600e multisolvent delivery system

Reversed-phase high-performance liquid Chromatography (RP-HPLC) was performed on a Waters Chromatograph based on a 600 E multi-solvent delivery system coupled to a Waters 2998 photodiode array detector (Waters, Vienna, Austria) and a Gabi gamma-detector (Raytest, RSM Analytische Instrumente GmbH, Straubenhardt, Germany). The processing of data and Chromatography were controlled by Empower Software (Waters, Milford, MA, USA). For quality control, aliquots of the radiolabeling solution were loaded on a Symmetry Shield RP18 cartridge column (5 μm, 3.9 mm × 150 mm, Waters, Eschborn, Germany), eluted with the following linear gradient: 100%A/0% B to 70%A/30% B in 5 min and then 70%A/30% B to 55%A/45% B in 60 min, whereby A = 0.1% TFA in H₂O (v/v) and B = MeCN (system 1). The radiochemical labeling yields exceeded 98%, and the radiochemical purity was >99%; therefore, radioligands were used without further purification in all subsequent experiments. Samples of [¹¹¹In]In/[⁶⁷Ga]Ga-GAS1/2/3 and [¹⁷⁷Lu]Lu-GAS1/3 were tested before and after the end of all biological experiments.
Handling of solutions containing beta-/gamma-emitting radionuclides was conducted by authorized personnel in compliance with European radiation safety guidelines. Licensed facilities were supervised by the Greek Atomic Energy Commission (GAEC, license #A/435/17092/2019 and #A/435/15767/2019).

This analysis was performed by the calibration of a Waters controller 600
HPLC-DAD analysis (Waters. Millipore Corp., Waters Chromatography Division,
Milford, Ma, USA). The HPLC system was equipped with a Waters 600E
multisolvent delivery system with a Waters W996 diode array detector,
autosampler (Waters 717 Plus), and Software Millenium 32 (Waters). The
analytical HPLC separations were carried out on a Zorbax eclipse RP C18 (150
mm x 46 mm x 5 μm) column. The mobile phase consisted of an isocratic
elution with methanol-acetonitrile-water (20:30:50) and a flow rate of 0.8
mL min^-1. The standard PTOX was acquired from Sigma.

The chromatographic system used was a Waters apparatus (Milford, MA, USA) consisting of a pump (600E Multisolvent Delivery System), an auto sampler (700 Wisp model) and a differential refractive index (RI) detector (Waters 2414 model). Elution was performed at room temperature in a Protein KW-804 column (8 mm × 300 mm, Waters), and the mobile phase was phosphate-buffered saline (300 mM NaCl, 25 mM disodium hydrogen phosphate, pH 7.0) at a flow rate of 1.0 mL min⁻¹, and injection volume 25 µL. The data was collected and analyzed using the Millennium 32^® chromatography program (Waters). This software was used for chromatogram integration and estimating monomer and dimer content through relative peak area percentage values.

Separation of tannic acid, cathacol, ferulic acid and vaniline in the crude extract of A. pallens was achieved on HPLC system equipped with a PDA Detector. Xterra MSC-18 column (7.8 × 100 mm, 5 µm) with octadecylsilane as a solid support and 600e Multi Solvent Delivery System from Waters (USA) was used to separate the components. The mobile phase consists of methanol + acetonitrile:phosphate (30:15:55, v/v) (pH 3.5) and flow rate of the mobile phase was kept at 1.0 ml/min with isocratic elution. A linear gradient elution was used and detection was done at 280 nm in this method. The separated alkaloids, flavonoids and phenolic content were initially identified by a direct comparison of their retention times with those of standards. The contents of all compounds were calculated from the peak areas of HPLC chromatograms from the three replicate samples and the output was given in the units of ppm. The results were converted from ppm to mg/g. The data were analysed and processed using the installed Empower 2 software.