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Standard Data Analysis - Integration ChemStation - Level 1 - Training.

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Presentation on theme: "Standard Data Analysis - Integration ChemStation - Level 1 - Training."— Presentation transcript:

1 Standard Data Analysis - Integration ChemStation - Level 1 - Training

2 Agilent Restricted Integration locates the peaks in a signal and calculates their size What is Integration

3 Agilent Restricted T he Integration Process consists of the following: construct initial baseline peak recognition – Start and end Time finds the apex of the peak baseline allocation (construction) peak area measurement Integration Process

4 Agilent Restricted 1 Slope and Curvature within limit 2 Slope and curvature above limit: peak? 3 Slope remains above limit: peak recognized 4 Curvature becomes negative: front inflection 5 Negative slope: Apex 6 Curvature becomes positive: rear inflection point 7 Slope and curvature within limit: end of peak? 8 Slope and curvature within limit: end of peak Start, End and Apex of a Peak

5 Agilent Restricted Fit highest data point and those on each side to a quadratic equation, solve for highest point Determining Peak Apex

6 Agilent Restricted BB BV VB BB BV VB Baseline Tick Marks The default baseline is constructed as a series of straight line segments between: the start of the baseline, the tick marks, and the end of the peak. Default Baseline Construction

7 Agilent Restricted Standard or Enhanced Integrator  The ChemStation includes two integrators: the Enhanced Integrator (default) and the Standard Integrator (Genie)  New methods by default use the Enhanced Integrator and can not be down graded to the Standard Integrator  Old methods can be updated to use the Enhanced Integrator or maintained using the Standard Integrator so customers can continue using validated methods.  The default method “defoldlc.m” has the Standard Integrator  The default method “def_lc.m” has the Enhanced Integrator

8 Agilent Restricted Both integrators co-exist so you may continue to use validated methods The Enhanced Integrator offers better:  baseline allocation on wandering baselines  rejection of noise peaks through the use of an initial peak height parameter  control of integration start and stop marks  peak allocation on noisy signals  peak shoulder allocation through second derivative calculations Standard or Enhanced Integrator

9 Agilent Restricted Loading Signals Select path, data file name and channel Select or ‘Load Signal’ from File Menu Show/Hid e the Signal Informatio n Choose to load all available or selected signals only

10 Agilent Restricted You may also integrate by: Selecting Integrate or Auto Integrate from the Integration menu. Selecting the Integration Tool. Running a method where the Run Time Checklist includes Data Analysis When a Signal is Loaded, Integration May Occur Automatically

11 Agilent Restricted Initial Values Threshold Peak Width Area Reject Shoulders Standard Integrator - Initial Values

12 Agilent Restricted Standard Integrator - Timed Events What timed events could you use for this integration?

13 Agilent Restricted Auto Integrate determines the optimum values for the initial threshold, peak width, and area reject by integrating the chromatogram twice.  Threshold - based on signal range and averaged baseline noise.  Peak Width - proportional average of the peak widths of the first five peaks.  Area Reject - 1/1000st of the greatest area found. Auto Integration - Standard Integrator

14 Agilent Restricted Select Integration Tasks Integrate Current Chromatogram Auto Integrate Integration Events Manual Integration Draw Peak Baseline Manual Integration Negative Peaks Manual Integration Tangent Skim Manual Integration Split Peaks Manual Integration Remove Peaks Integration Tools - Standard Integrator

15 Agilent Restricted The Enhanced Integrator offers better:  baseline allocation on wandering baselines  rejection of noise peaks through the use of an initial peak height parameter  control of integration start and stop marks  peak allocation on noisy signals  peak shoulder allocation through second derivative calculations Enhanced Integrator

16 Agilent Restricted You may update methods developed on previous revisions of the ChemStation to the new Enhanced Integrator. Once you have updated the new method, you cannot revert back to the old integrator. Enhanced Integrator

17 Agilent Restricted Enhanced Integrator UI Define integration parameters just by selecting and clicking on the chromatogram There are two sets of integration events : - intial events - time-based Use:Slope Sensitivity  to define peak sensitivity Peakwidth  to set an initial sampling interval for the integrator to distinguish peaks from baseline noise Area Reject  to filter small peaks Height Reject  to set noise rejection

18 Agilent Restricted Advanced Baseline Absorbance Time [min.] Options Off Advanced Integration Options On Advanced Integration Peak Baseline Peak Area Unassigned

19 Agilent Restricted Using Initial Parameters Peak Width Height Reject

20 Agilent Restricted Add Timed Events Delete a Timed Event Enhanced Integrator - Timed Events

21 Agilent Restricted Solvent peak (Skim peak) Tangent Peak Straight Line Calculation Exponential Calculation Tangent Skim

22 Agilent Restricted Shoulders occur when two peaks are so close together that no valley exists between them Peak with shoulders Tangent ShouldersDrop Line Shoulders Shoulder Detection

23 Agilent Restricted Enhanced Integrator - Auto Integration  Examines beginning and end regions to estimate noise.  Assigns initial Slope Sensitivity and Height Reject.  Assigns temporary Peak Width value for first pass integration.  Sets Area Reject to zero.  Performs trial integration, may be repeated several times.  Calculates Peak Width based on early eluting peaks.  Refines Slope Sensitivity and Height Reject.  Computes Area Reject from initial Peak Width and noise level.

24 Agilent Restricted Integration Tasks Integrate Current Chromatogram Auto Integrate Integration Events Set Baseline Now Set Baseline Hold Set Tail Tangent Skim Set Integration Set Slope Sensitivity Set Area Reject Set Fixed Peak Width Set Detect Shoulders Enhanced Integrator - Tool Bar Use Auto Integrate to find suitable integration events Only visible with integration events open

25 Agilent Restricted Save Events to Method Save Integration Events as Part of a Method

26 Agilent Restricted Manual Integration Manual integration events

27 Agilent Restricted Manual Integration Use Cursor tools to perform manual integration by selecting the required action and dragging a line over or clicking on the chromatogram (depending on the task) Zoom in using the zooming tools to better view your baseline The new integration results are immediately visible in the user interface If your chromatography is so demanding that you can’t use the the automatic integration process  Use the manual integration tool set

28 Agilent Restricted Option 1: Save Manual Integration to Method Select Integration, Copy Manual Events to Method Save Manual Integration - To the method

29 Agilent Restricted Step 1: Select View, Windows Functions, Save Window Save Manual Integration - To the Raw Data File

30 Agilent Restricted Note: manual changes are stored in the raw data file directory. Step 2: Click OK to use default name or rename Save Manual Integration - To the Raw Data File

31 Agilent Restricted Select View, Windows Functions, Load Window To Recall Manual Integration From Data File

32 Agilent Restricted Lab Time !!!!! Lab 5 - Integration


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