Browsing by Author "Saini, S."
Now showing 1 - 6 of 6
Results Per Page
Sort Options
Item Open Access An amperometric enzyme electrode for the detection of L-lactate(Cranfield University, 1997-06) Selkirk, Jane Yvonne; Turner, Anthony P. F.; Saini, S.The main tasks of this thesis were to evaluate a number of amperometric enzyme electrode chemistries for the selective and sensitive detection of L-lactate, and apply mass fabrication technologies to reproducibly manufacture sensors in a controllable manner. The sensors studied were based on the use of lactate oxidase with a range of modified-carbon electrodes. Noble metals, hexacyanoferrate (111) or Prussian Blue were used to modify carbon electrodes for the electro-catalytic determination of hydrogen peroxide, the product of the reaction of lactate oxidase with L-lactate. Tetrathiafulvalene was employed as an artificial mediator between the enzyme and the electrode. Polypyrrole was tested as a means of immobilising lactate oxidase and to achieve direct charge transfer to the underlying carbon electrode. The characteristics of the sensor responses to hydrogen peroxide, L-lactate and ascorbate were compared, in relation to the electrochemical electrode area. From this investigation, it was confirmed that screen-printed electrodes were more reproducible to manufacture than hand-fabricated electrodes. For screen-printed rhodinised-carbon electrodes, an operating potential of +400 mV (SCE) was selected. Interference from ascorbic acid and sensitivity to hydrogen peroxide were deten-nined to be 26 gA. mM-' CM-2 and 27 gA. mM-'. CM-2, respectively. Screen-printed carbon electrodes modified with platinum, rhodium or palladium were selected for further investigation. Rhodium on carbon performed the best in ten-ns of sensitivity and selectivity at low potentials, and different formations of rhodium-carbon complexes were studied. Although rhodium electroplated onto carbon screen-printed electrodes was examined, printing inks made from a preformed powder of rhodium on carbon-graphite proved to be the preferred route of electrode fabrication. Screen printing, ink-jet printing and Cavro solution deposition were employed to fabricate the amperometric enzyme electrodes. These sensors were composed of rhodinised carbon and lactate oxidase in a water-based electrode ink with a protective outer membrane layer. Each stage, from ink preparation to membrane composition, was developed empirically. The sensitivity, stability and reproducibility of the working electrode was improved by altering it to a homogeneous ink, consisting of carbon graphite powder, rhodinised carbon powder (5% Rh by weight), hydroxyethyl cellulose (2% w/v) and lactate oxidase in the weight ratio of 2: 8: 18: 1. A layer of cellulose acetate (2% w/v in a 1: 1 solution of acetone to cyclohexanone) and an outer coating of a polyurethane called Pellethane (I% to 4% w/v in dimethyl formarnide and tetrahydrofuran) improved the selectivity, sensitivity and detection range of the sensor, allowing it to operate in physiological solutions with reduced passivation from protein adsorption. The sensor design was revised to allow its passage through a catheter and operation within a blood vessel; it was manufactured on flexible material using screen printing and Cavro solution deposition techniques. These miniature sensors, with a working surface of 0.5 x 15 mm, were capable of linearly measuring lactate up to 3 mM in buffer solutions with an average sensitivity of 44.8 nA. mM-1 L- actate. To test the sensor operation in physiological solutions, a flow injection system was employed. A planar three-electrode card used in this system was manufactured using screen printing and Cavro solution deposition techniques. L-lactate concentrations up to 6.4 mM were sensitively and, after minor correction, accurately determined in undiluted plasma and whole blood samples. This thesis has therefore made progress toward mass fabricating an amperometric enzyme electrode device suitable for the deten-nination of L-lactate concentrations in vitro.Item Open Access At-line testing of chlorophenol and chloroanisole contaminants in commercial wine production.(Cranfield University, 2004-09) Nicholls, C. R.; Saini, S.; Setford, S.The research described in this thesis concerns the development of at-line test procedures for the detection of trace levels of chlorophenols and chloroanisoles in wine and related materials. Competitive ELISA assays were developed and optimised for pentachlorophenol and pentachloroanisole to enable the detection of chlorophenols and chloroanisoles in the range 0.1 to 100ng/ml in purified sample extracts, using antibodies supplied by the French consortium partner, Diaclone, together with synthesised conjugate materials, based on the enzyme horseradish peroxidase. The cross reactivity of the assay towards chlorophenol and chloroanisole congeners in wine was investigated and the pentachlorophenol assay was selected as the most efficient antibody to detect and quantify both chlorophenol and chloroanisole congeners. The use of synthetic receptors based on molecularly imprinting technology was also investigated for pentachlorophenol and pentachloroanisole, and a new assay format (Displacement Imprinted Polymer Analysis (DIPRA)) was established to measure chlorophenols in contaminated materials at a concentration range of 0.1 to 50 µg/ml, based on the displacement of a reporter molecule rebound to the synthetic receptor sites. Two alternative reporter molecules were synthesised by linking a pentachlorophenol derivative (2,3,5,6-tetrachloro-4-hydroxy phenoxy acetic acid) to the enzyme horseradish peroxidase or 7-amino-4-methylcoumarin-3-acetyl hydrazide. Alternative hyphenated sample extraction and purification methods based on solid phase extraction, steam distillation and liquid/liquid partition were evaluated to enable the ELISA and DIPRA test methods to be employed using the limited facilities of a local winery laboratory. The application of the procedures to the analysis of sample types such as wine, corks and packaging materials was undertaken to investigate the suitability of the test methods for inclusion in quality control and incident analysis protocols.Item Open Access Diagnostic technologies for wound monitoring(Cranfield University, 2006-03) Trill, Helen; Setford, S.; Saini, S.Chronic wound infections represent a worldwide problem, generating high morbidity and medical expense. Failure to control infections such as MRSA in the reparative process of a wound can cause disruption of normal anatomical structure and function, resulting in a chronic wound. Existing approaches to identifying infection largely involve surveying a range of physical parameters, and a limited use of non-invasive technologies. Evaluation is time consuming, and often results in inconsistencies in patient care. This project researches three possible alternative methodologies/technologies for the monitoring of wounds, by measuring components of wound fluid. Two of the three technologies are designed to be used by physicians and patients, similarly to commercially available home blood glucose test kits, and are based on the measurement of three biomarkers: glucose, ethanol and H2O2 in PBS, and in serum as surrogate wound fluid. The first is a voltammetric technique known as dual pulse staircase voltametry (DPSV), which produces peaks characteristic of particular analytes at an electrode. The second is an amperometric biosensor array, based on screen printed three electrode assembies of carbon, rhodinised carbon (glucose biosensor only) and Ag/AgCl reference. The glucose biosensor uses glucose oxidase enzyme as the biorecognition agent, the H2O2 biosensor is a mediated system using horseradish peroxidase enzyme and dimethylferrocene mediator, and the ethanol biosensor is a bienzyme mediated system utilising alcohol oxidase enzyme horseradish peroxidase enzyme and coupled dimethylferrocene mediator. Wounds are known to produce characteristic odours, therefore the third technology studied is a single sensor odour analyser with advanced data analysis to detect five commonly occuring wound bacteria, S.aureus, K.pneumoniae, S.pyogenes, E.coli and P.aeruginosa in growth media and surrogate wound fluid. This technology would be used as a 'near patient' monitoring system and is based on machine olfaction similar to that of a commercial electronic nose, but uses a single metal oxide sensor in combination with principle components analysis. DPSV scans of the individual analytes demonstrated distinctive peaks, exhibiting nonlinear relationships with concentration. A great deal of useful information was generated using this technique, however, limitations were discovered regarding repeatability and inter-analyte interference in mixtures. Limits of detection in surrogate wound fluid with the glucose biosensor, hydrogen peroxide biosensor, and ethanol biosensor were as follows: 169.5 µM glucose, 8.43 µM hydrogen peroxide, and 7.94 µM ethanol respectively (all at 99.7% confidence). Direct detection of ethanol from metabolically active S.aureus in surrogate wound fluid yielded a limit of detection of 1.23 x 108 CFU/ml at 99.7% confidence, and 19 µM in terms of ethanol specific response. The single sensor odour analyser demonstrated the ability to detect and discriminate between the three biomarkers, between five bacteria individually, and partial discrimination of paired bacteria (in broth and surrogate wound fluid). It was also found that S.aureus could be detected down to a cell density of 5x106CFU/ml in surrogate wound fluid, lower than that found for the biosensor concept.Item Open Access Electrochemical method for the rapid in situ screening of heavy metals in soil and water samples(2004-03) Cooper, Joanne; Setford, S.; Bolbot, J. A.; Saini, S.The aim of this work has been to develop a field-based electrochemical sensor for the detection of cadmium (Cd), lead (Pb), copper (Cu), zinc (Zn), arsenic (As) and mercury (Hg) in soil and water samples. In situ analysis, using rapid sample screening tools, can facilitate monitoring and remediation processes by minimising delays and substantially reducing costs. Disposable screen-printed electrodes (SPEs) have been fabricated using low-cost thick film technology. Carbon SPEs, coupled with anodic stripping voltammetry (ASV), provided a rapid, inexpensive in situ electrochemical screening sensor capable of simultaneously detecting Cd, Pb and Cu in soil and water samples in the microgram per liter range (pg I*1). A simplified soil extraction procedure, using 1 mol I'1 aqua regia and a three minute ultrasonic sample agitation, was developed for use with the sensor, addressing the lack of existing field-based soil extraction protocol. Extraction efficiency was evaluated using a soil certified reference material (CRM). Recoveries of 64%, 52% and 57% for Cd, Pb and Cu respectively were obtained, with a relative standard deviation (RSD) of <8% for all analytes (n=10). 82 soil samples were tested using the combined extraction + ASV sensor and compared against standard ICP-MS analysis. Correlation coefficients of 0.9782, 0.9728 and 0.9869 for Cd, Pb and Cu, respectively, indicated a good linear relationship between the two methods. A novel gold (Au) SPE, fabricated using a curable Au ink, was next evaluated for the detection of As and Hg. This is an improvement in the current state-of the-art as no surface modification or subsequent electrode adulteration to facilitate metal analyte depositioning is required. Furthermore, the Au ink can be cured at a low temperature (130°C), permitting electrode fabrication using disposable, cheap, plastic substrates. A patent has been filed. A limit of detection (LOD) of 2.7 pg I"1 and 1 pg I"1 for As3+ and Hg2+, respectively, was obtained following a 30 s depositioning. Speciation measurements for As were performed by reducing As5+ effectively over a short (10 min) reduction time using cysteine. An LOD of 42 pg f 1 following a 30 s depositioning was obtained. Initial studies also indicated the possibility of using the Au SPEs for measuring Cd, Pb and Cu, providing an alterative Hg-free procedure.Item Open Access Identification and carcinogenicity of polycyclic aromatic hydrocarbons in transformer oil.(Cranfield University, 2003-05) Pillai, Ilona G.; Saini, S.; Setford, S.; Heyward, R.Insulating oils are rich in polycyclic aromatic hydrocarbons (PAHs) which act as inhibitors of oil breakdown, but are believed to be the main source of oil mutagenicity when converted to their epoxide form by mammalian enzymes. The current industry- recognised measurement of oil risk, the IP 346 method, measures total aromatic content and therefore cannot be directly related to PAH risk. This thesis describes efforts made to establish the contribution of PAH species alone to total oil mutagenicity. Cont/d.Item Open Access A peroxide sensor for the food and beverages industry(2001-11) Moody, A. P.; Saini, S.Due to desirable environmental reasons, peroxides have replaced halogenated substances for disinfection purposes in the food and beverage industry. However, cost issues and the requirement to completely remove these agents after disinfection necessitates simple, low-cost and sensitive test methods with a wide dynamic range and on-line capability. The development and performance of such a method is detailed in this thesis. The novelty of this work was in the simplified production of the sensor by pre-mixing all active components before application onto a screen-printed graphite base electrode in a single step. Having established the principle of pre-mixing all the components, the proportions were optimised with regard to a balance between cost and required performance in the industrial application. Once this had been established, the performance and stability was evaluated in the laboratory, a comparative study was performed against other methods for hydrogen peroxide determination and finally the sensor was demonstrated to work in a small-scale industrial field trial. Some of the available mass-production processes for the sensor were considered for suitability and a recommendation made. The relative analytical performances of sensors produced manually and by the recommended process were compared to their visual appearance under a microscope and a possible quality control process suggested for the manufacturing stage. Finally, a scanning electrochemical microscope was commissioned and a method for producing ultramicroelectrodes for use with it developed. This was then used to study the variation of the sensor surface coating with the aim of facilitating the optimisation of the composition, production and quality control processes.