Size exclusion chromatography (SEC) columns are highly sensitive to operating conditions, including the composition of the mobile phase and the nature of analyzed samples. Improper handling and unsuitable method conditions can significantly reduce column lifetime. This guide summarizes common causes of column degradation and provides practical recommendations for extending column lifespan.<h3>Common Causes of Reduced Column Lifetime</h3>Several factors can negatively affect the performance and durability of SEC columns:<h4>1. Contamination of the Stationary Phase</h4><ul><li>Accumulation of impurities from samples or mobile phases</li><li>Microbial growth due to infrequent replacement of mobile phase</li></ul><h4>2. Stationary Phase Degradation</h4><ul><li>Caused by unsuitable pH conditions</li><li>Exposure to inappropriate temperatures</li><li>Use of harsh chemical agents</li></ul><h4>3. Improper Column Storage</h4><ul><li>Incorrect storage conditions may reduce column performance over time</li></ul><h4>4. Column Inlet Blockage</h4><ul><li>Particulates from insufficiently prepared samples can clog the column inlet</li></ul><h3>Preventive Measures and Best Practices</h3><h4>1. Preventing Contamination</h4><ul><li>Use high-quality solvents</li><li>Prepare buffer solutions daily</li><li>Filter both mobile phases and samples before use</li><li>Perform regular column cleaning</li></ul>Important note: Not all SEC columns can be reverse flushed. For example:<ul><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1534">Biozen dSEC-2</a> and Yarra SEC-X7 columns must not be backflushed</li></ul><h4>2. Protecting the Stationary Phase</h4><ul><li>Operate within recommended pH and temperature limits</li><li>When using strong denaturants (e.g., urea or SDS):<ul><li>Perform immediate and thorough water flushing after use</li></ul></li></ul><h4>3. Proper Column Storage</h4>Before storage always clean the column thoroughly<ul><li>Two storage options:<ul><li>Buffer with bacteriostatic agent (e.g., sodium azide)</li><li>No buffer: water with 20% methanol</li></ul></li></ul>When introducing methanol:<ul><li>Use low flow rates</li><li>Prevent pressure spikes that could damage the column</li></ul>When restarting the column:<ul><li>Consider that the column contains 20% methanol</li><li>Adjust flow conditions accordingly</li></ul><h4>4. Preventing Column Inlet Clogging</h4>Use protective components:<ul><li>Guard column</li><li>Precolumn</li></ul>These act as barriers to prevent contamination of the analytical column.<h5>Types of Protection:</h5><ul><li>Security guard cartridges<ul><li>Minimal added length</li></ul></li><li>Short precolumns<ul><li>Same stationary phase as analytical column</li></ul></li></ul><h5>Key Advantage:</h5><ul><li>Essential for columns that cannot be backflushed</li><li>Protects against:<ul><li>Poorly centrifuged samples</li><li>Particulate contamination</li><li>Dirty sample matrices</li></ul></li></ul><h3>Conclusion</h3>Extending the lifetime of SEC columns requires careful attention to:<ul><li>Sample preparation</li><li>Mobile phase quality</li><li>Operating conditions</li><li>Storage procedures</li><li>Use of protective accessories</li></ul>Implementing these practices helps prevent contamination, degradation, and mechanical damage, ensuring consistent column performance and longer service life.

Extending the Lifetime of Size Exclusion Columns

<h4>Let’s begin with a simple question: what do we actually mean by high throughput?</h4>In the laboratory, high throughput usually refers to the ability to analyze tens, hundreds, or even thousands of samples per day, either across a laboratory workflow or on a single analytical system. Achieving this requires more than just a fast method. It depends on reliable instrumentation, automation, sufficient sample capacity, and smart system design that minimizes idle time between runs.In this article, we look at the key requirements for high-throughput HPLC analysis and how they are addressed in the <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/2371">HTQC system</a>.<h4>Speed</h4>High-throughput HPLC workflows typically rely on short methods. This includes both the chromatographic runtime and the cycle time of the injection module.With the HTQC system, fast gradient applications up to 1200 bar can be implemented using two integrated UHPLC high-pressure gradient pumps.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Fig_1_2_HPG_pumps_of_HTQC_system_left_P_8_1_L_max_pressure_1240_bar_right_P_6_1_L_max_pressure_1000_bar_4d4873ff2f.jpg" alt="KNAUER: Fig. 1–2 HPG pumps of HTQC system, left: P 8.1L (max. pressure 1240 bar), right: P 6.1L (max. pressure 1000 bar)" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Fig_1_2_HPG_pumps_of_HTQC_system_left_P_8_1_L_max_pressure_1240_bar_right_P_6_1_L_max_pressure_1000_bar_4d4873ff2f.jpg 245w," sizes="100vw" width="1266" height="386">We will not discuss the full details of transferring classical HPLC methods to fast, high-resolution UHPLC methods here, but UHPLC implementation can dramatically reduce total analysis time.Let’s use a hypothetical example. Imagine Giorgia, a randomly chosen name, working in pharmaceutical quality control in 1998. She must have travelled back in time, because she would have been too young to work in a lab then, but the example still works. With a classical HPLC method, each sample takes approximately 75 minutes, including equilibration, analysis, and backflushing of a 250 × 4.6 mm ID column. This type of method has been common in QC laboratories around the world. During an eight-hour working day, Giorgia can analyze around six samples.Now let’s transfer the same method to UHPLC. The improvement is substantial. Analysis time, equilibration time, and backflushing time are all reduced to under six minutes per sample. Giorgia can now process around 90 samples in one working day.You may wonder why the HTQC system requires two UHPLC gradient pumps. We will come back to that shortly.The next way to increase speed is to reduce injector cycle time. Injection procedures and washing steps can sometimes take a significant amount of time. This is where the LH 8.1 liquid handler becomes important.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Fig_3_KNAUER_LH_8_1_Liquid_Handler_77ab506ca6.webp" alt="KNAUER: Fig. 3 KNAUER LH 8.1 Liquid Handler" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Fig_3_KNAUER_LH_8_1_Liquid_Handler_77ab506ca6.webp 129w," sizes="100vw" width="800" height="971">The LH 8.1 supports overlapped injections. This means that the next injection can already be prepared while the current analysis is still running. Necessary cleaning steps of the injection system are still maintained, but idle time is reduced.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Fig_4_Schematic_buildup_of_injections_with_and_without_overlapped_injections_af089ba652.webp" alt="KNAUER: Fig. 4 Schematic buildup of injections with and without overlapped injections" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Fig_4_Schematic_buildup_of_injections_with_and_without_overlapped_injections_af089ba652.webp 245w," sizes="100vw" width="1920" height="452">Let’s return to Giorgia. By using overlapped injections in her QC method, she saves about one minute per sample. This increases her capacity to approximately 105 samples per day.<h4>Sample Throughput</h4>A high-throughput system must also be able to handle and store a sufficient number of samples. Here, the LH 8.1 provides another major advantage. In the long-rail version, it can be equipped with up to three robotic coolers.With three coolers, each containing three drawers, the system can hold nine 10 × 13 sample trays. That means a total capacity of 1170 vials. The samples are also temperature controlled.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Fig_5_6_Robotic_Cooler_with_two_10x6_trays_LH_8_1_887_version_with_two_Robotic_Coolers_855b10c2a3.jpg" alt="KNAUER: Fig. 5–6 Robotic Cooler with two 10x6 trays, LH 8.1 (887 version) with two Robotic Coolers" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Fig_5_6_Robotic_Cooler_with_two_10x6_trays_LH_8_1_887_version_with_two_Robotic_Coolers_855b10c2a3.jpg 245w," sizes="100vw" width="1262" height="503">Combined with the low eluent consumption typical of UHPLC, this large sample capacity makes it possible to extend operation from an eight-hour workday to 24-hour operation.For Giorgia and her QC lab, this changes everything again. The same UHPLC method, including overlapped injections, can now run continuously around the clock. This allows approximately 320 samples to be processed per day.Is that enough? Maybe not. As the blog title suggests: we just can’t get enough.<h4>Automation</h4>In addition to a standard 6-port, 2-position injection valve, the HTQC system includes an additional valve. This special column-switching and backflushing valve enables two columns to be operated alternately within a single UHPLC system.While one column is used for analysis, the second column can be backflushed and equilibrated in the reverse flow direction at the same time. This is particularly useful for reducing matrix effects, which are often encountered in QC laboratories when complex sample matrices are analyzed. It also helps reduce overall method runtime.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Fig_7_Exemplary_flow_scheme_of_special_column_switching_valve_d837a71117.webp" alt="KNAUER: Fig. 7 Exemplary flow scheme of special column switching valve" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Fig_7_Exemplary_flow_scheme_of_special_column_switching_valve_d837a71117.webp 127w," sizes="100vw" width="735" height="905"><h4>Parallel Processing</h4>Remember the earlier question: why does the system use two UHPLC gradient pumps?The reason is now clear. The second gradient pump is required to backflush one column while the other column is connected to the analytical flow path through the special switching valve.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Fig_8_Schematic_illustration_of_column_cleaning_equilibration_and_analysis_with_special_backflush_valve_eae2e6f705.webp" alt="KNAUER: Fig. 8 Schematic illustration of column cleaning, equilibration and analysis with special backflush valve" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Fig_8_Schematic_illustration_of_column_cleaning_equilibration_and_analysis_with_special_backflush_valve_eae2e6f705.webp 245w," sizes="100vw" width="1920" height="281">Giorgia immediately ordered a second UHPLC pump and the special valve for her QC lab. She now has a complete HTQC system and is very pleased with the result.Why? Let’s calculate it one final time. With all automation features in place, Giorgia can clean one column while the other column analyzes a sample. This enables her to process an impressive 600 samples per day.Compared with the classical HPLC method she used in 1998, the daily sample throughput has increased from six samples to 600 samples. That is a 100-fold increase.<h4>Summary</h4>We started with a question, so let’s end with one too: what steps increased the throughput?First, the method was scaled down from classical HPLC to a fast UHPLC approach. Then, a second pump and a special column-switching and backflushing valve were added to the UHPLC setup. Together with the <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1454">LH 8.1 liquid handler</a> and its exceptional sample capacity, the HTQC system becomes a flexible and powerful configuration for high-throughput analysis.The system can also be coupled to nearly any detector used in HPLC. Because of its ability to operate at very high speed, it is especially well suited for mass spectrometric methods.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/KNAUER_Fig_9_Complete_flow_scheme_of_HTQC_system_with_highlighted_system_components_edef36cb42.webp" alt="KNAUER: Fig. 9 Complete flow scheme of HTQC system with highlighted system components" srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_KNAUER_Fig_9_Complete_flow_scheme_of_HTQC_system_with_highlighted_system_components_edef36cb42.webp 161w," sizes="100vw" width="800" height="775"><blockquote>For further information on this topic, please contact our authors: kramer@knauer.net, m.myrach@knauer.net</blockquote>

Introducing Our System Solution for High Throughput Quality Control – HTQC

News from LabRulezLCMS Library - Week 19, 2026

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Phenomenex is a global technology leader committed to developing novel analytical chemistry solutions that solve the separation and purification challenges of researchers in industrial, government and academic laboratories.

Phenomenex

Based in Berlin, KNAUER is a medium-sized, owner-managed company that has been serving the sciences since 1962. We develop and manufacture scientific instruments of superior quality for liquid chromatography, including: Analytical HPLC/UHPLC, Preparative HPLC, Fast protein liquid chromatography (FPLC), Multi-column chromatography/Simulated moving bed (SMB) chromatography. High pressure dosing, in-line detection systems, and osmometry are additional business areas.

KNAUER

Determination of nitrosamine content (Bioanalytic)

Our offer includes comprehensive development and validation of methods for determination of nitrosamines in pharmaceutical products. We perform the work according to the criteria and needs of the client based on the EMA note.

Determination of nitrosamines is one of the pharmaceutical industry's top priorities. After the presence of N-nitrosodimethylamine (NDMA) was first reported in an active pharmaceutical ingredient (API), the U.S. Food and Drug Administration (FDA) and the European Medicines Agency (EMA) mandated the identification and control of all APIs and finished products in which there is a risk of nitrosamines.The content of nitrosamines in drug products can be controlled primarily by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) analysis. More than that, a reliable analysis result in our offer consists of a well-chosen sample preparation procedure, a selective and sensitive analytical method, and the experience of the analyst, who selects the best method parameters at the optimization stage.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Determination_of_nitrosamine_content_Bioanalytic_2_ce4f440837.jpg" alt="Bioanalytic: Determination of nitrosamine content." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Determination_of_nitrosamine_content_Bioanalytic_2_ce4f440837.jpg 234w," sizes="100vw" width="1536" height="1024">We have gained experience working with leading pharmaceutical companies. As part of our ongoing work, we have developed analytical methods for small-molecule nitrosamines of 70-150 g/mol. These compounds are mainly formed during manufacturing processes and depend on the presence and type of excipients (e.g. NDBA - N-nitroso-dibutylamine). We have also developed methods for determining molecules with higher nitrosamine drug substance related impurities (NDSRIs). In our work, we focus on achieving the highest possible sensitivity, selectivity and precision. To this end, we individually select the parameters of the methods depending on the composition of the drug and the expected nitrosamines. Due to the above criteria and the required detection limits, we perform nitrosamine impurity determinations using LC-MS/MS kits based on <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?manufacturerItems=52&amp;search=QTRAP+5500%2B">QTRAP 5500+</a> and <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?manufacturerItems=52&amp;search=QTRAP+6500%2B">QTRAP 6500+</a> mass spectrometers (<a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?mainauthorItems=55">SCIEX</a>).<blockquote>Our offerOur offer includes comprehensive development and validation of methods for determination of nitrosamines in pharmaceutical products. We perform the work according to the criteria and needs of the client based on the EMA note.At the stage of sample preparation, we focus on:<ul><li>obtaining the highest possible recovery of nitrosamines (matrix purification);</li><li>limiting additional synthesis of impurities (selection of appropriate equipment and reagents);</li><li>ensuring the stability of the sample prepared for LC-MS/MS analysis.</li></ul>During the optimization of the LC-MS/MS method, we solve such problems as:<ul><li>selection of an appropriate internal standard;</li><li>separation of the impurity from the API;</li><li>obtaining the expected sensitivity of the analytical method.</li></ul>Quantitative analysis of nitrosamines requires sensitivity, precision and accuracy, which we provide to our customers.</blockquote><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Determination_of_nitrosamine_content_Bioanalytic_1_d629e00e62.jpg" alt="Bioanalytic: Determination of nitrosamine content." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Determination_of_nitrosamine_content_Bioanalytic_1_d629e00e62.jpg 103w," sizes="100vw" width="1130" height="1710">

Identification of the proteins (Bioanalytic)

We perform proteomic analyses on a state-of-the-art high-resolution ZenoTOF 7600 (SCIEX), in which the Zeno ion trap and EAD (Electron-Activated Dissociation) technology together make a powerful combination.

Protein identification is based on experimental measurement of peptide molecular weights and matching the amino acid sequence of the protein under study with protein sequences deposited in libraries (databases). Two approaches are most commonly used in proteomic studies: top-down and bottom-up. The top-down method involves analyzing full protein sequences. Bottom-up analyses, on the other hand, use digestion of proteins with proteolytic enzymes. The peptides obtained by digestion are analyzed using a mass spectrometer. Each analysis leads to a comparison of experimental data with data obtained by theoretical digestion of proteins described in the database.<img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Identification_of_the_proteins_Bioanalytic_2_6231b73f0d.jpg" alt="Bioanalytic: Identification of the proteins." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Identification_of_the_proteins_Bioanalytic_2_6231b73f0d.jpg 245w," sizes="100vw" width="1536" height="766"><h4>LC-MS/MS</h4>Tandem mass spectrometers are commonly used to determine protein sequences. They allow ion selection of pseudomolecular peptides and then fragmentation and measurement of the masses of the resulting fragments. Fragmentation using collision induced dissociation(CID ) is an invaluable tool in targeted and untargeted MS/MS analysis of small molecules. It is important to note that in proteomic analysis, data obtained using the aforementioned fragmentation method may leave gaps. For example, spectra obtained by CID fragmentation do not allow unambiguous differentiation of some amino acids: leucine from isoleucine (identical molar masses and CID fragmentation) and lysine from glutamine (very similar masses). For this reason, we perform proteomic analyses on a state-of-the-art high-resolution <a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/library?manufacturerItems=52&amp;search=+ZenoTOF+7600">ZenoTOF 7600</a> (SCIEX), in which the Zeno ion trap and EAD (Electron-Activated Dissociation) technology together make a powerful combination. It enables the creation of unique fragments to differentiate problematic amino acids and increase the sensitivity and specificity of the analysis.<blockquote>We perform analyses of:<ul><li>samples from solution;</li><li>samples from a gel;</li><li>samples from a complex matrix (by prior arrangement);</li><li>samples from lyophilizer;</li><li>samples previously prepared by the client.</li></ul>Qualitative analysis of proteins includes:<ul><li>To confirm the presence of a protein, the sample is subjected to digestion and the result obtained from experimental data is compared with theoretical data;</li><li>Comparison of experimental data on the masses of peptides included in the analyzed protein with the protein sequences described in the database;</li><li>Analysis and identification of the protein mixture;</li><li>Identification of post translational modifications (PTMs) (we use the tools: Skyline, Spectronaut).</li><li>Reporting of the result including, but not limited to, confirmation of the coverage of the protein sequence obtained in the test sample with the theoretical protein and the list of modified peptides, location and % of modifications.</li></ul>Comparative analyses include:<ul><li>Determining the composition of mixtures;</li><li>Comparison and profiling of sample series - protein identification and trend analysis.</li></ul>Our offer also includes:<ul><li>Support in the planning and execution of studies (the scope is determined individually);</li><li>Assistance in the development of extraction methods from complex matrices - biological fluids, plant tissue, animal tissues (the scope is determined individually);</li><li>Statistical analysis using tools: MarkerView (PCA), GraphPad Prism, Trends, Sciex OS;</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1294">Quantitative analysis of proteins</a>;</li><li><a target="_blank" rel="noopener noreferrer" href="https://lcms.labrulez.com/products/1296">Analysis and characterization of monoclonal antibodies</a>.</li></ul></blockquote><img src="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/Bioanalytic_Identification_of_the_proteins_Bioanalytic_1_97cf8c94b3.jpg" alt="Bioanalytic: Identification of the proteins." srcset="https://storage.googleapis.com/labrulez-bucket-strapi-h3hsga3/thumbnail_Bioanalytic_Identification_of_the_proteins_Bioanalytic_1_97cf8c94b3.jpg 117w," sizes="100vw" width="1500" height="2000">

Products from category Pharmaceutical analyses focused on LC/MS - page 1

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News from LabRulezLCMS Library - Week 19, 2026

Introducing Our System Solution for High Throughput Quality Control – HTQC

Extending the Lifetime of Size Exclusion Columns