How To Decide Tailing Factor Limit In HPLC And GC Analysis: Get Mastery In 3 Minutes

Understanding the Role of Tailing Factor in HPLC and GC: A Practical Perspective

The Tailing Factor is a critical parameter in chromatographic analysis, particularly in High-Performance Liquid Chromatography (HPLC) and Gas Chromatography (GC). It plays a vital role in system suitability testing by providing insight into the symmetry of chromatographic peaks, which directly affects both quantitative accuracy and resolution.

In this article, I’ll share practical, experience-based insights into the concept of the tailing factor – its calculation, significance in method development, and the various factors that influence it. By the end, you will have a clear understanding of:

• What the tailing factor is and how it is calculated
• Its importance and application in chromatographic method development
• Factors that contribute to peak tailing
• Industry-accepted criteria for evaluating peak tailing

Whether you’re a student, analyst, or method developer, this guide will help you better assess chromatographic performance and make informed decisions during method optimisation.

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Tailing Factor

The Tailing factor is defined as the distance from the front edge of the peak to the back edge, divided by the distance from the front edge to the centre line with all distances measured at 5% ( or 1/20) of the maximum peak height (see figure-1). It is also called USP tailing and it is denoted by T.

Figure-1

Where: X is the front half width, and Y is the back half width at 5% of the peak height

Tailing Factor Calculation Formula:

Fronting and Tailing in Peak

Note: In the above chromatogram, peak-2 is symmetrical, whereas peak-1 is deformed from the front side, and peak-3 is deformed from the tail side. Peak-1 is called the ideal peak, peak-2 is called the fronting peak and peak-3 is called the tailing peak.

The Fitting and Tailing tells about the symmetry /shape of the peak. Both peak shapes can negatively impact quantitation accuracy and resolution, and they are critical indicators in system suitability testing

Tailing Peak:

A tailing peak in chromatography refers to a peak where the descending part (the “tail”) extends more slowly than the ascending part. It causes asymmetry, making the peak skewed to the right.

• Cause: Often due to secondary interactions between analyte and stationary phase (e.g., residual silanol groups), column overloading, or poor column packing.
• Tailing Factor (TF): Used to quantify this effect; a value >1.0 indicates tailing.

Fronting Peak:

A fronting peak occurs when the ascending part of the peak is broader than the descending part, creating a skewed peak shape to the left.

• Cause: Commonly results from column overloading (too much sample) or instrumental issues like detector saturation or poor injection technique.
• Fronting Indicator: Tailing factor <0.9 may suggest fronting

Tailing Factor Limit: Acceptance Criteria

Practically, it is impossible to get a symmetrical peak in chromatographic analysis. Following are the acceptance criteria for the tailing factor:

• Peak with T ≤1.5 is preferable and it is accepted by all regulatory agencies
• But for typical molecules like T ≥ 2 is also acceptable by regulatory agencies

Factors affecting the tailing factor

• Structure of the molecule: Generally basic molecules show more tailing
• Shape of the peak: Broad peaks have more tailing compared to sharp peaks.
• pH of the mobile phase: Some of the molecules are highly pH sensitive and beyond beyond that pH shows tailing in the peak
• Buffer concentration: Tailing is directly proportional to the buffer concentration of the mobile phase.
• Solvents used in the mobile phase; Some of the molecules are highly solvent selective and show taling if other solvents are used. Generally, the mobile phase with Acetonitrile solvent peaks with no tailing or less tailing compared to the mobile phase with methanol.
• Column temperature: Tailing factors is directly proportional to the column temperature
• Elution time: T is inversely proportional to the peak elution time
• Column efficiency: T is directly proportional to the column efficiency
• Injection volume:T is inversely proportional to the peak injection volume
• Nature of the diluent:T also depends upon the nature of the diluent

Case Study: Addressing High Tailing Factor in HPLC Analysis of an API

Background:

A pharmaceutical lab was developing an HPLC method to quantify Ibuprofen in tablet formulations. During validation, the tailing factor for the ibuprofen peak consistently exceeded 2.0, failing the system suitability criteria (T ≤ 1.5).

Initial Method Conditions:

• Column: C18, 150 mm × 4.6 mm, 5 µm
• Mobile Phase: Water:Acetonitrile (60:40, v/v), pH 3.0 (adjusted with phosphoric acid)
• Flow Rate: 1.1 mL/min
• Injection Volume: 20 µL
• Detection: UV at 225 nm

Root Cause Investigation:

1. Column Aging: A fresh column slightly reduced tailing (to ~1.9), suggesting minor column degradation.
2. Analyte-Stationary Phase Interaction: Ibuprofen (a weak acid) possibly interacting with residual silanol groups on the stationary phase.
3. Mobile Phase pH: At pH 3.0, ibuprofen is partially ionized, increasing its interaction with residual silanol groups, leading to tailing.
4. Injection Solvent Mismatch: The sample was dissolved in a higher-strength solvent (100% acetonitrile), causing peak distortion.

Corrective Actions Taken:

• Changed pH of the aqueous phase to pH 2.5, where ibuprofen is mostly non-ionized.
• Switched to an end-capped C18 column to reduce silanol interactions.
• Adjusted sample solvent to match the mobile phase composition (60:40 Water:ACN).
• Reduced injection volume to 10 µL to minimise overload.

Results Summary:

Conclusion:

The high tailing factor was primarily due to silanol interaction and pH mismatch. Through pH adjustment, column selection, and proper sample preparation, the tailing factor was reduced to within acceptable limits, ensuring regulatory compliance and improved method robustness.

Conclusion

Maintaining a tailing factor (T ≤ 1.5) is an important part of system suitability criteria during chromatographic method development, ensuring accurate, precise, and regulatory-compliant results. If a higher tailing factor is observed, it should be supported with a scientifically sound justification to avoid potential regulatory concerns.

This concludes the overview of the tailing factor’s role in HPLC and GC. If you have any questions or need further clarification, feel free to leave a comment – I’ll respond promptly and prioritise your queries.

You may also want to check out other articles on my blog, such as:

• What is the Analytical Method Validation?
• The 7 Key Differences Between Validation and Verification

FAQs

More Questions Related to T

What is the symmetry factor in HPLC?

What can be used to reduce tailing in gas chromatography?

What causes tailing in chromatography?

What is the tailing peaks gc?

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What is usp tailing factor acceptance criteria?

Abbreviation

• T: Tailing factor

References

• Pharmaceutical analysis by James. W. Munson., page-109
• Thai Pharmacopeia