1998-5. Sample Overload — Friend or Foe?

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1998-5. Sample Overload — Friend or Foe?

May 1, 1998

Sample overload can be frustrating, but it also can be used to your advantage

LCGC 16(5) 442-446 (1998)

Nan S. Wilson, John R. Kern, and John W. Dolan

Tags: chiral, column types, detector problems, overload, related substances, sample overload, system suitability

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2022-5. Dealing with Wandering First-Dimension Peaks in 2D-LC Separations

2022-4. Essentials of LC Troubleshooting, Part II: Misbehaving Retention Times

2022-3. But Why Doesn’t It Get Better? Kinetic Plots for Liquid Chromatography, Part III: Pulling It All Together

2022-2. But Why Doesn’t It Get Better? Kinetic Plots for Liquid Chromatography, Part II: Making and Interpreting the Plots

2022-1. But Why Doesn’t It Get Better? Kinetic Plots for Liquid Chromatography, Part I: Basic Concepts

2021-12. Essentials of LC Troubleshooting, Part I: Pressure Problems

2021-11. But My Peaks Are Not Gaussian! Part III: Physicochemical Causes of Peak Tailing

2021-10. Retention Factor Is Independent of Pressure in LC, Right?

2021-9. But My Peaks Are Not Gaussian! Part II: Physical Causes of Peak Asymmetry

2021-8. But My Peaks Are Not Gaussian! Part I: Basic Concepts in Peak Shape, and the Effect of Extracolumn Connections

2021-7. Where Has My Efficiency Gone? Impacts of Extracolumn Peak Broadening on Performance, Part IV: Gradient Elution, Flow Splitting, and a Holistic View

2021-6. Where Has My Efficiency Gone? Impacts of Extracolumn Peak Broadening on Performance, Part III: Tubing and Detectors

2021-5. Where Has My Efficiency Gone? Impacts of Extracolumn Peak Broadening on Performance, Part II: Sample Injection

2021-4. Where Has My Efficiency Gone? Impacts of Extracolumn Peak Broadening on Performance, Part I: Basic Concepts

2021-3. Surfing on Mobile Phase, Part II: Impact of Mobile-Phase Composition Waves on Retention in LC

2021-2. Surfing on Mobile Phase, Part I: Origins of Mobile-Phase Composition Waves and Their Effects on Detector Baselines

2021-1. LC Troubleshooting Odds and Ends from 2020

2020-12. Tips for Liquid Chromatography Coupled with Charged Aerosol Detection

2020-11. Troubleshooting for Two-Dimensional Liquid Chromatography: Breakthrough in the Second Dimension

2020-10. Troubleshooting 101: Some Essential Principles of Effective Troubleshooting

2020-9. Rules of Thumb for Reversed-Phase LC: What’s In Your Chromatographic Mind?

2020-8. Troubleshooting LC Separations of Biomolecules, Part II: Passivation and Mobile-Phase Additives

2020-7. Recovering from a COVID-19 Shutdown: Tips and Tricks for Starting Up, Part II

2020-6. Recovering from a COVID-19 Shutdown: Tips and Tricks for Starting Up, Part I

2020-5. Liquid Chromatography at Room Temperature: When Should the Column Temperature Be Specified in a Method?

2020-4. Selectivity in Reversed-Phase Liquid Chromatography: 20 Years of the Hydrophobic Subtraction Model

2020-3. Troubleshooting LC Separations of Biomolecules, Part I: Background, and the Meaning of Inertness

2020-2. Effects of Buffer Capacity in LC, Part II: Visualization of pH Changes Inside the Column

2020-1. Effects of Buffer Capacity in Reversed-Phase Liquid Chromatography, Part I: Relationship Between the Sample- and Mobile-Phase Buffers

2019-12. Effects of Flow Rate on UV Detection in Liquid Chromatography

2019-11. Column Re-equilibration Following Gradient Elution: How Long is Long Enough? Part I: Reversed-phase and HILIC Separations of Small Molecules

2019-10. Minimizing Fluctuating Peptide Retention in 2D-LC: How to Address a Moving Target

2019-9. Mind the Diluent: Effects of Sample Diluent on Analyte Recovery in Reversed-Phase and HILIC Separations

2019-8. Pass the Salt: Evolution of Coupling Ion-Exchange Separations and Mass Spectrometry

2019-7. Mobile Phase Buffers in LC: Effect of Buffer Preparation Method on Retention Repeatability

2019-6. Mixing and Mixers in Liquid Chromatography, Part III: Solutions for Problems with Sample Diluents

2019-5. Methods from Mars? Coping with Chromatographic Legacies

2019-4. Reversed-Phase Liquid Chromatography and Water, Part II: Re-equilibration of the Stationary Phase Following Gradient Elution

2019-3. Looking for New Selectivity in Reversed-Phase Liquid Chromatography? A View of the Selectivity Landscape Over the Past Two Years

2019-2. Reversed-Phase Liquid Chromatography and Water, Part I: How Much is Too Much?

2019-1. What’s Trending in LC Troubleshooting?

2018-12. Letting the Chromatograph Talk, Part I: Looking for Troubleshooting Clues in Unexpected Places

2018-11. Mixing and Mixers in Liquid Chromatography—Why, When, and How Much? Part II, Injections

2018-10. Mixing and Mixers in Liquid Chromatography—Why, When, and How Much? Part I, The Pump

2018-9. Tips, Tricks, and Troubleshooting for Separations of Biomolecules, Part II: Contemporary Separations of Proteins by Size-Exclusion Chromatography

2018-8. Contaminants Everywhere! Tips and Tricks for Reducing Background Signals When Using LC–MS

2018-7. Tips, Tricks, and Troubleshooting for Separations of Biomolecules, Part I: Contemporary Reversed-Phase Protein Separations

2018-6. Peak Purity in Liquid Chromatography, Part III: Using Two-Dimensional Liquid Chromatography

2018-5. Fittings and Connections for Liquid Chromatography—So Many Choices!

2018-4. Peak Purity in Liquid Chromatography, Part II: Potential of Curve Resolution Techniques

2018-3. Eluent Preparation for Hydrophilic Interaction Liquid Chromatography, Part II: pH, Buffers, and Gradient Elution

2018-2. Peak Purity in Liquid Chromatography, Part I: Basic Concepts, Commercial Software, and Limitations

2018-1. Eluent Preparation for Hydrophilic Interaction Liquid Chromatography, Part I: Solvent Volumes and Buffer Counterions

2017-12. Resources for LC Practitioners in 2017: What’s on Your Bookshelf and in Your Web Browser?

2017-11s. The Best of LC Troubleshooting: A Farewell to John Dolan

2017-10a. 34 Years of LC Troubleshooting: An Interview with John Dolan

2017-10. All’s Well That Ends Well

2017-9. How to Take Maximum Advantage of Column-Type Selectivity

2017-8. Method Adjustment for Gradient Elution

2017-7. Column Care for the Long Haul — Considerations for Column Storage

2017-6. Method Adjustment the USP Way

2017-5. Count the Cost, Part III: Increasing Resolution by Changing Selectivity

2017-4. Count the Cost, Part II: Increasing Resolution by Increasing Retention

2017-3. Count the Cost, Part I: Increasing Resolution by Increasing Column Efficiency

2017-2. Filters and Filtration in Liquid Chromatography — What to Do

2017-1. Back to Basics: The Role of pH in Retention and Selectivity

2016-12. Failed System Suitability Test: A Case Study

2016-11. Reader’s Question: Early Eluted Peak

2016-10. Reader’s Questions: Gradient Ghost Peaks

2016-9. How Does It Work? Part V: Fluorescence Detectors

2016-8. How Does It Work? Part IV: Ultraviolet Detectors

2016-7. How Does It Work? Part III: Autosamplers

2016-6. How Does It Work? Part II: Mixing and Degassing

2016-5. How Does It Work? Part I: Pumps

2016-4. Retention Time Drift — A Case Study

2016-3. Column Plate Number and System Suitability

2016-2. Detective Work, Part IV: Chemical Problems with the Column — Chemical Attack

2016-1. Detective Work, Part III: Strong Retention and Chemical Problems with the Column

2015-12. Detective Work, Part II: Physical Problems with the Column

2015-11. Detective Work, Part I: Simplify the Choices

2015-10. How to Reduce Mobile-Phase Consumption

2015-9. LC Column Problems Everywhere

2015-8. Overload in Liquid Chromatography

2015-7. Distorted Peaks — A Case Study

2015-6. Internal Standard Calibration Problems

2015-5. Calibration Problems — A Case Study

2015-4. Be Careful of the Flow Rate

2015-3. Listen to the Data

2015-2. Slow Column Equilibration

2015-1. Seven Things to Avoid in the Liquid Chromatography Laboratory

2014-12. Column Protection: Three Easy Steps

2014-11. How Much Can I Inject? Part II: Injecting in Solvents Other than Mobile Phase

2014-10. How Much Can I Inject? Part I: Injecting in Mobile Phase

2014-9. Estimating Resolution for Marginally Separated Peaks

2014-8. How Much Retention Time Variation is Normal?

2014-7. Mobile-Phase Degassing: What, Why, and How

2014-6. UV Detector Problems

2014-5. Readers’ Questions: Calibration

2014-4. Problems with Large-Molecule Separations

2014-3. LC Method Scaling, Part II: Gradient Separations

2014-2. LC Method Scaling, Part I: Isocratic Separations

2014-1. Column Dead Time as a Diagnostic Tool

2013-12. Peak Fronting . . . Some of the Time

2013-11. What’s the Problem with the LLOQ – A Case Study

2013-10. Past, Present, and Future – 30 Years of LC Troubleshooting

2013-9. Variable Retention Times – A Case Study

2013-8. Gradient Elution, Part VI: Ghost Peaks

2013-7. Gradient Elution, Part V: Baseline Drift Problems

2013-6. Gradient Elution, Part IV: Dwell-Volume Problems

2013-5. Gradient Elution, Part III: Surprises

2013-4. Gradient Elution, Part II: Equivalent Separations

2013-3. Gradient Elution, Part I: Intuition

2013-2. The Importance of the Sample Matrix

2013-1. Making the Most of a Gradient Scouting Run

2012-12. Avoiding Refractive Index Detector Problems

2012-11. Evaluation of an LC Method

2012-10. The Role of the Injection Solvent

2012-9. Reader’s Questions

2012-8. Questions from Afar

2012-7. Troubleshooting Basics, Part IV: Peak Shape Problems

2012-6. When Should an Internal Standard Be Used?

2012-5. Extra Chromatographic Peaks – A Case Study

2012-4. Retention Time Changes

2012-3. Why Does an Improvement Make Things Worse?

2012-2. A Case of Sporadic LC Assay Results

2012-1. LC Trouble Everywhere

2011-12. Troubleshooting Basics, Part III: Retention Problems

2011-11. Locating Precision Problems

2011-10. Column Triage

2011-9. Troubleshooting Basics, Part II: Pressure Problems

2011-8. How Fast Can a Gradient Be Run?

2011-7. Troubleshooting Basics, Part I: Where to Start?

2011-6. The Case of the Too Big Little Peak

2011-5. Ghost Peak Investigation in a Reversed-Phase Gradient LC System

2011-4. Selectivity in Reversed-Phase LC Separations, Part IV: Pressure Selectivity

2011-3. Selectivity in Reversed-Phase LC Separations, Part III: Column-Type Selectivity

2011-2. A Picture is Worth a Thousand Words

2011-1. Selectivity in Reversed-Phase LC Separations, Part II: Solvent-Strength Selectivity

2010-12. Selectivity in Reversed-Phase LC Separations, Part I: Solvent-Type Selectivity

2010-11. UHPLC Tips and Techniques

2010-10. Are the Method Requirements Reasonable?

2010-9. Speeding Up Gradients

2010-8. System Contamination

2010-7. Where Did That Peak Come From?

2010-6. 300 and Counting

2010-5. Too Much Scrutiny

2010-4. Getting the Most Out of Calibration Standards

2010-3. Enhancing Signal-to-Noise

2010-2. Column Diameter, Linear Velocity, and Column Efficiency

2010-1. Too Little or Too Much

2009-12. LC Troubleshooting Strategies

2009-11. Kinetic Plots Made Easy

2009-10. Integration Problems

2009-9. Method or System?

2009-8. Too Many Changes

2009-7. Calibration Curves, Part V: Curve Weighting

2009-6. Calibration Curves, Part IV: Choosing the Appropriate Model

2009-5. Calibration Curves, Part III: A Different View

2009-4. Calibration Curves, Part II: What are the Limits?

2009-3. Calibration Curves, Part I: To b or Not to b?

2009-2. Testing Method Performance

2009-1. Problems from East and West

2008-12. LC Pumps

2008-11. Extracolumn Effects

2008-10. Drip, Drip, Drip…

2008-9. Odds and Ends from the Inbox

2008-8. Column Care

2008-7. Peak Shape Problems

2008-6. Check Valves and Acetonitrile

2008-5. How Do I …?

2008-4. Only Three Things

2008-3. Method Transfer Problems

2008-2. Ion Pairing — Blessing or Curse

2008-1. Flow-Rate Adjustment and System Suitability

2007-12. The Perfect Method, Part VII: The Gradient Shortcut

2007-11. The Perfect Method, Part VI: Make It Faster

2007-10. The Perfect Method, Part V: Changing Column Selectivity

2007-9. The Perfect Method, Part IV: Controlling Peak Spacing

2007-8. The Perfect Method, Part III: Adjusting Retention

2007-7. The Perfect Method, Part II: Where to Start?

2007-6. The Perfect Method, Part I: What is Your Goal?

2007-5. How Does Temperature Affect Selectivity?

2007-4. Variability — How to Control It

2007-3. Solvent Conservation

2007-2. How Little Can I See?

2007-1. Questions From India

2006-12. Method Reproducibility — A Case Study

2006-11. On-Column Sample Degradation

2006-10. Making Connections

2006-9. What’s Happening to My Column?

2006-8. Autosampler Carryover

2006-7. Gradient Performance Problems — A Case Study

2006-6. The Power of Mobile Phase Strength

2006-5. Dwell Volume Revisited

2006-4. It’s Gonna Break

2006-3. Chromatography Forum — Use the Braintrust

2006-2. Check Valve Problems

2006-1. Obtaining More Consistent Results

2005-12. The Role of Signal-to-Noise Ratio in Precision and Accuracy

2005-11. Extending Column Life

2005-10. LC Troubleshooting: The Basics

2005-9. A Baker’s Dozen

2005-8. Broad Peaks

2005-7. Reader Questions: Mobile Phase

2005-6. Peak Fronting, Column Life, and Column Conditioning

2005-5. Overload or Minor Peak?

2005-4. The Bermuda Triangle

2005-3. Split Peaks — A Case Study

2005-2. Extracolumn Effects

2005-1. Stay Away from the Cliffs!

2004-12. Retention Time Changes

2004-11. My Favorite Shortcuts

2004-10. Gradient Performance Checks

2004-9. Grandma’s Ham

2004-7. Divide and Conquer

2004-6. But I Can’t Change the Method

2004-5. System Suitability

2004-4. Parallel Chromatography — Double Your Money

2004-3. Success with Evaporative Light-Scattering Detection, Part II: Tips and Techniques

2004-2. Selecting the Best Curve Fit

2004-1. Broad Peaks

2003-12. Understanding Split Peaks

2003-11. Extracolumn Effects

2003-10. How Much Is Enough?

2003-9. Honoring Readers

2003-7. Why Do Peaks Tail?

2003-6. Now What’s Going On?

2003-5. Not Another Leak!

2003-4. Flow Rate and Peak Spacing

2003-3. LC Columns — The Top-10 List

2003-2. Success with Evaporative Light-Scattering Detection

2003-1. Mobile-Phase Buffers, Part III — Preparation of Buffers

2002-12. Mobile-Phase Buffers, Part II — Buffer Selection and Capacity

2002-11. Mobile-Phase Buffers, Part I — The Interpretation of pH in Partially Aqueous Mobile Phases

2002-10. The Hazards of Adjusting Gradients

2002-9. Performance Qualification of LC Systems

2002-7. Resolving Minor Peaks

2002-6. The Importance of Temperature

2002-5. Peak Tailing and Resolution

2002-4. Stability-Indicating Assays

2002-3. The Future of LC Troubleshooting — 10 Years Later

2002-2. Quiet, Please

2002-1. Ion Suppression in LC-MS/MS — A Case Study

2001-12. From the e-Mail Bag

2001-11. Retaining Polar Compounds

2001-10. Attacking Carryover Problems

2001-9. When Ideal Isn’t Practical

2001-7. Communicating with the Baseline

2001-6. Buffers and Baselines

2001-5. Autosamplers, Part II — Problems and Solutions

2001-4. Autosamplers, Part I — Design Features

2001-3. Reproducibility and Carryover — A Case Study

2001-2. Autosampler Carryover

2001-1. Unexpected Peaks

2000-12. Reducing Column Diameter in Gradient Elution — A Case Study

2000-11. The Problem of Late Elution — A Case Study

2000-10. Scaling Gradient LC Methods to LC-MS

2000-9. Reversed-Phase in 100% Water

2000-7. Stainless Steel Surfaces in LC Systems, Part II — Passivation and Practical Recommendations

2000-6. Stainless Steel Surfaces in LC Systems, Part I — Corrosion and Erosion

2000-5. Starting Out Right, Part VI — The Scouting Gradient Alternative

2000-4. Starting Out Right, Part V — Changing Column Conditions

2000-3. Starting Out right, Part IV — Additional Variables to Control Selectivity

2000-2. Starting Out Right, Part III — The Role of the Solvent in Controlling Selectivity

2000-1. Starting Out Right, Part II — Measuring Satisfaction

1999-12. Starting Out Right, Part I — Selecting the Tools

1999-11. Reader’s Questions: Carryover, Mobile-Phase Temperature, and Column Care

1999-10. Mobile-Phase Degassing — Why, When, and How

1999-9. Poor Peak Shape: A Case Study

1999-7. Priming Injections

1999-6. How Much Is Too Much?

1999-5. LC Troubleshooting On-Line with Chromatography Forum

1999-4. Lessons in Column Washing

1999-3. Late Elution and Carryover Peaks — A Case Study

1999-2. Basic Compounds — Starting on the Right Foot

1999-1.UV Detector Noise

1998-12. Column Temperature Effects in Gradient Elution

1998-11. Gradient Background Peaks — A Case Study