solvents which would have an impact on flow rate gradient separations in normal

Solvents which would have an impact on flow rate

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solvents, which would have an impact on flow rate, gradient separations in normal phase are much more difficult. Chromatography in the reverse phase also has the benefit that pH selectivity can be used to enhance separations. Aim Problem Statement: A Ketoprofen sample is suspected to be contaminated during manufacturing process - suspected blender contamination. Suspected components: Sulindac, Naproxen, Diclofenac and Phenylbutazone. SKS has added Uracil to the Composite (CP) sample for ease of calculating t0. In this investigation, Lucinda would like SopPharma to resolve the CP and check selectivity on different columns and to different solvent systems (depending on the group). Hypothesis: We hypothesise that AUC would remain stable and H would increase and W would decrease if we increased the flow rate. We hypothesised that an increase in flow rate would cause BP to increase. We hypothesised that an increase in flow rate would cause RT to decrease.
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HPLC CHROMATOGRAPHY 8 Specific Column allocated by SKS Column and Its Properties Solvents for mobile phases: Acetonitrile Methanol The MilliQ water system has been repaired and purified water can be used Formic acid Conditions: Flow rate : Variable Temperature: Variable Injection volume : Variable Wavelength : 254nm Material Delivered by SKS for this session of Investigation: Standard Sample (0.1mg/ml): Sulindac, Piroxicam, Ketoprofen, Naproxen, Diclofenac Na, Phenylbutazone standards have been provided by SKS to develop the method. The sample containing the standards will be called the Composite (CP) and each standard should be at 0.1mg/ml. SKS recommends to also use Uracil in the standard sample to simplify the various calculations. Standard Preparation Diluent for Stock Solution : 70% acetonitrile: 30% distilled water
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HPLC CHROMATOGRAPHY 9 Stock 1 Solution (2.5mg/ml): Accurately weigh approximately 50mg of each of the standards in an aluminium weighing boat and gently slide into a 20 ml volumetric flask. Dilute to volume with prepared diluent and invert the flask 20 times. Label the flask clearly as Stock 1. Stock 2 Solution (0.1mg/ml) Accurately pipette 400uL of each of the stock 1 into a 10 ml volumetric flask. Dilute to volume with prepared diluent and invert the flask 20 times to dissolve the standard sample. Label the flask clearly as Stock 2. Solvents for Mobile Phase: Acetonitrile or Methanol is the only available solvent at that stage. Formic acid will also be delivered and should be used at 0.1%. No water can be used at that stage as we understand that your MilliQ water system is currently down. Columns: 8 Columns have been sent to Sop-pharm Lab and are all C18- 150x4.6x5, some seem to be branded Pinnacle C18-150x4.6x5. In this separation method (Reversed Phase separation) there are three different PAH (polycyclic aromatic hydrocarbons) components. The naphthalene molecule is the least hydrophobic while the chrysene molecule with more carbon atoms and more aromatic rings is the most hydrophobic. For the mobile phases, water is the most polar followed by methanol then acetonitrile. THF and hexane are the non-polar for mobile phases. Silica is the most
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