Nexera Advanced Troubleshooting

Browser lacks capabilities required to play.

Upgrade or switch to another browser.

Loading…

Config.debug = false;

<div class="container"><h1 class="title">Troubleshooting System Peaks</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/ColumnContamination_SysPeaks.jpg" alt="Troubleshooting System Peaks"></div> Column contamination is caused from a buildup of material that is retained on the stationary phase. This buildup of contamination may cause retention time shift, extraneous peaks, loss of resolution and adverse affects to the peak shape. Particulate matter that is precipitated from the samples and buffers may also contaminate a column by plugging the column frit and column packing. These contaminants may arise from any of the following sources: *Samples that do not fully elute from the stationary phase using the current mobile phase.Most common with mobile phases that have a low organic ratio. * particulate matter in the samples or mobile phase that eventually block the column frit at the column inlet. * impurities in the mobile phase that strongly adsorb to the column packing. Most common with ion-pair chromatography and when mobile phase additives are added to low organic isocratic mobile phases. <div class="ts-question">Does the extra peak have a consistent retention time in every injection?</div> <div class="buttons"> <span class="btn-yes">[[Yes->Null Injection]]</span> <span class="btn-no">[[No->Column Contamination - Avoiding]]</span> </div> </div>

<div class="container"><h1 class="title">Null Injection</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/Null_Injection_SIL-30AC.jpg" alt="Troubleshooting System Peaks"></div> <h3>Make two consecutive null injections.</h3> It is possible to program the autosampler to run a normal analysis process without physically moving the injection mechanisms. Local Control - Examine the SIL display and verify that the vial numbers are not entered as (-1). Remote Control - Examine the data system software to verify that the vial number is not entered as (-1). <div class="ts-question">Is the extra peak still present in the chromatogram?</div> <div class="buttons"> <span class="btn-yes">[[Yes->HPLC System Flush]]</span> <span class="btn-no">[[No->Injection Sequence]]</span> <span class="btn-back"><<back "Back">></span> </div> </div>

<div class="container"><h1 class="title">Column Contamination - Avoiding</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/ColumnDecontamination.jpg" alt="Column Contamination - Avoiding"></div> Column contamination is caused from a buildup of material that is retained on the stationary phase. The buildup of contamination can be avoided by following some simple preventative procedures. * Sample cleanup - This could include filtration and solid extraction techniques. * Solvent quality - Always use HPLC grade solvents (or better) for mobile phase preparation Always filter all solutions that are used in mobile phase preparation. * Column maintenance - Incorporate a strong solvent (100% ACN) column flush in the regular column maintenance procedure. This causes any material retained on the stationary phase to be washed off the column's packing material. * Guard column - Incorporate the use of an in-line guard column between the injector and the column inlet. Guard columns collect any compounds that would have been retained in the analytical column, thus extending the life of the column. Guard columns are inexpensive when compared to the cost of a new analytical column. Monitor a peak parameter, such as width, to determine when the guard column should be changed. <div class="buttons"> <span class="btn-back"><<back "Back">></span> </div> </div>

<div class="container"><h1 class="title">HPLC System Flush</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/ColumnUnion.jpg" alt="HPLC System Flush"></div> Contaminants in the system tubing may create active sites to which sample components adhere, causing peak tailing and other chromatographic problems. In these instances, it is necessary to flush (clean) the system tubing to remove these contaminants and active sites. <ol> <li>Remove the column and replace it with a tubing union.</li> <li>Depending on the composition of the current mobile phase, flush the system with 100% filtered degassed water for 10 minutes at a flow rate of 3mL/min.</li> <li>Exchange the water for an organic solvent in which the suspected contaminant is soluble.</li> <li>Flush the system with 100% filtered degassed organic solvent for 10 minutes at a flow rate of 3mL/min.</li> <li>Replace the solvent with water and flush for 10 more minutes at a flow rate of 3mL/min.</li> <li>Purge the system with fully degassed and filtered mobile phase.</li> <li>Reinstall the column and equilibrate the system at the appropriate method parameters (flow rate, etc.).</li> </ol> <div class="buttons"> <span class="btn-back"><<back "Back">></span> </div> </div>

<div class="container"><h1 class="title">Injection Sequence</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/Blanks.jpg" alt="Injection Sequence "></div> <ol> <li>Make two consecutive injections from the LAB blank sample vial.</li> <li>Make two consecutive injections from the SYSTEM blank sample vial.</li> </ol> <img class="warning" style="float: left;" src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/virtual-advisor-warning-icon.png" alt="warning-icon"><strong>NOTE: When performing troubleshooting procedures it is generally a good idea to make more than one injection to look for recurring patterns that may be evident in a longer period than the normal run time.</strong> <div class="ts-question">Does the extra peak appear in the SYSTEM blank?</div> <span class="btn-link">[[SYSTEM Blank->SYSTEM Blank]]</span><br> <span class="btn-link">[[LAB Blank Prepare->LAB Blank Prepare]]</span><br> <div class="buttons"> <span class="btn-yes">[[Yes->LAB Blank]]</span> <span class="btn-no">[[No->Active Sites]]</span> <span class="btn-back"><<back "Back">></span> </div> </div>

<div class="container"><h1 class="title">Active Sites</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/Adsorb_ActiveSites.jpg" alt="Active Sites"></div> <h3>Active sites in the system may cause compounds to adhere temporarily before being washed off in a subsequent injection.</h3> It is important to be aware of the surfaces that the sample comes into contact with to ensure that the components of the flow line are non-reactive with the sample and mobile phase. A partial list of the surfaces that may contact the sample are listed below. <ul> <li>Sample preparation components (filters, pipettes, vials, etc.)</li> <li>Sample vial, septum and closure</li> <li>Sampling needle and sample loop</li> <li>Valve components</li> <li>Sample transfer tubing and tubing unions</li> <li>Column packing material</li> <li>Flow cell components</li> <li>Sample transfer tubing and tubing unions PEEK tubing is generally more resistant to chemical adsorption than stainless steel.</li> </ul> <div class="ts-question">Is the sample reacting with active sites in the system?</div> <span class="btn-link"><a href="/service-support/virtual-advisor/nexera/advanced-troubleshooting/troubleshooting-the-autosampler/index.html">Troubleshooting the Autosampler</a></span><br> <div class="buttons"> <span class="btn-yes">[[Yes->Active Sites System Components]]</span> <span class="btn-back"><<back "Back">></span> </div> </div>

<div class="container"><h1 class="title">LAB Blank Prepare</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/ColumnDecontamination.jpg" alt="LAB Blank"></div> <h3>Prepare and make two injections each of a LAB blank and a SYSTEM blank. </h3> The LAB blank solution is prepared to obtain a blank of the sample preparation process, i.e., the sample solution without the original sample. This is injected to look for peaks originating in the sample preparation process. Using an equivalent amount of the actual sample solvent, perform all of the handling techniques that are normally performed for each sample solution, without the original sample. <img class="warning" style="float: left;" src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/virtual-advisor-warning-icon.png" alt="warning-icon"><strong>NOTE: If samples are extracted, filtered, diluted etc., perform these procedures on the blank solution in the same manner.</strong> <div class="ts-question">Does the extra peak appear in the SYSTEM blank? </div> <span class="btn-link">[[SYSTEM Blank->SYSTEM Blank]]</span><br> <span class="btn-link">[[Injection Sequence->Injection Sequence]]</span><br> </div> <div class="buttons"> <span class="btn-yes">[[Yes->LAB Blank]]</span><span class="btn-no">[[No->Active Sites]]</span> <span class="btn-back"><<back "Back">></span> </div>

<div class="container"><h1 class="title">Sample Preparation</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/ColumnDecontamination.jpg" alt="Sample Preparation"></div> Depending on the complexity of the sample preparation procedure, including the extraction process, there are a various stages at which preparation errors can be made. Some common things to re-examine are listed below: <ul> <li>Clean glassware - glassware may be contaminated with substances that can absorb the analyte or keep it from being fully dissolved.</li> <li>Appropriate solvents - inappropriate solvents may not fully dissolve the sample.</li> <li>Correct measurements - ensure that the appropriate amount of sample was added to the sample vial.</li> <li>Proper filtration - the wrong filter could absorb the sample before it reaches the sample vial.</li> <li>Vial and septa type - the wrong vial type or septa type may absorb the sample from the sample solution. </li> </ul> Carefully examine the sample preparation and review the process for accuracy. <div class="buttons"> <span class="btn-back"><<back "Back">></span> </div> </div>

<div class="container"><h1 class="title">Active Sites System Components</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/Adsorb_ActiveSites.jpg" alt="Active Sites"></div> It is important to be aware of the surfaces that the sample comes into contact with to ensure that the components of the flow line are non-reactive with the sample and mobile phase. A partial list of the surfaces that may contact the sample are listed below. <ul> <li>Sample preparation components (filters, pipettes, vials, etc.)</li> <li>Sample vial, septum and closure</li> <li>Sampling needle and sample loop</li> <li>Valve components</li> <li>Sample transfer tubing and tubing unions</li> <li>Column packing material</li> <li>Flow cell components</li> <li>Sample transfer tubing and tubing unions PEEK tubing is generally more resistant to chemical adsorption than stainless steel.</li> </ul> PEEK tubing is generally more resistant to chemical adsorption than stainless steel. <div class="buttons"> <span class="btn-back"><<back "Back">></span> </div> </div>

<div class="container"><h1 class="title">SYSTEM Blank</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/Waste_SystemBlank.jpg" alt="SYSTEM Blank"></div> <h3>Prepare and make two injections each of a LAB blank and a SYSTEM blank.</h3> <ol> <li>Allow the instrument to equilibrate fully.</li> <li>Using a similar sample vial, collect the liquid from the detector outlet tubing (waste tubing)</li> </ol> <img class="warning" style="float: left;" src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/virtual-advisor-warning-icon.png" alt="warning-icon"><strong>NOTE: Use caution to avoid contamination of the inside or rim of the sample vial.</strong> <div class="ts-question">Does the extra peak appear in the SYSTEM blank?</div> <span class="btn-link">[[LAB Blank Prepare->LAB Blank Prepare]]</span><br> <div class="buttons"> <span class="btn-yes">[[Yes->LAB Blank]]</span> <span class="btn-no">[[No->Active Sites]]</span> <span class="btn-back"><<back "Back">></span> </div>

LAB Blank and LAB Blank Prepare have the same names in current nexera virtual advisor. I changed to avoid confusion. I also did not follow one of the back buttons they had oddly programmed. It went back to a different node than where it was sent from.

<div class="container"><h1 class="title">LAB Blank</h1> <div class="center"><img src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/nexera/troubleshooting-system-peaks/ColumnDecontamination.jpg" alt="LAB Blank"></div> <h3>Refer to the injections made in the previous question.</h3> The LAB blank solution is prepared to obtain a blank of the sample preparation process, i.e., the sample solution without the original sample. This is injected to look for peaks originating in the sample preparation process. Using an equivalent amount of the actual sample solvent, perform all of the handling techniques that are normally performed for each sample solution, without the original sample. <img class="warning" style="float: left;" src="https://www.ssi.shimadzu.com/sites/ssi.shimadzu.com/files/support/virtual-advisors/virtual-advisor-warning-icon.png" alt="warning-icon"><strong>NOTE: If samples are extracted, filtered, diluted etc., perform these procedures on the blank solution in the same manner.</strong> <div class="ts-question">Does the extra peak appear in the LAB blank?</div> <div class="buttons"> <span class="btn-yes">[[Yes->Sample Preparation]]</span> <span class="btn-back"><<back "Back">></span> </div> </div>