Introduction

An analytical HPLC system using microparticulate packing materials requires a lower system volume, higher operating pressure and high speed response. Rapid separation HPLC (RHPLC) is likely to become the new standard for high performance analysis, covering measurement in a wide range of applications from conventional HPLC to some types of UHPLC. The following is an analysis report for the measurement of fatty acids using a Unifinepak C18 column with the RHPLC system including a refractive index detector for a semi-micro scale measurement.

LC-4000 RHPLC system

Experimental

Chromatographic conditions
Column:   A) Unifinepak C18 (4.6 mmI.D. x 150 mmL, 5 µm), B) Unifinepak C18 (2.0 D. x 150 mmL, 1.9 µm), C) Unifinepak C18 (2.0 D. x 30 mmL, 1.9 µm)
Eluent :   0.1%TFA/Acetonitrile (20/80)
Flow rate:   A; 1.2 mL/min, B-C; 0.6 mL/min
Column temp.:   40ºC
Injection volume:   A; 10 µL, B-C; 2 µL
Standard Sample:   Fatty acids mixture (1 mg/mL per component)

Keywords

RHPLC, fatty acids, semi-micro, ODS, Refractive index detector

Results

Figure 1 shows chromatograms of fatty acids, which were measured using the LC-4000 series RHPLC system with conventional HPLC and sub 2 μm UHPLC columns. The refractive index detector which is optimized for semi-micro scale measurement allows the acquisition of chromatograms with improved peak shape and a higher resolution separation. Unifinepak offers a common packing material in a range of particle sizes, which is independent of the measurement method, such as column size.

Figure 1. Chromatogram of fatty acid standard (1: Myristic acid, 2: Linoleic acid, 3: Palmitic acid, 4: Stearic acid)

Figure 2 shows chromatograms for an 8-component fatty acid mixture using columns with two different particle sizes (A column: 5 µm, B column: 1.9 µm).

Figure 2. Chromatogram of 8-component fatty acid mixture (1: Caprylic acid, 2: Capric acid, 3: Lauric acid, 4: Myristic acid,   5: Linoleic acid, 6: Palmitic acid, 7: Elaidic acid, 8: Stearic acid)