Reactions were analyzed for coproheme, harderoheme, and heme b by HPLC on an Agilent 1100 series instrument with a Phenomenex Luna C18 3μm column (150 mm x 4.6 mm). The solvents were H2O with 0.1% TFA (trifluoroacetic acid) (Solvent A) and acetonitrile (ACN) with 0.1% TFA (Solvent B). All samples were run at a flow rate of 1.5 mL/min starting with isocratic 80% A for 3 min, followed by a linear gradient transitioning to 5% A over 12 min. This was followed by 5% A for 5 min and a final isocratic run of 20 % A for 3 min. UV/vis detection was at 411 nm. Standard curves (0 – 500 pmol) were generated for quantifying heme species via integration of the peak areas. Samples were quantified in triplicate and error bars are ± 1 standard deviation.
Luna c18 3μm column
Manufactured by Phenomenex
Sourced in United States
The Luna C18 3μm column is a reversed-phase high-performance liquid chromatography (HPLC) column. It features a 3μm particle size and a C18 stationary phase, providing high-efficiency separation of a wide range of analytes.
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Lab products found in correlation
2 protocols using luna c18 3μm column
1
Quantitative HPLC Profiling of Heme Species
2
HPLC Purification and Mass Spectrometry
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Bioactive fractions (100 μl; produced as described above) were semi-purified on a Waters® HPLC system equipped with a Luna C18 (3 μm) column (Phenomenex Corporation, Torrance, CA, USA) by isothermal (35°C) gradient elution (97% H2O/3% Acetonitrile to 100% Acetonitrile) at a 1.0 mL/min flow rate with a photodiode array detector. Molecular mass determinations were made with a Waters® 1525 system equipped with a 2767 sample manager and Micromass ZQ Mass Spectrometer. Spectra were analyzed using the MassLynxTM software system.
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