Reversed-phase (RP) is the dominating mode of HPLC and is used for separating small, predominantly neutral, molecules of different polarities with an extensive structural variety. RP HPLC is utilized within research & development and in quality control in an extremely wide range of industries, but especially within pharmaceutical, food & beverage, and environmental monitoring.
Diduco offers a range of services within RP method development and troubleshooting, and do also maintain an open-access instrument park for local collaboration partners.
The foundation of RP HPLC separations is hydrophobic partitioning which push the less polar molecules to be more distributed into the oil-like environment of the bonded stationary phase within RP columns, which often consists of octadecyl chains (i.e., C18). Differences between molecules in this distribution enables separation of them. In RP, a wide column selectivity range is available due to secondary interactions with the chemical structures of the stationary phase (e.g., C8, phenyl, biphenyl, fluorophenyl), or with the underlying silica base material and its end-capping.
Eluents in RP are based on mixtures of water and one or several organic solvents, typically methanol or acetonitrile, plus often a buffer or additive to control the pH and ionic strength. RP retention decrease with increasing organic solvent, and since the separation process works under a wide variety of conditions, elution can be performed from almost pure water to pure organic solvent, depending on the sample requirements. However, to maintain high reproducibility, gradient span in practise leaves a few percent admixture of the other component at each end point, typically while maintaining a constant pH and ionic strength throughout the analysis.
Instruments for HPLC are generally designed with RP mode separations in mind, and a range of detection options are available including UV light absorption, fluorescence (FL), refractive index (RI), evaporative light scattering (ELSD), charged aerosol (CAD), and mass spectrometry (MS), where the latter combination also has gained its own terminology with the abbreviation LC-MS. The recent few decades of developments within silica particle design and pump performance at ultra-high pressures have resulted in coining of the term UHPLC. This indicates an ability to accomplish separations at higher efficiency, with higher sensitivity, at higher speed and thus process a larger number of samples per time unit, but the underlying separation processes are identical to classical HPLC.
To learn more, browse the application examples below or contact Diduco to discuss your specific analysis.
The legislation in many countries allow cultivation of industrial hemp provided that the strains do not produce high amounts of tetrahydrocannabinol (THC) and other psychoactive cannabinoids classified as narcotic substances. These strains of cannabis sativa are instead typically rich in compounds such as cannabidiol (CBD) and cannabidiolic acid (CBDA).
The move towards a more sustainable world includes sourcing functional monomers for polymer synthesis from renewable natural products rather than from fossil-based sources. This green chemistry revolution often relies on the development of new reaction procedures efficiently catalysed by enzymes to involve fewer harmful chemicals.
Get in contact
I’ll be happy to answer any questions you might have regarding our products and services.
Tobias Jonsson
CEO, PhD in chemistry
Mobile: +46 705783490
E-mail: tobias@diduco.com
Diduco is a leading provider of professional services in high performance liquid chromatography and manufacturer of the Xenoic® XAMS chemical suppressor and ASUREX automatic regenerator for ion chromatography.
Diduco AB
Tvistevägen 48C
90736 Umeå
Sweden