Miltefosine: a novel internal standard approach to lysophospholipid quantitation using LC-MS/MS.

Understanding and determining levels of lysophospholipids (LPLs) is of increasing interest to the bioanalytical community as they may be targeted for preparative removal as a matrix interference or as a lead substance as a biomarker of disease. Studies monitoring levels of LPLs have used a range of approaches for quantitation whereby those using an internal standard have used either deuterated analogues of the target LPL or alternative LPLs containing an odd number of carbon atoms within its chain, which can be expensive and difficult to distinguish with other LPLs, respectively. A structural analogue, miltefosine, was investigated as a novel internal standard to quantify a selection of lysophosphatidylcholines (LPCs) of clinical interest. A reverse phase C18 LC-MS/MS method was characterised for 16:0-LPC, 18:1-LPC and 18:0-LPC, showing good sensitivity and linearity for all compounds, with limit of detection (LOD) values <1 µg/mL and R 2 ≥ 0.97. Quality control (QC) samples were studied to determine accuracy and precision of the method, with values <15% variation for each compound at multiple concentrations. As an example application, we have used this method to detect the amount of LPC breakthrough following solid phase extraction (SPE) of plasma to quantify LPCs as a target species and to remove them as matrix interferences under various conditions typical to clinical work. This study showed that changes in sample pH could adversely affect the capture of the LPCs and their contribution as matrix interferences, with 3.6 µg/mL of 18:1-LPC observed following plasma extraction. Graphical Abstract A novel internal standard approach to lysophospholipid quantitation in extracted plasma using miltefosine, with analysis by LC-MS/MS.

Photoelectrocatalytic Surfactant Pollutant Degradation and Simultaneous Green Hydrogen Generation.

For the first time, we demonstrate a photoelectrocatalysis technique for simultaneous surfactant pollutant degradation and green hydrogen generation using mesoporous WO3/BiVO4 photoanode under simulated sunlight irradiation. The materials properties such as morphology, crystallite structure, chemical environment, optical absorbance, and bandgap energy of the WO3/BiVO4 films are examined and discussed. We have tested the anionic type (sodium 2-naphthalenesulfonate (S2NS)) and cationic type surfactants (benzyl alkyl dimethylammonium compounds (BAC-C12)) as model pollutants. A complete removal of S2NS and BAC-C12 surfactants at 60 and 90 min, respectively, by applying 1.75 V applied potential vs RHE to the circuit, under 1 sun was achieved. An interesting competitive phenomenon for photohole utilization was observed between surfactants and adsorbed water. This led to the formation of H2O2 from water alongside surfactant degradation (anode) and hydrogen evolution (cathode). No byproducts were observed after the direct photohole mediated degradation of surfactants, implying its advantage over other AOPs and biological processes. In the cathode compartment, 82.51 µmol/cm2 and 71.81 µmol/cm2 of hydrogen gas were generated during the BAC-C12 and S2NS surfactant degradation process, respectively, at 1.75 V RHE applied potential.

Use of QuEChERS as a manual and automated high-throughput protocol for investigating environmental matrices.

Environmental pollution has strong links to adverse human health outcomes with risks of pollution through production, use, ineffective wastewater (WW) remediation, and/or leachate from landfill. 'Fit-for-purpose' monitoring approaches are critical for better pollution control and mitigation of harm, with current sample preparation methods for complex environmental matrices typically time-consuming and labour intensive, unsuitable for high-throughput screening. This study has shown that a modified 'Quick Easy Cheap Effective Rugged and Safe' (QuEChERS) sample preparation is a viable alternative for selected environmental matrices required for pollution monitoring (e.g. WW effluent, treated sludge cake and homogenised biota tissue). As a manual approach, reduced extraction times (hours to ∼20 min/sample) with largely reproducible (albeit lower) recoveries of a range of pharmaceuticals and biocidal surfactants have been reported. Its application has shown clear differentiation of matrices via chemometrics, and the measurement of pollutants of interest to the UK WW industry at concentrations significantly above suggested instrument detection limits (IDL) for sludge, indicating insufficient removal and/or bioaccumulation during WW treatment. Furthermore, new pollutant candidates of emerging concern were identified - these included detergents, polymers and pharmaceuticals, with quaternary ammonium compound (QAC) biocides observed at 2.3-70.4 mg/kg, and above levels associated with priority substances for environmental quality regulation (EQSD). Finally, the QuEChERS protocol was adapted to function as a fully automated workflow, further reducing the resource to complete both the preparation and analysis to <40 min. This operated with improved recovery for soil and biota (>62%), and when applied to a largely un-investigated clay matrix, acceptable recovery (88.0-131.1%) and precision (≤10.3% RSD) for the tested pharmaceuticals and biocides was maintained. Therefore, this preliminary study has shown the successful application of a high-throughput QuEChERS protocol across a range of environmental solids for potential deployment in a regulated laboratory.

QuEChERS: a simple extraction for monitoring quaternary ammonium biocide pollution in soils and antimicrobial resistance.

Quaternary ammonium compounds (QACs) are broad-spectrum disinfectants used in a range of everyday materials. Their high usage rates, limited regulation and reporting has meant their environmental release is largely uncontrolled and impact unknown. With links to antimicrobial resistance (AMR) and adsorption to wastewater solids (that are recycled), there is a need for more controlled disposal measures and monitoring. These environmental matrices are highly complex requiring methods that are often laborious and costly to undertake. Using a robust quantitative reversed-phase LC-MS/MS method, we have shown that an 'off the shelf' QuEChERS product can reliably extract (<10% RSD) aromatic and aliphatic QACs anticipated within municipal, industrial and agricultural waste from water and soil, with reduced matrix effects of 95.7-104.4% for recoveries of up to 53% from soil when combined with extract dilution. Therefore, unlike current literature, this work has shown that, with minimal development, the QuEChERS product can provide a rapid, effective and low cost preparation for quantifying QAC pollution and monitoring AMR.

An investigation of the utility of QuEChERS for extracting acid, base, neutral and amphiphilic species from example environmental and clinical matrices.

Accurate measurement of the composition of complex samples is key for the safety and efficacy of a range of products used in daily life, with sample preparation a critical step in this workflow. QuEChERS is one such method, however published protocols do not explicitly address acidic, basic, neutral, and amphiphilic species in a single protocol and often use extra steps or an alternative preparation to recover the breadth of chemical types. Our work addresses this need by investigating the use of QuEChERS for monitoring this wide range of chemistries within environmental solids and blood plasma, using a protocol that can accommodate both milliliter and microliter sample volumes. While published methods can require significant resource and time, our approach offers a reduction in preparation time (for environmental samples), with the "micro-QuEChERS" protocol offering a further reduction in cost. The analytical performance of these methods were assessed using reversed-phase LC-MS and showed good accuracy, precision, and sensitivity for the expected concentrations in the tested applications. Target analytes of variable lipophilicity/acidity were extracted and isolated from soil, with largely repeatable matrix effects < 15%RSD and recoveries of 39-100%. An initial "proof-of-concept" investigation using the "micro-QuEChERS" protocol showed reduced matrix enhancement (median value of 90%ME) for soil, and improved matrix effects and recovery (>65%) for blood plasma. This novel sample preparation method can therefore offer an improved approach with wider applicability providing "cleaner" extracts than other methods used for high-throughput clinical analysis.

Novel adenovirus-based vaccines induce broad and sustained T cell responses to HCV in man.

Currently, no vaccine exists for hepatitis C virus (HCV), a major pathogen thought to infect 170 million people globally. Many studies suggest that host T cell responses are critical for spontaneous resolution of disease, and preclinical studies have indicated a requirement for T cells in protection against challenge. We aimed to elicit HCV-specific T cells with the potential for protection using a recombinant adenoviral vector strategy in a phase 1 study of healthy human volunteers. Two adenoviral vectors expressing NS proteins from HCV genotype 1B were constructed based on rare serotypes [human adenovirus 6 (Ad6) and chimpanzee adenovirus 3 (ChAd3)]. Both vectors primed T cell responses against HCV proteins; these T cell responses targeted multiple proteins and were capable of recognizing heterologous strains (genotypes 1A and 3A). HCV-specific T cells consisted of both CD4+ and CD8+ T cell subsets; secreted interleukin-2, interferon-γ, and tumor necrosis factor-α; and could be sustained for at least a year after boosting with the heterologous adenoviral vector. Studies using major histocompatibility complex peptide tetramers revealed long-lived central and effector memory pools that retained polyfunctionality and proliferative capacity. These data indicate that an adenoviral vector strategy can induce sustained T cell responses of a magnitude and quality associated with protective immunity and open the way for studies of prophylactic and therapeutic vaccines for HCV.

Standardised analgesia packs after day case orthopaedic surgery.

Two of the advantages of day surgery are less disruption to patients' lives and the comfort of recovering at home. However, despite advances in analgesic and anaesthetic techniques, pain following day surgery is not well managed: recent studies have shown that between 30-60% of patients suffer moderate to severe pain during the first 24 hours after discharge home following day surgery (Beauregard et al 1998, McGrath et al 2004, Pavlin et al 2004). A significant proportion of patients (25-30%) continue to report pain of this severity at seven days following day surgery (Beauregard et al 1998, Watt-Watson 2004). This article reviews published studies of patient experiences of pain and analgesia consumption after day case surgery and provides a model for the introduction of standardised take-home analgesic packs.

After critical care: a study to explore patients' experiences of a follow-up service.

AIMS AND OBJECTIVES: To establish patients' experiences after discharge from critical care and to evaluate implementation of a follow-up service. BACKGROUND: Government recommendations advise critical care follow-up to prevent readmission and address problems after discharge. Admission to critical care results in significant psychological and physiological sequelae. DESIGN: A prospective, longitudinal and exploratory study of surgical cancer patients requiring >48 hours in critical care. Qualitative interviews were conducted and short questionnaires were used. METHODS: Patient Expert Advisory Groups were invited to participate in research design. Patients were visited in the ward at days 1 and 5 after discharge, invited to nurse-led follow-up clinic and interviewed at three and six months. Short questionnaires were administered at six and 12 months. FINDINGS: Twenty-seven patients participated in the study. All patients experienced benefit from the service. Emergent themes included: rehabilitation from critical care: physiological issues and needs, memories: real and unreal, uncertainty and fear and empathy. A core theme of reassurance was underlying through the research. Issues while in critical care included: the need for nursing presence, nightmares, delusions, confusion, fear of ward transfer, inability to remember, disorientation and being prepared for the experience. After discharge, issues shifted to longer term needs. Psychological support, in the form of the follow-up clinic, proved useful. The ability to move on with life after discharge varied and uncertainty about the future and their cancer had an impact upon this. Recovery was made easier through the follow-up clinic. Patients required reassurances that their experiences were valid and also wanted reassurances about their cancer. CONCLUSION: Critical care causes various difficulties for patients that may impinge on recovery. Incorporating patients into the design process helps identify needs more closely. Follow-up proved beneficial and highlighted the role nurses have in improving patient experiences after discharge from critical care. Nurses should be vigilant for both immediate and longer-term needs. RELEVANCE TO CLINICAL PRACTICE: This research into nurse-led follow-up clinics after critical care highlights an important, and often neglected, part of the critical illness continuum. Attending such clinics may help reassure patients after discharge from critical care.