Today, drug development studies are in increasing need of analytical instruments to measure target analytes in complex study matrices. Besides, these analytical tools should be able to analyze multiple study samples as rapidly as possible while complying with data quality requirements.
Combining liquid chromatography (HPLC) with mass spectrometry (MS) offers scientists a powerful bioanalytical tool to meet all stringent requirements. As liquid chromatography-mass spectrometry (LC-MS) assays are highly efficient and versatile, they are increasingly employed in numerous modern bioanalytical laboratories.
LC-MS testing involves the separation of target analytes followed by detection based on their mass-to-charge ratio. Although LC-MS services are relatively new compared to traditional analytical tools, their selectivity, accuracy, and sensitivity have made it a primary choice of detection technique for numerous analytes such as drug metabolites, food adulterants, pesticides, and natural product extracts. However, with any advanced technology, scientists face multiple challenges while setting up LC-MS testing for complex study matrices. Hence, the current article shares strategies and tips for overcoming some of these challenges.
Strategies and tips for LC-MS testing
LC-MS assays are highly suitable for analyzing polar and nonpolar compounds. These compounds range from very high molecular mass proteins to low molecular mass analytes. Coupling LC-MS assays with fragmentation spectra can help elucidate the structure of the target analytes.
However, LC-MS assays are expensive to operate, own and maintain. Besides, bioanalytical laboratories require expert professionals to run LC-MS assays and analyze the data. Moreover, LC-MS testing depends on the spectra obtained. These spectra, in turn, depend on experimental conditions, and hence, identifying data for comparison to a reference standard is limited.
Scientists must take adequate care when handling samples as mass spectrometers are a demolishing detecting system. Besides being a laboratory-based system, LC-MS testing can prove challenging to analyze reactive or unstable study samples.
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LC-MS assays can only analyze liquid samples. Hence, one must extract the analyte from the sample or dissolve it in a suitable solvent. Besides, sample preparation techniques, such as liquid-liquid or solid-phase extraction, are critical to extracting target analytes from the study material. This intermediate is essential to improve assay sensitivity and reduce system contamination. Contaminants such as metal ions and polyethylene glycol can interfere with LC-MS analysis. Some approaches to minimize these contaminants include:
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Using high-purity water solvent and reagents in mobile phases
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Using freshly prepared mobile phase
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Avoiding the use of detergent to clean glassware
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Using high-purity gasses
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Maintaining nitrogen generators and gas cylinders
Biological sample assessments can include the enhancement or separation of ionization due to other sample constituents. Hence, one must isolate the analyte of interest to minimize matrix effects. This measure makes the sample preparation step crucial for LC-MS testing. However, adequate care is necessary to avoid co-elution of interfering analytes.
Biological samples such as DNA and proteins may get lost due to non-specific binding. Adsorptions of target analytes with laboratory consumables, for example, microcentrifuge tubes, impact the precision and accuracy of LC-MS assays. Hence, researchers must use containers with low surface adsorption to minimize analyte loss. Besides, approaches such as adding blocking agents can help reduce the interaction of target analytes with containers.