Application Note: SPE Micro-HPLC with double throughput

Our neighbour CRO, Swiss BioQuant, use the Zirconium pump with a new scheme for loading and analyzing samples at the same time using the same binary pump. Dead times of the MS are greatly reduced, so they save a lot of instrument time while achieving exceptional sensitivity and precision.

Reference Assay

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LC Setup

The setup includes a novel technique (patent pending) of using a pump to serve a secondary flow path for each eluent. An external switch valve controls a split channel for each pump channel, which will not change the gradient on the analytical column thanks to the closed-loop flow control mode using a flow sensor for each channel right before mixing the eluents. The split flow is used to load the sample from the injector loop to the trapping column with the same pressure as the analytical flow. Its pressure and flow rate are determined by a restrictor capillary between the 10-port valve and waste.
Schema of the Zirconium Dual Trap Setup

In addition to the time savings of parallel loading and analyzing using two trapping columns, a big advantage is that switching the loaded trapping column in line with the analytical column does not produce a pressure drop, because the trapping column was already loaded under the same pressure.

Zirconium dual trap setup - detail

The ZircoFit capillary fitting system was used for all small-ID connections of the setup.

Instrument Methods

TSQ Quantum Instrument Method

Scan Events:
1: + c SRM Skimmer Offset 5, Micro Scans 1,
Parent Center Width Time CE
Mass transition Analyte: 544.180 to 361.100
Mass transition ISTD: 548.200 to 365.100
Collision energy [V]: 25
Positive Polarity
Spray Voltage [V] 3200
Capillary Temperature [°C] 350

LC Instrument Method

Analytical column: YMC, Triart C18, 150 x 0.3 mm
Trap columns: 2 x ZircoTrap™ with Reprospher  100 C18-TN 5 µm 8×1.1mm

Eluent A: water containing 0.1 % formic acid
Eluent B: acetonitrile containing 0.1 % formic acid


Wash cycle: 1.5 min ~15 µl/min ACN
Equilibration: 2.5 min ~15 µl/min H2O
Loading: 3 min H2O transfer from injection loop to trap column
No separate loading pump required!


Total flow: 5 µl/min
Pressure: ~150 bar

Equilibration time: 1 min
0 – 60 sec: 5% B isocratic
60 – 240 sec.: 5 to 95% B linear gradient
240 – 390 sec.: 95% isocratic


Chromatogram (complete). Click on picture for higher resolution.
Chromatogram – zoomed time scale. Click on picture for higher resolution.

The results in brief:

  • Same peak quality and reproducibility as with standard (splitless) Zirconium pumps
  • CV (area ratio): 3.1%
  • Gradient run time: 6.5 min
  • Total cycle time: ~7.5 min

The known precision of Zirconium gradients combined with optimized sample throughput is a very advantageous solution for this customer.