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Background Antivirals are needed to combat the COVID-19 pandemic, which is caused by SARS-CoV-2. The clinically-proven protease inhibitor Camostat mesylate inhibits SARS-CoV-2 infection by blocking the virus-activating host cell protease TMPRSS2. However, antiviral activity of Camostat mesylate metabolites and potential viral resistance have not been analyzed. Moreover, antiviral activity of Camostat mesylate in human lung tissue remains to be demonstrated. Methods We used recombinant TMPRSS2, reporter particles bearing the spike protein of SARS-CoV-2 or authentic SARS-CoV-2 to assess inhibition of TMPRSS2 and viral entry, respectively, by Camostat mesylate and its metabolite GBPA. Findings We show that several TMPRSS2-related proteases activate SARS-CoV-2 and that two, TMPRSS11D and TMPRSS13, are robustly expressed in the upper respiratory tract. However, entry mediated by these proteases was blocked by Camostat mesylate. The Camostat metabolite GBPA inhibited recombinant TMPRSS2 with reduced efficiency as compared to Camostat mesylate. In contrast, both inhibitors exhibited similar antiviral activity and this correlated with the rapid conversion of Camostat mesylate into GBPA in the presence of serum. Finally, Camostat mesylate and GBPA blocked SARS-CoV-2 spread in human lung tissue ex vivo and the related protease inhibitor Nafamostat mesylate exerted augmented antiviral activity. Interpretation Our results suggest that SARS-CoV-2 can use TMPRSS2 and closely related proteases for spread in the upper respiratory tract and that spread in the human lung can be blocked by Camostat mesylate and its metabolite GBPA. Funding NIH, Damon Runyon Foundation, ACS, NYCT, DFG, EU, Berlin Mathematics center MATH+, BMBF, Lower Saxony, Lundbeck Foundation, Novo Nordisk Foundation. more...

Objectives Camostat mesilate is a drug that is being repurposed for new applications such as that against COVID-19 and prostate cancer . This induces a need for the development of an analytical method for the quantification of camostat and its metabolites in plasma samples. Camostat is, however, very unstable in whole blood and plasma due to its two ester bonds. The molecule is readily hydrolysed by esterases to 4-(4-guanidinobenzoyloxy)phenylacetic acid (GBPA) and further to 4-guanidinobenzoic acid (GBA). For reliable quantification of camostat, a technique is required that can instantly inhibit esterases when blood samples are collected. Design and methods An ultra-high-performance liquid chromatography-tandem mass spectrometry method (UHPLC-ESI-MS/MS) using stable isotopically labelled analogues as internal standards was developed and validated. Different esterase inhibitors were tested for their ability to stop the hydrolysis of camostat ester bonds. Results Both diisopropylfluorophosphate (DFP) and paraoxon were discovered as efficient inhibitors of camostat metabolism at 10 mM concentrations. No significant changes in camostat and GBPA concentrations were observed in fluoride-citrate-DFP/paraoxon-preserved plasma after 24 h of storage at room temperature or 4 months of storage at −20 °C and −80 °C. The lower limits of quantification were 0.1 ng/mL for camostat and GBPA and 0.2 ng/mL for GBA. The mean true extraction recoveries were greater than 90%. The relative intra-laboratory reproducibility standard deviations were at a maximum of 8% at concentrations of 1–800 ng/mL. The trueness expressed as the relative bias of the test results was within ±3% at concentrations of 1–800 ng/mL. Conclusions A methodology was developed that preserves camostat and GBPA in plasma samples and provides accurate and sensitive quantification of camostat, GBPA and GBA by UHPLC-MS/MS. more...

For prospective investigation of drugs and metabolites in archaeological and contemporary dental calculus, a sensitive, broadly applicable ultra-high-performance liquid chromatography-tandem mass spectrometry method using pneumatically assisted electrospray ionisation (UHPLC-ESI-MS/MS) was developed. The dental calculus was treated with citric acid and the dissolution extracts were cleaned using weak and strong polymeric cation-exchange sorbents. The method was validated on hydroxyapatite for the analysis of 67 drugs and metabolites. Typically, the lower limits of quantification were in the range of 0.01–0.05 ng for the sample mass extracted. The general applicability of the method was tested using dental calculus material sampled from 10 corpses undergoing forensic autopsy. The calculus material was washed several times before dissolution to remove residual substances originating from saliva, gingival crevicular fluid and blood. The wash extracts and the calculus samples (cleaned calculus material) were analysed using the same instrumental conditions. The dry mass of the calculus samples ranged from 1 to 10 mg. The total number of drug detections was 131 in the dental calculus samples and 117 in the whole blood samples. From the analyses of the wash extracts and calculus samples, it was proven that drug residues were trapped in the interior of the calculus material. In 82 of the drug detections, the drug concentrations were higher in the dental calculus than in the blood. Among substances detected in the dental calculus but not in the blood were cocaine, heroin, 6-MAM and THCA-A. more...

Cytosolic DNA stimulates innate immune responses, including type I interferons (IFN), which have antiviral and immunomodulatory activities. Cyclic GMP-AMP synthase (cGAS) recognizes cytoplasmic DNA and signals via STING to induce IFN production. Despite the importance of DNA in innate immunity, the nature of the DNA that stimulates IFN production is not well described. Using low DNA concentrations, we show that dsDNA induces IFN in a length-dependent manner. This is observed over a wide length-span of DNA, ranging from the minimal stimulatory length to several kilobases, and is fully dependent on cGAS irrespective of DNA length. Importantly, in vitro studies reveal that long DNA activates recombinant human cGAS more efficiently than short DNA, showing that length-dependent DNA recognition is an intrinsic property of cGAS independent of accessory proteins. Collectively, this work identifies long DNA as the molecular entity stimulating the cGAS pathway upon cytosolic DNA challenge such as viral infections. more...

The stability of cannabinoids is complex and crucial for the assessment of impaired driving caused by cannabis. Therefore, the effect of antioxidants on the long-term stability of Δ 9-tetrahydrocannabinol (THC), cannabinol (CBN), cannabidiol (CBD), 11-hydroxy-Δ 9-tetrahydrocannabinol (THC-OH), and 11-nor-9-carboxy-Δ 9-tetrahydrocannabinol (THC-COOH) in whole blood samples preserved with fluoride citrate (FC) and fluoride oxalate (FX) mixtures was investigated at different temperatures. The measured concentrations of the cannabinoids in authentic whole blood preserved solely with FC or FX mixtures decreased significantly during prolonged storage at −20°C. On average, less than 5% of the initial concentrations of THC and CBD were recovered after 19 weeks of storage interrupted by 5 thawing/freezing cycles. The rate of decrease was greatest in FC-preserved blood. The repeated thawing/freezing of the samples accelerated the instability progression. At 5°C approximately 60% of the initial concentrations of THC and CBD were recovered after 19 weeks of storage. No significant decrease was observed in samples stored at −80°C during the test period of 5 months. The instability at −20°C was to a great extend avoided by adding 30 mM ascorbic acid (ASC) to the samples before storage. Samples preserved with a combination of the FX mixture and ASC showed no significant decrease in the recovered concentrations during a 5-month storage period interrupted by 6 thawing/freezing cycles. Samples preserved with a combination of the FC mixture and ASC showed almost similar improvements in cannabinoid stability. Other reducing agents such as sodium metabisulfite and glutathione also improved the stability in FX-preserved blood stored at −20°C. more...

Methylglyoxal (MG) is a toxic glycolytic by-product associated with increased levels of inflammation and oxidative stress and has been linked to ageing-related diseases, such as diabetes and Alzheimer's disease. As MG is a highly reactive dicarbonyl compound, forming both reversible and irreversible adducts with a range of endogenous nucleophiles, measuring endogenous levels of MG are quite troublesome. Furthermore, as MG is a small metabolite it is not very immunogenic, excluding conventional ELISA for detection purposes, thus only more instrumentally demanding LC-MS/MS-based methods have demonstrated convincing quantitative data. In the present work we develop a novel bifunctional MG capture probe as well as a high specificity monoclonal antibody to finally setup a robust reaction-based ELISA (ReactELISA) method for detecting the highly reactive and low-level (nM) metabolite MG in human biological specimens. The assay is tested and validated against the current golden standard LC-MS/MS method in human blood plasma and cell-culture media. Furthermore, we demonstrate the assays ability to measure small perturbations of MG levels in growth media caused by a small molecule drug buthionine sulfoximine (BSO) of current clinical relevance. Finally, the assay is converted into a homogenous (no-wash) AlphaLISA version (ReactAlphaLISA), which offers the potential for operationally simple screening of further small molecules capable of perturbing cellular MG. Such compounds could be of relevance as probes to gain insight into MG metabolism as well as drug-leads to alleviate ageing-related diseases. Keywords: Methylglyoxal, ELISA, Glyoxalase, Plasma, Cell culture, Buthionine sulfoximine more...

Buprenorphine (BUP) was not long-term stable in whole blood samples preserved with fluoride citrate (FC) and fluoride oxalate (FX) mixtures when stored at −20°C. On average, only half of the initial concentrations of BUP was recovered after 12 weeks of storage at −20°C when interrupted by 3–4 thaw/freeze cycles. Norbuprenorphine (NBUP) was less unstable; approximately 90% was recovered after 12 weeks of storage at −20°C. The instability was less at 5°C, but the formation of BUP and NBUP from their glucuronides was observed at that temperature, especially in FC-preserved blood. The substances were stable for at least 5 months when stored uninterrupted at −80°C. The instability of BUP and NBUP in FC- and FX-preserved whole blood stored at −20°C was eliminated when the samples were modified with 30 mM ascorbic acid (ASC) prior to storage. The mean recoveries were greater than 95% after a 5-month interrupted storage period at −20°C when modified with ASC. more...

Propofol (2,6-diisopropylphenol) is commonly used as an anaesthetic agent but is also abused for recreational purposes. Several cases of fatalities involving self-administered propofol have been reported. For rapid quantification of propofol and propofol β-d-glucuronide (propofol G) in clinical and forensic cases, an ultra-performance liquid chromatography-tandem mass spectrometry method using pneumatically assisted electrospray ionisation has been developed. The technique has been validated on both ante-mortem and post-mortem human whole blood. The proteins in the blood samples were removed by the addition of a mixture of methanol and acetonitrile, and the extract was cleaned up by solid phase extraction. The extract was concentrated in dimethyl sulphoxide. The system was calibrated using matrix-matched calibrants combined with isotope dilution. The lower limits of quantification were 0.01 and 0.02mg/L for propofol and 0.02 and 0.04mg/L for propofol G in ante-mortem and post-mortem whole blood, respectively. The relative intra-laboratory reproducibility standard deviation was less than 10% at concentrations of 0.2mg/L or higher. The mean true extraction recovery was 85% for propofol and 81% for propofol G. The trueness of the propofol determination expressed as the relative bias of the test results was within ±6% at concentration levels of 0.01-8.5mg/L. Propofol was less stable in blood stabilised with a citrate-EDTA-fluoride mixture than in blood stabilised with an oxalate-fluoride mixture. The stability was lower at -20°C than at 5°C and -80°C. Propofol G did not show instability under the storage conditions tested. more...

Innate immune activation by macrophages is an essential part of host defence against infection. Cytosolic recognition of microbial DNA in macrophages leads to induction of interferons and cytokines through activation of cyclic GMP-AMP synthase (cGAS) and stimulator of interferon genes (STING). Other host factors, including interferon-gamma inducible factor 16 (IFI16), have been proposed to contribute to immune activation by DNA. However, their relation to the cGAS-STING pathway is not clear. Here, we show that IFI16 functions in the cGAS-STING pathway on two distinct levels. Depletion of IFI16 in macrophages impairs cGAMP production on DNA stimulation, whereas overexpression of IFI16 amplifies the function of cGAS. Furthermore, IFI16 is vital for the downstream signalling stimulated by cGAMP, facilitating recruitment and activation of TANK-binding kinase 1 in STING complex. Collectively, our results suggest that IFI16 is essential for efficient sensing and signalling upon DNA challenge in macrophages to promote interferons and antiviral responses., The role of IFI16 as a DNA sensor is highly controversial. With support from a Nature Communications back-to-back publication from Almine et al. the authors here provide functional evidence that IFI16 is required for DNA sensing via the cGAS-STING pathway in human macrophages. more...

Direct analysis of Δ9-tetrahydrocannabinol (THC) and other cannabinoids in crude acetonitrile extracts of whole blood by liquid chromatography-tandem mass spectrometry using pneumatically assisted electrospray ionization (LC-ESI-MS-MS) was subjected to pronounced ion suppression from co-eluting phospholipids (PLs). The interferences were mainly caused by the lysophosphatidylcholine and lysophosphatidylethanolamine classes of PLs. The PLs were easily removed from crude extracts by filtration through a sorbent with Lewis acid properties, which typically increased the THC and cannabinol (CBN) signal intensities by a factor of 5. Based on this technique, a simple high-throughput LC-MS-MS method was developed for the determination of cannabinoids in 100 μL samples of whole blood. The lower limits of quantification were 0.2 μg/L for THC, CBN, cannabidiol (CBD) and Δ9-tetrahydrocannabinolic acid A (THCA-A) and 0.5 μg/L for 11-hydroxy-Δ9-tetrahydrocannabinol (THC-OH) and 11-nor-9-carboxy-Δ9-tetrahydrocannabinol (THC-COOH). The mean ion suppression levels after clean-up were 10% (THC), 9% (CBN), 17% (CBD), 0% (THC-OH), 2% (THC-COOH) and 9% (THCA-A) at blood concentration levels of 1-10 μg/L. The mean true extraction recoveries were 97% (THC), 101% (CBN), 101% (CBD), 98% (THC-OH), 95% (THC-COOH) and 90% (THCA-A) at the same concentration levels. The relative intra-laboratory reproducibility standard deviations were more...

There is a realization that the quality of teaching and learning has declined due to ineffective and inefficient monitoring of schools and this has provided an impetus for appraising the objectives which quality assurance agents of the inspectorate services is expected to achieve. A survey research-design was adopted for the study, two research questions and two null hypotheses, tested at (p < .05) level of significance, were used. Through a multistage procedure, three out of the five states were selected from South-East Zone. 371 head-teachers from public primary schools and 217 quality assurance agents of the inspec- torate services (evaluators) were sampled. A 27 item questionnaire titled Primary Schools Appraisal Questionnaire (PSAQ), covering different areas of school evaluation, was developed by the researchers to get information from both head-teachers of public primary schools and evaluators. Cronbach Alpha method was used to determine the internal consistency of the PSAQ. The computation yielded reliability coefficient of 0.97, for the general cluster of the instrument. The findings of the study showed that although evaluators em- bark on routine monitoring and make recommendations to government based on the results of their visits, they do not close schools that are not performing up to expectation. The study also showed that evaluators lack facilities and equipment for school monitoring. more...

A liquid-chromatography-tandem-mass-spectrometry method using pneumatically assisted electrospray ionisation (LC-ESI-MS/MS) was developed for the simultaneous determination of cathinone, methcathinone, ethcathinone, amfepramone, mephedrone, flephedrone, methedrone, methylone, butylone, cathine, norephedrine, ephedrine, pseudoephedrine, methylephedrine and methylpseudoephedrine in human live and post-mortem whole blood. The blood proteins were precipitated by the addition of methanol, and the extract was purified by ultrafiltration. The separation of diastereomeric ephedrines was achieved on an ethyl-linked phenyl column. Matrix-matched calibrants combined with the isotope dilution of selected substances were used for quantitative analysis. The relative intra-laboratory reproducibility standard deviations were generally better than 7% at concentrations of 20 μg/L, and the mean true recoveries were 87-106% in the concentration range of 10-250 μg/L. The detection limits were in the range of 0.5-3 μg/L. The cathinones were unstable in whole blood and sample extracts under neutral conditions, but the stability could be improved by the acidification of the sample matrix. more...

Metformin is an anti-diabetic drug in the biguanide class which also includes phenformin and buformin. Because of the potential adverse effects of the biguanides, a reliable liquid chromatography-tandem mass spectrometry method using pneumatically assisted electrospray ionization was developed for the quantification of the drugs in both live and post-mortem human whole blood. The blood proteins were precipitated by the addition of a mixture of methanol and acetonitrile, and the extract was cleaned up by cation-exchange solid-phase extraction to eliminate ion suppression effects. The separation was performed by hydrophilic interaction liquid chromatography. Matrix-matched calibrants combined with isotope dilution of metformin were used for calibration. The detection limits were 0.01 mg/L for metformin and phenformin and the relative intra-laboratory reproducibility standard deviations were less than 6% at concentrations of 1-10 mg/L. The mean true recoveries were greater than 86%. more...

A liquid chromatography–tandem mass spectrometry method using pneumatically assisted electrospray ionisation (LC–ESI-MS/MS) was developed for the simultaneous determination of paracetamol, naproxen, ibuprofen, etodolac, diclofenac, salicylic acid, lamotrigine, carbamazepine, 10-OH-carbazepine, phenytoin, phenobarbital, barbital, cyclobarbital, pentobarbital, amobarbital and caffeine in human live and post-mortem whole blood. The blood proteins were precipitated in a mixture of methanol and acetonitrile, and the extract was purified by ultrafiltration. The separation was performed on an ether-linked phenyl column with polar endcapping, and both negative and positive ESI were applied. Matrix-matched standards were used for calibration. The relative intra-laboratory reproducibility standard deviations were, in general, better than 8% at concentrations in the therapeutic range. The mean true recoveries were in the range 92–101% for the live blood and 84–101% for the post-mortem blood. more...

A rapid confirmatory liquid chromatographic/tandem mass spectrometic method was developed for determination of chloramphenicol in bovine milk. Chloramphenicol was extracted directly from milk by solid-phase extraction on a C18 cartridge. The extract was further cleaned up on neutral aluminium oxide. Three transition products were monitored in negative ion mode after atmospheric pressure chemical ionization. The detection capability related to the transition product of lowest abundance was 0.03 μg/kg. The mean recovery was 90% at levels of 0.1–0.2 μg/kg. The relative repeatability standard deviations were 4.3, 3.8, and 2.8% at levels of 0.1, 0.2, and 1.0 μg/kg, respectively. more...

The recruitment and retention strategy of the Department of Social Development (2006:23) postulates that there is a “lack of structured supervision and poor quality supervisors, who themselves also lack capacity to conduct professional supervision” in South Africa. Pieterse (1961), one of the first authors on supervision in South Africa, made similar claims close to five decades ago. These two comments give rise to the question: has social work in South Africa been practising supervision over the past five decades beyond historical bureaucratic discourses, and has it engaged with changing global, local and personal contexts to develop and sustain critically responsive practices? In other words: is social work supervision in South Africa keeping up with the times? By addressing this question, this paper seeks to respond to the call of the recruitment and retention strategy document (Department of Social Development, 2006) to evaluate current supervision practices. In this attempt the paper reports on research, drawing on an historical analysis of the international and local development of social work supervision and on a case study of current supervision practices in an NGO environment, in order to recommend a context-specific conceptual framework for future supervision practices. This paper thus aims to examine the interplay between the historical development, current practices and future challenges of social work supervision more...

Students enrolled in the laboratory animal science graduate program at MCP Hahnemann University seek to gain entrance to veterinary school or to manage an animal facility within an academic institution, pharmaceutical or biotechnology company conducting biomedical research. Ongoing interaction between faculty in the radiation oncology, radiation safety, and lab animal science disciplines revealed an acute need for radiation safety education for laboratory animal science students who will likely interact with researchers either designing and writing protocols for animal studies using radiation or radioactive materials, or veterinary staff who will use sources of radiation to diagnose and/or treat possible animal injuries and diseases. A core course in the Radiation Sciences graduate program was modified to address the needs of these students, instructing them in radiation safety, detection and counting instrumentation, and radiation biology. These fundamental areas were integrated to help the students gain a sound, basic knowledge of radiation and radioactive materials used in biomedical research. more...

A high-performance liquid chromatographic (HPLC) method based on solid phase extraction was developed for determination of metronidazole (MNZ) and its metabolite hydroxymetronidazole (MNZ-OH) in muscle and skin tissues of rainbow trout. Tinidazole (TNZ) was used as internal standard. The compounds were extracted with acetonitrile and the extract was evaporated and redissolved in a mixture of ethyl acetate and hexane (1:2). The extract was cleaned up by solid phase extraction on a silica cartridge. The extract was analysed by reverse phase gradient HPLC on a C18 column followed by ultraviolet detection at 325 nm. The limit of detection was 2.8 micrograms/kg for both compounds in muscle. The estimated limits of detection in skin tissue were 3 micrograms/kg for MNZ and 5 micrograms/kg for MNZ-OH. The mean recoveries of MNZ in muscle calculated without use of internal standard were 93% and 81% at levels of 10 micrograms/kg and 25-100 micrograms/kg respectively. The mean recovery of MNZ-OH in muscle was 79% at a level of 10-100 micrograms/kg. The mean relative repeatability standard deviations on spiked muscle tissue were 3.3% for MNZ and 3.2% for MNZ-OH at a level of 10-100 micrograms/kg. The method was applied to trout given feed containing MNZ in an aquaculture pilot plant. Residues of MNZ and MNZ-OH were detected in muscle and skin tissues shortly after the administration period but not 3 weeks later. more...

Vigabatrin, pregabalin, gabapentin and baclofen are γ-aminobutyric acid analogs that are used in the treatment of epileptic seizures (vigabatrin, pregabalin and gabapentin) and spasticity (baclofen). The intake of these drugs may induce adverse reactions and impair the ability of an individual to drive a vehicle. There have also been reports of cases of intoxication and fatalities from overdoses. For rapid and accurate quantification of these drugs in forensic cases, an ultraperformance liquid chromatography tandem mass spectrometry method using pneumatically assisted electrospray ionization has been developed. The technique has been validated on both ante- and postmortem human whole blood. The protein in the blood samples was removed by the addition of a mixture of methanol and acetonitrile, and the extract was ultrafiltered and diluted with acetonitrile. The separation was performed by hydrophilic interaction liquid chromatography. Calibration of the system was achieved through use of matrix-matched calibrants combined with isotope dilution. The lower limits of quantification were 0.02-0.04 mg/L, and the relative intra-laboratory reproducibility standard deviations were4 and 8% at concentrations of 10 and 1 mg/L, respectively. The mean true recoveries were89%. The trueness expressed as the relative bias of the test results was within ±7% at concentrations of 1-40 mg/L for vigabatrin, pregabalin and gabapentin and of 0.1-4 mg/L for baclofen. more...

A high-performance liquid chromatographic method based on C18 solid-phase extraction and ultraviolet detection at 323 nm of analytes derivatized with benzoic anhydride and 1,2,4-triazole mercuric chloride solution was developed for the simultaneous determination of amoxicillin, penicillin G (benzylpenicillin), ampicillin, oxacillin, cloxacillin and dicloxacillin residues in raw milk. The detection limit of the method was 1.3 microg/l for penicillin G; 1.4 microg/l for amoxicillin, oxacillin and cloxacillin, 1.5 microg/l for ampicillin and 2.7 microg/l for dicloxacillin. The mean recovery was 95-102% for amoxicillin, penicillin G and ampicillin, 92-98% for oxacillin and cloxacillin and 87-94% for dicloxacillin, measured by using an internal standard. The relative repeatability standard deviation was 4-9% on level 4-15 micro/l, respectively, 2-7% on level 30-40 microg/l. more...

Background: limited data exist on transitioning patients with psoriasis from conventional systemic agents to biologics. Objectives: the TRANSIT study aimed to assess the efficacy and safety of two methotrexate-to-ustekinumab transition strategies. Methods: patients with moderate-to-severe psoriasis and inadequate methotrexate response were randomized 1 : 1 to receive ustekinumab with immediate (arm 1) or 4-week gradual (arm 2) methotrexate withdrawal. Patients weighing ≤ 100 kg or > 100 kg received ustekinumab 45 mg or 90 mg, respectively. The primary endpoint was the frequency of adverse events (AEs) at week 12. Secondary endpoints included additional safety, efficacy and patient-reported outcomes. We report the 12-week efficacy and safety results. Results: overall, 244 patients in arm 1 and 245 in arm 2 were randomized and received ustekinumab. Four patients per arm discontinued the trial by week 12. At week 12 in arms 1 and 2, respectively, 61% and 65% of patients experienced an AE, 2·9% and 2·4% had a serious AE, and 1·2% and 0·4% had an AE leading to ustekinumab discontinuation. In arms 1 and 2, respectively, median Psoriasis Area and Severity Index (PASI) score decreased from 15·2 and 15·4 at baseline to 2·9 and 2·8 at week 12; 58% and 62% of patients achieved a 75% reduction from baseline in PASI score (PASI 75) at week 12; median baseline Dermatology Life Quality Index fell from 8 and 9 at baseline to 1 (both arms) at week 16. Conclusions: ustekinumab was well tolerated and effective in patients who had an inadequate response to methotrexate. Both transition strategies resulted in similar week 12 safety and efficacy outcomes. more...

Background: There are limited long-term, ‘real-world’ data on ustekinumab, or the effect of dose adjustment in suboptimal responders. Objectives: We describe 52-week data from TRANSIT, which initiated ustekinumab by licensed regimen and investigated exploratory dose adjustment. Methods: Patients with moderate-to-severe psoriasis and inadequate methotrexate response received ustekinumab, with immediate or gradual methotrexate withdrawal. Outcomes were similar between treatment arms at week 12 (primary endpoint), so week 52 data were pooled. Patients weighing ≤ 100 kg or > 100 kg were administered ustekinumab 45 or 90 mg, respectively. Patients weighing ≤ 100 kg without 75% improvement in Psoriasis Area and Severity Index (PASI 75) response at weeks 28 or 40 received a dose adjustment to 90 mg. The primary analysis used observed data. Results: Overall, 391 and 98 patients received ustekinumab 45 and 90 mg, respectively. Forty-four patients (9%) discontinued before week 52 (0·4% due to adverse events). At week 52 (in the overall population), 369 patients (83%) achieved a PASI score ≤ 5, and 341 patients (77%) achieved PASI 75; the median PASI score decreased from 15 at baseline to 1·8. At weeks 28 and 40, 84 and 31 patients, respectively, did not achieve PASI 75 and received a dose adjustment; by week 52, 35/82 (43%) and 15/31 (48%) of these patients, respectively, achieved PASI 75 (two discontinued between weeks 28 and 40). Conclusions: Ustekinumab showed sustained 1-year efficacy and was well tolerated when initially administered according to label. Adjusting the ustekinumab dose to 90 mg may result in clinically meaningful improvement in response in patients weighing ≤ 100 kg with suboptimal initial response. more...

A high-performance liquid chromatographic (HPLC) method based on solid-phase extraction was developed for the determination of cefazolin, cefoperazone, cefquinome and ceftiofur in raw bovine milk. The milk fat was removed by centrifugation and the cephalosporins were extracted in acetonitrile. The extract was cleaned up by solid-phase extraction on an octadecyl sorbent. The compounds were separated by ion-paired gradient HPLC on a phenyl column with ultraviolet detection at 270 nm. The limits of detection estimated by a conservative model were 11 microg/kg for cefazolin and cefoperazone and 7 microg/kg for cequinome and ceftiofur. The mean recoveries were 86-88% for cefazolin, 91-93% for cefoperazone, 69-72% for cefquinome and 84-88% for ceftiofur in the concentration range 20-200 microg/kg. more...

A recent American study identified clinical factors which effectively predicted those patients who would have significant findings on cranial computed tomography. It was proposed to apply these criteria in a UK setting and to determine whether modifications could be made to improve their efficiency.A prospective observational study was conducted over a four month period including all non-trauma adult patients referred from the accident and emergency (AE) department for urgent cranial computed tomography. Presenting symptoms and signs were analysed for ability to predict clinically significant computed tomography findings, namely: acute infarct, malignancy, acute hydrocephalus, intracranial haemorrhage, or intracranial infection.Sixty two patients were included; 22 (35%) had significant findings on computed tomography. Applying the original criteria (any of: age 60 years or older, focal neurology, headache with nausea or vomiting, altered mental status) to the study population showed that no clinically significant tomograms would have been omitted but only 11% fewer performed. Modifying the criteria by removing "age 60 years or older" and replacing "altered mental status" with a Glasgow coma score14, still ensured 100% sensitivity and would have resulted in 19% fewer scans being performed.Simple clinical criteria can be usefully applied to patients presenting to an AE department in this country to target patients most likely to have clinically significant findings on urgent cranial computed tomography. more...

A high-performance liquid chromatographic (HPLC) method based on solid-phase extraction (SPE) was developed for determination of amoxicillin, penicillin G (benzylpenicillin), ampicillin, oxacillin, cloxacillin, nafcillin and dicloxacillin in muscle, liver and kidney tissues of pigs and cattle. The compounds were extracted in aqueous solution by precipitation of organic materials with a mixture of sulphuric acid and sodium tungstate. The extract was cleaned up by SPE on a divinylbenzene-co- N -vinylpyrrolidone polymeric sorbent. Further clean-up was performed by liquid–liquid partition with diethyl ether. The extract was derivatised with benzoic anhydride and 1,2,4-triazole mercury (II) reagent. Chromatography was performed by reversed-phase gradient HPLC on a C 18 column with ultraviolet detection at 323 nm. The limits of detection estimated by a conservative model were in the range 8.9–11.1 μg/kg for amoxicillin, penicillin G, ampicillin, oxacillin, cloxacillin and nafcillin and 18.3–20.9 μg/kg for dicloxacillin. The mean recovery range was 66–77% for amoxicillin, 73–75% for penicillin G, 81–82% for ampicillin, 73–76% for oxacillin, 74–75% for cloxacillin, 66–72% for nafcillin and 58–65% for dicloxacillin. more...

A high-performance liquid chromatographic (HPLC) method based on solid phase extraction was developed for the simultaneous determination of fenbendazole (FBZ), fenbendazole sulfoxide (FBZ-SO) and fenbendazole sulfone (FBZ-SO2) in trout muscle and skin tissues. The compounds were extracted with acetonitrile and the extract was concentrated and cleaned up by solid phase extraction on C18 and CN cartridges. The extract was analysed by reversed-phase gradient HPLC on a C18 column followed by ultraviolet detection at 290 nm. The method's detection limits were 4.0 micrograms kg-1 for FBZ, 4.5 micrograms kg-1 for FBZ-SO and 3.8 micrograms kg-1 for FBZ-SO2. The mean recovery in muscle was 88% for FBZ, 94% for FBZ-SO and 92% for FBZ-SO2 at a level of 5-150 micrograms kg-1. The corresponding mean recoveries in skin tissue were 88, 81 and 86% at a level of 10-100 micrograms kg-1. The mean relative repeatability standard deviation was 9.2% at a level of 5 micrograms kg-1, 5.9% at a level of 10-100 micrograms kg-1 and 2.3% at a level of 150 micrograms kg-1. The method was applied to trout given feed containing FBZ in an aquaculture pilot plant. The three compounds FBZ, FBZ-SO and FBZ-SO2 were all detected in muscle and skin tissues shortly after administration. The concentrations were generally highest in skin tissue. more...