HPLC calculator v3.1
Developed by Davy GUILLARME [davy.guillarme@unige.ch]
With contribution of Dao NGUYEN, Serge RUDAZ, Jean-Luc VEUTHEY
Group of analytical sciences, School of Pharmaceutical Sciences,
University of Geneva, CMU building, Rue Michel Servet 1, 1211 Geneva 4, Switzerland
Method transfer in isocratic mode
Original method (1)
Original column geometry
Original exp. Conditions
Column length (L1) [mm]
Column diameter (dc1) [mm]
Particle size (dp1) [µm]
Flow-rate (F1) [µL/min]
Injection volume (Vinj1) [µL]
Analysis time [min]
Transferred method (2)
Transferred column geometry
Transferred exp. Conditions
Column length (L2) [mm]
Column diameter (dc2) [mm]
Particle size (dp2) [µm]
Flow-rate (F2) [µL/min]
—
Injection volume (Vinj2) [µL]
—
Analysis time [min]
—
Performance of transferred method
Efficiency
Pressure
Analysis time
Solvent consumption
—
—
—
—
Equations used (from Guillarme & Veuthey):
• F₂ = F₁ × (dc₂²/dc₁²) × (dp₁/dp₂) — Flow rate scaling (maintain u·dp constant)
• Vinj₂ = Vinj₁ × (dc₂²·L₂)/(dc₁²·L₁) — Injection volume proportional to column volume
• t₂ = t₁ × (F₁/F₂) × (dc₂²/dc₁²) × (L₂/L₁) — Analysis time scaling
• ΔP₂/ΔP₁ = (L₂/L₁) × (dp₁³/dp₂³) × (F₂/F₁) × (dc₁²/dc₂²) — Pressure (Darcy's law)
• V₂/V₁ = (F₂·t₂)/(F₁·t₁) — Solvent consumption
Developed by Davy GUILLARME [davy.guillarme@unige.ch] — With contribution of Dao NGUYEN, Serge RUDAZ, Jean-Luc VEUTHEY
Group of analytical sciences, School of Pharmaceutical Sciences, University of Geneva, Switzerland
Method transfer in gradient mode
Original method (1)
Original column geometry
Original exp. Conditions
L1 [mm]
dc1 [mm]
dp1 [µm]
Dwell vol. Vd1 [mL]
F1 [µL/min]
Vinj1 [µL]
Additional gradient information
Suggested reconditioning step [min]
—
Original gradient profile
Performance of transferred method
Optimal flow rate
Flow rate
—
Efficiency
—
Pressure
—
Analysis time
—
Solvent consumption
—
Transferred method (2)
Transferred column geometry
Transferred exp. Conditions
L2 [mm]
dc2 [mm]
dp2 [µm]
Dwell vol. Vd2 [mL]
F2 (optimal) [µL/min]
—
Vinj2 [µL]
—
Additional gradient information
Suggested reconditioning step [min]
—
Geometric transfer of the gradient profile
Optimized transfer (non-optimal flow rate)
Flow-rate (F2) [µL/min]
Performance with user flow-rate
Flow rate
—
Pressure
—
Analysis time
—
Gradient transfer rules: Isocratic steps (initial hold, re-equilibration) scale with column dead time ratio. Gradient segments maintain constant %B range while adjusting slope to preserve column volumes during gradient. Dwell volume differences may require an initial isocratic hold or injection delay.
Developed by Davy GUILLARME — University of Geneva
Chromatographic performance in isocratic mode
Chromatographic conditions
Stationary support
Length [mm]
Diameter [mm]
Particle size [µm]
Organic solvent
Temperature [°C]
% Organic
Flow-rate [µL/min]
Solute molecular mass
Injection volume [µL]
Detector time constant [ms]
Average retention factor (k)
Chromatographic performance
Optimal flow rate [µL/min]
—
Suggested working flow rate [µL/min]
—
N theoretical (plates)
—
N observed (UPLC™ only)
—
% N loss (instrumentation)
—
% N loss (non-optimal F)
—
% N loss (total)
—
ΔP observed [bar]
—
Knox curve — Efficiency vs Flow rate
Pressure vs Flow rate
Knox equation: h = A·ν1/3 + B/ν + C·ν    where ν = u·dp/Dm (reduced velocity).
Pressure (Darcy/Kozeny-Carman): ΔP ≈ 1.1 · φ · η · L · u / dp²   (column parameters; φ≈500–800 depending on stationary phase).
Extra-column band broadening accounts for: injection volume, detector cell volume & time constant, connecting tubing (127 µm ID assumed).
Developed by Davy GUILLARME [davy.guillarme@unige.ch] — With contribution of Dao NGUYEN, Serge RUDAZ, Jean-Luc VEUTHEY
School of Pharmaceutical Sciences, University of Geneva, Switzerland
Chromatographic performance in gradient mode
Chromatographic conditions (gradient mode)
Stationary support
Length [mm]
Diameter [mm]
Particle size [µm]
Organic solvent
Temperature [°C]
Initial %B
Final %B
Gradient time [min]
Flow-rate [µL/min]
Final hold time [min]
Re-equilibration time [min]
Solute molecular mass
Injection volume [µL]
Detector time constant [ms]
Dwell volume [µL]
Gradient performance
Column dead time t₀ [min]
—
Peak capacity (P or nc)
—
ΔP observed [bar]
—
Total cycle time [min]
—
Solvent consumption [mL]
—
Gradient elution profile
Peak capacity vs Gradient time
Peak capacity in gradient elution (Snyder/Dolan): nc ≈ 1 + (√N/4) · (Δ%B · S · t₀) / (tG · ΔΦ · ln10)   (approximate calculation using average retention model).
Total cycle time includes: dwell-volume delay, gradient segment, final hold, and re-equilibration.
Developed by Davy GUILLARME — University of Geneva
About HPLC Calculator

HPLC CALCULATOR v3.1

Software for chromatographic performance evaluation and method transfer

HPLC calculator is free to use and distribute.

Version 3.1 — August 2020

All rights reserved to the University of Geneva, Group of Analytical Sciences.

For in-depth discussion or information on the calculations used, please contact:

davy.guillarme@unige.ch

Update your version regularly from:
https://ispso.unige.ch/labs/fanal/hplc_calculator:en

This calculator is provided as-is, without any expressed or implied warranty. In no event will the author be held liable for any consequences/damages arising from the use of this calculator.

This program and the accompanied tutorial are FREEWARE. You can use it for personal, business, educational or any other need, subject to the following restrictions:

  1. You may re-distribute this program as part of any free software suite/package after informing the author. Commercial redistribution requires written consent.
  2. This notice may not be removed or altered from any distribution.
  3. No form of decompilation / disassembling of the calculator may be done.
  4. You may not misrepresent the origin of the software.

Web reproduction: faithfully ported from the original Excel workbook
Based on the equations of Guillarme, Veuthey et al., University of Geneva