Chromatography Theory Chapter 1 – Chromatography Peak Theory

This chapter introduces the common measurements and definitions used to describe chromatography peak theory. It provides the foundation for later topics such as data acquisition, integration, smoothing, and calibration.

Common Peak Measurements

• Start and end times: where the baseline meets the chromatogram or droplines.

• Peak area: area between the chromatogram and baseline.

• Peak height: greatest vertical distance between chromatogram and baseline.

• Retention time (RT): time at the peak’s apex.

Peak Width Measurements

• Typically measured at half height using horizontal lines, even when baselines slope.

• Front width and back width are defined by where the half-height line intersects the chromatogram.

• Width can also be measured at other fractions (e.g., 10%).

• In clusters, width estimation is less straightforward; sometimes area/height ratio is used instead.

Peak Shape Measurements

Chromatographers use peak shape to assess column performance. Real peaks deviate from the ideal Gaussian symmetrical shape.

Skew

• Ratio of back width (b) to front width (f).

• Symmetrical peak: skew = 1.0.

• Tail = skew > 1.0.

• Rarely, skew < 1.0 → “reverse tailing.”

Tailing Factor (TF)

• TF = (f + b) ÷ (2f).

• Related to skew: TF = 1 + skew ÷ 2.

• Symmetrical peak: TF = 1.0.

• TF always ≥ 0.5; >1.0 indicates tailing.

Kurtosis (Gaussian Factor)

• K = 1.83 × width@50% ÷ width@10%.

• Gaussian peak: K = 1.0.

• Wide near top: K > 1.0.

• Wide near baseline: K < 1.0.

Peak Resolution

Resolution compares separation of two peaks relative to their widths:

R = 2 × (t₂ – t₁) ÷ (1.7 × (w₂ + w₁))

• R > 2.0 = good separation.

• R < 1.5 = marginal separation.

Theoretical Plates (Column Efficiency)

N = 5.5 × (t ÷ w²)

• Higher N = better efficiency.

• Chromperfect uses width-at-half-height × 1.7 for baseline width, aligning with post-1996 USP standards.

Topic chromatography peak theory