Guidelines for Troubleshooting Chromatographic Peak Loss in Liquid Chromatography and LC-MS Systems

Introduction: The Prevalence of Peak Loss Phenomenon and Troubleshooting Approaches

In the analysis work of liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS), peak loss is one of the most troubling issues for analysts. This phenomenon can be caused by various factors, including instrument hardware failures, improper method parameter settings, errors in mobile phase preparation, sample handling problems, etc. When faced with sudden disappearance of chromatographic peaks, analysts often find themselves at a loss on how to proceed. This article systematically reviews various factors that may lead to peak loss and provides detailed troubleshooting steps and solutions to help analysts quickly identify the root cause.

The investigation into peak loss should follow the principle from simple to complex, from surface symptoms to underlying causes. It is recommended first to check the instrument pressure curve as this is the most intuitive indicator before sequentially investigating key components such as sampling systems, chromatographic columns, and detectors. During troubleshooting, it is crucial to distinguish between systemic failures (e.g., no peaks across all samples) or individual sample issues (e.g., only specific samples not producing peaks), which will significantly narrow down the scope of investigation. We will begin with an analysis of abnormal pressure followed by various fault diagnosis methods.

1. Analysis of Abnormal Pressure Curves and Fault Diagnosis

Pressure monitoring is a primary indicator for determining whether a liquid chromatography system operates normally. Modern chromatographic data systems typically have real-time pressure monitoring capabilities; comparing normal samples with abnormal ones often reveals clues about potential issues based on their pressure curves' anomalies.

1.1 Overall Decrease in Pressure Curve but Normal Fluctuation When observing an overall decrease in pressure while maintaining normal fluctuation patterns, this usually indicates a leakage issue within the system. Leaks may occur at points such as injectors, tubing connections or pump heads; among these areas injector leaks are common yet difficult for instruments to automatically detect since modern liquid systems primarily focus leak detection on high-pressure sections while injectors generally operate under low-pressure conditions. To verify if there’s leakage at the injector level can be determined by comparing main pass pressures against bypass pressures during operation mode where flow must traverse more components like quantitation loops or injection needles theoretically leading higher system pressures than those observed under bypass modes: First adjust system pressures approximately 50 bar above regular operating levels ensuring not exceeding maximum tolerances then switch states via injection valves—if main pass shows significant drops below bypass values confirm presence leakage problem present within injectors; injector leaks commonly arise due wear seals around needle seats six-way valve rotors seal failure loosened connections along quantitation rings requiring replacement parts resolution experienced operators may refer maintenance manuals perform replacements otherwise contacting professional engineers advisable following proper disassembly order prior re-calibrating positions ensuring accurate alignment post-replacement completion thereafter checking results stability after adjustments made appropriately... ... [Content continues detailing further diagnostic procedures related specifically towards resolving identified faults through structured methodologies addressing both mechanical integrity aspects involved operational efficiency concerns pertinent testing standards applied accordingly throughout processes outlined comprehensively.]