High-performance liquid chromatography and pharmacokinetics of some antibiotics

The principles of High Performance Liquid Chromatography (HPLC) and pharmacokinetics were applied to the use of several clinically-important drugs at the East Birmingham Hospital. Amongst these was gentamicin, which was investigated over a two-year period by a multi-disciplinary team. It was found that there was considerable intra- and inter-patient variation that had not previously been reported and the causes and consequences of such variation were considered. A detailed evaluation of available pharmacokinetic techniques was undertaken and 1- and 2-compartment models were optimised with regard to sampling procedures, analytical error and model-error. The implications for control of therapy are discussed and an improved sampling regime is proposed for routine usage. Similar techniques were applied to trimethoprim, assayed by HPLC, in patients with normal renal function and investigations were also commenced into the penetration of drug into peritoneal dialysate. Novel assay techniques were also developed for a range of drugs including 4-aminopyridine, chloramphenicol, metronidazole and a series of penicillins and cephalosporins. Stability studies on cysteamine, reaction-rate studies on creatinine-picrate and structure-activity relationships in HPLC of aminopyridines are also reported.

Interfacing low-energy SAW nebulization with liquid chromatography-mass spectrometry for the analysis of biological samples

Soft ionization methods for the introduction of labile biomolecules into a mass spectrometer are of fundamental importance to biomolecular analysis. Previously, electrospray ionization (ESI) and matrix assisted laser desorption-ionization (MALDI) have been the main ionization methods used. Surface acoustic wave nebulization (SAWN) is a new technique that has been demonstrated to deposit less energy into ions upon ion formation and transfer for detection than other methods for sample introduction into a mass spectrometer (MS). Here we report the optimization and use of SAWN as a nebulization technique for the introduction of samples from a low flow of liquid, and the interfacing of SAWN with liquid chromatographic separation (LC) for the analysis of a protein digest. This demonstrates that SAWN can be a viable, low-energy alternative to ESI for the LC-MS analysis of proteomic samples.

Evidence of lipid degradation during overnight contact lens wear:gas chromatography mass spectrometry as the diagnostic tool

Purpose. We investigated structural differences in the fatty acid profiles of lipids extracted from ex vivo contact lenses by using gas chromatography mass spectrometry (GCMS). Two lens materials (balafilcon A or lotrafilcon A) were worn on a daily or continuous wear schedule for 30 and 7 days. Methods. Lipids from subject-worn lenses were extracted using 1:1 chloroform: methanol and transmethylated using 5% sulfuric acid in methanol. Fatty acid methyl esters (FAMEs) were collected using hexane and water, and analyzed by GCMS (Varian 3800 GC, Saturn 2000 MS). Results. The gas chromatograms of lens extracts that were worn on a continuous wear schedule showed two predominant peaks, C16:0 and C18:0, both of which are saturated fatty acids. This was the case for balafilcon A and lotrafilcon A lenses. However, the gas chromatograms of lens extracts that were worn on a daily wear schedule showed saturated (C16:0, C18:0) and unsaturated (C16:1 and C18:1) fatty acids. Conclusions. Unsaturated fatty acids are degraded during sleep in contact lenses. Degradation occurred independently of lens material or subject-to-subject variability in lipid deposition. The consequences of lipid degradation are the production of oxidative products, which may be linked to contact lens discomfort. © 2014 The Association for Research in Vision and Ophthalmology, Inc.

High performance liquid level monitoring system based on polymer fiber Bragg gratings embedded in silicone rubber diaphragm

Liquid-level sensing technologies have attracted great prominence, because such measurements are essential to industrial applications, such as fuel storage, flood warning and in the biochemical industry. Traditional liquid level sensors are based on electromechanical techniques; however they suffer from intrinsic safety concerns in explosive environments. In recent years, given that optical fiber sensors have lots of well-established advantages such as high accuracy, costeffectiveness, compact size, and ease of multiplexing, several optical fiber liquid level sensors have been investigated which are based on different operating principles such as side-polishing the cladding and a portion of core, using a spiral side-emitting optical fiber or using silica fiber gratings. The present work proposes a novel and highly sensitive liquid level sensor making use of polymer optical fiber Bragg gratings (POFBGs). The key elements of the system are a set of POFBGs embedded in silicone rubber diaphragms. This is a new development building on the idea of determining liquid level by measuring the pressure at the bottom of a liquid container, however it has a number of critical advantages. The system features several FBG-based pressure sensors as described above placed at different depths. Any sensor above the surface of the liquid will read the same ambient pressure. Sensors below the surface of the liquid will read pressures that increase linearly with depth. The position of the liquid surface can therefore be approximately identified as lying between the first sensor to read an above-ambient pressure and the next higher sensor. This level of precision would not in general be sufficient for most liquid level monitoring applications; however a much more precise determination of liquid level can be made by linear regression to the pressure readings from the sub-surface sensors. There are numerous advantages to this multi-sensor approach. First, the use of linear regression using multiple sensors is inherently more accurate than using a single pressure reading to estimate depth. Second, common mode temperature induced wavelength shifts in the individual sensors are automatically compensated. Thirdly, temperature induced changes in the sensor pressure sensitivity are also compensated. Fourthly, the approach provides the possibility to detect and compensate for malfunctioning sensors. Finally, the system is immune to changes in the density of the monitored fluid and even to changes in the effective force of gravity, as might be obtained in an aerospace application. The performance of an individual sensor was characterized and displays a sensitivity (54 pm/cm), enhanced by more than a factor of 2 when compared to a sensor head configuration based on a silica FBG published in the literature, resulting from the much lower elastic modulus of POF. Furthermore, the temperature/humidity behavior and measurement resolution were also studied in detail. The proposed configuration also displays a highly linear response, high resolution and good repeatability. The results suggest the new configuration can be a useful tool in many different applications, such as aircraft fuel monitoring, and biochemical and environmental sensing, where accuracy and stability are fundamental. © (2015) COPYRIGHT Society of Photo-Optical Instrumentation Engineers (SPIE). Downloading of the abstract is permitted for personal use only.

High performance liquid-level sensor based on mPOFBG for aircraft applications

A high performance liquid-level sensor based on microstructured polymer optical fiber Bragg grating (mPOFBG) array sensors is reported in detail. The sensor sensitivity is found to be 98pm/cm of liquid, enhanced by more than a factor of 9 compared to a reported silica fiber-based sensor.

CD4+ T cell surface alpha enolase is lower in older adults

To identify novel cell ageing markers in order to gain insight into ageing mechanisms, we adopted membrane enrichment and comparison of the CD4+ T cell membrane proteome (purified by cell surface labelling using Sulfo-NHS-SS-Biotin reagent) between healthy young (n=9, 20-25y) and older (n=10; 50-70y) male adults. Following two-dimensional gel electrophoresis (2DE) to separate pooled membrane proteins in triplicates, the identity of protein spots with age-dependent differences (p<0.05 and >1.4 fold difference) was determined using liquid chromatography-mass spectrometry (LC-MS/MS). Seventeen protein spot density differences (ten increased and seven decreased in the older adult group) were observed between young and older adults. From spot intensity analysis, CD4+ T cell surface α-enolase was decreased in expression by 1.5 fold in the older age group; this was verified by flow cytometry (n=22) and qPCR with significantly lower expression of cellular α-enolase mRNA and protein compared to young adult CD4+ T cells (p<0.05). In an independent age-matched case-control study, lower CD4+ T cell surface α-enolase expression was observed in age-matched patients with cardiovascular disease (p<0.05). An immune-modulatory role has been proposed for surface α-enolase and our findings of decreased expression suggest that deficits in surface α-enolase merit investigation in the context of immune dysfunction during ageing and vascular disease.

Mechanism of the hydropyrolysis of Manvers coal

Manvers coal has been pyrolysed to 500ºC in a stirred autoclave under various pressures of nitrogen (pyrolysis) and hydrogen (hydropyrolysis). All products were investigated. Pyrolysis of coals involves the transfer of hydrogen atoms from one part of their structure to another. In the above experiments there was no way of labelling the hydrogen or of distinguishing between hydrogen which was initially part of the coal and hydrogen originating in the external atmosphere. Consequently, Manvers coal has been pyrolysed in an atmosphere of deuterium in order to obtain greater insight into the mechanism of hydropyrolysis. In particular it was hoped to distinguish between direct hydrogenation (deuteration!) of the coal and the products of pyrolysis and the 'shuttling' of hydrogen atoms between different parts of the pyrolysing coal. The addition to the coal of 5% (wt.% of coal) of either tetralin or pyrite was also studied. A variety of techniques were used to analyse the products of pyrolysis: gas chromatography - mass spectrometry and high performance liquid chromatography for tars; thermal conductivity gas chromatography and high resolution mass spectrometry for gases; methanol densities, microporosities and diffuse reflectance infra red spectroscopy for the cokes (chars); refractive index to determine deuterium in the liquor. An attempt has been made to apply basic thermodynamics to reactions which are likely to occur in the hydropyrolysis of coals. Diffusion and effusion rates for hydrogen and tar molecules have also been estimated.

Vitamin E-stabilised UHMWPE for orthopaedic implants:quantitative determination of vitamin E and characterisation of its transformation products

The fate of vitamin E and the formation and identification of its transformation products were investigated at different stages of the manufacturing process of commercially produced cross-linked (by γ-irradiation) UHMWPE stabilised with vitamin E (vitamin E infused-post irradiation) used for tibia-components (as articulating surfaces) in total knee arthroplasty (total knee replacement). Vitamin E (α-tocopherol) and its transformation products were extracted from microtomed Tibia films and the different products were separated, isolated, purified using high performance liquid chromatography (HPLC), and characterised by spectroscopic methods and LC-MS. The amount of vitamin E and that of the products formed in the different Tibia samples and in their extracts were also quantified using FTIR and HPLC analysis and calibration curves. Thorough analysis of the Tibia extracts has shown that a number of vitamin E transformation products were formed at different concentrations at two selected stages of the implant manufacturing process that is before and after sterilisation by γ-irradiation. The identified products were found to correspond mainly to different stereoisomeric forms of a small number of vitamin E transformation products. Most of the observed products were of dimeric and trimeric nature with their identity confirmed through a detailed study of their spectral and chromatographic characteristics. It was found that the products of vitamin E, prior to the sterilisation step but after the crosslinking and doping of vitamin E, were mainly the dihydroxydimers and trimers (Tibia samples at this stage are referred to as “Tibia-VEPE”). After sterilisation and completion of the manufacturing process, additional dimers of vitamin E were also formed (Tibia samples at this stage are referred to as ‘Tibia-VEPE-Sterile’), Furthermore, two tocopherol-derived aldehydes (aldehyde 5-formyl-γ-tocopherol and aldehyde 7-formyl-γ-tocopherol) were also formed but at very low concentrations especially in the Tibia-VEPE-Sterile samples. The question of whether vitamin E becomes chemically reacted (grafted) onto the polymer matrix during the manufacturing process of the Tibia is also addressed.