Required knowledge includes: sample preparation procedures including specialised techniques such as: handling unstable/hazardous chemicals and samples, fragile/labile biological material liquid-liquid extraction, solid-phase micro extraction, derivatisation, filtering and dilution/concentration principles for separation of analytes such as: chemical composition of stationary and mobile phases and their types of interaction selection of solvents based on polarity, viscosity, ultraviolet (UV) cut-off requirements for solvent purity and pre-treatment including filtration, degassing, buffering and modifying solvents isocratic and gradient elution flow programming (linear, concave and convex gradients and step) recovery and recycling of solvents separation by polarity: normal phase systems and hydrophilic (interaction chromatography) reverse phase systems and hydrophobic (interaction chromatography) order of elution in normal and reverse phase systems separation by charge: ion exchange chromatography(IEC) and ion chromatography (IC) ion suppression and ion pairing techniques non-suppressed systems separation based on molecular size: size exclusion chromatography (SEC) gel-permeation chromatography (GPC) relationship between retention time and molecular mass bioaffinity chromatography principles chromatography concepts and calculations involving: retention times, peak widths, peak asymmetry, capacity factor k' and resolution column selectivity, column efficiency (plates/m), optimum flow rate, minimum theoretical plate height, Van Deemter and related equations limit of detection, limit of quantitation and their application to quality control procedures operation, construction, selectivity, typical applications, troubleshooting and routine maintenance of LC columns including: (semi) preparation columns, packed columns and capillary columns column oven (role of temperature in achieving close separations) checking for leaks, changing of columns, lines and valves system flushing and conditioning and storage of columns operation, construction, selectivity, typical applications, troubleshooting and routine maintenance of LC sample introduction systemsincluding: manual and auto-injection, injector valves, solvent reservoirs, selector valves, gradient programmer, mixing manifolds and column switching pump designs such as reciprocating piston or diaphragm, pressures, flow rates operation, construction, selectivity, sensitivity, linear range, typical applications, troubleshooting and routine maintenance of LC detectors including: UV fixed wavelength and dispersion/diode array multi-wavelength detectors conductivity detector (ECD) electrochemical detector (ECD) fluorescence detectors refractive index (RI) evaporative light scattering (ELSD) mass spectrometry (LC-MS) (LC-MS-MS) using full scan or selective ion monitoring (SIM) flow splitting, tandem detectors (e.g. UV and MS) routine quality control procedures such as use of manual/computer calibration charts and/or standards computer control software for operating and optimising instrument (peak detection and integration, drift parameters, baseline correction and instrument/integrator zero) procedures for optimising instrument performance such as: optimising separation by changing solvent composition including use of mobile phase gradient computer control programs investigation of elution order in normal and reverse phase systems effects on instrumental outputs and analytical results by fine tuning injection, mobile phase flow rate, column pressures and changing column type or detector aligning MS interface cones steps in identifying and quantifying analytes including relative retention data, peak area normalisation and response factors calculation steps to give results in appropriate units and precision troubleshooting and maintenance procedures recommended by instrument manufacturer enterprise and/or legal traceability requirements relevant health, safety and environment requirements
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