Technical Interviews - How To Prepare For HPLC Troubleshooting Questions
Often the best cure is to prevent rather than to cure. However when working with a HPLC column, that is not always possible and so I have written out a 14 step process that you can talk about in an interview if you are asked how you troubleshoot this common analytical method in pharmaceuticals. Alongside the actual steps involved, I have included some notes on what soft skills that you are demonstrating by talking about these technical skills. Feel free to link back to these points in your interview if you are asked about those specific soft skills.
Here's a step-by-step guide to troubleshooting in High-Performance Liquid Chromatography (HPLC):
Define the Problem
Clearly identify the specific issue or error encountered in HPLC analysis. This could include poor peak shape, baseline noise, retention time shifts, or low sensitivity, among others. Knowing how to identify a problem can make the solving of that problem much more efficient.
Gather Information
Collect all relevant information related to the problem. This includes the chromatographic conditions (mobile phase composition, flow rate, column type), sample preparation method, instrument parameters (detector settings, column temperature), and any observations or symptoms associated with the issue. You will want to deomonstrate that you are intimately aware of the various different aspects of the machine and how they work together.
Preliminary Checks
Perform some initial checks to ensure that the basic setup is correct. Ensure that all connections are tight, the solvent bottles are properly filled, and the instrument is properly powered on and initialised. Sometimes we forget the basics and it’s all that’s needed to fix the problem and get on with our analysis. Showing this in an interview shows that you are aware of the potential time investment in overdoing your search for a solution to your problem. It will also demonstrate your thoroughness.
Check Chromatographic Conditions
Review the chromatographic conditions and parameters. Verify that the mobile phase composition and concentrations are correct, the flow rate is appropriate, and the column temperature is stable and within the recommended range. This is taking a further step beyond preliminary checks and further demonstrates your technical expertise with HPLC.
Inspect the Column
Examine the column for any physical damage or contamination. If necessary, flush the column with appropriate solvents or consider replacing it. This is a common issue when there is something wrong with a HPLC column and by being conscientious, you are making sure that your testing is done to the highest standard, whether that is to ISO or GMP standards.
Sample Preparation
Evaluate the sample preparation method to ensure it is appropriate for the analytes of interest. Check for possible issues such as incomplete dissolution, inadequate filtration, or improper dilution. This can be a tricky part of the interview as it’s possible that the sample from your client may not be possible to be analysed and this may be needed to communicated to your supervisor and this can lead your interviewer to ask you how you manage client’s expectations.
Detector Issues
If the problem is related to the detector, inspect the detector flow cell for air bubbles, leaks, or contamination. Check the detector settings, such as wavelength, sensitivity, or gain, to ensure they are properly configured.
Troubleshoot Gradient Problems
If using a gradient, verify that the gradient program is properly set up. Check the gradient proportioning valve, mixing efficiency, and pump pressure to ensure consistent solvent delivery.
Column Equilibration
Allow sufficient time for column equilibration before running samples. This helps ensure consistent baseline and reproducible results.
Systematic Parameter Adjustments
Make small adjustments to various parameters one at a time to isolate the issue. For example, modify the flow rate, change the mobile phase pH, or adjust the column temperature. Observe the impact of each adjustment on the chromatography and record the results. This will show again that you take your role seriously and that you are looking to rule out as much human error as possible.
Data Analysis
Analyse the chromatograms obtained during troubleshooting. Look for any patterns, changes, or anomalies that could help identify the root cause of the problem. Compare the obtained data with previous or expected results to identify any discrepancies. This will show that you are always looking to improve your abilities and are responsible for your own learning, showing your ambition and accountability.
Implement Corrective Actions
Based on the analysis of the troubleshooting data, select appropriate corrective actions. This may involve modifying instrument settings, optimizing chromatographic conditions, replacing or cleaning components, or adjusting sample preparation techniques.
Verify and Validate
After implementing the corrective actions, verify the effectiveness by running control samples or reference standards. Ensure that the problem has been resolved, and the chromatographic results are satisfactory. Document the troubleshooting process, including the identified problem, steps taken, and the outcome. This not only logs the work that you have done but also leaves a step by step process for you and your team if the same issue arises again.
Preventive Measures
Take preventive measures to minimize the recurrence of similar issues in the future. This may include routine maintenance of the HPLC system, regular column conditioning, using appropriate quality control checks, or updating standard operating procedures.
Remember, troubleshooting in HPLC requires a systematic approach and patience. It is essential to document each step of the troubleshooting process for future reference and to ensure reliable and consistent chromatographic results.
If you are someone who is recently graduated and wondering how to best talk about technical skills in a CV/Resume when most of your experience, make sure to check out my article how to do just that here.
