samples, with the highest contamination level with adjuncts. The median levels of AFB2 were 27.0 and 40.4 ng/mL for the group of beers with and without adjuncts, respectively. Among the positive samples, the median value for AFT and AFB1 were 40.9 and 16 ng/mL, respectively. Carcinogenic risk characterization was performed for exposure to AFB2 and AFT by estimating the margin of exposure (MOE). The MOE for elevated beer consumption (>160 mL/day) was 5303 for a beer with adjuncts and 3643 for pure-malt beers. In summary, the results found in this study demonstrate that elevated beer consumption may pose a risk of exposure to the carcinogenic effects of aflatoxins, especially AFB2. The consumption of pure malt beers produced and commercialized in Brazil represents an even higher risk than the consumption of beer with added adjuncts. This fact alone requires further investigation since it may suggest that the barley from which the malt is produced may contribute to the occurrence of aflatoxins in beer, at least in Brazil.

Inter-laboratory study on simultaneous quantification of ten trichothecenes in feed.

An inter-laboratory study was performed in eight laboratories to evaluate the simultaneous quantification method for HT-2 toxin (HT-2), T-2 toxin (T-2), diacetoxyscirpenol (DAS), neosolaniol (NES), 3-acetyldeoxynivalenol (3-AcDON), 15-acetyldeoxynivalenol (15-AcDON), deoxynivalenol (DON), deoxynivalenol-3-glucoside (D3G), nivalenol (NIV), and fusarenon-X (FUS-X) in feed. The mycotoxins in the samples were extracted with hydrous acetonitrile, purified using a multifunctional column (InertSep® VRA-3) and a phospholipid removal column (Hybrid SPE®-Phospholipid), and then quantified using liquid chromatography-tandem mass spectrometry (LC-MS/MS) with atmospheric pressure chemical ionisation mode. The mean recovery, repeatability, reproducibility, and Horwitz ratio from the inter-laboratory validation study were 99.8-109%, 3.1-9.8%, 4.3-9.8%, and 0.19-0.45, respectively, for type A trichothecenes (HT-2, T-2, DAS, and NES). Those values for type B trichothecenes (3-AcDON, 15-AcDON, DON, NIV, and FUS-X) were 89.9-116%, 3.4-9.1%, 5.6-14%, and 0.25-0.70, and the values for modified mycotoxin (D3G) were 78.2-96.7%, 3.5-6.4%, and 13-22%, respectively.

Dynamics of Naïve White Storks Ciconia Ciconia Migrating at the Central European Migratory Divide during 1947–2018

Flexibility in migratory behaviour may have significant consequences for the adaptability of birds to ongoing environmental changes. Shifts in arrival and departure dates, migration distance and even direction of migration have all been demonstrated over the last few decades in a variety of species, mostly passerines. Less well understood is the spatio-temporal dynamics of migration at so-called migratory divides, i.e. contact zones of populations with different migratory strategies. We used data on 58 White Storks Ciconia ciconia ringed as nestlings in a part of the Central European migratory divide in the Czech Republic during the period 1947–2018 and recovered during their first winter season in October to February, to test the hypotheses of (1) shortening migratory distance over the years and (2) a shift in the proportion of adopted migratory strategies being a proximate factor of changes in migration distance. Using regression analysis, we provided evidence of gradual linear decrease over the years in mean recovery distance of ringed nestlings during their first winter. Results of multinomial regression analysis showed that this decrease has been proximately driven by an increase in the proportion of storks recovered within 500 km from their natal site, a decreasing proportion of storks recovered on the eastern migratory flyway and an increasing proportion of storks recovered on the western migratory flyway. Evidence for Czech Storks migrating via the western migratory flyway first appeared in 2003. The apparent shift of young Storks to migrate via the western flyway from the Czech Republic is most likely a result of immigration by individuals from growing western populations. This leads to diluting of populations migrating via the eastern flyway with migrants using the western flyway. Our study provides evidence that behavioural flexibility may drive dynamics of migration at the migratory divide, with changes of similar magnitudes as those driven by genetic change, as reported in passerines.

Development for the Simultaneous Analytical Method of Furan and Alkyl Furans in Processed Foods

A simultaneous analytical method was developed for the determination of alkyl furans (Furan, 2-methylfuran, 3-methylfuran and 2,5-dimethylfuran) in processed foods by headspace-GC-MS. Single-laboratory validation data of furan, 2-methylfuran, 3-methylfuran and 2,5-dimethylfuran showed good precision and accuracy. The mean recoveries ranged from 92 to 116%, the intermediate precision (RSDi) ranged from 0.9 to 12.9%. The level of LOQ ranged from 0.5 to 1.2 μg/kg (coffee), from 3.5 to 4.1 μg/kg (soy sauce), from 0.4 to 1.3 μg/kg (other foods: clear apple juice, infant formula and baby food), respectively. This method has the sensitivity to detect low levels of furan and alkyl furans contaminated in various foods and is thus applicable to surveillance for risk management in food safety.

Development of Novel Microwell-Based Spectrofluorimetry and High-Performance Liquid Chromatography with Fluorescence Detection Methods and High Throughput for Quantitation of Alectinib in Bulk Powder and Urine Samples

Background and Objectives: This study presents the development and validation of the 96-microwell-based spectrofluorimetric (MW-SFL) and high performance liquid chromatography (HPLC) with fluorescence detection (HPLC-FD) methods for the quantitation of alectinib (ALC) in its bulk powder form and in urine samples. Materials and Methods: The MW-SFL was based on the enhancement of the native fluorescence of ALC by the formation of micelles with the surfactant cremophor RH 40 (Cr RH 40) in aqueous media. The MW-SFL was executed in a 96-microwell plate and the relative fluorescence intensity (RFI) was recorded by utilizing a fluorescence plate reader at 450 nm after excitation at 280 nm. The HPLC-FD involved the chromatographic separation of ALC and ponatinib (PTB), as an internal standard (IS), on a C18 column and a mobile phase composed of methanol:potassium dihydrogen phosphate pH 7 (80:20, v/v) at a flow rate of 2 mL min-1. The eluted ALC and PTB were detected by utilizing a fluorescence detector set at 365 nm for excitation and 450 nm for emission. Results: Validation of the MW-SFL and HPLC-FD analytical methods was carried out in accordance with the recommendations issued by the International Council for Harmonization (ICH) for the process of validating analytical procedures. Both methods were efficaciously applied for ALC quantitation in its bulk form as well as in spiked urine; the mean recovery values were ≥86.90 and 95.45% for the MW-SFL and HPLC-FD methods, respectively. Conclusions: Both methodologies are valuable for routine use in quality control (QC) laboratories for determination of ALC in pure powder form and in human urine samples.

Development and validation of a simple method for the determination of Atorvastatin calcium in pure and pharmaceutical formulations using spectrofluorimetry

A simple, accurate, precise, sensitive and selective spectrofluorimetric method was developed and validated for the determination of Atorvastatin calcium (ATV), an HMG-CoA reductase inhibitor, in its pure and tablet dosage form. The proposed method was based on direct measurement of the native fluorescence of ATV. Fluorescence analysis was accomplished by using an emission wavelength 385 nm after excitation at the wavelength of 270 nm in acetonitrile, without difficult preparation steps of the sample solution such as separation, extraction, pH adjustment or derivatization. All variables affecting the fluorescence intensity such as measurement time, temperature, and diluting solvent were investigated and optimized. Under the typical conditions, a validation study for linearity, range, accuracy, precision, selectivity and robustness of the proposed method was implemented according to ICH guidelines. The fluorescence intensity was linear over concentration range of (0.4–12) μg/ml (r = 0.9999), and the lower limits of detection and quantification were 0.079 and 0.24 μg/ml, respectively. Good accuracy and precision results were obtained through using the presented method with excellent mean recovery value 100.08 ± 0.32 which was in the acceptable range (98.0–102.0%), and RSD <2%, proving the precision of the developed method. Specificity was proved in the presence of excipients and Amlodipine besylate (AML) which encountered usually as combined drug with ATV. The developed method was successfully applied to the analysis of pharmaceuticals containing the mentioned drug with no interference from other drugs or dosage form additives, and the recoveries were in the range of 99.11 ± 0.75 to 100.89 ± 0.70. Furthermore, the obtained results were compared with reported HPLC method. Then, the t- and F- values were calculated and compared with the theoretical ones, which indicate good precision and high accuracy of the proposed method. Therefore, this method is valuable, reliable, and very suitable to be applied in routine quality control laboratories.

Ultrarapid Microwave-Assisted Synthesis of Fluorescent Silver Coordination Polymer Nanoparticles and Its Application in Detecting Alkaline Phosphatase Activity

Fluorescent silver coordination polymer nanoparticles (Ag-TPA CPNs) were synthesized using a combination of terephthalic acid (TPA) and silver nitrate via an ultrarapid microwave-assisted strategy within 15 min. The Ag-TPA CPNs displayed a high fluorescent quantum yield (QY = 20.19%) and large Stokes shift (~200 nm), with two emission peaks at 490 nm and 520 nm under an excitation wavelength of 320 nm. A fluorescent "turn-off" method using fluorescent Ag-TPA CPNs was applied to detect the alkaline phosphatase (ALP) activity on the basis of the ALP-catalyzed hydrolysis of ascorbic acid 2-phosphate (AA2P) to ascorbic acid (AA), and the AA product triggered the reduction of Ag+ ions into silver nanoparticles. The fluorescent lifetime of Ag-TPA CPNs decreased from 3.93 ms to 3.80 ms after the addition of ALP, which suggests that this fluorescent "turn-off" detection of ALP activity is a dynamic quenching process. The fluorescent intensity had a linear relationship with the concentration of ALP in the range of 0.2-12 mU/mL (r = 0.991) and with a limit of detection (LOD) of 0.07 mU/mL. It showed high selectivity in ALP detection towards metal ions and amino acids, as well as other enzymes such as horseradish peroxidase, glucose oxidase, tyrosinase, trypsin, lysozyme, and superoxides. When it was applied for the fluorescent "turn-off" detection of ALP activity in serum samples, mean recovery levels ranging from 99.5% to 101.2% were obtained, with relative standard deviations (RSDs) lower than 4% accuracy. Therefore, it is an efficient and accurate tool for analyzing ALP levels in biosamples.

A simple uric acid assay by using 3‐hydroxytyramine as a chromogenic colorimetric sensor in human serum samples: Density functional theory supported mechanistic approach

A simple, fast, sensitive, convenient, and economical accurate enzymatic colorimetric sensor is developed for the quantification of the clinically important biomarker uric acid (UA) in human serum samples. The method is based on the quantification of UA using 3-hydroxytyramine (3-HT) as a chromogenic substrate by using peroxidase (POD) and uricase (UOx). UOx catalyzes the oxidation of UA where H2O2 is generated in-situ to produce allantoin and CO2. The POD in oxidative coupling of peroxide in presence of 3-HT, forms an intense orange-colored product with λmax = 500 nm. The reaction was carried out in a citric acid-tripotassium-citrate buffer (12.5 mM) of pH = 6.8 at room temperature. Computational calculations were done by Gaussian-16 employing the B3LYP functional basis set. The standard curve for UA was found to be linear between 12.3–297.3 and 3.07–396.5 μM by rate and fixed time method, respectively. The LOD and LOQ for UA were 1.5 and 2.9 μM, respectively. Within-day and day-to-day precision was 1.50–3.07 and 3.10–4.16% (n = 7), respectively. The accuracy ranges for UA concentrations of 24.7, 198 and 297.3 μM were 87%–101.50%, 90%–105%, and 89%–107%, respectively. The UA recovery range by the proposed method was 98.00%–100.47% with a mean recovery of 99.5% and has A good correlation coefficient of 0.981 with the standard method.